Document 7OMDq3R0gwYBMvEnxZNDpYN2R

TRADE SECRET DuPont HL-1997-00598 AR226-3079 Study Title I nhalation Approximate Lethal Concentration (ALC) en R ats Laboratory Project ID: HL-1997-00598 Author: David P. Kelly, B.S. Study Completed on: August 29,1997 Performing Laboratory: E i. du Pont de Nemours and Company Haskell Laboratory for Toxicology and Industrial M edicine Elkton Road, P.O. B ox 50 Newark, Delaware 19714-0050 Medical Research Numbers not contain TSCA CBI Sanitized. Does Company Page 1 o f 11 alation Approximate Lethal Concentration (ALC) in Rats DuPont HL-1997-00598 Substance Tested: Svnonvm s/C odes: GENERAL INFORMATION Haskell Num berfsl: CAS Registry Num berfsi: H-22632 2 2 3 8 7 ,2 2 6 3 2 Physical Characteristics: Stability: water clear liquid The test substance appears to be stable under the conditions o f the study; no evidence o f instability was observed. Company Sanitized. Does not contain TSC CBI ilatioD Approximate Lethal Concentration (ALC) in Rats DuPont HL-1997-00598 fi GENERAL INFORM ATION (Continued) Sponsor: E.I. du Pont de Nemours and Company Wilmington, Delaware 19898 U .S .A . Study Initiated - Completed: February 13, 1997 - (see report cover page) In-Life Initiated - Completed: February 13, 1997 - July 9, 1997 A ll original data and the original o f this final report w ill be retained at Haskell Laboratory Newark, Delaware, or at Iron Mountain, 200 Todds Lane, Wilmington, Delaware 19802. ' Company Sanitized, Does not contain TSCA CBf lation Approximate Lethal Concentration (ALC) in Rats DuPont HL-1997-00598 SUMMARY Groups o f six male Crl:CD^^!D)BR rats each were exposed nose-only for a single four-hour period to a water mixture o ^ ^ ^ f c B f t i - 2 2 3 8 L and H-22622) in air. Test atmospheres were generated by aerosolizlRon, and concenftations o M ^ ^ ^ I ' e r e measured by gravimetric filter analysis. Aerosol concentrations werereported asw et and dry aerosol. Rats were weighed and observed for clinical signs o f toxicity during a 14- to 15-day recovery period. Rats were exposed to atmospheres it wet aerosol concentrations o f 320, 330, or 570 mg/m3, corresponding to dry aerosol concentrations o f 210,270, or 420 mg/m3. The mass median aerodynamic diam eter^fth eu ero so ls tested were 5.0,2.3, and 3.0 fim, respectively. Rats died following exposure concentrations o f 320 and 570 mg/m3 (wet), corresponding to 210 and 420 mg/m (dry). Deaths occurred immediately after exposure or within two days. No deaths occurred after exposure at concentrations o f 320 mg/m3 (wet) 270 mg/m3 (dry). h L^der the conditions o f this study, the approximate lethal concentration (ALC) for aerosolized 6 M H f e l H H ^ p 3 2 0 mg/m (wet aerosol), 210m g/m 3 (dry aerosol). On an acute inhalation basis and based on the dry aerosol concentratioE flfeH H H M M H ls considered to be moderatelv toxic (ALC 200-800 mg/m3). Conti &T Inhalation Approximate Lethal Concentration (ALC) in Rats DuPont HL-1997-00598 SIGNATURES Reviewed and Approved for Issue: ,_ David F^Kly, B.S. Research-Toxicologist Study Director STUDY PERSONNEL The following individuals were responsible for conduct o f the study: Management: Scott E. Loveless, Ph.D. Study Director: David P. Kelly, B.S. Primary Technicians: M. Ahmad Pulliam, B.S. Dwayne A. Lavoie, B.S. Toxicology Report Preparation: Maryanne M. Wilford, B.A. Date 2( The following individual was responsible for assessing the health status o f the animals on study: Laboratory Veterinarian: Charles E. Cover, V.M.D. !||P r Company Sanitized. o e s not contain Ts c a c b / -5 - talation Approximate Lethal Concentration (ALC) in Rats DuPont HL-1997-00598 INTRODUCTION The objective of this s tu & n y ^ ^ g te rm m e a four-hour inhalation approximate lethal concentration (ALC) o p H H n male rats. The ALC is defined as the lowest atmospheric concentration tested which caused the death o f one or more exposed rats either on the day of exposure or within at least 14 days following exposure. The inhalation route o f exposure was chosen based on the expected route o f potential human exposure and was requested by the sponsor. Data for this study was obtained using two protocols: an inhalation screen protocol (SOP AI005-T) and an approximate lethal concentration protocol (SOP AI001-T). MATERIALS AND M ETH ODS A. Test Substance The test | | b s t a n c f i B p | | i U l --222387 and H-2'2632)_-wwaas supplied by the sponsor as a mixture s was diluted t ^ j j ^ E t Haskell Laboratory to facilitate spraying. B. Animals Young adult, male, Crl.'CD (SD)BR rats were obtained from Charles River Breeding Laboratories. The rats were approximately seven to eight weeks old upon arrival. Rats have historically been used in safety evaluation studies for acute inhalation toxicity testing. The Crl:CD (SD)BR rat has been chosen based on consistently acceptable health status and the extensive experience w ith the strain at this laboratory. C. Anim al Husbandry 1. Quarantine and Animal Selection Rats were quarantined after arrival for approximately six days prior to testing. During the quarantine period, rats were weighed and observed for clinical signs o f disease three times. Rats were obtained from the general population o f stock rats released from quarantine and were selected for use on this study from those rats exhibiting a normal pattern o f weight gain and no overt signs o f disease. 