Document dOdgE67qYgQVd1nOgpj5Y2BB

AR226-2990 Study Titl STATIC, ACU TE, 96` HOUR LC50 O FH -19520 T O FA T H E A D MINNOW S, Pim ephales prom elas Data Requirem ent O rganisation for Econ om ic Co-O peration and D evelopm ent (O ECD ) G u ideline for Testing Chem icals: 203 Author Keith B. Pierson, Ph.D. Study Com pleted On May 25.1993 Perform ing Laboratory E. I. du Pont de Nem ours and Com pany H askell Laboratory for Toxicology and Industrial M edicine Elkton Road, Post O ffice Box 50 Newark, Delaw are 19714 Laboratory Project ID H askell Laboratory Report No. 593-92 Page 1 of 19 smpany Sanitized. Does not contain TSCA CBI Du Pont HLR 593-92 G O O D LA B O R A T O R Y PRACTICE CO M PLIAN CE STATEM EN T T h is study w as roridu cted according to E P A F iF R A and T S C A G ood Laboratory P ra ctice Standards (4000= 1160 and 40 C F R 7 9 2 .O E C D Prin ciples o f Good Laboratory P ra ctice (G(81 )30(Final), Annex 2), a n d M A F F Japan G ood Laboratory P ractice Standards (59 NohSan No. 3850) with the two exceptions noted below . Sclutrility an d stability of the test substance in dilution w ater and under storage conditions w ere not determ ined. Concentrations of the test su bstan ce in dilution w ater w ere m easured, but these m easurem ents w ere not m ade in accordance w ith G o o d Laboratory Practice Standards. Protocol am endm ents and/or S O P deviations appear in Appendix I. A reas of noncom piiance (if any) are a lso docum ented in the study records. No deviations existed that affected the scien tific validity of the study. Subm itter. E . I. du Pont de Nem ours and Com pany Sponsor: D u Pont Chem icals E . I. du Pont de Nem ours and Com pany STU D Y DIRECTOR: Keith B. Pierson, Ph.D. R esearch Toxicologist Environm ental S cien ces SfAV m 3 M o/Day/Year 2 Company Sanitized. Does not contain TSCA CBl Du Pont HLR 593-92 co to to oo oo oo oo TABLE O F CONTENTS Page G ood Laboratory Practice Com pliance S ta te m e n t......................................................2 T able of C o n te n ts............................................................................ 3 G eneral Inform ation......................................................................................................... 4 Su m m ary......................................................................................... ............................... 6 Q uality A ssu ra n ce D ocum entation................................................................................. 7 |a P u i ^ p o s o II. R e c o r d s ...........................................................................................................8 III. M aterials and M ethods A. T est Substance...... B. D ilution W a te r....... C . A nim al C u ltu re ...... D. T est M eth o d s......... E. C h em ical A n a ly s is . F. Statistical A n a ly s is . IV. R e su lts and D iscu ssio n ................................................................................... 10 V. C o n c lu s io n ......................... 10 VI. Literature C ite d ............ ..................................................................................10 VII. T a b le s. 1. C h em ical Characteristics of H askell 2. M easured Concentrations of H-19520 96-Hour, Static, Unaerated T est ... 