NTP TECHNICAL REPORT
 

ON THE
 

TOXICOLOGY AND CARCINOGENESIS
 

STUDIES OF
 

2-BUTOXYETHANOL
 

(CAS NO. 111-76-2) 

IN F344/N RATS AND B6C3F1 MICE 

(INHALATION STUDIES) 

NATIONAL TOXICOLOGY PROGRAM
 

P.O. Box 12233
 

Research Triangle Park, NC 27709
 

March 2000 

NTP TR 484
 

NIH Publication No. 00-3974
 

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES
 

Public Health Service
 

National Institutes of Health
 
FOREWORD
 

The National Toxicology Program (NTP) is made up of four charter agencies of the U.S. Department of 

Health and Human Services (DHHS): the National Cancer Institute (NCI), National Institutes of Health; the 

National Institute of Environmental Health Sciences (NIEHS), National Institutes of Health; the National 

Center for Toxicological Research (NCTR), Food and Drug Administration; and the National Institute for 

Occupational Safety and Health (NIOSH), Centers for Disease Control and Prevention. In July 1981, the 

Carcinogenesis Bioassay Testing Program, NCI, was transferred to the NIEHS. The NTP coordinates the 

relevant programs, staff, and resources from these Public Health Service agencies relating to basic and 

applied research and to biological assay development and validation. 

The NTP develops, evaluates, and disseminates scientific information about potentially toxic and hazardous 

chemicals. This knowledge is used for protecting the health of the American people and for the primary 

prevention of disease. 

The studies described in this Technical Report were performed under the direction of the NIEHS and were 

conducted in compliance with NTP laboratory health and safety requirements and must meet or exceed all 

applicable federal, state, and local health and safety regulations. Animal care and use were in accordance 

with the Public Health Service Policy on Humane Care and Use of Animals. The prechronic and chronic 

studies were conducted in compliance with Food and Drug Administration (FDA) Good Laboratory Practice 

Regulations, and all aspects of the chronic studies were subjected to retrospective quality assurance audits 

before being presented for public review. 

These studies are designed and conducted to characterize and evaluate the toxicologic potential, including 

carcinogenic activity, of selected chemicals in laboratory animals (usually two species, rats and mice). 

Chemicals selected for NTP toxicology and carcinogenesis studies are chosen primarily on the bases of 

human exposure, level of production, and chemical structure. The interpretive conclusions presented in this 

Technical Report are based only on the results of these NTP studies. Extrapolation of these results to other 

species and quantitative risk analyses for humans require wider analyses beyond the purview of these 

studies. Selection per se is not an indicator of a chemical’s carcinogenic potential. 

Listings of all published NTP reports and ongoing studies are available from NTP Central Data 

Management, NIEHS, P.O. Box 12233, MD E1-02, Research Triangle Park, NC 27709 (919-541-3419). 

The Abstracts and other study information for 2-year studies are also available at the NTP’s World Wide 

Web site: http://ntp-server.niehs.nih.gov. 
NTP TECHNICAL REPORT
 

ON THE
 

TOXICOLOGY AND CARCINOGENESIS
 

STUDIES OF
 

2-BUTOXYETHANOL
 

(CAS NO. 111-76-2) 

IN F344/N RATS AND B6C3F1 MICE 

(INHALATION STUDIES) 

NATIONAL TOXICOLOGY PROGRAM
 

P.O. Box 12233
 

Research Triangle Park, NC 27709
 

March 2000 

NTP TR 484
 

NIH Publication No. 00-3974
 

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES
 

Public Health Service
 

National Institutes of Health
 
2 2-Butoxyethanol, NTP TR 484 

CONTRIBUTORS
 

National Toxicology Program 

Evaluated and interpreted results and reported findings 

J.H. Roycroft, Ph.D., Study Scientist 

D.A. Bridge, B.S. 

J.R. Bucher, Ph.D. 

L.T. Burka, Ph.D. 

B.I. Ghanayem, Ph.D. 

J.R. Hailey, D.V.M. 

J.K. Haseman, Ph.D. 

R.R. Maronpot, D.V.M. 

A. Nyska, D.V.M. 

G.N. Rao, D.V.M., Ph.D. 

C.S. Smith, Ph.D. 

G.S. Travlos, D.V.M. 

K.L. Witt, M.S., Integrated Laboratory Systems 

Battelle Pacific Northwest Laboratories 

Conducted studies, evaluated pathology findings 

B.J. Chou, D.V.M., Ph.D., Principal Investigator 

J.A. Dill, Ph.D. 

S.L. Grumbein, D.V.M., Ph.D 

K.M. Lee, Ph.D. 

R.A. Miller, D.V.M., Ph.D. 

H.A. Ragan, D.V.M. 

S.E. Rowe, D.V.M., M.S. 

R.B. Westerberg, Ph.D. 

Experimental Pathology Laboratories, Inc. 

Provided pathology quality assurance 

J.F. Hardisty, D.V.M., Principal Investigator 

E.T. Gaillard, D.V.M., M.S. 

C.C. Shackelford, D.V.M., M.S., Ph.D. 

Dynamac Corporation 

Prepared quality assurance audits 

S. Brecher, Ph.D., Principal Investigator 

NTP Pathology Working Group 

Evaluated slides, prepared pathology report on rats 

(14 October 1997) 

M.P. Jokinen, D.V.M., Chairperson 

Pathology Associates International 

T. Amanuma, D.V.M., Observer 

National Toxicology Program 

R. Cattley, V.M.D., Ph.D. 

Chemical Industry Institute of Toxicology 

E.T. Gaillard, D.V.M., M.S. 

Experimental Pathology Laboratories, Inc. 

R.H. Garman, D.V.M., Observer 

Canadian Medical Association 

R.A. Herbert, D.V.M., Ph.D. 

National Toxicology Program 

J.R. Leininger, D.V.M., Ph.D. 

National Toxicology Program 

A. Radovsky, D.V.M., Ph.D. 

National Toxicology Program 

Evaluated slides, prepared pathology report on mice 

(25 September 1997) 

M.P. Jokinen, D.V.M., Chairperson 

Pathology Associates International 

R.H. Garman, D.V.M., Observer 

Canadian Medical Association 

J.E. Heath, D.V.M. 

Southern Research Institute 

R.A. Herbert, D.V.M., Ph.D. 

National Toxicology Program 

J.R. Leininger, D.V.M., Ph.D. 

National Toxicology Program 

J. Mahler, D.V.M., Ph.D. 

National Toxicology Program 

R.A. Miller, D.V.M., Ph.D. 

Battelle Pacific Northwest Laboratories 

A. Radovsky, D.V.M., Ph.D. 

National Toxicology Program 

C.C. Shackelford, D.V.M., M.S., Ph.D. 

Experimental Pathology Laboratories, Inc. 

A. Singer, D.V.M. 

Battelle Columbus Laboratories 
2-Butoxyethanol, NTP TR 484 3 

Analytical Sciences, Inc. 

Provided statistical analyses 

R.W. Morris, M.S., Principal Investigator 

K.P. McGowan, M.B.A. 

M.A. Mauney, M.S. 

N.G. Mintz, B.S. 

J.T. Scott, M.S. 

Biotechnical Services, Inc. 

Prepared Technical Report 

S.R. Gunnels, M.A., Principal Investigator 

J.R. Carlton, B.A. 

L.M. Harper, B.S. 

T.S. Kumpe, M.A. 

A.M. Macri-Hanson, M.A., M.F.A. 

M.L. Rainer, B.S. 
4 

CONTENTS
 

ABSTRACT ............................................................ 5 
 

EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTIVITY ............ 10 
 

TECHNICAL REPORTS REVIEW SUBCOMMITTEE ............................... 11 
 

SUMMARY OF TECHNICAL REPORTS REVIEW SUBCOMMITTEE COMMENTS ......... 12 
 

INTRODUCTION ........................................................ 15 
 

MATERIALS AND METHODS ............................................... 33 
 

RESULTS .............................................................. 43 
 

DISCUSSION AND CONCLUSIONS ........................................... 81 
 

REFERENCES .......................................................... 87 
 

APPENDIX A Summary of Lesions in Male Rats in the 2-Year Inhalation Study
 

of 2-Butoxyethanol ............................................. 95 
 

APPENDIX B Summary of Lesions in Female Rats in the 2-Year Inhalation Study
 

of 2-Butoxyethanol ............................................. 133 
 

APPENDIX C Summary of Lesions in Male Mice in the 2-Year Inhalation Study
 

of 2-Butoxyethanol ............................................. 167 
 

APPENDIX D Summary of Lesions in Female Mice in the 2-Year Inhalation Study
 

of 2-Butoxyethanol ............................................. 203 
 

APPENDIX E Genetic Toxicology ............................................. 239 
 

APPENDIX F Hematology and Bone Marrow Cellularity Results ....................... 249 
 

APPENDIX G Organ Weights and Organ-Weight-to-Body-Weight Ratios .................. 261 
 

APPENDIX H Chemical Characterization and Generation of Chamber Concentrations ......... 265 
 

APPENDIX I Ingredients, Nutrient Composition, and Contaminant Levels
 

in NIH-07 Rat and Mouse Ration ................................... 277 
 

APPENDIX J Sentinel Animal Program ........................................ 281 
 

APPENDIXK H-ras Codon 61 Mutation Spectra in Forestomach Neoplasms
 

from B6C3F1 Mice Exposed to 2-Butoxyethanol for 2 Years ................. 285 
 
 

 

5 

ABSTRACT
 

CH3-CH2-CH2-CH2-O-CH2-CH2-OH 

2-BUTOXYETHANOL 

CAS No. 111-76-2 

Chemical Formula: C6H14O2 Molecular Weight: 118.17 

Synonyms: 2-Butoxy-1-ethanol; m-butyl ether; butyl glycol; ethylene glycol monobutyl ether 

Trade name: Butyl Cellosolve 

2-Butoxyethanol is a member of a family of ethylene 

glycol monoalkyl ethers. It is used extensively as a 

solvent in surface coatings such as lacquers, 

enamels, varnishes, and latex paint; in paint thinners, 

paint stripping formulations, and inks; and in 

degreasers and industrial and household cleaners. 

2-Butoxyethanol was nominated for study because of 

its widespread use in industrial and consumer 

applications, the potential for exposure to workers and 

the general population, and the absence of chronic 

toxicity data. Male and female F344/N rats 

and B6C3F1 mice were exposed to 2-butoxyethanol 

(greater than 99% pure) by inhalation (primary route 

of human exposure) for 14 weeks or 2 years. Genetic 

toxicology studies were conducted in Salmonella 

typhimurium, cultured Chinese hamster ovary cells, 

and the bone marrow of male F344/N rats and 

B6C3F1 mice. 

14-WEEK STUDY IN RATS 

Groups of 10 male and 10 female rats were exposed 

to 2-butoxyethanol by inhalation at concentrations of 

0, 31, 62.5, 125, 250, or 500 ppm, 6 hours per day, 

5 days per week for 14 weeks. One female rat in the 

250 ppm group was killed moribund during week 8; 

four females in the 500 ppm group were killed moribund during week 1 and one during week 5. Final 

mean body weights of females exposed to 500 ppm 

were significantly less than those of the chamber controls. Clinical findings included abnormal breathing, 

pallor, red urine stains, nasal and eye discharge, 

lethargy, and increased salivation and/or lacrimation. 

Due to vascular thrombosis and infarction in the tail 

vertebrae of 500 ppm female rats, the tails became 

necrotic and either sloughed off or were chewed off. 

The primary effect on the hematopoietic system was 

an anemia characterized as macrocytic, normochromic, and regenerative in males exposed to 

125 ppm or greater and, to a greater extent, in all 

exposed groups of females. Compared to the 

chamber controls, kidney weights of males exposed to 

500 ppm and females exposed to 125 ppm or greater 

and liver weights of males exposed to 250 or 500 ppm 

and females exposed to 125 ppm or greater were significantly increased, and thymus weights of females 

exposed to 500 ppm were significantly less. In female 

rats killed moribund, there was considerable 

histologic evidence of thrombosis in tissues and 

organs including the nasal cavity, incisors, liver, 

lung, and heart. In addition to thrombosis, infarction 

occurred in the vertebrae of the tail resulting in 

necrosis and loss of the distal portion of the tail. 

There were also inflammation, necrosis, and 

ulceration of the forestomach; necrosis and 

centrilobular degeneration of the liver; renal tubule 
 

 

6 2-Butoxyethanol, NTP TR 484 

degeneration; and atrophy of the spleen and thymus. 

Exposure-related increases in the incidences of 

Kupffer cell pigmentation, forestomach inflammation 

and epithelial hyperplasia, bone marrow hyperplasia, 

splenic hematopoietic cell proliferation, and renal 

tubule pigmentation were observed in male and/or 

female rats surviving to the end of the study. 

14-WEEK STUDY IN MICE 

Groups of 10 male and 10 female mice were exposed 

to 2-butoxyethanol by inhalation at concentrations of 

0, 31, 62.5, 125, 250, or 500 ppm, 6 hours per day, 

5 days per week for 14 weeks. Two male and two 

female mice exposed to 500 ppm died and two males 

and two females were killed moribund during the first 

2 weeks of the study. Final mean body weights of 

125, 250, and 500 ppm male mice were significantly 

less than those of the chamber controls. Clinical 

findings were observed only in 500 ppm males and 

females that died or were killed moribund and 

included abnormal breathing, red urine stains, and 

lethargy. Hematologic evaluation indicated an anemia 

that was characterized as normocytic, normochromic, 

and regenerative in mice exposed to 62.5 ppm or 

greater; the anemia was more pronounced in females. 

Liver weights of males exposed to 500 ppm were significantly greater than the chamber controls. In mice 

either dying early or killed moribund, there were 

inflammation, necrosis, and ulceration of the forestomach; mediastinal pleura and peritoneal inflammation associated with the forestomach lesions; liver 

necrosis; renal tubule degeneration; atrophy of the 

spleen, thymus, and mandibular and mesenteric lymph 

nodes; and degeneration of the testis. Exposurerelated increases in the incidences of hematopoietic 

cell proliferation and hemosiderin pigmentation of the 

spleen, Kupffer cell hemosiderin pigmentation of the 

liver, inflammation and epithelial hyperplasia of the 

forestomach, and renal tubule hemosiderin pigmentation occurred in male and/or female mice 

surviving to the end of the study. 

2-YEAR STUDY IN RATS 

Groups of 50 male and 50 female rats were exposed 

to 2-butoxyethanol by inhalation at concentrations of 

0, 31.2, 62.5, or 125 ppm, 6 hours per day, 5 days 

per week for 104 weeks. For hematology and bone 

marrow analyses, additional groups of 27 male and 

27 female rats were exposed to 0, 62.5, or 125 ppm 

for evaluation at 3, 6, and 12 months and nine male 

and nine female rats were exposed to 31.2 ppm for 

evaluation at 3 (hematology only) and 6 months. 

Survival and Body Weights 

Survival of exposed male and female rats was similar 

to the chamber control groups. The mean body 

weights of females exposed to 125 ppm were generally less than the chamber control group. 

Hematology and Bone Marrow Cellularity 

The most consistent exposure-related effect on the 

hematopoietic system was an exposure concentrationrelated mild macrocytic, normochromic, regenerative 

anemia present at 3, 6, and 12 months, with females 

more affected than males. Significant increases in 

bone marrow cellularity and decreases in the myeloid/ 

erythroid ratio relative to the chamber controls were 

observed at all time points in females exposed to 

125 ppm, and a decrease in the myeloid/erythroid 

ratio was observed in males exposed to 125 ppm at 

12 months. 

Pathology Findings 

The incidence of benign or malignant pheochromocytoma (combined) of the adrenal medulla in 

females exposed to 125 ppm was not significantly 

increased compared to the chamber controls but 

exceeded the historical control range. Exposurerelated increases in the incidences of hyaline degeneration of the olfactory epithelium and Kupffer cell 

pigmentation of the liver were observed in male and 

female rats. 

2-YEAR STUDY IN MICE 

Groups of 50 male and 50 female mice were exposed 

to 2-butoxyethanol by inhalation at concentrations of 

0, 62.5, 125, or 250 ppm, 6 hours per day, 5 days 

per week for 104 weeks. For hematology and bone 

marrow analyses, additional groups of 30 male and 

30 female mice were exposed to 0, 62.5, 125, or 

250 ppm for evaluation at 3, 6, and 12 months. 

Survival and Body Weights 

Survival of male mice exposed to 125 or 250 ppm was 

significantly less than that of the chamber control 

group. The mean body weights of exposed males 
 

 

2-Butoxyethanol, NTP TR 484 7 

were generally less than those of the chamber control 

group during the last 6 months of the study. The 

mean body weights of exposed female mice were less 

than those of the chamber control group; the reductions were greater and occurred earlier than those 

observed in males. 

Hematology 

The most consistent exposure-related effect on the 

hematopoietic system was an exposure concentrationrelated minimal normocytic, normochromic, regenerative anemia present at 3, 6, and 12 months, with 

females affected slightly more than males. 

Pathology Findings 

In females exposed to 250 ppm, incidences of forestomach squamous cell papilloma and squamous cell 

papilloma or carcinoma (combined) were significantly 

increased relative to the chamber controls, and these 

incidences exceeded the ranges in historical chamber 

controls. In 2-butoxyethanol exposed males, there 

were possible exposure-related increases in the incidences of squamous cell papilloma of the forestomach, although the increases were not significant 

and the incidences were within the historical control 

range for chamber controls. Accompanying these 

neoplasms in females and, to a lesser extent, in males 

were exposure-related increases in the incidences of 

ulcer and epithelial hyperplasia of the forestomach. 

In male mice exposed to 250 ppm, the incidence of 

hemangiosarcoma of the liver was significantly 

increased relative to chamber controls and exceeded 

the range in historical controls; in addition, there 

were possible exposure-related increases in the incidence of hepatocellular carcinoma. Incidences of 

hemosiderin pigmentation in the Kupffer cells were 

significantly increased in 125 and 250 ppm males and 

all exposed groups of females. 

The incidences of splenic hematopoietic cell proliferation and hemosiderin pigmentation were generally increased in males and females, and the incidences of bone marrow hyperplasia were increased in 

males. The incidences of hyaline degeneration of the 

olfactory and respiratory epithelia of the nose were 

increased in female mice. 

GENETIC TOXICOLOGY 

2-Butoxyethanol did not induce mutations in any of 

the S. typhimurium strains tested, with or without 

induced hamster or rat liver S9. 2-Butoxyethanol 

induced cycle delay but did not induce either sister 

chromatid exchanges or chromosomal aberrations in 

cultured Chinese hamster ovary cells with or without 

S9. 2-Butoxyethanol did not induce micronuclei in 

bone marrow cells of male rats or mice administered 

the chemical by intraperitoneal injection three times at 

24-hour intervals. 

CONCLUSIONS 

Under the conditions of these 2-year inhalation 

studies, there was no evidence of carcinogenic 

activity* of 2-butoxyethanol in male F344/N rats 

exposed to 31.2, 62.5, or 125 ppm. There was 

equivocal evidence of carcinogenic activity of 

2-butoxyethanol in female F344/N rats based on the 

increased combined incidences of benign or malignant 

pheochromocytoma (mainly benign) of the adrenal 

medulla. There was some evidence of carcinogenic 

activity of 2-butoxyethanol in male B6C3F1 mice 

based on increased incidences of hemangiosarcoma of 

the liver. A marginal increase in the incidences of 

forestomach squamous cell papilloma and an increase 

in the incidences of hepatocellular carcinoma may 

have been exposure related. There was some 

evidence of carcinogenic activity of 2-butoxyethanol 

in female B6C3F1 mice based on increased incidences 

of forestomach squamous cell papilloma or carcinoma 

(mainly papilloma). 

Increased incidences of forestomach neoplasms in 

male and female mice occurred in groups in which 

ulceration and hyperplasia were also present. 

Exposure to 2-butoxyethanol caused a mild regenerative anemia and effects secondary to the anemia. 

* Explanation of Levels of Evidence of Carcinogenic Activity is on page 10. A summary of the Technical Reports Review Subcommittee 

comments and public discussion on this Technical Report appears on page 12. 
8 2-Butoxyethanol, NTP TR 484 

Summary of the 2-Year Carcinogenesis and Genetic Toxicology Studies of 2-Butoxyethanol
 

Male 

F344/N Rats 

Female 

F344/N Rats 

Male 

B6C3F1 Mice 

Female 

B6C3F1 Mice 

Concentrations 

in air 

Chamber control, 31.2, 

62.5, and 125 ppm 

Chamber control, 31.2, 

62.5, and 125 ppm 

Chamber control, 62.5, 

125, and 250 ppm 

Chamber control, 62.

125, and 250 ppm 

5, 

Body weights Exposed groups similar to 

the chamber control group 

125 ppm group less than 

the chamber control group 

Exposed groups generally 

less than the chamber 

control group 

Exposed groups less t

the chamber control g

han 

roup 

Survival rates 19/50, 11/50, 21/50, 24/50 29/50, 27/50, 23/50, 21/50 39/50, 39/50, 27/50, 26/50 29/50, 31/50, 33/50, 36/50 

Nonneoplastic 1/50, 

effects 

Nose: olfactory 

epithelium, hyaline 

degeneration (13/48, 

21/49, 23/49, 40/50) 

Nose: olfactory 

epithelium, hyaline 

degeneration (13/50, 

18/48, 28/50, 40/49) 

Forestomach: ulcer (1/50, 

2/50, 9/49, 3/48); 

epithelium hyperplasia 

(1/50, 7/50, 16/49, 21/48) 

Forestomach: ulcer (

7/50, 13/49, 22/50); 

epithelium hyperplasia 

(6/50, 27/50, 42/49, 

Liver: Kupffer cell 

pigmentation (23/50, 

30/50, 34/50, 42/50) 

Liver: Kupffer cell 

pigmentation (15/50, 

19/50, 36/50, 47/50) 

Liver: Kupffer cell 

pigmentation (0/50, 0/50, 

8/49, 30/49) 

Spleen: hematopoietic cell 

proliferation (12/50, 

11/50, 26/48, 42/49); 

hemosiderin pigmentation 

(0/50, 6/50, 45/48, 44/49) 

Bone Marrow: hyperplasia 

(0/50, 1/50, 9/49, 5/50) 

44/50) 

Liver: Kupffer cell 

pigmentation (0/50, 5

25/49, 44/50) 

Spleen: hematopoieti

proliferation (24/50, 

29/50, 32/49, 35/50); 

hemosiderin pigmenta

(39/50, 44/50, 46/49, 

48/50) 

/50, 

c cell 

tion 

Neoplastic effects None None Liver: hemangiosarcoma 

(0/50, 1/50, 2/49, 4/49) 

Nose: olfactory 

epithelium, hyaline 

degeneration (6/50, 1

11/49, 12/50); respira

epithelium, hyaline 

degeneration (17/50, 

35/50, 26/49, 23/50) 

Forestomach: squam

cell papilloma (0/50, 

2/50, 5/50); squamou

papilloma or carcino

ous 

ma 

4/50, 

tory 

1/50, 

s cell 

Uncertain 

findings 

None Adrenal Medulla: benign 

or malignant 

pheochromocytoma (3/50, 

4/50, 1/49, 8/49) 

Forestomach: squamous 

cell papilloma (1/50, 1/50, 

2/49, 2/49) 

Liver: hepatocellular 

carcinoma (10/50, 11/50, 

(0/50, 1/50, 2/50, 6/5

None 

0) 

Level of evidence 

of carcinogenic 

activity 

No evidence Equivocal evidence 

16/49, 21/49) 

Some evidence Some evidence 
2-Butoxyethanol, NTP TR 484 9 

Summary of the 2-Year Carcinogenesis and Genetic Toxicology Studies of 2-Butoxyethanol
 

Genetic toxicology 

Salmonella typhimurium gene mutations: Negative in strains TA97, TA98, TA100, TA1535, and TA1537, with and without S9 

Sister chromatid exchanges

 Cultured Chinese hamster ovary cells in vitro: Negative with and without S9 

Chromosomal aberrations

 Cultured Chinese hamster ovary cells in vitro: Negative with and without S9 

Micronucleated erythrocytes

 Rat bone marrow in vivo: Negative

 Mouse bone marrow in vivo: Negative 
10 2-Butoxyethanol, NTP TR 484 

EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTIVITY 

The National Toxicology Program describes the results of individual experiments on a chemical agent and notes the strength of the evidence 

for conclusions regarding each study. Negative results, in which the study animals do not have a greater incidence of neoplasia than control 

animals, do not necessarily mean that a chemical is not a carcinogen, inasmuch as the experiments are conducted under a limited set of 

conditions. Positive results demonstrate that a chemical is carcinogenic for laboratory animals under the conditions of the study and 

indicate that exposure to the chemical has the potential for hazard to humans. Other organizations, such as the International Agency for 

Research on Cancer, assign a strength of evidence for conclusions based on an examination of all available evidence, including animal 

studies such as those conducted by the NTP, epidemiologic studies, and estimates of exposure. Thus, the actual determination of risk to 

humans from chemicals found to be carcinogenic in laboratory animals requires a wider analysis that extends beyond the purview of these 

studies. 

Five categories of evidence of carcinogenic activity are used in the Technical Report series to summarize the strength of the evidence 

observed in each experiment: two categories for positive results (clear evidence and some evidence); one category for uncertain findings 

(equivocal evidence); one category for no observable effects (no evidence); and one category for experiments that cannot be evaluated 

because of major flaws (inadequate study). These categories of interpretative conclusions were first adopted in June 1983 and then revised 

in March 1986 for use in the Technical Report series to incorporate more specifically the concept of actual weight of evidence of 

carcinogenic activity. For each separate experiment (male rats, female rats, male mice, female mice), one of the following five categories is 

selected to describe the findings. These categories refer to the strength of the experimental evidence and not to potency or mechanism. 

•	 Clear evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing a dose-related (i) 

increase of malignant neoplasms, (ii) increase of a combination of malignant and benign neoplasms, or (iii) marked 

increase of benign neoplasms if there is an indication from this or other studies of the ability of such tumors to progress to 

malignancy. 

•	 Some evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing a chemical-related
 

increased incidence of neoplasms (malignant, benign, or combined) in which the strength of the response is less than that
 

required for clear evidence.
 

•	 Equivocal evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing a marginal increase 

of neoplasms that may be chemical related. 

•	 No evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing no chemical-related
 

increases in malignant or benign neoplasms.
 

•	 Inadequate study of carcinogenic activity is demonstrated by studies that, because of major qualitative or quantitative
 

limitations, cannot be interpreted as valid for showing either the presence or absence of carcinogenic activity.
 

When a conclusion statement for a particular experiment is selected, consideration must be given to key factors that would extend the actual 

boundary of an individual category of evidence. Such consideration should allow for incorporation of scientific experience and current 

understanding of long-term carcinogenesis studies in laboratory animals, especially for those evaluations that may be on the borderline 

between two adjacent levels. These considerations should include: 

• adequacy of the experimental design and conduct;
 

• occurrence of common versus uncommon neoplasia;
 

• progression (or lack thereof) from benign to malignant neoplasia as well as from preneoplastic to neoplastic lesions; 

•	 some benign neoplasms have the capacity to regress but others (of the same morphologic type) progress. At present, it is 

impossible to identify the difference. Therefore, where progression is known to be a possibility, the most prudent course 

is to assume that benign neoplasms of those types have the potential to become malignant; 

•	 combining benign and malignant tumor incidence known or thought to represent stages of progression in the same organ
 

or tissue;
 

• latency in tumor induction; 

•	 multiplicity in site-specific neoplasia; 

•	 metastases; 

•	 supporting information from proliferative lesions (hyperplasia) in the same site of neoplasia or in other experiments (same 

lesion in another sex or species); 

• presence or absence of dose relationships; 

•	 statistical significance of the observed tumor increase; 

•	 concurrent control tumor incidence as well as the historical control rate and variability for a specific neoplasm; 

•	 survival-adjusted analyses and false positive or false negative concerns; 

•	 structure-activity correlations; and 

• in some cases, genetic toxicology. 
 

2-Butoxyethanol, NTP TR 484 11 

NATIONAL TOXICOLOGY PROGRAM BOARD OF SCIENTIFIC COUNSELORS
 

TECHNICAL REPORTS REVIEW SUBCOMMITTEE
 

The members of the Technical Reports Review Subcommittee who evaluated the draft NTP Technical Report on 2-butoxyethanol on 

30 October 1998 are listed below. Subcommittee members serve as independent scientists, not as representatives of any institution, 

company, or governmental agency. In this capacity, subcommittee members have five major responsibilities in reviewing the NTP studies: 

• to ascertain that all relevant literature data have been adequately cited and interpreted, 

• to determine if the design and conditions of the NTP studies were appropriate, 

• to ensure that the Technical Report presents the experimental results and conclusions fully and clearly, 

• to judge the significance of the experimental results by scientific criteria, and 

• to assess the evaluation of the evidence of carcinogenic activity and other observed toxic responses. 

Gary P. Carlson, Ph.D., Chairperson 

School of Health Sciences 

Purdue University 

West Lafayette, IN 

A. John Bailer, Ph.D. 

Department of Mathematics and Statistics 

Miami University 

Oxford, OH 

Steven A. Belinsky, Ph.D. 

Inhalation Toxicology Research Institute 

Kirkland Air Force Base 

Albuquerque, NM 

James S. Bus, Ph.D. 

Health and Environmental Sciences 

Dow Chemical Company 

Midland, MI 

Linda A. Chatman, D.V.M.* 

Pfizer, Inc. 

Groton, CT 

John M. Cullen, V.M.D., Ph.D. 

Department of Microbiology, Parasitology, and Pathology 

College of Veterinary Medicine 

North Carolina State University 

Raleigh, NC 

* Did not attend 

Susan M. Fischer, Ph.D.* 

M.D. Anderson Cancer Center
 

The University of Texas
 

Smithville, TX
 

Thomas L. Goldsworthy, Ph.D.* 

Integrated Laboratory Systems 

Research Triangle Park, NC 

Stephen S. Hecht, Ph.D. 

University of Minnesota Cancer Centers 

Minneapolis, MN 

Michele Medinsky, Ph.D. 

Chemical Industry Institute of Toxicology 

Research Triangle Park, NC 

Jose Russo, M.D.* 

Fox Chase Cancer Center 

Philadelphia, PA 
 

 

 

 

12 2-Butoxyethanol, NTP TR 484 

SUMMARY OF TECHNICAL REPORTS REVIEW SUBCOMMITTEE COMMENTS
 

On 30 October 1998 the draft Technical Report 

on the toxicology and carcinogenesis studies 

of 2-butoxyethanol received public review by the 

National Toxicology Program’s Board of Scientific 

Counselors’ Technical Reports Review Subcommittee. 

The review meeting was held at the National Institute 

of Environmental Health Sciences, Research Triangle 

Park, NC. 

Dr. J.H. Roycroft, NIEHS, introduced the toxicology 

and carcinogenesis studies of 2-butoxyethanol by 

discussing the uses of the chemical and the rationale 

for study, describing the experimental design, reporting on the survival and body weight effects, and 

commenting on compound-related neoplastic and 

nonneoplastic lesions in rats and mice. In addition to 

the standard core study, a number of animals were 

assessed for hematologic parameters and bone marrow 

cellularity and myeloid/erythroid ratios. Additionally, 

animals were included in the design for toxicokinetic 

measures of 2-butoxyethanol and its principal 

metabolite, 2-butoxyacetic acid. The proposed conclusions for the 2-year studies were no evidence of 

carcinogenic activity in male F344/N rats, equivocal 

evidence of carcinogenic activity in female F344/N 

rats, and some evidence of carcinogenic activity in 

male and female B3C6F1 mice. 

Dr. Medinsky, a principal reviewer, agreed in principle with the proposed conclusions. Her concern 

was that the proposed conclusions for female rats and 

male mice were made based on differences in 

response of the test animals compared with historical 

control values and were not based on differences in 

response compared to respective controls. She asked 

what objective statistical measure of differences was 

used. Dr. Roycroft responded that, as a rule, neoplasm data are not compared statistically with 

historical control data because so many factors can 

vary from study to study. The concurrent controls are 

still considered the most appropriate control group. 

Dr. J.K. Haseman, NIEHS, said that many factors, 

such as whether there were increases in incidences of 

preneoplastic lesions, factored into a decision. 

Dr. Medinsky commented that one of the report’s 

strengths was the comprehensive section on the 

chemical disposition and toxicokinetics of 

2-butoxyethanol and 2-butoxyacetic acid and 

suggested a summary paragraph for the chemical 

disposition and toxicokinetics data (pages 19-20). 

Dr. Bailer, the second principal reviewer, agreed with 

the proposed conclusions for rats but not for mice. 

He thought that not enough consideration was given to 

the strong exposure-related trends in the neoplasm 

data in mice. He asked for clarification as to why the 

findings did not support a conclusion of clear 

evidence in mice. Dr. Roycroft said that benign and 

malignant neoplasms are analyzed independently and 

in combination, with the most important being the 

combined neoplasms. For male mice, the combined 

incidences of hepatocellular adenoma or carcinoma 

did not increase, and the incidences of carcinoma 

alone were within the historical control range. 

Dr. J.R. Hailey, NIEHS, noted that whether more 

emphasis is given to the combined neoplasm incidence 

depends somewhat on the neoplasm type. With liver 

neoplasms, there is a morphologic and biologic 

continuum of progression from foci to adenomas and 

to carcinomas. Further, it is often difficult to 

distinguish benign from malignant neoplasms. 

Dr. Cullen, the third principal reviewer, agreed with 

the proposed conclusions. He said that, in the liver of 

male mice, reliance on historical control incidence 

and lack of concordant increases in preneoplastic and 

benign lesions supported less than clear evidence, but 

the data appeared to reflect at least equivocal 

evidence. In female mice, he found the proposed 

conclusion regarding squamous papillomas appropriate. Dr. Cullen commented that some of the toxic 

effects attributed directly to action of the chemical 

might be addressed as secondary responses due to 

other insults created by the chemical, e.g., anemia in 

response to blood loss from gastric ulceration in 

female, and perhaps male, mice. 

Dr. T.R. Tyler, Chemical Manufacturers Association 

Ethylene Glycol Ethers Panel, stated that 

2-butoxyethanol has long been recognized primarily 

as a hemolytic agent, with humans being less 

susceptible than rodents. Regarding the forestomach 

neoplasms in female mice, he thought that some 
 

2-Butoxyethanol, NTP TR 484 13 

evidence was probably correct but likely irrelevant 

because there is no such organ in humans. Regarding 

the pheochromocytomas in female rats, he asked the 

Subcommittee to reconsider the designation of 

equivocal evidence as there were no statistically 

significant pairwise comparisons, the incidence was 

barelyoutside the historical control range, and there 

was no indication of increased incidences in males. 

Dr. R. Boatman, Eastman Chemical Company, 

thought that hemangiosarcomas of the liver in male 

mice represented a marginal or equivocal finding. He 

compared the results for this study with those from 

the NTP bioassay of p-nitroaniline (NTP, 1993a), for 

which similar incidences of hemangiosarcomas of the 

liver in male mice were classified as equivocal evidence. Further, he stated that the possibility that the 

study was compromised by Helicobacter infection 

could not be ruled out. Dr. Medinsky asked for staff 

comment on the p-nitroaniline study. Dr. Roycroft 

responded that the p-nitroaniline study was a gavage 

study and that the historical control range and high 

incidence for gavage studies at the time were slightly 

higher than the range and high incidence for the 

current inhalation study of 2-butoxyethanol. 

Dr. Medinsky moved that under the conditions of this 

study, the Technical Report on 2-butoxyethanol be 

accepted with revisions discussed and the conclusions 

as written. Dr. Bailer seconded the motion. 

Dr. Cullen asked whether a sentence could be added 

to the conclusion for male mice that there was an 

exposure-concentration related increase in the 

incidences of malignant hepatocellular neoplasms. 

Dr. Roycroft noted that the increased incidences of 

hepatocellular neoplasms in male mice could be added 

to the sentence about the marginal increases in the 

incidences of forestomach neoplasms. There was 

consensus that that addition would be acceptable. The 

motion was accepted with five yes votes with one 

abstention (Dr. Bus). 
14 2-Butoxyethanol, NTP TR 484 
 

 

15 

INTRODUCTION
 

CH3-CH2-CH2-CH2-O-CH2-CH2-OH 

2-BUTOXYETHANOL 

CAS No. 111-76-2 

Chemical Formula: C6H14O2 Molecular Weight: 118.17 

Synonyms: 2-Butoxy-1-ethanol; m-butyl ether; butyl glycol; ethylene glycol monobutyl ether 

Trade name: Butyl Cellosolve 

CHEMICAL AND PHYSICAL PROPERTIES 

2-Butoxyethanol is a member of a family of ethylene 

glycol monoalkyl ethers. It has a melting point of 

–70E C, a boiling point of 171E C, a density of 

0.9012 g/mL, and a vapor pressure of 0.88 mm Hg at 

25E C. It is miscible with water and soluble in 

mineral oil and most organic solvents. Although 

considered combustible, 2-butoxyethanol does not 

ignite readily and has a flash point of 60E C (closed 

cup) and flammability limits of 1.13% to 10.6% 

(Patty’s, 1994; Merck Index, 1996; HSDB, 1998). 

PRODUCTION, USE, 

AND HUMAN EXPOSURE 

2-Butoxyethanol is produced by reacting ethylene 

oxide with butyl alcohol. It may also be produced by 

reacting ethylene chlorohydrin or ethylene glycol with 

sodium hydroxide and dialkyl sulfide (HSDB, 1998). 

It is used extensively as a solvent in surface coatings 

such as lacquers, enamels, varnishes, and latex 

paint; in paint thinners, paint stripping formulations, 

and inks; and in degreasers and industrial and 

household cleaners (NIOSH, 1990; ATSDR, 1998; 

HSDB, 1998). It was estimated that in the United 

States in the 1970s, over 740 products contained 

2-butoxyethanol at an average concentration of 2.8% 

(Vincent et al., 1993). Over half of these products 

were used in the home. 2-Butoxyethanol concentrations in commercial products vary depending 

upon the product and its use. For example, 

2-butoxyethanol may reach concentrations of 35% for 

paint strippers, 5% for paint thinners, 21% for window cleaners, and 0.4% for inks (Vincent et al., 

1993; ATSDR, 1998). As a coupling agent, it stabilizes immiscible ingredients in metal cleaning 

products, textile lubricants, cutting oils, and hydraulic 

fluids. It is also used as a chemical intermediate in 

the production of acetate esters as well as 

phthalate and stearate plasticizers (HSDB, 1998). 

2-Butoxyethanol is the largest volume alkyl glycol 

ether produced, with an estimated 390 million pounds 

produced in the United States in 1992 (USITC, 1992). 

Because of its large production volume and widespread use, workers involved in the manufacture and 

formulation of products as well as consumers are 

potentially exposed to 2-butoxyethanol. According to 

the National Occupational Exposure Survey (1981-

1983), 1,680,768 workers were potentially exposed to 

2-butoxyethanol annually (NIOSH, 1990). The timeweighted average threshold limit value for 

2-butoxyethanol (skin notation) is 20 ppm (ACGIH, 
 

16 2-Butoxyethanol, NTP TR 484 

1999). The exposure limit of 2-butoxyethanol permitted by the Occupational Safety and Health Administration is 50 ppm (also with a skin designation), 

while The National Institute for Occupational Safety 

and Health (NIOSH) recommends a 10-hour timeweighted average of 5 ppm or 24 mg/m3 (also with a 

skin designation) (HSDB, 1998). NIOSH has 

estimated that most workplace exposures are below 

7 ppm; however, 2-butoxyethanol concentrations 

ranged from 0.04 to 367 ppm in various industrial 

operations in Belgium (Veulemans et al., 1987; 

NIOSH, 1990). 2-Butoxyethanol, because of its 

volatility and its solubility properties, enters the 

environment by air emissions, via leachate from 

municipal landfills and hazardous waste sites, and 

water runoff. 2-Butoxyethanol has been detected in 

drinking water from several major United States 

municipalities (concentrations not provided), groundwater (United States, 23 μg/L), surface water (Japan, 

5.68 μg/L), and industrial wastewater (United States, 

less than 100 μg/L). According to the National 

Ambient Volatile Organic Compounds Database (Shah 

and Singh, 1988), the average daily home indoor air 

concentration of 2-butoxyethanol is 0.214 ppb 

(1.0 μg/m3

); however, concentrations as high as 

8 μg/m3

 have been detected in homes in Italy. 

2-Butoxyethanol has been measured in air from 

commercial buildings at 34 μg/m3

, in commercial 

building exhaust at 13 μg/m3

, and in elevator shafts at 

19 μg/m3

 (ATSDR, 1998; HSDB, 1998). 

ABSORPTION, DISTRIBUTION, 

METABOLISM, AND ELIMINATION 

Experimental Animals 

Carpenter et al. (1956) first demonstrated that 

2-butoxyethanol was absorbed following short-term 

exposure of rats, rabbits, dogs, and guinea pigs to 100 

to 400 ppm 2-butoxyethanol and of human volunteers 

to 100 to 200 ppm 2-butoxyethanol. 2-Butoxyacetic 

acid was excreted in the urine, with most being 

excreted within 24 hours. Since then, the toxicokinetics and metabolism of 2-butoxyethanol have been 

extensively studied in humans and in several other 

species (primarily the male rat and predominantly the 

male F344/N rat) by the inhalation, oral, dermal, and 

intravenous routes of administration. Comprehensive 

reviews of this work have been published (ATSDR, 

1998; CIRP, 1996). 

2-Butoxyethanol, like the other alkyl glycol ethers, 

2-ethoxy- and 2-methoxyethanol, is readily absorbed 

from the lungs and gastrointestinal tract and through 

the skin, rapidly distributed into tissues, metabolized, 

and eliminated. The major route of metabolite 

elimination is urinary excretion, followed by 

exhalation of CO2. 2-Butoxyethanol, as with 

2-ethoxy- and 2-methoxyethanol, is oxidized in the rat 

liver by alcohol dehydrogenase (Johanson et al., 

1986a; Ghanayem et al., 1987a) to an intermediate 

aldehyde (butoxyacetaldehyde) that then undergoes 

further oxidation by aldehyde dehydrogenase 

(Ghanayem et al., 1987a) to the respective acid 

(2-butoxyacetic acid). A proposed metabolic scheme 

for 2-butoxyethanol (ATSDR, 1998) is shown in 

Figure 1. The pathways of 2-butoxyethanol metabolism, whether by oxidation, conjugation, or 

dealkylation, appear to be saturable, and the differences in metabolism noted for various routes of 

exposure may be due merely to differences in 

the internal dose achieved. In the male rat, 

2-butoxyacetic acid is the major metabolite of 

2-butoxyethanol, and its excretion in urine represents 

60% to 75% of the absorbed dose of 2-butoxyethanol 

regardless of whether the route of administration is 

dermal (Sabourin et al., 1992a), drinking water 

(Medinsky et al., 1990), gavage (Ghanayem et al., 

1987b), or inhalation (Jönsson and Steen, 1978; 

Sabourin et al., 1992b; Johanson, 1994). Route of 

administration, duration of exposure, dose, and 

differences in age have been shown to impact the 

urinary profile of 2-butoxyethanol metabolites and the 

overall elimination of absorbed 2-butoxyethanol. 

Route Comparison 

Ghanayem et al. (1987b) have shown that following 

gavage administration, in addition to 2-butoxyacetic 

acid in the urine, a glucuronide conjugate and sulfate 

conjugate are detected, although at much lower 

concentrations. The sulfate conjugate was observed 

only at low gavage doses (100 versus 500 mg/kg) and 

only in the first 8 hours of urine collection. 

Following inhalation exposure to 4.3, 49, or 438 ppm 

2-butoxyethanol, there was an exposure-related 

increase in the proportion of butoxyethanol 

glucuronide to 2-butoxyacetic acid and a decrease in 

the percentage of 2-butoxyethanol exhaled in CO2 

(Sabourin et al., 1992b). Ghanayem et al. (1987b) 

also showed that following a 500 mg/kg gavage dose 
2-Butoxyethanol, NTP TR 484 17 

FIGURE 1
 

Scheme for Metabolism of 2-Butoxyethanol (ATSDR, 1998)
 
 

 

 

18 2-Butoxyethanol, NTP TR 484 

of 2-butoxyethanol, the major biliary metabolite was 

the 2-butoxyethanol glucuronide conjugate, followed 

by 2-butoxyacetic acid; this was the opposite of what 

was observed in urine. Ethylene glycol has been 

detected in the urine of rats following exposure to 

2-butoxyethanol by inhalation (Sabourin et al., 

1992b), by dermal application (Sabourin et al., 

1992a), and in drinking water (Medinsky et al., 

1990). Although ethylene glycol was not detected by 

Ghanayem et al. (1987b), this may merely reflect the 

difference in the position of the 14C radiolabel on the 

2-butoxyethanol molecule; these authors’ label was on 

the butoxy moiety, whereas the ethanol moiety was 

used by the other authors. 

Bartnik et al. (1987) injected male Wistar rats subcutaneously with [14C]-2-butoxyethanol (118 mg/kg) 

and measured the elimination of radioactivity for 

72 hours. The primary route of elimination was via 

urine, with 78% of the administered dose appearing in 

the urine by 72 hours. Less than 1% was found in 

feces, while 10% was exhaled as 14CO2. In addition, 

Bartnik et al. administered 200 mg/kg of 

[

14C]-2-butoxyethanol topically to male and female 

Wistar rats and observed them for 48 hours. The 

highest amount of radioactivity in blood and plasma 

occurred 2 hours after application, with specific 

activity higher in plasma than blood. Measurements 

of 2-butoxyacetic acid in plasma followed the same 

qualitative course as 2-butoxyethanol. The authors 

concluded that the majority of 2-butoxyethanol absorbed was metabolized to 2-butoxyacetic acid. 

Approximately 29% and 25% of the administered 

dose was absorbed by males and females, respectively, with urine radioactivity accounting for 

23% and 20% of the applied dose; there were no 

significant differences between genders. Over 95% of 

the radioactivity excreted in the urine was eliminated 

in the first 24 hours. For in vitro skin penetration 

studies, Bartnik et al. (1987) demonstrated that 

skin penetration rates were two to three times greater 

in hairless rat skin than in pig or human skin, with no 

significant difference in penetration rates between pig 

and human skin. Dugard et al. (1984) showed in 

in vitro studies that the rates of absorption for three 

glycol ethers by human abdominal whole skin 

differed. 2-Methoxyethanol was the most readily 

absorbed (2.82 mg/cm2

 per hour), followed by 

2-ethoxyethanol (0.796 mg/cm2

 per hour), with 

2-butoxyethanol (0.198 mg/cm2

 per hour) being the 

least absorbed. These studies indicate that for human 

skin in vitro, where evaporation is eliminated, 

absorption decreases with increased chain length of 

the glycol ethers. 

Sabourin et al. (1992a) investigated the dermal 

absorption and elimination of 2-butoxy-[14C]ethanol as 

well as that of methoxyethanol and ethoxyethanol in 

male F344 rats for 72 hours following dosing. Each 

glycol ether was applied (unoccluded) at three different doses: for 2-butoxyethanol, 122, 367, or 

650 μmol per rat; for ethoxyethanol, 121, 387, or 

881 μmol per rat; and for methoxyethanol, 114, 342, 

or 1,027 μmol per rat. Within these dose ranges for 

all three glycol ethers, the absorption and metabolism 

of each were found to be linearly related to the dose 

applied dermally. Regardless of chain length or dose 

administered, there was no major difference in absorption (20% to 27%) of the ethers. The percentage 

of 2-butoxyethanol absorbed (20% to 25%) is similar 

to that for Wistar rats reported by Bartnik et al. 

(1987). The majority of the absorbed dose for all 

three glycol ethers was detected in urine; for 

2-butoxyethanol, this was 82% to 83%, for ethoxyethanol, 64% to 77%, and for methoxyethanol, 67% 

to 72%. The major urinary metabolite was the 

alkoxyacetic acid for each glycol ether. However, 

there were appreciable differences in the amount of 

alkoxyacetic acid for each. For 2-butoxyethanol, 

2-butoxyacetic acid represented 65% to 71% of the 

total urinary metabolites, while the ethoxyacetic acid 

represented 50% to 58% and the methoxyacetic acid 

represented 23% to 46%. Only urine from 

2-butoxyethanol-treated rats contained a glucuronide 

metabolite which accounted for 13% to 15% of the 

radioactivity in urine. Ethylene glycol was also 

detected in urine for all three ethers and represented 

4% to 6% for 2-butoxyethanol, 13% to 18% for 

ethoxyethanol, and 9% to 11% for methoxyethanol. 

The amount of 14CO2 eliminated in expired air was 

4% to 6% for 2-butoxyethanol, 4% to 8% for ethoxyethanol, and 6% to 11% for methoxyethanol. Although the relative amount of glycol ether absorbed 

was similar for all three glycol ethers, there were 

differences in metabolism and elimination of the 

three. 

Medinsky et al. (1990) also showed that, in drinking 

water studies, the fraction of the dose metabolized to 

ethylene glycol and CO2 was inversely proportional to 

chain length of the glycol ethers, 2-methoxy-, 

2-ethoxy-, and 2-butoxyethanol. The proportion of 
 

2-Butoxyethanol, NTP TR 484 19 

2-butoxyacetic acid and, to some extent, CO2 actually 

decreased with increasing drinking water concentrations of 290, 860, or 2,590 ppm 2-butoxyethanol, 

while ethylene glycol slightly increased For singleexposure inhalation studies, 2-butoxyacetic acid 

excretion has been shown to be linearly related to 

exposure concentration for up to 438 ppm 

2-butoxyethanol, whereas ethylene glycol in urine 

decreases with increasing exposure concentration 

(Sabourin et al., 1992b). In general, the amount of 

urinary glucuronide appears to increase with dose 

regardless of route. 

Age Comparison 

In single-dose gavage studies of 500 mg/kg 2-(1-[14C]-

butoxy)ethanol, Ghanayem et al. (1987c) showed that 

4- to 5-week-old male F344 rats exhaled a significantly higher percentage of CO2 than 9- to 

13-week-old rats. Young rats also excreted a significantly higher percentage of the administered dose 

in urine, which contained more 2-butoxyacetic acid 

and less glucuronide conjugates than did urine from 

older rats. In another study, to further determine a 

dose and age effect, the authors also compared 

the effect of intravenous administration of 

[

14C]-2-butoxyethanol (31.25, 62.5, or 125 mg/kg) on 

2-butoxyethanol toxicokinetics in 3- to 4-month-old 

and 12- to 13-month-old male F344 rats (Ghanayem 

et al., 1990). They found both dose- and age-related 

effects on 2-butoxyethanol and 2-butoxyacetic 

acid kinetics. In younger rats administered 

2-butoxyethanol intravenously, the area under 

the curve (AUC), maximum plasma concentration 

(Cmax), and systemic clearance (ClS) of 

2-butoxyethanol were dose dependent in that the AUC 

and Cmax increased with increasing dose while ClS decreased with increasing dose. At the same doses, Cmax 

and AUC of 2-butoxyethanol increased as a function 

of age. For 2-butoxyacetic acid, half-life (t1⁄2), AUC, 

and Cmax were increased relative to the dose of 

2-butoxyethanol given and to the age of the rats. For 

3- to 4-month-old rats, the t1/2 in blood was 

approximately 10 minutes for 2-butoxyethanol and 

3 hours for 2-butoxyacetic acid. 

Repeated Exposure 

The studies discussed previously primarily involved a 

single administration of radiolabeled 2-butoxyethanol 

in male rats. Johanson (1994) investigated the 

toxicokinetics of 2-butoxyethanol and 2-butoxyacetic 

acid in 4-month-old male rats following continuous 

inhalation exposure for 12 days to 20 or 100 ppm 

2-butoxyethanol. Daily analyses indicated that the 

kinetics of 2-butoxyethanol and 2-butoxyacetic acid 

were linear up to 100 ppm. 2-Butoxyethanol and 

2-butoxyacetic acid concentrations increased rapidly 

in tissue during the first 1 to 3 days and then more 

slowly during the rest of the study. At these concentrations there was no effect of exposure concentration on total blood clearance of 2-butoxyethanol 

(approximately 2.3 L/kg per hour) or the fraction of 

inhaled 2-butoxyethanol excreted in urine as 

2-butoxyacetic acid (approximately 65%). 

In conjunction with the long-term toxicology and 

carcinogenesis studies presented in this Technical 

Report, the National Toxicology Program performed 

toxicokinetic evaluations of 2-butoxyethanol and 

2-butoxyacetic acid in the blood of male and female 

F344/N rats and B6C3F1 mice following 1 day, 

2 weeks, and 3, 6, 12, and 18 months of exposure to 

2-butoxyethanol. 2-Butoxyacetic acid concentrations 

were determined in urine collected during the 

16 hours after exposure at each of these time points 

except day 1. In addition, a separate set of male and 

female mice were maintained in the control chamber 

until they were approximately 19 months old, at 

which time they were exposed to 125 ppm 

2-butoxyethanol for 3 weeks. The toxicokinetic 

parameters for 2-butoxyethanol and 2-butoxyacetic 

acid in blood, as well as the urine 2-butoxyacetic acid 

concentrations and the 19-month-old mouse study, are 

published by Dill et al. (1998). For male and female 

rats and mice, systemically absorbed 2-butoxyethanol 

was rapidly cleared from blood, independent of exposure concentration, throughout the study (t1⁄2 for rat, 

less than 10 minutes; t1⁄2 for mice, less than 5 minutes 

after 1 day of exposure). Increases in AUCs were 

proportional to increases in 2-butoxyethanol exposure 

concentrations, indicating linear 2-butoxyethanol 

kinetics. In contrast, the rate of 2-butoxyacetic acid 

elimination from blood decreased as the exposure 

concentration increased. Nonproportional increases 

in the 2-butoxyacetic acid AUC also indicated that 

2-butoxyacetic acid was eliminated following dosedependent, nonlinear kinetics. Overall, mice eliminated both 2-butoxyethanol and 2-butoxyacetic acid 

from blood faster than rats. Gender-related differences in 2-butoxyacetic acid elimination were most 

significant in rats, in that females were less efficient 

in clearing 2-butoxyacetic acid from the blood. 

Differences in renal excretion of 2-butoxyacetic acid 
 

 

20 2-Butoxyethanol, NTP TR 484 

are possibly responsible for the gender-related 

difference in the 2-butoxyacetic acid blood profiles in 

rats. With repeated exposure, the rates of elimination 

for both 2-butoxyethanol and 2-butoxyacetic acid 

decreased in both species, resulting in longer 

residence times of 2-butoxyethanol and 2-butoxyacetic 

acid in the blood. 2-Butoxyethanol was rapidly 

cleared from the systemic circulation of 19-month-old 

naive mice exposed to 125 ppm, indicating clearance 

profiles similar to those of young mice. However, 

old mice eliminated 2-butoxyacetic acid from blood 

about 10 times more slowly than young mice after 

1 day of exposure. This delayed elimination of 

2-butoxyacetic acid in old mice was less obvious after 

3 weeks of exposure compared to young mice exposed 

for 2 weeks, suggesting that there may be factors 

other than age that influenced the apparent difference 

in 2-butoxyacetic acid kinetics. Dill et al. (1998) concluded that the elimination kinetics of 2-butoxyethanol 

and 2-butoxyacetic acid following repeated 

2-butoxyethanol exposure appeared to be dependent 

on species, gender, age, number of exposures, and 

exposure concentration. 

In vitro Metabolism 

The metabolism of 2-butoxyethanol, 2-ethoxyethanol, 

and 2-methoxyethanol in F344 rat and human 

hepatocytes in vitro was reported by Green et al. 

(1996). Each glycol ether, 14C-labeled on the ethanol 

carbons, was incubated with rat or human hepatocytes 

for 4 hours at concentrations ranging from 0.02 to 

10 mM. The major metabolite in each species for all 

the glycol ethers was the respective alkoxyacetic acid, 

followed by ethylene glycol. For 2-butoxyethanol an 

additional metabolite, a glucuronide conjugate, was 

found with each species. The percentage of glycol 

ether converted to the alkoxyacetic acid increased 

with glycol ether concentrations up to 10 mM in rat 

hepatocytes. However, in human hepatocytes, the 

percentage actually decreased between 0.02 and 

0.2 mM. The following relative rate of alkoxyacetic 

acid formation from the glycols was determined: the 

2-butoxyethanol rate was greater than that of 

2-ethoxyethanol, followed by 2-methoxyethanol. This 

contrasted with ethylene glycol formation: 

2-methoxyethanol had a greater rate than 

2-ethoxyethanol, followed by 2-butoxyethanol. These 

findings are similar to those observed in vivo. Rat 

hepatocytes metabolized the glycol ethers at greater 

rates than human hepatocytes. For 2-butoxyethanol, 

at 0.2 mM, less than 1% of the 2-butoxyethanol 

remained after 4 hours in rat hepatocytes as opposed 

to about 55% in human hepatocytes. For 

2-methoxyethanol, at 0.2 mM, 36% remained 

unmetabolized after 4 hours compared to 80% in 

human hepatocytes. The maximum velocity (Vmax) for 

each glycol ether and the Michaelis constant (Km) for 

all but 2-butoxyethanol were considerably higher for 

alkoxyacetic acid production in rat than in human 

hepatocytes. The Vmax was 741 nmol/hour for 106

 rat 

hepatocytes and 113 nmol/hour for 106

 human 

hepatocytes for 2-butoxyethanol; for 2-ethoxyethanol, 

1,519 nmol/hour (rat) and 71 nmol/hour (human); and 

for 2-methoxyethanol, 1,511 nmol/hour (rat) and 

61 nmol/hour (human). The Km was 6.6 mM (rat) 

and 1.2 mM (human) for 2-ethoxyethanol and 

6.3 mM (rat) and 1.7 mM (human) for 

2-methoxyethanol. For 2-butoxyethanol, the Km for 

rat and human hepatocytes was the same, 0.9 mM. 

Physiologic Based Pharmacokinetic Models 

A number of physiologic based pharmacokinetic 

(PBPK) models have been developed based on shortterm 2-butoxyethanol exposures by various routes for 

male rats and humans. Most of the models (Johanson, 

1986; Johanson and Näslund, 1988; Shyr et al., 1993; 

Corley et al., 1994) have been thoroughly reviewed 

and models compared (ATSDR, 1998). In general, 

for these 2-butoxyethanol PBPK models, successive 

authors used data and results from previous authors 

and added new information to develop their models. 

Johanson (1986) and Johanson and Näslund (1988) 

developed a model for human exposure to 

2-butoxyethanol by inhalation. This model utilized 

human and perfused rat liver data. Shyr et al. (1993) 

based their PBPK model on toxicokinetic data from 

male F344 rats exposed to 2-butoxyethanol by dermal 

application, by inhalation, and in drinking water. 

This model also included a metabolic pathway for 

2-butoxyethanol. Corley et al. (1994) expanded the 

models of Johanson (1986) by modeling the 

disposition of 2-butoxyethanol and 2-butoxyacetic acid 

in humans and male F344 rats. This model included 

inhalation (whole body or nose/mouth only), oral 

(gavage or drinking water), intravenous infusion, and 

dermal (vapor or liquid) routes of administration; 

physiologic descriptions for rats; competing pathways 

for metabolism of 2-butoxyethanol; and measured 

partition coefficients determined by the authors for 

2-butoxyethanol and 2-butoxyacetic acid in several rat 

tissues and human blood. Corley et al. (1997) 

modified this model further to include dermal 
 

 

 

 

2-Butoxyethanol, NTP TR 484 21 

absorption of 2-butoxyethanol vapors for humans 

excluding secondary pathways for 2-butoxyethanol 

metabolism (conjugation and dealkylation) and 

including conditions for rest versus exercise. 

Toxicokinetic data from female rats or from male or 

female mice were not available to test against previous 

models. Lee et al. (1998), as part of the evaluation of 

the present 2-year inhalation studies with 

2-butoxyethanol in male and female rats and mice, 

reported a PBPK model for 2-butoxyethanol and 

2-butoxyacetic acid based on continuous exposure to 

2-butoxyethanol for 18 months. The model 

incorporated data and results from previous models 

and accommodated exposure to 2-butoxyethanol by 

inhalation (whole body or nose only), dermal, gavage, 

or intravenous administration. In addition, because of 

the time course of repeated exposure and the 

associated need for time-dependent effects on 

physiologic and biochemic parameters, a number of 

additional parameters, functions, and mass balance 

differential equations were incorporated. Tissue 

compartments were revised relative to previous 

models in that a muscle compartment was included 

into the poorly perfused compartment, a kidney 

compartment was added for 2-butoxyethanol, and 

spleen was added as a separate compartment. 

Humans 

Controlled Experiments 

In addition to 2-butoxyethanol occupational exposure 

assessments, there have been several controlled 

experiments whereby human volunteers have been 

exposed to 2-butoxyethanol percutaneously or by 

inhalation, and toxicokinetic assessments have been 

performed. Johanson et al. (1986b) exposed seven 

male volunteers to 20 ppm 2-butoxyethanol for 

2 hours during light exercise on a bicycle ergometer. 

Respiratory uptake, derived from measurements of 

expired air over time, was determined to be 

10.1 μmol/minute (1.2 mg/minute) or approximately 

57% of the amount of inspired 2-butoxyethanol. 

Within 1 to 2 hours, 2-butoxyethanol blood 

concentrations reached a plateau of 7.4 μmol/L. The 

elimination half-time was 40 minutes; mean residence 

time, 42 minutes; total blood clearance time, 

1.2 L/minute; and steady state volume of distribution, 

54 L. Less than 0.03% of the total 2-butoxyethanol 

uptake was excreted in urine, whereas 2-butoxyacetic 

acid excretion represented 17% to 55% of the total 

2-butoxyethanol uptake. In another experiment, 

Johanson and Johnsson (1991) exposed five male 

volunteers to 20 ppm 2-butoxyethanol for 2 hours 

during light exercise. Blood collected from a brachial 

vein at 2, 4, or 6 hours after exposure was analyzed 

for 2-butoxyacetic acid; concentrations ranged from 

approximately 22 to 60 μM with an average of 

approximately 45 μM reached within 2 to 4 hours. 

The half-life of 2-butoxyacetic acid in blood was 

estimated to be about 4 hours with a renal clearance 

of 23 to 39 mL per minute and an apparent volume of 

distribution of about 15 L, which the authors 

concluded represented considerable binding to blood 

proteins. 

Johanson et al. (1988) determined the percutaneous 

absorption of 2-butoxyethanol in five male volunteers 

by immersing two to four fingers of one hand in neat 

2-butoxyethanol for 2 hours. Blood was collected 

from the opposite hand (finger prick) during and up to 

4 hours after exposure. Urine was collected for 

24 hours and analyzed for 2-butoxyacetic acid. The 

skin of exposed fingers appeared to be more rigid, 

less elastic, and wrinkled. Within a few hours of 

exposure, a dry reticulate pattern with small fissures, 

some of which became erythematous, was observed 

but disappeared within 1 to 2 days. 2-Butoxyethanol 

was detected in the blood of all volunteers; however, 

the shape of the blood concentration profiles varied 

considerably between volunteers and experiments. 

Therefore, the percutaneous uptake of 7 to 

96 nmol/cm2

 per minute (geometric mean was 

20 nmol/cm2

 per minute) and the half-time of 

2-butoxyethanol decay of 0.6 to 4.8 hours was highly 

variable. 2-Butoxyacetic acid was detected in the 

urine of each volunteer with a peak concentration 

occurring 3 hours after exposure and an average halftime of 3.1 hours. The amount of 2-butoxyacetic acid 

excreted was also highly variable (87 to 313 μmol). 

2-Butoxyacetic acid excretion represented, on the 

average, 17% (2.5% to 39%) of the absorbed dose of 

2-butoxyethanol. 

To assess the percutaneous absorption component of 

inhalation exposure, four male volunteers breathed 

50 ppm 2-butoxyethanol through a respiratory valve 

for 2 hours (Johanson and Boman, 1991). After a 

1-hour nonexposure period, the volunteers’ entire 

bodies were exposed for 2 hours to 50 ppm 

2-butoxyethanol, followed by a 2-hour recovery 

period. The volunteers wore respirators to prevent 

inhalation exposure during the second 2 hours. In 

addition, each volunteer was also separately 
 

 

22 2-Butoxyethanol, NTP TR 484 

challenged throughout each study to two different 

chamber environments, i.e., 23E C and 29% relative 

humidity or 33E C and 71% relative humidity. Blood 

was collected periodically over the 7-hour period 

from a finger prick. Blood 2-butoxyethanol 

concentrations appeared to reach a steady state during 

the second hour of the mouth-only exposure at 

approximately 3 μM, with an apparent blood clearance of 3.8 L per minute and a respiratory uptake of 

11 μmol/minute. However, during percutaneous 

exposure, the concentration of 2-butoxyethanol 

increased to approximately 9 μM during the second 

hour of exposure, about three times higher than 

2-butoxyethanol blood concentration during the inhalation exposure period. Percutaneous absorption was 

31 μmol/minute, also about three times greater than 

the respiratory uptake. 2-Butoxyethanol half-life in 

blood following skin exposure averaged 34 minutes. 

In general, increased temperature and humidity 

increased blood 2-butoxyethanol concentrations during 

inhalation and dermal exposure; however, these differences were not statistically significant. The authors 

suggested that dermal uptake of 2-butoxyethanol 

accounts for approximately 75% of the total uptake 

during whole-body exposure. 

Corley et al. (1997) investigated the impact of blood 

sampling location on measurement of dermal 

absorption of 2-butoxyethanol vapors in humans. 

These studies were initiated to provide an explanation 

for the apparently enhanced dermal absorption in 

humans following dermal exposure to 2-butoxyethanol 

vapors observed by Johanson and Boman (1991) and 

those authors’ assertion that dermal absorption was a 

major contributor to total 2-butoxyethanol absorption 

in inhalation studies. Corley et al. compared blood 

collection by finger prick from the arm exposed to 

2-butoxyethanol (Johanson method) to collecting 

blood by venous catheter in the unexposed arm. Six 

human volunteers were exposed, arm only, to 50 ppm 

2-butoxyethanol for 2 hours and monitored for 

24 hours during which blood and urine were 

collected. Neither 2-butoxyethanol nor 2-butoxyacetic 

acid was detected in blood from the catheterized arm 

for the first 30 minutes of exposure. 2-Butoxyethanol 

was rapidly cleared from the blood with t1⁄2 of 

0.66 hour. 2-Butoxyacetic acid concentrations 

exceeded those of 2-butoxyethanol at all time points, 

and 2-butoxyacetic acid was cleared less rapidly from 

the blood (t1⁄2=3.3 hours). Peak blood concentrations 

were 0.037 μM for 2-butoxyethanol and 0.31 μM for 

2-butoxyacetic acid. Using the finger prick method of 

collection, the 2 hour, 2-butoxyethanol blood concentration was 35.5 μM, about 1,500 times that 

obtained from the unexposed catheterized arm. 

2-Butoxyethanol concentration in blood obtained from 

the finger prick was also about 37 times higher than 

the 2-butoxyacetic acid concentration (0.97 μM). 

This was the opposite of what was observed for the 

catheterized arm, where the blood 2-butoxyethanol 

concentration was roughly one tenth that of 

2-butoxyacetic acid. The 2-butoxyacetic acid 

concentrations were more consistent when the two 

collection methods were compared, i.e., within a 

factor of four of each other. In blood from the 

catheterized arm, elimination half-times for 

2-butoxyethanol (0.66 hours) and 2-butoxyacetic acid 

(3.3 hours) were consistent with the elimination halftimes of 0.66 hours for 2-butoxyethanol and 4 hours 

for 2-butoxyacetic acid obtained following inhalation 

exposure under exercise conditions (Johanson et al., 

1986b; Johanson and Johnsson, 1991). The authors 

concluded that blood collected by finger prick most 

likely represented both systemic concentration and 

local concentration of 2-butoxyethanol in venous 

blood draining the skin and was not an accurate 

representation of systemic 2-butoxyethanol 

absorption. 

Occupational Exposure 

A number of studies, which are detailed below, have 

provided biologic monitoring data following 

occupational exposure to 2-butoxyethanol. Occupations evaluated include those involved in varnish 

production, beverage packaging, automotive manufacturing and cleaning, office maintenance, and 

gravure printing. Preshift and postshift assessments 

were made. 2-Butoxyacetic acid concentration in 

urine was the primary end point for biologic 

monitoring. In some cases, only 2-butoxyacetic acid 

concentrations were reported. However, in others, 

free, conjugated, and total 2-butoxyacetic acid concentrations were stated. 

Angerer et al. (1990) evaluated exposure to 

2-butoxyethanol for 12 men working in a varnish 

production plant. Blood was collected at the end of 

each shift, whereas urine was collected prior to and 

following each shift. 2-Butoxyethanol workplace 

concentrations averaged 1.1 ppm (0.1 to 8.1 ppm). 

Blood 2-butoxyethanol concentrations averaged 
 

 

2-Butoxyethanol, NTP TR 484 23 

121.3 μg/L (5 to 570 μg/L). Urine 2-butoxyacetic 

acid, taken at the same time blood was drawn, 

averaged 10.5 mg/L (0.6 to 30.3 mg/L). Urinary 

2-butoxyacetic acid prior to starting a shift was 

considerably less, averaging 3.3 mg/L (0.1 to 

14.3 mg/L). Söhnlein et al. (1993) determined the 

extent of 2-butoxyethanol exposure in 15 men and 

four women employed in the production of varnishes 

or their quality control. Preshift Monday and postshift Tuesday measurements were made. In the 

production area, the concentration of 2-butoxyethanol 

in air was determined to be 0.5 ppm (preshift) 

and 0.6 ppm (postshift). Prior to starting a shift, the 

average urinary concentration of 2-butoxyacetic acid 

was 0.2 mg/L, whereas the postshift concentration 

was 16.4 mg/L, indicating significant absorption of 

2-butoxyethanol. 

Haufroid et al. (1997) monitored urinary 

2-butoxyacetic acid concentration in 31 male workers 

in a beverage packaging plant pre- and postshift. 

Twenty of the workers were transferring the decor to 

the cans, while the remainder were exposed to 

2-butoxyethanol while spraying varnish on the inside 

of the cans. Workplace exposure averaged 0.76 ppm 

2-butoxyethanol (0.37 to 1.29 ppm) for the decor 

transfer workers and 0.46 ppm (0.15 to 0.7 ppm) for 

the varnish sprayers. As in the studies reported 

previously, preshift urine 2-butoxyacetic acid concentrations were considerably lower than 

postexposure concentrations. For the decor transfer 

workers, preshift 2-butoxyacetic acid averaged 

0.7 mg/L (0 to 2.8 mg/L) as opposed to a postshift 

2-butoxyacetic acid average of 19.5 mg/L (0.9 to 

78.9 mg/L). Likewise, the varnish sprayers had 

comparable average 2-butoxyacetic acid 

concentrations of 1.0 mg/L (0 to 5.2 mg/L) for 

preshift and 14.2 mg/L (0.9 to 35.5 mg/L) for 

postshift measurements. These measurements for 

2-butoxyacetic acid are in close agreement with those 

reported by Angerer et al. (1990) and Söhnlein et al. 

(1993). 

Vincent et al. (1993) reported the occupational 

exposure to 2-butoxyethanol in municipal government 

workers who cleaned windows in automobiles and 

workers cleaning office windows. 2-Butoxyethanol 

concentrations in window cleaning products ranged 

from 0.9% to 21.2%. 2-Butoxyethanol concentrations 

monitored in the workplace air ranged from less than 

0.1 to 7.33 ppm for automobile cleaners and less than 

0.3 to 0.73 ppm for office cleaners. Preshift urinary 

concentrations ranged from less than 2 to 98.6 mg 

2-butoxyacetic acid/g creatinine for automobile 

washers versus less than 2 to 4.6 mg 2-butoxyacetic 

acid/g creatinine for office cleaners. End-shift concentrations were considerably higher, ranging from 

12.7 to 371 mg 2-butoxyacetic acid/g creatinine and 

2 to 3 mg 2-butoxyacetic acid/g creatinine for the two 

occupations, respectively. Pre- and postshift 

2-butoxyacetic acid concentrations for office workers 

were similar, indicating a low level of exposure 

during work hours and possible exposure to 

2-butoxyethanol in products outside the workplace. 

Rettenmeier et al. (1993) monitored the urine of six 

lacquerers at an automotive manufacturing plant to 

determine 2-butoxyethanol exposure. Although 

exposure conditions were stated as being similar, no 

workplace environmental monitoring was reported. 

Each worker used a 2-butoxyethanol-containing 

detergent to clean automotive body parts. Urine 

samples were collected from each worker at the end 

of a Friday shift and on Monday prior to beginning a 

new shift and analyzed for 2-butoxyacetic acid and 

2-butoxyacetic acid glutamine conjugate. The 

Monday samples contained only traces of either 

metabolite (data not reported). Postshift samples 

taken on the previous Friday contained both 

metabolites in almost equimolar quantities; however, 

the concentrations of each were highly variable 

between individuals although consistent for each 

individual. Concentrations ranged from 0.13 to 

5.91 mmol 2-butoxyacetic acid/L and 0.12 to 

2.45 mmol glutamine conjugate/L. The authors 

concluded that measurement of urinary 2-butoxyacetic 

acid alone was not sufficient for biologic monitoring 

of 2-butoxyethanol exposure in humans. 

In similar studies, Sakai et al. (1993) measured 

2-butoxyacetic acid concentrations at the end of 

an8-hour workday in the urine of nine workers 

engaged in gravure printing. The workers were 

exposed during the workday to an average 

2-butoxyethanol concentration of 0.64 ppm (0.4 to 

0.8 ppm). Urine 2-butoxyacetic acid concentrations 

averaged 3.92 mg 2-butoxyacetic acid/g creatinine 

(1.3 to 9.9 mg/g C). In another study, Sakai et al. 

(1994) assessed exposure of five workers throughout 

a workweek in a semiconductor factory where they 

were engaged in polymerization of resin dissolved in 

2-butoxyethanol. The workplace was monitored for 
 

 

 

24 2-Butoxyethanol, NTP TR 484 

2-butoxyethanol. Individual 2-butoxyethanol 

concentrations were not reported; however, exposures 

were reported to be less than 0.8 ppm. Urine 

concentrations of free and conjugated 2-butoxyacetic 

acid were measured. As expected, free and 

conjugated 2-butoxyacetic acid concentrations 

increased at the end of each workday, with a small 

amount of conjugated 2-butoxyacetic acid detected 

prior to the start of the next workday. During the 

first 2 days of the workweek, more conjugated 

2-butoxyacetic acid was excreted than toward the end 

of the week; however, conjugated 2-butoxyacetic acid 

always contributed the greatest portion of total 

2-butoxyacetic acid excreted, 71% (44% to 92%). 

The authors concluded that the decline in excretion of 

conjugated 2-butoxyacetic acid indicated that the 

metabolic capacity for 2-butoxyacetic acid conjugation 

was gradually depressed. 

TOXICITY 

Experimental Animals 

Extensive reviews of the toxicity of 2-butoxyethanol 

and other alkyl glycol ethers have been published 

(NIOSH, 1990; Patty’s, 1994; ATSDR, 1998; CIRP, 

1996; Ghanayem, 1996). In general, alkyl glycol 

ethers such as 2-methoxy-, 2-ethoxy-, and 

2-butoxyethanol have been shown to induce 

reproductive and developmental effects or hematologic effects. As the length of the alkyl chain 

increases, hematologic effects predominate, whereas 

as the alkyl chain length decreases, reproductive and 

developmental effects are more common. The 

alkoxyacetic acids (2-methoxy-, 2-ethoxy-, and 

2-butoxyacetic acid) are the primary metabolites of 

these alkyl glycol ethers and are considered to be the 

toxic agents. Inhibition of the metabolism of the 

glycol ether to its corresponding alkoxyacetic acid has 

been shown to reduce or abolish the resulting 

testicular atrophy, teratogenicity, and immunotoxicity 

of 2-methoxyethanol and the hemolytic effects of 

2-butoxyethanol. 

The LD50 values for 2-butoxyethanol vary widely 

depending on the route of administration, duration of 

exposure, and species involved. The oral LD50 for 

rats is about 1,500 mg/kg (530 to 3,000 mg/kg); for 

mice, 1,230 mg/kg; for rabbits, approximately 

350 mg/kg; and for guinea pigs, 1,200 mg/kg. The 

dermal LD50 for rabbits ranges from 72 to 638 mg/kg 

and for the guinea pig from 205 to 4,800 mg/kg. The 

LC50 values are 486 ppm for male and 450 ppm for 

female F344 rats in 4-hour inhalation studies and 

700 ppm in mice in 7-hour studies. The intravenous 

LD50 is 340 mg/kg for rats, 1,130 mg/kg for mice, 

and 280 mg/kg for rabbits (ITII, 1981; Tyler, 1984; 

Patty’s, 1994; ATSDR, 1998; CIRP, 1996). 

2-Butoxyethanol toxicity has been demonstrated in 

animals following inhalation, gavage, injection, or 

dermal exposure. In a series of experiments, Werner 

et al. (1943a,b,c), exposed dogs, mice, and rats to 

2-butoxyethanol vapors. Dogs were exposed to 

415 ppm 2-butoxyethanol 7 hours per day, 5 days per 

week for 12 weeks. A maximum decrease in 

erythrocyte count, hemoglobin concentration, and 

hematocrit value was observed after 4 to 6 weeks of 

exposure and continued until the end of the study. In 

other studies, white Swiss mice were exposed to 

concentrations of 390 to 1,210 ppm 2-butoxyethanol 

for 7 hours. Mortality occurred before the end of 

exposure for mice exposed to 770 ppm or greater and 

within 32 hours at concentrations above 390 ppm. 

Respiratory distress, hemoglobinuria, and splenic 

congestion were noted. Wistar rats were exposed to 

320 ppm 2-butoxyethanol 7 hours per day, 5 days per 

week for 5 weeks; significant decreases in hemoglobin 

concentration and erythrocyte count were observed 

after 1 week. 

Carpenter et al. (1956) exposed rats, mice, rabbits, 

dogs, monkeys, and guinea pigs to 100 to 400 ppm 

2-butoxyethanol for up to 90 days. One rhesus 

monkey exposed for 30 days to 210 ppm 

2-butoxyethanol exhibited increased erythrocyte 

fragility (4th exposure) and increased fibrinogen 

(14th exposure). At the end of the study, erythrocyte 

count and hemoglobin concentration were reduced to 

levels half that at the start of the study. A male and 

a female monkey exposed to 100 ppm 2-butoxyethanol 

for 90 days had increased erythrocyte fragility and decreased numbers of erythrocytes. The female was 

more severely affected. Erythrocyte fragility was 

observed in rats within 4 hours of exposure to 62 ppm 

2-butoxyethanol, whereas hemolysis and hemoglobinuria were observed within 2 to 3 hours in 

female rats exposed to 432 ppm. Females were 

usually more severely affected. Erythrocyte fragility 

was also observed in mice (100 ppm) and rabbits 

(125 ppm) following 7-hour exposure. The extent of 
 

 

 

 

 

2-Butoxyethanol, NTP TR 484 25 

increased erythrocyte fragility did not increase in mice 

when exposure to 100 ppm was continued for 

90 days. Dogs exposed to 385 ppm 2-butoxyethanol 

for at least 8 days also showed increased fragility, 

whereas guinea pigs exposed for 30 days to 500 ppm 

2-butoxyethanol did not show this effect. In addition, 

rats (107 ppm) and mice (200 ppm) had increased 

liver weights after 30 and 60 days exposure, respectively. Cloudy swelling of the liver and convoluted tubules of the kidney were observed in female 

rats. 

Dodd et al. (1983), in two separate experiments, 

exposed male and female F344 rats 6 hours per day to 

0, 20, 86, or 245 ppm 2-butoxyethanol for 9 days or 

0, 5, 25, or 77 ppm 2-butoxyethanol, 5 days per week 

for 13 weeks. Rats exposed to 245 ppm exhibited 

significant decreases in erythrocyte count, hemoglobin 

concentration, and mean cell hemoglobin concentration and increases in nucleated erythrocyte, 

reticulocyte, and lymphocyte counts at 9 days. Liver 

weights were increased. Similar but less marked 

effects were observed in rats exposed to 20 ppm. 

Female rats exposed to 77 ppm for 6 weeks had 

statistically significant decreases in erythrocyte count 

and hemoglobin concentration accompanied by an 

increase in mean cell hemoglobin value; these effects 

were still present at the end of the 13-week study, 

although they were less severe. Male rats exposed to 

77 ppm 2-butoxyethanol (after 66 exposures) had a 

5% decreased erythrocyte count, although the 

decrease was not statistically significant. No 

exposure-related effects on urine or serum chemistries, body or organ weights, or microscopic lesions 

were observed. 

Nelson et al. (1984), in a range-finding study prior to 

an inhalation teratology study, exposed three or 

four female Sprague-Dawley rats to 250 to 500 ppm 

2-butoxyethanol for 6.5 to 7 hours. Deaths occurred 

within 18 to 36 hours at all exposure concentrations 

with the majority occurring at 450 ppm and higher. 

Various degrees of hematuria were noted. In addition, the survivors from each concentration were 

maintained individually, and approximately 1 week 

past exposure it was noted that the distal half of the 

tail became necrotic and sloughed off or was chewed 

off. This same tail effect was reported by Dow 

Chemical Corporation in 1981 (presented in ATSDR, 

1998) in female F344 rats following a single gavage 

dose of 1,000 mg 2-butoxyethanol/kg body weight. 

Hardin et al. (1984), while conducting a dermal 

administration comparative teratology study in SPF 

Sprague-Dawley rats with several glycol ethers, 

observed overt toxicity in dams dosed with 

2-butoxyethanol. The glycol ethers were administered 

in equimolar doses with 0.35 mL neat 

2-butoxyethanol administered four times daily (every 

2.5 hours) on gestation days 7 to 16. During the first 

day of treatment, burgundy-colored urine was noted, 

and by the end of the first day, the dams were 

showing signs of ataxia. With subsequent treatments, 

ataxia led to inactivity and ultimately death for 10 of 

the 11 dams (days 3 through 7 of dosing). As 

reported previously, the rats’ tails blackened distally 

and were gradually eaten away as the apparent 

necrosis progressed. 

