National Cancer Institute 

CARCINOGENESIS 

Technical Report Series 

No. 40 

1978 

BIOASSAY OF 

HEXACHLOROPHENE 

FOR POSSIBLE CARCINOGENICITY 

CAS No. 70-30-4 

NCI-CG-TR-40 

U.S. DEPARTMENT OF HEALTH, EDUCATION, AND WELFARE 

Public Health Service 

National Institutes of Health 

Division of Cancer Cause and Prevention 

BIOASSAY OF 

HEXACHLOROPHENE 

FOR POSSIBLE CARCINOGENICITY 

Carcinogenesis Testing Program 

National Cancer Institute 

National Institutes of Health 

Bethesda, Maryland 20014 

U.S. DEPARTMENT OF HEALTH, EDUCATION, AND WELFARE

Public Health Service 

National Institutes of Health 

DHEW Publication No. (NIH) 78-840 

BIOASSAY OF 

HEXACHLOROPHENE 

FOR POSSIBLE CARCINOGENICITY 

Carcinogenesis Testing Program

Division of Cancer Cause and Prevention 

National Cancer Institute 

National Institutes of Health 

CONTRIBUTORS: This report presents the results of the bioassay of

hexachlorophene for possible carcinogenicity, conducted for the

Carcinogenesis Testing Program, Division of Cancer Cause and

Prevention, National Cancer Institute (NCI), Bethesda, Maryland.

The bioassay was conducted by Stanford Research Institute, Menlo

Park, California, initially under direct contract to NCI and

currently under a subcontract to Tracor Jitco, Inc., prime

contractor for the NCI Carcinogenesis Testing Program. 

The experimental design and doses were determined by Drs. R. R. 

Bates1

,2, D. C. L. Jones3, D. P. Sasmore3, G. W. Newell3, and R. M.

Elashoff4

, and Mr. W. E. Davis3. The principal investigator was

Dr. D. C. L. Jones; the technical supervisor of animal treatment,

observation, and data handling was Mr. W. E. Davis; necropsy and 

tissue fixation were supervised by Dr. D. P. Sasmore. 

Histopathologic examinations were performed by Dr. H. Elster5

,

and the diagnoses included in this report represent his interpretation. Neoplasms and compound-related hyperplastic lesions

were reviewed by Dr. W. M. Busey5

, who also prepared the

interpretive pathology summary included in this report. 

Animal pathology tables and animal survival tables were compiled

at EG&G Mason Research Institute7

. The statistical analyses were 

performed by Dr. J. R. Joiner8

, using methods selected for the

bioassay program by Dr. J. J. Gart9

. Chemicals used in this 

bioassay were analyzed at Stanford Research Institute, and the

analytical results were reviewed by Dr. C. W. Jameson8

. 

iii 
_________________________________________________________________ 

This report was prepared at Tracor Jitco8

 under the direction of 

NCI. Those responsible for the report at Tracor Jitco were Dr.

Marshall Steinberg, Director of the Bioassay Program; Dr. J. F.

Robens, toxicologist; Dr. R. L. Schueler, pathologist; Ms. L. A.

Waitz and Mr. W. D. Reichardt, bioscience writers; and Dr. E. W.

Gunberg, technical editor, assisted by Ms. Y. E. Presley. 

The statistical analysis was reviewed by one or more members of

the Mathematical Statistics and Applied Mathematics Section of

NCI9

: Dr. John J. Gart, Mr. Jun-mo Nam, Dr. Hugh M. Pettigrew, 

and Dr. Robert E. Tarone. 

The following other scientists at the National Cancer Institute

were responsible for evaluating the bioassay experiment, interpreting the results, and reporting the findings: 

Dr. Kenneth C. Chu 

D. Cipriano Cueto, Jr.

Dr. J. Fielding Douglas 

Dr. Dawn G. Goodman 

Dr. Richard A. Griesemer 

Mr. Harry A. Milman 

Dr. Thomas W. Orme 

Dr. Robert A. Squire 

Dr. Jerrold M. Ward 

1

Carcinogenesis Testing Program, Division of Cancer Cause and

Prevention, National Cancer Institute, National Institutes of

Health, Bethesda, Maryland. 

2

Now with the Office of the Commissioner, Food and Drug

Administration, Rockville, Maryland. 

3

Stanford Research Institute, Menlo Park, California. 

4

Department of Biomathematics, Center for the Health Science,

University of California, Los Angeles, California. 

iv 
5

Department of Pathology, David M. Brotman Memorial Hospital,

3828 Hughes Avenue, Culver City, California. 

6

Experimental Pathology Laboratories, Inc., P. 0. Box 474,

Herndon, Virginia. 

7

EG&G Mason Research Institute, 1530 East Jefferson Street,

Rockville, Maryland. 

8

Tracer Jitco, Inc., 1776 East Jefferson Street, Rockville,

Maryland. 

9

Mathematical Statistics and Applied Mathematics Section,

Biometry Branch, Field Studies and Statistics, Division of

Cancer Cause and Prevention, National Cancer Institute,

National Institutes of Health, Bethesda, Maryland. 

10Now with the Division of Comparative Medicine, Johns Hopkins

University, School of Medicine, Traylor Building, Baltimore,

Maryland. 

v 

 


 

untreated rats of each sex. lled at

n

SUMMARY
 

A bioassay of hexachlorophene for possible carcinogenicity was

conducted by administering the test chemical in feed to Fischer 

344 rats. 

Groups of 24 rats of each sex were administered hexachlorophene

at one of three doses, either 17, 50, or 150 ppm, for 105-106

weeks. Higher doses of 200-600 ppm, used in 8-week subchronic

studies, induced neuronal necrosis of the brain and clinical

signs of toxicity. Matched-control groups consisted of 24 

All surviving animals were ki

105-106 weeks. 

Mean body weights of the rats were unaffected by the hexachloro

phene, and no clinical signs of toxicity were recorded. Survival 

also was unaffected, and adequate numbers of animals survived,

permitting meaningful evaluation of the incidences of lateappearing tumors. 

No tumors were present in a statistically significant incidence

at any site in the treated rats. 

It is concluded that under the conditions of this bioassay,

hexachlorophene did not induce malignant or benign tumors i

Fischer 344 rats. 

­

vii 


 

.......... 37 

TABLE OF CONTENTS 

Page 

I. Introduction ............................................... 1 

II. Materials and Methods ...................................... 3 

A. Chemical ............................................. 3 

B. Dietary Preparation .................................. 3 

C. Animals .............................................. 4 

D. Animal Maintenance ................................... 5 

E. Subchronic Studies ................................... 6 

F. Design of Chronic Studies ............................ 7 

G. Clinical and Pathologic Examinations ................. 7 

H. Data Recording and Statistical Analyses .............. 9 

III. Results ................................................... 15 

A. Body Weights and Clinical Signs ..................... 15 

B. Survival ............................................ 15 

C. Pathology ........................................... 18 

D. Statistical Analyses of Results ..................... 19 

IV. Discussion ................................................ 21 

V. Bibliography .............................................. 23 

APPENDIXES
 

Table Bl Summary of the Incidence of Nonneoplastic Lesions

in Male Rats Fed Hexachlorophenein the Diet 

Appendix A Summary of the Incidence of Neoplasms in 

Rats Fed Hexachlorophene in the Diet ............... . 25 

Table Al Summary of the Incidence of Neoplasms in

Male Rats Fed Hexachlorophene in the Diet..... ....... 27 

Table A2 Summary of the Incidence of Neoplasms in

Female Rats Fed Hexachlorophene in the Diet .......... 30 

Appendix B Summary of the Incidence of Nonneoplastic 

Lesions in Rats Fed Hexachlorophene in the Diet.... .. 35 

ix 
	 

	 

in Rats Fed Hexachlo ene in the Diet.................. 
 

	 

	 

	 

	 

 in the Diet .............................................. 

