Document gDRVp9gBG4vM5144b9n18K7LL

FORDUPONTUSEONLY AR226-2962 Du Pont HLO 432-91 StudyTitle Skin Sens itization Test with in Guinea Pigs StudyCompletedOn July 31, 1991 ContractingLaboratory Pharmakon Research International, Inc . P .O . Box 313 Waverly, Pennsylvania 18471 Author Susan Armondi for Haskell E . I . du Pont de Nemours and Company Laboratory for Toxicology and Industrial Elkton Road, P . 0 . Box 50 Newark, Delaware 19714 Medicine Study Monitor Scott E . Loveless MedicalResearchNo . Laboratory ProjectID Haskell Laboratory Report No . 432-91 Company Sanitized Does not contain TSCA CBI Page 1 of 60 Du Pont PHARMAKON RESEARCH INTERNATIONAL, INC. WAVERLY, PENNSYLVANIA 18471 COMPLIANCE STATEMENT HLO 432-91 PHONE (717) 586.2411 FAX (717) 586-3450 This study was conducted in compliance with the Principles of Good Laboratory Practices (GLP) as promulgated by the following regulatory agencies: U.S. Food and Drug Administration, as stated in the Federal Register, 21 CFR Part 58, Friday, September 4, 1987. U.S. Environmental Protection Agency as stated in the Federal Register, 40 CFR, Part 792, Thursday August 17, 1989. U.S. Environmental Protection Agency as stated in the Federal Register, 40 CFR, Part 160, Thursday, August 17, 1989. Organization for Economic Co-operation and Development Guidelines for Testing Chemicals (OECD), ISBN 92-64 12221-4, adopted by the council at its 535th meeting on 12th May, 1981. Study No.: PH 424-DU-001-91 "To the best of my knowledge, this study was conducted in accordance with applicable Good Laboratory Practice regulations;. there were no deviations from these regulations that impacted on study conclusions." M{/}0/^C . iMf^cwL Study Director \\{iU^.WI Date^Jy7 Material Tested: Medical Research No. Haskell No.; Haskell Test Code No. Pharmakon Study No.: Physical Form: Purity: Composition: GENERAL INFORMATION Test Material Du Pont HLO 432-91 Synonyms: Other Codes; Contaminants: Stability: Material Tested; Purity: Synonyms: Other Codes: In the absence of visible evidence to the contrary, the test material was assumed to be stable under the conditions of administration. Positive Control Material N-Phenyl-1,4-Phenylenediamine 98% 1,4-Benzenediamine p-phenylenediamine PPD Aldrich Chemical Co., Lot ft05012MK rAldrich Chemical Co., Catalog #24139-3 3 - - e'osssany Sanfflzed. Does net co^a^i TSCA CBt GENERAL INFORMATION (CONT.) Du Font HLO 432-91 CAS Registry No.: 101-54-2 - Stability; In the absence of visible evidence to the contrary, the test material was assumed to be stable under the conditions of administration. Sponsor: Du Font Chemicals E. I. du Font de Nemours and Company Vilmington, Delaware Materials Submitted By: E. J. Greenwood Du Font Chemicals E. I. du Font de Nemdui-s arid Company Jackson Laboratory Deepwater, New Jersey Pharmakon Notebook No.: Study Initiated - Completed: 2/22/91 - 7/31/91 In-Life Phase Initiated - Completed; 4/29/91 - 6/7/91 There are 60 pages in this report. Distribution: K. D. Dastur (1) V. J. Brock (1) P. G. Gilby (1) E. J. Greenwood (1) N. C. Chromey/S. E. Loveless (1) ^'^"I'pai'i'y' .^sliiii'ZSfct .. DU FONT CENTRAL RESEARCH AND DEVELOPMENT HASKELL LABORATORY FOR TOXICOLOGY AND INDUSTRIAL MEDICINE Du Font HLO 432-91 July 31, 1991 E. J. Greenwood Du Pont Chemicals Jackson Laboratory Deepwater, New Jersey Skin Sensitization with' n Guinea Pigs Attached is a report from Pharmakon Research International, Inc. (PH 424-DU-001-91). Included in the report are test methods, results and conclusions. [----------^ J----1----^----jaatn intradermal induction concentration of 1% and chT&CU'neennegeeccoonncceennTTrraa1tions of 50% and 5% in distilled water, was tested on the clipped intact skin of 5 male and 5 female guinea pigs. Appropriate positive, negative, and vehicle control groups were used during this study. During the challenge phase-^iQ^rYlhema was observed in any test animals treated with either 50% or 5^||I^^^HU^ As was expected, a positive sensitization response (slight to severe erythema) was observed in 9 out of 10 positive control animals. In contrast, no dermal responses were observed in any of the 5 negative or 5 vehicle control animals when challenged with the test material, positive control material, and/or vehicle. Under the conditions of this studyy contact hypersensitivity in guinea pigs. yjdid not produce delayed SEL/alw Z-^<. 7-^ Scott E. Loveless, Ph.D. Senior Research Toxicologist ) GwiBanv SanW-'B'i f^w. ir? r^^iin TfiCA CB? DU Font PHARMAKON RESEARCH INTERNATIONAL, INC. WAVERLY, PENNSYLVANIA 18471 HLO 432-91 PHONE (717)586-2411 FAX (717) 586-3450 Skin Sensitization Test with H-18,790 in Guinea Pigs PH 424-DU-001-91 Submitted to E. I. Du Pont De Nemours Newark, Delaware ^Mn)^. lllmoOY^L Susan E. Annondi, LAT Study Director [^uL Datel/ mi ^ (\ I ^^^^/f^^&', /"i Test Facility Management _D__a__t_e//>7^//9^r '.^'^liipcil^' ^c&iiii.il-'^ii.iB i^'Q^s,- i'-" './wii--.*^.. i ^^s-^ ,,'*.?* Du Font HLO 432-91 Skin Sensitization Test with H-18.790 In Guinea Pigs SUMMARY Test Article H-18,790 at an induction concentration of 1.0% (w/v) and challenge concentrations of 50% and 5.0% (w/v) in distilled water was tested on the clipped, intact skin of male and female guinea pigs. p-Phenylenediamine (PPD) as a 35% (w/v) suspension in distilled water was used to demonstrate the ability of the test system to detect a skin sensitizer (positive control group). Distilled water served as a vehicle control. During the primary irritation phase, no dermal irritation was observed in the vehicle control animals. Slight erythema was observed at 24 hours in one animal receiving the test article at the 50% concentration. No erythema was observed at the 5.0% concentration sites. No erythema was observed in the positive control animals receiving PPD at a 35% concentration. During the challenge phase, no to severe erythema was observed in the positive control group treated with p-phenylenediamine at a 35% concentration. No signs of erythema were observed in the test article-treated animals at the 50 or 5.0% concentration sites at 24 or 48 hours. No signs of erythema were observed in the vehicle control animals. No signs of erythema were observed in the negative control animals receiving the test article, vehicle or the positive control articles. Under the conditions of this study, H-18,790 did not produce delayed contact hypersensitivity in guinea pigs at 50 or 5.0% concentrations. Work by: .^enarles BoyaVrskyr^ Technician ur Study Director: W^\(W ^ c ^(T^W^ Susan E. Armondi, LAT Reviewed and Approved for Issue:: i r f\ AM^C - ^rmo^L' Susan E. Armondi, LAT Study Director i) %?tipany Sanitized. Does no'i ccr^afn TSCA CPS Du Font HLO 432-91 QUALITY ASSURANCE UNIT STATEMENT Study No.: PH 424-DU-001-91 Study Director: Susan E. Armondi This study was conducted in compliance with the Good Laboratory Practice Regulations. The Quality Assurance Unit conducted the inspections listed below and reported the results to the study director and to management on the dates indicated. The following inspections were performed: Interval Date In Life Phase April 29. 1991 Reporting Phase July 18. 1991 Date QAU Report Issued: To Study Director July 18, 1991 To Management July 18, 1991 O ^ A ^ J ^ ^ ^ Quality i/ AssAirance ) i^'~,"--' "" ' " r fir" "----""" i ! ^ 4 ^ 3 , Dat^7 ]??/ V ^te^di^ 0^^.;^^^ ^ -'"i...--;., ^ k^'t.