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BACK TO MAIN -V 3 `. Study Title Acuta Toxicity of N2803-4 to tha Daphnid, Daphnla oagna Data Requirement! Guideline Number U.S. EPA-TSCA, Guideline 797.1300 Authora Timothy J. Ward Pater L. Kowalski Robert L. Boeri Study Completed November 28, 1995 Report Amended March 25, 1996 Sponsor 3m company Environmental Laboratory 935 Bush Avonue Building 2-3E-09 St. Paul, Minnesota 55144 ; ' i Testing Facility i T.R. Wilbury Laboratories, Inc. ' ' ` -- 'f 40 Doaks Lana Marblehead, Massachusetts 01945 Vi V. .. J .. '.#4/\\ ' ` i ; . V' T.R. wilbury study Number 895-TH ' page 1 of 22 BACK TO MAIN Amandaci 3/23/96 x. g o o d l a b o r a t o r y p r a c t i c e c o m p l i a n c e b t a s r n x m t Tbia study was conducted according to e p a Good Laboratory Practics Regulations (40 CPR 792), with the following exception. Tba stability of the tast substance was assumed but not verified .at t .r . wilbury Laboratories. Deviations from the study protocol and protocol amendments are presented in Section XXX of thin report. The final report was amended on Kerch 25, 1996 to include the date of the amendment on pages 1 and 3, to allow for signatures to the amended report on pages 2, 3, and 18, to modify the 48 hour EC50 on pages 6, 13, and 16, to modify the definition of the EC50 on page 12, and to include the amendment summary on this page. Mk Petr L. Kowalski Study Director Page 2 of 22 ||li'ilillilTIIMi BACK TO MAIN BACK TO MAIN SECTION > I. Good Laboratory Practice compliance statement II. Quality Assurance Statement III. Table of Contents IV. Index of Tables and Figures V. Summary VI. General Information VII. Introduction Vili. Methods and Materials IX. Results X. References XI. Signature Page XII. Protocol Amendments and Deviations Appendix A. Hater Quality Data from Toxicity Test PAGE 2 3 4 5 6 7 8 8 13 17 18 19 20 T.R. Wilbury study Number 895-TH Page 4 of 22 BACK TO MAIN Chemical characterization of a representative sample of dilution water used for the toxicity test with dsphnids, Daphnla magna, and N2803-4 Table 2. Test system summary for the toxicity teat with N2803-4 and daphnids, Daphnla magna Table 3. survival and sublethal effect data from the toxicity test with daphnids, Daphnla magna, and N2803-4 Table 4. Median lethal concentrations (Lc50s) and median effective concentrations (ECSOs) from the toxicity test with daphnids, Daphnla magna, and N2803-4 Table A.l. Dissolved oxygen concentration, conductivity, pH, and temperature measured during the toxicity test with daphnids, Daphnla magna, and N2803--4 Figure 1. Survival and sublethal effects of daphnids, Daphnla magna, exposed to N2803-4 for 48 hours 14 16 21 15 BACK TO MAIN The acuta toxicity of H2803-4 to tba dapbnid, Daphnla aagna, is described in this final report. Tba test was conducted for 3M Company for 48 bours from September 26 to 28, 1995, at T.R. Hilbutry Laboratories, Inc. in Karblehead, Massachusetts. It was conducted by Jeanne Magazu, Pater Kowalski, Jacqueline Nevius, Keven Stevens, Peter Nevius, Robert Boeri, and Timothy Hard according to the protocol developed for T.R. wilbury Study Number 895-TH. N2803-4 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 19.1-20.6*C. The dilution water was filtered deionized water collected at Marblehead, Massachusetts and adjusted to a hardness