Document 3V685N6Lx3ZDmeYmz85Ro4VJ

SPONSOR Elf Atochem S.A. Cours Michelet La Dfense 10 92091 Paris-la-Dfense CEDEX France AR226-3069 uuiNfUJNTIAL STUDY TITLE ACUTE ORAL TOXICITY IN RATS TEST SURSTANCF. STUDY DIRECTOR Stphane de Jouffrey STUDY COMPLETION DA TE 5 Februaiy 1997 PERFORMING TARORA TORY Centre International de Toxicologie (C.I.T.) Miserey - 27005 Evreux - France LABORATORY STUDY N1MRER 14886 TAR Company Sanitized. D oes not contain TSC CFj5 CONTENTS STATEMENT OF THE STUDY DIRECTOR OTHER SCIENTISTS INVOLVED IN THIS STUDY STATEMENT OF QUALITY ASSURANCE UNIT SUMMARY RESUME 1. INTRODUCTION 2. MATERIALS AND METHODS 2.1. TEST SUBSTANCE 2.1.1 Identification 2.1.2 Preparation 2.2. TEST SYSTEM 2.2.1 Animals 2.2.2 Environmental conditions 2.2.3 Food and water 2.3. TREATMENT 2.3.1 Fasting of the animals 2.3.2 Administration of the test substance 2.3.3 Date of treatment and duration of the study 2.4. CLINICAL EXAMINATIONS 2.4.1 Clinical signs and mortality 2.4.2 Bodv weight 2.5. NECROPSY 2.6. DATA EVALUATION 2.7. ARCHIVES 3. RESULTS 3.1. CLINICAL EXAMINATIONS 3.1.1 Clinical signs (table 1) 3.1.2 Mortality (table 1) 3.1.3 Body weight (treated animals: figure I, table 2) (historical control animals: figure 2, table 3) 3.2. PATHOLOGY (table 4) 4. CONCLUSION * jf 8 8 8 8 9 9 9 9 10 10 10 10 10 10 10 11 11 11 12 12 12 12 12 12 12 Safiilizefl. Does not contain TSC.* Figure 1: Body weight of treated rats (g) Figure 2: Body weight of C.I.T. historical control rats (g) Table 1: Individual clinical signs and mortality Table 2: Individual and mean body weight and weekly body weight change of treated rats (g) Table 3: Mean body weight and weekly body weight change of C.I.T. historical control rats (g) Table 4: Individual macroscopicexaminations at necropsy APPENDICES 1. Test article description 2. Diet formula 13 14 15 16 17 lg 19 20 22 and 23 4 STATEMENT OF THE STUDY DIRECTOR The study was performed in compliance with the following Principles of Good Laboratory Practice Regulations: . O.E.C.D. principles of Good Laboratory Practice, Decision Concerning Mutual Acceptance of Data in the Assessment of Chemicals, C(81)30(final) Annex 2. May 12, 1981. . Dcret N 90-206 du 7 mars 1990 concernant les Bonnes Pratiques de Laboratoire (Journal Officiel du 9 mars 1990), Ministre de l'Industrie et de l'Amnagement du Territoire. I declare that this report constitutes a true and faithful record of the procedures undertaken and the results obtained during the performance of the study. This study was performed at the Centre International de Toxicologie (C.I.T.), Miserey, 27005 Evreux, France. Toxicology S. d^yoitffrey Date: 5 February 1997 Study Director Doctor of Veterinary Medicine Head of Short-term and Environmental Toxicology OTHER SCIENTISTS INVOLVED IN THIS STUDY For Pharmacy: J. Richard Doctor of Pharmacy For Toxicology: C. Pelcot Study Supervisor Company Sanitized. D oes no! contain TSCA CBI STATEMENT OF QUALITY ASSURANCE UNIT 1. Specific study inspections Type of inspections Inspections Protocol Report 21 Oct. 96 20 Jan. 97 Dates Report to Study Director (*) 29 Oct. 96 20 Jan. 97 Report to Management (*) 29 Oct. 96 20 Jan. 97 - 2. Routine inspections performed on other studies of the same type according to a frequency defined in Q.A.U. procedures Inspected phase -- Inspections Treatment/test substance 14 Aug. 96 Preparation/test substance 25 Sept. 96 Dates Report to Study Director (*) 20 Aug. 96 25 Sept. 96 Report to . Management (*) 20 Aug. 96 25 Sept. 96 The inspections were performed in compliance with C.I.T. Quality Assurance Unit procedures and the Good Laboratory Practice Regulations. (*) The dates mentioned correspond to the dates of signature of audit reports by Study Director and Management. - 1< L. Valette-Talbi Date: 5 February 1997 Doctor of Biochemistry Head of Quality Assurance Unit and Scientific Archives i pSMpaif S'an'/feel, V m im eoMain TS'CCBI