1977 

National Cancer Institute 

CARCINOGENESIS 

Technical Report Series 

No. 11 

BIOASSAY OF 

TRISODIUM 

ETHYLEN EDIAMINETETRAACETATE 

TRIHYDRATE (EDTA) 

FOR POSSIBLE CARCINOGENICITY 

CAS No. 150-38-9 

NCI-CG-TR-11 

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

Public Health Service 

National Institutes of Health 
BIOASSAY OF
 

TRISODIUM ETHYLENEDIAMINETETRAACETATE TRIHYDRATE
 

FOR POSSIBLE CARCINOGENICITY
 

Carcinogen Bioassay and Program Resources Branch
 

Carcinogenesis Program
 

Division of Cancer Cause and Prevention
 

National Cancer Institute
 

National Institutes of Health
 

Bethesda, Maryland 20014
 

DREW Publication No. (NIH) 77-811
 

BIOASSAY OF
 

TRISODIUM ETHYLENEDIAMINETETRAACETATE TRIHYDRATE
 

For Possible Carcinogenicity
 

Carcinogenesis Program
 

Division of Cancer Cause and Prevention
 

National Cancer Institute
 

National Institute of Health
 

CONTRIBUTORS; This report presents the results of the bioassay of
 

trisodium ethylenediaminetetraacetate trihydrate (Na3EDTA-3H20)
 

for possible carcinogenicity, conducted by the Carcinogen
 

Bioassay and Program Resources Branch, Carcinogenesis Program,
 

Division of Cancer Cause and Prevention, National Cancer
 

Institute (NCI), Bethesda, Maryland. The bioassay was conducted
 

at Litton Bionetics, Inc., Kensington, Maryland, initially under
 

direct contract to the NCI and currently under a subcontract to
 

Tracor Jitco, Inc., the prime contractor for the NCI carcinogen
 

bioassay program.
 

The experimental design was determined, and the doses were
 

selected, by Drs. E. K. Weisburger* and J. H. Weisburger1

* ^.
 

Animal treatment and observations were supervised by Drs. B. M.
 

Ulland3>4 an(j F. M. Garner3, with the technical assistance of Mr.
 

J. D. Farmer3, Ms. H. I. Ruckenbrad3, Mr. D. J. Howard3, and Mr.
 

H. D. Thornett3.
 

Histopathology was performed at Litton Bionetics, Inc. by Drs. A.
 

de Paoli and J. Wosu for rats and by Dr. A. de Paoli for mice,
 

under the supervision of Dr. F. M. Garner, and the diagnoses
 

included in this report represent their interpretation.
 

Pathologists at NCI and Tracor Jitco, Inc. have reviewed the
 

pathology report and concur with the overall pathologic
 

evaluation of the study.
 

iii
 
Compilation of tables of individual animal survival and tables of
 

neoplasms and of nonneoplastic lesions was performed by EG&G
 

Mason Research Institute**. Statistical analyses were performed
 

by Dr. J. R. Joiner-*, using methods selected for the bioassay
 

program by Dr. J. J. Gart^.
 

This report was prepared at Tracor Jitco under the direction of
 

NCI. Those responsible for the report at Tracor Jitco were Dr.
 J. F. Robens^, toxicologist; Dr. S. S. Olin^, chemist; Ms. L. A.
 Waitz^, bioscience writer; and Dr. E. W. Gunberg^, technical
 

editor. The final report was reviewed by members of the
 

participating organizations^*3» ->.
 

^•Carcinogenesis Program, Division of Cancer Cause and Prevention,
 

National Cancer Institute, National Institutes of Health,
 

Bethesda, Maryland.
 

with the Nay lor Dana Institute for Disease Prevention,
 

American Health Foundation, Hammond House Road, Valhalla,
 

New York.
 

^Litton Bionetics, Inc., 5516 Nicholson Lane, Kensington,
 

Maryland.
 

with Hazleton Laboratories, Inc., 9200 Leesburg Turnpike,
 

Vienna, Virginia.
 

^Tracor Jitco, Inc., 1776 East Jefferson Street, Rockville,
 

Maryland.
 

Mason Research Institute, 1530 East Jefferson Street,
 

Rockville, Maryland.
 

'Mathematical Statistics and Applied Mathematics Section,
 

Field Studies and Statistics, Division of Cancer Cause and
 

Prevention, National Cancer Institute, National Institutes of
 

Health, Bethesda, Maryland.
 

iv
 
SUMMARY
 

A bioassay of the chelating agent, trisodium ethylenediaminetetraacetate trihydrate (Na3EDTA-3H20), for possible
 

carcinogenicity was conducted by administering the test material
 

in feed to Fischer 344 rats and B6C3F1 mice. The chemical was
 

administered to 50 males and 50 females of each species at low
 

and high concentrations, 3,750 and 7,500 ppm, for 103 weeks.
 

Matched-control groups were composed of 20 males and 20 females
 

of each species.
 

No compound-related signs of clinical toxicity were noted.
 

Although a variety of tumors occurred among test and control
 

animals of both species, no tumors were related to treatment.
 

Since survival was satisfactory and showed no consistent vari­ ation among test and control groups, the absence of treatment-


related tumors could not be attributed to early mortality.
 

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 9
 

H. Data Recording and Statistical Analyses 10
 

III. Results - Rats 13
 

A. Body Weights and Clinical Signs (Rats) 13
 

B. Survival (Rats) 13
 

C. Pathology (Rats) 16
 

D. Statistical Analyses of Results (Rats) 17
 

IV. Results - Mice 29
 

A. Body Weights and Clinical Signs (Mice) 29
 

B. Survival (Mice) 29
 

C. Pathology (Mice) 32
 

D. Statistical Analyses of Results (Mice) 33
 

V. Discussion 43
 

VI. Bibliography 47
 

APPENDIXES
 

Appendix A Summary of the Incidence of Neoplasms in
 

Rats Fed EDTA in the Diet 51
 

Table Al Summary of the Incidence of Neoplasms in
 

Male Rats Fed EDTA in the Diet 53
 

Table A2 Summary of the Incidence of Neoplasms in
 

Female Rats Fed EDTA in the Diet 57
 

vii
 
Page 

Appendix B Summary of the Incidence of Neoplasms in 

Mice Fed EDTA in the Diet 61 

Table Bl Summary of the Incidence of Neoplasms in 

Male Mice Fed EDTA in the Diet 63 

Table B2 Summary of the Incidence of Neoplasms in 

Female Mice Fed EDTA in the Diet 66 

Appendix C Summary of the Incidence of Nonneoplastic 

Lesions in Rats Fed EDTA in the Diet 69 

Table Cl Summary of the Incidence of Nonneoplastic 

Lesions in Male Rats Fed EDTA in the Diet... 71 

Table C2 Summary of the Incidence of Nonneoplastic 

Lesions in Female Rats Fed EDTA in the 

Diet '. 75 

Appendix D Summary of the Incidence of Nonneoplastic 

Lesions in Mice Fed EDTA in the Diet 79 

Table Dl Summary of the Incidence of Nonneoplastic 

Lesions in Male Mice Fed EDTA in the Diet... 81 

Table D2 Summary of the Incidence of Nonneoplastic 

Lesions in Female Mice Fed EDTA in the 

Diet 85 

Tables 

Table 1 Design of Chronic Study of EDTA 8 

Table 2 Analyses of the Incidence of Primary 

Tumors at Specific Sites in Male Rats 

Fed EDTA in the Diet 18 

Table 3 Analyses of the Incidence of Primary 

Tumors at Specific Sites in Female Rats 

Fed EDTA in the Diet 22 

Table 4 Analyses of the Incidence of Primary 

Tumors at Specific Sites in Male Mice 

Fed EDTA in the Diet 35 

viii 
Page
 

Table 5 Analyses of the Incidence of Primary 

Tumors at Specific Sites in Female Mice 

Fed EDTA in the Diet 38 

Figures 

Figure 1 Growth Curves for Rats Fed EDTA in the 

Diet 14 

Figure 2 Survival Curves for Rats Fed EDTA in the 

Diet 15 

Figure 3 Growth Curves for Mice Fed EDTA in the 

Diet 30 

Figure 4 Survival Curves for Mice Fed EDTA in the 

Diet 31 

ix
 

I. INTRODUCTION
 

Ethylenediaminetetraacetic acid (EDTA) is a synthetic aminopolycarboxylic acid chelating agent. It has been used extensively as a food additive to sequester trace metals that catalyze the
 

oxidation of oils, vitamins, and unsaturated fats and cause
 

rancidity, flavor changes, and discoloration (Furia, 1975).
 

Permissible levels of calcium disodium EDTA in food range from 25
 

to 800 ppm (Food and Drug Administration, 1974), and an
 

acceptable daily intake of 2.5 mg/kg has been established (Joint
 

FAO/WHO Expert Committee on Food Additives, 1974). EDTA has also
 

been used to control the interactions of trace metals in
 

formulations of liquid soaps, cosmetics, and Pharmaceuticals, in
 

metal working, in pulp and paper processing, in rubber and
 

polymer chemistry, and in textile processing and dyeing (Dow
 

Chemical Company, 1974). Calcium disodium EDTA has been used
 

primarily in the treatment of metal poisoning, although toxicity
 

to the renal proximal tubules necessitates precautions with this
 

therapy (Foreman et al., 1956).
 

EDTA was selected for testing because of structural and
 

functional similarity to another chelating agent, namely,
 

nitrilotriacetic acid trisodium salt (Na3NTA'H20), which also was
 

tested in this program.
 

1
 

II. MATERIALS AND METHODS

 

A. Chemical
 

The trisodium ethylenediaminetetraacetate trihydrate
 

(Na3EDTA-3H20) used in this study was supplied by Pfaltz & Bauer,
 

Inc., Flushing, N. Y. The chemical is hereinafter referred to
 

as EDTA. Its molecular weight is 410.84, of which 70% is
 

ethylenediaminetetraacetic acid. Identity of the material was
 

verified by infrared and nuclear magnetic resonance (NMR)
 

spectroscopy. The NMR analysis showed that the compound was in
 

the trihydrate form. The sodium content as determined by flame
 

emission spectrophotometry was 17.1%, compared to a theoretical
 

content of 16.8% for the trisodium salt trihydrate. Thin-layer
 

chromatography in two solvent systems produced a single spot as
 

visualized by iodine vapor, methyl red, and phenol red indicator
 

and amine-specific ferric chloride - potassium ferricyanide
 

spray.
 

B. Dietary Preparation
 

A 6-kilogram batch of dosed feed was prepared in a hooded
 

twin-shell blender two times a week for mice and three times a
 

week for rats. Preparations and chemicals were refrigerated at
 

4°C until they were used.
 

3
 
Analyses were performed, using FDA methods (Food and Drug
 

Administration, 1965) to determine the efficiency of the mixing
 

procedure and the stability of the test chemical in feed.
 

Samples were taken from the bottom and two wings of the blender,
 

and analyses were performed in triplicate; the error is reported
 

as the standard deviation. Recoveries were found to be 90.3 i
 

1.4% of the theoretical value at 7,500 ppm EDTA and 90.4 i 3.4%
 

of the theoretical value at 3,750 ppm. Analyses performed on
 

samples from the same batches after 12 days of storage at ambient
 

temperature gave 95.5 i 2.7% of the theoretical value at 7,500
 

ppm and 92.1 i 5.7% at 3,750 ppm. It was concluded from these
 

results that the preparations contained reasonably accurate
 

concentrations of EDTA and were mixed homogeneously, and that the
 

chemical was stable in feed for at least a week. When
 

concentrations of EDTA were rechecked near termination of the
 

chronic study, the accuracy of mixing and the homogeneity of the
 

mixtures was reconfirmed.
 

C. Animals
 

Inbred Fischer 344 rats were received from A. R. Schmidt,
 

Madison, Wisconsin, and hybrid B6C3F1 mice from the Charles River
 

Breeding Laboratories, Wilmington, Massachusetts, at 28 days of
 

age. These laboratories were under contract to the Division of
 

Cancer Treatment, NCI, to provide animals used for testing. On
 

4
 
arrival at the laboratory, the animals were quarantined for 14
 

days to acclimate them to the laboratory environment and to allow
 

observation of their physical condition. Animals were considered
 

acceptable for testing if they had no clinical signs of disease
 

and were within a weight range of 19-22 g for mice and 85-110 g
 

for rats at the end of the quarantine period. Animals were
 

weighed individually and segregated into equal weight groups.
 

Cage assignments were then made by choosing one animal from each
 

such group, so that the total animal weight was the same in each
 

cage.
 

