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Lito

DISCLAIMER NOTICE

THIS DOCUMENT IS BEST QUALITYPRACTICABLE. THE COPY FURNISHEDTO DTIC CONTAINED A SIGNIFICANTNUMBER OF PAGES WHICH DO NOTREPRODUCE LEGIBLY.

AD

Mammalian ToxicoloqicalEvaluation

of nfIMP and DCPD(Phase 3 - IMPA) 0 7kE1T C

A ELECTEFINAL REPORT ELET8NOV 1 9 1981

Francis J. Mecler, Sc.f. HMay I181

Supported hyU.S. ARMY MEDICAL RESEARCH AND DEVELOPMENT COMMAND

Fort Detrick, Frederick, MD 21701

Contract No. DAMn 17-77-C-7003;

Contract Officer's Technical Representative: Dr. Jack C. nacreEnvironmental Protection Research Division

U.S. Army Medical Bioergineerinq Research and Development LaboratoryFort Detrick, Frederick, Maryland 217ni

Litton Bionetics, Inc.5516 Nicholson Lane

Kensinqton, Maryland 20795

Approved for public release; distribution unlimited

The findings in this report are not to he construed as an officialDepartment of the Armv position unless so desionated hw other authorized documents.

I

SECURITY CLASSIFICATION OF THIS PAGE (Wen Date Entered)REPORT DOCUMENTATION PAGE READ INSTRUCTIONS

BEFORE COMPLETING FORMI. REPORT NUMBER 2. GOVT ACCESSION NO. 3. RECIPIENT'S CATALOG NUMBER

4. TITLE (andSubtitle . TYP OF REPORT& PfRIOD Cf RED4. TILE (ed Suhfil.)Final Report - Phasei -

Mammalian Toxicological Evaluation of DIMP and IMPA, DEC ot-Apt 80

DCPD (Phase III - IMPA) ______DEC _________80

6. PERFORMING ORG. REPORT NUMBER

7. AUTHOR(aJ 8. CONTRACT OR GRANT NUMBER(e)

Francis J. Mecler, Sc.D. DAD17-77-C-7003

9. PERFORMING ORGANIZATION NAME AND ADDRESS 10. PROGRAM ELEMENT. PROJECT. TASKLitton Bionetics, Inc. AREA & WORK UNIT NUMBERS

5516 Nicholson Lane 62720A. 3E162720A835.AA.002

J Kensington, MD 20795

1,. CONTROLLING OFFICE NAME AND AoRESS 12. REPORT DATE

US Army Medical Research and Development Command May 1981Ft. Detrick, Frederick, MD 21701 ,s. NUMBER OF PAGES

14. MONITORING AGENCY NAME & ADDRESS(If different from ontrolling Office) 15. SECURITY CLASS. (of thia report)

US Army Medical Bioengineering Research Unclassified

and Development Laboratory ,S,. DECLASSIFICATION/DOWNGRADING

Ft. Detrick, Frederick, MD 21701 SCHEDULE

IS. DISTRIBUTION STATEMENT (Wf this Repot)

Approved for public release; distribution unlimited.

17. DISTRIBUTION STATEMENT (of the ebstatga entered In Block 20. it different ir, Report)

Approved for public release; distribution unlimited

18. SUPPLEMENTARY NOTES

19. KEY WORDS (Continue on reverse aide It necessary end Identiffy by block number)

IMPA eye irritation rats/mice/rabbits

toxicity skin sensitization Ames assayacute bacterial mutagenesis 90 day toxicity testsoral subchronic toxicity0 armalI

4(k AWS'TIAC' CCwhu aw revrs std Nf sa OM IdevutofF by block ntmlbor)

IMPA *had oral LD50's of 7650 and 6070 mg/kg in male and female rats, respectiveland of 5620 and 6550 mg/kg in male and female mice. It was not irritant torabbit eyes. It was mildly irritant to rabbit skin but produced no systemic

toxicity at a dose of 2 mg/kg. IMPA did not induce dermal sensitization inguinea pigs. No evidence of toxicity followed administration of IMPA in thedrinking water at levels up to 3000 ppm to rats for 90 days. IMPA was notmutagenic to five strains of Salmonella indicator organisms in the Ames assayboth with and without rat liver activation.

DO I' 147 EDITION OF I NOV 65 IS OBSOLETE

SECURITY CLASSIFICATION OF THIS PAGE (W en D es Entered)

EXECUTIVE SUMMARY

ISUPRUPYL METHYLPHOSPHONIC ACID - SODIUM SALT

The oral LD5O values of sodium IMP (sodium isopropyl methylphosphonate)were calculated to be 7650 (6560-8920) and 6070 (4760-7740) mg/kg inmale and female rats, respectively, and 5620 (4530-6990) and 6550 (5140-8360)ing/kg in male and female mice, respectively.

Application of sodium IMP to the intact and abraded skin of rabbitsdt doses of 2.0 g/kg produced no signs of systemic toxicity. Mildskin irritation was evident.

The Eye Irritation test revealed no signs of irritation in any of theraboits' eyes which were treated with sodium IMP.

Sodium IMP did not induce dermal sensitization in the guinea pig.

After 9U days of administration of IMPA in the drinking water at levelsof 300, 1000 and 3000 ppm, no changes in body weight, food intake, waterintake, clinical chemistry or hematologic parameters were seen in treatedrats compared to controls.

Sodium IMP did not exert a mutagenic effect in any of five strains ofSalmonella indicator organisms in the Ames Assay when tested with orwithout rat liver activation.

%p-

G 0.

FOREWORD

The studies reported herein were an addition to Contract No. DAMD 17-77-7003.Under this addition acute and subchronic toxicity testinq were authorizedon IMPA.

In conducting the research described in this report, the investiqation(s)adhered to the "Revised 1978 Guide for the Care and Use of LahoratoryAnimals, U.S.H.E.W., PHS, DHEW Publication No. (NIH) 78-23".

The methods Qf killing animals at study terminations were intravenousoverdosage of pentobarbital sodium for rabbits, inhalation of chloroformfor rats, mice and guinea pigs used in acute studies and inhalation ofcarbon dioxide for rats used in the suhchronic study.

E. Ross Hart, Ph.D. was the original Principal Investiqator until hisretirement in December, 1979. He was replaced by Cipriano Cueto, Jr., Ph.D.who left LBI in May, 1980. In May, 1980 Francis J. Mecler, Sc.D., hecamethe Principal Investigator.

Citations of trade names in this report does not constitute an off'cialDepartment of Army endorsement or approval of the use of such items.

Or.

TABLE OF CONTENTS

PAGE

IMPA (ISUPRUPYL METHYLPHOSPHONIC ACID) SODIUM SALT

EXECUTIVE SUMMARY .. ............ ............ I

FURLWURU .. ..... ............... ........ 2

SECTION A - INTRODUCTION AND MATERIAL. ...... ........ 5

SECTION B - ACUTE URAL TOXICITY STUDY IN RATS. ...... .... 6

SECTION C - ACUTE ORAL TOXICITY STUDY IN MICES .. ..... .... 9

SECTION 0 - ACUTE DERMAL TOXICITY STUDY IN RABBITS .. ....... 12

SECTION E - PRIMARY EYE IRRITATION STUDY IN RABBITS. ....... 15

SkLTIUN F - DERMAL SENSITIZATION STUDY IN GUINEA PIGS. ...... 21

SECTION G - MICROBIAL MUTAGENESIS .. ............ ... 31

SECTION H - 9U VAY TOXICITY STUDY IN RATS .. ........... 41

APPENDIX A - INDIVIDUAL ANIMAL TOXICITY OBSERVATIONS .... 57

APPEliDIX B - PATHOLOGY REPORT .. ..... ........... 143

APPENDIX C - CHEMICAL ANALYSIS REPORT .. ..... ...... 257

UISTkIbUTIUN LIST .. ............ ............ 276

003

LIST OlF TABLES

PAG1EIMPA (ISOPROPYL METHYLPHOSPHONIr ACID)

TABLE

E-1 SCORING FOR EYE IRRITATION. .. ............... 18

F-2 OBSERVATIONS .. ...... ................. 24

G-3 TABULATION OF REVERTANTS IN AMES ASSAY USING SODIUM

ISOPROPYL METHYPHOSPHONATE WITH AND WITHOUT ACTIVATION 33

G-4 TABULATION OF REVERTANTS IN AMES ASSAY USING SODIUJMISOPROPYL METHYLPHOSPHONATE WITH AND WITHOUT ACTIVATION-REPEAT TEST. ...... .................. 34

H-5 MEAN BODY WEIGHTS ................... 4

H-6 MEAN FOOD INTAKE .. ..... ................ 46

H-7 MEAN DAILY LIQUID INTAKE. .. ................ 47

H-8 COMPOUND INTAKE. ....... ............... 48

H-9 MEAN CLINICAL HEMATOLOGY. .. ............ .... 49

H-10 MEAN CLINICAL CHEMISTRY .. ............. .... 52

H-il MEAN ORGAN WEIGHTS .. ...... .............. 54

H-12 MEAN ORGAN WEIGHT/BODY WEIGHT PERCENTAGES. ......... 55

H-13 CLINICAL SIGNS .. ...... ................ 58

H-14 INDIVIDUAL BODY WEIGHTS .. ............. .... 85

H-i5 INDIVIDUAL DAILY FOOD INTAKE. .. .............. 93

H-16 INDIVIDUAL DAILY LIQUID INTAKE .. ...... ....... 101

H-17 CLINICAL HEMATOLOGY. ....... ............. 109

H-18 CLINICAL CHEMISTRY .. ...... .............. 122

H-19 INDIVIDUAL ORGAN WEIGHTS. .. ............. .... 127

H-20 INDIVIDUAL ORGAN WEIGHT/BODY WEIGHT PERCENTAGES .... 135

0(

SECTInN A

INTRODUICTION ANP MATERIAL

ISOPROPYL METHYLPHOSPHONIC ACID

1. INTRODUCTION

The material, isopropyl methylphosphonic acid (IMPA), was submittedas the sodium salt for toxicologic evaluation. This material is abreakdown product of diisopropyl methylphosphonate (DIMP) and as suchrequired assessment of potential toxic hazard. DIMP has been foundin ground water under the Federally owned Rocky Mountain arsenal.IMPA has been identified there as well.

Assessment included determination of potential for bacterial muta-qenesis, eye irritation, dermal sensitization, acute dermal toxicity,acute oral toxicity and subchronic toxicity in rats.

2. MATERIAL

The materials used in these studies were three batches of isopropylmethylphosphonic acid, sodium salt, supplied by the U.S. Army MedicalBioengineering Research and Development Laboratory. The first lotwas used for all acute studies and to start the subchronic study.The other two lots were used to finish the subchronic study. Forthe acute studies the material was used as received but for the sub-chronic study the material was corrected to express the concentra-tions as the free acid.

The dermal sensitization study utilized Eastman 1-chloro-2,4-dinitro-benzene Lot No. F7A as a positive control. This was purchased throuaha scientific supply house and used as a 0.1% solution.

3. PURITY ANALYSIS

Isopropyl methylphosphonic acid was provided as the sodium salt in threebatches from the US Army Medical Bioenqineerinq Research and DevelopmentLaboratory. The laboratory's claim of 99% purity hy oas chromatooraphyesterification and titration was accepted and the white crystalline saltwas used without further treatment.

005

SECTInN R

ACUTE ORAL TOXICITY STUDY IN RATS


LBI PROJECT NO. 10734-12

SUMMARY

The oral LD50 values of the test material, IMPA, were 7650 and 6070 mq/knfor male and female rats, respectively.

I. OBJECTIVE

The objective of this study was to evaluate the acute toxicityof the test compound when administered hy oral qavaqe to male andfemale rats.

2. MATERIAL

Two qlass jars (550 q each) containing a white powder and labeled:

Isopropyl Methylphosphonic Acid, Sodium SaltBatch 1 #127818

were received from U. S. Army Medical Research and Development Commandby Litton Bionetics, Inc. (LBI), on December 28, 1978, and designatedas LBI No. 3427.

3. EXPERIMENTAL DESIGN

Young adult rats (weighinq 187 to 293 q and 56 to 67 days of aqe atthe time of treatment, February 21, 1979) of the Charles RiverCD strain [CRL:COBS CD (SD) BR] were obtained from the CharlesRiver Breedinq Laboratories, Inc., Portaqe, Michiqan. The animalswere acclimated to laboratory conditions for seven days in theRockville facility of the Department of Toxicoloqy. The animalswere individually housed in wire-bottom cages in temperature-controlled quarters under artificial illumination controlled toprovide a 12-hour light cycle. Water and Purina Laboratory Chowwere provided ad libitum with the exception of the night beforetreatment when-ood was removed from the caqes.

The test material as the salt, was dissolved in deionized water atconcentrations of 250 or 500 mq/ml. Sinqle graded doses of thetest material were administered by oral qavaqe to five rats of eachsex at each of four dose levels.

3. EXPERIMENTAL DESIGN (Continued)

The rats were observed frequently on the day of treatment and

daily thereafter. The animals were weighed on the day of treat-

ment, and on Days 7 and 14 following treatment. Necropsies were

performed on all animals that died during the study and on the

surviving animals that were killed 14 days after treatment.

4. RESULTS

The data have been summarized as follows below.

Mean Body WeightDose (g) Deaths Total

mg/k DayDay Mortality

0 7 14 0 1 2 3 4 5-14 Deaths/Treated

Males

4640 271 321 352 0/5

6810 235 282 328 1 1/5

10000 257 - - 5 5/5

14700 286 - - 5 5/5

Females

3160 206 231 24d 0/5

4640 211 241 248 1 1/5

6810 195 239 238 3 3/5

10000 203 - - 5 5/5

Based on the deaths occurring in the 14 days after treatment, and

using a modification of the method of Horn (Biometrics, V. 12,

p. 311, 1956), LD50 values of 7650 mg/kg (95% confidence limits:

6560-8920) and 6070 mg/kg (95% confidence limits: 4760-7740) for

male and female rats, respectively, were calculated.

Signs of toxicity included loose or liquid stool in several animals

at all dose levels. Reduced motor activity and prostration were

observed in some animals at doses greater than 3160 mg/kg. Ataxia

was seen in one male at the 10,000 mg/kg dose level,

one female at the 10,000 and 4640 mg/kg levels, and three females

at the 6810 mg/kg dose level. Additional observations consisted

of an opaque eye in one male at the 6810 mg/kg level, abnormal

breathing sounds in one female at the 10,000 mg/kg level, and

tremors in one female at the 4640 mg/kg dose level.

007

4. RESULTS (Continued)

No abnormal necropsy findings were observed in any of the survivors.Among rats that died, red or mottled lungs in some of the malesat the 10,000 and 14,700 mg/kg levels and some females at the 6810 mg/kglevel and one female at the 10,000 and 4640 mg/kg levels were noted.Distended, fluid-filled stomachs and intestines were observed in allanimals found dead. In addition some of the stomach linings werered. Dark spots on the thymus were seen in females at 6810 and10,000 mg/kg levels and males at the 10,000 and 14,700 mg/kg levels.

Thirty days after transmittal of this report, original data anda copy of the final report will be transferred to the LBI Archivist,5516 Nicholson Lane, Kensington, Maryland, for distribution to theproper repositories. A copy of this report and underlying data werereviewed by the LBI Quality Assurance Unit.

5. CONCLUSION

Following the oral administration of graded doses of the testcompound to fasted young adult rats, LD50 values of 7650 and6070 mg/kg were calculated for male and female rats, respectively.The central nervous system and gastrointestinal tract may bepossible sites of toxicity.

Submitted by:

I -r I ,hv . 72 ,F

David R. Damske, B.A. Francis J. Ylecie- DateTechnical Group Leader Senior ToxicologistDepartment of Toxicology Department of Toxicology

Reviewed by:

E. Ross Hart, Ph.D. atePrincipal Investigator

008

SECTION C

ACUTE ORAL TOXICITY STInfY IN MICF



SUMMARY

The oral LD50 values of the test material, IMPA, were 5620 and 6550 mo/kafor male and female mice, respectively.

1. OBJECTIVE

The objective of this study was to evaluate the acute toxicityof the test material when administered by oral qavaqe to male andfemale mice.

2. MATERIAL

Two glass jars (550 q each) containinq a white powder and labeled:


were received from U. S. Army Medical Research and DevelopmentCommand by Litton Rionetics, Inc. (LBI) on December 18, 1978, anddesignated as LBI No. 3427.


Young adult mice (weighinq 24 to 38 Q and nine weeks of aqe atthe time of treatment, February 6, 197q) of the Charles RiverCD-1 strain were obtained from the Charles River Breedinq Lahora-tories, Inc., Portaqe, Michiqan. The mice were acclimated tolaboratory conditions for 13 days in the Rockville facility ofthe Department of Toxicoloqy. The animals were individuallyhoused in wire-bottom caqes in temperature-controlled quartersunder artificial illumination controlled to provide a 12-hourlight cycle. Acidified water (pH 2.5) and Purina LaboratoryChow were provided ad libitum with the exception of the niahtbefore treatment when food was removed from the caqes.

The test material as a salt, was dissolved in deionized water at aconcentration of 500 mg/ml at 10,000 and 6810 mq/kq, and 250 mQ/mlat 4640 and 3160 mq/kq dose levels. Single qraded doses of thetest material were administered by oral qavage to five mice of eachsex at each of four dose levels.

.. 003


The mice were observed frequently on the day of treatment anddaily thereafter. The animals were weighed on the day of treat-ment, and on Days 7 and 14 following treatment. Necropsieswere performed on all animals that died during the study and onthe surviving animals that were killed 14 days after treatment.

4. RESULTS

The data have been summarized below.

Mean Body WeightDose (g) Deaths Total(mg/kg) Day Day Mortality

7 14 0 2 3 4-14 Deaths/Treated

Males

3160 31 36 35 0/54640 31 37 38 1 1/56810 34 37 39 3 1 4/510000 29 - - 5 5/5

Females

3160 26 31 32 0/54640 27 32 32 1 1/56810 27 33 33 2 2/5

10000 28 - - 5 5/5

Based on the deaths occurring in the 14 days after treatment,and using a modification of the method of Horn (Biometrics,V. 12, p. 311, 1956), LD50 values of 5620 mg/kg (95% confidencelimits: 4530-6990) and 6550 mg/kg (95X confidence limits:5140-8360) for male and female mice, respectively, were calculated.

Signs of toxicity included soft or liquid stool in some mice atall dose levels, except the 3160 mg/kg female mice. Reducedmotor activity and ataxia were observed in some mice at doselevels greater than 3160 mg/kg , with all male mice at the6810 mg/kg level affected and all mice at the 10,000 mg/kg levelwhich were alive at the one hour observation interval. Prostra-tion was observed in one female at the 6810 mg/kg level and twomales and one female at the 10,000 mg/kg level.

010


No abnormal necropsy findings were observed in any of the micesurviving for 14 days. Among mice that died, fluid-filled stomachand intestines were observed in all animals. Mottled or con-gested lungs were observed in three females at the 10,000 mg/kglevel and dark red areas on the lungs of one female at the4640 mg/kg dose level were observed.

Thirty days after transmittal of this report, original data fromthe Department of Toxicology will be transferred to the LBIArchivist, 5516 Nicholson Lane, Kensington, Maryland, for distri-bution to the proper repositories. A copy of this report wasreviewed by the LBI Quality Assurance Unit.

5. CONCLUSION

Following the oral administration of graded doses of the testmaterial to fasted young adult mice, LD5O values of 5620 and6550 mg/kg were calculated for male and female mice, respectively.The nervous system and gastrointestinal tract may be possiblesites of toxicity.

Submitted by:

David R. Damske, B.A. Francis J cler DateTechnical Group Leader Senior Toxi ologistDepartment of Toxicology Department of Toxicoloqy

Reviewed by:

obert P. Beliles, Ph.D. DateDirectorDepartment of Toxicology

'. O0t1

SECTION D

ACUTE DERMAL TOXICITY STUDY IN PARRITS



SUMMARY

Following the dermal application of 2.0 q/kq of the test material toyoung adult female rabbits, no deaths or signs of systemic toxicity wereobserved. The dermal LD50 was estimated to he greater than 2.0 g/kq.

1. OBJECTIVE

The objective of this study was to evaluate the acute systemictoxicity and the potential for skin irritation of the testmaterial following a single dermal application to six rabbits.

2. MATERIAL

Two glass jars (550 g each) containing a white powder and labeled:

Isopropyl Methylphosphonic Acid, Sodium SaltRatch 1 #127818

were received from U. S. Army Medical Research and DevelopmentCommand by Litton Bionetics, Inc. (LBI) on December 18, 1978,and desiqnated as LBI No. 3427.


Young adult female New Zealand White (albino) rabbits (weiqhinq2.5 to 3.0 kg and 14 weeks of age) were obtained from R and HRabbitry, Rockville, Maryland. The animals were acclimated tolaboratory conditions for 15 days in the Rockville facility ofthe Department of Toxicology. The animals were individuallyhoused in wire-bottom cages in temperature-controlled Quartersunder artificial illumination controlled to provde a 12-hour lightcycle. Water and Purina Rabbit Chow were provided ad lihitum.

Six rabbits were prepared by clipping the skin of the trunk freeof hair 24 hours prior to test material application. Threerabbits were further prepared by making epidermal abrasions every2-3 cm longitudinally over the areas to be exposed. The abrasionswere sufficiently deep to penetrate the stratum corneum, but notto disturb the derma. A single 2.0 q/ka dose of the test materialwas applied to the skin of the rabbits beginning February 6, 1979.

IL!


After application, the site was covered with gauze and Saranwrap, and the rabbits were placed in restraining collars for24 hours. At the end of 24 hours, the Saran wrap and collarswere removed. The gauze pad was then weighed to determine theamount of unabsorbed test material. The subjects were cleanedby thorough wiping and observed daily for gross signs of poison-ing and skin reactions for two weeks after testing. Body weightswere obtained initially, and at 7 and 14 days after treatment.The evaluation of skin reactions was based on the followingscoring system. All animals surviving 14 days after treatmentwere necropsied and any changes in the internal organs noted.

Erythema and Eschar Formation

No erythema 0.......................0Very slight erythema (barely perceptible). ........Well defined erythema ....... .................. 2Moderate to severe erythema .............. 3Severe erythema (beet redness) to slight eschar

formation (injuries in depth) ..... ............. 4

Total possible erythema score ..... .............. 4

Edema Formation

'o edema ..... ......................... 0Very slight edema (barely perceptible) ............. 1Slight edema (edges of area well defined bydefinite raising) ................... 2

Moderate edema (raised approximately 1 mm) 3........3Severe edema (raised more than 1 mm andextending beyond area of exposure) ................4

Total possible edema score .... ................

4. RESULTS

The results of the data have been summarized on page 3. On anaverage, 35% of the applied dose was unabsorbed and remainedon the skin and gauze pad after the Saran wrap was removed.

013


Body WeightsRabbit Dose (kg)Number (g/kg) Skin Day

0 7 T4

5204 2.0 Intact 2.7 3.0 3.15205 2.0 Intact 2.7 3.1 3.45206 2.0 Intact 2.6 3.0 3.0

5207 2.0 Abraded 2.6 2.9 2.85208 2.0 Abraded 2.5 3.1 3.45209 2.0 Abraded 3.0 3.2 3.5

Based on the absence of deaths occurring in the 14 days aftertreatment, the LD50 value was estimated to be greater than 2 g/kgfor female rabbits.

Both the intact and abraded rabbits showed very slight to well-defined erythema for one to five days after treatment. Observa-tions included one rabbit with a swollen left eyelid and a whitedischarge from the left eye, and one rabbit had hair loss at themid-dorsal cervical region. Observations at necropsy revealed novisible abnormalities.

Thirty days after transmittal of this report, original data fromthe Department of Toxicology will be transferred to the LBi Archivist,5516 Nicholson Lane, Kensington, Maryland, for distribution to theproper repositories. A copy of this report was reviewed by theLBI Quality Assurance Unit.

5. CONCLUSION

Following the dermal application of 2.0 g/kg of the test materialto young adult female rabbits, no mortalities or signs of systemictoxicity were observed. The dermal LD5O was estimated to be greaterthan 2.0 g/kg and the dermal response was judged to suggest a mildpotential for skin irritation.

Submitted by:

David R. Damske, B.A. Francis J Mecer Date

Technical Group Leader Senior ToxicologistDepartment of Toxicology Department of Toxicology

Reviewed by:

Robert P. Beliles, Ph.D. Date

DirectorDepartment of Toxicology

014

L

SECTInN E

PPIMARY EYE IRRITATION ANn CORROSIVENESS STIJOY IN RARRITS



SUMMARY

The test material, IMPA, was instilled (10 mq) into the eyes of albinorabbits. The eyes were examined at 1, 24, 48 and 72 hours, and at4 and 7 days after treatment. Based on the scorinq system used, thetest material was judged not to be irritating to the eye.

1. OBJECTIVE

The objective of this study was to investigate the possibilitythat direct contamination of the eye with the test material wouldbe followed by irritation and/or injury.

2. MATERIAL

Two glass jars (550 q each) containing a white powder and labeled:


were received from U. S. Army Medical Research and DevelopmentCommand by Litton Bionetics, Inc. (LBI) on December 18, 1978,and designated as LBI No. 3427.


Six female New Zealand White (albino) rabbits were obtained fromB and H Rabbitry, Rockville, Maryland. The animals were acclimatedto laboratory conditions for 14 days in the Rockville facility ofthe Department of Toxicoloqy. The rabbits weighed 2.3 to 2.8 kqat the time of treatment, February 5, 1979. The rabbits wereindividually housed in wire-bottom cages in temperature-controlledquarters under artificial illumination controlled to provide a12-hour light cycle. Water and Purina Rabbit Chow were Providedad libitum.

The eyes of the rabbits were examined with the aid of 2% sodiumfluorescein before testing to ensure that the eyes were withoutdefects or irritation. The animals were firmly held and 100 mqof the test material was instilled into one eye of each rabbit.The eyes were examined and araded according to the table onpage 2. The scores were recorded at 1, 24, 4P and 72 hours, andat 4 and 7 days.

" 015


CORNEA GRADE

No ulceration or opacity ...................................... 0Scattered or diffuse areas of opacity, details of I

iris clearly visible ....................................... 11Easily discernible translucent areas of opacity, 1,2

details of iris slightly obscured .......................... 2Nacreous areas of opacity, no details of iris visible, 1,2

size of pupil barely discernible ......... .............. 3 9Complete corneal opacity, iris not discernible .............. 41,2Ulceration, absence of a gross patch of corneal 2

epithelium ................................................. 4

IRIS

Normal ........................................................ 0Markedly deepened folds, congestion, swelling,

moderate circumcorneal injection (any of theseor combination of any thereof), iris still 1reacting to light (sluggish reaction is positive) .......... 1

No reaction to light, hemorrhage, gross destruc-tion (any or all of these) ................................. 2

CONJUNCTIVAE

Redness (refers to palpebral and bulbarconjunctivae excluding cornea and iris):

Vessels normal ................................................ 0Some vessels definitely injected .............................. IDiffuse, crimson red, individual vessels not

easily discernible ........................................ 21Diffuse beefy red ............................................ 3

Chemosis:

No swelling ................................................... 0Any swelling above normal (includes nictitating

membrane) .................................................. 1Obvious swelling with partial eversion 1

of lids ............................................ 21Swelling with lids about half closed ....................... 31Swelling with lids more than half closed....................41Ulceration or necrosis of palpebral and 2

bulbar conjunctivae or nictitating membrane..............4

1Grades considered positive for irritation.Grades considered positive for corrosiveness.In addition, grade I opacity evident for any6 or more days will also be considered ascorrosive.

^16

4. RESULTS

The detailed results of the ocular changes in the treated eyes ofeach rabbit have been included in the Appendix. No changes wereobserved in the untreated eyes. No ocular changes were observedin five of the six treated rabbits. One rabbit was observed toshow a Grade 1 reaction at the 24 hour observation interval forthe iris and redness of the conjunctivae.

Thirty days after transmittal of this report, original data fromthe Department of Toxicology will be transferred to the LBI Archivist,5516 Nicholson Lane, Kensington, Maryland, for distribution tothe proper repositories. A copy of this report was reviewed bythe LBI Quality Assurance Unit.

5. CONCLUSION

The test material, IMPA, was judged to be non-irritating wheninstilled into the eyes of rabbits.

Submitted by:

David R. Damske, B.A. F Date-Technical Group Leader Senior Toxi logistDepartment of Toxicology Department of Toxicology

Reviewed by:

Robert P. Beliles, Ph.D. DateDirectorDepartment of Toxicology

'017

TABLE E-1LITTON BIONETICS, INC.PROJECT NO. 10734-11

SCORING FOR EYE IRRITATION

RABBIT NUMBER: 5198D EYE TREATED: RIGHT

TIME OF OBSERVATION AFTER TREATMENTHOURS DAY

1 24 48 72 4T 7

CORNEA 0 0 0 0 0 0

IRIS 0 1 0 0 0 0

CONJCNCTIVAE

A. REDNESS 0 1 0 0 0 0

B. CHEMOSIS 0 0 0 0 0 0

RABBIT NUMBER: 5199D EYE TREATED: LEFT

TIME OF OBSERVATION AFTER TREATMENTHOURS DAY1 24 48 72 7

CORNEA 0 0 0 0 0 0

IRIS 0 0 0 0 0 0

CONJUNCTIVAE

A. REDNESS 0 0 0 0 0 0

B. CHEMOSIS 0 0 0 0 0 0

TABLE E-1 (CONTINUED)LITTON BIONETICS, INC.PROJECT NO. 10734-11



TIME OF OBSERVATION AFTER TREATMENT

HOURS DAYT 24 48 72

CORNEA 0 0 0 0 0 0

IRIS 0 0 0 0 0 0

CONJUNCTIVAE

A. REDNESS 0 0 0 0 0 0

B. CHEMOSIS 0 0 0 0 0 0


TIME OF OBSERVATION AFTER TREATMENT

HOURS DAY1 24 48 72 4

CORNEA 0 0 0 0 0 0

IRIS 0 0 0 0 0 0

CONJUNCTIVAE

A. REDNESS 0 0 0 0 0 0

B. CHEMOSIS 0 0 0 0 0 0

019

-kk" •Il l - - r . . . . . . . . . . ll i l I ,l . .

TABLE E-1 (CONTINUED)LITTON BIONETICS, INC.PROJECT NO. 10734-11



TIME OF OBSERVATION AFTER TREATMENTHOU.RS DAY1 24 48 72

CORNEA - 0 0 0 0 0 0

IRIS 0 0 0 0 0 0

CONJUNCTIVAE

A. REDNESS 0 0 0 0 0 0

B. CHEMOSIS 0 0 0 0 0 0


TIME OF OBSERVATION AFTER TREATMENTHOURS DAY1 24 48 72 _

CORNEA 0 0 0 0 0 0

IRIS 0 0 0 0 0 0

CONJUNCTIVAE

A. REDNESS 0 0 0 0 0 0

B. CHEMOSIS 0 0 0 0 0 0

;02o

SECTION F

GUINEA PIG SENSITIZATION STUDY



SUMMARY

Guinea piqs were injected intradermally with a 0.1% solution of thetest material in ten 0.1 ml injections over a course of three weeks inan attempt to induce sensitization. Two weeks after the last iniectiona challenge injection of 0.1 ml was made. Based on the observations ofthe reaction to the challenge injection, the test material was ,judqedto have no potential to induce sensitization in the quinea piq.

1. OBJECTIVE

The objective of this study was to evaluate the potential ofthe test material to cause skin sensitization in quinea piqs.

2. MATERIAL

Two glass jars (550 q each) containing a white powder and labeled:


were received from U. S. Army Medical Research and DevelopmentCommand by Litton Bionetics, Inc. (LBI), on December 18, 1978,and designated as LBI No. 3427.

A brown jar (100 g) containing an amber crystalline material andlabeled:

Eastman 1-chloro-2,4-dinitrobenzene

Lot No. F7A

was purchased from Scientific Products by Litton Bionetics, Inc.(LBI), and designated as LBI No. 3720.