SUMMARY At the r e q i^ ^ ^ y ^ ^ ^ o y iM ^ ^ ^ ^ ^ k J ^ D e f e n s e , France, the acute oral toxicity of the test s u b s t a n c e ^ j ^ f ^ H m ^ m m m H w a s evaluated in rats according to O.E.C.D. (No. 401, 24th February 1987) andE!c792/69/E.E.C., B,, 31st July 1992) guidelines. The study was conducted in compliance with the Principles of Good Laboratory Practice Regulations. Methods The test substance was administered by oral route to one group of ten fasted Sprague-Dawley rats (five males and five females). The test substance was administered in its original form, by gavage, at a dose of 2000 mg/kg, taking into consideration that its density was 1.07. Clinical signs, mortality and body weight gain were checked for a period of 14 days following the single administration of the test substance. All animals were subjected to necropsy. Results The general behaviour and body weight gain of the animals were not affected by treatment with the test substance. No death occurred at 2000 mg/kg. No abnormalities were observed at necropsy. Conclusion xperimental conditions, the oral LD0 of the test substance!_______ ______ as higher than or equal to 2000 mg/kg in rats. No signs of toxicity were observed at this dose. RESUME j^Ja^flefflande de Elf Atochem S.A., Paris-la-Dfense, France, la toxicit aigu du produit a t value par voie orale chez le Rat conformment aux li g n e ^ i r e c S c e ^ ^ ^ ^ ^ ^ ^ F ( N o . 401, 24th February 1987) et de la C.E.E. (92/69/C.E.E., B,, 31st July 1992). L'tude a t ralise conformment aux rgles de Bonnes Pratiques de Laboratoire. Mthodes Le produit a t administr par voie orale un groupe de 10 rats Sprague-Dawley (5 mles et 5 femelles) mis la dite hydrique. L'administration a t effectue avec le produit tel quel, par gavage, la dose de 2000 mg/kg, en tenant compte de la densit du produit (d = 1,07). Les signes cliniques, la mortalit et l'volution pondrale des animaux ont t suivis pendant une priode de 14 jours aprs l'administration unique du produit. Un examen anatomopathologique a t effectu sur tous les animaux. Rsultats Le comportement gnral et l'volution pondrale des animaux ne sont pas influencs par le traitement. La mortalit est nulle la dose de 2000 mg/kg. . L'autopsie des animaux ne met en vidence aucune anomalie macroscopique. Conclusion Dans nos conditions exprimentales, la DL0 du produit administr par voie orale chez le Rat est suprieure ou gale 2000 mg/kg. Aucun signe toxicit n'est observ cette dose. .. % Company Sanitized. D oes not contain TSCA CBl 1. INTRODUCTION The objective of this study was to evaluate the toxicity of the test substance following a single oral administration in rats. In the assessment of the toxic characteristics of a test substance, determination of acute oral toxicity is an initial step. It provides information on health hazards likely to arise from a short term exposure by the oral route in Man. The study was conducted in compliance with: . O.E.C.D. guideline No. 401,24th February 1987. . E.C. Directive No. 92/69/E.E.C., B,, 31st July 1992. 2. MATERIALS AND METHODS 2.1. TEST SUBSTANCE 2.1.1 Identification The test substance used in the study was supplied by Elf Atochem S.A. Documentation supplied by the Sponsor identified the test substance as follows: . name: - protocol and labelling: | . batch number: - protocol and labelling:^ . Elf Atochem filing number: j . description: dark brown liquic . container: one plastic flask . date of receipt: 11 October 1996 . storage conditions: at room temperature and protected from light. Data relating to the characterization of the test substance are documented in a test article description (presented in appendix 1) provided by the Sponsor. At the beginning of the study, the analytical certificate was not available. 