Document MMOQR7xq0LyO4eEGpzXv4K56j

ARARG 3237 | | || || | A226. 3239 DuPontsets `TRADE SECRET 'H-24678: Study Title Inhalation Rats Approximate Lethal Concentration (ALC) in Laboratory Project ID: DuPont-8684 AUTHOR: John R. Bamberger, M.A. STUDY COMPLETED ON: April 5, 2002 "PERFORMING LABORATORY: E.L du Pont de Nemours and Company Haskell Laboratory for Health and Environmental Sciences ENlekwtaornk,RoDaedl,aPw.aOr.e B1o9x71540-0050 - -- DompanySanhized. DocsnotcontainTSCA e531 124678 Inhalaion Approximate Lethal Concenation (ALC) in Rats Dupontsess CERTIFICATION We, the undersigned, declare that this report provides an accurate evaluation of data obtained from this study. EvaAluvaattioonnsiReePpaortiheadobgyy: ort.BRlenonmBAYCVMFPD. Pipe Resear Sint Hu,/2002 Bae eedbyStudy Diets TOR\BankSerge MA Research Scent 50gn an [ESP Bamtized. Dossrot conTtSCaAlCSr} -_ 124678: Inhtsion Approximate Lethal Concenaton (ALC) i Ras STUDY INFORMATION Substance esc WEA snonmsCoses:(NAD C-- 1 Haskell Number; 24678 Dupont set "TT Physical Characteristics: Amber brown liquid - Stability: Tcohneditteitonssubosfttahnecestaupdpyeanroedevtidbeencsetaobfleinusntdabeirlitthyewas observed. Sponsor: EL du Pont de Nemours and Company WUiSlmAington, Delaware 1989 Study Intiated/Completed: November 28, 2001 / (see report cover page) In-Life Iniisted/Completed: November 29, 2001 /December 27, 2001 ~ mr Company Sunitizec. 00snotcontain TSCA G2: ------etstrt-------- 124678: Ihalion Approimte Lethal Concenation (ALC) in Rats Dupont868 STUDY PERSONNEL Study Director: Management: Primary Technician: John R. Bamberger, M.A. Arthur J, O'Neill, B.S. Scott E. Loveless, Ph.D. Janice L. Connell, M.S., B.A., CLH. Christine B. Skoranski Pathologist: John F. Hansen, D.V.M., Ph.D. Management: Steven R. Frame, D.V.M., Ph.D. Toxicology Report Preparation: Maryanne M. Wilford, B.A. Laboratory Veterinarian: William Singleton, D.V.M., ACLAM. ~ -_-- CompanySanitized, poe #not continTSCAcay H-24675: Inhalation Approximate Lethal Concentration (ALC) in Rats DuPont8684 SUMMARY 4`T-hhroeuergpreoruiposdotfo 6aemraolsoelCsrolf:HC-D24(6S7D8)IiGnaSirBaRt rcaotnsceeantcrhatwieornes eoxfp2o3s,ed57n,oosre-1o2n0lymgfo/ram?s(idnrgyle, weight). Test atmospheres were generated by atomization, and concentrations of H-24678 were `measured gravimetrically. Rats were weighed and observed for clinical signs of toxicity during a 14-day recovery period. Surviving rats from the 57 mg/m? group underwent pathological evaluation at the end of the recovery period. `The mass median aerodynamic diameters for the aerosol tested ranged from 13 to 1.8 ym. Rats died following exposure to H-24678 at concentrations of 57 mg/m? or greater; deaths occurred within 1 day of exposure. Clinical signs of toxicity noted during the study included a diminished response to an auditory stimulus in rats during exposure to 57 or 120 mg/m. In addition, rats exposed at 120 mg/m? exhibited lethargy, lung noise, and labored breathing immediately following the exposure. One rat exposed at 23 mg/m? exhibited slight body-weight loss within 1 day of exposure (2.1% of pre-exposure body weight) but begantro egain weight by test day 2. Rats exposed at 57 mg/m? exhibited slight to moderate body-weight losses within 2 days of exposure (3.1 to 14% of pre-exposure body weight) but began to regain weight by test day 3. - Under the 57 mg/m'. coAncdciotridoinnsgotfothHiassksteuldly,Latbhoeraaptporryoxtiomxiactietylectlhaaslsicfoinccaetniotnrsa,tiHo-n2(4A67L8C)isfcoronHs-i2d4er6e7d8 tios be extremely toxic (ALC less than 0.08 mg/L) in male rats. --_--_----mmM Company Sanilized. DoesnotconTtSaCAiCnB 124675: Inhalation Approximate Lethal Concentration (ALC) in Rats Duo 8684 INTRODUCTION `The objective of this study was to determine a 4-hour inhalation approximate lethal concentration (ALC) of H-24678 in male rats. The ALC is defined as the lowest atmospheric concentration tested which caused the death of 1ormore exposed rats either on the day of exposure or within at least 14 days following exposure. The inhalation route of exposure was chosen based on the expected route of potential human exposure and was requested by the sponsor. MATERIALS AND METHODS A. Test Substance The test substance, H-24678, was supplied by the