D. Animal Maintenance
 

During the chronic study rats were housed four per cage, and
 

mice, five per cage in solid polycarbonate cages suspended on
 

moveable racks. All cages were lined with heat-treated hardwood
 

chip bedding and covered with filter paper over a wire mesh
 

screen. Clean cages and fresh bedding were provided twice a
 

week. Water bottles were sanitized and refilled with acidulated
 

water (pH 2.5) twice a week. Test diets were prepared with
 

Wayne^ Lab Blox Meal (Allied Mills, Inc.) that was also used as
 

feed for the control animals. Feed was available ad libitum and
 

replaced three times a week. Air in the animal rooms was changed
 

15 times per hour and exhausted through HEPA filters. Animal
 

rooms were negatively pressurized with respect to the clean hall
 

5
 
and positively pressurized with respect to the dirty hall. The
 

room air was maintained at 21-25°C and 45-55% humidity. Rooms
 

were illuminated by fluorescent lighting for 8 hours a day.
 

Rats and mice were housed in separate rooms. Matched controls
 

were housed with the respective test animals. Animals treated
 

with EDTA were maintained in rooms with animals of the same
 

species being treated with other chemicals as follows:
 

Rats
 

iodomethanesulfonic acid, sodium salt
 

3-hydroxy-(3 alpha, 5 beta)cholan-24-oic acid
 

3,3-dimethyl-2-oxethanone
 

Mice
 

hydroxytriphenylstannane
 

4,4'-diisocyanato-3,3'-dimethoxy-l,1'-biphenyl
 

N-(aminocarbonyl)-2-bromo-2-ethylbutanamide
 

N,N'-diethylthiourea
 

mono(2,2-dimethylhydrazine)butanedioic acid
 

iodomethanesulfonic acid, sodium salt
 

2,5-cyclohexadiene-l,4-dione, dioxime
 

3-hydroxy-(3 alpha, 5 beta)cholan-24-oic acid
 

4-amino-2-nitrophenol
 

E. Sub_cJjronic_^Studies
 

Feeding studies were conducted to estimate the maximum tolerated
 

doses in order to determine the high and low concentrations
 

(hereinafter referred to as "high doses" and "low doses") to be
 

administered in the chronic studies. The low doses given in the
 

chronic studies were 1/2 of the high doses. In the subchronic
 

6
 
studies EDTA was added to the animal feed at five dietary
 

concentrations: 4,640, 6,800, 10,000, 14,700, and 21,600 ppm.
 

The compound was provided in feed to experimental groups of five
 

male and five female animals of each species for 7 weeks,
 

followed by a 1-week period of observation.
 

During these studies soft stools, a sign of compound-related
 

toxicity (Foreman, 1953; Yang, 1964), were noted at 10,000 ppm
 

and above in male rats and at 14,700 ppm and above in female
 

rats. After 8 weeks, body weights of the treated rats at all
 

doses were comparable to the weights of the matched controls.
 

Gross pathologic examinations showed no signs of organ toxicity
 

in rats. In mice, depression of weight was noted in males at
 

21,600 ppm and in females at 6,800 ppm and above. One male
 

receiving 21,600 ppm died. There were no pathologic changes in
 

any animals detected at necropsy. The low and high doses for
 

both species were set at 3,750 and 7,500ppm.
 

F • Re

JLi&HJ°!J^°!liJ:Lj>tH,1!^
 

The design of the chronic studies, including both test and
 

matched-control groups, is shown in table 1.
 

77 
Table 1. Design of Chronic Studies of EDTA
 

Species, Sex
 

and Treatment
 

Group
 

RATS
 

Male
 

Matched-Control
 

Low-Dose
 

High-Dose
 

Female
 

Matched-Control
 

Low-Dose
 

High-Dose
 

MICE
 

Male
 

Matched-Control
 

Low-Dose
 

High-Dose
 

Female
 

Matched-Control
 

Low-Dose
 

High-Dose
 

Initial
 

No. of
 

Animals
 

20
 

50
 

50
 

20
 

50
 

50
 

20
 

50
 

50
 

20
 

50
 

50
 

EDTA
 

in Diet
 

(ppm)
 

0
 

3,750
 

7,500
 

0
 

3,750
 

7,500
 

0
 

3,750
 

7,500
 

0
 

3,750
 

7,500
 

Time on
 

Treated
 

(weeks)
 

0
 

103
 

103
 

0
 

103
 

103
 

0
 

103
 

103
 

0
 

103
 

103
 

Study
 

Untreated
 

(weeks)
 

104
 

1
 

1
 

104
 

1
 

1
 

104
 

1
 

1
 

104
 

1
 

1
 

8 
G. Clinical and Pathologic Examinations
 

All animals were observed twice daily £or signs of toxicity,
 

weighed at regular intervals, and palpated for masses at each
 

weighing. Those animals appearing moribund at the time of
 

clinical examination were killed and necropsied.
 

The pathologic evaluation consisted of gross and microscopic
 

examination of all major tissues, organs, or gross lesions taken
 

from killed animals and, when feasible, from animals found dead.
 

The following tissues and organs were routinely subjected to
 

microscopic examination: skin, lymph nodes, mammary gland,
 

salivary gland, bone marrow, trachea, lungs and bronchi, heart,
 

thyroids, parathyroids, esophagus, stomach, small intestine,
 

large intestine, liver, gallbladder, pancreas, spleen, kidneys,
 

adrenals, urinary bladder, prostate or uterus, testis or ovary,
 

brain, and pituitary. Occasionally additional tissues were also
 

examined microscopically. The different tissues were preserved
 

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

stained with hematoxylin and eosin. An occasional section was
 

subjected to special staining techniques for more definitive
 

diagnosis.
 

A few tissues were not examined for some animals, particularly
 

for those that died early. Also, some animals were missing,
 

cannibalized, or judged to be in such an advanced state of
 

9 
autolysis as to preclude histopathologic interpretation. Thus,
 

the number of animals for which particular organs, tissues, or
 

lesions were examined microscopically varies and does not
 

necessarily represent the number of animals that were placed on
 

experiment in each group.
 

H. Data Recordingjind 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, animal 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.
 

Probabilities of survival were estimated by the product limit
 

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

in the form of graphs. Deaths due to accident or scheduled
 

deaths are treated as censored observations and all other deaths
 

are uncensored. Statistical tests of differences in survival
 

between groups are compared using the method of Cox (1972) for
 

two groups and an extension of this method by Tarone (1975) for
 

more than two groups.
 

10
 
The incidence of neoplastic or nonneoplastic lesions is given as
 

the proportion of the number of animals bearing such lesions at a
 

specific anatomic site (numerator) to the number of animals
 

examined pathologically at that site (denominator). For the
 

organs and tissues in which most of the lesions appeared, the
 

denominators included only those animals for which such sites
 

were examined histologically. For tissues that required gross
 

observation for detection of lesions (e.g., skin or mammary
 

tumors), for lesions that appeared at several sites (e.g.,
 

lymphomas), or for tissues that were examined histologically only
 

when lesions were detected grossly, the denominators consisted of
 

the numbers of animals necropsied.
 

Statistical analysis of the incidence of tumors was made using
 

the Fisher exact test (Cox, 1970) to compare a control group to a
 

group of treated animals at each dose. In addition, the Armitage
 

and Cochran test for linear trend in proportions, with continuity
 

correction (Armitage, 1971), was used. This test, assuming a
 

linear trend, determines if the slope of the dose-response curve
 

is different from zero, at the 0.05 level of significance. The
 

method also calculates the level of probability of a departure
 

from linear trend.
 

A conservative adjustment, the Bonferroni inequality (Miller,
 

1966), was used for simultaneous comparison of several treated
 

11
 
groups with a control group. For the comparison of results
 

obtained with k different test doses with those for a control,
 

this correction requires a level of significance less than or
 

equal to 0.05/k for the overall comparison to be significant at
 

the 0.05 level. This adjustment was not made in the tables where
 

the Fisher exact test results are shown but is discussed in the
 

analysis when appropriate.
 

As an additional analysis, the exact 95% confidence interval for
 

the odds ratio (Cart, 1970) between each of the treated groups
 

and its control was calculated. The odds ratio' is pt(l-pc)/pc
 

(l-pt) where pt is the true binomial probability of tumor in a
 

treated animal and pc is the true spontaneous tumor probability
 

in the controls. The hypothesis of equality between the true
 

proportion of a specific tumor in a treated group and that in a
 

control is expressed by an odds ratio of 1 (one). Values in
 

excess of 1 (one) represent the condition of a larger proportion
 

in the treated group than in the control. The confidence
 

probabilities, Pt~Pc» where Pf-Pc 0 implies an odds ratio of 1
 

interval entries in the statistical tables of this report 

represent the conversion of each odds ratio to the difference in 

=

(one).
 

12
 
III. RESULTS - RATS
 

A

- Body Weights and Clinical Signs (Rats)^
 

Average body weights of treated male and female rats were
 

comparable to those of the matched controls throughout the study
 

(figure 1). No significant signs were observed among test
 

animals during the first year of the study. In the 6 months
 

preceding termination of the test, corneal opacities, ascites,
 

and urine stains, occurred in both treatment and control groups.
 

B. Survival plats)
 

Curves showing the probability of survival of treated and control
 

rats are shown in figure 2. The male rats exhibited a negative
 

dose-related trend in survival with the probability level of P =
 

0.103; the treated and control groups of male rats can thus be
 

considered as comparable to each other in survival. The female
 

rats also exhibited a negative dose-related trend in survival,
 

but in this case the effect was statisically significant (P =
 

0.029).
 

13
 
Figure 1. Growth Curves for Rats Fed EDTA in the Diet
 

14
 
Figure 2. Survival Curves for Rats Fed EDTA in the Diet
 

15
 
C. p.atholo^x_(Rats}
 

Histopathologic findings on neoplasms in rats are tabulated in
 

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

are tabulated in Appendix C, tables Cl and C2. The incidence of
 

neoplasms was high in the reproductive and endocrine systems and
 

lower in the hematopoietic, respiratory, integumentary, and
 

digestive systems. No neoplasms were observed in the nervous,
 

musculoskeletal, or urinary systems or in organs of special
 

sense.
 

Interstitial-cell tumors of the testes were observed in nearly
 

all male rats in each feeding group. This high incidence of
 

interstitial-cell tumors in both treated and control animals
 

reflects this commonly occurring age-related lesion in the male
 

Fischer 344 rat.
 

In females the distribution of neoplasms in the reproductive
 

system among control and treated rats was random, the tumors
 

occurred mainly in the uterus. The majority of these were
 

endometrial stromal polyps. However, one adenocarcinoma and one
 

leiomyosarcoma occurred in high-dose animals. An ovarian
 

cystadenoma was detected in a single low-dose rat.
 

A variety of endocrine tumors were found, some types occurring
 

only in treated animals. However, these tumors occurred in low
 

16
 
numbers and have frequently been seen in untreated animals in
 

other studies. Therefore, they are probably unrelated to treatment.
 

A number of tumors occurred in other organ systems of both sexes,
 

controls as well as treated animals. In some instances the
 

incidence of tumors in the controls exceeded that of the treated
 

animals. With the possible exception of endocrine tumors in the
 

males, no clear association between the incidence of tumors,
 

treatment, or sex could be established.
 

Inflammatory and degenerative changes were observed in about the
 

same frequency in all groups. These lesions appeared to be
 

related to age and not to the administration of the chemical.
 

In the judgment of the pathologist, the nature, incidence, and
 

severity of the lesions observed in this study provide no clear
 

evidence of carcinogenic effect in rats.
 

D. Statistical Analyses of Results (Rats)
 

Tables 2 and 3 list those tumors that occurred in more than 5% of
 

the rats in a given treated group or appeared in a larger
 

proportion in the treated group when compared to the matched
 

controls. There was no other control group whose environment,
 

period of test, and pathologic diagnosis were comparable to the
 

17
 
Table 2. Analyses of the Incidence of Primary Tumors at Spec

in Hale Rats Fed EDTA Trisodiuro Salt in the Diet3 

Matched LOI-J 

Topography: Morphology Control Dose 

Hematopoietic System: Leukemia, Malignant
 

Lymphoma and Lymph ocy tic Leukemia" 3/20 (0.15) 4/50 (0

I' Values0 N.S. N.S. 

95% Confidence Tntervald (-0.25,0

Weeks to First Observed Turaor 76 104 

Adrenal: Pheochromocytoma" 2/20 (0.10) 5/49 (0

P Values0 N.S. N.S. 

95% Confidence Intervald (-0.19,0

Weeks to First Observed Tumor 104 104 
Table 2. Analyses of the Incidence of Primary Tumors at Specific Sites

in Male Rats Fed EDTA TrisodiuH Salt in the Diet3
 

(continued)
 

Topography: Morphology
 

Thyroid: C-cell Adenoma^
 

P Values0
 

95% Confidence Intervald
 

Weeks to First Observed Tumor
 

Pituitary: Chromophobe Adenoma**
 

P Values0
 

95% Confidence Intervald
 

Weeks to First Observed Tumor
 

Matched
 

Control
 

0/17 (0.00)
 

N.S.
 

—
 

0/18 (0.00)
 

P - 0.089
 

—
 

Low
 

Dose
 

6/35 (0.17)
 

P - 0.080
 

(-0.07,1.00)
 

104
 

3/47 (0.06)
 

N.S.
 