2.1.2 Preparation - The test substance was administered in its original form. IPempfnfSantzsS. Does o eiVialhT bF 9 2.2. TEST SYSTEM 2.2.1 Animals Species, strain: rat, Sprague-Dawley ICO: OFA-SD (IOPS Caw). Reason for this choice: rodent species commonly requested by the international regulations for this type of study. Breeder: Iffa Crdo, 69210 L'Arbresle, France. Number and sex: one group of ten animals (five males and five females). Age/weight: on the day of treatment, the animals were approximately six weeks old, and had a mean body weight standard deviation of 175 4 g for the males and 142 5 g for the females. Acclimatization: at least five days before the beginning of the study. Identification of the animals: the animals were identified individually by earmarks or eamotches. 2.2.2 Environmental conditions During the acclimatization period and during the main test, the conditions in the animal room were as follows: . temperature: 21 2C . relative humidity: 30 to 70% . light/dark cycle: 12 h/12 h . ventilation: approximately 12 cycles/hour of filtered, non-recycled air. The temperature and relative humidity were recorded continuously and records retained. The housing conditions (temperature, relative humidity and ventilation) were checked monthly. The animals were housed in polycarbonate cages (48 cm x 27 cm x 20 cm). Each cage contained four to seven animals of the same sex during the acclimatization period and five rats of the same sex during the treatment period. Each cage contained dust-free sawdust (SICSA, 94142 Alfortville, France). Bacteriological analysis of the sawdust and detection of possible contaminants (pesticides, heavy metals) are performed periodically. 2.2.3 Food and water All the animals had free access to A04 C pelleted diet (U.A.R., 91360 Villemoisson-sur-Orge, France), except as noted in "2.3.1 Fasting of the animals". .Each batch of food was analysed (composition and contaminants) by the supplier. The diet formula is presented in appendix 2. Drinking water filtered by a F.G. Millipore membrane (0.22 micron) was provided ad libitum. Bacteriological and chemical analysis of the water and diet and detection of possible contaminants (pesticides, heavy metals and nitrosamines) are performed periodically. Results are archived at C.I.T. It was verified that no contaminants in the diet or water at levels likely to influence the outcome of the study were present. !?T7nri&&S55 1U 2.3. TREATMENT As the test substance was anticipated to be non-toxic at 2000 mg/kg, a limit test was performed by administering 2000 mg/kg of the test substance to one group of ten animals (five males and five females). 2.3.1 Fasting of the animals The animals were fasted for an overnight period of approximately 18 hours before dosing, but had free access to water. Food was given back approximately 4 hours after administration of the test substance. 2.3.2 Administration of the test substance The test substance was administered in its original form, taking into consideration that its density was 1.07. The administration was performed in a single dose by oral route using a stainless steel roundtipped probe (diameter: 18 G.2", Perfektum: Poffer & Sons Inc., New Hyde Park, New York 11040, U.S.A.) fitted to a 1 ml glass syringe (0.01 ml graduations, Record: Carrieri, 75005 Paris, France). The volume administered to each animal was adjusted according to body weight determined on the day of treatment. 2.3.3 Date of treatment and duration of the study The single administration was performed on 5 November 1996 in the morning (day 1) and was followed by a 14-day observation period until 19 November 1996 (day 15). 2.4. CLINICAL EXAMINATIONS The single administration was performed in the morning of day 1; it was followed by a 14-day observation period until day 15. 2.4.1 Clinical signs and mortality The animals were observed frequently during the hours following administration of the test substance, for detection of possible treatment-related clinical signs. Thereafter, observation of the animals was made at least once a day. Type, time of onset and duration of clinical signs were recorded for each animal individually. - Time of death was recorded individually, in terms of the number of hours or days after dosing. 