Document Yj4oEGwoD2YQVp43wJV4J4e2E

/ H U U -05 TOXICITY TO AQUATIC PLANTS TEST SUBSTANCE________________________________________________ Identity: Perfluorooctanoic acid, ammonium salt; may also be referred to as PFOA ammonium salt, Ammonium perfluorooctanoate, PFO, FC116, FC-126, FC-169, FC-143 or as the major component of FC1015. (Octanoic acid, pentadecafluoro-, ammonium salt, CAS # 3825-26-1) Remarks: The test sample is FC-1015. It's purity was not sufficiently characterized, though current information indicates is a 30% straight carbon chain version of FC-143 in 80% water. The 3M product lot number was "HOGE 205". The following summary applies to the test sample as a mixture of the test substance in water solution with incompletely characterized concentrations of impurities. Data may not accurately relate toxicity of the test sample with that of the test substance. Data was used to compare toxicity of the branched/straight chain ammonium perfluorooctanoate homologue mixture in FC-143 vs. what is supposed to be the 100% straight carbon chain ammonium perfluorooctanoate in FC-1015. METHOD:________________________________________________________ Method: OECD 201, U S E P A -T S C A , Buideline 797.1050 Type (test type): Acute Static GLP: No Year study performed: 1996 Species: Selenastrum capricornutum Source: Originally obtained from the University of Texas at Austin on January 10,1996, maintained in culture medium atT.R . Wilbury Laboratories, Inc, Marblehead, Massachusetts. Element basis: Algal cell counts (cells/ml), cell dry weights. Exposure period: 96 hours Statistical Methods: Probit analysis Analytical monitoring: pH and temperature. Test Conditions: Algal Nutrient Medium: Sterile enriched nutrient medium per USEPA 1978 guideline. This nutrient medium provided all mineral nutrients essential for algal growth and also served as the diluent for all algal operations. The pH of this synthetic algal medium was adjusted to 7.5 prior to use in assays. Stock and test solution preparation: A 3330 mg/L stock solution was prepared by diluting 3.33 grams of test substance in 1 liter of water. Aliquots 004071 were then added directly to dilution water in test vessels to create test solutions. Exposure vessels: 250 ml Erlenmeyers containing 100 ml test solution and capped with Inverted glass beakers. Agitation: Shaken continuously at 100 rpm. Number of replicates: 3 Initial cell loading: 1.0 x 104 cells/mL Number of concentrations: five plus a blank control Lighting: 400 foot-candles from continuous cool-white fluorescent lighting. Water Chemistry: pH range (0-96 hours): 7.5 - 10.8 (control exposure) 7.4 - 7.6 (3,330 mg/L exposure) Test temperature range (0-96 hours): 24.0 - 24.2 C (incubator) RESULTS________________________________________________________ Nominal concentrations: Blk control, 210, 430, 830,1670, and 3330 mg/L Algal Growth Response EC5ovalues Exposure (Contact) Hours 24 48 72 96 Cell-Count mg/L (95% C.l.) 2510 (1340->3330) >3330 2040 (1190->3330) 1980 (1710-2360) Average Specific Growth Rate mg/L (95% C.l.) 1700 (673->3300) >3330 >3330 >3330 Element values are based on nominal concentrations. Reversibility of Growth Inhibition: Aliquots of the 3330 mg/L test solution were diluted with algal medium and cultured for 72 hours. Based on growth observed in the recovery phase, the effect on algal growth was found to be alg static. Remarks: Testing was conducted on the mixture of the test substance as described in the test substance remarks field. The values reported apply to that mixture and not the test substance. CONCLUSIONS___________________________________________________ FC-1015 exhibits a 96-hour EC50 