(-0.09,1.00)
 

88
 
__ 

Table 2. Analyses of the Incidence of Primary Tumors at Specific Sites

in Male Rats Fed EDTA Trisodium Salt in the Diet3
 

(continued)
 

Topography: Morphology
 

Lung: Alveolar/Bronchiolar Adenoma
 

and Carcinoma'5
 

P Values0
 

95% Confidence Intervald
 

Weeks to First Observed Tumor
 

Liver: Uepatocellular Adenoma
 

and Neoplastic Nodule**
 

P Values0
 

95% Confidence Intervald
 

Weeks to First Observed Tumor
 

Matched
 

Control
 

1/18 (0.06)
 

N.S.
 

104
 

0/20 (0.00)
 

N.S.
 

Low
 

Dose
 

2/50 (0.04)
 

N.S.
 

(-0.14,0.06)
 

95
 

1/48 (0.02)
 

N.S.
 

(-0.05,1.00)
 

104
 
Table 2. Analyses of the Incidence of Primary Tumors at Specific 

in Male Rats Fed EDTA Trisodium Salt in the Dieta
 

(continued)
 

Matched Low 

Topography: Morphology Control ______ JDose. 

Testis: Interstitial-cell Tunorb 19/20 (0.95) 43/50 (0.86) 

P Values0 N.S. N.S. 

95% Confidence Intervald (-0.14,0.13) 

Weeks to First Observed Tumor 88 85 

^Treated groups received average doses of 3,750 and 7,500 ppm in feed.
 

"Number of tumor-bearing animals/number of animals examined at site (proportion).

c

Beneath the proportions for the matched-control group are the probability levels 

for the Cochran-Armitage test for dose-related trend in proportions when P is be

otherwise, N.S. (not significant) is indicated.
 

Beneath the proportions for the treated groups are the P values for the Fisher e

tional) test for the comparison of the treated groups with the matched-control g

is below 0.05; otherwise, N.S. is indicated.
 

<*95% confidence interval of the difference in proportions of treated group and ma

group.
 
Table 3. Analyses of the Incidence of Primary Tumors at Specific S

in Female Rats Fed EDTA Trisodium Salt in the Diet3
 

Topography: Morphology
 

Hematopoietic System: Malignant Lymphoma, »
 

Leukemia and Lymphocytic Leukemia"
 

P Values0
 

95% Confidence Intervald
 

Weeks to First 'Observed Tumor
 

Adrenal: Pheochromocytoma'3
 

P Values0
 

95% Confidence Intervald
 

Weeks to First Observed Tumor
 

Matched Low 

Control Dose 

1/20 (0.05)	 8/50 (0.16) 

N.S.	 N.S. 

(-0.11,0.18) 

104 80
 

1/20 (0.05)	 1/49 0.02) 

N.S.	 N.S. 

(-0.10,0.04) 

98 104 
Table 3. Analyses of the Incidence of Primary Tumors at Specif

in Female Rats Fed EDTA Trisodium Salt in the Diet3
 

(continued)
 

Matched Low 

Topography: Morphology Control Dose 

Thyroid: C-cell Adenomab 0/11 (0.00) 0/36 (0.0

P Values0 N.S. N.S. 

95% Confidence Interval*1 

Weeks to First Observed Tumor 

— —
 

Pituitary: Chromophobe Adenomab 6/19 (0.32) 10/48 (0

P Values0 N.S. N.S. 

95% Confidence Intervald (-0.37,0

Weeks to First Observed Tumor 95 104 
Table 3. Analyses of the Incidence of Primary Tumors at Specif

in Female Rats Fed EDTA Trisodium Salt in the Diet3 

(continued) 

Topography: Morphology
 

Lung: Alveolar/Bronchiolar Adenoma*5
 

P Values0
 

95% Confidence Intervald
 

Ueeks to First Observed Turaor
 

Liver: Me op las tic Nodule"
 

P Values0
 

95% Confidence Tntervald
 

Weeks to First Observed Tumor
 

Matched
 

Control
 

0/20 (0.00)
 

M.S.
 

—
 

0/20 (0.00)
 

N.S.
 

Low
 

Dose
 

3/48 (0.0

M.S.
 

(-0.08,1.

104
 

1/48 (0.

N.S.
 

(-0.05,1.00

104
 
Table 3. Analyses of the

in Female Rats

(continued) 

Topography: Morphology
 

Uterus: Endometrial Stromal Polypb
 

P Values0
 

95% Confidence Intervald
 

Weeks to First Observed Tumor
 

Mammary Gland: Fibroadenoma'3
 

P Values0
 

95% Confidence Intervald
 

Weeks to First Observed Tumor
 

 Incidence of Primary Tumors at Specific 

 Fed EDTA Trisodium Salt in the Dieta 

Matched Low 

Control Dose 

5/20 (0.25)	 6/50 (0.12) 

N.S.	 N.S. 

(-0.35,0.07

104	 96 

4/20 (0.20)	 3/50 (0.06) 

N.S.	 N.S. 

(-0.29,0.04) 

85	 96 

a

Treated groups received average doses of 3,750 and 7,500 ppm in feed.
 

^Number of tumor-bearing animals/number of animals examined at site (proportion)
Table 3. Analyses of the Incidence of Primary Tumors at Specif

in Female Rats Fed EDTA Trisodium Salt in the Diet3
 

(continued)
 

c

Beneath the proportions for the matched-control group are the probability lev

for the Cochran-Armitage test for dose-related trend in proportions when P is 

otherwise, N.S. (not significant) is indicated.
 

Beneath the proportions for the treated groups are the P values for the Fishe

tional) test for the comparison of the treated groups with the matched-control 

is below 0.05; otherwise, N.S. is indicated.
 

^95% confidence interval of the difference in proportions of treated group and 

group.
 
matched-control group used in this analysis, so no pooled-


control group was used.
 

No tumor appeared in a statistically significant (P < 0.05)
 

positive trend in either sex. The Cox test for positive trend
 

(unadjusted for time of appearance) has a probability level of P
 

= 0.089 for chromophobe adenoma of the pituitary, and the Fisher
 

exact test for the comparison of C-cell adenomas of the thyroid
 

of the low-dose and untreated males had a probability level of P
 

= 0.08. Except for these two results, all statistical tests
 

showed probability levels higher than 0.10.
 

As an additional statistical test, the 95% confidence interval of
 

each group was calculated and entered in the tables. The
 

implication of this interval is that in 95/100 (95%) of a large
 

number of similar experiments, the true difference between the
 

tumor rate for the treated group of animals and the rate for the
 

control group would be inside the interval calculated from the
 

experiment. In each of the intervals shown in the tables, zero
 

is included; this indicates the negative aspects of the results.
 

It should also be noted that each of the intervals has a positive
 

endpoint, indicating the theoretical possibility of tumor
 

induction by EDTA, which was not detected under the conditions of
 

this test.
 

27
 

IV. RESULTS - MICE
 

A. Body Weights and Clinical Signs (Mice)
 

In male mice only the high-dose group showed throughout most of
 

the test period a decrease in average body weight compared to the
 

controls (figure 3). In female mice average body weights of the
 

treatment groups were depressed in a dose-related manner during
 

the test period, although the effect was small (figure 3). Few
 

clinical signs of toxicity appeared in mice during the test.
 

Ataxia occurred in a low-dose male at 8 months, and ascites was
 

noted in mice of both sexes during the second year of the study.
 

B. Survival (Mice)
 

Curves for the probability of survival of treated and control
 

mice are shown in figure 4. There was no statistically
 

significant difference in survival between the different groups,
 

and this finding applied to both sexes. In male mice, 5/50 (10%)
 

of the low-dose group, 2/50 (4%) of the high-dose group, and 1/20
 

(5%) of the matched controls were accidentally killed or missing
 

at 26 weeks or before and are, therefore, censored at the time of
 

death in the survival curve. While there was a higher rate of
 

mortality in the treated male groups than in the control males,
 

29
 
Figure 3. Growth Curves for Mice Fed EDTA in the Diet
 

30
 
Figure 4. Survival Curves for Mice Fed EDTA in the Diet
 

31 
the highest rate was observed in the low-dose group, and therefore the Tarone test for positive trend was not significant.
 

No low-dose female mice died accidentally (compared to the five
 

low-dose males); however, 1/20 (5%) of the matched-control group
 

and 3/50 (6%) of the high-dose group were lost in this way during
 

the first 22 weeks of study and were censored from the survival
 

curve. The Tarone test for dose-related trend in the female mice
 

resulted in a nonsignificant probability.
 

C. Pathology (Mice)
 

Histopathologic findings on neoplasms in mice are tabulated in
 

Appendix B, tables Bl and B2; findings on nonneoplastic lesions
 

are tabulated in Appendix D, tables Dl and D2.
 

A variety of neoplasms was observed in both treated and control
 

animals. Each type observed has been encountered previously as a
 

spontaneous lesion in the mouse. However, the incidence of
 

neoplasms in all groups was high in the hematopoietic, endocrine,
 

digestive, and respiratory systems. The incidence of neoplasms
 

in other systems was variable.
 

With the exception of a splenic hemangioma in a control female
 

and a low-dose male, all of the tumors of the hematopoietic
 

system were malignant lymphomas or leukemias.
 

32
 
The distribution of endocrine tumors varied little between
 

treated and control mice; the bulk of such tumors were of the
 

pituitary gland, affecting 12 females and 2 males.
 

The incidence of hepatic neoplasms was considerably higher in
 

males than in females. The males appeared to have a significant
 

number of hepatocellular adenomas. However, the incidence of
 

these neoplasms in the low-dose (9/44 [20%]) and high-dose (6/47
 

[13%]) male groups was approximately the same as in the male
 

controls (2/19 [11%]).
 

Primary neoplasms of the respiratory system were observed in both
 

treated and control groups and were most: common in the high-dose
 

males. Even though the greatest incidence of pulmonary neoplasms
 

were found in the high-dose male mice (control 2/18 [11%],
 

low-dose 8/44 [18%], and high-dose 12/45 [26%]), the frequent
 

spontaneous occurrence of neoplasms in the lungs of mice suggest
 

that the distribution observed in this study is probably not
 

related to treatment.
 

In the judgment of the pathologist, the nature, incidence, and
 

severity of the lesions observed in this study provide no clear
 

evidence of carcinogenic effect of EDTA in mice.
 

D. Statistical Analyses of Results (Mice)
 

Tables 4 and 5 list those tumors which occurred in more than 5%
 

33
 
of the mice in a given treated group or appeared in a larger
 

proportion in the treated group when compared to the matched
 

controls. There was no other control group available whose
 

environment, period of test, and pathologic diagnosis were
 

comparable to the matched-control group used in this analysis, so
 

no pooled-control group was used. Tumors of the hematopoietic
 

system, lung, and liver were observed in over 10% of the
 

matched—control males, while tumors of the hematopoietic system
 

and pituitary gland were seen in over 10% of the matched-control
 

females. Although this relatively high incidence in the matched
 

controls may have concealed statistical evidence of
 

carcinogenicity, it should be noted (1) that there were no
 

significant differences in incidence among different treated
 

groups and (2) that these tumors have appeared spontaneously in
 

matched-control groups in other studies.
 

The only tumor system with a positive dose-related probability
 

level less than 0.10 was alveolar/bronchiolar carcinoma or
 

adenoma of the lung (P = 0.096). The Fisher exact test had a
 

probability level of over 0.10 in all comparisons between treated
 

and control groups. There is, therefore, no statistical evidence
 

of carcinogenicity of EUTA at the concentrations administered.
 

As an additional statistical test, the 95% confidence interval of
 

each group was calculated and entered in the tables. The
 

34
 
Table 4. Analyses of the Incidence of Primary Tumors

in Male Mice Fed EDTA Trisodium Salt in the

Topography: Morphology
 

Hematopoietic System: Malignant
 

Lymphoma"
 

P Values0
 

95% Confidence Intervald
 

Weeks to First Observed Tumor
 

Lung: Alveolar/Bronchiolar Adenoma
 

and Carcinoma"3
 

P Values0
 

95% Confidence Intervald
 

Weeks to First Observed Tumor
 

Matched
 

Control
 

2/20 (0.10)
 

N.S.
 

91
 

2/18 (0.11)
 

P = 0.096
 

105
 

Diet 3 

Low
 

Dose
 

7/46 (0.15)
 

N.S.
 

(-0.18,0.19)
 

73
 

8/44 (0.18)
 

N.S.
 

(-0.18,0.20)
 

99
 

 at Specific Si
Table 4. Analyses of the Incidence of Primary Tumors at Specific 

in Male Mice Fed EDTA Trisodium Salt in the Diet3 

(continued) 

Matched Low 

Topography: Morphology Control Dose 

Pituitary: Chromophobe Adenoma'3 1/13 (0.08) 0/19 (0.00) 

P Values0 N.S. N.S. 

95% Confidence Intervald (-1.00,0.05) 

Weeks to First Observed Tumor 105 

Liver: Hepatocellular Adenoma
 

and Carcinoma*5 3/19 (0.16) 10/44 (0.23) 

P Values0 N.S. N.S. 