2. Housing Rats were housed either singly or in pairs in suspended, stainless steel, wire-mesh cages. -6 - ilation Approximate Lethal Concentration (ALC) in R a ts________ DuPont HL-1997-00598 3. Animal Room Environment Animal rooms were maintained on a timer-controlled, 12-hour light/12-hour dark cycle. Environmental conditions o f the rooms were targeted to be within a temperature range of 23 1c and a relative humidity range o f 50 10%. Excursions outside these ranges were o f insufficient magnitude and/or duration to have adversely affected the validity o f the study. 4. Identification Each rat was assigned a unique six-digit identification number which corresponded to a numbered card affixed to the cage. Prior to exposure, the tail o f each rat was color-coded or numbered with water-insoluble markers so that individual rats could be identified after exposure. 5. Feed and Water Except during exposure, Purina Certified Rodent Chow #5002 and tap water from United Water Delaware were available ad libitum. 6. Health Monitoring Program Haskell Laboratory has an animal health monitoring program. The following procedures are performed periodically: W ater samples are analyzed for total bacterial counts, and the presence o f coliforms lead and other contaminants. 5' Feed samples are analyzed for the presence o f bacteria and fungi. Samples from freshly washed cages and cage racks are analyzed to ensure adequate sanitation by the cage washers. Haskell Laboratory uses certified animal feed. The feed is guaranteed by the manufacturer to meet specified nutritional requirements and to be free o f a list o f specified contaminants. The animal health monitoring program is administered by the laboratory animal veterinarian. Data are maintained separately from study records Mid are not included in the final report. Evaluation o f these data did not indicate any conditions that affected the validity o f the study. D. Study Design Three groups o f six male rats each were exposed to atmospheres o exposed nose-only for a single, four-hour period. in air. Rats were Rats were approximately eight to nine weeks old and weighed between 255 to 289 grams at the time o f exposure. Rats were observed for mortality and response to alerting stimuli during the exposure and observed for mortality and clinical signs o f toxicity immediately after they were removed from Company Sanfffeed Does not contain TSCA CBI 7- - F alation Approximate Lethal Concentration (ALC) in Rats DuPont HL-1997-00598 the restrainers following exposure. During a 14- to 15-day postexposure period, all surviving rats were observed each day for mortality, and were weighed and observed for clinical signs o f toxicity at least twice weekly. At the end o f the recovery period, all surviving rats were sacrificed by carbon dioxide asphyxiation and discarded. E. Inhalation Exposure System 1. Atmosphere Generation Chamber atmospheres were generated by aerosolization o f l ^ H K n air with a Spraying Systems nebulizer. The test substance was metorecUntcHtenebulizer with a Harvard Apparatus model 22 Syringe Infusion Pump. Filtered houseline air, introduced at the nebulizer, atomized the te&Lsubstajrce^id earned the aerosol into the exposure chamber. Chamberconcentrations c t t ^ ^ p | M v e r e controlled by varying the test substance feed rate to the atmosphere generator. *' Test atmospheres were exhausted through a bubbler, a dry-ice cold trap, and an MSA charcoal/HEPA filter cartridge prior to discharge into the fume hood. 2. Chamber Construction and Design The exposure chamber was constructed o f glass (cylindrical) with a nominal internal volume of 34 L. A dispersion plate at the chamber inlet promoted uniform chamber distribution o f 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 the polymethylmethacrylate faceplate o f the exposure chamber so that only the nose o f each rat extended into the chamber. F. Characterization o f Chamber Atmosphere 1. Test Substance Sampling and Analysis The atmospheric concentration { m ^ v a s determined by gravimetric analysis at approximately 30-minute intervals during exposure. Known volumes o f chamber atmospheres were drawn from the reference sampling port through a 25 mm filter cassette that contained a pre-weighed Gelman glass fiber (Type A/E) filter. After samples were collected, the