3. W ater Chem istry of the Control at Test S tart........... 4. P h ysica l and Chem ical C haracteristics of H -19520 T est S o lu tio n s......................................................... S' 5. O bserved M ortality ofF ath ead M inn ow s, Pim ephales prom elas, Exp osed to H - 1 9 5 2 0 p H | ^ H lf o r 96 H ours in a Static, Unaerated T e s t.................. . ................. ........... 6. O bserved Effects of H - 1 9 5 2 0 N ^ B H H H p n Fathead Minnows, Pim ephate^ rom elasJoi 96 H ours in a Static, Unaerated T e st-- H - 1 9 5 2 0 H H H H M B jC o n c e n tra tio n s C au sing M ortality to 50% (L cS o ^ ffa tn e a a fw in n o w s, Pim ephales prom elas, at S p e cific Tim e Intervals....................................................................... 11 11 12 13 14 16 17 18 VIII. A p p e n d ix........................................................................................................ 19 1. Protocol Am endm ents and S O P D eviation s............................................ 19 3 Company Sanitized. Does not contain TSCA CBI Du Pont H L R 593-92 GENERAL INFORMATION i : ! Haskei! Number; Euri& i H-19520 4 not contain TSCA CB! Com position; D u Pont H L R 593-92 Sponsor: Study Initiated/Completed: inrlifeL&riy Initiated/Com pleted: Du Pont C hem icals E. I. du Pont de N em ours and Com pany W ilm ington, D elaw are 19898 7-17-92 / 5-25-93 7-22-92 / 8-7-92 j 5 (Sompany Sanitized. Does? noi contain TSC CBf Du Pont HLR 593-92 STATIC. A C U T E , 96-H O U R LC5Q O F H-19520 T O FA T H EA D M INNOW S, PJmephsies prom elas SUM M ARY H-19520 exhibited slight toxicity in a 96-hour, unaerated, static, acute test using fathead minnows, Pim ephales promelas. Nom inai H -19520 concentrations were 0, 7 5 .4 ,1 1 6 ,1 7 9 ,2 7 5 ,4 2 3 ,6 5 0 , and 1000 m g /L M easured concentrations at 96 hours were 0 .1 4 ,7 2 .9 ,9 1 .7 ,8 6 .5 ,2 4 3 ,3 4 3 ,5 4 0 , and 907 mg/L. The 96-hour LC 50 based on m easured concentrations at 96 hours w as 276 mg/L with a 95% confidence interval of 237 - 317 m g/L W ork By: Z t b d w p tiu s* * * im p Robert W. Peterson, B. A. Toxicology A ssociate x J w j 93 M o/D ay/Y ear W ork By: J & f e b ' '*<*>*. (reffreYT. Turner, Sr. Toxicology A ssociate M o/Day/Year THIS R E P O R T IS A PP R O V E D FO R ISSUE. 4. Keith B. Pierson, Ph.D . Research Toxicologist and Study Director Environm ental S cie n ce s S /HA-t iM Z M o/D ay/Y ear 6 Company Sanitized. Does not contain TSCA CB8 Q U A LITY A S S U R A N C E D O CU M EN TATIO N (H -1 9 5 2 0 ) D ates o f Inspection: Conduct - 7/22/92 Records, Report(s) - 4/21-22/93 Fin ding s Reported to: Study Director - 4/23/93 M anagem ent - 5/24/93 Reported Donna R. Holt Q uality A ssu ran ce Auditor s la n ts 'D a li m 7 $ m m t Sanitized, Does not contain TSCA CBt Du Pont HLR 593-92 I. P U R P O S E The goal of this study w as to asse ss the acute toxicitv of H -19520 to fathead m innows, Pim ephales prom elas. II. R E C O R D S A ll data and the final report w ill be m aintained either at H askell Laboratory for T oxicology and Industrial M edicine (Newark, D elaw are) o r be archived at the Du Pont R eco rd s M anagem ent Center (W ilm ington, D elaw are). III. M A T E R IA L S A N D M E T H O D S A. Test Substance T he test substance is described under G eneral Information. A sto ck solution w as prepared by dissolving the test substance in dilution w ater (see below) and stirring for 30 m inutes. Visual inspection suggested that the test substance w as in solution in the stock. Aliquots of the sto ck solution w ere added to dilution w ater to prepare individual test solutions. E ach of the ab ove solution s