of 164 mg/L as Caco^. Aeration was not required to maintain dissolved oxygen concentrations above an acceptable level. Nominal concentrations of N2903-4 were 0 (control), 130, 220, 360, 600, and 1,000 mg/L. Daphnids used in the test rare less than 24 hours old at the start of the test. They were produced at T.R. Wilbury Laboratories and acclimated under test conditions for their entire life (cultures that produced test specimens were maintained at test conditions for more than 7 days). After 48 hours of exposure the control daphnids had an average wet weight (blotted) of 0.27 mg. All daphnids were in good condition at the beginning of the study. Exposure of daphnids to the test substance resulted in 100% survival at 24 bours and no surviving daphnids were immobilized. The 48 hour siedian lethal concentration (LC50) and median effective concentration (EC50) are 720 mg/L N2803-4 with a 95% confidence interval of 600 to 1,000 mg/L (no surviving daphnids were immobilized). The 48 hour no observed effect concentration (NOEC) is 360 mg/L. T.R. wilbury Study Number 895-TH VI. GENERAL INFORMATION Material Tested! Sponsor: Test Substance Shipped From: Reported Purity: Study Initiation Date: Experimental Start Date: Experimental completion Date: Study Completion Date: Archive location for the Biological Raw Data and a Copy of the Final Report: N2803-4 3M company Environmental laboratory 935 Bush Avenue Building 2-3E-09 St. Paul, Minnesota 55144 3M Company 3m Center st. Paul, Minnesota 55144-1000 100% August 16, 1995 (protocol signed) September 26, 1995 September 28, 1995 November 28, 1995 T.R. Wilbury Laboratories, Inc. Marblehead, Massachusetts T.R. Nilbury Study Number 895-TH Page 7 of 22 VII INTRODUCTION This study was sponsorsd by 3M Company, St Paul, Minnesota. Ths objactivs of ths study was to determine tbs 24 and 48 hour median Xathal concsntratlons (LC50s) and madian affsotiva concentrations (ECSOs) of tha tast substanca to tha dapbnid, Daphnla aagna, a frashwatar invartabrata, undar static conditions. Tha tast protocol was basad on u.s. e p a methods (U.S. e p a , 1993). Tha report contains sections that describe tha methods and materials employed in the study, and results of the investigation. The report also contains an appendix that presents water quality data collected during the test. VIII. METHODS AND MATERIALS TEST SUBSTANCE! The sample of N2803-4 (T.R. Wilbury Laboratories Sample Number 522) was delivered to T.R. wilbury Laboratories on August 14, 1995 in a cardboard box at ambient temperature. It was contained in an approximately 50 ml glass jar. The label attached to the bottle included the following information! "N2803-4*. N2803-4 (a white powder) 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 conditions. Unused test substance will be returned to the sponsor. DILUTION WATER! Water used for acclimation of test organisms and for all toxicity testing was deionized water collected at T.R. Wilbury Laboratories in Marblehead, Massachusetts. Water was adjusted to a hardness of 160 180 mg/L as CaCO, and stored in 500-gallon polyethylene tanks where it was aerated and continuously passed through a particle filter, ultraviolet sterilizer, and activated carbon. Results of chemical analysis of a representative sample of water are presented in Table 1. water used to initiate the final definitive