Page 

Table B2	 Summary of the Incidence of Nonneoplastic 

Lesions in Female Rats Fed Hexachlorophene 

in the Diet............................................. 40 

Appendix C Analyses of the Incidence of Primary Tumors 

roph 45
 

Table Cl	 Analyses of the Incidence of Primary Tumors 

in Male Rats Fed Hexachlorophene in the Diet ............. 47 

Table C2	 Analyses of the Incidence of Primary Tumors 

in Female Rats Fed Hexachlorophene in the Diet .......... 50 

TABLES 

Table 1	 Design of Hexachlorophene Chronic Feeding 

Studies in Rats ......................................... 8 

FIGURES 

Figure 1 Growth Curves for Rats Fed Hexachlorophene 

16 

Figure 2 Survival Curves for Rats Fed Hexachlorophene

 in the Diet ........................................................ 17

 

X 
I. INTRODUCTION 

Hexachlorophene (CAS 70-30-4; NCI C02653) is a chlorinated bisphenol which was widely used as an antiseptic prior to 1972. It is 

highly effective against gram-positive bacteria and many pathogenic 

fungi (Harvey, 1975). The chemical was used as a surgical scrubbing 

agent, in bathing newborns to prevent staphylococcal skin 

infection, and in many over-the-counter drugs such as mouthwashes, 

powders, cosmetics, soaps, and skin cleansing agents. In 1972, the 

Food and Drug Administration restricted the uses of hexachlorophene 

because of neurologic toxicity and deaths that occurred following 

its misuse on infants (FDA, 1972; FDA, 1976). 

Agricultural chemicals containing hexachlorophene are currently 

registered for use on three vegetables and a few ornamental plants 

to control mildew and bacterial spot. Limited industrial and 

household applications are also permitted for disinfectant purposes 

(EPA, 1975). 

This bioassay of hexachlorophene was conducted as a part of a 

larger study that was designed to assess the combined effects of a 

group of chemicals, including known or suspected carcinogens. Only 

the results of the study of the administration of hexachlorophene 

are reported herein. 

1 

 

 

 

 

 


 

II. MATERIALS AND METHODS 

A. Chemical 

Hexachlorophene is the generic name for 2,2'-methylenebis-(3,4,6­

trichlorophenol). The chemical was obtained in two batches (both 

from Lot No. 91387), from the Givaudan Corporation, Clifton, New 

Jersey. The purity of these batches was determined to be 98-100% 

by analyses at Stanford Research Institute. The melting point was 

161-162°C (literature: 161°C), and the elemental analyses (C, H, 

C1) were correct for C13H6Cl602, the molecular formula of 

hexachlorophene. The identity of the chemical was confirmed by 

nuclear magnetic resonance, infrared, and ultraviolet spectra, 

which were in agreement with the structure and matched the 

spectra given in the literature. No attempt was made to identify 

or quantitate impurities. 

The chemical was stored at room temperature in clear glass 

bottles. 

B. Dietary Preparation 

All diets were formulated every 2 weeks using Low Fat Lab Chow® 

(Ralston Purina Co., St. Louis, Mo.). A stock diet was prepared 

by first grinding the chemical to a fine powder and then mixing 

by hand a weighed amount with a small amount of feed. Corn oil 

3
 
 


 

and more feed were then added to give a final concentration of 

2,000 ppm hexachlorophene and 3% corn oil, and mixed in a Hobart 

blender for 30 minutes. Each stock diet was analyzed for content 

of hexachlorophene by a method involving extraction, Florisil® 

chromatography, and quantitation by gas-liquid chromatography. 

Concentrations of 2,000 ppm ± 200 ppm were considered acceptable 

for use in preparing test diets. Hexachlorophene at 2,000 ppm in 

the stock diet was found to be stable when held in rat feeders at 

room temperature for a 2-week period. 

To obtain test diets having appropriate concentrations of 

hexachlorophene, the stock diet was diluted, as required, with 

control diet containing 3% corn oil and mixed in a Hobart blender 

for 7 minutes. The stock and test diets were stored at room 

temperature in covered plastic containers. 

C. Animals 

Male and female Fischer 344 rats, obtained through contracts of 

the Division of Cancer Treatment, National Cancer Institute, were 

used in these bioassays. The rats were obtained from Simonsen 

Laboratory, Gilroy, California. On arrival at the laboratory, all 

animals were quarantined for 2 weeks as an acclimation period. 

Following this period, all males gaining less than 25 grams, all 

females gaining less than 15 grams, and all unhealthy 

4
 
animals were culled. The remaining animals were assigned to 

cages, one per cage, until each cage contained three animals. 

Cages were then numbered and assigned to control and treated 

groups using a computer-generated randomization table. Rats were 

ear-clipped for individual identification. 

D. Animal Maintenance 

All animals were housed in temperature- and humidity-controlled 

rooms. The temperature was maintained at 22°C with a range from 

21-24°C, and the relative humidity was maintained at approximately 45%. The room air was changed 10 times per hour and was 

maintained under positive pressure relative to the access halls. 

Fluorescent lighting provided illumination 12 hours per day. Food 

and water were available ad libitum. Drinking water was softened, 

filtered, sterilized with ultraviolet light, and supplied by 

means of an automatic watering system. 

The rats were housed three per cage in polycarbonate cages 

equipped with disposable polyester woven filter tops. Autoclaved 

hardwood chips (Iso-Dri®-, Becton, Dickinson, and Carworth, 

Warrensburg, N. Y) were used as bedding. The cages were changed, 

washed, and provided with fresh bedding twice per week. Filter 

tops were replaced once per month. 

5 
 


 

Rats fed hexachlorophene were housed in the same room as rats 

treated with aflatoxin B1 (CAS 1162-65-8), N-nitrosodipentylamine 

(CAS 13256-06-9), or Aroclor® 1254 (CAS 27323-18-8). 

E. Subchronic Studies 

Subchronic feeding studies were conducted with male and female 

Fischer 344 rats to estimate the maximum tolerated dose of 

hexachlorophene, on the basis of which low, mid, and high 

concentrations (hereinafter referred to as "low doses", "mid 

doses", and "high doses") were determined for administration in 

the chronic studies. In the subchronic studies, hexachlorophene 

was added to feed in concentrations of 50, 100, 200, 400, or 600 

ppm. Treated and control groups each consisted of 15 male and 15 

female rats. The chemical was provided in feed to the treated 

groups for 8 weeks. 

Animals treated at 600 ppm developed hindquarter paralysis and 

exophthalmos, and had a generally wasted appearance; by the end 

of the study, 12/15 males and 11/15 females at this dose had 

died. Histologically, neuronal necrosis of the brain was found in 

groups treated at 600 ppm, 400 ppm, and, to a lesser degree, 200 

ppm. Body weight gain in animals treated at 400 ppm was 76% of 

that of controls in males and 74% of that of controls in females. 

Weight gain was unaffected at lower doses, and no 

6
 
 


 

deaths occurred at doses of 400 ppm or lower. The low, mid, and 

high doses for the chronic studies were set at 17, 50, and 150 

ppm. 

F. Design of Chronic Studies 

The design of the chronic studies is shown in table 1. 

G. Clinical and Pathologic Examinations 

All animals were observed twice daily for signs of toxicity and 

palpated for masses at each weighing. Animals were weighed 

individually every other week for 12 weeks, and once every fourth 

week for the remainder of the study. Animals that were moribund 

at the time of clinical examination were killed and necropsied. 

The pathologic evaluation consisted of gross examination of major 

organs and tissues from killed animals and from animals found 

dead. The following tissues were routinely examined 

microscopically from both control and treated animals: lungs and 

bronchi, spleen, liver, kidney, pituitary, testis, and brain. In 

addition, esophagus, stomach, urinary bladder, thyroid, uterus, 

and ovary were examined from a majority of the control animals; 

these tissues were examined from treated animals only if a lesion 

was found at necropsy. Occasionally, additional tissues were 

examined microscopically. Gross lesions from all animals were 

7
 
Sex and 

Treatment 

Group

Initial 

No. of 

Animalsa

Hexachlorophene 

in Dietb

ppm

Treatedc 

(weeks) 

MALE 

Matched-Control 24 0 106 

Low-Dose 24 17 105-106 

Mid-Dose 24 50 106 

High-Dose 24 150 106 

FEMALE 

Matched-Control 24 0 105-106 

Low-Dose 24 17 105-106 

Mid-Dose 24 50 106 

High-Dose 24 150 106 

other. 

Table 1. Design of Hexachlorophene Chronic Feeding Studies in Rats 

Time on Study

Untreated 

( ) weeks

a

All animals were approximately 52 ± 3 days of age when

placed on study. 

b

All diets contained 3% corn oil. 

C

A11 animals were started on study within 2 days of each 

8
 
 
 


 

also examined microscopically. The different tissues were preserved 

in 10% buffered formalin, embedded in paraffin, sectioned, and 

stained with hematoxylin and eosin. Special staining techniques 

were utilized when indicated for more definitive diagnosis. 