cell count value of 1980 (1710-2360 mg/L) and a 96-hour E C5o growth rate value of >3330 mg/L. 004072 The 96-hour No Observed Effect Concentration (NOEC) is 210 mg/L for cell count and 430 mg/L for growth rate. The Lowest Observed Effect Concentration (LOEC) is 430 mg/L for cell count and 830 mg/L for growth rate. This test substance was determined to be algistatic. Submitter: 3M Company, Environmental Laboratory, P.O. Box 33331, St. Paul, Minnesota, 55133 DATA QUALITY___________________________________________________ Reliability: Klimisch ranking = 2. This study meets the criteria for quality testing. However, the study lacks information on purity of the test substance and actual measurements of the amount of test substance in solution. Also, no explanation is given as to why the 48-hour values were in excess of the 24 hour values. REFERENCES____________________________________________________ Test was conducted by T.R. Wilbury Laboratories, Inc., of Marblehead, Massachusetts at the request of the 3M Company, Lab Request number P1624, 1996. OTHER________________________________________________________ Last changed: 5/24/00 004073 ,iJCTiww(* p a w i p f | g K S I COPY AVAU ABLE DrLSLH Study Title Growth and Reproduction Toxicity Taat with FC-1015 And Ths rraahwatar Alga, salanaatnm caprlcornutum Guidalina Huabar U.S. EPA-TSCA, Guideline 797.1050 OECD, Guideline 201 Authora Timothy J. Ward Jacqueline H. Neviua Robert I. Boeri Study Completed May 15, 1996 Sponaor 3M company Environmental Laboratory 935 Bush Avenue Building 2-3E-09 St. Paul, Minnesota 55144 Seating Facility .;"7 b-i' T.R.Wilbury Laboratories, Inc. 40 Doaka Lana Marblehead, Maaaachuaatta 01945 Hi hM `<fJ' Page -I.tN'-'I.1,,,,.1,1 , -if J f y ^ ,f, I * .. t'j 'k4,j^U m um BEST COPY AVAII ABLE H H * H H I . TABU or CORTOTS SECTION I X. ' Table of Contents '. index of Tables and Figures XXX. summary IV. General information V. Introduction i n . Methods and Materials VII. Results Vi l i . References Signature Page Appendix A. Water Quality Data from The Toxicity Test FASE ,^ 2 1 ' .iiV 3 4, 5 6 6 11 16 17 18 B R T COPYAWIMBLE II. INDEX or TABLES AMD PIQURES Tabi* 1. chemical characterization of a representative sample of vater used to formulate test media for the toxicity test with the freshwater alga, Salanaatrum caprlcornutum, and FC-1015. Table 2. Test conditions summary for the toxicity teBt with FC-1015 and the freshwater alga, Salanaatrum caprlcornutum. Table 3. cell growth data from the toxicity test with FC-1015 and the freshwater alga, Salanaatrum caprlcornutum. Table 4. Average specific growth rate and percent change from the control from the toxicity test with the freshwater alga, solonaatrum caprlcornutum, and FC-1015. Table 5. Effective concentrations (EC10S, ECSOs, EC90s) from the toxicity test with the freshwater alga, selenastrum caprlcornutum, and FC-1015. Table A.l. Temperature measured during the toxicity test with FC-1015 and the freshwater alga, Selenastrum caprlcornutum. Table A.2. pH values measured during the toxicity test with FC-1015 and the freshwater alga, Selenastrum caprlcornutum. Figure 1. Growth of Selenastrum caprlcornutum exposed to FC-1015 for 96 hours. PAGE 7 9 12 14 15 19 20 13 iji , iff a i' 't'' IV> ' V * ' ' n - ,< 4 wilbury study Number 1029-TH , w Page 3 of 20 {J , f ` , 0407 RttCOPYAVAIimt III. BUMMARX The toxicity of FC-1015 to the fraahwator alga, fiaianaatrcun ctprlcornutum, is daacribad in this final report. The test was conducted for 3K company for 96 hours from April 19 to 23, 1996 at T.R. wilbury Laboratories, Zno. in Marblehead, Massachusetts. It was conducted according to procedures of the U.s. Environmental