Thirteen female guinea pigs (weighing 302 to 445 q at the time oftreatment, February 7, 1979) of the Hartley strain were obtainedfrom Charles River Breeding Laboratories, Inc., Kingston, New York.The animals were acclimated to laboratory conditions for 14 daysin the Rockville facility of the Department of Toxicology. Theanimals were individually housed in wire-hottom cages in temperature-controlled quarters under artificial illumination controlled toprovide a 12-hour liqht cycle. Water and Purina Guinea Piq Chowwere provided ad libitum.

S0211


One flank of each animal was shaved 24 hours prior to treatment.The animals were divided into two groups. Group 1, consistingof four guinea pigs, received 2,4-dinitro-l-chlorobenzene 0.1%as a positive control. Group 2, consisting of nine guineapigs, received 0.1 ml of a 0.1% solution of the test materialin normal saline. The test solutions were injected intradermallyinto the shaved flank, three times per week (Monday, Wednesdayand Friday) for 10 injections. Fifteen days after the tenthinjection, achallenge injection was made to a site below thesensitization site.

At each sensitizing injection an evaluation of the reactionfrom the previous injection was made. The site of the challengeinjection was evaluated 24 and 48 hours after the challenge.The evaluation of the reaction sites included measurement ofreaction diameters with calipers and estimation of height ofreaction. Color of the reaction site was noted. The guineapigs were observed daily for mortality.

4. RESULTS

The results of the individual observations have been detailed inAppendix Table F-2.

Observations of the test group guinea pigs' reactions revealedno reaction during the sensitizing period. Very slight reactionsconsisting of very mild erythema and very slight edema were observed24 hours after the challenge injection. Two guinea pigs werefound dead prior to completion of dosing but these deaths werenot considered to be related to test material administration.Observations of the positive control guinea pigs' reactionsrevealed redness and swelling after the fourth sensitizing injectionwith the degree of redness and diameter of the reaction increasingwith subsequent injections. The response to the challenge injectionwas greater than the average sensitization reaction. Based on thereaction of the guinea pigs to the challenge injection of thetest material, the test material was judged to have no potentialto cause skin sensitization in the guinea pig.

Thirty days after transmittal of this report, original data anda copy of the final report will be transferred to the LBI Archivist,5516 Nicholson Lane, Kensington, Maryland, for distribution to theproper repositories. A copy of this report and underlying datawere reviewed by the LBI Quality Assurance Unit.

L02

5. CONCLUSION

Guinea pigs were injected intradernally with a 0.17. solution ofthe test material in ten 0.1 ml injections over a course ofthree weeks in an attempt to induce sensitization. Two weeks afterthe last injection, a challenge injection was made. Based on theobservations of the reaction to the challenge injection, the testmaterial was judged to have no potential to induce sensitizationin the guinea pig.

Submitted by:

David R. Damske, B.A. Francis J. Mcleff DateTechnical Group Leader Senior ToxicologistDepartment of Toxicology Department of Toxicology

Reviewed by:

E. P Hart, Ph.D. DatePrincipal Investigator

LITTON BIONETICS, INC.

PROJECT NO. 10734-14

TABLE F-2

OBSERVATIONS

ANIMAL NUMBER: 5439 POSITIVE CONTROL

DOSE OBSERVATIONS(ML) DATE DIAMETER (MM) HEIGHT (MM) COLOR

0.1 2-09-79 3x3 1 02-12-79 3x4 2 02-14-79 3xlO 2 02-16-79 4x15 0 SLIGHT RED2-19-79 8xll 0 SEVERE RED2-21-79 5xi0 2 SEVERE RED2-23-79 5x10 3 SEVERE RED2-26-79 6x6 2 SEVERE RED2-28-79 7x6 2 SEVERE RED

CHALLENGE DOSE

0.1 3-16-79 6x6 2 SEVERE RED3-17-79 8x5 2 SEVERE RED



0.1 2-09-79 2x2 0 02-12-79 0 0 02-14-79 3xlO 0 02-16-79 7x10 I SLIGHT RED2-19-79 15x8 1 02-21-79 15x8 I SLIGHT RED2-23-79 8x18 I SLIGHT RED2-26-79 10x3 1 SLIGHT RED2-28-79 7x1O 1 SEVERE RED

CHALLENGE DOSE

0.1 3-16-79 7x7 2 RED3-17-79 7x6 2 RED

M4.

LITTON BIONETICS, INC.PROJECT NO. 10734-14

TABLE F-2 (CONTINUED)

OBSERVATIONS


DOSE OBSERVATIONS(ML) DATE DIAMETER (MMI HEIGHT (MM) COLOR

0.1 2-09-79 lxI 1 SLIGHT RED2-12-79 lxl 1 SLIGHT RED2-14-79 3x9 1 SLIGHT RED2-16-79 1Oxi0 2 SLIGHT RED2-19-79 1Oxi0 1 02-21-79 1Ox1O 2 SLIGHT RED2-23-79 10x12 2 SEVERE RED2-26-79 10x6 1 SEVERE RED2-28-79 10x7 1 SEVERE RED

CHALLENGE DOSE

0.1 3-16-79 8x6 2 MODERATE RED3-17-79 7x7 2 MODERATE RED



0.1 2-09-79 0 0 02-12-79 3x2 1 02-14-79 3x8 1 02-16-79 5x8 2 SLIGHT RED2-19-79 5x17 1 02-21-79 7x12 1 SLIGHT RED2-23-79 5x10 1 SLIGHT RED2-26-79 5xi0 1 SLIGHT RED2-28-79 5x12 2 SLIGHT RED

CHALLENGE DOSE

0.1 3-16-79 7x7 2 MODERATE RED3-17-79 7x7 2 RED

V. -.




OBSERVATIONS

ANIMAL NUMBER: 5443 IMPA


0.1 2-09-79 0 0 0

2-12-79 0 0 0

2-14-79 0 0 0

2-16-79 0 0 0

2-19-79 0 0 0

2-21-79 0 0 0

2-23-79 0 0 0

2-26-79 0 0 0

2-28-79 0 0 0

CHALLENGE DOSE

0.1 3-16-79 2x2 I SLIGHT RED

3-17-79 0 0 0


DOSE OBSERVATIONS(ML) DATE DIAMET MM) HEIGHT (MM) COLOR

0.1 2-09-79 0 0 0

2-12-79 0 0 0

2-14-79 0 0 0

2-16-79 0 0 0

2-19-79 0 0 0

2.-21-79 0 0 0

2-23-79 0 0 0

2-26-79 a

aFound dead.



OBSERVATIONS


DOSE OBSERVATIONS -EGTM(ML) DATE

________________

0.1 2-09-79 00 02-12-79 0 0 02-14-79 0 0 02-16-79 000

2-1-7 00 02-19-79 0 0 02-23-79 0 0 02-26-79 0002-28-79 0 0 0

CHALLENGE DOSE

0.1 3-16-79 403 1 WELL DEFINED RED

3-17-79 4x3 0 SLIGHT RED


DOSE OBSERVATIONS

(ML) DATE DIAMETER_____

0.1 2-09-79 0 00

2-12-79 0 00

2-14-79 0 00

2-16-79 0 00

2-19-79 0 00

2-21-79 0 00

2-23-79 0 00

2-26-79 0 00

2-28-79 0 00

CHALLENGE DOSE

0.1 3-16-79 0 00

3-17-79 0 00




OBSERVATIONS


DOSE OBSERVATIONS(ML) DATE DIAMETER CMM) HEIGHT (MM) COLOR

0.1 2-09-79 0 0 02-12-79 0 0 0

2-14-79 0 0 0

2-16-79 0 0 02-19-79 0 0 02-21-79 0 0 0

2-23-79 0 0 02-26-79 0 0 0

2-28-79 0 0 0

CHALLENGE DOSE

0.1 3-16-79 2x2 0 SLIGHT RED

3-17-79 0 0 0


DOSE OBSERVATIONS(ML) DATE DIAMETER7MM) HEIGHT (MM) COLOR

0.1 2-09-79 0 0 0

2-12-79 0 0 0

2-14-79 0 0 0

2-16-79 0 0 0

2-19-79 0 0 0

2-21-79 0 0 02-23-79 0 0 02-26-79 0 0 0

2-28-79 0 0 0

CHALLENGE DOSE

0.1 3-16-79 1x2 0 SLIGHT RED

3-17-79 0 0 0

LITTON BIONET,40,, INC.PROJECT NO. 10734-14


OBSERVATIONS


DOSE OBSERVATIONS(ML) DATE DIAMETERMM) HEIGHT (MM) COLOR

0.1 2-09-79 0 0 0

2-12-79 0 0 0

2-14-79 0 0 0

2-16-79 0 0 0

2-19-79 0 0 0

2-21-79 0 0 0

2-23-79 0 0 0

2-26-79 0 0 0

2-28-79 0 0 0

CHALLENGE DOSE

0.1 3-16-79 4x4 1 SLIGHT RED

3-17-79 0 0 0


DOSE OBSERVATIONS(ML) DATE DIAMERHEIGHT MM) COLOR

0.1 2-09-79 0 0 0

2-12-79 0 0 0

2-14-79 0 0 0

2-16-79 0 0 0

2-19-79 0 0 0

2-21-79 0 0 0

2-23-79 0 0 0

2-26-79 0 0 0

2-28-79 0 0 0

CHALLENGE DOSE

0.1 3-16-79 8x5 1 SLIGHT RED

3-17-79 0 0 0

.. . m I n . .... ... ... . .. . . . . ... , ,.,,,m,, ., ,n -- r.. ...... .. "... .... . 0 2 3 . .



OBSERVATIONS



0.1 2-12-79 0 0 0

2-14-79 0 0 0

2-16-79 0 0 0

2-19-79 0 0 0

2-21-79 0 0 0

2-23-79 0 0 0

2-26-79 0 0 0

2-28-79 0 0 0

3-02-79 0 0 0

3-05-79 a

aFound dead.

L 030

SECTION G

MICROBIAL MUTAGENESIS

ISUPROPYL METHYLPHOSPHiONIC ACID

LBI PROJECT NO. 10734

I. SPONSOR: U.S. Army

i1. M~ATERIAL

A. Identification: IMPA-isopropyl methylphosphonic acid, Sodium Salt

B. Date Received: January 22, 1979

C. Physical Description: White fluffy powder

III. TYPE OF ASSAY: Ames Salmonella/Microsome Plate Test

tV. PROTOCOL NO.: OMT-1O0

V. RESULTS

The results of this assay are presented in Tables G-3 and G-4.

'/1. INTERPRETATION OF RESULTS AND CONCLUSIONS

The test compound was examined for mutagenic activity in aseries of in vitro microbial assays employing Salmonellaindicator organisms. The comoound was tested directly andin the presence of liver microsomal enzyme oreparationsfrom Aroclor-induced rats.

The compound was tested over a series of concentrationssuch that there was either quantitative or qualitativeevidence of some chemically-induced physiological effectsat the high dose level. The low dose in all cases was be-low a concentration that demonstrated any toxic effect.The dose range employed for the evaluation of this comooundwas from 1.0 ug to 1000 ug per plate.

The test compound did not exhibit toxicity in the initialtest; therefore.a repeat test was conducted at additionaldoses of 2500 ug and 5000 ;g per plate with the strainsTA-1537, TA-98 and TA-lO0. Slight toxicity was observedat 5000 ug per plate with the strain TA-1537.

The results of the tests conducted on the compound in theabsence of a metabolic activation system were all negative.The test was repeated with TA-1537 and TA-1538 because oflow solvent control values for TA-1538 and because of theoverall increased number of revertantover the solventcontrols with TA-1537. The repeat tests were also negative.

>031

VI. INTERPRETATION OF RESULTS AND CONCLUSIONS (Continued)

The results of the tests conducted on the compound in thepresence of a rat liver activation system were all negative.

The test compound IMPA-ISOPROPYL METHYLPHOSPHONIC ACID, SodiumSalt did not demonstrate genetic activity in any of the assaysconducted in this evaluation and was considered not mutagenicunder these test conditions.

Submitted by:

Study Director

4 .5 1t 3. /Ps2".R. Jagannath, Ph.D. DateSection ChiefSubmammalian GeneticsDepartment of Geneticsand Cell Biology

Reviewed by:

.r usick, Ph.D. 4DirectorDepartment of GenetIcsand Cell Biology

2

: 0302

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PROTOCOL

1. PURPGSE

The purpose of this study was to evaluate the test material forgenetic activity in a microbial assay with and without theaddition of mammalian metabolic activation preparations.

2. MATERIALS

A. Indicator Microorganisms

Salmonella typhimurium TA-1535TA-1537TA-1538TA-98TA-lO0

B. Activation System

I. Reaction Mixture

Component Final Concentration/ml

TPN (Sodium salt) 4 umolGlucose-6-phosphate 5 WmolSodium phosphate (dibasic) 100 tmolMgCl 2 8 umolKCL 33 umolHomogenate S9 fraction 0.1 ml

2. S9 Homogenate

A 9,000 x j supernatant prepared from Sprague-Dawleyadult male rat liver induced by Aroclor 1254 lot #BI0-82was purchased from Biological Products, Litton Bionetics,Inc. and was used in these assays.

S035

2. MATERIALS (Continued)

C. Positive Control Chemicals

The chemicals used for positive controls in the nonactivation andactivation assays are given in the Tables of Section V. Results.

0. Solvent

The solvent employed to prepare the stock solution of the testchemical is given in the Tables of Section V. Results.All dilutions of the test chemical were made using this solvent.


A. Plate Test (Agar Incorporation)

Approximately 10 cells from an overnight culture of each indicatorstrain were added to separate test tubes containing 2.0 ml moltenagar supplemented with biotin and a trace of nistidine. For non-activation tests, at least 4 dose levels of the test compoundwere added to the contents of the appropriate tubes and pouredover the surfaces of selective agar plates. In activation tests,at least 4 dose levels of the test chemical were added to theappropriate tubes with cells. Just prior to pouring, an aliquotof reaction mixture (0.5 ml containing the 9,000 x 2 liver homo-genate) was added to each of the activation overlay tubes, whichwere then mixed, and the contents poured over the surface of aminimal agar plate and allowed to solidify. The plates were incu-bated for 48 hrs at 371C and scored for the number of coloniesgrowing on each plate. The concentrations of all chemicals aregiven in the Tables of Section V. Results. Positive controls usingboth directly active positive chemicals and those that reouiremetabolic activation were run with each assay.

Certain classes of chemicals known to be mutagens and carcinogens do notproduce detectable responses using the standard Ames agar incorporationmethod. Some dialkyl nitrosamines and certain substituted hydrazines aremutagenic in suspension assays, but not in the plate assay. Chemicalsof these classes should be screened in a suspension assay.

"' 036

I.

3. EXPERIMENTAL CESIGN (Continued)

B. Recording and Presentina Data

The numbers of colonies on each plate were counted and recorded onprinted forms. These raw data were analyzed in a comouter programand reported on a printout. The results are presented as rever-tants oer plate for each indicator strain employed in tne assay.The positive and solvent controls are orovided as referencepoints. Other relevant data are provided on the comouter printout.

4. EVALUATION CRITERIA

Plate test data consist of direct revertant colony counts obtained froma set of selective agar plates seeded with populations of mutant cellssuspended in a semisolid overlay. Because the test chemical and thecells are incubated in the overlay for 2 days, and a few cell divisionsoccur during the incubation period, the test is semiquantitative innature. Although these features of the assay reduce the quantitationof results, they provide certain advantages not contained in a quantita-tive suspension test:

The small number of cell divisions permits potential mutagensto act on replicating DNA, which is often more sensitive thannonreplicating DNA.

The combined incubation of the compound and the :ells in theoverlay permits constant exposure of the indicator cells for2 days.

A. Survivinq Populations

Plate test procedures do not permit exact quantitation of the numbero cells surviving chemical treatment. At low concentrations of thetest chemical, the surviving population on the treatment plates isessentially the same as that on the negative control plates. Athigh concentrations, the surviving population is usually reduced bysome fraction. Our protocol normally emplo)y several doses rangingover 2 or 3 log concentrations, the highest of these doses beingselected to show slight toxicity as determined by subjective criteria.

B. Dose Response Phenomena

The demonstration of dose-related increases in mutant counts is animportant criterion in establishing mutagenicity. A factor thatmight modify dose-response results for a mutagen would be the selec-tion of doses that are too low (usually mutagenicity and toxicityare related). If the highest dose is far lower than a toxic concen-tration, no increases may be observed over the dose range selected.

a037

EVALUATTON CRIT RIA (Continued)

B. Dose-Resoonse Phenomena

Conversely, if the lowest dose employed is highly cytotoxic,the test chemical may kill any mutants that are induced, andthe compound will not appear to be mutagenic.

C. Control Tests

Positive and negative control assays are conducted with eachexperiment and consist of direct-acting mutagens for nonactivationassays and mutagens that require metabolic biotransformation inactivation assays. Negative controls consist of the test compoundsolvent in the overlay agar together with the other essential com-ponents. The negative control plate for each strain gives a refer-ence point to which the test data are compared. The positive controlassay is conducted to demonstrate that the test systems are functionalwith known mutagens.

D. Evaluation Criteria for Ames Assay

Because the procedures used to evaluate the utagenicity of thetest chemical are semiquantitative, the criteria used to determinepositive effects are inherently subjective and are based primarilyon a historical data base. Most data sets are evaluated using thefollowing criteria:

I. Strains TA-1535, TA-1537 and TA-1538

If the solvent control value is within the normal range, achemical that produces a positive dose response over threeconcentrations with Jhe highest increase equal to thre-etimes thesolvent control value is considered to be mutagenic.

2. Strains TA-g8 and TA-400

If the solvent control value is within the normal range, achemical that produces a positive dose response over threeconcentrations with the highest increase equal to t'ice thesolvent control value for TA-10 and 2-3 times the solventcontrol value for strain TA-g8 is considered to be matagenic.For these strains, the dose-response increase should startat approximately the solvent control value.

03SL~ALI

4. EVALUATION CRITERIA (Continued'

D. Evaluation Criteria for Ames Assay

3. Pattern

Because TA-1535 and TA-10 are both derived from the sameparental strain (G-46) and because TA-1538 and TA-98 are bothderived from the same parental strain (03052), there is abuilt-in redundancy in the microbial assay. In general, thetwo strains of a set respond to the same mutagen and such apattern is sought. It is also anticipated that if a givenstrain, e.g., TA-1537, responds to a mutacen in nonactivationtests, it will generally do so in activation tests (the con-verse of this relationship is not expected). While similarresponse patterns are not required for all mutagens, they canbe used to enhance the reliability of an evaluation decision.

4. Reproducibility

If a chemical produces a response in a single test thatcannot be reproduced in one or more additional runs, theinitial positive test data lose significance.

The preceding criteria are not absolute, and other extenuating

factors may enter into a final evaluation decision. However,these criteria are applied to the majority of situations and arepresented to aid those individuals not familiar with this proce-dure. As the data base is increased, the criteria for evaluationcan be more firmly established.

E. Relationship Between Mutagenicity and Carcinogenicity

It must be emphasized that the Ames Salmonella/Microsome Plate Testis not a definitive test for chemical carcinogens. It is recognized,however, that correlative and functional relationships have beendemonstrated between these two endpoints. The results of comparativetests on 300 chemicals by McCann et al. (1975) show an extremelygood correlation between results of microbial mutagenesis tests andin vivo rodent carcinogenesis assays.

All evaluations and interpretation of the data presented in thisreport are based only on the demonstration, or lack, of mutagenicactivity.

039

REFEREICES

Ames, B.N., McCann, J. and Yarasake, E. (1975). Methods for detectingcarcinogens and rnutagens with the Salmonella/ ramalian-microsome muta-genicity test. Mutation Res. 31, P34.~

McCann, J., Choi, E., Yamasaki, E. and Ames. B.N. (1975). Detection ofcarcinogens as mutagens in the Salmonella/miicrosome test: Assay of 300chemicals. Proc. Nat. Acad. ScT7=7"M55-:5139.

~.040

SECTION H

90-DAY ORAL TOXICITY STUDY IN RATS

IMPA


SUMMARY

Sprague Dawley-derived albino rats, both male and female, were presentedsodium IMPA in the drinking water at levels of 300, 1000 and 3000 ppm expressedas free acid. Concurrent controls were presented deionized water adjusted topH 2.5 with hydrochloric acid. Evaluation of the physical condition of therats, the body weights, the food intake, the liquid intake, hematologicparameters, clinical chemistry parameters, organ weight analysis and gross andmicroscopic examination of the tissues revealed no adverse effect of testmaterial administration at these levels.

1. OBJECTIVE

The objective of this study was to evaluate the oral toxicity of the testmaterial when administered to rats in the drinking water over a 3 monthperi od.

2. MATERIAL

Three shipments of test material consisting of 5 glass jars containing awhite powder and labeled Isopropyl Methylphosphonic acid, sodium salt (IMPA)were received from USAMBDRL by Litton Bionetics, Inc. (LBI) and designated asfollows:

LBI Date Label Number Amount UsageNumber Received Information of Jars Received (g) Dates

3427 12/18/78 Batch 1 #127818 2 550 each 10/23/80 - 12/07/803427A 11/13/80 Notebook No. 761-50 1 590 11/17/80 - 12/14/806488 12/05/80 761-67 120280 2 1010 and 420 12/15/80 - 01/23/81

The test material was stored at room temperature with the identity, purityand stability determined by the sponsor. Stability and correctness of eachformulation were determined by the LBI Chemistry Department. A newformulation was prepared every 7 to 14 days.

041


One hundred eighty weanling rats, 90 male and 90 female[CRL:COBS CD(SD)BR] were obtained from the Charles River BreedingLaboratories, Inc., Portage, Michigan on Septer.ber 24, 1980. Theanimals were acclimated to laboratory cOnditions for 29 days in theRockville facility of the Department of Toxicology before treatmentwas initiated, October 23, 1980 and were 51 days old on this date.The rats were individually housed in stainless steel wire-bottomcages in a temperature controlled animal room. A 12 hour lightcycle was maintained via artificial illumination. The feed, PurinaLaboratory Chow 5001®, was provided ad libitum except the nightprior to bleeding and terminal necropsy -wen the feed was removedfrom the cage.

The rats were randomly assigned by the use of a computer generatedrandom numbers table into their respective cages upon arrival intothe facility. All study animals were individually identified bythe use of cage tapes and ear tags (F series) as listed below.Extra animals were discarded from the study.

Averaqe Test MaterialGroup Animal Numbers Dose Intake (mn/kn/dav)Number Males Females (ppm as free acid) Males Females

1 8610-8629 9695-8714 0 0 02 8630-8649 8715-8734 300 26.3 34.R3 8650-8669 8735-8754 1000 97.n 14n.74 8670-8689 8755-8774 3000 293.1 406.0

The treated animals received the test material [the amount mixedwas corrected for sodium salt (1.159)] in their deionized drinkingwater which was acidified to a pH 2.5 with hydrochloric acid at thedoses listed above 7 days for 13 weeks. The control rats weregiven acidified deionized water.

All animals were observed twice daily for mortality and overt signsof toxicity throughout the study. Detailed observations, bodyweights, food consumption and liquid consumption (3 day period)were determined weekly. An ophthalmoscopic examination by a staffveterinarian was performed during the acclimation period to insureall study animals were free of eye abnormalities prior to studyinitiation. An ophthalmoscopic examination required during thefinal week of study was omitted.

Clinical pathology determinations were performed on the first8 rats/sex/group by orbital sinus bleeding at Weeks 4, 8 andtermination of the study. The same animals were used for allintervals. Clinical hematology was determined at all intervals andconsisted of:

erythrocyte count hemoglobinleukocyte count hematocritdifferential leukocyte count reticulocyte count

>042


Clinical chemistry determinations were performed at terminationonly and consisted of:

Glucose Serum Glutamic OxaoaceticBlood Urea Nitrogen TransaminaseSerum Glutamic Pyruvic BilirubinTransaminase PhosphorusCreatinine Calcium

A complete gross necropsy was performed on all animals at thetermination of the study. Carbon dioxide was used to kill therats. The following organs and tissues were taken and preserved in10% buffered formalin.

Tissues Saved/Examined

Any Unusual Lesions EsophagusBrain (Cerebrum, Cerebellum,* Stomach

Brainstem) Small Intestine (Duodenum, Ileum,Spinal Cord (2 sections, Jejunum)

thoracic, cervical) Large Intestine (Colon, Cecum)Eye (Left) Adrenal Glands*Pituitary PancreasSalivary Gland (Submandibular) Liver (2 sections)*Heart* Kidneys*Thymus Ovaries/Testes*Thyroid* ProstateLungs (with Mainstem Bronchi) Uteri (Corpus, Cervix)Trachea Skin (Mammary area)Spleen* Skeletal Muscle Rectus FemorisBone (with marrow) Sternum Sciatic NerveMesenteric Lymph Node Urir- ', Bladder

Body weights were taken and organ weights indicated by the asteriskwere weighed at the termination of the study.

Histopathology was performed on the listed organs and tissues ofall rats at the control and 3000 ppm dose levels. In addition, anyunusual lesions or observable effects were examined microscopicallyfrom the animals of the other two dose levels.

Statistical analysis, when comparing the differences between thetreated and control animals, was performed using Dunnett's t-test.A probability value of <0.05 was used as a basis for significance.

043

4. RESULTS

The analytical chemistry method development and results of analyseshave been included in Appendix C.

Observation of the rats during the course of the study revealed nosigns indicative of an adverse effect of test material administration.The individual animal observations have been included in Appendix A,Table H-13.

Body weight analysis revealed a slight difference between controland high dose level rats. At Week 11 the high dose males had adecreased body weight compared to control which was statisticallysignificant. There were no other statistically significantdecreases and based on the other dosages levels the decrease wasnot felt to be a test material effect. The mean body weights havebeen tabulated in Table H-5. The individual data have been includedin Appendix A, Table H-14.

Analysis of average daily food intake revealed decreases in thehigh dose males compared to control which were statistically signi-ficant at sporadic points throughout the study. The high dosefemales exhibited very slight decreases in food intake which werenot statistically significant. The increases seen at the otherdosage levels led to the conclusion the decreased food intake wasnot due to test material administration. The mean food intake datahave been tabulated in Table H-6, while the individual data havebeen presented in Appendix A, Table H-15.

No differences were observed in liquid intake which would indicaterejection of the dosed water; in fact some increases in water intakewere seen in treated rats compared to the controls. The mean liquidintake data have been tabulated in Table H-7, while the individualdata have been presented in Appendix A, Table H-16.

An approximation of dosage based on body weight liquid intake andthe mean analyzed concentrations of the drinking water. These datahave been presented in Table H-8.

Clinical hematology determinations for Weeks 4, 8 and 12 revealedno effect of test material administration on the parameters examined.The mean values have been tabulated in Table H-9, while the individualdata have been presented in Appendix A, Table H-17.

Clinical chemistry determinations mean values have been presentedin Table H-10. The individual data have been presented in Appendix A,Table H-18. There were no differences observed between control andtreated groups for any of the parameters examined.

044

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052


The organs weighed at the terminal necropsy were analyzed both asabsolute weights and as percentages of body weight. The mean weightshave been tabulated in Table H-11 and mean percentages in Table H-12. The individual organ weights and percentages have been includedin Appendix A, Tables H-19 and H-20, respectively. Kidneys weightswere higher than controls for mid dose male and female rats. Thedifferences were statistically significant for the absolute weightsbut not so when expressed as a percentage of body weight. No otherdifferences were observed other than a statistically significantincrease in absolute heart weight seen in the low dose females.These increases were not related to dosage and were felt not toindicate an effect of test material administration.

Microscopic examination of the tissues taken at necropsy revealedno findings which would indicate any adverse effect occurred due toadministration of the test material. The report of the Pathologist,Richard H. Cardy, D.V.M., has been included in Appendix B.

Thirty days after transmittal of this report, original data and acopy of the final report will be transferred to the LBI Archivist,1330 Piccard Drive, Rockville, Maryland for distribution to theproper repositories, A copy of this report and underlying datawere reviewed by the LBI Quality Assurance Unit prior to submissionto the sponsor.

K053

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5. CONCLUSION

Sprague Dawley-derived albino rats, both male and female, werepresented sodium IMP in the drinking water at levels of 300, 1000and 3000 ppm expressed as free acid. Concurrent controls werepresented deionized water adjusted to pH 2.5 with hydrochloricacid. Evaluation of the physical condition of the rats, the bodyweights, the food intake, the liquid intake, hematologic parameters,clinical chemistry parameters, organ weight analysis and gross andmicroscopic examination of the tissues revealed no adverse effectof test material administration at these levels.