Tyler (1984) gave mice gavage doses of 500, 1,000, 

or 2,000 mg/kg 2-butoxyethanol 5 days per week for 

5 weeks. All mice receiving 2,000 mg/kg died. Mice 

treated with 500 mg/kg or higher had reduced 

erythrocyte counts. Grant et al. (1985) gavaged male 

F344 rats with 500 or 1,000 mg 2-butoxyethanol/kg 

body weight daily for 4 consecutive days and then 

observed them for 3 weeks. Spleen, liver, kidney, 

and thymus weights were increased for up to a week 

after dosing. Only liver and spleen weights remained 

elevated at day 22. Mean body weight gain was 

reduced only at 1,000 mg/kg. Hematologic effects 

observed on the fourth day of dosing included 

decreases in erythrocyte count, hemoglobin concentration, and hematocrit value and marked elevated 

mean cell volume, mean cell hemoglobin value, and 

reticulocyte count. These effects returned to normal 

during the recovery period, and by day 22, only an 

increase in mean cell volume was noted. Bone 

marrow hyperplasia and a marked increase in splenic 

extramedullary hematopoiesis were observed. Unlike 

2-butoxyethanol, 2-methoxyethanol given to rats at 

100 and 500 mg/kg during the same period had a 

minimal effect on erythrocytes. However, exposure 

to 2-methoxyethanol did result in significantly reduced 

leukocyte counts, as represented by reductions in 

neutrophils and lymphocytes that remained until day 

22. Treatment with 2-methoxyethanol resulted in 

reduced spleen, kidney, testes, and thymus weights, 

with testes weight remaining significantly reduced at 

day 22. Bone marrow was hemorrhagic in the 

500 mg/kg group at day 1 and appeared to be 

associated with sinus endothelial cell damage. 

Normal extramedullary hematopoiesis of the spleen 
 

 

 

 

 

 

26 2-Butoxyethanol, NTP TR 484 

was abolished at the end of dosing and had returned 

to normal by day 22. Of importance was the severity 

of the testicular atrophy, which included disruption of 

the normal tubular architecture, degenerative changes 

in spermatocytes, and absence of spermatozoa in the 

epididymides. Thymic lymphocyte depletion was 

noted only at day 1. 

In gavage studies of longer duration, Krasavage 

(1986) dosed male COBS CD (SD) BR rats with 0, 

222, 443, or 885 mg/kg 2-butoxyethanol, 5 days per 

week for 6 weeks. There was a dose-dependent 

decrease in mean body weight gain. Although rats 

exposed to 2-butoxyethanol ate less than controls early 

in the study, only those treated at 885 mg/kg had 

significantly reduced feed consumption. Relative 

liver, kidney, and spleen weights were increased, 

primarily in the 885 mg/kg dose group. At the end of 

dosing, there were dose-related decreases in erythrocyte counts, hemoglobin concentrations, and mean 

cell hemoglobin concentrations with increases in mean 

cell volumes and mean cell hemoglobin values. 

Hepatocytomegaly was observed at 885 mg/kg. In 

addition, there was hemosiderin accumulation in the 

liver (885 mg/kg only) and proximal convoluted 

tubules of the kidney (all doses). Minimal to mild 

hyperkeratosis and acanthosis were observed in the 

stomach (forestomach or glandular stomach was not 

specified). Dosing did not affect the testes, thymus, 

bone marrow, or leukocytes. These results are 

consistent with those of Grant et al. (1985) using male 

F344 rats, except that those authors observed bone 

marrow hyperplasia. 

In drinking water studies, male Sprague-Dawley rats 

were exposed to 0, 2,000, or 6,000 ppm 

2-butoxyethanol or 2-methoxyethanol, and female 

Sprague-Dawley rats were exposed to 0, 1,600, or 

4,800 ppm 2-butoxyethanol or 2-methoxyethanol for 

21 days (Exon et al., 1991). Male and female rats 

exposed to the highest concentration of either chemical consumed significantly less water. Male rats 

exposed to high doses of either chemical and females 

exposed to both doses of 2-butoxyethanol gained 

significantly less weight than controls. The thymus 

was significantly smaller in male and female rats 

exposed to 2-methoxyethanol, while the testis weight 

in males was greatly reduced at the high dose. No 

significant organ weight effects were noted in rats 

drinking water containing 2-butoxyethanol. No 

2-butoxyethanol-related lesions were observed in the 

thymus, liver, kidney, or testis, whereas atrophy was 

noted in the thymus of 2-methoxyethanol-treated rats. 

The NTP (1993b) conducted 13-week drinking water 

studies in male and female F344/N rats and B6C3F1 

mice with 750 to 6,000 ppm 2-butoxyethanol. Estimates of compound consumption were 70 to 

500 mg/kg for rats and 100 to 1,300 mg/kg for mice. 

All rats and mice survived to the end of the study. 

Exposure-related reductions in mean body weight gain 

were seen in rats and mice. There was no effect on 

the testis weight of rats or mice. Although thymic 

weights were reduced in rats, these reductions were 

significantly less severe than those observed following 

exposure to either 2-methoxyethanol or 

2-ethoxyethanol. In addition, male and female rats 

had exposure-related increased relative liver and 

kidney weights. There were increased incidences of 

bone marrow hyperplasia; pigmentation and hematopoiesis of the spleen; and pigmentation, degeneration, 

and cytoplasmic alteration of the liver in rats. There 

were no treatment-related histopathologic findings in 

mice. Histopathologic effects in rats were consistent 

with effects on the hematopoietic system which 

resulted in a progressive anemia. The anemia caused 

by 2-butoxyethanol was characterized as macrocytic, 

hypochromic, and regenerative with increased bone 

marrow cellularity and marginal thrombocytopenia. 

The ethylene glycol alkyl ethers have been shown to 

affect the hematopoietic system. For 2-butoxyethanol, 

this effect is the primary toxicologic event in animals. 

Over the last 50 years, a number of scientists have 

investigated these effects (Werner et al., 1943a,b,c; 

Carpenter et al., 1956; Dodd et al., 1983; Grant 

et al., 1985; Krasavage, 1986; Bartnik et al., 1987; 

Ghanayem, 1989; Ghanayem and Sullivan, 1993; 

Ghanayem et al., 1987a,b,c, 1989, 1990, 1992). 

A number of comprehensive reviews describe the 

2-butoxyethanol-associated hematologic toxicity in 

animals compared to humans (Patty’s, 1994; ATSDR, 

1998; CIRP, 1996; Ghanayem, 1996); therefore, only 

a summary of the 2-butoxyethanol-induced hematotoxicity is provided here. 2-Butoxyethanol causes 

hemolysis of erythrocytes in vivo leading to an anemia 

and secondary effects on other organs (liver, 

kidney, spleen, and bone). Following exposure, 

2-butoxyethanol caused increased osmotic fragility of 

erythrocytes (Carpenter et al., 1956) resulting in 

increased sensitivity to osmotic lysis. The hemolytic 

effects of 2-butoxyethanol are exerted by its major 
 

 

2-Butoxyethanol, NTP TR 484 27 

metabolite, 2-butoxyacetic acid (Carpenter et al., 

1956; Bartnik et al., 1987; Ghanayem et al., 1987a). 

Inhibition of alcohol dehydrogenase by pyrazole or 

aldehyde dehydrogenase by cyanamide (Ghanayem 

et al., 1987a) or administration of either ethanol, 

n-propanol, or n-butanol with 2-butoxyethanol (Morel 

et al., 1996) inhibits the hemolytic effect of 

2-butoxyethanol. There is an apparent species sensitivity to the 2-butoxyethanol-induced lysis; rats and 

mice are more sensitive, followed by rabbits and 

primates. Pigs, dogs, cats, guinea pigs, and humans 

are relatively insensitive (Ghanayem and Sullivan, 

1993). Several studies have shown that females of 

several species, including monkeys (Carpenter et al., 

1956) and rats (Carpenter et al., 1956; Dodd et al., 

1983; NTP, 1993b), are more susceptible than males 

to the hematologic effects of 2-butoxyethanol administration. Whether females of other species are more 

sensitive than males is unknown because most of the 

toxicity and mechanism studies have been conducted 

only in males. In addition, it has been reported that 

young rats are less sensitive to hemolysis caused by 

2-butoxyethanol exposure than older rats (Ghanayem 

et al., 1987c). It has been shown that repeated 

exposure or pretreatment with high doses of 

2-butoxyethanol may result in a tolerance due to 

selective loss of older erythrocytes followed by an 

increase in newly formed erythrocytes that appear to 

be less sensitive (Ghanayem et al., 1992; Sivarao and 

Mehendale, 1995). 

In rats, lysis is preceded by erythrocyte swelling, 

decreased adenosine triphosphate levels (Ghanayem 

et al., 1989), and reduced deformability (Udden and 

Patton, 1994). The erythrocytes have a tendency to 

agglutinate and release hemoglobin, which forms 

visible precipitates (Udden and Patton, 1994). 

Erythrocyte morphologic alterations of stomatocytosis, spherocytosis, fragmentation, and formation 

of ghost cells have been reported (Ward et al., 1989; 

Udden and Patton, 1994). Human erythrocytes 

in vitro are less susceptible than rat erythrocytes to 

hemolysis and deformability by 2-butoxyacetic acid 

(Bartnik et al., 1987; Ghanayem, 1989; Udden, 

1994). In addition, when 2-butoxyacetic acid was 

incubated with erythrocytes obtained from individuals 

with hereditary spherocytosis or sickle cell disease, 

two human disorders marked by chronic hemolysis, 

there was no increase in hemolysis, no changes in 

mean cell volume or morphology, and no changes in 

deformability (Udden, 1994). 

Secondary to the 2-butoxyethanol-induced hemolysis 

and depending on the dose of 2-butoxyethanol, there 

may be increased incidences of hematopoietic cell 

proliferation in the spleen, pigmentation/hemosiderin 

accumulation in the Kupffer cells of the liver and 

proximal convoluted tubules of the kidney, and 

hyperplasia and increased cellularity of the bone 

marrow with a reticulocytosis and increase in nucleated erythrocytes and hemoglobinuria (Werner et al., 

1943a,b,c; Carpenter et al., 1956; Dodd et al., 1983; 

Grant et al., 1985; Krasavage, 1986; Ghanayem, 

1987c; NTP, 1993b). In addition, in vitro studies 

have shown that exposure to 2-butoxyethanol may 

have a direct toxic effect on hematopoiesis (Ruchaud 

et al., 1992). 

Humans 

Workers have reported eye, skin, and respiratory tract 

irritations as well as headache, dizziness, 

lightheadedness, and nausea following exposure to 

2-butoxyethanol (ATSDR, 1998). Several acute 

poisoning cases have been reported. RambourgSchepens et al. (1988) reported a suicide case in 

which a 50-year-old woman ingested 250 to 500 mL 

of window cleaner containing 12% 2-butoxyethanol. 

When admitted to the hospital, she was described as 

comatose with labored respiration. Metabolic 

acidosis, hypokalemia, rise in serum creatinine, 

hemoglobinuria, and oxaluria were observed. The 

hemoglobinuria was paralleled by reduced erythrocyte 

count, hematocrit value, and hemoglobin 

concentration. Urinary 2-butoxyacetic acid and 

oxalate were measured during 8 days in the hospital. 

2-Butoxyethanol was excreted the greatest the first 

24 hours after ingestion (approximately 2.5 g 

2-butoxyethanol/g creatinine), whereas 2-butoxyacetic 

acid excretion peaked 2 days after ingestion at 40 g 

2-butoxyacetic acid/g creatinine and was still measurable 1 week after ingestion. On arrival, urinary 

oxalate was high (40 g/g creatinine). The authors 

concluded that the high urinary concentrations 

of 2-butoxyethanol were a result of overload of the 

two main metabolic pathways for 2-butoxyethanol. 

They also concluded that 2-butoxyethanol was also 

hydrolyzed to ethylene glycol, which was further 

metabolized to oxalate. ATSDR (1998) estimated that 

the dose was between 467 and 933 mg/kg. 

In another suicide attempt (Gijsenbergh et al., 1989), 

a 23-year-old woman weighing 64 kg ingested 250 to 

500 mL of a window cleaner containing 12.7% 
 

 

 

 

 

28 2-Butoxyethanol, NTP TR 484 

2-butoxyethanol and 3.2% ethanol. The dose was 

estimated at 400 to 500 mg/kg 2-butoxyethanol. On 

arrival, the patient was comatose with obstructive 

respiration and was placed on a respirator. Blood 

evaluation was normal; however, within an hour, she 

was in metabolic acidosis, and hemodialysis was 

started. On the second day, her hemoglobin concentration decreased from 11.9 g/L to 8.9 g/L at the 

same time she was excreting hemoglobin in her urine. 

2-Butoxyethanol and 2-butoxyacetic acid concentrations were measured in the dialysis fluid, and 

2-butoxyacetic acid was measured in the urine. The 

half-life of 2-butoxyethanol was 210 minutes. 

2-Butoxyacetic acid in urine reached a maximum 

(about 7.5 g/g creatinine) 24 hours after arrival. Unlike the first case, there was no oxaluria. The authors 

concluded that hemodialysis eliminating the 

2-butoxyethanol in the blood prevented the oxaluric 

situation observed in the previous case. 

Bauer et al. (1992) reported a case of a 53-year old 

male alcoholic who attempted suicide by drinking 

approximately 500 mL of a household cleaning fluid 

(9.1% 2-butoxyethanol, 2.5% ethanol). The patient 

was comatose and in metabolic acidosis and 

hypoxemia. Serum 2-butoxyethanol concentration 

was 5.28 g/L. 2-Butoxyethanol was not detected in 

gastric lavagate or urine. Hemoglobin (9.4 g/dL) and 

hematocrit (25%) were reduced. Respiratory support 

and hemodialysis were employed, and the patient 

recovered. 

Several controlled inhalation studies with 

2-butoxyethanol have been conducted in humans. 

Carpenter et al. (1956) exposed human volunteers to 

an atmosphere containing 98, 113, or 195 ppm 

2-butoxyethanol for 4 or 8 hours. No evidence of 

erythrocyte fragility, hemolysis, or a systemic toxic 

effect was reported. The volunteers complained of 

nasal and eye irritation, and some reported nausea, 

headaches, a disagreeable metallic taste, and 

occasional belching. 2-Butoxyacetic acid concentrations in the urine of individuals exposed to 98 ppm 

ranged from 75 to 250 mg. 

Johanson et al. (1986b) exposed seven male 

volunteers to an atmosphere containing 20 ppm 

2-butoxyethanol for 2 hours during light exercise on 

a bicycle ergometer. None of the volunteers 

complained of or showed evidence of adverse effects 

that could be attributed to 2-butoxyethanol exposure. 

There was no effect of exposure on pulmonary 

ventilation, respiratory frequency, or heart rate. In 

another study, Johanson and Boman (1991) orally 

exposed four male volunteers to 50 ppm 

2-butoxyethanol for 2 hours, followed by 1 hour with 

no exposure and then by 2 hours of dermal exposure 

to 50 ppm 2-butoxyethanol to determine percutaneous 

absorption of 2-butoxyethanol vapors. No treatmentrelated toxicity was observed. Johanson et al. (1988) 

reported the percutaneous exposure of five men to 

neat 2-butoxyethanol. The volunteers placed four 

fingers in a jar of 2-butoxyethanol for 2 hours. The 

skin of the exposed fingers appeared to be more rigid, 

less elastic, and wrinkled after exposure. Within a 

few hours of exposure, a dry reticulate pattern with 

several fissures, some of which became erythematous, 

was observed but disappeared within 1 to 2 days. No 

other treatment-related effect was observed. 

Greenspan et al. (1995) exposed 201 adults to 0.2 mL 

2-butoxyethanol (10% aqueous solution, the highest 

concentration used in cosmetic products) on the back. 

The dosed area was occluded. Testing included 

dosing, removal of the patch within 24 hours, evaluating the exposed area at 48 hours, and application 

of an identical patch to the same location. The 

subjects received nine such applications over a 6-week 

period. By the end of the 6 weeks, 25% of the subjects showed slight or definite erythema. 

Followingapplication to a naive area at 6 weeks, slight 

erythema was observed in seven subjects at 48 hours 

and 12 at 72 hours; one subject had definite erythema 

at 72 hours. No other effects were noted. 

REPRODUCTIVE 

AND DEVELOPMENTAL TOXICITY 

Experimental Animals 

The short-chain alkyl glycol ethers, 2-methoxyethanol 

and 2-ethoxyethanol, and their alkoxyacetic acid 

metabolites are potent male reproductive toxicants and 

teratogens in several species (Hardin, 1983; Hardin 

et al., 1984, 1987; Nagano et al., 1984; Wier et al., 

1987; Morrissey et al., 1989; Heindel et al., 1990; 

Schwetz and Harris, 1993). However, neither 

2-butoxyethanol nor 2-butoxyacetic acid has clearly 

been shown to cause reproductive effects. 

There was no effect of 2-butoxyethanol exposure on 

the male rat or mouse reproductive system in several 
 

 

 

 

2-Butoxyethanol, NTP TR 484 29 

repeated exposure studies via inhalation, gavage, or 

drinking water (Dodd et al., 1983; Nagano et al., 

1984; Grant et al., 1985; Krasavage, 1986; Exon 

et al., 1991; NTP, 1993b). 

Foster et al. (1987) compared the in vivo and in vitro 

testicular effects produced by the alkoxyacetic 

acid metabolites of 2-butoxy-, 2-ethoxy-, and 

2-methoxyethanol. Wistar rats were gavaged once 

with 0, 174, 434, or 868 mg/kg 2-butoxyacetic acid; 

137, 342, or 684 mg/kg 2-ethoxyacetic acid; and 118, 

296, or 592 mg/kg 2-methoxyacetic acid and followed 

for 14 days. All doses of all three acids caused 

decreased mean body weight gain during the first 

2 days with recovery by day 14. Animals dosed with 

868 mg/kg 2-butoxyacetic acid showed evidence 

of hematuria throughout the study. Only 

2-methoxyacetic acid (592 mg/kg) caused a significant 

decrease in testis weight. Histologic evidence of 

testicular damage was observed in all 2-methoxyacetic 

acid groups; similar effects in 2-ethoxyacetic acid 

groups were less severe than those in 2-methoxyacetic 

acid groups. No evidence of testicular damage was 

observed in 2-butoxyacetic acid treated groups. 

Likewise, in an in vitro study with mixed cultures of 

Sertoli and germ cells, 2-methoxyacetic acid and, to 

a lesser extent, 2-ethoxyacetic acid enhanced germ 

cell loss. 2-Butoxyacetic acid at equimolar concentrations had no effect on the cultured cells. All acids 

were tested at 0 to 10 mM. For the testis, glycol 

ether-induced toxicity was more apparent in the 

shorter alkyl chain alkoxyacetic acids; 

2-methoxyacetic acid was more sensitive than 

2-ethoxyacetic acid, followed by 2-butoxyacetic acid. 

Heindel et al. (1990) reported the effect of 

2-butoxyethanol treatment in drinking water (0%, 

0.5%, 1.0%, or 2.0%) to male and female Swiss 

(CD-1®) mice in a 98-day continuous breeding study. 

In a 2-week pilot study, males and females treated 

with 2.5% or greater 2-butoxyethanol lost weight, and 

mortality occurred at 5.0%. In the continuous breeding study, 13 of 20 females in the 2.0% group died 

compared to 6 in the 1.0% group. Male mice given 

1.0% or 2.0% 2-butoxyethanol lost weight during the 

study, and females in the 2.0% group gained considerably less than controls. Treated mice consumed 

less water than controls. At doses causing maternal 

toxicity, there was a reduction in the number of live 

pups per litter, proportion of pups born alive, and live 

pup weight. Following the continuous breeding 

study, a crossover mating study using F0 mice from 

control and 1.0% 2-butoxyethanol groups was performed. In female mice treated with 1.0% 

2-butoxyethanol and mated with control males, there 

was a significant reduction in the fertility index 

(number fertile/number with copulatory plugs) and a 

reduction in the number of live pups per litter. There 

was no effect in mating index (number with copulatory plugs/number cohabited). Other than reduced 

body weight, there were no treatment-related effects 

on sperm indices or histopathology in male mice 

treated with 1.0% 2-butoxyethanol. In addition to 

body weight reduction, female mice treated with 1.0% 

2-butoxyethanol had increased liver weights. There 

were no histopathologic effects or effects on estrous 

cycle stages or average cycle length. However, 7 of 

13 females had cycles longer than 7 days compared to 

controls in which 9 of 38 were abnormal. Because of 

the lack of sufficient pups from the first generation 

study at 1.0%, the second generation study was 

conducted using the 0.5% group. There were no 

treatment-related effects on mating or fertility indices 

or other reproductive parameters. The only effects 

related to 2-butoxyethanol exposure were increased 

liver weights in males and females and increased 

kidney weights in females. 

A number of developmental toxicity studies, both 

in vivo and in vitro, have been conducted with 

2-butoxyethanol. Hardin et al. (1987) reported the 

effects of 2-butoxyethanol gavage treatment 

(1,180 mg/kg per day) in Swiss (CD-1®) mice during 

gestation days 6 and 13. The dose selected was the 

LD10 determined by a previous pilot study. During 

the study, 10 of the 50 dams died and the survivors 

had a significant reduction in weight gain when 

compared to controls. The number of viable litters 

was significantly reduced from controls. There were 

no treatment-related effects on neonatal response 

variables. Schwetz and Harris (1993) reported the 

results of two developmental toxicity studies in rats 

treated with 2-butoxyethanol. Rats were gavaged 

with 30 to 200 mg/kg 2-butoxyethanol during 

gestation days 9 to 11 or 30 to 300 mg/kg during days 

11 to 13. Maternal toxicity, but not developmental 

toxicity, was observed in both studies at all doses. 

Wier et al. (1987) investigated the potential 

teratogenic and postnatal growth effects in Swiss 

(CD-1®) mice given 0, 350, 650, 1,000, 1,500, or 
 

 

 

 

 

30 2-Butoxyethanol, NTP TR 484 

2,000 mg 2-butoxyethanol/kg by gavage during 

gestation days 8 to 14. Mortality occurred in all six 

dams in the 2,000 mg/kg group and three of six in the 

1,500 mg/kg group. There were significant increases 

in resorptions in the surviving 1,000 and 1,500 mg/kg 

females. There was no evidence of 2-butoxyethanolrelated toxicity during the postnatal assessment. 

Nelson et al. (1984) exposed pregnant SpragueDawley rats to 150 or 200 ppm 2-butoxyethanol by 

inhalation 7 hours per day during gestation days 7 to 

15. Hematuria was noted in the dams only on day 1 

of exposure. There were no other treatment-related 

effects in the dams, and there were no teratogenic 

effects due to exposure to 2-butoxyethanol. 

In a teratology study reported by Tyl et al. (1984), 

pregnant F344 rats and New Zealand white rabbits 

were exposed to 0, 25, 50, 100, or 200 ppm 

2-butoxyethanol on gestation days 6 to 15 (rats) or 6 

to 18 (rabbits). Significant reductions in rate of 

weight gain and water consumption were observed in 

dams exposed to 200 ppm. Feed consumption was 

decreased in dams exposed to 100 or 200 ppm 

2-butoxyethanol. Also of interest is that during the 

exposure period, dams exposed to 200 ppm had tails 

that were deemed discolored and ulcerated. These 

tails were missing later, as previously reported. 

Following sacrifice on gestational day 21, gravid 

uterine weight was significantly reduced and absolute 

and relative spleen weights and relative kidney weight 

were elevated in the 200 ppm group when compared 

to controls. In addition, dams exposed to 100 or 

200 ppm 2-butoxyethanol had reduced erythrocyte 

counts and increased mean cell hemoglobin concentration. Hematocrit value was significantly increased only in dams exposed to 200 ppm. The 

authors concluded that these findings are consistent 

with destruction of mature erythrocytes and release of 

immature and/or young erythrocytes into the peripheral circulation. For rabbit dams there was 

treatment-related mortality, depressed body weight 

during the exposure period, and increases in spontaneous abortions. There was a significant decrease 

in gravid uterine weight in dams exposed to 200 ppm. 

There was no effect of exposure on hematologic 

parameters as observed in rats. For rats exposed to 

200 ppm 2-butoxyethanol, the number of viable 

implants and percentage of live fetuses per litter were 

reduced. The number of nonviable implants, due to 

early resorptions, was increased. There were no 

significant increases in external, visceral, skeletal, or 

total malformations in the fetuses due to treatment. 

However, there was evidence of retarded skeletal 

ossification at 100 and 200 ppm. For rabbits, as with 

rats at 200 ppm 2-butoxyethanol, there was a significant reduction in the number of viable implants per 

litter; however, there was no effect of exposure on the 

number of nonviable implants. The authors 

concluded that exposure to 2-butoxyethanol resulted 

in maternal, embryonic, and fetal toxicity in rats at 

100 or 200 ppm 2-butoxyethanol; maternal and 

embryonic toxicity in rabbits at 200 ppm; and no 

teratogenicity in rats or rabbits. 

In a comparative developmental toxicity study, Hardin 

et al. (1984) tested five glycol ethers, including 

2-butoxyethanol, administered to SPF SpragueDawley rats dermally. The original design was to test 

all five ethers at equimolar concentrations during 

gestation days 7 to 16; however, during the first 

replicate, 2-butoxyethanol at 0.35 mL was determined 

to be overtly toxic to the dams. In a subsequent 

replicate, the 2-butoxyethanol dose was reduced to 

0.12 mL and, like the other glycol ethers, was administered four times a day (at 2.5 hour intervals) on 

gestation days 7 to 16. 2-Butoxyethanol at this dose 

caused no maternal toxicity, was not toxic to the 

embryo or fetus, and did not cause teratogenic effects. 

Diethylene glycol monoethyl ether (0.35 mL) caused 

a reduction in weight gain but, like 2-butoxyethanol, 

was not toxic to the embryo or fetus and was not 

teratogenic. 2-Ethoxyethanol (0.25) and ethylene 

glycol monoethyl ether acetate (0.35 mL) caused 

reduced weight gain in dams associated with completely resorbed litters and significantly fewer live fetuses per litter. Fetal body weights were reduced and 

visceral malformations and skeletal variations were 

increased when compared to controls. 

Giavini et al. (1993) compared the embryotoxic 

potential of 2-butoxyethanol and 2-butoxyacetic acid, 

2-ethoxyethanol and 2-ethoxyacetic acid and 

2-methoxyethanol and 2-methoxyacetic acid in an 

in vitro system whereby 9.5-day-old embryos from 

CD rats were incubated with each glycol ether or 

alkoxy acetic acid for 48 hours. Final media 

concentrations for 2-butoxyethanol were 3.12, 6.25, 

12.5, or 25 mM and for 2-butoxyacetic acid were 0.4, 

0.8, 1.6, or 3.2 mM. For 2-butoxyethanol, 

embryonic development was blocked at 25 mM, and 

exposure to 12.5 mM 2-butoxyethanol resulted in 
 

2-Butoxyethanol, NTP TR 484 31 

severe dysmorphogenic effects (inhibition of rotation 

and severe reduction of telencephalic vesicles). 

General embryotoxic effects, as shown by reduction 

of the somite number and of protein/embryo, were 

observed at 6.25 mM. There were no effects of 

2-butoxyethanol at 3.12 mM. Incubation with 

2-butoxyacetic acid resulted in abnormal embryos 

with morphologic alterations of brain vesicle size. 

Reduction in somite number and total protein/embryo 

was observed at all incubation concentrations of 

2-butoxyacetic acid except at 0.4 mM 2-butoxyacetic 

acid, the no-effect level for all types of effects. Of 

importance is that in in vitro situations embryotoxicity 

increased with increased alkyl chain length of the 

glycol ether. 2-Butoxyethanol was more active than 

2-ethoxyethanol, followed by 2-methoxyethanol, but 

the opposite was observed for the corresponding 

alkoxyacid; 2-methoxyacetic acid was more sensitive 

than 2-ethoxyacetic acid, followed by 2-butoxyacetic 

acid. Moreover, the embryotoxic potency of each 

acid was considerably higher than that of the corresponding ether. 

In another study, Bowden et al. (1995) cultured 

10-day-old Sprague-Dawley rat embryos for 48 hours 

with 2-butoxy-, 2-ethoxy-, or 2-methoxyethanol at 

media concentrations of 0.3, 0.5, 0.75, or 

1.0 mg/mL. 2-Butoxyethanol caused marginal 

reduction in growth and developmental parameters at 

0.3 mg/mL. At all doses, the embryos had poor yolk 

sac circulation, thin allantois, twisted flexion, 

incomplete fusion, and/or irregular formation of the 

caudal neural tube and brain, irregular posterior 

neuropore, and growth-retarded forelimb buds. At 

1.0 mg/mL, five of 10 embryos died. As reported by 

Giavani et al. (1993), embryo toxicity was directly 

related to increased alkyl chain length of the glycol 

ether (2-butoxyethanol was more active than 

2-ethoxyethanol, followed by 2-methoxyethanol). The 

results of embryo culture with glycol ethers were the 

opposite of what was observed in inhalation and 

dermal in vivo rat studies, in which embryo toxicity 

was directly related to decreased chain length of 

glycol ethers; 2-methoxyethanol was more active than 

2-ethoxyethanol, followed by 2-butoxyethanol (Hardin 

et al., 1984; Nelson et al., 1984), which may be 

because the glycol ethers are rapidly metabolized to 

the alkoxyacids in vivo. 

Humans 

No information on the reproductive or developmental 

toxicity of 2-butoxyethanol in humans was found in 

the available literature. 

CARCINOGENICITY 

No information on the carcinogenicity of 

2-butoxyethanol in experimental animals was found in 

the available literature; additionally, no epidemiologic 

studies or case reports examining the relationship 

between exposure to 2-butoxyethanol and cancer in 

humans were found in the literature. 

GENETIC TOXICITY 

Published information on the genotoxicity of 

2-butoxyethanol, recently reviewed by Elliot and 

Ashby (1997), indicates that the chemical is not 

mutagenic, consistent with the absence of structural 

alerts to genotoxicity (Tennant and Ashby, 1991). 

Results of Salmonella typhimurium gene mutation 

assays with 2-butoxyethanol were negative in the 

presence and in the absence of induced hamster or rat 

liver S9 (Zeiger et al., 1992). Positive results were 

reported (Hoflack et al., 1995) with 2-butoxyethanol 

in a later S. typhimurim test with strain TA97a 

[closely related to TA97, which was used in the 

Zeiger et al. (1992) study and shows the same 

response as TA97 to mutagens that produce frameshift alterations], but an independent replication of the 

study using a well-characterized sample of 

2-butoxyethanol was unable to duplicate the positive 

response (Gollapudi et al., 1996). In addition, 

Gollapudi et al. (1996) tested 2-butoxyethanol for 

mutagenicity in TA100 and in Escherichia coli 

WP2 uvrA, and no mutagenic activity was detected. 

2-Butoxyethanol did not induce gene mutations in 

cultured Chinese hamster ovary AS52 cells in the 

absence of S9 activation and a major metabolite of 

2-butoxyethanol, 2-butoxyacetaldehyde, also gave 

negative results in this assay (Chiewchanwit and Au, 

1995). 

Keith et al. (1996) reported that no increase in DNA 

adducts was detected by 32P postlabeling in the brain, 

liver, kidney, testis, or spleen of Sprague-Dawley rats 

following oral administration of 120 mg/kg 

2-butoxyethanol. No increases in the frequencies of 
 

32 2-Butoxyethanol, NTP TR 484 

sister chromatid exchanges or micronucleated lymphocytes were observed in peripheral blood lymphocytes 

of varnish plant workers exposed to 2-butoxyethanol 

and other glycol ethers (Söhnlein et al., 1993). 

In summary, 2-butoxyethanol was not mutagenic in 

bacterial or mammalian cells in vitro, and no evidence 

of DNA damage was detected in vivo in rats or in 

exposed workers. This data set is limited, but all 

evidence supports the conclusion that 2-butoxyethanol 

is nonmutagenic. 

STUDY RATIONALE 

The Consumer Product Safety Commission and the 

United Auto Workers International Union nominated 

2-butoxyethanol for study because of its widespread 

use in industrial and consumer applications, the 

potential for exposure to workers and the general 

population, and the absence of chronic toxicity data. 

Inhalation was chosen as the route of exposure 

because human exposure occurs primarily by this 

route. 
 

 

33 

MATERIALS AND METHODS
 

PROCUREMENT 

AND CHARACTERIZATION 

OF 2-BUTOXYETHANOL 

2-Butoxyethanol was obtained from Dow Chemical 

U.S.A. (Plaquemine, LA) in two lots. 

Lot QP-911021-26D1 was used during the 14-week 

studies, and lot QP-921215-26D2 was used during the 

2-year studies. Identity and purity analyses were 

conducted by the study laboratory. Reports on 

analyses performed in support of the 2-butoxyethanol 

studies are on file at the National Institute of 

Environmental Health Sciences. 

Both lots of the chemical, a clear, colorless liquid, 

were identified as 2-butoxyethanol by infrared and 

nuclear magnetic resonance (proton and C13) spectroscopy. The infrared spectrum was consistent with 

that expected for the structure, with the literature 

spectra (Aldrich, 1981, 1983), and/or with those of a 

reference sample obtained from Aldrich Chemical 

Company (Milwaukee, WI). Purity of each lot was 

determined by elemental analysis, Karl Fischer water 

analysis, titrations for acid and peroxide content, and 

gas chromatography with flame ionization detection. 

For both lots, elemental analyses for carbon, hydrogen, and oxygen were in agreement with the theoretical values for 2-butoxyethanol. Karl Fischer 

water analysis indicated 0.02% water for 

lot QP-911021-26D1 and 0.0254% water for 

lot QP-921215-26D2. Titrations indicated 0.001% 

acidity (as acetic acid) and 105 ppm peroxide for lot 

QP-911021-26D1, well within the acceptable limits of 

0.02% acid and 5,000 ppm peroxide; titrations 

indicated less than 0.003% acetic acid and less than 

1,000 ppm peroxide for lot QP-921215-26D2. 

Gas chromatographic analysis of lot QP-911021-26D1 

indicated one major peak and one impurity with an 

area of 0.1% relative to the major peak area. Gas 

chromatographic analysis of lot QP-921215-26D2 

indicated one major peak and three impurities with 

areas greater than 0.1% of the major peak area; these 

impurities were tentatively identified as ethylene 

glycol, 2-ethyl-2-hexenal, and 2-ethyl-1-hexanol by 

gas chromatography/mass spectroscopy with electron 

impact ionization detection. Major peak comparisons 

of each lot relative to a reference sample were performed by gas chromatography; results indicated a 

purity of 100.8% for lot QP-911021-26D1 and 99.2% 

for lot QP-921215-26D2 relative to the reference 

sample. The overall purity of each lot was determined to be greater than 99%. 

Accelerated stability studies of the bulk chemical were 

performed by Midwest Research Institute (MRI, 

Kansas City, MO) (MRI, 1984). These studies 

indicated that 2-butoxyethanol is stable as a bulk 

chemical for 2 weeks when stored protected from 

light at temperatures up to 60E C. To ensure stability, 

the bulk chemical was stored at room temperature 

during the 14-week studies and at approximately 

16E C during the 2-year studies in stainless steel 

containers. Throughout the studies, stability was 

monitored by titrations for acid and peroxide and by 

gas chromatography with flame ionization detection. 

No degradation of the bulk chemical was detected. 

VAPOR GENERATION 

AND EXPOSURE SYSTEM 

2-Butoxyethanol was held in a stainless-steel reservoir 

under a nitrogen blanket. A liquid micrometering 

pump was used to pump 2-butoxyethanol into a glass 

column filled with glass beads and heated by flexible 

electric heat tape encircling the column. Vapor 

temperature was monitored at the top of the condenser 

column by a temperature sensor. The total output of 

the generator was calculated from the metered nitrogen flow and the 2-butoxyethanol vapor pressure at 

the exit temperature. 

To prevent 2-butoxyethanol from condensing while in 

transport to the exposure room, the Teflon® transport 

line was heated. The vapor was mixed with heated 

HEPA- and charcoal-filtered air before entering a 

short vapor distribution manifold. An automatic 
 

 

 

 

 

 

34 2-Butoxyethanol, NTP TR 484 

controller maintained a constant flow in the distribution manifold. 

Compressed-air pumps delivered the vapor from the 

distribution manifold to the exposure chambers via 

individual temperature-controlled Teflon® delivery 

lines. A three-way valve between the distribution line 

and each chamber directed vapor to the exposure 

chamber exhaust until a stable concentration of 

2-butoxyethanol vapor was collected in the 

distribution line. At each chamber, the vapor was 

further diluted with conditioned, HEPA- and 

charcoal-filtered chamber air to the appropriate 

2-butoxyethanol concentration. 

The study laboratory designed the inhalation exposure 

chamber (Harford Systems Division of Lab Products, 

Inc., Aberdeen, MD) so that uniform vapor concentrations could be maintained throughout the chamber 

with the catch pans in place. The total active mixing 

volume of each chamber was 1.7 m3

. A small-particle 

detector (Type CN, Gardner Associates, Schenectady, 

NY) was used with and without animals in the exposure chambers to ensure that 2-butoxyethanol vapor, 

and not aerosol, was produced. No particle counts 

above the minimum resolvable level (approximately 

200 particles/cm3

) were detected. 

PILOT STUDIES 

Pilot studies were conducted to determine the maximum concentration of 2-butoxyethanol vapor that 

could be generated without the presence of 2-butoxyethanol aerosols and to determine the acute toxicity of 

2-butoxyethanol in male and female rats and mice at 

those concentrations. Aerosols were generated at 

approximately 800 ppm; 4-week-old F344/N rats and 

B6C3F1 mice were exposed to 31 to 700 ppm 

2-butoxyethanol by inhalation for 3 to 14 days. 

VAPOR CONCENTRATION MONITORING 

Chamber concentrations of 2-butoxyethanol were 

monitored with an on-line gas chromatograph. 

Samples were drawn from each chamber approximately every 16 minutes (14-week studies) or 

30 minutes (2-year studies) during exposures by a 

computer-controlled, 12-port, stream select valve. 

The on-line gas chromatograph was calibrated by a 

comparison of chamber concentration data to data 

from grab samples analyzed by an off-line gas 

chromatograph; the grab samples were collected in 

bubblers containing water. The off-line gas 

chromatograph was calibrated with gravimetrically 

prepared standards of 2-butoxyethanol during the 

14-week and 2-year studies. Summaries of the 

chamber concentrations for the 14-week and 2-year 

studies are in Tables H2 and H3. 

CHAMBER ATMOSPHERE 

CHARACTERIZATION 

At a chamber airflow rate of 15 air changes per hour, 

the theoretical value for the time to achieve 90% of 

the target concentration after the beginning of vapor 

generation (T90) and the time for the chamber concentration to decay to 10% of the target concentration 

after vapor generation was terminated (T10) was 

approximately 12.5 minutes. Prior to and during the 

14-week and 2-year studies, T90 and T10 ranges were 

determined with and without animals. A T90 value of 

12 minutes was selected for all studies. 

Studies of 2-butoxyethanol degradation and monitoring for impurities were conducted throughout the 

studies by gas chromatography; chamber concentration uniformity was maintained throughout the 

14-week and 2-year studies, and no significant degradation of 2-butoxyethanol was observed. 

14-WEEK STUDIES 

The 14-week studies were conducted to evaluate the 

cumulative toxic effects of repeated exposure to 

2-butoxyethanol and to determine the appropriate 

exposure concentrations to be used in the 2-year 

studies. 

Male and female F344/N rats and B6C3F1 mice were 

obtained from Taconic Laboratory Animals and 

Services (Germantown, NY). On receipt, the rats and 

mice were approximately 4 weeks old. Animals were 

quarantined for 11 or 12 days and were 6 weeks old 

on the first day of the studies. Before initiation of the 

studies, five male and five female rats and mice were 

randomly selected for parasite evaluation and gross 

observation for evidence of disease. Serologic analyses were performed on five male and five female 

sentinel rats and mice on day 10 and on five male and 
 

 

2-Butoxyethanol, NTP TR 484 35 

five female control rats and mice at the end of the 

studies using the protocols of the NTP Sentinel 

Animal Program (Appendix J). 

Groups of 10 male and 10 female rats and mice were 

exposed to 2-butoxyethanol at concentrations of 0, 31, 

62.5, 125, 250, or 500 ppm by inhalation, 6 hours 

plus T90 (12 minutes) per day, 5 days per week for 

14 weeks. Water was available ad libitum; feed was 

available ad libitum except during exposure periods. 

Rats and mice were housed individually. Animals 

were weighed initially, weekly, and at the end of the 

studies. Clinical findings were recorded weekly and 

at the end of the studies. Details of the study design 

and animal maintenance are summarized in Table 1. 

At the end of the 14-week studies, rats and mice were 

anesthetized with a CO2:O2 mixture, and blood was 

collected from the retroorbital sinus for hematology 

analyses and placed in tubes containing potassium 

EDTA as the anticoagulant. Hematology determinations, including erythrocyte, leukocyte, and 

platelet counts, hemoglobin concentration, packed cell 

volume, mean cell volume, mean cell hemoglobin, 

and mean cell hemoglobin concentration, were performed on an Ortho ELT-8/ds 9000 hematology 

analyzer (Ortho Diagnostic Systems, Westwood, 

MA). Hematocrit was also determined manually 

using a Damon/IEC microcapillary centrifuge and 

reader (International Equipment Company, Needham 

Heights, MA). Leukocyte differential and nucleated 

erythrocyte counts were determined by light microscopic examination of blood films stained with 

Wright’s stain on a Wescor Stainer (Wescor, Logan, 

UT). Smears made from preparations of equal 

volumes of new methylene blue and whole blood and 

incubated at room temperature for 20 minutes were 

examined microscopically, using the Miller disc 

method, for the quantitative determination of 

reticulocytes. 

A necropsy was performed on all animals. The heart, 

right kidney, liver, lungs, right testis, and thymus 

were weighed. Tissues for microscopic examination 

were fixed and preserved in 10% neutral buffered 

formalin, processed and trimmed, embedded in 

paraffin, sectioned to a thickness of 5 to 6 μm, and 

stained with hematoxylin and eosin. A complete 

histopathologic examination was performed on chamber control and 500 ppm rats and mice and on 

250 ppm female rats. Target organs were identified 

and evaluated to a no-observable-adverse-effect level. 

Table 1 lists the tissues and organs routinely 

examined. 

2-YEAR STUDIES 

Study Design 

Groups of 50 male and 50 female rats and mice were 

exposed to 2-butoxyethanol by inhalation, 6 hours 

plus T90 (12 minutes) per day, 5 days per week for 

104 weeks. Rats were exposed to 0, 31.2, 62.5, or 

125 ppm, and mice were exposed to 0, 62.5, 125, or 

250 ppm. 

For hematology and bone marrow analyses, additional 

groups of 27 male and 27 female rats were exposed to 

0, 62.5, or 125 ppm 2-butoxyethanol, nine male and 

nine female rats were exposed to 31.2 ppm 

2-butoxyethanol, and 30 male and 30 female mice 

were exposed to 0, 62.5, 125, or 250 ppm 

2-butoxyethanol. Nine male and nine female rats 

exposed to 0, 62.5, or 125 ppm 2-butoxyethanol and 

10 male and 10 female mice from each exposure 

group were evaluated at 3, 6, or 12 months; nine male 

and nine female rats exposed to 31.2 ppm 

2-butoxyethanol were evaluated at 3 (hematology 

only) and 6 months. 

Source and Specification of Animals 

Male and female F344/N rats and B6C3F1 mice were 

obtained from Taconic Laboratory Animals and 

Services for use in the 2-year studies. Rats and mice 

were quarantined for 18 days before the beginning of 

the studies. Five male and five female rats and mice 

were randomly selected for parasite evaluation and 

gross observation of disease. Rats and mice were 7 to 

8 weeks old at the beginning of the studies. The 

health of the animals was monitored during the studies 

according to the protocols of the NTP Sentinel Animal 

Program (Appendix J). 

Animal Maintenance 

Rats and mice were housed individually. Water was 

available ad libitum; feed was available ad libitum 

except during exposure periods. Cages were rotated 

weekly. Further details of animal maintenance are 

given in Table 1. Information on feed composition 

and contaminants is provided in Appendix I. 
 

 

 

 

 

36 2-Butoxyethanol, NTP TR 484 

Clinical Examinations and Pathology 

All animals were observed twice daily. Body weights 

were recorded initially; body weights and clinical 

findings were recorded monthly from week 5 through 

week 89 (rats) or week 93 (mice) and every 2 weeks 

from week 92 (rats) or week 94 (mice) until the end 

of the studies. 

At 3, 6, and 12 months, the rats and mice selected as 

part of the hematology studies were anesthetized with 

a CO2:O2 mixture, and blood was collected from the 

retroorbital plexus and placed in tubes containing 

potassium EDTA as the anticoagulant. As in the 

14-week studies, 3- and 6-month hematology 

determinations were performed on an Ortho ELT-8/ds 

9000 hematology analyzer; a Roche COBAS Helios 

analyzer (Roche Diagnostic Systems, Inc., Montclair, 

NJ) was used at 12 months. Leukocyte differential 

and nucleated erythrocyte counts were determined by 

light microscopic examination of blood films stained 

with Wright’s stain using a Wescor Stainer. Bone 

marrow samples were taken by flushing the cells from 

the femur into tubes containing tissue culture media. 

Marrow cellularity was enumerated using a 

hydrodynamically focused electronic impedance 

system (Coulter Model ZH,, Coulter Electronics, Inc., 

Hialeah, FL). Cytological evaluations of bone 

marrow cell morphology and myeloid/erythroid ratios 

were performed by microscopic examination of 

Wright’s-stained, cytocentrifuged preparations from 

bone marrow samples collected for cellularity counts. 

Hematology and bone marrow cellularity parameters 

measured are listed in Table 1. 

Complete necropsies and microscopic examinations 

were performed on all core study rats and mice. At 

necropsy, all organs and tissues were examined for 

grossly visible lesions, and all major tissues were 

fixed and preserved in 10% neutral buffered formalin, 

processed and trimmed, embedded in paraffin, 

sectioned to a thickness of 5 to 6 μm, and stained with 

hematoxylin and eosin for microscopic examination. 

For all paired organs (e.g., adrenal gland, kidney, 

ovary), samples from each organ were examined. 

Tissues examined microscopically are listed in 

Table 1. 

Microscopic evaluations were completed by the study 

laboratory pathologist, and the pathology data were 

entered into the Toxicology Data Management 

System. The slides, paraffin blocks, and residual wet 

tissues were sent to the NTP Archives for inventory, 

slide/block match, and wet tissue audit. The slides, 

individual animal data records, and pathology tables 

were evaluated by an independent quality assessment 

laboratory. The individual animal records and tables 

were compared for accuracy, the slide and tissue 

counts were verified, and the histotechnique was 

evaluated. For the 2-year studies, a quality assessment pathologist evaluated slides from all tumors and 

all potential target organs, which included the bone 

marrow, forestomach, kidney, liver, lung, nose, and 

spleen of all male and female rats and mice; the 

adrenal gland and clitoral gland of female rats; and 

the preputial gland, prostate gland, skin (prepuce), 

testis, and urinary bladder of male mice. 

The quality assessment report and the reviewed slides 

were submitted to the NTP Pathology Working Group 

(PWG) chairperson, who reviewed the selected tissues 

and addressed any inconsistencies in the diagnoses 

made by the laboratory and quality assessment 

pathologists. Representative histopathology slides 

containing examples of lesions related to chemical 

administration, examples of disagreements in diagnoses between the laboratory and quality assessment 

pathologists, or lesions of general interest were 

presented by the chairperson to the PWG for review. 

The PWG consisted of the quality assessment 

pathologist and other pathologists experienced in 

rodent toxicologic pathology. This group examined 

the tissues without any knowledge of dose groups or 

previously rendered diagnoses. When the PWG 

consensus differed from the opinion of the laboratory 

pathologist, the diagnosis was changed. Final diagnoses for reviewed lesions represent a consensus 

between the laboratory pathologist, reviewing pathologist(s), and the PWG. Details of these review 

procedures have been described, in part, by Maronpot 

and Boorman (1982) and Boorman et al. (1985). For 

subsequent analyses of the pathology data, the decision of whether to evaluate the diagnosed lesions for 

each tissue type separately or combined was generally 

based on the guidelines of McConnell et al. (1986). 

Toxicokinetic Studies 

The study design and results of toxicokinetic studies 

conducted utilizing special study rats and mice during 

the 2-year studies are provided in Dill et al. (1998) 

and Lee et al. (1998). 
2-Butoxyethanol, NTP TR 484 37 

TABLE 1 

Experimental Design and Materials and Methods in the Inhalation Studies of 2-Butoxyethanol 

14-Week Studies 2-Year Studies 

Study Laboratory 

Battelle Pacific Northwest Laboratories 

(Richland, WA) 

Strain and Species 

Rats: F344/N 

Mice: B6C3F1 

Animal Source 

Taconic Laboratory Animals and Services (Germantown, NY) 

Time Held Before Studies 

11 days (males) or 12 days (females) 

Average Age When Studies Began 

6 weeks 

Date of First Exposure 

23 (males) or 24 (females) March 1992 

Duration of Exposure 

6 hours plus T90 (12 minutes) per day, 5 days per week, for 

14 weeks 

Date of Last Exposure 

Rats: 22 (males) or 23 (females) June 1992 

Mice: 24 (males) or 25 (females) June 1992 

Necropsy Dates 

Rats: 23 (males) or 24 (females) June 1992 

Mice: 25 (males) or 26 (females) June 1992 

Average Age at Necropsy 

19 weeks 

Size of Study Groups 

10 males and 10 females 

Method of Distribution 

Animals were distributed randomly into groups of approximately 

equal initial mean body weight. 

Animals per Cage 

Method of Animal Identification 

Tail tattoo 

Battelle Pacific Northwest Laboratories 

(Richland, WA) 

Rats: F344/N 

Mice: B6C3F1 

Taconic Laboratory Animals and Services (Germantown, NY) 

18 days 

Rats: 7-8 weeks 

Mice: 7 weeks 

Rats: 23 August 1993 

Mice: 26 July 1993 

6 hours plus T90 (12 minutes) per day, 5 days per week, for 

104 weeks 

Rats: 18 August 1995 

Mice: 21 July 1995 

Rats: 21-23 August 1995 

Mice: 24-28 July 1995 

Rats: 111-112 weeks 

Mice: 111 weeks 

Core study: 50 males and 50 females 

Hematology and bone marrow analyses: 

Rats: 27 males and 27 females (0, 62.5, or 125 ppm), or 

9 males and 9 females (31.2 ppm) 

Mice: 30 males and 30 females 

Same as 14-week studies 

1 

Tail tattoo 

1 
38 2-Butoxyethanol, NTP TR 484 

TABLE 1 

Experimental Design and Materials and Methods in the Inhalation Studies of 2-Butoxyethanol 

14-Week Studies 2-Year Studies 

Diet 

NIH-07 open formula pelleted diet (Zeigler Brothers, Inc., 

Gardners, PA), available ad libitum, except during exposure 

periods; changed weekly 

Water 

Tap water (City of Richland municipal supply) softened at 

Battelle and delivered via automatic watering system (Edstrom 

Industries, Waterford, WI), available ad libitum 

Cages 

Stainless steel wire-bottom cages (Hazleton Systems, Inc., 

Aberdeen, MD), changed weekly 

Chamber Air Supply Filters 

Single HEPA (Northland Filter Systems International, 

Mechanicville, NY) and charcoal (RSE, Inc., New Baltimore, 

MI) 

Chambers 

Stainless steel chambers (Lab Products, Inc., Harford System 

Division, Aberdeen, MD), changed weekly 

Chamber Environment 

Temperature: 23.9E-24.3E C 

Relative humidity: 55%-56% 

Room fluorescent light: 12 hours/day 

Chamber air changes: 15/hour 

Exposure Concentrations 

0, 31, 62.5, 125, 250, or 500 ppm 

Type and Frequency of Observation 

Observed twice daily; animals were weighed initially, weekly,
 

and 

at the end of the studies. Clinical findings were recorded weekly
 

and at the end of the studies.
 

Method of Sacrifice 

Asphyxiation with 70% CO2 

Necropsy 

Necropsy performed on all animals. Organs weighed were heart, 

right kidney, liver, lung, right testis, and thymus. 

Same as 14-week studies 

Same as 14-week studies 

Same as 14-week studies 

Single HEPA (Flanders Filters, Inc., San Rafael, CA) and purafil
 

(Environmental Systems, Lynnwood, WA)
 

Same as 14-week studies
 

Temperature: 23.7E-24.1E C (rats) or 24.0E-24.3E C (mice)
 

Relative humidity: 57%-58% (rats) or 54%-56% (mice)
 

Room fluorescent light: 12 hours/day
 

Chamber air changes: 15/hour
 

Rats: 0, 31.2, 62.5, or 125 ppm
 

Mice: 0, 62.5, 125, or 250 ppm
 

Observed twice daily; animals were weighed initially and body
 

weights and clinical findings were recorded monthly from week 5
 

through week 89 (rats) or week 93 (mice) and every 2 weeks from
 

week 92 (rats) or week 94 (mice) until the end of the studies.
 

Asphyxiation with 70% CO2 

Necropsy performed on core study animals. 
2-Butoxyethanol, NTP TR 484 39 

TABLE 1 

Experimental Design and Materials and Methods in the Inhalation Studies of 2-Butoxyethanol 

14-Week Studies 2-Year Studies 

Clinical Pathology 

Blood was collected from the retroorbital sinus of all animals 

surviving to the end of the studies for hematology analyses. 

Hematology: automated hematocrit; manual hematocrit; 

hemoglobin concentration; erythrocyte, reticulocyte, and platelet 

counts; mean cell volume; mean cell hemoglobin; mean cell 

hemoglobin concentration; total leukocyte count and differentials; 

and morphologic assessment of erythrocytes, platelets, and 

leukocytes 

Histopathology 

Complete histopathology was performed on 0 and 500 ppm rats, 

250 ppm female rats, and 0 and 500 ppm mice. In addition to 

gross lesions and tissue masses, the following tissues were 

examined: adrenal gland, bone with marrow, brain, clitoral 

gland, esophagus, gallbladder (mice), heart, large intestine 

(cecum, colon, rectum), small intestine (duodenum, jejunum, 

ileum), kidney, larynx, liver, lung, lymph nodes (mandibular, 

mesenteric, bronchial, mediastinal), mammary gland, nose, 

ovary, pancreas, parathyroid gland, pituitary gland, preputial 

gland, prostate gland, salivary gland, spleen, stomach 

(forestomach and glandular), testis (with epididymis and seminal 

vesicle), thymus, thyroid gland, trachea, urinary bladder and 

uterus. In addition, the bone marrow, forestomach, kidney, liver, 

and spleen of male rats; nose, salivary gland, tail, and thymus of 

female rats; and the kidney, liver, lung, lymph nodes (mandibular 

and mesenteric), stomach, testis, and thymus of male and female 

mice were examined at all other exposure levels. 

Blood was collected from the retroorbital plexus of nine male and 

nine female rats and 10 male and 10 female mice designated for 

hematology analyses at 3, 6, and/or 12 months (except 31.2 ppm 

rats). Femurs were collected from these same animals for bone 

marrow analyses. 

Hematology: automated hematocrit; manual hematocrit; 

hemoglobin concentration; erythrocyte, reticulocyte, and platelet 

counts; mean cell volume; mean cell hemoglobin; mean cell 

hemoglobin concentration; total leukocyte count and differentials; 

and morphologic assessment of erythrocytes, platelets, and 

leukocytes 

Bone Marrow: total nucleated bone marrow cell count and 

myeloid/ erythroid ratio 

Complete histopathology was performed on all core study rats and 

mice. In addition to gross lesions and tissue masses, the following 

tissues were examined: adrenal gland, bone with marrow, brain, 

clitoral gland, esophagus, gallbladder (mice), heart, large intestine 

(cecum, colon, rectum), small intestine (duodenum, jejunum, 

ileum), kidney, larynx, liver, lung, lymph nodes (mandibular, 

mesenteric, bronchial, mediastinal), mammary gland (except male 

mice), nose, ovary, pancreas, parathyroid gland, pituitary gland, 

preputial gland, prostate gland, salivary gland, spleen, stomach 

(forestomach and glandular), testis (with epididymis and seminal 

vesicle), thymus, thyroid gland, trachea, urinary bladder, and 

uterus. 

STATISTICAL METHODS 

Survival Analyses 

The probability of survival was estimated by the 

product-limit procedure of Kaplan and Meier (1958) 

and is presented in the form of graphs. Animals 

found dead of other than natural causes or missing 

were censored from the survival analyses; animals 

dying from natural causes were not censored. Statistical analyses for possible dose-related effects on 

survival used Cox’s (1972) method for testing two 

groups for equality and Tarone’s (1975) life table test 

to identify dose-related trends. All reported P values 

for the survival analyses are two sided. 

Calculation of Incidence 

The incidences of neoplasms or nonneoplastic lesions 

are presented in Tables A1, A4, B1, B5, C1, C5, D1, 

and D5 as the numbers of animals bearing such 

lesions at a specific anatomic site and the numbers of 

animals with that site examined microscopically. For 

calculation of statistical significance, the incidences of 

most neoplasms (Tables A3, B3, C3, and D3) and all 

nonneoplastic lesions are given as the numbers of 

animals affected at each site examined microscopically. However, when macroscopic examination 

was required to detect neoplasms in certain tissues 
 

 

 

 

 

40 2-Butoxyethanol, NTP TR 484 

(e.g., harderian gland, intestine, mammary gland, and 

skin) before microscopic evaluation, or when neoplasms had multiple potential sites of occurrence 

(e.g., leukemia or lymphoma), the denominators 

consist of the number of animals on which a necropsy 

was performed. Tables A3, B3, C3, and D3 also give 

the survival-adjusted neoplasm rate for each group 

and each site-specific neoplasm. This survivaladjusted rate (based on the Poly-3 method described 

below) accounts for differential mortality by assigning 

a reduced risk of neoplasm, proportional to the third 

power of the fraction of time on study, to animals that 

do not reach terminal sacrifice. 

Analysis of Neoplasm 

and Nonneoplastic Lesion Incidences 

The Poly-k test (Bailer and Portier, 1988; Portier and 

Bailer, 1989; Piegorsch and Bailer, 1997) was used to 

assess neoplasm and nonneoplastic lesion prevalence. 

This test is a survival-adjusted quantal-response 

procedure that modifies the Cochran-Armitage linear 

trend test to take survival differences into account. 

More specifically, this method modifies the denominator in the quantal estimate of lesion incidence to 

approximate more closely the total number of animal 

years at risk. For analysis of a given site, each 

animal is assigned a risk weight. This value is one if 

the animal had a lesion at that site or if it survived 

until terminal sacrifice; if the animal died prior to 

terminal sacrifice and did not have a lesion at that 

site, its risk weight is the fraction of the entire study 

time that it survived, raised to the kth power. 

This method yields a lesion prevalence rate that 

depends only upon the choice of a shape parameter 

for a Weibull hazard function describing cumulative 

lesion incidence over time (Bailer and Portier, 1988). 

Unless otherwise specified, a value of k=3 was used 

in the analysis of site-specific lesions. This value was 

recommended by Bailer and Portier (1988) following 

an evaluation of neoplasm onset time distributions for 

a variety of site-specific neoplasms in control F344 

rats and B6C3F1 mice (Portier et al., 1986). Bailer 

and Portier (1988) showed that the Poly-3 test 

gave valid results if the true value of k was anywhere 

in the range from 1 to 5. A further advantage of the 

Poly-3 method is that it does not require lesion 

lethality assumptions. Variation introduced by the use 

of risk weights, which reflect differential mortality, 

was accommodated by adjusting the variance of the 

Poly-3 statistic as recommended by Bieler and 

Williams (1993). 

Tests of significance included pairwise comparisons of 

each exposed group with controls and a test for an 

overall exposure-related trend. Continuity-corrected 

tests were used in the analysis of lesion incidence, and 

reported P values are one sided. Values of P greater 

than 0.5 are presented as 1-P with the letter N added 

to indicate a lower incidence or negative trend in 

neoplasm occurrence relative to the control group 

(e.g., P=0.99 is presented as P=0.01N). 

Analysis of Continuous Variables 

Two approaches were employed to assess the significance of pairwise comparisons between exposed 

and control groups in the analysis of continuous 

variables. Organ and body weight data, which have 

approximately normal distributions, were analyzed 

with the parametric multiple comparison procedures 

of Dunnett (1955) and Williams (1971, 1972). Blood 

and bone marrow hematology data, which have 

typically skewed distributions, were analyzed using 

the nonparametric multiple comparison methods of 

Shirley (1977) and Dunn (1964). Jonckheere’s test 

(Jonckheere, 1954) was used to assess the significance 

of the dose-related trends and to determine whether a 

trend-sensitive test (Williams’ or Shirley’s test) was 

more appropriate for pairwise comparisons than a test 

that does not assume a monotonic dose-related trend 

(Dunnett’s or Dunn’s test). Prior to statistical analysis, extreme values identified by the outlier test of 

Dixon and Massey (1951) were examined by NTP 

personnel, and implausible values were eliminated 

from the analysis. Average severity values were 

analyzed for significance with the Mann-Whitney U 

test (Hollander and Wolfe, 1973). Treatment effects 

were investigated by applying a multivariate analysis 

of variance (Morrison, 1976) to the transformed data 

to test for simultaneous equality of measurements 

across exposure concentrations. 

Historical Control Data 

Although the concurrent control group is always the 

first and most appropriate control group used for 

evaluation, historical control data can be helpful in the 

overall assessment of neoplasm incidence in certain 

instances. Consequently, neoplasm incidences from 

the NTP historical control database, which is updated 
 

 

 

2-Butoxyethanol, NTP TR 484 41 

yearly, are included in the NTP reports for neoplasms 

appearing to show compound-related effects. 

QUALITY ASSURANCE METHODS 

The 14-week and 2-year studies were conducted in 

compliance with Food and Drug Administration Good 

Laboratory Practice Regulations (21 CFR, Part 58). 

In addition, as records from the 2-year studies were 

submitted to the NTP Archives, these studies were 

audited retrospectively by an independent quality 

assurance contractor. Separate audits covered completeness and accuracy of the pathology data, 

pathology specimens, final pathology tables, and a 

draft of this NTP Technical Report. Audit procedures 

and findings are presented in the reports and are on 

file at NIEHS. The audit findings were reviewed and 

assessed by NTP staff, and all comments were resolved or otherwise addressed during the preparation 

of this Technical Report. 

GENETIC TOXICOLOGY 

The genetic toxicity of 2-butoxyethanol was assessed 

by testing the ability of the chemical to induce 

mutations in various strains of Salmonella 

typhimurium, sister chromatid exchanges and chromosomal aberrations in cultured Chinese hamster ovary 

cells, and increases in the frequency of micronucleated erythrocytes in bone marrow of male rats 

and mice. The protocols for these studies and the 

results are given in Appendix E. 

The genetic toxicity studies of 2-butoxyethanol are 

part of a larger effort by the NTP to develop a 

comprehensive database that would permit a critical 

anticipation of a chemical’s carcinogenicity in experimental animals based on numerous considerations, 

including the molecular structure of the chemical and 

its observed effects in short-term in vitro and in vivo 

genetic toxicity tests (structure-activity relationships). 

These short-term genetic toxicity tests were originally 

developed to clarify mechanisms of chemical-induced 

DNA damage growing out of the earlier 

electrophilicity/mutagenicity relationship proposed by 

(Miller and Miller, 1977) and the somatic mutation 

theory of cancer (Straus, 1981; Crawford, 1985). 

Therefore, the information obtained from these tests 

applies only to mutagenic carcinogens. 

For mutagenic carcinogens, there is a strong 

correlation between a chemical’s potential for DNA 

reactivity, mutagenicity in Salmonella, and carcinogenicity in rodents. The combination of DNA 

reactivity and Salmonella mutagenicity is highly 

correlated with the induction of carcinogenicity in 

multiple species and genders of rodents and at 

multiple tissue sites (Ashby and Tennant, 1991). Data 

from NTP studies show that a positive response in 

Salmonella is the most predictive in vitro test for 

rodent carcinogenicity (89% of the Salmonella mutagens are rodent carcinogens) and that there is no 

complementarity among the in vitro genetic toxicity 

tests (Tennant et al., 1987; Zeiger et al., 1990). That 

is, no battery of tests that included the Salmonella test 

improved the predictivity of the Salmonella test alone. 

Although other in vitro genetic toxicity tests correlate 

less well with rodent carcinogenicity compared with 

the Salmonella test, these other tests can provide 

useful information on the types of DNA and chromosomal effects induced by the chemical under 

investigation. 

The predictivity for carcinogenicity of a positive 

response in in vivo bone marrow chromosome 

aberration or micronucleus tests appears to be less 

than the Salmonella test (Shelby et al., 1993; Shelby 

and Witt, 1995). Positive responses in long-term 

peripheral blood micronucleus tests have not been 

formally evaluated for their predictivity for rodent 

carcinogenicity. However, because of the theoretical 

and observed associations between induced genetic 

damage and adverse effects in somatic and germ cells, 

the determination of in vivo genetic effects is 

important to the overall understanding of the risks 

associated with exposure to a particular chemical. 
42 2-Butoxyethanol, NTP TR 484 
 

 

43 

RESULTS
 

RATS 

14-WEEK STUDY 

Six female rats were killed moribund during the 

study. One female rat in the 250 ppm group was 

killed moribund during week 8; four females in the 

500 ppm group were killed moribund during week 1 

and one during week 5 (Table 2). All other animals 

survived to the end of the study. The final mean body 

weights and body weight gains of females exposed to 

500 ppm were significantly less than those of the 

chamber controls. Clinical findings were most prevalent in rats exposed to 125, 250, or 500 ppm and 

included abnormal breathing, pallor, red urine stains, 

TABLE 2 

nasal and eye discharge, lethargy, and increased 

salivation and/or lacrimation. In addition, all females 

exposed to 500 ppm displayed tail lesions consisting 

of alternating bands of dark purplish blue with 

blanched white bands in approximately the distal onethird of the tail. This progressed to self-mutilation 

(chewing off) and/or sloughing of this portion of the 

tail. These findings were most prevalent during the 

first 2 weeks of the study; however, all females 

exposed to 500 ppm lost the distal portion of the tail. 

Survival and Body Weights of Rats in the 14-Week Inhalation Study of 2-Butoxyethanol
 

Mean Body Weightb (g) Final Weight 

Concentration Survivala Initial Final Change Relative to Controls 

(ppm) (%) 

Male 

0 10/10 132 ± 2 355 ± 7 223 ± 7 

31 10/10 128 ± 4 364 ± 6 235 ± 4 103 

62.5 10/10 132 ± 3 367 ± 3 235 ± 3 103 

125 10/10 130 ± 3 346 ± 8 217 ± 8 97 

250 10/10 129 ± 2 349 ± 6 220 ± 5 98 

500 10/10 132 ± 2 357 ± 5 225 ± 5 101 

Female 

0 10/10 112 ± 2 217 ± 5 105 ± 4 

31 10/10 112 ± 2 211 ± 3 99 ± 2 97 

62.5 10/10 111 ± 2 206 ± 4 95 ± 3 95 

125 10/10 111 ± 1 211 ± 4 100 ± 3 97 

250 

500 

9/10c 

5/10d 

111 ± 2 

110 ± 2 

210 ± 7 

197 ± 4* 

99 ± 5 

89 ± 3* 

97 

91 

*	 Significantly different (P#0.05) from the chamber control group by Williams’ or Dunnett’s test a Number of animals surviving at 14 weeks/number initially in group b	 Weights and weight changes are given as mean ± standard error. Subsequent calculations are based on animals surviving to the end of 

the study. 

Week of death: 8 

d Week of death: 1, 1, 1, 1, 5 

c 
 

44 2-Butoxyethanol, NTP TR 484 

The hematology data are listed in Tables 3 and F1. 

An exposure concentration-related anemia, evidenced 

by decreases in automated and manual hematocrit 

values, hemoglobin concentrations, and erythrocyte 

counts, occurred in the 125 ppm or greater males and 

all exposed groups of females. Females appeared to 

be slightly more sensitive to 2-butoxyethanol than 

males. Besides the anemia in all exposed female 

groups, the gender difference was evidenced by 

approximately 25% to 35% decreases in hematocrit 

value and hemoglobin concentration and a 44% 

decrease in erythrocyte count for the 500 ppm females 

compared to approximately 20% to 25% decreases in 

hematocrit value and hemoglobin concentration and a 

34% decrease in erythrocyte count for the 500 ppm 

males. The anemia was characterized as macrocytic, 

normochromic, and responsive. Evidence of macrocytosis was demonstrated by increases in the mean 

cell volumes, which occurred in males exposed to 

125 ppm or greater and females exposed to 62.5 ppm 

or greater. Increased mean cell hemoglobin values 

occurred concurrently with the increased mean cell 

volumes. Normochromic erythrocytes were evidenced by the lack of change in the mean cell 

hemoglobin concentrations. Evidence of an erythropoietic response was demonstrated by increases in the 

reticulocyte and nucleated erythrocyte counts which 

occurred in males exposed to 125 ppm or greater and 

females exposed to 62.5 ppm or greater. 

Microscopic evaluation of blood smears of rats in the 

500 ppm groups revealed increased numbers of 

polychromatophilic erythrocytes; occasional microcytes also were observed. Decreases in leukocyte 

counts, characterized by decreased lymphocyte and 

monocyte counts, occurred in males exposed to 

125 ppm or greater. Platelet counts increased in 

females exposed to 125 or 500 ppm. 
 

2-Butoxyethanol, NTP TR 484 45 

TABLE 3 

Selected Hematology Data for Rats in the 14-Week Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 31 ppm 62.5 ppm 125 ppm 250 ppm 500 ppm 

Male 

n 10 10 10 10 10 10 

Automated hematocrit

 (mL/dL) 45.5 ± 0.4 43.8 ± 0.7 45.1 ± 0.4 42.7 ± 0.5** 38.4 ± 0.4** 34.9 ± 0.3** 

Manual hematocrit (%) 46.8 ± 0.5 45.8 ± 0.6 47.0 ± 0.4 44.5 ± 0.5** 41.1 ± 0.3** 37.3 ± 0.4** 

Hemoglobin (g/dL) 

Erythrocytes (106

/μL) 

Reticulocytes (106

/μL) 

15.5 ± 0.1 

9.05 ± 0.08 

0.16 ± 0.02 

14.8 ± 0.3 

8.71 ± 0.14* 

0.17 ± 0.03 

15.4 ± 0.1 

8.91 ± 0.06 

0.15 ± 0.02 

14.5 ± 0.2** 

8.01 ± 0.08** 

0.30 ± 0.04** 

13.1 ± 0.1** 

7.10 ± 0.07** 

0.48 ± 0.06** 

11.7 ± 0.1** 

5.97 ± 0.05** 

0.68 ± 0.07** 

Nucleated erythrocytes

 (103

/μL) 0.04 ± 0.02 0.05 ± 0.01 0.04 ± 0.03 0.11 ± 0.03 0.17 ± 0.04** 0.20 ± 0.06* 

Mean cell volume (fL) 50.4 ± 0.3 50.2 ± 0.2 50.7 ± 0.2 53.1 ± 0.2** 53.8 ± 0.3** 58.5 ± 0.3** 

Mean cell hemoglobin

 (pg) 17.1 ± 0.1 17.0 ± 0.1 17.3 ± 0.1 18.1 ± 0.1** 18.4 ± 0.1** 19.5 ± 0.1** 

Mean cell hemoglobin

 concentration (g/dL) 

Leukocytes (103

/μL) 

Lymphocytes (103

/μL) 

Monocytes (103

/μL) 

33.9 ± 0.2 

6.70 ± 0.29 

5.34 ± 0.26 

0.19 ± 0.04 

33.7 ± 0.2 

6.29 ± 0.38 

4.99 ± 0.33 

0.19 ± 0.05 

34.2 ± 0.2 

6.13 ± 0.33 

4.82 ± 0.28 

0.11 ± 0.03 

33.9 ± 0.1 

5.87 ± 0.24* 

4.72 ± 0.22 

0.08 ± 0.02* 

34.1 ± 0.2 

5.72 ± 0.36* 

4.65 ± 0.20* 

0.06 ± 0.02** 

33.4 ± 0.1 

5.34 ± 0.17** 

3.93 ± 0.42** 

0.08 ± 0.04* 

Female 

n 10 10 10 10 9 5 

Automated hematocrit

 (mL/dL) 46.7 ± 0.3 44.7 ± 0.5** 43.6 ± 0.5** 40.5 ± 0.3** 37.4 ± 0.3** 31.9 ± 0.6** 

Manual hematocrit (%) 48.5 ± 0.5 46.0 ± 0.5** 45.2 ± 0.5** 42.9 ± 0.4** 40.0 ± 0.3** 36.2 ± 0.6** 

Hemoglobin (g/dL) 

Erythrocytes (106

/μL) 

Reticulocytes (106

/μL) 

15.6 ± 0.1 

8.48 ± 0.05 

0.13 ± 0.02 

15.0 ± 0.1** 

8.08 ± 0.07** 

0.10 ± 0.01 

14.6 ± 0.1** 

7.70 ± 0.08** 

0.16 ± 0.02 

13.6 ± 0.1** 

6.91 ± 0.05** 

0.26 ± 0.04* 

12.5 ± 0.1** 

6.07 ± 0.04** 

0.34 ± 0.04** 

10.5 ± 0.3** 

4.77 ± 0.15** 

0.40 ± 0.11** 

Nucleated erythrocytes 

 (103

/μL) 0.04 ± 0.02 0.05 ± 0.02 0.12 ± 0.03* 0.18 ± 0.07 0.61 ± 0.24** 0.73 ± 0.27** 

Mean cell volume (fL) 55.1 ± 0.3 55.3 ± 0.2 56.4 ± 0.2** 58.7 ± 0.2** 61.6 ± 0.2** 66.8 ± 0.9** 

Mean cell hemoglobin

 (pg) 18.4 ± 0.1 18.6 ± 0.2 19.0 ± 0.0** 19.6 ± 0.1** 20.6 ± 0.1** 22.0 ± 0.1** 

Mean cell hemoglobin 

 concentration (g/dL) 

Platelets (103

/μL) 

33.4 ± 0.1 

573.5 ± 19.5 

33.6 ± 0.3 

576.1 ± 31.6 

33.6 ± 0.1 

583.5 ± 13.3 

33.6 ± 0.2 

657.0 ± 25.7* 

33.4 ± 0.1 

611.6 ± 25.6 

32.9 ± 0.2 

719.6 ± 52.9* 

* Significantly different (P#0.05) from the chamber control group by Dunn’s or Shirley’s test 

** P#0.01 a Mean ± standard error. Statistical tests were performed on unrounded data. 

Kidney weights of males exposed to 500 ppm and Female rats that were killed moribund during the 

females exposed to 125 ppm or greater and liver study exhibited a number of histopathologic changes 

weights of males exposed to 250 or 500 ppm and (Table 4). Thrombosis occurred in a number of 

females exposed to 125 ppm or greater were sig- tissues in females exposed to 500 ppm. The thromnificantly greater than those of the chamber controls bosis was associated with areas of infarction in the tail 

(Table G1). Thymus weights of females exposed to and necrosis in the incisors and liver. Thrombosis 

500 ppm were significantly less. was present in the atrium of the heart, in the blood 
46 2-Butoxyethanol, NTP TR 484 

TABLE 4 

Incidences of Selected Nonneoplastic Lesions in Female Rats Killed Moribund 

in the 14-Week Inhalation Study of 2-Butoxyethanol 

250 ppm 500 ppm 

Tail, Vertebraea 1 5 

Thrombosisb 0 4 (2.0)c 

Infarct 0 5 (3.0) 

Bone Marrow, Necrosis 0 5 (2.0) 

Femur 1 3 

Thrombosis 0 2 (1.0) 

Bone Marrow, Hyperplasia 1 (2.0) 1 (2.0) 

Tooth, Incisor 1 5 

Dental Pulp, Thrombosis 0 4 (2.0) 

Odontoblast, Degeneration 0 4 (2.0) 

Nasal Cavity 1 5 

Thrombosis 0 3 (2.0) 

Lung 1 5 

Thrombosis 0 3 (1.0) 

Heart, Atrium 1 5 

Thrombosis 0 1 (2.0) 

Forestomach 1 5 

Inflammation 0 3 (1.7) 

Necrosis 0 2 (1.5) 

Ulcer 0 2 (2.0) 

Hyperplasia 0 1 (1.0) 

Liver 1 5 

Necrosis 1 (1.0) 4 (2.0) 

Thrombosis 0 3 (1.0) 

Centrilobular, Degeneration 0 4 (2.0) 

Kupffer Cell, Pigmentation 1 (1.0) 1 (1.0) 

Kidney, Renal Tubule 1 5 

Degeneration 0 4 (2.0) 

Pigmentation 1 (1.0) 2 (1.0) 

Spleen 1 5 

Atrophy 0 1 (2.0) 

Hematopoietic Cell Proliferation 0 2 (1.5) 

Thymus 1 5 

Atrophy 0 4 (2.5) 

a Number of animals killed moribund 

b Number of animals with lesion 

Average severity grade of lesions in affected animals: 1=minimal, 2=mild, 3=moderate, 4=marked c 
 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 47 

vessels of the lamina propria of the nasal septum of 

the most anterior nasal section (Plate 1), in central 

veins of the liver associated with large foci of 

necrosis, in the lung, and in blood vessels of the 

dental pulp (Plates 2 and 3), which was associated 

with focal odontoblast degeneration. In addition, 

thrombi were noted within the metaphyseal vessels of 

the femur and the intramedullary and periosteal blood 

vessels in the vertebrae of the tail. There were also 

areas of necrosis within the marrow. The remaining 

portion of the tails in these females had infarcts in the 

cortical and trabecular bone of the coccygeal 

vertebrae (Plate 4). This was also noted in females 

surviving to the end of the study. Many of the 

osteocytes within the cortical and especially the 

trabecular bone exhibited nuclear pyknosis/ 

fragmentation to complete loss of nuclear staining, 

often appearing as ghost cells. Growth plate and 

articular chondrocytes also exhibited loss of nuclear 

staining consistent with cell death. In the female rat 

that was killed moribund on day 32, there were 

additional changes noted in the coccygeal vertebrae. 

These included necrosis of the marrow (including 

adipocytes) and the presence of growth arrest lines. 

On the growth plate side of the growth arrest lines, 

there was no evidence of necrosis; however, on the 

diaphyseal side of the growth arrest lines, there was 

evidence of widespread marrow and bone infarction. 

The longitudinal growth in the area of infarction was 

reduced or completely stopped and was associated 

with growth plate degenerative changes. Affected 

marrow was infiltrated by macrophages (foreign body 

type inflammation) in response to fat necrosis. 

In the tail vertebrae of female rats in the 500 ppm 

group that were killed at the end of the study, there 

were lesions consistent with prior infarction with 

transient or complete growth arrest. In the most 

severely affected vertebrae, there was growth plate 

degeneration with no evidence of renewed 

longitudinal growth, indicating irreversible growth 

plate injury. These vertebrae exhibited marrow 

necrosis that extended to the growth plate, capping of 

the growth plate with a dense layer of bone, and 

degeneration of the growth plate cartilage. In other 

vertebrae, renewed longitudinal growth was 

evidenced by the presence of growth arrest lines. The 

presence of a single growth arrest line at the end of 

each vertebra is consistent with a single ischemic 

insult, and its presence also confirmed renewed 

longitudinal growth which could only occur if blood 

flow was normalized after the insult. The space 

between the arrest line and growth plate represents the 

period of time between resuming longitudinal growth 

and terminal sacrifice. 

Also observed in rats killed moribund during the 

study were atrophy of the spleen and thymus, characterized by depletion of lymphocytes from the white 

pulp of the spleen and from the cortex of the thymus; 

inflammation, necrosis, ulceration, and hyperplasia of 

the forestomach; centrilobular degeneration of the 

liver characterized by fatty change, presence of 

granular eosinophilic debris, single cell necrosis, and 

some neutrophil infiltration; and renal tubule degeneration in which the affected tubules, located mainly 

in the cortex, had dilated lumens with flattened and/or 

necrotic epithelium. The luminal contents of the 

tubules stained intensely red and in some tubules 

resembled crystalline hemoglobin. Renal changes are 

consistent with a hemoglobinuric nephrosis. 

The histopathologic lesions noted in rats at terminal 

sacrifice were similar between males and females and 

were consistent with hemolytic anemia and hemoglobinuria (Table 5). Many of these effects were the 

same as those observed in females killed moribund 

during the study. Minimal hematopoietic cell proliferation of the spleen, primarily erythroid, was noted 

in female rats exposed to 62.5 ppm or greater and in 

male rats exposed to 250 ppm or greater compared to 

the chamber controls. The incidences of bone marrow hyperplasia in males exposed to 250 ppm or 

greater and in females exposed to 62.5 ppm or greater 

were significantly increased. An exposure 

concentration-related increase in the incidence of 

pigmentation of the hepatic Kupffer cells was noted in 

males and females exposed to 125 ppm or greater and 

in the females exposed to 62.5 ppm. In addition, 

there was a concentration-related deposition of pigment in the renal cortical tubules in males and females 

exposed to 250 or 500 ppm and in 125 ppm females. 

Liver and kidney pigmentation stained positive for 

Perls’ Prussian blue stain for iron, which is consistent 

with hemosiderin. Minimal forestomach inflammation 

and epithelial hyperplasia were noted in male rats 

exposed to 250 or 500 ppm. In addition, one female 

exposed to 250 ppm and one female exposed to 

500 ppm had epithelial hyperplasia of the 

forestomach. 
 

48 2-Butoxyethanol, NTP TR 484 

TABLE 5 

Incidences of Selected Nonneoplastic Lesions in Rats at Terminal Sacrifice 

in the 14-Week Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31 ppm 62.5 ppm 125 ppm 250 ppm 500 ppm 

Male 

Livera 10 10 10 10 10 10 

Kupffer Cell, Pigmentationb 0 0 0 7** (1.0)c 10** (1.0) 10** (1.0) 

Bone Marrow 10 10 10 10 10 10 

Hyperplasia 0 0 0 0 10** (1.7) 10** (2.7) 

Spleen 10 10 10 10 10 10 

Hematopoietic Cell Proliferation 0 1 (1.0) 0 0 10** (1.0) 10** (1.0) 

Kidney 10 10 10 10 10 10 

Renal Tubule, Pigmentation 0 0 0 0 8** (1.0) 10** (1.0) 

Forestomach 10 10 10 10 10 10 

Inflammation 0 0 0 0 2 (1.0) 2 (1.0) 

Epithelium, Hyperplasia 0 0 0 0 3 (1.7) 2 (1.0) 

Female 

Tail, Vertebrae 10 10 10 10 9 5 

Infarct 0 0 0 0 0 5** (3.0) 

Bone Marrow, Necrosis 0 0 0 0 0 5** (2.0) 

Liver 10 10 10 10 9 5 

Kupffer Cell, Pigmentation 0 0 10** (1.0) 10** (1.0) 9** (1.0) 5** (1.0) 

Bone Marrow 10 10 10 10 9 5 

Hyperplasia 0 0 8** (1.0) 10** (2.0) 9** (2.5) 5** (3.0) 

Spleen 10 10 10 10 9 5 

Hematopoietic Cell Proliferation 0 0 1 (1.0) 10** (1.0) 8** (1.0) 5** (1.4) 

Kidney 10 10 10 10 9 5 

Renal Tubule, Pigmentation 0 0 0 10** (1.0) 9** (1.0) 5** (1.0) 

Forestomach 10 10 10 10 9 5 

Epithelium, Hyperplasia 0 0 0 0 1 (1.0) 1 (1.0) 

** Significantly different (P#0.01) from the chamber control group by the Fisher exact test a Number of animals examined microscopically b Number of animals with lesion 

Average severity grade of lesions in affected animals: 1=minimal, 2=mild, 3=moderate, 4=marked 

Exposure Concentration Selection Rationale: The erythrocyte cell indices was less than 10% from the
 

hematologic effects in rats exposed to 125 ppm chamber controls. Based on the hematologic effects,
 

were not considered sufficiently severe to preclude the 2-butoxyethanol exposure concentrations selected for
 

use of this concentration in a 2-year study. In the 2-year inhalation study in rats were 31.2, 62.5,
 

addition, 62.5 ppm was a no-effect concentration for and 125 ppm.
 

male rats, and in females the overall change in
 

c 
 

2-Butoxyethanol, NTP TR 484 49 

2-YEAR STUDY 

Survival	 Body Weights and Clinical Findings 

Estimates of 2-year survival probabilities for male and Mean body weights of all exposed groups of males 

female rats are shown in Table 6 and in the and of females exposed to 31.2 or 62.5 ppm were 

Kaplan-Meier survival curves (Figure 2). Survival of generally similar to those of the chamber controls 

exposed male and female rats was similar to the (Figure 3; Tables 7 and 8). The mean body weights 

chamber control groups. of females exposed to 125 ppm were generally less 

than those of the chamber control groups from 

week 17 until the end of the study. No clinical 

findings were attributed to 2-butoxyethanol exposure. 