Protection Agency (1993) and the OECD (1984). FC-101S was supplied by the sponsor. The test was performed under static conditions with five concentrations of test substance and a dilution water control at a temperature of 24 t 2*C. Nominal concentrations of FC-1015 were 0 (control), 210, 430, 830, 1,670, and 3,330 mg/L. The dilution water was sterile enriched media adjusted to a pH of 7.5. Exposure of algae to the test substance for 96 hours resulted in a median effective concentration (EC50) of 1,980 mg/L FC-1015 (95t confidence interval - 1,710 to 2,360 mg/L) when calculated using the number of cells/ml, and >3,330 mg/L when calculated using the average specific growth rate. The 96 hour no observed effect concentration (NOEC) is 210 rag/L and the lowest observed effect concentration (LOEC) is 430 mg/L when calculated using the number of cells/ml. The NOEC is 430 mg/L and the LOEC is 830 when calculated with the average specific growth rate. -**pa*%r~* B B T COPY AVAU ABLE xv. onm ub imforkatiom 1 E Material Tested: Sponsor : Test substance Shipped From: Reported purity: Experimental Start Date: Experimental completion Date: Archive Location for the Raw Data and a Copy of the Final Report: rc-1015 -' 3M company Environmental Laboratory 935 Bush Avenue Building 2-3B-09 St. Paul, Minnesota 55106 3M Company Environmental Laboratory 935 Bush Avanua Building 2-3E-09 St. Paul, Minnesota 55106 100 April 19, 1996 April 23, 1996 T.R. Wilbury Laboratories, Xnc. Marblehead, Massachusetts *I t | i K s r c o p y V in j/ |BL V INTRODUCTION Thi* tudy waa aponaorad by 3H Company. Tba objective of the atudy wee to determine the 24, 48, 72, and 96 hour effective concentratlona (EClOa, EC50a, EC90a) and no obaerved affect concentration (NOEC) of the teat aubatance to the freahwater alga, Selenaatrum capricornutum, under atatic conditiona. The teat vaa conducted following U .s . EPA (TSCA) and OECD guidelinea but at the Sponaor'e raqueat it waa not conducted according to Good Laboratory Practice (CLP) rulaa. The report containa sections that deacribe the methoda and materials employed in the atudy and reaulta of the inveatigation. The report also containa an appendix that preaenta water quality data collected during the teat. VI. METHODS AMD MATERIALS TEST SUBSTANCES The sample of FC-1015 (T.R. wilbury Laboratories sample Number 604) was delivered to T.R. Wilbury Laboratories on April 4, 1996. The sample waa delivered in a metal can within a cardboard box at ambient temperature. The teat aubatance waa contained in 1 250-ml plastic bottle. The label attached to the bottle contained the following informations "FC1015, (LR P1624), See USDS, Susan A. Beach, 2-3E-09, (612) 778-7452, 4/2/48". FC-1015 (a clear liquid) was supplied by 3M Company, St Paul, Minnesota. Prior to use the test substance was stored in the dark at room temperature. The test substance was assumed to have a purity of 100 active ingredient and to be stable under storage and testing conditiona. Unused test substance is returned to the sponsor. DILUTION WATER! Water used for acclimation of test organisms and for all toxicity testing was sterile enriched siedia (U.S. EPA, 1978; T.R. Wilbury standard Operating Procedure number 6) adjusted to a target pH of 7.5 prior to use. Characterization of a representative sample of water used to formulate media is presented in Table 1. TEST ORGANISM! m : Algae used for the test (Selenaatrum capricornutum, u t e x 1648) was ''' from a culture originally procured from the Culture Collection of Algae at > the University, of Texas at Austin and delivered to T.R. wilburyLaboratories!on January 10,!1996. The culture was transferred to sterile