Submitted by:

David R. Damske, B.A. TateGroup LeaderDepartment of Toxicology

Hands J. JBcle4,Sc.D.,. DateStudy DirectorDepartment of Toxicology

Reviewed by:

Dougla . Craig, Ph. " DateDirecto\Department of Toxicolo

056

SECTION H

APPENDIX A

* 057

T,]X:CCLOGY CBSERVATIICN RrFCRT

TABLE H-13

CLINICAL SIGNSPROJECT NUPBE-R - 1C73415

WEEK ANIMAL OBSERVATIONS : QUALIFI-O

NO. *& DOSE GROUP/SEX * NLMBER ***** COMMENTS

000 C PP" FEMALE 08711 EAR TAG MISSING

EAR TAG REPLACED : EAR-RIGHT

EAR TAG NO* 8710E

G01 MALE C8615 EAR TAG INFECTTON : EAR-RtGHT

08623 EAR TAG ImFECTION EAR-P, HT

FEMALE 08659 EAR TAG INFECTION : EAR-RIGHT

08710 EAR TAG INFECTION : EAR-Rr$HT

Tic Ppm MALE 08630 EAR TAG INCECTION : EAR-RIGHT

Cq632 EAR TAG I1vECTION : EAQRIGHT

08637 EAR TAG IFECTION : FAR-O'GHT

130 PPM 08651 EAR TAG INFECTION : EAr-P:GHT

0866_ 0ISCCLCRED URINE

RECFEMALE 08742 rAP TAG M!SSING

EAR TAG q_0DLAC-_D EaQ-LEPT

•' ,,,- 8742F

Zom0 FPP MaLEE 08676 EAR TAG INFECTION ! EAP-OIGHT

08686 EAR TAG INFECTION EAR-RIGHT

002 3 PPM MALE 08615 EAR TAG INFECTION EAO-RIGHT

FEMALE 08699 EAR TAG INFECTION EAR-RIGHT

08710 EAR TAG INFECTION EAR-RIGHT

330 FFM MALE 08633 EAR TAG INFECTION : EAR-RIGHT

086 7 EAR TAG INFECTISN : EAR-RIGHT

08628 EAR TAG INFECTION : EAR-RIGHT


1000 FPM 086!1 EAR TAG INFECTION : EAR-RIGHT

G8655 EAR TAG INFECTION : EAR-RtGHT




08669 EAR TAG INFECTION : EAP-RIGHT

3000 PPM 08676 EAR TAG INFECTION : EAP-RIGHT

C8683 EAR TAG INFEC'bON : EAR-RIGHT08686 EAR TAG INFECTION : EAR-RIGHT

C33 1 PPM UALE 08613 EAR TAG INFECTION : EAR-QIGHT

09615 EAR TAG INFECTION : cAR-PIGHT

300 FPM C4620 EAR TAG INF-CTION : EAR-OGHT096!2 CRUST : EYE-LFT:AqOUN

05I

TlXLCCL''FY CgS-,VATICN REFCRT

TABLE H-13 (CONTINUED)

CLINICAL SIGNSPROJECT NLPSER - 1C73415

WEEK ANIMAL OBSERVATIONS QUALIFIEPNC. oo DOSE GR'.UPfSEX * NLMBER . COWENTS

"33 300 PPM MALE 08635 EAR TAG INFECTION : EAR-RIGHTC8637 EAR TAG INFECTION : EAR-RIGHT08638 EAR TAG INECTtON : EAR-PrIGHT08639 EAR TAG INFECT'ION : EAP-PIGHT

FEVALE C1726 EAR TAG ISSING1000 PPq MALE 08651 EAR TAG INFECTION : EAP-PTGHT

096!3 EAR TAG INFECTION : CAP-P:GHT08655 EAR TAG INFECTION : EAP-RIGHT086E2 EAR TAG INFECTION : EAR-RIGHT08663 EAR TAG INFECTION . FAR-RIGHT08667 EAR TAG INFECTION : EAR-RIGHT

08669 EAR TAG INFECTTON : E60-OrGHTFEMALE 08739 EAR TAG INFECTION : EAR-RIGHT

3030 PPM HALE 08682 EAR TAG INcECTrOJ : EAR-RIGHT08683 -AR TAG INFr-CTION . EAR-RIGHT08686 EAR TAG INPECTION : EAR-RIGHT

FEMALE u8764 TAR TAG INFECTION t EAR-RIGHT

314 PPM MALE 18610 FASTED : OVERNIGHTSLCCC SUBMITTED CC: HEP GPAMEAR TAG INFECTION : EAR-RIGHT

08611 FASTED : OVTRN4GHTBLOOC SUBMITTED FOR HEMOGPAMEAR TAG IINFECTION : EAP-RIqHT

0612 FASTED : OVERNIGHTBLOOD SUBMITTED FOR HE"OGRAM

C8613 FASTED : OVErNIGHTOLCC SUBAITTEC FCP MEMOGRAMEAR TAG INFECTION : EAR-PIGHT

08614 FASTED : OVERNIGHT -J

BLOOD SUBMITTED FC' HEPOGPAM08615 FASTED : OVERNIGHT

BLOOD SUBMITTED FOR HEMOGRAM01616 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR HEPOGPAM08617 FASTED : OVERN:GHT

BLOOC SUBMITT-D FOR HEMOGOAMFEMALE 0869 FASTED : OVERNIGHT

BLOOD SUBMITTED FOR HEMOGPlMf08696 FASTED : OVERNIGHT

BLOOD SUBMITTED FO HEUOGvAM086S7 FASTED : IVERMIGHT

BLOCC SUBUITTED FnQ HEMOGCAM0869 FASTED : OVERNIGHT

~ .T

T:XIC%LGY CBSERVA'ICI .EFCRT

3

TABLE H-13 (CONTINUED)CLINICAL SIGNSPRCj'crC NL9ER - 1.'73415

W--K ANIMAL OBSERVATIONS : GUALTFIE

NO. &* DOSE GROUP/SEX * N4L4ER *.... COMMENTS *** ***"

o4 3 PP FEMALE BLOOD SUBMITTEC FOR HEMOGPAMCRUST EY--RIGHT:ARCUNO

.. **, . r

08699 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR HEVOGOAM08700 FASTED : OVERNIGHT

9LOOC SU9MITTED FOR HEMOGRAM

0701 FASTED : OVERNIGHTBLOOC SUBMITTED FOR HEMOGRAM

08702 FASTED . OVERNIGHT

OLOCC SUB"ITTED FOR HEP0I*RAMCRUST : EYE-RIGHT:tROUNDRED

,q714 EAR TAG MISSINGEAR TAG R-PLACED : AR-PIGHT

330 PPM MLALE 0863C FASTED : OVERNIGHT8LOOC SUBMITT-ED FOP HEMOGPAM

EAR TAG INFECTION :. Sg-prGHT

0C631 FASTED : OVERNIGHT

LOO SUBMITTED FOR HEMOGRAM


BLOCC SUBMITTED FOR HEMOGRAMCRUST : EYES:AROUND

C8633 FASTED : OVERNIGHTBLCOC SUBMITTED FOR HE*OGRAM

08634 FASTED : OVERIrGHTBLOOC SUBMITTED FOR HEMOGRAM

EAR TAG INFECTION : EAR-RIGHT

08635 FASTED : OVERNIGHTBLOOD SUpoITTED FOR HEMOGOAMEAR TAG INFECrION : EAQ-0 tGHT

08616 FASTED : OVERNIGHTRLOC SUBMITTED FOR HEMOGPAM

08637 FASTED : OVERNIGHTBLOOC SUBMITTED FOR HEMOGPAM


08648 EAR TAG INFECTION : EAQ-RIGHT

FEMALE 08715 FASTED : OVERNIGHTBLOOC SUP8!ITTED FOq HEfdOGP.AM

C871 FASTEO : OVER:IGHT8LOT0 SUBMITTED FOR HEOGPAM

08717 FASTED : OVERN1IGHT

060

TCXICCL^JGY IBSERV,1T!CN .REF1RT


CLINICAL SIGNSPRIJECT NLOSER - 10734115

WEEK ANIMAL OBSERVAT!ONS : aUALIFIEPNO. -~ 3CSE GROUP/SEX * NLPSER *'.C04mENTS

a34~ 300 PPM FEMALE BLOOD SUBMITTED FOR HEMOGRAM08718 FASTED : OVERNIGHT

RLOOC SUBMITTED FOP HEMflGPAP08119 FASTED : OVERNIGHT

BL0CC SUBM4ITTED FOR HEMOGOAMoq720 FASTED :OVERNdIGHT

C71 BLOOD SURMITTT:0F!: H ?lG~

BLOOD SUBMITTED FOq HEVOGPAMC8722 FASTED : OVERNIGHT

E10CC SUBMIZTTED FOP HEPMCGRAM08724 EAR TAG INFECTION :EAR-RIGHT08726 EAR TAG ?!SSING

rAR TAG REPLACED : EAQ-LEFT#.~ 87260

a8728 EAR TAG INFECTION 1 EAR-PIGHT1301 PPM MALE 08650 FASTED : OVERNIGHT

BLOOD SUBMITTED FOP HEMOGPAM086!1 FASTED VDOH

BLOOD SUBMITTED FCR HEPOGOAM08652 FASTED : OVERNIGHT

BLOOD SURMITTEO FOR HEOGA086!3 FASTED :OVERNIGHT

BLOOD SURMITTED FCP HENOGPAMEAR TAG INFECTION :EAR-rGHT

C86!4 FASTED :OVERNIGHTOLCOC SUBMITTED FOP HEPMOGPAM

08655 FASTED :OVERNIGHTBLOO SUBMITTED FOR HEMOGPA'4EAR TAG INFECTION :EAP-PIcHT

086!6 FASTED : OVERNIGHTBLOOD SUBM4ITTED FOR HEmOGQAM

G8657 FASTED : OVERNIGHTBLCOC SUBM4ITTED FOP HEMCGRAMEAR TAG INFECTION :EAP-RTf.HT

08659 EAR TAG I'JFECTION : EAP-PIGHT08662 EAR TAG INFECT:ON : EAR-PGHT19663 EAR TAG INFECTION :ED-RIGHT09665 EAR TAG INFECTION : EAR-RIGHT08667 EAR TAG INFECTION :EAR-RIGHT08667 EAR TAG INFECTION :EAR-RIGHT

FEMALE 00735 9ASTr-C : OVEP'IGHTSLOOC SUBMITTED FCC HEMOGDAM

08736 FASTED: OVERNIGHT

061

TCYICCLCGY CRSE'RVATICN REFCRT


CLINICAL SIGNSPROJECT NLPBER - IC73415

UE:-KANIMA*L OBSERVATIONS C UALIFIER

No. ** DOSE GROUPISEX & NLMgER COMmENTS **..**

004 1000 PPM FEMALE BLOOD SUBMITTED FOq HEPOGPAM

09737 FASTED : OVERNIGHTBLOOD SUBMITTED FOR HEP'OGRAM

C87!8 FASTED : OVERNIGHTBLOOD SUBMITTED FOR HEmOG;;AM

EAR TAG INFECTION : EAR-P!GHT

C37!1 FASTED : OVERNIGHT9100C SUBMITTE0 FOP NEPOGPAM

CgT40 FASTED :OVERNIGHT

ELCOC supmrTyrn FCo HEvOSCAM01741 FASTED : 'YVEQNIGHT

9L0O SUBMITTED FOR PEMOGPAM9742 FASTED : OVERNIGHT

RLCCC SUemITTED FOO PEPOGPAP

300 PPM MALE 03670 FASTED :OVERNIGHTBLOOC SUBM!TTE-D FOR HEMOGPAM

C1671 FASTED : OVrPNIGHT

ELCOC SUBMITTED FCO HEA.M046'72 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR HEMOGPAM

C8673 FASTED : OVERN7GHT

BLOCC SUBMITTED FOP HEPOGPAM

09674 FASTED : OVERNIGHTBLOOC SUBMITTED FOP HEMOGRAM

08675 FASTED : VERsIGHTRLCOC SUBMITTED FOP HEPOGPAM

09676 FASTED : VERNIGHT

BLOOC SUB41TTED FOq HM'MOGPAM

08617 FASTED : OVERNTGHT

BLTOC SUBMITTED FOP H E ?O0GPA M

08679 EAR TAG INJFECTTON : AP-01? HT

08679 EAR TAG INFECTTON : EAP-R7GHT

08682 EAR TAG INFECTION :EAR-RTI'HT


08686 EAR TAG INFECTION :EAR-RIGHTFEMALE 08755 FASTED : OVFPRNIGHT

PLCCC SUBmITTED Fic HEUCrvPA~


BLOOC SUBMITTED FOR MEu GPAM

087!7 FASTED : OVFQNIGHT

BLGO SUBMITTED FOR H 0 ~~A F087!8 FASTED :OVEDNIGHT

9LOOD SUR"!TTE0 FOO H!M OGDAM

097!9 FASTED :OVERNIGH'

TCXICCLOGY CBSERVATION R-PCRT

TABLE H-13 (CONTINUED)CLINICAL SIGNSPQ)JECT NUPER - 1C73415

WEEK ANIMAL OBSFRVATIONS QUALIFIER

NO. w* DOSE GRCUPISEX * NLMBER " COMMENTS

004 3000 PPM FEMALE BLOOC SUBMITTED FOR HEMOGPA"08760 FASTED : OVERNIGHT

BLOOC SUBMITTEO FCC HE OGPAM

08761 FASTED : OVENIGHTBLCOC SURMrTTED FOR HEwOGPAM

08762 FASTED : OVE9NIGHT

RLCOC SUBuITTED FOP HEMCGOAM

09764 EAR TAG INFECTION : EAP-RTGHT

075 J PP" MALE 0861S EAR TAG INFECTION : 7-AP-RIr H T

C0611 EAR TAG INCECTION : EA-0IrHT


08625 EAR TAG INFECTION : EAQ-R.GHT

330 PPM -6310 EAR TAG INFECTION : EA-RIGHT

C96!2 CRUST : EY--RIrHT:A'P0UNO,**** RED08634 EAR TAG IcrECTION : -AR-RIGHT08639 EAR TAG INVECTION : EAR-Q:GHT0 6!9 EAR TAG INCECTION " EAP-RIGHT

FEMALE 08728 EAR TAG INFECTION : EAR-RIGHT

08732 EAR TAG INCECTION : EAR-P:GHT

1000 FPM MALE 08653 EAR TAG INFECTION : EAR-RIGHT

0865n EAR TAG INFECTION ! EAR-0IGHT

08657 EAR TAG IN-ECTION : EAR-RIGHT

08659 EAR TAG INFECTION : EAR-RIGHT08663 EAR TAG INFECTION : EAP-RIGH

T

08664 EAR TAG MISSING08665 EAR TAG INFECTION : EAR-RIGHT

38667 EAR TAG INFECTION : EAP-RIGHr

FEMALE 087!9 EAR TAG INFECTION : EAP-GISHT

3000 PPM MALE 08618 EAR TAG INFECTION : EAP-PIGHT

08679 EAR TAG IFECTION : EAR-QIGHT





C8689 EAR TAG INFECTION : EAR - R IGH T

106 3 PPM MALE 08610 EAR TAG INFECT ION : EAR-RIGHT

08611 TH:CKENED EAR(S) : EAR-OIGHT

CR61. EAR TAG INFECTION : EAR-RIGHT

Cq614 EAR TAG INFECTION : EAR-RG HT

08615 ER TAG INFECTION : r'AP-PIGHT

08624 THICKENED rARCS) : 1AP-019HT

063

TOXICOLOGY OBSERVATION QEFCRT

TABLE H-13 (CONTINUED)CLiNICAL SIGNSPROJECT NLPRER - 1C73415

WEEK ANIMAL OBSERVATIONS QUALIFIER

NO. -" DOSE GROUP/SEX * NLMBER -**. COMMENTS

006 0 PPM MLE 0862 EAR TAG INFECTION : EAP-PIGHT300 PPM 08630 EAR TAG INFECTION : EAR-RIGHT

08632 DISCHARGE : EYE-RIGHT086!4 EAR TAG INFECTION : EAq-RIGHT

C6! EAR TAG INFECTION : EAR-RIGHT

08637 EAR TAG INFECTION : EAR-QIGHT08639 EAR TAG INFECTION : EAR-RIGHT

08642 EAR TAG INFECTION : EAR-PISHT08649 EAR TAG INFECTION : EAR-PIGHT08649 THICKENED EAR(S) : EAq-RIGOT

FEMALE 08726 LACRIMATION : EYE-cIGHT08728 EAR TAG INFECTION : EAP-OIGHT

C8732 THICKENED rAR(S) : FAR-RIGHT

1030 PPM MALE 09653 EAR TAG INFECTION : EAR-RIGHTS8653 EAR TAG INFECTION : ES-R!GHTC86!7 EAR TAG INFECTION : EA-PoIGHT

086!9 EAR TAG INCECTION : EAR-DIGHT2662 EAR TAG IN'=ECTION : E30-O'GHT

C9663 EAR TAG INFECTION : EAR-RIGHT08664 EAR TAG MISSING

08665 EAR TAG INFECTION : EAR-PIGHT08667 THICKENED EAR(S) : EAR-RIGHT08669 THICKENED EAR(S) : EAR-OIGHT

FEMALE 007!8 EAR TAG INFECTION : EAR-DIGHT

0740 THICKENED EAR(S) : EARS-RCTHi0oo PPM MALE C?671 LOCAL HAIR LOSS : LIMPS-FOPE

C8678 EAR TAG II=ECTION : EAR-ZIGHT

C8679 EAR TAG INFECTION : EAR-RIGHT

08680 EAR TAG INFECTION : EAR-QIGHT08681 THICKENED EAR(S) : EAR-RIGHT08682 EAR TAG INFECTION : EAQ-RIGHT08683 THICKENED EAR(S) : EAR-RIGHT08685 EAR TAG INFECTION : EAR-RIGHT


08688 THICKENED EAR(S) : EAR-RIGHTFEMALE 08755 ROUGH HAIR CCAT

CPT63 EAR TAG MISSING01764 EAR TAG INFECTION : E1 0 -QIGHTaq7£6 LACRIMATION : EYES-BOTH08772 EAR TAG INFECTION : EAD-0IGHT

017 3 PPM MALE 0910 EAR TAG INFECTION : EAR-RIGHT02611 THICKENED TAR(S) EAP-01GHT09613 EAP TAG INFECTION : EAP-P!GH4

064

TCXICOLCGY 08BSER'AlCN RF.CRj

3

TABLE H-13 (CONTINUED)CLINICAL SIGNSPROJECT NLMBER - 1'73419

WEEK ANIMAL OBSERVATIONS Q CUALIFIEDNC- * QSE GRCUP/SSX * NL!8ER COMMENTS *

007 0 PP" PALE 08614 EAR TAG INFECTION : EAR-RIGHTM86I5 EAR TAG INFECTION : EAR-RIGHTCe62! EAR TAG I!'PFCTION EAP-PInHT

300 PF" 08630 EAR TAG INFECTION : EAR-RiGHT

38632 CRUST : EYE-LCFT:AROUN0•*t.*. RED

86!4 EAR TAG VFECTION : ;-'R-4IGHT0863 EAR TAG INFECTION : EAP-qIGHT08617 EAR TAG INcECTtON : EAI-PIGHT

C86! EAR TAG INFECTION : -AP-RIGHTCR642 EAR TAG INFECTION : EAR-RIGHT

09643 EAR TAG INFECTION : SAP-OIGHTC8648 EAR TAG INFECTION : EAR-RIGHT

FEMALE 08732 ,rAR TAG INCECTION : EAR-RIrHT1001 PPM MALE 086!2 EAR TAC PISS IP

08653 EAR TAG INFECTION : EA;- OHTC86!5 EAR TAG INFEC"ION : FAR-DIGmT18651 EAR TAG INFECTION : E4 0-PIGH "

C?6!9 EAR TAI I*IFECTION : EAR-RIGHT08662 EAR TAG INFECTION : EAR-RIGHT08663 EAR TAG INFECTION : EbR-RqHT

CRUST : EYE-RIGHT:APOtAO*. ,, REC

08664 EAR TAG MISSINGEAR TAG REPLACED EAR-RIGHT

08665 EAR TAG I'JFECT:ON : EAR-RIGHT08667 EAR TAG INFECTION : EAQ-PIGHT08663 EAR TAG INFECTION EAR-RIGHT

FErALE 081!7 CCNVULSIONS-CLONICEXCESSIVE SALIVATION

08738 EAR TAG INFECTION EAR-RIGHT08740 THICKENED fAR(S) : SAR-RIGHT

3000 PPM PALE 08671 LOCAL HAIR LCSS : LIMPS-FCRE08679 EAR TAG INFECTION t EAR-RIGHT086T EAR TAG rNFECTION : EAR-RIGHT08610 EAR TAG INECTrION EAD-RIGHT18681 EAR TAG INFECTION SAR-RIGHT

08682 EAR TAG INF"CT:ON : EAR-PIGHT08685 EAR TAG INPECTION : EAP-RIGHTC2686 EAR TAG INFECTIMN FAR-Ir-HT08688 EAR TAG INFECTION EAO-RICHT

FEMALE 08755 ROUGH HAIR CCAT

08763 EAR TAG MtSSING

065

TOEECL0GY OBSE-RVATICN RSPCRT

9

TABLE H-.13 (CONTINUED)CLINICAL SIGNSPROJECT NL"BER - 1- 73419

WJEEK A A IMAL OBSERVATIONS : UALIFIF;N1). ** DOSE GROUPISEX t NLP9ER COMM4ENTS

ofl 3000 PPM FEMALE EAR TAG REPLACED :EAR-LEFT

08764 EAR TAG INFECTION :EAP-PIGHT08770 EAR TAG INFECTION :EAP-PIGHTC8772 EAR TAG INFECTION 1 EAP-PISHT

GIB a PP" PALE 08610 FASTtED : OVERNIGHT9LOOO SUBMTTTED FOR HEMOGPAM

38611 FASTED : OVERNIGHTeLCOC SU9MITTED FCO HEPCGP.AM

C8612 FASTED : 3VERNIGHTRLOOC SURMITTVED FOR HEMOGPAM

CP613 FASTED :OVTRNIGH'81000 SUB'MITTEDO FCP MEViOGPAM

01614 FASTEO OVERNJIGHTBLOOC SUBM4ITTED FIR HEMOGRAM

08615 FASTrED : 'VrRP.IGH T

SLCOC SUBMITTED FOO HEP0'$;AmEAR TAG rlNFECr:'ON ADIH

08616 FASTED : OVEPMIGHTPLOOC SUBOITTrO FCR HEP'CGPAT


9LOOC SUBMITTrED FOO HET MOG0AP08624 SAR TAG INFECTION : EAR-RIGHT0862n TEETH CUT

EAR TAG INFECTTON : EAR-RIGHTCRUST :EYE-LE:FT:APOUND

***RD

FEPALE 08695 FASTED :OVERNIGHTBLOOD SUBMITTED 900 HEMOGDAM

08696 FASTED : OVERNIGHTBLOOC SUBMITTED FOR HE"OGF.AM

08697 FASTED :OVEPNIGHTBLOOD SUBMITTED FOR HEOGPAM

G8698 FASTED : OVERNIGHTBLOOD SUBMITTED FOR HEOG'rM

C86S9 FASTED : VYFNIGHTBLOOD SUBMITTED FCP MEPSOGOAM

28700 FAST!ED : OVERNIjGHTBt.OOC SUIBMITTED FOR HEMOGFAM

08701 FASTED : OVERNIGHTBLCC SUBMITI-70 FOR HEPOGPAM

08702 FASTED : 0V9:R'IIGHT

RLOOD SU9M4TTTE') FOP HEMOG04M

T3XICOLOGY OBSERVATICN REF RT

TABLE H-13 (CONTINUED)CLINICAL SIGNSPROJECT NLMBER - 1 .73419

WEEK ANIMAL OBSERVATIONS aUALIFIERNO. -" DOSE GROUP/SEX * NUMBER **-** COMMENTS

008 a PPM FEMALE 08704 EAR TAG INFECTION : EAR-RIGHT300 PPM VALE 08630 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR HEMOGPAM

EAR TAG INFECTION : EA- IGHT08631 FASTED : OVERNIGHT

BLCCC SUBMITTED FOR HEMOGPAM38632 FASTED : OVERNIGHT

BLOOD SUBMITTED FOR HEMOGPAM0RS33 FASTED : OVERNIGHT

BLOOC SUBMITTE-D FO P HFMOGPAM

0863 4 FASTED : OVERNIGH T

BLOOC SUBMITTED FOP HEMOGPAMEAR TAG INFECTION : EAR-RIGHT


BLOCC SUB"ITTrO PIP HEMOGAM08636 FASTED : OVERNIGHT

BLCOC SURMITTED FOR HEMOGPAM

CRUST : EYT-R:GHT:AROLND

C9637 FASTED OVEPNIGHT

LOOD SUBMITTED FO? HEMOGPAMEAR TAG INFECTION EAP-RIGHT

053639 EAR TAG INFECTION EAR-RIGHT08642 EAR TAG INFECTION EAR-RIGHT08646 EAR TAG MISSING

EAR TAG REPLACED EAP-LEFT**** 8646F

08648 EAR TAG INFECTION EAR-QIGHTFETMALE 08715 FASTED : OVERNIGHT

RLOOC SUBMITTED FOR HEMOGRAM08716 FASTED : IVTRNIGHT

BLOOC SUBMITTED FCR HEMOGDAM08717 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR HEMOGPAM08718 FASTEO : OVERNIGHT

BLOOD SUBMITTED FCQ HEPOGPAM08719 FASTED : OVERNIGHT

BLOOD SUBMITTED FOR HEMOGPAMCRUST EYZ-RrGHT:AqC2ND


BLOOC SUFMITTEO FOP HEvfGcAvC8721 FASTED : OVERNIGHT

BLGOC SUBMITTED FOR HEMOGQAM

067

TOXICOLOGY OBSERVAlIION REPORT

TABLE H1-13 (CONTINUED)CLINICAL SIGNSPROJECT NUMBER - IC73415

WEEK ANIMAL OBSrRVATIONS : UAL!FIEPNO. ** DOSE GROUP/SEX * VJUP'ER *. COM4MENTS

008 300 FPM FEMALE 08722 FASTED : OVFRN!GHTSLOOD SUBMrTTE:D FOP HEMOGOAMCRUST EYE-RrGHT:AROUNO

0*.RED

1300 PPM MALE 08650 FASTED :OVE RNISHTBLOOC SUBMITTrED FOR HEMOGOAM

08651 FASTED : OVERN~IGHTBLOOD SUBWITTEDr FIR HMOSPAMEAR TAG INFECTION :EPR-RIqHT

08652 FASTEO ' VSRNIGHTBLOOC SUBMITTED FOR HEv0GPAMEAR TAG MISSINIEAR TAG REPLACED :EAR-L'7T4SF*NB52A

CRUST EYr-RlGHT:&0OUN0

09653 FAST70 : fVSRNIGHTBLCO SUBMTTTEI FCQ HEMOGPAM

33654 FASTED : JVSRNIGHTBLOOC SUBMITTED0 FOR HEKOGRA4M

CR6!5 FASTED : 'IVERNIGHTBLOOC SUBMITTED FOr HEMOGDAMEAR TAG INFECTION :EAR-RIGHlT

38656 FASTED : OVERNIGHTBLOOD SUBMITTED FOO HEMOGPAM

0865T FASTEO OVERNIGHTBLOOD SUBMITTED FOR HE,"OGPAM

086t9 EAR TA6 INFECTION :EAP-PTGHT08661 EAR TAG INFECTION :EAP-RIGHT0166? EAR TAG INFECTION : EAR-RIGHTa9663 TEETH CUT

PAR TAG INFECTION :EAR-PIGHTERUST : EYE-RIGHT:A;ROLNO

***RED08665 EAR TAG INFECTION :EAR-RIGHT08607 EAR TAG INFECTION :EAR-Rlr-HT08669 EAR TAG INFECTION :EAR-P:GHT

FEMALE 08735 FASTED :OVERNIGHTBLOOD SUBMITTED FOR HEPOGDAM

087!6 FASTrD :OVERNIGHTLCV3C SUBMITTED FICR HEPOGRAM

087!7 FASTED : IVERMIGHT

8LOOC SUBMITTEO0 FOR HEMOGPAM19734 FAST7D : VED%IGHT

TCXICCLOGY COBSERVATION REFCRT

2


CLINICAL SIGNSPR3JECT NUMBER - J.a73419

WEEK ANI1AL OBS'RVATIONS : QUALIFIEQNO. a. DOSE GROUP/SEX * NLOBER ..... COM'4vET,

008 1000 PPM FEMALE BLOOO SUBMITTED FOR HEMOGRA4

EAR TAG INFECTION : EAR-RIGHTCRUST , EYE-RIGHT:AROUND

.1739 FASTED : OVERNIGHTSLOOC SUBMITTED FOR HEMOGRAMEYE CPACITY-PIGHT cYECRUST EY_-RIGHT:A9OUNO

•**' ,- BROWN08743 FASTED : OVERNIGHT

SLOC SUBMITTF) F0 HSMOGPAMC8741 FASTED : OVERNIGHT

BLOO SUBMITTED FOR HEOGPAMa8142 FASTED : OVERNIGHT

BLCCC SUBMITTED F0o HSMOGPAM3000 PPm VALE 08670 FASTED : OVERNIGHT

BLOOD SUBMITTED FOP HEmOGRAM08671 FASTED : OVERNIGHT

BLOCC SUBMITTED FIR HEOJGRAM39672 FASTED : OVERNIGHT

SLOOC SURMITTEO FOR HEOGRAM08673 FASTED : OVERNIGHT

BLOCC SUBMITTED FOR HEP'3;PAM08674 FASTED : OVERNIGHT

BLOOD SUBMITTED FOR HEMOGDAM08675 FASTED : OVERNIGHT

BLCCC SUBMITTEr' FCQ HEUOrPA.M08676 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR HE"OGPAM

08677 FASTED : OVERNIGHTBLCOC SUBMITTE1 F0 HEMOGPAM

08679 EAR TAG INFECTION : EAR-RIGHT08680 EAR TAG INFECTION : EAR-RIGHT08681 EAR TAG INFECTION : EAR-0IGHT08622 EAR TAG INFECTION : EAR-RIGHT08685 EAR TAG INFECTION : EAR-RIGHT

08686 EAR TAG INFECTION : EAR-QIGHTFEM0LE 08759 FASTED : OVERNIGHT

BLOCC SUBMITTED FOR HEMOGRAM08756 FASTED : OVERNIGHT

BLOOC SUBMITTED FOr HEMOGRAM

08717 FASTED : OVERNIGHTBLOOD SUBMITTED FOP HEPOGPAM


069

TOXICOLOGY ORBSRVA1ON REPrRT

3

TABLE H-13 (CONTINUED)CLINICAL SIGNSPRCJECT NLBER - 1C73415

WEAK NIMAL OBSrRVATIONS QUALIFIEQNO. ** DOSE GROUP/SEX * NUMBER , COMMENTS

008 2000 PPM FEMALE BLOOD SUBMITTED FOR HEPOGPAM08759 FASTED : OVERN!GHT

BLOOD SUBMITTED FOR HE.MOGRAM08160 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR HE"OGcAM00761 FASTED : OVEPN:GHT

BLOOC SUBMITTED FOR HEMOGPAM08162 FASTED : OVERNIGHT

BLOOCD SUBMITTED FOP HFMOGDAMCRUST : rYE-qIGHT:A0LUNDRCD

08764 EAR TAG INFECTION : EAP-PIGHTa872 EAR TAG INFECTION t EAR-RIGHT

039 1 pp" MALE 0861 EAR TAG INFECTION : .AR-RIGHTC8625 TEETH CUT

CRUST : EYE-LEFT:APOUNO• -,*. RF C

OR627 EAR TAG INFECTION : EAR-PIGHT3GO PPM 086 3 EAR TAG INFECTION : EAP-DIGHT

08634 EAR TAG INFECTION : EAP-RIGHT086V EAR TAG INFECTION : EAR-RIGHT08642 EAR TAG INFECTION : EAR-PIGHT

C8649 EAR TAG INFECTION : EAR-RtGHT

1000 PPM 086'51 EAR TAG INFECTION : EAR-RIGHT08655 EAR TAG INFECTION : EaR-RIGHT08659 EAR TAG INFECTION EAP-RIGHT08661 EAR TAG INFECTION : EAR-RIGHT08663 TEETH CUT

EAR TAG INFECTION : EAR-RIGHT08665 EAR TAG INFECTION : EAR-RIGHT08667 EAR TAG INFECTION : EAP-RIGHT08669 EAR TAG INFECTION " EAR-RIGHT

FEMALE 08735 EAR TAG INFECTION : EAR-RIGHT08738 EAR TAG INFECTION : EAR-PIIHTCe739 EYE OPACITY-RIGHT LYE

CRUST : EYE-RIGHT:AROUND***.. RROWN

3000 PPM MLE 08611 LOCAL HAIR LOSS : LIMES-FOPE08679 EAR TAG INFECTION : EaR-RIGHT08680 EAR TAG INPECTION : EAR-QIGHT08682 E.AR TAG INFECTION : EAR-PIGHT0869!, EAR TAG INFECTION : EAP-RIGHT08686 EAR TAG INFECTION I EA-RIGHT

070

TOXICOL3GY OBSERVATI^N REFCRT


CLINICAL SIGNSPR13JECT NLPBER - t:73415

WEEK ANIMAL OBSERVATIONS OUAL:FIENC. ** DOSE GROUP/SEX * NLPBER *'' COMMENTS

339 3000 PPM FEMALE 08764 EAR TAG INFECTION : EAR-RIGHT08772 EAR TAG INFECTION : EAR-RIGHT0C714 CRUST : EYE-LFFT:AQOUhO

RED

010 0 PPM MALE 092515 EAR TAG INFECTION : EAR-PI'HT08625 TEETH CUT

08627 EAR TAG INFECTION : EAP-RIGHT300 PON 396!4 EAR TAG TNFECTION : EAR-OIGHT

C9637 EAR TAG INFECTION : EAR-RIGHT08642 EAR TAG INFECTION : EAP-R:GHT

1000 PPM 08651 EAR TAG INJFECTION : EJR-RIGHT08655 EAR TAG INFECTION : EAR-RIGHT08659 EAR TAG INFECTION : EAR-RIGHT08661 EAR TAG INFECTION : EAP-RTGHT086E3 TEETH CUT08667 EAR TAG INFECTION : EAR-RTIGHT

FEMALE 08739 EYE OPACITY-RIGHT FYECRUST : EYV-PIGHT:AROUtOBROWN

3000 PPM mALE 08670 EAR TAG MISSING08671 LOCAL HAIR LOSS : LIMBS-FOPE08679 EAR TAG INFECTION : EAR-RIGHT08680 TEETH CUT

CRUST : EYE-LEFT:AROUND• **.* RED08682 EAR TAG INFECTION : EAR-PIGHT08685 EAR TAG INFECTION : EAR-RIGHT08686 EAR TAG INFECTION : EAR-RGHT

FEMALE 08766 EAR TAG MISSING08772 EAR TAG INFECTION : EAR-RIGHT

011 3 PPm PALE 08612 EYE DISCHARGE : EYE-LEFT• **** CLEAR08q615 EAR TAG INFECTION : EAR-RIGHT

SMALL ULCERATION(<ICM) : EAR-RIGHT:RELCo**** W/ CRUST

08617 EAR TAG MISSING08625 TEETH CUT

EAR TAG INFECTION : EAR-RIGHT08627 EAR TAG INFECTION : EAR-RIGHT

FEMALE C8707 EAR TAG INFECTION : EAR-RIGHT08711 TEETH CUT

CRUST : EYE-LEFT:A9QOUNDBROWN

071

TCXICOLOGY OBSERVATICNJ REFCRT

TABLE H-13 (CONTINUED)CLINICAL SIGNSPROJECT NLO8ER - lC73415

WEEK ANIMAL 08SERVATIoNS CUALIFIEPNO. **DOSE GROUP/SEX * NILMBER ** COMMENTS

0ll a PPM FEMALE 08712 TEETH CUT300 PPM PALE 08630 TEETH CUT

EAR TAG INEFECTION : EAR-RIGHTC9634 EAR TAG INFECTION : EAR-RIGHT08637 EAR TAG INFECTION :EAR-RIGHT08638 TEETH CUT01642 EAR TAG INFECTION :EAP-RIGHT0647 EAR TAG INFECTION :EAR-RI6HT