TABLE 6 

Survival of Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Male 

Animals initially in study 50 50 50 50 

Moribund 25 31 24 21 

Natural deaths 6 8 5 5 

Animals surviving to study termination 19 11 21 24 

Percent probability of survival at end of studya 

Mean survival (days)b 

38 

660 

22 

650 

42 

654 

48 

669 

Survival analysisc P=0.115N P=0.196 P=0.772N P=0.393N 

Female 

Animals initially in study 50 50 50 50 

Moribund 18 21 23 26 

Natural deaths 3 2 4 3 

Animals surviving to study termination 29 27 23 21 

Percent probability of survival at end of study 58 54 46 42 

Mean survival (days) 688 689 660 678 

Survival analysis P=0.106 P=0.832 P=0.238 P=0.159 

a Kaplan-Meier determinations b	 Mean of all deaths (uncensored, censored, and terminal sacrifice) 

The result of the life table trend test (Tarone, 1975) is in the chamber control column, and the results of the life table pairwise 

comparisons (Cox, 1972) with the chamber controls are in the exposed group columns. A negative trend or lower mortality in an 

exposure group is indicated by N. 

c 
 

50 2-Butoxyethanol, NTP TR 484 

FIGURE 2
 

Kaplan-Meier Survival Curves for Male and Female Rats
 

Exposed to 2-Butoxyethanol by Inhalation for 2 Years
 
2-Butoxyethanol, NTP TR 484 51 

FIGURE 3 


Growth Curves for Male and Female Rats
 

Exposed to 2-Butoxyethanol by Inhalation for 2 Years
 
52 2-Butoxyethanol, NTP TR 484 

TABLE 7 

Mean Body Weights and Survival of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Weeks Chamber Control 31.2 ppm 62.5 ppm 125 ppm 

on Av. Wt. No. of Av. Wt. Wt. (% of No. of Av. Wt. Wt. (% of No. of Av. Wt. Wt. (% of No. of 

Study (g) Survivors (g) controls) Survivors (g) controls) Survivors (g) controls) Survivors 

1 164 50 164 100 50 162 98 50 164 100 50 

5 263 50 263 100 50 261 99 50 261 99 50 

9 308 50 317 103 50 314 102 50 316 103 50 

13 349 50 359 103 50 353 101 50 349 100 50 

17 384 50 389 102 50 387 101 50 377 98 50 

21 399 50 409 102 50 408 102 50 399 100 50 

25 414 50 422 102 50 418 101 50 413 100 50 

29 431 50 438 102 50 435 101 50 428 99 50 

33 446 50 455 102 50 452 102 50 441 99 50 

37 464 50 472 102 50 470 101 50 459 99 50 

41 468 50 479 102 50 481 103 50 465 99 50 

45 477 50 492 103 50 494 104 50 478 100 50 

49 484 50 501 104 50 498 103 50 483 100 50 

53 492 49 509 103 50 508 103 49 489 99 50 

57 502 49 518 103 50 517 103 47 493 98 50 

61 503 49 516 103 50 520 103 47 498 99 50 

65 500 49 514 103 50 519 104 47 489 98 50 

69 499 49 506 101 49 520 104 45 489 98 49 

73 502 49 518 103 48 529 105 44 495 99 47 

77 508 46 511 101 47 525 104 44 494 97 44 

81 504 44 516 102 43 522 104 42 496 99 43 

85 511 40 521 102 39 531 104 39 503 99 41 

89 506 37 516 102 37 524 104 38 497 98 38 

92 494 35 495 100 31 515 104 35 478 97 37 

94 510 28 496 97 27 516 101 34 487 96 35 

96 505 28 501 99 23 510 101 33 488 97 32 

99 489 26 488 100 20 507 104 29 484 99 31 

100 493 24 478 97 18 508 103 28 482 98 30 

102 492 22 476 97 15 505 103 25 481 98 27 

104 497 19 452 91 14 507 102 21 480 97 25 

Mean for weeks 

1-13 271 276 102 273 101 273 101 

14-52 441 451 102 449 102 438 99 

53-104 500 502 100 517 103 490 98 
2-Butoxyethanol, NTP TR 484 53 

TABLE 8 

Mean Body Weights and Survival of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Weeks Chamber Control 31.2 ppm 62.5 ppm 125 ppm 

on Av. Wt. No. of Av. Wt. Wt. (% of No. of Av. Wt. Wt. (% of No. of Av. Wt. Wt. (% of No. of 

Study (g) Survivors (g) controls) Survivors (g) controls) Survivors (g) controls) Survivors 

1 120 50 119 99 50 118 99 50 119 99 50 

5 164 50 164 100 50 163 100 50 162 99 50 

9 182 50 181 99 50 182 100 50 177 97 50 

13 194 50 193 100 50 194 100 50 189 98 50 

17 210 50 209 99 50 207 98 50 201 95 50 

21 218 50 218 100 50 216 99 50 207 95 50 

25 227 50 228 101 50 224 99 50 216 95 50 

29 239 50 237 99 50 234 98 50 222 93 50 

33 243 50 245 101 50 241 99 50 228 94 50 

37 253 50 257 102 50 250 99 49 237 94 50 

41 264 50 267 101 50 261 99 49 244 93 50 

45 276 50 278 101 50 273 99 49 258 94 50 

49 290 50 293 101 50 286 99 49 268 92 50 

53 300 50 301 101 50 294 98 49 273 91 50 

57 312 50 316 101 50 308 99 48 284 91 50 

61 323 49 327 101 50 322 100 48 291 90 50 

65 321 49 329 102 50 322 100 48 291 91 50 

69 325 49 330 102 50 322 99 46 297 92 48 

73 335 49 341 102 49 335 100 45 309 92 48 

77 342 48 346 101 49 337 99 44 313 91 48 

81 343 45 342 100 47 340 99 42 317 92 46 

85 350 44 356 102 44 345 99 39 321 92 45 

89 353 43 360 102 42 350 99 38 324 92 42 

92 359 41 364 102 41 352 98 36 322 90 40 

94 362 39 363 101 40 352 97 34 321 89 38 

96 361 37 373 103 37 353 98 33 325 90 34 

99 354 37 373 106 35 352 100 32 321 91 31 

100 355 35 373 105 33 351 99 29 323 91 28 

102 354 35 376 106 31 351 99 26 328 93 22 

104 367 30 368 100 30 353 96 25 331 90 22 

Mean for weeks 

1-13 165 164 99 164 99 162 98 

14-52 247 248 100 244 99 231 94 

53-104 342 349 102 338 99 311 91 
 

 

 

54 2-Butoxyethanol, NTP TR 484 

Hematology and Bone Marrow Cellularity 

The hematology data for rats are listed in Tables 9 

and F2. As in the 14-week study, inhalation of 

2-butoxyethanol by rats in the 2-year study resulted in 

the development of a persistent and exposure-related 

macrocytic, normochromic, responsive anemia. The 

anemia, evidenced by decreases in automated and 

manual hematocrit values, hemoglobin concentrations, 

and erythrocyte counts, occurred at 3, 6, and 

12 months in 62.5 ppm females and 125 ppm males 

and females. An anemia also occurred in 31.2 ppm 

females at 3 and 6 months, and there was evidence of 

an anemia in 62.5 ppm males at 12 months. Evidence 

of macrocytosis was demonstrated by increases in the 

mean cell volumes, accompanied by elevations in the 

mean cell hemoglobin values. The observed increases 

in reticulocyte and nucleated erythrocyte counts in 

males and/or females would be consistent with an 

erythropoietic response to the anemia. Increases in 

bone marrow cellularity occurred in 125 ppm females 

at all time points. Microscopic examination of the 

bone marrow preparations revealed approximate 15% 

to 35% decreases in the myeloid/erythroid (M/E) ratio 

for 125 ppm males and females during the study. 

Females exposed to 62.5 ppm generally had reduced 

M/E ratios of 10% to 30% during the study. Cytologically, morphologic alterations were observed and 

megakaryocytes were present in all exposed groups. 
2-Butoxyethanol, NTP TR 484 55 

TABLE 9 

Selected Hematology and Bone Marrow Cellularity Data for Rats at 3, 6, and 12 Months 

in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Male 

Hematology 

n 

3 Months 9 9 9 9 

6 Months 9 8 9 8 

12 Months 8 0b 9 9 

Automated hematocrit (mL/dL) 

3 Months 46.2 ± 0.3 48.5 ± 0.4 46.1 ± 0.4 43.5 ± 0.3** 

6 Months 47.1 ± 0.3 46.5 ± 0.6 47.4 ± 0.7 44.3 ± 0.4** 

12 Months 46.1 ± 0.3 — 44.4 ± 0.7** 41.4 ± 1.1** 

Manual hematocrit (%) 

3 Months 44.9 ± 0.2 46.9 ± 0.5 44.8 ± 0.4 42.9 ± 0.5* 

6 Months 47.2 ± 0.2 46.4 ± 0.5 47.2 ± 0.6 44.3 ± 0.7** 

12 Months 47.8 ± 0.4 — 45.9 ± 0.8* 42.9 ± 1.2** 

Hemoglobin (g/dL) 

3 Months 15.0 ± 0.1 15.5 ± 0.1 15.0 ± 0.1 14.2 ± 0.1** 

6 Months 15.2 ± 0.1 15.1 ± 0.2 15.3 ± 0.2 14.4 ± 0.1** 

12 Months 15.2 ± 0.0 — 14.7 ± 0.2** 13.4 ± 0.3** 

Erythrocytes (106

/μL) 

3 Months 8.99 ± 0.06 9.19 ± 0.06 8.84 ± 0.09 8.01 ± 0.06** 

6 Months 9.02 ± 0.08 8.85 ± 0.10 9.05 ± 0.14 8.16 ± 0.07** 

12 Months 8.88 ± 0.08 — 8.39 ± 0.15** 7.43 ± 0.20** 

Reticulocytes (106

/μL) 

3 Months 0.12 ± 0.02 0.14 ± 0.01 0.14 ± 0.02 0.20 ± 0.01** 

6 Months 0.08 ± 0.01 0.09 ± 0.01 0.08 ± 0.01 0.14 ± 0.01** 

12 Months 0.11 ± 0.03 — 0.13 ± 0.02 0.19 ± 0.03 

Nucleated erythrocytes (103

/μL) 

3 Months 0.01 ± 0.01 0.04 ± 0.03 0.02 ± 0.02 0.05 ± 0.02 

6 Months 0.05 ± 0.02 0.00 ± 0.00* 0.01 ± 0.01 0.02 ± 0.01 

12 Months 0.05 ± 0.02 — 0.15 ± 0.10 0.04 ± 0.03 

Mean cell volume (fL) 

3 Months 51.3 ± 0.3 52.8 ± 0.1** 52.0 ± 0.2* 54.2 ± 0.3** 

6 Months 52.3 ± 0.5 52.5 ± 0.2 52.4 ± 0.2 54.4 ± 0.2** 

12 Months 52.0 ± 0.2 — 52.9 ± 0.3* 55.8 ± 0.2** 

Mean cell hemoglobin (pg) 

3 Months 16.7 ± 0.1 16.9 ± 0.0 16.9 ± 0.1 17.7 ± 0.0** 

6 Months 16.9 ± 0.1 17.1 ± 0.1* 16.9 ± 0.1 17.6 ± 0.1** 

12 Months 17.1 ± 0.2 — 17.5 ± 0.1 18.1 ± 0.1** 

Mean cell hemoglobin concentration (g/dL) 

3 Months 32.5 ± 0.1 32.0 ± 0.1* 32.4 ± 0.1 32.6 ± 0.1 

6 Months 32.3 ± 0.3 32.5 ± 0.1 32.3 ± 0.1 32.4 ± 0.1 

12 Months 33.0 ± 0.2 — 33.1 ± 0.2 32.5 ± 0.2 
56 2-Butoxyethanol, NTP TR 484 

TABLE 9 

Selected Hematology and Bone Marrow Cellularity Data for Rats at 3, 6, and 12 Months 

in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Male (continued) 

Bone Marrow Cellularity 

n 

3 Months 9 0 9 9 

6 Months 9 9 9 9 

12 Months 9 0 9 9 

Nucleated bone marrow cells (106

/femur) 

3 Months 82.9 ± 3.1 — 82.5 ± 2.6 88.8 ± 3.7 

6 Months 103.0 ± 3.0 102.8 ± 7.4 104.3 ± 4.7 110.6 ± 3.4 

12 Months 111.5 ± 6.4 — 102.6 ± 3.3 128.1 ± 3.1 

Myeloid/erythroid ratio 

3 Months 1.232 ± 0.056 — 1.063 ± 0.059 1.023 ± 0.062 

6 Months 0.958 ± 0.042 1.122 ± 0.076 1.118 ± 0.101 0.826 ± 0.045 

12 Months 1.107 ± 0.126 — 1.042 ± 0.058 0.720 ± 0.048** 

Female 

Hematology 

n 

3 Months 8 9 9 9 

6 Months 9 9 9 9 

12 Months 9 0 8 9 

Automated hematocrit (mL/dL) 

3 Months 48.9 ± 0.2 47.3 ± 0.5** 44.9 ± 0.5** 43.0 ± 0.4** 

6 Months 46.0 ± 0.4 41.1 ± 1.5* 42.2 ± 0.9** 40.0 ± 0.3** 

12 Months 44.2 ± 0.3 — 43.7 ± 0.3 40.7 ± 0.4** 

Manual hematocrit (%) 

3 Months 46.5 ± 0.5 46.1 ± 0.5 43.3 ± 0.5** 42.2 ± 0.5** 

6 Months 45.8 ± 0.4 41.9 ± 1.4* 43.1 ± 0.9** 41.4 ± 0.3** 

12 Months 45.4 ± 0.2 — 45.3 ± 0.3 42.3 ± 0.4** 

Hemoglobin (g/dL) 

3 Months 15.5 ± 0.1 14.8 ± 0.2** 14.3 ± 0.2** 13.7 ± 0.1** 

6 Months 15.2 ± 0.1 13.7 ± 0.5** 13.9 ± 0.3** 13.2 ± 0.1** 

12 Months 14.9 ± 0.1 — 14.6 ± 0.1 13.5 ± 0.1** 

Erythrocytes (106

/μL) 

3 Months 8.52 ± 0.03 8.10 ± 0.10** 7.54 ± 0.08** 7.08 ± 0.05** 

6 Months 8.40 ± 0.07 7.50 ± 0.25** 7.54 ± 0.15** 6.89 ± 0.05** 

12 Months 7.81 ± 0.05 — 7.42 ± 0.06** 6.75 ± 0.05** 

Reticulocytes (106

/μL) 

3 Months 0.13 ± 0.01 0.16 ± 0.02 0.18 ± 0.02 0.20 ± 0.02* 

6 Months 0.06 ± 0.01 0.08 ± 0.01 0.12 ± 0.01** 0.17 ± 0.01** 

12 Months 0.06 ± 0.01 — 0.11 ± 0.02* 0.12 ± 0.02** 

Nucleated erythrocytes (103

/μL) 

3 Months 0.10 ± 0.03 0.18 ± 0.02 0.09 ± 0.03 0.26 ± 0.05* 

6 Months 0.05 ± 0.03 0.02 ± 0.02 0.03 ± 0.01 0.04 ± 0.02 

12 Months 0.07 ± 0.02 — 0.05 ± 0.02 0.25 ± 0.08* 

Mean cell volume (fL) 

3 Months 57.4 ± 0.2 58.3 ± 0.2** 59.6 ± 0.4** 60.7 ± 0.4** 

6 Months 54.8 ± 0.3 54.8 ± 0.4 56.0 ± 0.3* 58.2 ± 0.2** 

12 Months 56.8 ± 0.2 — 58.8 ± 0.3** 60.3 ± 0.3** 
2-Butoxyethanol, NTP TR 484 57 

TABLE 9 

Selected Hematology and Bone Marrow Cellularity Data for Rats at 3, 6, and 12 Months 

in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Female (continued) 

Hematology (continued) 

n 

3 Months 8 9 9 9 

6 Months 9 9 9 9 

12 Months 9 0 8 9 

Mean cell hemoglobin (pg) 

3 Months 18.2 ± 0.0 18.3 ± 0.1 18.9 ± 0.1** 19.3 ± 0.1** 

6 Months 18.1 ± 0.1 18.3 ± 0.1 18.4 ± 0.1 19.2 ± 0.1** 

12 Months 19.1 ± 0.1 — 19.7 ± 0.1** 20.0 ± 0.1** 

Mean cell hemoglobin concentration (g/dL) 

3 Months 31.7 ± 0.1 31.3 ± 0.2 31.8 ± 0.2 31.8 ± 0.2 

6 Months 33.2 ± 0.2 33.4 ± 0.2 32.9 ± 0.1 33.1 ± 0.1 

12 Months 33.7 ± 0.2 — 33.6 ± 0.2 33.2 ± 0.2 

Bone Marrow Cellularity 

n 

3 Months 8 0 9 9 

6 Months 9 9 9 9 

12 Months 9 0 8 9 

Nucleated bone marrow cells (106

/femur) 

3 Months 62.8 ± 4.4 — 68.4 ± 3.7 84.0 ± 2.0** 

6 Months 63.6 ± 2.3 64.8 ± 4.8 64.0 ± 4.5 89.7 ± 2.4** 

12 Months 74.3 ± 4.8 — 81.8 ± 4.4 110.1 ± 5.6** 

Myeloid/erythroid ratio 

3 Months 1.060 ± 0.084 — 0.840 ± 0.036 0.744 ± 0.044** 

6 Months 1.137 ± 0.061 1.037 ± 0.066 0.788 ± 0.040** 0.708 ± 0.025** 

12 Months 0.846 ± 0.047 — 0.766 ± 0.031 0.690 ± 0.046* 

* Significantly different (P#0.05) from the chamber control group by Dunn’s or Shirley’s test 

** P#0.01 a Mean ± standard error. Statistical tests were performed on unrounded data. b Not examined at this exposure concentration 
 

 

58 2-Butoxyethanol, NTP TR 484 

Pathology and Statistical Analyses 

This section describes the statistically significant or 

biologically noteworthy changes in the incidences of 

neoplasms and/or nonneoplastic lesions of the adrenal 

medulla, nose, liver, and spleen. Summaries of the 

incidences of neoplasms and nonneoplastic lesions, 

individual animal tumor diagnoses, statistical analyses 

of primary neoplasms that occurred with an incidence 

of at least 5% in at least one animal group, and 

historical incidences for the neoplasms mentioned in 

this section are presented in Appendix A for male rats 

and Appendix B for female rats. 

Adrenal Medulla: The incidences of benign or malignant pheochromocytoma (combined) occurred with a 

positive trend in females; however, the incidence in 

females exposed to 125 ppm was not significantly 

increased relative to the chamber controls (Tables 10 

and B3) but exceeded the range for historical controls 

from 2-year inhalation studies (Tables 10 and B4). 

One pheochromocytoma in the 125 ppm female group 

was malignant and another, while benign, was bilateral (Tables 10 and B1). The incidence of medullary hyperplasia was slightly, although not significantly, greater in females in the 125 ppm group than 

in the chamber controls (Tables 10 and B5). 

The primary criterion used to distinguish pheochromocytoma from medullary hyperplasia was the 

presence of mild to moderate compression of the 

adjacent tissue. Most of the pheochromocytomas 

were small and not substantially larger than the more 

severe grades of adrenal medullary hyperplasia. 

Nose: Incidences of hyaline degeneration of the 

olfactory epithelium were significantly increased in all 

exposed groups of males (chamber control, 13/48; 

31.2 ppm, 21/49; 62.5 ppm, 23/49; 125 ppm, 40/50; 

Table A4) and in females exposed to 62.5 or 125 ppm 

(13/50, 18/48, 28/50, 40/49; Table B5); the severity 

of this lesion was not affected by exposure. This 

change has been shown to occur in control and 

exposed animals (Morgan and Harkema, 1996) and is 

considered to be the most common age-related change 

in the nasal passages of rats (St. Clair and Morgan, 

1992). In exposure-related cases, this change has 

been proposed to have an adaptive/protective role 

(Buckley et al., 1985). This lesion consisted of 

intracytoplasmic accumulation of homogeneous 

eosinophilic material, either unilaterally or involving 

both sides of the nose. The change was of minimal 

severity and was generally confined to the olfactory 

epithelium lining the dorsal meatus of level II, 

although in more severe cases it was also present in 

the olfactory epithelium of the ethmoid turbinates in 

level III. Two neoplasms in the nose were observed 

in this study: a chondroma in a 31.2 ppm male and 

an adenoma in a 62.5 ppm male (Table A1). Due to 

the sporadic occurrence of these neoplasms and the 

lack of any preneoplastic change in the nasal 

epithelium, they are considered to be incidental and 

likely not related to 2-butoxyethanol exposure. 

Liver: The incidences of Kupffer cell pigmentation 

were significantly increased relative to the chamber 

controls in males (23/50, 30/50, 34/50, 42/50; 

Table A4) and females (15/50, 19/50, 36/50, 47/50; 

Table B5) exposed to 62.5 or 125 ppm; the severities 

of this lesion were increased in the 125 ppm groups 

(males: 1.3, 1.5, 1.5, 2.0; females: 1.4, 1.5, 1.4, 

2.0). 

Spleen: Incidences of splenic fibrosis were significantly increased relative to the chamber controls in 

males exposed to 62.5 or 125 ppm (11/50, 14/50, 

19/50, 20/50; Table A4). 
 

59 

c 

2-Butoxyethanol, NTP TR 484 

TABLE 10 

Incidences of Neoplasms and Nonneoplastic Lesions of the Adrenal Medulla in Female Rats 

in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Number Examined Microscopically 

Hyperplasiaa 50 11 (1.9)b 

50 

11 (2.3) 

49 

8 (2.1) 

49 

17 (2.5) 

Benign Pheochromocytoma, Bilateral 0 0 0 1 

Benign Pheochromocytoma (includes bilateral)c 

Overall rated 3/50 (6%) 4/50 (8%) 1/49 (2%) 7/49 (14%) 

Adjusted ratee 

Terminal ratef 

6.9% 

1/29 (3%) 

9.2% 

1/27 (4%) 

2.6% 

1/22 (5%) 

16.7% 

1/21 (5%) 

First incidence (days) 554 584 730 (T) 638 

Poly-3 testg P=0.090 P=0.499 P=0.353N P=0.138 

Malignant Pheochromocytoma 0 0 0 1 

Benign or Malignant Pheochromocytomah 

Overall rate 3/50 (6%) 4/50 (8%) 1/49 (2%) 8/49 (16%) 

Adjusted rate 6.9% 9.2% 2.6% 18.9% 

Terminal rate 1/29 (3%) 1/27 (4%) 1/22 (5%) 1/21 (5%) 

First incidence (days) 554 584 730 (T) 612 

Poly-3 test P=0.044 P=0.499 P=0.353N P=0.086 

(T)Terminal sacrifice 

a	 Number of animals with lesion 

b	 Average severity grade of lesions in affected animals: 1=minimal, 2=mild, 3=moderate, 4=marked 

Historical incidence for 2-year inhalation studies with chamber control groups (mean ± standard deviation): 47/889 (5.3% ± 3.9%); 

range, 0%-13% d	 Number of animals with neoplasm per number of animals with adrenal gland examined microscopically e Poly-3 estimated neoplasm incidence after adjustment for intercurrent mortality f Observed incidence at terminal kill 

g	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the 

P values corresponding to pairwise comparisons between the chamber controls and that exposed group. The Poly-3 test accounts for 

differential mortality in animals that do not reach terminal sacrifice. A lower incidence in an exposure group is indicated by N. h	 Historical incidence: 57/889 (6.4% ± 3.5%); range, 2%-13% (also includes complex and unspecified pheochromocytoma) 
 

 

60 

c 

2-Butoxyethanol, NTP TR 484 

MICE 

14-WEEK STUDY 

Two male and two female mice exposed to 500 ppm were significantly less than those of the chamber 

died and two male and two female mice were killed controls. Clinical findings were observed only in 

moribund during the first 2 weeks of the study; all males and females exposed to 500 ppm that died or 

other mice survived until the end of the study were killed moribund and included abnormal 

(Table 11). The final mean body weights and body breathing, red urine stains, and lethargy. 

weight gains of 125, 250, and 500 ppm male mice 

TABLE 11 

Survival and Body Weights of Mice in the 14-Week Inhalation Study of 2-Butoxyethanol 

Mean Body Weightb (g) Final Weight 

Concentration Survivala Initial Final Change Relative to Controls 

(ppm) (%) 

Male 

0 10/10 24.6 ± 0.3 36.4 ± 0.8 11.8 ± 0.7 

31 10/10 24.6 ± 0.2 35.7 ± 0.6 11.1 ± 0.4 98 

62.5 10/10 24.8 ± 0.3 36.2 ± 0.6 11.4 ± 0.5 99 

125 10/10 24.7 ± 0.4 34.3 ± 0.7* 9.6 ± 0.5* 94 

250 10/10 24.3 ± 0.3 34.2 ± 0.5* 10.0 ± 0.6* 94 

500 6/10c 24.3 ± 0.2 32.0 ± 0.5** 7.7 ± 0.6** 88 

Female 

0 10/10 19.3 ± 0.2 29.3 ± 0.6 10.0 ± 0.5 

31 10/10 19.6 ± 0.2 29.7 ± 0.8 10.1 ± 0.8 101 

62.5 10/10 19.6 ± 0.2 30.2 ± 0.9 10.6 ± 0.8 103 

125 10/10 19.5 ± 0.2 29.3 ± 0.6 9.8 ± 0.6 100 

250 10/10 19.7 ± 0.2 30.2 ± 0.4 10.5 ± 0.5 103 

500 6/10c 19.4 ± 0.4 28.3 ± 0.6 8.5 ± 0.6 97 

* Significantly different (P#0.05) from the chamber control group by Williams’ or Dunnett’s test 

** P#0.01 a Number of animals surviving at 14 weeks/number initially in group b Weights and weight changes are given as mean ± standard error. Subsequent calculations are based on animals surviving to the end of 

the study. 

Week of death: 1, 1, 1, 2 

The hematology data are listed in Tables 12 and F3. 2-butoxyethanol. As in the rats, the anemia in mice 

Similar to the rats, an exposure-concentration-related was responsive, evidenced by increased reticulocyte 

anemia, evidenced by decreases in automated and counts; however, unlike the rats, the morphologic 

manual hematocrit values, hemoglobin concentrations, classification was normocytic and normochromic. 

and erythrocyte counts, occurred in males exposed to Normocytic and normochromic erythrocytes were 

125 ppm or greater and in all exposed groups of demonstrated by the lack of change in the mean cell 

females. Because the anemia occurred in all exposed volumes and mean cell hemoglobin concentrations, 

female groups and was slightly more pronounced than respectively. Platelet counts increased in 500 ppm 

in the males, females appeared to be more sensitive to males and females and 250 ppm females. 
 

 

2-Butoxyethanol, NTP TR 484 61 

TABLE 12 

Selected Hematology Data for Mice in the 14-Week Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 31 ppm 62.5 ppm 125 ppm 250 ppm 500 ppm 

n 10 10 10 10 10 6 

Male 

Automated hematocrit

 (mL/dL) 47.7 ± 1.0 48.8 ± 0.4 48.0 ± 0.6 47.1 ± 0.5 44.2 ± 0.3** 35.1 ± 1.4** 

Manual hematocrit (%) 47.3 ± 1.0 48.3 ± 0.4 47.6 ± 0.5 46.6 ± 0.4 44.2 ± 0.4** 36.3 ± 1.4** 

Hemoglobin (g/dL) 

Erythrocytes (106

/μL) 

Reticulocytes (106

/μL) 

15.7 ± 0.4 

9.71 ± 0.22 

0.21 ± 0.03 

16.0 ± 0.1 

10.04 ± 0.08 

0.22 ± 0.03 

15.9 ± 0.1 

9.77 ± 0.10 

0.21 ± 0.02 

15.4 ± 0.1** 

9.47 ± 0.06* 

0.32 ± 0.03* 

14.4 ± 0.1** 

8.90 ± 0.07** 

0.45 ± 0.04** 

11.4 ± 0.4** 

7.21 ± 0.23** 

0.79 ± 0.20** 

Mean cell volume (fL) 49.1 ± 0.4 48.5 ± 0.3 49.0 ± 0.4 49.7 ± 0.4 49.8 ± 0.4 48.3 ± 0.9 

Mean cell hemoglobin

 (pg) 16.2 ± 0.1 16.0 ± 0.1 16.2 ± 0.1 16.2 ± 0.0 16.2 ± 0.1 15.8 ± 0.2 

Mean cell hemoglobin

 concentration (g/dL) 

Platelets (103

/μL) 

33.0 ± 0.2 

922.5 ± 29.9 

32.8 ± 0.3 

878.0 ± 22.1 

33.0 ± 0.2 

894.0 ± 23.7 

32.7 ± 0.2 

933.5 ± 30.0 

32.5 ± 0.2 

1,001.3 ± 46.4 

32.5 ± 0.3 

1,176.7 ± 78.2* 

Female 

Automated hematocrit

 (mL/dL) 47.1 ± 0.4 46.6 ± 0.3 46.4 ± 0.3 45.4 ± 0.5* 42.0 ± 0.4** 35.8 ± 0.7** 

Manual hematocrit (%) 46.2 ± 0.3 45.9 ± 0.3 45.8 ± 0.3 45.1 ± 0.2** 42.3 ± 0.4** 37.8 ± 1.0** 

Hemoglobin (g/dL) 

Erythrocytes (106

/μL) 

Reticulocytes (106

/μL) 

15.7 ± 0.1 

9.72 ± 0.05 

0.18 ± 0.02 

15.4 ± 0.1* 

9.55 ± 0.06* 

0.21 ± 0.03 

15.4 ± 0.1* 

9.51 ± 0.06* 

0.19 ± 0.02 

14.8 ± 0.1** 

9.18 ± 0.05** 

0.29 ± 0.02** 

13.7 ± 0.1** 

8.57 ± 0.06** 

0.47 ± 0.04** 

11.6 ± 0.1** 

7.35 ± 0.07** 

1.17 ± 0.28** 

Mean cell volume (fL) 48.3 ± 0.3 48.8 ± 0.2 48.8 ± 0.2 49.5 ± 0.5 49.0 ± 0.3 48.8 ± 1.0 

Mean cell hemoglobin

 (pg) 16.1 ± 0.1 16.0 ± 0.1 16.2 ± 0.1 16.1 ± 0.1 16.0 ± 0.0 15.8 ± 0.1 

Mean cell hemoglobin 

 concentration (g/dL) 

Platelets (103

/μL) 

33.3 ± 0.2 

838.0 ± 19.0 

33.0 ± 0.3 

779.7 ± 29.5 

33.2 ± 0.2 

854.7 ± 18.1 

32.6 ± 0.2 

930.3 ± 44.1 

32.6 ± 0.2 

1,032.1 ± 44.1** 

32.4 ± 0.4* 

1,179.0 ± 75.6** 

* Significantly different (P#0.05) from the chamber control group by Dunn’s or Shirley’s test 

** P#0.01 a Mean ± standard error. Statistical tests were performed on unrounded data. 

Microscopic evaluation of blood smears of mice in the 

500 ppm groups revealed increased numbers of 

polychromatophilic erythrocytes. 

Absolute and relative liver weights of 500 ppm males 

and relative liver weights of 250 ppm males and 

500 ppm females were significantly greater than those 

of the chamber controls (Table G2). Male and female 

mice exposed to 500 ppm that either died or were 

killed moribund during the study exhibited a number 

of histopathologic changes (Table 13). Male mice had 

ulceration and necrosis of the forestomach. Female 

mice had full-wall-thickness forestomach necrosis, 

and one female had an ulcer in the glandular stomach. 

Acute inflammation surrounded the necrotic or 

ulcerative lesions. In addition to the focal inflammation, suppurative inflammation was present in the 

peritoneum of two males and on the mediastinal 

pleura of two males and two females. These lesions 

were considered to be secondary to gastric ulceration 

and/or necrosis with extension of the inflammation 

through the wall or perforation of the wall. 
62 

c 

2-Butoxyethanol, NTP TR 484 

TABLE 13 

Incidences of Selected Nonneoplastic Lesions in Mice that Died or Were Killed Moribund 

in the 14-Week Inhalation Study of 2-Butoxyethanol 

500 ppm 

Male Female 

Forestomacha 4 4 

Inflammationb 3 (2.0)c 3 (2.0) 

Necrosis 2 (2.0) 3 (3.0) 

Ulcer 2 (3.0) 0 

Epithelium, Hyperplasia 1 (1.0) 0 

Glandular Stomach 4 4 

Ulcer 0 1 (2.0) 

Peritoneum 4 4 

Suppurative Inflammation 2 (2.0) 0 

Mediastinum 4 4 

Suppurative Inflammation 2 (2.0) 2 (2.0) 

Spleen 4 4 

Atrophy 4 (2.3) 3 (3.0) 

Thymus 4 4 

Atrophy 2 (3.5) 2 (3.0) 

Lymph Nodes 4 4 

Atrophy 2 (2.0) 2 (2.5) 

Liver 4 4

 Necrosis 0 1 (3.0) 

Kidney 4 4 

Renal Tubule, Degeneration 4 (1.8) 3 (1.3) 

Epididymis 4 — 

Necrosis 1 (1.0) — 

Testis 4 — 

Degeneration 2 (2.0) — 

a Number of animals that died or were killed moribund 

b Number of animals with lesion 

Average severity grade of lesions in affected animals: 1=minimal, 2=mild, 3=moderate, 4=marked 

In addition, lymphoid atrophy of the spleen, thymus, 

and mesenteric and mandibular lymph nodes occurred 

in males and females. Renal cortical degeneration 

and some necrosis were noted and were characterized 

by glandular eosinophilic debris in the lumen of the 

cortical tubules and pyknotic nuclei. There were also 

testicular degeneration and necrosis of the epididymis 

in male mice. 

The types of lesions noted in mice at terminal 

sacrifice were similar between males and females 

(Table 14). Epithelial hyperplasia and inflammation 

of the muscularis or serosa of the forestomach 

occurred in females exposed to 125 ppm or greater. 

The minimal to mild forestomach inflammation 

consisted of focal infiltration of mixed mononuclear 

cells. Two male mice exposed to 500 ppm had 
 

63 

c 

2-Butoxyethanol, NTP TR 484 

TABLE 14 

Incidences of Selected Nonneoplastic Lesions in Mice at Terminal Sacrifice 

in the 14-Week Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31 ppm 62.5 ppm 125 ppm 250 ppm 500 ppm 

Male 

Spleena 

Hematopoietic Cell Proliferationb 

10 

0 

10 

0 

10 

0 

10 

2 (1.0)c 10 9** 

6 

(1.2)6** (2.0) 

Pigmentation, Hemosiderin 0 0 0 10** (1.0) 10** (1.0) 6** (1.0) 

Liver 10 10 10 10 10 6 

Kupffer Cell, Pigmentation, Hemosiderin 0 0 0 0 0 6** (1.0) 

Forestomach 10 10 9 10 10 6 

Epithelium, Hyperplasia 0 0 0 1 (1.0) 0 2 (1.0) 

Kidney 10 10 10 10 10 6 

Renal Tubule, Pigmentation, Hemosiderin 0 0 0 0 0 5** (1.0) 

Female 

Spleen 10 10 10 10 10 6 

Hematopoietic Cell Proliferation 0 0 0 0 1 (2.0) 6** (2.0) 

Pigmentation, Hemosiderin 0 0 0 0 10** (1.0) 6** (1.0) 

Liver 10 10 10 10 10 6 

Kupffer Cell, Pigmentation, Hemosiderin 0 0 0 0 10** (1.0) 6** (1.0) 

Forestomach 10 10 10 10 10 6 

Inflammation 0 1 (1.0) 0 2 (1.0) 4* (1.0) 4** (1.3) 

Epithelium, Hyperplasia 1 (1.0) 2 (1.5) 5 (1.2) 9** (1.8) 10** (1.7) 4** (2.0) 

Kidney 10 10 10 10 10 6 

Renal Tubule, Pigmentation, Hemosiderin 0 0 0 0 0 6** (1.0) 

* Significantly different (P#0.05) from the chamber control group by the Fisher exact test 

** P#0.01 a Number of animals with organ examined microscopically b Number of animals with lesion 

Average severity grade of lesions in affected animals: 1=minimal, 2=mild, 3=moderate, 4=marked 

epithelial hyperplasia of the forestomach, but the 

inflammatory process observed in females was not 

present. Extramedullary hematopoietic cell 

proliferation, primarily erythroid, and hemosiderin 

pigmentation of the spleen were present in males 

exposed to 125 ppm or greater and in females exposed 

to 250 or 500 ppm. 

Hemosiderin pigmentation in Kupffer cells was the 

only change observed in the livers of males exposed 

to 500 ppm and females exposed to 250 or 500 ppm. 

The incidences of renal tubule hemosiderin pigmentation in males and females exposed to 500 ppm 

were significantly increased. 

Exposure Concentration Selection Rationale: Mortality and significant hematologic effects observed at 

500 ppm precluded the use of this concentration in the 

2-year study; however, the hematologic effects in 

mice exposed to 250 ppm and the minimal inflammatory and hyperplastic lesions of the forestomach in 

female mice exposed to 250 ppm (although only 
64 2-Butoxyethanol, NTP TR 484 

slightly less severe than those in 500 ppm females 

surviving to terminal sacrifice) were not considered 

sufficiently severe to preclude use of this concentration in a 2-year study. In addition, there were 

minimal exposure-related effects in male mice 

exposed to 125 or 250 ppm. Whenever possible in 

inhalation studies, males and females of a species for 

each concentration are housed in the same chamber 

for economic reasons. No space was available for 

mice in the rat chambers. Therefore, the 

2-butoxyethanol concentrations selected for the 2-year 

inhalation study in mice were 62.5, 125, and 

250 ppm. 
2-Butoxyethanol, NTP TR 484 65 

2-YEAR STUDY 

Survival	 Body Weights and Clinical Findings 

Survival of male mice exposed to 125 or 250 ppm was The mean body weights of exposed male mice were 

significantly less than that of the chamber control generally less than those of the chamber control group 

group; survival of all other exposed groups of mice during the last 6 months of the study (Figure 5 and 

was similar to the chamber controls (Table 15 and Table 16). Mean body weights of exposed female 

Figure 4). mice were less than those of the chamber control 

group; the reductions were greater and occurred 

earlier than those observed in males (Figure 5 and 

Table 17). No clinical findings were attributed to 

2-butoxyethanol exposure. 

TABLE 15 

Survival of Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Male 

Animals initially in study	 50 50 50 50 

Moribund 7 8 13 10 

Natural deaths 4 3 10 14 

Animals surviving to study termination 39 39 27 26 

Percent probability of survival at end of studya 78 78 54 52 

Mean survival (days)b 697 713 665 651 

Survival analysisc	 P=0.001 P=1.000N P=0.021 P=0.015 

Female 

Animals initially in study	 50 50 50 50 

Accidental deathd 1 0 0 0 

Moribund 14 16 12 10 

Natural deaths 6 3 5 4 

Animals surviving to study termination 29 31 33 36 

Percent probability of survival at end of study 60 62 66 72 

Mean survival (days) 688 674 667 694 

Survival analysis	 P=0.254N P=1.000N P=0.741N P=0.324N 

a Kaplan-Meier determinations b	 Mean of all deaths (uncensored, censored, and terminal sacrifice) 

The result of the life table trend test (Tarone, 1975) is in the chamber control column, and the results of the life table pairwise 

comparisons (Cox, 1972) with the chamber controls are in the exposed group columns. A negative trend or lower mortality in an 

exposure group is indicated by N. d	 Censored from survival analyses 

c 
 

66 2-Butoxyethanol, NTP TR 484 

FIGURE 4
 

Kaplan-Meier Survival Curves for Male and Female Mice
 

Exposed to 2-Butoxyethanol by Inhalation for 2 Years
 
2-Butoxyethanol, NTP TR 484 67 

FIGURE 5 


Growth Curves for Male and Female Mice
 

Exposed to 2-Butoxyethanol by Inhalation for 2 Years
 
68 2-Butoxyethanol, NTP TR 484 

TABLE 16 

Mean Body Weights and Survival of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Weeks Chamber Control 62.5 ppm 125 ppm 250 ppm 

on Av. Wt. No. of Av. Wt. Wt. (% of No. of Av. Wt. Wt. (% of No. of Av. Wt. Wt. (% of No. of 

Study (g) Survivors (g) controls) Survivors (g) controls) Survivors (g) controls) Survivors 

1 25.2 50 25.5 101 50 25.3 100 50 25.6 102 50 

5 29.3 50 29.1 99 50 28.6 98 50 29.0 99 50 

9 31.4 50 31.9 102 50 31.7 101 50 31.0 99 50 

13 34.4 50 34.3 100 50 33.9 99 50 33.6 98 49 

17 36.3 50 36.5 101 50 35.8 99 50 35.2 97 49 

21 39.2 50 39.3 100 50 38.5 98 50 36.7 94 49 

25 41.0 50 41.2 101 50 40.9 100 50 39.4 96 49 

29 42.4 50 41.8 99 50 41.5 98 50 40.4 95 49 

33 44.1 50 42.5 96 50 42.9 97 50 41.4 94 49 

37 45.8 50 44.5 97 50 44.3 97 50 42.3 92 49 

41 46.6 50 46.0 99 50 45.5 98 50 43.8 94 49 

45 48.0 50 47.9 100 50 47.0 98 50 46.1 96 48 

49 48.3 50 48.6 101 50 48.3 100 50 46.9 97 48 

53 48.4 50 48.4 100 50 48.0 99 50 47.0 97 47 

57 49.3 49 48.3 98 50 48.4 98 50 47.2 96 46 

61 49.9 49 50.0 100 50 50.2 101 49 47.5 95 45 

65 50.2 49 49.7 99 50 51.3 102 48 48.5 97 44 

69 50.2 48 49.5 99 50 50.3 100 47 48.4 96 43 

73 51.1 48 50.1 98 50 50.7 99 46 48.9 96 43 

77 50.5 48 50.1 99 50 50.0 99 45 50.1 99 43 

81 51.0 46 47.5 93 49 49.6 97 43 47.3 93 43 

85 51.5 45 49.9 97 49 50.7 98 38 48.0 93 43 

89 51.8 44 49.3 95 48 50.5 98 35 48.4 93 39 

93 51.5 42 48.2 94 47 49.7 97 34 48.1 93 36 

94 51.1 42 48.3 95 46 49.6 97 33 47.2 92 36 

96 52.0 39 48.2 93 45 48.6 94 32 46.8 90 34 

98 52.1 39 48.5 93 42 49.6 95 30 47.4 91 32 

100 52.1 39 48.7 94 42 50.1 96 29 47.8 92 28 

102 52.2 39 48.0 92 39 49.1 94 27 46.9 90 26 

104 51.2 39 46.8 91 39 47.7 93 27 46.1 90 26 

Mean for weeks 

1-13 30.1 30.2 100 29.9 99 29.8 99 

14-52 43.5 43.1 99 42.7 98 41.4 95 

53-104 50.9 48.8 96 49.7 98 47.7 94 
2-Butoxyethanol, NTP TR 484 69 

TABLE 17 

Mean Body Weights and Survival of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Weeks Chamber Control 62.5 ppm 125 ppm 250 ppm 

on Av. Wt. No. of Av. Wt. Wt. (% of No. of Av. Wt. Wt. (% of No. of Av. Wt. Wt. (% of No. of 

Study (g) Survivors (g) controls) Survivors (g) controls) Survivors (g) controls) Survivors 

1 20.4 50 20.3 100 50 20.1 99 50 19.8 97 50 

5 24.4 50 24.5 100 50 24.6 101 50 24.5 100 50 

9 27.0 50 27.2 101 50 28.0 104 50 27.2 101 50 

13 29.7 50 29.0 98 50 29.6 100 49 29.1 98 50 

17 32.3 50 31.1 96 50 32.0 99 49 30.8 95 50 

21 34.5 50 33.0 96 50 33.5 97 49 32.0 93 50 

25 36.7 50 35.8 98 50 36.8 100 49 35.0 95 50 

29 39.2 50 36.6 93 50 37.8 96 48 35.3 90 50 

33 41.2 50 36.7 89 50 38.4 93 48 35.9 87 50 

37 42.7 50 39.2 92 49 39.3 92 48 36.5 86 50 

41 44.1 50 41.8 95 49 42.1 96 48 38.3 87 50 

45 46.6 50 44.8 96 49 45.1 97 48 41.3 89 50 

49 47.8 50 46.2 97 49 47.1 99 48 42.7 89 50 

53 48.3 50 46.5 96 49 48.5 100 48 43.3 90 50 

57 49.9 49 47.2 95 47 47.6 95 48 43.7 88 50 

61 52.2 49 50.2 96 47 51.1 98 46 43.6 84 50 

65 53.5 49 50.1 94 47 51.8 97 46 45.1 84 49 

69 53.6 49 49.6 93 45 51.0 95 45 44.5 83 49 

73 53.5 49 49.6 93 45 50.8 95 45 44.0 82 47 

77 53.9 48 50.4 94 44 50.8 94 44 46.2 86 46 

81 54.2 47 50.0 92 43 50.5 93 43 44.2 82 46 

85 55.1 42 49.0 89 43 50.2 91 41 43.7 79 44 

89 54.5 40 49.8 91 40 49.8 91 40 44.3 81 43 

93 54.7 38 49.0 90 39 50.6 93 39 43.8 80 42 

94 54.5 37 48.6 89 39 50.1 92 39 43.6 80 41 

96 54.2 37 47.6 88 39 49.4 91 38 43.1 80 38 

98 53.3 37 48.5 91 36 49.1 92 38 43.7 82 37 

100 53.0 36 48.9 92 35 49.0 93 38 44.4 84 37 

102 52.2 35 47.4 91 35 47.6 91 37 43.4 83 36 

104 50.7 30 47.5 94 31 46.8 92 33 42.8 84 36 

Mean for weeks 

1-13 25.4 25.3 100 25.6 101 25.2 99 

14-52 40.6 38.4 95 39.1 96 36.4 90 

53-104 53.0 48.8 92 49.7 94 44.0 83 
 

 

 

70 2-Butoxyethanol, NTP TR 484 

Hematology and Bone Marrow Cellularity 

The hematology data are listed in Tables 18 and 

F4. As in the 14-week study, inhalation of 

2-butoxyethanol by mice in the 2-year study resulted 

in the development of a persistent, exposure-related, 

responsive anemia. The anemia, evidenced by decreases in automated and manual hematocrit values, 

hemoglobin concentrations, and erythrocyte counts, 

occurred at 3, 6, and 12 months in 125 and 250 ppm 

male and female mice; there also was evidence indicating an anemia at 6 months in 62.5 ppm females. 

The erythropoietic response was demonstrated by 

increased reticulocyte counts in 125 and 250 ppm 

male and female mice at 3 and 6 months and 250 ppm 

females at 12 months. The reticulocyte response 

ameliorated for 125 and 250 ppm males and 125 ppm 

females at 12 months. In general, the anemia was 

normocytic and normochromic and was demonstrated 

by the lack of changes in the mean cell volumes and 

mean cell hemoglobin concentrations. At 12 months, 

however, a minimal increase in the mean cell volume, 

suggesting a macrocytosis, occurred in 250 ppm 

females. A thrombocytosis, evidenced by increased 

platelet counts, occurred in 250 ppm males and 

females at 3, 6, and 12 months. Platelet counts also 

were increased in 125 ppm females at 6 months and 

62.5 ppm females and 125 males and females at 

12 months. Increased neutrophil counts occurred in 

125 ppm males and females at 6 months and 125 and 

250 ppm females at 12 months. Unlike the rats, there 

were no increases in the total bone marrow cell counts 

or decreases in the M/E ratios of exposed mice at any 

time point. 
2-Butoxyethanol, NTP TR 484 71 

TABLE 18 

Selected Hematology and Bone Marrow Cellularity Data for Mice at 3, 6, and 12 Months 

in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Male 

Hematology 

n 

3 Months 10 10 10 10 

6 Months 10 10 10 10 

12 Months 10 10 9 10 

Automated hematocrit (mL/dL) 

3 Months 46.3 ± 0.4 46.3 ± 0.3 44.9 ± 0.5* 42.5 ± 0.2** 

6 Months 48.1 ± 0.4 47.6 ± 0.5 46.6 ± 0.3* 43.4 ± 0.4** 

12 Months 48.8 ± 0.5 50.1 ± 2.0 47.0 ± 1.0 42.4 ± 0.5** 

Manual hematocrit (%) 

3 Months 47.5 ± 0.3 47.3 ± 0.5 46.0 ± 0.4* 43.7 ± 0.2** 

6 Months 48.1 ± 0.4 48.1 ± 0.4 47.2 ± 0.4 44.5 ± 0.3** 

12 Months 47.9 ± 0.4 48.7 ± 1.9 46.4 ± 1.0 42.1 ± 0.4** 

Hemoglobin (g/dL) 

3 Months 15.2 ± 0.1 15.3 ± 0.1 14.7 ± 0.2 13.8 ± 0.1** 

6 Months 15.7 ± 0.2 15.6 ± 0.1 15.2 ± 0.1** 14.3 ± 0.1** 

12 Months 15.7 ± 0.1 16.0 ± 0.7 14.9 ± 0.4* 13.6 ± 0.2** 

Erythrocytes (106

/μL) 

3 Months 9.61 ± 0.22 9.83 ± 0.06 9.41 ± 0.11 8.95 ± 0.05** 

6 Months 9.88 ± 0.10 9.79 ± 0.08 9.58 ± 0.07* 9.09 ± 0.07** 

12 Months 9.58 ± 0.07 9.73 ± 0.49 9.36 ± 0.32* 8.33 ± 0.10** 

Reticulocytes (106

/μL) 

3 Months 0.07 ± 0.01 0.07 ± 0.01 0.08 ± 0.01 0.13 ± 0.01** 

6 Months 0.05 ± 0.01 0.07 ± 0.01 0.09 ± 0.01** 0.17 ± 0.01** 

12 Months 0.06 ± 0.02 0.06 ± 0.02 0.11 ± 0.02 0.07 ± 0.02 

Mean cell volume (fL) 

3 Months 48.6 ± 1.2 47.1 ± 0.2 47.7 ± 0.2 47.4 ± 0.3 

6 Months 48.8 ± 0.4 48.6 ± 0.3 48.6 ± 0.3 47.8 ± 0.2 

12 Months 50.9 ± 0.3 51.7 ± 0.5 50.3 ± 0.8 51.1 ± 0.6 

Mean cell hemoglobin (pg) 

3 Months 15.9 ± 0.4 15.5 ± 0.0 15.7 ± 0.0 15.4 ± 0.1 

6 Months 15.9 ± 0.1 16.0 ± 0.1 15.9 ± 0.1 15.7 ± 0.1 

12 Months 16.4 ± 0.1 16.5 ± 0.2 16.0 ± 0.3 16.3 ± 0.2 

Mean cell hemoglobin concentration (g/dL) 

3 Months 32.8 ± 0.1 33.0 ± 0.1 32.8 ± 0.1 32.4 ± 0.1 

6 Months 32.6 ± 0.2 32.9 ± 0.2 32.7 ± 0.2 32.9 ± 0.2 

12 Months 32.2 ± 0.2 32.0 ± 0.1 31.7 ± 0.3 31.9 ± 0.1 

Platelets (103

/μL) 

3 Months 904.2 ± 18.3 888.2 ± 11.7 869.8 ± 13.4 940.6 ± 18.9 

6 Months 988.3 ± 16.8 955.4 ± 34.1 1,028.8 ± 20.6 1,075.2 ± 17.1** 

12 Months 

Segmented neutrophils (103

/μL) 

831.2 ± 38.1 997.6 ± 37.8** 1,116.6 ± 69.5** 1,112.8 ± 39.4** 

3 Months 0.67 ± 0.11 0.73 ± 0.08 0.81 ± 0.09 0.95 ± 0.15 

6 Months 0.43 ± 0.08 0.43 ± 0.04 0.75 ± 0.09* 0.62 ± 0.09 

12 Months 0.80 ± 0.04 0.84 ± 0.12 0.80 ± 0.13 0.70 ± 0.08 
72 2-Butoxyethanol, NTP TR 484 

TABLE 18 

Selected Hematology and Bone Marrow Cellularity Data for Mice at 3, 6, and 12 Months 

in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Male (continued) 

Bone Marrow Cellularity 

n 

3 Months 10 10 10 10 

6 Months 10 10 10 10 

12 Months 10 10 9 10 

Nucleated bone marrow cells (106

/femur) 

3 Months 23.4 ± 0.8 22.8 ± 0.6 22.1 ± 1.1 23.5 ± 0.9 

6 Months 23.8 ± 0.9 23.3 ± 1.0 23.5 ± 1.2 24.3 ± 1.4 

12 Months 32.9 ± 0.9 28.9 ± 0.9 30.1 ± 1.3 30.0 ± 1.5 

Myeloid/erythroid ratio 

3 Months 1.99 ± 0.10 2.11 ± 0.10 1.98 ± 0.13 2.02 ± 0.06 

6 Months 1.88 ± 0.08b 2.24 ± 0.10 2.33 ± 0.09** 2.10 ± 0.09 

12 Months 1.78 ± 0.12 2.15 ± 0.07 2.34 ± 0.07** 2.06 ± 0.10 

Female 

Hematology 

n 

3 Months 10 10 10 9 

6 Months 10 10 10 10 

12 Months 10 10 10 10 

Automated hematocrit (mL/dL) 

3 Months 48.9 ± 0.5 48.2 ± 0.3 46.0 ± 0.4** 42.8 ± 0.4** 

6 Months 48.2 ± 0.5 46.2 ± 0.6* 45.7 ± 0.3** 42.5 ± 0.3** 

12 Months 47.4 ± 0.5 46.9 ± 0.5 43.7 ± 0.6** 42.4 ± 0.4** 

Manual hematocrit (%) 

3 Months 49.3 ± 0.5 48.9 ± 0.4 46.2 ± 0.5** 43.7 ± 0.5** 

6 Months 48.1 ± 0.6 46.6 ± 0.5* 45.7 ± 0.2** 42.8 ± 0.2** 

12 Months 46.9 ± 0.4 46.3 ± 0.4 43.8 ± 0.4** 41.8 ± 0.3** 

Hemoglobin (g/dL) 

3 Months 15.5 ± 0.2 15.3 ± 0.1 14.6 ± 0.1** 13.4 ± 0.3** 

6 Months 15.6 ± 0.2 14.9 ± 0.2** 14.7 ± 0.1** 13.6 ± 0.1** 

12 Months 15.4 ± 0.1 15.0 ± 0.1* 14.3 ± 0.1** 13.6 ± 0.1** 

Erythrocytes (106

/μL) 

3 Months 9.89 ± 0.09 9.68 ± 0.07 9.23 ± 0.09** 8.58 ± 0.07** 

6 Months 9.71 ± 0.15 9.33 ± 0.12* 9.19 ± 0.06** 8.68 ± 0.05** 

12 Months 9.32 ± 0.09 9.14 ± 0.08 8.50 ± 0.12** 8.08 ± 0.09** 

Reticulocytes (106

/μL) 

3 Months 0.05 ± 0.00 0.06 ± 0.01 0.09 ± 0.01** 0.16 ± 0.01** 

6 Months 0.05 ± 0.01 0.06 ± 0.01 0.09 ± 0.01** 0.14 ± 0.01** 

12 Months 0.10 ± 0.01 0.14 ± 0.02 0.15 ± 0.02 0.24 ± 0.03** 

Mean cell volume (fL) 

3 Months 49.3 ± 0.3 49.7 ± 0.2 49.8 ± 0.3 49.8 ± 0.1 

6 Months 49.8 ± 0.6 49.5 ± 0.3 49.8 ± 0.4 49.0 ± 0.3 

12 Months 50.9 ± 0.3 51.3 ± 0.3 51.5 ± 0.2 52.4 ± 0.3** 
2-Butoxyethanol, NTP TR 484 73 

TABLE 18 

Selected Hematology and Bone Marrow Cellularity Data for Mice at 3, 6, and 12 Months 

in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Female (continued) 

Hematology (continued) 

n 

3 Months 10 10 10 9 

6 Months 10 10 10 10 

12 Months 10 10 10 10 

Mean cell hemoglobin (pg) 

3 Months 15.7 ± 0.1 15.8 ± 0.0 15.8 ± 0.1 15.7 ± 0.1 

6 Months 16.1 ± 0.2 16.0 ± 0.1 16.0 ± 0.1 15.7 ± 0.1 

12 Months 16.6 ± 0.1 16.5 ± 0.1 16.8 ± 0.1 16.8 ± 0.1 

Mean cell hemoglobin concentration (g/dL) 

3 Months 31.8 ± 0.1 31.7 ± 0.1 31.7 ± 0.1 31.4 ± 0.1* 

6 Months 32.4 ± 0.2 32.3 ± 0.2 32.1 ± 0.1 32.1 ± 0.1 

12 Months 32.6 ± 0.2 32.2 ± 0.2 32.7 ± 0.2 32.0 ± 0.2 

Platelets (103

/μL) 

3 Months 835.8 ± 27.5 832.9 ± 26.9 849.9 ± 12.4 921.3 ± 23.3* 

6 Months 938.6 ± 27.2 942.5 ± 34.9 1,010.6 ± 24.9* 1,064.6 ± 12.9** 

12 Months 778.1 ± 28.2 837.6 ± 14.0* 844.5 ± 28.6** 952.9 ± 12.1** 

Segmented neutrophils (103

/μL) 

3 Months 0.49 ± 0.11 0.38 ± 0.05 0.50 ± 0.07 0.59 ± 0.05 

6 Months 0.50 ± 0.05 0.83 ± 0.25 1.05 ± 0.31* 0.74 ± 0.07 

12 Months 0.56 ± 0.06 0.79 ± 0.14 0.75 ± 0.06* 0.88 ± 0.09** 

Bone Marrow Cellularity 

n 

3 Months 10 10 10 9 

6 Months 10 10 10 10 

12 Months 10 10 9 10 

Nucleated bone marrow cells (106

/femur) 

3 Months 23.0 ± 0.9 22.7 ± 1.0 23.2 ± 0.7 22.9 ± 0.9 

6 Months 26.5 ± 0.9 26.3 ± 1.6 24.6 ± 1.2 26.9 ± 0.5 

12 Months 32.7 ± 1.0 31.0 ± 1.9 34.7 ± 1.0 35.8 ± 1.0 

Myeloid/erythroid ratio 

3 Months 1.72 ± 0.12 2.03 ± 0.10 2.19 ± 0.10* 1.52 ± 0.09c 

6 Months 1.81 ± 0.06 1.94 ± 0.13 2.08 ± 0.07* 1.90 ± 0.07 

12 Months 1.53 ± 0.10 1.98 ± 0.08* 1.94 ± 0.13 1.59 ± 0.11 

* Significantly different (P#0.05) from the chamber control group by Dunn’s or Shirley’s test 

** P#0.01 a Mean ± standard error. Statistical tests were performed on unrounded data. b n=9
 

n=10
 c 
 

 

74 2-Butoxyethanol, NTP TR 484 

Pathology and Statistical Analysis 

This section describes the statistically significant or 

biologically noteworthy changes in the incidences of 

neoplasms and/or nonneoplastic lesions of the forestomach, liver, spleen, bone marrow, nose, and urogenital system. Summaries of the incidences of neoplasms and nonneoplastic lesions, individual animal 

tumor diagnoses, statistical analyses of primary neoplasms that occurred with an incidence of at least 5% 

in at least one animal group, and historical incidences 

for the neoplasms mentioned in this section are presented in Appendix C for male mice and Appendix D 

for female mice. 

Forestomach: The incidences of squamous cell 

papilloma and squamous cell papilloma or carcinoma 

(combined) occurred with a positive trend in females, 

and the incidences in females exposed to 250 ppm 

were significantly increased relative to the chamber 

controls (Tables 19 and D3). These incidences exceeded the ranges for historical controls (Tables 19 

and D4a). In male mice exposed to 125 or 250 ppm, 

the incidences of squamous cell papilloma also exceeded the range for historical controls (Tables 19 and 

C4a). A squamous cell carcinoma was observed in a 

male mouse exposed to 125 ppm; this mouse also had 

a papilloma. Squamous cell papilloma had the morphology typical of these neoplasms in B6C3F1 mice 

and consisted of multiple branching papillary 

projections, composed of a thick layer of epithelium 

overlying a fibrous tissue core, radiating from a basal 

stalk (Plate 5). The squamous cell carcinoma was 

characterized by the invasion of cords or clusters of 

neoplastic cells through the forestomach wall (Plates 6 

and 7). Incidences of ulcer were significantly increased relative to the chamber controls in males 

exposed to 125 ppm and in all exposed groups of 

females (Tables 19, C5, and D5). Ulcer consisted of 

a defect in the forestomach wall that penetrated the 

full thickness of the forestomach epithelium. The 

ulcerated area frequently contained accumulations of 

inflammatory cells and debris. Incidences of epithelial hyperplasia, usually focal, were significantly 

increased in all exposed groups of males and females. 

Frequently, particularly in the females, the hyperplasia was associated with ulceration. Hyperplasia 

consisted of an increased thickness of the stratified 

squamous epithelium of the forestomach; this was 

accompanied in some cases by an increase in the 

thickness of the keratinized layer (Plate 8). 
 

2-Butoxyethanol, NTP TR 484 75 

TABLE 19 

Incidences of Neoplasms and Nonneoplastic Lesions of the Forestomach in Mice 

in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Male 

Number Necropsied 

Ulcera 50 1 (1.0)b 

50 

2 (1.5) 

50 

9** (1.6) 

50 

3 (1.7) 

Epithelium, Hyperplasia 1 (3.0) 7* (2.3) 16** (1.8) 21** (2.3) 

Number Examined Microscopically 50 50 49 48 

Squamous Cell Papilloma 1 1 2 2 

Squamous Cell Carcinoma 0 0 1 0 

Squamous Cell Papilloma or Carcinomac 1 1 2 2 

Female 

Number Necropsied 50 50 50 50 

Ulcer 1 (3.0) 7* (1.3) 13** (1.5) 22** (1.4) 

Epithelium, Hyperplasia 6 (1.8) 27** (2.0) 42** (2.4) 44** (2.9) 

Squamous Cell Papillomad 

Overall ratee 

Adjusted ratef 0/50 (0%) 

0.0% 

1/50 (2%) 

2.4% 

2/50 (4%) 

4.8% 

5/50 (10%) 

11.2% 

Terminal rateg 

First incidence (days) 

Poly-3 testh 

0/29 (0%) 

—i 

P=0.008 

1/31 (3%) 

731 (T) 

P=0.495 

2/33 (6%) 

731 (T) 

P=0.231 

3/36 (8%) 

582 

P=0.034 

Squamous Cell Carcinoma 0 0 0 1 

Squamous Cell Papilloma or Carcinomaj 

Overall rate 0/50 (0%) 1/50 (2%) 2/50 (4%) 6/50 (12%) 

Adjusted rate 0.0% 2.4% 4.8% 13.4% 

Terminal rate 0/29 (0%) 1/31 (3%) 2/33 (6%) 4/36 (11%) 

First incidence (days) — 731 (T) 731 (T) 582 

Poly-3 test P=0.002 P=0.495 P=0.231 P=0.017 

(T)Terminal sacrifice 

* Significantly different (P#0.05) from the chamber control group by the Poly-3 test 

** P#0.01 a Number of animals with lesion 

b Average severity grade of lesions in affected animals: 1=minimal, 2=mild, 3=moderate, 4=marked c Historical incidence for 2-year inhalation studies with chamber control groups (mean ± standard deviation): 5/970 (0.5% ± 0.9%); 

range, 0%-2% d	 Historical incidence: 7/973 (0.7% ± 1.0%); range, 0%-2% e Number of animals with neoplasm per number of animals with forestomach examined microscopically f Poly-3 estimated neoplasm incidence after adjustment for intercurrent mortality 

g Observed incidence at terminal kill 

h	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the 

P values corresponding to pairwise comparisons between the chamber controls and that exposed group. The Poly-3 test accounts for 

differential mortality in animals that do not reach terminal sacrifice. i	 Not applicable; no neoplasms in animal group j Historical incidence: 9/973 (0.9% ± 1.1%); range, 0%-3% 
 

 

 

 

76 2-Butoxyethanol, NTP TR 484 

Liver: The incidences of hemangiosarcoma occurred 

with a positive trend in male mice. The incidence of 

hemangiosarcoma in males exposed to 250 ppm was 

significantly increased relative to the chamber controls 

and exceeded the range of historical controls 

(Tables 20 and C3). Hemangiosarcomas had a morphological structure similar to that of spontaneously 

occurring hemangiosarcomas, such as the presence of 

atypical endothelial cells that form capillary and 

cavernous channels and the presence of solid cellular 

masses and local invasion. Two of the four 250 ppm 

mice with hemangiosarcomas also had hemangiosarcomas in either the bone marrow and heart or 

bone marrow and spleen. Due to the extreme infiltration in the bone marrow and spleen, it was not 

possible to determine whether these were the primary 

or metastatic foci for the hemangiosarcomas. The 

incidences of hepatocellular carcinoma occurred with 

a positive trend in male mice. The incidence of 

hepatocellular carcinoma in males exposed to 

250 ppm was significantly increased relative to the 

chamber controls. 

In assessing potential chemical-related increases in the 

incidences of liver neoplasms, benign and malignant 

neoplasms are routinely analyzed independently and 

in combination. Although the independent analyses 

provide useful information, the most important 

analysis for determining potential chemical-related 

effects in the liver is the combined analysis (hepatocellular adenoma and carcinoma). There was no 

difference in the combined incidence of hepatocellular 

neoplasms between chamber control and exposed 

groups (Tables 20, C3, and C4). While the statistically significant increased incidence of hepatocellular carcinoma in 250 ppm males may suggest a 

progression toward malignancy, the incidence of 

hepatocellular carcinoma in this group is well within 

the historical control range. Additionally, there were 

significant decreases in the incidences of hepatocellular adenoma in 125 and 250 ppm female mice, 

while the combined hepatocellular neoplasm incidences were not different between groups. Also, 

there was no increase in the incidence of potential 

preneoplastic lesions (e.g., altered foci). The decreased incidences of hepatocellular adenoma in 125 

and 250 ppm females were interpreted as normal 

variations based upon chance rather than effects 

associated with exposure to 2-butoxyethanol. The 

increased incidence of hepatocellular carcinoma in 

250 ppm males may have been caused by exposure to 

2-butoxyethanol. 

In a retrospective evaluation, a bacterial organism, 

Helicobacter hepaticus, has been identified in the 

livers of mice from several 2-year carcinogenicity 

studies conducted by the NTP (Hailey et al., 1998). 

In the studies in which significant liver disease was 

attributable to H. hepaticus, an association with 

increased incidences of liver neoplasms (hepatocellular neoplasms and hemangiosarcoma of the liver) 

was demonstrated. The most definitive method of 

identifying the presence of H. hepaticus is with 

polymerase chain reaction-based assays on fresh or 

frozen liver tissue. However, in other studies in 

which liver disease was associated with the presence 

of the organism, the use of silver stains (e.g., 

Steiner’s modification of Warthin-Starry) has also 

been effective in identifying a helical organism 

consistent with H. hepaticus. Fresh or frozen liver 

tissue was not available from this study; therefore, 

sections of liver from six males from the chamber 

control group and eight from the 250 ppm group were 

stained with Steiner’s modification of the WarthinStarry silver stain and evaluated for H. hepaticus. 

While inflammation, karyomegaly, and oval cell 

hyperplasia are nonspecific liver lesions that can be 

observed under a number of circumstances, they are 

consistent components of the liver disease observed in 

male mice infected with H. hepaticus. Therefore, to 

maximize the potential for identifying H. hepaticus in 

this study, the 14 animals were selected for staining 

based on the presence of these lesions in the liver. 

No organisms consistent with H. hepaticus were 

identified in any of the 14 animals evaluated; 

therefore, H. hepaticus was not considered to have 

been a factor in the development of liver neoplasms in 

this study. 

Incidences of hemosiderin pigmentation in the Kupffer 

cells were significantly increased in males exposed to 

125 or 250 ppm and in all exposed groups of females 

(Tables 20, C5, and D5). 
2-Butoxyethanol, NTP TR 484 77 

TABLE 20 

Incidences of Neoplasms and Nonneoplastic Lesions of the Liver in Mice in the 2-Year Inhalation Study 

of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Male 

Number Examined Microscopically 

Kupffer Cell, Pigmentation, Hemosiderina 50 0 

50 

0 

49 

8** (1.0)b

49 

 30** (1.2) 

Hemangiosarcomac 

Overall rated 0/50 (0%) 1/50 (2%) 2/49 (4%) 4/49 (8%) 

Adjusted ratee 

Terminal ratef 

First incidence (days) 

0.0% 

0/39 (0%) 

—h 

2.1% 

0/39 (0%) 

670 

5.0% 

1/27 (4%) 

704 

10.0% 

2/26 (8%) 

454 

Poly-3 testg P=0.014 P=0.511 P=0.211 P=0.046 

Hepatocellular Adenoma 

Hepatocellular Carcinomai 

Hepatocellular Adenoma or Carcinomaj 

22 

10

30

18 

11 

24 

18 

16 

31 

17 

21** 

30 

Female 

Number Examined Microscopically 50 50 49 50 

Kupffer Cell, Pigmentation, Hemosiderin 0 5* (1.0) 25** (1.0) 44** (1.0) 

Hemangiosarcoma 

Hepatocellular Adenomak 

0 

16 

1 

8

0 

7* 

0 

8* 

Hepatocellular Carcinoma 10 12 13 10 

Hepatocellular Adenoma or Carcinoma 22 16 18 18 

* Significantly different (P#0.05) from the chamber control group by the Poly-3 test 

** P#0.01 a Number of animals with lesion 

b Average severity grade of lesions in affected animals: 1=minimal, 2=mild, 3=moderate, 4=marked c Historical incidence for 2-year inhalation studies with chamber control groups (mean ± standard deviation): 14/968 (1.5% ± 1.5%); 

range, 0%-4% d	 Number of animals with neoplasm per number of animals microscopically examined e Poly-3 estimated neoplasm incidence after adjustment for intercurrent mortality f Observed incidence at terminal kill 

g	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the 

P values corresponding to pairwise comparisons between the chamber controls and that exposed group. The Poly-3 test accounts for 

differential mortality in animals that do not reach terminal sacrifice. h	 Not applicable; no neoplasms in animal group i	 Historical incidence: 247/968 (25.7% ± 10.4%); range, 11%-48% j Historical incidence: 503/968 (52.2% ± 16.4%); range, 20%-86% k Historical incidence: 191/968 (19.8% ± 10.2%); range, 8%-43% 

Spleen: Incidences of hematopoietic cell proliferation increased in all exposed groups of males and in 

in males exposed to 125 or 250 ppm and females females exposed to 125 or 250 ppm. Hematopoietic 

exposed to 250 ppm were significantly increased cell proliferation and hemosiderin pigmentation were 

relative to the chamber controls (Tables 21, C5, and attributed to the primary hemolytic effect of 

D5). This lesion consisted of excess production of 2-butoxyethanol, which was followed by regenerative 

cells with no apparent change in M/E ratios. Inci- hyperplasia of the hematopoietic tissue. 

dences of hemosiderin pigmentation were significantly 
 

 

 

78 2-Butoxyethanol, NTP TR 484 

TABLE 21 

Incidences of Nonneoplastic Lesions of the Spleen in Mice in the 2-Year Inhalation Study of 

2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Male 

Number Examined Microscopically 

Hematopoietic Cell Proliferationa 50 12 (1.8)b 

50 

11 (2.5) 

48 

26** (2.3) 

49 

42** (2.4) 

Hemosiderin Pigmentation 0 6* (1.0) 45** (1.6) 44** (1.9) 

Female 

Number Examined Microscopically 50 50 49 50 

Hematopoietic Cell Proliferation 24 (2.5) 29 (2.4) 32 (2.2) 35* (2.1) 

Hemosiderin Pigmentation 39 (1.5) 44 (1.8) 46** (1.9) 48** (2.0) 

* Significantly different (P#0.05) from the chamber control group by the Poly-3 test 

** P#0.01 a Number of animals with lesion 

b Average severity grade of lesions in affected animals: 1=minimal, 2=mild, 3=moderate, 4=marked 

Bone Marrow: Incidences of hyperplasia in males 

exposed to 125 or 250 ppm were significantly 

increased relative to the chamber controls (0/50, 1/50, 

9/49, 5/50; Table C5). 

Nose: The incidences of hyaline degeneration were 

increased relative to the chamber controls in the 

olfactory epithelium (6/50, 14/50, 11/49, 12/50) and 

in the respiratory epithelium (17/50, 35/50, 26/49, 

23/50) of exposed groups of females (Table D5); 

however, the severities of these lesions were similar 

to those in the chamber controls (olfactory: 1.3, 1.1, 

1.3, 1.2; respiratory: 1.1, 1.3, 1.2, 1.1). In males, 

the incidences of hyaline degeneration were similar to 

those of the chamber controls in the olfactory epithelium (1/50, 2/50, 3/48, 1/48) and in the respiratory 

epithelium (4/50, 10/50, 5/48, 5/48), as were the 

severities (olfactory: 1.0, 1.0, 1.3, 1.0; respiratory: 

1.0, 1.1, 1.4, 1.0) (Table C5). These lesions 

consisted of intracytoplasmic accumulation of homogenous eosinophilic material. The affected olfactory 

epithelium was located primarily in nasal levels I and 

II. These lesions were seen as brightly eosinophilic 

globules in the cytoplasm of affected respiratory 

epithelial cells and sustentacular cells, with the 

olfactory cells being involved most often. 

Urogenital System: In the kidney, incidences of 

glomerulosclerosis and hydronephrosis in males 

exposed to 125 ppm and of chronic inflammation in 

males exposed to 250 ppm were significantly 

increased relative to the chamber controls (Tables 22 

and C5). The incidences of inflammation of the 

preputial and prostate glands in males exposed to 

250 ppm were significantly increased. Incidences of 

chronic inflammation and ulcer of the prepuce skin 

were significantly increased in males exposed to 125 

or 250 ppm compared to the chamber controls, as 

were the incidences of inflammation of the urinary 

bladder. The incidence of ulcer of the transitional 

epithelium of the urinary bladder in males in the 

125 ppm group was significantly increased. These 

inflammatory changes in the urinary tract are indicative of an ascending urinary tract infection and are 

consistent with a genito-urinary condition described in 

the literature as mouse urologic syndrome. In 

inhalation studies, the incidence of mouse urologic 

syndrome is particularly high in mice housed individually in wire mesh cages (Everitt et al., 1988). It is 

likely that this condition was exacerbated by the 

irritative effects of 2-butoxyethanol exposure directly 

or by the presence of 2-butoxyethanol metabolites in 

urine, or by both. 
79 

c 

2-Butoxyethanol, NTP TR 484 

TABLE 22 

Incidences of Nonneoplastic Lesions of the Urogenital System in Male Mice in the 2-Year Inhalation Study 

of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Kidneya 

Glomerulosclerosisb 

50 

4 (1.3)c 50 4 (1.3) 

47 

11* (1.3) 

50 

9 (1.4) 

Hydronephrosis 1 (2.0) 0 6* (1.8) 5 (2.2) 

Inflammation, Chronic Active 0 1 (2.0) 2 (2.5) 4* (3.0) 

Preputial Gland 49 49 49 49 

Inflammation 2 (1.0) 7 (2.3) 6 (2.3) 8* (2.1) 

Prostate Gland 49 49 48 45 

Inflammation 0 2 (2.0) 3 (2.3) 4* (1.8) 

Skin, Prepuce 50 50 49 50 

Inflammation, Chronic Active 2 (3.5) 3 (3.7) 13** (3.7) 8* (3.9) 

Ulcer 0 3 (2.7) 11** (2.8) 8** (2.5) 

Urinary Bladder 50 50 46 45 

Inflammation 0 2 (3.5) 5* (3.4) 4* (3.8) 

Transitional Epithelium, Ulcer 0 1 (3.0) 4* (2.8) 2 (3.0) 

* Significantly different (P#0.05) from the chamber control group by the Poly-3 test 

** P#0.01 a Number of animals with organ examined microscopically b Number of animals with lesion 

Average severity grade of lesions in affected animals: 1=minimal, 2=mild, 3=moderate, 4=marked 

GENETIC TOXICOLOGY 

2-Butoxyethanol (100 to 10,000 μg/plate) did not 

induce mutations in any of the Salmonella 

typhimurium strains tested (TA97, TA98, TA100, 

TA1535, and TA1537), with or without induced 

hamster or rat liver S9 (Table E1; Zeiger et al., 

1992). In tests for induction of chromosomal damage 

in Chinese hamster ovary cells in vitro, 

2-butoxyethanol induced cell cycle delay (an 

indication of cytotoxicity) but did not induce either 

sister chromatid exchanges (Table E2) or chromosomal aberrations (Table E3) with or without S9. In 

the chromosomal aberrations test without S9, a 

weakly positive response was obtained in the second 

trial at the highest dose tested (5,000 μg/mL), but this 

response was not reproduced in a third trial and the 

test results were concluded to be negative overall. 

Due to the cell cycle delay caused by 2-butoxyethanol 

in the trials conducted without S9, a delayed harvest 

was used to increase the number of cells available for 

analysis. In vivo, no induction of micronuclei was 

observed in polychromatic erythrocytes in bone 

marrow of rats or mice treated with 2-butoxyethanol 

(Tables E4 and E5). Rats received up to 450 mg 

2-butoxyethanol/kg body weight three times at 

24-hour intervals via intraperitoneal injection; two out 

of five rats administered 450 mg/kg died. Mice were 

treated by the same protocol. All mice receiving 

550 mg/kg survived, whereas 100% mortality 

occurred in the 1,100 mg/kg dose groups. 
80 2-Butoxyethanol, NTP TR 484 
2-Butoxyethanol, NTP TR 484 

PLATE 1 PLATE 2 

Blood vessel thrombosis (arrow) in the incisors dental pulp in a 

female F344/N rat killed moribund 4 days following initiation of 

exposure to 500 ppm 2-butoxyethanol in the 14-week inhalation 

study. H&E; 25×. 

Thrombosis of small caliber arteries in the submucosa of the septum 

at level I of the nasal cavity of a female F344/N rat killed moribund 4 

days following initiation of exposure to 500 ppm 2-butoxyethanol in 

the 14-week inhalation study. H&E; 50×. 

PLATE 3 PLATE 4 

Bone infarction at the level of growth plate and primary spongiosa 

characterized by presence of necrotic osteocytes (arrows) and necrosis of the bone-lining cells in a female F344/N rat killed moribund 4 

days following initiation of exposure to 500 ppm 2-butoxyethanol in 

the 14-week inhalation study. H&E; 25×. 

Odontoblast degeneration (arrow) in the incisor tooth of a female 

F344/N rat killed moribund 4 days following initiation of exposure to 

500 ppm 2-butoxyethanol in a 14-week inhalation study. H&E; 80×. 
2-Butoxyethanol, NTP TR 484 

PLATE 5 

Squamous cell papilloma in the forestomach of a female B6C3F1 

mouse exposed to 250 ppm 2-butoxyethanol by inhalation for 2 

years. Note the presence of multiple branching papillary projections. 

H&E; 4×. 

PLATE 6 

Squamous cell carcinoma in the forestomach of a female B6C3F1 

mouse exposed to 250 ppm 2-butoxyethanol by inhalation for 2 years. 

H&E; 8×. 

PLATE 7 

Squamous cell carcinoma in the forestomach (same animal as presented in Plate 6). Note clusters of stratified squamous cells that have 

invaded the submucosa and muscular layer. H&E; 20× 

PLATE 8 

Mucosal ulceration associated with moderate squamous cell hyperplasia and mild submucosal inflammation in the forestomach of a 

male B6C3F mouse exposed to 250 ppm 2-butoxyethanol by inhala­ 1 

tion for 2 years. H&E; 25× 
 

 

 

 

 

81 

DISCUSSION AND CONCLUSIONS
 

The Consumer Product Safety Commission and the 

United Auto Workers International Union nominated 

2-butoxyethanol for study because of its use in many 

industrial and consumer applications, the potential 

exposure of workers and the general population, and 

the absence of chronic toxicity data. 2-Butoxyethanol 

was evaluated for toxicity and carcinogenicity in 

14-week and 2-year studies in male and female 

F344/N rats and B6C3F1 mice utilizing whole body 

inhalation as the route of exposure. 

In previous studies, short-term exposure of 

2-butoxyethanol to rats and mice caused hemolytic 

anemia and subsequent effects on the hematopoietic system by the 2-butoxyethanol metabolite 

2-butoxyacetic acid (ATSDR, 1998; Ghanayem, 

1996); the duration of these studies was less than 

14 weeks and primarily involved rats. In the current 

studies, exposure of male and female rats and mice to 

2-butoxyethanol for periods of 14 weeks or 2 years 

also caused a regenerative hemolytic anemia and 

subsequent effects on the hematopoietic system in rats 

and mice. In addition to the hemolytic effect, 

2-butoxyethanol exposure for 2 years caused increases 

in the incidences of neoplasms and nonneoplastic 

lesions. 