^^i *>'?,, Du Font HLO 432-91 INTRODUCTION The purpose of this study was to evaluate the potential of H-18,790 to produce delayed hypersensitivity or allergic reactions when applied to the skin of guinea pigs. Sensitization was defined as a significant score increase at challenge over the response observed after the primary application of the test material to the test guinea pigs, or the response observed in the negative controls. A significant score increase defined as a 2-or-more step increase (e.g., from 0 to 2, from 1 to 3, etc.) has been used at Haskell Laboratory as the criterion in this study protocol to identify compounds that are sensitizers. The sensitivity of the test system to detect chemical sensitizers was evaluated with pphenylenediamine. This study was conducted according to EPA Good Laboratory Practice Regulations as listed on page 2 of this report. Areas of noncompliance are documented in the study records. No deviations existed that significantly affected the validity of the study. MATERIAL AND METHODS A. Animal Husbandry Young adult male and female Duncan Hartley albino guinea pigs were received from BuckberG Lab Animals, Tomkins Cove, New York. The guinea pigs were housed singly in suspended, stainless steel, h" wire-mesh cages. Each guinea pig was assigned a unique identification card on which the animal number, study number, dose level and sex were recorded and affixed to the cage. Guinea pigs were ear tagged. Purina Certified Guinea Pig Chow" #5026 and water were available ad libitum. Guinea,pigs were acclimated and observed for general health during a period of approximately five days. Animal rooms were maintained on a timer-controlled, 12-hour light/12-hour dark cycle. Environmental conditions of the rooms were targeted for a temperature of 22 3C (66- 77F) and relative humidity of 30 - 70%. Any excursions outside these ranges were of small magnitude and/or brief duration and did not adversely affect the validity of the study. B. Protocol A copy of the signed original protocol is attached as Appendix A. A preliminary range-finding test was conducted to estimate the primary irritation potential of the test material. The results of the range-finding study were used to select the exposure concentrations for the main study. The main sensitization study consisted of 3 phases: a primary irritation phase, an induction phase and a challenge phase. During each phase, skin responses wtheere sstucdoyre, d boadcycowrdeiingghtsto wtehree sreycsotermdedprewseeenktleyd. in Table I. During Cwitarw anW?eri. does y.'s'f confab TSCA CPi Du Pont HLO 432-91 The topical range-finding test was conducted on 2 males and 1 female guinea pigs ranging in weight from 300 to 358 grams. Aliquots (approximately 0.05 mL) of 1.0%, 10%, 50% and 100% (w/v) suspensions of the test material in distilled water were applied and lightly rubbed onto four separate test sites on the clipped, intact skin of the back of each animal. The primary irritation phase was conducted in 10 guinea pigs (5 males and 5 females), weighing from 403 to 442 grams, by applying and lightly rubbing in 1 drop (approximately 0.05 mL) of 50% and 5.0% (w/v) suspensions of the test material in distilled water onto two separate sites of the clipped, intact skin of each animal. In addition, 10 positive control guinea pigs (5 males and 5 females), weighing from 401 to 451 grams, were treated by applying and lightly rubbing in 1 drop of a 35% (w/v) suspension of p-phenylenediamine in distilled water onto the shaved, intact left shoulder skin of each animal. Five vehicle control guinea pigs (2 males and 3 females), weighing from 401 to 439 grams, were treated with distilled water in the same manner at one site only. Dermal responses were scored approximately 24 and 48 hours after application of the test material. Two days after the primary dermal irritation phase, the induction phase of the study was initiated using the same 10 test guinea pigs in which primary irritation had been evaluated. Induction consisted of a series of 4 intradermal applications (1 each week) of 0.1 mL of the test material at a 1.0% (w/v) concentration in 0.9% saline. A virgin site on the back of each animal was used for each induction. The same induction procedure was followed for the 10 positive control guinea pigs using 0.1 ml of a 1.0% (w/v) solution of p-phenylenediamine in saline. The vehicle control animals were similarly treated with 0.1 ml saline. Skin responses were evaluated approximately 24 hours after each induction. Two weeks after the last induction treatment, the test guinea pigs were challenged for sensitization by applying and lightly rubbing in 1 drop of 50% and 5.0% (w/v) suspensions of the test material in distilled water onto two separate sites of clipped, intact shoulder skin. The sites used during challenge were the same used on each animal during the primary dermal irritation phase, respectively. The positive control animals were challenged for sensitization by applying and lightly rubbing in 1 drop of a 35% (w/v) suspension of p-phenylenediamine distilled water onto two separate sites of clipped, intact skin. The vehicle control animals were treated at two sites with distilled water, in the same manner. Also, 3 male and 2 female previously untreated guinea pigs, weighing 410 to 438 grams at study initiation, were treated by applying 1 drop each of 50% and 5.0% (w/v) suspensions of the test material in distilled water onto two separate sites of clipped, intact skin and served as negative control animals. In ^CWp&s'iy'^cSiiiiiSc.-.. ^-s-s :i^-;-">"i:.... i^.^-.^.:.!- 10 Du Font HLO 432-91 addition, these animals received 0.05 ml of distilled water and 0.05 ml of p-phenylenediamine at 35%. These animals also served as negative control animals for the vehicle and positive control materials. C. Record Retention All raw data and the final report will be stored in the archives of Pharmakon Research International, Inc., Waverly, PA. RESULTS AND CONCLUSIONS Weekly body weights are presented in Appendix B. A copy of raw data is presented in Appendix C. In the topical range-finding test, no to mild erythema was observed. Based upon the results of the dose-range-finding study, 50% proved to be mildly irritating and as per protocol, the dose concentrations used for the primary- irritation phase were 50% and 5.0%. The dose concentration used for induction was 1% and as per protocol, the dose concentrations chosen for challenge were 50% and 5.0%. During the primary irritation phase, no signs of erythema were observed in the test article-treated animals at the 5.0% concentration site. Slight erythema was observed in one animal at the 50% concentration site at 24 hours. No erythema was observed in the positive control animals. No erythema was observed in the vehicle control animals receiving distilled water. Individual animal data are presented in Table II. III. During the induction phase, no signs of erythema were observed in the test article-treated animals. No to mild erythema with and without edema was observed in the positive-control animals. No signs of erythema were observed in the vehicle control animals. Individual animal data from the induction phase are presented in Table Dermal responses observed during the challenge phase are summarized in the following table (page 12). Individual animal data are presented in Table IV. ACT/RP6 t) Company Sair.iizese. Does "oi