95% Confidence Intervald (-0.20,0.23) 

Weeks to First Observed Tumor 103 84 
Table 4. Analyses of the Incidence of Primary Tumors at Specific S

in Male Mice Fed EDTA Trisodium Salt in the Diet3
 

(continued)
 

Matched Low 

Topography: Morphology Control Dose 

Thyroid: Follicular-cell Adenoma
 

and Carcinoma'5 0/10 (0.00) 1/29 (0.03) 

P Values0 N.S. N.S. 

95% Confidence Intervald (-0.09,1.00) 

Weeks to First Observed Tumor 104 

—
 

a

Treated groups received average doses of 3,750 and 7,500 ppm in feed.
 

^Number of tumor-bearing animals/number of animals examined at site (proportion).
 

c

Beneath the proportions for the matched-control group are the probability levels 

for the Cochran-Armitage test for dose-related trend in proportions when P is bel

otherwise, N.S. (not significant) is indicated.
 

Beneath the proportions for the treated groups are the P values for the Fisher ex

tional) test for the comparison of the treated groups with the matched-control gr

is below 0.05; otherwise, N.S. is indicated.
 

^95% confidence interval of the difference in proportions of treated group and mat

group.
 
Table 5. Analyses of the Incidence of Primary Tumors at Speci

in Female Mice Fed EDTA Trisodium Salt in the Diet3 

Matched Low 

Topography: Morphology Control Dose 

Hematopoietic System: Malignant
 

Lymphomab 5/19 (0.26) 11/49 (0.

P Values0 N.S. N.S. 

95% Confidence Intervald (-0.30,0

Weeks to First Observed Tumor 85 99 

Lung: Alveolar/Bronchiolar Adenoma^5 0/19 (0.00) 3/47 (0.

P Values0 N.S. N.S. 

95% Confidence Intervald (-0.09,1

Weeks to First Observed Tumor 105 

—
 
Table 5. Analyses of the Incidence of Primary Tumors at Specific 

in Female Mice Fed EDTA Trisodium Salt in the Diet3 

(continued) 

Topography: Morphology
 

Pituitary: Chromophobe	 Adenoma**
 

P Values0
 

95% Confidence Intervald
 

Weeks to First Observed Tumor
 

Liver: Hepatocellular Adenoma
 

and Carcinoma*5
 

P Values0
 

95% Confidence Interval0*
 

Weeks to First Observed Tumor
 

Matched Low 

Control Dose 

2/12 (0.17)	 6/34 (0.18

N.S.	 N.S. 

(-0.31,0.1

105 105 

0/19 (0.00)	 1/46 (0.02

N.S.	 N.S. 

(-0.05,1.0

105 

—
 
Table 5. Analyses of the Incidence of Primary Tumors at Specif

in Female Mice Fed EDTA Trisodium Salt in the Dieta
 

(continued)
 

Matched Low 

Topography: Morphology Control Dose 

Thyroid: Follicular-cell Adenoma
 

and Carcinoma^3 1/12 (0.08) 3/33 (0.0

P Values0 N.S. N.S. 

95% Confidence Interval4 (-0.23,0.

Weeks to First Observed Tumor 105 99 

a

Treated groups received average doses of 3,750 and 7,500 ppm in feed.
 

^Number of tumor-bearing animals/number of animals examined at site (proportio

c

Beneath the proportions for the matched-control group are the probability lev

for the Cochran-Armitage test for dose-related trend in proportions when P is 

otherwise, N.S. (not significant) is indicated.
 

Beneath the proportions for the treated groups are the P values for the Fishe

tional) test for the comparison of the treated groups with the matched-control 

is below 0.05; otherwise, N.S. is indicated.
 

^95% confidence interval of the difference in proportions of treated group and 

group.
 
implication of this interval is that in 95/100 (95%) of a large
 

number of similar experiments, the true difference between the
 

tumor rate for the treated group of animals and the rate for the
 

control group would be inside the interval calculated from the
 

experiment. In each of the intervals shown in the tables, zero
 

is included; this indicates the negative aspects of the results.
 

It should also be noted that each of the intervals has a positive
 

endpoint, indicating the theoretical possibility of tumor
 

induction by EDTA, which was not detected under the conditions of
 

this test.
 

41
 

V. DISCUSSION
 

EDTA and its salts are poorly absorbed following oral administration, due to dissociation and subsequent precipitation caused by
 

gastric pH (Foreman et al., 1953). EDTA salts have shown
 

relatively little toxicity in feeding studies, perhaps because of
 

the poor absorption. Toxicity varies according to the specific
 

salt: disodium EDTA is more toxic than either calcium disodium
 

EDTA or lead disodium EDTA (Reuber and Schmieler, 1962), and
 

chromium EDTA is less toxic than calcium EDTA (Ahrens and
 

Aronson, 1.971).
 

Early in its clinical use, EDTA showed potential nephrotoxicity,
 

specifically tubular hydropic degeneration, following parenteral
 

administration (Foreman et al., 1956). However, Doolan et al.
 

(1967) questioned the relative hazards of this effect clinically
 

and reported that in rats tubular vacuolization was not
 

accompanied by significant elevation of serum creatinine or urea
 

nitrogen. The cause of the renal vacuolization is not known. It
 

may be a reflection of the induction of pinocytosis by
 

Na3EDTA«3H20 (Schwartz et al., 1970) or of the presence of a
 

foreign nonmetabolizable substance (Doolan et al., 1967). In
 

studies of younger rats, 4 weeks of age, Reuber (1967) reported
 

43
 
that hydropic change of the parenchymal cells of the liver rather
 

than of the kidney was predominant.
 

The chronic toxicity of f^EDTA'Sl^O used in this bioassay has
 

not been previously invest igated. However, in two feeding
 

studies in rats, using concentrations of N32EDTA as high as 5% of
 

the diet (50,000 ppm) over a ttro-year period, no
 

treatment-related effects were found on gross or histopathologic
 

examination of organs or tissues (Yang and Chan, 1964); toxic
 

manifestations were limited to diarrhea in animals receiving the
 

highest dose of EDTA, reduced weight gain in some of the treated
 

groups, and increased blood coagulation in one of the studies but
 

not the other. The absence of adverse effects following oral
 

administration was also shown for rats receiving 1,000, 2,500,
 

and 5,000 ppm Cat^EDTA-ll^O in feed for two years (Oser, 1963).
 

In the present bioassay there was no evidence of carcinogenicity
 

of EUTA at the concentrations administered, and no tumors or
 

lesions of the kidney or other organs were related to the
 

treatment. Survival of all groups of animals of both species was
 

good; thus, the lack of appearance of treatment-related tumors
 

could not be attributed to early mortality. It should be noted
 

that the confidence intervals for all tumor sites include a
 

positive value; this indicates that the possibility of
 

tumorigenicity is not theoretically precluded. However, under
 

44
 
the conditions of this study, using concentrations of 3,500 ppm
 

and 7,500 ppm in feed, Na3EDTA'3H20 was not demonstrated to be
 

carcinogenic in rats or mice.
 

45
 

VI. BIBLIOGRAPHY
 

Ahrens, F. A. and Aronson, A. L. A comparative study of the
 

toxic effects of calcium and chromium chelates of
 

ethylenediaminetetraacetate in the dog. Toxicol. Appl.
 

Pharmacol. 18:10-25, 1971.
 

Armitage, P. Statistical Methods jji Medical Research, John
 

Wiley and Sons, New York, 1971, p. 135.
 

Berenblum, I., ed. Care inogenic ity Testing, UICC Technical
 

Report Series, Vol. 2. International Union Against Cancer,
 

Geneva, 1969.
 

Cox, D. R, Analysis £f_ Binary Data, Methuen, London, 1970, pp.
 

61-65.
 

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

Soc. B 34:187-220, 1972.
 

Doolan, P. D. , Schwartz, S. L., Hayes, J. R. , Mullen, J. C. and
 

Cummings, N. B. An evaluation of the nephrotoxicity of
 

ethylenediaminetetraacetate and diethylenetriaminepentaacetate in the rat. Toxicol. Appl. Pharmacol. 10;481-500,
 

1967.
 

Dow Chemical Company, Keys ^p_ Chelation with Versene Chelating
 

Agents. Dow Chemical Co., Midland, Michigan, 1974.
 

Food and Drug

Analytical 

Manual, Wa

 Administration,

shington, D.C, 19

 U.S.D.H.E.W.

65. 

 Food Additives 

Food and Drug Administration, U.S.D.H.E.W. Calcium disodium 

EDTA. Code of Federal Regulations, 21 CFR 10.1 Sec.
 

121.1017, 1974.
 

Foreman. H. , Vier, M. and Magee, M. The metabolism of
 

C^^-labeled ethylenediatninetetraacetic acid in the rat. J_._
 

Biol. Chem. 203:10451053, 1953.
 

Foreman, H., Finnegan, C. and Lushbaugh, C. C. Nephrotoxic
 

hazard from uncontrolled edathamil calcium-disodium therapy.
 

J.A.M.A. 160:1042-1046, 1956.
 

47
 
Furia, R. E., ed. Handbook of Food Additives, 2nd ed. CRC Press,
 

Cleveland, Ohio, 1975.
 

Gart, J. J. Point and interval estimation of the common odds
 

ratio in the combination of 2 x 2 tables with fixed
 

marginals. Biometrika 57:471-475, 1970.
 

Joint FAO/WHO Expert Committee on Food Additives. Toxicological
 

Evaluation of Certain Food Additives with £ Review c?f
 

General Principles and of_ Specifications. FAO Nutrition
 

Meeting Report Series 53. World Health Organization, Rome,
 

1974.
 

Joint FAO/WHO Expert Committee on Food Additives. Toxicological
 

Evaluation o£ Some Food Additives Including Anticaking
 

Agents, Antimicrobials, Antioxidants, Emulsifiers, and
 

Thickening Agents. FAO Nutrition Meeting Report Series 53A.
 

World Health Organization, Rome, 1974.
 

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

incomplete observations. J. Amer. Statist. Assn.
 

_53_:457-481, 1958.
 

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

Peters, J. A. Carcinogenesis bioassay data system. J_._
 

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

Miller, R. G., Jr. Simultaneous Statistical Inference,
 

McGraw-Hill, New York, 1966.
 

Oser, B. L. , Oser, M. and Spencer, H. C. Safety evaluation
 

studies of calcium EDTA. Toxicol. Appl. Pharmacol.
 

_5:142-162, 1963.
 

Reuber, M. D. Hepatic lesions in young rats given calcium
 

disodium edetate. Toxicol. Appl. Pharmacol. 11:321-326,
 

1967.
 

Reuber, M. D. and Schmieler, G. C. Edetate kidney lesions in
 

rats. Arch. Environ. Health 5:430-436, 1962.
 

Schwartz, S. L. , Johnson, C. B. and Doolan, P. D. Study of the
 

mechanism of renal vacuologenesis induced in the rat by
 

ethylenediaminetetraacetate: Comparison of the cellular
 

activities of calcium and chromium chelates. Molec.
 

Pharmacol. 6:54-60, 1970.
 

48
 
Tarone, R. E. Tests for trend in life table analysis.
 

Biometrika 62:679-682, 1975.
 

Yang, S. S. and Chan, M. S. Summaries of toxicological data:
 

Toxicology of EDTA. Food Cosmet. Toxicol. 2:763-767, 1964.
 

49
 

APPENDIX A
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS
 

IN RATS FED EDTA IN THE DIET
 

51
 

TABLE A1

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE RATS 

FED EDTA IN THE DIET 

CONTROL (UNTB) LOW DOSE
 

ANIMAi.3 INITIALLY IN STUDY 20 50
 

ANIMALS NECFOPSIED 20 50
 

ANIMALS EXAMINED HISTOPAT HOLOGICALLY 20 50
 

INTEGUMENTABY SYSTEM
 

*SKIN (20) (50)
 

SEBACEOUS ADENOMA 1 (2S)
 

KERATOACANTHOMA 1 (5X)
 

*SUBCtJT TISSUE (20) (50)
 

SQUAMOUS CELL CARCINOMA 1 (2*)
 

FIBBOMA 1 <5X) 3 (6%)
 

FIBBOSABCOMA 1 (2X)
 FIBBOADENOMA
 

BESPIHATOBY SYSTEM
 

ILUNG (18) (50)
 

SQUAMOUS CELL CABCINOMA 1 (2%)
 

ALVEOLAfi/BEONCHIOLAH ADENOMA 1 (6X) 1 (2X)
 

ALVEOLAR/BBONCHIOLAH CARCINOMA 1 12%)
 

CORTICAL CARCINOMA, METASTATIC
 

NONCHROMAFFIN PAB AG ANGL IOM A , MET
 

FIBHOSABCOMA 1 (2%)
 

HEMATOPOIETIC SYSTEM
 

*MULTIPLE OBGANS (20) (50)
 LEUKEMIA, DOS 3 (15X) 3 (6*)
 LYMPHOCYTIC LEUKEMIA 1 (2*)
 

CIRCULATORY SYSTEM
 

«HEABT __ (18) (U8)
 —
 aiyEOLA.S^BRONC.HIOL.AR.SAJCOOMA.^.. J_J2X1___.
 