A few of the tissues selected by design from some animals were not 

examined. Also, some animals were judged to be in such an advanced 

state of autolysis as to preclude histopathologic evaluation. Thus, 

the number of animals from which particular organs or tissues were 

examined microscopically varies, and does not necessarily represent 

the number of animals that were placed on study in each group. 

H. Data Recording and Statistical Analyses 

Pertinent data on this experiment have been recorded in an automatic data processing system, the Carcinogenesis Bioassay Data 

System (Linhart et al., 1974). The data elements include descriptive information on the chemicals, animals, experimental design, 

clinical observations, survival, body weight, and individual 

pathologic results, as recommended by the International Union 

Against Cancer (Berenblum, 1969). Data tables were generated for 

verification of data transcription and for statistical review. 

These data were analyzed using the statistical techniques 

9
 

 

described in this section. Those analyses of the experimental 

results that bear on the possibility of carcinogenicity are 

discussed in the statistical narrative sections. 

Probabilities of survival were estimated by the product-limit 

procedure of Kaplan and Meier (1958) and are presented in this 

report in the form of graphs. Animals were statistically censored 

as of the time that they died of other than natural causes or 

were found to be missing; animals dying from natural causes were 

not statistically censored. Statistical analyses for a possible 

dose-related effect on survival used the method of Cox (1972) for 

testing two groups for equality and Tarone's (1975) extensions of 

Cox's methods for testing for a dose-related trend. One-tailed P 

values have been reported for all tests except the departure from 

linearity test, which is only reported when its two-tailed P 

value is less than 0.05. 

The incidence of neoplastic or nonneoplastic lesions has been 

given as the ratio of the number of animals bearing such lesions 

at a specific anatomic site (numerator) to the number of animals 

necropsied (denominator). 

The purpose of the statistical analyses of tumor incidence is to 

determine whether animals receiving the test chemical developed a 

significantly higher proportion of tumors than did the control 

10
 

 

animals. As a part of these analyses, the one-tailed Fisher exact 

test (Cox, 1970) was used to compare the tumor incidence of a 

control group with that of a group of treated animals at each 

dose level. When results for a number of treated groups (k) are 

compared simultaneously with those for a control group, a 

correction to ensure an overall significance level of 0.05 may be 

made. The Bonferroni inequality (Miller, 1966) requires that the 

P value for any comparison be less than or equal to 0.05/k. In 

cases where this correction was used, it is discussed in the 

narrative section. It is not, however, presented in the tables, 

where the Fisher exact P values are shown. 

The Cochran-Armitage test for linear trend in proportions, with 

continuity correction (Armitage, 1971), was also used. Under the 

assumption of a linear trend, this test determines if the slope 

of the dose-response curve is different from zero at the onetailed 0.05 level of significance. Unless otherwise noted, the 

direction of the significant trend is a positive dose relationship. This method also provides a two-tailed test of departure 

from linear trend. 

A time-adjusted analysis was applied when numerous early deaths 

resulted from causes that were not associated with the formation 

of tumors. In this analysis, deaths that occurred before the 

first tumor was observed were excluded by basing the statistical 

11
 

 

tests on animals that survived at least 52 weeks, unless a tumor 

was found at the anatomic site of interest before week 52. When 

such an early tumor was found, comparisons were based exclusively 

on animals that survived at least as long as the animal in which 

the first tumor was found. Once this reduced set of data was 

obtained, the standard procedures for analyses of the incidence 

of tumors (Fisher exact tests, Cochran-Armitage tests, etc.) were 

followed. 

When appropriate, life-table methods were used to analyze the 

incidence of tumors. Curves of the proportions surviving without 

an observed tumor were computed as in Saffiotti et al. (1972). 

The week during which an animal died naturally or was sacrificed 

was entered as the time point of tumor observation. Cox's methods 

of comparing these curves were used for two groups; Tarone's 

extension to testing for linear trend was used for three groups. 

The statistical tests for the incidence of tumors which used 

life-table methods were one-tailed and, unless otherwise noted, 

in the direction of a positive dose relationship. Significant 

departures from linearity (P < 0.05, two-tailed test) were also 

noted. 

The approximate 95 percent confidence interval for the relative 

risk of each treated group compared to its control was calculated 

from the exact interval on the odds ratio (Gart, 1971). The 

12
 
 


 

relative risk is defined as Pt/pc where Pt is the true binomial 

probability of the incidence of a specific type of tumor in a 

treated group of animals and pc is the true probability of the 

spontaneous incidence of the same type of tumor in a control 

group. The hypothesis of equality between the true proportion of 

a specific tumor in a treated group and the proportion in a 

control group corresponds to a relative risk of unity. Values in 

excess of unity represent the condition of a larger proportion in 

the treated group than in the control. 

The lower and upper limits of the confidence interval of the 

relative risk have been included in the tables of statistical 

analyses. The interpretation of the limits is that in 

approximately 95% of a large number of identical experiments, the 

true ratio of the risk in a treated group of animals to that in a 

control group would be within the interval calculated from the 

experiment. When the lower limit of the confidence interval is 

greater than one, it can be inferred that a statistically 

significant result (P < 0.025 one-tailed test when the control 

incidence is not zero, P < 0.050 when the control incidence is 

zero) has occurred. When the lower limit is less than unity, but 

the upper limit is greater than unity, the lower limit indicates 

the absence of a significant result while the upper limit 

indicates that there is a theoretical possibility of the 

13
 
induction of tumors by the test chemical, which could not be 

detected under the conditions of this test. 

14 
 
 


 

III. RESULTS
 

A. Body Weights and Clinical Signs 

Hexachlorophene at the doses used in this bioassay had no effect on 

the mean body weights of the rats (figure 1). At week 24, an 

intercurrent respiratory infection occurred in the colony, but 

weights of animals in this bioassay were scarcely affected and 

recovery occurred within 4 weeks without treatment for infection. 

No other clinical signs were recorded. 

B. Survival 

The Kaplan and Meier curves estimating the probabilities of 

survival for male and female rats fed hexachlorophene in the diet 

at the doses of this study, together with those of the controls, 

are shown in figure 2. 

In neither sex is the Tarone test result significant for positive 

dose-related trend in mortality. In male rats, 79% of the high-dose 

group, 88% of the mid-dose and matched-control groups, and 67% of 

the low-dose group lived to termination of the study. In females, 

75% of the high-dose group, 79% of the mid-dose group, 50% of the 

low-dose group, and 63% of the matched-control group survived to 

termination of the study. Sufficient numbers of male 

15
 
!;! 

~ 

:1: 

" w 

3: 

>-

Q 

0 

.. 

z 

"' w 

:E 

!;! 

~ 

:1: 

!1 

w 

3: 

>-

c 

0 

.. 

z 

"' w 

:E 

"" 

,., 

'"' 

,., 

'" 

"'" 

MALE RATS 

[Jo'IATCHED CONTROL 

OLOWDOSE 

0Mt000SE 

AHIGH DOSE 

. 

....... 

.:t,~~~,,~--~'~,----~-----;,:~~~:"'~~~------~----~----"-----r----_j -r TIM 

60 

E ON STUDY 70 IWEEKSI ao 90 

,., 

"" 

""' 

"" 

,., 

"" 

,., 

" 

" " "' "' ~ "' TIME ON STUDY IWEEKS~o "' 

FEMALE RATS 

QMATCHED CONTROL 

QLOWDOSE 

0MtDOOSE 

i:I.HIGH DOSE 

00 "" "' 

Figure 1. Growth Curves for Rats Fed Hexachlorophene in the Diet 

16 
'"" 

•• 

0.00 

" • 0.70 

~ 

> 

, • ... • 

• 0 

~ 000 

0 

• • 0.40 

• 0 

• • ooo MALE RATS 

0 MATCHEOCONTROL 

0.00 0 LOW DOSE 

0 MID DOSE 

fi HIGH DOSE 

.. L 

0.00 

015304560 9C105 

TIME ON STUDY !WEEKSI 

,_00 f""" .. _ .. , .... @-,..--·~·-··-·-·····--··- -, D---- -----~·! ()- --, 

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45 60 75 90 

TIME ON STUDY (WEEKS) 

Figure 2. Survival Curves for Rats Fed Hexachlorophene in the Diet 

17 
 


 

and female rats were available for analyses of the incidences of 

late-appearing tumors. 