uoniam TSCA CBS 11 Du Font HLO 432-91 Summary of Skin Responses: Challenge Phase Responses Positive Control _____PPD_____ 35% 24 hr 48 hr Test Animals H-18.790 50% 24 hr 48 hr 5.0% 24 hr 48 hr No erythema 1/10 1/10 10/10 10/10 10/10 10/10 Slight erythema 2/10 4/10 0/10 0/10 0/10 0/10 Mild erythema 5/10 3/10 0/10 0/10 0/10 0/10 Moderate erythema 1/10 1/10 0/10 0/10 0/10 0/10 Severe erythema with necrosis 1/10 1/10 0/10 0/10 0/10 0/10 Responses Negative Control H-18.790 PPD 50% 5.0% 24 hr 48 hr 24 hr 48 hr 24 hr 48 hr Vehicle Distilled Water 24 hr 48 hr No erythema 5/5 5/5 5/5 5/5 5/5 5/5 5/5 5/5 ^ No erythema Vehicle Control___ 24 hr 48 hr 5/5 5/5 At challenge, H-18,790 did not produce delayed contact hypersensitivity at 50 or 5.0% concentrations. Individual animal data are presented in Table V. 6) 12 Du Font HLO 432-91 TABLE I SCORING SYSTEM USED TO EVALUATE SKIN RESPONSES Score Skin Reaction ' 0 No erythema or Edema 1 Slight erythema (barely perceptible, usually nonconfluent) 2 Mild erythema (well defined, usually confluent) 3 Moderate erythema 4 Severe erythema (beet redness, with or without edema, eschar formations or necrosis) If edema, blanching, necrosis or eschar formation occurred, they were indicated using the following code: ED = Edema N = Necrosis E = Eschar B = Blanching SN == Superficial Necrosis Cemsany Sanitizad. iSoes wi conia?n TRCA CB 13 GUINEA PIG NUMBER 1186 1102 1103 1104 1105 1106 1107 1189 1190 1110 Du Font HLO 432-91 TABU3 II PRIMARY IRRI"CATION P HASF, SKIN RESPONSES OBSERVE D IN TE;T GUINEA 1?IGS FOLLOWING TOPICAL E3<POSURE 'TO H-18,79 0 LEFT FRONT 50% 24 hr 48 hr 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 . RIGHT FRONT 5.0% 24 hr 48 hr 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 14 Du Font HLO 432-91 TABLE II (Cont^d) PRIMARY IRRITATION PHASE SKIN RESPONSES OBSERVED IN VEHICLE CONTROL GUINEA PIGS FOLLOWING TOPICAL EXPOSURE TO DISTILLED WATER GUINEA PIG NUMBER 1111 LEFT FRONT Distilled Water 24 hr______48 hr 1112 1113 1114 1191 Company Sanitized. Deas not contain TSCA CS! 15 Du Font HLO 432-91 TABLE II fCont'd) PRIMARY I]y^ITAT:[ON PITASR SKIN RESPONSES OBSE;RVED I N POS ITIVE C ONTROL GUINEA PIGS FOLLOWING TOPICS\L EXP(3SURE TO p-PIIENYLENEDIAMINE GUINEA PIG NUMBER 1121 1122 1123 1187 1125 1193 1194 1128 1129 1195 LEFT FRONT 35% 24 hr 48 hr 0 0 0 0 0 0 0 0 0 0 0 0 0 0- 0 0 0 0 0 0 -^o'i'4?aKy ^s'&tiiU^.v.'CL. Liiot-s i;--- .;i-i^aii; 's &<-,.i G-j.. 16 Du Pont HLO 432-91 TABLE III TNIAUCTION PHASE SKIN RESPONSES OBSERVED IN TEST GUIN EA PIGS 24 HOURS FOLLOWING INTRADERMAL ADMINISTRATION OF 1.0% H-18.790 GUINEA PIG NUMBER 1186 1102 1103 1104 1105 1106 1107 1189 1190 1110 Week 1 Dose #1 (LEFT) 0 0 0 0 0 0 0 0 0 0 Week 2 Dose #2 (RIGHT) 0 0 0 0 0 0 0 0 0 0 Week 3 Dose #3 (LEFT ') 0 0 0 0 0 0 0 0 0 0 Week 4 Dose # 4 (RIGHT) 0 0 0 0 0 0 0 0 0 0 Ccwtpany Samiizea. Dees no? contain TSCA CB! 17 Du Font HLO 432-91 TABLE III (continued) INDUCTION PHASE SKIN RESPONSES OBSERVED IN TEST GUINEA PIGS 24 HOURS FOLLOWING INTRADERMAL ADMINISTRATION OF 0.9% SALINE GUINEA PIG NUMBER 1111 1112 1113 1114 1191 Week 1 Dose #1 (LEFT) Week 2 Dose #2 (RIGHT) Week3 Dose #3 (LEFT) Week 4 Dose #4 (RIGHT) ) 18 Du Font HLO 432-91 TABL]S III fcontinu Rrt) HEDUCTION PHASE SKIN RE;SPONSES OBSER;VED IN TEST GIJINEA PIGS 24 HOURS FOLLOWING INTRADERMAL A.DMINISTRATION OF D-PHENYLEN EDIAMINI^ GUINEA PIG NUMBER Week 1 Dose #1 (LEFT) Week 2 Dose #2 (RIGHT) Week 3 Dose #3 (LEFT) Week 4 Dose #4 (RIGHT) 1121 0 2ED 0 1 1122 0 2ED 0 1 1123 0 1 1 1 1187 1 1 1 0 1125 1 1 1ED 1 1193 0 1 0 0 l-^ 1194 0 1ED 1ED 1 1128 0 1 1 0 1129 1 1 2ED 1 1195 0 1 0 1 ED = edema Company Sanitized. Does not contain TSCA CB( 19 GUINEA PIG NUMBER 1186 1102 1103 1104 1105 1106 1107 1189 1190 1110 Du Font HLO 432-91 TABLE IV CHALLENGE PHASE SKIN RESPONSES OBSERVED IN TEST GUINEA PIGS FOLLOWING TOPICAL APPLICATION OF H-18,790 LEFT FRONT 50% 24 hr 48 hr 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 RIGHT FRONT 5.0% 24 hr 48 hr 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 ^ .f^-'.'.^WS, Ol^'"" ;->;,(. -.-:W, 20 Du Font HLO 432-91 TABLE IV (cont'd) CHALLENGE PHASE SKIN RESPONSES OBSERVED IN VEHICLE CONTROL GUINEA PIGS FOLLOWING TOPICAL APPLICATION OF DISTILLED WATER GUINEA PIG NUMBER 1111 1112 1113 1114 1191 LEFT FRONT Distilled water 24 hr 48 hr Sompany Sanitized. Does noi contain TSCA CBt 21 GUINEA PIG NUMBER 1116 1117 1118 1119 1192 1 Du Font HLO 432-91 TABLE IV (Cont'd) CHATi LENG E PHASE SKIN RESPONSES OBS ERVE D IN NEGATIVE CONTROL GUINEA PIGS FOLLOWING T OPICAL APPLICATION OF H-18,790 DISTILLED WATER AND D-PHENYL13NEDIAMINE - LEFT FRONT H-18,790 50% 24 hr 48 hr 0000 0000 00 RIGHT FRONT H-18 ,790 5. 0% 24 hr 48 hr 0 0 0 0 0 0 0 0 0 0 LEFT REAR Di stilled Water 100% 24 hr 48 hr 0 0 0 0 0 0 0 0 0 0 RIGHT REAR PPD 35% 24 hr 48 hr 0000 0000 00 22 Du Font HLO 432-91 TABLE IV fCont'd) CHAT, T,ENGE PHASE SKIN RESPONSES OBSERVED IN POSITIVE CONTROL GUINEA PIGS FOLLOWING TOPICAL APPLICATION OF p -PHENYLENEDIAMINE GUINEA PIG NUMBER 1121 1122 1123 1187 1125 1193 1194 1128 1129 1195 LEFT FRONT 35% 24 hr 48 hr 2 2 2 2 4 3 2 1 2 1 3 4 1 1 2 2 1 1 - 0 0 ^ y Company Sanitized. Does no? conta'n TSCA CBi 23 Du Font HLO 432-91 APPENDIX A PROTOCOL 24 Du Font HLO 432-91 PHARMAKON RESEARCH INTERNATIONAL. WAVERLY. PENNSYLVANIA 18471 Protocol - 424D INC. Skin Sensitization Test in Guinea Pigs PHONE (7171586-2411 FAX (717)586.3450 Sponsor: E.I. duPont de Nemours Haskell Laboratory for Toxicology and Industrial Medicine Newark, DE 19714 Testing Facility: Phannakon Research International, Inc. Waverly, Pennsylvania 18471 ' . Test Facility S.O.P. No.: Study Ho.: Purpose of the Study; PH-424 To be assigned upon study initiation. (it+HZ-^-OU -<y < -c] | This study is designed to evaluate the dermal sensitization potential of the test material in guinea pigs. Ownership of the Study; Study Monitor: Study Director: g.A.U. Responsible Personnel: Dates of Performance: The sponsor owns the study. All raw data, analysis, ^'^^ill '5^ and reports are the property of the sponsor. . JLo-<^ Pk.D. DaJ.mres E. -Ma^ek, Ph.D., E.I. duPont de Nemours Susan Armondi, LAT, Phannakon Research International, Inc. Leslie J. Pinnell, M.S. Study will begin within one month of the receipt of the test chemical and authorized protocol. Good Laboratory Practices Statement: This study will be conducted following the U.S. Environmental Protection Agency Toxic Substances Control Act Good Laboratory Practice Standards (40 CFR Part 160) or Good Laboratory Practice Standards (40 CE-R Part 792) . IACUC Statement: This protocol has been reviewed by the Institutional Animal Care and Use Committee (IACUC) and complies with acceptable standard animal welfare and humane care. Tentative Date of Submission of Draft Report: Within three weeks following the completion of the in-life phase of the study. 