» NUMBER OF ANIMALS BITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBEE OF ANIMALS NECROPSIED
 

HIGH DOSE
 

50
 

50
 

50
 

(50)
 

(50)
 

1 (2X)
 

Ct 9)
 

3 (6X)
 

1 (2X)
 

1 (2X)
 

(50)
 

H <8X)
 

(<*•))
 

_—_•__—.
 

53
 
TABLE A1. MALE RATS: NEOPLASMS (CONTINUED)

 

HIGH DOSE
 

(50)
 

1 (2*)
 

(U9)
 

2 (<»*)
 

(UI»)
 

5 (11*)
 

(50)
 

1 (2*)
 

1 (2*)
 

1 (2*)
 « (8*)
 

(38)
 

1 (3*)
 

3 (8*)
 

(21)
 

(«8)
 

2 <"•*)
 

(50)
 

l|U (88X)
 

(50)
 

1_U21_­

DIGESTIVE SYSTEH
 

ILIVEB
 HEPATOCELLULAB ADENOMA
 

NEOPLASTIC NODULE
 

ISMALL INTESTINE
 

LEIOMYOSARCOMA
 

URINARY SYSTEH
 

HONE
 

INDOCRINE SYSTEM
 

IPITUITAHY
 

CHROHOPHOBE ADENOMA
 

IADBENAL
 

COBTICAL ADENOMA
 CORTICAL CARCINOMA
 

NONCHBOMAPFIH PARAGANGLIOM A , HAL
 

PHEOCHHOMOCYTOHA
 

ITHYBOID
 

FOLLICULAB-CELL ADENOMA
 

C-CELL ADENOMA
 

IPARATHYBOID
 

ADENOMA, NOS
 

IPANCPEATIC ISLETS
 

ISLET-CELL ADENOMA
 

REPRODUCTIVE SYSTEM
 

tTESTIS
 

INTERSTITIAL-CELL TUMOR
 

NERVOUS SYSTEM
 

•CEREBRUM
 

__A5IBO£YT21U
 

CONTBOL(UNTB)
 

(20)
 

(20)
 

(18)
 

(20)
 

1 (5*)
 

2 (10X)
 

(17)
 

(12)
 

(19)
 

(20)
 

19 (95%)
 

(20)
 

LOW DOSE
 

(i»8)
 

1 (2*)
 

(50)
 

(»7)
 

3 (6*)
 

(i»9)
 

5 (10*)
 

(35)
 

6 (17*)
 

(2«)
 

1 («*)
 

C»7)
 

3 (6*)
 

(50)
 

43 (86*)
 

(50)
 

» NUMBER OF ANIMALS KITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECBOPSIED
 

54
 
TABLE A1 MALE RATS: NEOPLASMS (CONTINUED) 

»BRA1 N
 

GLIOMA.
 

SPECIAL SENSE ORGANS
 

NONE
 

MUSCULOSKEI.ETAI. SYSTEM
 

NONE
 

BODY CAVITIES
 

*BODY CAVITIES
 

MLSOTHELIONA,
 

*PLEURA
 

ALVEOLAR/BRONCHIOLAR CARCINOMA,
 

ALL OIHES SYSTEMS
 

THORACIC CAVITY
 

PIBROSARCOBA
 

ANIMA L DISPOSITION SUMMAR Y
 

ANIMALS INITIALLY IN STUDY
 

NATURA L DEATHS
 

MORIBUND SACRIFICE
 

SCHEDULED SACRIFICE
 

ACCIDENTALLY KILLED
 

ThFNINAL SACRIFICE
 ANIMA L MISSING
 

S INCLUDES iUTOLJfZED ANIMUS
 

CONTROL (UNTR) LOW DOSE HIGH DOSE 

(20) (50) (50) 

2 (IX) 

(20)
 

(20)
 

20
 

1
 

It
 

15
 

(50)
 

(50)
 

1 ,(2X)
 

1
 

50
 

6
 

It
 

40
 

(50)
 

2 («)
 

(50)
 

50
 

1
 

a
 

145
 

« NUMUE R OF ANIMAL S WITH TISSUE EXAMINED MICROSCOPICALLY
 * NUMBER OF ANIMALS NECROPSIED
 

55
 
TABLE A1 MALE RATS: NEOPLASMS (CONTINUED) 

CONTROL (UNTR) LOW DOSE HIGH DOSE
 

"IUMOR SUMMAR Y 

TOTAL ANIMAL S WITH PRIMARY TUMORS* 

TOTAL PRIMARY TUMORS 

20 

28 

<4H 

77 

47 

78 

TOTAL ANIMALS WITH BENIGN TUMORS 

TOTAL BENIGN TUMORS 

19 

25 

U7 

67 

U5 

6<* 

TOTAL ANIMAL S WITH MALIGNANT TUMORS 

TOTAL MALIGNANT TUMORS 

3 

3 

7 

10 

1 1 

11 

TOTAL ANIMALS WITH SECONDARY TUMORS* 

TOTAL SECONDARY TUMORS 

1 

2 

2 

2 

TOTAL ANIMALS WITH TUMORS UNCERTAIN­ BENIGN OR MALIGNANT 

TOTAL UNCERTAIN TUMORS 

3 

3 

TOTAL ANIMAL S WITH TUMORS UNCERTAINPRIMARY OR METASTATIC 

TOTAL UNCERTAIN TUMORS 

* PRIMARY TUJiORS: ALL TUMORS EXCEPT SE iCONDARY TUMORS 

* SECONDARY TUMORS: METASTATIC TUMORS OR TUMORS INVA SIVE INTO AN AD JACENT ORGAN
 

56
 
TABLE A2 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE RATS 

FED EDTA IN THE DIET 

ANIMAL S INITIAU.Y IN STUDY
 

ANIMALS NECROPSIED
 

ANIMAL S EXAMINE D H ISTOP ATHOLOG ICA LL Y
 

INTEGUMENTARY SYSTEM
 

*SUBCIJT TISSUE
 

SUUAMOUS CELL CARCINOMA
 

FIBROADENOMA
 

RESPIRATORY SYSTEM
 

*LARYNX
 

SQUAHOUS CELL PAPILLOMA
 

tLUNO
 

ALVEOLAR/BROSICHIOLAR ADENOMA
 

IIEMATOPOIETIC SYSTEM
 

*MULTIPLE ORGANS
 

MALIGNANT LYMPHOMA,
 

LEUKFMIA.NCS
 

LYMPHOCYTIC LEUKEMIA
 

ILYMPH NODE
 

MALIGNAN T LYMPHOMA,
 

CIRCULATORY SYSTEM
 

NONE
 

DIGESTIVE SYSTEM
 

ILIVER
 

NEOPLASTIC NODULE
 

CONTROL (UNTR)
 

20
 

20
 

20
 

(20)
 

1 (5%)
 

(20)
 

(20)
 

(20)
 

1 (5%)
 

(20)
 

(20)
 

.__


LOW UOSF HIGH DOSE
 

50 50
 

50 50
 

50 50.
 

(50) (50)
 

1 <2S)
 

1 (2X) 2 (IK)
 

(50)	 (50)
 

1 (2X)
 

(18)	 (MS)
 

3 ((,%) 2 (tX)
 

(50)	 (50)
 

2 (UX)
 

4 (8X)
 

2 (!»%>
 

(46)	 (i»7)
 

1 (2«)
 

C*9) (119)
 

1 J2X1
 

*	 NUMBER OF ANIMALS WITH TISSUE EXAMINE D MICROSCOPICALLY
 

*	 NUMBE R OF ANIMAL S NECROPSIED
 

57
 
TABLE A2 FEMALE RATS: NEOPLASMS (CONTINUED) 

HIGH DOSE
 

(50)
 

(50)
 

11 (22X)
 

(U8)
 

3 (6%)
 

(37)
 

1 (3*)
 

1 <3X>
 

(50)
 

1 (2X)
 

3 (6X)
 

(50)
 

1 (2%)
 

1 (2%)
 7	 (1HX)
 

(47)
 

1 (2«)
 

(50)
 

(50)
 

1 J2S1­

•SHALL	 INTESTINE
 

LEIOMYOMA
 

URINAR Y SYSTEM
 

NONE
 

ENDOCRINE SYSTEM
 

•PITUITARY
 

CHROMOPHOBE ADENOM A
 

• ADRENA L
 

PIIPOCHROMOCYTOMA
 

»THYhOID
 

FOLLICULAR-CELL CARCINOMA
 

e-CPLL ADENOMA
 

REPRODUCTIVE SYSTEM
 

*MAMMAR Y .rtLAND
 

ADENOMA, NOS
 

INTRADUCTAL PAVILLOMA
 

INTRADUCTAL CARCINOMA
 

FIBROADENOMA
 

IUTERIJ5
 

ADENOCAKCINOMA,
 

LL.IOMYOSARCCMA
 

ENDOMETBIAL STROMAL POLYP
 

»OVAR Y
 

CYSTADENOflA,
 

NERVOU S SYSTEM
 

•CEREBRUM
 

GLIOMA,
 

SPECIAL SENSE ORGANS
 

*EAP LOBULE
 

Ni.UROFIBRO»1A
 

CONTROL (UNTF)
 

(20)
 

(19)
 

6(32%)
 

(20)
 

1 (5X)
 

(11)
 

(20)
 

1 (5%)
 

14 (20X)
 

(20)
 

5 (25%)
 

(18)
 

(19)
 

1 (5%)
 

(20)
 

LOW DOSE
 

(50)
 

1 (2S)
 

(1»8)
 

10 (2U)
 

(U9)
 

1 (2X)
 

(36)
 

(50)
 

1 (2X)
 

1 (6X)
 

(50)
 

6 (12«)
 

(<«9)
 

(50)
 

(50)
 

* NUMBE R OF ANIMAL S WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMAL S NECROPSIEP
 

58
 
TABLE A2 FEMALE RATS: NEOPLASMS (CONTINUED)

 

CONTROL (UNTR) LOW DOSE HIGH DOSE
 

MUSCULOSKELETAL SYSTEM
 

NONE
 

BODY CAVITIES
 

NONE
 

ALL OTHER SYSTEMS
 

NONE
 

ANIMAL DISPOSITION SUMMARY
 

ANIMALS INITIALLY IN STUDY
 

NATURAL DEATHS
 

MORIBUND SACRIFICE
 

SCHEDULED SACRIFICE
 

ACCIDENTALLY KILLED
 

TERMINAL SACRIFICE
 

ANIMAL MISSING
 

d INCLUDES AIITOLYZED ANIMALS
 

TUMOR SUMMAR Y
 

TOTAL ANIMALS KITH PRIMARY TUMORS*
 

TOTAL PRIMARY TUMORS
 

TOTAL ANIMALS WITH BENIGN TUMORS
 TOTAL BENIGN TUMORS
 

TOTAL ANIMALS WITH MALIGNANT TUMORS
 

TOTAL MALIGNANT TUMORS
 

TOTAL ANIMALS WITH SECONDARY TUMORS
 

TOTAL SECONDARY TUMORS
 

TOTAL ANIMALS WITH TUMORS UNCERTAIN
 

HENIGN OR MALIGNANT
 

TOTAL UNCERTAIN TUMORS
 

TOTAL ANIMALS WITH TUMORS UNCERTAIN
 PRIrtAR V OR HFTASTATIC
 

TOTAL UNCERTAIN TUMORS
 

20 50 50 

2 2 1 

3 <l 2 

15 114 47 

1U 28 27 

20 38 35 

13 22 27 

17 27 31 

2 9 '1 

3 10 4 

« 

. 