toxicity test had a hardness of 164 mg/L and an alkalinity of 106 mg/L as Caco3. This water contained less than 1 mg/L total organic carbon and less than 0.1 mg/L total residual chlorine. TEST ORGANISM! Juvenile daphnids, Daphnla magna, employed as test organisms were less than 24 hours old at the start of the test. They were from a single source and were identified using an appropriate taxonomic key. Test speciswns were produced by adult daphnids that were maintained under test conditions at the T.R. wilbury facility for more than 7 days. T.R. Wilbury study Number 895-th Psge 8 of 22 BACK TO MAIN Table X. chemical characterization of a rapraaantativa sample of dilution watar uaad for tha toxicity tast with daphnida, Daphnia magna, and M2803-4. Parameter1 Unit of Maasurement 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 Monocrotophos PCBb 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 pg/L pg/L pg/L pg/L pg/L pg/L pg/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 0.5 2 0.5 2.0 2.0 2.0 0.5 ND2 KO ND ND ND ND ND 0.03 ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND Notes 1 1. Metals, nitrate, chloride, fluoride, total organic carbon, total phosphorus, organochlorine pesticides, organophosphorus pesticides, and PCBs were measured in deionized water (used to formulate dilution water) that was collected during August, 1995 and analyzed by Pace, Inc. as part of routine water quality testing. ': 2. ND not datsctad at or above the detection limit. T.R. wilbury study Number 895-TH ? BACK TO MAIN The original culture was obtained on June 1, 1994 from Aquatic Research Organism, Hampton, Hew Hampshire. Measurements made during the 7 days before the start of the test Indicated that the culture temperature ranged from 20.1 to 21.0*c and the dissolved oxygen concentration was always at least 8.6 mg/L. The culture produced young prior to day 12 and a subsample of adults produced at least 3 young/dapbnid/day for 7 days before the start of the test (reproduction data for the 24 hours prior to the start of the test cannot be verified). During acclimation daphnids were not treated for disease, they were free of apparent sickness, injuries, and abnormalities at the beginning of the test and there was no mortality during the 48 hours preceding the start of the test. The culture was supplied with a yeast/trout chow mix (Lot# DC02) and the freshwater alga, Smlenamtrxm caprlcornutua, daily during acclimation. Daphnids were not fed during the test. control daphnids, blotted and weighed at the conclusion of the toxicity test, averaged 0.27 mg and the loading rate during the definitive toxicity test was approximately 10.8 mg/L. TOXICITY TESTISO: A static screening test was conducted from August 23 to 26, 1995 with five concentrations of test substance and a dilution water control, nominal concentrations of H28C3-4 were 0.10, 1.0, 10, 100, and 1,000 mg/L. After 48 hours of exposure there was lOOt survival in the control and at 0.10, 1.0, 10, and 100 mg/L and 04 survival at 1,000 mg/L. A definitive test was conducted from September 6 to 8, 1995. It was conducted with 5 nominal concentrations of N2803-4> 130, 220, 360, 600, and 1,000 mg/L. After 48 hours there was 1004 survival at 130, 220, and 360 mg/L, 204 survival at 600 mg/L (all surviving daphnids exposed to 600 mg/L were immobilized at 48 