w as kept in a container with minim al head space sealed with S aran Wrap to avoid potential lo ss of the test substance through volatility. Stab ility and solubility of H-19520 in dilution water and under storage conditions w ere not determ ined. 8. Dilution Water W ater (Table 1) originated from the H askell Laboratory well. C . Animal Culture Fathead m innow s, Pim ephales promelas, w ere reared in continuously flowing w ell water at H askell Laboratory. Adult fish resided in 39 L aq uaria at approxim ately 25C and were fed frozen brine shrim p Artem ia sn. Golden G ate Brand, Kordon D ivision, Novalek, Inc., Hayward, C A ) g enerally tw ice daily, once on Saturday and o nce on Sunday. Eggs w ere deposited on polyvinyl chloride tile s coated with sand. T iles with eggs w ere aerated and incubated at approxim ately 25C in a w ater bath. Larvae w ere reared in 39 L aquaria at approxim ately 21 C and were fed newly hatched Artem ia sp. (Aquarium Products, G len Burnie, MD) generally tw ice d aily on w eekdays and d aily on w eekends. Ju ve n iles w ere held either in 272-L circular, fib erg lass tanks or in 39 L g la ss aq uaria (23-cm depth; 50 (L) x 25 (W) x 31 cm (H)) at approxim ately 21 C and w ere fed newly hatched Artem ia sp. and/or frozen brine shrim p generally tw ice d aily on weekdays, once on Saturday and o nce on Sunday. Sickn ess, injury, and abnorm ality were not observed. A nim als were identified by labels on aquaria and test vessels. 8 Company Sanitized. Does not contain TSCA CBt Du Pont HLR 593-92 D. Test Methods [1] Seven test concentrations and a control were used. Nom inal concentrations were 0 ,7 5 .4 ,1 1 6 ,1 7 9 ,2 7 5 ,4 2 3 ,6 5 0 , and 1000 Control fish ranged from 1.1 to 1.6 cm standard length (mean 1.3 cm) and 0.019 to 0.056 g wet weight (mean 0.030 g). Ertenm eyer g la ss flasks [4.4 liter; 21 (base) x 35 (height) cm] containing 4.4 L of test solution (35-cm depth) were em ployed a s test cham bers. Each flask w as totally filled to m inim ize head space and w as covered with Saran Wrap to prevent lo s s of potentially volatile test substance com ponents. Ten fish w ere added to each replicate using random num bers (one replicate per concentration, total 10 fish p e r concentration); Control loading w as 0:067 g/L at test conclusion. T e st solution s ranged from 20.8 to 21.4C (mean 21.0C). Fish w are not fed 27 hours prior to nor during the te s t A photoperiod of 16 hours light (161 to 215 Lux) versu s 8 hours darkness w as em ployed with 25 m inutes of transitional light (1 - 2 Lux) preceding and following the 16 hour light interval. M ortality and behavioral effects w ere noted daily. D issolved oxygen (YSI M odel 54A, Yellow Springs Instruments, Yellow Springs, O H ) and pH (Com ing Model 61OA, Com ing Scientific Instruments, M edfield, M A) were m easured in the water control and all test concentrations before fish w ere added at the beginning of the test, daily thereafter and at total m ortality. Tem perature w as m easured (Am erican Society of Testing M aterials registered m ercury thermometer) in the water control and all test concentrations before fish w ere added at the beginning of the test, daily thereafter, at the end of the test and at total mortality. Total alkalinity (Hach kit, Hach Chem ical Com pany, Loveland, CO ), ED T A hardness (Hach kit, H ach Chem ical Com pany, Loveland, C O ), and conductivity (YSI M odel 33, Yellow Springs Instruments, Y ellow Springs, O