In the current studies, a concentration-dependent 

anemia occurred in exposed rats and mice; rats 

were more severely affected than mice, and females 

more severely than males. The anemia in rats was 

characterized as macrocytic, normochromic, and 

responsive and was evidenced by increases in mean 

cell volumes, no changes in mean cell hemoglobin 

concentrations, and an erythropoietic response 

demonstrated by increased reticulocyte and nucleated 

erythrocyte counts. However, in mice, the anemia 

was considered normocytic. The 14-week female rats 

were more sensitive to the hemolytic effects of acute 

2-butoxyethanol exposure; female rats removed early 

had thrombosis in the blood vessels of a number of 

tissues, including the heart atrium, nasal septum and 

turbinates, liver, lung, incisors, and femur. In some 

tissues, the thrombosis was severe and/or associated 

with infarction, such as in the blood vessels of the 

dental pulp of the incisors, where there was odontoblast degeneration, and in the vessels of the tail 

vertebrae, where infarction led to necrosis and ultimate loss of the distal portion of the tail. Loss of the 

distal portion of the tail was also noted in prior 

reports (Hardin et al., 1984; Nelson et al., 1984; Tyl 

et al., 1984) of inhalation, dermal, and gavage studies 

with 2-butoxyethanol. It is proposed that 

2-butoxyethanol at concentrations of 500 ppm and 

greater (as used in pilot studies) produces an acute 

disseminated thrombosis and bone infarction in male 

and female rats as a result of severe acute hemolysis 

and reduced deformability of the erythrocytes or 

through anoxic damage to endothelial cells that 

compromises blood flow. In surviving female rats, 

there were lesions in the tail consistent with prior 

infarction, transient or complete growth arrest of the 

vertebrae, and, in the most severely affected animals, 

there was growth plate degeneration with no evidence 

of renewed longitudinal growth, indicating 

irreversible growth-plate injury. A number of effects 

on the hematopoietic system in rats and mice surviving to the end of the 14-week studies were 

consistent with the regenerative anemia, including 

excessive splenic extramedullary hematopoiesis and 

hemosiderosis, hemosiderin accumulation in hepatic 

Kupffer cells and renal cortical tubules, and bone 

marrow hyperplasia (rats). At the end of 2 years, 

only the pigmentation in the liver was present in 

exposed rats, while hematopoietic cell proliferation 

and pigmentation of the spleen and pigmentation in 

the liver occurred in exposed mice. 

In the 2-year studies, 2-butoxyethanol continued to 

affect the circulating erythroid mass, inducing a 

responsive anemia. In rats, the anemia was considered mild and persisted with no apparent progression or amelioration of severity from 3 months to 

the final blood collection at 12 months. Although the 

anemia in mice, for the most part, was minimal and 

characterized as normocytic, normochromic, and 

responsive, there were changes with duration of 

exposure. By 12 months, the reticulocytosis was 

ameliorated in males and was only present in 125 ppm 

females. It has been demonstrated that after an initial 
 

 

 

 

 

82 2-Butoxyethanol, NTP TR 484 

acute hemolytic event, rats become resistant to the 

hemolytic effects of 2-butoxyethanol (Ghanayem 

et al., 1992; Sivarao and Mehendale, 1995). The 

assumption is that the response is nonspecific and 

is related to the increase in the number of immature erythrocytes. As immature erythrocytes age, 

the increased resistance diminishes (Sivarao and 

Mehendale, 1995). The findings in this 2-year study 

also suggest that as the immature erythrocytes age, 

they become susceptible to the hemolytic effects of 

2-butoxyethanol and are hemolyzed, which results in 

a persistent anemia. Apparently, there is a balance 

between the release of immature erythrocytes to the 

circulation and the aging process so that at any particular time, only a limited number of erythrocytes are 

susceptible to hemolysis; thus, the anemia is persistent 

without any dramatic changes in severity. The 

macrocytosis observed in rats was attributed to the 

number of larger reticulocytes in the circulation and 

is consistent with an erythropoietic response. 

Reticulocytosis can result in transitory increases in 

mean cell volumes and mean cell hemoglobin values 

(Duncan and Prasse, 1986); thus, the increases in 

mean cell volumes and mean cell hemoglobin values 

in rats is consistent with the reticulocytosis that 

occurred in response to the anemia. There were 

increases in bone marrow cellularity in 125 ppm 

female rats at all time points that were consistent with 

a bone-marrow response to anemia. In the 13-week 

drinking water study with 2-butoxyethanol, total bone 

marrow cell counts were increased in rats (NTP, 

1993b). The bone marrow cytocentrifuge preparations, in general, revealed decreases in the myeloid 

to erythroid (M/E) ratios for 125 ppm males and 

females and 62.5 ppm females throughout the 2-year 

study. In the present study, the severities of the M/E 

ratio depressions were consistent with the persistent, 

responsive anemia at each time point. Unlike the rats, 

there were no increases in the total bone marrow cell 

counts or decreases in the M/E ratios of exposed mice 

at any time point. 

As noted previously, female rats are more sensitive to 

the 2-butoxyethanol-induced hemolysis than are males 

(Carpenter et al., 1956; Dodd et al., 1983; NTP, 

1993b). The gender difference in rats is consistent 

with the kinetics observed in this 2-year study 

(Dill et al., 1998). The hemolytic effects of 

2-butoxyethanol exposure are caused by 

2-butoxyacetic acid, the major metabolite of 

2-butoxyethanol (Carpenter et al., 1956; Bartnik 

et al., 1987; Ghanayem, 1987a). The toxicokinetic 

data for male and female rats reported for the 2-year 

study (Dill et al., 1998; Lee et al., 1998) indicate that 

there are definite gender differences in rats, especially 

in the elimination of 2-butoxyacetic acid. Female rats 

eliminated 2-butoxyacetic acid more slowly from the 

blood, as indicated by the smaller elimination rate 

constant, longer elimination half-life, and larger area 

under the blood concentration-versus-time curve. In 

addition, female rats excreted significantly less 

2-butoxyacetic acid in urine than did males; this 

reduced renal excretion of 2-butoxyacetic acid may be 

the cause of higher blood concentrations. The 

observed maximum blood concentrations of 

2-butoxyacetic acid for females were greater than for 

males at each exposure concentration and time point. 

Therefore, female rats had considerably more 

2-butoxyacetic acid in the blood at any given time 

point to produce a greater hemolytic effect than did 

males. Griffin et al. (1997) showed similar gender 

differences with male and female Sprague-Dawley 

rats dosed orally with 2,4-dichlorophenoxyacetic acid. 

Male rats were able to clear 2,4-dichlorophenoxyacetic acid from the plasma much more rapidly than 

were females, suggesting that female rats were 

exposed to a greater dose for a longer period of time 

than were males. The difference in clearance was 

attributed to a gender difference in anion transport in 

the kidney. The slight differences in the incidences of 

anemia in male and female mice may also be 

explained by kinetics but less clearly than for rats 

(Dill et al., 1998). Like female rats, female mice 

tended to have greater blood concentrations of 

2-butoxyacetic acid than did male mice. Unlike 

female rats, female mice excreted slightly more 

2-butoxyacetic acid than did male mice. However, 

there was no significant difference between male and 

female mice in the overall rate of elimination or the 

half-life of 2-butoxyacetic acid. 

In the 2-year rat study, survival rates were not affected by 2-butoxyethanol exposure. Mean body 

weights of exposed male rats were generally similar 

to those of the chamber controls, whereas mean body 

weights of 125 ppm females were slightly less than 

those of chamber controls for much of the study. The 

only potentially positive neoplastic finding in rats was 

a marginal increase in the incidence of benign or 

malignant pheochromocytomas (combined) of the 

adrenal gland in 125 ppm female rats; the incidences 

occurred with a positive trend in females. The inci-
 

 

 

2-Butoxyethanol, NTP TR 484 83 

dence (8/49) was not significantly greater than that of 

the chamber controls (3/50); however, the eight neoplasms observed in the 125 ppm group exceeded the 

overall historical control incidence in inhalation 

studies (6.4%) and slightly exceeded the highest 

incidence observed in any one inhalation control 

group (13%) or noninhalation control group (14%). 

Pheochromocytomas are relatively common in male 

rats, occurring with an historical control incidence of 

33%. In the 125 ppm female rats, one animal had a 

malignant and one animal had a bilateral benign 

pheochromocytoma. The primary criterion used to 

diagnose pheochromocytomas, in contrast to medullary hyperplasia, was the presence of mild to moderate compression of the adjacent tissue. Most of the 

pheochromocytomas observed were small and not 

substantially larger than the more severe grades of 

adrenal gland medullary hyperplasia. In addition, the 

incidences of medullary hyperplasia were not significantly increased in exposed rats. Overall, the 

slight increase in incidences of pheochromocytoma 

was considered an equivocal finding and could not be 

attributed with certainty to 2-butoxyethanol exposure. 

In the 2-year mouse study, survival rates were 

reduced in 125 or 250 ppm male mice. Exposed 

males generally had slightly lower mean body weights 

than chamber controls during the last 6 months of the 

study, whereas exposed females weighed less than 

chamber controls earlier in the study and had a 

greater reduction in weight. 2-Butoxyethanol 

exposure resulted in a concentration-related increase 

in the incidences of squamous cell papilloma or 

carcinoma (combined) of the forestomach. The 

incidence in 250 ppm females, in addition to being 

significantly increased compared to that in the 

chamber controls, exceeded the range for historical 

inhalation controls. Forestomach neoplasms are rare 

in B6C3F1 mice; for the contemporary historical 

controls for other routes of administration, 8% is the 

highest incidence observed in a chamber control 

group. Although forestomach inflammation and 

necrosis were more prevalent in mice removed during 

the first 9 days of the 14-week study, hyperplasia and 

inflammation were present at the end of the study 

(females only), indicating that the forestomach 

continued to be affected by repeated 2-butoxyethanol 

exposure. With repeated exposure in the 2-year 

study, there were significant concentration-related 

increases in the incidences of forestomach hyperplasia 

and ulceration. The hyperplasia was focal and often 

associated with the ulceration. A direct association of 

neoplasia with ulceration and hyperplasia was not 

shown in this study although it is hypothesized that 2-

butoxyethanol exposure-induced irritation caused the 

inflammatory and hyperplastic effects in the 

forestomach, and that the neoplasia was associated 

with a continuation of the injury/degeneration process 

and was, therefore, related to 2-butoxyethanol 

exposure. Assessment of ras mutations in 

forestomach neoplasms from male and female mice 

exposed to 2-butoxyethanol showed that the mutation 

frequency was similar to that of spontaneously 

occurring forestomach neoplasms (Appendix K). 

In addition, there was no difference in the spectrum of 

ras mutations between males and females. The 

mechanism of forestomach exposure is not clear, and 

the role of preening or mucocilliary clearance of the 

respiratory tract in the exposure is unknown. A direct 

effect of 2-butoxyethanol exposure on the forestomach 

is suggested by the work of Ghanayem et al. (1987b); 

48 hours after gavage administration of 14C-labeled 

2-butoxyethanol, the highest concentration of the label 

was in the forestomach, with approximately equal 

concentrations in the liver, kidney, lung, glandular 

stomach, and spleen. The glandular stomach contained only one-third that of the forestomach, which 

may indicate that for 2-butoxyethanol exposure there 

is a different reactivity and/or absorption in the two 

parts of the stomach. 

For male mice exposed to 2-butoxyethanol, there 

were increases in the incidences of squamous cell 

papilloma or carcinoma (combined) of the 

forestomach. Although the incidences were low and 

not significantly increased at any concentration, the 

incidences in the 125 and 250 ppm mice exceeded the 

historical control range. Squamous cell papillomas 

and carcinomas (one in the 125 ppm group) are as 

rare in males as in females. Squamous cell carcinomas of the forestomach have not been observed in 

inhalation historical control male mice. For contemporary historical controls in inhalation studies, the 

highest number of papillomas observed was one, and 

for papillomas and carcinomas combined for other 

routes, three were seen in one untreated control 

group; otherwise, one was the highest observed for 

any route. Forestomach inflammation, necrosis, and 

hyperplasia were observed in mice removed early in 

the 14-week studies. Inflammatory and hyperplastic 

changes were observed at the end of the 14-week 
 

 

 

 

 

 

84 2-Butoxyethanol, NTP TR 484 

study; however, they were more prevalent and 

severe in females than in males. With repeated 

exposure in the 2-year study, there was a significant, 

concentration-related increase in the incidences of 

forestomach hyperplasia. The incidences of ulcers 

were also increased in exposed males. The increased 

incidences of forestomach neoplasms may have been 

associated with 2-butoxyethanol exposure because of 

the rarity of these neoplasms, the increased incidences 

of nonneoplastic lesions supporting the neoplasia, the 

observation of nonneoplastic lesions in the 14-week 

studies, and most importantly, the fact that there was 

a similar, but more severe effect in female mice. The 

forestomach has not been a target for many inhalation 

studies. However, forestomach hyperplasia in male 

and female mice has been associated with inhalation 

exposure to vapors such as acetonitrile (NTP, 1996), 

1,3-butadiene (NTP, 1993c), and chloroprene (NTP, 

1998). Although the incidences of forestomach 

neoplasms were increased in exposed animals for each 

chemical, only the increases in the 1,3-butadiene and 

chloroprene studies were considered treatment 

related. The incidences for acetonitrile were low and 

did not exceed the historical control ranges. 

2-Butoxyethanol exposure caused a concentrationrelated increase in the incidences of hemangiosarcoma 

of the liver in male mice that was significant in the 

250 ppm group. Although the incidence was low, it 

exceeded the historical control incidence. For contemporary inhalation historical controls, no more than 

two hemangiosarcomas have been observed in any 

one control group, whereas as many as three have 

been observed in dosed feed and drinking water 

controls. The hemangiosarcomas were 

morphologically similar to spontaneously occurring 

hemangiosarcomas in that there were atypical 

endothelial cells that formed capillary and cavernous 

channels and solid cellular masses with local invasion. 

Two of the four 250 ppm mice with 

hemangiosarcomas also had hemangiosarcomas in 

other tissues; one mouse had hemangiosarcomas in the 

heart and bone marrow, whereas another had 

hemangiosarcomas in the bone marrow and spleen. 

The one 62.5 ppm male mouse with a 

hemangiosarcoma in the liver also had one in the bone 

marrow and one in the spleen. In animals with 

hemangiosarcomas at more than one site, the site of 

origin could not be determined by histologic evaluation. All of these tissues are involved in hematopoiesis and have been shown to be affected by the 

hemolytic activity of 2-butoxyethanol. Because 

hemangiosarcomas are rare neoplasms that were 

found in all exposed groups, but not in chamber 

controls, and because the incidence of the lesion 

exceeded the historical control rates for inhalation 

chamber controls, these neoplasms were considered 

related to 2-butoxyethanol exposure. The 

pathogenesis of this neoplasm in rodents is unknown. 

Although there were statistically significant increases 

in the incidences of hemosiderin pigmentation in 

hepatic Kupffer cells in male mice exposed to 125 or 

250 ppm, the severities of this lesion were considered 

minimal. The incidences of pigmentation were increased in female mice at all exposure concentrations, 

and 2-butoxyethanol exposure did not result in 

increased incidences of hemangiosarcoma in female 

mice. In the four male mice exposed to 250 ppm that 

had hemangiosarcomas, only three of the four had 

hemosiderin pigmentation in the liver. An association 

of hemosiderin deposition in the liver and liver 

neoplasms (adenomas, carcinomas, or hemangiosarcomas) was not found in the 79 male and 

103 female mice from the 2-year NTP studies in 

which the liver was a site of chemical-related neoplasms. At least for male mice, it does not appear 

that an accumulation of hemosiderin and possible related oxidative stress alone were the cause of the liver 

neoplasms. Exposure of mice to 2-butoxyethanol 

caused a significant concentration-related increase in 

the incidences of hematopoietic cell proliferation and 

hemosiderin deposition in the spleen; however, it did 

not cause increases in neoplasms in the spleen as one 

might expect if there were an association of 

hemosiderin accumulation and neoplasm response. 

2-Butoxyethanol exposure caused a concentrationrelated increase in the incidences of hepatocellular 

carcinoma in male mice that was significant in the 

250 ppm group. However, this incidence (43%) was 

within the historical control range for inhalation 

studies (11% to 48%). The incidences of hepatocellular adenoma or carcinoma (combined) were not 

increased in exposed groups of male mice and were 

within the historical control range for inhalation 

studies (20% to 86%). Hepatocellular adenomas and 

carcinomas are relatively common neoplasms in male 

B6C3F1 mice; the morphology of the neoplasms was 

similar in exposed and chamber control mice. As discussed previously, the only exposure-related nonneoplastic effect in the liver was hemosiderin pigmentation in the Kupffer cells. In addition, in female 
 

 

 

2-Butoxyethanol, NTP TR 484 85 

mice exposed to 125 or 250 ppm there were actually 

decreases in the incidences of hepatocellular 

neoplasms. In assessing potential chemical-related 

increases in the incidences of liver neoplasms, benign 

and malignant neoplasms are routinely analyzed 

independently and in combination. Although the 

independent analysis provides useful information, the 

most important analysis for determining potential 

chemical-related effects in the liver is the combined 

analysis of hepatocellular adenoma and carcinoma. 

The principal advantage of the independent analysis is 

for evaluation of progression to a more malignant 

state which may occur along with an overall increase 

in the incidence of neoplasms or in the absence of an 

overall increase (as in this study). Whether the 

21 hepatocellular carcinomas in male mice exposed to 

250 ppm versus the 10 in the chamber control group 

and the positive trend in the incidences of 

hepatocellular carcinoma in male mice in general 

were caused by exposure to 2-butoxyethanol is 

uncertain. 

CONCLUSIONS 

Under the conditions of these 2-year inhalation 

studies, there was no evidence of carcinogenic 

activity* of 2-butoxyethanol in male F344/N rats 

exposed to 31.2, 62.5, or 125 ppm. There was 

equivocal evidence of carcinogenic activity of 

2-butoxyethanol in female F344/N rats based on the 

increased combined incidences of benign or malignant 

pheochromocytoma (mainly benign) of the adrenal 

medulla. There was some evidence of carcinogenic 

activity of 2-butoxyethanol in male B6C3F1 mice 

based on increased incidences of hemangiosarcoma of 

the liver. A marginal increase in the incidences of 

forestomach squamous cell papilloma and an increase 

in the incidences of hepatocellular carcinoma may 

have been exposure related. There was some 

evidence of carcinogenic activity of 2-butoxyethanol 

in female B6C3F1 mice based on increased incidences 

of forestomach squamous cell papilloma or carcinoma 

(mainly papilloma). 

Increased incidences of forestomach neoplasms in 

male and female mice occurred in groups in which 

ulceration and hyperplasia were also present. 

Exposure to 2-butoxyethanol caused a mild regenerative anemia and effects secondary to the anemia. 

* Explanation of Levels of Evidence of Carcinogenic Activity is on page 10. A summary of the Technical Reports Review Subcommittee 

comments and public discussion on this Technical Report appears on page 12. 
86 2-Butoxyethanol, NTP TR 484 
 

87 

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The Aldrich Library of Infrared Spectra (1981). 

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Chemical Company, Inc., Milwaukee, WI. 

The Aldrich Library of NMR Spectra (1983). 2nd ed. 

(C.J. Pouchert, Ed.), p. 191. Aldrich Chemical 

Company, Inc., Milwaukee, WI. 

American Conference of Governmental Industrial 

Hygienists (ACGIH) (1999). 1999 Threshold Limit 

Values for Chemical Substances and Physical Agents 

and Biological Exposure Indices. ACGIH, 

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2-ethoxyethanol and 2-ethoxyethyl acetate and the 

genotoxic effects of these glycol ethers. Int. Arch. 

Occup. Environ. Health 64, 479-484. 

Straus, D.S. (1981). Somatic mutation, cellular differentiation, and cancer causation. JNCI 67, 233-241. 

Tarone, R.E. (1975). Tests for trend in life table 

analysis. Biometrika 62, 679-682. 

Tennant, R.W., and Ashby, J. (1991). Classification 

according to chemical structure, mutagenicity to 

Salmonella and level of carcinogenicity of a further 

39 chemicals tested for carcinogenicity by the U.S. 

National Toxicology Program. Mutat. Res. 257, 

209-227. 

Tennant, R.W., Margolin, B.H., Shelby, M.D., 

Zeiger, E., Haseman, J.K., Spalding, J., 

Caspary, W., Resnick, M., Stasiewicz, S., 

Anderson, B., and Minor, R. (1987). Prediction of 

chemical carcinogenicity in rodents from in vitro 

genetic toxicity assays. Science 236, 933-941. 
 

 

 

94 2-Butoxyethanol, NTP TR 484 

Tyl, R.W., Millicovsky, G., Dodd, D.E., 

Pritts, I.M., France, K.A., and Fisher, L.C. (1984). 

Teratologic evaluation of ethylene glycol monobutyl 

ether in Fischer 344 rats and New Zealand white 

rabbits following inhalation exposure. Environ. 

Health Perspect. 57, 47-68. 

Tyler, T.R. (1984). Acute and subchronic toxicity of 

ethylene glycol monobutyl ether. Environ. Health 

Perspect. 57, 185-191. 

Udden, M.M. (1994). Hemolysis and deformability 

of erythrocytes exposed to butoxyacetic acid, a 

metabolite of 2-butoxyethanol: II. Resistance in red 

blood cells from humans with potential susceptibility. 

J. Appl. Toxicol. 14, 97-102. 

Udden, M.M., and Patton, C.S. (1994). Hemolysis 

and deformability of erythrocytes exposed to 

butoxyacetic acid, a metabolite of 2-butoxyethanol: 

I. Sensitivity in rats and resistance in normal humans. 

J. Appl. Toxicol. 14, 91-96. 

United States International Trade Commission 

(USITC) (1992). Synthetic Organic Chemicals. 

United States Production and Sales, 1992. 

Investigation 332-135: Report of the U.S. International Trade Commission on Domestic Production 

and Sales of Synthetic Organic Chemicals and Raw 

Materials from Which They Are Made. USITC 

Publication No. 2720. U.S International Trade 

Commission, Washington, DC. 

Veulemans, H., Groeseneken, D., Masschelein, R., 

and Van Vuem, E. (1987). Survey of ethylene glycol 

ether exposures in Belgian industries and workshops. 

Am. Ind. Hyg. Assoc. J. 48, 671-676. 

Vincent, R., Cicolella, A., Subra, I., Rieger, B., 

Poirot, P., and Pierre, F. (1993). Occupational 

exposure to 2-butoxyethanol for workers using 

window cleaning agents. Appl. Occup. Environ. Hyg. 

8, 580-586. 

Ward, S., Blair, P.C., and Ghanayem, B.I. (1989). 

Hematologic effects of 2-butoxyethanol (BE) in vivo 

and its effects on the morphology of rat erythrocytes. 

Toxicologist 9, 288 (Abstr.). 

Werner, H.W., Mitchell, J.L., Miller, J.W., and 

von Oettingen, W.F. (1943a). Effects of repeated 

exposure of dogs to monoalkyl ethylene glycol ether 

vapors. J. Ind. Hyg. Toxicol. 25, 409-414. 

Werner, H.W., Mitchell, J.L., Miller, J.W., and 

von Oettingen, W.F. (1943b). The acute toxicity of 

vapors of several monoalkyl ethers of ethylene glycol. 

J. Ind. Hyg. Toxicol. 25, 157-163. 

Werner, H.W., Nawrocki, C.A., Mitchell, J.L., 

Miller, J.W., and von Oettingen, W.F. (1943c). 

Effects of repeated exposures of rats to vapors of 

monoalkyl ethylene glycol ethers. J. Ind. Hyg. 

Toxicol. 25, 374-379. 

Wier, P.J., Lewis, S.C., and Traul, K.A. (1987). A 

comparison of developmental toxicity evident at term 

to postnatal growth and survival using ethylene glycol 

monoethyl ether, ethylene glycol monobutyl ether, 

and ethanol. Teratog. Carcinog. Mutagen. 7, 55-64. 

Williams, D.A. (1971). A test for differences 

between treatment means when several dose levels are 

compared with a zero dose control. Biometrics 27, 

103-117. 

Williams, D.A. (1972). The comparison of several 

dose levels with a zero dose control. Biometrics 28, 

519-531. 

Zeiger, E., Haseman, J.K., Shelby, M.D., 

Margolin, B.H., and Tennant, R.W. (1990). Evaluation of four in vitro genetic toxicity tests for 

predicting rodent carcinogenicity: Confirmation of 

earlier results with 41 additional chemicals. Environ. 

Mol. Mutagen. 16 (Suppl. 18), 1-14. 

Zeiger, E., Anderson, B., Haworth, S., Lawlor, T., 

and Mortelmans, K. (1992). Salmonella mutagenicity 

tests: V. Results from the testing of 311 chemicals. 

Environ. Mol. Mutagen. 19 (Suppl. 21), 2-141. 
95 

APPENDIX A 

SUMMARY OF LESIONS IN MALE RATS 

IN THE 2-YEAR INHALATION STUDY 

OF 2-BUTOXYETHANOL 

TABLE A1 Summary of the Incidence of Neoplasms in Male Rats 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 96 

TABLE A2 Individual Animal Tumor Pathology of Male Rats 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 100 

TABLE A3 Statistical Analysis of Primary Neoplasms in Male Rats 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 122 

TABLE A4 Summary of the Incidence of Nonneoplastic Lesions in Male Rats 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 128 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

96 2-Butoxyethanol, NTP TR 484 

TABLE A1 

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 25 31 24 21 

Natural deaths 6 8 5 5 

Survivors 

Terminal sacrifice 19 11 21 24 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Esophagus (50) (50) (50) (50) 

Intestine large, colon (47) (47) (47) (48) 

Intestine large, cecum (46) (47) (47) (46) 

Intestine small, duodenum (46) (47) (49) (49) 

Intestine small, ileum (45) (45) (46) (47) 

Liver (50) (50) (50) (50) 

Carcinoma, metastatic, islets, pancreatic 1 (2%) 

Hemangiosarcoma, metastatic, spleen 1 (2%) 

Hepatocellular carcinoma 1 (2%) 1 (2%) 

Hepatocellular adenoma 1 (2%) 2 (4%) 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Sarcoma, metastatic, uncertain primary site 1 (2%) 

Mesentery (8) (6) (15) (8) 

Hemangiosarcoma, metastatic, spleen 1 (7%) 

Sarcoma 1 (7%) 

Sarcoma, metastatic, spleen 1 (7%) 

Oral mucosa (3) (1) (1) 

Gingival, fibrosarcoma 1 (100%) 

Pharyngeal, squamous cell papilloma 3 (100%) 

Pancreas (50) (50) (50) (50) 

Adenoma 1 (2%) 3 (6%) 

Salivary glands (50) (50) (50) (50) 

Stomach, forestomach (50) (50) (50) (50) 

Stomach, glandular (49) (49) (50) (50) 

Tongue (1) (1) 

Squamous cell papilloma 1 (100%) 1 (100%) 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Endocrine System 

Adrenal cortex (50) (50) (49) (50) 

Adenoma 2 (4%) 1 (2%) 

Carcinoma 1 (2%) 

Adrenal medulla (50) (50) (49) (50) 

Pheochromocytoma malignant 2 (4%) 3 (6%) 

Pheochromocytoma benign 12 (24%) 15 (30%) 9 (18%) 12 (24%) 

Bilateral, pheochromocytoma benign 3 (6%) 5 (10%) 4 (8%) 4 (8%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 97 

TABLE A1 

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Endocrine System (continued) 

Islets, pancreatic (50) (50) (50) (50) 

Adenoma 5 (10%) 2 (4%) 3 (6%) 3 (6%) 

Adenoma, multiple 1 (2%) 1 (2%) 

Carcinoma 4 (8%) 1 (2%) 3 (6%) 

Pituitary gland (49) (49) (48) (47) 

Pars distalis, adenoma 24 (49%) 29 (59%) 24 (50%) 25 (53%) 

Thyroid gland (50) (46) (48) (48) 

C-cell, adenoma 7 (14%) 3 (7%) 7 (15%) 10 (21%) 

C-cell, carcinoma 1 (2%) 3 (6%) 2 (4%) 

Follicular cell, adenoma 1 (2%) 3 (6%) 

Follicular cell, carcinoma 1 (2%) 1 (2%) 

General Body System 

Peritoneum (2) (1) 

Tissue NOS (1) 

Sarcoma 1 (100%) 

Genital System 

Epididymis (50) (50) (50) (50) 

Preputial gland (46) (47) (50) (43) 

Adenoma 3 (7%) 1 (2%) 5 (10%) 5 (12%) 

Carcinoma 5 (11%) 1 (2%) 2 (4%) 1 (2%) 

Prostate (49) (49) (50) (50) 

Adenoma 1 (2%) 4 (8%) 1 (2%) 

Adenoma, multiple 1 (2%) 

Seminal vesicle (47) (44) (46) (48) 

Adenoma 1 (2%) 

Testes (50) (50) (50) (50) 

Bilateral, interstitial cell, adenoma 34 (68%) 38 (76%) 36 (72%) 31 (62%) 

Interstitial cell, adenoma 7 (14%) 4 (8%) 8 (16%) 10 (20%) 

Hematopoietic System 

Bone marrow (50) (50) (50) (49) 

Lymph node (10) (10) (8) (10) 

Sarcoma, metastatic, skin 1 (10%) 

Lymph node, bronchial (36) (38) (41) (34) 

Lymph node, mandibular (44) (47) (47) (46) 

Lymph node, mesenteric (50) (49) (50) (50) 

Hemangiosarcoma, metastatic, spleen 1 (2%) 

Sarcoma, metastatic, spleen 1 (2%) 

Lymph node, mediastinal (44) (44) (45) (44) 

Hemangiosarcoma, metastatic, spleen 1 (2%) 

Spleen (50) (50) (50) (50) 

Fibroma 1 (2%) 1 (2%) 

Hemangiosarcoma 1 (2%) 

Sarcoma 1 (2%) 1 (2%) 

Thymus (45) (44) (44) (39) 

Thymoma malignant 1 (2%) 1 (3%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

98 2-Butoxyethanol, NTP TR 484 

TABLE A1 

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Integumentary System 

Mammary gland (44) (40) (44) (45) 

Carcinoma 1 (3%) 

Fibroadenoma 2 (5%) 1 (3%) 3 (7%) 1 (2%) 

Fibroadenoma, multiple 1 (2%) 

Skin (50) (50) (50) (50) 

Basal cell adenoma 2 (4%) 1 (2%) 

Basal cell carcinoma 2 (4%) 1 (2%) 

Keratoacanthoma 3 (6%) 3 (6%) 4 (8%) 2 (4%) 

Keratoacanthoma, multiple 1 (2%) 

Squamous cell papilloma 1 (2%) 1 (2%) 4 (8%) 1 (2%) 

Sebaceous gland, adenoma 2 (4%) 

Subcutaneous tissue, fibroma 2 (4%) 2 (4%) 2 (4%) 3 (6%) 

Subcutaneous tissue, fibroma, multiple 1 (2%) 

Subcutaneous tissue, fibrosarcoma 1 (2%) 

Subcutaneous tissue, fibrosarcoma, multiple 1 (2%) 

Subcutaneous tissue, hemangiosarcoma 1 (2%) 

Subcutaneous tissue, lipoma 1 (2%) 1 (2%) 2 (4%) 

Subcutaneous tissue, sarcoma 2 (4%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Osteosarcoma 1 (2%) 1 (2%) 

Skeletal muscle (1) 

Sarcoma 1 (100%) 

Nervous System 

Brain (50) (50) (50) (50) 

Astrocytoma benign 1 (2%) 

Astrocytoma malignant 1 (2%) 

Carcinoma, metastatic, Zymbal’s gland 1 (2%) 

Glioma malignant 2 (4%) 

Meningioma malignant 1 (2%) 

Respiratory System 

Lung (50) (50) (50) (50) 

Alveolar/bronchiolar adenoma 1 (2%) 2 (4%) 4 (8%) 

Alveolar/bronchiolar carcinoma 1 (2%) 1 (2%) 1 (2%) 

Carcinoma, metastatic, islets, pancreatic 1 (2%) 

Carcinoma, metastatic, preputial gland 1 (2%) 

Hemangiosarcoma, metastatic, spleen 1 (2%) 

Histiocytic sarcoma 1 (2%) 1 (2%) 

Osteosarcoma, metastatic, uncertain

 primary site 1 (2%) 1 (2%) 

Pheochromocytoma malignant, metastatic,

 adrenal medulla 1 (2%) 1 (2%) 

Sarcoma, metastatic, skin 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 99 

TABLE A1 

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Respiratory System (continued) 

Nose (48) (49) (49) (50) 

Lateral wall, adenoma 1 (2%) 

Turbinate, chondroma 1 (2%) 

Pleura (1) 

Trachea (50) (49) (50) (48) 

Special Senses System 

Zymbal’s gland (1) (1) (2) 

Carcinoma 1 (100%) 1 (100%) 2 (100%) 

Urinary System 

Kidney (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 

Mesenchymal tumor malignant 1 (2%) 

Sarcoma 1 (2%) 

Renal tubule, adenoma 1 (2%) 2 (4%) 

Renal tubule, oncocytoma benign 1 (2%) 

Transitional epithelium, carcinoma 1 (2%) 

Urinary bladder (49) (50) (50) (50) 

Transitional epithelium, papilloma 2 (4%) 

Systemic Lesions 

Multiple organsb (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 1 (2%) 

Leukemia mononuclear 29 (58%) 31 (62%) 33 (66%) 30 (60%) 

Lymphoma malignant 1 (2%) 

Mesothelioma benign 1 (2%) 1 (2%) 1 (2%) 1 (2%) 

Mesothelioma malignant 3 (6%) 1 (2%) 

Neoplasm Summary 

Total animals with primary neoplasmsc 50 50 50 50 

Total primary neoplasms 173 161 187 163 

Total animals with benign neoplasms 49 49 49 49 

Total benign neoplasms 123 114 131 119 

Total animals with malignant neoplasms 40 40 41 38 

Total malignant neoplasms 50 47 56 44 

Total animals with metastatic neoplasms 3 3 3 2 

Total metastatic neoplasms 4 3 8 3 

Total animals with malignant neoplasms

 of uncertain primary site 2 1 

a Number of animals examined microscopically at the site and the number of animals with neoplasm b Number of animals with any tissue examined microscopically 

Primary neoplasms: all neoplasms except metastatic neoplasms c 
 

 

 

 

 

 

 

 

 

 

 

 

 

100 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

3555555555566666666666666 

Number of Days on Study 5222556666901133444444788 

4019883458790718126889337 

0000000000000000000000000 

Carcass ID Number 3441242320010421113035443 

0943431502744667203640701 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Intestine large, colon +A+A+++++A+++++++++++++++ 

Intestine large, rectum +A+++++++A+++++++++++++++ 

Intestine large, cecum + A + A + + A + + A +++++++++++++++ 

Intestine small, duodenum + A + A +++++A++++++A++++++++ 

Intestine small, jejunum + A + A +++++A+A++++A++++++++ 

Intestine small, ileum +A+A++A++A++++++A++++++++ 

Liver +++++++++++++++++++++++++ 

Carcinoma, metastatic, islets, pancreatic X 
 

Hepatocellular adenoma
 

Sarcoma, metastatic, uncertain primary site X
 

Mesentery + + + + + 

Oral mucosa ++ + 

Pharyngeal, squamous cell papilloma X X X 

Pancreas +++++++++++++++++++++++++ 

Adenoma 

Salivary glands +++++++++++++++++++++++++ 

Stomach, forestomach +++++++++++++++++++++++++ 

Stomach, glandular +++++++++++++++++++++++++ 

Tongue 

Squamous cell papilloma 

Tooth + + 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adenoma X 

Adrenal medulla +++++++++++++++++++++++++ 

Pheochromocytoma benign X X X X 

Bilateral, pheochromocytoma benign X X 

Islets, pancreatic +++++++++++++++++++++++++ 

Adenoma X 

Carcinoma X X 

Parathyroid gland +++++++++++++++++++++++++ 

Pituitary gland +++++++++++++++++++++++++ 

Pars distalis, adenoma XX XX X X XXXXXX X 

Thyroid gland +++++++++++++++++++++++++ 

C-cell, adenoma X 

C-cell, carcinoma 

Follicular cell, adenoma 

Follicular cell, carcinoma 

General Body System 

None 

+: Tissue examined microscopically M: Missing tissue X: Lesion present 

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 101 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

6677777777777777777777777 

Number of Days on Study 8900012222222222222223333 

7728889999999999999990000 

0000000000000000000000000 Total 

Carcass ID Number 4232300001112223334440112 Tissues/ 

5563281356892897891289157 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Intestine large, colon +++++++++++++++++++++++++ 47 

Intestine large, rectum +++++ I +++++++++++++++++ I + 46 

Intestine large, cecum +++++++++++++++++++++++++ 46 

Intestine small, duodenum +++++++++++++++++++++++++ 46 

Intestine small, jejunum +++++++++++++++++++++++++ 45 

Intestine small, ileum +++++++++++++++++++++++++ 45 

Liver +++++++++++++++++++++++++ 50 

Carcinoma, metastatic, islets, pancreatic 1 

Hepatocellular adenoma X 1 

Sarcoma, metastatic, uncertain primary site 1 

Mesentery + ++ 8 

Oral mucosa 3 

Pharyngeal, squamous cell papilloma 3 

Pancreas +++++++++++++++++++++++++ 50 

Adenoma X 1 

Salivary glands +++++++++++++++++++++++++ 50 

Stomach, forestomach +++++++++++++++++++++++++ 50 

Stomach, glandular ++++++++++++++++++++++ ++ 49 

Tongue + 1 

Squamous cell papilloma X 1 

Tooth + 3 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 50 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 50 

Adenoma X 2 

Adrenal medulla +++++++++++++++++++++++++ 50 

Pheochromocytoma benign X X X X X X X X 12 

Bilateral, pheochromocytoma benign X 3 

Islets, pancreatic +++++++++++++++++++++++++ 50 

Adenoma X X X X 5 

Carcinoma X X 4 

Parathyroid gland +++++++++++++M++M+++++++M 47 

Pituitary gland ++++++++++++++++++++++ ++ 49 

Pars distalis, adenoma XXXXX XX XXX X 24 

Thyroid gland +++++++++++++++++++++++++ 50 

C-cell, adenoma X X X X X X 7 

C-cell, carcinoma X 1 

Follicular cell, adenoma X 1 

Follicular cell, carcinoma X 1 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

102 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

3555555555566666666666666 

Number of Days on Study 5222556666901133444444788 

4019883458790718126889337 

0000000000000000000000000 

Carcass ID Number 3441242320010421113035443 

0943431502744667203640701 

Genital System 

Epididymis +++++++++++++++++++++++++ 

Preputial gland +++++++++++++++++M+++++++ 

Adenoma X X 

Carcinoma X 

Prostate +++++++++++++++++++++++++ 

Adenoma 

Seminal vesicle +++++++++A++++++A++++++++ 

Adenoma 

Testes +++++++++++++++++++++++++ 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X 

Interstitial cell, adenoma X X X X 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 

Lymph node + + + + + + 

Lymph node, bronchial + M +++++++++MM+M+M+M++MM++ 

Lymph node, mandibular +++++++++++++++++MM++++++ 

Lymph node, mesenteric +++++++++++++++++++++++++ 

Lymph node, mediastinal +++++++++++++++M+++++++M+ 

Spleen +++++++++++++++++++++++++ 

Thymus ++++++++M++++M+++++++++++ 

Integumentary System 

Mammary gland ++++++++M++++++++++++++++ 

Fibroadenoma X X 

Fibroadenoma, multiple 

Skin +++++++++++++++++++++++++ 

Basal cell adenoma 

Basal cell carcinoma 

Keratoacanthoma X 

Squamous cell papilloma 

Subcutaneous tissue, fibroma X 

Subcutaneous tissue, fibrosarcoma, multiple X 

Subcutaneous tissue, hemangiosarcoma X 

Subcutaneous tissue, lipoma 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Osteosarcoma 

Nervous System 

Brain +++++++++++++++++++++++++ 

Glioma malignant X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 103 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

6677777777777777777777777 

Number of Days on Study 8900012222222222222223333 

7728889999999999999990000 

0000000000000000000000000 Total 

Carcass ID Number 4232300001112223334440112 Tissues/ 

5563281356892897891289157 Tumors 

Genital System 

Epididymis +++++++++++++++++++++++++ 50 

Preputial gland ++++++M++++++M++M++++++++ 46 

Adenoma X 3 

Carcinoma X X X X 5 

Prostate ++++++++++++++++++++++ ++ 49 

Adenoma X 1 

Seminal vesicle ++++++++++++++++++++++ ++ 47 

Adenoma X 1 

Testes +++++++++++++++++++++++++ 50 

Bilateral, interstitial cell, adenoma X X X XXXXXXXXXXXXXXX XXXX 34 

Interstitial cell, adenoma X X X 7 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 50 

Lymph node + + + + 10 

Lymph node, bronchial + M +++++++M+++++++++M+MM+M 36 

Lymph node, mandibular +++M+++++++++++M+++++++MM 44 

Lymph node, mesenteric +++++++++++++++++++++++++ 50 

Lymph node, mediastinal +++++++M++++++++M+++M++M+ 44 

Spleen +++++++++++++++++++++++++ 50 

Thymus +MM++++++++++++++++M+++++ 45 

Integumentary System 

Mammary gland ++M++++M+++M+++++++M++++M 44 

Fibroadenoma 2 

Fibroadenoma, multiple X 1 

Skin +++++++++++++++++++++++++ 50 

Basal cell adenoma X X 2 

Basal cell carcinoma X X 2 

Keratoacanthoma X X 3 

Squamous cell papilloma X 1 

Subcutaneous tissue, fibroma X 2 

Subcutaneous tissue, fibrosarcoma, multiple 1 

Subcutaneous tissue, hemangiosarcoma 1 

Subcutaneous tissue, lipoma X 1 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 50 

Osteosarcoma X 1 

Nervous System 

Brain +++++++++++++++++++++++++ 50 

Glioma malignant X 2 
 

 

 

 

 

 

 

 

104 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

3555555555566666666666666 

Number of Days on Study 5222556666901133444444788 

4019883458790718126889337 

0000000000000000000000000 

Carcass ID Number 3441242320010421113035443 

0943431502744667203640701 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Carcinoma, metastatic, islets, pancreatic 

Histiocytic sarcoma 

Osteosarcoma, metastatic, uncertain primary site 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

X 

+A+++++++A+++++++++++++++ 

+++++++++++++++++++++++++ 

Special Senses System 

Eye 

Zymbal’s gland 

Carcinoma

+ 

X 

+ 

Urinary System 

Kidney 

Renal tubule, adenoma 

Urinary bladder 

Transitional epithelium, papilloma 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Leukemia mononuclear 

Lymphoma malignant 

Mesothelioma benign 

+++++++++++++++++++++++++ 

X 

X X X X X X X XXXXXX X X 

X 

X 
 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 105 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

6677777777777777777777777 

Number of Days on Study 8900012222222222222223333 

7728889999999999999990000 

0000000000000000000000000 Total 

Carcass ID Number 4232300001112223334440112 

5563281356892897891289157 

Tissues/ 

Tumors 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Carcinoma, metastatic, islets, pancreatic 

Histiocytic sarcoma 

Osteosarcoma, metastatic, uncertain primary site 

Nose 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

X 

+++++++++++++++++++++++++ 

50 

50 

1 

1 

1 

1 

48 

Trachea +++++++++++++++++++++++++ 50 

Special Senses System 

Eye 

Zymbal’s gland 

Carcinoma 

+ 2 

1 

1 

Urinary System 

Kidney 

Renal tubule, adenoma 

Urinary bladder 

Transitional epithelium, papilloma 

+++++++++++++++++++++++++ 

X 

++++++++++++++++++++++ ++ 

X 

50 

1 

49 

2 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Leukemia mononuclear 

+++++++++++++++++++++++++ 

X XXXXXX XX XX XX X 

50 

1 

29 

Lymphoma malignant 

Mesothelioma benign 

1 

1 
 

 

 

 

 

 

 

 

 

 

106 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 31.2 ppm 

4455555555556666666666666 

Number of Days on Study 6723334677791112233344555 

5993364156920770703869222 

2222222222222222222222222 

Carcass ID Number 3532443410314022120040113 

6074052728982529462431793 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Intestine large, colon ++A+++++++++++++++++++++A 

Intestine large, rectum ++A++++++++A+++++++++++++ 

Intestine large, cecum + + + + + + + + + + A A + + + + + + + + + + + + + 

Intestine small, duodenum +++++++++++++A++++++++++A 

Intestine small, jejunum + + + + + + + + + + A A + A + + + + + + + + + + A 

Intestine small, ileum ++++++++++ A A + A ++++++++++ A 

Liver +++++++++++++++++++++++++ 

Hepatocellular carcinoma 

Hepatocellular adenoma X 

Mesentery + + + 

Oral mucosa + 

Gingival, fibrosarcoma X 

Pancreas +++++++++++++++++++++++++ 

Salivary glands +++++++++++++++++++++++++ 

Stomach, forestomach +++++++++++++++++++++++++ 

Stomach, glandular +++++++++++A+++++++++++++ 

Tooth + 

Cardiovascular System 

Blood vessel 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adrenal medulla +++++++++++++++++++++++++ 

Pheochromocytoma malignant X 

Pheochromocytoma benign X X X 

Bilateral, pheochromocytoma benign X 

Islets, pancreatic +++++++++++++++++++++++++ 

Adenoma X 

Carcinoma 

Parathyroid gland ++M+++M+++++++++++++++++A 

Pituitary gland ++++++++++++++++++++++++A 

Pars distalis, adenoma XXXXXXXXX X X XXXX X 

Thyroid gland ++++++++++ A A ++++++++++++ A 

C-cell, adenoma X 

General Body System 

Tissue NOS + 

Sarcoma X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 107 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 31.2 ppm 

6666666777777777777777777 

Number of Days on Study 5667789000122222222222233 

9464783244623399999999900 

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 Total 

Carcass ID Number 2 2 4 1 3 3 0 3 0 1 4 1 1 2 0 0 2 2 2 3 4 4 4 1 3 Tissues/ 

1 7 4 3 0 8 7 4 3 5 6 6 1 5 6 9 0 3 8 5 1 8 9 0 1 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + A + + + + + + + + + + + + + + + + 47 

Intestine large, rectum + + + + + + + + + + + I + + + + + + + + + + + + + 47 

Intestine large, cecum + + + + + + + + A + + + + + + + + + + + + + + + + 47 

Intestine small, duodenum + + + + + + + + A + + + + + + + + + + + + + + + + 47 

Intestine small, jejunum + + + + + + + + A + + + + + + + + + + + + + + + + 45 

Intestine small, ileum + + + + + + + + A + + + + + + + + + + + + + + + + 45 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hepatocellular carcinoma X 1 

Hepatocellular adenoma X 2 

Mesentery + + + 6 

Oral mucosa 1 

Gingival, fibrosarcoma 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Tooth 1 

Cardiovascular System 

Blood vessel + + 2 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma malignant X 2 

Pheochromocytoma benign X X X X X X X X X X X X 15 

Bilateral, pheochromocytoma benign X X X X 5 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 2 

Carcinoma X 1 

Parathyroid gland + + + + + + + + + + + + + + + + + + + + + + M + + 46 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Pars distalis, adenoma X X X X X X X X X X X X X 29 

Thyroid gland + + + + + + + + A + + + + + + + + + + + + + + + + 46 

C-cell, adenoma X X 3 

General Body System 

Tissue NOS 1 

Sarcoma 1 
 

 

 

 

 

 

 

 

108 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 31.2 ppm 

4455555555556666666666666 

Number of Days on Study 6723334677791112233344555 

5993364156920770703869222 

2222222222222222222222222 

Carcass ID Number 3532443410314022120040113 

6074052728982529462431793 

Genital System 

Epididymis +++++++++++++++++++++++++ 

Preputial gland ++++M+++++++++M++++++++++ 

Adenoma 

Carcinoma 

Prostate +++++++++++A+++++++++++++ 

Seminal vesicle ++ A +++++++ A A +++++++++ A ++ A 

Testes +++++++++++++++++++++++++ 

Bilateral, interstitial cell, adenoma X X XXX X XXXXXXX X X 

Interstitial cell, adenoma X X 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 

Lymph node + + + + + + 

Lymph node, bronchial M + M +++M+++++++M+M+MM+++++ 

Lymph node, mandibular ++++M+++++++++M++++++++++ 

Lymph node, mesenteric +++++++++++++++++++++++++ 

Lymph node, mediastinal +++++++++++++++++++++M+++ 

Spleen +++++++++++++++++++++++++ 

Thymus ++M++++++M+++++++++++++M+ 

Integumentary System 

Mammary gland +++++++M+++++++++++++M+M+ 

Carcinoma 

Fibroadenoma 

Skin +++++++++++++++++++++++++ 

Basal cell adenoma 

Basal cell carcinoma X 

Keratoacanthoma X X 

Squamous cell papilloma 

Subcutaneous tissue, fibroma X 

Subcutaneous tissue, lipoma X 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Skeletal muscle 

Sarcoma 

Nervous System 

Brain +++++++++++++++++++++++++ 

Astrocytoma malignant 

Carcinoma, metastatic, Zymbal’s gland X 

Meningioma malignant X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 109 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 31.2 ppm 

6666666777777777777777777 

Number of Days on Study 5667789000122222222222233 

9464783244623399999999900 

2222222222222222222222222 Total 

Carcass ID Number 2241330301411200222344413 Tissues/ 

1743087435661569038518901 Tumors 

Genital System 

Epididymis +++++++++++++++++++++++++ 50 

Preputial gland ++++++++++ ++++++++++++++ 47 

Adenoma X 1 

Carcinoma X 1 

Prostate +++++++++++++++++++++++++ 49 

Seminal vesicle ++++++++A++++++++++++++++ 44 

Testes +++++++++++++++++++++++++ 50 

Bilateral, interstitial cell, adenoma XXXXX XXXX XXXXXXXXXXXXXX 38 

Interstitial cell, adenoma X X 4 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 50 

Lymph node + + + + 10 

Lymph node, bronchial ++++M+++M++++++++MM++M+++ 38 

Lymph node, mandibular ++++++++++++M++++++++++++ 47 

Lymph node, mesenteric +++++++++++++++++M+++++++ 49 

Lymph node, mediastinal + + M + M + + M +++++++++++++M++M 44 

Spleen +++++++++++++++++++++++++ 50 

Thymus M+++++++++M++++++++++++M+ 44 

Integumentary System 

Mammary gland ++M+++M++M++MM+ I +++M+++++ 40 

Carcinoma X 1 

Fibroadenoma X 1 

Skin +++++++++++++++++++++++++ 50 

Basal cell adenoma X 1 

Basal cell carcinoma 1 

Keratoacanthoma X 3 

Squamous cell papilloma X 1 

Subcutaneous tissue, fibroma X 2 

Subcutaneous tissue, lipoma 1 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 50 

Skeletal muscle + 1 

Sarcoma X 1 

Nervous System 

Brain +++++++++++++++++++++++++ 50 

Astrocytoma malignant X 1 

Carcinoma, metastatic, Zymbal’s gland 1 

Meningioma malignant 1 
 

 

 

 

 

110 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 31.2 ppm 

4455555555556666666666666 

Number of Days on Study 6723334677791112233344555 

5993364156920770703869222 

2222222222222222222222222 

Carcass ID Number 3532443410314022120040113 

6074052728982529462431793 

Respiratory System 

Larynx +++++++++++A+++++++++++++ 

Lung +++++++++++++++++++++++++ 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Osteosarcoma, metastatic,

 uncertain primary site 

Pheochromocytoma malignant, metastatic, 

adrenal medulla X 

Nose +++++++++++A+++++++++++++ 

Turbinate, chondroma 

Trachea +++++++++++A+++++++++++++ 

Special Senses System 

Zymbal’s gland + 

Carcinoma X 

Urinary System 

Kidney +++++++++++++++++++++++++ 

Sarcoma 

Renal tubule, adenoma X 

Transitional epithelium, carcinoma 

Urinary bladder +++++++++++++++++++++++++ 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 

Leukemia mononuclear X X X XXXXX XXXX X XXX 

Mesothelioma benign X 
 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 111 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 31.2 ppm 

6666666777777777777777777 

Number of Days on Study 5667789000122222222222233 

9464783244623399999999900 

2222222222222222222222222 Total 

Carcass ID Number 2241330301411200222344413 

1743087435661569038518901 

Tissues/ 

Tumors 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Osteosarcoma, metastatic, 

uncertain primary site 

Pheochromocytoma malignant, metastatic, 

adrenal medulla 

Nose 

Turbinate, chondroma 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X 

X 

X 

+++++++++++++++++++++++++ 

X 

+++++++++++++++++++++++++ 

49 

50 

2 

1 

1 

1 

49 

1 

49 

Special Senses System 

Zymbal’s gland 

Carcinoma 

1 

1 

Urinary System 

Kidney 

Sarcoma 

Renal tubule, adenoma 

Transitional epithelium, carcinoma 

Urinary bladder 

+++++++++++++++++++++++++ 

X 

X 

X 

+++++++++++++++++++++++++ 

50 

1 

2 

1 

50 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma benign 

+++++++++++++++++++++++++ 

X X X X X XXXX XX X X XX 

50 

31 

1 
 

 

 

 

 

 

 

 

 

 

 

112 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

3334445555566666666666677 

Number of Days on Study 4786784466811224566778900 

0071242915427274866377413 

4444444444444444444444444 

Carcass ID Number 1240032133111002414132132 

6003928541420643188787952 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Intestine large, colon +A+A+++++++++++++++++++++ 

Intestine large, rectum +A+A+++++++++++++++++++++ 

Intestine large, cecum + A + A +++++++++++++++++++++ 

Intestine small, duodenum + + + A +++++++++++++++++++++ 

Intestine small, jejunum + A + A + + A ++++++++++++++++++ 

Intestine small, ileum +A+A++A++++++++++++++++++ 

Liver +++++++++++++++++++++++++ 

Hemangiosarcoma, metastatic, spleen
 

Histiocytic sarcoma
 

Mesentery + ++ + +++ + 

Hemangiosarcoma, metastatic, spleen 

Sarcoma 

Sarcoma, metastatic, spleen 

Oral mucosa
 

Pancreas +++++++++++++++++++++++++ 
 

Adenoma 

Salivary glands +++++++++++++++++++++++++ 

Stomach, forestomach +++++++++++++++++++++++++ 

Stomach, glandular +++++++++++++++++++++++++ 

Tongue 

Squamous cell papilloma 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++A+++++++++++++++++++++ 

Adenoma X 

Carcinoma X 

Adrenal medulla +++A+++++++++++++++++++++ 

Pheochromocytoma malignant X X 

Pheochromocytoma benign X X X X 

Bilateral, pheochromocytoma benign X X 

Islets, pancreatic +++++++++++++++++++++++++ 

Adenoma X 

Adenoma, multiple 

Parathyroid gland ++++++++++++++++++++M++++ 

Pituitary gland +++A+++++++++++++++++++++ 

Pars distalis, adenoma X XX XX XX X XX X 

Thyroid gland +A+++++++++++++++++++++++ 

C-cell, adenoma X X 

C-cell, carcinoma X 

Follicular cell, carcinoma 

General Body System 

Peritoneum + 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 113 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

7777777777777777777777777 

Number of Days on Study 0011222222222222222233333 

8868999999999999999900000 

4444444444444444444444444 Total 

Carcass ID Number 0312002222333344444500144 Tissues/ 

7614581569037924579012336 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Intestine large, colon ++A++++++++++++++++++++++ 47 

Intestine large, rectum +++++++++++++++++++++++++ 48 

Intestine large, cecum + + A ++++++++++++++++++++++ 47 

Intestine small, duodenum +++++++++++++++++++++++++ 49 

Intestine small, jejunum + + A ++++++++++++++++++++++ 46 

Intestine small, ileum ++A++++++++++++++++++++++ 46 

Liver +++++++++++++++++++++++++ 50 

Hemangiosarcoma, metastatic, spleen X 1 

Histiocytic sarcoma X 1 

Mesentery +++ + ++ + 15 

Hemangiosarcoma, metastatic, spleen X 1 

Sarcoma X 1 

Sarcoma, metastatic, spleen X 1 

Oral mucosa + 1 

Pancreas +++++++++++++++++++++++++ 50 

Adenoma X X X 3 

Salivary glands +++++++++++++++++++++++++ 50 

Stomach, forestomach +++++++++++++++++++++++++ 50 

Stomach, glandular +++++++++++++++++++++++++ 50 

Tongue + 1 

Squamous cell papilloma X 1 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 50 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 49 

Adenoma 1 

Carcinoma 1 

Adrenal medulla +++++++++++++++++++++++++ 49 

Pheochromocytoma malignant X 3 

Pheochromocytoma benign X X X X X 9 

Bilateral, pheochromocytoma benign X X 4 

Islets, pancreatic +++++++++++++++++++++++++ 50 

Adenoma X X 3 

Adenoma, multiple X 1 

Parathyroid gland +++++++++++++++++M++++M++ 47 

Pituitary gland ++++++ I ++++++++++++++++++ 48 

Pars distalis, adenoma X X X X X X X X X X X X X 24 

Thyroid gland ++A++++++++++++++++++++++ 48 

C-cell, adenoma X X X X X 7 

C-cell, carcinoma X X 3 

Follicular cell, carcinoma X 1 

General Body System 

Peritoneum + 2 
 

 

 

 

 

 

 

114 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

3334445555566666666666677 

Number of Days on Study 4786784466811224566778900 

0071242915427274866377413 

4444444444444444444444444 

Carcass ID Number 1240032133111002414132132 

6003928541420643188787952 

Genital System 

Epididymis +++++++++++++++++++++++++ 

Preputial gland +++++++++++++++++++++++++ 

Adenoma X X X 

Carcinoma X 

Prostate +++++++++++++++++++++++++ 

Adenoma 

Seminal vesicle +A+A+++++++++A+++++++++++ 

Testes +++++++++++++++++++++++++ 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X 

Interstitial cell, adenoma X X X X X X 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 

Lymph node + + + + + + + 

Lymph node, bronchial + + M ++++M+MM++++++M+++M+++ 

Lymph node, mandibular +++++++++++++++++++++++++ 

Lymph node, mesenteric +++++++++++++++++++++++++ 

Hemangiosarcoma, metastatic, spleen 

Sarcoma, metastatic, spleen 

Lymph node, mediastinal +++++++++M+++++++++M+++++ 

Hemangiosarcoma, metastatic, spleen 

Spleen +++++++++++++++++++++++++ 

Fibroma 

Hemangiosarcoma 

Sarcoma 

Thymus +++++++++++++++M+M+M+M+++ 

Thymoma malignant 

Integumentary System 

Mammary gland ++M+M+++++++++++++M++++++ 

Fibroadenoma X X 

Skin +++++++++++++++++++++++++ 

Keratoacanthoma X X X 

Squamous cell papilloma 

Sebaceous gland, adenoma 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, fibrosarcoma 

Subcutaneous tissue, lipoma 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Nervous System 

Brain +++++++++++++++++++++++++ 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 115 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

7777777777777777777777777 

Number of Days on Study 0011222222222222222233333 

8868999999999999999900000 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 Total 

Carcass ID Number 0 3 1 2 0 0 2 2 2 2 3 3 3 3 4 4 4 4 4 5 0 0 1 4 4 Tissues/ 

7 6 1 4 5 8 1 5 6 9 0 3 7 9 2 4 5 7 9 0 1 2 3 3 6 Tumors 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X X 5 

Carcinoma X 2 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X XX X 4 

Seminal vesicle + + A + + + + + + + + + + + + + + + + + + + + + + 46 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X X X X X X X X X X X 36 

Interstitial cell, adenoma X X 8 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node + 8 

Lymph node, bronchial + + M +++M+++M++++++++++++++ 41 

Lymph node, mandibular + + + + M + + + + + + + + + + + M + + M + + + + + 47 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma, metastatic, spleen X 1 

Sarcoma, metastatic, spleen X 1 

Lymph node, mediastinal + + + + M + + + + + + + M + + + + + + + + M + + + 45 

Hemangiosarcoma, metastatic, spleen X 1 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Fibroma X 1 

Hemangiosarcoma X 1 

Sarcoma X 1 

Thymus + + + + + I + + + + + + + + + + + + + + + + M + + 44 

Thymoma malignant X 1 

Integumentary System 

Mammary gland + + + + + + + + + M + + + + + + M + + M + + + + + 44 

Fibroadenoma X 3 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Keratoacanthoma X 4 

Squamous cell papilloma X X X X 4 

Sebaceous gland, adenoma X X 2 

Subcutaneous tissue, fibroma X X 2 

Subcutaneous tissue, fibrosarcoma X 1 

Subcutaneous tissue, lipoma X X 2 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 
 

 

 

 

 

 

 

116 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

3334445555566666666666677 

Number of Days on Study 4786784466811224566778900 

0071242915427274866377413 

4444444444444444444444444 

Carcass ID Number 1240032133111002414132132 

6003928541420643188787952 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hemangiosarcoma, metastatic, spleen 

Histiocytic sarcoma 

Pheochromocytoma malignant, metastatic, 

adrenal medulla 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

X 

Nose +++++++++++++++++++++++++ 

Lateral wall, adenoma 

Pleura + 

X 

Trachea +++++++++++++++++++++++++ 

Special Senses System 

Eye 

Zymbal’s gland 

Carcinoma 

+ 

X 

Urinary System 

Kidney 

Histiocytic sarcoma 

Mesenchymal tumor malignant 

Renal tubule, oncocytoma benign 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Leukemia mononuclear

+++++++++++++++++++++++++ 

X X X X X X X X X X XXXXXXX 

Mesothelioma benign 

Mesothelioma malignant X X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 117 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

7777777777777777777777777 

Number of Days on Study 0011222222222222222233333 

8868999999999999999900000 

4444444444444444444444444 Total 

Carcass ID Number 0312002222333344444500144 

7614581569037924579012336 

Tissues/ 

Tumors 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hemangiosarcoma, metastatic, spleen 

Histiocytic sarcoma 

Pheochromocytoma malignant, metastatic, 

adrenal medulla 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X X 

X 

X 

X 

50 

50 

4 

1 

1 

1 

1 

Nose ++A++++++++++++++++++++++ 49 

Lateral wall, adenoma 

Pleura 

1 

1 

Trachea +++++++++++++++++++++++++ 50 

Special Senses System 

Eye 

Zymbal’s gland 

Carcinoma 

+ 

+ 

X 

+ 2 

2 

2 

Urinary System 

Kidney 

Histiocytic sarcoma 

Mesenchymal tumor malignant 

Renal tubule, oncocytoma benign 

Urinary bladder 

+++++++++++++++++++++++++ 

X 

X 

X 

+++++++++++++++++++++++++ 

50 

1 

1 

1 

50 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Leukemia mononuclear 

+++++++++++++++++++++++++ 

X 

X X X X X X X X X X X XXXXX 

50 

1 

33 

Mesothelioma benign 

Mesothelioma malignant X 

X 1 

3 
 

 

 

 

 

 

 

 

 

 

 

118 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

4445555555666666666666777 

Number of Days on Study 4890113778112345556999001 

4158794569178872496045388 

6666666666666666666666666 

Carcass ID Number 0210433113111010440231222 

1527414580160894522877971 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Intestine large, colon ++++++++++++++++++++++A++ 

Intestine large, rectum ++++++++ I +++++++++++++A++ 

Intestine large, cecum ++++++A++++A++++++++++A++ 

Intestine small, duodenum ++++++++++++++++++++++A++ 

Intestine small, jejunum ++++++A++++A++++++++++A++ 

Intestine small, ileum +++++++++++A++++++++++A++ 

Liver +++++++++++++++++++++++++ 

Hepatocellular carcinoma X 

Hepatocellular adenoma 

Mesentery + + 

Pancreas +++++++++++++++++++++++++ 

Salivary glands +++++++++++++++++++++++++ 

Stomach, forestomach +++++++++++++++++++++++++ 

Stomach, glandular +++++++++++++++++++++++++ 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adrenal medulla +++++++++++++++++++++++++ 

Pheochromocytoma benign X X X X 

Bilateral, pheochromocytoma benign X X 

Islets, pancreatic +++++++++++++++++++++++++ 

Adenoma X 

Adenoma, multiple 

Carcinoma 

Parathyroid gland +M+M+++++++M++++++++++M++ 

Pituitary gland ++++++++++++++++++++ I +A++ 

Pars distalis, adenoma X XXX X XX XX X X 

Thyroid gland +++++++++++A++++++++++A++ 

C-cell, adenoma X 

C-cell, carcinoma 

Follicular cell, adenoma 

General Body System 

Peritoneum + 

Genital System 

Epididymis +++++++++++++++++++++++++ 

Preputial gland ++++++++++++++++M+++M++++ 

Adenoma X X 

Carcinoma 

Prostate +++++++++++++++++++++++++ 

Adenoma 

Adenoma, multiple 

Seminal vesicle +++++++++++A++++++++++A++ 

Testes +++++++++++++++++++++++++ 

Bilateral, interstitial cell, adenoma X XXX X XXXXX X 

Interstitial cell, adenoma X X X X X X X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 119 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

7777777777777777777777777 

Number of Days on Study 2222222222222222223333333 

3999999999999999990000000 

6666666666666666666666666 Total 

Carcass ID Number 3000012223344444451223334 Tissues/ 

8356943462501367803023699 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Intestine large, colon ++++++++++++++M++++++++++ 48 

Intestine large, rectum ++++++++++ I ++++++++++++++ 47 

Intestine large, cecum A ++++++++++++++++++++++++ 46 

Intestine small, duodenum +++++++++++++++++++++++++ 49 

Intestine small, jejunum A ++++++++++++++++++++++++ 46 

Intestine small, ileum A++++++++++++++++++++++++ 47 

Liver +++++++++++++++++++++++++ 50 

Hepatocellular carcinoma 1 

Hepatocellular adenoma X 1 

Mesentery + ++ + + + 8 

Pancreas +++++++++++++++++++++++++ 50 

Salivary glands +++++++++++++++++++++++++ 50 

Stomach, forestomach +++++++++++++++++++++++++ 50 

Stomach, glandular +++++++++++++++++++++++++ 50 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 50 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 50 

Adrenal medulla +++++++++++++++++++++++++ 50 

Pheochromocytoma benign X X X X X X X X 12 

Bilateral, pheochromocytoma benign X X 4 

Islets, pancreatic +++++++++++++++++++++++++ 50 

Adenoma X X 3 

Adenoma, multiple X 1 

Carcinoma X X X 3 

Parathyroid gland ++++++M+++++++++++M++++++ 44 

Pituitary gland ++++++++++++++++++++++M++ 47 

Pars distalis, adenoma X X XX XXXXX X XX XX 25 

Thyroid gland +++++++++++++++++++++++++ 48 

C-cell, adenoma X X X X X X X X X 10 

C-cell, carcinoma X X 2 

Follicular cell, adenoma X X X 3 

General Body System 

Peritoneum 1 

Genital System 

Epididymis +++++++++++++++++++++++++ 50 

Preputial gland ++M+++++MM++++M++++M+++++ 43 

Adenoma X X X 5 

Carcinoma X 1 

Prostate +++++++++++++++++++++++++ 50 

Adenoma X 1 

Adenoma, multiple X 1 

Seminal vesicle +++++++++++++++++++++++++ 48 

Testes +++++++++++++++++++++++++ 50 

Bilateral, interstitial cell, adenoma X XXXX XXXXXXXXXXXXXX X 31 

Interstitial cell, adenoma X X X 10 
 

 

 

 

 

 

 

 

 

 

120 2-Butoxyethanol, NTP TR 484 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

4445555555666666666666777 

Number of Days on Study 4890113778112345556999001 

4158794569178872496045388 

6666666666666666666666666 

Carcass ID Number 0210433113111010440231222 

1527414580160894522877971 

Hematopoietic System 

Bone marrow ++++++++++++++++++++++A++ 

Lymph node + + + + + + + + 

Sarcoma, metastatic, skin X 

Lymph node, bronchial + + + + + M + + M M + + M M M + + + M M + + M I + 

Lymph node, mandibular ++++++++++++++M++++++++M+ 

Lymph node, mesenteric +++++++++++++++++++++++++ 

Lymph node, mediastinal +++++++++++++++++M+++++M+ 

Spleen +++++++++++++++++++++++++ 

Fibroma 

Sarcoma 

Thymus +M+++++M+M+M++++M+M+++M++ 

Thymoma malignant 

Integumentary System 

Mammary gland +++++++++++M++++++M++++++ 

Fibroadenoma 

Skin +++++++++++++++++++++++++ 

Keratoacanthoma X 

Keratoacanthoma, multiple 

Squamous cell papilloma 

Subcutaneous tissue, fibroma X 

Subcutaneous tissue, fibroma, multiple 

Subcutaneous tissue, sarcoma X X 

Musculoskeletal System 

Bone 

Osteosarcoma 

+++++++++++++++++++++++++ 

Nervous System 

Brain 

Astrocytoma benign 

+++++++++++++++++++++++++ 

X 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar carcinoma 

Carcinoma, metastatic, preputial gland 

Sarcoma, metastatic, skin 

Nose 

Trachea 

++++++++++++++++++++++A++ 

+++++++++++++++++++++++++ 

X 

+++++++++++++++++++++++++ 

+++++++++++A++++++++++A++ 

Special Senses System 

Eye 

Urinary System 

Kidney 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma benign 

Mesothelioma malignant 

+++++++++++++++++++++++++ 

X X X X XXXXXXXXXXXXXXX 

X 

X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 121 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

7777777777777777777777777 

Number of Days on Study 2222222222222222223333333 

3999999999999999990000000 

6666666666666666666666666 Total 

Carcass ID Number 3000012223344444451223334 

8356943462501367803023699 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Lymph node 

Sarcoma, metastatic, skin 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Fibroma 

Sarcoma 

Thymus 

Thymoma malignant 

+++++++++++++++++++++++++ 

+ + 

+++MMM+++++M+++M+++++M+++ 

+++++++++++MM++++++++++++ 

+++++++++++++++++++++++++ 

M +++++M+++M++++M+++++++++ 

+++++++++++++++++++++++++ 

X 

X 

++++++MMM+++++M++++++++++ 

X 

49 

10 

1 

34 

46 

50 

44 

50 

1 

1 

39 

1 

Integumentary System 

Mammary gland 

Fibroadenoma 

Skin 

Keratoacanthoma 

Keratoacanthoma, multiple 

Squamous cell papilloma 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, fibroma, multiple 

Subcutaneous tissue, sarcoma 

++++M+++++M+++++++++++++M 

X 

+++++++++++++++++++++++++ 

X 

X 

X 

X X 

X 

45 

1 

50 

2 

1 

1 

3 

1 

2 

Musculoskeletal System 

Bone 

Osteosarcoma 

+++++++++++++++++++++++++ 

X 

50 

1 

Nervous System 

Brain 

Astrocytoma benign 

+++++++++++++++++++++++++ 50 

1 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar carcinoma 

Carcinoma, metastatic, preputial gland 

Sarcoma, metastatic, skin 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

X 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

49 

50 

1 

1 

1 

50 

48 

Special Senses System 

Eye + 1 

Urinary System 

Kidney 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

50 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma benign 

Mesothelioma malignant 

+++++++++++++++++++++++++ 

X X X X X X X XXXX 

50 

30 

1 

1 
122 2-Butoxyethanol, NTP TR 484 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Adrenal Medulla: Benign Pheochromocytoma 

Overall ratea 15/50 (30%) 20/50 (40%) 13/49 (27%) 16/50 (32%)
 

Adjusted rateb 36.5% 50.3% 32.8% 38.3%
 

Terminal ratec 7/19 (37%) 6/11 (55%) 6/21 (29%) 9/24 (38%)
 

First incidence (days) 558 533 658 611
 

Poly-3 testd P=0.424N P=0.140 P=0.452N P=0.522
 

Adrenal Medulla: Malignant Pheochromocytoma 

Overall rate 0/50 (0%) 2/50 (4%) 3/49 (6%) 0/50 (0%) 

Adjusted rate 0.0% 5.4% 7.7% 0.0% 

Terminal rate 0/19 (0%) 0/11 (0%) 1/21 (5%) 0/24 (0%) e First incidence (days) — 630 622 — 

Poly-3 test P=0.506N P=0.227 P=0.119 —f 

Adrenal Medulla: Benign or Malignant Pheochromocytoma 

Overall rate 15/50 (30%) 21/50 (42%) 15/49 (31%) 16/50 (32%)
 

Adjusted rate 36.5% 52.4% 37.5% 38.3%
 

Terminal rate 7/19 (37%) 6/11 (55%) 7/21 (33%) 9/24 (38%)
 

First incidence (days) 558 533 622 611
 

Poly-3 test P=0.413N P=0.100 P=0.557 P=0.522
 

Liver: Hepatocellular Adenoma or Carcinoma 

Overall rate 1/50 (2%) 3/50 (6%) 0/50 (0%) 2/50 (4%) 

Adjusted rate 2.6% 8.1% 0.0% 4.9% 

Terminal rate 0/19 (0%) 1/11 (9%) 0/21 (0%) 1/24 (4%) 

First incidence (days) 708 617 — 666 

Poly-3 test P=0.572 P=0.290 P=0.500N P=0.516 

Lung: Alveolar/bronchiolar Adenoma 

Overall rate 1/50 (2%) 2/50 (4%) 4/50 (8%) 0/50 (0%)
 

Adjusted rate 2.6% 5.5% 10.1% 0.0%
 

Terminal rate 1/19 (5%) 2/11 (18%) 3/21 (14%) 0/24 (0%)
 

First incidence (days) 729 (T) 729 (T) 472 —
 

Poly-3 test P=0.341N P=0.482 P=0.183 P=0.492N
 

Lung: Alveolar/bronchiolar Adenoma or Carcinoma 

Overall rate 1/50 (2%) 3/50 (6%) 5/50 (10%) 1/50 (2%) 

Adjusted rate 2.6% 8.2% 12.7% 2.5% 

Terminal rate 1/19 (5%) 2/11 (18%) 4/21 (19%) 1/24 (4%) 

First incidence (days) 729 (T) 723 472 729 (T) 

Poly-3 test P=0.508N P=0.285 P=0.104 P=0.752N 

Mammary Gland: Fibroadenoma 

Overall rate 3/50 (6%) 1/50 (2%) 3/50 (6%) 1/50 (2%) 

Adjusted rate 7.7% 2.7% 7.6% 2.5% 

Terminal rate 1/19 (5%) 0/11 (0%) 1/21 (5%) 1/24 (4%) 

First incidence (days) 641 677 565 729 (T) 

Poly-3 test P=0.289N P=0.325N P=0.661N P=0.294N 
2-Butoxyethanol, NTP TR 484 123 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Mammary Gland: Fibroadenoma or Carcinoma 

Overall rate 3/50 (6%) 2/50 (4%) 3/50 (6%) 1/50 (2%) 

Adjusted rate 7.7% 5.4% 7.6% 2.5% 

Terminal rate 1/19 (5%) 1/11 (9%) 1/21 (5%) 1/24 (4%) 

First incidence (days) 641 677 565 729 (T) 

Poly-3 test P=0.242N P=0.527N P=0.661N P=0.294N 

Oral Cavity (Oral Mucosa): Squamous Cell Papilloma 

Overall rate 3/50 (6%) 0/50 (0%) 0/50 (0%) 0/50 (0%)
 

Adjusted rate 7.5% 0.0% 0.0% 0.0%
 

Terminal rate 0/19 (0%) 0/11 (0%) 0/21 (0%) 0/24 (0%)
 

First incidence (days) 609 — — —
 

Poly-3 test P=0.041N P=0.133N P=0.123N P=0.115N
 

Oral Cavity (Oral Mucosa or Tongue): Squamous Cell Papilloma 

Overall rate 4/50 (8%) 0/50 (0%) 1/50 (2%) 0/50 (0%)
 

Adjusted rate 10.1% 0.0% 2.6% 0.0%
 

Terminal rate 1/19 (5%) 0/11 (0%) 1/21 (5%) 0/24 (0%)
 

First incidence (days) 609 — 729 (T) —
 

Poly-3 test P=0.032N P=0.070N P=0.185N P=0.058N
 

Pancreas: Adenoma 

Overall rate 1/50 (2%) 0/50 (0%) 3/50 (6%) 0/50 (0%)
 

Adjusted rate 2.6% 0.0% 7.8% 0.0%
 

Terminal rate 1/19 (5%) 0/11 (0%) 3/21 (14%) 0/24 (0%)
 

First incidence (days) 729 (T) — 729 (T) —
 

Poly-3 test P=0.470N P=0.510N P=0.304 P=0.492N
 

Pancreatic Islets: Adenoma 

Overall rate 5/50 (10%) 2/50 (4%) 4/50 (8%) 4/50 (8%)
 

Adjusted rate 12.7% 5.4% 10.3% 9.9%
 

Terminal rate 2/19 (11%) 0/11 (0%) 2/21 (10%) 3/24 (13%)
 

First incidence (days) 610 561 694 654
 

Poly-3 test P=0.524N P=0.234N P=0.507N P=0.480N
 

Pancreatic Islets: Carcinoma 

Overall rate 4/50 (8%) 1/50 (2%) 0/50 (0%) 3/50 (6%) 

Adjusted rate 10.3% 2.7% 0.0% 7.5% 

Terminal rate 2/19 (11%) 1/11 (9%) 0/21 (0%) 3/24 (13%) 

First incidence (days) 673 729 (T) — 729 (T) 

Poly-3 test P=0.483N P=0.195N P=0.060N P=0.483N 

Pancreatic Islets: Adenoma or Carcinoma 

Overall rate 9/50 (18%) 3/50 (6%) 4/50 (8%) 7/50 (14%)
 

Adjusted rate 22.7% 8.0% 10.3% 17.3%
 

Terminal rate 4/19 (21%) 1/11 (9%) 2/21 (10%) 6/24 (25%)
 

First incidence (days) 610 561 694 654
 

Poly-3 test P=0.452N P=0.069N P=0.117N P=0.371N
 
124 2-Butoxyethanol, NTP TR 484 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Pituitary Gland (Pars Distalis): Adenoma 

Overall rate 24/49 (49%) 29/49 (59%) 24/48 (50%) 25/47 (53%) 

Adjusted rate 55.2% 65.0% 55.8% 59.0% 

Terminal rate 6/18 (33%) 7/11 (64%) 10/20 (50%) 14/23 (61%) 

First incidence (days) 520 479 340 495 

Poly-3 test P=0.522 P=0.223 P=0.562 P=0.442 

Preputial Gland: Adenoma 

Overall rate 3/46 (7%) 1/47 (2%) 5/50 (10%) 5/43 (12%) 

Adjusted rate 8.4% 2.9% 12.5% 14.4% 

Terminal rate 1/16 (6%) 1/11 (9%) 2/21 (10%) 2/19 (11%) 

First incidence (days) 610 729 (T) 370 519 

Poly-3 test P=0.141 P=0.314N P=0.417 P=0.336 

Preputial Gland: Carcinoma 

Overall rate 5/46 (11%) 1/47 (2%) 2/50 (4%) 1/43 (2%) 

Adjusted rate 14.0% 2.9% 5.2% 3.0% 

Terminal rate 1/16 (6%) 1/11 (9%) 1/21 (5%) 1/19 (5%) 

First incidence (days) 564 729 (T) 701 729 (T) 

Poly-3 test P=0.089N P=0.104N P=0.180N P=0.112N 

Preputial Gland: Adenoma or Carcinoma 

Overall rate 8/46 (17%) 2/47 (4%) 7/50 (14%) 6/43 (14%) 

Adjusted rate 21.9% 5.8% 17.5% 17.3% 

Terminal rate 2/16 (13%) 2/11 (18%) 3/21 (14%) 3/19 (16%) 

First incidence (days) 564 729 (T) 370 519 

Poly-3 test P=0.559N P=0.049N P=0.421N P=0.425N 

Prostate Gland: Adenoma 

Overall rate 1/49 (2%) 0/49 (0%) 4/50 (8%) 2/50 (4%) 

Adjusted rate 2.7% 0.0% 10.3% 5.0% 

Terminal rate 1/18 (6%) 0/11 (0%) 3/21 (14%) 2/24 (8%) 

First incidence (days) 729 (T) — 708 729 (T) 

Poly-3 test P=0.275 P=0.508N P=0.186 P=0.524 

Skin: Squamous Cell Papilloma 

Overall rate 1/50 (2%) 1/50 (2%) 4/50 (8%) 1/50 (2%) 

Adjusted rate 2.6% 2.7% 10.3% 2.5% 

Terminal rate 0/19 (0%) 0/11 (0%) 4/21 (19%) 1/24 (4%) 

First incidence (days) 708 723 729 (T) 729 (T) 

Poly-3 test P=0.563 P=0.749 P=0.176 P=0.753N 

Skin: Keratoacanthoma 

Overall rate 3/50 (6%) 3/50 (6%) 4/50 (8%) 3/50 (6%) 

Adjusted rate 7.6% 8.0% 9.9% 7.4% 

Terminal rate 2/19 (11%) 0/11 (0%) 1/21 (5%) 2/24 (8%) 

First incidence (days) 529 579 340 617 

Poly-3 test P=0.567N P=0.645 P=0.515 P=0.647N 
2-Butoxyethanol, NTP TR 484 125 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Skin: Squamous Cell Papilloma or Keratoacanthoma 

Overall rate 4/50 (8%) 4/50 (8%) 8/50 (16%) 4/50 (8%)
 

Adjusted rate 10.2% 10.6% 19.8% 9.8%
 

Terminal rate 2/19 (11%) 0/11 (0%) 5/21 (24%) 3/24 (13%)
 

First incidence (days) 529 579 340 617
 

Poly-3 test P=0.544 P=0.622 P=0.184 P=0.627N
 

Skin: Basal Cell Adenoma or Carcinoma 

Overall rate 4/50 (8%) 2/50 (4%) 0/50 (0%) 0/50 (0%)
 

Adjusted rate 10.4% 5.4% 0.0% 0.0%
 

Terminal rate 4/19 (21%) 1/11 (9%) 0/21 (0%) 0/24 (0%)
 

First incidence (days) 729 (T) 638 — —
 

Poly-3 test P=0.015N P=0.355N P=0.059N P=0.054N
 

Skin: Squamous Cell Papilloma, Keratoacanthoma, Basal Cell Adenoma, or Basal Cell Carcinoma 

Overall rate 8/50 (16%) 6/50 (12%) 8/50 (16%) 4/50 (8%)
 

Adjusted rate 20.3% 15.8% 19.8% 9.8%
 

Terminal rate 6/19 (32%) 1/11 (9%) 5/21 (24%) 3/24 (13%)
 

First incidence (days) 529 579 340 617
 

Poly-3 test P=0.145N P=0.410N P=0.590N P=0.156N
 

Skin (Subcutaneous Tissue): Fibroma 

Overall rate 2/50 (4%) 2/50 (4%) 2/50 (4%) 4/50 (8%)
 

Adjusted rate 5.1% 5.4% 5.2% 9.9%
 

Terminal rate 1/19 (5%) 0/11 (0%) 2/21 (10%) 3/24 (13%)
 

First incidence (days) 646 617 729 (T) 638
 

Poly-3 test P=0.241 P=0.680 P=0.693 P=0.355
 

Skin (Subcutaneous Tissue): Fibroma or Fibrosarcoma 

Overall rate 3/50 (6%) 2/50 (4%) 3/50 (6%) 4/50 (8%)
 