hours) and 04 survival at 1,000 mg/L. The test was considered invalid and repeated because the production of at least 3 young/female/day by adults in the culture used to initiate the toxicity test could not be verified from the raw data. The final definitive toxicity test was performed from September 26 to 28, 1995, according to T.R. Wilbury protocol 895-TH (Acute Toxicity of K2803-4 To The Daphnid, Daphnla magna). The protocol, which was signed by the study director on August 16, 1995, is based on procedures of the u.s. Environmental Protection Agency (1993). A summary of the test system is presented in Table 2. The test was conducted at 20 t 1C with five concentrations of test substance and a dilution water control, nominal concentrations of the test substance were: 0 mg/L (control), 130, 220, 360, 600, and 1,000 mg/L. A 1,000 mg/L stock solution was formulated by combining 2.0 g of test substance and dilution water in a Class A volumetric flask and adjusting the final stock solution volume to 2,000 ml. The stock solution was mixed and then appropriate amounts were added to dilution water in test vessels. T.R. Wilbury Study Humber 895-TH Page 10 of 22 Amended 3/25/96 Twenty dapbnlda wara indiscriminately and equally diatributad among two raplicataa of aacb treatment. Tba taat waa performed in 300 ml glaaa baakara that containad 250 ml of taat aolution (watar daptb waa approximataly 9 cm). Taat vaaaala tiara randomly arrangad during tha 48 hour taat (a random numbara tabla waa uaad to aalact tba location of aacb vaaaal) and thay wara looaaly covarad. A 16 hour light and B hour dark photoperiod with a 15 minuta tranaition period waa automatically maintained with cool-white fluoraacant lighta that provided a light intensity of 110 footcandlea. Aeration waa not required to maintain diaaolved oxygen concentrationa above acceptable levela. Tha number of aurviving organiama and the occurrence of aublethal effecta (immobilization, loaa of equilibrium, loaa of reflex, excitability, diacoloration, or change in behavior) were determined viaually and recorded initially and after 24 and 48 houra. Dead teat organiama were removed when firat observed. Diaaolved oxygen (YSI Model 18 meter; inatrument number 1), pH (Beckman model pHI 12 meter; inatrument number 124), conductivity (Cole Parmer meter, inatrument number 94), and temperature (Beckman model pHI 12 meter; inatrument number 124, probe pET4) were meaaured and recorded daily in each test chamber that contained live animals. The temperature in a beaker of water incubated among the teat vessels was recorded continuously during the teBt. STATISTICAL METHODS I Results of the toxicity teat were interpreted by standard statistical techniques. Computer methods (Stephan, 1983) were used to calculate LC50s and EC50s. All calculations were performed by using nominal concentrationa of test substance and the number of live or immobilized daphnids. The no observed effect concentration ia the highest concentration of test substance that did not cause any toxicant-related mortality or sublethal effecta. T.R wilbury study Number 895-TH Page 12 of 22 BACK TO MAIN BACK TO MAIN Table 3. Survival and sublathal affect data from the toxicity test with daphnida, Daphnla magna, and N2803-4. Nominal of N2803-4 (mg/L) Rep. Number Alive 0 24 48 hr hr hr Number Affected 0 24 hr hr 48 