enriched media `identical.to media uaed for this teat and maintained at:: test conditions' for.;at;,least 14 daya before the definitive test. ..The. subsample of algae 'used`,,to inoculate media at the start of the definitive! test came,'from, an 8 day old culture. identification of the cult organisms was verified using an appropriate taxonomic key 00407 BESTCOPY AVAUABLE m Tabla 1. chemical characterization of a representative aazipla of water used to formulate test media for the toxicity teat with the freshwater alga, Sslsnsstrum capricornutum, and rc-1015. Parameter1 Unit of Measurement! Detection Limit Measured Value Metals Aluminum Arsenic Boron Cadmium chromium Cobalt Copper Iron Lead Mercury Nickel Silver Zinc Nitrate chloride Fluoride Total organic Carbon Total Phosphorus Organochlorine pesticides Toxaphene Organophosphorus Pesticides Dimethoate TEPP Monocrotophoa PCBa mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L as n mg/L mg/L mg/L mg/L 0.1 0.01 0.5 0.0002 0.01 0.03 0.005 0.03 0.005 0.0003 0.03 0.02 0.02 0.05 1 0.1 1 0.03 NDJ ND ND HO ND ND ND 0.03 ND ND ND ND ND ND ND ND ND pg/L pg/L 0.5 2 ND ND pg/L pg/L pg/L pg/L pg/L ; 0.5 ' ND 2.0 ND 2.0 ND 2.0 v ND o.5|ipi<r> : . % - *V 1> 1 i. . , * * >V,.: . during;August, 1995 and oiitine water quality limit. ` r ,ii' * 1 C040  B rcB pyAtoj/jiB ( filli TOXZCITT TESTI The toxicity tost was performed iron April 19 to 23, 1996, according to procaduraa of tha U.s. Environmental Protaction Agency (1993) and tha OECD (1984), A summary of tha tast conditions is prassntad in Table 2, A screening tast was not conducted because nominal concentrations for tha definitive tast were provided by tha sponsor. The tast was conducted at a target temperature of 24 t 2*C with five concentrations of test substance and a dilution water control. Kosdnal concentrations of the test substance were: 0 mg/L (control), 210, 430, 830, 1,620, and 3,330 mg/L. A 3,330 mg/L stock solution was formulated by adding 3.33 g of test substance to a 1,000 ml class A volumetric flask and filling the flask to the mark with sterile algal media. The stock solution was mixed and appropriate amounts were added directly to dilution water in test vessels to formulate teat media. Algae were distributed among three replicateB of each treatment at the rate of approximately 10,000 cella/ml. The test was performed in 2S0 ml glass Erlenmeyer flasks that contained 100 ml of test solution. Test vessels ware capped with inverted glass beakers and randomly arranged on a rotary shaker adjusted to 100 rpm in an incubator during the test (a random numbers table was used to select the location of each vessel). A 24 hour light and 0 hour dark photoperiod was automatically maintained with cool-white fluorescent lights that provided a light intensity of 400 footcandles. The number of algal cella/ml in each test vessel and the occurrence of relative size differences, unusual cell shapes, colors, flocculations, adherence of cells to test containers, or aggregation of cells was determined visually by means of direct microscopic examination with a hemocytometar. Cell counts were made and recorded daily during the 96 hour test. The determination of whether toxic effects were algistatic or algicidal was performed at the conclusion of the toxicity test (0.5 ml of media from each 3,330 mg/L vessel were transferred to a vessel containing 100 ml of fresh media without FC-1015 and incubated for 72 hours under test conditions). Temperature of the incubator was measured and recorded daily (thermometer number 3794), and pH (Beckman model pHI 12 meter; instrument number 144) was determined in each test vessel at the beginning and end of