H) of the control water were m easured at the beginning of the test. A continuously-recording thermometer in a control vessel w as used to ch e ck for tem perature variation during the 96-hour test. E. Chemical Analysis A n alyses w ere conducted after the test ended at the Du Pont Jackson Laboratory at Cham bers W orks, Deepwater, NJ. Sam ples and standards were com pletely com busted in a hydrogen/oxygen W ickboid Torch. The resulting w ater w as then analyzed for total fluoride concentration using an Orion fluoride ion selective electrode. Concentrations were calculated from the standards using either a three or four point calibration curve. F . S ta tis tic a l A n a ly s is The 24- and 48-hour LC 5 0 s were calculated by the S A S Probit method [2]. The 72- and 96-hour LC 5 0 s were calculated by using the moving average angle technique [3]. 9 Sfitnpapx Sanitized-Dges not contain TSCA CBf Du Pont HLR 593-92 IV. R E S U L T S A N D D IS C U S S IO N A rangefinding te st with five fish per concentration w as conducted with a control and test concentrations of 0 .1 ,1 , TO. 100, and 1000 m g/L M ortalities w ere 0 ,0 ,0 , 0 . 0. and 100%, respectively. Tw o definitive stu d ies w ere conducted. D ata from th e first were considered invalid becaMS o f low dissolved oxygen concentrations. Data from the secon d a re reported herein. N om inal concentrations in the definitive test were 0 ,7 5 .4 ,1 1 6 ,1 7 9 ,2 7 5 ,4 2 3 , 650. and 1000 m o /L M easured concentrations at 96 hours were 0 .1 4 ,7 2 .9 , 91 7, 8 6 .5 ,2 4 3 ,3 4 3 , 540, and 907 mg/L (Table 2). T he discrepancy between the m easured and nom inal concentrations m ay stem from use of a stock solution in w hich the test substance w as not truly in solution. A ll chem ical and p h ysical param eters in the final definitive test (Tables 3 and 4) w ere within expected ranges except on day 4. D issolved oxygen concentrations dropped a s low a s 53% (minimum) o f saturation at 72.9 mg/L. However, these observations w ere not o f biological significance sin ce fathead m innow s ca n su rvive at m uch low er d issolved oxygen concentrations for extended periods. B ehavioral o b servations appear in Table 6. The 96-hour LC50 based upon m easured concentrations at 96 hours w a s 276 mg/L with a 95% confidence interval o f 237 - 317 m g/L (Tables 5 and 7). V. C O N C LU SIO N H -19520 w as slig h tly toxic to fathead minnows, Pim ephales promelas, in a 96-hour, static, unaerated test. VI. LIT E R A T U R E C ITED 1. O rganisation for Econom ic Co-O peration and Developm ent (O ECD ). G uideline fo r T esting Chem icals: 203. 4 April 1984.23 2. Finney, D. J. 1971. Probit Analysis, 3rd Ed., Cam bridge University P ress, p. 333. 3. Peltier, W J ' and C . I. W eber, Eds. 1985. M ethods for m easuring the acute toxicity of effluents to freshwater and m arine organism s. EPA/600/4-85/013. United States Environm ental Protection Agency, Environm ental M onitoring and Support Laboratory, Cincinnati, O hio. Appendix E. 10 Company Sanitized. Does not contain TSCA CBI Du Pont HLR 593-92 STATIC, ACU TE, 96-HO UR LC 50 O F H-19520 T O FATH EAD M INNOW S, Pim ephales prom elas TABLE 1 C hem ical Characteristics of H askell Laboratory W ell W ater1* Param eter C o n c e n tra tio n BO D, mg/L C O D , mg/L D O C, mg/L T O C 2, mg/L Total Kjeldahl N, m g/L Am m onia N, mg/L Turbidity, N TU Phenolics, mg/L Color, apparent C o /P t7 S o lid s Total suspended, mg/L Aluminum, mg/L