2.4.2 Body weight ., The animals were weighed individually just before administration of the test substance on day 1 and then on days 8 and 15. The body weight gain of the treated animals was compared to a reference curve of C.I.T. control animals with the same initial body weight. I .Company Sanitized. D oes no! contain TSCA C5*' 11 2.5. NECROPSY On day 15, all animals were killed by C 02 inhalation in excess and a macroscopic examination was performed. After opening the thoracic and abdominal cavities, a macroscopic examination of the main organs (digestive tract, heart, kidneys, liver, lungs, pancreas, spleen and any other organs with obvious abnormalities) was performed. In case of macroscopic lesions, organ samples were taken and preserved in 10% buffered formalin. No microscopic examination was performed. 2.6. DATA EVALUATION Evaluation of the toxicity of the test substance following a single oral administration in rats should include the relationship, if any, between the animals' exposure to the test substance and the incidence and severity of all abnormalities including behavioural and clinical abnormalities, macroscopic lesions, body weight changes, mortality and any other toxic effects. .' 2.7. ARCHIVES The study documentation and materials, namely: . protocol and possible amendments, . raw data, . . correspondence, . final report and possible amendments, are stored in the archives of C.I.T., Miserey, 27005 Evreux, France, for five years after the end of the in vivo phase of the study. At .the end of this period, the study documentation will be returned to the Sponsor. i i om pmy S'anWze?.'Urtss'Tfo!esftafi 3. RESULTS 3.1. CLINICAL EXAMINATIONS 3.1.1 Clinical signs (table 1) No clinical signs were observed during the study. 3.1.2 Mortality (table 1) No death occurred during the observation period. 3.1.3 Body weight (treated animals: figure 1, table 2) (historical control animals: figure 2, table 3) The body weight gain of the animals was not influenced by treatment. 3.2. PATHOLOGY (table 4) Macroscopic examination of the main organs of the animals revealed no apparent abnormalities. 4. CONCLUSION Under our experimental conditions, the oral LD0 of the test substance ^ ----------- **^was higher than or equal to 2000 mg/kg in rats. No signs of toxicity were observed ^ t this dose. Company Sanitized. D oes Hot contain TSCA CBI 13 jgPffgsny Sanitfoe. Does not contaln TSCA C'! Figure 2: Body weight of C.I.T. historical control rats (g) e.g.: C.I.T. Historical data of animals dosed by the oral route: results of control animals from October 1990 to January 1996. i Company Sanitized. D oes not contain TSCA CBI Table 1: Individual clinical signs and mortality Dose Time (mg/kg) Animals Males Females Mortality Clinical signs 2000 30 min 1 1-2-4 h f 01-02-03-04-05 01-02-03-04-05 No D 2 to D 15j None min: minutes h : hour D : day 15 li i Kpmpafty Sanitized. Does hot contain TSCA CB 16 Table 2. Individual and mean body weight and weekly body weight change of treated rats (g) Dose mg/kg Volume , Sex ml/kg Animalc -- 1 Days ---------------- -- (1) 8 (1) 15 2000 1.87 Male 01 178 68 246 64 310 02 175 71 246 61 307 03 170 84 254 63 317 04 172 75 247 62 309 05 178 62 240 64 304 M 175 72 247 63 309 SD 4 8 5 1 5 2000 1.87 Female 01 02 03 04 05 M SD 143 136 150 141 139 142 5 39 48 33 61 48 46 11 00 42 .224 184. 