the test.,' The'temperature in a representative vessel of water incubated with the test^vessels was continuously recorded. { i /Y V !is3&6 nm il a> it; a T.R. Wilbury study Humber 1029-TH Page 8 of 20 r `V V ; I J B V * 'f 1 i %t 1 BEST COPY AVAII ABU BEST COPY AVAII ABLE STATISTICAL KITHODSI The average pacific growth rata was calculatad as tha natural log of tha numbar of cells/ml at 24, 40, 72, and 96 hours minus tha natural log of tha numbar of calls/sil at 0 hours divided by tha exposure period. Tha percent change from tha control was calculatad by subtracting the treatment average specific growth rate from the control average specific growth rata, dividing the difference by the average specific growth rate in the control, and multiplying that value by 100. Results of the toxicity test were interpreted by standard statistical techniques, when possible. The probit method (Stephan, 1983) was used for calculating EC10, EC50, and EC90 values (the probit method was used despite cautionary statements because ECSO values compared well to values compared to other methods). All calculations were performed using the number of cells/ml and the average specific growth rates and nominal concentrations of FC-1015. The no observed effect concentration (NOEC) weib determined using a parametric one-way analysis of variance (ANOVA) and the number of cells/ml and the average specific growth rate in each test vessel at the end of the test. bestcoptavahable VIZ. RESULTS Insoluble material was not obaacvad in tast vaaaala during tha taat. Tha ' algal . population grow vail, raaulting in an average of 1.471.000 cells/ml in tha control aftar 96 hours (Tabla 3). Water quality throughout tha tast was within accaptabla limit (Appendix A ) . Tha range of incubator temperatures was 24.0 to 24.2*c (Tabla A.l). Tha pH of test madia was not significantly affacted by tha tast substance at tha beginning of tha tast (Tabla A.2). Biological data generated by the acute toxicity tast with rc-1015 are presented in Tables 3 and 4. The growth curve for algae exposed to the test substance for 96 hours is presented in Figure 1 (the slope of the dose-response curve is 2.2 based on calls/ml at 96 hours). The EC10, EC50, and EC90 values for algae exposed to FC-1015 are presented in Table 5. The 96 hour EC50 (and associated 95 percent confidence limits) is 1,980 mg/L (1,710 and 2,360 mg/L) when calculated using the number of cells par ml, and >3,330 mg/L when calculated using tha avaraga specific growth rata. No effects (size differences, unusual cell shapes, colors, flocculations, adherence of cells to test containers, or aggregation of cells) were noted during the test. The 96 hour NOEC is 210 mg/L and the LOEC is 430 mg/L FC-1015, when calculated using the number cells/sil and the 96 hour NOEC is 430 mg/L and the LOEC is 830 mg/L FC-1015, when calculated using the average specific growth rate. The determination of whether toxic effects were algistatic or algicidal was performed at the conclusion of the toxicity test. A 0.5 ml aliquot of media from each 3,330 mg/L vessel was transferred to a vessel containing fresh siedia without FC-1015 and incubated for 72 hours under test conditions. Algae increased from <10,000 cells/ml to 830.000 cells/ml, indicating that the effect of FC-1015 at this concentration was algistatic rather than algicidal. BESTCOPY AVAII ABLE Table 3. Call growth data iron tha toxicity taat with rc-1015 and tha freshwater alga, salanaatrun caprlcornuturn. Noainal concantration of FC-1015 (ng/i<) Replicata 0 (control) 210 430 830 1,670 3,330 * \*t' 1 2 3 mean i 2 3 mean % control 1 2 3 mean % control 1 2 3 mean % control 1 2 3 mean control 1 2 3 mean * control Number of cella/ol X 101 (hour) 0 24 48 72 96 10 32 10 32 10 34 10 33 10 30 10 30 10 34 10 31 100 94 10 30 10 40 10 28 10 33 100 100 10 30 10 24 10 28 10 27 100 82 10 20 10 22 10 16 10 19 100 58 10 10 10 10 100 V 16 12 14 14 42 ....... 