FEMALE 08722 EAR TAG INFECTION :EAP-QIGHT00723 1EYE oISCHAPGE : YE-LEFT***** CLEAR0!725 SKIN FLAKY :TAIL08728 SKIN FLAKY TAIL

lion PPM M'ALE 08651 EAR TAG INFECTION 1 EAR-2IGHT0896!S EAR TAG INFECTION : AR-R'GHT06657 EAR TAG INFECTION :EAP-RIGHT08659 EAR TAG INFECTION :EAR-RIGHT08661 EAR TAG INFECTION :EAR-RIGHT08662 TEETH CUT08663 TEETH CUT

EAR TAG INFECTION :EAR-RPIGIH08665 EAR TAG INFECTION : EAP-RIGHTC8666 EAR TAG INFECTION :EAR-RIGHT08667 EAR TAG INFECTION :EAQ-RIGHT

FEMALE 08739 LOCAL HAIR LCSS :LIMB-FORELEFTCRUST : EYE-RIGHTON CORNEACRUST : EYE-RIGHT:APO'ND

***BROUN08748 SKIN FLAKY : TAIL

ULCEFATION : MULTIPLE SITES*.'TAIL

3000 PPM MALE 08679 EAR TAG REPLACED :EAR-LEFT***.* 8670A08671 LOCAL HAIR LOSS :LIMES-FOOE09673 TEETH CUT

EAR TAG INOECTION :EAR-RIGHT09679 EAR TAG INFECTION :EAR-RIGHT08684" TEETH CUT

CRUST : EYE-RIGHT:3ROUND08682 EAR TAG INFECTION :EAR-RIGHT08693 EAR TAA INFECTION : EAR-RIfHT08685 EAR TAG INFECTION : EAR-RIGHT08686 EAR TAG INFECT1I : EAR-QIGHT

072

T3XICGLOGY CBSTRVTION REPORT

16

TABLE H-13 (CONTINUED)CLINICAL SIGNSPROJECT NtMHER - 1G7341g

WEEK ANIMAL OBSERVATIONS : CUALIFIEPNO. ** DOSE GROUP/SEX * NL"BER .. *." COMMENTS

011 3000 FPM MALE 08688 EAR TAG INFECTION : EAP-RIGHT08689 EAR TAG INFECTION : EAR-RIGHT

FEMALE 08761 SMALL ULCERATON(<ICV) : MULTIPLE SITES****w TAIL

08764 EAR TAG IMCECTTON 1 EAP-0GHT

08766 EAR TAG REPLACED : EAF-LEFT**a'* 87664

012 0 PPM MALE 08612 CRUST : EYE-LEFT:AROUND08615 EAR TAC- INFECTION : EAR-RIGHT

SMALL ULCERATICN(<1Cm) : EAR-RIGHT:REL(W*b.** WI CRUST

09616 CRUST : EYr-LEFT:APOUND08617 EAR TAG REPLACED EAR-RIGHTa.... 8617N

08625 TEETH CUTEAR TAG I'FECTION ! EAR-RtGHT

C8627 EAR TAG INFECTION EAR-PIGHTFEMALE 08711 TEETH CUT

08712 TEETH CUT08714 TEETH CUT

301 PFM PALE 08630 THICKENED EAR(S) E AR-RIGHT08634 EAR TAG INFECTION : EAR-RIGHT08637 EAR TAG INFECTION : EAR-RIGHT08642 EAR TAG INFECTION : EAR-PIGHT08647 EAR TAG INFECTION : EAR-RIGHT

FEMALE 08722 EAR TAG INFECTION : EAR-DIGHT08725 SKIN FLAKY : TAIL08729 SKIN FLAKY : TAIL

1000 PPM MALE 086!1 EAR TAG INFECTION : EAR-RIGHT08657 EAR TAG INFECTION : EAR-RIGHT08661 EAR TAG INFECTION : EAR-RIGHT08662 TEETV CUT08663 TEETH CUT

EAR TAG INFECTION : EAR-RIGHT08665 EAR TAG INFECTTON : EAR-RIGHT08666 EAR TAG INFECTION : EAR-RIGHT08667 EAR TAG INFECTION : EAR-RIGHT

FEMALE 08735 EAR TAG INFECTION : EAF-PIGHT087!7 THICKENED EAR(S) : EARS-BOTH08739 LOCAL HAIR LOSS 1 LIMS-FRE

LICAL HAIR LOSS : EYE-RIGHT:AROUNDCRUST : EY-RIGHT

..... R C, ON CORNEA

~073

TOXICOLOGY OBSERVATION REPORT

17

TABLE H-13 (CONTINUED)CLINICAL SIGNSPROJECT NLMBER - 1C73415

WEEK ANIMAL ORSERVATIONS : GUALIFIERNO. *' DOSE GROUP/SEX * NLMBER "*. COMMENTS **..*.,

012 1000 PPM FEMALE 08740 THICKENED EAR(S) : EARS-ROTHENLARGED : EAR-RIGHT

08748 SKIN ABNORMAL : TAILFLAKYSMALL MASS(<ICM) : NOSE:ABOVEFIRIASMALL SCA8(<lCM) : TAILSEVERAL

C97!4 EAR TAG INFECTION : EAP-RIGHT3000 PPM mALE 08671 LOCAL HAIR LC3S : LIMES-FCPE

08673 EAR TAG INFECTION " EAP-P:GHT28679 EAR TAG INFECTION : EAR-RIGHT08680 TEETH CUT

CRUST : EYE-RIGHT:AROUNO- REC

C8681 THICKENED EAR(S) : EAR-RIGHT0.682 THICKENED EAR(S) : EAR-RIGHT08683 EAR TAG I4FECTION : EAR-q!GHT28685 EAR TAG INFECTION : EAP-RIGHT08686 EAR TAG INFECTION : EAP-RIGHT00688 THICKENED EAR(S) : :7AR-RIGHT

08689 THICKENED EAR(S) : EAR-RIGHIFEMALE 08760 THICKENED TAR(S) : EAR-RIGHT

08761 SMALL SCAB(<lCM) : TAIL***** SEVERAL08764 EAR TAG INFECTION : EAR-RIGHT

013 0 PPM MALE 08610 FASTED : OVERNIGHTBL0OC SUBMITTED FOR C'EM AND HEMnqPAV

08611 FASTED : OVERNIG"'BLOOC SUBMITTr3 - CHEM AND HEMCGQAM

08612 FASTED : OV " AG."ELCOC SUBMT- _u FC- -HEM AND HEMCGRAM

08613 FASTED : OVERNIGHTBLOOD SUBMITTED FOR CHEM AND HEMCG9AM

00614 FASTED : OVERNIGHTeLOOC SUBMITT-D FOR CHEM AND HMCGVAM

08615 FASTED : OVERNIGHTBLOOD SU MITTED FOR CHEM AND HEMCGRAM

08615 FASTED : OVERNIGHTBLGO SUBMITTED FOP CHEM AND HEMCGPAM

09617 FASTED : OVERNIGHTBLOOC SUBMITTED FOR CHEM AND H-MCGPAMTERMINAL KILL

074

TOXICOLOGY OBSERVATION REFRT

I8

TABLE H-13 (CONTINUED)CLINICAL SIGNSPRqJECT NUMBER - 1C73415

WEEK AhIMAL OBSERVATIONS : OUALIFIEDNO. ** DOSE GROUP/SEX * NLPBER **w.. COM4ENTS

013 a PP" MALE FASTED : OVERNIGHT08618 TERMINAL KILL

FASTED : OVERNIGHT08619 TERMINAL KILL

FASTED : IVERNrGHT01620 TERMINAL KILL


FASTED : OVERNIGHTC8622 TrRMIXAL KILL



FASTED : OVERNIGH T

OQ62' TERMINAL KILLFASTED : OVERNIGHT

08626 TERMINAL KILLFASTED : OVERNIGHT

C8627 TERMINAL KILLFASTED : OVERN'IGHT

G8628 TERMINAL KILLFASTED : OVERNIGHT


FEMALF 08695 FASTED : OVERNIGHT

BLOOD SUBMITTED FOP CHEM AND HFMCGRAMC86S6 FASTED : OVERNIGHT

BLOOD SUBMITTED FO' CHEM AND HErCGVAM08657 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR CHEM AND HEMOGRAM08698 FASTED : OVERNIGHT

BLOOC SUBMITTEO FOR CHEM AND HEMCGFAM08699 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR CHEM AND HFMCGOAM08700 FASTED : OVERNIGHT

BLOOC SUBMITTEO FOR CHEM AND HFMOGPAM08701 FASTED : OVERNIGHT

BLCOD SUBMITTEO FOP CPEM AND HEMCGOAM08702 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR CHEM AID HEMCGPAMTERMINAL KILLFASTED : OVERNIGHT

28703 TERMINAL KILL

075

TZEICCLOGY OBERVATICN REFCRT

19


CLINICAL SIGNSPROJECT NQPBER - 1073415

WEEK AhIMAL OBSERVATIONS OUALIFIEFNO. ** DOSE GROUP/SEX * NLBER ....- COMMENTS

013 1 PP" FEMALE FASTED : OVERNIGHT08704 TERMINAL KILL


FASTED : OVERNIGHT0870i TERMINAL KILL




FASTED : OVERNIGHT






300 PPM MALE 08620 FASTED : OVERNIGHTBLOOD SUBMITTED FOR CHEM AND HEMOGRAM

08631 FASTED : OVERNIGHTBLOOD SUBMITTED FOR CHEM AND HEMCGqAM

086!2 FASTED : OVERNIqHTBLCOD SUBMITTED POO CIHE4 AND HEMCIPAM

08633 FASTED : OVERNIGHTBLOOD SUBMITTED P OO CHEM AND HEMCGPAM

08634 FASTED : OVERNIGHTBLOOD SUBMITTED FOR CHEM AND HEMOGPAM

08635 FASTED : OVERNIGHTBLOOC SUBMITTED FOR CHEM AND HEPCGPAP

08636 FASTED : OVERNIGHTBLOOD SUBMITTED F09 CHEM AND HFMOGPAMTERMINAL KILLFASTED : OVERNIGHT

08637 FASTED : OVERNIGHTBLOOD SUBMITTED FOR CHEM AND HFMOGOAMTtRMINAL KILLFASTED : !VERNIGHT

08631 TERWINAL KILL

07(i

TOX[CILOGY CBSERVATICN REFRT

TABLE H-13 (CONTINUED)CLINICAL SIGNSPROJECT NUMMER - 1G73415

WEEK ANIMAL OBSERVATIONS QUALIFIEPNC. *D DOSE GROUP/SEX * NLMBER *.... COMMENTS .*..*.

013 300 PPM MALE FASTED : OVERNIGHT08639 TERMINAL KILL

FASTED : OVEPNIGHT08640 TERMINAL KILL

FASTED : OVERNIGHTC8641 TERMINAL KILL

FASTED : OVEPN-IGHT



0644 TERMINAL KILL





FASTED : 0VrEQNIGHT08649 TERMINAL KILL

FASTED : OVERNIGHTFEMALE 08715 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR CHEM AND HEMOGPAP08716 FASTED : OVERNIGHT

BLOOD SUBMITTED FOR CHEM ANC HEMOGFAM08717 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR CHEM AND HEMCGPA!A0871 FASTED : OVERNIGHT

BLOCC SUBMITTFO FOq CHEu AND HEMCGRAP08719 FASTED : OVERNIGHT

BLOOC SUBMITTED FOP CHEM AND HEMCGPA!08720 FASTED : OVERNIGHT

BLOOC SUBMITTED FCP CHEM A,*! Hr"RCPA"08721 FASTED : OVERNIGHT

9LOOC SURMITTED FO9 CHEM AND HE'CGQAv08722 FASTED : OVERNIGHT

LOCC SURWITTED FOU CHEM A' D HFMCGCANTERMINAL KILLFASTED : OVERNIGHT

0q21 TERMINAL KILLFASTr : OVERN:IGHT

G0724 TERMINAL KILL

077

TCECCLCGY CBSERVAIICN qEPCRT


CLINICAL SIGNSPRIJECT NLMBEQ - IC73415

WEE ANIMAL OBSERVATIONS : QUALIFIER

NO. *. DOSE GROUP/SEX * NUMBER **" COMMENTS

013 !30 PPM FEMALE FASTED : OVERNIGHT08725 TERMINAL KILL

FASTED : OVERNIGHT0RT26 TERMINAL KILL

FASTED : OVERNIGHT



09729 TERMINAL KILLFASTED : OVERNIGH

T

087!0 TERMINAL KILLFASTED : OVERNIGHT


097!2 TERMINAL KILLFASTED : OVEV~eIGH

T

Z1733 TERMINAL K:LLFASTED : OVERN:GHT

C9714 TERMINAL KILLFASTED : OVERNIGHT

1000 PPM MALE 08650 FISTED t OVEr'tIGHTRLOOO SUBMITTED FOR CPE- Ai1 HE ClQAM

t86!1 FASTED : OVERNIGHT

PLCOD SU8MITTED FOO CVEP AND HEMCGRAM

786!2 FASTED : OVERNIGHT

BLOGC SUB&$TTF[ FCP CAEM AND HMMCTQRA

a86! FASTED : CVERNIGHT

BLOOC SUBMITTED FOR CHEM AND HEMCGqv&


BLOOD SUB"ITTED FGC CPEM AD HTCqPAv


BLOOD SUB9ITTED FOR CHEM AND HEMCGPAM

0965S FASTED : OVERNIGHTBLOCC SUB"ITT7eD Fq CVEM 4 1 HSMCGv

C9657 FASTED : OVERNIGHT

BLCCC SUBMITTEI FOR CHEM A%'O HEMCGPAm

TERMINAL KILL

FASTED : OVERNIGHT

38658 TERMINAL K!LLFASTSC t *VERNIGHT

G6!9 TERMINAL KILL

FASTED : OVEPNIGHT

01660 TERMINAL KILL

078

TnXICOLOGY OBSERVATION REFCRT

22


CLINICAL SIGNSPRCJECT NLBER - lC73'15

WEEK ANIMAL OBSERVAT!ONS QUALIFIERNO. e DOSE GROUP/SEX * NLMBER * COMMENTS

013 1000 PPM MALE FASTED : OVERNIGHT


08662 TERMINAL KILL




FASTED : OVERNIGHT

08666 TERMINAL K:LLFASTED : OVERNIGHT

08667 TERMINAL KILLFASTED : OVERNIGHr

C8668 TERPINAL KILLFASTED : OVERNIGHT


FEMALE C2735 FASTED : OVERNIGHTELOCC SU8MITTT0 FOR CHE" A: HrMCCPAW

C8736 FASTED : OVERNIGHTBLOOD SUBMITTED FOR CHEM AND HEPCGPAM

08737 FASTED : OVERNIGHT8L(OC SUR"ITTED FCR CHE" AND HEMCGPAP

08718 FASTED : OVERNIGHTBLOOD SUBMITTED FOP CHEM AND HEMOGRAM

38739 FASTED : OVERNIGHTBLOCC SUBMITTED FOR CHEM AND HEMCGRAM

38740 FASTED : OVERNIGHTBLOOD SUBMITTED FOR CHEM AND HEMCGPAM

01741 FASTED : OVERNIGHTOLOOC SUBMITrEO FOR CHEM AND HEMCGRA"TERMINAL KILLFASTED : OVERNIGHT

08742 FASTED : OVERNIGHTBLCGC SUBMITTED FOR CHEM AND HEM CGPAMTERMINAL KILLFASTED : OVERNIGHT


Oe44 TERMINAL KILLFASTED : OVERNIGHT

*9745 TERMINAL K:LL

~ 079

TOXICCLOCi OBSZRVATICN REFCRT

23

TABLE H-13 (CONTINUED)CLINICAL SIGNSPROJECT NUMBER - 1073415

WEEK ANIMAL OBSERVATIONS QUALIFIERNO. ** DOSE GROUPfSEX * NLMBER .-- *' COMMENTS

013 13Oa PPM FEMALE FASTED : OVERNIGHTC8746 TERMINAL KILL

FASTED : OVERNIGHT


08748 TERMINAL KILLFASTED : OVSRNIGHT

1874q TERMINAL KILLFASTED : OVERNIGHT


08751 TERMINAL KILLFASTED : 0VFR VGHT

C27!2 TERMINAL KILLFASTED : OVEQNIGHT

03753 TERMINAL K:LL

FASTED : OVERNIGHTCR,754 TERMINAL KILL

FASTED : OVERNIGHTiGOo PPM MAL- 08670 FASTED : *VERNIGHT

ELCOC SUBMITTED FCq CHEM AND HrWCGQAM08671 FASTED : OVERNIGHT

BLOOC SUBMITTED FOR CHEW AMND HEMCGRAM08672 FASTED : OVERNIGHT

BLOOC SUOMITTEI FIO CHEM AND HEMCGOAM0P673 FASTED : OVERNIGHT

BLOOE SUBMITTnD FOR CHEM AND HEMOGaAM08674 FASTED : OVERNIGHT

RLOOC SUBMITTED FOR CHEM AND HEPCGqA"08675 FASTED : OVERNIGHT

9LOOC SUBMITTED FIR CHEV A'1D HEMCGPAP08676 FASTED : OVERNIGH T

BLCOC SUBMITTED FOR CHEW AND HE"CGRA"08677 FASTED : OVERNIGHT

9LCOC SUBMITTED FOR CHEW AND HEMPCGAPTERMINAL KILLFASTED : OVERNIGHT

08579 TERiNAL KILLFASTED : OVERNIGHT


38680 TERMINAL KILLcASTEC : IVERNIGH

19681 TE0 MINAL KILL

Obo

TCXICCLOG OBSETRATICN REFCRI

4

TABLE H-13 (CONTINUED)CLINICAL SIGNSPRIJECT NLiPSER - 1073415

UEeK ANIMAL OBSERVATIONS QUALIFIEDNC. * 3CZE GROUP/SEX * JLUBER * COMMENTS

013 3000 PPM MALE FASTED : OVERNIGHT08682 TERM!NAL KILL

FASTEC : OVERNIGHT08683 TERMINAL KILL


FASTEC : OVERNIGHTC8685 TERMINAL KILL

FASTED : OVERNIGHT

08686 TERMINAL KILLFASTED : OVERNIGH T




FEMALE 09557 FASTED : nVERNIGHT

BLOOC SURMITTED FOR CHEM AND HrMCGPAMC87 6 FASTED : OVERNIGHT

BLOOC SURMTTTED FOR CHEM AND HEMCGOAM

08757 FASTED : OVERNIGHTBLOOC SUBMITTED FOR CHEM AND HEMOGPAM

093 FASTED : OVERNIGHTBLOOC SUBMITTED FOR CHEM AND HFMOGRAM

08759 FISTED : OVERNIGHT

BLOOD SUBMITTED FOP CHEM AND HEMC'PAM08760 FASTED : OVERNIGHT

BLOOD SUBMITTED FOR CHEM AND HEMCGPAM

08761 FASTEO : OVERNIGHTBLOOC SUBMITTED FOR CHEM AND HEPGAV

08762 FASTED : OVERNIGHTBLOOD SURITTEO FOR CHEM AND HEMCGOAM

TERMINAL KILL


FASTFC : OVERNIGHT

OA764 TERMINAL KILLFASTED : OVERNIGHT

OP65 TERMINAL KILL

FASTED : OVERNIGHT

0876; TERMINAL KILLFASTEC : OVrRNIGHT

03767 TERPINAL KILL

of),3

TOXICOLOGY 0BSEIVATION REFPCT

'5

TABLE H-13 (CONTINUED)CL:NrCA.L SIGNSPRIJECT NLP ER - IC73415

W K ANIMAL OBSERVATIONS : GUALIFIE O

NO. ** OOSE GROUP/SZX a NLMRER **** CO"ENTS

013 0OO0 PPM FEMALE FASTED : OVERNIGHT08768 TERMINAL KILL




VASTEC : OVERNIGHT0@772 TERMINAL KILL

FASTEO : OVSRNIGH T

C8773 TERMINAL KILLFASTEC : OVERNIGHT

C87T TERmINAL KILLFASTED OVERNIGHT

014 0 PPM wALE C8610 TERMINAL KILLFASTED : OVERNIGHT

C8611 TERMINAL KILLFASTEC : OVERNIGHT

08612 TERMINAL KILL

FASTED : OVERN:GHTC8613 TERMINAL KILL

FASTED : OVERNIGHTQ8614 TERMINAL KILL

FASTEC : OVERNIGHT0e615 TERMINAL KILL


FASTED : OVERNIGHTFEMALE 08695 TERMINAL KLL


FASTED : OVERNIGHY08697 TERMINAL KILL

FASTED : OVERNIGHT08698 TrRMINAL KILL

FASTEO : !VERNIGHT08699 TERMINAL KILL

FASTEO : OVERNIGHT08TI TERMINAL KILL

FASTEC : IVSRNIGHTC9711 TERMINAL KILL

FASTEO : OVeRNIGHT

*019oz

TCXICOLClY CBSERVATICN REFCRT

TABLE H-13 (CONTINUED)CLINICAL SIGNSPROJECT NUPBER - 1.73415

WEEK ANIMAL OBSERVATIONS GUALIFIERNO. * DOSE GRCUP/SEX * NLPOER " COMMENTS

014 300 PPM MALE 08630 TERMINAL KILLFASTED : OVERNIGHT

0R611 TERMINAL KILL

FASTED : OVERNIGHT


09633 TERMINAL KILL


FASTED : OVERNIGHT

0a635 TERMINAL KILLFASTED : OVERNIGHT

FEMALE 08715 TrRMINAL KILLFASTED : OVERNIGHT

08716 TERMINAL KILL

FASTED : OVERNIGHT


08718 TERRMINAL KILLFASTED : OVERNIGHT

08711 TERMINAL KILL

FASTEO : OVERNIGHT08720 TERMINAL KILL

FASTED : OVERNIGHT

08721 TERMINAL KILLFASTED : OVERNIGH

1000 PPM MALE 00650 TERMINAL KILLFASTED : IVERNIGHT

096!1 TERMiNAL KILLFASTED : OVERNIGHT

086!2 TERMINAL KILL

FASTED : OVERNIGHT

08653 TERMINAL KILL

FASTED : OVERNIGHT

086!4 TTRMINAL KILLFASTED : OVERNIGHT


0B6! TERMINAL KILL

FASTED : OVERNIGHT

FEMALE 08735 TERMINAL KILLFAST7 : OVERNIGHT

C9 7 TERMINAL KILL

FASTED CVER4IGHT

C13

T3XI:LOGY O8SrPVAT!N REPCRT

7


CLINICAL SIGNSPRCJECT NLPSEQ - 1C71415

WEEK AhIMAL OBSERVATIONS QUAL!FIEr

NO. ** DOSE GROUP/SEX * NLOSER * COMMENTS .*****

114 L300 PPp FEMALE 087!7 TERMINAL KILLFASTED : OVERNIGHT


087!9 TERMINAL KILL


FASTED : IVERNIGHT

3000 PPM MALE 08671 TERMINAL KILLFASTED : OVERNIGHT

c8671 TERMINAL KILLFISTEC : IVERNIGHT

08672 TERMINAL KILLFASTED : OVEPNIGHT

C8673 TERMINAL K:LLCASTEO : IVERNIGHT

08674 TERMINAL KILLFASTED : OVERMIGHT



FEMALE 087!5 TERMINAL KILLFASTED : OVERNIGHT

09756 TERMINAL KILLFASTED : OVERAIGHT

08757 TERMINAL KILLFASTEC : OVERNIGHT


087!9 TERMINAL K:LL

FASTEO : OVERNIGHT

38760 TERMINAL KILL

FASTED : OVERNIGHT


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TABLE H-I7

CLINICAL HEMATOLOGY

KEY

WBC = Leukocyte Count 1 3 n

RBC = ~Erythrocyte Count 16/

HGB = Hemoglobin Vl%

HCT = Hematocrit%RET = Reticulocyte %.BN Band Neutrophils %.SG = Segmented NeutrophilS %.LY = Lymphocytes %.MO = MonocyteS %.

EO Eosinophils %.BS = Basophils %.IM = Immature%OT = Other

* - p<0.05 as compared to controls: Dunnett's t-test.

I = Quantity not sufficient

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LITTOIN BIONETICS, INC.PRUJECT NO. 10734-15

TABLE H-18

CLINICAL CHEM1ISTRY

KEY g d

BNBlood Urea Nitrogen mg /d 1CAN Calcium mg/dl

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SGPT s Serum Glutamic-pyruvic Transaminase mU/mi

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SECTION H

APPENDIX B

, 143

SPONSOR: U.S. Army R&D

MATERIAL: Isopropyl Methylphosphonic acid, sodium salt

SUBJECT: FINAL PATHOLOGY REPORT3-Month Oral Toxicity Study in RatsLBI Project No. 10734-15

METHODOLOGY

Pathological examinations were conducted on 20 male and 20 female CRL:COBSCS(SD)BR rats which were exposed (or not exposed) to IMPA in the fLed forapproximately 13 weeks at each of the following dose levels: Group 1, 0 ppm(control); Group 2, 300 ppm; Group 3, 1000 ppm; and Group 4, 3000 ppm. Allanimals survived and at the end of the exposure period, each animal waskilled by C02 intoxication and subjected to a detailed necropsy examination.The most recent clinical observations for each animal were reviewed atnecropsy, and all observations of gross abnormalities were recorded directlyonto the necropsy sheet as encountered. Liver, heart, spleen, kidneys, ad-renals, gonads, and brain were weighed from all animals immediately aftersacrifice. Following gross examination appropriate samples of the follow-ing tissues were fixed in 10% neutral buffered formalin: Any unusual lesions,brain (cerebrum, cerebellum, brainstem), spinal cord (thoracic, cervical),eye (left), pituitary, salivary gland (submandibular), heart, thymus, thyroid,lungs (with mainstem bronchi), trachea, spleen, bone (with marrow) sternum,mesenteric lymph node, esophagus, stomach, small intestine (duodenum, ileum,jejunum), large intestine (colon, cecum), adrenal glands, pancreas, liver(2 sections), kidneys, ovaries/testes, prostate, uteri (corpus, cervix),skin (mammary area), skeletal muscle (rectus femoris), sciatic nerve, andurinary bladder. These tissues from all Group 1 and 4 animals were embeddedin paraffin, sectioned at five to seven microns, stained with hematoxylinand examined microscopically.1

GROSS FINDINGS

There were few gross lesions observed. Those that were present were notunusual for rats of this age and strain and had no particular relevanceto the study. In general, the rats appeared to be quite free from disease.

HISTOPATHOLOGY

The results of microscopic examination of the tissues are presented in TheTables of Microscopic Findings. Ocassional tissues were lost in processingand not available for microscopic examination; thus, the number of any onetissue examined does not necessarily correspond with the number of animalsnecropsied. No neoplasms were observed in the examined animals. A varietyof inflammatory and degenerative changes occurred randomly in both treatedand/or control rats. Most common of these lesions was lymphocytic inflamma-tory infiltration around pulmonary vessels. The occurred usually as mild or

144.

moderate cuffing, the cause of which was not determined but probably is of viralorigin. A second lesion of unknown etiology but commonly observed in rats,particularty-nales, was myocardial degeneration or myocardiopathy. Finally,renal mineralization was noted, usually as a focal change in the medulla orpelvis of kidneys, while hydronephrosis was present in some of these animalsmost likely the cause of pelvic urolithiasis.

CONCLUSIONS

No changes or lesions associated with treatment were noted in examined animals.Lesions which were present occurred randomly in both treated and control ani-mals and are of the kinds commonly seen in rats of this age and strain.

SUBMIT BY:

'Richard H. Mrdy, D.V.M. / DateDirector, Department of Pathology

Necropsies were conducted under the direct supervision of this pathologist.Initial histopathologic examinations were done by Dr. Alexander DePaoli.Histologic review of all slides from 25% of the animals (selected randomly)and review of selected lesions was conducted by me. My interpretation ofhistologic material from these animals was not different from Dr. Depaoli's.This report was prepared by me from data obtained during the gross andhistooathologic examinations.

145


LESEND

Animal disposition codes

E = escapedF = found deadI = interim killK = killed accidentallyM = moribund killT = terminal kill

Tissue status codes

+ = tissue examined, lesions foundX = tissue examined, ro lesion foundI = insufficient tissue present for evaluationA = autolysis too severe for microscopic evaluationC = cannibalizedS = section and/or tissue not presentP = paired organs, one missingN = tissue not examinedE = excluded by protocol

Lesion grades (non-tumor Datho!ogy, Tumor counts (tumor patnologv* = not graded

I = minimal 1 = one occurrence2 = mild 2 = two occurrences

3 = moderate 3 = three occurrences4 = marked 4 = four occurrences5 = severe 5 = five occurrences

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LITTON BIONETICS, INC.PROJECT NO. 10734-15 INDIVI3UAL ANIMAL EVALUATION

ANIMAL NOa 9610 SEX: MALE PATHOLIGY REFERENCE N". 81-0405_1CROUP 1 - UNTREATED CONTROL SPECIES: RATSCOSE AM'WUNT: 3 PPM STRAIN: COBS CO(SO) BRUE-"KS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLIERMINAL WEIGHT: 494.O3O GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:

ANIMAL APPEARED NORMAL GRGSSLY

MICROSCOPIC FINDINGS:

STOMACHINFLAMMATION9 NOS

(MILD )SPLEEN

HEMATOPOIESIS (EXTRAMEOULLARY)(MILD )

LUNGSBLOOD VESSEL

LYMPHOCYTIC INFLAMMATORY INFILTRAT_

LITTON eIONETICS, INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8611 SEX: MALE PATHOLMGY REFERENCE NO. 81-O402GROUP I - UNTREATEC CONTROL sPFCrES: RATSCOSE AMCUNT: 0 PPM STRAIN: C')RS CO(Sl)) PQ6EEKS ON STUDY: 13 S'URCE: CHARLES RIVER (CPCTAGE)CATE OF DEATH: 01-23-81 TYPE IF O-ATH: TERMINAL KILLTERMINAL WEIGHT: 454.000 GRAMS PETHVJ OF KILL: CARBCN oCXI."

