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EC number: 202-297-4
CAS number: 94-04-2
IntroductionA study was performed to assess the chronic toxicity of the test item to Daphnia magna. The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (2012) No 211, "Daphnia magna Reproduction Test" referenced as Method C.20 of Commission Regulation (EC) No. 440/2008.
MethodsPreliminary solubility work conducted indicated that it was not possible to obtain a testable solution of the test item using traditional methods of preparation e.g. ultrasonication and high shear mixing.A preliminary media preparation trial (Labcorp Study Number: 8461247) indicated that a dissolved test item concentration of approximately 14 mg/L was obtained from a saturated solution method of preparation indicating this to be the limit of water solubility of this item under test conditions.Due to the potentially volatile nature of the test item, testing was conducted in completely filled and sealed test vessels in order to minimize possible losses due to volatilization.Based on the results of a preliminary range-finding test, Daphnia magna were exposed (10 replicates of a single daphnid per group) to solutions of the test item at nominal concentrations of 0.45, 1.0, 2.2, 4.8 and 10% v/v saturated solution for a period of 21 days. The test solutions were renewed daily throughout the test. The test item solutions were prepared by shaking in excess (100 mg/L) of the test item in test water at approximately 150 rpm, at a temperature of 24 °C for 24 hours. After the shaking period any undissolved test item was removed by filtration through a 0.2 μm Gelman Acrocap filter (first approximate 100 mL was discarded in order to pre-condition the filter) to give a 100% v/v saturated solutions of the test item. This saturated solution was then further diluted to give the 10, 4.8 and 2.2% v/v saturated solutions of the test item. The 10% v/v saturated solution was also further diluted to give the 1.0 and 0.45 % v/v saturated solutions of the test item.The numbers of live and dead adult Daphnia and young daphnids (live and dead) were determined daily. The Daphnia were fed daily with an algal suspension, supplemented for the first 4 days of the test with a GEMMA Micro 300 fish food suspension.
ResultsAnalysis of the fresh test preparations on Days 0, 6, 13 and 20 showed measured test concentrations to range from 0.037 to 1.1 mg/L. A decline in measured test concentration of the aged test preparations on Days 1, 7, 14* and 21 was observed to be between 0.0060 and 0.73 mg/L. As not all test concentrations were maintained within ± 20% of the measured initial test concentration it was considered appropriate to calculate the results based on the geometric mean measured test concentration in order to give a “worst case” analysis of the data. The geometric mean measured test concentrations were determined to be 0.023, 0.052, 0.10, 0.32 and 0.71 mg/L.Exposure of Daphnia magna to the test item gave the following results based on the geometric mean measured test concentrations:
Exposure of Daphnia magna to the test item gave the following results based on the geometric mean measured test concentrations:
Endpoint Concentration mg/L(95% confidence limits)ImmobilizationEC10 >0.71 (not determined)EC20 >0.71 (not determined)EC50 >0.71 (not determined)No Observed Effect Concentration 0.71Lowest Observed Effect Concentration Not determined
Endpoint Concentration mg/L(95% confidence limits)Body LengthEC10 0.26 (0.18 – 0.35)EC20 0.64 (0.43 – 0.94)EC50 >0.71 (not determined)No Observed Effect Concentration 0.023Lowest Observed Effect Concentration 0.052
Endpoint Concentration mg/L(95% confidence limits)ReproductionEC10 0.31 (0.28 – 0.34)EC20 0.40 (0.36 – 0.42)EC50 0.63 (0.59 – 0.67)No Observed Effect Concentration 0.32Lowest Observed Effect Concentration 0.71
* = Due to a sampling error the Day 14 (old) concentration could not be determined for the 4.8 % v/v saturated solution samples and as such could not be used in order to determine the geometric mean concentration.† = It was not possible to calculate 95% confidence limits for the ECx value as the data generated did not fit the models available for the calculation of the confidence limits
The chronic EC10 for freshwater daphnia is 0.26 mg/L.
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