Adjusted rate 7.6% 5.4% 7.8% 9.9%
 

Terminal rate 1/19 (5%) 0/11 (0%) 3/21 (14%) 3/24 (13%)
 

First incidence (days) 609 617 729 (T) 638
 

Poly-3 test P=0.369 P=0.525N P=0.656 P=0.517
 

Skin (Subcutaneous Tissue): Fibroma, Fibrosarcoma, or Sarcoma 

Overall rate 3/50 (6%) 2/50 (4%) 3/50 (6%) 5/50 (10%)
 

Adjusted rate 7.6% 5.4% 7.8% 12.2%
 

Terminal rate 1/19 (5%) 0/11 (0%) 3/21 (14%) 3/24 (13%)
 

First incidence (days) 609 617 729 (T) 576
 

Poly-3 test P=0.229 P=0.525N P=0.656 P=0.379
 

Testes: Adenoma 

Overall rate 41/50 (82%) 42/50 (84%) 44/50 (88%) 41/50 (82%)
 

Adjusted rate 89.9% 92.1% 93.4% 88.2%
 

Terminal rate 19/19 (100%) 11/11 (100%) 21/21 (100%) 22/24 (92%)
 

First incidence (days) 529 533 370 481
 

Poly-3 test P=0.399N P=0.501 P=0.386 P=0.529N
 
126 2-Butoxyethanol, NTP TR 484 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Thyroid Gland (C-cell): Adenoma 

Overall rate 7/50 (14%) 3/46 (7%) 7/48 (15%) 10/48 (21%)
 

Adjusted rate 17.9% 8.7% 18.4% 25.5%
 

Terminal rate 5/19 (26%) 1/11 (9%) 5/21 (24%) 9/24 (38%)
 

First incidence (days) 648 533 658 611
 

Poly-3 test P=0.120 P=0.204N P=0.597 P=0.292
 

Thyroid Gland (C-cell): Carcinoma 

Overall rate 1/50 (2%) 0/46 (0%) 3/48 (6%) 2/48 (4%) 

Adjusted rate 2.6% 0.0% 8.0% 5.2% 

Terminal rate 1/19 (5%) 0/11 (0%) 2/21 (10%) 2/24 (8%) 

First incidence (days) 729 (T) — 703 729 (T) 

Poly-3 test P=0.263 P=0.525N P=0.295 P=0.501 

Thyroid Gland (C-cell): Adenoma or Carcinoma 

Overall rate 7/50 (14%) 3/46 (7%) 9/48 (19%) 11/48 (23%)
 

Adjusted rate 17.9% 8.7% 23.6% 28.1%
 

Terminal rate 5/19 (26%) 1/11 (9%) 6/21 (29%) 10/24 (42%)
 

First incidence (days) 648 533 658 611
 

Poly-3 test P=0.066 P=0.204N P=0.370 P=0.208
 

Thyroid Gland (Follicular Cell): Adenoma 

Overall rate 1/50 (2%) 0/46 (0%) 0/48 (0%) 3/48 (6%) 

Adjusted rate 2.6% 0.0% 0.0% 7.7% 

Terminal rate 1/19 (5%) 0/11 (0%) 0/21 (0%) 3/24 (13%) 

First incidence (days) 729 (T) — — 729 (T) 

Poly-3 test P=0.089 P=0.525N P=0.505N P=0.305 

Thyroid Gland (Follicular Cell): Adenoma or Carcinoma 

Overall rate 2/50 (4%) 0/46 (0%) 1/48 (2%) 3/48 (6%) 

Adjusted rate 5.2% 0.0% 2.7% 7.7% 

Terminal rate 2/19 (11%) 0/11 (0%) 1/21 (5%) 3/24 (13%) 

First incidence (days) 729 (T) — 729 (T) 729 (T) 

Poly-3 test P=0.255 P=0.265N P=0.509N P=0.501 

All Organs: Benign or Malignant Mesothelioma 

Overall rate 1/50 (2%) 1/50 (2%) 4/50 (8%) 2/50 (4%) 

Adjusted rate 2.6% 2.7% 10.1% 4.9% 

Terminal rate 0/19 (0%) 0/11 (0%) 2/21 (10%) 0/24 (0%) 

First incidence (days) 597 652 461 589 

Poly-3 test P=0.349 P=0.749 P=0.181 P=0.516 

All Organs: Mononuclear Cell Leukemia 

Overall rate 29/50 (58%) 31/50 (62%) 33/50 (66%) 30/50 (60%)
 

Adjusted rate 63.6% 69.4% 73.5% 64.6%
 

Terminal rate 11/19 (58%) 6/11 (55%) 14/21 (67%) 11/24 (46%)
 

First incidence (days) 354 533 387 444
 

Poly-3 test P=0.541 P=0.352 P=0.204 P=0.547
 
127 

c 

2-Butoxyethanol, NTP TR 484 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

All Organs: Benign Neoplasms 

Overall rate 49/50 (98%) 49/50 (98%) 49/50 (98%) 49/50 (98%) 

Adjusted rate 99.8% 99.5% 98.7% 99.5% 

Terminal rate 19/19 (100%) 11/11 (100%) 21/21 (100%) 24/24 (100%) 

First incidence (days) 520 479 340 481 

Poly-3 test P=0.748N P=0.999N P=0.771N P=1.000N 

All Organs: Malignant Neoplasms 

Overall rate 41/50 (82%) 40/50 (80%) 41/50 (82%) 38/50 (76%) 

Adjusted rate 85.5% 84.9% 88.4% 80.1% 

Terminal rate 16/19 (84%) 8/11 (73%) 17/21 (81%) 16/24 (67%) 

First incidence (days) 354 465 387 444 

Poly-3 test P=0.270N P=0.588N P=0.455 P=0.328N 

All Organs: Benign or Malignant Neoplasms 

Overall rate 50/50 (100%) 50/50 (100%) 50/50 (100%) 50/50 (100%) 

Adjusted rate 100.0% 100.0% 100.0% 100.0% 

Terminal rate 19/19 (100%) 11/11 (100%) 21/21 (100%) 24/24 (100%) 

First incidence (days) 354 465 340 444 

Poly-3 test — — — — 

(T)Terminal sacrifice 

a Number of neoplasm-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for adrenal gland, 

liver, lung, pancreas, pancreatic islets, pituitary gland, preputial gland, prostate gland, testes, and thyroid gland; for other tissues, denominator is number 

of animals necropsied. b	 Poly-3 estimated neoplasm incidence after adjustment for intercurrent mortality 

Observed incidence at terminal kill 

d	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values 

corresponding to pairwise comparisons between the chamber controls and that exposed group. The Poly-3 test accounts for differential mortality in 

animals that do not reach terminal sacrifice. A negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group f Value of statistic cannot be computed. 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

128 2-Butoxyethanol, NTP TR 484 

TABLE A4 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 25 31 24 21 

Natural deaths 6 8 5 5 

Survivors 

Terminal sacrifice 19 11 21 24 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Intestine large, colon (47) (47) (47) (48) 

Parasite metazoan 4 (9%) 4 (9%) 4 (9%) 4 (8%) 

Intestine large, rectum (46) (47) (48) (47) 

Parasite metazoan 1 (2%) 2 (4%) 6 (13%) 3 (6%) 

Perforation 1 (2%) 

Intestine large, cecum (46) (47) (47) (46) 

Necrosis 1 (2%) 

Parasite metazoan 6 (13%) 7 (15%) 4 (9%) 5 (11%) 

Intestine small, duodenum (46) (47) (49) (49) 

Necrosis 1 (2%) 

Intestine small, ileum (45) (45) (46) (47) 

Inflammation, acute 1 (2%) 

Inflammation, chronic active 1 (2%) 

Parasite metazoan 1 (2%) 1 (2%) 1 (2%) 1 (2%) 

Liver (50) (50) (50) (50) 

Angiectasis 1 (2%) 1 (2%) 2 (4%) 1 (2%) 

Basophilic focus 26 (52%) 21 (42%) 25 (50%) 27 (54%) 

Clear cell focus 10 (20%) 7 (14%) 13 (26%) 9 (18%) 

Cyst 1 (2%) 

Degeneration, cystic 7 (14%) 6 (12%) 15 (30%) 6 (12%) 

Eosinophilic focus 6 (12%) 4 (8%) 3 (6%) 4 (8%) 

Fatty change 7 (14%) 10 (20%) 6 (12%) 3 (6%) 

Hematopoietic cell proliferation 2 (4%) 3 (6%) 1 (2%) 

Hepatodiaphragmatic nodule 1 (2%) 2 (4%) 1 (2%) 

Mixed cell focus 1 (2%) 1 (2%) 1 (2%) 

Necrosis 7 (14%) 9 (18%) 4 (8%) 11 (22%) 

Regeneration 1 (2%) 4 (8%) 

Thrombosis 1 (2%) 1 (2%) 1 (2%) 

Bile duct, hyperplasia 28 (56%) 29 (58%) 30 (60%) 36 (72%) 

Centrilobular, necrosis 4 (8%) 9 (18%) 5 (10%) 4 (8%) 

Kupffer cell, pigmentation 23 (46%) 30 (60%) 34 (68%) 42 (84%) 

Mesentery (8) (6) (15) (8) 

Fat, necrosis 8 (100%) 6 (100%) 11 (73%) 8 (100%) 

Oral mucosa (3) (1) (1)

 Pharyngeal, hyperplasia, squamous 1 (100%) 

Pancreas (50) (50) (50) (50) 

Atrophy 25 (50%) 25 (50%) 26 (52%) 22 (44%) 

Basophilic focus 3 (6%) 2 (4%) 5 (10%) 2 (4%) 

Hyperplasia 3 (6%) 5 (10%) 2 (4%) 2 (4%) 

a Number of animals examined microscopically at the site and the number of animals with lesion 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 129 

TABLE A4 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Alimentary System (continued) 

Salivary glands (50) (50) (50) (50) 

Artery, mineralization 1 (2%) 

Stomach, forestomach (50) (50) (50) (50) 

Mineralization 2 (4%) 

Necrosis 1 (2%) 

Ulcer 7 (14%) 9 (18%) 3 (6%) 3 (6%) 

Epithelium, hyperplasia 5 (10%) 8 (16%) 4 (8%) 4 (8%) 

Stomach, glandular (49) (49) (50) (50) 

Inflammation, acute 1 (2%) 

Mineralization 1 (2%) 3 (6%) 2 (4%) 4 (8%) 

Necrosis 4 (8%) 8 (16%) 4 (8%) 10 (20%) 

Tooth (3) (1) 

Inflammation, chronic active 3 (100%) 1 (100%) 

Malformation 1 (33%) 

Cardiovascular System 

Blood vessel (2) 

Aorta, mineralization 2 (100%) 

Heart (50) (50) (50) (50) 

Cardiomyopathy 42 (84%) 44 (88%) 39 (78%) 41 (82%) 

Artery, mineralization 2 (4%) 

Atrium, thrombosis 1 (2%) 6 (12%) 4 (8%) 3 (6%) 

Endocrine System 

Adrenal cortex (50) (50) (49) (50) 

Atrophy 1 (2%) 3 (6%) 2 (4%) 

Hyperplasia 23 (46%) 27 (54%) 25 (51%) 30 (60%) 

Hypertrophy 7 (14%) 7 (14%) 8 (16%) 4 (8%) 

Necrosis 1 (2%) 1 (2%) 1 (2%) 

Vacuolization cytoplasmic 2 (4%) 2 (4%) 4 (8%) 4 (8%) 

Adrenal medulla (50) (50) (49) (50) 

Hyperplasia 24 (48%) 22 (44%) 31 (63%) 27 (54%) 

Islets, pancreatic (50) (50) (50) (50) 

Hyperplasia 2 (4%) 4 (8%) 1 (2%) 

Parathyroid gland (47) (46) (47) (44) 

Hyperplasia 4 (9%) 7 (15%) 2 (4%) 3 (7%) 

Pituitary gland (49) (49) (48) (47) 

Cyst 1 (2%) 

Mineralization 1 (2%) 

Pars distalis, hyperplasia 16 (33%) 12 (24%) 16 (33%) 14 (30%) 

Pars intermedia, hyperplasia 1 (2%) 1 (2%) 

Thyroid gland (50) (46) (48) (48) 

C-cell, hyperplasia 23 (46%) 31 (67%) 32 (67%) 17 (35%) 

Follicular cell, hyperplasia 1 (2%) 1 (2%) 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

130 2-Butoxyethanol, NTP TR 484 

TABLE A4 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Genital System 

Epididymis (50) (50) (50) (50) 

Granuloma sperm 1 (2%) 1 (2%) 1 (2%) 

Preputial gland (46) (47) (50) (43) 

Hyperplasia 3 (6%) 4 (9%) 

Inflammation, chronic active 1 (2%) 5 (11%) 2 (4%) 1 (2%) 

Prostate (49) (49) (50) (50) 

Hyperplasia 6 (12%) 9 (18%) 4 (8%) 8 (16%) 

Inflammation, chronic active 4 (8%) 3 (6%) 2 (4%) 1 (2%) 

Seminal vesicle (47) (44) (46) (48) 

Hyperplasia 1 (2%) 

Mineralization 2 (5%) 

Testes (50) (50) (50) (50) 

Atrophy 2 (4%) 2 (4%) 1 (2%) 2 (4%) 

Artery, inflammation, chronic active 1 (2%) 

Interstitial cell, hyperplasia 5 (10%) 2 (4%) 5 (10%) 8 (16%) 

Hematopoietic System 

Bone marrow (50) (50) (50) (49) 

Hyperplasia 5 (10%) 4 (8%) 4 (8%) 6 (12%) 

Necrosis 1 (2%) 

Lymph node, mandibular (44) (47) (47) (46) 

Infiltration cellular, plasma cell 1 (2%) 

Lymph node, mediastinal (44) (44) (45) (44) 

Infiltration cellular, plasma cell 1 (2%) 

Spleen (50) (50) (50) (50) 

Fibrosis 11 (22%) 14 (28%) 19 (38%) 20 (40%) 

Hematopoietic cell proliferation 8 (16%) 8 (16%) 6 (12%) 5 (10%) 

Hemorrhage 1 (2%) 1 (2%) 

Hyperplasia, focal, lymphoid 1 (2%) 

Necrosis 2 (4%) 3 (6%) 3 (6%) 3 (6%) 

Pigmentation, hemosiderin 45 (90%) 45 (90%) 40 (80%) 44 (88%) 

Integumentary System 

Mammary gland (44) (40) (44) (45) 

Galactocele 1 (2%) 1 (2%) 

Hyperplasia 1 (2%) 

Skin (50) (50) (50) (50) 

Cyst epithelial inclusion 1 (2%) 

Hyperkeratosis 1 (2%) 1 (2%) 3 (6%) 

Inflammation, chronic active 2 (4%) 1 (2%) 4 (8%) 

Subcutaneous tissue, thrombosis 1 (2%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Fibrous osteodystrophy 1 (2%) 3 (6%) 

Hyperostosis 2 (4%) 2 (4%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 131 

TABLE A4 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Nervous System 

Brain (50) (50) (50) (50) 

Compression 1 (2%) 

Hemorrhage 1 (2%) 

Hydrocephalus 1 (2%) 

Mineralization 1 (2%) 1 (2%) 

Thrombosis 1 (2%) 

Respiratory System 

Lung (50) (50) (50) (50) 

Hemorrhage 1 (2%) 

Infiltration cellular, histiocyte 1 (2%) 1 (2%) 2 (4%) 

Inflammation, chronic active 1 (2%) 3 (6%) 3 (6%) 

Inflammation, suppurative 1 (2%) 

Mineralization 2 (4%) 

Necrosis 1 (2%) 

Thrombosis 1 (2%) 

Alveolar epithelium, hyperplasia 5 (10%) 13 (26%) 6 (12%) 14 (28%) 

Artery, infiltration cellular, histiocyte 2 (4%) 

Artery, mediastinum, mineralization 2 (4%) 

Mediastinum, inflammation, chronic 1 (2%) 

Nose (48) (49) (49) (50) 

Inflammation, suppurative 7 (15%) 7 (14%) 10 (20%) 5 (10%) 

Thrombosis 7 (15%) 13 (27%) 11 (22%) 10 (20%) 

Olfactory epithelium, degeneration, hyaline 13 (27%) 21 (43%) 23 (47%) 40 (80%) 

Olfactory epithelium, metaplasia 4 (8%) 5 (10%) 2 (4%) 3 (6%) 

Respiratory epithelium, hyperplasia 4 (8%) 4 (8%) 6 (12%) 1 (2%) 

Respiratory epithelium, metaplasia, squamous 1 (2%) 

Trachea (50) (49) (50) (48) 

Infiltration cellular, polymorphonuclear 1 (2%) 

Special Senses System 

Eye (2) (2) (1) 

Cataract 2 (100%) 2 (100%) 1 (100%) 

Retina, atrophy 2 (100%) 2 (100%) 1 (100%) 

Urinary System 

Kidney (50) (50) (50) (50) 

Hydronephrosis 1 (2%) 

Infarct 1 (2%) 2 (4%) 1 (2%) 6 (12%) 

Mineralization 2 (4%) 

Nephropathy 48 (96%) 50 (100%) 49 (98%) 50 (100%) 

Cortex, cyst 1 (2%) 1 (2%) 

Pelvis, inflammation, acute 1 (2%) 

Renal tubule, hyperplasia 3 (6%) 1 (2%) 1 (2%) 

Renal tubule, necrosis 2 (4%) 
132 2-Butoxyethanol, NTP TR 484 
133 

APPENDIX B 

SUMMARY OF LESIONS IN FEMALE RATS 

IN THE 2-YEAR INHALATION STUDY 

OF 2-BUTOXYETHANOL 

TABLE B1 

TABLE B2 

TABLE B3 

TABLE B4 

TABLE B5 

Summary of the Incidence of Neoplasms in Female Rats 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

Individual Animal Tumor Pathology of Female Rats 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

Statistical Analysis of Primary Neoplasms in Female Rats 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

Historical Incidence of Adrenal Gland Pheochromocytoma 

in Chamber Control Female F344/N Rats ................................ 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

134 

138 

158 

161 

162 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

134 2-Butoxyethanol, NTP TR 484 

TABLE B1 

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 18 21 23 26 

Natural deaths 3 2 4 3 

Survivors 

Terminal sacrifice 29 27 23 21 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Intestine large, colon (49) (50) (50) (49) 

Histiocytic sarcoma 1 (2%) 

Intestine large, rectum (44) (47) (48) (47) 

Intestine large, cecum (47) (49) (48) (48) 

Intestine small, duodenum (49) (49) (49) (48) 

Histiocytic sarcoma 1 (2%) 

Intestine small, jejunum (47) (49) (47) (48) 

Histiocytic sarcoma 1 (2%) 

Intestine small, ileum (47) (50) (47) (47) 

Histiocytic sarcoma 1 (2%) 

Liver (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 

Mesentery (12) (9) (8) (3) 

Hemangioma 1 (13%) 

Sex cord stromal tumor, malignant,

 metastatic, ovary 1 (8%) 

Pancreas (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 

Salivary glands (50) (50) (50) (49) 

Histiocytic sarcoma 1 (2%) 

Stomach, forestomach (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 

Stomach, glandular (50) (50) (49) (50) 

Histiocytic sarcoma 1 (2%) 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 

Endocrine System 

Adrenal cortex (50) (50) (50) (50) 

Adenoma 1 (2%) 2 (4%) 

Histiocytic sarcoma 1 (2%) 

Osteosarcoma, metastatic, bone 1 (2%) 

Bilateral, carcinoma 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 135 

TABLE B1 

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Endocrine System (continued) 

Adrenal medulla (50) (50) (49) (49) 

Osteosarcoma, metastatic, bone 1 (2%) 

Pheochromocytoma malignant 1 (2%) 

Pheochromocytoma benign 3 (6%) 4 (8%) 1 (2%) 6 (12%) 

Bilateral, pheochromocytoma benign 1 (2%) 

Islets, pancreatic (50) (50) (49) (50) 

Adenoma 1 (2%) 1 (2%) 

Carcinoma 1 (2%) 

Pituitary gland (50) (50) (49) (49) 

Histiocytic sarcoma 1 (2%) 

Pars distalis, adenoma 33 (66%) 32 (64%) 30 (61%) 20 (41%) 

Thyroid gland (50) (48) (49) (50) 

Histiocytic sarcoma 1 (2%) 

C-cell, adenoma 6 (12%) 2 (4%) 5 (10%) 5 (10%) 

C-cell, carcinoma 2 (4%) 2 (4%) 

Follicular cell, adenoma 1 (2%) 1 (2%) 

Follicular cell, carcinoma 1 (2%) 1 (2%) 

General Body System 

Peritoneum (1) 

Genital System 

Clitoral gland (46) (45) (44) (48) 

Adenoma 6 (13%) 3 (7%) 4 (9%) 4 (8%) 

Carcinoma 3 (7%) 1 (2%) 3 (7%) 2 (4%) 

Histiocytic sarcoma 1 (2%) 

Ovary (50) (50) (50) (50) 

Granulosa cell tumor benign 1 (2%) 

Granulosa-theca tumor malignant 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Sex cord stromal tumor, malignant 1 (2%) 

Uterus (50) (50) (50) (50) 

Carcinoma 1 (2%) 

Deciduoma benign 1 (2%) 

Deciduoma NOS 1 (2%) 

Hemangiosarcoma 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Polyp stromal 6 (12%) 3 (6%) 5 (10%) 2 (4%) 

Polyp stromal, multiple 1 (2%) 

Hematopoietic System 

Bone marrow (50) (50) (50) (50) 

Lymph node (5) (3) (6) (2) 

Histiocytic sarcoma 1 (17%) 

Renal, histiocytic sarcoma 1 (17%) 

Renal, osteosarcoma, metastatic, bone 1 (20%) 

Lymph node, bronchial (33) (36) (34) (26) 

Histiocytic sarcoma 1 (3%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

136 2-Butoxyethanol, NTP TR 484 

TABLE B1 

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Hematopoietic System (continued) 

Lymph node, mandibular (49) (43) (46) (46) 

Histiocytic sarcoma 1 (2%) 

Lymph node, mesenteric (48) (50) (50) (49) 

Histiocytic sarcoma 1 (2%) 

Lymph node, mediastinal (38) (43) (39) (28) 

Histiocytic sarcoma 1 (3%) 

Spleen (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 

Thymus (48) (47) (49) (44) 

Histiocytic sarcoma 1 (2%) 

Integumentary System 

Mammary gland (50) (50) (50) (49) 

Carcinoma 3 (6%) 4 (8%) 4 (8%) 3 (6%) 

Fibroadenoma 19 (38%) 16 (32%) 10 (20%) 12 (24%) 

Fibroadenoma, multiple 4 (8%) 8 (16%) 10 (20%) 3 (6%) 

Histiocytic sarcoma 1 (2%) 

Skin (49) (50) (50) (50) 

Basal cell carcinoma 1 (2%) 1 (2%) 

Keratoacanthoma 2 (4%) 

Squamous cell papilloma 2 (4%) 

Subcutaneous tissue, fibroma 1 (2%) 1 (2%) 

Subcutaneous tissue, lipoma 1 (2%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Chordoma 1 (2%) 

Osteosarcoma, multiple 1 (2%) 

Skeletal muscle (1) (2) 

Hemangioma 1 (100%) 

Sarcoma 1 (50%) 

Nervous System 

Brain (50) (50) (50) (50) 

Astrocytoma malignant 1 (2%) 

Glioma malignant 1 (2%) 

Histiocytic sarcoma 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 137 

TABLE B1 

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Respiratory System 

Larynx (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 

Lung (50) (50) (50) (49) 

Carcinoma, metastatic, mammary gland 1 (2%) 

Carcinoma, metastatic, thyroid gland 1 (2%) 1 (2%) 

Chordoma, metastatic, bone 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Osteosarcoma, metastatic, bone 1 (2%) 

Sarcoma, metastatic, skeletal muscle 1 (2%) 

Nose (50) (48) (50) (49) 

Histiocytic sarcoma 1 (2%) 

Special Senses System 

Zymbal’s gland (1) (1) 

Carcinoma 1 (100%) 1 (100%) 

Urinary System 

Kidney (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 

Osteosarcoma, metastatic, bone 1 (2%) 

Sex cord stromal tumor, malignant,

 metastatic, ovary 1 (2%) 

Renal tubule, carcinoma 1 (2%) 

Urinary bladder (50) (50) (49) (50) 

Histiocytic sarcoma 1 (2%) 

Systemic Lesions 

Multiple organsb (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 

Leukemia mononuclear 18 (36%) 21 (42%) 23 (46%) 24 (48%) 

Mesothelioma malignant 1 (2%) 

Neoplasm Summary 

Total animals with primary neoplasmsc

 47 48 50 48 

Total primary neoplasms 112 99 108 98 

Total animals with benign neoplasms 42 43 40 37 

Total benign neoplasms 83 69 72 58 

Total animals with malignant neoplasms 26 26 30 32 

Total malignant neoplasms 28 30 36 40 

Total animals with metastatic neoplasms 2 1 4 

Total metastatic neoplasms 7 1 4 

Total animals with uncertain neoplasms-

 benign or malignant 1 

Total uncertain neoplasms 1 

a Number of animals examined microscopically at the site and the number of animals with neoplasm b Number of animals with any tissue examined microscopically 

Primary neoplasms: all neoplasms except metastatic neoplasms c 
 

 

 

 

 

 

 

 

 

138 2-Butoxyethanol, NTP TR 484 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

4555555666666667777777777 

Number of Days on Study 0035579113455880111123333 

0834854778824788088820000 

1111111111111111111111111 

Carcass ID Number 3310423124444133201430111 

3837982657802275965149147 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Intestine large, colon ++++++A++++++++++++++++++ 

Intestine large, rectum I +++++A+++++++++ I ++++++++ 

Intestine large, cecum ++++++A+++A+++++A++++++++ 

Intestine small, duodenum ++++++A++++++++++++++++++ 

Intestine small, jejunum ++++++A+++A+++++A++++++++ 

Intestine small, ileum ++++++A+++A+++++A++++++++ 

Liver +++++++++++++++++++++++++ 

Mesentery + + + ++ + 

Sex cord stromal tumor, malignant, 

metastatic, ovary X 

Pancreas +++++++++++++++++++++++++ 

Salivary glands +++++++++++++++++++++++++ 

Stomach, forestomach +++++++++++++++++++++++++ 

Stomach, glandular +++++++++++++++++++++++++ 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adenoma X 

Osteosarcoma, metastatic, bone X 

Adrenal medulla +++++++++++++++++++++++++ 

Osteosarcoma, metastatic, bone X 

Pheochromocytoma benign X X 

Islets, pancreatic +++++++++++++++++++++++++ 

Adenoma X 

Parathyroid gland +++++++++++++++++M+++++++ 

Pituitary gland +++++++++++++++++++++++++ 

Pars distalis, adenoma X X XX X XXXXXX X 

Thyroid gland +++++++++++++++++++++++++ 

C-cell, adenoma X X X 

Follicular cell, adenoma 

Follicular cell, carcinoma 

General Body System 

None 

+: Tissue examined microscopically M: Missing tissue X: Lesion present 

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined 
 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 139 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

7777777777777777777777777 

Number of Days on Study 3333333333333333333333333 

0000001111111111111111111 

1111111111111111111111111 Total 

Carcass ID Number 1233450000001122222233444 Tissues/ 

8119401234580902346706356 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Intestine large, colon +++++++++++++++++++++++++ 49 

Intestine large, rectum ++++ I I ++++++++++++++++ I ++ 44 

Intestine large, cecum +++++++++++++++++++++++++ 47 

Intestine small, duodenum +++++++++++++++++++++++++ 49 

Intestine small, jejunum +++++++++++++++++++++++++ 47 

Intestine small, ileum +++++++++++++++++++++++++ 47 

Liver +++++++++++++++++++++++++ 50 

Mesentery + + + ++ + 12 

Sex cord stromal tumor, malignant, 

metastatic, ovary 1 

Pancreas +++++++++++++++++++++++++ 50 

Salivary glands +++++++++++++++++++++++++ 50 

Stomach, forestomach +++++++++++++++++++++++++ 50 

Stomach, glandular +++++++++++++++++++++++++ 50 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 50 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 50 

Adenoma 1 

Osteosarcoma, metastatic, bone 1 

Adrenal medulla +++++++++++++++++++++++++ 50 

Osteosarcoma, metastatic, bone 1 

Pheochromocytoma benign X 3 

Islets, pancreatic +++++++++++++++++++++++++ 50 

Adenoma 1 

Parathyroid gland +++++++++++MM+++M++++++++ 46 

Pituitary gland +++++++++++++++++++++++++ 50 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X X 33 

Thyroid gland +++++++++++++++++++++++++ 50 

C-cell, adenoma X X X 6 

Follicular cell, adenoma X 1 

Follicular cell, carcinoma X 1 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

 

 

140 2-Butoxyethanol, NTP TR 484 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

4555555666666667777777777 

Number of Days on Study 0035579113455880111123333 

0834854778824788088820000 

1111111111111111111111111 

Carcass ID Number 3310423124444133201430111 

3837982657802275965149147 

Genital System 

Clitoral gland 

Adenoma 

Carcinoma 

Ovary 

Sex cord stromal tumor, malignant 

Uterus 

Deciduoma benign 

Deciduoma NOS 

Polyp stromal 

++++M++++++M++++++++++M++ 

X 

X 

+++++++++++++++++++++++++ 

X 

+++++++++++++++++++++++++ 

X 

X X 

Hematopoietic System 

Bone marrow 

Lymph node 

Renal, osteosarcoma, metastatic, bone 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+++++++++++++++++++++++++ 

+ + + + + 

X 

+++++M++++++M+MMMM+++MM+M 

+++++++++++++++++++++++++ 

+++++++++++++++++++++M+++ 

+ + M ++++++++++++MM+M++++M+ 

+++++++++++++++++++++++++ 

+++++++++++M++++++++M++++ 

Integumentary System 

Mammary gland 

Carcinoma 

Fibroadenoma

Fibroadenoma, multiple 

Skin 

Basal cell carcinoma 

Keratoacanthoma 

+++++++++++++++++++++++++ 

X 

X X X X X X X X X 

X 

+++++++++++++++++++++++++ 

X 

Musculoskeletal System 

Bone 

Osteosarcoma, multiple 

+++++++++++++++++++++++++ 

X 

Nervous System 

Brain +++++++++++++++++++++++++ 

Respiratory System 

Larynx 

Lung 

Osteosarcoma, metastatic, bone 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 141 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

7777777777777777777777777 

Number of Days on Study 3333333333333333333333333 

0000001111111111111111111 

1111111111111111111111111 Total 

Carcass ID Number 1233450000001122222233444 

8119401234580902346706356 

Tissues/ 

Tumors 

Genital System 

Clitoral gland 

Adenoma 

Carcinoma 

Ovary 

Sex cord stromal tumor, malignant 

Uterus 

Deciduoma benign 

Deciduoma NOS 

Polyp stromal 

++++++++++++++++M++++++++ 

X X X X X 

X X 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

X X X X 

46 

6 

3 

50 

1 

50 

1 

1 

6 

Hematopoietic System 

Bone marrow 

Lymph node 

Renal, osteosarcoma, metastatic, bone 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+++++++++++++++++++++++++ 

+++M++MMMMM+M++++++M+++++ 

++++++M++++++++++++++++++ 

++++++++++++++++M++++++++ 

+++++++MMMM++++M+++M+M+++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

5 

1 

33 

49 

48 

38 

50 

48 

Integumentary System 

Mammary gland 

Carcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Basal cell carcinoma 

Keratoacanthoma 

+++++++++++++++++++++++++ 

X X 

X X X X X X X X X X 

X X X 

++++++++M++++++++++++++++ 

X 

X 

50 

3 

19 

4 

49 

1 

2 

Musculoskeletal System 

Bone 

Osteosarcoma, multiple 

+++++++++++++++++++++++++ 50 

1 

Nervous System 

Brain +++++++++++++++++++++++++ 50 

Respiratory System 

Larynx 

Lung 

Osteosarcoma, metastatic, bone 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

50 

1 

50 

50 
 

 

 

142 2-Butoxyethanol, NTP TR 484 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

4555555666666667777777777 

Number of Days on Study 0035579113455880111123333 

0834854778824788088820000 

1111111111111111111111111 

Carcass ID Number 3310423124444133201430111 

3837982657802275965149147 

Special Senses System 

None 

Urinary System 

Kidney +++++++++++++++++++++++++ 

Osteosarcoma, metastatic, bone X 

Sex cord stromal tumor, malignant, 

metastatic, ovary X 

Urinary bladder +++++++++++++++++++++++++ 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 

Leukemia mononuclear X X X X X X X X X 
 

 

 

 

2-Butoxyethanol, NTP TR 484 143 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

7777777777777777777777777 

Number of Days on Study 3333333333333333333333333 

0000001111111111111111111 

1111111111111111111111111 Total 

Carcass ID Number 1233450000001122222233444 Tissues/ 

8119401234580902346706356 Tumors 

Special Senses System 

None 

Urinary System 

Kidney +++++++++++++++++++++++++ 50 

Osteosarcoma, metastatic, bone 1 

Sex cord stromal tumor, malignant, 

metastatic, ovary 1 

Urinary bladder +++++++++++++++++++++++++ 50 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 50 

Leukemia mononuclear X X X X XXXXX 18 
 

 

 

 

 

 

144 2-Butoxyethanol, NTP TR 484 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 31.2 ppm 

4555556666666666677777777 

Number of Days on Study 9456680113555668900022233 

5431446078225677117822900 

3333333333333333333333333 

Carcass ID Number 2242041433013543122114201 

3156339008754091875017952 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Intestine large, colon +++++++++++++++++++++++++ 

Intestine large, rectum +++ I ++++ I +++A++++++++++++ 

Intestine large, cecum +++++++++A+++++++++++++++ 

Intestine small, duodenum +++++++++A+++++++++++++++ 

Intestine small, jejunum +++++++++A+++++++++++++++ 

Intestine small, ileum +++++++++++++++++++++++++ 

Liver +++++++++++++++++++++++++ 

Mesentery ++ ++ + + 

Pancreas +++++++++++++++++++++++++ 

Salivary glands +++++++++++++++++++++++++ 

Stomach, forestomach +++++++++++++++++++++++++ 

Stomach, glandular +++++++++++++++++++++++++ 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adrenal medulla +++++++++++++++++++++++++ 

Pheochromocytoma benign X X X 

Islets, pancreatic +++++++++++++++++++++++++ 

Carcinoma 

Parathyroid gland +++++++++++++++++M+++++++ 

Pituitary gland +++++++++++++++++++++++++ 

Pars distalis, adenoma XXXXXX XXX X XXXX 

Thyroid gland +++++++++A++A++++++++++++ 

C-cell, adenoma X X 

General Body System 

None 

Genital System 

Clitoral gland ++++++++++M++++++++++++++ 

Adenoma X X 

Carcinoma 

Ovary +++++++++++++++++++++++++ 

Uterus +++++++++++++++++++++++++ 

Polyp stromal 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 

Lymph node + + 

Lymph node, bronchial + + M + MMMM++++M++++++M+++++ 

Lymph node, mandibular +++++++++++M++++++M++++++ 

Lymph node, mesenteric +++++++++++++++++++++++++ 

Lymph node, mediastinal + + M ++++M+++++MM++++++++++ 

Spleen +++++++++++++++++++++++++ 

Thymus ++++++++++++++M++++++++++ 
 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 145 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 31.2 ppm 

Number of Days on Study 

7777777777777777777777777 

3333333333333333333333333 

0000000000111111111111111 

Carcass ID Number 

3333333333333333333333333 

1112234444000000122333334 

4674831246124689302256798 

Total 

Tissues/ 

Tumors 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+ + + 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

50 

47 

49 

49 

49 

50 

50 

9 

50 

50 

50 

50 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma benign X 4 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma X 1 

Parathyroid gland + + + + + + + M + + + + + + + + + + + + + + + + + 48 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X 32 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 48 

C-cell, adenoma 2 

General Body System 

None 

Genital System 

Clitoral gland ++M+++MM+++++M+++++++++++ 45 

Adenoma X 3 

Carcinoma X 1 

Ovary +++++++++++++++++++++++++ 50 

Uterus +++++++++++++++++++++++++ 50 

Polyp stromal X X X 3 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 50 

Lymph node + 3 

Lymph node, bronchial +++MM+++MM+++++M++++M+M++ 36 

Lymph node, mandibular M M +++++++++M++++M+++++M++ 43 

Lymph node, mesenteric +++++++++++++++++++++++++ 50 

Lymph node, mediastinal +++++++++M++++++M+++++M++ 43 

Spleen +++++++++++++++++++++++++ 50 

Thymus M++++++++++++++++++++++M+ 47 
 

 

 

 

 

 

 

 

146 2-Butoxyethanol, NTP TR 484 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 31.2 ppm 

4555556666666666677777777 

Number of Days on Study 9456680113555668900022233 

5431446078225677117822900 

3333333333333333333333333 

Carcass ID Number 2242041433013543122114201 

3156339008754091875017952 

Integumentary System 

Mammary gland +++++++++++++++++++++++++ 

Carcinoma X X X X 

Fibroadenoma X X X X X X X X 

Fibroadenoma, multiple X X X 

Skin +++++++++++++++++++++++++ 

Basal cell carcinoma 

Subcutaneous tissue, fibroma X 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Nervous System 

Brain +++++++++++++++++++++++++ 

Astrocytoma malignant X 

Respiratory System 

Larynx +++++++++++++++++++++++++ 

Lung +++++++++++++++++++++++++ 

Nose +++++++++A++A++++++++++++ 

Trachea +++++++++++++++++++++++++ 

Special Senses System 

Eye ++ 

Zymbal’s gland + 

Carcinoma X 

Urinary System 

Kidney +++++++++++++++++++++++++ 

Urinary bladder +++++++++++++++++++++++++ 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 

Leukemia mononuclear XXXXX X XX X XXXX X X 
 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 147 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 31.2 ppm 

Number of Days on Study 

7777777777777777777777777 

3333333333333333333333333 

0000000000111111111111111 

Carcass ID Number 

3333333333333333333333333 

1112234444000000122333334 

4674831246124689302256798 

Total 

Tissues/ 

Tumors 

Integumentary System 

Mammary gland 

Carcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Basal cell carcinoma 

Subcutaneous tissue, fibroma 

+++++++++++++++++++++++++ 

X X X X X X X X 

X X X X X 

+++++++++++++++++++++++++ 

X 

50 

4 

16 

8 

50 

1 

1 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 50 

Nervous System 

Brain 

Astrocytoma malignant 

+++++++++++++++++++++++++ 50 

1 

Respiratory System 

Larynx 

Lung 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

50 

48 

50 

Special Senses System 

Eye 

Zymbal’s gland 

Carcinoma 

++ + + + 7 

1 

1 

Urinary System 

Kidney 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

50 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

+++++++++++++++++++++++++ 

X X X X X X 

50 

21 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

148 2-Butoxyethanol, NTP TR 484 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

2344455555556666666666677 

Number of Days on Study 4966924478881344568899900 

7022694443699389668814958 

5555555555555555555555555 

Carcass ID Number 1111230010342042440414413 

4203547892263388041155779 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Intestine large, colon +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Intestine large, rectum ++++++++++++++++++++++A++ 

Intestine large, cecum A + + + A ++++++++++++++++++++ 

Intestine small, duodenum A ++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Intestine small, jejunum A + + + A +++++++++++++++++A++ 

Histiocytic sarcoma X 

Intestine small, ileum A+++A+++++++++++++++++A++ 

Histiocytic sarcoma X 

Liver +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Mesentery + +++ + 

Hemangioma X 

Pancreas +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Salivary glands +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Stomach, forestomach +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Stomach, glandular A ++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Cardiovascular System 

Heart 

Histiocytic sarcoma 

+++++++++++++++++++++++++ 

X 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adenoma 

Histiocytic sarcoma X 

Bilateral, carcinoma 

Adrenal medulla +++++++++++++++++++++++++ 

Pheochromocytoma benign 

Islets, pancreatic +++++++++++++++ I +++++++++ 

Parathyroid gland +++++++M+++++++MM+++M++++ 

Pituitary gland A++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Pars distalis, adenoma X X X X X X X X X X X X 

Thyroid gland A++++++++++++++++++++++++ 

Histiocytic sarcoma X 

C-cell, adenoma X X 

C-cell, carcinoma X 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 149 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

7777777777777777777777777 

Number of Days on Study 2233333333333333333333333 

2400000000000111111111111 

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 Total 

Carcass ID Number 4 3 0 0 0 1 1 1 2 2 2 3 4 0 2 2 2 2 3 3 3 3 3 4 5 Tissues/ 

3 3 4 5 6 1 6 8 0 2 7 6 9 9 1 4 6 9 0 1 5 7 8 2 0 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Intestine large, rectum + + + M + + + + + + + + + + + + + + + + + + + + + 48 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Histiocytic sarcoma 1 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Histiocytic sarcoma 1 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Histiocytic sarcoma 1 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Mesentery + ++ 8 

Hemangioma 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Histiocytic sarcoma 1 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X X 2 

Histiocytic sarcoma 1 

Bilateral, carcinoma X 1 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + M + + + 49 

Pheochromocytoma benign X 1 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Parathyroid gland + + + + + M + M M + + + + + + + + + + + M + + + + 42 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Histiocytic sarcoma 1 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X 30 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Histiocytic sarcoma 1 

C-cell, adenoma X X X 5 

C-cell, carcinoma X 2 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

150 2-Butoxyethanol, NTP TR 484 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

2344455555556666666666677 

Number of Days on Study 4966924478881344568899900 

7022694443699389668814958 

5555555555555555555555555 

Carcass ID Number 1111230010342042440414413 

4203547892263388041155779 

Genital System 

Clitoral gland +++++++++++++++++M+++M+++ 

Adenoma 

Carcinoma X X 

Histiocytic sarcoma X 

Ovary +++++++++++++++++++++++++ 

Granulosa cell tumor benign 

Histiocytic sarcoma X 

Uterus +++++++++++++++++++++++++ 

Hemangiosarcoma X 

Histiocytic sarcoma X 

Polyp stromal X 

Polyp stromal, multiple 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 

Lymph node + + + + + + 

Histiocytic sarcoma X 

Renal, histiocytic sarcoma X 

Lymph node, bronchial ++++++++++M+++++++MM+++M+ 

Histiocytic sarcoma X 

Lymph node, mandibular ++++++++++++++++M++M+++++ 

Histiocytic sarcoma X 

Lymph node, mesenteric +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Lymph node, mediastinal +++++++++M++++++++M++++++ 

Histiocytic sarcoma X 

Spleen +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Thymus +++++++++++++M+++++++++++ 

Histiocytic sarcoma X 

Integumentary System 

Mammary gland +++++++++++++++++++++++++ 

Carcinoma X X X 

Fibroadenoma X X X X X 

Fibroadenoma, multiple X X 

Histiocytic sarcoma X 

Skin +++++++++++++++++++++++++ 

Subcutaneous tissue, lipoma X 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Skeletal muscle + 

Hemangioma X 

Nervous System 

Brain +++++++++++++++++++++++++ 

Glioma malignant X 

Histiocytic sarcoma X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 151 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

7777777777777777777777777 

Number of Days on Study 2233333333333333333333333 

2400000000000111111111111 

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 Total 

Carcass ID Number 4 3 0 0 0 1 1 1 2 2 2 3 4 0 2 2 2 2 3 3 3 3 3 4 5 Tissues/ 

3 3 4 5 6 1 6 8 0 2 7 6 9 9 1 4 6 9 0 1 5 7 8 2 0 Tumors 

Genital System 

Clitoral gland + M + + + M + + M + + + + + + + + + + + + + + M + 44 

Adenoma X X XX 4 

Carcinoma X 3 

Histiocytic sarcoma 1 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Granulosa cell tumor benign X 1 

Histiocytic sarcoma 1 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma 1 

Histiocytic sarcoma 1 

Polyp stromal X X X X 5 

Polyp stromal, multiple X 1 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node 6 

Histiocytic sarcoma 1 

Renal, histiocytic sarcoma 1 

Lymph node, bronchial M M M + M + M M M + M + + + M + M M + M + + + + + 34 

Histiocytic sarcoma 1 

Lymph node, mandibular + + + + + + + + + + + + + + M + M + + + + + + + + 46 

Histiocytic sarcoma 1 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Lymph node, mediastinal + MMMM++++MM++++MM+++++++M 39 

Histiocytic sarcoma 1 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Thymus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Histiocytic sarcoma 1 

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma X 4 

Fibroadenoma X X X X X 10 

Fibroadenoma, multiple X X X X X X X X 10 

Histiocytic sarcoma 1 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Subcutaneous tissue, lipoma 1 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Skeletal muscle 1 

Hemangioma 1 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Glioma malignant 1 

Histiocytic sarcoma 1 
 

 

 

 

 

 

 

 

152 2-Butoxyethanol, NTP TR 484 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

2344455555556666666666677 

Number of Days on Study 4966924478881344568899900 

7022694443699389668814958 

5555555555555555555555555 

Carcass ID Number 1111230010342042440414413 

4203547892263388041155779 

Respiratory System 

Larynx +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Lung +++++++++++++++++++++++++ 

Carcinoma, metastatic, thyroid gland 

Histiocytic sarcoma X 

Nose +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Trachea +++++++++++++++++++++++++ 

Special Senses System 

Eye A + + 

Urinary System 

Kidney +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Urinary bladder A++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Leukemia mononuclear X X X X X XXXXXXX X X 
 

 

 

 

 

2-Butoxyethanol, NTP TR 484 153 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

7777777777777777777777777 

Number of Days on Study 2233333333333333333333333 

2400000000000111111111111 

5555555555555555555555555 Total 

Carcass ID Number 4300011122234022223333345 Tissues/ 

3345616802769914690157820 Tumors 

Respiratory System 

Larynx +++++++++++++++++++++++++ 50 

Histiocytic sarcoma 1 

Lung +++++++++++++++++++++++++ 50 

Carcinoma, metastatic, thyroid gland X 1 

Histiocytic sarcoma 1 

Nose +++++++++++++++++++++++++ 50 

Histiocytic sarcoma 1 

Trachea +++++++++++++++++++++++++ 50 

Special Senses System 

Eye + 3 

Urinary System 

Kidney +++++++++++++++++++++++++ 50 

Histiocytic sarcoma 1 

Urinary bladder +++++++++++++++++++++++++ 49 

Histiocytic sarcoma 1 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 50 

Histiocytic sarcoma 1 

Leukemia mononuclear X X X X X X X X X 23 
 

 

 

 

 

 

 

 

 

 

 

 

154 2-Butoxyethanol, NTP TR 484 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

4455566666666666666666666 

Number of Days on Study 6734711112345666668888999 

2231100274862233675999445 

7777777777777777777777777 

Carcass ID Number 4414403230211211232234221 

6518292084642778864579236 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Intestine large, colon +++++++++++++A+++++++++++ 

Intestine large, rectum ++++++++++ I ++A++++++++++A 

Intestine large, cecum +++++++++++++A++++++++++A 

Intestine small, duodenum +++++++++++++A++++++++++A 

Intestine small, jejunum +++++++++++++A++++++++++A 

Intestine small, ileum +++A+++++++++A++++++++++A 

Liver +++++++++++++++++++++++++ 

Mesentery + + 

Oral mucosa 

Pancreas +++++++++++++++++++++++++ 

Salivary glands +++++M+++++++++++++++++++ 

Stomach, forestomach +++++++++++++++++++++++++ 

Stomach, glandular +++++++++++++++++++++++++ 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adrenal medulla ++++++++++++++++++++++++M 

Pheochromocytoma malignant X 

Pheochromocytoma benign X X X X X 

Bilateral, pheochromocytoma benign X 

Islets, pancreatic +++++++++++++++++++++++++ 

Adenoma 

Parathyroid gland +++A++M++++M+++++++++++++ 

Pituitary gland +++A+++++++++++++++++++++ 

Pars distalis, adenoma X X X X X X X X X X 

Thyroid gland +++++++++++++++++++++++++ 

C-cell, adenoma X X X 

C-cell, carcinoma 

Follicular cell, adenoma 

Follicular cell, carcinoma X 

General Body System 

Peritoneum + 

Genital System 

Clitoral gland ++++++++++++++++M++++++++ 

Adenoma X 

Carcinoma X 

Ovary +++++++++++++++++++++++++ 

Granulosa-theca tumor malignant 

Uterus +++++++++++++++++++++++++ 

Carcinoma X 

Polyp stromal X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 155 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

7777777777777777777777777 

Number of Days on Study 0002333333333333333333333 

1559000000011111111111111 

7777777777777777777777777 Total 

Carcass ID Number 4440022334400000111133335 Tissues/ 

4016519353712378035901490 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Intestine large, colon +++++++++++++++++++++++++ 49 

Intestine large, rectum +++++++++++++++++++++++++ 47 

Intestine large, cecum +++++++++++++++++++++++++ 48 

Intestine small, duodenum +++++++++++++++++++++++++ 48 

Intestine small, jejunum +++++++++++++++++++++++++ 48 

Intestine small, ileum +++++++++++++++++++++++++ 47 

Liver +++++++++++++++++++++++++ 50 

Mesentery + 3 

Oral mucosa + 1 

Pancreas +++++++++++++++++++++++++ 50 

Salivary glands +++++++++++++++++++++++++ 49 

Stomach, forestomach +++++++++++++++++++++++++ 50 

Stomach, glandular +++++++++++++++++++++++++ 50 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 50 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 50 

Adrenal medulla +++++++++++++++++++++++++ 49 

Pheochromocytoma malignant 1 

Pheochromocytoma benign X 6 

Bilateral, pheochromocytoma benign 1 

Islets, pancreatic +++++++++++++++++++++++++ 50 

Adenoma X 1 

Parathyroid gland +++++M++++M++++++M++++++M 43 

Pituitary gland +++++++++++++++++++++++++ 49 

Pars distalis, adenoma XX X XXX XX X X 20 

Thyroid gland +++++++++++++++++++++++++ 50 

C-cell, adenoma X X 5 

C-cell, carcinoma X X 2 

Follicular cell, adenoma X 1 

Follicular cell, carcinoma 1 

General Body System 

Peritoneum 1 

Genital System 

Clitoral gland ++++++++++++++++++++++++ 48 

Adenoma X X X 4 

Carcinoma X 2 

Ovary +++++++++++++++++++++++++ 50 

Granulosa-theca tumor malignant X 1 

Uterus +++++++++++++++++++++++++ 50 

Carcinoma 1 

Polyp stromal X 2 
 

 

 

 

 

 

 

 

 

 

 

 

156 2-Butoxyethanol, NTP TR 484 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

4455566666666666666666666 

Number of Days on Study 6734711112345666668888999 

2231100274862233675999445 

7777777777777777777777777 

Carcass ID Number 4414403230211211232234221 

6518292084642778864579236 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+++++++++++++++++++++++++ 

+ 

M +++++M+M++M+MM++MMMMM+M+ 

+++++M+++++++++++++++++++ 

++++++++++++++M++++++++++ 

+++++M+M+++++MM++MM+M+MMM 

+++++++++++++++++++++++++ 

+++++++++++++M++M++++++++ 

Integumentary System 

Mammary gland 

Carcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Squamous cell papilloma 

Subcutaneous tissue, fibroma 

+++A+++++++++++++++++++++ 

X 

X X X X X X X 

X 

+++++++++++++++++++++++++ 

X 

Musculoskeletal System 

Bone 

Chordoma 

Skeletal muscle 

Sarcoma 

+++++++++++++++++++++++++ 

+ + 

X 

Nervous System 

Brain +++++++++++++++++++++++++ 

Respiratory System 

Larynx 

Lung 

Carcinoma, metastatic, mammary gland 

Carcinoma, metastatic, thyroid gland 

Chordoma, metastatic, bone 

Sarcoma, metastatic, skeletal muscle 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

+++++++++++++A+++++++++++ 

+++++++++++++++++++++++++ 

Special Senses System 

Eye 

Zymbal’s gland 

Carcinoma 

+ 

Urinary System 

Kidney 

Renal tubule, carcinoma 

Urinary bladder 

+++++++++++++++++++++++++ 

X 

+++++++++++++++++++++++++ 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

+++++++++++++++++++++++++ 

XXXXXXX XXX XX XXXXX 

X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 157 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

7777777777777777777777777 

Number of Days on Study 0002333333333333333333333 

1559000000011111111111111 

7777777777777777777777777 Total 

Carcass ID Number 4440022334400000111133335 

4016519353712378035901490 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+++++++++++++++++++++++++ 

+ 

+ M ++++MMMMMM++++++MMMM+M+ 

+ M ++++++++++++++++++M+M++ 

+++++++++++++++++++++++++ 

+++++MMMMMM+++++++MMMMM+M 

+++++++++++++++++++++++++ 

+++++++++++++M+M+M++++++M 

50 

2 

26 

46 

49 

28 

50 

44 

Integumentary System 

Mammary gland 

Carcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Squamous cell papilloma 

Subcutaneous tissue, fibroma 

+++++++++++++++++++++++++ 

X X 

X X X X X 

X X 

+++++++++++++++++++++++++ 

X 

X 

49 

3 

12 

3 

50 

2 

1 

Musculoskeletal System 

Bone 

Chordoma 

Skeletal muscle 

Sarcoma 

+++++++++++++++++++++++++ 

X 

50 

1 

2 

1 

Nervous System 

Brain +++++++++++++++++++++++++ 50 

Respiratory System 

Larynx 

Lung 

Carcinoma, metastatic, mammary gland 

Carcinoma, metastatic, thyroid gland 

Chordoma, metastatic, bone 

Sarcoma, metastatic, skeletal muscle 

Nose 

Trachea 

+++++++++++++++++++++++++ 

++++++++++++++++++++ ++++ 

X 

X 

X 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

49 

1 

1 

1 

1 

49 

50 

Special Senses System 

Eye 

Zymbal’s gland 

Carcinoma 

+ 

X 

1 

1 

1 

Urinary System 

Kidney 

Renal tubule, carcinoma 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

1 

50 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

+++++++++++++++++++++++++ 

X X X X X X X 

50 

24 

1 
158 2-Butoxyethanol, NTP TR 484 

TABLE B3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Adrenal Medulla: Benign Pheochromocytoma 

Overall ratea 3/50 (6%) 4/50 (8%) 1/49 (2%) 7/49 (14%)
 

Adjusted rateb 6.9% 9.2% 2.6% 16.7%
 

Terminal ratec 1/29 (3%) 1/27 (4%) 1/22 (5%) 1/21 (5%)
 

First incidence (days) 554 584 730 (T) 638
 

Poly-3 testd P=0.090 P=0.499 P=0.353N P=0.138 

Adrenal Medulla: Benign or Malignant Pheochromocytoma 

Overall rate 3/50 (6%) 4/50 (8%) 1/49 (2%) 8/49 (16%)
 

Adjusted rate 6.9% 9.2% 2.6% 18.9%
 

Terminal rate 1/29 (3%) 1/27 (4%) 1/22 (5%) 1/21 (5%)
 

First incidence (days) 554 584 730 (T) 612
 

Poly-3 test P=0.044 P=0.499 P=0.353N P=0.086 

Clitoral Gland: Adenoma 

Overall rate 6/46 (13%) 3/45 (7%) 4/44 (9%) 4/48 (8%)
 

Adjusted rate 14.9% 7.8% 11.8% 10.1%
 

Terminal rate 5/27 (19%) 1/23 (4%) 3/20 (15%) 3/20 (15%)
 

First incidence (days) 688 722 722 646
 

Poly-3 test P=0.385N P=0.262N P=0.479N P=0.374N
 

Clitoral Gland: Carcinoma 

Overall rate 3/46 (7%) 1/45 (2%) 3/44 (7%) 2/48 (4%)
 

Adjusted rate 7.5% 2.6% 8.7% 5.0%
 

Terminal rate 2/27 (7%) 1/23 (4%) 1/20 (5%) 1/20 (5%)
 

First incidence (days) 722 730 (T) 583 663
 

Poly-3 test P=0.523N P=0.320N P=0.594 P=0.503N
 

Clitoral Gland: Adenoma or Carcinoma 

Overall rate 9/46 (20%) 3/45 (7%) 7/44 (16%) 6/48 (13%)
 

Adjusted rate 22.4% 7.8% 20.2% 15.0%
 

Terminal rate 7/27 (26%) 1/23 (4%) 4/20 (20%) 4/20 (20%)
 

First incidence (days) 688 722 583 646
 

Poly-3 test P=0.395N P=0.066N P=0.521N P=0.285N
 

Mammary Gland: Fibroadenoma 

Overall rate 23/50 (46%) 24/50 (48%) 20/50 (40%) 15/50 (30%)
 

Adjusted rate 49.9% 53.5% 48.2% 33.9%
 

Terminal rate 14/29 (48%) 15/27 (56%) 12/23 (52%) 5/21 (24%)
 

First incidence (days) 508 584 462 462
 

Poly-3 test P=0.043N P=0.445 P=0.522N P=0.088N
 

Mammary Gland: Carcinoma 

Overall rate 3/50 (6%) 4/50 (8%) 4/50 (8%) 3/50 (6%) 

Adjusted rate 6.9% 9.1% 9.8% 7.2% 

Terminal rate 2/29 (7%) 0/27 (0%) 1/23 (4%) 1/21 (5%) 

First incidence (days) 638 606 462 663 

Poly-3 test P=0.571N P=0.507 P=0.464 P=0.642 
2-Butoxyethanol, NTP TR 484 159 

TABLE B3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Mammary Gland: Fibroadenoma or Carcinoma 

Overall rate 25/50 (50%) 27/50 (54%) 23/50 (46%) 18/50 (36%)
 

Adjusted rate 53.8% 58.9% 54.3% 40.4%
 

Terminal rate 15/29 (52%) 15/27 (56%) 13/23 (57%) 6/21 (29%)
 

First incidence (days) 508 584 462 462
 

Poly-3 test P=0.070N P=0.386 P=0.568 P=0.137N
 

Pituitary Gland (Pars Distalis): Adenoma 

Overall rate 33/50 (66%) 32/50 (64%) 30/49 (61%) 20/49 (41%)
 

Adjusted rate 72.8% 69.1% 69.5% 45.6%
 

Terminal rate 22/29 (76%) 19/27 (70%) 16/23 (70%) 9/21 (43%)
 

First incidence (days) 575 584 496 462
 

Poly-3 test P=0.002N P=0.430N P=0.452N P=0.005N
 

Skin: Squamous Cell Papilloma, Keratoacanthoma, or Basal Cell Carcinoma 

Overall rate 3/50 (6%) 1/50 (2%) 0/50 (0%) 2/50 (4%) 

Adjusted rate 6.9% 2.3% 0.0% 4.8% 

Terminal rate 2/29 (7%) 1/27 (4%) 0/23 (0%) 0/21 (0%) e First incidence (days) 710 730 (T) — 695 

Poly-3 test P=0.457N P=0.307N P=0.135N P=0.520N 

Thyroid Gland (C-cell): Adenoma 

Overall rate 6/50 (12%) 2/48 (4%) 5/49 (10%) 5/50 (10%)
 

Adjusted rate 13.8% 4.8% 12.6% 11.9%
 

Terminal rate 3/29 (10%) 0/27 (0%) 3/23 (13%) 2/21 (10%)
 

First incidence (days) 688 701 694 612
 

Poly-3 test P=0.514 P=0.145N P=0.564N P=0.523N
 

Thyroid Gland (C-cell): Adenoma or Carcinoma 

Overall rate 6/50 (12%) 2/48 (4%) 7/49 (14%) 6/50 (12%)
 

Adjusted rate 13.8% 4.8% 17.5% 14.3%
 

Terminal rate 3/29 (10%) 0/27 (0%) 4/23 (17%) 3/21 (14%)
 

First incidence (days) 688 701 649 612
 

Poly-3 test P=0.341 P=0.145N P=0.435 P=0.598 

Uterus: Stromal Polyp 

Overall rate 6/50 (12%) 3/50 (6%) 6/50 (12%) 2/50 (4%)
 

Adjusted rate 13.7% 7.0% 15.2% 4.8%
 

Terminal rate 5/29 (17%) 3/27 (11%) 5/23 (22%) 1/21 (5%)
 

First incidence (days) 554 730 (T) 705 662
 

Poly-3 test P=0.182N P=0.249N P=0.549 P=0.149N
 

All Organs: Mononuclear Cell Leukemia 

Overall rate 18/50 (36%) 21/50 (42%) 23/50 (46%) 24/50 (48%)
 

Adjusted rate 38.5% 44.3% 51.5% 51.2%
 

Terminal rate 9/29 (31%) 7/27 (26%) 8/23 (35%) 5/21 (24%)
 

First incidence (days) 400 495 390 533
 

Poly-3 test P=0.117 P=0.358 P=0.145 P=0.149 
160 

c 

2-Butoxyethanol, NTP TR 484 

TABLE B3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

All Organs: Benign Neoplasms 

Overall rate 42/50 (84%) 43/50 (86%) 40/50 (80%) 37/50 (74%) 

Adjusted rate 88.1% 91.3% 88.9% 78.9% 

Terminal rate 25/29 (86%) 25/27 (93%) 21/23 (91%) 17/21 (81%) 

First incidence (days) 508 584 462 462 

Poly-3 test P=0.059N P=0.418 P=0.588 P=0.163N 

All Organs: Malignant Neoplasms 

Overall rate 26/50 (52%) 26/50 (52%) 30/50 (60%) 32/50 (64%) 

Adjusted rate 54.7% 54.1% 63.2% 67.1% 

Terminal rate 14/29 (48%) 9/27 (33%) 9/23 (39%) 9/21 (43%) 

First incidence (days) 400 495 247 533 

Poly-3 test P=0.085 P=0.555N P=0.261 P=0.148 

All Organs: Benign or Malignant Neoplasms 

Overall rate 47/50 (94%) 48/50 (96%) 50/50 (100%) 48/50 (96%) 

Adjusted rate 94.0% 96.0% 100.0% 97.4% 

Terminal rate 26/29 (90%) 25/27 (93%) 23/23 (100%) 20/21 (95%) 

First incidence (days) 400 495 247 462 

Poly-3 test P=0.212 P=0.500 P=0.119 P=0.363 

(T)Terminal sacrifice 

a Number of neoplasm-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for adrenal gland, 

clitoral gland, pituitary gland, thyroid gland, and uterus; for other tissues, denominator is number of animals necropsied. b	 Poly-3 estimated neoplasm incidence after adjustment for intercurrent mortality 

Observed incidence at terminal kill 

d	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values 

corresponding to pairwise comparisons between the chamber controls and that exposed group. The Poly-3 test accounts for differential mortality in 

animals that do not reach terminal sacrifice. A negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group 
2-Butoxyethanol, NTP TR 484 161 

TABLE B4 

Historical Incidence of Adrenal Gland Pheochromocytoma in Chamber Control Female F344/N Ratsa 

Incidence in Controls 

Benign Complex Malignant Benign, Complex, 

or Malignantb 

Historical Incidence at Battelle Pacific Northwest Laboratories 

Acetonitrile 1/48 0/48 0/48 1/48 

Chloroprene 3/49 0/49 0/49 3/49 

Cobalt sulfate heptahydrate 2/48 0/48 0/48 2/48 

Furfuryl alcohol 4/50 0/50 1/50 5/50 

Hexachlorocyclopentadiene 6/47 0/47 0/47 6/47 

Isobutene 3/50 1/50 0/50 4/50 

Isobutyraldehyde 1/49 0/49 0/49 1/49 

Isoprene 1/50 0/50 1/50 2/50 

Molybdenum trioxide 5/49 1/49 0/49 6/49 

Nitromethane 1/49 1/49 0/49 2/49 

Ozone 6/50 0/50 0/50 6/50 

Tetrafluoroethane 4/50 0/50 0/50 4/50 

Tetrahydrofuran 0/50 0/50 2/50 2/50 

Overall Historical Incidence 

Total (%) 47/889 (5.3%) 5/889 (0.6%) 5/889 (0.6%) 57/889 (6.4%) 

Mean ± standard deviation 5.3% ± 3.9% 0.6% ± 1.2% 0.6% ± 1.1% 6.4% ± 3.5% 

Range 0%-13% 0%-4% 0%-4% 2%-13% 

a Data as of 12 November 1997 

b Includes data for unspecified pheochromocytomas 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

162 2-Butoxyethanol, NTP TR 484 

TABLE B5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 18 21 23 26 

Natural deaths 3 2 4 3 

Survivors 

Terminal sacrifice 29 27 23 21 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Intestine large, colon (49) (50) (50) (49) 

Necrosis 1 (2%) 

Parasite metazoan 4 (8%) 5 (10%) 2 (4%) 3 (6%) 

Intestine large, rectum (44) (47) (48) (47) 

Parasite metazoan 1 (2%) 4 (9%) 3 (6%) 4 (9%) 

Intestine large, cecum (47) (49) (48) (48) 

Inflammation, chronic 1 (2%) 

Parasite metazoan 8 (17%) 6 (12%) 4 (8%) 5 (10%) 

Intestine small, ileum (47) (50) (47) (47) 

Inflammation, chronic active 1 (2%) 1 (2%) 

Liver (50) (50) (50) (50) 

Angiectasis 2 (4%) 2 (4%) 1 (2%) 

Basophilic focus 39 (78%) 41 (82%) 41 (82%) 38 (76%) 

Clear cell focus 12 (24%) 8 (16%) 5 (10%) 9 (18%) 

Degeneration, cystic 1 (2%) 

Eosinophilic focus 13 (26%) 10 (20%) 6 (12%) 5 (10%) 

Fatty change 11 (22%) 12 (24%) 12 (24%) 6 (12%) 

Hematopoietic cell proliferation 4 (8%) 5 (10%) 1 (2%) 

Hepatodiaphragmatic nodule 9 (18%) 6 (12%) 3 (6%) 1 (2%) 

Inflammation, granulomatous 1 (2%) 

Mixed cell focus 6 (12%) 2 (4%) 4 (8%) 4 (8%) 

Necrosis 3 (6%) 4 (8%) 3 (6%) 3 (6%) 

Regeneration 1 (2%) 3 (6%) 

Vacuolization cytoplasmic, focal 1 (2%) 

Bile duct, hyperplasia 8 (16%) 8 (16%) 8 (16%) 5 (10%) 

Centrilobular, necrosis 2 (4%) 1 (2%) 1 (2%) 5 (10%) 

Kupffer cell, pigmentation 15 (30%) 19 (38%) 36 (72%) 47 (94%) 

Mesentery (12) (9) (8) (3) 

Artery, inflammation, chronic active 1 (13%) 

Fat, necrosis 12 (100%) 9 (100%) 6 (75%) 3 (100%) 

Oral mucosa (1) 

Pharyngeal, hyperplasia, squamous 1 (100%) 

Pancreas (50) (50) (50) (50) 

Atrophy 17 (34%) 15 (30%) 13 (26%) 13 (26%) 

Basophilic focus 2 (4%) 5 (10%) 4 (8%) 

Hyperplasia 1 (2%) 1 (2%) 

Salivary glands (50) (50) (50) (49) 

Atrophy 1 (2%) 1 (2%) 1 (2%) 

a Number of animals examined microscopically at the site and the number of animals with lesion 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 163 

TABLE B5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Alimentary System (continued) 

Stomach, forestomach (50) (50) (50) (50) 

Mineralization 1 (2%) 

Ulcer 2 (4%) 4 (8%) 7 (14%) 3 (6%) 

Epithelium, hyperplasia 3 (6%) 5 (10%) 6 (12%) 2 (4%) 

Stomach, glandular (50) (50) (49) (50) 

Mineralization 3 (6%) 3 (6%) 2 (4%) 8 (16%) 

Necrosis 2 (4%) 3 (6%) 4 (8%) 2 (4%) 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Cardiomyopathy 34 (68%) 31 (62%) 34 (68%) 30 (60%) 

Atrium, thrombosis 1 (2%) 2 (4%) 

Endocrine System 

Adrenal cortex (50) (50) (50) (50) 

Accessory adrenal cortical nodule 1 (2%) 

Atrophy 1 (2%) 1 (2%) 

Degeneration, cystic 1 (2%) 3 (6%) 

Hyperplasia 28 (56%) 24 (48%) 19 (38%) 20 (40%) 

Hypertrophy 18 (36%) 11 (22%) 9 (18%) 7 (14%) 

Necrosis 3 (6%) 4 (8%) 3 (6%) 

Vacuolization cytoplasmic 7 (14%) 9 (18%) 1 (2%) 5 (10%) 

Adrenal medulla (50) (50) (49) (49) 

Hyperplasia 11 (22%) 11 (22%) 8 (16%) 17 (35%) 

Islets, pancreatic (50) (50) (49) (50) 

Hyperplasia 2 (4%) 

Parathyroid gland (46) (48) (42) (43) 

Hyperplasia 1 (2%) 

Pituitary gland (50) (50) (49) (49) 

Angiectasis 1 (2%) 1 (2%) 1 (2%) 

Pars distalis, hyperplasia 15 (30%) 14 (28%) 11 (22%) 23 (47%) 

Pars intermedia, hyperplasia 1 (2%) 

Thyroid gland (50) (48) (49) (50) 

C-cell, hyperplasia 37 (74%) 41 (85%) 36 (73%) 29 (58%) 

Follicular cell, hyperplasia 1 (2%) 1 (2%) 2 (4%) 

General Body System 

None 

Genital System 

Clitoral gland (46) (45) (44) (48) 

Hyperplasia 7 (15%) 4 (9%) 2 (5%) 2 (4%) 

Ovary (50) (50) (50) (50) 

Cyst 9 (18%) 7 (14%) 3 (6%) 4 (8%) 

Hyperplasia 1 (2%) 

Inflammation, granulomatous 1 (2%) 1 (2%) 

Interstitial cell, hyperplasia 1 (2%) 

Uterus (50) (50) (50) (50) 

Prolapse 1 (2%) 

Endometrium, hyperplasia, cystic 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

164 2-Butoxyethanol, NTP TR 484 

TABLE B5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Hematopoietic System 

Bone marrow (50) (50) (50) (50) 

Atrophy 1 (2%) 

Hyperplasia 5 (10%) 2 (4%) 2 (4%) 6 (12%) 

Hyperplasia, histiocytic 2 (4%) 1 (2%) 1 (2%) 

Lymph node (5) (3) (6) (2) 

Renal, infiltration cellular, plasma cell 1 (17%) 

Spleen (50) (50) (50) (50) 

Fibrosis 2 (4%) 6 (12%) 7 (14%) 7 (14%) 

Hematopoietic cell proliferation 11 (22%) 17 (34%) 6 (12%) 4 (8%) 

Hemorrhage 1 (2%) 1 (2%) 

Necrosis 1 (2%) 

Pigmentation, hemosiderin 48 (96%) 48 (96%) 48 (96%) 49 (98%) 

Thymus (48) (47) (49) (44) 

Cyst 1 (2%) 

Integumentary System 

Mammary gland (50) (50) (50) (49) 

Galactocele 2 (4%) 

Hyperplasia, atypical 1 (2%) 

Skin (49) (50) (50) (50) 

Cyst 1 (2%) 

Hyperkeratosis 1 (2%) 1 (2%) 

Inflammation, chronic active 1 (2%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Hyperostosis 9 (18%) 10 (20%) 8 (16%) 9 (18%) 

Inflammation, chronic 1 (2%) 

Skeletal muscle (1) (2) 

Hemorrhage 1 (50%) 

Nervous System 

Brain (50) (50) (50) (50) 

Hydrocephalus 1 (2%) 

Mineralization 1 (2%) 

Necrosis 1 (2%) 

Respiratory System 

Larynx (50) (50) (50) (50) 

Epiglottis, metaplasia, squamous 1 (2%) 4 (8%) 2 (4%) 1 (2%) 

Lung (50) (50) (50) (49) 

Foreign body 1 (2%) 

Infiltration cellular, histiocyte 2 (4%) 1 (2%) 

Inflammation, chronic active 11 (22%) 11 (22%) 9 (18%) 13 (27%) 

Inflammation, granulomatous 1 (2%) 

Thrombosis 1 (2%) 

Alveolar epithelium, hyperplasia 17 (34%) 7 (14%) 8 (16%) 5 (10%) 

Artery, infiltration cellular, histiocyte 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 165 

TABLE B5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 31.2 ppm 62.5 ppm 125 ppm 

Respiratory System (continued) 

Nose (50) (48) (50) (49) 

Inflammation, chronic 3 (6%) 1 (2%) 

Inflammation, suppurative 9 (18%) 8 (17%) 5 (10%) 4 (8%) 

Thrombosis 2 (4%) 4 (8%) 6 (12%) 5 (10%) 

Olfactory epithelium, degeneration, hyaline 13 (26%) 18 (38%) 28 (56%) 40 (82%) 

Olfactory epithelium, metaplasia 3 (6%) 2 (4%) 3 (6%) 

Olfactory epithelium, metaplasia, squamous 1 (2%) 

Respiratory epithelium, hyperplasia 2 (4%) 2 (4%) 1 (2%) 

Respiratory epithelium, metaplasia, squamous 1 (2%) 3 (6%) 1 (2%) 

Trachea (50) (50) (50) (50) 

Infiltration cellular, polymorphonuclear 1 (2%) 

Special Senses System 

Eye (7) (3) (1) 

Cataract 5 (71%) 2 (67%) 1 (100%) 

Degeneration 1 (33%) 

Hemorrhage 1 (14%) 

Retina, atrophy 5 (71%) 1 (33%) 1 (100%) 

Urinary System 

Kidney (50) (50) (50) (50) 

Infarct 1 (2%) 2 (4%) 3 (6%) 

Inflammation, suppurative 1 (2%) 

Nephropathy 47 (94%) 48 (96%) 43 (86%) 46 (92%) 

Papilla, necrosis 1 (2%) 

Pelvis, inflammation, acute 1 (2%) 

Renal tubule, hyperplasia 1 (2%) 1 (2%) 

Renal tubule, necrosis 2 (4%) 

Urinary bladder (50) (50) (49) (50) 

Inflammation, granulomatous 1 (2%) 
166 2-Butoxyethanol, NTP TR 484 
167 

APPENDIX C 

SUMMARY OF LESIONS IN MALE MICE 

IN THE 2-YEAR INHALATION STUDY 

OF 2-BUTOXYETHANOL 

TABLE C1 

TABLE C2 

TABLE C3 

TABLE C4a 

TABLE C4b 

TABLE C5 

Summary of the Incidence of Neoplasms in Male Mice 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

Individual Animal Tumor Pathology of Male Mice 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

Statistical Analysis of Primary Neoplasms in Male Mice 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

Historical Incidence of Forestomach Squamous Cell Papilloma 

in Chamber Control Male B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 

Historical Incidence of Liver Neoplasms in Chamber Control Male B6C3F1 Mice . . . . . 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

168 

172 

192 

195 

196 

197 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

168 2-Butoxyethanol, NTP TR 484 

TABLE C1 

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 7 8 13 10 

Natural deaths 4 3 10 14 

Survivors 

Terminal sacrifice 39 39 27 26 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Intestine large, cecum (48) (48) (44) (43) 

Intestine small, jejunum (48) (47) (43) (38) 

Histiocytic sarcoma 1 (3%) 

Intestine small, ileum (48) (47) (43) (39) 

Carcinoma 1 (2%) 

Liver (50) (50) (49) (49) 

Carcinoma, metastatic, islets, pancreatic 1 (2%) 

Hemangiosarcoma 1 (2%) 2 (4%) 4 (8%) 

Hepatocellular carcinoma 5 (10%) 8 (16%) 12 (24%) 16 (33%) 

Hepatocellular carcinoma, multiple 5 (10%) 3 (6%) 4 (8%) 5 (10%) 

Hepatocellular adenoma 13 (26%) 10 (20%) 13 (27%) 14 (29%) 

Hepatocellular adenoma, multiple 9 (18%) 8 (16%) 5 (10%) 3 (6%) 

Hepatocholangiocarcinoma 1 (2%) 

Histiocytic sarcoma 2 (4%) 

Mesentery (5) (3) (3) (1) 

Histiocytic sarcoma 1 (100%) 

Oral mucosa (1) 

Squamous cell carcinoma 1 (100%) 

Pancreas (50) (50) (47) (47) 

Hemangioma 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Salivary glands (50) (50) (48) (50) 

Stomach, forestomach (50) (50) (49) (48) 

Histiocytic sarcoma 1 (2%) 

Squamous cell carcinoma 1 (2%) 

Squamous cell papilloma 1 (2%) 1 (2%) 2 (4%) 2 (4%) 

Stomach, glandular (50) (50) (46) (47) 

Adenoma 1 (2%) 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Hemangiosarcoma 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 169 

TABLE C1 

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Endocrine System 

Adrenal cortex (50) (50) (49) (50) 

Hepatocellular carcinoma, metastatic, liver 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Capsule, adenoma 2 (4%) 2 (4%) 2 (4%) 

Adrenal medulla (49) (50) (49) (49) 

Pheochromocytoma malignant 1 (2%) 

Islets, pancreatic (50) (50) (47) (46) 

Adenoma 2 (4%) 

Carcinoma 1 (2%) 

Thyroid gland (50) (48) (49) (50) 

Follicular cell, adenoma 2 (4%) 1 (2%) 1 (2%) 1 (2%) 

General Body System 

None 

Genital System 

Epididymis (50) (50) (49) (50) 

Hemangiosarcoma 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Leiomyoma 1 (2%) 

Prostate (49) (49) (48) (45) 

Seminal vesicle (50) (49) (46) (46) 

Histiocytic sarcoma 1 (2%) 

Testes (50) (50) (49) (50) 

Interstitial cell, adenoma 2 (4%) 

Hematopoietic System 

Bone marrow (50) (50) (49) (50) 

Hemangiosarcoma 1 (2%) 2 (4%) 

Histiocytic sarcoma 1 (2%) 

Lymph node (2) (1) (1) 

Schwannoma malignant, metastatic, spinal cord 1 (50%) 

Lymph node, bronchial (38) (35) (33) (35) 

Hepatocholangiocarcinoma, metastatic, liver 1 (3%)

 Lymph node, mandibular (24) (26) (32) (25) 

Lymph node, mesenteric (50) (50) (47) (43) 

Histiocytic sarcoma 1 (2%) 

Lymph node, mediastinal (40) (33) (40) (37) 

Hepatocholangiocarcinoma, metastatic, liver 1 (3%) 

Histiocytic sarcoma 1 (3%) 

Spleen (50) (50) (48) (49) 

Hemangiosarcoma 1 (2%) 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Mast cell tumor NOS 1 (2%) 

Thymus (43) (43) (36) (37) 

Histiocytic sarcoma 1 (3%) 
 

 

 

 

 

 

 

 

 

 

170 2-Butoxyethanol, NTP TR 484 

TABLE C1 

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Integumentary System 

Skin 

Hemangioma

Prepuce, histiocytic sarcoma

Subcutaneous tissue, hemangiosarcoma

(50) 

 1 (2%) 

 1 (2%) 

(50) (49) 

 1 (2%) 

(50) 

Musculoskeletal System 

Bone 

Hemangiosarcoma

(50) (50) 

 1 (2%) 

(50) (50) 

Nervous System 

Brain 

Spinal cord

Schwannoma malignant

(50) 

 (2) 

 1 (50%) 

(50) (49) (50) 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma

Alveolar/bronchiolar adenoma, multiple

Alveolar/bronchiolar carcinoma

Alveolar/bronchiolar carcinoma, multiple

Hepatocellular carcinoma, metastatic, liver

Hepatocholangiocarcinoma, metastatic, liver

Histiocytic sarcoma

Mediastinum, hepatocholangiocarcinoma,

 metastatic, liver

Nose 

(50) 

(50) 

 7 (14%)

 2 (4%)

 5 (10%)

 7 (14%)

 1 (2%) 

 1 (2%) 

(50) 

(50) 

(50) 

 7 (14%)

 2 (4%)

 5 (10%)

(50) 

(48) 

(49) 

 8 (16%)

 2 (4%) 

 2 (4%)

 1 (2%) 

 5 (10%)

(48) 

(49) 

(50) 

 8 (16%) 

 3 (6%) 

 3 (6%) 

 1 (2%) 

(48) 

Special Senses System 

Harderian gland

Adenoma

 (3)

 3 (100%)

 (3)

 3 (100%)

 (2)

 2 (100%)

 (2) 

 2 (100%) 

Urinary System 

Kidney 

Histiocytic sarcoma

Urinary bladder 

Histiocytic sarcoma

(50) 

(50) 

(50) 

(50) 

(47) 

(46) 

(50) 

 1 (2%) 

(45) 

 1 (2%) 

Systemic Lesions 

Multiple organsb 

Histiocytic sarcoma

Lymphoma malignant

(50) 

 1 (2%)

 1 (2%)

(50) 

 3 (6%) 

(50) (50) 

 2 (4%) 
171 

c 

2-Butoxyethanol, NTP TR 484 

TABLE C1 

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Neoplasm Summary 

Total animals with primary neoplasmsc 42 33 36 41 

Total primary neoplasms 62 55 60 69 

Total animals with benign neoplasms 30 28 26 28 

Total benign neoplasms 42 32 37 33 

Total animals with malignant neoplasms 19 16 19 28 

Total malignant neoplasms 20 22 23 36 

Total animals with metastatic neoplasms 9 5 5 4 

Total metastatic neoplasms 12 6 5 4 

Total animals with benign or malignant neoplasms

 of uncertain primary site 1 

Total uncertain neoplasms 1 

a Number of animals examined microscopically at the site and the number of animals with neoplasm b Number of animals with any tissue examined microscopically 

Primary neoplasms: all neoplasms except metastatic neoplasms 
 

 

 

 

 

 

 

 

172 2-Butoxyethanol, NTP TR 484 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

3455566666677777777777777 

Number of Days on Study 7556612466622222222222222 

4961581237999999999999999 

0000000000000000000000000 

Carcass ID Number 4151214331100001122333334 

2408200395725672945125681 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Gallbladder MM+M++++A+M+++++++M++++++ 

Intestine large, colon ++++++++A++++++++++++++++ 

Intestine large, rectum +++++++++++++++++++++++++ 

Intestine large, cecum + + A +++++A++++++++++++++++ 

Intestine small, duodenum + + A ++++++++++++++++++++++ 

Intestine small, jejunum + + A +++++A++++++++++++++++ 

Intestine small, ileum +++++M++A++++++++++++++++ 

Liver +++++++++++++++++++++++++ 

Hepatocellular carcinoma X X X X X 

Hepatocellular carcinoma, multiple X X X X X 

Hepatocellular adenoma X X X X X X 

Hepatocellular adenoma, multiple X 

Hepatocholangiocarcinoma X 

Mesentery +
 

Pancreas +++++++++++++++++++++++++ 
 

Salivary glands +++++++++++++++++++++++++ 
 

Stomach, forestomach +++++++++++++++++++++++++ 
 

Squamous cell papilloma X 

Stomach, glandular +++++++++++++++++++++++++ 

Adenoma 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Capsule, adenoma X 

Adrenal medulla +++++++++++++M+++++++++++ 

Islets, pancreatic +++++++++++++++++++++++++ 

Adenoma X X 

Parathyroid gland +M++M+MM+M++M++M++MM+M++M 

Pituitary gland +++++++++++++++++++++++++ 

Thyroid gland +++++++++++++++++++++++++ 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Epididymis +++++++++++++++++++++++++ 