sponsoras an amber brown liquid. The test substance was assumed to be stable throughout the exposure phase of the study; no evidence of instability was observed. B. Animals `Young adult, male Crl:CD(SD)IGS BR rats were received from Charles River Laboratories, Inc., Raleigh, North Carolina. The rats were approximatel7y weeks old on the dayofarrival Rats have historically been used in safety evaluation studies for inhalation toxicity testing. The Crl:CD(SD)IGS BR rat was selected based on consistently acceptable health status and on extensive experience with the strain at Haskell Laboratory. C. Animal Husbandry 1. Quarantine and Animal Selection Rats were quarantined after arrival for approximately 1 week prior to testing. During the quarantine period, rats were weighed and observed for clinical signs of disease 3 times. Rats were obtained from the general population of stock rats released from quarantine and were selected for use on this study from those rats exhibiting a normal pattem of weight gain and no overt signs of disease. 2 Housing Rats were housed either singly or in pairs in stainless steel, wire-mesh cages suspended above. cage boards. BompanyS8aniizea, 085 Hot contain Tsogy -- 1.24675: Inhalation Approximate Lethal Concentration (ALC) n Rats Dupont8684 - 3. Animal Room Environment Animal rooms were maintained on a timer-controlled, 12-hour light/12-hour dark cycle. `Environmental conditions of the rooms were targeted to be within a temperature range of i2n3su+ff1iciCenatndmaagnreiltautdiveeahnudm/iodridtuyrartainogne toofh5a0ve+ a1d0v%e.rseElxycuafrfseicotnesd otuhtesviadleidtihteysoefrtahnegesstuwdeyr.e of 4. Identification cEaarcdharfaftiwxeadstaostshiegnceadgea. uPnriiqouret6o-edxigpiotsuirdee,ntitfhictaatiilonofneuamcbherratwhwiacshccoodrerdeswpitohndweadtetro-ainnsuomlubbelreed `markers so that individual rats could be identified after exposure. 5. Feed and Water `tEaxpcweapttedrurfirnogmeUxnpiotseudreW,aPteMrIDNeultraiwtairoen wInetreernaavtaiiolnaabl,leIandc. lCiebrittiufmi.ed Rodent LabDiet 5002 and 6. Health Monitoring Program AfosllsopweciinfgiperdoicnetdhuereHsasakreelpleLrafboorrmaetdorpyerainoidimcaalllhyeatlothenasnudreenthvaitrocnomnetnatmailnamnotniletvoerlisngarperboeglroawm, the _ those that would be expected to impact the scientific integrity of the study: + aWnadteorthsearmcpolnetsamarienaanntaslyzed for total bacterial counts, and the presence of coliforms, lead, Feed samples are analyzed for total bacterial, spore, and fungal counts. + Samples from freshly washed cages and cage racks are analyzed to ensure adequate sanitation by the cagewashers. Certified animal feed is used, guaranteed by the manufacturer to meet specified nutritional requirements and not to exceed stated maximum concentrations of key contaminants, including specified heavy metals, aflatoxin, chlorinated hydrocarbons, and organophosphates. The presence of these contaminants below the maximum concentration stated by the manufacturer would not be expected to impact the integrity of the study. `The animal health and environmental monitoring program is administered by the attending laboratory animal veterinarian. Evaluation of these data did not indicate any conditions that affected the validity of the study. D. Study Design _ `Three groupsof 6 male rats each were exposed to aerosol atmospheres of H-24678 in air. Rats were exposed nose-only foar single, 4-hour period. BompanySanitized. Dossnot contain Tsca co; _-- 24676: Inhalstion Approximate Lethal Concentration (ALC) in Rats DuPont8684 ~ Rats were approximately 8 or 9 weeks old and weighed between 211 and 316 grams at the time of exposure. Rats were observed for mortality and response to alerting stimuli during each exposure and observed for mortality and clinical signs of toxicity immediately after they were removed from the restrainers following exposure. During a 14-day postexposure period, all surviving rats were observed each day for mortality,andwere weighed and observed for clinical signs of toxicity daily until weight gains were observed then once weekly thereafter. At the end of the recovery period, all surviving rats were sacrificed by carbon dioxide asphyxiation. Three rats from the. 57 mg/m group underwent a gross pathology examination. Tissues