GROSS ORSERVATIONS:

ANIMAL APPEARED NORMAL GROSSLY

PICROSCOPIC FINDINGS:

STOMACHINFLAMMATION. NOS

(MINIMAL)LUNGS

BLOOD VESSELLYMPHOYTIC INFLAMMATOPY INFILTRATE

LITTON BIONETICS, INC.FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8612 SPX: MALE PATHOLOGY REFEREN'E NO. 81-04C53EROUP 1 - UNTREATEO CONTROL SPECIES: RATSDOSE AMOUNT: a PPM STRAIN: COBS CD(SD) RRbEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL RILLTERMINAL WEIGHT: 481.000 GRAMS METHOD OF KILL: CARBON DIOXIOI

GROSS OBSERVATIONS:LEFT EYE

RED CRUST AROUND LEFT EYE



(MINIMAL)SPLEEN

HEMATOPOIESIS (EXTRA4EOULLARY)(MILD )

LEFT EYETISSUE EXAMINED, NO LESIONS FOUND

HTTON PIONETICS3 INC.OJECT NO* I734-15 INOIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 861' SEX: MALE PATHOLOGY REFERENCE NO. 81-04G;4GRJUP I - UNTREATED CONTROL SPECIES: RATSCOSE AMOUNT: 0 PPM STRAIN: CORS CD(SO) BR6EEKS ON STUOY: 1i SOURCE: CHARLES RIVER (PCOTAGE)CATE OF DEATH: 01-23-81 TYPE OF DFATH: TERNiNAL KILLTERMINAL WEIGHT: 464.003 GRAMS METHOD OF KILL: CARBON 0ICXI'E

GROSS OBSERVATIONS:

ANIMAL APPEARED NCRMAL GROSSLY

PICROSCCPIC FINDINGS:

STOMACHINFLAMMATION, NOS

(MINIMAL)PROSTATE

INFLAMMATIlNo NOS(MODERATE)

jmI

LITTON BIONETICS, INC.PROJECT NO* 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8614 SEX: MALE PATHOLOGY REFE-RENCE NO* 81-01#C55EROUP 1 - UNTREATED CONTROL SP--CIES: RATSCOSS ANCLUNT: a PPM STiAIN: COBS CO(S0) 44hEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCPTAGE)CAT7 OF DEATH: 01-23-81 TYPE 5F DEATH: TERVINAL KILLTERMINAL WEIGHT: 501.000 GRAMS METHOD OF KILL: CAPBCJ DICXrOE

GROSS OBSERVATIONS:


PICROSCGPIC FINDINGS:

PANCREASINFLAMMATION, FOCAL

STOMACH(MLINFLAMMATION, NOS

(MILD

LITTON SIONETICS9 INC.FROJECT NO. 10734-l! INoIVIDUAL AN!MAL EVALUATION

ANI"4L NO. 8615 SEX: MALE PATHIL"GY REFERENCE NO. 81-046GROUP 1 - UNTREATED CONT ROL S~rECItS: RATSCOSr AMCUNT: a PPMl s?*4!4.: CCeS CocSO) op6ZErk3 ON STUDY: 13 SCURC7: CHARLES RIVER (ORTAGS)CATr- OF DEATH: 01-23-81 TYPE 'F* DEATH: TERMIP4AL KILLTERRINAL WEIGHT: 374.030 GRAMS PrETHOD OF-KILL: CARBON DIOXInE

GROSS OBSERVATIONS:

SKINSCAB ON SKIN BELOW RIGHT EAR (1.0 CM IN DIAMETER), RIGHT rAPTAG

INFECTIONLUNGS

PINPOINT WHITE FOCI ON ALL LOBES


SPLEENHEMATOPOIESIS (EXTRAMEOULLARVI

(MILDSKIN

INFLAMMATION, NOS(MODERATE)

LUNGSBLOOD VESSEL

LY'4PHOCYTIC INFLA"PATr)RY INFILTRATE

LIrTTON sIQNETICSIgcFROJECT NO.-10754-15; INDIVIDUAL ANIMAL EVALUATIONANIMAL N't. 8616 SEX: MALE PATHj)LOGY REFERENCE N('. 81-04ts?GROUP I -' UNTREATED CONTROL SPECIES: RATSCOSE AMOUNT: a PPM STRAIN: COSS CODSO RRUEEXS ON STUDY: 13 SOURCE: CHARLES RIVER (PCTrvE)LATE OF DEArH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 437.oOO GRAMS PZTHO0 OF KILL: CAR80N DIOXIDE

GROSS OBSERVATIONS:


FICROSCCPIC FINDINGS:

HEAR TDEGENERATION,9 NOS

STOMACH(MLINFLAMMATION, NOS

(MILDLUNGS

HISTIOCYTCSIS(MILD

COMMENTS:

CRUST AROUND LEFT EYE NOT APPARENT AT NECROPSY

LITTON BIGNETICS,INC.PROJECT -10a IQ? -is TNDXVIO3UAL AN!"AL EVALUATION

ANIMAL NO* 8617 SEX: MALE PATHOLOGY PEF EEE N". 81-960'82EROUP 1 - UNTREATED CONTROL SP7CIrS: RATSCOSE AMCUNT: 0 PPM STRAIN: CCBS CtO(SO) ROIsEES ON STUOY: 13 SO)URCE: CHARLES RIV!P (PORTAG-1CATE OF OEATH1 01-22-81 TYPE 09 DEATH: T;"R"'NAL KILLTERMINAL WEIHT: 46000 O GRAMS "STMOD OF KILL: CARRO4 DICXIDC

GROSS OBSERVATIONS:

ANIMAL APPEARED NORM4AL GROSSLY

PICROSCoprC FINDINGS:


(MILOLUNGS

BLOOD VESSELLY4PHOCYTIC INFLAMMATORY INFILTRATE

IISSUE(S) MISSING OR SEVERELY AUTOLYZED:

TH Y MUSSECTION AND/OR TISSUE NOT PRESENT

COMMENTS:

RIGHT EARTAG REPLACED (ALSO 38617)

jbo

LITTON BIONETICS9 INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL No. 8618 SEX: MALE PATHOLOGY REFERENCE NO. 81-04C59GROUP I - UNTREATED CONTROL SPECIES: RATSrOSE AMOUNT: 0 PPt4 STRAIN: COBS CO(SD) RD6EEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAr:-)DATr OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 502.030 GRAMS M9ETHOO OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:


ICROSCOPIC FINDINGS:

STOPACHINFLAMMATION, NOS

(MILD )LUNGS

BLOOD VESSrLMINERALIZATION

(MINIMAL)BLOOD VESSELLYMPHOCYTIC INFLAMMATORY INFILTRAT _

LITTON BIONETICS I NC,ROJECT NO. 107 q-11 INDIVIDUAL AN14AL EVALUATION

ANIMAL NO* 8619 SEX: MALE PATHOLIGY REFERENCE NO* 81-04CSOGROUP 1 - UNTREATED CONTROL SPI.CIES: RATSCOSE AMOUNT: 0 PPM STRAIN: CIBS CO(SO) PRWEEKS ON STUDY: 13 SIURCE: CHARLES RTVER (PCRTAGE)CATE OF OCAT,4* 01-22-81 TYPE OF DrATH: TERMINAL KILLTERMINAL WEIGAT: 512.000 GRAMS MTHO0 OF KILL: CARBON IOwI3E

GROSS OBSERVATIONS:



STO10ACHINFLAMMATION, NOS

(ILO ISPL .N

HENATOPOIrSIS (EXTRAMECULLARY)(MILD I

LUNGSBLOOD VESSEL

LY4PHOCYTIC INFLAMPATORY INFILTRATEKIDNEYS

HYORONEPHROS IS(MILD I

1.61

LITTON BIONE TICS, 1.PROJECT NO. 107 !r-19 INDtV!UAL ANIMAL EVALUATION

ANIPAL NO. 8620 SEX: MALE PATHOLOGY PEFERENCE NO* 81-04061GROUP 1 - UNTREATED CONTROL S~cCIES: RATSCOSE AMOUNT: a PPOM SRA:N: C09S CD(SO) 90UEEKS ON~ STUOY: 13 SOUR-E: CHARLES RIVER (PCPTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERmI'JAL KILLTERMINAL WEIGHT: 466,000 SPAMqS METHOO OF KILL: CAPBON. DIOXIDE

GROSS OBSERVATIONS:

ANIMAL APPEARED hORMAL GOOSSLY

MI1CROSCOPIC FINDINGS:


(MILDLUNGS

BLOOD VESSELLYMPHOCYTic INFLAMMATORY INFILTRATE

KID0N EY SHYORONEPHRCSI S

(MODERATE)

LITTON PIONETICS ICPROJECT O.1T-1! iheivIDuIL ANIMAL EVALUATION

ANIMAL NO* 8621 SEX: MALE PATHOLOGY AEFrREJCE NO. 81-04C62GROUP 1 - UNTREATED CONTROL SPECIES: RATSCOSr AMOUNT: 0 PIPM STQArN: cops COsn0 99hCE~iS ON STUOY: 1! SOURCr: CHARLES RIVER (PCRTAGE)CAf-- OF DEATH: 01-22-81 TYPE 6F DEATH: TFAMINAL KILLTERMINAL WEIGHT: 495,00 GOA"' MZTH00 OF KILL: CARBIN Orl'XIT~E

GROSS OBSERVATIONS:



PANCREASINFLAMMATION, FOCAL(MINIMAL)

STOMACHINFLAMMAT13N9 NOS

(MILDLUNGS

BLOOD VESSELLYMPMOCYTIC INFLA4PATORY INFILTRATE

K10D4EYSINFLAM'4ATION9 FOCAL

(MINIMAL

LITTON RION.TICSt INC.FROJEC NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO* 8622 SEX: MALE PATHOLOGY REFERENCE NO. 81-04'63GROUP 1 - UNTREATEO CONTROL SPEC S: RATSCOSE AMCUNT: 0 PPM STRAIN: COBS Co(So) RR6EEKS ON STUOY: 13 SOURCE: CHARLES RIVER (PCRTAGE)CAT- OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERINAL WEIGHT: 478.O00 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:


FICROSCOPIC FINDINGS:

PAN CREASINFLAMMATICN9 FOCAL(MILO I


(MILO I

LITT-', FtO',4.7IC;, 1' :.

AJ~A~ :.i621 S7X: NLL P 'j"4%y F:'..-4-4~jp 1 - -' T C ~ Lr~T3

W-KS 9'. .TJY: 13 ':LS - (PCO '.)' - OF DE--' : ._.,, ] Y -" 3 -:' . "";'f"%lL K(:Li.TE'RMINAL W-I1,H.-: 44. 3,.; '04w; -T tr KILL.: CARR'%, I' ; " r

GR3SS O SEQ'J"T7'"I$:

AI TMAL APPE:llrD %!Qw-L G~l';SLl"I:ROSCCPIC Fjl"N;Gc:

STI).CH

(LIL )M: ST!£-,:YT.lS![3

LY ,.'.YT;C I rL* 4MATDDY t'ILTR '--TTST;'r

#As Z rTF{ .AL

(m- ,r'aL )

113

RLITTON BONETICS, INC.

TROJCT NO,* 1034-15 INOIVIDUAL ANEMAL EVALUATION

AINAL NO. 8624 SEX: MALE PATHOLOGY PEFERENC- N1. 81-04965GROUP 1 - UNTREATED CONTROL SPECIES: RATSDOSE AMOUNT: O PPM STRAIN: COBS CD(SO) BpWEEKS ON STUOY: 1 SOURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-21-81 TYr OF DEATH: TrRMI'NAL KILL7ERMINAL WEIGHT: 504.o3Q GRAMS METHOD OF KILL: CARBON DlnXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSY

RIGHT EARTAG INFECTION


STOMACHINFLAMATION, NOS

(MINIMAL )

TISSUE(S) MISSING OR SEVERELY AUTOLYZEO:

THYMUSSECTION AND/CR TISSUE NOT PRESENT

LITTON BIONETICS. INC.PROJECT NO. 10734-1S INDIVIDUAL AnIMAL EVALUATION

ANIMAL NO. 8625 SEX: MALE PATHOLOGY REFERENCE NO. 81-04066CROUP 1 - UNTREATED CONTROL SOrCIES: PATSCOSE AMCUNT: 0 PPM STRAIN COBS CP(SO) BRhoEEKS ON STUDY: 13 SOURCE: CHARLES R(VER (POPTAGECATE OF DEATH: 01-21-81 TYPE OF DEATH: T-RMINAL KILLIERMINAL WEIGHT: 35,,000 GRAMS METHOO OF KILL: CARBON OIcXI-

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYTEETH CUTCRUST AND SWELLING AT EARTAG SITE

LUNGSDAMAGErO (NOTED AT TRIMMING)


HEARTDEGENERATION, NOS

(MODERATE)LUNGS

TISSUE EXAMINED, NO LESIONS FOUND

LITTON RIONETICSv INC.FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUAT13N

ANIMAL NO* 8626 SEX: MALE PATHOLOGY REFERENCE NO. 81-04067GROUP I - UNTREATED CONTROL SPCIES: PATSCOSE AMOUNT: a PPM STRAIN: COSS CO(SD) 9RUEEKS ON STUDY: 13 SCURCE: CHARLES RIVER (PCPTAGr)CATE OF OEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL UEIGHT: 4!S.000 GRAMS METHOD OF KILL: CARBON OICX10D-

GROSS OBSERVATIONS:

KIDNEYSLEFT KIONEY DILATED,9 CONTAINS GREEN FLUID IN PELVIC AREA



(MINIMALLUNGS

BLOOD VESSELLYMPHOCYTIC INFLAMMATORY INFILTRATE

KIDNEYSHYORONEPHROSIS

INTERSTITIAL TISSUE OF KIDNEYMULTIFOCAL

INFLAMMATiaN, INTERSTITIAL(MILDO

LITTON 8IONET ICSICFROJECT NO. 10734-1I5. iPhIvIDUAL ANIMAL EVALUATION

ANIMAL NO. 8627 SEX: MALE PATHOLOGY REFrRENCE NO. PI-040'-8GROUP I - U4TREATED CONTROL SPECIES: RATSCOSE AMOUNT: a PPM STRAIN: C17S CO(SO) PRU7EKS ON STUDY: 13 SIURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-21-81 TYPE OF DEATH:- TER"INAL KILLTERMINAL WEIGHT: 539000 GRAMS METHOD OF KILL: CARB':N CICXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCr AT NrCROPSYEARTAG INFECTION



(MILDSPLEEN

HE4ATOPOIESIS (EXTRAMEDULLARY)(MILO

LUNGSBLOOD VESSEL

LYMPHOCYTIC INFLAMMATORY INFILTRA-rc

TISSUE(S) MISSING OR SEVERELY AUTILYZED:

MAMMARY GLANOSECTION ANDIGR 'ISSUE NOT PRESENT

LITTON eIONETICS9 IC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8628 SEX: MALE PATHOLOGY REFERENCE N% 81-04C69GROUP 1 - UNTREATEO CONTROL SPECIES: RATSDOSE AMOUNT: 0 PPM STRAIN: COBS CO(SO BRWEEKS ON STUDY: 13 SOURCE: CHARLES RIVEP (PCRTA'E)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERM104AL KILLTERMINAL WEIGHT: 479.000 GRAMS PETHOD OF KILL: CARBON Drx1IOE

GROSS OBSERVATIONS:

ANIMAL APPEARED %ORMAL GROSSLY


PANCREASPANCREATIC ACINUS

ATROPHY9 NCS(MILD I

CECU4INFLAMMATION* NOS

(MILO ILUN3S

HISTIOCYTOSIS(MIN:MAL I

BLOOD VESSELMINERALIZATION

(MINIMAL)KIOUCYS

KIONtYfCORTEXCYST, NOS

TISSUE(S) MISSING OR SEVERELY AUTOLYZED:

LEFT EYESECTION ANDICR TISSUE NOT PRESENT

LITTON BIONETICS* INC.FPROJECT NO. 13734-15 INDIVIOUAL ANIMAL EVALUATION

#NImAL NO* 8629 SEX: MALE PATHOLOGY REFERENCE NO. 81-040706ROUP 1 - UNTREATEO CONTROL SPECIES: RATSDOSE AMOUNT: 0 PPM STRAIN: CLPS C0(SV) RRbEEKS ON STUDY: 13 SOURCE: CHARL'S RIVER (PORTAGE)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 424.000 GRAMS METHOD OF KILL: CARBCN DICXIDE

GROSS OBSERVATIONS:

ANIMAL APPEARED NORMAL GRs LY

PzCROScOPIC FINDINGS:

STOMACHINFLAMMATION* NOS

(MINIMAL)LUNGSBL00 VESSEL

LYMPHOCYTIC INFLAMMATORY INFILTRATEKIDNEYS

HYORONEPHRSI S(MILD I

LITTON BIONETICS, INC.FRCJ!ECT NO. lT4-l5 INDIVIDUAL ANI4AL EVALUATION

ANIMAL NC. 8633 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 " LOW DOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: CORS CO(SD) RR6EEKS ON STUOY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERlqINAL WEIGHT: q72.000 GRAMS PETHOO OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:GENERAL APPEARANCE AT NECROP~f

RIGHT EAR THICKENED

MICROSCOPIC FINOINGS:

MICRCSCOPIC EVALUATION NCT REQUIREO

LITTON BIONETICSv INC.PROJECT NO. 13734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO* 8631 S.X: MALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOW DOSE SPECIES: RATSCOSr AMfUNT: 300 PPm ST.AIN: COnS C'(SO) 9REEKS ON STUDY: 13 SOURC-: CHARLSS RIVER (PORTAGE)LATE OF DEATH: 01-23-81 TYPE 5F= OEATH: 'TFRIONAL KILLTERMINAL WEIGHT: 460.000 GRAM9S METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:



MICROSCOPIC EVALUATION NOT REQUIRED

18'7

LITTON 8:ONETICS, INC.FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUAT!ON

ANIMAL NO. 8632 SEX: "ALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOW OOSE SPeCIES: RATSCOSE ANCUNT: 300 PPM STRAIN: COBS CD(SO) RP6EEKS ON STUDY: 13 SOURCE: CHARLES QIVrR (POPTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILL7ERMINAL WEIGHT: 486.000 GRA4S METHOD OF KILL: CARBON OIOXI3£

GROSS OBSERVATIONS:



MICROSCCPIC EVALUATION NCT REQUIRED

LITTON PIONETICS9 INC.FROJECT NOe 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIOAL NO. 8633 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOW DOSE SECIES: RATSCOSE AMOUNT: 30C PPM STRAIN: COBS CD(SD) BR6EEKS ON STUDY: 13 SOURCE: CHARLES RIVED (PCPTArE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 453,030 GRAMS METHOD OF KILL: CARBON DIcXIE

GROSS OBSERVATIONS:



MICROSCOPIC EVALUATION NOT REQUIREO

LITTON BIONETICS, INC.FROJECT NO. 10734-1! INDIVIDUAL ANIMAL EVALUATION

ANIAL NO. 8634 SEX: MALE PATHOLOGY REFERENCE N'o 0- 0GROUP 2 - LOW oOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CO(SOI B0WEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGE)CATE OF OEATH4: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 459.000 GRAMS METHOD OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYRIGHT EARTAG INFEC7ITO,



LITTON BIONETICSo INC.FROJECT NO. 10734-15 I1hDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8635 SEX: MALE PATHOLOGY REFERENCE PIN. 0- 0GROUP 2 - LOW DOSE SPECIES: PATSCOSE AMOUNT: 300 PPM STRAIN: COBS CD(SD) SRkEEKS ON STUDY: 13 SOURCE: CHARLES RIVEP (POR'AGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMRNL WEIGHT: 455.000 GRAMS METHOD OF KILL: CARBON or('xIx

GROSS OBSERVATIONS:




... J ... " ... " ...... ... .. .... . i .. .. . .. l ll I I 1 II ...... .. . .. .... .....

LITTON B1ONETICSPROJECT No. 10714-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8636 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0EROUP 2 - LOU DOSE SPECIES: RATSDOSE AMLUNT: 300 PPM STRAIN: COSS CD(SO) 80mEERS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATZ OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL VEIGHT: 411.000 GRAMS METHOD OF KILL: CARBON OICXIOE

GROSS OBSERVATIONS:



MICROSCOPIC EVALUATION NCT REQUIRED

LITTON BIONETICS, INC.PROJECT NO. 10?3 -15 IscIvIouAL ANIMAL EVALUATION

ANIMAL NO. 8637 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOW DOSE: SPrCIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CO(SO) BRhEEKS ON STUOY: 1! SOURCE: CHARLES RIVER (PCRTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TFRMINAL KILLTERMINAL WEIGHT: 411.OO GRAPS METHOD OF KILL: CARBON DIIXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYRIGHT EARTAG INFECTION


MICROSCOPIC EVALUATIZN NCT REQUIRED

1h

LITTON BIONETICS, INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATIONANIPAL NC. 8638 SEX: "ALE PATHOLO.Y PEFERENCE Nn,, C- 0GROUP 2 - LOW DOSE SPECrES: RATSCOSt AMGUNT: 300 PPM S-riAIN: CMBS CD(SO) ROWE7KS ON STUDY: 13 SOURCE: CHARLES RIVER IPORTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 471.00 GRAMS METHOD OF KILL: CARBON DICXI).

GROSS OBSERVATIONS:





FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8639 SEX: 4ALE PATHOLOGY PEFERENCE NC. 0- 0GROUP 2 - LOW DOSE SPECIES: RATSCOSE AMuUNT: 30a PPM STRAIN: COBS CO(SO) OFhEEKS ON STUDY: 13 SOURCe: CHARLES RIVER (PCRTAGE)DATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 503,000 GRAMS METHOO OF KILL: CAPBC'4 OIOXIr

GROSS OBSERVATIONSt



MICRCSCOPIC EVALUATION NCT REQUIRED

. ,_ 10

ILITTON BIONETICS, INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NOD 8640 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOW, DOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CO(SO) BRUEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGr)EATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 414.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:

ANIMAL APPEARED NORMAL GRISSLY

PICROSCcPIC FINDINGS:


LITTON BIONETICSI INC.FROJECT NO* 107 34-1 INDIVIDUAL ANIMAL EVALUATION

ANIAL NO. 8641 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOU DOSE SPECIES: RATSCOSE AMCUNT: 300 PPM STiAIN: COBS CO(SO) BRUEEKS ON STUDY: 13 SIURCE: CHARLES RIV-R (PCIRTASE)CATE OF DEATH: 01-22-81 TYCE OF DEATH: TERPOINAL KILL'TERMINAL WEIGHT: 538°030 GRAMS PETHOO OF KILL: CARBON 0I(IDE

GROSS OBSERVATIONS:

ANIMAL APPEARED NORMAL GPOSSLY



"--"-iiii . ... i .. .. ... { . ... . -

LITTON RION .TICS, INC.FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATTON

ANIMAL NO. 8642 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOWI DOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CO(SO) 8BUEcKS ON STU3Y: 13 SOURCE: CHARLES RIVER (PCQTAC-E)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 429.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




LITTON BIONETICS9 INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIPAL NO* 8643 SEX: MALE PATHOLOGY REFERENCE NO* 0- 0GROUP 2 - LOW COSE SPECIES: RATSCOSE AMGUNT: 300 PP" STRA11: COBS CO(SD) RPWEEKS ON STUDY: 13 SOURCE: CHARLES RtVER (POPTAGE)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILL7ERMINAL WEIGHT: 536.000 GRAM3 METHOO OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




193

LITTON BLONETICS 1 N'FR Od CT 0~. 101H4- IND~IVrDUAL ANIMAL EVALUATION

ANIMAL hi'. 8644 SEX: MALE PATHDL-lGY REFERENCE N1, 0- aGROUP 2 - LOW DOSE SPECIES: RATSCOSS AMOUNT: 30C PPM STRAINJ: COSS CO(SD) RoUEEKS ON STUDY: 13 SCURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERR!NAL KILLTERMINAL WEIGHT: 369.000 GRAMS METHOD OF KILL: CARBON OIOXID.-

CROSS OBSERVATIONS:




LITTON SIONETICS3,INC.PROJECT NO. 10 O5-1 5 INDIVIDUAL ANTMAL EVALUATION

A~fMAL NO. 8645 SEX: MALE PATHOL!n6Y REFERENCE NO. 0- 04ROUP 2 - LOW DOSE SPECI':S: RATSCOSE AMCUNT: 303 PPM STRAIN: COBS co(so) PRUErKS ON STUDY: 13 SOURCE: CHARLFS RIVER CPORTAGE )CATE OF DEATH: 01-21-81 TYPE OF DEATH: TrRUIT*AL KILLTERMINAL WEIGHT: 4899000 GRAMS METHOD OF KILL: CAPSON OIOXIDE-

GROSS OBSERVATIONS:

ANIMAL APPEARED NURMAL GROSSLY

PTCROSCCPIC FrNoINeS:

MICROSCOPIC EVALUATION N"'T REQUIRED

1934

LITTON BION'TICS INC.FROJECT NO. 1O73S-1! INOIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8646 SEX: MALE P4THOLOGY REFERENCE NCe - 0GROUP 2 - LOW DOSE SPECIES: OATSCOSE AMCUNT: 300 PPM STRArN: COBS CO(SD) BR6EEKS ON STUDY: 13 SOURCE: CHARLES RIVER 10CPTAGE)CATE OF DEATH: 01-21-81 TYPE OF OEATH: TERMINAL KILLTERMINAL WEIGHT: 461.000 GRAM3 METH3D OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




LITTON BIONETICS1 INC.

FROJECT NC. 1073€-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8647 SEX: MALE PATHOLOGY REFERENCE NO. a- 0EROUP 2 - LOW DOSE SPECIES: PATSCOS _ AMOUNT: 300 PPM STRAIN: COBS COCSO) BPbEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-21-81 TYPE OF OATH: TERMINAL KILLTERMINAL MEIGHT: 4449000 GRAMS METHOD OF KILL: CARBON DIOXIDe'

GROSS OBSERVATIONS:




.15

LITTON BICNETI CS IhC.FROJECT NO. 1 a734-15 INDIVIDUAL ANIMAL EVALUATION

ANITMAL NO. 8648 SEX: MALE PATHOLOGY REFERENCE NO. 0- 9GROUP 2 - LOW OSE SPECIrS: PATSCOSE A'4CUNT: 3013 PPM STRAIR: CCBS CO(SD) OQWEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCPrAGE)CATE OF DEATH: 01-21-81 TYDE OF DEATH: TFRmINAL KILL7ERMINAL WEIGHT: *a5oO00 GRAMS META03 OF KILL: CARBON DIOXIDE

GROSS oeSERVATIONS:


MICV -OPIC FINDINGS:

[CROSCOPIC EVALUATION NCT REQUIRED

LITTON sIONETICS9 INC.FROJECT NO. 10734-15 INDIVIDUAL ANJIMAL EVALUATION

ANIMAL NC. 8649 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOW DOSE SPE-CIEs: RATSCOSE AMOUNT: 300 PPM ST4AIN: COBS CO(SD) RR6EEKS ON STUDY: 13 SOURCE: CHARLES RIVER (POQTAGZ)CATE OF DEATH: 01-21-81 TYPE OF OFATH: TERMINAL KILLTERMINAL WEIGHT: 462.oOO GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




lf.9

LITTON RIONETICS9 INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATIflN

ANIMAL NO. 8650 SEX: MALE PATHOLOGY REFERENCE NO. 0- aGROUP 3 - MEDIUM DOSE SP;CIES: RATSCOSE AMOUNT: IG0 PPM STI.AIN: COBS CD(SOV BP6EEKS ON~ STUDY: 13 SOURCE: CHARLES RIVEP (PCPTAr)CATE OF DEATH: 01-23-81 TYPE CF DEATH: TERMINAL KILLTERMINAL WEIGH4T: 4470000 GRAMS METHO0 OF-KILL: CARBON DICNtDE

GROSS OBSERVATIONS:



MICROSCOPIC EVLUATION NOT REQUIRED

LITTON sIONETICSt INC*PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATICN

ANIMAL NO. 8651 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0CROUP 3 - ME0IUP DOSE SPECIES: RATSCOSr AMCUNT: loco PPM STRAIN: COBS CD(SD) BRWEEiS ON STUDY: 13 Sn'URCE: CHARLES RIVER (PCPTAGE)CATE OF DEATH: 01-23-81 TYPE 00 DEATH: TSRMINAL KILLTERMINAL WEIGHT: 540.000 rGRAMS PETHOD Of KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYRIGHT EARTAG, INFECTION



I L-----

LITTON IONETICSI INC.PROJECT IO. 107 4-15 INOIVIOUAL AmIrMAL EVALUATION

ANIMAL NO 8652 SEW: MALE PATHOLOGY REFEPRENCE NO. a- aGROUP 3 - MEDIUM DOSE SPE'CIES: RATSCOSZ AMOUNT: 1000 PPM STRAIN: COBS CDtSD) REEKS ON STUIY: 13 SaURCZ: CHARLES RIVER (PCRTAGE)

CATE OF DEATH: 01-23-81 TYPE OP DEATH! TERMINAL KILLTERMINAL WEIGHT: 412.000 GRAMS METHOD OF KILL: CARBON4 DIOXIOE

GROSS OPSERVATIONS:


FICROSCOPrC FINDINGS:


LITTON BIONETICS9 INC.PROJECT NO* 10?34-15 INOIVI3UAL ANIMAL EVALUATION

ANIMAL NO. 8653 SEx: MALE PATHOLOGY REFERENCE NO. C- VGROUP 3 - MEDIUM DOSE SPErCISS: RATSCOSE AMOUNT: 10C PPM STRAIN: COBS C1)(SO) PRbo-:KS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAE)ATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 453.000 GRAMS METHOD OF KILL: CARBON 017XI3E

GROSS OBSERVATIONS:



MICROSCOPIC EVALUATION NOT REQUIRZ-D

LITTON PIONETICS, INC.FROJECT NO. 1 T34 115 INCIV;DUAL ANIMAL EVALUATION

ANIMAL NO. 8654 SEX: MALE PATHOLOGY REFERENCS NO. 0-GROUP 3 - MEDIUM DOSE SPECIES: RATSCOSE AMOUNT: 1002 PP" STRAIN: COBS Co(tO) spEEKXS ON STUDY: 13 SIURCC: CHARLES RIVER (PCRTAGS)CATE OF DEAT4: 01-23-81 TYPE OF DEATH: TIRW!NAL XILLTERMINAL WEIGHT: 447.000 GRAMS PETHOL OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




LITTON RIONETICS. INC.PROJECT NO. 10134-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL 40. 8655 SEX: MALE PATHOLOGY REFERENCE NO. 0- CGROUP 3 - MEDIUM DOSE SPECIES: RATSDOSr AMOUNT: 1000 PPM STRAIN: COBS CO(SO) BWEzKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGF)tATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAi WEIGHT: 463.0 0 GRAwS METHOO OF KILL: CARRON DIOXIOE

GROSS OBSERVATIONS:


PICROSCOPIC FINOINGS:

MICROSCOP!C EVALUATION NOT REQUIRED

.

19

LITTON eIGNETICS, INC*FROJECT NO. 10134-15 INDIvIDUAL ANIMAL EVALUATION

ANIMAL NO. 8656 SEX: MALE PfATHOLOGY REFERENCE Nl). 0- CGROUP 3 - ME-DIUM DOSE SPrCICS: RATSLOSE AMCUNT: 1000 PPM STRA!N: COSS Cn(SD) RPWEEKS ON STUDY: 13 SCURCE: CHARLES QTVER (PCPTASE-)DATE OF DEATH: 01-23-81 TYPE OF DFATH: TrRW'rNAL KILLTERMINAL WEIGHT: 521.000 GRAMS MTrHOC OF KILL: CAR8ON orOxi:)r-

GROSS OBSERVATIONS:


MICROSCOPIC FIN!DINGS:

MICRCSCOPIC EVALUATION NCT REQUIRED

LITTON PIONETICS3,1NC.PROJECT NO. 107 415 INDIVIDUAL ANIMAL EVALUATION

ANIPAL NO. 865? SEX: MALE PATHOLOGY REFERENCE NO. 0- 0GRIUP 3 - MEDIUM DOSE SPECIES: RATSCOSF AMCUNT: 1001 PPM STRAIN: COSS CDCSD) ORUEEKS ON STUDY: 13 SOURCE: CHARLE:S RIVER (PCPTAGE)CATE OF DEATH: 01-22-81 TYPE 17F DEATH: TERMiiNAL KILLTERMINAL WEIGHT: 452.000 GRAMS PETHOO OF KILL: CARBON OrCXI~r-

GROSS OBSERVATIONS:

GENERAL APPEARANCE, AT NECROPSYRIGHT EARTAG INFECTION



zoo

LITTON BIONETICS9 INC.FROjECT NO. 10734-15 INOIV.'OUAL ANIMAL EVALUAT!ON

ANIMAL NO* 8658 SEX: MALE PATHOLOGY REFERQENCE- NO. 0- 0CROUP 'R - MzDIUM DOSE SP;CISS: RATSCOSr- AMCUNT: i0oo PPM STRAI': COS co(SO) BPimEEKS ON STUDY: 13 SOURCE: CHARLES RIVrR (PORTAGE)CATE OF DEATH: 01-22-81 TYPr OF DEATH: TERMINAL KILLTERPINAL WEIGHT: 489.000 GRAMS PETHOD OF KILL: CARBONJ oIcxlOE

GROSS OBSERVATIONS:




LITTON BIONETI CSICFROJECT NO. 03-15 INDIVIaUAL ANI4AL EVALUATION

ANIMAL NO. 8659 SEX: MALE PATHOLOGY REFERENCE NO* 0- 0GROUP 3 - MEDIUM DOSE SPECIES: RATSCOST AMOUNT: 1OO00 PPM STRAIN: COSS C0(SD) RR6EEXS ON STUDY: 13 SnURCE: CHARLES RIVER cPORTASE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 488e000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OOSERVATIONS:




201

K - -

S LITTON BI04ETICS3,INC.