C. Pathology 

Histopathologic findings on neoplasms in rats are summarized in 

Appendix A, tables Al and A2; findings on nonneoplastic lesions are 

summarized in Appendix B, tables B1 and B2. 

A variety of neoplastic processes were observed in both the control 

and treated rats. Interstitial-cell tumors of the testes were the 

most frequently observed neoplasm in the control and treated males. 

Leukemia, generally of the granulocytic type, was the next most 

frequently observed neoplastic process in both the male and female 

animals. The incidence of this neoplasm was comparable among the 

control and treated groups. The following neoplasms were randomly 

distributed throughout the control and treated rats: squamous-cell 

carcinomas of the skin, alveolar/ bronchiolar adenomas of the lung, 

adenomas of the liver, transitional-cell carcinoma of the urinary 

bladder, adenomas of the pituitary, adenomas of the thyroid, 

adenomas and fibro-adenomas of the mammary gland, and endometrial 

stromal polyps of the uterus. 

The results of the microscopic examination indicate that the 

administration of hexachlorophene at the three doses used in this 

18
 
 

study did not have a carcinogenic effect in the Fischer 344 

rat. With the exception of interstitial-cell tumors of the testes 

and granulocytic leukemia, the incidence of neoplasia in both the 

control and treated rats was relatively low. 

D. Statistical Analyses of Results 

Tables Cl and C2 in Appendix C contain the statistical analyses 

of the incidences of those primary tumors that were observed in 

at least 5% of one or more of the treated groups. 

In male rats, the results of the Cochran-Armitage test for 

positive dose-related trend in the proportions of fibrosarcoma of 

the subcutaneous tissue and of mesothelioma, NOS (not otherwise 

specified), of the tunica vaginalis are significant (P < 0.050), 

but none of the results of the Fisher exact test are significant 

in any treated group when compared with controls. There is no 

other incidence of tumors at any specific site which is 

statistically significant in either sex. 

In each of the intervals of relative risk, shown in the tables, 

the value of one is included; this indicates the absence of 

positive significant results. It should also be noted that each 

of the intervals has an upper limit greater than one, indicating 

the theoretical possibility of the induction of tumors by hexa19 
chlorophene, which could not be detected under the conditions of 

this test. 

20 
 


 

IV. DISCUSSION 

At the doses used in this bioassay, hexachlorophene had no effect 

on mean body weights of Fischer 344 rats of either sex, and no 

clinical signs of toxicity were recorded. Survival also was 

unaffected at the doses used. Adequate numbers of rats in treated 

and control groups were available for analyses of the incidences 

of late-appearing tumors. 

In the subchronic study, neuronal necrosis of the brain was found 

in animals treated for 8 weeks at 200, 400, or 600 ppm, and the 

animals at 600 ppm also showed numerous clinical signs of 

toxicity. However, even at the highest dose used in the chronic 

study, i.e., 150 ppm, there were no effects on the nervous 

system, either clinically or histopathologically. Brains from all 

animals were routinely examined histologically; however, no other 

effort was made to demonstrate lesions of the nervous system. No 

tumors at any site were present in the treated rats in a 

statistically significant incidence. 

No previous studies of the carcinogenicity of hexachlorophene 

have been reported. However, toxicity studies in rats (Kimbrough, 

1976) have demonstrated neurotoxic signs (nervousness and 

hindquarter paralysis) and morphological changes (vacuolization 

of white matter) in the brain and spinal cord, following 

21
 

 

either oral (500 ppm, 10-14 weeks) or dermal (24 mg/kg) 

administration of hexachlorophene. Reversibility of toxic signs 

after discontinuing administration of hexachlorophene has also 

been reported (Kimbrough, 1971). 

It is concluded that under the conditions of this bioassay, 

hexachlorophene did not induce malignant or benign tumors in 

Fischer 344 rats. 

22
 
 

 

 

 

 

 

 

 


 


 

V. BIBLIOGRAPHY
 

Armitage, P., Statistical Methods, in Medical Research, John Wiley

& Sons, Inc., New York, 1911, pp. 362-365. 

Berenblum, I. , ed. , Carcinogenicity Testing A Report of the 

Panel of Carcinogenicity of the Cancer Research Commission,

of the UICC, Vol. 2, International Union Against Cancer, 

Geneva, 1969. 

Cox, D. R., Regression models and life tables. J. R. Statist. 

Soc. B. 34:187-220, 1912. 

Cox, D. R., Analysis of Binary Data, Methuen & Co., Ltd., London, 

1910, pp. 48-52. 

Environmental Protection Agency, EPA Compendium of Registered

Pesticides; Vol. II, U. S. Government Printing Office, 

Washington, D.C. 1975, pp. M-23-00.01 - M-23-00.03. 

Food and Drug Administration, Hexachlorophene, as a component of

drug and cosmetic products. Code of Federal Regulations

21:81-83, 1976. 

Food and Drug Administration, Hexachlorophene as a component in 

drug and cosmetic products for human use. Federal Register 

31:20160-20164, 1972. 

Gart, J. J., The comparison of proportions: a review of 

significance tests, confidence limits and adjustments for

stratification. Rev. Int. Stat. Inst. 39:148-169, 1971. 

Harvey, S. C., Antiseptics and disinfectants. In: The 

Pharmacological Basis of Therapeutics, Goodman, L. S. and

Gilman, A. G., eds., Macmillan Publishing Co., Inc., New

York, 1975, pp. 990-992. 

Kaplan, E. L, and Meier, P., Nonparametric estimation from 

incomplete observations. J. Am. Statist. Assoc. 53:457-481,

1958. 

Kimbrough, R. D., Hexachlorophene: toxicity and use as an 

antibacterial agent. In: Essays in Toxicology Vol. 7,

Academic Press, New York, 1976, pp. 100-120. 

23
 
 

 

 


 

Kimbrough, R. D. and Gainer, T. B., Hexachlorophene effects on

rat brain. Arch Environ Health 23:114-118, 1971. 

Linhart, M. S., Cooper, J. A., Martin, R. L., Page, N. P., and

Peters, J. A., Carcinogenesis bioassay data system. Comp and 

Biomed. Res. 7:230-248, 1974. 

Miller, R. G., Jr., Simultaneous Statistical Inference McGrawHill Book Co., New York, 1966, pp. 6-10. 

Saffiotti, U., Montesano, R., Sellakumar, A. R., Cefis, F. and

Kaufman, D. G., Respiratory tract carcinogenesis in hamsters

induced by different numbers of administrations of benzo (a)

pyrene and ferric oxide. Cancer Res 32:1073-1081, 1972. 

Tarone, R. E., Tests for trend in life table analysis. Biometrika

62(3):679-682,,1915. 

24
 

 


 


 


 

APPENDIX A

SUMMARY OF THE INCIDENCE OF NEOPLASMS
 

IN RATS FED HEXACHLOROPHENE IN THE DIET
 

25
 

----------------------------------------------------------------------------- ------------------------------------------------------------------------------------------------

TABLE A1. 


SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE 


RATS FED HEXACHLOROPHENE IN THE DIET 


CONTROL LOW DOSE MID DOSE HIGH DOSE 

ANIMALS INITIALLY IN STUDY 24 24 24 24 

ANIMALS NECROPSIED 24 24 24 24 

ANIMALS EXAMINID HISTOPATHOLOGICALLY 24 24 24 24 

INTEGUMENTARY SYSTEM 

*SKIN (2 4) (24) (24) (24) 


SQUAMOUS CELL CARCINOMA 1 (4 %) 


*SUBCUT TISSUE (2 4) (24) (24) (24) 

BASAL-CELl CARCINOMA 1 (4'11) 

FIBROSARCCIIA 3 (13'11) 

RESPIRATORY SYSTEM 

HUNG (24) (24) (24) (24) 


ALVEOLAR/ERONCHIOLAR ADENOMA 2 (8'11) 2 (8'11) 


ANGIOSARCOMA, METASTATIC 1 (4'11) 


HEMATOPOIETIC SYSTEM 

*MULTIPLE ORGANS (24) (24) (24) (24) 

GRANULOCY~IC LEUKEMIA 5 (21'11) 5 ( 21'11) 3 (13'11) 4 ( 17'11) 

#SPLf.EN (23) (24) (23) (24) 

HMIARTOMA 1 ('I'll) 

CIRCULATORY SYS~EII 

NONE 

DIGESTIVE SYSiEM 

tSALIVARY GLAND (1) (1) ____!N§lQ~A~~~~A-------------------------1-JlQQil_______________________________________________ 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY * NUMBER OF ANIMAlS NECROPSIED 