were taken from these 3 rats and further processed for microscopic pathology examination. E. Inhalation Exposure System 1. Aunosphere Generation Chamber atmospheres were generated by atomization of the test substance in air with a Spraying Systems Nebulizer. The test substance was metered into the nebulizer with a Harvard Apparatus `model 22 Syringe Infusion Pump. Filtered, high-pressure air, metered into the nebulizer by a Brooks model 1355 Sho-Rate Rotometer, carried the resulting atmosphere into the exposure: chamber. Chamber concentrations of test substance were controlled by varying the test substance --~ feed rate to the atmosphere generator. Test atmospheres were exhausted through a dry-ice cold trap followed by an MSA charcoal/HEPA filter cartridge prior to discharge into the fume hood 2. Chamber Construction and Design `The exposure chamber was constructedof glass (cylindrical) with a nominal intemal volume of 29L. A polycarbonate baffle inside the chamber promoted uniform chamber distribution of the test atmosphere. 3. Exposure Mode During exposure, rats were individually restrained in perforated stainless steel cylinders with conical nose pieces. `The restrainers were inserted into a polymethylmethacrylate faceplate which was attached to the exposure chamberso that only the nose of each rat extended into the chamber. F. Characterization of Chamber Atmosphere 1. Test Substance Sampling and Analysis --- "The atmospheric concentration of H-24678 was determined by gravimetric analysis at approximately 30-minute intervals during each exposure. Known volumes of chamber I `Company Sanitized. Does nol contain TSCA CB 1.24678: Inhalation Approximate Lethal Concentration (ALC) in Ras DuPont 8684 ~ atmosphere were drawn from the sampling port through a 25 mim filer cassette that contained a pre-weighed Gelman glass fiber (Type A/E) filter. The filters were weighed on a Cahn model Ca-tm3o0spMhiecrroibcacloanncceent,raptliaocneodfiHna-d2e4s6s7i8cawtaosr cfaolrc1uldaatye,d afnrdomretwheeidgihfefderoennctehienstahmeeprbea-laancned.poTsth-e `sampling dry filter weights divided by the volume of chamber atmosphere sampled. A sample to determine particle size distribution (mass median aerodynamic diameter and percent particles less than 10 um diameter) was taken during each exposure with a Sierra Series 210 Cyclone Preseparator/Cascade Impactor and Sierra Series 110 Constant Flow Air Sampler." 2. Environmental Monitoring Chamber airflow was set at the beginning of each exposure to achieve at least 12 air changes per hour. The airflow was monitored continually with acalibrated Brooks model 1355 Sho-Rate Rotometer and recorded initially and whenever changes were made during the exposure. Chamber airflow was inadvertently not recorded during the 23 or 120 mg/m? exposures. Chamber temperature was targeted at 22. 2C. The temperature was monitored continually with a glass alcohol thermometer and recorded 4 times during each exposure. Chamber relative humidity was targeted at 50 + 10%. The relative humidity was measured with an Omega model RHS100C Digital Psychrometer and recorded 3 times during each exposure. Chamber oxygen concentration was targeted to be at least 19%. The oxygen concentration was measured with a Biosystems model 3100R Oxygen Analyzer and recorded 3 times during each exposure. G. Anatomic Pathology Evaluation `The 3 surviving animals in the 57 mg/m group had the lungs, trachea, pharynx/larynx, and nose grossly examined at sacrifice. In addition, th tissues from these rats were processed for histology and microscopically examined. --~ Company Sanitized," po,Does notcontain sc cup Ee H-24678: Inhalation Approximate Lethal Concentration (ALC) in Rats DuPont 8684 RESULTS AND DISCUSSION A. Exposure Conditions (Table 1) Animals were exposed to H-24678 at concentrations of 23, 57, or 120 mg/m'. The. concentrations reported are based on the solids component of the test substance. The solvent `component of the test substance was measured by gas chromatography during each exposure `The mean concentration of solvent measured during the study was less than 53 ppm. This concentration is negligible compared to the reported acute inhalation toxicity of the solvent. Therefore, the concentrations reported do not