1 

1 

_ 

« PRIMAR Y TDHOSS: ALt TUMORS EXCEPT SECONDARY TUMORS
 

» SECuNDAPV TUMORS: BETASTATIC TUMORS OR TUMOKS INVASIVE INTO A M ADJACENT OKGAH
 

59
 

APPENDIX B
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS
 

IN MICE FED EDTA IN THE DIET
 

61
 

TABLE B1

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE 

MICE FED EDTA IN THE DIET

 

CONTROL (UNTR) LOU DOSE
 

ANIMALS INITIALLY IN STUDY 20 50
 ANIMALS HISSING U
 ANIMALS NECEOPSIED 20 U6
 ANIMALS EXAMINED HISTOPATHOLOG ICA LLY 20 it 5
 

INTEGUHENTAPY SYSTEM
 

*SKIN (20) (46)
 

NL'UROPIBROMA
 

RESPIRATORY SYSTEM
 

UUNG (18) <<»<•)
 

A1.VEOLAR/BRONCHIOLAR ADENOMA 2 (11*) 5 (11X)
 

ALVEOLAR/BEONCHIOLAR CARCINOMA 3 <7X)
 

HEMATOPOIETIC SYSTEM
 

*MULTIPLE ORGANS (20) (46)
 

MALIGNANT LYMPHOMA, 3 (7*)
 

MALIGNANT LYMPHOMA, HISTIOCYTIC 1 <2X)
 

LDIIKEMIA.NGS
 

*MEDIASTINUM (20) (U6)
 

MALIGNANT LYMPHOMA,
 

MALIGNANT LYMPHOMA, LYMPHOCYTIC 1 (5*)
 

ISPLtEN (18) (39)
 

HtMANGIOMA 1 (3*)
 

MALIGNANT LYMPHOMA, 1 (6*) 1 (3<)
 

IMESENTERIC L. NODE (18) (38)
 

MALIGNANT LYMPIIONA, 1 (3«)
 

•SMALL	 INTESTINE (19) (11)


MALIGNANT LYMPHOMA, 1 (2«)
 

CIRCULATORY SYSTEM
 

_ NQNE _
 

« NUMBTR OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMUES OF ANIMALS NECROPSIED
 

HIGH DOSE
 

50
 

2
 

US
 

U0
 

C*8)
 

1 (2*)
 

(«5)
 

11 (24()
 

1 <2X)
 

(08)
 

5 (10»)
 

1 <2X)
 

(«8)
 

1 (2X)
 

(42)
 

1 (2X)
 

(42)
 

(17)
 

63 
TABLE B1 MALE MICE: NEOPLASMS (CONTINUED) 

HIGH DOSE
 

(47)
 

7 (15X)
 

3 (6X)
 

(U8)
 

1 <2X)
 

(t6)
 

(26)
 

1 (tX)
 

(33)
 

1 (3X)
 

(«H)
 

(HU)
 

1 <2X)
 

DIGESTIVE SYSTEM
 

tLIVEP
 

IIEPATOCELLULAR ADENOMA
 

HEPATOCELLULAR CARCINOMA
 

*ANUS
 

SQUAMOUS CELL CARCINOMA
 

URINARY SYSTEM
 

»KIDNEY
 

TUBULAR-CELL ADENOCAHCINOM A
 

ENDOCRINE SYSTEM
 

•PITUITARY
 

CHROMOPHOBE ADENOMA
 

ITHYROID
 

FOLLICULAR-CELL ADENOMA
 

tPANCREATIC ISLETS
 

ISLET-CELL ADENOMA
 

REPRODUCTIVE SYSTEM
 

#TESTIS
 

INTERSTITIAL-CELL


NERVOUS SYSTEM
 

NONE
 

SPECIAL SENSE ORGANS
 

NONE
 

 TUMOR
 

CONTROL (UNTR)
 

(19)
 

2 (11X)
 

1 (5*)
 

(20)
 

(19)
 

1 <5X)
 

(13)


1 (8X)
 

(10)
 

(19)
 

(19)
 

1 (5X)
 

LOW DOSE
 

<UI»)
 

9 (20%)
 

1 (2*)
 

(U6)
 

(43)
 

(19)
 

(29)
 

1 OX)
 

(38)
 

1 (3X)
 

(l»2)
 

MUSCULOSKELETAL SYSTEM
 

HON£ _ _ _
 

* NUMBFR OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

64
 
TABLE B1 MALE MICE: NEOPLASMS (CONTINUED)

 

CONTROL (UNTR) LOW DOSE HIGH DOSE 

BODY CAVITIES 

NONE 

ALL OTHER SYSTEMS 

NONE 

ANIMA L DISPOSITION SUMMARY 

ANIMALS INITIALLY IN STUDY 

NATURAL DEATHS 

MORIBUND SACRIFICE 

SCHEDULED SACRIFICE 

ACCIDENTALLY KILLED 

TtRMINAL SACRIFICE 

ANIMAL MISSING 

20 

2 

1 

17 

50 

10 

1 

1 

3U 

U 

50 

f> 

1 

«1 

2 

3 INCLUDES AUTOLYZED ANIMALS 

TUMOR SUMMAR Y 

TOTAL ANIMALS WITH PRIMARY TUMORS* 

TOTAL PRIMARY TUMORS 

9 

10 

2U 

28 

27 

35 

TOTAL ANIMALS WITH BENIGN TUMORS 

TOTAL BENIGN TUMORS 

5 

6 

15 

17 

18 

22 

TOTAL ANIMALS WITH MALIGNANT TUMORS 

TOTAL MALIGNANT TUMORS 

14 

U 

11 

11 

13 

13 

TOTAL ANIMALS WITH SECONDARY TUMORS* 

TOTAL SECONDARY TUMORS 

TOTAL ANIMALS WITH TUMORS UNCERTAINBENIGN OR MALIGNANT 

TOTAL UNCFRTA1N TUMORS 

TOTAL ANIMALS WITH TUMORS UNCERTAINPRIMARY OP METASTATIC 

TOTAL UNCERTAIN TUMORS 

* PRIMARY TUMORS: ALL TUMORS EXCEPT SE COHDARY TUMORS t SECONDARY TUMORS: METASTATIC TUHOSS OR TUMORS INVA: 3IVB INTO AN A l QJACENT ORGAN
 

65
 
TABLE B2

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE 

MICE FED EDTA IN THE DIET 

CONTROL (UNTR) LOW DOSE HIGH DOSE
 

ANIMALS INITIALLY IN STUDY 20 50 50
 ANIMALS MISSING 1 3
 ANIMALS NECKOPSIED 19 M9 M7
 ANIMALS EXAMINED HISTOPATHOLOG ICA LLY 19 US U7
 

INTEGUMENTARY SYSTEM
 

NONE.
 

RESPIKATOHY SYSTEM
 

ILUNG (19) (l»7) <»5)
 

ALVEOLAR/BRONCHIOLAH ADENOMA 3 <6») H (9*)
 

HEMATOPOIETIC SYSTEM
 

*MULIIPLE ORGANS (19) C»9) (<|7)
 

MALIGNANT LYMPHOMA, 5 (26X) 9 (18X) 9 (19%)
 

MALIGNANT LYMPHOMA, HISTIOCYTIC 1 (2X)
 

LEUKEBIA.NOS 1 (5%) 1 (2X)
 

•SPLEEN	 (19) (U6) («6)
 

HEMANGIOMA 1 (5*)
 

MALIGNANT LYMPHOBA, 1 (2*) 2 (<»*)
 

*LTVER (19) («6) (")
 

MALIGNANT LYMPHOMA, 1 (2«)
 

CIRCULATORY SYSTEM
 

NONE
 

CIGESTIVE SYSTEM
 

ILIVEF (19) («6) (17)
 

HEPATOCELLULAR ADENOMA 1 (2») 1 (2X)
 

ICECUM (19) («3) (46)
 _ _LJI2MI05i££Oj!i_ ._ 1 J5J1
 

* NUMBER OF ANIMALS KITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

66
 
TABLE B2 FEMALE MICE: NEOPLASMS (CONTINUED) 

HIGH DOSE
 

(17)
 

(42)
 

(29)
 

1 (14*)
 

(44)
 

(34)
 

1 <3X)
 

(43)
 

(46)
 

URINARY SYSTEM
 

(KIDNEY
 

TUBULAR-CELL ADENOCARCINOM A
 

tURINAR Y BLADDER
 

HEMANGIOMA
 

ENDOCRINE SYSTEM
 

(PITUITARY
 

CHROMOPHOBE ADENOMA
 

tADRENAL
 

CORTICAL ADENOMA
 

ITHYEOID
 

FOLLICULAR-CELL ADENOMA
 

FOLLICULAH-CELL CARCINOMA
 

REPRODUCTIVE SYSTEM
 

tOVARY
 

CYSTADENOMA,
 

NERVOUS SYSTEM
 

•BRAIN
 

MENINGIONA
 

SPECIAL SENSE ORGANS
 

NONE
 

MUSCULOSKELETAL SYSTEM
 

NONE
 

BODY CAVITIES
 

NONt
 

CONTROL (IJNTH) LOW DOSE
 

(11)	 <«7)
 

1 (5X)
 

(17)	 (38)
 

1 (6%)
 

(12)	 (3U)
 

2 (17X) 6 (1BX)
 

(19)	 (<*1)
 

1 (2*)
 

(12) (33)
 

3 (9X)
 

1 (8X)
 

(16)	 C»2)
 

1 (2*)
 

(18)	 (t7)
 

1 (2X)
 

»	 NUHBPR OF ANIMALS HITH TISSUE EXAMINED MICROSCOPICALLY
 

• NUMBER OF ANIMALS NECROPSIED
 

67
 
TABLE B2 FEMALE MICE: NEOPLASMS (CONTINUED) 

CONTROL (UNTR) LOW DOSE HIGH DOSE
 

ALL OTHER SYSTEMS
 

NONE
 

ANIMA L DISPOSITION SUMMAR Y
 

ANIMALS INITIALLY IN STUDY
 

NATURAL DEATHS
 

MORIBUND SACRIFICE
 

SCHEDULED SACRIFICE
 

ACCIDENTALLY KILLED
 

TERMINAL SACRIFICE
 

ANIMAL MISSING
 

S> INCLUDES AUTOLYZED ANIMALS
 

TUMOR SUMMAR Y
 

TOTAL ANIMALS WITH PRIMARY TUMORS*
 

TOTAL PRIMARY TUMORS
 

TOTAL ANIHALS WITH BENIGN TUMORS
 

TOTAL BENIGN TUMORS
 

TOTAL ANIMALS WITH MALIGNANT TUMORS
 

TOTAL MALIGNANT TUMORS
 

TOTAL ANIMALS WITH SECONDARY TUMORS*
 

TOTAL SECONDA8Y TUMORS
 

TOTAL ANIMAL S WITH TUMORS UNCERTAINBENIGN OR MALIGNANT
 

TOTAL UNCERTAIN TUMORS
 

TOTAL ANIMALS WITH TUMOBS UtlCEHTAIN-


PBIMARY OR METASTATIC
 

TOTAL UNCERTAIN TUMORS
 

20 50 50 

2 7 6 

3 3 11 

1U 140 37 1 3 

12 24 23 

13 27 23 

i| 11 10 

4 15 10 

9 12 13 

9 12 13 

» PRIMAR Y TUMORS: ALL TUMORS EXCEPT SECONDARY TUMORS
 

~t SECONDARY TUMORS: METASTATIC TUMORS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN
 

68
 
APPENDIX C
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN RATS FED EDTA IN THE DIET
 

69
 

TABLE C1 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN MALE RATS FED EDTA IN THE DIET 

CONTROL (UNTB) LOU DOSE
 

ANIMALS INITIALLY IN STUDY 20 50
 

ANIMALS NECROPSIED 20 50
 

ANIMALS EXAMINED HISTOPATHOLOGICALLY 20 50
 

INTEGUMENTARY SYSTEM
 

*SKIN (20) (50)
 

EPIDERMAL INCLUSION CYST 1 (2*)
 

FIBBOSIS
 

BESPIRATOBY SYSTEM
 

ILUNG (1<t) (50)
 

INFLAMMATION, INTERSTITIAL 2 (11%) 1 (2X)
 

PNEUMONIA, CHRONIC HUBINE 2 ("»«)
 

HYPEBPLASIA, ADENONATOUS
 

HYPERPLASIA, ALVEOLAR EPITHELIUM 1 (2*)
 

HSBATOPOIETIC SYSTEM
 

IUOHE HARROW (20) C»6)
 

HYPERPLASIA, GRANULOCYTIC
 

ISPLEEN (20) C*8)
 

FIBBOSIS, FOCAL 1 (2*)
 

PIBB03IS, DIFFUSE 1 (5*)
 

HYPEBPLASIA, LYNPHOID 1 (2«)
 

IHANDIBULAR L. NODE (19) (US)
 

HYPERPLASIA, CYSTIC
 

HYPF.HPLASIA, PLASMA CELL
 

HYPERPLASIA, RETICULUH CELL 1 (5X) 1 (9«)
 

HYPERPLASIA, LYMPHOID 1 (2«)
 

IHESENTEBIC L. NODE (U) C«5)
 

ATROPHY,
 

HYPEBPLASIA, PLASMA CELL
 

JJ](EefieU5U*_£m£SlI,il]LfiUJ. 2_-UJSl fi_JU2l
 

* NUMBER OF ANIMALS KITH TISSUE EXAMINED MICROSCOPICALLY
 * NUMBER OF ANIMALS NECROPSIED
 

HIGH DOSE
 

50
 

50
 

50
 

(50)
 

1 (2%)
 

(U9)
 

2 (»%)
 

1 (2«)
 

1 (2X)
 

(i«6)
 

1 <2X)
 

("»9)
 

(U6)
 

1 (2%)
 

2 (4X)
 

2 (<*X)
 

1 (2X)
 

(U6)
 

1 (2X)
 

1 <2X)
 

Jl_J2Si_
 

71
 
TABLE C1 MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

HIGH DOSE
 

(6)


1 (17*)
 

(19)
 

1 <2X)
 

(19)
 

1 <2X)
 

18 (37X)
 

(50)
 

1 (2X)
 

1 (2X)
 

2 (H%)
 

1 <2X)
 

(18)
 

1 (2X)
 

1 (2X)
 

(18)
 

2 (IX)
 

1 <2X)
 

(19)
 

(19)
 

(19)
 

(19)


1 <2X)
 

2 <4X)
 

(15)
 

6 J13JL
 

ITHYMUS
 

HYPERPLASIA, RETICULUM CELL
 

HYPERPLASIA, LYMPHOID
 

CIRCULATORY SYSTEM
 

tHEART
 

PEEIABTERITIS
 

•MYOCARDIUM
 

INFLAMMATION, FOCAL
 

PIBROSIS
 

DIGESTIVE SYSTEM
 

ILIVER
 

CIRRHOSIS,
 

NECROSIS, DIFFUSE
 

METAMORPHOSIS FATTY
 BASOPHILIC CYTOPLASM ALTERATION
 

(PANCREAS
 

FIBROSIS, DIFFUSE
 

ATROPHY,
 

ATROPHY, FOCAL
 

IPANCPRATIC ACINUS
 

ATROPHY,
 

ATROPHY, FOCAL
 

ISTOMACH
 

INFLAKMATICN.
 