080489 \) Company Sam-Sized. Dues not contain TSCA CB? 25 Protocol - 424D 2 Skin Sensitization Test in Guinea Pigs Du Font HLO 432-91 Records Maintained: Raw Data: Record Retention; Analytical Chemistry: Species: Strain: Supplier; . (Source): Sex; ^L Weight at Initiation: All raw data, final reports, documentation and protocol will be maintained in the Phannakon Central Files. Amendments to protocol Feed Lot Number Body weights, initial, weekly and final Compound preparation Volume administration Observed signs Observed skin reactivity Maintained in a Standard Pharmakon' Notebook All raw data and completed notebooks. Analysis and stability of the test article and test article/carrier mixture is the responsibility of the sponsor. If requested by the sponsor, Phannakon Research International, Inc. will, through its subcontractor, conduct appropriate analytical analysis and will indicate the additional cost involved following receipt and evaluation of the appropriate analytical method. In the case where a satisfactory method is not provided, Pharmakon Research International, Inc. or its subcontractor, at additional cost to the sponsor, will develop appropriate methods. If the sponsor elects to analyze the test article/carrier mixtures, Pharmakon Research International, Inc. will collect the sample at appropriate designated intervals during the study and transmit the samples to the sponsor for analysis. TfcST SYSTEM Guinea pig Hartley BuckberG Lab Animals, Tonkins Cove, New York or from any other U.S.D.A acceptable source. Male and Female (the sex used will be documented in the study records and final report). Young Adult 400 - 650 grams 080489 GTiV-^Sn'i'fl?., 26 Du Font HLO 432-91 Protocol - 424D 3 Skin Sensitization Test in Guinea Pigs No. on Study: Method and Justification for Randomization: Acclimation Period: System of Identification: Research Facility Registration: Animal Rooms: Housing; Sanitization: Food: Dose-Range-finding: 3 Vehicle control: 5 (at the discretion of the Study Monitor, this group may be excluded from the study.) Negative control: 5 (if re-challenge is necessary, a second set of 5 naive, age-matched negative control animals will be used.) Test article: 10 Positive control: 10 (at the discretion of the Study Director after consultation with the Study Monitor, this group may be excluded from the study.) Treatment groups will be housed by vertical cage positioning. Any animals showing evidence of clinical signs during acclimation will be eliminated from randomization. Randomization is carried out by use of a computer generated random number table. Before treatment commences, all guinea pigs will be weighed, ranked according to body weight and assigned to treatment groups using a table of random numbers so that each treatment group will have a similar distribution according to body weight. Guinea pigs If beyond the extremes of the body weight range will not be assigned to treatment groups. there is a statistically significant difference in mean body weight between any two groups of the same sex after allocation to the treatment groups, substitution from spare animals will be used to correct the situation. Minimum; five (5) days Cages marked with an animal group number, sex and treatment group. Guinea pigs are ear tagged. HUSBANDRY U.S.D.A. Registration No. 23-107 under the Animal Welfare Act 74: SC 2131 et seq. Separate isolation by test system. Light cycle - 12 hours light, 12 hours dark. Temperature/Humidity - Maintained at a temperature of 22 "C 3-0 and a humidity of 30 to 70%. Guinea pigs are housed individually in stainless steel '2" wire nesh cages. Size in accordance with the "Guide for the Care and Use of Laboratory Animals" of the Institute of Laboratory Animal Resources, National Research Council. Waste material will be removed daily. Cages and feeders will be sanitized every two weeks. p Purina Certified Guinea Pig Chow S5026 or acceptable substitute will be provided ad libitum. Feeders are 080489 "^ Company Sanitized. Does not contain TSCA CS;' 27 Protocol - 424D 4 Skin Sensitization Test in Guinea Pigs Food Analysis: Water: Hater Analysis: designed to reduce soiling, bridging, and scattering. The potential effects of dietary contaminants have been considered, and based on the manufacturer's data, contaminant levels are believed to be within acceptable limits. There are no contaminants expected to be present in the dietary material known to be capable of interfering with the purpose or conduct of the study. Availability - fresh tap water, ad libitum. Water is monitored for contaminants at periodic intervals according to Standard Operating Procedure Rationale for Test System; Rationale for Dose Selection: Justification for Test System; Compound Preparation; Absorption of the Test Material: METHODS This study is intended to provide information on the health hazards likely to arise from exposure to the test material by the dermal route; skin contact is a possible worker and consumer exposure route. Based upon the. results of the dose-range finding study and/or at the sponsor's request. According to EPA Health Effect Test Guidelines EPA 560/6-82-001. The guinea pig has historically been used in the evaluation of chemicals for dermal sensitization. Liquids Liquids will be applied as received except when specific instructions request dilution or mixing. The use of a vehicle for.'dilution will be described in the study records and final report. Solids The test material will be pulverized if appropriate. The test material may be moistened with distilled water or a suitable vehicle to ensure good contact with the skin. When a vehicle is used, the influence of the vehicle on penetration of the skin by the test substance will be considered. Semi-Solids Extremely viscous liquids nay be heated to approximately 50C to dosing, with the approval of the Study Monitor, The procedure used will be documented in the study records and included in the final report. For the purpose of this study, all of the test material applied to the test sites is assumed to be absorbed by the test system. All calculations and the evaluation of effects will be based on the applied dose. 080489 28 Du Font HLO 432-91 Protocol - 424D 5 Skin Sensitization Test in Guinea Pigs Vehicle: The choice of vehicle varies, depending on the nature of the test substance. Commonly used vehicles are distilled water, saline, ethanol:distilled water (80:20, v/v), dimethyl phthalate, acetone:dimethyl phthalate (1:9, v/v) or propylene glycol. The vehicle used will be documented in the study records and the final report. Volume Administration: Approximately 0.05 ml/dose Route of Administration: The test and control materials will be applied topically and/or injected intradennally, to the clipped, intact shoulder skin. Rationale for Route of Adminis tration: Skin contact is a possible worker and consumer exposure route. No. and Description Dose-range-finding: 3 of Animals per Vehicle control: 5 (at the discretion of the Study Dose Group: Director, this group nay be excluded from the study). Negative control: 5 (if re-challenge is necessary, a second set of 5 naive, age-matched negative control animals will be used). D Test Article: 10 Positive control: 10 (at the discretion of the Study Director, after consultation with the Study Monitor, Method of Study this group may be excluded from the study). Performance: Dose-range-finding Study Prior to initiation of the main study, a dose-range-finding study will be performed. In order to evaluate the irritation potential of the test material, 3 animals will be subjected to preliminary studies as follows: Animals will be closely clipped over the back and sides. The test material concentrations will be based on the solubility of the test compound (solids will be tested at the maximum soluble concentration and at 2 or 3 lower concentrations). Liquids will be tested at 100% (as received or as prepared per the Sponsor's instructions) and at 2 or 3 lower concentrations. Each guinea pig may be dosed with up to 4 different concentrations at different sites (l concentration/ site). In some cases where the solid is not soluble in a common solvent it will be tested as received by topical application. One drop (approximately 0.05 ml) of each concentration will be applied directly on the appropriate test site and rubbed gently with the end of the dosing syringe to cover an area of approximately 25 mm in diameter. 