Antimony, mg/L Arsenic, mg/L Beryllium , mg/L Boron4, mg/L Cadm ium 4, mg/L Calcium , mg/L C hloride3- mg/L Chromium, mg/L Cobalt, mg/L Copper, mg/L Cyanide, mg/L Iron3, mg/L Fluoride3, mg/L <2 <10 1.4 1.8 0.8 <0.2 <1.0 <0.050 <5 <4 <0.100 <0.060 <0.010 <0.001 0.020 <0.002 24.3 6.2 <0.010 <0.020 <0.010 <0.025 0.001 <0.1 Param eter Lead, mg/L M agnesium , mg/L M B A S/LA S , mg/L M ercury, mg/L N ickel, mg/L Nitrite3, mg/L Nitrate3, mg/L Phosphate3, mg/L Potassium , mg/L Selenium , mg/L Silver, mg/L Sodium , mg/L Sulfate3, mg/L Sulfide, mg/L Zinc, mg/L V olatile priority p o llu ta n ts A cid extractable priority pollutants Base/neutral priority pollutants P e sticid e s/P C B s O rg an op h o sp h ate pesticides5, pg/L C o n c e n tra tio n < 0 .0 7 5 3.7 < 0.05 < 0 .0 0 0 2 0 < 0.02 0 <0.1 1.9 <0.1 1.7 < 0 .0 0 5 < 0 .0 1 0 8.2 5.0 <0.10 < 0 .0 5 8 ND6 ND6 ND6 ND6 <0.50 1Date of sam ple collection 26 May 1992 unless indicated otherwise, an alyses performed at Environm ental Testing and Certification Corporation, Edison, New Jersey; 2D ate of sam ple collection 1 Ju ly 1992, perform ed at Du Pont Engineering T est Center, Newark, Delaware; 3Date of collection 10 Ju ne 1992, analyses performed at Du Pont Chem icals, Jackson Laboratory, Deepwater, New Jersey; 4Below the Published Method Detection Lim it (BMDL); sAnalyses perform ed at Hazleton Laboratories, Inc., M adison, Wl; 6None detected; 7Units based on cobalt/platinum reference. l1 Company Sanitized. Does not contain TSCA CBS Du Pont HLR 593-92 STATIC, A CU TE, 96-H O UR LC50 O F H-19520 T O FA T H EA D M INNOW S, Pim ephales prom elas TABLE 2 M easured Concentrations o f H-195201 in a 96-Hour, Static, Unaerated Test * N om inal Concentration (mg/L) M easured Concentration (mg/L) Start End C o n tro l 75.4 116 179 275 423 650 1000 0.00 69.7 93.3 132 209 278 455 815 0.14 72.9 91.7 86.5 243 343a 540a 907a aSam ples taken at tim e of total mortality. l2 Company Sanitized. Does not contain TSCA CBj Du Pont HLR 593-92 STATIC, ACUTE, 96-HOUR LC50 O F H-19520 TO FATHEAD MINNOWS, Pimephales promelas TABLE 3 W ater Chem istry of the Control at T est Start Total A lkalinity (mg/L a s C aC O s) 80 ED TA Hardness (mg/L a s C a C 03) 85 C onductivity (pm hos/cm ) 175 I3 fipmpany Sanitized. Does not contain TSCA CBI Du Pom HLR 593^92 . r. _ STATIC, ACU TE, 96-H CU R LC50 Q F H-19520 T O FA T H EA D MINNOW S, Pimephades prom olas TABLE 4 Ph ysical and Cham inal Hhaaracteristics of H -1 95201 est Solutions M easured C o n c e n tra tio n (mg/L) Tim e o Hr 24 Hr 48 Hr 72 Hr 96 Hr D issolved O xvoen. m? /l Control (0.14) 72.9 91.7 86.5 243 343 540 907 QH Control (0.14) 72.9 91.7 86.5 243 343 540 907 8.5 8.5 8.6 8.6 8.5 8.6 8.5 8.5 7.5 7.5 7.5 7.5 7.5 7.5 7.5 7.5 7.3 7.4 7.4 7.4 7.5 7.9 8.1 8.2 7.4 7.4 7.4 7.4 7.4 7.4 7.5 7.5 6.7 6.6 6.7 6.7 6.6 7.2 .a . 7.4 7.4 7.4 7.4 7.4 7.4 . 6.3 5.9 6.4 6.4 6.8 6.8 7.4 7.4 7.4 7.4 7.4 7.4 5.3 4.8 5.4 5.3 5.3 7.4 7.4 7.4 7.4 7.4 _ aDash (-) indicates that reading was not made due to total mortality. 