»SMALL INTESTINE
 

HYPERPLASIA, LYMPHOID
 

*DUODENUM
 

HYPERPLASIA, EPITHELIAL
 

*ILEUM'
 

INFLAMMATION,
 

HYPERPLASIA, LYMPHOID
 

#LAR*GE INTESTINE
 

NEMA.TODIASIS __ __ _
 

CONTROL (UNTR)
 

(1)
 

1 (100X)
 

(18)
 

(18)
 

4(22%)
 

(20)
 

(19)
 

(19)
 

(20)
 

(20)
 

1 (5%)
 

(20)
 

(20)
 

(16)
 

.-I 16X)
 

LOH DOSE
 

<fr)
 

(«8)
 

1 12X)
 

(H8)
 

10 (21X)
 

(K8)
 

U (8X)
 

5 (10X)
 

<<»7)
 

1 (2%)
 

1 (2%)
 

(«)
 

1 (2«)
 

1 (2X)
 

(50)
 

1 (2X)
 

(50)
 

(50)
 

1 (2«)
 

(50)
 

3 (6X)
 

(M8)
 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBES OP ANIMALS NECROPSIED
 

72
 
TABLE C1 MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL (UNT8) LOW DOSE
 

•COLON	 (16) (48)
 

FIBROSIS, DIFFUSE 1 (6X)
 

HYPERPLASIA, LYHPHOID <» <8X)
 

UBINABY SYSTEM 

•KIDNEY 

PYELONEPHRITIS, 

INFLAMMATION CHBONIC 

NEPHROPATHY 

(20) 

11 (55X) 

(49) 

1 (2*) 

24 (49X) 

2 <4X) 

•URINARY BLADDER 

INFLAMMATION, (15) <<H> 

1 <2X) 

ENDOCRINE SYSTEM
 

•ADRENAL CORTEX (20) (49)
 

METAMORPHOSIS FATTY 1 (2X)
 

MPOIDOSIS 1 (2X)
 

•ADRENAL	 MEDULLA (20) (49)
 

HYPEF.PLASIA, NODULAB 1 (2X)
 

•THYBOID	 (17) (35)
 

HYPERPLASIA, C-CELL
 

REPRODUCTIVE SYSTEM
 

*PBEPUTIAL GLAND (20) (50)
 

ABSCESS, 1 <2X)
 

•PROSTATE	 (17) <U5)
 

INFLAMMATION, 2 <4X)
 

HYPERPLASIA, EPITHELIAL 1 (6*)
 

NERVOUS SYSTEM
 

•CEREBRUM	 (20) (50)
 

HC.MORRHAGIC CYST	 1 (2X)
 

•BRAIN	 (20) (50)
 

• NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

« NUMBER OF ANIMALS NECROPSIED
 

HIGH DOSE
 

(15)
 

(50)
 

31 (62X)
 

1 (2X)
 

(36)
 

(50)
 

4 (8X)
 

(50)
 

(38)
 

1 <3X)
 

(50)
 

(48)
 

2 (4X)
 

(50)
 

(50)
 

73
 
TABLE C1 MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED)

 

NECROSIS,
 

SPECIAL SENSE ORGANS
 

NONE
 

BUSCULOSKELETAL SYSTEM
 

NONE
 

BODY CAVITIES
 

•HEStNTEHY
 

NECROSIS, FAT
 

ALL OTHER SYSTEMS
 

•MULTIPLE ORGANS
 

LEIIKENOID REACTION
 

SPECIAL MORPHOLOGY SUMMARY
 

NO LESION REPORTED
 

CONTROL (UNTS) LOW DOSE HIGH DOSE
 

1 (2%)
 

(20) (50) (50) 

1 (2*) 

(20) (50) (50) 

2 <U») 

2 

74 
TABLE C2 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN FEMALE RATS FED EDTA IN THE DIET 

HIGH DOSE
 

50
 

50
 

50
 

(16)
 

1 <2X)
 

C»8)
 

1 (2«)
 

a <sx)
 

1 (2X)
 

(1.9)
 

1 <2X)
 

C«7)
 

1 (2X)
 

(«7)
 

1 (2*)
 

1 (2»)
 

6 (13X)
 

3 (6«)
 

(17)


1 (2X)
 

8 (17X)
 

J_JUS.1_.
 

ANIMALS INITIALLY IN STUDY
 

ANIMALS NECBOPSIED
 

ANIMALS EXAMINED HISTOPATHOLOGICALLY
 

INTEGUMENTARY SYSTEM
 

NONE
 

RESPIRATORY SYSTEM
 

*TRACHEA
 INFLAMMATION CHRONIC
 

tLUNG
 

INFLAMMATION, INTERSTITIAL
 

PNEUMONIA, CHRONIC HURINE
 

HYPERPLASIA. EPITHELIAL
 

HYPFRPLASIA, ALVEOLAR EPITHELIUM
 

HEHATOPOIETIC SYSTEM
 

•BONE MARROH
 

HYl'ERPLASIA,
 

HYPERPLASIA, GSANULOCYTIC
 

•LYMPH	 NODE
 

HYPERPLASIA, RETICULUM CELL
 

IMANDIBULAR L. MODE
 

INFLANHATICN CHRONIC
 

HYPERPLASIA, PLASMA CELL
 

HYPERPLASIA, RETICULUfl CELL
 

HYPERPLASIA, LYMPIIOID
 

IMESKNTEHIC L. NODE
 

HYPERPLASIA, CYSTIC
 

HYPERPLASIA, HETICULUM CELL
 

flXEEBm3IA*_..lIflPJQID
 

CONTROL (UNT8)
 

20
 

20
 

20
 

(19)
 

(20)
 

1 (5X)
 

3 (15*)
 

1 (5*)
 

(19)
 

1 <5«)
 

(20)
 

(20)
 

1 (5*1
 

(20)
 

1.JSS1
 

LOW DOSE
 

50
 

50
 

50
 

(50)
 

(H8)
 

2 (1%)
 

6	 (13X)
 

(50)
 

1 UX)
 

C»6)
 

(16)
 

1 <2X)
 

(U6)
 

1 (2X)
 

2 <<•*)
 

J_Ji«l
 

« NUMBER OP ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OP ANIMALS NECHOPSIED
 

75
 
TABLE C2 FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED)

 

CONTROL (UNTE) LOW DOSE
 

CIRCULATORY SYSTEM
 

•HEART	 (19) (16)
 PERIARTEEITIS 1 (2X)
 

•MYOCARDIUM	 (19) (16)
 

FIBPOSIS 2 (11X) « (9*)
 

DIGESTIVE SYSTEM
 

•LIVER	 (20) (19)


METAMOBPHOSIS FATTY 1 (2X)
 

EASOPHILIC CYTOPLASM ALTERATION 2 (10*) 13 (27*)
 

• LIVER/CENTRILOBU1.AR (20) (19)
 

NECROSIS. 1 <5X)
 

•BILE DUCT (20) (50)
 HYPERPLASIA, 1 <2X)
 

•PANCREAS	 (18) (18)'
 

FIBROSIS. FOCAL 1 (6X) 1 (2X)
 

ATROPHY, FOCAL 1 (2X)
 

•PANCSEATIC	 DUCT (18) (18)
 

DILATATION,
 

•PANCREATIC	 ACINUS (18) (18)
 

FIBROSIS, FOCAL
 

ATROPHY, 2 (IX)
 

ATROPHY, FOCAL
 

•SMALL	 INTESTINE (20) (50)


HYPERPLASIA, LYNPIIOID 1 (2X)
 

•ILEUM	 (20) (50)


HYPERPLASIA, LYMPHOID	 2 (IX)
 

•LARGE	 INTESTINE (19) (16)
 

NEMATODIASIS 7 (37X)
 

• COLON	 (19) (16)
 

» NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMAL S NEC30PSIED
 

76
 

HIGH DOSE
 

<«1)
 

(11)


6	 (14X)
 

(19)


1 (2X)
 

15	 (31X)
 

(19)
 

(50)


1 (2X)
 

(19)
 

(19)
 

1 (2X)
 

(19)


1 <2X)
 

1 (2X)
 

(50)
 

(50)


1 (2X)
 

(18)
 

(18)
 
TABLE C2 FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED)

 

CONTROL (IINTS) LOW DOSE 

URINARY SYSTEM
 

#KIDNEY (20) (50)


HYDRONEPHROSIS 1 (2X)
 

INFLAMMATION, INTERSTITIAL 1 (2X)
 

INFLAMMATION CHRONIC 2 (10X) 5 (10X)
 

INFLAMMATION, GRANULOMATOUS 1 (2%)
 

HYPEBPLASIA, TUBULAR CELL 1 (5%)
 

ENDOCRINE SYSTEM
 

•PITUITARY (19) («8)
 

CYST, « (8%)
 MULTIPLE CYSTS 1 (2*)
 

HEMORRHAGIC CYST 1 <2X)
 

ANGIECTASIS 1 (5X)
 

tADRENAL (20) (t9)
 

CYST. 1 (2X)
 

HEMORRHAGE 1 (5%)
 

HEMORBHAGIC CYST 1 (2X)
 

METAMORPHOSIS FATTY 1 (5X)
 

LIPOIDOSIS 1 <2X)
 

*ADRENAL CORTEX (20) («9)
 

METAMORPHOSIS FATTY
 

LIPOIDOSIS 1 (5X) 1 (2X)
 

IIYtERPLASIA, 1 (2X)
 

HYPF8PLASIA, FOCAL 1 (2X)
 

ITHYEOID (11) (36)
 

HYPERPLASIA, C-CELL 1 (9*) 1 O*)
 

IPANCREATIC ISLETS (18) (t8)
 

HYPERPLASIA,
 

REPRODUCTIVE SYSTEM 

tUTEBUS 

CYST, 

PYOMETRA 

FIBROSIS 

(20) 

1 (5*) 

(50) 

1 (2X) 

tCERVIX UTERI 

-CYST* __. 