Preputial gland ++++++++++M++++++++++++++ 

Prostate +++++++++++++++++++++++++ 

Seminal vesicle +++++++++++++++++++++++++ 

Testes +++++++++++++++++++++++++ 

+: Tissue examined microscopically M: Missing tissue X: Lesion present 

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined 
 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 173 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

7777777777777777777777777 

Number of Days on Study 2223333333333333333333333 

9990000000000000000000011 

0000000000000000000000000 Total 

Carcass ID Number 4440000112222222333444401 Tissues/ 

3561489130136789047478936 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Gallbladder ++++++M+++++++++++++++M++ 42 

Intestine large, colon +++++++++++++++++++++++++ 49 

Intestine large, rectum +++++++++++++++++++++++++ 50 

Intestine large, cecum +++++++++++++++++++++++++ 48 

Intestine small, duodenum +++++++++++++++++++++++++ 49 

Intestine small, jejunum +++++++++++++++++++++++++ 48 

Intestine small, ileum +++++++++++++++++++++++++ 48 

Liver +++++++++++++++++++++++++ 50 

Hepatocellular carcinoma 5 

Hepatocellular carcinoma, multiple 5 

Hepatocellular adenoma X XXXX X X 13 

Hepatocellular adenoma, multiple X X X X XXXX 9 

Hepatocholangiocarcinoma 1 

Mesentery + ++ + 5 

Pancreas +++++++++++++++++++++++++ 50 

Salivary glands +++++++++++++++++++++++++ 50 

Stomach, forestomach +++++++++++++++++++++++++ 50 

Squamous cell papilloma 1 

Stomach, glandular +++++++++++++++++++++++++ 50 

Adenoma X 1 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 50 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 50 

Capsule, adenoma X 2 

Adrenal medulla +++++++++++++++++++++++++ 49 

Islets, pancreatic +++++++++++++++++++++++++ 50 

Adenoma 2 

Parathyroid gland M+MM++MMM++MMM+M++MM++++M 26 

Pituitary gland +++++++++M+++++++++++++++ 49 

Thyroid gland +++++++++++++++++++++++++ 50 

Follicular cell, adenoma X X 2 

General Body System 

None 

Genital System 

Epididymis +++++++++++++++++++++++++ 50 

Preputial gland +++++++++++++++++++++++++ 49 

Prostate +++++M+++++++++++++++++++ 49 

Seminal vesicle +++++++++++++++++++++++++ 50 

Testes +++++++++++++++++++++++++ 50 
 

 

 

 

 

 

 

 

 

 

174 2-Butoxyethanol, NTP TR 484 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

3455566666677777777777777 

Number of Days on Study 7556612466622222222222222 

4961581237999999999999999 

0000000000000000000000000 

Carcass ID Number 4151214331100001122333334 

2408200395725672945125681 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 

Lymph node 

Schwannoma malignant, metastatic, spinal cord 

Lymph node, bronchial 

Hepatocholangiocarcinoma, metastatic, liver 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Hepatocholangiocarcinoma, metastatic, liver 

Spleen 

Thymus 

+ + 

X 

+++++++++MM++M+++++++++M+ 

X 

M + MMMM++++M++MMM++M++M+++ 

+++++++++++++++++++++++++ 

+++M+++M+++++++++++++M++M 

X 

+++++++++++++++++++++++++ 

++M+++++MMM++++M+++++++++ 

Integumentary System 

Mammary gland 

Skin 

MMMMMMMMMMMMMMMMMMMMMMMMM 

+++++++++++++++++++++++++ 

Prepuce, histiocytic sarcoma 

Subcutaneous tissue, hemangiosarcoma X 

X 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Skeletal muscle + 

Nervous System 

Brain +++++++++++++++++++++++++ 

Spinal cord 

Schwannoma malignant 

+ 

X 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Hepatocholangiocarcinoma, metastatic, liver 

Mediastinum, hepatocholangiocarcinoma,

 metastatic, liver 

Nose 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X X X X 

X X X 

X X X X X X X 

X 

X 

+++++++++++++++++++++++++ 

Pleura 

Trachea +++++++++++++++++++++++++ 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 175 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

7777777777777777777777777 

Number of Days on Study 2223333333333333333333333 

9990000000000000000000011 

0000000000000000000000000 Total 

Carcass ID Number 4440000112222222333444401 

3561489130136789047478936 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 50 

Lymph node 

Schwannoma malignant, metastatic, spinal cord 

Lymph node, bronchial 

Hepatocholangiocarcinoma, metastatic, liver 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Hepatocholangiocarcinoma, metastatic, liver 

Spleen 

Thymus 

M ++++++++M+MMM++M+M++++M+ 

M + M + + M + MMM++MM+MMMMM+++MM 

+++++++++++++++++++++++++ 

M + M +++++++++MM++M++M+++++ 

+++++++++++++++++++++++++ 

+++M+++++++++++++++M+++++ 

2 

1 

38 

1 

24 

50 

40 

1 

50 

43 

Integumentary System 

Mammary gland 

Skin 

M M M M M M M M M M M M M M M M M M M M M M M MM 

+++++++++++++++++++++++++ 50 

Prepuce, histiocytic sarcoma 

Subcutaneous tissue, hemangiosarcoma 

1 

1 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 50 

Skeletal muscle 1 

Nervous System 

Brain +++++++++++++++++++++++++ 50 

Spinal cord 

Schwannoma malignant 

+ 2 

1 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Hepatocholangiocarcinoma, metastatic, liver 

Mediastinum, hepatocholangiocarcinoma, 

metastatic, liver 

Nose 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X 

X X 

X X 

+++++++++++++++++++++++++ 

50 

50 

7 

2 

5 

7 

1 

1 

50 

Pleura + 1 

Trachea +++++++++++++++++++++++++ 50 
 

 

 

176 2-Butoxyethanol, NTP TR 484 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

3455566666677777777777777 

Number of Days on Study 7556612466622222222222222 

4961581237999999999999999 

0000000000000000000000000 

Carcass ID Number 4151214331100001122333334 

2408200395725672945125681 

Special Senses System 

Ear 

Eye 

Harderian gland + 

Adenoma X 

Urinary System 

Kidney +++++++++++++++++++++++++ 

Urinary bladder +++++++++++++++++++++++++ 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Lymphoma malignant X 
 

 

 

 

 

2-Butoxyethanol, NTP TR 484 177 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 

Chamber Control 

7777777777777777777777777 

Number of Days on Study 2223333333333333333333333 

9990000000000000000000011 

0000000000000000000000000 Total 

Carcass ID Number 4440000112222222333444401 Tissues/ 

3561489130136789047478936 Tumors 

Special Senses System 

Ear + 1 

Eye + 1 

Harderian gland + + 3 

Adenoma X X 3 

Urinary System 

Kidney +++++++++++++++++++++++++ 50 

Urinary bladder +++++++++++++++++++++++++ 50 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 50 

Histiocytic sarcoma 1 

Lymphoma malignant 1 
 

 

 

 

 

 

 

 

 

178 2-Butoxyethanol, NTP TR 484 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

5666666677777777777777777 

Number of Days on Study 4124578800022222222222222 

9119601115799999999999999 

2222222222222222222222222 

Carcass ID Number 1131041423200001122223333 

5641784983924698934572678 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Gallbladder A++M++M+A++++++++++++++++ 

Intestine large, colon +++++++++++++++++++++++++ 

Intestine large, rectum ++++++++A+M++++++++++++++ 

Intestine large, cecum A +++++++A++++++++++++++++ 

Intestine small, duodenum ++++++++A+A++++++++++++++ 

Intestine small, jejunum A +++++++A+A++++++++++++++ 

Intestine small, ileum A+++++++A+A++++++++++++++ 

Carcinoma X 

Liver +++++++++++++++++++++++++ 

Hemangiosarcoma X 

Hepatocellular carcinoma X X X X X 

Hepatocellular carcinoma, multiple X X 

Hepatocellular adenoma X X X X X X 

Hepatocellular adenoma, multiple X X X X X 

Mesentery + + 
 

Pancreas +++++++++++++++++++++++++ 
 

Salivary glands +++++++++++++++++++++++++ 
 

Stomach, forestomach +++++++++++++++++++++++++ 
 

Squamous cell papilloma 

Stomach, glandular +++++++++++++++++++++++++ 

Tooth + + 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Hepatocellular carcinoma, metastatic, liver 

Capsule, adenoma X X 

Adrenal medulla +++++++++++++++++++++++++ 

Pheochromocytoma malignant X 

Islets, pancreatic +++++++++++++++++++++++++ 

Parathyroid gland +M+++MMMMM+++M++MM++M+++M 

Pituitary gland ++++++M++++++++++++++++++ 

Thyroid gland +++++++M+++M+++++++++++++ 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Epididymis +++++++++++++++++++++++++ 

Preputial gland +++++++++++++++++++++++++ 

Prostate +++++++++++++++++++++++++ 

Seminal vesicle A++++++++++++++++++++++++ 

Testes +++++++++++++++++++++++++ 
 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 179 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

7777777777777777777777777 

Number of Days on Study 2222333333333333333333333 

9999000000000000000000111 

2222222222222222222222222 Total 

Carcass ID Number 3445000011122233344444124 Tissues/ 

9360135823712601501245007 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Gallbladder +++++++++++++++M+++++++++ 45 

Intestine large, colon +++++++++++++++++++++++++ 50 

Intestine large, rectum +++++++++++++++++++++++++ 48 

Intestine large, cecum +++++++++++++++++++++++++ 48 

Intestine small, duodenum +++++++++++++++++++++++++ 48 

Intestine small, jejunum +++++++++++++++++++++++++ 47 

Intestine small, ileum +++++++++++++++++++++++++ 47 

Carcinoma 1 

Liver +++++++++++++++++++++++++ 50 

Hemangiosarcoma 1 

Hepatocellular carcinoma X X X 8 

Hepatocellular carcinoma, multiple X 3 

Hepatocellular adenoma X X X X 10 

Hepatocellular adenoma, multiple X X X 8 

Mesentery + 3 

Pancreas +++++++++++++++++++++++++ 50 

Salivary glands +++++++++++++++++++++++++ 50 

Stomach, forestomach +++++++++++++++++++++++++ 50 

Squamous cell papilloma X 1 

Stomach, glandular +++++++++++++++++++++++++ 50 

Tooth + + 4 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 50 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 50 

Hepatocellular carcinoma, metastatic, liver X 1 

Capsule, adenoma 2 

Adrenal medulla +++++++++++++++++++++++++ 50 

Pheochromocytoma malignant 1 

Islets, pancreatic +++++++++++++++++++++++++ 50 

Parathyroid gland +M+++++M++MM++M+MMM+++M+M 29 

Pituitary gland +++++++++++++++++++++++++ 49 

Thyroid gland +++++++++++++++++++++++++ 48 

Follicular cell, adenoma X 1 

General Body System 

None 

Genital System 

Epididymis +++++++++++++++++++++++++ 50 

Preputial gland ++++++++++M++++++++++++++ 49 

Prostate +++++++++++++++++++++++M+ 49 

Seminal vesicle +++++++++++++++++++++++++ 49 

Testes +++++++++++++++++++++++++ 50 
 

 

 

 

 

 

 

 

 

 

180 2-Butoxyethanol, NTP TR 484 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

Number of Days on Study 

5666666677777777777777777 

4124578800022222222222222 

9119601115799999999999999 

Carcass ID Number 

2222222222222222222222222 

1131041423200001122223333 

5641784983924698934572678 

Hematopoietic System 

Bone marrow 

Hemangiosarcoma 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Hemangiosarcoma 

Mast cell tumor NOS 

Thymus 

+++++++++++++++++++++++++ 

X 

+ 

M +++M+M++++++M++++MM+++M+ 

M M + + M + M + + MMMMM+M+M++++M++ 

+++++++++++++++++++++++++ 

+++M++M+++M+M+M+++MMM++++ 

+++++++++++++++++++++++++ 

X 

++++M++M++M+M++++++++++++ 

Integumentary System 

Mammary gland 

Skin 

MMMMMMMMMMMMMMMMMMMMMMMMM 

+++++++++++++++++++++++++ 

Musculoskeletal System 

Bone 

Hemangiosarcoma 

+++++++++++++++++++++++++ 

X 

Nervous System 

Brain +++++++++++++++++++++++++ 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X X X 

X X 

X X X 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

Special Senses System 

Harderian gland 

Adenoma 

Zymbal’s gland 

+ 

X 

+ 

Urinary System 

Kidney 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

Systemic Lesions 

Multiple organs 

Lymphoma malignant 

+++++++++++++++++++++++++ 

X X 
 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 181 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

Number of Days on Study 

7777777777777777777777777 

2222333333333333333333333 

9999000000000000000000111 

Carcass ID Number 

2222222222222222222222222 

3445000011122233344444124 

9360135823712601501245007 

Total 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Hemangiosarcoma 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Hemangiosarcoma 

Mast cell tumor NOS 

Thymus 

+++++++++++++++++++++++++ 

++++++M+MM+M+++MMM++++++M 

+ + M +++MMM++++M++M+MMMMM+M 

+++++++++++++++++++++++++ 

+ M + M ++++++++M+MM+++MMM++M 

+++++++++++++++++++++++++ 

X 

+++M+M+++++M+++++++++++++ 

50 

1 

1 

35 

26 

50 

33 

50 

1 

1 

43 

Integumentary System 

Mammary gland 

Skin 

M M M M M M M M M M M M M M M M M M M M M M M MM 

+++++++++++++++++++++++++ 50 

Musculoskeletal System 

Bone 

Hemangiosarcoma 

+++++++++++++++++++++++++ 50 

1 

Nervous System 

Brain +++++++++++++++++++++++++ 50 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X X 

X X 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

50 

7 

2 

5 

50 

50 

Special Senses System 

Harderian gland 

Adenoma 

Zymbal’s gland 

+ 

X 

+ 

X 

3 

3 

1 

Urinary System 

Kidney 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

50 

Systemic Lesions 

Multiple organs 

Lymphoma malignant 

+++++++++++++++++++++++++ 

X 

50 

3 
 

 

 

 

 

 

 

182 2-Butoxyethanol, NTP TR 484 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

4445555555555666666667777 

Number of Days on Study 2350136688889014567890022 

1072173723357087530184599 

4444444444444444444444444 

Carcass ID Number 3331423110111324442035400 

9839485185065110359100167 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Gallbladder A+M+A++A+++MA A+A+++++++++ 

Intestine large, colon A+A+A+++++++A++++++++++++ 

Intestine large, rectum A+A+A+++++++A++++++++++++ 

Intestine large, cecum A + A ++++++++A A A+A+++++++++ 

Intestine small, duodenum A + A + A ++++++A A++A+++++++++ 

Intestine small, jejunum A + A + A ++++++A A++A+++++A+++ 

Intestine small, ileum A+A+A++++++A A++A+++++A+++ 

Liver A++++++++++++++++++++++++ 

Hemangiosarcoma X 

Hepatocellular carcinoma X X X X X X X X X X 

Hepatocellular carcinoma, multiple X X 

Hepatocellular adenoma X X X X X 

Hepatocellular adenoma, multiple X 

Mesentery + 

Oral mucosa 

Squamous cell carcinoma 

Pancreas A+A+A++++++++++++++++++++ 

Hemangioma 

Salivary glands A++++++++++++++M+++++++++ 

Stomach, forestomach A++++++++++++++++++++++++ 

Squamous cell carcinoma 

Squamous cell papilloma 

Stomach, glandular A + A + A +++++++A++++++++++++ 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex M++++++++++++++++++++++++ 

Capsule, adenoma X 

Adrenal medulla M++++++++++++++++++++++++ 

Islets, pancreatic A+A+A++++++++++++++++++++ 

Parathyroid gland M+M+M+MM++M+MM+M+++MM+MM+ 

Pituitary gland A+A+++++++++++++++++M+M++ 

Thyroid gland M++++++++++++++++++++++++ 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Epididymis A++++++++++++++++++++++++ 

Penis 

Preputial gland A++++++++++++++++++++++++ 

Prostate A+A++++++++++++++++++++++ 

Seminal vesicle A+A+A+++++++A++++++++++++ 

Testes A++++++++++++++++++++++++ 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 183 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

7777777777777777777777777 

Number of Days on Study 2222222222333333333333333 

9999999999000000000001111 

4444444444444444444444444 Total 

Carcass ID Number 0122233444000111223440223 Tissues/ 

8702527279249234676683344 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Gallbladder ++++++++++++M++++++++M+++ 40 

Intestine large, colon +++++++++++++++++++++++++ 46 

Intestine large, rectum +++++++++++++++++++++++++ 46 

Intestine large, cecum +++++++++++++++++++++++++ 44 

Intestine small, duodenum +++++++++++++++++++++++++ 44 

Intestine small, jejunum +++++++++++++++++++++++++ 43 

Intestine small, ileum +++++++++++++++++++++++++ 43 

Liver +++++++++++++++++++++++++ 49 

Hemangiosarcoma X 2 

Hepatocellular carcinoma X X 12 

Hepatocellular carcinoma, multiple X X 4 

Hepatocellular adenoma X X X X X X X X 13 

Hepatocellular adenoma, multiple X X X X 5 

Mesentery + + 3 

Oral mucosa + 1 

Squamous cell carcinoma X 1 

Pancreas +++++++++++++++++++++++++ 47 

Hemangioma X 1 

Salivary glands +++++++++++++++++++++++++ 48 

Stomach, forestomach +++++++++++++++++++++++++ 49 

Squamous cell carcinoma X 1 

Squamous cell papilloma X X 2 

Stomach, glandular +++++++++++++++++++++++++ 46 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 50 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 49 

Capsule, adenoma X 2 

Adrenal medulla +++++++++++++++++++++++++ 49 

Islets, pancreatic +++++++++++++++++++++++++ 47 

Parathyroid gland +++M+++++MMMMM+MM++M++MM+ 26 

Pituitary gland +++++++++++++++++++++++++ 46 

Thyroid gland +++++++++++++++++++++++++ 49 

Follicular cell, adenoma X 1 

General Body System 

None 

Genital System 

Epididymis +++++++++++++++++++++++++ 49 

Penis + 1 

Preputial gland +++++++++++++++++++++++++ 49 

Prostate +++++++++++++++++++++++++ 48 

Seminal vesicle +++++++++++++++++++++++++ 46 

Testes +++++++++++++++++++++++++ 49 
 

 

 

184 2-Butoxyethanol, NTP TR 484 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

4445555555555666666667777 

Number of Days on Study 2350136688889014567890022 

1072173723357087530184599 

4444444444444444444444444 

Carcass ID Number 3331423110111324442035400 

9839485185065110359100167 

Hematopoietic System 

Bone marrow 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

A++++++++++++++++++++++++ 

A M M ++++M+MMM++MM+M+MM+M++ 

A + M +++M+++MM+++M+++++M+++ 

A + M ++++++++++++++M+++++++ 

A +++M++++M+++M+++++M+++++ 

A+A++++++++++++++++++++++ 

A+M+M++++M+++++++M+M++M++ 

Integumentary System 

Mammary gland 

Skin 

Hemangioma 

MMMMMMMMMMMMMMMMMMMMMMMMM 

A++++++++++++++++++++++++ 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Nervous System 

Brain A++++++++++++++++++++++++ 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Alveolar/bronchiolar carcinoma, multiple 

Hepatocellular carcinoma, metastatic, liver 

Nose 

Trachea 

A+++A++++++++++++++++++++ 

A++++++++++++++++++++++++ 

X X X X X 

X 

X 

X X X 

A+++A++++++++++++++++++++ 

A+A+A++++++++++++++++++++ 

Special Senses System 

Eye 

Harderian gland 

Adenoma 

Urinary System 

Kidney 

Urinary bladder 

A+A++++++++++A+++++++++++ 

A+A+A++++++++A+++++++++++ 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 
 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 185 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

Number of Days on Study 

7777777777777777777777777 

2222222222333333333333333 

9999999999000000000001111 

Carcass ID Number 

4444444444444444444444444 

0122233444000111223440223 

8702527279249234676683344 

Total 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+++++++++++++++++++++++++ 

++++++++M++++M++++MM+++++ 

M + M M + + M + M M + M + + M ++++M+MM++ 

+++++++++++++++++++++++++ 

+++++++++M+M++MM+++M+++++ 

+++++++++++++++++++++++++ 

+++M+M+M+M+++M+++++++++MM 

49 

33 

32 

47 

40 

48 

36 

Integumentary System 

Mammary gland 

Skin 

Hemangioma 

M M M M M M M M M M M M M M M M M M M M M M M MM 

+++++++++++++++++++++++++ 

X 

49 

1 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 50 

Nervous System 

Brain +++++++++++++++++++++++++ 49 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Alveolar/bronchiolar carcinoma, multiple 

Hepatocellular carcinoma, metastatic, liver 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X X 

X 

X 

X 

X X 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

48 

49 

8 

2 

2 

1 

5 

48 

47 

Special Senses System 

Eye 

Harderian gland 

Adenoma 

+ 

X 

+ 

+ 

X 

1 

2 

2 

Urinary System 

Kidney 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

47 

46 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 50 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

186 2-Butoxyethanol, NTP TR 484 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 250 ppm 

Number of Days on Study 

0333444566666666666666777 

7167045901122366778899012 

3284224643439835004444219 

Carcass ID Number 

6666666666666666666666666 

1431033513244040240102430 

7085236042952579298346147 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Histiocytic sarcoma 

Intestine small, ileum 

Liver 

Carcinoma, metastatic, islets, pancreatic 

Hemangiosarcoma 

Hepatocellular carcinoma 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Mesentery 

Histiocytic sarcoma 

Pancreas 

Histiocytic sarcoma 

Salivary glands 

Stomach, forestomach 

Histiocytic sarcoma 

Squamous cell papilloma 

Stomach, glandular 

+++++++++++++++++++++++++ 

+A A+A A A A A A+A+A+A++M ++++M+ 

+++++++A+++++++A+++++++++ 

+++++ A A A +++++++ A +++++++++ 

++++ A A A A A++A+++A +++++++++ 

+ A + + A A A A A++A+++A +++++++++ 

+ A A + AAAAAA+A+A+A +++++++A+ 

X 

+A A A A A A A A++A+A+A +++++++++ 

+++++++A+++++++++++++++++ 

X X 

XXXXX X X X X X X 

X X X X 

X X X X X X X 

X 

X X 

+ 

X 

+ + + + + + A A + + + + + + + A + + + + + + + + + 

X 

+++++++++++++++++++++++++ 

++++++ A A +++++++++++++++++ 

X 

X 

+ + + + + + A A + + + + + + + A + + + + + + + + + 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Hemangiosarcoma X 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Adrenal medulla +++++++++++++++++++++++++ 

Islets, pancreatic ++++++ A A +++++++ A +++++++++ 

Carcinoma 

Parathyroid gland +M+++M+MM++++MM+MMM+++++M 

Pituitary gland ++++++M++++++++++++++++++ 

Thyroid gland +++++++++++++++++++++++++ 

Follicular cell, adenoma X 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 187 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 250 ppm 

7777777777777777777777777 

Number of Days on Study 2222222222223333333333333 

9999999999990000000000011 

6666666666666666666666666 Total 

Carcass ID Number 1111222233440001122334423 Tissues/ 

0689137859381361204014657 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Gallbladder ++M++M+++++++++++++++++++ 35 

Intestine large, colon +++++++++++++++++++++++++ 48 

Intestine large, rectum +++++++++++++++++++++M+++ 45 

Intestine large, cecum +++++++++++++++++++++++++ 43 

Intestine small, duodenum +++++M+++++++++++++++++++ 41 

Intestine small, jejunum +++++++++++++++++++++++++ 38 

Histiocytic sarcoma 1 

Intestine small, ileum +++++++++++++++++++++++++ 39 

Liver +++++++++++++++++++++++++ 49 

Carcinoma, metastatic, islets, pancreatic X 1 

Hemangiosarcoma X X 4 

Hepatocellular carcinoma X X X X X 16 

Hepatocellular carcinoma, multiple X 5 

Hepatocellular adenoma X X X X X X X 14 

Hepatocellular adenoma, multiple X X 3 

Histiocytic sarcoma 2 

Mesentery 1 

Histiocytic sarcoma 1 

Pancreas +++++++++++++++++++++++++ 47 

Histiocytic sarcoma 1 

Salivary glands +++++++++++++++++++++++++ 50 

Stomach, forestomach +++++++++++++++++++++++++ 48 

Histiocytic sarcoma 1 

Squamous cell papilloma X 2 

Stomach, glandular +++++++++++++++++++++++++ 47 

Cardiovascular System 

Heart 

Hemangiosarcoma 

+++++++++++++++++++++++++ 50 

1 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 50 

Histiocytic sarcoma 1 

Adrenal medulla +++++++++++++++++++++++M+ 49 

Islets, pancreatic +++++++++++++++M+++++++++ 46 

Carcinoma X 1 

Parathyroid gland M++MMM++++++M++MMM++++M++ 31 

Pituitary gland +++++++++++M+++++++++++++ 48 

Thyroid gland +++++++++++++++++++++++++ 50 

Follicular cell, adenoma 1 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

 

188 2-Butoxyethanol, NTP TR 484 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 250 ppm 

0333444566666666666666777 

Number of Days on Study 7167045901122366778899012 

3284224643439835004444219 

6666666666666666666666666 

Carcass ID Number 1431033513244040240102430 

7085236042952579298346147 

Genital System 

Epididymis 

Hemangiosarcoma 

Histiocytic sarcoma 

Leiomyoma 

Preputial gland 

Prostate 

+++++++++++++++++++++++++ 

X 

+++++++M+++++++++++++++++ 

+++++A+A+++++++A+++++++++ 

Seminal vesicle ++++++ A A A ++++++ A +++++++++ 

Histiocytic sarcoma 

Testes 

X 

+++++++++++++++++++++++++ 

Interstitial cell, adenoma 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 

Hemangiosarcoma 

Histiocytic sarcoma 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Histiocytic sarcoma 

Lymph node, mediastinal 

Histiocytic sarcoma 

Spleen 

Hemangiosarcoma 

Histiocytic sarcoma 

Thymus 

Histiocytic sarcoma 

X 

X 

+ 

+++++++++M++MM+M++M+MM+++ 

+ M + M + MMM+MM++++MM++MMMMM+ 

++++M++A+++++M+A+++++++M+ 

X 

+ M + MMM+++++M++++++++++++M 

X 

+++++++A+++++++++++++++++ 

X 

++++MM+++MM++M++++M+M+++M 

X 

Integumentary System 

Mammary gland 

Skin 

MMMMMMMMMMMMMMMMMMMMMMMMM 

+++++++++++++++++++++++++ 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Nervous System 

Brain +++++++++++++++++++++++++ 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Histiocytic sarcoma 

Nose 

+++++++A+++++++++++++++++ 

+++++++++++++++++++++++++ 

X X X X 

X X 

X 

+++++++A+++++++A+++++++++ 

Trachea +++++++A+++++++++++++++++ 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 189 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 250 ppm 

7777777777777777777777777 

Number of Days on Study 2222222222223333333333333 

9999999999990000000000011 

6666666666666666666666666 Total 

Carcass ID Number 1111222233440001122334423 

0689137859381361204014657 

Tissues/ 

Tumors 

Genital System 

Epididymis 

Hemangiosarcoma 

Histiocytic sarcoma 

Leiomyoma 

Preputial gland 

Prostate 

+++++++++++++++++++++++++ 

X 

X 

+++++++++++++++++++++++++ 

++++M++++++M+++++++++++++ 

50 

1 

1 

1 

49 

45 

Seminal vesicle +++++++++++++++++++++++++ 46 

Histiocytic sarcoma 

Testes +++++++++++++++++++++++++ 

1 

50 

Interstitial cell, adenoma X X 2 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 50 

Hemangiosarcoma 

Histiocytic sarcoma 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Histiocytic sarcoma 

Lymph node, mediastinal 

Histiocytic sarcoma 

Spleen 

Hemangiosarcoma 

Histiocytic sarcoma 

Thymus 

Histiocytic sarcoma 

X 

+ M + + M +++++++MM+++M+MMM+++ 

M M + + M ++++++++MMMM+M++M+MM 

++++M+++++M++++++++++++++ 

M +++M++++++M+++++MMMM++++ 

+++++++++++++++++++++++++ 

X 

M++++++M++++++++++M++M+M+ 

2 

1 

1 

35 

25 

43 

1 

37 

1 

49 

1 

1 

37 

1 

Integumentary System 

Mammary gland 

Skin 

M M M M M M M M M M M M M M M M M M M M M M M MM 

+++++++++++++++++++++++++ 50 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 50 

Nervous System 

Brain +++++++++++++++++++++++++ 50 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Histiocytic sarcoma 

Nose 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X X X 

X X X 

X 

+++++++++++++++++++++++++ 

49 

50 

8 

3 

3 

1 

48 

Trachea +++++++++++++++++++++++++ 49 
 

 

 

190 2-Butoxyethanol, NTP TR 484 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 250 ppm 

0333444566666666666666777 

Number of Days on Study 7167045901122366778899012 

3284224643439835004444219 

6666666666666666666666666 

Carcass ID Number 1431033513244040240102430 

7085236042952579298346147 

Special Senses System 

Harderian gland 

Adenoma 

Urinary System 

Kidney +++++++++++++++++++++++++ 

Histiocytic sarcoma X 

Urinary bladder +A++++A A A++++++A +++++++++ 

Histiocytic sarcoma X 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 

Histiocytic sarcoma X X 
 

 

2-Butoxyethanol, NTP TR 484 191 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 250 ppm 

7777777777777777777777777 

Number of Days on Study 2222222222223333333333333 

9999999999990000000000011 

6666666666666666666666666 Total 

Carcass ID Number 1111222233440001122334423 Tissues/ 

0689137859381361204014657 Tumors 

Special Senses System 

Harderian gland + + 2 

Adenoma X X 2 

Urinary System 

Kidney +++++++++++++++++++++++++ 50 

Histiocytic sarcoma 1 

Urinary bladder +++++++++++++++++++++++++ 45 

Histiocytic sarcoma 1 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 50 

Histiocytic sarcoma 2 
192 2-Butoxyethanol, NTP TR 484 

TABLE C3 

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Harderian Gland: Adenoma 

Overall ratea 

Adjusted rateb 3/50 (6%) 

6.7% 

3/50 (6%) 

6.4% 

2/50 (4%) 

5.0% 

2/50 (4%) 

5.1% 

Terminal ratec 3/39 (8%) 3/39 (8%) 2/27 (7%) 2/26 (8%) 

First incidence (days) 

Poly-3 testd 729 (T) 

P=0.440N 

729 (T) 

P=0.640N 

729 (T) 

P=0.555N 

729 (T) 

P=0.559N 

Liver: Hemangiosarcoma 

Overall rate 0/50 (0%) 1/50 (2%) 2/49 (4%) 4/49 (8%) 

Adjusted rate 0.0% 2.1% 5.0% 10.0% 

Terminal rate 0/39 (0%) 0/39 (0%) 1/27 (4%) 2/26 (8%) 

First incidence (days) —e 670 704 454 

Poly-3 test P=0.014 P=0.511 P=0.211 P=0.046 

Liver: Hepatocellular Adenoma 

Overall rate 22/50 (44%) 18/50 (36%) 18/49 (37%) 17/49 (35%) 

Adjusted rate 48.6% 37.2% 43.8% 40.3% 

Terminal rate 21/39 (54%) 13/39 (33%) 14/27 (52%) 10/26 (39%) 

First incidence (days) 642 549 582 368 

Poly-3 test P=0.345N P=0.180N P=0.407N P=0.283N 

Liver: Hepatocellular Carcinoma 

Overall rate 10/50 (20%) 11/50 (22%) 16/49 (33%) 21/49 (43%) 

Adjusted rate 20.8% 22.9% 35.9% 45.9% 

Terminal rate 3/39 (8%) 7/39 (18%) 5/27 (19%) 7/26 (27%) 

First incidence (days) 374 621 430 312 

Poly-3 test P=0.002 P=0.500 P=0.080 P=0.007 

Liver: Hepatocellular Adenoma or Carcinoma 

Overall rate 30/50 (60%) 24/50 (48%) 31/49 (63%) 30/49 (61%) 

Adjusted rate 61.9% 48.9% 67.5% 64.8% 

Terminal rate 22/39 (56%) 17/39 (44%) 16/27 (59%) 14/26 (54%) 

First incidence (days) 374 549 430 312 

Poly-3 test P=0.225 P=0.137N P=0.362 P=0.469 

Lung: Alveolar/bronchiolar Adenoma 

Overall rate 9/50 (18%) 7/50 (14%) 10/49 (20%) 8/50 (16%) 

Adjusted rate 19.9% 14.7% 24.2% 19.5% 

Terminal rate 8/39 (21%) 6/39 (15%) 5/27 (19%) 4/26 (15%) 

First incidence (days) 642 649 537 402 

Poly-3 test P=0.445 P=0.353N P=0.413 P=0.591N 

Lung: Alveolar/bronchiolar Carcinoma 

Overall rate 5/50 (10%) 2/50 (4%) 3/49 (6%) 3/50 (6%) 

Adjusted rate 11.1% 4.2% 7.6% 7.6% 

Terminal rate 5/39 (13%) 1/39 (3%) 3/27 (11%) 3/26 (12%) 

First incidence (days) 729 (T) 649 729 (T) 729 (T) 

Poly-3 test P=0.446N P=0.195N P=0.427N P=0.429N 
2-Butoxyethanol, NTP TR 484 193 

TABLE C3 

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Lung: Alveolar/bronchiolar Adenoma or Carcinoma 

Overall rate 14/50 (28%) 8/50 (16%) 12/49 (24%) 11/50 (22%) 

Adjusted rate 30.9% 16.9% 29.0% 26.9% 

Terminal rate 13/39 (33%) 7/39 (18%) 7/27 (26%) 7/26 (27%) 

First incidence (days) 642 649 537 402 

Poly-3 test P=0.524 P=0.087N P=0.515N P=0.430N 

All Organs: Hemangiosarcoma 

Overall rate 1/50 (2%) 1/50 (2%) 2/50 (4%) 5/50 (10%) 

Adjusted rate 2.2% 2.1% 5.0% 12.4% 

Terminal rate 1/39 (3%) 0/39 (0%) 1/27 (4%) 3/26 (12%) 

First incidence (days) 729 (T) 670 704 454 

Poly-3 test P=0.019 P=0.749N P=0.459 P=0.079 

All Organs: Hemangioma or Hemangiosarcoma 

Overall rate 1/50 (2%) 1/50 (2%) 4/50 (8%) 5/50 (10%) 

Adjusted rate 2.2% 2.1% 10.0% 12.4% 

Terminal rate 1/39 (3%) 0/39 (0%) 3/27 (11%) 3/26 (12%) 

First incidence (days) 729 (T) 670 704 454 

Poly-3 test P=0.020 P=0.749N P=0.145 P=0.079 

All Organs: Malignant Lymphoma 

Overall rate 1/50 (2%) 3/50 (6%) 0/50 (0%) 0/50 (0%) 

Adjusted rate 2.2% 6.3% 0.0% 0.0% 

Terminal rate 0/39 (0%) 1/39 (3%) 0/27 (0%) 0/26 (0%) 

First incidence (days) 621 611 — — 

Poly-3 test P=0.179N P=0.324 P=0.526N P=0.527N 

All Organs: Benign Neoplasms 

Overall rate 30/50 (60%) 28/50 (56%) 26/50 (52%) 28/50 (56%) 

Adjusted rate 65.9% 57.2% 60.2% 63.4% 

Terminal rate 28/39 (72%) 21/39 (54%) 17/27 (63%) 17/26 (65%) 

First incidence (days) 642 549 537 368 

Poly-3 test P=0.534N P=0.252N P=0.361N P=0.487N 

All Organs: Malignant Neoplasms 

Overall rate 19/50 (38%) 16/50 (32%) 19/50 (38%) 28/50 (56%) 

Adjusted rate 38.6% 32.4% 42.4% 59.4% 

Terminal rate 10/39 (26%) 8/39 (21%) 7/27 (26%) 12/26 (46%) 

First incidence (days) 374 549 430 312 

Poly-3 test P=0.008 P=0.332N P=0.435 P=0.031 
 

 

194 

c 

2-Butoxyethanol, NTP TR 484 

TABLE C3 

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

All Organs: Benign or Malignant Neoplasms 

Overall rate 42/50 (84%) 33/50 (66%) 36/50 (72%) 41/50 (82%) 

Adjusted rate 84.9% 66.4% 76.1% 85.3% 

Terminal rate 32/39 (82%) 24/39 (62%) 18/27 (67%) 21/26 (81%) 

First incidence (days) 374 549 430 312 

Poly-3 test P=0.278 P=0.025N P=0.192N P=0.596 

(T)Terminal sacrifice 

a Number of neoplasm-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for liver and lung; for 

other tissues, denominator is number of animals necropsied. b	 Poly-3 estimated neoplasm incidence after adjustment for intercurrent mortality 

Observed incidence at terminal kill 

d	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values 

corresponding to pairwise comparisons between the chamber controls and that exposed group. The Poly-3 test accounts for differential mortality in 

animals that do not reach terminal sacrifice. A negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group 
2-Butoxyethanol, NTP TR 484 195 

TABLE C4a 

Historical Incidence of Forestomach Squamous Cell Papilloma in Chamber Control Male B6C3F1 Micea 

Incidence in Controls 

Historical Incidence at Battelle Pacific Northwest Laboratories 

1,3-Butadiene 1/50 

Acetonitrile 0/50 

Chloroprene 1/50 

Cobalt sulfate heptahydrate 0/50 

Furfuryl alcohol 0/50 

Hexachlorocyclopentadiene 0/50 

Isobutene 1/50 

Isobutyraldehyde 1/50 

Molybdenum trioxide 0/50 

Nitromethane 0/50 

Ozone 0/50 

Tetrahydrofuran 0/50 

Overall Historical Incidence 

Total (%) 5/970 (0.5%) 

Mean ± standard deviation 0.5% ± 0.9% 

Range 0%-2% 

a Data as of 16 October 1997; no carcinomas observed 
196 2-Butoxyethanol, NTP TR 484 

TABLE C4b 

Historical Incidence of Liver Neoplasms in Chamber Control Male B6C3F1 Micea 

Incidence in Controls 

Hemangiosarcoma Hepatocellular Hepatocellular Hepatocellular 

Adenoma Carcinoma Adenoma or Carcinoma 

Historical Incidence at Battelle Pacific Northwest Laboratories 

1,3-Butadiene 0/50 13/50 11/50 21/50 

Acetonitrile 1/50 13/50 7/50 19/50 

Chloroprene 2/50 22/50 24/50 43/50 

Cobalt sulfate heptahydrate 2/50 22/50 23/50 38/50 

Furfuryl alcohol 0/50 13/50 15/50 28/50 

Hexachlorocyclopentadiene 0/50 19/50 7/50 24/50 

Isobutene 1/50 20/50 13/50 30/50 

Isobutyraldehyde 1/49 12/49 17/49 27/49 

Molybdenum trioxide 0/50 20/50 12/50 30/50 

Nitromethane 1/50 17/50 16/50 29/50 

Ozone 0/50 23/50 12/50 30/50 

Tetrahydrofuran 1/50 24/50 14/50 35/50 

Overall Historical Incidence 

Total (%) 14/968 (1.5%) 302/968 (31.2%) 247/968 (25.5%) 503/968 (52.0%) 

Mean ± standard deviation 1.5% ± 1.5% 31.3% ± 11.1% 25.7% ± 10.4% 52.2% ± 16.4% 

Range 0%-4% 7%-48% 11%-48% 20%-86% 

a Data as of 16 October 1997 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 197 

TABLE C5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 7 8 13 10 

Natural death 4 3 10 14 

Survivors 

Terminal sacrifice 39 39 27 26 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Gallbladder (42) (45) (40) (35) 

Degeneration, hyaline 1 (2%) 

Inflammation 1 (2%) 

Intestine small, duodenum (49) (48) (44) (41) 

Inflammation 1 (2%) 

Epithelium, hyperplasia 1 (2%) 

Peyer's patch, hyperplasia 1 (2%) 

Intestine small, jejunum (48) (47) (43) (38) 

Peyer's patch, hyperplasia 1 (2%) 1 (3%) 

Intestine small, ileum (48) (47) (43) (39) 

Inflammation 1 (2%) 

Peyer's patch, hyperplasia 1 (2%) 1 (2%) 

Liver (50) (50) (49) (49) 

Angiectasis 1 (2%) 

Basophilic focus 1 (2%) 3 (6%) 

Clear cell focus 4 (8%) 5 (10%) 3 (6%) 

Degeneration, fatty 1 (2%) 2 (4%) 4 (8%) 

Eosinophilic focus 14 (28%) 10 (20%) 18 (37%) 12 (24%) 

Hematopoietic cell proliferation 1 (2%) 1 (2%) 2 (4%) 2 (4%) 

Hepatodiaphragmatic nodule 1 (2%) 

Infarct 1 (2%) 

Inflammation 6 (12%) 3 (6%) 6 (12%) 6 (12%) 

Karyomegaly 2 (4%) 1 (2%) 

Necrosis 6 (12%) 2 (4%) 7 (14%) 9 (18%) 

Bile duct, cyst 1 (2%) 1 (2%) 

Kupffer cell, pigmentation, hemosiderin 8 (16%) 30 (61%) 

Oval cell, hyperplasia 1 (2%) 1 (2%) 2 (4%) 3 (6%) 

Mesentery (5) (3) (3) (1) 

Angiectasis 1 (33%) 

Fat, necrosis 5 (100%) 2 (67%) 2 (67%) 

Pancreas (50) (50) (47) (47) 

Atrophy 10 (20%) 4 (8%) 2 (4%) 4 (9%) 

Cytoplasmic alteration 1 (2%) 1 (2%) 

Degeneration, hyaline 1 (2%) 

Inflammation 1 (2%) 

Duct, cyst 1 (2%) 

Salivary glands (50) (50) (48) (50) 

Inflammation 1 (2%) 1 (2%) 1 (2%) 

a Number of animals examined microscopically at the site and the number of animals with lesion 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

198 2-Butoxyethanol, NTP TR 484 

TABLE C5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Alimentary System (continued) 

Stomach, forestomach (50) (50) (49) (48) 

Cyst 1 (2%) 

Hyperplasia 1 (2%) 

Mineralization 1 (2%) 

Ulcer 1 (2%) 2 (4%) 9 (18%) 3 (6%) 

Epithelium, hyperplasia 1 (2%) 7 (14%) 16 (33%) 21 (44%) 

Stomach, glandular (50) (50) (46) (47) 

Degeneration, hyaline 1 (2%) 

Inflammation, suppurative 1 (2%) 

Mineralization 1 (2%) 1 (2%) 1 (2%) 2 (4%) 

Ulcer 1 (2%) 2 (4%) 

Epithelium, hyperplasia 4 (9%) 1 (2%) 

Tooth (4) 

Malformation 4 (100%) 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Cardiomyopathy 47 (94%) 48 (96%) 47 (94%) 47 (94%) 

Artery, inflammation 1 (2%) 

Atrium, thrombosis 1 (2%) 1 (2%) 

Endocrine System 

Adrenal cortex (50) (50) (49) (50) 

Degeneration, cystic 1 (2%) 

Hematopoietic cell proliferation 1 (2%) 1 (2%) 

Hyperplasia 5 (10%) 9 (18%) 9 (18%) 9 (18%) 

Hypertrophy 38 (76%) 28 (56%) 27 (55%) 27 (54%) 

Capsule, hyperplasia 2 (4%) 3 (6%) 

Adrenal medulla (49) (50) (49) (49) 

Hyperplasia 1 (2%) 2 (4%) 3 (6%) 

Islets, pancreatic (50) (50) (47) (46) 

Hyperplasia 6 (12%) 3 (6%) 2 (4%) 

Inflammation 1 (2%) 

Parathyroid gland (26) (29) (26) (31) 

Cyst 1 (3%) 

Pituitary gland (49) (49) (46) (48) 

Hemorrhage 1 (2%) 

Pars distalis, cyst 1 (2%) 

Pars distalis, hyperplasia 5 (10%) 1 (2%) 2 (4%) 1 (2%) 

Thyroid gland (50) (48) (49) (50) 

Follicular cell, hyperplasia 5 (10%) 8 (17%) 6 (12%) 5 (10%) 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 199 

TABLE C5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Genital System 

Epididymis (50) (50) (49) (50) 

Granuloma sperm 1 (2%) 

Hyperplasia 1 (2%) 

Inflammation 4 (8%) 4 (8%) 1 (2%) 4 (8%) 

Inflammation, granulomatous 1 (2%) 

Penis (1) 

Inflammation 1 (100%) 

Preputial gland (49) (49) (49) (49) 

Ectasia 1 (2%) 1 (2%) 1 (2%) 

Inflammation 2 (4%) 7 (14%) 6 (12%) 8 (16%) 

Prostate (49) (49) (48) (45) 

Inflammation 2 (4%) 3 (6%) 4 (9%) 

Epithelium, hyperplasia 1 (2%) 1 (2%) 

Seminal vesicle (50) (49) (46) (46) 

Inflammation 1 (2%) 2 (4%) 3 (7%) 

Testes (50) (50) (49) (50) 

Atrophy 5 (10%) 4 (8%) 6 (12%) 4 (8%) 

Mineralization 3 (6%) 

Interstitial cell, hyperplasia 1 (2%) 

Hematopoietic System 

Bone marrow (50) (50) (49) (50) 

Fibrosis 1 (2%) 

Hyperplasia 1 (2%) 9 (18%) 5 (10%) 

Thrombosis 1 (2%) 

Lymph node (2) (1) (1) 

Renal, hyperplasia 1 (100%) 

Lymph node, bronchial (38) (35) (33) (35) 

Hyperplasia 2 (5%) 1 (3%) 

Lymph node, mandibular (24) (26) (32) (25) 

Hyperplasia 1 (4%) 1 (3%) 1 (4%) 

Lymph node, mesenteric (50) (50) (47) (43) 

Amyloid deposition 1 (2%) 

Angiectasis 2 (4%) 2 (4%) 1 (2%) 

Hematopoietic cell proliferation 1 (2%) 3 (6%) 1 (2%) 

Hyperplasia 7 (14%) 7 (14%) 4 (9%) 4 (9%) 

Spleen (50) (50) (48) (49) 

Hematopoietic cell proliferation 12 (24%) 11 (22%) 26 (54%) 42 (86%) 

Hyperplasia, lymphoid 5 (10%) 8 (16%) 3 (6%) 1 (2%) 

Pigmentation, hemosiderin 6 (12%) 45 (94%) 44 (90%) 

Thymus (43) (43) (36) (37) 

Atrophy 1 (2%) 1 (2%) 1 (3%) 1 (3%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

200 2-Butoxyethanol, NTP TR 484 

TABLE C5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Integumentary System 

Skin (50) (50) (49) (50) 

Granuloma 1 (2%) 

Inflammation 1 (2%) 

Pigmentation, melanin 1 (2%) 

Prepuce, hyperplasia 1 (2%) 2 (4%) 

Prepuce, inflammation, chronic active 2 (4%) 3 (6%) 13 (27%) 8 (16%) 

Prepuce, ulcer 3 (6%) 11 (22%) 8 (16%) 

Subcutaneous tissue, inflammation,

 granulomatous 1 (2%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Fibrous osteodystrophy 1 (2%) 4 (8%) 1 (2%) 

Nervous System 

Brain (50) (50) (49) (50) 

Inflammation, chronic 1 (2%) 

Necrosis 1 (2%) 

Vacuolization cytoplasmic 1 (2%) 1 (2%) 

Meninges, infiltration cellular, mononuclear

 cell 1 (2%) 1 (2%) 

Respiratory System 

Larynx (50) (50) (48) (49) 

Inflammation, suppurative 1 (2%) 

Squamous epithelium, hyperplasia 3 (6%) 

Lung (50) (50) (49) (50) 

Hematopoietic cell proliferation 1 (2%) 1 (2%) 

Hemorrhage 5 (10%) 5 (10%) 5 (10%) 3 (6%) 

Infiltration cellular, histiocyte 6 (12%) 1 (2%) 4 (8%) 4 (8%) 

Inflammation, chronic 1 (2%) 1 (2%) 

Pigmentation, hemosiderin 1 (2%) 

Alveolar epithelium, hyperplasia 6 (12%) 1 (2%) 2 (4%) 4 (8%) 

Artery, inflammation 1 (2%) 

Nose (50) (50) (48) (48) 

Inflammation, suppurative 2 (4%) 6 (12%) 2 (4%) 2 (4%) 

Polyp, inflammatory 1 (2%) 1 (2%) 

Glands, hyperplasia 1 (2%) 

Olfactory epithelium, atrophy 4 (8%) 4 (8%) 3 (6%) 

Olfactory epithelium, degeneration, hyaline 1 (2%) 2 (4%) 3 (6%) 1 (2%) 

Respiratory epithelium, degeneration, hyaline 4 (8%) 10 (20%) 5 (10%) 5 (10%) 

Respiratory epithelium, metaplasia, squamous 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 201 

TABLE C5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Special Senses System 

Ear (1) 

Inflammation, granulomatous 1 (100%) 

Eye (1) (1) 

Cornea, inflammation, chronic 1 (100%) 

Urinary System 

Kidney (50) (50) (47) (50) 

Glomerulosclerosis 4 (8%) 4 (8%) 11 (23%) 9 (18%) 

Hydronephrosis 1 (2%) 6 (13%) 5 (10%) 

Inflammation 1 (2%) 

Inflammation, chronic active 1 (2%) 2 (4%) 4 (8%) 

Metaplasia, osseous 2 (4%) 

Mineralization 1 (2%) 2 (4%) 

Nephropathy 48 (96%) 45 (90%) 40 (85%) 37 (74%) 

Capsule, inflammation 1 (2%) 

Cortex, cyst 4 (8%) 1 (2%) 6 (12%) 

Pelvis, inflammation, chronic active 1 (2%) 1 (2%) 

Renal tubule, hyperplasia 2 (4%) 2 (4%) 1 (2%) 

Renal tubule, mineralization 4 (8%) 2 (4%) 3 (6%) 2 (4%) 

Renal tubule, necrosis 1 (2%) 1 (2%) 

Renal tubule, pigmentation 3 (6%) 

Urinary bladder (50) (50) (46) (45) 

Inflammation 2 (4%) 5 (11%) 4 (9%) 

Transitional epithelium, hyperplasia 2 (4%) 1 (2%) 1 (2%) 

Transitional epithelium, ulcer 1 (2%) 4 (9%) 2 (4%) 
202 2-Butoxyethanol, NTP TR 484 
203 

APPENDIX D 

SUMMARY OF LESIONS IN FEMALE MICE 

IN THE 2-YEAR INHALATION STUDY 

OF 2-BUTOXYETHANOL 

TABLE D1 

TABLE D2 

TABLE D3 

TABLE D4a 

TABLE D4b 

TABLE D5 

Summary of the Incidence of Neoplasms in Female Mice 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

Individual Animal Tumor Pathology of Female Mice 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

Statistical Analysis of Primary Neoplasms in Female Mice 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

Historical Incidence of Forestomach Neoplasms 

in Chamber Control Female B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 

Historical Incidence of Liver Neoplasms in Chamber Control Female B6C3F1 Mice . . . 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 

204 

208 

228 

232 

233 

234 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

204 2-Butoxyethanol, NTP TR 484 

TABLE D1 

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Accidental death 1 

Moribund 14 16 12 10 

Natural deaths 6 3 5 4 

Survivors 

Terminal sacrifice 29 31 33 36 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Gallbladder (43) (43) (46) (46) 

Intestine small, duodenum (45) (49) (47) (47) 

Polyp adenomatous 1 (2%) 

Intestine small, jejunum (46) (49) (47) (48) 

Carcinoma 1 (2%) 

Intestine small, ileum (46) (50) (47) (48) 

Carcinoma 1 (2%) 

Liver (50) (50) (49) (50) 

Hemangiosarcoma 1 (2%) 

Hepatocellular carcinoma 7 (14%) 10 (20%) 11 (22%) 9 (18%) 

Hepatocellular carcinoma, multiple 3 (6%) 2 (4%) 2 (4%) 1 (2%) 

Hepatocellular adenoma 9 (18%) 5 (10%) 5 (10%) 8 (16%) 

Hepatocellular adenoma, multiple 7 (14%) 3 (6%) 2 (4%) 

Hepatocholangiocarcinoma 2 (4%) 1 (2%) 

Histiocytic sarcoma 1 (2%) 1 (2%) 1 (2%) 

Osteosarcoma, metastatic, bone 1 (2%) 

Mesentery (10) (4) (5) (9) 

Hemangioma 1 (10%) 

Hemangiosarcoma 1 (25%) 

Osteosarcoma 1 (20%) 

Sarcoma, metastatic, skin 2 (20%) 

Pancreas (49) (50) (49) (50) 

Salivary glands (48) (50) (49) (50) 

Parotid gland, hemangioma 1 (2%) 

Stomach, forestomach (50) (50) (49) (50) 

Hemangioma 1 (2%) 

Squamous cell carcinoma 1 (2%) 

Squamous cell papilloma 1 (2%) 2 (4%) 5 (10%) 

Stomach, glandular (48) (50) (49) (49) 

Tongue (1) (1) 

Squamous cell carcinoma 1 (100%) 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Hepatocholangiocarcinoma, metastatic, liver 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 205 

TABLE D1 

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Endocrine System 

Adrenal cortex (50) (50) (50) (50) 

Adrenal medulla (50) (50) (49) (50) 

Pheochromocytoma benign 3 (6%) 1 (2%) 1 (2%) 

Bilateral, pheochromocytoma benign 1 (2%) 

Islets, pancreatic (50) (50) (49) (50) 

Adenoma 2 (4%) 1 (2%) 

Pituitary gland (50) (49) (48) (49) 

Pars distalis, adenoma 5 (10%) 8 (16%) 8 (17%) 4 (8%) 

Pars distalis, carcinoma 1 (2%) 

Pars intermedia, adenoma 1 (2%) 

Thyroid gland (50) (50) (50) (49) 

Follicular cell, adenoma 1 (2%) 2 (4%) 5 (10%) 1 (2%) 

Follicular cell, carcinoma 1 (2%) 

General Body System 

None 

Genital System 

Ovary (50) (49) (49) (49) 

Cystadenoma 2 (4%) 1 (2%) 3 (6%) 

Granulosa cell tumor benign 1 (2%) 

Hemangioma 1 (2%) 

Teratoma benign 2 (4%) 

Teratoma malignant 1 (2%) 

Uterus (50) (50) (49) (50) 

Adenoma 1 (2%) 

Hemangiosarcoma 2 (4%) 

Histiocytic sarcoma 1 (2%) 1 (2%) 

Polyp stromal 1 (2%) 2 (4%) 4 (8%) 2 (4%) 

Sarcoma stromal 1 (2%) 

Hematopoietic System 

Bone marrow (50) (50) (50) (50) 

Hemangiosarcoma 1 (2%) 1 (2%) 1 (2%) 

Lymph node (4) (7) (6) (1) 

Sarcoma, metastatic, skin 1 (25%) 

Renal, sarcoma, metastatic, skin 1 (25%) 

Renal, teratoma malignant, metastatic, ovary 1 (17%) 

Lymph node, bronchial (41) (39) (41) (37) 

Hepatocholangiocarcinoma, metastatic, liver 1 (3%) 

Lymph node, mandibular (37) (38) (40) (38) 

Histiocytic sarcoma 1 (3%) 

Lymph node, mesenteric (47) (48) (49) (50) 

Hepatocholangiocarcinoma, metastatic, liver 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Sarcoma, metastatic, skin 1 (2%) 

Lymph node, mediastinal (31) (40) (38) (34) 

Hepatocholangiocarcinoma, metastatic, liver 1 (3%) 

Spleen (50) (50) (49) (50) 

Hemangiosarcoma 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Thymus (46) (41) (46) (48) 

Hepatocholangiocarcinoma, metastatic, liver 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

206 2-Butoxyethanol, NTP TR 484 

TABLE D1 

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Integumentary System 

Mammary gland (50) (50) (50) (50) 

Carcinoma 1 (2%) 3 (6%) 

Skin (50) (50) (50) (50) 

Sebaceous gland, adenoma 1 (2%) 

Subcutaneous tissue, hemangioma 1 (2%) 

Subcutaneous tissue, hemangiosarcoma 1 (2%) 1 (2%) 

Subcutaneous tissue, sarcoma 2 (4%) 1 (2%) 

Subcutaneous tissue, sarcoma, multiple 1 (2%) 1 (2%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Hemangiosarcoma 1 (2%) 

Osteosarcoma 1 (2%) 

Skeletal muscle (1) 

Hemangiosarcoma 1 (100%) 

Nervous System 

Brain (50) (50) (50) (50) 

Carcinoma, metastatic, pituitary gland 1 (2%) 

Respiratory System 

Lung (50) (50) (50) (50) 

Alveolar/bronchiolar adenoma 5 (10%) 2 (4%) 4 (8%) 1 (2%) 

Alveolar/bronchiolar adenoma, multiple 2 (4%) 

Alveolar/bronchiolar carcinoma 2 (4%) 1 (2%) 

Carcinoma, metastatic, harderian gland 1 (2%) 

Hepatocellular carcinoma, metastatic, liver 4 (8%) 3 (6%) 6 (12%) 1 (2%) 

Hepatocholangiocarcinoma, metastatic, liver 1 (2%) 1 (2%) 

Histiocytic sarcoma 1 (2%) 1 (2%) 

Osteosarcoma, metastatic, bone 1 (2%) 

Sarcoma, metastatic, skin 1 (2%) 1 (2%) 

Mediastinum, hepatocholangiocarcinoma,

 metastatic, liver 1 (2%) 

Mediastinum, osteosarcoma, metastatic, bone 1 (2%) 

Mediastinum, sarcoma, metastatic, skin 1 (2%) 

Nose (50) (50) (49) (50) 

Special Senses System 

Harderian gland (6) (4) (3) (4) 

Adenoma 4 (67%) 3 (75%) 2 (67%) 3 (75%) 

Carcinoma 1 (17%) 1 (25%) 1 (33%) 2 (50%) 
 

 

207 

c 

2-Butoxyethanol, NTP TR 484 

TABLE D1 

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Urinary System 

Kidney 

Hepatocholangiocarcinoma, metastatic, liver

Histiocytic sarcoma

Urinary bladder 

(50) 

 1 (2%) 

(47) 

(50) 

 1 (2%) 

(49) 

(49) 

(48) 

(50) 

(50) 

Systemic Lesions 

Multiple organsb 

Histiocytic sarcoma

Lymphoma malignant

(50) 

 1 (2%)

 7 (14%)

(50) 

 1 (2%)

 9 (18%)

(50) 

 2 (4%) 

 10 (20%)

(50) 

 7 (14%) 

Neoplasm Summary 

Total animals with primary neoplasmsc 

Total primary neoplasms 

Total animals with benign neoplasms

Total benign neoplasms 

Total animals with malignant neoplasms 

Total malignant neoplasms 

Total animals with metastatic neoplasms 

Total metastatic neoplasms 

43 

76 

31 

44 

27 

32 

9 

18 

38 

63 

23 

30 

26 

33 

4 

4 

38 

76 

27 

42 

28 

34 

7 

7 

35 

52 

18 

27 

22 

25 

3 

8 

a Number of animals examined microscopically at the site and the number of animals with neoplasm b Number of animals with any tissue examined microscopically 

Primary neoplasms: all neoplasms except metastatic neoplasms 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

208 2-Butoxyethanol, NTP TR 484 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: Chamber Control 

3555555566666677777777777 

Number of Days on Study	 7266778901244801112233333 

6115276061159903551501111 

1111111111111111111111111 

Carcass ID Number 3112314043013202442230011 

8191262395886678675014602 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Gallbladder M++++A+A A+++++++++A+A++++ 

Intestine large, colon ++++++++++++++++++++A++++ 

Intestine large, rectum +++++A+A+++++++++++++++++ 

Intestine large, cecum ++++++++++++++++++++A++++ 

Intestine small, duodenum +++++A+A A+++++++++A+A++++ 

Intestine small, jejunum +++++A+A A+++++++++++A++++ 

Carcinoma X 

Intestine small, ileum +++++A+A A+++++++++++A++++ 

Liver +++++++++++++++++++++++++ 

Hepatocellular carcinoma X X X X 

Hepatocellular carcinoma, multiple X X X 

Hepatocellular adenoma X X X X 

Hepatocellular adenoma, multiple X X X X X 

Hepatocholangiocarcinoma X 

Histiocytic sarcoma 

Osteosarcoma, metastatic, bone X 

Mesentery + + + ++ 

Hemangioma 

Sarcoma, metastatic, skin X X 

Pancreas +++++++M+++++++++++++++++ 

Salivary glands +M+++++++++++M+++++++++++ 

Stomach, forestomach +++++++++++++++++++++++++ 

Hemangioma X 

Stomach, glandular +++++A+A+++++++++++++++++ 

Tongue + 

Squamous cell carcinoma 	 X 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adrenal medulla +++++++++++++++++++++++++ 

Pheochromocytoma benign X 

Islets, pancreatic +++++++++++++++++++++++++ 

Adenoma 

Parathyroid gland +MM++++MMM+M+M++MM+M++M++ 

Pituitary gland +++++++++++++++++++++++++ 

Pars distalis, adenoma X X X 

Pars distalis, carcinoma X 

Thyroid gland +++++++++++++++++++++++++ 

Follicular cell, adenoma X 

General Body System 

None 

+: Tissue examined microscopically M: Missing tissue	 X: Lesion present 

A: Autolysis precludes examination	 I: Insufficient tissue Blank: Not examined 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 209 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: Chamber Control 

7777777777777777777777777 

Number of Days on Study	 3333333333333333333333333 

1111122222222222222233333 

1111111111111111111111111 Total 

Carcass ID Number 1123500001222334444412334 Tissues/ 

4724012595379031345834790 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Gallbladder ++++++++++++++++++++M++++ 43 

Intestine large, colon +++++++++++++++++++++++++ 49 

Intestine large, rectum ++++++++++++++++M++++++++ 47 

Intestine large, cecum +++++++++++++++++++++++++ 49 

Intestine small, duodenum +++++++++++++++++++++++++ 45 

Intestine small, jejunum +++++++++++++++++++++++++ 46 

Carcinoma 1 

Intestine small, ileum +++++++++++++++++++++++++ 46 

Liver +++++++++++++++++++++++++ 50 

Hepatocellular carcinoma X X X 7 

Hepatocellular carcinoma, multiple 3 

Hepatocellular adenoma X X X X X 9 

Hepatocellular adenoma, multiple X X 7 

Hepatocholangiocarcinoma X 2 

Histiocytic sarcoma X 1 

Osteosarcoma, metastatic, bone 1 

Mesentery + + + + + 10 

Hemangioma X 1 

Sarcoma, metastatic, skin 2 

Pancreas +++++++++++++++++++++++++ 49 

Salivary glands +++++++++++++++++++++++++ 48 

Stomach, forestomach +++++++++++++++++++++++++ 50 

Hemangioma 1 

Stomach, glandular +++++++++++++++++++++++++ 48 

Tongue 1 

Squamous cell carcinoma 1 

Cardiovascular System 

Heart +++++++++++++++++++++++++ 50 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 50 

Adrenal medulla +++++++++++++++++++++++++ 50 

Pheochromocytoma benign X X 3 

Islets, pancreatic +++++++++++++++++++++++++ 50 

Adenoma X X 2 

Parathyroid gland MMMM+++MM+++M++++M+++++++ 31 

Pituitary gland +++++++++++++++++++++++++ 50 

Pars distalis, adenoma X X 5 

Pars distalis, carcinoma 1 

Thyroid gland +++++++++++++++++++++++++ 50 

Follicular cell, adenoma 1 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

210 2-Butoxyethanol, NTP TR 484 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: Chamber Control 

3555555566666677777777777 

Number of Days on Study 7266778901244801112233333 

6115276061159903551501111 

1111111111111111111111111 

Carcass ID Number 3112314043013202442230011 

8191262395886678675014602 

Genital System 

Clitoral gland M++++++++++M+M+MM+++++M++ 

Ovary +++++++++++++++++++++++++ 

Cystadenoma X X 

Hemangioma X 

Uterus +++++++++++++++++++++++++ 

Histiocytic sarcoma 

Polyp stromal 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 

Hemangiosarcoma X 

Lymph node + + 

Sarcoma, metastatic, skin X 

Renal, sarcoma, metastatic, skin X 

Lymph node, bronchial +++M++M++++++++++M++MMM++ 

Lymph node, mandibular + M + + M + + M ++++MM+M+M+++MM++ 

Lymph node, mesenteric +++M++++++++++++++++M++++ 

Hepatocholangiocarcinoma, metastatic, liver X 

Sarcoma, metastatic, skin X 

Lymph node, mediastinal M + M M + + M + M + M ++++M++++M+M++ 

Spleen +++++++++++++++++++++++++ 

Hemangiosarcoma 

Histiocytic sarcoma 

Thymus +M+++++++M+++++++++++++++ 

Integumentary System 

Mammary gland +++++++++++++++++++++++++ 

Skin +++++++++++++++++++++++++ 

Subcutaneous tissue, sarcoma X X 

Subcutaneous tissue, sarcoma, multiple X 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Hemangiosarcoma X 

Osteosarcoma X 

Skeletal muscle + 

Hemangiosarcoma X 

Nervous System 

Brain +++++++++++++++++++++++++ 

Carcinoma, metastatic, pituitary gland X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 211 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: Chamber Control 

7777777777777777777777777 

Number of Days on Study	 3333333333333333333333333 

1111122222222222222233333 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 Total 

Carcass ID Number 1 1 2 3 5 0 0 0 0 1 2 2 2 3 3 4 4 4 4 4 1 2 3 3 4 Tissues/ 

4 7 2 4 0 1 2 5 9 5 3 7 9 0 3 1 3 4 5 8 3 4 7 9 0 Tumors 

Genital System 

Clitoral gland + + + + + + M M + + + + + + + M + + + + + + + + + 41 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Cystadenoma 2 

Hemangioma 1 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma X 1 

Polyp stromal X 1 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma 1 

Lymph node + + 4 

Sarcoma, metastatic, skin 1 

Renal, sarcoma, metastatic, skin 1 

Lymph node, bronchial + + + + + + + + + + + + M + M + + + + + + + + + M 41 

Lymph node, mandibular + + + + + + + + + + + M + + M + M + + M + + + + + 37 

Lymph node, mesenteric + + + + + + + + + + + + M + + + + + + + + + + + + 47 

Hepatocholangiocarcinoma, metastatic liver 1 

Sarcoma, metastatic, skin 1 

Lymph node, mediastinal + M M + + M + + M +++MMM++++M+M++M 31 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X 1 

Histiocytic sarcoma X 1 

Thymus + + + + + + + + + + + + + + + + + + + + M + + M + 46 

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Subcutaneous tissue, sarcoma 2 

Subcutaneous tissue, sarcoma multiple 1 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma 1 

Osteosarcoma 1 

Skeletal muscle 1 

Hemangiosarcoma 1 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma, metastatic, pituitary gland 1 
 

 

 

 

 

 

 

 

 

 

 

212 2-Butoxyethanol, NTP TR 484 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: Chamber Control 

3555555566666677777777777 

Number of Days on Study	 7266778901244801112233333 

6115276061159903551501111 

1111111111111111111111111 

Carcass ID Number 3112314043013202442230011 

8191262395886678675014602 

Respiratory System 

Larynx 	 +++++++++++++++++++++++++ 

Lung +++++++++++++++++++++++++ 

Alveolar/bronchiolar adenoma X 

Alveolar/bronchiolar adenoma, multiple X 

Hepatocellular carcinoma, metastatic, liver X X X 

Hepatocholangiocarcinoma, metastatic, liver X 

Osteosarcoma, metastatic, bone X 

Sarcoma, metastatic, skin X 

Mediastinum, osteosarcoma, metastatic, bone X 

Mediastinum, sarcoma, metastatic, skin X 

Nose 	 +++++++++++++++++++++++++ 

Trachea 	 +++++++++++++++++++++++++ 

Special Senses System 

Eye 

Harderian gland 

Adenoma 

Carcinoma 

+ 

Urinary System 

Kidney 

Hepatocholangiocarcinoma, metastatic, liver 

Urinary bladder 

+++++++++++++++++++++++++ 

X 

+++++A+A++++++++++++A++++ 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant 

+++++++++++++++++++++++++ 

X X X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 213 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: Chamber Control 

7777777777777777777777777 

Number of Days on Study	 3333333333333333333333333 

1111122222222222222233333 

1111111111111111111111111 Total 

Carcass ID Number 1123500001222334444412334 

4724012595379031345834790 

Tissues/ 

Tumors 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Hepatocellular carcinoma, metastatic, liver 

Hepatocholangiocarcinoma, metastatic, liver 

Osteosarcoma, metastatic, bone 

Sarcoma, metastatic, skin 

Mediastinum, osteosarcoma, metastatic, bone 

Mediastinum, sarcoma, metastatic, skin 

Nose 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X X X 

X 

X 

+++++++++++++++++++++++++ 

50 

50 

5 

2 

4 

1 

1 

1 

1 

1 

50 

Trachea +++++++++++++++++++++++++ 50 

Special Senses System 

Eye 

Harderian gland 

Adenoma 

+ 

X 

+ 

++ 

X 

+ 

X 

+ 

X 

1 

6 

4 

Carcinoma X 1 

Urinary System 

Kidney 

Hepatocholangiocarcinoma, metastatic, liver 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

1 

47 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant 

+++++++++++++++++++++++++ 

X 

X X X X 

50 

1 

7 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

214 2-Butoxyethanol, NTP TR 484 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

2334455566666667777777777 

Number of Days on Study 5675734900477791112333333 

5926244246702642253111111 

3333333333333333333333333 

Carcass ID Number 4133204101201340212001233 

6202234863887972456490915 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Gallbladder + + + + + + + + + M M + + + + + + + + + + + + I + 

Intestine large, colon +++++++++++++++++++++++++ 

Intestine large, rectum +++M+++++++++++++++++++++ 

Intestine large, cecum +++++++++++++++++++++++++ 

Intestine small, duodenum ++++++++++++++++A++++++++ 

Intestine small, jejunum ++++++++++++++++A++++++++ 

Intestine small, ileum +++++++++++++++++++++++++ 

Carcinoma X 

Liver +++++++++++++++++++++++++ 

Hemangiosarcoma 

Hepatocellular carcinoma X X X X X X X X 

Hepatocellular carcinoma, multiple X 

Hepatocellular adenoma X X 

Hepatocellular adenoma, multiple X X 

Histiocytic sarcoma X 

Mesentery + + + 

Hemangiosarcoma X 

Pancreas +++++++++++++++++++++++++ 

Salivary glands +++++++++++++++++++++++++ 

Stomach, forestomach +++++++++++++++++++++++++ 

Squamous cell papilloma X 

Stomach, glandular +++++++++++++++++++++++++ 

Tooth + 

Cardiovascular System 

Blood vessel 

Heart 

+ 

+++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adrenal medulla +++++++++++++++++++++++++ 

Pheochromocytoma benign X 

Islets, pancreatic +++++++++++++++++++++++++ 

Adenoma 

Parathyroid gland +M+M+M+M++++++M+M++MM++M+ 

Pituitary gland ++++++++++++++++++++++++ I 

Pars distalis, adenoma X 

Thyroid gland +++++++++++++++++++++++++ 

Follicular cell, adenoma X X 

Follicular cell, carcinoma 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 215 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

7777777777777777777777777 

Number of Days on Study 3333333333333333333333333 

1112222222222222222333333 

3333333333333333333333333 Total 

Carcass ID Number 4440011122333344445012223 Tissues/ 

0395714913347812580160576 Tumors 

Alimentary System 

Esophagus +++++++++++M+++++++++++++ 49 

Gallbladder ++++MM++++++++++++MM+++++ 43 

Intestine large, colon +++++++++++++++++++++++++ 50 

Intestine large, rectum +++++++++++++++++++++++++ 49 

Intestine large, cecum +++++++++++++++++++++++++ 50 

Intestine small, duodenum +++++++++++++++++++++++++ 49 

Intestine small, jejunum +++++++++++++++++++++++++ 49 

Intestine small, ileum +++++++++++++++++++++++++ 50 

Carcinoma 1 

Liver +++++++++++++++++++++++++ 50 

Hemangiosarcoma X 1 

Hepatocellular carcinoma X X 10 

Hepatocellular carcinoma, multiple X 2 

Hepatocellular adenoma X X X 5 

Hepatocellular adenoma, multiple X 3 

Histiocytic sarcoma 1 

Mesentery + 4 

Hemangiosarcoma 1 

Pancreas +++++++++++++++++++++++++ 50 

Salivary glands +++++++++++++++++++++++++ 50 

Stomach, forestomach +++++++++++++++++++++++++ 50 

Squamous cell papilloma 1 

Stomach, glandular +++++++++++++++++++++++++ 50 

Tooth 1 

Cardiovascular System 

Blood vessel 

Heart +++++++++++++++++++++++++ 

1 

50 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 50 

Adrenal medulla +++++++++++++++++++++++++ 50 

Pheochromocytoma benign 1 

Islets, pancreatic +++++++++++++++++++++++++ 50 

Adenoma X 1 

Parathyroid gland +M++M+M++M+M++M++++M++++M 33 

Pituitary gland +++++++++++++++++++++++++ 49 

Pars distalis, adenoma X X X X X X X 8 

Thyroid gland +++++++++++++++++++++++++ 50 

Follicular cell, adenoma 2 

Follicular cell, carcinoma X 1 

General Body System 

None 
 

 

 

 

 

 

 

 

 

216 2-Butoxyethanol, NTP TR 484 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

2334455566666667777777777 

Number of Days on Study 5675734900477791112333333 

5926244246702642253111111 

3333333333333333333333333 

Carcass ID Number 4133204101201340212001233 

6202234863887972456490915 

Genital System 

Clitoral gland 

Ovary 

Cystadenoma 

Uterus 

M++M+++M++++M+MM+++++++++ 

+++++++M+++++++++++++++++ 

+++++++++++++++++++++++++ 

Adenoma 

Histiocytic sarcoma 

Polyp stromal 

X 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Histiocytic sarcoma 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+ + + + + 

MMMM++++++++++M+M++++++++ 

+++++M++++M+M++++++M+M+MM 

X 

+++M++++++++++++++++++++M 

+ M + M +++++++M+++++M+++++MM 

+++++++++++++++++++++++++ 

M++++M+M+M++M+++M++++++++ 

Integumentary System 

Mammary gland 

Carcinoma 

+++++++++++++++++++++++++ 

X 

Skin +++++++++++++++++++++++++ 

Subcutaneous tissue, hemangiosarcoma 

Subcutaneous tissue, sarcoma 

Subcutaneous tissue, sarcoma, multiple X 

X 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Nervous System 

Brain +++++++++++++++++++++++++ 

Spinal cord + 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Histiocytic sarcoma 

Sarcoma, metastatic, skin 

Nose 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

X X X 

X 

X 

+++++++++++++++++++++++++ 

Trachea +++++++++++++++++++++++++ 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 217 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

7777777777777777777777777 

Number of Days on Study 3333333333333333333333333 

1112222222222222222333333 

3333333333333333333333333 Total 

Carcass ID Number 4440011122333344445012223 

0395714913347812580160576 

Tissues/ 

Tumors 

Genital System 

Clitoral gland 

Ovary 

Cystadenoma 

Uterus 

+++++++M++++++++++++++++M 

+++++++++++++++++++++++++ 

X 

+++++++++++++++++++++++++ 

42 

49 

1 

50 

Adenoma X 1 

Histiocytic sarcoma 

Polyp stromal X X 

1 

2 

Hematopoietic System 

Bone marrow +++++++++++++++++++++++++ 50 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Histiocytic sarcoma 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+ + 

+ M +++++M+++M+++++M++++M++ 

++++++++M+++++++++MM+M+M+ 

+++++++++++++++++++++++++ 

+++M++++++++++++++MM++M++ 

+++++++++++++++++++++++++ 

+++M++M++++++M+++++++++++ 

7 

39 

38 

1 

48 

40 

50 

41 

Integumentary System 

Mammary gland 

Carcinoma 

+++++++++++++++++++++++++ 50 

1 

Skin +++++++++++++++++++++++++ 50 

Subcutaneous tissue, hemangiosarcoma 

Subcutaneous tissue, sarcoma 

Subcutaneous tissue, sarcoma, multiple 

X 1 

1 

1 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 50 

Nervous System 

Brain +++++++++++++++++++++++++ 50 

Spinal cord 1 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Histiocytic sarcoma 

Sarcoma, metastatic, skin 

Nose 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X 

X 

+++++++++++++++++++++++++ 

50 

50 

2 

2 

3 

1 

1 

50 

Trachea +++++++++++++++++++++++++ 50 
 

 

 

 

218 2-Butoxyethanol, NTP TR 484 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

Number of Days on Study 

2334455566666667777777777 

5675734900477791112333333 

5926244246702642253111111 

Carcass ID Number 

3333333333333333333333333 

4133204101201340212001233 

6202234863887972456490915 

Special Senses System 

Harderian gland + 

Adenoma X 

Carcinoma 

Urinary System 

Kidney 

Histiocytic sarcoma 

Urinary bladder 

+++++++++++++++++++++++++ 

X 

++++++++++++++M++++++++++ 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant 

+++++++++++++++++++++++++ 

X 

X X X X X X 
 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 219 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 62.5 ppm 

7777777777777777777777777 

Number of Days on Study 3333333333333333333333333 

1112222222222222222333333 

3333333333333333333333333 Total 

Carcass ID Number 4440011122333344445012223 Tissues/ 

0395714913347812580160576 Tumors 

Special Senses System 

Harderian gland + + + 4 

Adenoma X X 3 

Carcinoma X 1 

Urinary System 

Kidney +++++++++++++++++++++++++ 50 

Histiocytic sarcoma 1 

Urinary bladder +++++++++++++++++++++++++ 49 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 50 

Histiocytic sarcoma 1 

Lymphoma malignant X X X 9 
 

 

 

 

 

 

 

 

 

 

 

 

 

220 2-Butoxyethanol, NTP TR 484 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

0134455555666777777777777 

Number of Days on Study 7990534779269122233333333 

2678674021768533311111111 

5555555555555555555555555 

Carcass ID Number 4022120211140113300111233 

9359842749626020479357615 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 

Gallbladder +A++A A+++++A+++++++++++++ 

Intestine large, colon +A+++++++++A+++++++++++++ 

Intestine large, rectum +A+++++++++A+++++++++++++ 

Intestine large, cecum + A + + + A +++++A+++++++++++++ 

Intestine small, duodenum + A + + A ++++++A+++++++++++++ 

Polyp adenomatous 

Intestine small, jejunum + A + + A ++++++A+++++++++++++ 

Intestine small, ileum +A++A++++++A+++++++++++++ 

Liver +A+++++++++++++++++++++++ 

Hepatocellular carcinoma X X X X X X 

Hepatocellular carcinoma, multiple X 

Hepatocellular adenoma X X 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma X 

Mesentery + + 

Osteosarcoma X 

Pancreas +A+++++++++++++++++++++++ 

Salivary glands ++++++++M++++++++++++++++ 

Parotid gland, hemangioma 

Stomach, forestomach +A+++++++++++++++++++++++ 

Squamous cell papilloma 

Stomach, glandular + A +++++++++++++++++++++++ 

Cardiovascular System 

Blood vessel 

Heart 

A 

+++++++++++++++++++++++++ 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adrenal medulla +++++++++++A+++++++++++++ 

Bilateral, pheochromocytoma benign 

Islets, pancreatic +A+++++++++++++++++++++++ 

Parathyroid gland ++++M++MM+M++++++++M+++++ 

Pituitary gland +AM++++++++++++++++++++++ 

Pars distalis, adenoma X X X 

Pars intermedia, adenoma X 

Thyroid gland +++++++++++++++++++++++++ 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Clitoral gland M+MM++M++M+++M++++++++++M 

Ovary +++++++++++++++++++++++++ 

Cystadenoma X X 

Teratoma benign X 

Teratoma malignant X 

Uterus +A+++++++++++++++++++++++ 

Hemangiosarcoma X 

Polyp stromal X X 

Sarcoma stromal X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 221 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

7777777777777777777777777 

Number of Days on Study 3333333333333333333333333 

1111122222222222223333333 

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 Total 

Carcass ID Number 3 3 4 4 4 0 0 1 2 2 2 3 3 3 4 4 4 5 0 0 2 2 3 4 4 Tissues/ 

6 7 1 6 7 1 5 1 0 1 2 2 3 9 0 3 5 0 4 8 3 8 8 4 8 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Gallbladder + + + + + + + + + + + + + + + + + + + + + + + + + 46 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Polyp adenomatous X 1 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Hepatocellular carcinoma X X X X X 11 

Hepatocellular carcinoma, multiple X 2 

Hepatocellular adenoma X X X 5 

Hepatocellular adenoma, multiple X X 2 

Histiocytic sarcoma 1 

Mesentery + + + 5 

Osteosarcoma 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Parotid gland, hemangioma X 1 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Squamous cell papilloma X X 2 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Cardiovascular System 

Blood vessel 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Bilateral, pheochromocytoma benign X 1 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Parathyroid gland + + + + + + + M + M + + M + + + + + + + + + + + + 42 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Pars distalis, adenoma X X X X X 8 

Pars intermedia, adenoma 1 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Follicular cell, adenoma X X X X X 5 

General Body System 

None 

Genital System 

Clitoral gland + + + + + + + M + + + M + + M + + + + M + + M + + 38 

Ovary + + + + + M + + + + + + + + + + + + + + + + + + + 49 

Cystadenoma X 3 

Teratoma benign X 2 

Teratoma malignant 1 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Hemangiosarcoma X 2 

Polyp stromal X X 4 

Sarcoma stromal 1 
 

 

 

 

 

 

 

 

 

 

222 2-Butoxyethanol, NTP TR 484 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

0134455555666777777777777 

Number of Days on Study 7990534779269122233333333 

2678674021768533311111111 

5555555555555555555555555 

Carcass ID Number 4022120211140113300111233 

9359842749626020479357615 

Hematopoietic System 

Bone marrow 

Hemangiosarcoma 

Lymph node 

Renal, teratoma malignant, metastatic, ovary 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Histiocytic sarcoma 

Lymph node, mediastinal 

Spleen 

Thymus 

+++++++++++++++++++++++++ 

+ + + + + 

X 

M A + M ++++++M+M++++++M++++M 

++++M+++M+++M++MMMMM+++++ 

+ A +++++++++++++++++++++++ 

+ + M M +++++M++M+M++M++++++M 

+A+++++++++++++++++++++++ 

M+++++M++M+++++++++++++++ 

Integumentary System 

Mammary gland 

Skin 

Subcutaneous tissue, hemangioma 

Subcutaneous tissue, hemangiosarcoma 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Nervous System 