filters were immediately weighed to determine the wet aerosol concentration. After overnight drying in a dessicator, the filter samples were weighed to determine the dry aerosol weight. The filters were weighed on a Cahn model C-30 Microbalance. The atmospheric concentration was calculated from the difference in the pre- and post-sampling filter weights d i v M e c ^ ^ h ^ ^ volume o f chamber atmosphere sampled. Company sanitized. o e s not contain Ts c a CBI lation Approximate Lethal Concentration (ALC) in Rats DuPont HL-1997-00598 Samples to determine particle size distribution (mass median aerodynamic diameter and percent particles less than 3 fim diameter) were taken with a Sierra Series 210 Cyclone Preseparator/Cascade hnpactor and Sierra Series 110 Constant Flow Air Sampler.05 2. Environmental Monitoring Chamber airflow was set at the beginning o f each exposure to achieve at least 12 air changes per hour. The airflow was monitored continually with a calibrated Brooks model 1355 Sho-Rate Rotometer and recorded one to four times during exposure. Chamber temperature was targeted at 22 2C. The temperature was monitored continually with an Omega model 650 Thermocouple Thermometer and recorded two to five times during exposure. Chamber relative humidity was targeted at 50 10%. The relative humidity was measured with an Omega Digital Psychrometer and recorded two to three times during exposure. Chamber oxygen concentration was targeted to at least 19%. The oxygen concentration was measured with a Biosystems model 3 100R Oxygen Analyzer and recorded two to three times during exposure. Co, ,pany Sanitized Ooes not cons * se a e g | -9 - alation Approximate Lethal Concentration (ALC) in Rats DuPont HL-1997-00598 \ RESULTS A. Exposure Conditions (Table 1) Anima! SJWf re exp,osed i H P H t concentrations o f 210,270, or 420 mg/m3. The aerosols generated during the 270 mg/m exposure were considered to be marginally respirable in rats as the mass median aerodynamic diameters (MMAD) was 5.0 un. TABLE 1 CHARACTERIZATION OF TEST ATMOSPHERES AND ASSOCIATED ANIMAL MORTALITY AEROSOL CONG iNTRATION img/m3)a AEROSOL SIZE MORTALITY Wet Filters Mean 1 S.D. Range Dry Filters Mean _S ^j Range MMAD Percent (# deaths/ n (fimf GSDC <3 ttmd # exposed) 320 98 2 1 0 -4 8 0 270 87 1 6 0 -4 0 0 8 5.0 3.1 330 35 290 - 390 210 25 190 - 260 8 2.3 2.9 66 570 500 2 1 0 -1 8 0 0 420 250 150 - 940 12 3.0 2.3 53 0 /6 5 /6 6 /6 a Represents the mean, standard deviation (S.D ,)rand range for each exposure, based on n samples per exposure. Values are reported to two significant figures. F b The MMAD (Mass M edian Aerodynamic Diameter) is based oni o<ne particle size sample taken during each exposure. c Geometric Standard Deviation. d Percent aerosol mass having aerodynamic equivalent diameters o f less than 3' /mi Chamber temperature ranged from 20 to 23C, chamber relative humidity ranged from 76 to 86%, chamber airflow ranged from 15 to 25 L/min, and the oxygen concentration was 21%. The / chamber relative humidity was high because o f the aerosolized water content o f the test mixture. M ortality, C linical Signs, and Body W eights Deaths occurred ai two days o f exposure. oncentrations o f 210 mg/m3 or greater. Rats died within A diminished alerting response was observed near the end o f the 270 mg/m3 exposure. Following exposures, when the rats were removed from the chambers, the clinical signs of toxicity observed included irregular respiration, lung noise, lethargy, and wet perineum. " ''""'" V Sa0ilfeed. Doss 001contain tscacsi 10 lalation Approximate Lethal Concentration (ALC) in Rats DuPont HL-1997-00598 Surviving rats showed slight to severe weight loss on the day after exposure. Normal weight gain rates were resumed within one week after exposure. C O N C L U SIO N hi these studies, the difference in aerosol concentrations between a lethal level, where 5 o f 6 rats died, and a non-lethal level was minimal. It is speculated that the reason rats died at 330 mg/m3 (wet aerosol), 210 mg/m (dry), and none died at 320 mg/m3 (wet), 270 mg/m3 (dry), was that the aerosol particle was larger (5 pm) in the 270 mg/m3 experiment and smaller in the 210 mg/m3 experiment. Although both aerosol sizes are inhalable, the smaller aerosol would reach deeper in the lung, and possibly cause more acute damage. From an industrial health standpoint it is recommended that production o f small aerosols o f this material be minimized. Jnder the conditions o f this shidy, the approximate lethal concentration (ALC) for aerosolized 320 mg/m (wet aerosol), 21feng/m^_(dry aerqgpl). On an acute inhalation (" lo o m Z T Sl ^ moderately REFERENCES 1. Calculation described in Sierra Instruments, Inc., Bulletin 7-79-219IM, Instruction Manual: Series 210 Ambient Cascade Impactors and Cyclone Preseparators. Company Sanitized. Does not contain TSCA CBi