FROJECT NO. 10? s3 INDIVIDUAL ANIrAL EVALUATIONANIMAL N3* 8660 SEX: MAL-E PATHOLOGY REF '-NC - NC. 0- 0

CROUP 3 - MEDIUp DOSE SPRCIES: RATS

COSE ANGUNT: looc PPN STRAIN: CBS R RCIS) HiR

WEEKS ON STUOY: 13 SIUR-Et CHARLES RiveR (PORTAGE)

CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILL

TERPI'dAL WEIGHT: 468.000 GRAMS WETHOD OF KILL: CARBOJ DIOXIDE

GROSS OBSERVATIONS:




LITTON BIONETICS9 INC.FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL GO. 8661 SEX: MALE PATHOLOGY REFERENCE Nfo. - 0

GROUP 3 - NEDIUP DOSE SPECIFS: RATS

ROSE ANOUNT: 1000 PPM STRAIN: COBS CO(S01 RR

UEEKS ON STUJY: 13 SOURCE: CHARLES PIVR (PCRTAGr)

CATE OF DEATH: 01-22-81 TY0 E OF DEATH: TERMIIIAL KILL

TERMINAL WEIGHT: 482.000 GRAMS PET4OD OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:


MICROSCOPIC FINDXNGS:

MICROSCCPIC EVALUATION NCT REQUIRED

Zo~w

LITTON BIONETICS, INC.FROJECT NO. 10734-15 !NDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8662 SEX: MALE PATHILnGY REVERENCE NC. a- 0GROUP 3 - MEDIUM DOSE SPECIES: RATSCOS! AMOUNT: 1000 PPM ST4ArI: COBS Co(SO) RehEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (POPTAG5)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TER'4INAL KILLTERMINAL WEIGHT: 492.000 GRAMS METHOD OF KILL: CARBON DICXZDE

GROSS OBSERVATIONS:GENERAL APPEARANCE AT NECROPSY

TEETH CUT



LITTON BIONETICS. INC.FROJECT NO. 10754-15 INDIVIDUAL ANIMAL EVALUATIONLIOS N AM O N T S 100 PP S PEI E: C B D S )ANIMAL NO. 8663 SEX: MALE PATHOLOGY REFERENCE NO. 0- CGROUP 3 - MDIUT DOSE SPECIES: RATSCOSE AMOUNT: 1000 PPM STRAIN: COBS CO(SD) BQ

WEEKS ON STUDY: 1! SOURCE: CHARLES RTVER CPCPTAGE )

DATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 422.000 GRAMS METHJO OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS :

GENERAL APPEARANCE AT NECROPSYEARTAG rNFECTIONTEETH CUT



0

Z0;3

I ILITTON BIONETICS, NC.FROJECT NO. 1073#-15 IINDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8664 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0GROUP 3 - MEDIUM OOSE_ SPECIES: RATSCOSE AMOUNT: 1000 PPM STRAIN: COBS CO(SO) BRhEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATE OF DEAT4: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 5029000 GRAMS MITHOO OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:



MICROSCOPIC EVALUATIO14 NCT REQUIRED

LITTON BIONETICS, INC.PROJECT NO. 1071t15 INDIVIDUAL ANIMAL eVALUATION

ANIMAL NO. 8665 SEX: MALE PATHOLOGY REFERENCE NC. 0- 0GROUP 3 - MEDIUM DOSE SPECIES: RATSCOS. AMOUNT: 1000 PPM STRAIN: COSS CO(SO) BVEEKS ON STUDY: 13 SIURCE: CHARLES RIVER (PCRTAr' )CATE OF DEATH: 01-21-81 TYPE OF OATH: TERMINAL KILLTERMINAL WEIGHT: 547.000 GRAMS METHOD OF KILL: CARBON' DIOXIDE

GROSS OBSERVATIONS:




ZOI

LITTON BIONETICS9 INC.FROJECT NO. 10734-15 ISDIVIDUAL ANIMAL EVALUATION

ANIMAL kh. 8666 SEX: MALE PATHOLOGY REFERENCE Nfw 0- 0C-ROUP 3 - MEDIUM DOSE SPrCIES: RATSCOSE AMOUNT: 10o0 PPM STRAIN: COBS CD(SO) 80WEEKS O STUOY: 13 SIURCE: CHARLES RIVER (PORTAGE)CATr OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 410*030 CZAWM METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT 4ECROPSYCRUST AND SWELLINq AT EARTAG SITE


MICROSCOPIC EVALUATION NOT REQUIVEO

LITTON PIONETICSs INC.FROJECT NO. 10134 15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8667 SEX: MALE PATHOL'!GY REFERENCE NO. 0- CGROUP 3 - MEDIUM DOSE SOrCIES: RATSCOSE AMOUNT: 1000 PPM STRAIN: COBS CD(SDI aQ

WEEKS ON STUDY: 13 SOURCE: CHARLES PIVER (PCRTAGE)CATE OF DEATH: 01-21-B1 TYPE OF DFATH: TERM!NAL KILLTERMINAL WEIGHT: 4980O0O GRAMS METHOD OF KILL: CARBON DIOXIT

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYEARTAG INFECTION


NICROSCOPIC EVALUATION NCT REQUIRED

20

205

LITTON SIONETICS9 INC.FROJECT NCO 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL N-. 8668 SEX: MALE PATHOLIGY REFERENCE NO. 0-cROUP 3 - MEDIUM DOSE SPECIES: RATSCOSE AMCUNT: 1000 PPM STRAIN: CORS CD(SD) BRbEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAG7)CATE OF DEATH: 01-21-81 TYPE OF DEATH. TER;INAL KILLTERMINAL MEIGHT: 515.000 GRAMS METHOD OF KILL: CARBON DIOXIDF

GROSS OBSERVATIONS:

LUNG3PINPOINT WHITE FOCI ON ALL LOBES



LITTON EIONETICS9 INC.FR3JECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8669 SEX: MALE PATHOLOGY REFERENCE NO. 0- 0GROUP 3 - MEOIUM DOSS SPrCIES: OATSCOSE ANCUNT: 1000 PPM STQAIN: COBS CO(SO) BRWEEKS ON STUDY: 13 SOURCE: CHARLES R!VER (PORTAGE)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGH': 553.030 GRAMS PETHOI OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:


MICROSCOPIC FINOINGS:


206

LITTON RIONETICS, INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIM4AL NC. 8,67C SEX: MALE PATHOLOGY REFERENCE MT. 81-04G91ERIUP 4 - HIGH DOSE SPE-CIES: RATS-COSE AMOUNT: 3O,30 PPM STRAIN: COBS CD(SO) BpIoEEKS ON STUDY: 13 SgURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: T--RMINAL KILLTERMINAL WEIGHT: 416.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




SPLEEN(MLHEMATOPOIESIS (EXTRAMEDULLARYI

(MILOLUNGS

BLOOD VESSELLYMPHOCYTic INFLAMMATORY INFILTRATE

KIDNEYSFO CAL

MINERALIZATION(MINIMA.

LjTT ON e IONETICS,,INC.F OJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8671 SEX: MALE PATM3LOGY REFERENCE NO. 81-0*092lGROUP 4 - HIGH DOSE SPECIES: RATSCOSE AMOUNT: 3000 PPM STRAIN: CCRS CD(SO0 99WEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: 'ERMTNAL KILLTERMINAL WEIGHT: 446.000 GRA14S PETHOO OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYHAIR LOSS* FCRE LIPBS



(MILDOSPLEEN

HEMATOPOIESIS (EXTRAMEDULLARY)(MILD

LUNGSBLOOD VESSEL

MINERALIZATION(MINIMAL


207

LITTON BIONETlCS INC.FROJECT NO* 10T74-1! INDIVIDUAL ANIMAL EVALUATIONANIMAL 1N0- 8672 SEX: MALE PATHOLOGY REFERENCE Nmo 81-04093

GR3UP 4 - HIGH DOSE SPSC15S: RATS

COSE AMOUNT: 3000 PP" STRAIN: COBS CO(SD) R

%EEKS Oh STUOY: 13 SOURCe: CHARLES RIVER (PCRTAqE)

CATE OF OEATH: 01-23-81 TYPE 8F DEATH: TFRqINAL KILL

TERMINAL WEIGHT: 418o000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:



STOMACHINFLAPMATION9 NOS

(MILDLUNGS

BLOOD VESSELMINERALIZATION

(MINIMAL)

SITTON BIONETICS, INC.ROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8673 SEX: MALE PATHOLOGY REFERENCE NO. B1-04094

GROUP 4 - HIGH DOSE SPECIES: RATSLOSE AMOUNT: 300 PPM STqATN: CnBS CO(SO) BR

%ErKS ON STUDY: 13 SIURCE: CHARLES RIVER (PCRTAGE)

CATE OF DEATH: 01-23-81 TYPE OF DF4TH: TERMINAL KILL

TERMIINAL WEIGHT: 503.000 GRAMS METHOD OF KILL: CARBON DIOXIOr

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYEARTAG SITE SWOLLEN, ENCRUSTED

MICRf,3COPIC FINDINGS:

HEART OEGENERATICN, NOS

(MILD PSPLEEN

HEMATOPOIESIS tEXTRAMEDULLARY)(MILO

LUNGSBLOOD VESSEL

LYMPHOCYTIC INFLAMPATORY INFILTRATE

A.08

LITTON BIONETICSj IhC.PROJECT NO. 101 -15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO, 8674 SEX: MALE PATHOLOGY REFERENCE NO. 81-040!5CROVP 4 - HIGH DOSE SPECIES: RATSCOSE AMCUNT: 3000 PPM STRAIN: COBS CO(SO) 9Q6EEKS ON STUDY: 13 SURCE: CHARLES P[VrR (PORTAG_)CATE OF DEATH: 01-25-81 TY0 E OF OEATH: TFRMINAL KILLTERMINAL WEIGHT: 506o000 GRA MS METHD OF KILL: CARRON DIOXIDE

GROSS ORSERVATIOlS :

ANIMAL APPEARED hORMAL GROSSLY


STOMACHINFLAMMATION, NOS(MILD )

LUNGSBLOOD VESSEL


BLOOD VESSELLYMPHOCYTIC INFLARwATORY INFrLTRATS

KIDNEYSHYORONEPHRCSIS

PROSTATEINFLAMMATICN, NOS

(MILD )


ADRENAL SLANDSSECTION AND/OR TISSUE NOT PRESENT

MAMMARY GLANOSECTION AND/OR TISSUE NOT PRESENT

LITTON BIONETICS% INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATIOJN

ANIMAL NO. 8675 SEX: MALE PATHOLOGY REFERENCE NO. 81-04096CROUP 4 - HIGH COSE SPECIES: RATS-OSE AMOUNT: 3000 PPM STRAIN: COBS CD(SD) BPUEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 4246000 GRAMS MPTHOD OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYSUBCUTANEOUS THICK RED FLUID* RIGHT EYERED CRUST AROUND RIGHT EYE


HEARTDEGENERATICN9 NOS

(MINIMALSTOMACH

INFLAMMATION, NOS(MILD

SPLEENHEMATOPOIESIS (EXTRAMEDULLARY)

(MILD ILUNGS



MAMmARY GLANDSECTION ANOfOR TISSUE NCT PRESENT ZC,)

LITTON 8IONETICSj INC.FR3JECT NO* 107 ~15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8676 SEX: MALE PATHOLOGY REFERENCE P40. 81-04097GROUP 4 - HIGH DOSE S~rCIrS: RATSCOSE AMCUNT: 3000 PPM STRAIFN: Cf(RS CD(SO) APUEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 457*000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYSUBCUTANEOUS ACCUPULATION OF BLOOD AOOVE SKULLRIGHT EYE ULCERATE0, YELLCW9 PROTRUDING



(MINIMALSPLEEN

"EMATOPOIESIS (EXTRAMEOULLARY)(MILO

LEFT EYEINFLAMMATION, ACUTE

EYE/CORNEAINFLAMMATION, ACUTE

(SEVERE

LITTON BIONETICSv INC.FROJECT NO. 10734-15 INDIVIDUAL ANIL EVALUATION

ANIMAL NC. 8677 SEX: MALE PATHOLOGY REFERENCE NC. 81-C4096GROUP 4 - HIGH DOSE S~rCIES: RATSDOSE AMOUNT: 3000 PPM STRAIN: COSS CO(S ' RUEEKS ON STUDY: 1! SrIURCF: CHARLES PIVS9 (PCPTACE)CATE OF DEATH: 01-22-81 TYPE OF OSATH: TFP0!NAL KILLTERMINAL WEIGHT: 404.000 GRAMS METHOD OF KILL: CARSON DiIrVYiE

GROSS OBSERVATIONS:



STOMACHINFLAMPATION, NOS

(MILOCECUM

INFLAMATION, NOS(MILO

LUNGSBLOOD VESSEL

LYMPHOCYTIC INFLAMPATODY INFILTRAT:-

LITTON BIONETICS9 INC.FROJECT NC. 10734-15 IhOIVIOUAL ANIMAL EVALUATION

ANIPAL NC. 8678 SEX: MALE PITHILOGY REFERENCE N'). B1-04C' iGR'UP 4 - HIGH D0Sr SPECIES: RATSLOSE AMCUNT: 3000 PPii STAAV: COSS CDCSD) F9UEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TFRMPJNAL KILLTERPINAL WEIGHT: 463oOO0 GRAMS PETHOD OF KILL: CARBON DIoxiDE

GROSS OBSERVATIONS:


P[CROSCCPIC FINDINGS:


(MILOLUNGS SICYOS

(MILOBLOOD VESSEL

LYMPHOCYTIC INFLAMMATORY INFILTRATEKID%~E YS

1(IONEY/CORTEXCYST, NOS

LITTON 8IONETICS# INVC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8679 SEX: MALE PATHOLOGY REFERENCE NO*. 81-041COGROUP 4 - HIGH DOSE SPECIES: RATSGOSE AMCUNT: 3000 PPM STRAIN: CnHS CO~So) So6E:'KS ON STUDY: 13 SOUJRCE: CHARLc:S PIVER (PORTAGE)CAT'E OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLIERMINAL WEIGHT: 435.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

CROSS OBSERVATI0M:

GENZRAL APPEARANCE AT NECROPSYRIGHT EARTAG INFECTION



(MINIMAL

LITTON PIONETICS, INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8680 SEX: MALE PATHOLOGY REFERENCE N)0 81-04101GROUP 4 - HIGH DOSE SPECIES: RATSCOSE AMCUNT: 3000 PPM STRAIN: COBS CD(SO) 80WEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGE)DATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 429.000 GRAMS METHOD OF KILL: CARBON DIOXI!r

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYTEETH CUTRED CRUST AROUND RIGHT EYE



(MILO )CECUM

INFLAMMATION, NOS(MILD )


(MILD )LUNGZBLOOD VESSEL



THYMUSSECTION AND/OR TISSUE NOT PRESENT

LITTON BIONETICS9 INC.PROJECT NO. 10734-15 I&OIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8681 SEX: MALE PATHOLOGY REFERENCE NO. 81-04C2CROUP 4 - HIGH DOSE SPECIES: RATSCOSE AMOUNT: 3000 PPM STRAIN: COBS CO(SO) RWEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (POFTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERVINAL KILLTERMINAL WEIGHT: 4449000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYRIGHT EAR THICKENEC



(MILDLUNGS

INFLAMMATION, NOS(MINIMAL)

PROSTATEINFLAMMATION, NOS

(MILO )

-- ]2

LITTON RION 'TICS* INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO* 8682 SEX: M&LE PATHOLOGY REFERENCE N%. 81-04103GRaup 4 - HIGH DOSE SP7CIES: RATSCOSE AMOUNT: 300C PPM STRAI'): CCSS CD(SD) RO'L6EEKS ON STUDY: 13 SOUJRCE: CHARLES PIVFR (PCRTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERPINAL WEIGHT: 454.000 GRAMS METHOD OF KILL: CARBON DIOXICE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYRIGHT EAR THICKENED



(MINIMAL)LUNGS

BLOOD VJESSELMINERALIZATION

(MILDBLOOD VESSEL

LYMPH0CYTIC INFLAM-ATORY INFILTRATE

LITTON BIONETICSI.PRG4ECT NO. 10341N5S INCIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8683 SEX: MALE PATHOLOGY REFERENCE N13. 81-04104GROUP 4 - HIGH DOSE SPECIES: PATSCOSr AMOUNT: 3000 Pom STPAt'J: COBS CtlSOD SP6EEKS ON STUOY: 13 SIURCE: CHARLIES P!VEP (PCRTAGE)CATE OF DEAT4: 01-22-81 TYPE flF DEATH: TFRMTNAL KILLTERMINAL WEIGHT: 448.000 GRAMS PETHOO OF KILL: CA0B"% CICX)t'E

GROSS OBSERVATIONS:




(MINIMAL3STOMAC H

INFLAMMATION, NOS(MILO

CECU MINFLAMMATION, NOS

(MILD )SPLEEN

HEMATOPOIESIS (EXTRAMEDULLAPY)(MILD

LUNGSINFLAMMATION, FOCAL

(MINIMAL3BLOOD VESSEL


LITTON BIONETICS IiNC.PROJECT NO* 1073415 IOIV!DUAL ANIMAL EVALUATION

ANIMAL h~o 8684 SEX: MALE PATHOL )GY REFERENCE NO. 81-041C5GROUP 4 - HIGH OOSE SPtCIES: RATSCOSE AMOUNT: 3O000 PPM STRAIN: COBS COCSO) BRUEEKS Oh STUDY: 13 SOURCE: CHARLES RIVER (PCRTAG7)CATE OF DEATH: 01-21-81 TYPE OF 07ATH: TERMINAL KILLTERMINAL WEIGHT: 402e030 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:



PA NCRE ASINFLAMMATIaN, FOCAL

(MINIMALCE CUP


SPENHEMATOPOIESIS (EXTRAMEDULLARY)

KIOEYS(MINIMAL3

HYURONEPHROSI S(MODERATE)

TISSUrE(S) MISSING OR SEVERELY AUTOLYZEO:

PITLITARYSECTION AND/OR TISSUE NOT PRESENT

LITTON BrONETICS, INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NC* 8685 SEX: MALE PATHOLOGY REFERENCE NO. 81-04106GROUP 4 - HIGH DOSE SPECIES: RATSCOSE AMCUNT: 3000 PPM STRAIN: CnBS CO(SO) SRUEEKS ON STUDY: 13 SOURCE: CHARLTS R!VER (PORTAGE)CATE O)F DEATH: 01-21-81 TYPE OF DEATH: TtRMINAL KILLTERMINAL WEIGHT: 388.000 GRAMS M7ETHOD OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:



LUNGSBLOOD VESSEL

LYMPHOCYTIC INFLAMPATORY INFILTRATEKIDNEYS

INFLAMMATION, FOCAL(MODERATE)

LITTON BIONETICS. INC*PRcOJECT NO. 10734-15 INDIVIOUAL ANIMAL EVALUATION

ANIMAL NO- 8686 SEX: MALE PATHOLOGY PEFERENCE N'o 81-04107GROUP 4 - HIGH COSE SPECIrS: RATSCOSE AMOUNT: 3000 PPM STRAN: COSS CD(SD) BR

IEEKS ON STUDY: 13 SIURCE: CHARLES RVEP (PORTAGE$

CATE OF DEATH: 01-21-81 TYPE OF DEATH: TFRMINAL KILL

TERMINAL WEIGHT: 429.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NrCROPSY

EARTAG INFECTION


STOMACHINFLAPMATION, NOS

(MILD )LUNGS

BLOOD VESSELLYMPHOCYTIC INFLAMPATORY INFILTRATE

LITTON BIONETIC3, INC.PROJECT NO. 10 34-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8687 SEX: MALE PATHOLOGY REF- RENCE NO. 81-041CCROUP 4 - HIGH OOS!: SPeCIeS: PATSDOSE AMUUNT: 3003 ppM STRAIN: COBS CD(SD) BPIEEKS ON STUDY: 13 Sn'URCE: CHARLIS PrVER (PORTAGE)CATE OF DEATH: 01-21-01 TYPE OF DEATH: TERmINIAL KILLTERMINAL WEIGHT: 429.000 GRAMS METHOD OF KILL: CARSCN DIOXIDES

GROSS OBSERVATIONS:




(MILD IKIDNEYS

KIDNEYfPELVISMINERALIZATION

(MILD )

G" ,42,,.

LITTON BIONETICS, INC.PROJECT NO. 10734-15 IhDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8688 SEX: MALE PATHOLOGY REFERENCE NO. 81-14109GROUP 4 - HIGH OSE SP-CIES: RATSCOSE AMOUNT: 3000 PPM STRAIN: coos CD(SD) BPWEEKS ON STUOY: 13 SOURCE: CHARLES RIVER (PCPTAGE)CATE OF DEATH: 01-21-81 TYPE OF DrATH= TERMINAL KILLTERMINAL WEIGHT: 503.000 GRAMS P-THOD 0: KILL: CARB'N DIOXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYRIGHT EAR THICKENED


STOMACHINIFLAMMATION, NOS

(MILD )LUNGS


LITTON RIONETICS, INC.FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NC. 8689 SEX: MALE PATHOLOGY REF-RENCE NO. 81-04110GROUP 4 - HIGH DOSE SPECIES: RATSCOSE AMOUNT: 3000 PPp STRAIN: COBS Co(S0) BP6EEKS ON STUDY: 13 SOURCE: CHARLES RIVEP (PORTAGIE)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLIERMINAL WEIGHT: 369.000 GRAMS METHOD OF KILL: CARBON DIOXIO

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYRIGHT EAR THICKENEC, EARTAG INFECTION

LUNGSPINPOINT WHITE FOCI ON ALL LCFES


STOMACHINFLAMMATION, NOS(MINIMAL


(MILD )LUNGS

HISTIOCYTOSIS(MINIMAL )


MAMMARY GLANDSECTION AND/OR TISSUE NOT PRESENT


ANIMAL NO. 8695 SEX: FEMALE PATHOLOGY REFERENCE N1. 81-04071GROUP 1 - UNTREATED CONTROL SorCIES: RATSCOSE AMCUNT: a PPM STRAIN: CCBS CO(SO) RRkEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TER4INAL KILLIERMINAL WEIGHT: 249e000 GRAMS METHOD OF KILL: CARBON DIOXIDE

EROSS OBSERVATIONS:




(MILD )SPLEEN

HENATOPOIESIS (EXTRAMEOULLARY)(MODERATE)


ANIMAL NO. 8696 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04C72GROUP I - UNTREATED CONTROL SPECIES: RATSCOSE AMOUNT: 0 PPM STRAIN: COB CO0S0) RRkEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGE)CATE OF DEATH: 01-23-81 TYPE Cf: DEATH: TERMINAL KILLTERMINAL WEIGHT: 0.000 GRAMS METHOD OF KILL: CARBON OIrIXI)E

GROSS OBSERVATIONS:


FICROSCGPIC FINDINGS:


(MINIMAL)SPLEEN

HEMATOPOIESIS (EXTRAMEOULLARY)(MODERATE)

V-4

LITTON BIONETICS, INC*FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO* 8697 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04C7 3CROUP 1 - UNTREATED CONTROL SPECIES: RATSCOSE AMOUNT: C PPM STRAIN: COSS CD(SD) BRbE7KS ON STUDY: 13 SOURCE: CHARLES P:'VER (PCPTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH:- TERMINAL KILLTERMINAL WEIGHT: 272.000 GRAMS METHOD OF KILL: CARBON OICXICr-

GROSS OBSERVATIONS:



LIVERINFLAMMATION, MULTIFOCAL

(MINIMAL IHEMATOPOIESIS (EXTPAMEOULLARY)

(MINIMALSTOMACH

INFLAMMATION, NOSSPLEEN(MINIMAL

HEMATOPOIESIS (EXTRAMEOULLARY)(MODERATE)

LUNGSFOCAL

MIST IOCT TOSI S(MINIMAL

KIDNEYSINFLAMMATION, CHRONIC FOCAL

(MINIMAL

LITTON BIONETICS, INC.FROJECT NO* 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8698 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04CY4CROUP 1 - UNTREATED CONTROL SPECIES: RATSCOSE AM(;UNT: i PPM STRAIN: COSS CO(SD) BRUEEKS ON STUDY: 13 SIURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-23-81 TYP ' OF DEATH: TERMTNAL KILLTERPI'dAL WEIGHT: 215.000 GRAMS METHOD OF KILL: CAPB,IN DIOXIDE

GROSS OBSERVATIONS:

LUNGSLUNGS SMALL AND MOTTLED (NOTED AT TRIMMING)


ST01MAC HINFLAMMATION, NOS

(MIhIMALSPLEEN

HEMATOQOIESIS (EXTRAMEDULLARY)(MODERATE)

'qESENTERIC LYMPH NODEPLA SMACY 10515

(MODERATE)LUNGS

INFLAMMATION, CHRONIC FOCAL(MODERATE)

KIDNEYSHYDRONEPHRCSIS

KIDNEY/PELVISCALCULUS, NOS

LITTONT BN.~ 107 . INDIVIDUAL ANIMAL EVALUATIONj

ANIMAL NO- 8699 SEX: FEMALE PATHOLOGY REFERENCE NO* 81-O47'5

GROUP I - UNTREATED CONTROL SPECIES: RATSCOSE AMOUNT: 0 PPM STRAIN: COBS Cn-CSOI RR

6EEKS ON STUDY: 13 SOURCE: CHARLES RIVER cPCPTAGCJCATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILL

TERMINAL WEIGHT: 66e000 GRAMS PETH10 OF KILL: CARBON uroxivE

GROSS OBSERVATIONS:

ANIMAL APPEARED NCRFAL GROSSLY


HEART DEGENERATION, NOS

STOACH(MINIMAL )IINFLAMMATION, NOS

(MILD )

LEMHYPERPLASIA, LYMPHOID(MODERATE)

SPLEEN HEATOPOIESIS (EXTRAMEDULLART)

LUNGS (MODERATE)LNSPNEUMONIA, CHRONIC MURINE

(MILD )

LITTON BIONETICS, INC.FROJECT NO. 10734-15 INDIVIDUAL ANJIMAL EVALUATION

ANIMAL No. 8702 SEX: FEMALE PATHOLOGY REFERENCE N-- 81-04076C-ROUP 1 _ UNTREATED CONTROL SPrCIZS: RATSCOSE AMCUNT: 0 PPM STAAIN: COPS CO(S'J) SP

UEEKS ON STUDY: 13 SOURCE: CMARLTS RIVER (POOTAGE)

CATE- OF DEATH: 01-23-81 TYPE OF DEATH: TERMV4AL KILL

TERMINAL WEIGHT: 296.000 GRAMS FETHOC Op KILL: CARe8?N nIrE

GROSS OBSERVATIONS:

LVRSMALL YELLOW NODULES ON LEFT MEDIAN LOBE (NOTED AT TRIMMING)


LVRMETAMORPHOSIS, FATTY(MINIMAL


(MILDLUNGS

FOCALHISTIOCYTOSIS

(MINIMAL)KIDNEYS

HYORONEPHRCSI S

LITTON BIONETICS, INC.FR3JECT NO. 107 i-1s INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8701 SEX: FEMALE PATHOLOGY REFERENCS Nl. 81-0477GROUP 1 - UNTREATED CONTROL SPECIES: PATSDOSE AMCUNT: 0 PPM STRAIN: COBS CD(SO) 806EEKS ON STUOY: 13 SOURCE: CHARLES RIVER (POPTAGS)CATE OF DEATH: 01-23-81 TYPE OF OEATH: TERMINAL KILL1ERMINAL WEIGHT: 231.000 GRAMS METHOO OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:


MICROSCOPIC FINDINGS'

STOPACHIN LAM4ATION, NO3

(MINIMAL )SPLEEN

HEMATOPOIESIS (EXTRA4EOULLARY)(MILD )

LITTON BIONETICS, IjNC.FRO ECT NO* 107 4-15 TNOIVIOUAL ANIMAL EVALUATION

ANIMAL NO. 8702 SEX: FEMALE PATHOLOGY REFERENCr NO. 81-04078GROUP 1 - UNTREATED CONTROL SPECIES: RATSCOSE AMCUNT: 0 PPM STRAIN: COBS CD(SO) BRhEEKS ON STUDY: 13 S3URCE: CHARLES RIVER (PCRTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 228.000 GRAMS METHOD OF KILL: CARBON OICXIDE

GROSS OBSERVATIONS:




(MINIMALSPLEEN

HEMOSIDERCSIS(MILD 1

LUNGSHISTIOCYTOSIS

(MINIMALBLOOD VESSEL

LYMPHOCYTIC INFLAMMATORY INFILTRATE

220

LITTON RIONETICSS INC.FROJECT NO* 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NC. 8703 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04C79GROUP I - UNTREATED CONTROL SPECIES: PATSCOSE AMOUNT: 0 PPM STRAIN: COBS CD(SD) B8UEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCPTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL MEIGHT: 24.loOO GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:



STOMACHINFLAMPATION, NOS

(MINIMAL )SPLEENHEMOSIDEROSIS

(MILD )LUNGS

HISTIOCYTOSIS(MINIMAL )

LITTON BIONETICS, INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8704 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04080EROUP I - UNTREATED CONTROL SPECIES: RATSCOSE ANCUNT: 0 PPM STRAIN: COBS CO(SO) BRbEEKS ON STUDY: 13 SIURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL VEIGHT: 273.000 GRAMS FETHRO OF KILL: CARBON Oc"XIo

GROSS OBSERVATIONS:



SPLEENHEMOSIDEROSIS

(MODERATE)LUNGS


~z1

ILITTON BIONETICs INC.FROJECT NC. 10I134-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8705 SEX: FEMALE PATHOLIGY REFERENCE NO. 81-04081GROUP 1 - UNTREATED CONTROL SPECIES: RATSCOSE AMOUNT: 0 PPM STRAIN: COBS CD(SD) BRLEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAG )CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERPINAL WEIGHT: 239.000 GRAMS METHOD OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:




(MINIMAL)SPLEEN

HEMOSIDEROSIS(MODERATE)

LUNGSHISTIOCYTOSIS

(MINIMAL)BLOOD VESSEL


NEPHROPATHY(MINIMAL)


THYPOIDSECTION ANDOCR TISSUE NOT PRESENT

LITTON BIONETICS INC.FROJECT NC. 1073-15 INOIVIDUAL ANIMAL EVALUATrIN

ANIMAL NO. 8706 SEX: FEMALE PATHOLOGY REFERENCE N'. 81-04092GROUP 1 - UNTREATEC CONTROL S0 CIES: RATSCOSE AMOUNT: C PPM STRAIN: COBS Co(So) BP6EEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCPTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 249.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:



LIVERMULTIFOCAL

LYMPHOCYTIC INFLAMMATORY INFILTRATE(MINIMAL)

SPLEENHENOSIDERCSIS

(MILD IKIDNEYS

FOCALMINERALIZATION

(MINIMAL )


LEFT EYESECTION ANDIOR TISSUE NOT PRESENT

LITTON BIONETICS# INC.PROJECT NO. 10754-15 INDIVIDUAL AN:RAL EVALUATTON

ANIMAL NC. ST07 SEX: FEMALE PATHILOGY REFERENCE N'. 81-04083GROUP 1 - UNTREATED CONTROL SPeCIVS: RATSCOSE AMIUNT: 0 PPM STAAIN: COBS CD(SD) SOhEEKS ON STUDY: 13 SfURCE: CHARL-S RIV

r R (PCRTAt-E)CATE OF DEATH: 01-22-81 TYPE or DEATH: TERMINAL KILLTERMINAL WEIGHT: 234.O03 GRAMS METHOD OF KILL: CAPBON D(CXIOE

GROSS OBSERVATIONS:



STOMACHINFLAMNATTON9 NOS

(MILD )SPLEEN

HE4OSIOEROS IS(MILO )

LUNGSBLOOD VESSEL

LYMPHOCYTIC INFLAMWATORY INFILTRA'E

LITTON BIONET CS, INC.FROJECT NO. 1074-1- INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8708 SEX* FEMALE Pk'HOLOGY REFERENCE NO. 81-04CS46ROUP I - UNTREATEO CONTROL SozCIrS: RATSCOS _ AMOUNT: 0 PPM STRAIN: COeS CO(SO) BR

kE.KS ON STUDY: 13 SOURC:: CHARLES RIV'G (PCRTA'E)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLIERMINAL WEIGHT: 248.000 GRAMS METHOD Of KILL: CARBCJ DIOXIDE

GROSS OBSERVATIONS:




(MILD )SPLEEN

HE4MOSIOEROSI S(MILD I

LUNGSBLOOD VESSEL

LYMPHOCYTIC INFLAMMATORY INFILTRATF

L

rX

k_ J

r LjTTON qZONETICS. IN*.