(20) 

1 J521 _. (50) 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

77
 

H I G H DO S E
 

(48)
 

1 (2X)
 

(50)
 

1 (2*)
 

1 (BX)
 

C«8)
 

(U8)
 

1 (2X)
 

1 (2X)
 

(37)
 

(1*9)
 

1 (2X)
 

(50)
 

1 <2X)
 

1 (2X)
 

(50)
 
TABLE C2 FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED)

 

ABSCESS,
 

IUTEKUS/ENDOMF.TRIOM
 

INFLAMMATION,
 

INFLAMMATION, SOPPURATIVE
 

INFLAMMATION, VESICULAR
 

INFLAMMATION, CHRONIC SDPPOBATIV
 

*OVAKY/OVIDUCT
 

INFLAMMATION,
 

tOVARY
 CYST,
 

FIBHOSIS
 

PIGMENTATION,
 

NERVOUS SYSTEM
 

#BRAIN
 

HYDROCEPHALOS,
 

PERIVASCULAH CUFFING
 

SPECIAL SENSE ORGANS
 

NONE
 

MUSCULOSKELETAL SYSTEM
 

NONE
 

BODY CAVITIES
 

NONE
 

ALL OTHER SYSTEMS
 

NONE
 

SPECIAL MORPHOLOGY SUMMARY
 

NO LESION EEPORTED
 

CONTROL (UNTR)
 

(20)
 

3 (15*)
 

(20)
 

1 (5X)
 

(18)
 

(19)
 

1 <5X)
 

LOW DOSE
 

1 (2X)
 

(50)
 

3 (6X)
 

U (8X)
 

3 (6X)
 

1 (2)1)
 

(50)
 

1 <2X)
 

(49)
 

3 (6X)
 

(50)
 

1 (2X)
 

3
 

HIGH DOSF
 

(50)
 

3 (6X)
 

3 (6X)
 

1 (2X)
 

1 (2X)
 

(50)
 

(17)
 

5 (11X)
 

1 (2X)
 

1 (2X)
 

(50)
 

1 (2X)
 

1 <2X)
 

4
 

78
 
APPENDIX D
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC
 

LESIONS IN MICE FED EDTA IN THE DIET
 

79
 

TABLE D1 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN MALE MICE FED EDTA IN THE DIET 

CONTROL (UNTR) LOW DOSE HIGH DOSE
 

ANIMALS INITIALLY IN STUDY 20 50 50
 

ANIMALS MISSING It 2
 

ANIMALS NECROPSIEI) 20 U6 18
 

ANIMALS EXAMINED IIISTOPATHOLOGICALLlf 20 as 48
 

INTEGUMENTARY SYSTEM
 

NONE
 

RESPIRATORY SYSTEM
 

II LUNG (18) (<ti») (i»5)
 EDEMA, 1 (2*)
 PNEUMONIA, CHRONIC HUBINE 2 <5X) 5 (11*)
 PERIARTERITIS 1 (6%)
 IIYPERPLASIA, FOCAL 1 <2X)
 LYMPHOCYTOSIS 1 (2X)
 

HEMATGPOIETIC SYSTEM 

IBONE MARROW (19) (36) («5) HYPERPLASIA, HEN ATOPOIETIC 1 (3«> 

*SPLEEN 

AMYLOIDOSIS 

(18) (39) 

1 <3X) 

C<2) 

IIYPERPLASIA, LYMPHOID 1 (2X) 

»SPLENIC CAPSULE (18) (39) (<*2) FIBROSIS 1 (2X) 

ILYMPII NODE (18) (38) C42) HYPERPLASIA, 2 (5*) 

IflANDIBULAR L. NODE (18) (38) <t2) HYPERPLASIA, RETICULUM CELL 1 (2X) 

IMESENTERTC L. NODE (18) (38) (U2) 

_

HEMORRHAGE 

 INFLAMMATION., _ . _ 1 J6«l_ _. 

1 <3X) .._ _J_J2J1__ 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS HECROPSIED
 

81 
TABLE D1 MALE MICE: NONNEOPLASTIC LESIONS (CONTINUED)

 

CONTROL (UNTR) LOU DOSE HIGH DOSE
 

HYPERPLASIA. 1 <6<) 1 (3X) 1 (2*)
 

HYPEBPLASIA, RETICULUM CELL 1 (2X)
 

CIRCULATORY SYSTEM
 

•HEART	 (19) («3) C*5)
 

MINERALIZATION 1 (2%)
 

•MYOCARDIUM	 (19) (U3) <t5)
 

INFLAMMATION, FOCAL 1 (2X)
 

FIBROSIS, FOCAL 1 (2X)
 

•PULMONARY	 ARTERY (20) <«6) (U8)
 

HYPERTROPHY, 1 (2*)
 

DIGESTIVE SYSTEM
 

•LIVER	 (19) (i»lt) (U7)
 

INFLAMMATION. FOCAL 1 (5%) 1 (2X)
 

LYHPHOCYTIC INFLAH INFILTRATE 1 (2«)
 

INFLAMMATION CHRONIC 1 <2X)
 

NECROSIS, 2 (5X) 1 (2*)
 

MLTAMOHPHOSIS FATTY 1 (5%)
 

HYPERPLASIA, NODIILAH 1 (2*)
 

HYPERPLASIA, FOCAL 1 (2%)
 

ANGIECTASIS 2 (5*) 1 <2X)
 

LYMPHOCYTOSIS 1 <2X)
 

*GALLDLADDER (20) C»6) (US)
 

INFLAMMATION, VESICULAR 1 (2«)
 

•PANCREAS	 (19) (38) (DM)
 

NECROSIS, 1 (2X)
 

•PANCREATIC ACINUS (19) (38) (UU)
 

ATROPHY, FCCAL 1 (3*)
 

•STOMACH	 (19) («D (U7)
 

INFLAMMATION, 1 (2*)
 

INFLAMMATION, FOCAL 1 (5%)
 

tSMALL INTESTINE (19) (U1) (47)
 

AUSCESS, 1 (2*)
 

_A«-YLO!fiOS!S __ __ __ J_J251_ _._„ —
 

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

82
 
TABLE D1 MALE MICE: NONNEOPLASTIC LESIONS (CONTINUED)

 

CONTROL (UNTB) LOH DOSE
 

HYPEfcPLASIA, LYMPIIOID
 

IlHIMAfcY SYSTEM
 

•KIDNEY	 (19) (U3)
 

HYDRONEPHROSIS
 

PYELONEPHRITIS, 1 (2X)
 

SCAB 1 (2*)
 

AMYI.OIDOSIS 1 (2X)
 

METAPLASIA, OSSEOUS 1 (2*)
 

(URINARY BLADDER (19) (38)
 

INFLAMMATION, 1 (3*)
 

INDOCRINE SYSTEM
 

•ADRtNAL	 (17) (36)
 

AMYLOIDOSIS 1 <3X)
 

•THYKOID	 (10) (29)
 

HYPERPLASIA, FOLLICUL AR -CELL 2 (7*)
 

REPRODUCTIVE SYSTEM
 

•PROSTATE	 (19) (39)
 

INFLAMMATION, 1 (3*)
 

NERVOUS SYSTEM
 

•BRAIN	 (19) (U2)
 

INFLAMMATION, CHRONIC FOCAL
 

SPECIAL SENSE ORGANS
 

NONE
 

MUSCULOSKELETAL SYSTEM
 

NONE	 __ ...
 

» NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 « NUMtER OP ANIMALS NECROPSIEI)
 

HIGH DOSE
 

1 (2X)
 

(16)
 

1 (2X)
 

1 (2X)
 

(11)
 

(HI)
 

(33)
 

<* (12X)
 

C*1)
 

(U6)
 

1 <2X)
 

83
 
TABLE D1 MALE MICE: NONNEOPLASTIC LESIONS (CONTINUED)

 

BODY CAVITIES
 

'MEDIASTINUM
 

THROMBOSIS,
 

ALL OTHER SYSTEMS
 

NONE
 

SPECIAL MORPHOLOGY SUMMARY
 

NO LESION FEEORTED
 

ANIMA L MISSING/NO NECROPSY PERP
 

AUTOLYSIS/NECROPSY PESF/NO HISTO
 

CONTROL (UNTR) LOU DOSE HIGH DOSE
 

(20) <«6) (H8)
 

1 (2X)
 

7 13 11*
 

a 2
 

1
 

84
 
TABLE D2 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC 

LESIONS IN FEMALE MICE FED EDTA IN THE DIET 

1IIUH DOSE
 

50
 

3
 

U7
 

47
 

(US)
 

1 (2%)
 

16 (36X)
 

(16)
 

1 (2«)
 

1 (2%)
 

1 (2«)
 

(15)
 

2 (4X)
 

(15)
 

1 (2*)
 

(15)
 

1 <2X)
 

1 <2X)
 

(17)
 

J.J2J1 .
 

ANIMALS INITIALLY IN STUDY
 

ANIMAL S MISSING
 

ANIMALS NECROPSIED
 

ANIMALS EXAMINE D HISTOPATHOLOG 1 CA LLV
 

INTECUMSHTARY SYSTEM
 

NONE
 

RESPIRATORY SYSTEM
 

KLUNli
 

INFLAMMATION, FOCAL
 

INFLAMMATICN, INTERSTITIAL
 

PNEUMONIA, CHRONIC MURINE
 

METAPLASIA, OSSEOUS
 

HEHATOPOIETIC SYSTEM
 

*SPLEEN
 

FIBPOSIS
 

INFAKCT,
 

AMYLOID,
 

•LYMPH	 NODE
 

TNFLAMMATICN,
 

HYPERPLASIA,
 

#MANUIBULAH L. NODE
 

AUSCESS,
 

#MEStNTERIC L. NODE
 

IHFLAMMATICN,
 

HYPERPLASIA, PETICOLUM CELL
 

CIRCULATORY SYSTEM
 

tHEAKT
 

_ .MINERALIZATION _ _ .
 

CONTPOL (UNTR)
 

20
 

1
 

19
 

19
 

(19)
 

1 (5*)
 

t (21S)
 

(19)
 

(18)
 

(13)
 

(18)
 

(19)
 

LOW DOSE
 

50
 

U9
 

US
 

(47)
 

1U (30%)
 

1 (2<)
 

(«6)
 

("")


1 (2K)
 

2 (5*)
 

(i»«)
 

(4U)
 

('•7)
 

1 Jim - ...
 

# NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBE R OF ANIMALS NECROPSIED
 

85
 
TABLE D2 FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED)

 

CONTROL (UNTH) LOU DOSE
 

•MYOCARDIUM	 (15) (l»7)
 

INFLAMMATION, 1 (2«)
 

*PULbONASY ARTERY (19) (<»S)
 

HYPERTROPHY, 1 (5%)
 

*3PLLNIC ARTEB Y (19) C«9)
 

DILATATION,
 

DIGESTIVE SYSTEM
 

•LIVKR	 (19) (Uf,)
 

INFLAMMATION, FOCAL
 

INFLAMMATIO N CHRONIC 1 (2*)
 

IIYPEKI'LASIA, NODULAR 1 (2«)
 

•PANCREAS	 (19) («6)
 

ATFOPIIY, 1 (2X)
 

•PANCREATIC	 ACINUS (19) («6)
 

CYSTIC DUCTS 1 (2X)
 

•STOrtACH	 (19) C*6)
 

INFLAMHATICN, ACUTE FOCAL
 

•GASTRIC	 SEROSA (19) (i»6)
 

INFLAMMATIO N CHRONIC 1 (2X)
 

•LARGE	 INTESTINE (19) C»3)
 

NEMATODIASIS 1 (5X)
 

URINAR Y SYSTEM
 

•KIDNEY	 (19) («7)
 

HYDRONEPHPOSIS 2 («X)
 

LiMPHOCYTIC INFLAH INFILTRATE 1 (2X)
 

ENDOCRINE SYSTEM
 

•ADRENAL CORTEX (19) (U1)
 

LIPOIDOSIS 1 (2X)
 

•THYhOID	 (12) 03)
 

HXPERPLASTA^ NODULAR__	 1 1331_
 

* NUM^fR OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMAL S NECROPSIEP
 

86
 

HIGII DOSE
 

(«7)
 

C<7)
 

<«7)
 

1	 (2X)
 

(«7)
 

1 (2S)
 

(«6)
 

(K6)
 

(U6)
 

1 (2*)
 

(«t)
 

(«6)
 

(tt7)
 

(i»i»)
 

(31)
 
TABLE D2 FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

HliJI I DOSE
 

5 (15*)
 

(Kb)
 

1 (2*)
 

(46)
 

14 (9X)
 

(i46)
 

1 (2X)
 

("3)
 

7 (16*)
 

(47)
 

1 (2*)
 

(147)
 

1-J2X)
 

I1YPERPLASIA, FOCAL
 

HYPERPLASIA, FOLLICULAR-CELI,
 

BEPROliUCTIVE SYSTEM
 

HUTEhUS
 

CYST,
 

*»T BE (IS/EN DOM ETKIUM
 CYST,
 

*OVAliY/OVIDIICT
 

INFLAMMATICH, SUPPU8ATIVE
 

ABSCESS,
 

»OVAEY
 

CYST,
 

NERVOUS SYSTEM
 

NONE
 

SPECIAL SENSE OBGANS
 

NONi
 

MUSCIILOSKELETAL SYSTEM
 

NONE
 

BODY CAVITIES
 

*MESENTE8Y.
 

NECROSIS, FAT
 

ALL OTHER SYSTEMS
 

•MULTIPLE OBGANS
 

PE.RIARTEDITIS
 

COHTEOL (UNTH)
 

3 (25*)
 

(19)
 

(1r*)
 

1 <5X)
 

(19)
 

(16)
 

2 (13X)
 

(19)
 

(19)
 

LOW DOSE
 

1 (3«)
 

7 (21*)
 

(16)
 

1 (2X)
 

(H6)
 

B (17X)
 

(146)
 

1 (2%)
 

<t2)
 

« (10X)
 

(1(9)
 

(19)
 

* NUrtbFR OF ANIMALS WITH TISSUE hXAMINED MICROSCOPICALLY
 * MI«L,E8 OF ANIMALS NECROPSIFD
 

87
 
TABLE D2 FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED)

 

CONTROL (UNTR) LOW DOSE HIGH DOSE
 22-2056 22-2051 22-2052
 

SPECIAL MORPHOLOGY SUMMARY
 

q
 NO LESION BEPORTED 3 6
 

ANIMA L MISSING/NO NECROPSY PEPF 1 2
 

AUTOLYSIS/NECROPSY PEKF/HO KISTO 1
 

AJTOLYSIS/NO NECROPSY PERFORMED 1
 

<r U. S. GOVERNMENT PRINTING OFFICE : 1977 241-161/3082 

88