hr 0 (control) 1 10 10 2 10 10 10 10 000 000 130 1 10 10 10 00 0 2 10 10 10 00 0 220 1 10 10 10 000 2 10 10 10 000 360 1 10 10 10 000 2 10 10 10 000 600 1 10 10 8 008 2 10 10 9 00 9 1,000 1 10 10 2 10 10 0 0 0 10 -- 0 10 " Note: Affected daphnide were immobilized at 24 hours and less active than all others at 48 hours. T.R. wilbury study Number 895-TH Page 14 of 22 BACK TO MAIN Figure 1. Survival and aublethal effects of daphnids, Daphnla jnagna, exposed to N2803-4 for 48 hours. i- ' : 1 T.R. Wilbury study Humber 895-TH Page IS of 22 Amended 3/25/96 Tabla 4. Nadian lathal concentration* (LC50a) and median affective concentrations (EC50a) from the toxicity teat with daphnids, Daphnla magna, and N2803-4. Exposure period LC50/EC50 95 percent confidence limit Calculation method LC50 24 hours >1,000 mg/L -- -- 48 hours 720 mg/L 600 to 1,000 mg/L Binomial EC50 24 hours 780 mg/L O o to to 1,000 mg/L Binomial 48 hours 720 mg/L 600 to 1,000 mg/L Binomial T.R. wilbury Study Number 895-TH Page 16 of 22 x. uruuncn ASTM. 1986. Standard Practice toe Conducting Acuta Toxicity Taata with Fishes, Macroinvertebrates, and Amphibians. E-729--80. 2B Annual Book of Standards. Stephan, C.E. 1983. Computer Program for the Calculation of LC50 Values. U.S. EPA. Duluth, MN. Personal Communication. U.S. EPA. 1993. Toxic substances Control Act Test Guidelines; rinal Rules. Section 797.1300. U.S. EPA. 1993. 40 CFR Part 792. Toxic substances Control Act (TSCA); Good Laboratory Practice Standards. Code of roderal Regulations, Title 40, July 1, 1993. j. ' ) i : i T.R. Wilbury study Number 895-TH : Page 17 of 22 BACK TO MAIN Keven M. stevena Biologist T.R. wilbury study Number 895-th Page 18 of 22 BACK TO MAIN The protocol va* amended to specify the nominal concentration* for the definitive test. The culture produced young prior to day 12 and a subBample of adult* produced at least 3 young/daphnid/day for 7 day* before the start of the test, however, reproduction data for the 24 hour* prior to the start of the test cannot be verified. This deviation did not affect the outcome of the test and no other deviations were made from the protocol. BACK TO MAIN Appendix Al HATER QUALITY DATA FROM TOXICITY TEST T.R. Hllbury study Number 895-TH Page 20 of 22 BACK TO MAIN Table X. 1. Dieaolved oxygen conoantration, conductivity, pH, and tamparatura meaaured during tha toxicity taat with dapbnida, Daphnla aagna, and N2803-4. Nominal Concentration of N2803-4 (mg/L) Diasolved Oxygan (mg/L) ----------------------- 0 24 4B Rep hr hr hr Conductivity (pmhos/cm) ----------------------- 0 24 48 hr hr hr 0 (control) i 8.7 -.7 8.4 2 8.7 8.7 8.4 130 1 8.7 8.7 8.4 2 8.7 8.7 8.4 220 1 8.7 8.7 8.4 2 8.7 8.7 8.4 360 1 8.7 8.7 8.1 2 8.7 8.7 8.0 600 1 8.7 8.7 8.2 2 8.7 8.7 7.9 1,000 1 8.7 8.7 8.1 2 8.7 8.7 8.3 540 560 550 540 560 560 560 560 570 560 560 580 580 580 590 580 580 590 600 600 610 600 600 610 650 640 650 650 640 660 720 710 720 720 710 720 T.R. Wilbury study Number 895-TH Page 21 of 22 BACK TO MAIN Tabla A.l. continuad. Nominal Concantration o N2803-4 (mg/L) Rep. PH 0 24 hr hr 48 hr 0 (control) 1 8.3 8.0 8.0 2 8.3 8.0 8.1 130 1 8.3 8.0 8.1 2 8.3 8.0 8.2 220 1 8.3 8.0 8.2 2 8.3 8.0 8.2 360 1 8.3 8.0 8.2 2 8.3 8.0 8.2 600 1 8.2 8.0 8.2 2 8.2 8.0 8.2 1,000 1 8.1 8.0 8.2 2 8.1 8.0 8.2 Tamparatura fc> 0 24 48 hr hr hr 19.1 20.4 20.1 19.1 20.5 20.0 19.2 