136 146 128 137 , 664 662 696 674 154 724 146 650 144 712 148 695 108 103 138 620 140 598 148 662 142 627 104 93 156 400 160 484 150 436 155 440 113 65 140 404 142 402 138 380 140 395 l ( f 2 59 88 '8 4 ' 80 84 i. 280 232 250 254 38 1,480 1,518 1,416 1,471 1,442 1,496 1,466 1,468 100 1,338 1,280 1,452 1,357 92 1,190 1,096 1,168 1,151 78 798 862 870 843 57 460 454 488 467 32 BESTCOPyAVAII ABLE 0 24 Control 830 mg/L 48 Hours -210mg/L -1,670 mg/L 72 96 430 mg/L -3,330 mg/L W TCOPYAM IIAB E Tabi* 4 Average pacific growth rat* and parcant change iron tha control iron tha toxicity taat with tha fraahwatar alga, Salenaatruo capricornutum, and re-1015. Nominal concantration of FC-1015 (mg/L) Average spacific Growth Rata 24hr 48hr 72hr 96hr Percent change Prom Control 24hr 48br 72hr 96hr 0 (control) 210 430 330 1,670 3,330 0.050 0.055 0.053 0.052 0.047 0.056 0.059 0.052 0.050 0.055 0.057 0.051 0.041 0.057 0.053 0.049 0.027 0.055 0.051 0.046 0.014 0.044 0.045 0.040 6 -2 -2 0 0022 13 -4 9 6 46 0 12 12 72 20 22 23 [ *i ; ASMS*] 8B T COPY m \l ABLE Tabi* S. Effective concentrations (EClOs, EC50s, EC90a) fron tha toxicity taat with tha freshwater alga, Salunaitrua capricornutum, and rc-1015. Tima EC Valua (mg/L) 95 Parcant Confidanca Limita (mg/L) Calculatad Using tha Euabar of Cells/ml 24 hours 48 hours 72 hours 96 hours EC10 524 EC50 2,510 EC90 a >3,330 EC50 m >3,330 EC10 m 448 EC50 2,040 EC90 m >3,330 EC10 a 533 EC 50 xt 1,980 EC90 as >3,330 <210 - 1,010 1,340 - >3,330 >3,330 >3,330 <210 - 814 1,190 - >3,330 >3,330 417 - 643 1,710 - 2,360 >3,330 calculated Using tha Average Specific Growth Rate 24 hours EC10 a 541 EC50 st 1,700 EC90 - >3,330 EC50 a >3,330 EC 10 a 1,330 EC50 a >3,330 EC90 - >3,330 EC10 a 1,430 EC50 a >3,330 EC90 a >3,330 i f, <210 - 1,110 673 - >3,330 2,260 - >3,330 >3,330 970 - 1,770 >3,330 >3,330 ,860 >3,330 >3,330' * 0040S BEST COPY AVAIl ABLE Vili. REFERENCES OECD. 1981. OECD Guidelines for Testing of Chesiicels. Annex 2. OECD principles of Good Laboratory Practice. Adopted 1 June 1981. OECD. 1984. OECD Guidelines for Testing of Chesiicals. Section 2i Effects on Biotic Systems. Method 201, Alga Growth Inhibition Test. * Adopted 4 April 1984. Stephan, C.E. 1983. Computer Program for Calculation of LC50 Values. Personal communication. U.S. EPA. 1978. The Selenaatrum caprlcornutum Prints Algal Bottle Test. EPA-600/9-78-018. Environmental Research Laboratory, Corvallis, . Oregon. : U.S. EPA. 1993. 40 c f r part 797. Toxic Substances control Act Test Guidelines; Final Rules. Section 797.1050. U.S. EPA. 1993. 40 CFR Part 792. Toxic Substances Control Act (TSCA); Good Laboratory Practice standards; Final Rule. 0040S BESTCOPY AVAII ABLE IX. SIONAXURS PUOI Xt,(M(U.(Mt, % 0 ( . U l - Jacqueline M. Hisvius Study Director Biologist - (Pi-ti 1 ( ftiiiS <( Petjr L. Kowalski Aqiuatic 'TJoxicologist -5 .< it Jeanne p . HagazV !) Biologist 'J 5-15% v l(U kct.a Ki'-it o 'fci PorI ( Keven M. sr Biologist Robert L. Boeri Coauthor S jS d k . BESTCOPY AVAII ABLE K SrCO Pir AVAIl ABLE COPYAVJUlABLE Tabi A.2. pH valu swaaurad during tha toxicity tat with rc-1015 and th iraahvatar alga, 5lnaatnin otpeleorautum. Nominal Concentration of rc-1015 (ag/L) Replicata PH Initial Pinal 0 (control) 210 430 830 1,670 3,330 1 2 3 1 2 3 1 2 3 1 2 3 1 2 3 1 2 7.5 10.7 7.5 10.8 7.5 10.8 7.4 9.9 7.4 9.8 7.4 10.0 7.4 6.5 7.4 6.4 7.4 6.4 7.4 6.1 7.4 7.0 7.4 7.4 7.4 7.6 7.4 7.6 7.4 7.4 7.4 7.6 7.4 7.5