27 

TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE MID DOSE HIGH DOSE 

fLIVER (22) (23) (23) 


ADENOMA, NOS 1 (5") , ( 4ll) 


I COLON (2) (2) 

ADENOMATOU5 POLYP, NOS 1 (50") 

URINARY SYSTEM 

NCNE 

ENDOCRINE SYS1EM 

NONi 

REPPODUCTIVE SYSTEM 

*MAMIIARY GLANt (24) (24) (24) (24) 


F:i.BROMA 1 ( 4 %) 


LIPOMA (4%) 


nESTIS (24) (24) (24) (24) 

INTERSTITIAL-CELL TUllO!\ 24 ( 1 00%) 24 ( 100%) 24 ( 11)0%) 24 ( 100%) 

NERVOUS SYSTU 

IBPAIN (24) (24) 


GLIOIIA, NOS 


SPECIAL SENSE ORGANS 

NONi 

IIUSCULOSKELETAL SYSTEM 

NONE 

EODY CAVITIES 

*PERITONEUM (24) (24) (24) (24) 

____n~~Qitl~11£nA~_MQ~ ____________________1_J!~l-----------------·--------------------------------

t NU!!dER OF ANIMALS WITH TISSUE EXAMINED ~ICROSCOPICALLY 

* NU~BER OF ANIMALS NECROPSIED 

28 

TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE MID DOSE HIGH DOSE 

*TONICA VAGINALIS M~SOTHELICMA, NOS (24) (24) (24) (24) 

2 (8%) 

ALL OTHER SYS!t~S 

*~ULTIPLE ORGANS 

MESOTHELICMA, MALIGNANT 

(24) 

1 (4%) 

(24) (24) (24) 

1 (4%) 

ANIMAL DISPOSITION SUMMARY 

ANIMALS INITIALLY IN STUDY 

NATURAL DEATHiil 

MORIBUND SACRIFICE 

SCHEDOLEC SACRIFICE 

ACCIDENTALlY KILLED 

TERMINAL SACRIFICE 

ANIMAL MISSING 

24 

3 

21 

24 

3 

5 

16 

24 

2 

1 

21 

24 

1 

4 

19 

ill INCLUDES AUTCLYZED ANIMALS 

'IUI!OR SUMMARY 

TOTAL ANII!ALS WITH PRIMARY 

TOTAL PRIMARY TUMORS 

TUMORS* 24 

37 

24 

29 

24 

31 

24 

38 

TOTAL ANIMALS WITH BENIGN 

TOTAL BENIGN TUI!ORS 

TUMORS 24 

27 

24 

24 

24 

27 

24 

28 

TOTAL ANIMAlS WITH I!ALIGNANT 

TOTAL MALIGNANT TUMORS 

TUMORS 9 

9 

5 

5 

4 

4 

7 

8 

TOTAL ANIMALS WITH SECONDARY 

TOTAL SECCNDARY TUMORS 

TUMORS I 1 

1 

TOTAL ANII!ALS WITH TUMORS 

BENIGN OR MALIGNANT 

TOTAL UNCERTAIN TUI!ORS 

UNCERTAIN1 

1 

2 

2 

TOTAL ANII!ALS WITH TUMORS 

PRIMARY OR METASTATIC 

TOTAL UNCERTAIN TUMORS 

UNCERTAIN­

* PRIMARY TUMORS: ALL TUMORS EXCEPT SECONDARY TOI!ORS t SECONDARY TUMORS: METASTATIC TUMORS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN 

29 

------------------------------------------------------------------------------------------------

TABLE A2. 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE RATS 


FED HEXACHLOROPHENE IN THE DIET 


----------------------·------------------------------------------------------------------------­

CONTROL LOW DOSE MID DOSE HIGH DOSE 

ANIMALS INITIAllY IN STUDY 24 24 24 24 

ANIMALS N!CROPSI!C 24 24 24 24 

ANIMALS EXAMINED HISTOPATHOLOGICALLY 24 24 24 23 

---------~---------------~---------------------------------------------------------------------

INTEGUMENTARY SYS~!M 

*SUBCUT TISSUE (24) (24) (24) (24)

SQUAMOUS CELL CARCINOMA 1 (4ll)

ADENOCARCINOMA, NOS 1 (!Ill) 

CYSTADENCMA, NOS (4ll) 

FIBROMA 1 (4ll) (4ll) 

FIBROUS HISTIOCYTOMA, MALIGNANT 1 (4ll) 

LIPOMA (4ll) 

RESPihATORY SYS~E" 

'TRACHEA (22) (4) (1) 


FIBROSARCOMA 1 (25ll) 


HUNG (24) (23) (24) (23) 

ADENOCARCINOMA, NOS, METASTATIC 1 (!Ill) 

ALVEOLAR;EEONCHIOLAR ADENOMA 2 (8ll) 

PHEOCHROMCCYTOMA, METASTATIC 1 (4ll)

FIBROUS HISTIOCYTOMA, METASTATIC 1 (4ll) 

HEMATOPOIETIC SYSTEM 

*MULTIPLE ORGANS (24) (24) (24) (24) 

GRANULOCYTIC LEUKEMIA 8 (33ll) 6 (25ll) 10 (42ll) 3 ( 13ll) 

#LYMPH NODE (3) (5) (II) (9) 

ADENOCARCINOMA, NOS, METASTATIC 1 ( 11ll) 

HUNG (24) (23) (24) (23) 


GRANULOCYiiC LEUKEMIA 1 (4ll) 


CIRCULATORY SYSTEM 

__.l!Qll_________________________.______________________________________________ 

t NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMALS NECROPSIED 

30 

--------------------------------------------------------------------- -------------------------------------------------------------------------------------------------

TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE MID DOSE HIGH DOSE 

DIGESTIVE SYST!~ 

NONE 

URINARY SYST!~ 

'KIDNEY (24) (22) (24) (23) 

ADENOIIA, NOS 1 (4ll) 

IORINARY BLAttER ( 18) 

TRANSITIONAL-CELL PAPILLO~A 1 (6ll) 

TRANSITIONAl-CELL CARCINOIIA 1 (6ll) 

ENDOCRINE SYS!EII 

tPITUITARY 

ADENOMA, NOS 

(24) 

3 ( 13 "l 

(23) 

2 (9") 

(24) 

4 (17") 

(23) 

4 (17") 

#ADRENAL 

PHEOCHROIIOCYTCIIA 

PHEOCHROIIOCYTCIIA, IIALIGNANT 

(2) 

(50") 

( 1) 

1 (100ll) 

( 1) 

#THYROID 

ADENOIIA, NOS 

ADENOCARCINCMA, NOS 

(20) 

2 (10") 

(2) (1) 

1 (1 00") 

(1) 

(100") 

REPRODUCTIVE SYSTEM 

*MAIIIIARY GIANI (24) (24) (24) (24) 

ADENOMA, NCS 2 (8") 1 ( 4ll) 

ADENOCARCINCIIA, NOS 1 (4") 

CYSTADENCf.A, NOS (4ll) 

FIBROMA 3 ( 13") 

FIBROADENOMA 4 (17") (4") 

*GENITAL SY STEll (211) (24) (24) (24) 

SQUAIIOUS CELL CARCINOMA 1 ( 4") 

#UTERUS (22) (12) (11) (10) 

ENDOMETRIAL STROMAL POLYP 13 (59") 9 (75") 7 (64") 8 (80") ----~!~Q~~1EJ!1-~1BQ~!~_SAE~OIIA__________j_j~Jl__________________________________________ 

I NUMBER OF ANIIIALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUIIBER OF ANIIIALS NECROPSIED 

31 

TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 


CONTROL LOW DOSE MID DOSE HIGH DOSE 

fOVARY (23) (2) 


GRANULOSA-CELL CARCINOMA 1 ( 50li\) 


NERVOUS SYSTEM 

NONE 

SPECIAL SENSE ORGANS 

NONt: 

MUSCULOSKELETAl SYSTEM 

NONE 

PODY CAVITIES 

NON£ 

AlL OTHFR SYSTEMS 

*MULTIPLE ORGANS (2 4) (24) (24) 