FLOJECT NO. JMC 4-1. IthDIVIOUAL ANIMAL EVALUATION

ANIMAL NO. 3709 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04085GROUP 1 - UNTREATED CONTROL SPECIESt RATSCOSE AMOUNT: 0 PPM STRAIN: COSS CO(SOf RM

hEEKS Oh STUDY: 13 SOURCE: CHARLES RIVEP (PCRTArnrCATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMIN&L KILLIERMINAL WEIGHT: 245.030 GRAMS METHOO OF KILL: CARBON OTOXIOE

GROSS OBSERVATIONS:


VICROSCGPIC FINDINGS:

STOMACHINFLAMMATIONt NOS

(MILD )

LITTON SIONETICSI INC.FROJECT NO. 10t 15 INDIVIOUAL ANIMAL EVALUATION

ANIMAL NO. 8710 SEX: FEMALE PATHOLOGY REFRENCE NO. 81-04C86GROUP 1 - UNTREATED CONTROL SPECl'_S= OATSCOSE AMOUNT: 0 PPM STRAIN: COBS CO(SD) PR6EEKS ON STUDY: 13 SOURCE: CHARLES RIVER IPCRTAGE)CAT_ OF DEATH: 01-21-81 TYPE OF DEATH: TERINAL KILLTERMINAL WEIGHT- 256.000 GRAMS METHOD OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:




(MILD I

TISSUE(S? mIssrsNG OR SEVERELY AUTOLYZEO:

MAMMARY GLANDSECTION ANOtOR TISSUE NOT PRESENT

24-

LITTON B! ONETICSi,*INC,FRO~JECT NO* 107I3 15 INOIVIDUAL ANIMAL EVALUATIONANIMiAL Nnl. 8711 SEX: FEMALE PATHO)LOGY REFERENCE NO. 81-0408iGROUP 1 - UNTREATED CONTROL SPrCIES: RATSCOSE AMCUNT: 0 PPo STRAIN: COSS C0(So BQWEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTA95)CATE OF DEATH: 01-21-81 TYPE OF DEATH4: TERMINAL KILLTERMINAL WEIGHT: 210.000 GRAMS METHOO OF KILL: CARBON DIOXIDr-

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYTEETH CUT


STOMACHINFLAPMATION, NOS

SPLEEN(MINIMAL )

HENOSIDERCSIS(MILD )

LUNGSBLOOD VESSEL


INFLAMMATION, FOCAL(MINIMAL)

KIONEYfPELVI SINFLAMMATION, CHRONIC

(MINIM4AL )

LITTON BIONETICS IC.FRJCO. 107 135 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8712 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-014C88GRaUP 1 - UNTREATED CONTROL SPECIES: RATSCOSE AMOUNT: 0 PPM STRAIN: CORS COCSO) BR6EEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGE)CAT-- OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 216.000 GRAMS FETHOD OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYTEETH CUT


STOMACHINFLAMMATION,9 NOS

(MILDSPLEEN

HE MO SIDE ROS IS(MILD

RECTUS FEMORIS MUSCLEDEGENERATION, NOS

(MINIMALLUNGS

BLCOD VESSELLYMPMfICYTIC INFLAMMATORY INFILTRATE

KIDNEYSKIDNEY/CORTEX

CYST, NOS

LITTON FION-TICS, INC.PROJECT NO* 10734-15 INOIVIDUAL ANIMAL EVALUATION

ANIMAL NO* 8713 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-040R9GROUP 1 - UNTREATED CONTROL SPECIES: RATSCOSE AMOUNT: 0 PPM STRAIN: CCBS CD(SD) BRUEEKS ON STUDY: 13 SOURCE: CHARLES RIV-R (PCRTAGe')CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERM!NAL KILLTERMINAL WEIGHT: 234.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




(MILD )

LITTON BIOETICS, INC.FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NC. 8714 SEX: FEMALE PATHOLOGY REFERENCE NC. 81-04.90GROUP 1 - UNTREATED CONTROL SPECIES: RATSCOSE AMOUNT: 0 PPM STqA!N: COBS CD(SD) BPbEEKS ON STUDY: 13 SOURCS: CHARLES RIVER (PO0TAGE)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 237.000 GRAMS METHOD OF KILL: CARBON oIOXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSY

TEETH CUT



(MILDLUNGS


• Z6

.. . F m . . .' - . . . . . . - " :. . . . . . . -. . . . . . .. .. . . .. . . ' . :. . . ."

LITTON RIONETICS9 INC.PROJECT %0. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NC. 8715 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOW DOSE SPE.CIFS: QATSCOS- AMOUNT: 330 PPM STRAIN: COBS CD(SO) RPUEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTA--)CATE OF DEATH: 01-23-81 TYPE 0F DEATH: TERMINAL KILLTERMINAL WEIGHT: 243.000 GRAMS METHOD OF KILL: CARRON DIOXIOE

GROSS OBSERVATIONS:

ANIMAL APPEARED NORMAL GPTSSLY



LITTON BIONETICsv INC.FROJECT NO. 10734-15 INDIVIDUAL AN:MAL EVALUATION

ANIMAL NO. 8716 SEX: FEMALE PATHOLOGY REFERENCE N3. 0- 0CROUP 2 - LOW COSE SPECIES: RATSCOSE AMCUNT: 30C PPM STRAIN: COS CO(SO) RQ'E~KS ON STUDY: 13 SOURCE: CHARLES RIVER (DOPTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL iEIGHT: 24:3.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




ooEN


ANIMAL NO. 8717 SEX: FEMALE PATHOLOGY REFERENCE NC. 0- 0GROUP 2 - LOW DOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CD(SO) BRWEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCPTAG--)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 260.000 GRAMS METHOD OF KILL: CARBON DIOXID-E

GROSS OBSERVATIONS:




LiTTON 8IONETICSj INC.F OJECT NO. 107 415 INDIV13UAL ANIMAL EVALUATION

ANIMAL NO. 8718 SEX: FEMALE PATHILOGY REFERENCE NO. 0- cGR')UP 2 - LOW DOSE SPTCTES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CD(SD) BR6TEKS ON STUOY: 13 SOURCE: CHARLES RIVER (PCRTAr-E)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 283.000 GRAMS METHOD OF KILL: CARBON' Dif!XT07

GROSS OBSERVATIONS:




LITTON BIONETICS, INC*FRD.JECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANT14AL NO. 8719 SEX: FEMALE PATHOLOGY REFERENCE NO. C- 0GROUP 2 - LOU DOSE SP7CIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COSS CO(SO) SOWEEKS ON STUDY: 13 S'nURCE: CHARLES RIVER (PCRTAG--)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERM!NAL KILLIERMINAL WEIGHT: 241,030 GRA"S PMETHOD OF KILL: CAPBON OI41XIDL

GROSS OBSERVATIONS:


MICROSCaPrC FIPEDINGS:


LITTON BIONETICS, INC.PROJECT NO* 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8720 SEX: FEMALE PATHOLOGY REFrRENCE NO. a- 0GROUP 2 - LOU DOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CDtSD) RRWEEKS ON STUOY: 13 SO'JRCF CHARLES RIVER (PCOTAGE)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMITNAL KILLTERMINAL WEIGHT: 240,000 GRAMS METHOD OF KILL: CARaO'i orexr!OE

GROSS OBSERVATIONS:



MICROSCOPIC EVALUAT!ON NOT REQUIRED

LITTON BIONETICS3 INC.FROJECT NO. 1074-15 INIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8721 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOW DOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CO(SD) RnUEEKS ON STUOY: 13 SOURC: CHARLES RIVER (ODTAG)CATE OF DEATH: 01-23-81 TYPE IF DEATH: TERMINAL KILLTERMINAL WEIGHT: 253.000 GRAMS MzETHOD OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:

ANIMAL APPEARED NCRMAL GROSSLY



LITTON HIONETICS, INC.FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8722 SEX: FEMALE PATHOLOGY REFERENCE N. 0- 0GROUP 2 - LOW DOSE SPECI-S: RATSCOSE A4GUNT: 300 PPM STRAINt C0BS CD(SO) POWEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGE)RATE OF DEATH: 01-22-81 TYPE OF DEATH: TER,"TNAL KILLTERMINAL WEIGHT: 208.000 GRAMS METHOD OF KILL: CARBONJ DIOXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NlCROPSY

RIGHT EAR ENLARGED, EARTAG SITE SWOLLEN

1ICROSCOPIC FINDINGS:

MICROSCOPIC EVALUATION NOT REQU!Qr-

230

I

LITTON BIONETICS9 INC.PROJECT NO- 10734-15 INDTVIoUAL ANIMAL EVALUATION

ANIMAL NGC 8723 SEX: FEMALE PA T H)LOGY REFERENCE N0. C- 0CROUP 2 - LOW DOSE SPECIES: PAlSCOSE AmOUNT: 300 PPM STRAIN: COSS CO(SO) B8UEEKS 0 STUOYT 13 SOUqCE: CHARLES P!VrR (PCRTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TEPINAL KILLTERMINAL UEIGHT: 259.000 GRAMS METHOD OF KILL: CAPBON OICXID!

GROSS OBSERVATIONS:

UTERUS BOTH HORNS FILLED VITH CLEAR FLUID rPICROSCOPIC FINDINGS:


LITTON SIONETICS9 INC.FROJECT NO* 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8724 SEX: FEMALE PATHOLOGY REFERENCF NCo. 0- 0GROUP 2 - LOU DOSE SPECIES: RATSCOSE AMCUNT: 300 PPM STRAIN: COBS CD(SD) OR6EtKS ON STUOY: 13 SOURCe: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL UEIGHT: 220.000 GRAMS M7THOD OF KILL: CARBON DIOXIDE

GROSS OESERVATIONS:



MICROSCOPIC EVALUATION NCT REQUIQED

231

LITTON 9IONETICS, INC.PROJECT NO. 1075-1t INDIVIDUAL ANIMAL EVALUATION

ANIMAL N3, 8725 SEX: FEMALE PATHOLOGY PEFERENCE NO, 0- 0GROUP 2 - LOW DOSE SP!ECIES: OATSDOSE AMOUNT: 30C PP" STRAIN: COBS CDOSD) RQ

6EEKS ON STUDY: 13 S"IURCE: CHARLES PIVER (PCrTAE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TER1INAL KILLTERMINAL .WEIGHT: 241.000 GRAMS METHOD OF KILL: CARBO'l DICXID:-

GROSS OBSERVATIONS:

SKINFLAKY ON TAIL



LITTON BIONETICS, INC*PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUAT!ON

ANIMAL NO. 8726 SEX: FEMALE PATHOLOGY REFERENCE NO* 0- 0GROUP 2 - LOU DOSE SPECI'ES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS Cl(SD) B6EKS ON STUDY: 13 SOUPCE: CHARLE S RIVER (POPTAS:)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERM14NL KILLTERMINAL WEIGHT: 297.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:LUNG5

NUMEROUS PINPOINT UHITE FOCI ON ALL LOBES



~2;3Z

LITTON SIONETICSv INC,PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUA T ION

ANIMAL NOD 8721 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOW COSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: CORS CO(St BRbE:'KS ON STUDY: 13 SOURCE: CHARLES RIVER (PCOTASE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TFRMINAL KILLTERMINAL WEIGHT: 223.000 GRAMS METHOD OF KILL: CARBON DICXI'E

GROSS OBSERVATIONS:





ANIMAL NO. 8728 SEX: FEMALE PATHOLOGY REFERENCE NO* 0- 0GROUP 2 - LOW DOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CD(S) BP6E-KS ON STUDY: 13 SOURCE: CHARLES DIVFR (PCRTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL MEIGHT: 277.000 GRAMS METHOO OF KILL: CARBr4 DIOXIDE

GROSS OBSERVATIONS:

SKINSKIN FLAKY ON TAIL



233

LITTON SIONETICS. INCoFROJECT NO. 107# -15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO* 8729 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- 0GROUP 2 - LOW DOSE SPrCIES: RATSCOSE AMOUNT: 300 PPM ST4AIN: CORS CD(SO) OR

UESON STUDY: 13 SOURCE: CHARLES RIVER (PCPTAGEICATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL XILLTERMINAL WEIGH4T: 2eg.OaO GRAMS METH30 OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




LITTON BIONETICS9 INC.FROJECT NO* 10734-15 IhOIVIOUAL ANIMAL EVALUATION

ANIMAL NO. 8730 SEX: FEMALE PATHOLOGY REFE-RENCE NO. 0- 0GROUP 2 - LOU DOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COSS CO(SD) BR6EEKS ON STUDY: 13 SIURCE: CHARL!:S RIVER (OCRTAGEtCATE OF DEATH: 01-21-8i TYPE OF DEATH: TSRMINAL KILLTERMINAL WEIGHT: 335e000 GRAMS METMOD OF KILL: CARBON DI'oxIDE

GROSS OBSERVATIONS:



mICROSCOPIC EVALUATION NOT REQUIRED

234


ANIMAL NO* 8T31 SEX: FEMALE PATHOLOGY REFERENCE NO* 0- 0GROUP 2 - LOW DOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CO(SD) BRUEEKS ON STUDY: 13 SOURCE: CHARLSS PTVER (PCRrAGE)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMTNAL KILLTERMINAL WEIGHT: 218.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:


PICROSCOPIC FINOIKGS:


LITTON BroNcTrcs. Ic.FROJECT NO. 107,-s I4DIVIDUAL ANIMAL EVALUATICN

ANIMAL NO. 8732 SEX: FEMALE PATHOLOGY REFERENCE No. 0- aGROUP 2 - LOW DOSE SPECIES: RATSLOSE AMOUNT: 300 PPM STRAIN: COBS CO(SO' ORWC -KS ON STU3Y: 13 SOUJRCE : CHARLES RIVER (P CRTA3EICATE OF OEATH: 01-21-81 TYPE OF OEATH: TERMINAL KILLTERMINAL WEIGHT: 281.000 GRAMS METHOD OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:




23S

' ITTN PIONETICS, INC.OROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO- 8733 SEX: FEMALE PATHOLOGY REFERENCE N. 0- cGROUP 2 - LOW DCSE SPZC!ES: RATSCOSE AMOUNT: 300 PPM STRAIN: COBS CD(SD BPWEEKS ON STUDY: 13 SOURCE: CHARLES RIVcq (PCPTAGE)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 245.000 GRAMS METHOI OF KILL: CARBON DOTXIDE

GROSS OBSERVATIONS:

LUNGSPINPOINT WHITE FOCI CN ALL LOBES



LITTON BIONETICS, INC.FROJCT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8734 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- aCROUP 2 - LOW DOSE SPECIES: RATSCOSE AMOUNT: 300 PPM STRAIN: COSS CO(SD) BPrEEKS ON STUIY: 13 SOURCE: CHARLES RIVER (PORTAGE)CAT- OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERINAL WEIGHT: 266.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




236

LITTON BIONETICS9 INC*FROJECT NC. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8735 SEX: FEMALE PATHOLOGY REFERENC: Nis. 0- aCROU 3 M~pUP OSESPECIES: RATS

GOUP AMUT -o PPIU MDS STRAIN: COSS CD(SD) OR.IorS AON TU: 10PPSORE: CHARLES RIVER (PORTAGE)

CATE OF DEATH: 01-23-81 TYPE OF DEATH: TVRM1NAL KILLTERMINAL WEIGHT: 243 .030 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




LITTON SIONETICS, INC.FROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO* 8736 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- 0GROUP 3 - mEoIum DOSE SPECIES: RATS-COSE AMCUNT: looc PPM STRAIN: COSS CD(Sc) RUEEKS ON STUDY: 13 SO)URCE: CH4ARLES RIVrp (PCPTA'VE-)CATE OF DEATH: 01-23-81 TYPE OF DEATH: TE-RM'NAL KILLTERMINAL WEIGHIT. 250.000 GRAMS MrTHOO OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:


PICROSCOPIC FINDINGs:


LITTON BIONETICS9 INC*FROJECT NO* 10734-15 INDIVIDUAL ANTMAL EVALUATION

ANIMAL NO. 8737 SEX: FEMALE PATHOLOGY REFERENCE Nf . 0- CGROUP 3 - MEDIUM DOSE SPECIES: RATSCOSE AMOUNT: 1000 PPM STRAIN: COPS CD(SD) BPsE7KS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)EATE OF CEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 281.000 GRAMS METHOD OF KILL: CARBON DI'XIDE

GROSS OBSERVATIONS:

EARSTHICKENED




ANIMAL NO. 8738 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- 0GROUP 3 - MEOIUP DOSE SPECIES: RATSCOSE AMCUNT: 1000 PPM STRAIN: COBS CO(SO) RRWEEKS ON STU)Y: 13 SIURCE: CHARLES R!VER (PCRTASE)EATE OF DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 292.000 GRAMS MSTHOO OF KILL: CARBON DICXIc

GROSS OBSERVATIONS:



MICPOSCOPIC EVALUATION NOT REQUIRED

238

LITTON PIONETICS9 INC.FROJECT NO. IG734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL N~s 8739 SEX: FEMALE PATHOLO3GY REFERENCE 40. 0- 0GROUP 3 - MEDIUMA DOSE SPECIES: RATSCOSr AMOUNT: 160i PPM STRAIN: COS CD(SOi 86EEKS ON STUOY: 13 SOURCE: CHARLES RIVER (PCRTA7E)LATE OF DEATH: 01-23-8I TYPE OF GEATH: TIERMINAL KILLTERMINAL WEIGHT: 210*030 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NECROPSYHAIR LOSS, FORELIMBS

RIGHT EYE"AIR LOSS AROUND AND RED CRUST ON CORNEA



L.TT:% PI)NrTIC7* INC.

AN'M-'L Q. 741 S X: FrvAL: Pr40L-,ly RL5-R N-. c-

6I-:Ki *?. STU:Y: 13 SJ~C:CHAz Dlvrz cprvp--,

GEN.AL AP-'YE - AT *jl:CoflD Y

pICROSC~pjC rFIr:gG.;:

MC)S:P: :VALUATI'- NJCT REOUIRE-

239


ANIMA, Nl 8741 SEX: FEMALE PATHOLOGY PEFERENCE NO, 0- 0GROUP 3 - MEDIUM DOSE SPECIES: RATSCOSE AMCUNT: 1000 PPM STRAIN: COBS CD(SD) BP6EEKS ON STUDY: 13 SIURCE: CHARLES RIVEP (PCRTAG')CATE OF DEATH: 01-22-81 TYPE OF DEATH: TTRMINAL KILLTERMINAL WEIGHT: 257.000 GRAMS METHOD OF KILL: CARBON DrIOD

CROSS OBSERVATIONS:

ANIMAL APPEARED N4ORMAL CROSSLY



LITTON RIONETICS, INC.PROJECT 40o 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8742 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- 0GROUP 3 - MEDIUM DOSE SPECIES: RATSCOS!: AMOUNT: 1003 PPM STRAIN: COBS CD(SD 00U7EKS ON STUDY: 13 SIURCE: CHARLE'S RIVER (PCRTAr7E)RATE OF DEATH: 01-22-81 TYPE OF DEATH:- TERMINAL KILLTERMI4AL WEIGHT: 251.000 GRAMS METHOD OF-KILL: CARBON DIrcxIDE

GROSS OBSERVATIONS:

ANIMAL APPEARED NORFAL GROSSLY

PMICROSCOPIC FINDINGS:


240

LITTON OICNETICS9 INC.FROJECT NO. 1O734-1! INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. a743 SEX: FEALE PATHOLOGY PEFERENCS NC 0- CGROUP 3 - MEDIUM DOSgE SCECIES: RATSCOSE ANCUNT: 1000 PP7J STRAIN: COBS CD(SD) OR6EEKS ON STUDY: 13 SOURCE: CHARLES RIVc.R (PORTAGE)CATE OF CEATH: 01-22-81 TYPE OF DEATH: TERMINAL K!LLTERMINAL WEIGHT: 29T.00 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OeSERVATIONS:




LITTON BIONETICS, INC.FROJECT NO* 10734-15 INDIVIOUAL ANIMAL EVALUATION

ANIMAL NO. 8744 SEX: FEMALE PATHOLOGY PEFFRENCE NO. 0- 0GROUP 3 - MEDIUm DOSE SPECIES: RATSCOSE AMOUNT: 1000 PPM STRAIA: COBS CD(SD) PR6EEKS Oft STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-22-81 TYPE MCF DEATH: TERMINAL KILLTERMINAL WEIGHT: 245*000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




241

LITTON 9IONETICS INC.FROJECT NO. 107 -1c INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8745 SEX: FEMALE PATHOLOGY REFEQENCE NO. a-C-ROUP 3 - MEDIUM DOSE SPECIES: RATSCOSE AMOUNT: i001 PPM STRAIN: CTRS CD(sD) RpEEKS ON STUDY: 13 SOURCE: CHARLES R!V-R (PCRTArE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TFR!NAL KILLTERMINAL WEIGHT: 219.000 GRAMS METHO9 OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:


MICROSCCPIC FINDINGS:

MICROSCOPIC EVALUATION NCT REQUIREC

LITTON BIONETICS, INC.PROJECT NC. 10734-15 INOIVIDUAL ANTMAL EVALUATION

ANIMAL NO. 8746 SEX: FEMALE PATHOLOGY REFERENCE No. 0- 0GROUP 3 - MEDIUM DOSE SPECIES: RATSCOSE AMGUNT: lOOC PPM STRAIN: COBS CD(SO) B8hEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 274.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




242

LITTON RIONETICS, INC.PRO3JECT NO. 10734- 15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8747 SEX: FEMALE PATHOLOGY REFERENCE N9* 0- aGROUP 3 - MEDIUM DOSE SPECIES: RATSCOSE AMCUNT: 1000 PP" STRAIN: CCBS CO(SD) BQWEEKS ON STUDY: 13 SOUR E: CHARLES PIVER (PCPTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERUINAL KILLTERMINAL WEIGHT: 234.000 GRAMS METHOO OF KILL: CARBON OIOXIOD-

GROSS OBSERVATIONS:



MICROSCOPIC EVALUATIoN NCT REQUIRED

LITTON SIONETICS9 INC*PROJECT NO. 10734-15 INDIV-IUAL ANIMAL EVALUATION

ANIMAL NO. 8748 SEX: FEMALE PATHOLOGY PEFERENCE NO. 0- aGROUP 3 - MEDIUM OISE SPECIES: RATSCOSE AMGUNT: 10G0 PPM STRAIN: COBS CD(SO) BPWEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATE OF DEATH: 01-21-81 TYPE V~F DEATH: TR~r1NAL KILLTERMINAL WEIGHT: 270.000 GRAMS MET4O0 OF KILL: CARBON DICYIDE

GROSS OBSERVATIONS:




COMMENTS:

MASS ABOVE NOSE ANC FLAKY* SCABBY TAIL NOT APPARENT AT NECPOSPT

2~43

LITTON SIONETICSi,!INC.FROJECT NO. 10734is. INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8749 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- cGROUP 3 - MEDIUM DOSE SPrCIrS: RATSCOSE AMOUNT: 1001 PPM STRAIN: COSBS CD(SO) BPWEEKS ON STUDY: 13 SOU RCE: CHARLES RIVER (PCPTAGE-)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TTRpqrNAL KILLTERMINAL WEIGHT: 214.000 GRAMS METHOD OF KILL: CAR BON DI X IDE

GROSS OBSERVATIONS:




LITTON PIONETICS, INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8750 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- 0GROUP 3 - MEDIUM DOS!E SPECIES: RATSCOSE AMOUNT: 1003 PPM STRAIN: COBS CD(SO) BRWEEKS ON STUDY: 13 SCURCE: CHARLES RIVER (PCRTAGE')CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 257.000 GRAMS METHOD OF KILL: CARBON DICXIDE

GROSS OBSERVATIONS:




244

LITTON PIONETICSv INC.PROJECT NO. 10734-15 INDIVIDUAL A41MAL EVALUATION

ONIMAL NO. 8751 SEX: FEMALE PATHOLIGY REFERENCE NO. 0- 0GROUP 3- MEDIUM OOS-- SP.CIES: RATSCOSE &MCUNT: 1000 PPM STRAIN: COBS CO(cOI BWEEKS ON STUDY: 13 S'IURCE: CHARLrS RIVER (PCPTAGE)EATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 216.000 GRAMS METHOD OF KILL: CARBON. DIOXIDE

GROSS OBSERVATIONS:




LITTON BIONETICS9 INC.FROJECT NO* 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL hO, 8752 SEX: FEMALE PATHOLOGY REFERENCE NO. 0- 0GROUP 3 - MEDIUM Dosr SPECIES: RATSEOSE AMCUNT: 1003 PPM STRAIN: CCBS Co(SO)6EEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE)CATE OF OEATH: 01-21-81 TYPE OF DEATH: TER'4NAL KILLTERMINAL WEIGHT: 266.000 GRAMS METHOO OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




245 i

LITTON BIONETICS9 INC.FROJECT NO. 10734-15 tsoivrOUAL ANIMAL EVALUArION

ANINAL hl. 8753 SEX: FEMALE PATHOLOGY REFERENCE: N-'.* a-GROUP 3 - MEDIUM DOSE SPECIES: RATSCOSE AMOUNT: 1003 PP" STRAIN: C06S COC D) 8PWEEKS ON STUDY: 13 SOURCE: CHARLES RrV:*P (POPTAG:)CATE OF DEATH: 01-21-81 TYPE IF DEATH: TERi4INAL KILL-TfERMINAL WEIGHT: 288.OG0 GPAMS METHOD OF KILL: CARBON OICXIDE

GROSS OBSERVATIONS:




LITTON rIONETICS, I&:.FROJECT NC, 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL %O. 87'5k SEX: FEMALE PATHOLOGY REFERENCE NO* 0- 0GROUP 3 - MED)IUM DOSE SPr'CITS: RATSCOSE AMCUNT: 1000 ppm STRAI'N; COBS CO(SO) BRUErKS ON4 STUDY: 13 SOURCE: CHARLES RIV:R (PCRTAGE)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TER;INAL KILLIERMINAL WEIGHT: 242.0C0 GRAMS M:ETHOD OF KILL: CARBON OIOXI E

GROSS OBSERVATIONS:

GENERAL APPEARANCE AT NrCROPSYRIrHT EAR TAG INFECTION

PICROSCOPIC FINOINGS:

MICPOSCOPIC EVALUATrON NCT REQUIRED

246

LITTON 0IONETICS, INC*PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NC. 8755 SEX: FEMALE PATHOLOGY REFERENCIE NO. el-C4111GROUP 4 - HIGH DOSE SPECIES: RATSCOSE AMOUNT: 300C PPM STRAIN: COBS CO(SD) 0QWEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGS)CATE Of DEATH: 01-23-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 203.000 GRAM4S METHOD OF KILL: CARBON DIVXID

GROSS OBSERVATIONS:

OVARIESPLACED IN JAR OF FCRMULA FOR 13 MINUTES, FOUND, OVARIES LOST

BETWEEN COLLECTION A40 WEIGHING,



SPLEEN ML

HEMATOPOIESIS CEXTRAMECULLARY)(MODERATE)

LUNG SBLOOD VESSEL

LYM4PHOCYTIC INFLA14PATORY INFILTRATEOVARIES

TISSUE EXAMINED, NO LESIONS FOUND

LITTON BIONETICS9 INC.PROJECT NO. 10734-1! INDIVIDUAL ANIMAL EVALUATION

ANIMAL ND. 8756 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04112GROUP 4 - HIGH DOSE SPrCItS: RATSCOSE AMCUNT: 3000 PPM STRAIN: COSS CD(SD) opisErKS ON STUDY: 13 SCURCE: CHARLES RIVER (PCRTAGE)CATE OF DEATH: 01-23-81 TYPE OF CFATH: TERMINAL KILL7ERMINAL WEIGHT: 272s000 GRAMS METHOO OF KILL: CARBON DIOXI3M

GROSS OBSERVATIONS:



SPLEEkNEMATOPDIESIS CEXTRAMEDULLARY)

(MODERATE)LEFT EYE

PERIORSIXTAL REGIONINFLA"MATION, NOS

(MODERATE)KIDNEYS


CORPUS UTERIDILATATION, NOS

247

LITTON BIONETICSICPROJECT NO. 1O7i4l~l,! INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO, 8757 SEX: FEMALE: PATHOLI GY REFERENCE Nn. 81-04113GROUP 4 - HIGH DOSESPCE:RTCOSE AMOUNT: 3000 PPM4 SPRCINS: RASS D )R

6EES O STDY: 13S'IURCE: CHARLES RIVEP (PCOTAqv)CATE OF DEATH: 011-23-81 TYPE OF DEATH: TERMINAL KILL-TERMINAL WEIGHT: 257.000 GRAMS FETHOD OF-KILL: CARS044 DIOXIDE

GROSS OBSERVATIONS:




SPLEEN(MLHEMATOPOIESIS (EXTRAMEOULLARY)

(MODERlATE)

LITTON BIONETICS, INC.PROJECT NO. 1073t-15 INDIVIDUAL ANIM4AL EVALUATION

ANIMAL NO. 8758 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04114CROUP 4 - HIGH WiSE SPECIES: RATSCOSE AMOUNT: 300C PPM STRAIN: COBS CO(SD) OR6EEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PORTAGE$CATE OF DEATH: 01-23-81 TYPE nW DEATH:- TERMIN-AL KILLTERMINAL WEIGHT: 216*OCO GRAMS PETHOD OF KILL: CARBCN OICYIDE

GROSS OBSERVATIONS:




(MILOSPLEEN

HERArOPOIESIS (EXTRAMEDULLARY)(MODERATE)

LUNGSBLCOD VESSEL

LY4PHOCYTIC INFLAMPATORY INFILTRATE

OITON BIUP4ETTCS ITNC. IeNVIVIOUAL ANIMAL EVALUATIONF~ECT NO. 107 41

ANIMAL NO* 8759 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04115GROUP 4 - HIGH OOSE sp:rcIES: RATSCOSE AMCIT: 330a PPM STRAP4: cons com) v~kEEKS ON STUDY: 13 SOURCE: CHARLCS RIVEP (PORTAGE$CATE OF DEATH: 01-23-81 TYPE OF DEATH. TERMINAL KILLTERMINAL MEIGHT: 229.000 6RAMS PET&IOO OF XILL: CARBON DIOXIDE

GROSS OBSERVArIOIS.