080489 company Sanitised. Does not cor.tain 1'SC'A C'y, 29 Protocol - 424D Skin Sensitization Test in Guinea Pigs Du Font HLO 432-91 Observations for signs of dermal irritation will be recorded at 24 and 48 hours after application. At each observation, all treated sites will be scored according to Table I. ^ ^ -UA l<;t,ltar sl'i'ilA- +o non-i'n<Ti+>'^i C<ft~ 1*4- *>ThA< "~- Primary Irritation Phase For the test guinea pigs there will be 2 sites per animal in the area between the shoulder and the middle of the back. One site w^ll receive tha.-^j.-ghoot ^''"^rrl^<<q.qnn<<rraattiioonn--JJ'hi<h is jj^ir'-^'"^1v^^^'''^r'f"e^n"'^-m nT^^iyiyy''"Tj.^r~rta~.'-LaLxn'j 'as determined from the dose-range-finding study. The second site will receive one-tenth of that concentration. Each site will receive 3. drop (approximately 0.05 ml) of the appropriate concentration, rubbed gently into the clipped skin with the end of the dosing syringe to cover an area approximately 25 nan in diameter. The vehicle control guinea pigs, if used, will be treated at 1 site as above using the vehicle only. The positive control guinea pigs will be treated at 1 site with the positive control. The negative control guinea pig will remain untreated until the challenge phase. Observations for signs of dermal irritation will be recorded at 24. and 48 hours after application. Sensitization Phase (Induction) The first induction application will be made after the 48-hour dermal observation is made for the primary irritation phase. If the test material is soluble in appropriate vehicle, e.g., 0.9% saline or dimethyl phthlate, induction will be performed by intradermal injection. If the test material is insoluble in the commonly used vehicles a topical induction procedure will be used. The hair at the appropriate application site will be clipped on the day prior to each application or, if a topical induction procedure is employed, at least weekly during induction. After clipping, the area to be treated may be delineated for the convenience of and at the discretion of the study director or the experimenter. The hair will be clipped as needed during the study for evaluation of the dermal irritation. A total of 4 intradermal injections will be made, 1 per week at 1 week intervals. A virgin site will be used for each injection. A 0.1 ml aliquot of a 1% solution of the test material in an acceptable solvent (e.g., 0.9% saline, dimethyl phthalate, etc.) will be administered by intradermal injection to the clipped sacral area of each test guinea pig. If distilled water is used as a vehicle in the primary irritation phase, 0.9% saline will be used for the intradermal injections. The vehicle control guinea pigs will be 080489 fc) 30 Du Font HLO 432-91 Protocol - 424D 7 Skin Sensitization Test in Guinea Pigs Method Validation/ Positive Control: Evaluation of Dermal Response: treated as above using the vehicle only. The positive control guinea pigs will be treated as above using the positive control material. Topical Administration A total of 9 applications will be made to the sacral area, 3 applications per week for 3 weeks. One drop (approximately 0.05 ml) of a minimally irritating concentration of the test material in the acceptable solvent (e.g., dimethyl phthalate, 0.9% saline, etc. will be applied directly to the skin and then gently rubbed in with the end of the syringe, to cover an area approximately 25 mm in diameter on each test guinea pig. The vehicle control guinea pigs will receive similar treatments using the vehicle only. The positive control quinea pigs will receive similar treatments using the positive control article. Challenge Phase Approximately 14 days after the last induction treatment, the challenge treatment will be administered. The test material will be administered in the same manner as in the primary irritation phase, treating with fche same concentrations at the same sites. Following the same procedure the negative control animals will be treated with the same concentrations of the test material, and nay be treated with the vehicle and positive control material (up to 4 separate test sites). The vehicle control animals will be treated with the vehicle only. The positive control animals will receive the positive control article. If the test results are conclusive and the test material has either clearly caused allergic sensitization or is not a sensitizer, the test is terminated and the guinea pigs are euthanized. If the reactions are less than conclusive, a second challenge (rechallenge) may be performed approximately 1 week after the initial challenge using the.same test guinea pigs and additional naive control guinea pigs. The vehicle and positive control animals may not be rechallenged. The procedures and test system will be evaluated with a known dermal sensitizer to confirm the validity of the test. Evaluation of the test system with a known dermal sensitizer may not be conducted concurrently with every study, but testing with a known dermal sensitizer will be performed periodically. Dermal evaluations will be made between 23-25 hours and/or 47-49 hours following treatment, but the data will be reported as the 24- or 48-hour evaluations. 080489 Company Saniiizea. Does not contain TSCA CBi 31 Du Font HLO 432-91 Protocol - 424D 8 Skin Sensitization Test in Guinea Pigs Interpretation of Results: Sensitization Intervals a. Primary Irritation Test - Dermal evaluations will be made approximately 24 and 48 hours after the test material application. b. Induction - Dermal evaluations will be made approximately 24 hours after each induction. treatment. c. Challenge and Rechallenge (if required) - Dermal evaluations will be made approximately 24 and 48 hours after the test material applications. All test sites will be graded on a, scale of 0 to 4 as follows: TABLE I Score Skin.Reaction 0 Mo erythema or edema 1 Slight erythema (barely perceptible, usually nonconfluent) 2 Mild erythema (well defined, usually confluent) 3 Moderate erythema 4 Severe erythema (beet redness, with or without edema, eschar formation or necrosis) If edema, blanching, necrosis, superficial necrosis or eschar formation occur, they will also be indicated using the following code; ED = Edema N = Necrosis E c Eschar B c Blanching SH " Superficial Necrosis Sensitization is defined as a 2-or-more step increase (e.g., from 0 to 2, from 1 to 3, etc.) in dermal irritation scores in the test animals over those observed in the negative controls at challenge. When the test and negative control animals have similar dermal irritation scores, the test material will not be considered a sensitizer. The rating of a sensitizer is as follows: Ratio of Responsers Degree of Sensitization 1 to 2/10 Weak Sensitizer 3 to 4/10 Mild Sensitizer 080489 32 Du Font HLO 432-91 Protocol - 424D 9 'Skin Sensitization Test in Guinea Pigs Necropsy; Euthanasia: Statistical