SQUAMOUS CELL CARCINOMA 


THORAX 
 S~DAMOUS CELL CARCINOMA 


OMENTUM 


LIPOSARCOMA 


ANIMAL DISfOSIHCN SUMMARY 

ANIMALS INI'IIALlY IN STUDY 24 24 24 24 

NATURAL DEATHii> 1 4 4 2 

MOFIBUND SACRIFICE 8 8 1 4 

SCHEDULED SACRIFICE 

ACCIDENTAllY KILLED 

T<.FMINAL SACRIFICE 1 5 12 1 c 18 

ANIMAL MISSING 

~-llih1QQ~2-AU1~1XZIQ_AlilMA1~--------------------·--------------------------------------------

NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY 

* NUMBER OF ANIMAlS NECROPSIED 

32 

----------------------------------------------------------------------------------------------

TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

----------------------------------------------------------------------------------------------· .CONTROL LOW DOSE MID DOSE HIGH DOSE 

TUMOR SUMMARY 

TOTAL ANIMALS WITH PRIMARY 

TOTAL PRIBARY TUMORS 

TUMORS* 21 

38 

17 

28 

21 

31 

13 

19 

TOTAL ANIMAlS WITH BENIGN 

TOTAL EE~IGN 1UMORS 

TUMORS 16 

24 

12 

18 

15 

19 

11 

13 

TOTAL ANIMALS WITH MALIGNANT 

TOTAL MALIGNANT TUMORS 

TUMORS 12 

14 

8 

10 

12 

12 

5 

6 

TOTAL ANIMALS WITH SECONDARY 

TOTAL SECCNDARY TUMORS 

TUMORSI 1 

1 

1 

1 

1 

2 

TOTAL ANIMALS WITH TUMORS 

BENIGN OR MALIGNANT 

TOTAL UNCERTAIN TUMORS 

UNCERTAINTOTAL ANIMAlS WITH TUMORS 

PRIMARY OR METASTATIC 

TOTAL UNCERTAIN TUMORS 

UNCERTAIN­

* PRIMARY TUMORS: ALL TUMORS EXCEPT SECONDARY TUMORS t SECONDARY TUMORS: METASTATIC TUMORS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN 

33 


APPENDIX B 


SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS 


IN RATS FED HEXACHLOROPHENE IN THE DIET 


35 


TABLE 81. 


SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS 


IN MALE RATS FED HEXACHLOROPHENE IN THE DIET 


CONTROL LOW DOSE MID DOSE HIGH DOSE 

ANIMALS INITIALLY IN STUDY 24 24 24 24 

ANIMALS NECROPSIED 24 24 24 24 

ANIMALS EXAMINED HISTOPATHOLOGICALLY 24 24 24 24 

INTEGUMENTARY SYS!EM 

*SUBCUT TISSUE (24) (24) (24) (24) 


N.i:CROSIS, NOS 1 (4 'II) 


RESPIRATORY SYSTEM 

ILUNG/BRONCHUS (24) (24) (24) (24) 

BRONCHIECHSIS 1 (4'11) 

HUNG (2 4) (24) (24) (24) 

ATELECTASIS 2 (8'11) 4 ( 17'11) 1 (4'11) 

CONGESTICN, NOS 7 (29'11) 12 (50'11) 12 (50'11) 11 (46'11) 

INFLAMMATICN, NOS 2 (8'11) 1 (4 ll) 1 (41) 

INFLAMMATICN, FOCAL 1 (41) 

ABSCESS, NOS 1 (4ll) 1 (4'11) 1 (4 'II) 

HEMATOPOIETIC SYSTEM 

I SPLEEN (23) (24) (23) (24) 

HEMATOPOIESIS 1 (41) 

#LYMPH NODE ( 1) (1) ( 1) (6) 

LYMPHANGIECTASIS 1 (1001) 5 (83'11) 

CIRCULATORY SYSTEM 

IIIYOCARDIUM (1) 


l'IBROSIS 1 (1 OOll) 


DIGESTIVE SYSTEII 

ISALIVARY GLAND (1) (1) ____ lllk!~~A1l~B~-]Q2 ____________________________________l_j1QQ!L_______________________________

t NUIIBER OF ANIMALS WITH TISSUE EXAIIINED IIICROSCOPICALLY * NUIIBER OF ANIMALS NECROPSIED 

37 

TABLE 81. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 


CONTROL LOW DOSE 

INFLAMMA!ICN, NOS 	 1 (50") 

*SEIUNAL VESICLE (24) (24) 

DILATATICN, NOS 1 (4'll) 

HYPERPLASIA, NOS 1 ( 4'll) 

ITES'liS (24) (24) 


li.TBOPHY, NOS 2 (8") 


*EPIDIDYMIS (24) (24) 

INFLA!I!IATICN, NOS 1 ( 4%) 

NERVOUS SYSTEM 

NONE 

SPECIAL SENSE CRGANS 

NONE 

MUSCULOSKELETAL SYSTEM 

*MUSCLE OF EACK (24) (24) 

INFLAMMA!ICN, NOS 1 ( 4%) 

EODY CAVITIES 

*INGUINAL REGION (24) (24) 

NECROSIS, FAT 4 (17") 5 ( 21%) 

*MESENTERY (24) (24) 

NECROSIS, HEMORRHAGIC 1 (4~) 

ALL OIHER SYStEMS 

NONE 

SPECIAL MORPHOLOGY SUM!IABY 

NCN.i:. 

t NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY * 	NUMBER OF ANIMALS NECROPSIED 

MID DOSE HIGH DOSE 

(24) 	 (24) 

1 (4 "> 1 (4'll) 

(24) (24) 

(24) (24) 

(2 4) (24) 

(24) 	 (24) 

E ( 3 3%) 6 (25%) 

(2 4) (24) 

38 

TABLE 81. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

==================================================================================-------------­

CONTROL LOW DOSE MID DOSE HIGH DOSE 

tLIVER 

COHGESTIOII, NOS 

INFLAMMATICN, NOS 

INFLAMMATION, FOCAL 

ABSCESS, NCS 

HEPATITIS, TOXIC 

NECROSIS, !lOS 

NECROSIS 1 lOCA.L 

HYPERPLASIA, NODULAR 

AHGIECTASIS 

HEMATOPOHSIS 

(22) 

(5ll) 

(51) 

(23) 

7 

1 

2 

1 

(30$) 

(Ill) 

(Ill) 

(91) 

(Ill) 

(Ill) 

(211) 

1 

1 

1 

(II$) 

(Ill) 

(Ill) 

(Ill) 

(23) 

2 (91) 

(lill) 

(Ill) 

(Ill) 

fiSTOdACH 

DIVERTICULUM 

EDEMA, NOS 

(23) 

1 (II$) 

(8) 

(131) 

(1) (3) 

IGASTRIC MUCOSA 

HYPERPLASIA, NOS 

(23) (8) (1) 

1 (100ll) 

(3) 

fiCO LON 

A :rYPIA, NOS 

(2) (2) 

1 (501) 

fiCECU!I 

EDEMA, NOS 

INFLAMIIATICN, !lOS 

(2) 

1 (501) 

(2) 

(SOl) 

URINA!lY SYSTEM 

I KIDNEY (22) (211) (211) (211) 

HYDRONEPHROSIS 1 (Ill) 

CYST, NOS 1 (5$) 

tKIDNEY/PELVIS (22) (211) (211) (211) 

HEMORRHAGE 1 (Ill) 

ENDOCRINE SYS!!II 

tPITUITARY (211) (211) (211) (23) 

CONGESTION, NOS 1 (Ill) 

REPRODUCTIVE SYSTEM 

tPROSTATE (2) (3) 

__.J!!l.llUl~.!!.._l!Q~--------------------------------------------------Lj.l.}Jl_ 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY * NUMBER OF ANIMAlS NECROPSIED 

39 

TABLE 82. 


SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE 


RATS FED HEXACHLOROPHENE IN THE DIET 


CONTROL LOW DOSE MID DOSE HIGH DOSE 

ANIMALS INITIALLY IN STUI:Y 24 24 24 24 

ANIMALS NECROfSIEt 24 24 24 24 

ANIMALS EXAMINED HISTOPATHOLOGICALLY 24 24 24 23 

INTEGUMENTARY SYS!EM 

*SKIN (24) (24) (24) (24) 


EPIDERMAl INCLUSION CYST 1 (4ll) 


RESPIRATORY SYSTEM 

HUNG/BRONCHUS (24) (23) (24) (23) 


BRONCHIEC!ASIS 1 


INFlAMIIATICN, NOS 1 (4ll) 
 (4 "' 

#LUNG (24) (23) (24) (23) 

AIELECTASIS 2 (Bll I 2 ( 9ll) 3 (13ll) 

CONGESTION, NOS 7 (29ll) 8 (35ll) 4 (17ll) 12 (52ll) 

INFLAIIIIATICN, NOS 1 (4ll) 

ABSCESS, NOS 1 (4ll) 

HEIIATOPOIETIC SYSTEM 

tSPLEEN (24) (22) (2 1) (23) 

CONGESTION, NOS 1 ( 4ll) 

INFLAMMATICN, GRANULOMATOUS 1 (LI ll) 

FIBROSIS 1 ( 4ll) 

HEMATOPOIESIS 2 (8ll) 

tLYMPH NODE (3) (5) (4) (9) 

LYMPHANGIECTASIS 1 (3 3ll) 1 (20ll) 1 (2 5ll) 6 (67ll) 

INFLAMIIATICN, GRANULOMATOUS 1 (33ll) 

CIRCULATORY SYSTEM 

_ru:uu:_________________________________________________________________________:____ 

t NUIIBER OF ANIIIALS WITH TISSUE EXAIIINED MICROSCOPICALLY 

* NUMBER OF ANI!!ALS NECROPSIED 

40 

TABLE 82. FEMALE RATS: NONNEOPLASTIC LESIONS(CONTINUED) 


CONTROL LOW DOSE MID DOSE HIGH DOSE 

VIGESTIVE SYSiEM 

#LIVER (24) (23) (23) (20) 

CONGESTION, NOS 1 (4%) 1 (4%) 3 ( 15%) 

INFLAIIIIATICN, NOS 1 (4%) (9%) 1 (5%) 

ABSCESS, NCS 3 ( 13%) (9%) 

INFLAIIIIATICN, GRANULOIIA10US 3 ( 13%) 1 (4%) (1 3'11) 

HYPERPLASIA, NODULAR 1 (4 'I!) 2 ( 9:1) 

ANGIECTASIS 1 (5%) 

ICOLONIC SEECSA ( 1) (3) 

INFLAIIIIATICN, NOS 1 (33%) 

ADHESION, NOS 1 (33%) 

ICECUII ( 1 ) (3) 

INFLAMMATION, NOS 3 ( 100%) 

ADHESION, NOS (33%) 

HYPERPLASIA, lYMPHOID (33%) 

URINAf<Y SYSTEM 

~KIDNEY (24) (22) (24) (23) 


PYELONEPHRITIS, NOS 1 (4 %) 


tURINARY BLArrER ( 1 8) 


POLYP, INFlAMMATORY 1 (6%) 


ENDOCRINE SYStEM 

~PITUITARY (24) (23) (2 4) (23) 

CYST, NOS 1 (4%) 1 (4'11) 3 ( 13%) 

CONGESTICN, NOS 2 (8'1!) 2 ( 9%) 1 (4%) 2 (9%) 

HEMORRHAGE 1 (4'11) 3 ( 13%) 1 (4 %) 1 (4'11) 

IITHYROID (20) (2) ( 1) ( 1) 


INFLA!IIIATICN, NOS 2 (1 0%) 


REPRODUCTIVE SYSTEM 

liUTERUS (22) (12) (11) (10) 

____Hl~BQ~E1]~---------------------------l-l2~1----------l-l§!L__________J_l2~L----------l-llQ!l_ 

t NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY * NUMBER OF ANIMALS NECROPSIED 

41 

TABLE 82. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE MID DOSE HIGH DOSE 

PYOMETRA 4 (18:11) (9:11) 2 (20:11) 

ABSCESS, NOS 1 ( 10:11) 

INFARCT, NOS 1 (9%) 

#CERVIX U!EEI (22) (12) (11) (10) 


CYST, NOS 1 (9%) 


#UTERUS/ENDOMETRIUM (22) (12) ( 11) (1 0) 


HYPERPLASIA, CYSTIC 2 (9%) 1 (8:11) 


#UTERUS/MYOMETRIUM (22) (12) (11) (1 0) 


INFLAMMATICN, NOS 1 (8%) 


#OVARY (23) (2) 


CONGESTION, NOS 1 (50%) 


NERVOUS SYSTEM 

NONE 

SPECIAL SENSE CRGANS 

NONE 

MUSCULOSKELETAl SYSTEM 

NONE 

EODY CAVITIES 

NONE 

ALL OTHER SYSTEMS 

*MULTIPLE ORGANS (24) (24) (24) (24) 

INFARCT, NCS 1 (4%) 

ADIPOSE TISSUE ____Ill1A~~Ail~~~-~Q2____________________________________________________________________ l _____ _ 

t NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY * NUMBER OF ANIMALS NECROPSIED 

42 

--------------------------------------------------------------- -----------------------------------------------------------------------------------------------

TABLE 82. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE MID DOSE HIGH DOSE 

SPECIAL MORPBCIOGY SUMMARY 

NO LESION EE~ORTED 

AUTO/NECRO~SY/NO RISTO 

t NUMBER OP * NU!BER OP 

ANIMAlS 

ANIMAlS 

WITH TISSUE 

NECROPSIED 

EXAMINED !ICROSCOPICALLY 

43 


APPENDIX C 


ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS 


IN RATS FED HEXACHLOROPHENE IN THE DIET 


45 


Table Cl. Analyses of the Incidence of Primary Tumors in Hale Rats Fed 


Hexachlorophene in the Dieta 


Topography: Morphology 

Subcutaneous Tissue: 

Fibrosarcomab 

P Valuesc,d 

Relative Risk (Hatched Control)f 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Matched Low Mid High 

Control Dose Dose Dose 

0/24 (0) 0/24 (0) 0/24 (0) 3/24 (13) 

p = 0.009 N.S. N.S. N.S. 

Infinite 

0.624 

Infinite 

71 

Lung: Alveolar/Bronchiolar 

Adenomab 2/24 (8) 0/24 (0) 2/24 (8) 0/24 (0) 

P Valuesc,d N.S. N.S. N.S. N.S. 

Relative Risk (Matched Control)f 

Lower Limit 

Upper Limit 

0.000 

0.000 

3.283 

1.000 

0.078 

12.790 

0.000 

0.000 

3.283 

Weeks to First Observed Tumor 73 106 
Table Cl. Analyses of the Incidence of Primary Tumors in Male Rats Fed 

Hexachlorophene in the Dieta 

(continued) 

Matched Low Mid High 

Topography: Morphology Control Dose Dose Dose 

Hematopoietic System: 

Leukemiab 5/24 ( 21) 5/24 ( 21) 3/24 (13) 4/24 (17) 

P Valuesc,d N.S. N.S. N.S. N.S. 

Relative Risk (Matched Control)£ 1.000 0.600 0.800 

Lower Limit 0.263 0.104 0.181 

Upper Limit 3. 777 2.720 3.255 

Weeks to First Observed Tumor 92 83 86 92 

.+::­

00 Testis: Interstitial-cell 

Tumorb 24/24 (100) 24/24 (100) 24/24 (100) 24/24 (100) 

p Valuesc,d N.S. N.S. N.S. N. S. 

Relative Risk (Matched Control)£ 


Lower Limit 


Upper Limit 


Weeks to First Observed Tumor 73 83 86 71 
Table Cl. Analyses of the Incidence of Primary Tumors in Hale Rats Fed 

Hexachlorophene in the Dieta 

(continued 

Topography: Morphology 

Matched 

Control 

Low 

Dose 

Mid 

Dose 

High 

Dose 

Tunica Vaginalis: 

Mesothelioma, Nosh 0/24 (0) 0/24 (0) 0/24 (0) 2/24 {8) 

P Valuesc,d p = 0.042 N.S. N.S. N.S. 

Relative Risk (Matched Contro

Lower Limit 

Upper Limit 

l)f Infinite 

0.305 

Infinite 

Weeks to First Observed Tumor 106 

~ 

~ aTreated groups received doses of 17, 50, or 150 ppm in feed. 

bNumber of tumor-bearing animals/number of animals necropsied (percent). 