ANIMAL APPEAREO NORMAL GROSSLY

PICROSCOPIC F114DINGS:

STOMACHINFLAMMATION, NO0S

(MILDPITUITARY

CYST# NOSS PL EEN

HEMATOPOIESIS (EXTRAMEDULLARY)(MILD )

LUNGSBLOOD VESSEL

LYRPHOCYTIC INFLAMMATORY INFILTRATE

IISSUERS MILAND RSVRLYATLZD

MISSUE(S MGLSND RSVRLYATLZDSECTION AND/OR TISSUS NT PRES~

FROJEC8O. 107 4 -15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. B760 SEX: FEMAILE PATHOLOGY PEFERENCC NO* 81-0*116GROUP 4 - HIGH DOSE SPECIES: RATSrOSE AMOUNT: 3000 PPM STRAIN: COBS CD(SO) RRaEEKS ON STuOY: 13 SIURCC: CHARLCS RIVER (PORTAGE)CATE OF DEATH: 01-23-81 TYPE OF OEATI: TERMINAL KILLTERMINAL WEIGHT: 252.000 SRAMS METHO0D OF KILL: CARBON DIOXIZE

CROSS OBSERVATIONSt

GENERAL APPEARANCE AT NECROPSYRIGH7 EAR THICKENEC

KIDNEYSWHITE GRITTY MATERIAL IN LEFT KIDNEY* OILATE0 PELVIS


STOMACHINFLAP"ATIO4, NOS

SP E ATOPOrESIS (EXTRAME0ULLARY)(MILO

KIDNEYSHVORO~NEPHR OS? S

(MILD )

LITTON BIONETIC ,IC

FROJECT NO. j1oi3- INDIVIDUAL ANIMAL EVALUATIONANIMAL NO. 8761 SEX: FEMALE PATHOLIGY REFERENCE Nr,. 81-04117GROUP 4 - HIGH DOSE ScrCIES: PATSCOSS AMOUNT: 3000 PPM SRAIN: COSS CD(SO) RR6 EEKS ON STUDY: 13 SOURCE : CHARLES RIVER (PCRTAGE)CAT- OF DEATH: 01-23-81 TYPE- 3F DrATH: TERMINAL KILLTERRI 'NAL WEIGHT: 253.0aO GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:



STOMACHINFLAPMATICN, NOS

SPLEEN(MLHEMATOPOIESIS (EXTRAMEDULLARY)

(MODERATE)MESENTERIC LYMPH NODE

INFLAMMATION, GRANULOMATOUS(MODERATE)

LUNGSBLOOD VESSEL

LYMPH3CYTIC INFLAMPATORY INFILTRATE


MAMPARY GLANISECTION AND/OR TISSUE NOT PRESENT

COMMENTS:

SMALL SCABS ON TAIL NOT APPARENT AT NECROPSY

LITTON BIONETICS, .FROJECT NO. 1 0731-59 INDIVIDUAL ANIMAL EVALUAT ION

ANIMAL NO* 8762 SEX: FEMALE PATHOLIGY REFERENCE NO. 81-04118EROUP 4 - HIGH DOSE SPECIES: RATSCOSE AMCUNT.' 3000 PPM STRAIN: COBS CD(sD3 sD6EEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRrA^-E)CATE OF DEATH: 01-22-81 TYPE CF DEATH: TERMINAL KILLTERMINAL WEIGHT: 245e030 GRAPS PETHOD OF KILL: CARBO1 DIOXIDE

GROSS OBSERVATIONS:




(MILDOSPLESN

HEMATOPOIESIS (EXTRA4MEDULLA4Y)(MILDO

LUNGSBLOOD VESSEL

LYMPHOCYTIC INFLAMPATORY INFILTRATEKIDNEYSK;DNEYfPELVIS

MINERALIZATION(MODE:RATE)

LrTTON srONETICS INC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8763 SEX: FEMALE PATHOLOGY REFERENCE Nn. 81-04119GROUP 4 - HIGH DOSE SPrCES: RATSCOSE AMOUNT: 300C PPM STRAIN: COSS CC(-O) 8

=

UEEKS O4 STUDY: 13 SOURCE: CHARLES P!VER (POPTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERMINAL KILLTERMINAL WEIGHT: 238.000 GRAMS METHOD OF KILL: CARgON DICXIOE

GROSS OBSERVATIONS:



STOMACHINFLAMMATION, NOS(MILD )

LUNGSBLOOD VESSEL


LITTON BIONETIC3S INC.PROJECT NO. 10734-15 INDIVTOUAL ANIMAL EVALUATION

ANIMAL NO. 8764 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04120GROUP 4 - HIGH DOSE SPEC17S: RATSCOSE ANCUNT: 3000 PPM STRAIN: COPS CO(SD) HRhEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PCRTAGE)DATE OF DEATH: 01-22-81 TYPE 09 07ATH: TERMINAL KILLT7RPZNAL WEIGHT: 251.000 GRAMS PETHOD OF KILL: CARBON DIOXIDE

GROS% c BSERVATzONS:



STOMACHINFLAMMATION9 NOS

(MILD )LUNGS


251

HTTON PIONETICSS INC.ROJECT NO. 10734-1 5 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8765 SEX: FEMALE PATHOLOGY REFERENCE Nl. 81-04121GROUP 4 - HIGH DOSF SPECIES: RATSLOSE AMOUNT: 3000 PPM STRAIN: COBS CO(SO) BUEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (PrRTAGE)CATE OF DEATH: 01-22-81 TYPE OF DEATH: TERmrNAL KILLTERMINAL WEIGHT: 199.000 GRAMS METHOD OF KILL: CARRON DIOXIDE

EROSS OBSERVATIONS:



STOPACHINFLAMMATION, NOS

(MINIMAL)KIDNEYS

KIDNEYIPELVISINFLAMMATION, NOS

(MODERATE)

LITTON BIONETICS INC.PROJECT NO. 107345-1 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8766 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04122GROUP 4 - 44IGH DOSE SPeCIES: RATSCOSE AMOUNT: 3000 PPM STRAIN: CCBS CD(SO) B6ESKS ON STUDY: 13 SIURCE: CHARLES RIVER (PCPTA'E)CATr OF DEATH: 01-22-81 TYPE OF OrATH: TERMINAL KILLTERMINAL WEIGHT: 262.000 GRAMS PETHOD OF KILL: CARBON DOOXIDr

GROSS OBSERVATIONS:

ANIPAL APPEARED NORMAL GROSSLY



(MILO ILUNGS


tITTON BION.TICS, INC.PROJECT NO* 10?1,#-15 IhO[V[OUAL ANIMAL EVALUATION

ANIMAL NO, 8767 SEX* FEMALS PATH"LOGY REFEqECE ,40. 81-04123sROiP 4 - HIGH O0sE SPeCIES? RATSCOSE AMOUNT: 3000 PPM STIN.: COBS CO(SO) RRUIEEKS ON STUIT: 13 SOURCE: CHARLES PIVER (PORTAGECArE OF DEATH: 01-22-81 TYPE OF DEATH: T-RMJ'NAL KILL7ERMINAL WEISHT: 211.000 GRAMS METHOO OF KILL: CARBON DIOXI E

GROSS OBSERVATIONS:

ANIMAL APPEAREO NORMAL GROSSLY

PICROSCQPIC FINOrNAS:

LIVERLIVER/CENTR ILOBULAR

BASOPHILIC STIPPLING(MILD )

STOOACHIIVOLAMeATZON9 NOS

(MILO )LUNGSBLOOD VESSEL

LYMPHOCYTIC INFLAMMATORY 1NFILTRATE

LITTON BTONETICS INC. ,LPROJECT NDo C 0.iq-$1- Ik]VrUAL CN.IPL EVALUATION

ANIMAL NO. 8768 SEX: FEMALE PA 'rOLOGT REFERENCE NO. 81-04124GROUP 4 - HIGH OOSE SPECrES: RATSCOSE AMOUNT: 3000 PPM STAI4N: COBS CO(SO) 80,EEKS ON STUDY: 13 SOURCE: CHARLES QIVER (PCRTAGEiCATE OF DEATH: 01-21-81 TYPE OF DEATH: TTRMINAL KILLTERMINAL WEZGHT: 209,000 SPAMS MEHOO OF KILL: CARBON 014OXIDE

GROSS OBSERVATIONS:


MICROSCOPIC FINDINGS?

STOMACHINFLAN4ATION, NOS

(MILD )PITUITARY

CYST, NOSSPLEEN

HEMOSZDEROSIS(MILD )

LUNGSBLOOD VESSEL


253

LITTON BIONETICS, INC*FROJECT NO* 10714-15 INDIVIDUAL ANIMAL EVALuATrON

ANIMAL NO* 8769 SEX: FEMALE PA'HOLOGY REFERENCE NO. 81-04125GROUP 4 - HIGH DOSE SPECIES: RATSCOSE AMOUNT: 3000 PP" STRAIN: COBS CO(SO) RPWEEKS ON STUDY: 13 SOURCE: CHARLrS RIVER (PCRTAGE)CATE OF CEATH: 01-21-81 TYPE OF DEATH: TrRMTNAL KILLTERMINAL WEIGHT: 230,000 GRAMS METHOD OF KILL: CARBON OIOXIDE-

CROSS OBSERVATIONS:

THYROIDREDUCED


LIVERINFLAMRATION, FOCAL

(MILDSTOMACH

INFLAMMATIONt NOS(MILO

THYROID TISSUE EXAMINEO, NO LESIONS FOUND

SPLErNHEMOSIDEROSrS

(mrLO )LUNGS


LITTON BIONETICS, INC.ROJECT NO. 10T -i5 IOIVIDUAL ANIMAL EVALUATIOM

ANIPAL NO. 8770 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04126GROUP 4 - HIGH DOSE sPECrES: RATSCOSE AMOUNT: 30G PPM STRAIN: CISS CO(S D) POWEEKS ON STUDY: 13 SIURCE: CHARLES, RIVER (POPTAGE)CAT- OF DEATH: 01-21-81 TYPE OF DEATH: TSRMINAL KILL1ERMINAL WEIGHT: 255.000 GRAM METHOD OF KILL: CARBON oOXIoE

GROSS OBSERVATIONS:


rICROSCOPrC FINDZNGS:


(MILDSPLEEN

HEMOSIOEROSIS(MILO )

LUNGSBLOOD VESSEL


254

LITTON PIONETICS9 INC*FROJECT NO* 1O734-15 INOrVIOUAL ANIMAL EVALUATION

ANIMAL NO. 8771 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04127GROUP 4 - HIGH DOSE SPFCIFS: RATSCOSE AMOUNT: 3D0O PPM STRAIN: COtS CO(SO) RR6EEKS ON STUDY: 13 SOURCE: CHARLES RIVCR (PCPTAGE)

CATE OF DEATH: 01-21-81 TYPE OF DEATH: TERMINAL KILLIERMINAL WEIGHT: 249.000 GRAMS PETHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:


PICROSCOPIC FINDINGS*

HEARTDEGENERATION% NOS

(MINIMALSTOMACH

INFLAMRMATION9 NOS(MILD )

SPLEENHEMOSIOEROS!S

(MILO )

JTTONBO S IC

ON 8Noi-is INDIVIOUJAL ANIMAL EVALUATrON

ROJCT 8INEOtVs

ANIMAL NO. 8772 SEX: FEMALE P4THOLOSY REFERENCE Nl. 81-04128CROUP 4 - HIGH DOSE SPECIES: RATSCOSE AMOUNT: 3000 PPM STRAtN: COBS CO(SO) ORLEEKS ON STUDY: 13 SOURCe: CHARLES RIVER (PORTASE)ATE OF DEATH: 01-21-81 TYPE OF DEATH: TvRMNAL KILLTERMINAL WEIGHT: 236.000 GRAMS PETHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATIONS:




(MILDSPLEENHEMOSIDERCSIS

(MILO )LUNGS

BLOOD VESSELLYMPH3CYTIC INFLAMPATORY INFILTRATE

BLOOD VESSELFOCAL

MINERALIZATION(MILD )

LITTON BIONETICS, lhC.PROJECT NO. 10734-15 INDIVIDUAL ANIMAL EVALUATION

ANIMAL NO. 8773 SEX: FEMALE PATHOLOGY RTFERENCE N. B1-04129GROUP 4 - HIGH DOSE SPECIES: RATSCOSE AMOUNT: 300C PPM STRAI'!: COBS CO(SO) RcU ZKS ON STUDY: 13 SnURCZ: CHARLES RIVZR (PORTASE)CATE OF DEATH: 01-21-81 TYPE OF DEATH: TrRM!NAL KILLTERMINAL WEIGHT: 211.000 GRAMS METHOD OF KILL: CARBON DIOXIDE

GROSS OBSERVATInNS:




(MILO )LUNGSBLOOD VESSEL


FOCALINFLAMMATION, INTERSTITIAL

(MILD )

LITTON BIONETICS 41NC.POJECT NO* 107-15 INDIVIDUAL ANIMAL VALUATICJ

ANI"AL N'., 8174 SEX: FEMALE PATHOLOGY REFERENCE NO. 81-04130C-qOUP 4 - HIGH DOSE SPECIES: RATSrZSE AMCUNT: 3000 PPM STRAIN: COBS CO(SO) BRWIEEKS ON STUDY: 13 SOURCE: CHARLES RIVER (POPTAGe)CATE OF DEATH: 01-21-81 TYPE IF DEATH: TERMINAL KILLTERMINAL WEIGHT: 285.000 GRAMS METHOD OF KILL: CARBO'I DIOXIDE!

GROSS OBSERVATIONS:



STOMACHINFLAMMATICN, NOS

(MODERATE)CECUM


RECTUS F-MORIS MUSCLEMULTIFICAL

DEGENERATION, NOS(MINIMAL )

LUNGSBLOOD VESSEL


KICNEY/PELVISMINERALIZATION

(MILD )KIDNEY/PELVIS

INFLAPMATION, ArUTE(MILD )

25

L*~hL

SECTION H

APPENDIX C

257

DETERMINATION OF

ISOPROPYL METHYLPHOSPHONIC ACID(IMPA) IN'EXPERIMENTAL ANIMALDRINKING WATER FORMULATIONS:

CHEMICAL METHODS AND REULTS

FINAL CHEMISTRY REPORT

SUBMITTED TO:

ENVIRONMENTAL PROTECTION DEPARTMENT

U.S. ARMY MEDICAL BIOENGINEERING

RESEARCH AND DEVELOPMENT LABORATORY

FORT DETRICK, FREDERICK, MD. 21D71

ATTENTION: SGRD-UBG

CONTRACT NO.: DAMD-17- 77-70 03

SUBMITTED BY:

DEPARTMENT OF CHEMISTRYLITTON BIONETICS, INC.

5516 NICHOLSON LANE

KENSINGTON, MD 20795

AUGUST 10, 1981

SPONSOR: Environmental Protection DepartmentU.S. Army Medical Bioengineering Research and DevelopmentLaboratory

MATERIAL: Isopropyl Methylphosphonic Acid (IMPA) Sodium SaltLBI #3427 and 6488

SUBJECT: CHEMISTRY FINAL REPORTDetermination of IMPA in Experimental Animal DrinkingWater FormulationsLBI Project No. 10734

SUMMARY

Chemical analyses of the dosed drinking water prepared for toxicologicaltesting were performed to determine dose concentration and stability ofthe chemical. The analytical method used was that implemented by LittonBionetics, Inc. following a review meeting with representatives of thesponsor; the method involves formation of a volatile derivative usingdiazomethane. The method used is a modification of a method contributedby the Sponsor's research and development laboratory.

The dose verification analyses of the drinking water formulations showedthat for the most part, the formulations were near intended concentrations.Six preparations yielded results lower than the pre-study limit (85% oftarget). The study director was informed in each instance of low assayresults. A copy of the analytical method, and a discussion of the sensi-tivity and accuracy of the method is included.

SPONSOR: Environmental Protection DepartmentU.S. Army Medical Bioengineering Research and DevelopmentLaboratory

MATERIAL: Isopropyl Methylphosphonic Acid (IMPA) Sodium SaltLBI #3427 and 6488

SUBJECT: CHEMISTRY FINAL REPORTDetermination of IMPA in Experimental Animal Drinking WaterFormulationsLBI Project No. 10734

1. OBJECTIVE

The purpose of the chemistry study was to analyze the drinking waterformulations used in toxicological testing. The aqueous preparationwas also tested for stability.

2. EXPERIMENTAL METHODS AND PROCEDURES

A. Methods

Attempts were made to develop a method for the analysis of IMPAadded to animal feed, but these were unsuccessful. This workis summarized in Chemistry attachment A of this section. Theroute of administration was changed from dosed feed to drinkingwater, with the sponsor's approval.

Analyses of the drinking water formulations were performed byacidifying an aliquot of the water and diluting it with methanol.A small volume of the diluted sample was then treated with anether solution of diazomethane to form the methyl ester of IMPA.The volatile ester was then analyzed by gas chromatography usinga flame photometric detector with a phosphorus-specific filter.The internal standard used for the gas chromatography was triethylphosphate. A detailed version of the analytical method is pre-sented in attachment B of this section.

B. Procedures

Homogeneity testing of the formulations was not considerednecessary since solutions, by definition, are homogeneous.

Stability of the chemical in the acidified drinking water atambient temperatures was determined by analysis of single sub-samples of formulations taken from the carboys at time of for-mulation, and again at the end of time use for dosing animals.

The results of these two sets of analyses are listed in Table 1.

260

B. Procedures (Continued)

Dose verification analysis of the drinking water was originallyscheduled for the first three formulations and, if no problemsin formulation were detected, for every third formulation there-after.

However, indications of consistently low formulation were foundfor the second and third sets of preparations, so the ChemistryDepartment advised the Study Director to authorize analysis ofthe fourth formulation (see Table 2). When analysis of themix of 12/02/80 showed consistently low formulation (74.6 to79.0% of intended concentration), it was decided to assay thetwo remaining sets of formulations, so that results for allsix batches would be available. A complete summary of resultsof dose verifications is given in Table 2.

3. RESULTS AND CONCLUSIONS

The analytical data for testing the stability of sodium isopropylmethyl phosphonate in water shows some decrease in the test compoundconcentration at the two higher doses after room temperature storagefor a total of 25 days. Since new formulations were mixed aboutevery 14 days, the stability of the test compound in the drinkingwater appears adequate for the time of use for dosing. Twelve ofthe formulation were within limits of ±15% of intended concentration.However, the IMPA concentration of six of the drinking water formu-lations was below -15% of target.

Some individual assay values may be unreliable because the flamephotometric (phosphorous-specific) detector requires very stableflow rates of hydrogen, air and oxygen for a constant sensitivityto be achieved. The detector was serviced by an authorized instru-ment company service person; other than an insensitivity of thephotomultiplier tube, no problems were found in the operation ofthe device. It was necessary, throughout the study, to operate thedetector at or near its most sensitive setting. This operationintroduced considerable noise into the baseline, which in turncaused variations in the integration of the peak areas for themethyl ester of IMPA ano for the internal standard.

The precision of the method proved to be poorer than first foundwhen the written method (attachment B) was prepared. For this reason,average results of duplicate assays are given in Tables 1 and 2 andanalyses with exceptionally poor precision (which had to be includeddue to limted sample size) are footnoted.

Despite the possibility of some analytical error introduced by base-line noise problems, the overall results given in Table 2 show thatat all dose levels, average concentrations of IMPA found were rearintended levels. For the 300 ppm target level, the grand mean cf

261

the % of target results (n=6) in Table 2 is 87.0%. The correspondingstandard deviation for all formulations is ±7.7%. For the 1000 ppmtarget level, the grand mean (n=6) is 93.0% of target, with a standarddeviation of ±8.7%. For the 3000 ppm formulation, the grand mean(n=6) is 91.9% of target with a standard deviation of ±15.9%. Thismeans experimental animals were dosed at around 90% of the intendedconcentration over the time span of the study.

Summary report prepared by:

Lynn'S. Reed, Ph.D. DateHea , GC/MS Section

Revision of 8/11/81 prepared by:

irry R. Fitzge ld, Ph.D.Head, Analytical Chemistry DateSection

Apprdved y:

,-Frank Boyd-'Ph.B- __. B/Director, Dept. of Chemistry Date

262

SECTION H

APPENDIX C

CHEMISTRY ATTACHMENT A

METHOD DEVELOPMENT FOR THE ANALYSIS OF

IMPA SODIUM SALT IN FEED

263

METHOD DEVELOPMENT FOR THE ANALYSIS OF NaIMPA IN FEED

The experimental design consisted of the following:

1. Extract IMPA from feed2. Separate IMPA from interferences (from feed) by chromatography3. Detect and quantitate IMPA present

Analytical problems were found in all three areas.

1. EXTRACTION

The sodium salt of IMPA was found to dissolve only in water andmethanol (July 1979). Other solvents tried were tetrahydrofuran,acetonitrile, pyridine, dimethylformamide, dimethyl sulfoxide, acetoneand ethyl acetate. Extraction of an acidified aqueous solution ofNaIMPA with n-butanol gave a recovery of 20%.

2. CHROMATOGRAPHY

A. Derivatization

An attempt was made to reproduce the method developed by SRI,involving ion exchange chromatography, lyophilization, andformation of the methyl ester of IMPA. The recovery was 0.4%.Concurrently, several other derivatization methods were tried,including ion pair extraction and alkylation (ref: 1976, Aldri-chemica Acta, 9:35; 1971, Acta Pharm. Sueica, 8:113) solid-liquidphase transfer reaction (ref: Ludurkow, Metal, 1978, AngeroChem. Inst. Ed., 17(1):62; Van Heerden, F.R., et al., 1978,Tetrahedron Letters #7, 661), crown ether catalyzed alkylation(ref: Davis, B., 1977, Anal. Chem., 832). Attempts were madeto form esters using methylating and silylating agents as wellas dibromoacetophenone and pentafluorobenzyl bromide (PFB).Many of these reagents can be used successfully with the freeacid, but the sodium salt is more resistant to esterification.An identifiable product was formed from the crown ether catalyzedreaction of PFB and a solution of NaIMPA. However, this deriva-tive is not stable. IMPA dosed feeds gave method recoveriesranging from 87 to 43% (August 1979) and from 62 to 10% (August1980).

B. Gas Chromatography

Derivatives made with the free acid were found to tail badlyon the original SRI GC column. A more suitable stationaryphase was found by testing several possibilities.

Fp

2 6 4 j

3. DETECTION AND QUANTITATION

A phosphorus specific (flame photometric) detector was used forthe gas chromatographic analysis. There was adequate sensitivityand specificity for the pentafluorobenzyl derivative of IMPA usingthis detector. However, the excess derivatizing agent or its combus-tion products collected on the detector exhaust port - and possiblyon the optical filter as well - causing flow restrictions whichmight alter the response of or damage the detector. The possibilityof developing an HPLC method utilizing ultraviolet detection wasexamined but not considered feasible because of the difficulty ofderivatizing IMPA and because of the high background UV absorbanceusually seen for feed extracts.

After meeting with the sponsors, it was decided that the route ofadministration would be changed from feed to drinking water(August 1980). An adequate method for the analysis of NaIMPA indrinking water was then developed and implemented.

SECTION H

APPENDIX C

CHEMISTRY ATTACHMENT B

METHOD FOR DETERMINATION OF IMPA

SODIUM SALT IN DRINKING WATER

266

DETERMINATION OF IMPA MONOSODIUM SALT IN DRINKING WATER

1. PRINCIPLE

An aliquot of the water sample is acidified and diluted with methanol.A small volume of the dilution is treated with ethereal diazomethaneto form the methyl ester of IMPA, which is analyzed by gas chromato-graphy using a flame photometric detector with a phosphorus filter.Triethyl phosphate serves as the internal standard.

2. REAGENTS

Deionized or distilled water

Potassium hydroxide - reagent grade

Diethyl ether - Burdick and Jackson, grade suitable for pesticideanalysis

N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) - Aldrich Chemical Co.,Cat. No. 12994-1

Isopropyl methylphosphonic acid (IMPA) reference standard - LBI #6362,supplied by U.S. Army Medical Bioengineering Research and DevelopmentLaboratory, (Dr. Clarence W.R. Wade, research chemist).

Triethyl phosphate (TEP) internal standard - Aldrich Chemical Co.,Cat. No. T6,110-7

Methanol - Burdick and Jackson, grade suitable for pesticide analysis

Hydrochloric acid, concentrated - reagent grade

3. MATERIALS AND SUPPLIES

Analytical balance accurate to 0.01 mg

Top-loading balance accurate to 0.01 g

Erlenmeyer flask, 125 ml size, with cork stopper

Graduated cylinder: 100 ml size

Volumetric flasks: 10, 25, 50, 100, 200 and 1000 ml sizes

Volumetric pipettes: 1, 4, 5, 10, 15, 20, 25 and 50 ml sizes

Graduated pipettes: 5 and 10 ml sizes

Micropipettor, 200 pl size - SMI (Scientific Manufacturing Industries)or equivalent (positive displacement).

267

3. MATERIALS AND SUPPLIES (Continued)

Hewlett-Packard Model 5840 gas chromatograph, microprocessor controlled,with flame photometric detector (phosphorus filter), Model 7672Aautosampler and associated data system.

Chromatographic column - 1.8 m x 2 mm I.D. silanized glass packedwith 10% FFAP on 80/100 mesh Supelcoport.

Autosampler vials, 1 ml capacity (Wheaton Scientific, Millville, NJ08332, Stock No. 223682) with Teflon-lined aluminum caps (StockNo. 224211).

Crimper for above vials (Wheaton Stock No. 224301).

4. PREPARATION OF ETHEREAL DIAZOMETHANE

Weigh, to nearest 0.01 g, 4 g of KOH into 125 ml Erlenmeyer flask.Dissolve in 10 ml water.

Cool in ice/water bath and add 60 ml diethyl ether.

Weigh, to nearest 0.01 g, 0.75 g of MNNG. Add in small incrementsto Erlenmeyer flask in ice bath, swirling frequently and cappingwith cork stopper between additions.

After all MNNG has been added, stopper and let stand one hour.When not being used, store in freezer. (It is not necessary toseparate the ethereal solution of diazomethane from the lower aqueouslayer).

NOTE: Successful generation of diazomethane imparts an intenseyellow color to the ether phase. If this color is notobserved, the MNNG may have decomposed.

CAUTION: Diazomethane can undergo explosive polymerization. Neveruse old, etched glassware or glass-stoppered glasswarefor its preparation.

5. PREPARATION OF CALIBRATION STANDARDS (Unless otherwise noted, use

volumetric glassware for all preparations)

a. Stock Internal Standard Solution (Target: 6 mg/ml)

Weigh, to nearest 0.01 g, 0.6 g of TEP into 100 ml flask.Dissolve in and dilute to volume with methanol.

268

5. PREPARATION OF CALIBRATION STANDARDS (Continued)

b. Working Internal Standard Solution (IS) (Target: 6 ppm)

Dilute 1.0 ml of stock solution, above, to 1000 ml with methanol.

c. Stock IMPA Standard Solution (Target: 2 mg/ml)

Weigh, to nearest 0.01 mg, 100 mg of IMPA reference standardinto 50 ml flask. Dissolve in and dilute to volume with methanol.

d. Intermediate IMPA Standard Sulution (Target: 100 ppm)

Dilute 5.0 ml of stock solution, above, to 100 ml with methanol.

e. Stock Methylated IMPA Standard Solution (Target: 40 ppm as IMPA)

Transfer 4.0 ml intermediate solution, above, to 10.0 ml flask.Add 3 ml (graduated pipette) ethereal diazomethane. (Morediazomethane solution may be added as necessary to obtain apersistent yellow color). Swirl to mix, stopper and let standone hour. Dilute to volume with methanol.

f. Working Methylated IMPA Standard Solution (Target: 2 ppm)

Dilute 5.0 ml stock methylated solution, above, to 100 ml withmethanol.

g. Chromatographic Calibration Standards

Combine aliquots of working methylated IMPA and working ISstandard solutions indicated below to 100 ml with methanol:

Working Methylated Working IS Final IMPAIMPA Standard Standard Target Concentration

ml ml ppm

5.0 20.0 0.10010.0 20.0 0.20025.0 20.0 0.50050.0 20.0 1.00

6. PREPARATION OF SAMPLES FOR ANALYSIS

NOTE: (Although the test material is the monosodium salt ofIMPA, the concentrations of dosage formulations areexpressed in terms of the free acid).

269

6. PREPARATION OF SAMPLES FOR ANALYSIS (Continued)

(Unless noted otherwise, use volumetric glassware for all prepara-tions).

Transfer aliquots of sample solutions as indicated below to 200 mlflasks. Add 4 ml (graduated pipette) of concentrated HC and diluteto volume with methanol. Mix well. (Since dilution of water withmethanol results in contraction of volume, it will be necessary toadd additional methanol to the mark; mix well again.)

Dose or Aliquot Taken Target IMPATreatment Level For Dilution Concentration in Dilution

ppm ml ppm

0 50.0 0300 50.0 75

1000 15.0 753000 5.0 75

With micropipettor, transfer duplicate 200 p1 aliquots of each dilutedsample to 25.0 ml flasks. Add 4 ml (graduated pipette) ethereal diazo-methane. (More diazomethane solution may be added as necessary toobtain a persistent yellow color.) Swirl to mix, stopper and letstand one hour. Add 5.0 ml internal standard solution and dilute tovolume with methanol.

7. GAS CHROMATOGRAPHIC ANALYSIS

Transfer aliquots of calibration standards and methylated samplesto autosampler vials and cap closed. Load autosampler, beginningand ending run with a complete set of chromatographic standards.Since "ghost" peaks have been observed in solvent injections followingstandard injections, it would be advisable to place two or threesolvent "samples" in the autosampler after the first set of standardsto clear the system of residual IMPA and TEP.

Suggested chromatographic parameters: (May be modified to obtainapproximate retention times given below.)

TEMP 1 125 0CTIME 1 0.00 minINJ TEMP 225 0CAUX TEMP 245 0C

CHT SPD 0.50 cm/minZERO 10.0ATTN 2t 2AUX SGNL A*SLP SENS 0.05

*Column connected from injector B to FPO.

2

7. GAS CHROMATOGRAPHIC ANALYSIS (Continued)

AREA REJ 10000000FLOW B 30.0 ml/min

1.00 AREA REJ 01.50 IFN 97.00 STOP

Hydrogen : 200 rotometer units (24 psig)Air 50 rotometer units (24 psig)Oxygen : 20 rotometer units (24 psig)

Check chromatographic performance with a 3 p1 injection of the higheststandard (1.00 ppm). Retention times of IMPA and the internal standardshould be approximately 2.7 and 5.0 minutes, respectively.

Enter the options as indicated below. Set the "Change Run" at 0and the "Stop Run" to correspond to the number of analyses, includingthe number of solvent injections to clear the "ghost" peaks.

OPTN #1. ID: 10734-152. DATE: (Current)

TIME: (Current)3. INJ/BTL, STROKE: 1,2 (approximately 3 pI injection)4. EXT ALARM AMT5. DIL FACTOR: 1.0000 E+ 06. RF UNC PKS: 0.0000 E+ 07. RPRT UNC PEAKS (NO)8. TRANSMIT RPRT (NO)9. SUPPRESS RPRT (NO)

Press "Start Run" to initiate analysis.

8. CALCULATIONS

For standards and samples alike, calculate the area ratio, IMPA/TEP,using the area responses obtained by gas chromatographic analysis.

Using linear regression analysis, establish a calibration curverelating ppm of the GLC standards to their respective area ratios.The analytical run will be considered acceptable based on a correla-tion coefficient of at least 0.99. (Criteria for slope and interceptcannot be established because they depend upon the relative amountsof IMPA and T' in the standards.)

271

8. CALCULATIONS (Continued)

Based on the calibration curve and the area ratios of the samples,calculate the concentration (ppm) of IMPA in the methylated samplesolutions analyzed.

Calculate the concentration of IMPA in the dosage formulation fromthis formula:

ppm IMPA C x V-A xVV3 V1

where C concentration of IMPA in methylatedsolution analyzed (ppm)

V, = aliquot of dosage formulation takenfor analysis (ml).

V2 = volume to which V, was diluted(200 ml).

V3 = volume of dilution taken formethylation (0.200 ml)

V4 = final volume of methylated solution(25.0 ml)

Since V2 , V3 and V4 are constants, the expression reduces to:

Cppm IMPA in dosage formulation = x 25,000

Replication Check (Precision):

For any given concentration of IMPA, calculate:

% agreement = (Result for subsample A - Result for subsample B) x 100

Mean result of two subsamples

: 10% or smaller*

*Note added at end of study. In some isolated analyses, agreement be-tween duplicates was worse than 10%. Results were included in thisreport if no more sample was available for reanalysis.

Calcuiate percent of target concentration for each dosage formulation:

% of target Mean assay value of IMPA x 100%Target concentration of IMPA

272

Method developed by: Anproved for use by:

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