Analysis; Rechallenge; Positive Control Article: Supplier (Source)! Special Handling Instructions; Analysis of Purity; How Supplied; Rationale for Positive Control Article: Ratio of Responsers Degree of Sensitization 5 to 6/10 Moderate Senitizer 7 and above Strong Senitizer This rating scale is not fixed rigidly. Scientific evaluation of the data and judgement will be used to decide whether a material will be classified as a weak, mild, etc. sensitizer. Pathological evaluations of test animals will not ordinarily be performed. Exceptions will be noted in the study records and final report- Animals that surive the test period will be euthanized according to the Testing Facility's Euthanasia SOP and incinerated. Statistical analyses of data will not be performed, unless requested by the sponsor. The sponsor will be notified of animals that are considered sensitized. Verbal instructions for rechallenge will be given by the Sponsor, followed by written confirmation. All test article animals, along with an additional group of four naive guinea pigs, can be rechallenged six (6) days after primary challenge, but not before. The negative and positive control groups will not be rechallenged. POSITIVE CONTROL ARTICLE M-Phenyl-1,4 phenylenediamine (PPD) Aldrich Chemical Company, Milwaukee, Wisconsin Standard precautions The purity is the responsibility of the manufacturer. 100 gm bottle As per sponsor's request. ATXPL4 ) 080489 Sompany SanRfzed. Dc.es not cor;'am TSCA CBF 33 Protocol - 424D1 10 Skin Sensitization Test in Guinea Pigs Du Porit HLO 432-91 Test Article (Name or Code): Chemical Abstract No. or Code No.; Analysis of Purity/Stability; Carrier Mixtures; Analysis of the purity and stability of the test article is the responsibility of the sponsor. Analysis for stability, uniformity.ana correctness of concentration of the test article in the carrier is the responsibility of the sponsor. __ Return Test Article Carrier Mixtures to the Sponsor y Dispose of Test Article Carrier Mixtures Person to whom carrier mixtures should be sent; ^________________________________________ Shipping Instructions; AMENDMENTS ^ y/<!.c;Ji Li^AV^^. rr^^^V^t APPROVAL OF PROTOCOL Date ___^ 2-1 Nl_______ Study Monitor Date ___9-'^L^\______ ^"^ Director ATXPL4 A^^^ c/-g^^>_____ /lU<3Q/VOG Lh\(r^^C^ . ^^SiVf ^y^::., ,,, ^ ^ ci,^;.,fn , s; 34 Du Porit HLO 432-91 ii APPENDIX A Test Article Information Identification: Test Article (Name or Code); Lot or Sample Ho: H-in^ Physical Descript Purity:_____| | Expiration Datey Density/Specific~Gravity:__ Solubility (check one): Water Ethanol_______] Other (please specify)^ Chemical Classification: Flammable DMSO Corrosive Other II Storage Information: Material Storage (check one): Room Temperature < ; Refrigerator Freezer ; Other (specify) III Handling Information; Known Hazards; *--^t-a^(. a(-^ yr.TO^- Precautions: Routine use of protective clothing includes laboratory coats, latex gloves, dust masks, and safety glasses. Other recommended In precautions (\vs~^ \'^aW^\-\ t/'y.o.Avu'^ Case of Emergency Related to this substance, contact: Qn-^ mK f\-^ "' e^y Sc<-t-- ^. Love-UiJ (person) of "D^Y^-/CC^D (company/division) at 3>-^--4?'t-4^oQ) (phone number) IV Disposition; All materials will be returned to the sponsor three months following submission of the final report to the sponsor. Person and address to whom test articles are to be returned. f^fPr Kame: Address: VI Signature: ATXPL4 A^Li^7\ ^T^----1! -^ Date: 3-/-2-//9/ \) Company Sanitized. Dcea not contain! TSCA CBt Du Font HLO 432-91 APPENDIX B WEEKLY BODY WEIGHTS (a) 36 ^^^S-RV ^um[]:,^ p- ., . ,,,, .,,.., . '/_.-.....;; ;..-!'. ^-.y; Du Font HLO 432-91 Animal Number Sex Initial Week II Test Group 1186 1102 1103 1104 1105 1106 1107 1189 1190 1110 M 428 M 441 M 428 M 442 M 412 F 403 F 409 F 414 F 406 F 432 488 515 486 ' 505 463 460 435 441 446 476 WEEKLY BODY WEIGHTS fq) Week III Week IV Week V 574 630 692 592 561 721 545 611 685 575 631 685 503 544 593 511 558 599 503 531 573 481 523 547 495 530 530 518 578 607 Week VI 746 777 717 727 638 628 619 590 607 654 Final 765 795 750 755 661 638 599 . 606 628 672 Company Sanitizect', Goes not contain TSCA cai 37 Du Font HLO 432-91 Animal Number Sex Initial Positive Control Group 1121 1122 1123 1187 1125 1193 1194 1128 1129 1195 M 451 M 411 M 431 M 434 M 443 F 402 F 407 F 420 F 444 F 401 WEEKLY BODY WEIGHTS (q) Week II Week III Week IV 544 611 668 465 511 564 496 545 616 462 521 543 502 574 635 458 540 573 448 485 534 467 538 584 479 529 526 435 467 517 Week V 740 623 673 593 705 649 585 648 611 563 Week VI Final 804 830 669 680 737 736 612 604 741 780 676 722 615 638 678 651 629 626 593 613 38 Du Font HLO 432-91 Animal Number Sex Initial Vehicle Control Group 1111 1112 1113 1114 1191 M 418 M 439 F 423 F 436 F 401 WEEKLY BODY WEIGHTS (a) Week II Week III Week IV 488 548 596 503 580 570 436 486 497 453 496 529 454 480 516 Week V 647 638 497 558 540 Week VI Final 680 695 637 634 528 528 600 584 556 576 toh Coirnpany Sanitized. ;.oes not contain TSCA CB 39 Du Pont HLO 432-91 Animal Number Sex Initial Negative Control 1116 1117 1118 1119 1192 M 438 M 414 M 410 F 411 F 427 WEEKLY BODY WEIGHTS fcr) Week II Week III Week IV Week V 532 626 696 770 481 599 608 672 478 546 614 680 477 521 578 618 481 523 569 611 Week VI Final 834 840 726 722 706 731 658 676 650 659 D 40 DU Font HLO 432-91 APPENDIX C COPY OF RAW DATA 41 Sonipany Saniiizeti, Does not contain TSCA QQ[ UU fonx MljU 4J2-91 PHARMAKON RESEARCH INTERNATIONAL, INC. N.B.tf ;V^ Page # ,^/ c Skin Sensitization Test in Guinea Pigs Dose-Range-Finding Study Sponsor: /"r^ <//^r/- ^/g- /^W^ Purpose: -7^ e^/cufff. ~+fle ^e/^a^ i/l fUl/K'^p'yS Study No.: WI/AV-W-OO/''?/ 2r/7S, Aza-/70^ ftafcy^fa. / a^-Mc -^f- /na^y,< / Method: fcfe/'h ft/ska / ^^ Date of Initiation: Date of Termination: -/^^^ / " g ' Test Article: // CMf4trf//l Description --..--, ---- y^^----^ /^'^ W^) Amount Received: Date Submitted: g^4^/7/ / ' ; AT/i .y^ r l V~~~ ^ v e i s : . / . n,.^..-0-.j.-5 & ^ n r / / 0 0 % Vehicle: ------^^^Dose ^fe^^^^7^^/ Route of Admini s tra tion; -rffi Animal P.O. %: ^//-^^7f/// Species: (flU'ne^ ft7 Food Lot {t:0/g>.V^/V Strain; ^f-ffpj__________ Identification: uS^ &t<^^.,7!/ ^ No. of Animals on StudyiO_________ Sex: Male_^______ Scaled ^^^C'jS^f^V^eJb) Light Cycle Checked: '^//9/ Animals Clipped; '^/Y/giL-________ Compound Preparation:_____/0^ ^ ^)-^'^/^fc^^ ^^7^^^^ ___________,5?^ ^^J^. ^ /^^ -7 J2^^ f^ ^^^m f.o% z=7 ^ ^ ^H^ ^ Wf.^ ^g^j^^_________ ^^A^^ fijfl H^ ^//o^r^. 1 r A . 7 / . ^ . / , / / - / . i / J / fc ^--' " /y-J fi f^'"'r--/--/^<'/^}'/ (/ / A^S --" / " ^ r ^--^^--Uy^--------^--/--4/--- ATABL3 (1) ^ ^ Inv'yeastt-ji.gyeai it-otJir. A V^)//^ . //l/)7l07ir/L Study Director Date' ^//^/ Date &.> Du Porit HLO 432-91 PHARHAKON RESEARCH INTERNATIONAL, INC. Skin Sensitization Test in Guinea Pigs TW N.B.fl Page i C Dose-Range-Finding Study In this test, both sides of-the. animal are shaved and exposed to four concentrations of the test material. The different concentrations are applied concurrently to four different test sites. A given concentration is applied to a different test site on each guinea pig so.as to minimize variation in.. response due to dfferent skin locations. The format for application of the test art-iole is-described.below;. ....--_--.-.--- .. . --.-.__. _.. . . . ....._. ....,,_ ^~7 Figure C Figure D Site Site 21 =fO.