CBeneath the incidence of tumors in the control group is the probability level for the CochranArmitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the 

incidence of tumors in a treated group is the probability level for the Fisher exact test for 

the comparison of that treated group with the matched-control group when P < 0.05; otherwise, 

not significant (N.S.) is indicated. 

dA negative trend (N) indicates a lower incidence in a treated group than in the control group. 

eThe probability level for departure from linear trend is given when P < 0.05 for any comparison. 

fThe 95% confidence interval of the relative risk between each treated group and the matchedcontrol group. 
Table C2. Analyses of the Incidence of Primary Tumors in Female Rats Fed 


Hexachlorophene in the Dieta 


Topography: Morphology 

Matched 

Control 

Low 

Dose 

Mid 

Dose 

High 

Dose 

Lung: Alveolar/Bronchiolar 

Adenomab 0/24 (0) 0/24 (0) 2/24 (8) 0/24 (0) 

P Valuesc,d N.S. N.S. N.S. N.S. 

Relative Risk (Matched Control)f 

Lower Limit 

Upper Limit 

Infinite 

0.305 

Infinite 

Weeks to First Observed Tumor 106 

VI 

0 Hematopoietic System: 

Leukemiab 8/24 (33) 6/24 (25) 11/24 (46) 3/24 ( 13) 

p Valuesc,d N.S. N.S. N.S. N.S. 

Relative Risk (Matched Control)f 

Lower Limit 

Upper Limit 

0.750 

0.254 

2.075 

1.375 

0.619 

3.160 

0.375 

0.073 

l. 350 

Weeks to First Observed Tumor 74 84 92 88 
Table C2. Analyses of the Incidence of Primary Tumors in Female Rats Fed 

Hexachlorophene in the Dieta 

continued 

Topography: Morphology 

Pituitary: Adenoma, Nosh 

P Valuesc,d 

Relative Risk (Matched Control)f 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Matched 

Control 

3/24 (13) 

N.S. 

101 

Low 

Dose 

2/24 (8) 

N.S. 

0.667 

0.060 

5.292 

88 

Mid 

Dose 

4/24 (17) 

N.S. 

1. 333 

0.254 

8.185 

106 

High 

Dose 

4/24 ( 17) 

N.S. 

1. 333 

0.254 

8.185 

86 

Mammary Gland: Fibromab 0/24 (0) 3/24 (13) 0/24 (0) 0/24 (0) 

P Valuesc,d N.S. N.S. N.S. N.S. 

Departure from Linear Trende p = 0.018 

Relative Risk (Matched Control)f Infinite 

Lower Limit 0.624 

Upper Limit Infinite 

Weeks to First Obseryed Tumor 98 ~~~~~~~--------------------------~-------------------------------------
Table C2. Analyses of the Incidence of Primary Tumors 

Hexachlorophene in the Dieta 

(continued) 

Matched Low 

ToEograEhy: MorEhology Control Dose 

Mammary Gland: Fibroadenomab 0/24 (0) 4/24 ( 17) 

P Valuesc,d N.S. N.S. 

Departure from Linear Trende p = 0.022 

Relative Risk (Matched Control)£ Infinite 

Lower Limit 0.961 

Upper Limit Infinite 

Weeks to First Observed Tumor 66 V1 

N 

Mammary Gland: Adenoma, NOS, 

Cystadenoma, NOS, or Fibroadenomab 2/24 (8) 4/24 (17) 

p Valuesc,d N.S. N.S. 

Relative Risk (Matched Control)£ 2.000 

Lower Limit 0.319 

Upper Limit 20.335 

Weeks to First Observed Tumor 105 66 

in Female Rats 

Mid 

Dose 

1/24 (4) 

N.S. 

Infinite 

0.055 


Infinite 


103 

2/24 (8) 

N.S. 

1.000 

0.078 

12.790 

103 

Fed 

High 

Dose 

0/24 (0) 

N.S. 

1/24 (4) 

N.S. 

0.500 

0.009 

8.943 

106 
Table C2. Analyses of the Incidence of Primary Tumors in Female Rats Fed 

Hexachlorophene in the Dieta 

continued) 

Topography: Morphology 

Matched 

Control 

Low 

Dose 

Uterus: Endometrial 

Stromal Polypb 13/24 (54) 9/24 (38) 

P Valuesc,d N.S. N.S. 

Relative Risk (Matched Control)f 

Lower Limit 

Upper Limit 

0.692 

0.334 

1.398 

Weeks to First Observed Tumor 42 88 

Mid 

Dose 

High 

Dose 

7/24 (29) 8/24 (33) 

N.S. N.S. 

0.539 

0.231 

1.179 

0.615 

0.281 

1.290 

106 88 

U1 

w arreated groups received doses of 17, 50, or 150 ppm in feed. 

bNumber of tumor-bearing animals/number of animals necropsied (percent). 

CBeneath the incidence of tumors in the control group is the probability level for the CochranArmitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the 

incidence of tumors in a treated group is the probability level for the Fisher exact test for 

the comparison of that treated group with the matched-control group when P < 0.05; otherwise, 

not significant (N.S.) is indicated. 

dA negative trend (N) indicates a lower incidence in a treated group than in the control group. 

eThe probability level for departure from linear trend is given when P < 0.05 for any comparison. 

frhe 95% confidence interval of the relative risk between each treated group and the matchedcontrol group. 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cancer Institute's bioass ro am to identi and evaluate chemical

minutes of the Sub ou s meetin at which Hexachl hene was reviewed. 

that this have been the case for the treated female rats. The 

who die before terminal sacrifice in the total animal count. A staff 


 

Review of the Bioassay of Hexachlorophene* for Carcinogenicity by the

Data Evaluation/Risk Assessment Subgroup of the Clearinghouse on

Environmental Carcinogens 

November 28, 1977 

The Clearinghouse on Environmental Carcinogens was established in

May, 1976 under the authority of the National Cancer Act of 1971 (P.L.

92-218). The purpose of the Clearinghouse is to advise on the National

ay p gr fy

carcinogens in the environment to which humans may be exposed. The 

members of the Clearinghouse have been drawn from academia, industry, 

organized labor, public interest groups, State health officials, and 

quasi-public health and research organizations. Members have been 

selected on the basis of their experience in carcinogenesis or related 

fields and, collectively, provide expertise in organic chemistry, biochemistry, biostatistics, toxicology, pathology, and epidemiology, 

Representatives of various Governmental agencies participate as ad hoc 

members. The Data Evaluation/Risk Assessment Subgroup of the

Clearinghouse is charged with the responsibility of providing a peer

review of NCI bioassay reports on chemicals studied for 

carcinogenicity. In this context, below is the edited excerpt from the 

gr p' g orop 

Hexachlorophene had been used as a local antibiotic until it was

shown to cause neurologic toxicity and deaths in babies. 

Hexachlorophene was tested in rats as part of another study designed to

investigate the combined effects of chemicals. None of the treated 

groups showed an increased tumor incidence compared to the controls. 

As a negative study, one member said that it may be deficient

since only one species was used instead of the usual two. He added that 

greater emphasis should have been given to the histopathology of the

central nervous system. He also opined that it would have been more

appropriate to have tested Hexachlorophene by topical application than

by the oral route. Despite the shortcomings, the reviewer agreed with

the staff's conclusion that Hexachlorophene was not carcinogenic under

the conditions of test. 

A staff member pointed out that when no lesions are found at an 

organ site, the tissue count is not given in the report. He suggested 

may

Subgroup recommended that the table showing the neoplasms be changed to

include tissue counts in the absence of tumors. 

One Subgroup member questioned the validity of including animals 

member said that animals are included 

55
 
 

 

 

 

 

 

 

 

 

 

exce r. Garfinkel, wh sed it. 

in the denominator only when they live long enough to have been at

risk (usually 75 weeks) or until the time that the first tumor was

detected, whichever is earlier. 

It was moved that the bioassay report on Hexachlorophene be

accepted. The motion was seconded and approved by all present 

pt M o oppo 

Members present were: 

Gerald N. Wogan (Chairman), Massachusetts Institute of Technology 

Lawrence Garfinkel, American Cancer Society 

Henry C. Pitot, University of Wisconsin Medical Center

George Roush, Jr., Monsanto Company 

Verald K. Rowe, Dow Chemical U.S.A. 

Michael B. Shimkin, University of California at San Diego 

Louise Strong, University of Texas Health Sciences Center

John H. Weisburger, American Health Foundation 

* Subsequent to this review, changes may have been made in the 

bioassay report either as a result of the review or other

reasons. Thus, certain comments and criticisms reflected in

the review may no longer be appropriate. 

*U.S. GO VE RNMENT PRINTING OFFICE: 1978 260-899/3020 1-3 

56 

DHEW Publication No. (NIH) 78-840