14 ggmpany Sanitized. Does not contain TSCA CBI Du Pont HLR 593-92 STATIC, A C U T E , 96-H O U R LC50 O F H-19520 T O FA T H E A D M INNOW S, Pim ephales prom elas TABLE 4, Continued P h ysicalI and C hem ical C h aracteristics of H-19521 est Solutions M easured C o n c e n tra tio n (mg/L) Tim e O Hr 24 Hr 48 Hr 72 Hr 96 Hr Tem perature. C C ontrol (C 14) 72.9 91.7 86.5 243 343 540 907 21.4 21.4 21.3 21.3 21.3 21.2 21.3 21.2 21.1 21.1 21.1 21.1 21.1 21.0 21.1 21.1 20.9 20.9 20.9 20.9 20.9 20.9 .a 20.8 20.8 20.8 20.8 20.8 20.8 - 20.8 20.8 20.8 20.8 20.8 - Dash (-) indicates that reading w as not made due to total mortality. I5 jmpany Sanitized. Does npuontain TSCA CM Du Pont HLR 593-92 STATIC, ACUTE, 96-HOUR LC5 O F H-19520 TO FATHEAD MINNOWS, Pimephales promelas TABLE 5 O bserved M ortali of Fathead Minnows, Pim ephales prom elas, Exposed to H-19520I r 96 Hours in a Static, U naerated T est M easu.ed C o n c e n tra tio n (mg/L) Control (0.14) 72.9 91.7 86.5 243 343 540 907 Cum ulative M ortality in Percent3 24 Hours 0 0 0 0 10 80 100 100 48 Hours 0 0 0 0 10 80 100 100 72 Hours 0 0 0 0 10 100 100 100 96 Hours 0 0 0 0 10 100 100 100 aThere w ere ten fish per test concentration at test start. 16 Company Sanitized. Does not contain TSCA'CBI Du Pont HLR 593-92 STATIC, A C U T E, 96-H O U R LC50 O F H-19520 T O FA T H EA D MINNOW S, Pim ephales prom elas TABLE 6 O bserved Effects of H - 1 9 5 2 0 H H B H V | p n fa th e a d Minnows, Pim ephales promelas, for 96 H ours in a static. Unaerated Test M easured C o n c e n tra tio n (mg/L) Number of Fish W ith Effect 24 Hours 48 Hours 72 Hours 96 Hours Control (0.14) 72.9 91.7 86.5 243 343 540 907 0/10 0/10 0/10 0/10 0/9 ia.b/2 .c - 0/10 0/10 0/10 0/10 0/9 1b/2 - 0/10 0/10 0/10 0/10 0/9 - 0/10 0/10 0/10 0/10 0/9 - aAt the surface. bPartial lo ss o f equilibrium . cDash (-) indicates that no reading w as m ade because of total mortality. 17 Company Sanitized. Does not iowhudt Du Pont HLR 593-92 STATIC. ACUTE, 96-HOUR LC 50O F H-19520 TO FATHEAD MINNOWS* Pimephales promelas TABLE 7 H-19520U | H ^ | H j C o n c e n t r a t io n s C ausing M ortality to 50% (LC50) o f Fatheaa M innow s, rtm ephales promelas, at S p ecific Tim e Intervals Tim e 24 Hours LC50 (m g/La) 299 95% Lim its (m g/La) 260c 346c S lo p e 14.2 Y -in te rc e p t -35.2 48 Hours 299 260c 346 14.2 -35.2 72 Hours 276 237d 317d N/Ab N/A 96 Hours 276 237d 317d N/A N/A aB ased upon m easured concentrations at 96 hours. bN ot applicable. S lo pe and Y-intercept not calculated by the m oving average an gle technique. 95% fid u cial interval. d95% confidence limit. .I 18 t-oiTipany Sanitized. Does not contain ISCA CBI  Du Pont H LR 593-92 STATIC, ACUTE, 96-HOUR LC50 OF H-19520 TO FATHEAD MINNOWS, Pimephales promelas APPEN D IX 1 PR O T O C O L AM EN D M EN TS and S O P D EVIATIO N S PROTOCOL AMENDMENT NUMBER ONE a. ft is not necessary to weigh fish and determ ine test cham ber loading prior to test start. b. Stability and solubility of the test substance in dilution w ater and under storage conditions will not be determined. c. Sam ples w ill be taken from each replicate at test start and end for chem ical an alysis of the test substance concentration. d. A y e s ^ n d surviving fish will be sacrificed according to aquatic S O p jl e. Fish w ill be starved for 24 hours prior to test start. S O P DEVIATIONS a. D issolved oxygen concentrations on day 4 were less than 60% of saturation, and this observation w as not reported to the Study Director. ' b. The tem perature of the water bath w as not recorded. c. Tem peratures and equilibrium tim es for individual test cham bers were not recorded. PROTOCOL AMENDMENT NUMBER TWO a. The test start date on the short-term study protocol sheet (7-23-92) w as incorrect. The start date should have been 7-22-92 a s noted in the correction m ade by Robert Peterson on 7-21-92. 19 upany Sanitized. Does not contain TSCA CBI