%/ ^ = ... Site Site 3 4 = = y/e f^- l O O ',/ O . -W, ... c 1-4: Test Sites Animal No. Sex / ^ ^ ^ J ^ - . . Test article and dose //-/^ Q. 1.0,10,5& a^\ IW. Initial Wt. (gins) ^ ^ w 0^r Timte of Dose ^ Wf^ ^llol^lA A 24 Hour Scores Site Site Site Site Final 1 2 3 4 Wt. (gins) 0 0 / ^ ^1' ^ -r2. 0 ^ 0 ^D\/ ^L /mA-t^n^, ^^> ,^/3 /^/^y^ //// /^/ra. / U-- ' 48 Hour Scores '/-" 0\n 0 / o\a ^ / n0^ 0 ^/ W-7/ 5^ '' r L ATAUL3(2) Company Sanitized. Ocas not con'am TSCA CBi 43 uu i^onr uijU 432-91 PHARMRKON RESEARCH INTERNATIONAL, INC. WO N.B.S_ 3^ Page 1 c Skin Sensitization Test in Guinea Pigs Ti tie: -S/t^ Se^.'^iT/'S'0^'"" Te.sT' ^ 6-u^efi /^/c.^ r. Sponsor: ~. c{^> Ty^r cfe ^e-^o^es -rX;5^r--c/y if e/efff^ff^ ^ fv^t^^ Purpose: TC^T- ^i^-r^f.^c. ,'^ cn^fM p/rj . / ?"/p-- ?r^-^ /*<'--<;-/<""< o-^ T^C Study No.; 7'r/-y,iy-T>'u' - Cro/-?/ Method: /fg/M ^ P^rpe^C ^/O Technical Initiation: '//J^4/ Technical Termination: uji \ I/ Test Article; H -/&'7?0 Lot Number:_--__--__i--_f--\) T/--ft Descriptio: Vehicle: ^ ~v ^ ,. Amount Submitted: /</3. y/^ TW ^infS.' tr ^ Receipt Date: ^^^3-7/_______ Animal P.O. S .^OII-D^lTllit- S/jll 0^17^ ~ / Positive Control ^-^J-^AfwA Q^/SW Lot Number: Description. /?/A^^^^^3'_ Vehicle: ^.^^^/^.^t. )/(/ Route of Administration llvwJ?ifmm ^9" if ' 0 '^~ Toxicology Lab. No.: Species: ^(j^f^ p,c___________ ^S^/M^I^D^///} Height Range: '^Z>-^f& g-^^^y____ "o. of Animals on Study: ^/)wiMOO ) c Scale ft :Jo ^.-n.^.rj ^^^ -^^ ^.Sg^ J Animals Clipped: //^^" -//?^y S'/?/^ y/si/t/^ p/^/^ Strain: //^i/s'T't.&y_________ Food Lot S; fj Sex: Male /S' Female Light Cycle Checked; W^/^ ?/7^iy,1<r//J,/, Fonnat for Primary Irritation, Induction and Challenge Phases 1-2 Primary Irritation and Challenge 3-6 Sensitization Phase (Induction) c ATABLE3 c/'^.^y/./^j j^/)^&y^/ ^^stigator/// m^W^. (^frrifft^ Study Director ^/1/?L Date Date WSft.siyScii'l.-.ii.SsL ..,.,::.:: .._., , ,^^.,;^ Du Font HLO 432-91 Notebook 0 ) L/70 ' p^e. " W c ..PHARMAKOM RESEARCH INTERNATIONAL, INC. Denial Sensitization Study: Guinea Pig Maximization Test ... ....... -._.-..,.....GENERAL OBSERVATIONS ._ ... _ ............. Study No.: .-. Compound: Date >^^,-_//^^'^^^^^ ---'- '' :^j(/^/^/^ ^/gy- /^/- Wl-.tfl Sponsor ^^Ai.^J /^ ~/g''79A_____-_A_N__= __(a_p_p_e_ar_s__n_o_rm__al_) ______________I_n_i_t_i_a_l_s_' ~//^<'~//^7//^. - /^^-^/^^/^/^^^^-z^^^^ ^^7 /i/i^^iC-^ ////i/W^- -r-.;^.i/',r,/ryf^---c,.^,y/-//y:-^-^^^i ^^ ,'/j 0^i.'-^^A^)-!^-- - ^^-t- ^../T^^o.z,^ ^^ x^ A '^^'.^-^^i y..'. 7' - - -/ 7 7^ :- - C -------------------------^ / / / / / / / / / / / / / / L // Company Sanitized. Does Ws contain TSCA C-B; 45 / Du Font HLO 432-91 N.B. . Pageft ;W) ,-^< PHARMAKON RESEARCH INTERNATIONAL, INC. Skin Sensitization Test in Guines Pigs Test Article Preparation Study No.: T^-^? y-b^-cra ^^/ - ________ Tes^t" Article r-^--/^.^^ ' ---- - 'sponsor-: y~.T. ^vT?Jr^e. /^.^^^J- ^ ^ M ^ M ^ s ^ ^ ^ <^2^/^^^ prinary Irritation 'Phase--(Week I) = /^ .)7?^y^7^?3^^^^ ^^..^^J^^^^^^^^^^.^^^ J-....J^;/- P. ... / ^ ^ ^ J ^ ^ ^ y ^ ^ r '^>^ ^-^^^^Y^^^^^^^^ ^^^^^^^^^^/^^^ ^^<^^^e -;-- - ^ ^/"^ ^ , ^^ 4 - r ^ ^ 7 4-^A induction (Week I) = {u, / %^ ^VYW/) ^ y^^/j^^ y. ^. ^,/: ^ {IM. ^ O.^'M^^ 'fr^ r^fmO.//^-?^^ ^Q^ ^w'-^^y^/^^/y^/^^^^^w^^^^ induction (Week II) = (^/^-^/ffff^^ ^^/^^&^ f^ ^t^^ //g^^ ^Q9^^A^^^^ " ^^^^^''^^ .^o^ /nn^A -^/W/A ^ ^ 7 ^ ^ /_______ _f[^^ f^d^C^^/^n Inducfaion (Week III) -.(w. I.O^-? /W/W/) //0 ^ Gl/6f/ft ^ ^'/^< ^S- ^W^. /m^ -7 o. y / ^%-^ fii ffD^.^ IPA\U' 'Tc^Afaw^Jc/To^g^Pc'^^/^o.A)^. l/ ' 6 ^ K T ~ s c ATABL3(3) ) ^(-'ti'i'&Hi^' ^^^'i^.e-c,. Kc;a& !";f.; c^.i^t'7 T^'C^ C'' 46 Du Font HLO 432-91 f M N.B. p^s " .3^) PHARMAKON RESEARCH INTERNATIONAL, INC. Skin Sensitization Test in Guinea Pigs Test Article Preparation (Continued) Study No.: ^-yg. </- J)u - O-D/- ff________________ Test Article: /^-- /&~7'/C>_______:_____Sponsor :^2~^/?.jr ^-e /t/e,~, ec//ss Induction (Week IV) : (w, / 0'k --? /Pff/vm^)6/JM^''/>yM^/^ ff. S/O ^iK" ///^--7 ^^^./^^r^^^ ' pfT)^ ^- /^^y^^//^' ^7^1^^?^^^^^^^^^V/^ ^/^/^^^ ^^^7 '//9gfa^ Challenge (Week VI): (<&/^^ '^'^ ^/^/g^^^^^^^ S>W ^^^^^^.^//^^^ C /^^ ^ff^a^. / <) ^^^X^^-'? ^O^WMV^- ^J^? ^JJ^g^ ^7^A^^^/U?^ W. ^- /^^^^"7^^^%Zi>ti^ ' ^ ^ ' / /^ Rechallenge (Week VII):______ c ATABL3(4) Company Sanifized. Does not co^:ain TSCA CBs 47 Du Font HLO 432-91 Notebook Page It <'^T C PHARMAKOH RESEARCH INTERNAIOMAL, INC. Skin Sensitization Test in Guinea Pigs ' - - -Body Weights (grams)' --Sponsor: <$:Z' l.'Clffwt' j'." ^'''wjui" Test Article: ^i-l^fi. ' ^ Study Number; f-'f^t,^y/,)'l,c-iWf '^' Lot Number: O^Dl'lCft.bU Animal ^urnbe r ~:'^1 a 7 ^ TiS-- S ex c -7 -^i -^l^ -t 7^^7. 7^ ///^? - ^ ^/7 /'^ /^ nitial leek-1- Vleek 1-1 .^.... ^/- - ^ '/^?- - .6^- Week III - ^ ^6. - W eek- IV ^11 ^eek V M eek VI Iff^-- 7^1 .. ^<r Final 750 W- W l.,^l. . ^ . .. ^1 - G<lS'-- 75T w- ^... .^ ^3 66.^^ ^/ ^:... Jl6^ ^... -^/.^ .- W ../.^ .. ^ ^ -M^ ^/ ^ a - //^.7..... ^ ^11 ... .^5. .-.- M 6^. W -- ^?3 - ^ 6./9 W ^D ^1 //^ ^7 ^s ^^ .v^ ^- o^ ^. ^7.. ^ !lir. Q- w 1ft ^ ^^7 W a ^ W ^ ^ w^ -^/ ^7 ,./^ - . ^ff/.y s-ir-y^ . f:^g ^ . . -/3-^ y:jc^ iw ^ j'-^y y (^di^ ^ 5l3--7/ft/ ^5'-^ S-e- - /^<4 I/ ... .... . - - L ATnBL3(5) ^^~^ t ^/^) ^&^&^ A^^5. ^/7/y/ ^^l^^es'tiqator '^ / /'Drfce ihf^i^rL t,hllf Study Director TaTe 48 Du Font HLO 432-91 Notebook 8 (1(0 ,^7 PHARMAKON RESEARCH INTERNAIONAL, INC. Page It C Skin Sensitization Test in Guinea Pios Body Weights (grams) Sponsor: F-r.^.'t-htTP'^ WFfViMjg;) Test Article; i'i - (<?}(', Study Nuinber:'p3^^-Cii\-ooi - Q/ Lot Number: f^nf- Hnnlir^UJ Animal lumber | S ex II I][nitial | feek I Week VFW- ?.C(, \\\\ (^ ;ll,"? ^ G m3 -t- ^ l>9' ^ ^,L- Hi^ </^ ^3 ^^ ^^ l ^-91 'I w ^3 ^ W ^ ?% Week III fr'eek IV ^ ^ <<^ ^ w ^r^ ^ ^ ^7 ^ ^"/^ j'A'-^ /%^ Week V ^V7 ^3f V?7 ^ ^vo 5/^ Wleek VI Final /^ (^ ^57 W J^ 5^ boo W ^ S7^ ^-ms-v^^ ^W C ?'^/T^|<;.; v Inf.. JIC7 ;l(9 ^.9 !l^ ^,. ^ /r' ^3^ r7 ^I'l l?' Hio 0 -r ^11 ^ ^7 t-SH-'jl T MSWff^ ^5 ^1 4l? ^77 ^i ^-6.;7/ f'^,, ^^ ^ ^7^ ^ W ^^> ^-e6^^ ^/-/ M ^ 6ff^/ ,^ n^D 67^ (^ ^^r ^/( ^^hl ^ - W W 7o^ w ^ 1^) ^ ,^7^ ^ C^9 ^sM-y^. ^/ ^ D ATABL3(5) '^/' ./ ^> / / w-'/v^ ..^^y s^^-^ ^/~7/. " ^fvestigator ^ ./'D^te/ /UOT^S(.JAf^cmr'L/./7/9/ Studv Director Dace Gewwm' SanHssea. DOBS not cor'ts'n TSCA CBt 49 Du Poht HLO 432-91 PHARMAKOH RESEARCH INTERNAIONAL, INC. Notebook ft mo Page S ss. Skin Sensitization Test in Guinea Pigs Body Weights (grams) \\\w-m-oo^iw MABL3(5) ^ f e ^ ^Sw^^.^y_^/7^ _^Ifn(vveessttiiqqaactor - ^ AA ^< DDaaee' '^ /^ KMOj^(o. UAw^rLi f.h^/ Study Director----^te' ---- 50 ^y r r \ PHARMAKON RESEARCH .INTERNATIONAL, INC. Individual Animal Scores During Primary Irritation Phase Sponsor: c.T~ JW^iT jY JVgrt^jjg.-^ I Study, Number i,7^a'</--.hc^-00( -9/ Test Article: -<| \<S]<^0 . '' [ Lot Number; '" ^flA ! ': ' _______ i iI Notebook # E^age;_ .7TFp..^^rra^rL^ Animal No. Sex /^ / cr , /^63 ^ //^3 ^ iic4 6" /i0^ <r llt)L, lio'l ^ ll^l ^ \\% ^ J(/0 ^ i_____! Dose Cone. ,___________; Left Shoulder .ii.,.:! ' /^-^r^ ) ; , i^^ ^- ^> Dose Cone. ; Right Shoulder ; r--\ I^^ . '--<-1. ----! -- i , , Time of ' ;24 ; ^ 0 Dose ".^6P^ Hours ^ ':c> 0 1 ''/ 0 1 '0 (} . i i Tims of Reading i '//'AS"^. !' ; i ' ; ^' "^^ ^ -\, <C^ ..^ ^ ; ^ ; ^- <^' ^ \ . . <^- -i^>-------- \0 D 1 0 h1 !0 0 i 0 . 0 ' \ -r 0 P t i i 1 ; 1 ^ '.^fi^ / 0 6 ' /-'A>^< W - ^ -/ C:^(c^. lW///, -y -."3o ''// /lU-A : i , ' i i i -8i 1Hours fQ.^, ^ j 0\ 0 \ c>\ 0 .'