Brain +++++++++++++++++++++++++ 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Histiocytic sarcoma 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X 

X X X X 

X 

+++++++++++A+++++++++++++ 

+++++++++++++++++++++++++ 

Special Senses System 

Harderian gland 

Adenoma 

Carcinoma 

+ 

X 

+ 

X 

Urinary System 

Kidney 

Urinary bladder 

+A+++++++++++++++++++++++ 

+A+++++++++A+++++++++++++ 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant 

+++++++++++++++++++++++++ 

X 

X X X X X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 223 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 125 ppm 

7777777777777777777777777 

Number of Days on Study 3333333333333333333333333 

1111122222222222223333333 

5555555555555555555555555 Total 

Carcass ID Number 3344400122233344450022344 

6716715101223903504838848 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Hemangiosarcoma 

Lymph node 

Renal, teratoma malignant, metastatic, ovary 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Histiocytic sarcoma 

Lymph node, mediastinal 

Spleen 

Thymus 

+++++++++++++++++++++++++ 

X 

+ 

+++++M+++++M+++++++++++++ 

+++M++++++++++++++M++++++ 

+++++++++++++++++++++++++ 

X 

M +++++++MMM++++++++++M+++ 

+++++++++++++++++++++++++ 

+++++++++++++++M+++++++++ 

50 

1 

6 

1 

41 

40 

49 

1 

38 

49 

46 

Integumentary System 

Mammary gland 

Skin 

Subcutaneous tissue, hemangioma 

Subcutaneous tissue, hemangiosarcoma 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X 

50 

50 

1 

1 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 50 

Nervous System 

Brain +++++++++++++++++++++++++ 50 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Histiocytic sarcoma 

Nose 

Trachea 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X 

X 

X X 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

50 

4 

1 

6 

1 

49 

50 

Special Senses System 

Harderian gland 

Adenoma 

Carcinoma 

+ 

X 

3 

2 

1 

Urinary System 

Kidney 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

49 

48 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant 

+++++++++++++++++++++++++ 

X 

X X X X X 

50 

2 

10 
 

 

 

 

 

 

 

 

 

 

224 2-Butoxyethanol, NTP TR 484 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 250 ppm 

Number of Days on Study 

4445556666666777777777777 

3882881255568033333333333 

7599228136602811111111111 

Carcass ID Number 

7777777777777777777777777 

1031341102420300111222223 

0391405877748219237023893 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma 

Hepatocholangiocarcinoma 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Squamous cell carcinoma 

Squamous cell papilloma 

Stomach, glandular 

Tongue 

+++++++++++++++++++++++++ 

+ + A A + + + + A + + + + + + + + + + + + + + + + 

+++++++++++++++++++++++++ 

++++++++M++++++++++++++++ 

+ + A A +++++++++++++++++++++ 

+ + A A ++++A++++++++++++++++ 

+ + A A +++++++++++++++++++++ 

++ A A +++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X X X X X 

X 

X X 

X 

+ + ++ + 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

X X X 

+ + A ++++++++++++++++++++++ 

Cardiovascular System 

Blood vessel 

Heart 

Hepatocholangiocarcinoma, metastatic, liver 

+ 

+++++++++++++++++++++++++ 

X 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 

Adrenal medulla +++++++++++++++++++++++++ 

Pheochromocytoma benign 

Islets, pancreatic +++++++++++++++++++++++++ 

Parathyroid gland M+M+++M+M+++++++MM+++++++ 

Pituitary gland +++++++++++++++++++++++++ 

Pars distalis, adenoma X X 

Thyroid gland +++++++++++++++++++++++++ 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Clitoral gland M+++M+++++++++++++M++++++ 

Ovary ++++++++++++++++++++++++M 

Granulosa cell tumor benign X 

Uterus +++++++++++++++++++++++++ 

Polyp stromal 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 225 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 250 ppm 

7777777777777777777777777 

Number of Days on Study 3333333333333333333333333 

1111122222222222222333333 

7777777777777777777777777 Total 

Carcass ID Number 3444400011222333445013344 Tissues/ 

5345824546156078190691626 Tumors 

Alimentary System 

Esophagus +++++++++++++++++++++++++ 50 

Gallbladder +++++++++++++++M+++++++++ 46 

Intestine large, colon +++++++++++++++++++++++++ 50 

Intestine large, rectum +++++++++++++++++++++++++ 49 

Intestine large, cecum +++++++++++++++++++++++++ 48 

Intestine small, duodenum +++++++++++++++++++++++++ 47 

Intestine small, jejunum +++++++++++++++++++++++++ 48 

Intestine small, ileum +++++++++++++++++++++++++ 48 

Liver +++++++++++++++++++++++++ 50 

Hepatocellular carcinoma X X X 9 

Hepatocellular carcinoma, multiple 1 

Hepatocellular adenoma X X X X X X 8 

Hepatocholangiocarcinoma 1 

Mesentery + + + + 9 

Pancreas +++++++++++++++++++++++++ 50 

Salivary glands +++++++++++++++++++++++++ 50 

Stomach, forestomach +++++++++++++++++++++++++ 50 

Squamous cell carcinoma X 1 

Squamous cell papilloma X X 5 

Stomach, glandular +++++++++++++++++++++++++ 49 

Tongue + 1 

Cardiovascular System 

Blood vessel + 2 

Heart +++++++++++++++++++++++++ 50 

Hepatocholangiocarcinoma, metastatic, liver 1 

Endocrine System 

Adrenal cortex +++++++++++++++++++++++++ 50 

Adrenal medulla +++++++++++++++++++++++++ 50 

Pheochromocytoma benign X 1 

Islets, pancreatic +++++++++++++++++++++++++ 50 

Parathyroid gland +MM++++M++++++++++++M++++ 40 

Pituitary gland ++++++++++++++++++++M++++ 49 

Pars distalis, adenoma X X 4 

Thyroid gland ++++++++++++++++++++M++++ 49 

Follicular cell, adenoma X 1 

General Body System 

None 

Genital System 

Clitoral gland +++++++++++++++++++++++++ 47 

Ovary +++++++++++++++++++++++++ 49 

Granulosa cell tumor benign 1 

Uterus +++++++++++++++++++++++++ 50 

Polyp stromal X X 2 
 

 

 

 

 

 

 

 

 

 

 

226 2-Butoxyethanol, NTP TR 484 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 250 ppm 

4445556666666777777777777 

Number of Days on Study 3882881255568033333333333 

7599228136602811111111111 

7777777777777777777777777 

Carcass ID Number 1031341102420300111222223 

0391405877748219237023893 

Hematopoietic System 

Bone marrow 

Hemangiosarcoma 

Lymph node 

Lymph node, bronchial 

Hepatocholangiocarcinoma, metastatic, liver 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Hepatocholangiocarcinoma, metastatic, liver 

Spleen 

Thymus 

Hepatocholangiocarcinoma, metastatic, liver 

+++++++++++++++++++++++++ 

+ 

M ++++M+MM+++++M+++++M+++M 

X 

M +++++++++++M+M++M+++++++ 

+++++++++++++++++++++++++ 

++++++++M+++M+++++++M+M+M 

X 

+++++++++++++++++++++++++ 

++++++++M++++++++++++++M+ 

X 

Integumentary System 

Mammary gland 

Carcinoma 

Skin 

Sebaceous gland, adenoma 

+++++++++++++++++++++++++ 

X X 

+++++++++++++++++++++++++ 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 

Nervous System 

Brain +++++++++++++++++++++++++ 

Respiratory System 

Larynx +++++++++++++++++++++++++ 
 

Lung +++++++++++++++++++++++++ 
 

Alveolar/bronchiolar adenoma 

Carcinoma, metastatic, harderian gland X 

Hepatocellular carcinoma, metastatic, liver X 

Hepatocholangiocarcinoma, metastatic, liver X 

Mediastinum, hepatocholangiocarcinoma,

 metastatic, liver X 

Nose +++++++++++++++++++++++++ 

Trachea +++++++++++++++++++++++++ 

Special Senses System 

Eye ++ 

Harderian gland ++ 

Adenoma X 

Carcinoma X X 

Urinary System 

Kidney +++++++++++++++++++++++++ 

Urinary bladder +++++++++++++++++++++++++ 

Systemic Lesions 

Multiple organs +++++++++++++++++++++++++ 

Lymphoma malignant X X X X X X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 227 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol: 250 ppm 

7777777777777777777777777 

Number of Days on Study 3333333333333333333333333 

1111122222222222222333333 

7777777777777777777777777 Total 

Carcass ID Number 3444400011222333445013344 

5345824546156078190691626 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Hemangiosarcoma 

Lymph node 

Lymph node, bronchial 

Hepatocholangiocarcinoma, metastatic, liver 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Hepatocholangiocarcinoma, metastatic, liver 

Spleen 

Thymus 

Hepatocholangiocarcinoma, metastatic, liver 

+++++++++++++++++++++++++ 

X 

M M + + M M +++++++M+++++++++M+ 

M + M M + + M M ++++++M+++++M+++M 

+++++++++++++++++++++++++ 

M M + M + MMM+++M+M++++M++MM++ 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

1 

1 

37 

1 

38 

50 

34 

1 

50 

48 

1 

Integumentary System 

Mammary gland 

Carcinoma 

Skin 

Sebaceous gland, adenoma 

+++++++++++++++++++++++++ 

X 

+++++++++++++++++++++++++ 

X 

50 

3 

50 

1 

Musculoskeletal System 

Bone +++++++++++++++++++++++++ 50 

Nervous System 

Brain +++++++++++++++++++++++++ 50 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Carcinoma, metastatic, harderian gland 

Hepatocellular carcinoma, metastatic, liver 

Hepatocholangiocarcinoma, metastatic, liver 

Mediastinum, hepatocholangiocarcinoma, 

metastatic, liver 

Nose 

Trachea 

++++++++++++++++++++M++++ 

+++++++++++++++++++++++++ 

X 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

49 

50 

1 

1 

1 

1 

1 

50 

50 

Special Senses System 

Eye 

Harderian gland 

Adenoma 

Carcinoma 

+ 

+ 

X 

+ 

+ 

X 

4 

4 

3 

2 

Urinary System 

Kidney 

Urinary bladder 

+++++++++++++++++++++++++ 

+++++++++++++++++++++++++ 

50 

50 

Systemic Lesions 

Multiple organs 

Lymphoma malignant 

+++++++++++++++++++++++++ 

X 

50 

7 
228 2-Butoxyethanol, NTP TR 484 

TABLE D3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Adrenal Medulla: Benign Pheochromocytoma 

Overall ratea 

Adjusted rateb 3/50 (6%) 

7.0% 

1/50 (2%) 

2.4% 

1/49 (2%) 

2.4% 

1/50 (2%) 

2.3% 

Terminal ratec 2/29 (7%) 0/31 (0%) 1/33 (3%) 1/36 (3%) 

First incidence (days) 

Poly-3 testd 

713 

P=0.238N 

592 

P=0.311N 

731 (T) 

P=0.321N 

731 (T) 

P=0.297N 

Harderian Gland: Adenoma 

Overall rate 4/50 (8%) 3/50 (6%) 2/50 (4%) 3/50 (6%) 

Adjusted rate 9.3% 7.2% 4.8% 6.8% 

Terminal rate 4/29 (14%) 3/31 (10%) 2/33 (6%) 2/36 (6%) 

First incidence (days) 731 (T) 731 (T) 731 (T) 656 

Poly-3 test P=0.405N P=0.514N P=0.348N P=0.484N 

Harderian Gland: Adenoma or Carcinoma 

Overall rate 5/50 (10%) 4/50 (8%) 3/50 (6%) 4/50 (8%) 

Adjusted rate 11.6% 9.5% 7.1% 9.0% 

Terminal rate 5/29 (17%) 4/31 (13%) 3/33 (9%) 2/36 (6%) 

First incidence (days) 731 (T) 731 (T) 731 (T) 656 

Poly-3 test P=0.408N P=0.516N P=0.369N P=0.478N 

Liver: Hepatocellular Adenoma 

Overall rate 16/50 (32%) 8/50 (16%) 7/49 (14%) 8/50 (16%) 

Adjusted rate 35.8% 18.7% 16.7% 18.0% 

Terminal rate 9/29 (31%) 5/31 (16%) 6/33 (18%) 7/36 (19%) 

First incidence (days) 586 456 715 582 

Poly-3 test P=0.048N P=0.057N P=0.035N P=0.045N 

Liver: Hepatocellular Carcinoma 

Overall rate 10/50 (20%) 12/50 (24%) 13/49 (27%) 10/50 (20%) 

Adjusted rate 22.3% 27.5% 29.8% 21.9% 

Terminal rate 5/29 (17%) 5/31 (16%) 10/33 (30%) 5/36 (14%) 

First incidence (days) 376 592 537 582 

Poly-3 test P=0.483N P=0.375 P=0.288 P=0.583N 

Liver: Hepatocellular Adenoma or Carcinoma 

Overall rate 22/50 (44%) 16/50 (32%) 18/49 (37%) 18/50 (36%) 

Adjusted rate 47.8% 36.1% 41.2% 39.1% 

Terminal rate 13/29 (45%) 8/31 (26%) 14/33 (42%) 12/36 (33%) 

First incidence (days) 376 456 537 582 

Poly-3 test P=0.307N P=0.175N P=0.336N P=0.260N 

Lung: Alveolar/bronchiolar Adenoma 

Overall rate 7/50 (14%) 2/50 (4%) 4/50 (8%) 1/50 (2%) 

Adjusted rate 16.3% 4.8% 9.5% 2.3% 

Terminal rate 7/29 (24%) 2/31 (7%) 2/33 (6%) 1/36 (3%) 

First incidence (days) 731 (T) 731 (T) 715 731 (T) 

Poly-3 test P=0.032N P=0.083N P=0.272N P=0.027N 
2-Butoxyethanol, NTP TR 484 229 

TABLE D3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Lung: Alveolar/bronchiolar Adenoma or Carcinoma 

Overall rate 7/50 (14%) 4/50 (8%) 5/50 (10%) 1/50 (2%) 

Adjusted rate 16.3% 9.5% 11.9% 2.3% 

Terminal rate 7/29 (24%) 3/31 (10%) 3/33 (9%) 1/36 (3%) 

First incidence (days) 731 (T) 676 715 731 (T) 

Poly-3 test P=0.027N P=0.271N P=0.394N P=0.027N 

Mammary Gland: Carcinoma 

Overall rate 0/50 (0%) 1/50 (2%) 0/50 (0%) 3/50 (6%) 

Adjusted rate 0.0% 2.4% 0.0% 6.7% 

Terminal rate 0/29 (0%) 0/31 (0%) 0/33 (0%) 1/36 (3%) e First incidence (days) — 694 — 621 

Poly-3 test P=0.045 P=0.495 —f P=0.125 

Ovary: Cystadenoma 

Overall rate 2/50 (4%) 1/49 (2%) 3/49 (6%) 0/49 (0%)
 

Adjusted rate 4.6% 2.4% 7.3% 0.0%
 

Terminal rate 1/29 (3%) 1/31 (3%) 3/32 (9%) 0/35 (0%)
 

First incidence (days) 721 731 (T) 731 (T) —
 

Poly-3 test P=0.236N P=0.514N P=0.477 P=0.237N
 

Pituitary Gland (Pars Distalis): Adenoma 

Overall rate 5/50 (10%) 8/49 (16%) 8/48 (17%) 4/49 (8%)
 

Adjusted rate 11.5% 19.6% 18.9% 9.2%
 

Terminal rate 3/29 (10%) 8/30 (27%) 7/33 (21%) 2/35 (6%)
 

First incidence (days) 565 731 (T) 570 660
 

Poly-3 test P=0.334N P=0.233 P=0.255 P=0.502N
 

Pituitary Gland (Pars Distalis): Adenoma or Carcinoma 

Overall rate 6/50 (12%) 8/49 (16%) 8/48 (17%) 4/49 (8%)
 

Adjusted rate 13.7% 19.6% 18.9% 9.2%
 

Terminal rate 3/29 (10%) 8/30 (27%) 7/33 (21%) 2/35 (6%)
 

First incidence (days) 565 731 (T) 570 660
 

Poly-3 test P=0.251N P=0.335 P=0.361 P=0.372N
 

Skin: Sarcoma 

Overall rate 3/50 (6%) 2/50 (4%) 0/50 (0%) 0/50 (0%)
 

Adjusted rate 6.8% 4.6% 0.0% 0.0%
 

Terminal rate 0/29 (0%) 0/31 (0%) 0/33 (0%) 0/36 (0%)
 

First incidence (days) 521 369 — —
 

Poly-3 test P=0.039N P=0.510N P=0.127N P=0.118N
 

Stomach (Forestomach): Squamous Cell Papilloma 

Overall rate 0/50 (0%) 1/50 (2%) 2/50 (4%) 5/50 (10%)
 

Adjusted rate 0.0% 2.4% 4.8% 11.2%
 

Terminal rate 0/29 (0%) 1/31 (3%) 2/33 (6%) 3/36 (8%)
 

First incidence (days) — 731 (T) 731 (T) 582
 

Poly-3 test P=0.008 P=0.495 P=0.231 P=0.034
 
230 2-Butoxyethanol, NTP TR 484 

TABLE D3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Stomach (Forestomach): Squamous Cell Papilloma or Carcinoma 

Overall rate 0/50 (0%) 1/50 (2%) 2/50 (4%) 6/50 (12%)
 

Adjusted rate 0.0% 2.4% 4.8% 13.4%
 

Terminal rate 0/29 (0%) 1/31 (3%) 2/33 (6%) 4/36 (11%)
 

First incidence (days) — 731 (T) 731 (T) 582
 

Poly-3 test P=0.002 P=0.495 P=0.231 P=0.017
 

Thyroid Gland (Follicular Cell): Adenoma 

Overall rate 1/50 (2%) 2/50 (4%) 5/50 (10%) 1/49 (2%) 

Adjusted rate 2.3% 4.8% 11.9% 2.3% 

Terminal rate 1/29 (3%) 1/31 (3%) 5/33 (15%) 1/35 (3%) 

First incidence (days) 731 (T) 712 731 (T) 731 (T) 

Poly-3 test P=0.581 P=0.492 P=0.095 P=0.761N 

Thyroid Gland (Follicular Cell): Adenoma or Carcinoma 

Overall rate 1/50 (2%) 3/50 (6%) 5/50 (10%) 1/49 (2%) 

Adjusted rate 2.3% 7.1% 11.9% 2.3% 

Terminal rate 1/29 (3%) 2/31 (7%) 5/33 (15%) 1/35 (3%) 

First incidence (days) 731 (T) 712 731 (T) 731 (T) 

Poly-3 test P=0.537N P=0.296 P=0.095 P=0.761N 

Uterus: Stromal Polyp 

Overall rate 1/50 (2%) 2/50 (4%) 4/50 (8%) 2/50 (4%) 

Adjusted rate 2.3% 4.8% 9.5% 4.5% 

Terminal rate 1/29 (3%) 2/31 (7%) 3/33 (9%) 2/36 (6%) 

First incidence (days) 731 (T) 731 (T) 723 731 (T) 

Poly-3 test P=0.406 P=0.491 P=0.171 P=0.508 

Uterus: Stromal Polyp or Stromal Sarcoma 

Overall rate 1/50 (2%) 2/50 (4%) 4/50 (8%) 2/50 (4%) 

Adjusted rate 2.3% 4.8% 9.5% 4.5% 

Terminal rate 1/29 (3%) 2/31 (7%) 3/33 (9%) 2/36 (6%) 

First incidence (days) 731 (T) 731 (T) 723 731 (T) 

Poly-3 test P=0.406 P=0.491 P=0.171 P=0.508 

All Organs: Hemangioma 

Overall rate 3/50 (6%) 0/50 (0%) 2/50 (4%) 0/50 (0%)
 

Adjusted rate 6.9% 0.0% 4.7% 0.0%
 

Terminal rate 1/29 (3%) 0/31 (0%) 1/33 (3%) 0/36 (0%)
 

First incidence (days) 572 — 591 —
 

Poly-3 test P=0.118N P=0.126N P=0.513N P=0.117N
 

All Organs: Hemangiosarcoma 

Overall rate 2/50 (4%) 2/50 (4%) 3/50 (6%) 1/50 (2%) 

Adjusted rate 4.6% 4.7% 7.1% 2.3% 

Terminal rate 1/29 (3%) 1/31 (3%) 3/33 (9%) 1/36 (3%) 

First incidence (days) 689 670 731 (T) 731 (T) 

Poly-3 test P=0.392N P=0.686 P=0.487 P=0.493N 
231 

c 

2-Butoxyethanol, NTP TR 484 

TABLE D3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

All Organs: Hemangioma or Hemangiosarcoma 

Overall rate 5/50 (10%) 2/50 (4%) 5/50 (10%) 1/50 (2%) 

Adjusted rate 11.4% 4.7% 11.8% 2.3% 

Terminal rate 2/29 (7%) 1/31 (3%) 4/33 (12%) 1/36 (3%) 

First incidence (days) 572 670 591 731 (T) 

Poly-3 test P=0.124N P=0.231N P=0.611 P=0.100N 

All Organs: Malignant Lymphoma 

Overall rate 7/50 (14%) 9/50 (18%) 10/50 (20%) 7/50 (14%) 

Adjusted rate 16.1% 20.8% 23.0% 15.2% 

Terminal rate 5/29 (17%) 5/31 (16%) 5/33 (15%) 3/36 (8%) 

First incidence (days) 572 544 397 437 

Poly-3 test P=0.448N P=0.384 P=0.292 P=0.568N 

All Organs: Benign Neoplasms 

Overall rate 31/50 (62%) 23/50 (46%) 27/50 (54%) 18/50 (36%) 

Adjusted rate 67.4% 53.0% 61.3% 39.8% 

Terminal rate 19/29 (66%) 18/31 (58%) 22/33 (67%) 14/36 (39%) 

First incidence (days) 565 456 196 582 

Poly-3 test P=0.007N P=0.112N P=0.345N P=0.005N 

All Organs: Malignant Neoplasms 

Overall rate 27/50 (54%) 26/50 (52%) 28/50 (56%) 22/50 (44%) 

Adjusted rate 56.9% 56.4% 60.2% 45.3% 

Terminal rate 12/29 (41%) 12/31 (39%) 17/33 (52%) 11/36 (31%) 

First incidence (days) 376 369 72 437 

Poly-3 test P=0.132N P=0.563N P=0.456 P=0.172N 

All Organs: Benign or Malignant Neoplasms 

Overall rate 43/50 (86%) 38/50 (76%) 38/50 (76%) 35/50(70%) 

Adjusted rate 87.7% 81.1% 80.0% 71.0% 

Terminal rate 24/29 (83%) 23/31 (74%) 26/33 (79%) 22/36 (61%) 

First incidence (days) 376 369 72 437 

Poly-3 test P=0.023N P=0.268N P=0.219N P=0.033N 

(T)Terminal sacrifice 

a Number of neoplasm-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for adrenal gland, 

liver, lung, ovary, pituitary gland, and thyroid gland; for other tissues, denominator is number of animals necropsied. b Poly-3 estimated neoplasm incidence after adjustment for intercurrent mortality 

Observed incidence at terminal kill 

d	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values 

corresponding to pairwise comparisons between the chamber controls and that exposed group. The Poly-3 test accounts for differential mortality in 

animals that do not reach terminal sacrifice. A negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group f Value of statistic cannot be computed. 
232 2-Butoxyethanol, NTP TR 484 

TABLE D4a 

Historical Incidence of Forestomach Neoplasms in Chamber Control Female B6C3F1 Micea 

Incidence in Controls 

Squamous Cell Papilloma Squamous Cell Carcinoma Squamous Cell Papilloma 

or Carcinoma 

Historical Incidence at Battelle Pacific Northwest Laboratories 

1,3-Butadiene 0/50 0/50 0/50 

Acetonitrile 1/49 0/49 1/49 

Chloroprene 0/50 1/50 1/50 

Cobalt sulfate heptahydrate 1/50 0/50 1/50 

Furfuryl alcohol 0/50 0/50 0/50 

Hexachlorocyclopentadiene 0/49 0/49 0/49 

Isobutene 0/50 0/50 0/50 

Isobutyraldehyde 0/50 0/50 0/50 

Molybdenum trioxide 0/50 0/50 0/50 

Nitromethane 1/50 0/50 1/50 

Ozone 0/50 0/50 0/50 

Tetrahydrofuran 1/50 0/50 1/50 

Overall Historical Incidence 

Total (%) 7/973 (0.7%) 2/973 (0.2%) 9/973 (0.9%) 

Mean ± standard deviation 0.7% ± 1.0% 0.2% ± 0.6% 0.9% ± 1.1% 

Range 0%-2% 0%-2% 0%-3% 

a Data as of 16 October 1997 
2-Butoxyethanol, NTP TR 484 233 

TABLE D4b 

Historical Incidence of Liver Neoplasms in Chamber Control Female B6C3F1 Micea 

Incidence in Controls 

Hemangiosarcoma Hepatocellular Hepatocellular Hepatocellular 

Adenoma Carcinoma Adenoma or Carcinoma 

Historical Incidence at Battelle Pacific Northwest Laboratories 

1,3-Butadiene 1/49 11/49 4/49 15/49 

Acetonitrile 0/49 4/49 7/49 9/49 

Chloroprene 1/50 17/50 4/50 20/50 

Cobalt sulfate heptahydrate 1/50 8/50 12/50 18/50 

Furfuryl alcohol 0/50 7/50 9/50 14/50 

Hexachlorocyclopentadiene 0/49 5/49 4/49 9/49 

Isobutene 0/47 20/47 5/47 23/47 

Isobutyraldehyde 1/49 9/49 6/49 12/49 

Molybdenum trioxide 0/50 9/50 19/50 23/50 

Nitromethane 2/50 14/50 10/50 19/50 

Ozone 0/50 20/50 15/50 27/50 

Tetrahydrofuran 0/50 12/50 6/50 17/50 

Overall Historical Incidence 

Total (%) 8/968 (0.8%) 191/968 (19.7%) 149/968 (15.4%) 302/968 (31.2%) 

Mean ± standard deviation 0.8% ± 1.3% 19.8% ± 10.2% 15.4% ± 8.1% 31.3% ± 10.6% 

Range 0%-4% 8%-43% 8%-38% 18%-54% 

a Data as of 16 October 1997 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

234 2-Butoxyethanol, NTP TR 484 

TABLE D5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Accidental death 1 

Moribund 14 16 12 10 

Natural deaths 6 3 5 4 

Survivors 

Terminal sacrifice 29 31 33 36 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Esophagus (50) (49) (50) (50) 

Epithelium, hyperplasia 1 (2%) 

Intestine large, colon (49) (50) (48) (50) 

Inflammation 1 (2%) 

Epithelium, hyperplasia 1 (2%) 

Intestine large, cecum (49) (50) (47) (48) 

Epithelium, hyperplasia 1 (2%) 

Intestine small, duodenum (45) (49) (47) (47) 

Inflammation 2 (4%) 

Necrosis 1 (2%) 

Epithelium, hyperplasia 1 (2%) 2 (4%) 2 (4%) 

Intestine small, jejunum (46) (49) (47) (48) 

Inflammation 1 (2%) 

Necrosis 1 (2%) 

Intestine small, ileum (46) (50) (47) (48) 

Peyer’s patch, hyperplasia 1 (2%) 

Liver (50) (50) (49) (50) 

Angiectasis 1 (2%) 2 (4%) 

Basophilic focus 1 (2%) 1 (2%) 

Clear cell focus 2 (4%) 1 (2%) 1 (2%) 

Degeneration, fatty 2 (4%) 1 (2%) 1 (2%) 1 (2%) 

Eosinophilic focus 10 (20%) 6 (12%) 10 (20%) 5 (10%) 

Hematopoietic cell proliferation 5 (10%) 7 (14%) 1 (2%) 1 (2%) 

Hyperplasia, lymphoid 1 (2%) 

Infarct 1 (2%) 

Inflammation 3 (6%) 5 (10%) 4 (8%) 4 (8%) 

Necrosis 2 (4%) 8 (16%) 7 (14%) 3 (6%) 

Thrombosis 1 (2%) 

Bile duct, cyst 1 (2%) 

Endothelial cell, hyperplasia 1 (2%) 

Kupffer cell, pigmentation, hemosiderin 5 (10%) 25 (51%) 44 (88%) 

Mesentery (10) (4) (5) (9) 

Fat, necrosis 7 (70%) 3 (75%) 4 (80%) 9 (100%) 

Pancreas (49) (50) (49) (50) 

Atrophy 6 (12%) 4 (8%) 2 (4%) 8 (16%) 

Cytoplasmic alteration 1 (2%) 2 (4%) 2 (4%) 

Hyperplasia, lymphoid 1 (2%) 

Inflammation 3 (6%) 

Duct, cyst 1 (2%) 1 (2%) 

Duct, inflammation 1 (2%) 

a Number of animals examined microscopically at the site and the number of animals with lesion 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 235 

TABLE D5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Alimentary System (continued) 

Stomach, forestomach (50) (50) (49) (50) 

Cyst epithelial inclusion 1 (2%) 

Erosion 1 (2%) 

Ulcer 1 (2%) 7 (14%) 13 (27%) 22 (44%) 

Epithelium, hyperplasia 6 (12%) 27 (54%) 42 (86%) 44 (88%) 

Stomach, glandular (48) (50) (49) (49) 

Inflammation, suppurative 1 (2%) 

Mineralization 1 (2%) 

Ulcer 1 (2%) 1 (2%) 1 (2%) 1 (2%) 

Epithelium, hyperplasia 1 (2%) 1 (2%) 1 (2%) 

Tongue (1) (1) 

Epithelium, hyperplasia 1 (100%) 

Tooth (1) 

Inflammation 1 (100%) 

Cardiovascular System 

Blood vessel (1) (2) 

Inflammation 1 (50%) 

Mineralization 1 (100%) 

Heart (50) (50) (50) (50) 

Cardiomyopathy 46 (92%) 45 (90%) 44 (88%) 45 (90%) 

Mineralization 1 (2%) 

Artery, inflammation 1 (2%) 

Endocrine System 

Adrenal cortex (50) (50) (50) (50) 

Amyloid deposition 1 (2%) 

Degeneration, cystic 2 (4%) 

Hematopoietic cell proliferation 1 (2%) 2 (4%) 

Hyperplasia 1 (2%) 

Hypertrophy 3 (6%) 2 (4%) 6 (12%) 5 (10%) 

Capsule, hyperplasia 1 (2%) 

Adrenal medulla (50) (50) (49) (50) 

Hyperplasia 1 (2%) 3 (6%) 3 (6%) 2 (4%) 

Islets, pancreatic (50) (50) (49) (50) 

Hyperplasia 1 (2%) 

Pituitary gland (50) (49) (48) (49) 

Pars distalis, angiectasis 1 (2%) 4 (8%) 

Pars distalis, hyperplasia 15 (30%) 15 (31%) 16 (33%) 20 (41%) 

Pars intermedia, hyperplasia 1 (2%) 2 (4%) 1 (2%) 1 (2%) 

Thyroid gland (50) (50) (50) (49) 

Inflammation 1 (2%) 

Follicle, cyst 1 (2%) 

Follicular cell, hyperplasia 11 (22%) 8 (16%) 11 (22%) 11 (22%) 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

236 2-Butoxyethanol, NTP TR 484 

TABLE D5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Genital System 

Ovary (50) (49) (49) (49) 

Angiectasis 2 (4%) 2 (4%) 1 (2%) 

Atrophy 1 (2%) 2 (4%) 2 (4%) 1 (2%) 

Cyst 11 (22%) 12 (24%) 19 (39%) 16 (33%) 

Degeneration, fatty 1 (2%) 

Infiltration cellular, mast cell 1 (2%) 

Inflammation 1 (2%) 1 (2%) 

Thrombosis 1 (2%) 2 (4%) 

Corpus luteum, hyperplasia 1 (2%) 

Germinal epithelium, hyperplasia 1 (2%) 1 (2%) 

Interstitial cell, hyperplasia 1 (2%) 

Uterus (50) (50) (49) (50) 

Adenomyosis 1 (2%) 

Angiectasis 2 (4%) 3 (6%) 1 (2%) 

Hydrometra 2 (4%) 2 (4%) 1 (2%) 3 (6%) 

Hyperplasia, cystic 2 (4%) 2 (4%) 6 (12%) 

Inflammation 1 (2%) 

Lymphangiectasis 1 (2%) 

Thrombosis 1 (2%) 

Arteriole, hyperplasia 1 (2%) 

Hematopoietic System 

Bone marrow (50) (50) (50) (50) 

Fibrosis 1 (2%) 

Hyperplasia 2 (4%) 4 (8%) 2 (4%) 3 (6%) 

Hyperplasia, megakaryocyte 1 (2%) 

Lymph node (4) (7) (6) (1) 

Angiectasis 1 (25%) 1 (14%) 

Hyperplasia 1 (25%) 

Iliac, hyperplasia 1 (25%) 

Pancreatic, hyperplasia 1 (14%) 

Lymph node, bronchial (41) (39) (41) (37) 

Hematopoietic cell proliferation 1 (2%) 

Hyperplasia 1 (2%) 6 (15%) 4 (10%) 

Hyperplasia, histiocytic 1 (3%) 

Lymph node, mandibular (37) (38) (40) (38) 

Hyperplasia 1 (3%) 2 (5%) 2 (5%) 1 (3%) 

Lymph node, mesenteric (47) (48) (49) (50) 

Hematopoietic cell proliferation 1 (2%) 1 (2%) 1 (2%) 

Hyperplasia 1 (2%) 2 (4%) 3 (6%) 3 (6%) 

Lymph node, mediastinal (31) (40) (38) (34) 

Hyperplasia 2 (6%) 2 (5%) 5 (13%) 

Hyperplasia, lymphoid 1 (3%) 

Spleen (50) (50) (49) (50) 

Hematopoietic cell proliferation 24 (48%) 29 (58%) 32 (65%) 35 (70%) 

Hyperplasia, lymphoid 6 (12%) 15 (30%) 12 (24%) 10 (20%) 

Pigmentation, hemosiderin 39 (78%) 44 (88%) 46 (94%) 48 (96%) 

Thymus (46) (41) (46) (48) 

Atrophy 1 (2%) 2 (5%) 

Hyperplasia, lymphoid 1 (2%) 2 (5%) 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2-Butoxyethanol, NTP TR 484 237 

TABLE D5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Integumentary System 

Mammary gland (50) (50) (50) (50) 

Hyperplasia 2 (4%) 2 (4%) 1 (2%) 2 (4%) 

Skin (50) (50) (50) (50) 

Atrophy 1 (2%) 

Inflammation, chronic active 1 (2%) 

Necrosis 1 (2%) 

Pinna, inflammation, chronic 1 (2%) 

Subcutaneous tissue, edema 1 (2%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Arthrosis 1 (2%) 

Fibrous osteodystrophy 22 (44%) 28 (56%) 25 (50%) 21 (42%) 

Maxilla, fracture 1 (2%) 

Nervous System 

Brain (50) (50) (50) (50) 

Cyst epithelial inclusion 1 (2%) 

Hemorrhage 1 (2%) 1 (2%) 1 (2%) 

Inflammation, chronic 1 (2%) 

Necrosis 1 (2%) 

Meninges, infiltration cellular,

 mononuclear cell 2 (4%) 2 (4%) 

Respiratory System 

Larynx (50) (50) (50) (49) 

Inflammation, suppurative 1 (2%) 3 (6%) 1 (2%) 

Squamous epithelium, hyperplasia 2 (4%) 8 (16%) 3 (6%) 5 (10%) 

Lung (50) (50) (50) (50) 

Hemorrhage 10 (20%) 2 (4%) 6 (12%) 6 (12%) 

Infiltration cellular, histiocyte 3 (6%) 2 (4%) 3 (6%) 2 (4%) 

Inflammation 1 (2%) 

Metaplasia, osseous 1 (2%) 

Pigmentation, hemosiderin 1 (2%) 

Alveolar epithelium, hyperplasia 2 (4%) 2 (4%) 2 (4%) 

Mediastinum, inflammation 1 (2%) 

Perivascular, inflammation 2 (4%) 

Nose (50) (50) (49) (50) 

Inflammation, chronic active 1 (2%) 1 (2%) 

Inflammation, suppurative 2 (4%) 3 (6%) 1 (2%) 

Olfactory epithelium, atrophy 2 (4%) 4 (8%) 2 (4%) 4 (8%) 

Olfactory epithelium, degeneration, hyaline 6 (12%) 14 (28%) 11 (22%) 12 (24%) 

Olfactory epithelium, metaplasia 1 (2%) 1 (2%) 

Respiratory epithelium, degeneration, hyaline 17 (34%) 35 (70%) 26 (53%) 23 (46%) 

Special Senses System 

Eye (1) (4) 

Degeneration 1 (25%) 

Cornea, inflammation, chronic 1 (100%) 3 (75%) 
 

 

 

 

 

 

238 2-Butoxyethanol, NTP TR 484 

TABLE D5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber 

Control 62.5 ppm 125 ppm 250 ppm 

Urinary System 

Kidney (50) (50) (49) (50) 

Glomerulosclerosis 7 (14%) 2 (4%) 8 (16%) 8 (16%) 

Inflammation, chronic active 1 (2%) 

Metaplasia, osseous 1 (2%) 1 (2%) 3 (6%) 4 (8%) 

Nephropathy 34 (68%) 39 (78%) 38 (78%) 33 (66%) 

Pigmentation, hemosiderin 1 (2%) 1 (2%) 

Renal tubule, degeneration 1 (2%) 

Renal tubule, hyperplasia 1 (2%) 

Renal tubule, mineralization 1 (2%) 

Urinary bladder (47) (49) (48) (50) 

Inflammation 1 (2%) 1 (2%) 
239 

APPENDIX E
 

GENETIC TOXICOLOGY
 

SALMONELLA TYPHIMURIUM MUTAGENICITY TEST PROTOCOL ............................ 240 
 

CHINESE HAMSTER OVARY CELL CYTOGENETICS PROTOCOLS ............................ 240 
 

RAT AND MOUSE BONE MARROW MICRONUCLEUS TEST PROTOCOL ........................ 241 
 

EVALUATION PROTOCOL .................................................... 242 
 

RESULTS ............................................................... 242 
 

TABLE E1 Mutagenicity of 2-Butoxyethanol in Salmonella typhimurium ................... 243 
 

TABLE E2 Induction of Sister Chromatid Exchanges in Chinese Hamster Ovary Cells
 

by 2-Butoxyethanol ............................................... 245 
 

TABLE E3 Induction of Chromosomal Aberrations in Chinese Hamster Ovary Cells 

by 2-Butoxyethanol ............................................... 246 
 

TABLE E4 Induction of Micronuclei in Bone Marrow Polychromatic Erythrocytes
 

of Male Rats Treated with 2-Butoxyethanol by Intraperitoneal Injection ........... 248 
 

TABLE E5 Induction of Micronuclei in Bone Marrow Polychromatic Erythrocytes
 

of Male Mice Treated with 2-Butoxyethanol by Intraperitoneal Injection .......... 248 
 
 

240 2-Butoxyethanol, NTP TR 484 

GENETIC TOXICOLOGY 

SALMONELLA TYPHIMURIUM MUTAGENICITY TEST PROTOCOL 

Testing was performed as reported by Zeiger et al. (1992). 2-Butoxyethanol was sent to the laboratory as a 

coded aliquot from Radian Corporation (Austin, TX). It was incubated with the Salmonella typhimurium 

tester strains TA97, TA98, TA100, TA1535, and TA1537 either in buffer or S9 mix (metabolic activation 

enzymes and cofactors from Aroclor 1254-induced male Sprague-Dawley rat or Syrian hamster liver) for 

20 minutes at 37E C. Top agar supplemented with L-histidine and d-biotin was added, and the contents of 

the tubes were mixed and poured onto the surfaces of minimal glucose agar plates. Histidine-independent 

mutant colonies arising on these plates were counted following incubation for 2 days at 37E C. 

Each trial consisted of triplicate plates of concurrent positive and negative controls and five doses of 

2-butoxyethanol. In the absence of toxicity, 10,000 μg/plate was selected as the high dose. 

In this assay, a positive response is defined as a reproducible, dose-related increase in histidine-independent 

(revertant) colonies in any one strain/activation combination. An equivocal response is defined as an 

increase in revertants that is not dose related, is not reproducible, or is not of sufficient magnitude to 

support a determination of mutagenicity. A negative response is obtained when no increase in revertant 

colonies is observed following chemical treatment. There is no minimum percentage or fold increase 

required for a chemical to be judged positive or weakly positive. 

CHINESE HAMSTER OVARY CELL CYTOGENETICS PROTOCOLS 

Testing was performed as reported by Galloway et al. (1987). 2-Butoxyethanol was sent to the laboratory 

as a coded aliquot by Radian Corporation. It was tested in cultured Chinese hamster ovary (CHO) cells for 

induction of sister chromatid exchanges (SCEs) and chromosomal aberrations (Abs), both in the presence 

and absence of Aroclor 1254-induced male Sprague-Dawley rat liver S9 and cofactor mix. Cultures were 

handled under gold lights to prevent photolysis of bromodeoxyuridine-substituted DNA. Each test consisted 

of concurrent solvent and positive controls and of three doses of 2-butoxyethanol. In the SCE test, the 

highest testable dose of 2-butoxyethanol, in the absence of S9, was limited by toxicity to 3,000 (Trial 1) or 

3,500 μg/mL (Trial 2); with S9, no toxicity was observed and the high dose was limited to 5,000 μg/mL. 

In the Abs test, the high dose was limited to 5,000 μg/mL. A single flask per dose was used, and tests 

yielding equivocal or positive results were repeated. 

Sister Chromatid Exchange Test: In the SCE test without S9, CHO cells were incubated for 26 hours with 

2-butoxyethanol in supplemented McCoy’s 5A medium. Bromodeoxyuridine (BrdU) was added 2 hours 

after culture initiation. After 26 hours, the medium containing 2-butoxyethanol was removed and replaced 

with fresh medium plus BrdU and Colcemid, and incubation was continued for 2 hours. Cells were then 

harvested by mitotic shake-off, fixed, and stained with Hoechst 33258 and Giemsa. In the SCE test with 

S9, cells were incubated with 2-butoxyethanol, serum-free medium, and S9 for 2 hours. The medium was 

then removed and replaced with medium containing serum and BrdU and no 2-butoxyethanol. Incubation 

proceeded for an additional 26 hours, with Colcemid present for the final 2 hours. Harvesting and staining 

were the same as for cells treated without S9. All slides were scored blind and those from a single test 

were read by the same person. Fifty second-division metaphase cells were scored for frequency of 

SCEs/cell from each dose level. Since a significant chemical-induced cell cycle delay was seen, incubation 

time was lengthened to ensure a sufficient number of scorable (second-division metaphase) cells. 

Statistical analyses were conducted on the slopes of the dose-response curves and the individual dose points 

(Galloway et al., 1987). An SCE frequency 20% above the concurrent solvent control value was chosen as 
 

2-Butoxyethanol, NTP TR 484 241 

a statistically conservative positive response. The probability of this level of difference occurring by chance 

at one dose point is less than 0.01; the probability for such a chance occurrence at two dose points is less 

than 0.001. An increase of 20% or greater at any single dose was considered weak evidence of activity; 

increases at two or more doses resulted in a determination that the trial was positive. A statistically 

significant trend (P<0.005) in the absence of any responses reaching 20% above background led to a call 

of equivocal. 

Chromosomal Aberrations Test: In the Abs test without S9, cells were incubated in McCoy’s 5A medium 

with 2-butoxyethanol for 8.5 hours; Colcemid was added and incubation continued for 2 hours. The cells 

were then harvested by mitotic shake-off, fixed, and stained with Giemsa. For the Abs test with S9, cells 

were treated with 2-butoxyethanol and S9 for 2 hours, after which the treatment medium was removed and 

the cells were incubated for 8.5 hours in fresh medium, with Colcemid present for the final 2 hours. Cells 

were harvested in the same manner as for the treatment without S9. The harvest time for the Abs test was 

based on the cell cycle information obtained in the SCE test: if cell cycle delay was anticipated, the 

incubation period was extended. 

Cells were selected for scoring on the basis of good morphology and completeness of karyotype 

(21 ± 2 chromosomes). All slides were scored blind and those from a single test were read by the same 

person. One or two hundred first-division metaphase cells were scored at each dose level. Classes of 

aberrations included simple (breaks and terminal deletions), complex (rearrangements and translocations), 

and other (pulverized cells, despiralized chromosomes, and cells containing 10 or more aberrations). 

Chromosomal aberration data are presented as percentage of cells with aberrations. To arrive at a statistical 

call for a trial, analyses were conducted on both the dose response curve and individual dose points. For a 

single trial, a statistically significant (P#0.05) difference for one dose point and a significant trend 

(P#0.015) were considered weak evidence for a positive response; significant differences for two or more 

doses indicated the trial was positive. A positive trend test in the absence of a statistically significant 

increase at any one dose resulted in an equivocal call (Galloway et al., 1987). Ultimately, the trial calls 

were based on a consideration of the statistical analyses as well as the biological information available to the 

reviewers. 

RAT AND MOUSE BONE MARROW MICRONUCLEUS TEST PROTOCOL 

Preliminary range-finding studies were performed. Factors affecting dose selection included chemical 

solubility and toxicity and the extent of cell cycle delay induced by 2-butoxyethanol exposure. The standard 

three-exposure protocol is described in detail by Shelby et al. (1993). Published toxicity information was 

used to select doses. Male F344/N rats and B6C3F1 mice were injected intraperitoneally three times at 

24-hour intervals with 2-butoxyethanol dissolved in phosphate-buffered saline; the total dosing volume was 

0.4 mL. Solvent control animals were injected with 0.4 mL of phosphate-buffered saline. The positive 

control animals received injections of cyclophosphamide. The animals were killed 24 hours after the final 

injection, and blood smears were prepared from bone marrow cells obtained from the femurs. Air-dried 

smears were fixed and stained; 2,000 polychromatic erythrocytes (PCEs) were scored for the frequency of 

micronucleated cells in each of five animals per dose group. 

The results were tabulated as the mean of the pooled results from all animals within a treatment group plus 

or minus the standard error of the mean. The frequency of micronucleated cells among PCEs was analyzed 

by a statistical software package that tested for increasing trend over dose groups with a one-tailed CochranArmitage trend test, followed by pairwise comparisons between each dosed group and the control group 

(ILS, 1990). In the presence of excess binomial variation, as detected by a binomial dispersion test, the 

binomial variance of the Cochran-Armitage test was adjusted upward in proportion to the excess variation. 
 

242 2-Butoxyethanol, NTP TR 484 

In the micronucleus test, an individual trial is considered positive if the trend test P value is less than or 

equal to 0.025 or if the P value for any single dose group is less than or equal to 0.025 divided by the 

number of dose groups. A final call of positive for micronucleus induction is preferably based on 

reproducibly positive trials (as noted above). Ultimately, the final call is determined by the scientific staff 

after considering the results of statistical analyses, the reproducibility of any effects observed, and the 

magnitudes of those effects. 

EVALUATION PROTOCOL 

These are the basic guidelines for arriving at an overall assay result for assays performed by the National 

Toxicology Program. Statistical as well as biological factors are considered. For an individual assay, the 

statistical procedures for data analysis have been described in the preceding protocols. There have been 

instances, however, in which multiple aliquots of a chemical were tested in the same assay, and differing 

results were obtained among aliquots and/or among laboratories. Results from more than one aliquot or 

from more than one laboratory are not simply combined into an overall result. Rather, all the data are 

critically evaluated, particularly with regard to pertinent protocol variations, in determining the weight of 

evidence for an overall conclusion of chemical activity in an assay. In addition to multiple aliquots, the 

in vitro assays have another variable that must be considered in arriving at an overall test result. In vitro 

assays are conducted with and without exogenous metabolic activation. Results obtained in the absence of 

activation are not combined with results obtained in the presence of activation; each testing condition is 

evaluated separately. The summary table in the Abstract of this Technical Report presents a result that 

represents a scientific judgement of the overall evidence for activity of the chemical in an assay. 

RESULTS 

2-Butoxyethanol (100 to 10,000 μg/plate) did not induce mutations in any of the S. typhimurium strains 

tested (TA97, TA98, TA100, TA1535, and TA1537), with or without induced hamster or rat liver S9 

(Table E1; Zeiger et al., 1992). In tests for induction of chromosomal damage in CHO cells in vitro, 

2-butoxyethanol induced cell cycle delay (an indication of cytotoxicity) but did not induce either SCEs 

(Table E2) or Abs (Table E3) with or without S9. In the Abs test without S9, a weakly positive response 

was obtained in the second trial at the highest dose tested (5,000 μg/mL), but this response was not 

reproduced in a third trial and the test results were concluded to be negative overall. Due to the cell cycle 

delay caused by 2-butoxyethanol in the trials conducted without S9, a delayed harvest was used to increase 

the number of cells available for analysis. In vivo, no induction of micronuclei was observed in PCEs in 

bone marrow of rats or mice treated with 2-butoxyethanol (Tables E4 and E5). Rats received up to 

450 mg/kg 2-butoxyethanol three times at 24-hour intervals via intraperitoneal injection; two out of five rats 

administered 450 mg/kg dose died. Mice were treated by the same protocol. All mice receiving 550 mg/kg 

survived, whereas 100% mortality occurred in the 1,100 mg/kg dose groups. 
2-Butoxyethanol, NTP TR 484 243 

TABLE E1 

Mutagenicity of 2-Butoxyethanol in Salmonella typhimuriuma 

Revertants/plateb 

Strain Dose 

(μg/plate) 

!S9 + hamster S9 

10% 30% 5% 

+rat S9 

10% 30% 

TA100 0 

100 

333 

1,000 

3,333 

10,000 

164 ± 5.5 

156 ±11.6 

157 ± 7.1 

165 ± 17.0 

166 ± 2.6 

112 ± 7.8 

153 ± 9.5 

157 ± 3.2 

161 ± 12.7 

156 ± 1.8 

151 ± 4.8 

132 ± 15.5 

161 ± 7.5 

166 ± 3.2 

157 ± 17.5 

150 ± 1.5 

151 ± 13.3 

149 ± 12.1 

172 ± 6.1 

155 ± 8.3 

167 ± 8.5 

169 ± 4.7 

156 ± 3.3 

142 ± 9.2 

111 ± 2.8 

127 ± 12.1 

99 ± 5.0 

96 ± 3.7 

150 ± 2.3 

119 ± 4.0 

Trial summary 

Positive controlc Negative 

428 ± 30.9 

Negative 

930 ± 56.0 

Negative 

731 ± 52.2 

Negative 

471 ± 17.0 

Negative 

621 ± 9.8 

TA1535 0 

100 

333 

1,000 

3,333 

10,000 

30 ± 4.9 

39 ± 0.3 

33 ± 4.3 

25 ± 3.2 

25 ± 3.2 

22 ± 2.5 

14 ± 0.9 

13 ± 0.6 

12 ± 1.8 

8 ± 1.8 

13 ± 4.0 

7 ± 2.5 

14 ± 2.6 

12 ± 1.5 

14 ± 3.4 

12 ± 2.0 

12 ± 0.6 

10 ± 1.0 

12 ± 0.3 

10 ± 4.7 

11 ± 1.3 

11 ± 0.7 

8 ± 0.9 

11 ± 0.6 

13 ± 0.3 

14 ± 1.5 

14 ± 0.3 

12 ± 2.5 

10 ± 0.3 

10 ± 2.0 

Trial summary 

Positive control 

Negative 

585 ± 26.0 

Negative 

203 ± 10.1 

Negative 

698 ± 29.8 

Negative 

195 ± 16.0 

Negative 

186 ± 4.5 

TA1537 0 

100 

333 

1,000 

3,333 

10,000 

11 ± 3.2 

13 ± 2.6 

13 ± 1.9 

10 ± 1.9 

9 ± 1.3 

14 ± 2.4 

13 ± 1.5 

14 ± 2.1 

7 ± 1.2 

12 ± 1.5 

10 ± 2.3 

11 ± 1.3 

13 ± 3.4 

11 ± 1.8 

8 ± 1.2 

9 ± 3.3 

12 ± 4.1 

7 ± 0.6 

Trial summary 

Positive control 

Negative 

742 ± 61.5 

Negative 

64 ± 3.8 

Negative 

49 ± 2.9 

TA97 0 

100 

333 

666 

1,000 

1,666 

3,333 

10,000 

180 ± 15.1 

178 ± 4.9 

190 ± 8.4 

214 ± 3.7 

190 ± 2.7 

181 ± 1.8 

171 ± 10.4 

170 ± 18.0 

169 ± 3.0 

204 ± 6.9 

172 ± 11.5 

148 ± 10.3 

180 ± 3.0 

210 ± 8.2 

197 ± 5.2 

193 ± 3.3 

164 ± 0.7 

130 ± 4.1 

183 ± 11.9 

177 ± 8.9 

187 ± 2.0 

154 ± 9.5 

169 ± 10.3 

161 ± 19.1 

178 ± 6.6 

195 ± 8.5 

195 ± 16.5 

195 ± 15.1 

184 ± 6.4 

166 ± 22.1 

198 ± 11.3 

215 ± 13.2 

210 ± 5.0 

170 ± 15.2 

149 ± 11.4 

178 ± 2.9 

Trial summary 

Positive control 

Negative 

799 ± 76.2 

Negative 

285 ± 14.7 

Negative 

465 ± 20.5 

Negative 

494 ± 22.3 

Negative

355 ± 13.1 

Negative 

308 ± 8.8 
244 

c 

2-Butoxyethanol, NTP TR 484 

TABLE E1 

Mutagenicity of 2-Butoxyethanol in Salmonella typhimurium 

Revertants/plate 

Strain Dose !S9 + hamster S9 + rat S9 

(μg/plate) 10% 30% 10% 30% 

TA98 0 25 ± 2.3 19 ± 0.6 32 ± 1.9 34 ± 1.9 40 ± 0.6 

100 24 ± 3.0 26 ± 1.0 22 ± 3.4 33 ± 3.5 35 ± 4.7 

333 22 ± 2.5 20 ± 0.9 28 ± 2.0 22 ± 3.4 37 ± 5.7 

1,000 25 ± 5.0 27 ± 0.6 28 ± 0.9 24 ± 3.2 34 ± 1.2 

3,333 21 ± 2.8 26 ± 2.9 30 ± 1.2 27 ± 1.7 34 ± 2.3 

10,000 11 ± 1.5d 21 ± 4.3 27 ± 1.2 23 ± 2.8 42 ± 1.2 

Trial summary Negative Negative Negative Negative Negative 

Positive control 488 ± 48.6 933 ± 29.6 528 ± 35.3 355 ± 7.4 135 ± 6.9 

a Study was performed at SRI International. The detailed protocol and these data are presented by Zeiger et al. (1992). 0 μg/plate was 

the solvent control. 

b	 Revertants are presented as mean ± standard error from three plates. 

The positive controls in the absence of metabolic activation were sodium azide (TA100 and TA1535), 9-aminoacridine (TA97 and 

TA1537), and 4-nitro-o-phenylenediamine (TA98). The positive control for metabolic activation with all strains was 2-aminoanthracene. d	 Slight toxicity 
2-Butoxyethanol, NTP TR 484 245 

TABLE E2 

Induction of Sister Chromatid Exchanges in Chinese Hamster Ovary Cells by 2-Butoxyethanola 

Total No. of SCEs/ Relative 

Concentration Cells Chromo- No. of Chromo- SCEs/ Hrs Change of SCEs/ 

Compound (μg/mL) Scored somes SCEs some Cell in BrdU Chromosomeb 

(%) 

!S9 

Trial 1 

Summary: Equivocal 

Mediumc 50 1,016 418 0.41 8.4 26.0 

Mitomycin-Cd 0.001 50 1,017 568 0.55 11.4 26.0 35.75 

0.010 5 103 150 1.45 30.0 26.0 253.98 

2-Butoxyethanol	 1,510 50 1,004 410 0.40 8.2 31.0e !0.74 

2,220 50 998 453 0.45 9.1 31.0e 10.33 

3,000 50 1,013 496 0.48 9.9 31.0e 19.01 

P=0.001f 

Trial 2 

Summary: Negative 

Medium 50 1,027 485 0.47 9.7 26.0 

Mitomycin-C 0.001 50 1,015 626 0.61 12.5 26.0 30.60 

0.010 5 102 202 1.98 40.4 26.0 319.36 

2-Butoxyethanol	 2,500 50 1,007 531 0.52 10.6 36.0e 11.66 

3,000 50 1,009 541 0.53 10.8 36.0e 13.54 

3,500 50 1,007 551 0.54 11.0 36.0e 15.86 

P=0.010 

+S9 

Summary: Negative 

Medium 50 1,006 491 0.48 9.8 26.0 

Cyclophosphamided 0.4 50 1,038 705 0.67 14.1 26.0 39.16 

2.0 5 102 128 1.25 25.6 26.0 157.11 

2-Butoxyethanol 500 50 1,019 485 0.47 9.7 26.0 !2.48 

1,670 50 1,015 479 0.47 9.6 26.0 !3.31 

5,000 50 1,026 497 0.48 9.9 26.0 !0.75 

P=0.563 

a Study was performed at Litton Bionetics, Inc. The detailed protocol is presented by Galloway et al. (1987). SCE=sister chromatid 

exchange; BrdU=bromodeoxyuridine b	 SCEs/chromosome in treated cells versus SCEs/chromosome in solvent control cells 

Solvent control 

d Positive control e Since 2-butoxyethanol induced a delay in the cell division cycle, harvest time was extended to maximize the number of second-division 

metaphase cells available for analysis. f	 Significance of SCEs/chromosome tested by the linear regression trend test versus log of the dose 

c 
246 2-Butoxyethanol, NTP TR 484 

TABLE E3 

Induction of Chromosomal Aberrations in Chinese Hamster Ovary Cells by 2-Butoxyethanola 

Compound Concentration 

(μg/mL) 

Total Cells 

Scored 

Number 

of Aberrations 

Aberrations/ 

Cell 

Cells with 

Aberrations (%) 

!S9 

Trial 1 

Harvest time: 10.5 hours 

Summary: Negative 

Mediumb 200 7 0.04 3.5 

Mitomycin-Cc 0.25 

0.75 

200 

25 

22 

14 

0.11 

0.56 

10.5 

36.0 

2-Butoxyethanol 2,513 

3,750 

5,000 

200 

200 

100 

3 

2 

0 

0.02 

0.01 

0.00 

1.5 

1.0 

0.0 

P=0.991d 

Trial 2 

Harvest time: 20.5 hourse 

Summary: Weakly positive 

Medium 100 0 0.00 0.0 

Mitomycin-Cf 0.05 

0.08 

25 

200 

22 

16 

0.88 

0.08 

36.0 

5.0 

2-Butoxyethanol 2,513 

3,750 

5,000 

100 

100 

100 

4 

1 

8 

0.04 

0.01 

0.08 

3.0 

1.0 

7.0* 

P=0.007 

Trial 3 

Harvest time: 20.7 hourse 

Summary: Negative 

Medium 100 1 0.01 1.0 

Mitomycin-C 0.05 

0.08 

100 

25 

27 

15 

0.27 

0.60 

22.0 

40.0 

2-Butoxyethanol 4,500 

4,700 

5,000 

100 

100 

100 

1 

3 

2 

0.01 

0.03 

0.02 

1.0 

3.0 

2.0 

P=0.215 
247 

c 

2-Butoxyethanol, NTP TR 484 

TABLE E3 

Induction of Chromosomal Aberrations in Chinese Hamster Ovary Cells by 2-Butoxyethanol 

Compound Concentration Total Cells Number Aberrations/ Cells with 

(μg/mL) Scored of Aberrations Cell Aberrations (%) 

+S9 

Harvest time: 12.5 hours 

Summary: Negative 

Medium 200 6 0.03 3.0 

Cyclophosphamidec 7.5 200 20 0.10 8.0 

37.5 25 10 0.40 36.0 

2-Butoxyethanol	 2,513 100 1 0.01 1.0 

3,750 200 8 0.04 3.5 

5,000 200 6 0.03 3.0 

P=0.368 

* Positive response (P#0.05) versus the solvent control a Study was performed at Litton Bionetics, Inc. The detailed protocol is presented by Galloway et al. (1987). b Solvent control
 

Positive control
 d Significance of percent cells with aberrations tested by the linear regression trend test versus log of the dose e Due to a significant 2-butoxyethanol-induced cell cycle delay, incubation time prior to addition of Colcemid was lengthened to provide 

sufficient first-division metaphase cells at harvest. f Based on the observed responses, the positive control doses were apparently switched at the time of dosing. 
248 

c 

2-Butoxyethanol, NTP TR 484 

TABLE E4 

Induction of Micronuclei in Bone Marrow Polychromatic Erythrocytes of Male Rats Treated 

with 2-Butoxyethanol by Intraperitoneal Injectiona 

Compound Dose 

(mg/kg) 

Number of Rats

 with Erythrocytes Scored 

Micronucleated PCEs/1,000 PCEsb 

Phosphate-buffered salinec 5 1.9 ± 0.2 

Cyclophosphamided 7.50 5 21.0 ± 0.4 

2-Butoxyethanol 7.03 

14.06 

28.12 

56.25 

112.50 

225.00 

450.00 

5 

5 

5 

5 

5 

5 

3 

1.6 ± 0.3 

2.1 ± 0.8 

2.2 ± 0.3 

1.3 ± 0.3 

1.7 ± 0.3 

1.2 ± 0.2 

2.2 ± 0.6

 P=0.570e 

a Study was performed at Environmental Health Research and Testing, Inc. The detailed protocol is presented by Shelby et al. (1993). 

PCE=polychromatic erythrocyte b Mean ± standard error
 

Solvent control
 d Positive control e Significance of micronucleated PCEs/1,000 PCEs tested by the one-tailed trend test, significant at P#0.025 (ILS, 1990) 

TABLE E5 

Induction of Micronuclei in Bone Marrow Polychromatic Erythrocytes of Male Mice Treated 

with 2-Butoxyethanol Intraperitoneal Injectiona 

Compound Dose 

(mg/kg) 

Number of Mice

 with Erythrocytes Scored 

Micronucleated PCEs/1,000 PCEsb 

Phosphate-buffered salinec 5 2.5 ± 0.2 

Cyclophosphamided 10.00 5 12.9 ± 1.3 

2-Butoxyethanol 17.19 

34.38 

68.78 

137.50 

275.00 

550.00 

5 

5 

5 

5 

5 

5 

2.6 ± 0.9 

2.3 ± 0.3 

3.2 ± 0.9 

3.8 ± 0.8e 

3.7 ± 0.4 

2.8 ± 0.4

 P=0.236f 

a Study was performed at Environmental Health Research and Testing, Inc. The detailed protocol is presented by Shelby et al. (1993). 

PCE=polychromatic erythrocyte b Mean ± standard error 

Solvent control 

d Positive control e P=0.05 (pairwise comparison to the solvent control, P#0.004 required for significance) f Significance of micronucleated PCEs/1,000 PCEs tested by the one-tailed trend test, significant at P#0.025 (ILS, 1990) 

c 
249 

APPENDIX F 

HEMATOLOGY 

AND BONE MARROW CELLULARITY RESULTS 

TABLE F1 Hematology Data for Rats in the 14-Week Inhalation Study of 2-Butoxyethanol ...... 250 

TABLE F2 Hematology and Bone Marrow Cellularity Data for Rats 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 251 

TABLE F3 Hematology Data for Mice in the 14-Week Inhalation Study of 2-Butoxyethanol ..... 255 

TABLE F4 Hematology and Bone Marrow Cellularity Data for Mice 

in the 2-Year Inhalation Study of 2-Butoxyethanol ......................... 256 
250 2-Butoxyethanol, NTP TR 484 

TABLE F1 

Hematology Data for Rats in the 14-Week Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 31 ppm 62.5 ppm 125 ppm 250 ppm 500 ppm 

Male 

n 10 10 10 10 10 10 

Automated hematocrit (mL/dL) 45.5 ± 0.4 43.8 ± 0.7 45.1 ± 0.4 42.7 ± 0.5** 38.4 ± 0.4** 34.9 ± 0.3** 

Manual hematocrit (%) 46.8 ± 0.5 45.8 ± 0.6 47.0 ± 0.4 44.5 ± 0.5** 41.1 ± 0.3** 37.3 ± 0.4** 

Hemoglobin (g/dL) 

Erythrocytes (106

/μL) 

Reticulocytes (106

/μL) 

Nucleated erythrocytes (103

/μL) 

15.5 ± 0.1 

9.05 ± 0.08 

0.16 ± 0.02 

0.04 ± 0.02 

14.8 ± 0.3 

8.71 ± 0.14* 

0.17 ± 0.03 

0.05 ± 0.01 

15.4 ± 0.1 

8.91 ± 0.06 

0.15 ± 0.02 

0.04 ± 0.03 

14.5 ± 0.2** 

8.01 ± 0.08** 

0.30 ± 0.04** 

0.11 ± 0.03 

13.1 ± 0.1** 

7.10 ± 0.07** 

0.48 ± 0.06** 

0.17 ± 0.04** 

11.7 ± 0.1** 

5.97 ± 0.05** 

0.68 ± 0.07** 

0.20 ± 0.06* 

Mean cell volume (fL) 50.4 ± 0.3 50.2 ± 0.2 50.7 ± 0.2 53.1 ± 0.2** 53.8 ± 0.3** 58.5 ± 0.3** 

Mean cell hemoglobin (pg) 17.1 ± 0.1 17.0 ± 0.1 17.3 ± 0.1 18.1 ± 0.1** 18.4 ± 0.1** 19.5 ± 0.1** 

Mean cell hemoglobin

 concentration (g/dL) 

Platelets (103

/μL) 

Leukocytes (103

/μL) 

Segmented neutrophils (103

/μL) 

Lymphocytes (103

/μL) 

Monocytes (103

/μL) 

Eosinophils (103

/μL) 

33.9 ± 0.2 

482.3 ± 20.4 

6.70 ± 0.29 

1.12 ± 0.09 

5.34 ± 0.26 

0.19 ± 0.04 

0.05 ± 0.02 

33.7 ± 0.2 

487.6 ± 31.3 

6.29 ± 0.38 

1.08 ± 0.09 

4.99 ± 0.33 

0.19 ± 0.05 

0.03 ± 0.01 

34.2 ± 0.2 

499.0 ± 26.7 

6.13 ± 0.33 

1.12 ± 0.08 

4.82 ± 0.28 

0.11 ± 0.03 

0.08 ± 0.03 

33.9 ± 0.1 

469.1 ± 20.8 

5.87 ± 0.24* 

1.04 ± 0.09 

4.72 ± 0.22 

0.08 ± 0.02* 

0.02 ± 0.01 

34.1 ± 0.2 

486.2 ± 21.9 

5.72 ± 0.36* 

0.99 ± 0.23 

4.65 ± 0.20* 

0.06 ± 0.02** 

0.02 ± 0.01 

33.4 ± 0.1 

533.9 ± 35.9 

5.34 ± 0.17** 

1.32 ± 0.32 

3.93 ± 0.42** 

0.08 ± 0.04* 

0.01 ± 0.01 

Female 

n 10 10 10 10 9 5 

Automated hematocrit (mL/dL) 46.7 ± 0.3 44.7 ± 0.5** 43.6 ± 0.5** 40.5 ± 0.3** 37.4 ± 0.3** 31.9 ± 0.6** 

Manual hematocrit (%) 48.5 ± 0.5 46.0 ± 0.5** 45.2 ± 0.5** 42.9 ± 0.4** 40.0 ± 0.3** 36.2 ± 0.6** 

Hemoglobin (g/dL) 

Erythrocytes (106

/μL) 

Reticulocytes (106

/μL) 

Nucleated erythrocytes (103

/μL) 

15.6 ± 0.1 

8.48 ± 0.05 

0.13 ± 0.02 

0.04 ± 0.02 

15.0 ± 0.1** 

8.08 ± 0.07** 

0.10 ± 0.01 

0.05 ± 0.02 

14.6 ± 0.1** 

7.70 ± 0.08** 

0.16 ± 0.02 

0.12 ± 0.03* 

13.6 ± 0.1** 

6.91 ± 0.05** 

0.26 ± 0.04* 

0.18 ± 0.07 

12.5 ± 0.1** 

6.07 ± 0.04** 

0.34 ± 0.04** 

0.61 ± 0.24** 

10.5 ± 0.3** 

4.77 ± 0.15** 

0.40 ± 0.11** 

0.73 ± 0.27** 

Mean cell volume (fL) 55.1 ± 0.3 55.3 ± 0.2 56.4 ± 0.2** 58.7 ± 0.2** 61.6 ± 0.2** 66.8 ± 0.9** 

Mean cell hemoglobin (pg) 18.4 ± 0.1 18.6 ± 0.2 19.0 ± 0.0** 19.6 ± 0.1** 20.6 ± 0.1** 22.0 ± 0.1** 

Mean cell hemoglobin 

 concentration (g/dL) 

Platelets (103

/μL) 

Leukocytes (103

/μL) 

Segmented neutrophils (103

/μL) 

Lymphocytes (103

/μL) 

Monocytes (103

/μL) 

Eosinophils (103

/μL) 

33.4 ± 0.1 

573.5 ± 19.5 

6.58 ± 0.23 

1.17 ± 0.12 

5.15 ± 0.19 

0.18 ± 0.04b 

0.03 ± 0.01 

33.6 ± 0.3 

576.1 ± 31.6 

6.99 ± 0.40 

1.24 ± 0.15 

5.53 ± 0.31 

0.17 ± 0.05 

0.05 ± 0.02 

33.6 ± 0.1 

583.5 ± 13.3 

7.26 ± 0.41 

1.25 ± 0.12 

5.75 ± 0.35 

0.22 ± 0.05 

0.04 ± 0.02 

33.6 ± 0.2 

657.0 ± 25.7* 

6.88 ± 0.23 

1.20 ± 0.13 

5.45 ± 0.26 

0.18 ± 0.05 

0.04 ± 0.02 

33.4 ± 0.1 

611.6 ± 25.6 

7.64 ± 0.48 

1.24 ± 0.16 

6.19 ± 0.42 

0.19 ± 0.04 

0.03 ± 0.01 

32.9 ± 0.2 

719.6 ± 52.9* 

7.21 ± 0.45 

1.37 ± 0.22 

5.75 ± 0.27 

0.06 ± 0.03 

0.04 ± 0.02 

* Significantly different (P#0.05) from the chamber control group by Dunn’s or Shirley’s test 

** P#0.01 a Mean ± standard error. Statistical tests were performed on unrounded data. b n=9 
 

2-Butoxyethanol, NTP TR 484 251 

TABLE F2 

Hematology and Bone Marrow Cellularity Data for Rats in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber Control 31.2 ppm 62.5 ppm 125 ppm 

Male 

Hematology 

n 

3 Months 9 9 9 9 

6 Months 

12 Months 

9 

8 

8 

0b 

9

9 

8 

9 

Automated hematocrit (mL/dL) 

3 Months 46.2 ± 0.3 48.5 ± 0.4 46.1 ± 0.4 43.5 ± 0.3** 

6 Months 47.1 ± 0.3 46.5 ± 0.6 47.4 ± 0.7 44.3 ± 0.4** 

12 Months 46.1 ± 0.3 — 44.4 ± 0.7** 41.4 ± 1.1** 

Manual hematocrit (%) 

3 Months 44.9 ± 0.2 46.9 ± 0.5 44.8 ± 0.4 42.9 ± 0.5* 

6 Months 47.2 ± 0.2 46.4 ± 0.5 47.2 ± 0.6 44.3 ± 0.7** 

12 Months 47.8 ± 0.4 — 45.9 ± 0.8* 42.9 ± 1.2** 

Hemoglobin (g/dL) 

3 Months 15.0 ± 0.1 15.5 ± 0.1 15.0 ± 0.1 14.2 ± 0.1** 

6 Months 15.2 ± 0.1 15.1 ± 0.2 15.3 ± 0.2 14.4 ± 0.1** 

12 Months 

Erythrocytes (106/μL) 

15.2 ± 0.0 — 14.7 ± 0.2** 13.4 ± 0.3** 

3 Months 8.99 ± 0.06 9.19 ± 0.06 8.84 ± 0.09 8.01 ± 0.06** 

6 Months 9.02 ± 0.08 8.85 ± 0.10 9.05 ± 0.14 8.16 ± 0.07** 

12 Months 

Reticulocytes (106/μL) 

8.88 ± 0.08 — 8.39 ± 0.15** 7.43 ± 0.20** 

3 Months 0.12 ± 0.02 0.14 ± 0.01 0.14 ± 0.02 0.20 ± 0.01** 

6 Months 0.08 ± 0.01 0.09 ± 0.01 0.08 ± 0.01 0.14 ± 0.01** 

12 Months 

Nucleated erythrocytes (103/μL) 

0.11 ± 0.03 — 0.13 ± 0.02 0.19 ± 0.03 

3 Months 0.01 ± 0.01 0.04 ± 0.03 0.02 ± 0.02 0.05 ± 0.02 

6 Months 0.05 ± 0.02 0.00 ± 0.00* 0.01 ± 0.01 0.02 ± 0.01 

12 Months 0.05 ± 0.02 — 0.15 ± 0.10 0.04 ± 0.03 

Mean cell volume (fL) 

3 Months 51.3 ± 0.3 52.8 ± 0.1** 52.0 ± 0.2* 54.2 ± 0.3** 

6 Months 52.3 ± 0.5 52.5 ± 0.2 52.4 ± 0.2 54.4 ± 0.2** 

12 Months 52.0 ± 0.2 — 52.9 ± 0.3* 55.8 ± 0.2** 

Mean cell hemoglobin (pg) 

3 Months 16.7 ± 0.1 16.9 ± 0.0 16.9 ± 0.1 17.7 ± 0.0** 

6 Months 16.9 ± 0.1 17.1 ± 0.1* 16.9 ± 0.1 17.6 ± 0.1** 

12 Months 17.1 ± 0.2 — 17.5 ± 0.1 18.1 ± 0.1** 

Mean cell hemoglobin concentration (g/dL) 

3 Months 32.5 ± 0.1 32.0 ± 0.1* 32.4 ± 0.1 32.6 ± 0.1 

6 Months 32.3 ± 0.3 32.5 ± 0.1 32.3 ± 0.1 32.4 ± 0.1 

12 Months 

Platelets (103/μL) 

33.0 ± 0.2 — 33.1 ± 0.2 32.5 ± 0.2 

3 Months 477.7 ± 6.9 432.1 ± 15.7 486.2 ± 8.8 473.3 ± 10.6 

6 Months 501.2 ± 11.1 572.4 ± 36.1 602.3 ± 71.8 597.6 ± 35.6 

12 Months 

Leukocytes (103/μL) 

762.6 ± 33.5 — 789.2 ± 21.8 748.8 ± 26.9 

3 Months 5.98 ± 0.34 12.37 ± 0.26** 6.71 ± 0.33 6.40 ± 0.32 

6 Months 6.69 ± 0.54 7.83 ± 0.62 7.61 ± 0.77 6.79 ± 0.29 

12 Months 

Segmented neutrophils (103/μL) 

6.45 ± 0.24 — 6.54 ± 0.48 6.10 ± 0.53 

3 Months 1.78 ± 0.34 1.62 ± 0.26 1.09 ± 0.11* 1.23 ± 0.10 

6 Months 1.93 ± 0.27 2.42 ± 0.68 2.33 ± 0.66 1.47 ± 0.16 

12 Months 1.83 ± 0.23 — 1.58 ± 0.25 1.57 ± 0.40 
252 2-Butoxyethanol, NTP TR 484 

TABLE F2 

Hematology and Bone Marrow Cellularity Data for Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber Control 31.2 ppm 62.5 ppm 125 ppm 

Male (continued) 

Hematology (continued) 

n 

3 Months 

6 Months 

12 Months 

9 

9 

8 

9 

8 

0 

9 

9 

9 

9 

8 

9 

Lymphocytes (103

/μL) 

3 Months 

6 Months 

5.18 ± 0.26 

12 Months 

Monocytes (103

/μL) 

3 Months

6 Months 

12 Months 

Eosinophils (103

/μL) 

3 Months 

6 Months 

12 Months 

4.07 ± 0.17 

4.54 ± 0.18

 0.10 ± 0.03 

0.10 ± 0.05 

0.00 ± 0.00

0.04 ± 0.02 

0.07 ± 0.02 

0.09 ± 0.02

10.51 ± 0.29** 

4.59 ± 0.30

 — 

0.11 ± 0.04 

0.10 ± 0.03 

— 

0.13 ± 0.05 

0.06 ± 0.03 

— 

5.47 ± 0.30 

 5.24 ± 0.29 

4.88 ± 0.35 

0.06 ± 0.03 

0.13 ± 0.02 

0.02 ± 0.02 

0.08 ± 0.02 

0.07 ± 0.03 

0.07 ± 0.02 

5.13 ± 0.26 

5.08 ± 0.34 

4.46 ± 0.19 

0.04 ± 0.01 

0.11 ± 0.05 

0.01 ± 0.01 

0.03 ± 0.01 

0.04 ± 0.02 

0.06 ± 0.02 

Bone Marrow Cellularity 

n 

3 Months 

6 Months 

12 Months 

9 

9 

9 

0 

9 

0 

9 

9 

9 

9 

9 

9 

Nucleated bone marrow cells (106

/femur) 

3 Months 

6 Months 

12 Months 

Myeloid/erythroid ratio 

3 Months 

6 Months 

12 Months 

82.9 ± 3.1

103.0 ± 3.0 

111.5 ± 6.4

1.232 ± 0.056

0.958 ± 0.042 

1.107 ± 0.126

 — 

102.8 ± 7.4 

— 

— 

1.122 ± 0.076 

— 

82.5 ± 2.6 

104.3 ± 4.7 

102.6 ± 3.3 

1.063 ± 0.059 

1.118 ± 0.101 

1.042 ± 0.058 

88.8 ± 3.7 

110.6 ± 3.4 

128.1 ± 3.1 

1.023 ± 0.062 

0.826 ± 0.045 

0.720 ± 0.048** 
2-Butoxyethanol, NTP TR 484 253 

TABLE F2 

Hematology and Bone Marrow Cellularity Data for Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber Control 31.2 ppm 62.5 ppm 125 ppm 

Female 

Hematology 

n 

3 Months 8 9 9 9 

6 Months 9 9 9 9 

12 Months 9 0 8 9 

Automated hematocrit (mL/dL) 

3 Months 48.9 ± 0.2 47.3 ± 0.5** 44.9 ± 0.5** 43.0 ± 0.4** 

6 Months 46.0 ± 0.4 41.1 ± 1.5* 42.2 ± 0.9** 40.0 ± 0.3** 

12 Months 44.2 ± 0.3 — 43.7 ± 0.3 40.7 ± 0.4** 

Manual hematocrit (%) 

3 Months 46.5 ± 0.5 46.1 ± 0.5 43.3 ± 0.5** 42.2 ± 0.5** 

6 Months 45.8 ± 0.4 41.9 ± 1.4* 43.1 ± 0.9** 41.4 ± 0.3** 

12 Months 45.4 ± 0.2 — 45.3 ± 0.3 42.3 ± 0.4** 

Hemoglobin (g/dL) 

3 Months 15.5 ± 0.1 14.8 ± 0.2** 14.3 ± 0.2** 13.7 ± 0.1** 

6 Months 15.2 ± 0.1 13.7 ± 0.5** 13.9 ± 0.3** 13.2 ± 0.1** 

12 Months 14.9 ± 0.1 — 14.6 ± 0.1 13.5 ± 0.1** 

Erythrocytes (106

/μL) 

3 Months 8.52 ± 0.03 8.10 ± 0.10** 7.54 ± 0.08** 7.08 ± 0.05** 

6 Months 8.40 ± 0.07 7.50 ± 0.25** 7.54 ± 0.15** 6.89 ± 0.05** 

12 Months 7.81 ± 0.05 — 7.42 ± 0.06** 6.75 ± 0.05** 

Reticulocytes (106

/μL) 

3 Months 0.13 ± 0.01 0.16 ± 0.02 0.18 ± 0.02 0.20 ± 0.02* 

6 Months 0.06 ± 0.01 0.08 ± 0.01 0.12 ± 0.01** 0.17 ± 0.01** 

12 Months 0.06 ± 0.01 — 0.11 ± 0.02* 0.12 ± 0.02** 

Nucleated erythrocytes (103

/μL) 

3 Months 0.10 ± 0.03 0.18 ± 0.02 0.09 ± 0.03 0.26 ± 0.05* 

6 Months 0.05 ± 0.03 0.02 ± 0.02 0.03 ± 0.01 0.04 ± 0.02 

12 Months 0.07 ± 0.02 — 0.05 ± 0.02 0.25 ± 0.08* 

Mean cell volume (fL) 

3 Months 57.4 ± 0.2 58.3 ± 0.2** 59.6 ± 0.4** 60.7 ± 0.4** 

6 Months 54.8 ± 0.3 54.8 ± 0.4 56.0 ± 0.3* 58.2 ± 0.2** 

12 Months 56.8 ± 0.2 — 58.8 ± 0.3** 60.3 ± 0.3** 

Mean cell hemoglobin (pg) 

3 Months 18.2 ± 0.0 18.3 ± 0.1 18.9 ± 0.1** 19.3 ± 0.1** 

6 Months 18.1 ± 0.1 18.3 ± 0.1 18.4 ± 0.1 19.2 ± 0.1** 

12 Months 19.1 ± 0.1 — 19.7 ± 0.1** 20.0 ± 0.1** 

Mean cell hemoglobin concentration (g/dL) 

3 Months 31.7 ± 0.1 31.3 ± 0.2 31.8 ± 0.2 31.8 ± 0.2 

6 Months 33.2 ± 0.2 33.4 ± 0.2 32.9 ± 0.1 33.1 ± 0.1 

12 Months 33.7 ± 0.2 — 33.6 ± 0.2 33.2 ± 0.2 

Platelets (103

/μL) 

3 Months 556.4 ± 14.3 686.6 ± 78.4 591.7 ± 25.3 562.1 ± 25.7 

6 Months 576.3 ± 9.4 563.0 ± 16.6 558.4 ± 18.8 591.9 ± 18.0 

12 Months 663.3 ± 18.2 — 635.0 ± 12.5 702.3 ± 16.0 

Leukocytes (103

/μL) 

3 Months 7.20 ± 0.51 11.79 ± 1.02** 7.18 ± 0.43 7.17 ± 0.22 

6 Months 6.27 ± 0.37 6.48 ± 0.33 6.03 ± 0.39 6.66 ± 0.29 

12 Months 4.74 ± 0.30 — 4.31 ± 0.51 4.70 ± 0.17 

Segmented neutrophils (103

/μL) 