36 220 183 45 228 202 44 246 187 25 212 188 38 226 8 8 13 ( I) = Body weight gain M = Mean SD = Standard Deviation llTfSCA'CBl 17 Table 3: Mean body weight and weekly body weight change of C.I.T. historical control rats (g) BODY WEIGHT OF CONTROL RATS (g) Dose mg/kg Volume ml/kg Sex 0 10 Male M : i SD n 0 10 Female M SD n M : mean SD: standard deviation n : number of animals Days 18 185 262 8 13 90 90 155 201 9 15 90 90 15 317 20 90 225 19 90 BODY WEIGHT CHANGE OF CONTROL RATS (g) Dose mg/kg 0 Volume ml/kg 10 Sex Male 0 10 Female M : mean SD: standard deviation M' SD M SD Days 1 to 8 77 9 46 10 8 to 15 55 13 25 11 i e.g.: C.I.T. Historical data of animals dosed by the oral route: results of control animals from October 1990 to January 1996. .Company Sanitized. D oes not contain TSCA CBI Table 4: Individual macroscopic examinations at necropsy 18 Dose Time mg/kg 2000 D 15 D : day Animals Males Females 01-02-03-04-05 01-02-03-04-05 Macroscopic abnormalities None t ^3/1$Smitized. Does Hoi contain TSCA CS? 19 I APPENDICES *.i `K 'ww 20 1. Test article description Company Saniti W I-D o e s noi conia,,TSCArr,.  TOXICOLOGY DEPARTMENT CONFDOENTLAL 10 October 1996 elf atochem s.a, La dfense 10, cedex 42 92091 Paris-la-Dfense, France TEST ARTICLE DESCRIPTION 21 Test article name Chemical name CAS number EINECS number 3urity IOrigin and batch Batch Elf Atochem filing number PHYSICAL AND CHEMICAL PROPERTIES Appearance Melting point Boiling point Flash point Solubility brownish liquid -22C 95C 50C water TOXICOLOGICAL INFORMATIONS AND USE SAFETY See safety data sheet STORAGE AND DISPOSAL Storage Expiry date Disposal in dark and at room temperature December 1997 incineration J ^ p a n y - s anii;2Bd - -. u* . no! con!ain TSCA CP' 2. Diet formula 23 Ref: A04 COMPLETE DIET RAT AND MOUSE MAINTENANCE DIET Appearance: 15 mm diameter pellets or powder Conditioning: 25 kg double paper bag with aluminium on the outside Daily portion: Rat 18-25 g, Mouse 5-10 g, water ad libitum. FORMULA % Cereals and cereal biproducts.... 88 Vegetable protein (soya bean meal, yeast) ................................ 7 Animal protein (fish) ..................... 2 Vitamin and mineral mixture ......... 3 AVERAGE ANALYSIS % Calorific value (KCal/kg)........... 2900 Moisture.......................................... 12 Proteins.......................................... 17 Lipids............................................... 3 Carbohydrates (N.F.E.)............... 58,7 Fibre ................................................ 4 Minerals (ash) ................................ 5 MINERALS (calculated in mg/kg) Nat CMV vai. vai. Total P ............. 5900 Ca .......... 3300 K ............. 6700 Na .......... 300 M g .......... 1900 Mn ......... 50 F e ............ 90 Cu .......... 15 Zn .......... 40 Co .......... T I .............. 0.3 0 5000 0 1600 100 40 150 15 45 1.5 `0 5900 8300 6700 1900 2000 90 240 30 85 1.5 0.3 AMINO ACID VALUES (calculated in mg/kg) Arginine...................................... 9800 Cystine........................................ 2300 Lysine .......................................... 8500 Methionine.................................. 3200 Tryptophan.................................. 1900 Glycine......................................... 8100 FATTY ACID VALUES (calculated in mg/kg) Palmitic acid................................ 2600 Palmitoleic acid .......................... Traces Stearic acid.................................. 500 Oleic acid .................................... 8000 Linoleic a c id ................................ 14500 Linolenic acid .............. Traces VITAMINS (calculated per kg) Nat CMV vai. vai. Total Vitamin A Traces Vitamin D3 Traces Vitamin B1 6 mg Vitamin B2 2 mg Vitamin B3 10 mg Vitamin B6 1.3 mg Vitamin B12 0.01 mg Vitamin E 15 mg Vitamin K3 0.25 mg Vitamin PP 60 mg Folic acid . 0.5 mg Biotin 0.04 mg Choline 1200 mg 7 5 0 0 IU 1500IU 1 mg 4.5 mg 6.5 mg 1.3 mg 0.01 mg 15 mg 2.25 mg 15 mg 0 mg 0 mg 400 mg 7 5 0 0 IU 1500 IU 7 mg 6.5 mg 16.5 mg 2.6 mg 0.02 mg 30 mg 2.5 mg 75 mg 0.5 mg 0.04 mg 1600 mg Available under quality "Control Ref.: A04 C " U.A.R., 7 me Gallieni, 91360 Villemoisson -Tel: 69.04.03.57 - Fax : 69.04.81.97 (Ref. Doc. UAR: 1992) Pmpany Sanitized. D oes not contain TSCA CBI