.Time o 'Readin --i --f""\--/Q--^T i ' i \ > 0 i-4 /) ; ! & D a : 0 1 \ ! 1' ^ 0 a 0 \ 6. & !' ^\ 1-'^ --5G-^^'?n/^ " \ i i i i i ; . ^ \ ' i !' i . 1 1 i 1 ! ; i I 1 ' '1 ; . ' ; .' Initials ATABL3(9 11 " i .'$>-: (ji-^t-AAo\A(^, \^AJ^\~ jl^\^^ ; i 1 i ' \'- : i . ^ 'i \ ; ( i 1 , ; " ' ' . . ).. . i i ,' \ i ,' . r r \ PHARMAKON RESEARCH INTERNATIONAL, INC. Individual Animal Scores During Primary Irritation Phase Sponsor; F-f-JVi^n-^L }/Fm^i^^ Study Number :^(W^^->)f,l. -CGl -9/____________ 'Notebook Test Article: ^ -('?,79<0__________Lot Number:_______/^/J_________________' Pa9e VFh.oL^ Animal No. HI, (11,9 11^ 1|1<1 1^ OHL^.L Sex ^ o7 ^ ^ 0 Dose Cone. - ^-^^ Left Shoulder ^ (c>. ^C^ (^ ^ s ^^ ^X ^ ^^ ^ ^ c Dose Right Cone. y Shoulde/ / / / V/ -- Time of Dose /'.^W \ / .V /:^/in^i H^- M '11 . 24 Hours 0 0 . 0 0 0 Time of Reading' 1:^^ \ / - \y' ^ ^ /,' 93^. 48 Hour's 0: 0: Q : 0 ; 0 \ -it- Time Rea /' / '/r/ / V?^^ - Co^ L ilii^ ^ ^ ^ in-7 ^ / / irl: (7 / / K/? U X / //^ 4- / / ^^- i i i : '' Initials ATABL3(9) PHARMAKON RESEARCH ...i-ERNATIOMAL, INC. Individual Animal Scores During Primary Irritation Phase Sponsor; (Sjr.Kd^fft h9. V^rmjU^ Study number: 4^4ot^-g0l -^/ f ^ WDUG/^/ Notebook tt / Test Article: U-Vg'.l^O_________Lot Number:_______//U__________',_____ Page ^>f9 ^^E-WH-to.-, Animal No. Sex 151 (7 i^a ^ na3 S IW s 1135 ^ \l^ J. liQ4 na^ 9 1^9 ^ 1^6 ^ Dose Cone. Left Shoulder .----^ (c^ ^ ^ <^ ^N ^ ^ ^ ^ ^.^ ^>^ ^ ^ \ ^^ ~ N-----^ Dose Cone. Right Shoulder Time of Dose 24 Hours ;^ --------------/ ------/ 0 0 / D / 0 / 0 0 C> -7/----- D / -\ / 0 / / /^?l:3t^ c> ^-Sft-""' /^,s Time of Reading 1:33^ i \ -/---- l'.Sls^ ^.30-^ 48 Hours 6 0 Time of Reading /,/'1 o 0 0 0 0 0 0 -^ ^ 0 -^^ ^-/ /' d6- ^ Initials - ATABL3(9) r Sponsor: <f.-Z" ,^/g.j Je /Je^oc'/tS Tust Article:^,/- /.g^^ PI1ARMAKON r\ RESEARCH , INTERNATIONAL, INC.. Individual Animal Scores During Induction Study Number: /^y^^--^V- o-o/-y/ Lot Number:_________yK/4 Notebook t Page 0 Animal No. Sex 7^m< nc/-^- ll^U (^ 11^ ^ /r IIC -. IIC-1 ^ ///.^ ^ i^ ^6- -7- I/C-1 <f; I ii<r, i: r' (',' //%. -t ///a (..1 7- Dose Cone. Time of Dose Week I 24, Hrs. Time of Reading Time of Dose Meek 24 Hrs. II Time of Reading W qek :' III Time , Time . .. j of 24 of Dose Hrs. Reading i [ Wee Time of Dose A'3D/^ 0 /,'c^/>4 ^cr 0 3!: io/l^i //?:-SO 0 <9 ^ ^^ C> 0 f--^^ '0 (0 0 0 0 -^^. 0 H3 0 \ ^ -^ . -J ^ )C^ fill- / ^ c> 0 0 V 0 >. i Y m% \/ ^^ \f ^ ^.77, 0 /A5.V. 0 S:i<e ^ /f:s7 J'- 3 '^/TU^ s/k-ft/ ^Cfcl S'/' /Bt ^-------------- ' ----(. 0 ' 11:16 A^ //; Off, t D ! 0 i 0 ; 1 0 ! 0 : ( 0 . ,.I*.. i 0 ^ ^ // : ' / \f ^w ^ 7y:,^5/^! If-l^ i -/L -y/ W^. ' Initials. ATABL3(10) Sponsor: (ST.^O^/^ ^ f^^W^Cs Test Article; ^ 1'^lQO PIIARMAKON RESEARCH INTERNATIONAL, INC. Individual Animal Scores During Induction Study Number = ^H ./^- A/A -mi 9/ Lot Number:_________/^J^ Notebook Page Animal No. Sex Wllf^ . r^ct, III! ^ ^ //r? /7 111^ lll4 ^ I1'// V Dose Cone. Time of Dose Week I 24 Hrs. Time of Reading .^ '?> /.'37/9 '/ 0 >-' s^ 0 l:^ ^ 1 ^ 1^w/.v^^ 0 \ / 0 0 l-'y7^ - ^//^ Time of Dose 9.^^. v Cf:^ ^--S-.f/ /^ Week 24 Hrs. II Time of Reading 0 $':Z.7 ^ 0 0 \ / > 0 1:3o A-^ ^ ^ Time Wq|ek III of 24 Dose | Hrs. Time of Reading /A-5^ r> 0 //.^^ V 0 1 / ~\7' c> //-.fto 0 ir.^s^ W ^<5'/ '^-^ ^ W Time of Dose l/'.ILff ~\7~ n /you kWA cft-ffyy ^Rp<-fl^CoiA-fr 11^ 0^ iir-l ^ /(^ /7 //(^ ^ //^ 4; -.----- t Initial!; ATABL3(10) PIIARHAKON RESEARCH INTERNATIONAL, INC. Sponsor: j~J>^..)7r.l). ^)H^LiZ,3 Test Article: -ti - l^nQp Individual Animal Scores During Induction Study Number ::''^^'^'t/^^>44 jX^-CQt -'9< WW/l/-60^/Noteboo^ ft Lot Number: 3 ^/A____________ Page tt Animal No. +^H-1\ l/ai li^-> Jf^ 1^-7 l'<05 li 93 ll^l 11^'S i,.^ 11^ Sex l\-^t ^' (f:7 ^ 6-^ L"' ^ ^ ^ c- (^ -r Dose Cone. (^.. \^">"**s. ^> ^Y 3^)'^1 ^^ -t4 ^ 3T^4^^ ^ ^ Time of Dose Week I 24 Hrs. /w^? ^> ^) (3 / / 0 0 ^ \^ 1 /'w 0 W ^- 0--(V- Time of Reading /'.y^?^ \ / /.30^ ^w^ Time of Dose ' 0< \ /Wf : ^T MS^S Week 24 Hrs. II Time of Reading 2. c-o 2. <?o & '3/ /hvi / / / / ^ / / \ / / ' 3- 37/^ s-/> ^/^> III Wqek Time of 24 Dose Hrs. Time of Reading ///^ 6? ll--^^ 0 1 1 / W 0 i &y / ^^ ^s^ //.7'3 ^{^i v / f ^ //.. 0?^ ^'A ,-^/ /A^ Week Time ofi ". Dose ' H //;/?^ 0 \y f^ //,'W Q Initiali S'oi ^tdawa^. ATApr.3(10) . r r^ PHARHAKON RESEARCH INTERNATIONAL, INC. Individual Animal Scores after Challenge Test Article - //^^ ^ L ot Nuinbe r: /U//P Page . ^- / ,, ' f 7^7/mW>U' Dose Animal Cone. No. Sex Left Shoulder 11^ ^ i/^. ^ //^7 (r iinH rr li^ ^ I/O^ ^ 'r^r--\^ .^ ^ r^ ^ ^ \^fl /Ifln f ^ ,i/^ ? ^ ^r' ^ n^ ///o ^ \^^ Dose Cone. Right Shoulder ^K x^\^' \ ^ ^ ^^ ^ r \ ^y^\ - '~--"" Challenge Scores Tirae of 24 Dose A^SA fl'.^ 0 0 0 0 Time of Reading ^Hrls.. A.-y^ 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 > 0 0 0 0 0 0 0 0 0 0 \ / 00 \ / 00 M ^ ^ 0 0 ll'^ ^ /o:5'i*^ 0 0 ' ^^ ^iAj Q? 71-' /[n/JU. )Ao^jicL'^r d Initials ATAQL3(10) . Time of Reading /o.-ys/i^ \Y/ /0:fJ/^ M7//^^'a^" Dose Dose Cone. Cone. Time Left . Right of 24, Shoulder 'Shoulder Dose Hrs i ' -^ ^ \ )/. i ^l; ^ \ f^. r>'\J y \VT \ \y\ / VA ' / . \xl\U / / : / Recha Sco Time of Read / / / F /^ PHARMAKOM RESEARi-n INTERNATIONAL, INC. Sponsor- (^ ^ /li/Jr^jfW Teat Ar tide = ^-/^ In dividual Animal Scores after Challenge W/MW, s tudy Num her: ot Nuinbe - ^W-^/-^/- ?/ Wt- Challenge Noieb00. ,, Page . 3 Recha Scores Sco Dose Dose Dose Dose Animal Cone. Cone. Time Time Time Cone. Cone. Time Tinic Ho. //// ///a //i-\ //w //Cfl Sex Left Right of 24 of 48 of Left Right of Shoulder Shoulder Dose Hrs. Reading Hrs. Reading Shoulder Shoulder Dose -^ ^ ^7 ^^ -/f ^^ ^ t<( ^ n:^ 0 /O'-Sl/^ 0 /Sl-'-fl^ 0 Q ^ 0 0 0 0 ^ ^' ^^^. A- \^ ^/ \y ^ W 0 WSft^ IWsW^ ^ ^ 0 /o-.fi^ ^/r ^ ^i 24 : 0^^ .Hrs ^Read Initials ATAUL3UO) p ^ PHARMAKON RESEARi-n INTERNATIONAL, INC. Individual Animal Scores after Challenge Sponsor^ ( c/< /)(^/7)n7'/rff /////w>^/>. s tudy Number; ^-^^ /wWy-f^\ -^9/-^/ Teyt Ar cicle = ot Nuinbe r: Notebook # Page ft ^w^ / ' Animal 7 Ly^^V^/tl-^- f Dose Cone. No. Sex Left Shoulder llt!^ ^ ^^s tin cfl SJ-1 ^1 ^ >~ Vi ///^ ^ ^ ^> ?^ 11^ ? " 11^ ^ <^ ^(^.p- ^^ /^ . Dose Cone. Right Shoulder ?^ ^S> i ^ '-^i 'Si o^ rJ ^^ ^ ^ ^ ^? <^ ^- z-- Time of Dose //^ \ ///:3/ ^ Challenge Scores ^h Mrs. 0 0 0 o Time of Reading ^f l r s . l/:oTf^ P 0 0 0 Time of Reading /o:y7^ f) 0 0 00 V 0 0 0 //i)?/hYl 0 Cf/{ 'ffl 0 \ / 0 0 If-01 ^ i't /?/ -r Mz^ Dose Cone. 31'ruuldur' ^^ <- <^ ^ ^ ^ ^ ^ ^ ^ Dose Cone. Time R19h^ of Slioi.il dor Dose -4 ^ --------' ^----^1 ----------^ ^ ^ t TH^cha Sco Time ^ of Hrs Read 0 o fl'O 0 0 0 0 \/ 0 0 0 0 // -^ ^ ^ w^ ^<A ^//^) 01 -- ^-'fy A. ' /IA/M^ /}, ' W/ ^/7Z r^ '-"^'.^t ^ 1^ /i.W, L 6r 1 Initials ATAHL3(10) /-' /^ PHABMAKON RESEAIo-n INTERNATIONAL, INC. Sponsor: C,.^/-.((^l/t Test Article ^l^:/-/^ ftW-^AL- , /^\ ^--- ^ /,<^%^^e Dose Dose Animal Cone. Cone. No. Sex Left Right / Time / of 1/^1 ^ 11^ Z 11^3 -S^ ,//W /IS-T ^ ^f /^3 ? ^ ^ 1/9^ J. / '//^ f //^ JJZ- ^'^ // \/ //^ Shoulder Shouldefr Dose / 1/^0 / ^ ^< ^>^ / ^ ^ c^- \ 1 ' '^T^ ^ ^ //:'/,7 w t/sfl^ Initials ATABL3(10) ot Nuinbe - KVfl- Challenge Scores 24 Hrs. ^ x ^ Time of Reading 11:10^ 9. ^ 3 y ;?- \/ / c> //'/^ ^ &/'; /7 Time 48 of Hrs. Reading ?- //0//f^ ^3 / / ^ / ^ / \^ ^0 //:<37ft) ^1 , Page . W Dose Cone. Left Shoulder Dose Cone. Right Shoulder Time of Dose Rechall Score Time 24 of Hrs Readin -/ ^ / / f / 7^- / / / / / / / / / /