3 Months 1.44 ± 0.30 2.50 ± 0.88 1.19 ± 0.13 1.23 ± 0.08 

6 Months 1.63 ± 0.41 1.38 ± 0.28 1.13 ± 0.10 1.26 ± 0.14 

12 Months 1.07 ± 0.08 — 1.03 ± 0.12 1.00 ± 0.09 
254 2-Butoxyethanol, NTP TR 484 

TABLE F2 

Hematology and Bone Marrow Cellularity Data for Rats in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber Control 31.2 ppm 62.5 ppm 125 ppm 

Female (continued) 

Hematology (continued) 

n 

3 Months 

6 Months 

12 Months 

8 

9 

9 

9 

9 

0 

9 

9 

8 

9 

9 

9 

Lymphocytes (103

/μL) 

3 Months 

6 Months 

12 Months

Monocytes (103

/μL) 

3 Months 

6 Months 

12 Months

Eosinophils (103

/μL) 

3 Months 

6 Months 

12 Months

5.61 ± 0.27 

4.32 ± 0.18 

 3.63 ± 0.29

0.05 ± 0.03 

0.23 ± 0.04 

 0.02 ± 0.01

0.09 ± 0.03 

0.08 ± 0.02 

 0.03 ± 0.01

9.15 ± 0.34** 

4.70 ± 0.35 

— 

0.02 ± 0.02 

0.30 ± 0.04 

— 

0.10 ± 0.03 

0.08 ± 0.02 

— 

5.81 ± 0.32 

4.65 ± 0.32

3.21 ± 0.39 

0.10 ± 0.04 

0.16 ± 0.03 

0.01 ± 0.01 

0.08 ± 0.03 

0.10 ± 0.04 

0.04 ± 0.01 

5.82 ± 0.19 

 5.21 ± 0.22* 

3.67 ± 0.14 

0.04 ± 0.02 

0.17 ± 0.03 

0.00 ± 0.00 

0.06 ± 0.02 

0.02 ± 0.01 

0.04 ± 0.01 

Bone Marrow Cellularity 

n 

3 Months 

9 

6 Months 

12 Months 

9 

9 

8 

9 

0 

0 

9 

8 

9 

9 

9 

Nucleated bone marrow cells (106

/femur) 

3 Months 

6 Months 

89.7 ± 2.4** 

12 Months 

Myeloid/erythroid ratio 

3 Months 

6 Months 

12 Months 

62.8 ± 4.4

74.3 ± 4.8

1.060 ± 0.084

1.137 ± 0.061 

0.846 ± 0.047

 — 

63.6 ± 2.3 

— 

— 

1.037 ± 0.066 

— 

68.4 ± 3.7 

64.8 ± 4.8 

81.8 ± 4.4 

0.840 ± 0.036 

0.788 ± 0.040** 

0.766 ± 0.031 

84.0 ± 2.0** 

64.0 ± 4.5 

110.1 ± 5.6** 

0.744 ± 0.044** 

0.708 ± 0.025** 

0.690 ± 0.046* 

* Significantly different (P#0.05) from the chamber control group by Dunn’s or Shirley’s test 

** P#0.01 a Mean ± standard error. Statistical tests were performed on unrounded data. b Not examined at this exposure concentration 
2-Butoxyethanol, NTP TR 484 255 

TABLE F3 

Hematology Data for Mice in the 14-Week Inhalation Study of 2-Butoxyethanola 

Chamber 

Control 31 ppm 62.5 ppm 125 ppm 250 ppm 500 ppm 

n 10 10 10 10 10 6 

Male 

Automated hematocrit (mL/dL) 47.7 ± 1.0 48.8 ± 0.4 48.0 ± 0.6 47.1 ± 0.5 44.2 ± 0.3** 35.1 ± 1.4** 

Manual hematocrit (%) 47.3 ± 1.0 48.3 ± 0.4 47.6 ± 0.5 46.6 ± 0.4 44.2 ± 0.4** 36.3 ± 1.4** 

Hemoglobin (g/dL) 

Erythrocytes (106

/μL) 

Reticulocytes (106

/μL) 

Nucleated erythrocytes (103

/μL) 

15.7 ± 0.4 

9.71 ± 0.22 

0.21 ± 0.03 

0.00 ± 0.00b 

16.0 ± 0.1 

10.04 ± 0.08 

0.22 ± 0.03 

0.00 ± 0.00 

15.9 ± 0.1 

9.77 ± 0.10 

0.21 ± 0.02 

0.00 ± 0.00 

15.4 ± 0.1** 

9.47 ± 0.06* 

0.32 ± 0.03* 

0.00 ± 0.00 

14.4 ± 0.1** 

8.90 ± 0.07** 

0.45 ± 0.04** 

0.00 ± 0.00 

11.4 ± 0.4** 

7.21 ± 0.23** 

0.79 ± 0.20** 

0.00 ± 0.00 

Mean cell volume (fL) 49.1 ± 0.4 48.5 ± 0.3 49.0 ± 0.4 49.7 ± 0.4 49.8 ± 0.4 48.3 ± 0.9 

Mean cell hemoglobin (pg) 16.2 ± 0.1 16.0 ± 0.1 16.2 ± 0.1 16.2 ± 0.0 16.2 ± 0.1 15.8 ± 0.2 

Mean cell hemoglobin

 concentration (g/dL) 

Platelets (103

/μL) 

Leukocytes (103

/μL) 

Segmented neutrophils (103

/μL) 

Lymphocytes (103

/μL) 

Monocytes (103

/μL) 

Eosinophils (103

/μL) 

33.0 ± 0.2 

922.5 ± 29.9 

2.76 ± 0.27b 

0.41 ± 0.05b 

2.32 ± 0.24b 

0.01 ± 0.00b 

0.01 ± 0.01b 

32.8 ± 0.3 

878.0 ± 22.1 

2.27 ± 0.21 

0.30 ± 0.04 

1.94 ± 0.19 

0.01 ± 0.01 

0.02 ± 0.01 

33.0 ± 0.2 

894.0 ± 23.7 

2.04 ± 0.25 

0.30 ± 0.05 

1.72 ± 0.24 

0.01 ± 0.01 

0.01 ± 0.00 

32.7 ± 0.2 

933.5 ± 30.0 

2.54 ± 0.19 

0.39 ± 0.07 

2.12 ± 0.16 

0.02 ± 0.01 

0.02 ± 0.01 

32.5 ± 0.2 

1,001.3 ± 46.4 

1.91 ± 0.22 

0.28 ± 0.07 

1.61 ± 0.17 

0.00 ± 0.00 

0.01 ± 0.01 

32.5 ± 0.3 

1,176.7 ± 78.2* 

2.13 ± 0.26 

0.18 ± 0.02* 

1.94 ± 0.28 

0.01 ± 0.01 

0.00 ± 0.00 

Female 

Automated hematocrit (mL/dL) 47.1 ± 0.4 46.6 ± 0.3 46.4 ± 0.3 45.4 ± 0.5* 42.0 ± 0.4** 35.8 ± 0.7** 

Manual hematocrit (%) 46.2 ± 0.3 45.9 ± 0.3 45.8 ± 0.3 45.1 ± 0.2** 42.3 ± 0.4** 37.8 ± 1.0** 

Hemoglobin (g/dL) 

Erythrocytes (106

/μL) 

Reticulocytes (106

/μL) 

Nucleated erythrocytes (103

/μL) 

15.7 ± 0.1 

9.72 ± 0.05 

0.18 ± 0.02 

0.00 ± 0.00 

15.4 ± 0.1* 

9.55 ± 0.06* 

0.21 ± 0.03 

0.00 ± 0.00 

15.4 ± 0.1* 

9.51 ± 0.06* 

0.19 ± 0.02 

0.00 ± 0.00 

14.8 ± 0.1** 

9.18 ± 0.05** 

0.29 ± 0.02** 

0.00 ± 0.00 

13.7 ± 0.1** 

8.57 ± 0.06** 

0.47 ± 0.04** 

0.00 ± 0.00 

11.6 ± 0.1** 

7.35 ± 0.07** 

1.17 ± 0.28** 

0.00 ± 0.00 

Mean cell volume (fL) 48.3 ± 0.3 48.8 ± 0.2 48.8 ± 0.2 49.5 ± 0.5 49.0 ± 0.3 48.8 ± 1.0 

Mean cell hemoglobin (pg) 16.1 ± 0.1 16.0 ± 0.1 16.2 ± 0.1 16.1 ± 0.1 16.0 ± 0.0 15.8 ± 0.1 

Mean cell hemoglobin 

 concentration (g/dL) 

Platelets (103

/μL) 

Leukocytes (103

/μL) 

Segmented neutrophils (103

/μL) 

Lymphocytes (103

/μL) 

Monocytes (103

/μL) 

Eosinophils (103

/μL) 

33.3 ± 0.2 

838.0 ± 19.0 

2.68 ± 0.18 

0.26 ± 0.04 

2.38 ± 0.16 

0.01 ± 0.00 

0.03 ± 0.01 

33.0 ± 0.3 

779.7 ± 29.5 

2.77 ± 0.17 

0.28 ± 0.03 

2.46 ± 0.16 

0.01 ± 0.00 

0.02 ± 0.01 

33.2 ± 0.2 

854.7 ± 18.1 

2.46 ± 0.09 

0.24 ± 0.05 

2.19 ± 0.09 

0.00 ± 0.00 

0.03 ± 0.01 

32.6 ± 0.2 

930.3 ± 44.1 

2.48 ± 0.13 

0.23 ± 0.03 

2.21 ± 0.11 

0.02 ± 0.01 

0.02 ± 0.01 

32.6 ± 0.2 

1,032.1 ± 44.1** 

2.52 ± 0.10 

0.25 ± 0.04 

2.24 ± 0.11 

0.01 ± 0.01 

0.02 ± 0.01 

32.4 ± 0.4* 

1,179.0 ± 75.6** 

3.10 ± 0.33 

0.35 ± 0.08 

2.71 ± 0.27 

0.01 ± 0.01 

0.03 ± 0.02 

* Significantly different (P#0.05) from the chamber control group by Dunn’s or Shirley’s test 

** P#0.01 a Mean ± standard error. Statistical tests were performed on unrounded data. b n=9 
256 2-Butoxyethanol, NTP TR 484 

TABLE F4 

Hematology and Bone Marrow Cellularity Data for Mice in the 2-Year Inhalation Study of 2-Butoxyethanola 

Chamber Control 62.5 ppm 125 ppm 250 ppm 

Male 

Hematology 

n 

3 Months 10 10 10 10 

6 Months 10 10 10 10 

12 Months 10 10 9 10 

Automated hematocrit (mL/dL) 

3 Months 46.3 ± 0.4 46.3 ± 0.3 44.9 ± 0.5* 42.5 ± 0.2** 

6 Months 48.1 ± 0.4 47.6 ± 0.5 46.6 ± 0.3* 43.4 ± 0.4** 

12 Months 48.8 ± 0.5 50.1 ± 2.0 47.0 ± 1.0 42.4 ± 0.5** 

Manual hematocrit (%) 

3 Months 47.5 ± 0.3 47.3 ± 0.5 46.0 ± 0.4* 43.7 ± 0.2** 

6 Months 48.1 ± 0.4 48.1 ± 0.4 47.2 ± 0.4 44.5 ± 0.3** 

12 Months 47.9 ± 0.4 48.7 ± 1.9 46.4 ± 1.0 42.1 ± 0.4** 

Hemoglobin (g/dL) 

3 Months 15.2 ± 0.1 15.3 ± 0.1 14.7 ± 0.2 13.8 ± 0.1** 

6 Months 15.7 ± 0.2 15.6 ± 0.1 15.2 ± 0.1** 14.3 ± 0.1** 

12 Months 15.7 ± 0.1 16.0 ± 0.7 14.9 ± 0.4* 13.6 ± 0.2** 

Erythrocytes (106

/μL) 

3 Months 9.61 ± 0.22 9.83 ± 0.06 9.41 ± 0.11 8.95 ± 0.05** 

6 Months 9.88 ± 0.10 9.79 ± 0.08 9.58 ± 0.07* 9.09 ± 0.07** 

12 Months 9.58 ± 0.07 9.73 ± 0.49 9.36 ± 0.32* 8.33 ± 0.10** 

Reticulocytes (106

/μL) 

3 Months 0.07 ± 0.01 0.07 ± 0.01 0.08 ± 0.01 0.13 ± 0.01** 

6 Months 0.05 ± 0.01 0.07 ± 0.01 0.09 ± 0.01** 0.17 ± 0.01** 

12 Months 0.06 ± 0.02 0.06 ± 0.02 0.11 ± 0.02 0.07 ± 0.02 

Mean cell volume (fL) 

3 Months 48.6 ± 1.2 47.1 ± 0.2 47.7 ± 0.2 47.4 ± 0.3 

6 Months 48.8 ± 0.4 48.6 ± 0.3 48.6 ± 0.3 47.8 ± 0.2 

12 Months 50.9 ± 0.3 51.7 ± 0.5 50.3 ± 0.8 51.1 ± 0.6 

Mean cell hemoglobin (pg) 

3 Months 15.9 ± 0.4 15.5 ± 0.0 15.7 ± 0.0 15.4 ± 0.1 

6 Months 15.9 ± 0.1 16.0 ± 0.1 15.9 ± 0.1 15.7 ± 0.1 

12 Months 16.4 ± 0.1 16.5 ± 0.2 16.0 ± 0.3 16.3 ± 0.2 

Mean cell hemoglobin concentration (g/dL) 

3 Months 32.8 ± 0.1 33.0 ± 0.1 32.8 ± 0.1 32.4 ± 0.1 

6 Months 32.6 ± 0.2 32.9 ± 0.2 32.7 ± 0.2 32.9 ± 0.2 

12 Months 32.2 ± 0.2 32.0 ± 0.1 31.7 ± 0.3 31.9 ± 0.1 

Platelets (103

/μL) 

3 Months 904.2 ± 18.3 888.2 ± 11.7 869.8 ± 13.4 940.6 ± 18.9 

6 Months 988.3 ± 16.8 955.4 ± 34.1 1,028.8 ± 20.6 1,075.2 ± 17.1** 

12 Months 

Leukocytes (103

/μL) 

831.2 ± 38.1 997.6 ± 37.8** 1,116.6 ± 69.5** 1,112.8 ± 39.4** 

3 Months 3.63 ± 0.41 3.99 ± 0.37 3.92 ± 0.33 4.01 ± 0.44 

6 Months 2.35 ± 0.35 2.09 ± 0.24 2.69 ± 0.27 2.41 ± 0.26 

12 Months 3.27 ± 0.17 3.19 ± 0.48 3.06 ± 0.26 2.98 ± 0.17 

Segmented neutrophils (103

/μL) 

3 Months 0.67 ± 0.11 0.73 ± 0.08 0.81 ± 0.09 0.95 ± 0.15 

6 Months 0.43 ± 0.08 0.43 ± 0.04 0.75 ± 0.09* 0.62 ± 0.09 

12 Months 0.80 ± 0.04 0.84 ± 0.12 0.80 ± 0.13 0.70 ± 0.08 

Lymphocytes (103

/μL) 

3 Months 2.89 ± 0.31 3.17 ± 0.29 3.05 ± 0.26 3.00 ± 0.32 

6 Months 1.87 ± 0.28 1.63 ± 0.22 1.88 ± 0.19 1.72 ± 0.22 

12 Months 2.45 ± 0.15 2.34 ± 0.38 2.23 ± 0.18 2.27 ± 0.14 
2-Butoxyethanol, NTP TR 484 257 

TABLE F4 

Hematology and Bone Marrow Cellularity Data for Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber Control 62.5 ppm 125 ppm 250 ppm 

Male (continued) 

Hematology (continued) 

n 

3 Months 10 10 10 10 

6 Months 10 10 10 10 

12 Months 10 10 9 10 

Monocytes (103

/μL) 

3 Months 0.00 ± 0.00 0.00 ± 0.00 0.01 ± 0.01 0.00 ± 0.00 

6 Months 0.05 ± 0.02 0.03 ± 0.01 0.06 ± 0.02 0.06 ± 0.02 

12 Months 0.01 ± 0.02 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 

Eosinophils (103

/μL) 

3 Months 0.07 ± 0.02 0.10 ± 0.02 0.04 ± 0.01 0.06 ± 0.02 

6 Months 0.01 ± 0.00 0.00 ± 0.00 0.01 ± 0.01 0.01 ± 0.01 

12 Months 0.01 ± 0.01 0.01 ± 0.01 0.02 ± 0.01 0.01 ± 0.01 

Bone Marrow Cellularity 

Nucleated bone marrow cells (106

/femur) 

3 Months 23.4 ± 0.8 22.8 ± 0.6 22.1 ± 1.1 23.5 ± 0.9 

6 Months 23.8 ± 0.9 23.3 ± 1.0 23.5 ± 1.2 24.3 ± 1.4 

12 Months 32.9 ± 0.9 28.9 ± 0.9 30.1 ± 1.3 30.0 ± 1.5 

Myeloid/erythroid ratio 

3 Months 1.99 ± 0.10 2.11 ± 0.10 1.98 ± 0.13 2.02 ± 0.06 

6 Months 1.88 ± 0.08b 2.24 ± 0.10 2.33 ± 0.09** 2.10 ± 0.09 

12 Months 1.78 ± 0.12 2.15 ± 0.07 2.34 ± 0.07** 2.06 ± 0.10 
258 2-Butoxyethanol, NTP TR 484 

TABLE F4 

Hematology and Bone Marrow Cellularity Data for Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber Control 62.5 ppm 125 ppm 250 ppm 

Female 

Hematology 

n 

3 Months 10 10 10 9 

6 Months 10 10 10 10 

12 Months 10 10 10 10 

Automated hematocrit (mL/dL) 

3 Months 48.9 ± 0.5 48.2 ± 0.3 46.0 ± 0.4** 42.8 ± 0.4** 

6 Months 48.2 ± 0.5 46.2 ± 0.6* 45.7 ± 0.3** 42.5 ± 0.3** 

12 Months 47.4 ± 0.5 46.9 ± 0.5 43.7 ± 0.6** 42.4 ± 0.4** 

Manual hematocrit (%) 

3 Months 49.3 ± 0.5 48.9 ± 0.4 46.2 ± 0.5** 43.7 ± 0.5** 

6 Months 48.1 ± 0.6 46.6 ± 0.5* 45.7 ± 0.2** 42.8 ± 0.2** 

12 Months 46.9 ± 0.4 46.3 ± 0.4 43.8 ± 0.4** 41.8 ± 0.3** 

Hemoglobin (g/dL) 

3 Months 15.5 ± 0.2 15.3 ± 0.1 14.6 ± 0.1** 13.4 ± 0.3** 

6 Months 15.6 ± 0.2 14.9 ± 0.2** 14.7 ± 0.1** 13.6 ± 0.1** 

12 Months 15.4 ± 0.1 15.0 ± 0.1* 14.3 ± 0.1** 13.6 ± 0.1** 

Erythrocytes (106

/μL) 

3 Months 9.89 ± 0.09 9.68 ± 0.07 9.23 ± 0.09** 8.58 ± 0.07** 

6 Months 9.71 ± 0.15 9.33 ± 0.12* 9.19 ± 0.06** 8.68 ± 0.05** 

12 Months 9.32 ± 0.09 9.14 ± 0.08 8.50 ± 0.12** 8.08 ± 0.09** 

Reticulocytes (106

/μL) 

3 Months 0.05 ± 0.00 0.06 ± 0.01 0.09 ± 0.01** 0.16 ± 0.01** 

6 Months 0.05 ± 0.01 0.06 ± 0.01 0.09 ± 0.01** 0.14 ± 0.01** 

12 Months 0.10 ± 0.01 0.14 ± 0.02 0.15 ± 0.02 0.24 ± 0.03** 

Mean cell volume (fL) 

3 Months 49.3 ± 0.3 49.7 ± 0.2 49.8 ± 0.3 49.8 ± 0.1 

6 Months 49.8 ± 0.6 49.5 ± 0.3 49.8 ± 0.4 49.0 ± 0.3 

12 Months 50.9 ± 0.3 51.3 ± 0.3 51.5 ± 0.2 52.4 ± 0.3** 

Mean cell hemoglobin (pg) 

3 Months 15.7 ± 0.1 15.8 ± 0.0 15.8 ± 0.1 15.7 ± 0.1 

6 Months 16.1 ± 0.2 16.0 ± 0.1 16.0 ± 0.1 15.7 ± 0.1 

12 Months 16.6 ± 0.1 16.5 ± 0.1 16.8 ± 0.1 16.8 ± 0.1 

Mean cell hemoglobin concentration (g/dL) 

3 Months 31.8 ± 0.1 31.7 ± 0.1 31.7 ± 0.1 31.4 ± 0.1* 

6 Months 32.4 ± 0.2 32.3 ± 0.2 32.1 ± 0.1 32.1 ± 0.1 

12 Months 32.6 ± 0.2 32.2 ± 0.2 32.7 ± 0.2 32.0 ± 0.2 

Platelets (103

/μL) 

3 Months 835.8 ± 27.5 832.9 ± 26.9 849.9 ± 12.4 921.3 ± 23.3* 

6 Months 938.6 ± 27.2 942.5 ± 34.9 1,010.6 ± 24.9* 1,064.6 ± 12.9** 

12 Months 778.1 ± 28.2 837.6 ± 14.0* 844.5 ± 28.6** 952.9 ± 12.1** 

Leukocytes (103

/μL) 

3 Months 3.11 ± 0.16 2.79 ± 0.19 3.52 ± 0.19 4.10 ± 0.45 

6 Months 3.04 ± 0.16 3.78 ± 0.39 3.62 ± 0.25 3.78 ± 0.36 

12 Months 2.45 ± 0.22 2.74 ± 0.36 3.32 ± 0.22 3.12 ± 0.25 

Segmented neutrophils (103

/μL) 

3 Months 0.49 ± 0.11 0.38 ± 0.05 0.50 ± 0.07 0.59 ± 0.05 

6 Months 0.50 ± 0.05 0.83 ± 0.25 1.05 ± 0.31* 0.74 ± 0.07 

12 Months 0.56 ± 0.06 0.79 ± 0.14 0.75 ± 0.06* 0.88 ± 0.09** 

Lymphocytes (103

/μL) 

3 Months 2.57 ± 0.12 2.38 ± 0.16 2.96 ± 0.17 3.46 ± 0.40 

6 Months 2.43 ± 0.15 2.84 ± 0.27 2.45 ± 0.25 2.94 ± 0.28 

12 Months 1.87 ± 0.18 1.91 ± 0.24 2.46 ± 0.17 2.21 ± 0.18 
259 

c 

2-Butoxyethanol, NTP TR 484 

TABLE F4 

Hematology and Bone Marrow Cellularity Data for Mice in the 2-Year Inhalation Study of 2-Butoxyethanol 

Chamber Control 62.5 ppm 125 ppm 250 ppm 

Female (continued) 

Hematology (continued) 

n 

3 Months 10 10 10 9 

6 Months 10 10 10 10 

12 Months 10 10 10 10 

Monocytes (103

/μL) 

3 Months 0.00 ± 0.00 0.01 ± 0.00 0.01 ± 0.01* 0.00 ± 0.00 

6 Months 0.05 ± 0.02 0.05 ± 0.02 0.05 ± 0.02 0.05 ± 0.02 

12 Months 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 

Eosinophils (103

/μL) 

3 Months 0.06 ± 0.02 0.03 ± 0.01 0.05 ± 0.02 0.05 ± 0.02 

6 Months 0.06 ± 0.02 0.05 ± 0.03 0.07 ± 0.02 0.06 ± 0.02 

12 Months 0.03 ± 0.01 0.04 ± 0.01 0.10 ± 0.03 0.03 ± 0.02 

Bone Marrow Cellularity 

n 

3 Months 10 10 10 9 

6 Months 10 10 10 10 

12 Months 10 10 9 10 

Nucleated bone marrow cells (106

/femur) 

3 Months 23.0 ± 0.9 22.7 ± 1.0 23.2 ± 0.7 22.9 ± 0.9 

6 Months 26.5 ± 0.9 26.3 ± 1.6 24.6 ± 1.2 26.9 ± 0.5 

12 Months 32.7 ± 1.0 31.0 ± 1.9 34.7 ± 1.0 35.8 ± 1.0 

Myeloid/erythroid 

3 Months 1.72 ± 0.12 2.03 ± 0.10 2.19 ± 0.10* 1.52 ± 0.09c 

6 Months 1.81 ± 0.06 1.94 ± 0.13 2.08 ± 0.07* 1.90 ± 0.07 

12 Months 1.53 ± 0.10 1.98 ± 0.08* 1.94 ± 0.13 1.59 ± 0.11 

* Significantly different (P#0.05) from the chamber control group by Dunn’s or Shirley’s test 

** P#0.01 a Mean ± standard error. Statistical tests were performed on unrounded data. b n=9
 

n=10
 
260 2-Butoxyethanol, NTP TR 484 
261 

APPENDIX G 

ORGAN WEIGHTS AND 

ORGAN-WEIGHT-TO-BODY-WEIGHT RATIOS 

TABLE G1 

TABLE G2 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats 

in the 14-Week Inhalation Study of 2-Butoxyethanol ........................ 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice 

in the 14-Week Inhalation Study of 2-Butoxyethanol ........................ 

262 

263 
262 2-Butoxyethanol, NTP TR 484 

TABLE G1 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats in the 14-Week Inhalation Study 

of 2-Butoxyethanola 

Chamber 

Control 31 ppm 62.5 ppm 125 ppm 250 ppm 500 ppm 

Male 

n 10 10 10 10 10 10 

Necropsy body wt 363 ± 7 371 ± 6 375 ± 4 357 ± 9 355 ± 7 366 ± 6 

Heart 

Absolute 1.033 ± 0.023 1.097 ± 0.030 1.030 ± 0.015 1.002 ± 0.020 1.028 ± 0.012 1.106 ± 0.016 

Relative 2.84 ± 0.04 2.95 ± 0.04 2.75 ± 0.04 2.81 ± 0.04 2.90 ± 0.05 3.03 ± 0.04* 

R. Kidney 

Absolute 1.210 ± 0.021 1.265 ± 0.035 1.242 ± 0.020 1.224 ± 0.032 1.265 ± 0.028 1.441 ± 0.029** 

Relative 3.33 ± 0.03 3.40 ± 0.05 3.31 ± 0.05 3.43 ± 0.06 3.56 ± 0.07** 3.94 ± 0.05** 

Liver 

Absolute 12.663 ± 0.304 13.243 ± 0.331b 13.529 ± 0.245 13.015 ± 0.378 14.665 ± 0.543** 16.428 ± 0.384** 

Relative 34.86 ± 0.49 35.83 ± 0.56b 36.07 ± 0.67 36.45 ± 0.76 41.32 ± 1.46** 44.99 ± 1.03** 

Lung 

Absolute 1.709 ± 0.046 1.901 ± 0.095 1.846 ± 0.064 1.746 ± 0.045 1.697 ± 0.046 1.637 ± 0.042 

Relative 4.71 ± 0.10 5.10 ± 0.20 4.91 ± 0.14 4.90 ± 0.11 4.79 ± 0.14 4.48 ± 0.10 

R. Testis 

Absolute 1.429 ± 0.017 1.455 ± 0.024 1.420 ± 0.014 1.397 ± 0.026 1.431 ± 0.022 1.383 ± 0.065 

Relative 3.94 ± 0.08 3.92 ± 0.06 3.78 ± 0.02 3.92 ± 0.09 4.04 ± 0.07 3.81 ± 0.21 

Thymus 

Absolute 0.323 ± 0.012 0.317 ± 0.011 0.337 ± 0.019 0.288 ± 0.013 0.292 ± 0.009 0.290 ± 0.011 

Relative 0.89 ± 0.03 0.86 ± 0.03 0.90 ± 0.05 0.81 ± 0.03 0.82 ± 0.03 0.79 ± 0.03 

Female 

n 10 10 10 10 9 5 

Necropsy body wt 222 ± 5 216 ± 4 211 ± 4 214 ± 4 210 ± 7 201 ± 4* 

Heart 

Absolute 0.715 ± 0.013 0.703 ± 0.011 0.715 ± 0.014 0.724 ± 0.010 0.731 ± 0.021 0.750 ± 0.009 

Relative 3.22 ± 0.05 3.26 ± 0.05 3.39 ± 0.07* 3.39 ± 0.05* 3.49 ± 0.07** 3.73 ± 0.03** 

R. Kidney 

Absolute 0.744 ± 0.023 0.750 ± 0.012 0.761 ± 0.017 0.818 ± 0.015** 0.821 ± 0.027** 0.908 ± 0.015** 

Relative 3.35 ± 0.07 3.48 ± 0.06 3.61 ± 0.08** 3.83 ± 0.05** 3.91 ± 0.06** 4.51 ± 0.04** 

Liver 

Absolute 7.111 ± 0.194 7.052 ± 0.168 7.100 ± 0.207 7.855 ± 0.188* 8.309 ± 0.336** 8.692 ± 0.147** 

Relative 31.98 ± 0.53 32.71 ± 0.61 33.63 ± 0.65 36.73 ± 0.49** 39.58 ± 1.01** 43.22 ± 0.74** 

Lung 

Absolute 1.232 ± 0.040 1.247 ± 0.059 1.185 ± 0.022 1.265 ± 0.039 1.217 ± 0.049 1.112 ± 0.019 

Relative 5.55 ± 0.17 5.81 ± 0.33 5.62 ± 0.10 5.93 ± 0.19 5.79 ± 0.12 5.53 ± 0.12 

Thymus 

Absolute 0.250 ± 0.008 0.249 ± 0.010 0.241 ± 0.011 0.246 ± 0.011 0.246 ± 0.009 0.187 ± 0.011** 

Relative 1.13 ± 0.03 1.15 ± 0.03 1.14 ± 0.04 1.15 ± 0.04 1.18 ± 0.04 0.93 ± 0.06** 

* Significantly different (P#0.05) from the chamber control group by Williams’ or Dunnett’s test 

** P#0.01 a Organ weights (absolute weights) and body weights are given in grams; organ-weight-to-body-weight ratios (relative weights) are given as mg organ 

weight/g body weight (mean ± standard error). b n=9 
2-Butoxyethanol, NTP TR 484 263 

TABLE G2 

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice in the 14-Week Inhalation Study 

of 2-Butoxyethanola 

Chamber 

Control 31 ppm 62.5 ppm 125 ppm 250 ppm 500 ppm 

n 10 10 10 10 10 6 

Male 

Necropsy body wt 37.6 ± 0.9 37.0 ± 0.7 38.1 ± 0.7 35.9 ± 0.8 35.2 ± 0.7* 33.3 ± 0.4** 

Heart 

Absolute 0.167 ± 0.003 0.169 ± 0.005 0.176 ± 0.007 0.164 ± 0.005 0.173 ± 0.008 0.165 ± 0.004 

Relative 4.45 ± 0.11 4.57 ± 0.12 4.62 ± 0.18 4.58 ± 0.14 4.93 ± 0.22 4.95 ± 0.09 

R. Kidney 

Absolute 0.329 ± 0.007 0.314 ± 0.017 0.333 ± 0.009 0.314 ± 0.006 0.319 ± 0.008 0.315 ± 0.013 

Relative 8.74 ± 0.21 8.50 ± 0.48 8.76 ± 0.27 8.77 ± 0.21 9.07 ± 0.15 9.45 ± 0.32 

Liver 

Absolute 1.682 ± 0.044 1.603 ± 0.034 1.751 ± 0.053 1.684 ± 0.031 1.768 ± 0.050 1.838 ± 0.049* 

Relative 44.77 ± 0.99 43.27 ± 0.43 45.97 ± 1.21 47.05 ± 1.24 50.28 ± 0.97** 55.20 ± 1.29** 

Lung 

Absolute 0.236 ± 0.019 0.223 ± 0.003 0.233 ± 0.007 0.219 ± 0.006 0.222 ± 0.006 0.220 ± 0.007 

Relative 6.28 ± 0.51 6.03 ± 0.09 6.12 ± 0.17 6.10 ± 0.10 6.32 ± 0.15 6.61 ± 0.22 

R. Testis 

Absolute 0.113 ± 0.002 0.118 ± 0.003 0.114 ± 0.002 0.115 ± 0.002 0.115 ± 0.002 0.107 ± 0.007 

Relative 3.02 ± 0.09 3.19 ± 0.05 2.99 ± 0.06 3.21 ± 0.05 3.28 ± 0.07 3.22 ± 0.22 

Thymus 

Absolute 0.042 ± 0.003 0.041 ± 0.003 0.042 ± 0.003 0.039 ± 0.002 0.036 ± 0.002 0.037 ± 0.002 

Relative 1.12 ± 0.08 1.11 ± 0.07 1.11 ± 0.08 1.10 ± 0.07 1.03 ± 0.06 1.12 ± 0.07 

Female 

Necropsy body wt 31.3 ± 0.7 32.5 ± 0.9 31.4 ± 0.8 30.9 ± 0.7 31.0 ± 0.4 28.9 ± 0.8 

Heart 

Absolute 0.135 ± 0.003 0.144 ± 0.004 0.141 ± 0.003 0.141 ± 0.003 0.135 ± 0.004 0.147 ± 0.002 

Relative 4.32 ± 0.08 4.47 ± 0.17 4.51 ± 0.12 4.59 ± 0.12 4.36 ± 0.14 5.08 ± 0.12** 

R. Kidney 

Absolute 0.220 ± 0.006 0.220 ± 0.006 0.224 ± 0.006 0.224 ± 0.003 0.220 ± 0.005 0.233 ± 0.011 

Relative 7.07 ± 0.26 6.81 ± 0.21 7.18 ± 0.24 7.28 ± 0.14 7.09 ± 0.12 8.04 ± 0.22** 

Liver 

Absolute 1.495 ± 0.050 1.557 ± 0.059 1.485 ± 0.052 1.539 ± 0.057 1.575 ± 0.045 1.620 ± 0.078 

Relative 47.80 ± 1.21 48.12 ± 1.76 47.43 ± 1.29 49.93 ± 1.58 50.71 ± 0.93 55.89 ± 1.63** 

Lung 

Absolute 0.213 ± 0.004 0.218 ± 0.003 0.218 ± 0.002 0.223 ± 0.004 0.217 ± 0.004 0.210 ± 0.007 

Relative 6.83 ± 0.18 6.75 ± 0.16 6.98 ± 0.13 7.25 ± 0.15 7.00 ± 0.12 7.25 ± 0.08 

Thymus 

Absolute 0.053 ± 0.002 0.055 ± 0.003 0.048 ± 0.002 0.051 ± 0.003 0.047 ± 0.003 0.051 ± 0.002 

Relative 1.70 ± 0.05 1.70 ± 0.07 1.53 ± 0.06 1.65 ± 0.08 1.51 ± 0.08 1.77 ± 0.10 

* Significantly different (P#0.05) from the chamber control group by Williams’ or Dunnett’s test 

** P#0.01 a Organ weights (absolute weights) and body weights are given in grams; organ-weight-to-body-weight ratios (relative weights) are given as mg organ 

weight/g body weight (mean ± standard error). 
264 2-Butoxyethanol, NTP TR 484 
265 

APPENDIX H
 

CHEMICAL CHARACTERIZATION AND
 

GENERATION OF CHAMBER CONCENTRATIONS
 

PROCUREMENT AND CHARACTERIZATION OF 2-BUTOXYETHANOL .......................... 266 
 

VAPOR GENERATION AND EXPOSURE SYSTEM ....................................... 267 
 

VAPOR CONCENTRATION MONITORING ........................................... 268 
 

CHAMBER ATMOSPHERE CHARACTERIZATION ...................................... 268 
 

FIGURE H1 Infrared Absorption Spectrum of 2-Butoxyethanol ......................... 271 
 

FIGURE H2 Nuclear Magnetic Resonance Spectrum of 2-Butoxyethanol .................... 272 
 

TABLE H1 Gas Chromatography Systems Used in the 14-Week and 2-Year Inhalation Studies
 

of 2-Butoxyethanol ............................................... 273 
 

FIGURE H3 Schematic of the Vapor Generation and Delivery System
 

in the 14-Week and 2-Year Inhalation Studies of 2-Butoxyethanol ............... 274 
 

TABLE H2 Summary of Chamber Concentrations in the 14-Week Inhalation Studies
 

of 2-Butoxyethanol ............................................... 275 
 

TABLE H3 Summary of Chamber Concentrations in the 2-Year Inhalation Studies
 

of 2-Butoxyethanol ............................................... 275 
 
266 2-Butoxyethanol, NTP TR 484 

CHEMICAL CHARACTERIZATION AND 

GENERATION OF CHAMBER CONCENTRATIONS 

PROCUREMENT AND CHARACTERIZATION OF 2-BUTOXYETHANOL 

2-Butoxyethanol was obtained from Dow Chemical U.S.A. (Plaquemine, LA) in two lots. 

Lot QP-911021-26D1 was used during the 14-week studies, and lot QP-921215-26D2 was used during the 

2-year studies. Identity and purity analyses were conducted by the study laboratory. Reports on analyses 

performed in support of the 2-butoxyethanol studies are on file at the National Institute of Environmental 

Health Sciences. 

Both lots of the chemical, a clear, colorless liquid, were identified as 2-butoxyethanol by infrared and 

nuclear magnetic resonance (proton and C13) spectroscopy. The spectra were consistent with those expected 

for the structure, with the literature spectra (Aldrich, 1981, 1983), and/or with those of a reference sample 

obtained from Aldrich Chemical Company (Milwaukee, WI). The infrared and proton nuclear magnetic 

spectra are presented in Figures H1 and H2. 

The purity of each lot was determined by elemental analysis (performed by Huffman Laboratories, Inc., 

Golden, CO, for lot QP-911021-26D1 and Galbraith Laboratories, Inc., Knoxville, TN, for 

lot QP-921215-26D2), Karl Fischer water analysis, titrations for acid and peroxide content, and gas 

chromatography. Free acid content was determined by titration with sodium hydroxide to the 

phenolphthalein endpoint. For the determination of peroxide content, samples were dissolved in sulfuric 

acid; molybdate solution and excess potassium iodide were then added, and the samples were allowed to sit 

in the dark for 15 minutes. Starch indicator was added, and the samples were titrated to the colorimetric 

endpoint with approximately 0.005 N sodium thiosulfate. Several gas chromatography systems were used 

to characterize the 2-butoxyethanol used in the 14-week and 2-year studies; the details of the systems are 

given in Table H1. Gas chromatography with flame ionization detection using system A was used for area 

percent analysis of the bulk chemical. Gas chromatography with flame ionization detection using system B 

(14-week studies) or system C (2-year studies) was used to determine percent purity relative to reference 

material. Additionally, the ethylene oxide content was measured with system B. Impurities in the bulk 

chemical were characterized using gas chromatography with mass spectrometry (system D). 

For lot QP-911021-26D1, elemental analyses for carbon, hydrogen, and oxygen were in agreement with the 

theoretical values for 2-butoxyethanol. Karl Fischer water analysis indicated 0.02% water. Titrations 

indicated 0.001% acidity (as acetic acid) and 105 ppm peroxide, well within the acceptable limits of 0.02% 

acid and 5,000 ppm peroxide set for these studies. Gas chromatography indicated a major peak and one 

impurity with an area of 0.1% of the major peak area; less than 200 ppm ethylene oxide was detected. 

Concurrent analyses were performed on the reference material obtained from Aldrich Chemical Company. 

Results of elemental analyses were similar. The reference sample contained 0.12% water, 0.04% acetic 

acid, 5,537 ppm peroxide, and less than 200 ppm ethylene oxide; no impurities with areas of 0.1% or 

greater relative to the major peak area were detected by gas chromatography. Major peak comparisons of 

lot QP-911021-26D1 relative to the reference sample were performed by gas chromatography with flame 

ionization detection using system C with toluene as an internal standard and with a final temperature of 

265E C; results indicated a purity of 100.8% for lot QP-911021-26D1 relative to the reference material. 

The overall purity of lot QP-911021-26D1 was determined to be greater than 99%. 

For lot QP-921215-26D2, results of elemental analyses for carbon, hydrogen, and oxygen were in 

agreement with the theoretical values for 2-butoxyethanol. Karl Fischer water analysis indicated 0.0254% 

water. Titrations indicated less than 0.003% acetic acid and less than 1,000 ppm peroxide. Gas 

chromatography using system C indicated a major peak and three impurities with areas greater than 0.1% of 
2-Butoxyethanol, NTP TR 484 267 

the major peak area. The ethylene oxide content was below the limit of detection (132 ppm). The 

reference material obtained from Aldrich Chemical Company and lot QP-911021-26D1 were analyzed 

concurrently with lot QP-921215-26D2 by gas chromatography. No impurities with areas of 0.1% or 

greater relative to the major peak area were detected in the reference sample; lot QP-911021-26D1 

contained 0.12% ethylene glycol and approximately 0.03% 2-ethyl-2-hexenal. Major peak comparisons of 

lot QP-921215-26D2 relative to the reference sample were performed by gas chromatography as described 

for lot QP-911021-26D1; results indicated a purity of greater than 99.2% for lot QP-921215-26D2 relative 

to the reference sample. Three impurities with areas greater than 0.1% relative to the major peak area were 

identified in lot QP-921215-26D2; these impurities were tentatively identified with gas chromatography/ 

mass spectrometry using system D as ethylene glycol (0.10%), 2-ethyl-2-hexenal (0.19%), and 

2-ethyl-1-hexanol (0.11%) coeluting with 2-(2-ethoxyethoxy)ethanol. The overall purity of 

lot QP-921215-26D2 was determined to be greater than 99%. 

Accelerated stability studies of the bulk chemical were performed by Midwest Research Institute (MRI, 

Kansas City, MO) on a lot of 2-butoxyethanol not used in the current studies (MRI, 1984). Samples were 

analyzed by gas chromatography with flame ionization detection, a 20% SP2100/0.1% Carbowax 1500 on 

100/120 Supelcoport glass column, a nitrogen carrier gas at a flow rate of 70 mL/minute, and an oven 

temperature of 130E C. n-Decane was added as an internal standard. These studies indicated that 

2-butoxyethanol is stable as a bulk chemical for 2 weeks when stored protected from light at temperatures 

up to 60E C. To ensure stability, the bulk chemical was stored at room temperature during the 14-week 

studies and at approximately 16E C during the 2-year studies in stainless steel containers. Throughout the 

studies, stability was monitored by titrations for acid and peroxide and by gas chromatography with flame 

ionization detection by both area percent and relative purity methods. No degradation of the bulk chemical 

was detected. 

VAPOR GENERATION AND EXPOSURE SYSTEM 

A diagram of the vapor generation and delivery system is shown in Figure H3. 2-Butoxyethanol was held 

in a stainless steel reservoir under a nitrogen blanket. A liquid micrometering pump was used to pump 

2-butoxyethanol into a glass column filled with glass beads and heated by flexible electric heat tape 

encircling the column. Heated nitrogen entered the column from below, vaporized the 2-butoxyethanol, 

and carried it to a condenser column that was used to control generated vapor concentration. 

2-Butoxyethanol that was not vaporized by the incoming heated nitrogen was collected along with any 

lower-boiling impurities in a flask below the glass-bead-filled column. Vapor temperature was monitored at 

the top of the condenser column by a temperature sensor. The total output of the generator was calculated 

from the metered nitrogen flow and the 2-butoxyethanol vapor pressure at the exit temperature. 

To prevent 2-butoxyethanol from condensing while in transport to the exposure room, the Teflon® transport 

line was heated. The vapor was mixed with heated, HEPA- and charcoal-filtered air before entering a short 

vapor distribution manifold. An automatic controller maintained a constant flow in the distribution 

manifold. 

From the distribution manifold, individual temperature-controlled Teflon® delivery lines carried the vapor 

to the exposure chambers. Flow to each chamber was produced by compressed-air pumps located at the 

chamber end of each delivery line. A three-way valve between the distribution line and each chamber 

directed vapor to the exposure chamber exhaust until a stable concentration of 2-butoxyethanol vapor 

collected in the distribution line. At each chamber, the vapor was further diluted with conditioned, filtered 

chamber air to the appropriate 2-butoxyethanol concentration. 
268 2-Butoxyethanol, NTP TR 484 

The study laboratory designed the inhalation exposure chamber (Harford Systems Division of Lab Products, 

Inc., Aberdeen, MD) so that uniform vapor concentrations could be maintained throughout the chamber 

with the catch pans in place. The total active mixing volume of each chamber was 1.7 m3

. A small particle 

detector (Type CN, Gardner Associates, Schenectady, NY) was used with and without animals in the 

exposure chambers to ensure that 2-butoxyethanol vapor, and not aerosol, was produced. No particle 

counts above the minimum resolvable level (approximately 200 particles/cm3

) were detected. 

VAPOR CONCENTRATION MONITORING 

Chamber concentrations of 2-butoxyethanol were monitored with on-line gas chromatography/flame 

ionization detection using system E (14-week studies) or system F (2-year studies) (Table H1). A 12-port 

stream select valve coupled the on-line gas chromatograph and the exposure chambers. Each chamber was 

sampled approximately every 16 minutes (14-week studies) or every 30 minutes (2-year studies). The online gas chromatograph was checked daily for drift against an on-line standard of 2-butoxyethanol in 

nitrogen; the standard was prepared with a diffusion tube generator (Model 360, Thermo Electron, 

Hopkinton, MA). The on-line gas chromatograph was calibrated by comparing chamber concentration data 

to data collected from grab samples, which were collected with water-filled bubblers. The volumes of gas 

were sampled at a constant flow rate ensured by a calibrated critical orifice. Grab samples were analyzed 

by an off-line gas chromatograph with flame ionization detection (system G), which was calibrated using 

gravimetrically prepared standards of 2-butoxyethanol in water. No significant impurities were detected 

thus demonstrating the purity of the 2-butoxyethanol vapor in the exposure chambers reflected the purity of 

the starting material. Summaries of the chamber concentrations for the 14-week and 2-year studies are in 

Tables H2 and H3. 

CHAMBER ATMOSPHERE CHARACTERIZATION 

The theoretical value for the time to achieve 90% of the target concentration after the beginning of vapor 

generation (T90) and the time for the chamber concentration to decay to 10% of the target concentration 

after vapor generation was terminated (T10) was approximately 12.5 minutes at a chamber airflow rate of 15 

air changes per hour. Prior to and during the 14-week and 2-year studies, T90 and T10 ranges were 

determined with and without animals. The T90 values ranged from 8 to 12 minutes without animals and from 

9 to 28 minutes with animals. The T10 values ranged from 8 to 14 minutes without animals and from 10 to 

19 minutes with animals. A T90 value of 12 minutes was selected for all studies. 

The uniformity of vapor concentration in the inhalation exposure chambers without animals was evaluated 

before each of the studies began; concentration uniformity with animals present in the chambers was also 

measured once during the 14-week studies and periodically during the 2-year studies. Vapor concentration 

was determined with the on-line gas chromatograph, with the 12-port sampling valve disabled to allow 

continuous monitoring from a single line; samples were taken from several positions in each chamber. 

Chamber concentration uniformity was maintained throughout the 14-week and 2-year studies. 

The persistence of 2-butoxyethanol in the chamber following exposure was determined by monitoring the 

concentration overnight in the 500 ppm chamber in the 14-week studies, the 125 ppm chamber in the 2-year 

rat study, and the 250 ppm chamber in the 2-year mouse study, with and without animals present. In the 

14-week studies, the concentration decreased to 1% of the target concentration within 54 minutes. In the 

2-year rat study, the concentration decreased to 1% of the target concentration within 195 minutes with 

animals present and within 29 minutes without animals. In the 2-year mouse study, the concentration 

decreased to 1% of the target concentration within 36 minutes with animals present and within 26 minutes 

without animals. The increase in decay time in the 2-year rat study was attributed to the presence of rats in 

the chambers and not to residual 2-butoxyethanol in the monitoring system. 
2-Butoxyethanol, NTP TR 484 269 

Before and during all studies, 2-butoxyethanol from the vapor generator reservoir, collection flask, 

distribution line, and chambers was tested for stability with gas chromatography using a 2-butoxyethanol 

standard as a reference. Results indicated that 2-butoxyethanol was stable in the reservoir for up to 21 

days. 2-Butoxyethanol vapor was analyzed for volatile contaminants, peroxide, and acetic acid. 

Before and during the 14-week studies, samples from the 0, 31, and 500 ppm exposure chambers were 

analyzed for ethylene oxide by the on-line gas chromatograph. Gas bag standards of ethylene oxide were 

volumetrically prepared and analyzed; ethylene oxide in the standards was detected at concentrations as low 

as 0.6 ppm. No ethylene oxide was detected in any exposure chamber; the concentration in the exposure 

chambers was estimated to be less than 0.1% (0.7 ppm). One to four impurities with a combined area of 

less than 0.03% (prestudy testing) or 0.1% relative to the major peak area were detected in the 500 ppm 

chamber by the on-line gas chromatograph. No impurities were found in the 31 ppm chamber. Samples 

were also collected from the 0, 31, and 500 ppm chambers and the vapor distribution line in solid-phase 

adsorbent glass sampling tubes. Each tube consisted of a front and a rear portion packed with adsorbent 

(ORBO Supelpak 20F). Any 2-butoxyethanol that escaped from the front portion was collected in the rear 

portion; samples were desorbed in 1 mL methanol before analysis using system A. Percent breakthrough 

was calculated as the ratio of the amount collected on the rear portion to the amount collected on the front 

portion. A breakthrough of 10% or less (prestudy testing) or 35% or less was observed for all chamber 

samples. Ethylene glycol was present as an impurity at 2% or less by weight; additional impurities were 

also observed with areas of up to 0.1% of the major peak area. These impurities were observed at similar 

concentrations in the bulk chemical. Samples were also collected from the 0, 31, and 500 ppm chambers 

and distribution line in acetonitrile-filled bubblers and analyzed for impurities and degradation products by 

gas chromatography/mass spectrometry using system D with an initial temperature of 50E C. These 

samples were analyzed against gravimetrically prepared standards of ethylene glycol, 1-butanol, 

2-methyl-2-propanol (prestudy testing only), diethyleneglycol, 2-(2-ethoxy ethoxy) ethanol (prestudy testing 

only), and 2-(2-butoxy ethoxy) ethanol (14-week studies only) in acetonitrile. Samples of the bulk chemical 

were analyzed concomitantly. Results indicated that 1-butanol, diethylene glycol, 2-(2-ethoxy ethoxy) 

ethanol, and 2-(2-butoxy ethoxy) ethanol concentrations were less than 0.2%; ethylene glycol was detected 

in all samples collected during the studies. Additional impurities, tentatively identified as a branched 

alkane, 2-ethyl-2-hexenal, 2-ethyl-1-hexenol, and 2-ethyl-2-hexen-1-ol, were detected, and the 

concentrations were estimated to be less than 0.2% in one or more samples during prestudy testing and 

during the 14-week studies. 

Before the 2-year studies, samples from the reservoir, condensation flask, distribution line, and exposure 

chambers were collected with acetonitrile-filled glass bubblers and analyzed by gas chromatography/mass 

spectrometry using system D for identity and gas chromatography with flame ionization detection using 

systems B and H for area percent. Samples of the bulk chemical were also analyzed. Additional samples 

collected with solid-phase sampling tubes were extracted with methanol and analyzed by gas 

chromatography with flame ionization detection using system H. Distribution line and exposure chamber 

samples contained impurities which were tentatively identified as 1-(2-methoxyethoxy) butane and 

2-ethyl-2-hexenal. These impurities and ethylene glycol, 2-ethylhexanal, 2-ethyl-1-hexanol, 

2-(2-ethoxyethoxy) ethanol, and 2-(2-butoxyethoxy) ethanol were identified in the reservoir and collection 

flask; 2-ethyl-2-hexen-1-ol was also tentatively identified. The concentrations of impurities which were 

previously identified in the bulk chemical did not increase from that determined in the bulk chemical. The 

concentration of 2-ethyl-2-hexenal was approximately 0.13% in the collection flask and approximately 

0.25% in all other samples. Ethylene glycol was also detected in the collection flask at a concentration of 

0.21% and in the reservoir at a concentration of 0.13%. The concentration of 2-ethyl-1-hexanol in the 

collection flask was 0.16%. The concentrations of all other impurities were 0.1% or less. During the 

2-year studies, bubbler samples and sorbent tube samples were collected from the exposure system and 

analyzed by gas chromatography with flame ionization detection using systems C (for bubblers, with 

modified temperature program) and H (for sorbent tube samples). The most concentrated impurity was 
270 2-Butoxyethanol, NTP TR 484 

2-ethyl-2-hexenal (approximately 0.25%); n-butanol, ethylene glycol, 2-ethylhexanal, 2-ethyl-1-hexanol, 

2-(2-ethoxyethoxy) ethanol, and 1-(2-methoxyethoxy) butane were detected at concentrations of 0.1% or 

less. The concentrations of peroxide, ethylene oxide, and acetic acid in the reservoir and collection flask 

were analyzed before and during the 2-year studies as described for the bulk chemical analyses; 

concentrations were well below the maximum allowable concentrations. Ethylene oxide concentrations 

were also measured in samples from the distribution line and exposure chambers. Samples were collected 

with adsorbent tubes (ORBO-78, Supelco) containing a hydrogen bromide-impregnated sorbent that derived 

2-bromoethanol from ethylene oxide. The primary and secondary beds of the sampling tube were mixed 

with sodium carbonate and extracted with a 50:50 solution of acetonitrile and toluene. Gas bag standards 

were prepared with ethylene oxide in methylene chloride. The samples were analyzed for 2-bromoethanol 

by gas chromatography with electron capture detection (system I). Ethylene oxide concentrations were less 

than 0.06% in the exposure chambers and distribution line. In all studies, the concentration of each 

impurity, relative to 2-butoxyethanol, detected in all distribution line and exposure chamber samples was 

the same as that detected in the reservoir, indicating that the impurities were neither concentrated nor 

dispersed by the generation system. 
2-Butoxyethanol, NTP TR 484 271 

FIGURE H1 

Growth Curves for Male and Female Mice 

Exposed to 2-Butoxyethanol by Inhalation for 2 Years 
272 2-Butoxyethanol, NTP TR 484
 

FIGURE H2 

Nuclear Magnetic Resonance Spectrum of 2-Butoxyethanol 
2-Butoxyethanol, NTP TR 484 273 

TABLE H1 

Gas Chromatography Systems Used in the 14-Week and 2-Year Inhalation Studies of 2-Butoxyethanola 

Detection Oven 

System Column Carrier Gas Temperature Program 

System A 

Flame ionization DB WAX, 30 m × 0.53 mm, 

1 μm film (J&W Scientific, 

Folsom, CA) 

System B 

Flame ionization DB-1701, 60 m × 0.32 mm, 

1 μm film (J&W Scientific) 

System C 

Flame ionization DB-1701, 60 m × 0.25 mm, 

1 μm film (J&W Scientific) 

System D 

Mass spectrometry with Rtx-1701, 30 m × 0.25 mm, 

electron impact ionization 1 μm film (Restek, Bellefonte, 

(29 to 200 amu) PA) 

System E 

Flame ionization DB-1701, 15 m × 0.53 mm, 

1 μm film (J&W Scientific) 

System F 

Flame ionization Rtx-200, 30 m × 0.53 mm, 

3 μm film (Restek) 

System G 

Flame ionization DB WAX, 15 m × 0.53 mm, 

1 μm film (J&W Scientific) 

System H 

Flame ionization Rtx-200, 30 m × 0.53 mm, 

3 μm film (Restek) 

System I 

Electron capture DB WAX, 30 m × 0.53 mm, 

1 μm film (J&W Scientific) 

Helium at 11 mL/minute 

Helium at 1.1 mL/minute 

Helium at 2.4 mL/minute 

Helium at 6 psi head pressure 

Nitrogen at 20 mL/minute 

Nitrogen at 50 mL/minute 

Helium at 15 mL/minute 

Helium at 6 psi head pressure 

Nitrogen at 9 mL/minute 

30E C for 5 minutes, then 

5E C/minute to 130E C, then 

10E C/minute to 230E C, held 

for 5 minutes 

60E C for 4 minutes, then 

12E C/minute to 250E C 

60E C for 4 minutes, then 

12E C/minute to 265E C, held 

for 5 minutes 

80E C for 5 minutes, then 

5E C/minute to 250E C, held 

for 1 minute (cool-on-column 

injection) 

Isothermal at 110E C 

Isothermal at 125E C 

95E C for 0.5 minutes, then 

20E C/minute to 165E C 

(14-week studies) or 

20E C/minute to 180E C 

(2-year studies) 

55E C for 0.5 minutes, then 

5E C/minute to 100E C, held 

for 0.1 minutes, then 

20E C/ minute to 250E C, held 

for 2 minutes (cool-on-column 

injection) 

Isothermal at 150E C 

a All gas chromatographs were manufactured by Hewlett-Packard (Palo Alto, CA). 
274 2-Butoxyethanol, NTP TR 484 

FIGURE H3 

Schematic of the Vapor Generation and Delivery System 

in the 14-Week and 2-Year Inhalation Studies of 2-Butoxyethanol 
2-Butoxyethanol, NTP TR 484 275 

TABLE H2 

Summary of Chamber Concentrations in the 14-Week Inhalation Studies of 2-Butoxyethanol 

Target Concentration 

(ppm) 

Total Number of Readings Average Concentrationa 

(ppm) 

Rat Chambers 

31 

62.5 

125 

250 

500 

1,387 

1,386 

1,385 

1,388 

1,413 

31.0 ± 1.2 

62.1 ± 1.8 

125 ± 5.9 

249 ± 11 

497 ± 19 

Mouse Chambers 

31 

62.5 

125 

250 

500 

1,431 

1,430 

1,429 

1,432 

1,457 

31.0 ± 1.2 

62.1 ± 1.8 

125 ± 5.8 

249 ± 11 

497 ± 19 

a Mean ± standard deviation 

TABLE H3 

Summary of Chamber Concentrations in the 2-Year Inhalation Studies of 2-Butoxyethanol 

Target Concentration 

(ppm) 

Total Number of Readings Average Concentrationa 

(ppm) 

Rat Chambers 

31.2 

62.5 

125 

5,717 

5,728 

5,399 

30.8 ± 1.8 

62.3 ± 4.7 

125.0 ± 7.8 

Mouse Chambers 

62.5 

125 

250 

5,403 

5,402 

5,362 

62.7 ± 3.7 

125 ± 8.1 

248 ± 18.7 

a Mean ± standard deviation 
276 2-Butoxyethanol, NTP TR 484 
277 

APPENDIX I 

INGREDIENTS, NUTRIENT COMPOSITION, 

AND CONTAMINANT LEVELS 

IN NIH-07 RAT AND MOUSE RATION 

TABLE I1 

TABLE I2 

TABLE I3 

TABLE I4 

Ingredients of NIH-07 Rat and Mouse Ration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 

Vitamins and Minerals in NIH-07 Rat and Mouse Ration . . . . . . . . . . . . . . . . . . . . . 

Nutrient Composition of NIH-07 Rat and Mouse Ration . . . . . . . . . . . . . . . . . . . . . . 

Contaminant Levels in NIH-07 Rat and Mouse Ration . . . . . . . . . . . . . . . . . . . . . . . 

278 

278 

279 

280 
 

 

278 2-Butoxyethanol, NTP TR 484 

TABLE I1 

Ingredients of NIH-07 Rat and Mouse Rationa 

Ingredientsb Percent by Weight 

Ground #2 yellow shelled corn 24.50 

Ground hard winter wheat 23.00 

Soybean meal (49% protein) 12.00 

Fish meal (60% protein) 10.00 

Wheat middlings 10.00 

Dried skim milk 5.00 

Alfalfa meal (dehydrated, 17% protein) 4.00 

Corn gluten meal (60% protein) 3.00 

Soy oil 2.50 

Dried brewer’s yeast 2.00 

Dry molasses 1.50 

Dicalcium phosphate 1.25 

Ground limestone 0.50 

Salt 0.50 

Premixes (vitamin and mineral) 0.25 

a NCI, 1976; NIH, 1978 b Ingredients were ground to pass through a U.S. Standard Screen No. 16 before being mixed. 

TABLE I2 

Vitamins and Minerals in NIH-07 Rat and Mouse Rationa

 Amount Source 

Vitamins

 A 5,500,000 IU Stabilized vitamin A palmitate or acetate

 D3 4,600,000 IU D-activated animal sterol

 K3 2.8 g Menadione 

d-"-Tocopheryl acetate 20,000 IU

 Choline 560.0 g Choline chloride

 Folic acid 2.2 g

 Niacin 30.0 g 

d-Pantothenic acid 18.0 g d-Calcium pantothenate

 Riboflavin 3.4 g

 Thiamine 10.0 g Thiamine mononitrate

 B12 4,000 μg

 Pyridoxine 1.7 g Pyridoxine hydrochloride

 Biotin 140.0 mg d-Biotin 

Minerals

 Iron 120.0 g Iron sulfate

 Manganese 60.0 g Manganous oxide

 Zinc 16.0 g Zinc oxide

 Copper 4.0 g Copper sulfate

 Iodine 1.4 g Calcium iodate

 Cobalt 0.4 g Cobalt carbonate 

a Per ton (2,000 lb) of finished product 
 

2-Butoxyethanol, NTP TR 484 279 

TABLE I3 

Nutrient Composition of NIH-07 Rat and Mouse Ration 

Mean ± Standard 

Nutrient Deviation Range Number of Samples

 Protein (% by weight) 22.95 ± 0.49 22.1 ) 23.6 23

 Crude fat (% by weight) 5.39 ± 0.19 5.00 ) 5.80 23

 Crude fiber (% by weight) 3.12 ± 0.37 2.60 ) 4.30 23

 Ash (% by weight) 6.24 ± 0.15 5.72 ) 6.54 23 

Amino Acids (% of total diet)

 Arginine 1.273 ± 0.083 1.100 ) 1.390 12

 Cystine 0.307 ± 0.068 0.181 ) 0.400 12

 Glycine 1.152 ± 0.051 1.060 ) 1.220 12

 Histidine 0.581 ± 0.029 0.531 ) 0.630 12

 Isoleucine 0.913 ± 0.034 0.867 ) 0.965 12

 Leucine 1.969 ± 0.053 1.850 ) 2.040 12

 Lysine 1.269 ± 0.050 1.200 ) 1.370 12

 Methionine 0.436 ± 0.104 0.306 ) 0.699 12

 Phenylalanine 0.999 ± 0.114 0.665 ) 1.110 12

 Threonine 0.899 ± 0.059 0.824 ) 0.985 12

 Tryptophan 0.216 ± 0.146 0.107 ) 0.671 12

 Tyrosine 0.690 ± 0.091 0.564 ) 0.794 12

 Valine 1.079 ± 0.057 0.962 ) 1.170 12 

Essential Fatty Acids (% of total diet)

 Linoleic 2.389 ± 0.223 1.830 ) 2.570 11

 Linolenic 0.273 ± 0.034 0.210 ) 0.320 11 

Vitamins

 Vitamin A (IU/kg) 6,657 ± 338 5,500 ) 7,260 23

 Vitamin D (IU/kg) 4,450 ± 1,382 3,000 ) 6,300 4 

"-Tocopherol (ppm) 35.24 ± 8.58 22.5 ) 48.9 12

 Thiamine (ppm) 17.54 ± 3.70 14.0 ) 26.0 22

 Riboflavin (ppm) 7.78 ± 0.899 6.10 ) 9.00 12

 Niacin (ppm) 98.73 ± 23.21 65.0 ) 150.0 12

 Pantothenic acid (ppm) 32.94 ± 8.92 23.0 ) 59.2 12

 Pyridoxine (ppm) 9.28 ± 2.49 5.60 ) 14.0 12

 Folic acid (ppm) 2.56 ± 0.70 1.80 ) 3.70 12

 Biotin (ppm) 0.265 ± 0.046 0.190 ) 0.354 12

 Vitamin B12 41.6 ± 18.6 10.6 ) 65.0 12

 Choline (ppm) 2,955 ± 382 2,300 ) 3,430 11 

Minerals

 Calcium (%) 1.16 ± 0.06 1.03 ) 1.27 23

 Phosphorus (%) 0.89 ± 0.03 0.84 ) 0.97 23

 Potassium (%) 0.886 ± 0.059 0.772 ) 0.971 10

 Chloride (%) 0.531 ± 0.082 0.380 ) 0.635 10

 Sodium (%) 0.316 ± 0.031 0.258 ) 0.370 12

 Magnesium (%) 0.165 ± 0.010 0.148 ) 0.180 12

 Sulfur (%) 0.266 ± 0.060 0.208 ) 0.420 11

 Iron (ppm) 348.0 ± 83.7 255.0 ) 523.0 12

 Manganese (ppm) 93.27 ± 5.62 81.7 ) 102.0 12

 Zinc (ppm) 59.42 ± 9.73 46.1 ) 81.6 12

 Copper (ppm) 11.63 ± 2.46 8.09 ) 15.4 12

 Iodine (ppm) 3.49 ± 1.14 1.52 ) 5.83 11

 Chromium (ppm) 1.57 ± 0.53 0.60 ) 2.09 12

 Cobalt (ppm) 0.81 ± 0.27 0.49 ) 1.23 8 
 

 

 

 

 

 

 

 

 

280 2-Butoxyethanol, NTP TR 484 

TABLE I4 

Contaminant Levels in NIH-07 Rat and Mouse Rationa 

Mean ± Standard 

Deviationb Range Number of Samples 

Contaminants

 Arsenic (ppm) 0.53 ± 0.17 0.10 ) 0.80 23

 Cadmium (ppm) 0.05 ± 0.02 0.04 ) 0.13 23

 Lead (ppm) 0.23 ± 0.06 0.20 ) 0.40 23

 Mercury (ppm)c <0.02 23

 Selenium (ppm) 0.34 ± 0.10 0.10 ) 0.50 23

 Aflatoxins (ppb) <5.0 23

 Nitrate nitrogen (ppm)d 7.28 ± 2.45 2.90 ) 11.0 23

 Nitrite nitrogen (ppm)d 1.45 ± 0.91 0.30 ) 3.50 23

 BHA (ppm)e 1.07 ± 0.95 0.01 ) 5.0 23

 BHT (ppm)e 1.73 ± 1.17 0.18 ) 5.0 23

 Aerobic plate count (CFU/g) 128,000 ± 127,186 11,000 ) 460,000 23

 Coliform (MPN/g) 155 ± 582 3 ) 2,800 23 

Escherichia coli (MPN/g) 7 ± 3.5 3 ) 10 23 

Salmonella (MPN/g) Negative 23

 Total nitrosoamines (ppb)f 12.20 ± 3.90 4.0 ) 23.0 23 

N-Nitrosodimethylamine (ppb)f 10.55 ± 3.64 3.0 ) 21.0 23 

N-Nitrosopyrrolidine (ppb)f 1.66 ± 0.56 1.0 ) 2.9 23 

Pesticides (ppm) 

"-BHC <0.01 23 

$-BHCe <0.02 23 

(-BHC <0.01 23 

*-BHC <0.01 23

 Heptachlor <0.01 23

 Aldrin <0.01 23

 Heptachlor epoxide <0.01 23

 DDE <0.01 23

 DDD <0.01 23

 DDT <0.01 23

 HCB <0.01 23

 Mirex <0.01 23

 Methoxychlor <0.05 23

 Dieldrin <0.01 23

 Endrin <0.01 23

 Telodrin <0.01 23

 Chlordane <0.05 23

 Toxaphene <0.10 23

 Estimated PCBs <0.20 23

 Ronnel <0.01 23

 Ethion <0.02 23

 Trithion <0.05 23

 Diazinon <0.10 23

 Methyl parathion <0.02 23

 Ethyl parathion <0.02 23

 Malathion 0.13 ± 0.17 0.02 ) 0.83 23

 Endosulfan I <0.01 23

 Endosulfan II <0.01 23

 Endosulfan sulfate <0.03 23 

a CFU=colony-forming units; MPN=most probable number; BHC=hexachlorocyclohexane or benzene hexachloride b For values less than the limit of detection, the detection limit is given as the mean. 

All values were less than the detection limit. The detection limit is given as the mean. d Sources of contamination: alfalfa, grains, and fish meal e Sources of contamination: soy oil and fish meal f All values were corrected for percent recovery. 

c 
281 

APPENDIX J
 

SENTINEL ANIMAL PROGRAM
 

METHODS .............................................................. 282 
 

RESULTS ............................................................... 284 
 
 

 

282 2-Butoxyethanol, NTP TR 484 

SENTINEL ANIMAL PROGRAM
 

METHODS 

Rodents used in the Carcinogenesis Program of the National Toxicology Program are produced in optimally 

clean facilities to eliminate potential pathogens that may affect study results. The Sentinel Animal Program 

is part of the periodic monitoring of animal health that occurs during the toxicologic evaluation of chemical 

compounds. Under this program, the disease state of the rodents is monitored via serology on sera from 

extra (sentinel) animals in the study rooms. These animals and the study animals are subject to identical 

environmental conditions. The sentinel animals come from the same production source and weanling 

groups as the animals used for the studies of chemical compounds. 

Serum samples were collected from randomly selected rats and mice during the 14-week and 2-year studies. 

Blood from each animal was collected and allowed to clot, and the serum was separated. The samples were 

processed appropriately and sent to Microbiological Associates, Inc. (Bethesda, MD), for determination of 

antibody titers. The laboratory serology methods and viral agents for which testing was performed are 

tabulated below; the times at which blood was collected during the studies are also listed. 

Method and Test Time of Analysis 

RATS 

14-Week Study 

ELISA

 PVM (pneumonia virus of mice) 

 RCV/SDA (rat coronavirus/

 sialodacryoadenitis virus) 

Sendai 

Study termination

Study termination

Study termination 

Hemagglutination Inhibition

 H-1 (Toolan’s H-1 virus) 

 KRV (Kilham rat virus) 

Study termination

Study termination 

2-Year Study 

ELISA 

Mycoplasma arthritidis 

Mycoplasma pulmonis 

 PVM 

 RCV/SDA 

Sendai 

Study termination 

Study termination

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination 

Immunofluorescence Assay

 M. arthritidis Study termination 

Hemagglutination Inhibition

 H-1 

KRV 

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination 
2-Butoxyethanol, NTP TR 484 283 

Method and Test 

MICE 

14-Week Study 

ELISA

 Ectromelia virus 

 EDIM (epizootic diarrhea of infant mice) 

 GDVII (mouse encephalomyelitis virus) 

 LCM (lymphocytic choriomeningitis virus) 

 MHV (mouse hepatitis virus) 

 Mouse adenoma virus-FL 

 PVM 

 Reovirus 3 

Sendai 

Immunofluorescence Assay

 EDIM 

Hemagglutination Inhibition

 K (papovavirus) 

 MVM (minute virus of mice) 

 Polyoma virus 

2-Year Study 

ELISA

 Ectromelia virus 

EDIM 

 GDVII 

LCM 

 Mouse adenoma virus-FL 

MHV 

M. pulmonis 

 PVM
 

 Reovirus 3
 

Sendai
 

Immunofluorescence Assay

 Ectromelia virus 

EDIM 

 Mouse adenoma virus-FL 

M. arthritidis 

 PVM 

Sendai
 

 MCMV (mouse cytomegalovirus)
 

Hemagglutination Inhibition

 K 

 MVM 

 Polyoma virus 

Time of Analysis
 

Study termination

Study termination

Study termination

Study termination

Study termination

Study termination

Study termination

Study termination

Study termination 

Study termination 

Study termination

Study termination

Study termination 

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination

Study termination

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination 

6 months

Study termination

12 months and study termination

Study termination

12 months

Study termination

Study termination 

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination

6, 12, and 18 months, study termination
 
284 2-Butoxyethanol, NTP TR 484 

RESULTS 

All test results were negative. 
285 

APPENDIX K
 

H-RAS CODON 61 MUTATION SPECTRA IN
 

FORESTOMACH NEOPLASMS FROM B6C3F1 MICE
 

EXPOSED TO 2-BUTOXYETHANOL 

FOR 2 YEARS
 

Robert C. Sills, T. Ton, H. Lily Hong, Joe Roycroft,
 

and Theodora R. Devereux
 

National Institute of Environmental Health Sciences
 

Research Triangle Park, North Carolina
 

INTRODUCTION ........................................................... 286 

MATERIALS AND METHODS ................................................... 286 

RESULTS ............................................................... 287 

DISCUSSION ............................................................. 288 

REFERENCES ............................................................ 289 

TABLE K1 Patterns of H-ras Mutations in Forestomach Neoplasms from B6C3F1 Mice ........ 290 
286 2-Butoxyethanol, NTP TR 484 

H-RAS CODON 61 MUTATION SPECTRA 

IN FORESTOMACH NEOPLASMS FROM B6C3F1 MICE 

EXPOSED TO 2-BUTOXYETHANOL FOR 2 YEARS 

INTRODUCTION 

Various routes of chemical administration including oral, gavage, and inhalation may induce high 

incidences of forestomach neoplasms in rodent carcinogenicity bioassays. The mechanism of many of these 

forestomach carcinogens is unknown. The objective of this research is to investigate the effects of 

inhalation exposure in contributing to genetic alterations in cancer related genes, and to examine the factors 

underlying susceptibility of the forestomach to tumorigenesis. To date, there are no published data on the 

mutation spectrum and frequency of ras mutations in both spontaneous and chemical-induced forestomach 

neoplasms of B6C3F1 mice. 

Forestomach neoplasms occur in B6C3F1 mice with a typical incidence of 0% to 2% in chamber control 

males and 0% to 3% in chamber control females in 2-year inhalation studies. Molecular analysis of rodent 

neoplasms for genetic alterations in cancer genes, such as the ras proto-oncogene, provides additional 

mechanistic information to help distinguish spontaneous neoplasms from chemical-induced neoplasms. For 

example, chemical-induced neoplasms in mice may have a high frequency of proto-oncogene activation, 

particularly by point mutations in codons 12, 13, and 61 of H- or K-ras genes (Devereux et al., 1991; Sills 

et al., 1995; Hong et al., 1997). The frequency of ras activation in these neoplasms is often greater than 

that detected in neoplasms occurring in control animals, and there is evidence for chemical specificity in the 

pattern of mutations. The specific types of oncogene-activating mutations induced by a chemical carcinogen 

often agree with what is expected based on the DNA adducts formed by the agent. Even for “nongenotoxic 

carcinogens,” patterns of ras gene mutations in neoplasms can give clues about the mechanism of 

tumorigenesis (Devereux et al., 1993; Maronpot et al., 1995). 

MATERIALS AND METHODS 

Forestomach Neoplasms: Male and female B6C3F1 mice were exposed to 0, 62.5, 125, or 250 ppm 

2-butoxyethanol by inhalation for 6 hours per day, 5 days per week for 2 years. At necropsy, forestomach 

neoplasms were fixed in 10% neutral buffered formalin, routinely processed, embedded in paraffin, 

sectioned to a thickness of 5 μm, and stained with hematoxylin and eosin. Subsequently, six unstained 

serial sections (10 μm thick) were prepared from paraffin blocks containing forestomach papillomas or 

carcinomas for isolation of DNA for polymerase chain reaction (PCR)-based assays. In order to isolate 

adequate amounts of DNA, forestomach neoplasms greater that 1 mm in diameter were identified for 

analysis. A total of 25 paraffin-embedded neoplasms was examined for genetic alterations in the H- and 

K-ras genes. This included 14 neoplasms from 2-butoxyethanol-exposed mice and 11 neoplasms from 

control mice from various NTP studies. Also, 14 forestomach neoplasms from mice exposed to 

1,3-butadiene were evaluated for H- and K-ras mutations. 

DNA Isolation: The DNA isolation procedure is described by Marmur (1961) and Sills et al. (1995). The 

paraffin-embedded tissue was deparaffinized and rehydrated before digestion with proteinase K (Wright and 

Manos, 1990). DNA was extracted with phenol and chloroform and precipitated with ethanol. DNA was 

quantified by optical density at 260 nm and 200 ng/μL was used for amplification. 

DNA Amplification: DNA was amplified by PCR (Saiki et al., 1988; Sills et al., 1995); details of the use 

of nested primers are described by Devereux et al. (1991, 1993). 
2-Butoxyethanol, NTP TR 484 287 

Restriction Fragment Length Polymorphic Identification: For identification of H-ras mutations at codon 61, 

restriction fragment length polymorphism (RFLP) was used, and most of exon 2 surrounding codon 61 was 

amplified (Sukumuar and Barbacid, 1990). The sense primer used for amplification of exon 2 was 

5!-GACATCTTAGACACAGCAGTT-3!. A restriction site for MSE I, XbaI, or TaqI enzyme (New 

England Biolaboratory, Beverly, MA) is created by the presence of a C to A, A to T, or A to G mutation, 

respectively, in the first or second base of codon 61. By using this technique, one can detect codon 61 

AAA, CTA, and CGA mutations by MSEI, XbaI and TaqI digestion, respectively; the normal sequence 

(CAA) of codon 61 is not cut by these enzymes. The reaction was incubated at 37E C (for MSEI or XbaI) 

or 60E C (for Taq I) for 2 hours. Fifteen μL of the mixture with bromophenol blue dye was loaded onto 

the 6% acrylamide TBE gel (8 × 8 cm × 1 mm; 15 wells) (Novex, San Diego, CA). The gel was run at 

100 volts for 1 hour on the Novex gel electrophoresis unit. Gels were stained with a 5 μg/mL solution of 

ethidium bromide for 20 minutes and then destained in distilled water. Ethidium bromide-stained bands 

were visualized using a 312 nm ultraviolet viewing box and were photographed. 

Single-Strand Conformation Polymorphism Analysis (SSCP): Single-strand conformation analysis (Orita 

et al., 1989) was performed with PCR products into which ["-

33P]dATP was incorporated during the inner 

amplification. For the first exon of K-ras, 10% acrylamide gel containing 10% glycerol and 1X tris-borateEDTA buffer was electrophoresed at room temperature with constant power at 8 watts for 16 hours on a 

Model S2 sequencing gel electrophoresis apparatus (Bethesda Research Labs, Gaithersburg, MD). For the 

second exon of H-ras, 12% acrylamide gel with 5% glycerol in 1X tris-borate-EDTA buffer was used at 

35 watts in a 4E C cold room for 5 hours. 

Direct Sequencing: Direct sequencing of the amplified second exon of the K-ras gene was performed as 

described by Tindall and Stankowski (1989) using previously described sequencing primers (Devereux 

et al., 1991). 

RESULTS 

In order to determine if the 2-butoxyethanol-induced neoplasms contained an H-ras mutation profile similar 

to that observed with “spontaneous” neoplasms, sample groups of 12 and two neoplasms consisting of 

adenomas and carcinomas respectively, from various exposure groups, and 11 spontaneous neoplasms from 

the chamber controls (various studies), were evaluated by PCR amplification of H-ras exon 2 followed by 

RFLP analysis for codon 61 mutations in the B6C3F1 mouse (Table K1). SSCP was used as an alternative 

screening method for detection of H- or K-ras mutations in DNA and mutations were confirmed by direct 

sequencing. A similar frequency (57%, 8/14) of H-ras mutations was detected in forestomach neoplasms 

when compared to the frequency (45%, 5/11) detected in spontaneous forestomach neoplasms from B6C3F1 

mice (Table K1). In addition, the primary H-ras mutation in forestomach neoplasms from the 

2-butoxyethanol study and spontaneous neoplasms was a CGA mutation. There were no differences in the 

mutation frequency and spectrum between exposure groups (Table K1) or between benign and malignant 

forestomach neoplasms (data not shown). H-ras exon 1 or K-ras exon 1 and 2 mutations were not detected 

in forestomach neoplasms following exposure to 2-butoxyethanol. 

Compared to the 2-butoxyethanol study, the mutation frequency detected in 1,3-butadiene forestomach 

neoplasms was 38% (9/24) (Table K1), which included 6 chemical-specific CTA codon 61 H-ras mutations, 

and three spontaneous CGA mutations. 
288 2-Butoxyethanol, NTP TR 484 

DISCUSSION 

In order to gain insight into the mechanism of 2-butoxyethanol induced forestomach carcinogenesis, H-ras 

oncogene mutation spectra in forestomach neoplasms from male and female B6C3F1 mice exposed to 

2-butoxyethanol for 2 years were compared. Codon 61 mutations in the H-ras gene were detected in 57% 

of the forestomach neoplasms induced by exposure to 2-butoxyethanol and 45% of the spontaneous 

neoplasms examined. In addition, the mutation profile within these chemical-induced forestomach 

neoplasms did not differ significantly from that in spontaneous neoplasms. This is the first study which 

demonstrates that the formation of both chemical-induced and spontaneous forestomach neoplasms in 

B6C3F1 mice is associated with activation of the H-ras gene. The high frequency of activated H-ras genes 

detected in spontaneous forestomach neoplasms suggests that this gene is important in neoplasm formation 

in the B6C3F1 mouse. 

Results from this study suggests that the activation of the H-ras gene in the 2-butoxyethanol-induced 

forestomach neoplasms is not directly related to chemical exposure. Our findings indicate that 

2-butoxyethanol may act in the forestomach to promote clonal growth of forestomach cells which were 

initiated spontaneously and which primarily contain an activated H-ras gene. 

The similar mutation frequency and spectra of H-ras mutations detected in 2-butoxyethanol and spontaneous 

forestomach neoplasms is consistent with that seen with nongenotoxic agents in other organ systems 

(Devereux et al., 1993; Maronpot et al., 1995). Nongenotoxic agents by definition are not positive in 

in vitro assays and, therefore, the parent chemical or metabolites do not react with the DNA directly to 

cause mutations (Green, 1991). The findings of a similar frequency of ras mutations at codon 61 are 

consistent with the lack of mutagenicity in the Salmonella typhimurium assay and are consistent with the 

hypothesis that 2-butoxyethanol may be acting as a nongenotoxic carcinogen. In addition, the finding of no 

increase in the frequency of micronucleated erythrocytes in peripheral blood samples supports this theory. 

Unlike the 2-butoxyethanol forestomach study, the finding of six H-ras codon 61 CTA mutations in 

1,3-butadiene forestomach neoplasms and no CTA mutations in spontaneous forestomach neoplasms suggest 

that this is a chemical-specific mutation, consistent with the genotoxic properties of 1,3-butadiene (Melnick 

and Huff, 1992; Melnick and Kohn, 1995). 
 

2-Butoxyethanol, NTP TR 484 289 

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290 2-Butoxyethanol, NTP TR 484 

TABLE K1 

Patterns of H-ras Mutations in Forestomach Neoplasms from B6C3F1 Mice 

Codon 61 

Activated (Normal=CAA) 

Treatment H-ras (%) CTA CGA 

Controla 5/11 (45) 0 5 

2-Butoxyethanolb 8/14 (57) 0 8 

1,3-Butadienec 9/24 (38) 6 3 

a Study controls combined with historical spontaneous forestomach neoplasms of control B6C3F1 mice 

b	 Male and female B6C3F1 mice were exposed to 62.5, 125, or 250 ppm 2-butoxyethanol by inhalation for 6 hours per day, 5 days per 

week for 2 years. c Male and female B6C3F1 mice were exposed to 6.25 to 625 ppm 1,3-butadiene by inhalation for 2 years.