include the solvent component of the test substance. The atmospheres generated in this study were considered to be respirable in rats, as the mass median aerodynamic diameters (MMAD) ranged from 1.3 to 1.8 pm. Chamber temperature ranged from 22 to 23C, chamber relative humidity ranged from 40 to 45%, chamber airflow was 20 L/min, and the oxygen concentration was 21%. Chamber airflow `was inadvertently not recorded during the 23 or 120 mg/m? exposures. -- B. Mortality, Clinical Signs, and Body Weights Deaths occurred at concentrations of 57 mg/m? H-24678 or greater. Threeof 6rats died at a `concentration of 57 mg/m? and all rats died at 120 mg/m. Rats died within 1 day of exposure. Clinical signsof toxicity noted during the study included a diminished response to an auditory. stimulus in rats during exposure to 57 or 120 mg/m. In addition, rats exposed at 120 mg/m? exhibited lethargy, lung noise, and labored breathing immediately following the exposure. One rat exposed at 23 mg/m? exhibited slight body-weight loss within 1 day of exposure (2.1% of pre-exposure body weight) but began to regain weight by test day 2. Ras exposed at 57 mg/m? exhibited slight to moderate body-weight losses within 2 daysofexposure (3.1 to 14% of pre-exposure body weight) but began to regain weight by test day 3. C. Anatomic Pathology Evaluation (Appendix A) 1. Gross Pathology There were no gross pathologic observations in any of the tissues examined. _ 2. Microscopic Pathology `There were no microscopic pathologic observations in any of the tissues examined. - Company Sanifira Ae .. 24678: nhation Approximate Lethal Concenration (ALC) in Rats Dupont8684 - CONCLUSIONS Under the conditions of this study, the approximate lethal concentration (ALC) for H-24678 is. 57 mg/m. According to Haskell Laboratory toxicity classifications, H-24678 is considered to be extremely toxic (ALC less than 0.08 mg/L) in male rats. RECORDS AND SAMPLE STORAGE Specimens(ifapplicable), raw data, and the final report will be retained at Haskell Laboratory, Newark, Delaware, or at Iron Mountain Records Management, Wilmington, Delaware REFERENCES 1. CSearliccusla2t1i0onAdmebsicernibteCdaisncSaideerrIamIpnascttrourmsenatnsd, CIyncc.l,oBnuellPerteisncp7a-r7a9t-o2rs1.9IM, Instruction Manual. _ - Compan ' Saitized. Does notcontain Tscacey H-24676: Inhalation Approximate Lethal Concentration (ALC) in Rats DuPont8684 TABLE | CHARACTERIZATION OF TEST ATMOSPHERES AND ASSOCIATED ANIMAL MORTALITY "AEROSOL CONCENTRATION (mg/m)* AEROSOL SIZE MORTALITY MMAD Percent| (# deaths/ Mean SD. Range n| (um GSD <10pum'| #exposed) 23 37 14-27 8| 14 19-100 0/6 57 13 2.77 8 18 20 -100 3/6 120 1110-140 8] 13 19-100 6/6 a pReerpreexspeonstusret.heVmaelaune,ssrteapnrdeasredntdedvyiawteioingh(t5oDf)t,hantestrasnugbestfaonrceeaacnhdeaxpeosruerpeo,rtbeads0ed2onsingnsifaimcpalnets b TfihgeurMesMAD (Mass Median Aerodynamic Diameter) is based on 1 particle size sample taken GdueroimnegtreiacchSetaxnpdoasrudreDeviation. 0 Percent arosol mass having aerodynamic equivalent diameters of les than 10 um. - CompanySanitized. DoesnotcontainTSCAC . To H-24678: Inhalation Approximate Lethal Concentration (ALinCRa)ts DuPont 8684 -- APPENDIX A Individual Animal Pathology Data _ Company Sanitized. Does not contain TSCA C.. Be L1L224367687:5InInhhaallaotiioonnAApprpoxipmatreLLeeothhaallxCCoonnicceenntmtrraattaiioonnt((AALeLCC))iinnRRaattss ______ Individual Animal Pathology Date Concentration: 57 mg/m Sex: wales Animal Ref Microscopic & Macroscopic Findings eet KATnieilrnmlaielndaloinssDsaiacgyrnief+dic1eo4ff from necropsy Gross Pathology Ho MaTcGrSos,copTiRcACHAEbRm,ommPaRlAiRtEyNXoLbAseRrAve,d H OSE Histopathology No MLiUcNrGoSs,copTiRAcCHAEbRn,ormPaRlAiRtEyN OLbsAeRrv,ed N: OSE sane TKAneiirlmmlaielndaloinsSDsaaicygrnief:dic1eo4ff from neczopsy Gross rathology No M`aLcUrNoGsS,copTiRcACHAEbRn,ormPaRlAiRtYyA/oLbAsReYsNvYe,d N osE Histopathology Ho MiTcONrGoSs,copTiRcACKAEbRn,ormPaRAlRiEtNyS,OLbsAerRv,ed NOSE esaons nTKeiirlnmlieandalo{5nssDaiacgrynief:dic1eo4ff from necropsy Gross pathology : No NaNcGrSos,copTiRcACHAEbRn,orPmAalRiRtYy oLbsAerRv,ed NOSE Histopathology No MiLcUNrGoSs,copTiRcACHAEbAn,ormPaHlAiRtVyNXO/bLsAeRrIvXe,d NOSE DDuuPpooms866884s --_----m `CompanySaniizeq,Dpoe,s otcontainTSCAcay as