20.4 20.0 19.3 20.4 20.0 19.2 19.2 20.4 20.4 20.0 20.1 19.3 20.5 20.1 19.2 20.6 20.0 19.4 20.5 i9.9 19.3 20.5 20.0 19.5 20.4 20.0 19.4 20.5 20.0 T.R, wilbury study Number 895-TB Page 22 of 22 V/^/ V I / V U I err viTT unnni iml lmuumm \ un ARCHIVED FINAL REPORT BACK TO MAIN LAB REQUEST NO. N2803 REQUESTOR NAME: SAB/GOEBEL/PICKETT DEPARTMENT: 3048 PROJECT NO: BI703004UQ DATE RECEIVED: 08/01/1995 DESC: FC-143 AND RELATED PRODUCTS ECOTOX WORK CONTRACT LAB(S): TRWIL EXP COMP DATE: 10/04/1995 DATE COMPLETED: 03/27/1996 PROJECT LEAD: PHONE NO: 3M FAX NO: 651-778-6176 Samples received from S.J. Martin. Samples 1-5 sent to TRWIL on 8/11/95. An additional 4L of sample 5 sent on 8/22/95. Sample 1 = Tetrabutyl ammonium hydroxide, 40% in Mater. Sample 2 = L-13492 (45% solids, 27% Mater, 27% IPA) Sample 3 = FC-26, 100% solids. Dissolve in 50:50 Mater:IPA solution. Sample 4 = FC-143, 100% solids. Sample 5 = L-13856, 15% solids. Sample 6 = L-13856, diluted by a factor of 15, tested as an effluent. Sample #1 (Tetrabutylammonium hydroxide) - a neM bottle Mas ordered from Aldrich because TR Wilbury needed more sample. This Mas lot 05922BN, red label, cat. # 17,878,0. sab 9/11/95 NOTE: All data for sample 3 (FC-26) is reported in terms of FC-26, and not as a 50/50 mixture Hith isopropanol. sab 11/21/95 NOTE: For sample #5, only a range-finding test Mas run for toxicity to algae. The requestor canceled the request for testing so no definitive test Mas run. The results for the range finder are as folloMs: Test Cone. Percent Difference in Avg. Specific GroMth Rate mg/1 96-hrs Expsure 0.1 2 1.0 18 10 4 1 0 0 32 1000 100 EC50 (95% C.I.) 160 (100 - 1000) mg/L. sab 2/16/96 For N2803-6, the testing Mas canceled before a definitive study Mith fish could be completed. Results reflect testing Mith a single concentration of 13% v/v, Mhich shoMed no mortality after 96-hours, sab 3/14/96 SAMPLE DATE CODE DESCRIPTION RESULT 1 08/11/95 TB NH40H Lot 10317KQ TRWIL 48-HR-EC50 *DAPHNIA ACUTE TEST 27 MG/L MDL or (95% C.I.) (23 - 32) U U rrm R C T LAD DM IA 04/01/2001 LAB REQUEST NO. N2803 SAMPLE DATE CODE ENVIRONMENTAL LABORATORY ARCHIVED FINAL REPORT CONTINUED DESCRIPTION RESULT BACK TO MAIN 2 MDL or (95% C. I .) 1 08/11/95 TB NH40H Lot 10317KQ TRWIL 96-HR-LC50 *FISH ACUTE TEST - 96 HOUR 720 MG/L TRWIL IC50-ALGAE *ALGAE ASSAY - IC50 3.7 MG/L (600 - 1000) (1.8 - 7.7) 2 L-13492 Lot 2 TRWIL 48-HR-EC50 *DAPHNIA ACUTE TEST TRWIL 96-HR-LC50 *FISH ACUTE TEST - 96 HOUR TRWIL IC50-ALGAE *ALGAE ASSAY - IC50 62 MG/L 960 MG/L 8.4 MG/L (36-100) (830->1000) (5.9 - 14) 3 FC-26 Lot 135 TRWIL 48-HR-EC50 *DAPHNIA ACUTE TEST TRWIL 96-HR-LC50 *FISH ACUTE TEST - 96 HOUR TRWIL IC50-ALGAE *ALGAE ASSAY - IC50 360 MG/L 140 MG/L. 90 MG/L (300 - 500) (110 - 180) (63 - 130) 4 FC-143 Lot 427 TRWIL 48-HR-EC50 *DAPHNIA ACUTE TEST TRWIL 96-HR-LC50 *FISH ACUTE TEST - 96 HOUR TRWIL IC50-ALGAE *ALGAE ASSAY - IC50 720 MG/L 740 MG/L >1000 MG/L (600 - 1000) (660 - 830) (-) 5 08/16/95 L-13856 As-is TRWIL 48-HR-EC50 *DAPHNIA ACUTE TEST TRWIL 96-HR-LC50 *FISH ACUTE TEST - 96 HOUR TRWIL IC50-ALGAE *ALGAE ASSAY - IC50 190 MG/L (130-600) >1000 MG/L (-) SEE COMMENT ABOVE 6 L-13856 1:15 diluted TRWIL 48-HR-EC50 *DAPHNIA ACUTE TEST TRWIL 96-HR-LC50 *FISH ACUTE TEST - 96 HOUR TRWIL IC50-ALGAE *ALGAE ASSAY - IC50 * = CONTRACT LAB DATA 1.2 % V/V (1.0 - 1.7) >13 % V/V (-) TESTING CANCELED