APPENDIX I Toxicity Identification Evaluation Reports for Chollas ...

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Toxicity Identification Evaluation of Sweetwater River Stormwater Using Selenastrum capricornutum August 2006 TIEs are initiated when standard toxicity testing demonstrates toxicity in an effluent sample that has previously been toxic to test organisms. Standard toxicity test methods sometimes rely on sublethal endpoints, such as C. dubia reproduction as indicators of chronic toxicity, and require substantially more time and resources to evaluate than methods that rely exclusively on a mortality endpoint. In addition, the USEPA guidelines do not provide specific guideline for test procedures using each toxicity test species, given the large number of test organisms. Therefore, the USEPA’s TIE documents are used as guidance for conducting TIEs because it may not be possible or cost-effective to strictly adhere to these protocols. In addition, the USEPA protocols were initially designed for TIEs using whole effluent samples, and not the more variable and less predictable stormwater samples. Thus, modifications for efficiently conducting TIEs on stormwater samples using specific test organisms, and for specific site conditions may sometimes include the following: changes in test volumes, test duration, replicate number, number of test concentrations, and reduction in frequency of test solution renewal. Phase I test procedures are designed to identify obvious alterations in effluent toxicity, which may be achieved using modified chronic test methods. 2.2 INITIAL TOXICITY TESTING SUMMARY FOR SWEETWATER STORMWATER Sweetwater River has been identified as a TIE site due to the evidence of benthic community impacts and the persistent toxicity to Selenastrum capricornutum. During the October 18, 2005 storm event, stormwater from this site caused significant toxicity measured as reduced growth of S. capricornutum. The IC 50 at 96 hours was estimated to be 95.7% of the sample concentration. The NOEC was 50% of the sample concentration. Weston initiated TIE testing on November 1, 2005 with this sample utilizing the manipulations listed above. During the January 2, 2006 and February 19, 2006 storm events, stormwater from this site did not cause toxicity to S. capricornutum. The IC 50 at 96 hours was estimated to be greater than 100% of the stormwater sample. The NOEC was 100%. Because there was no toxicity in these stormwater samples TIE testing was not performed. 3. MATERIALS AND METHODS 3.1 TEST PROCEDURES All Selenastrum capricornutum baseline tests and TIE tests following sample manipulation followed general methods described in the U.S. EPA guidance manual, “Short-term Methods for Estimating the Chronic Toxicity of Effluents and Receiving Waters to Freshwater Organisms” (USEPA, 2002). 3.1.1 Toxicity Test Using Selenastrum capricornutum Stormwater tests for chronic toxicity using the freshwater algae S. capricornutum were run according to the USEPA protocol (EPA/821/R/02/013). Briefly, the sample and the control water were spiked with equal amounts of nutrients and subsequently filtered to 0.45 µm to remove any unicellular algae that might be present prior to test initiation. The concentration series was prepared, and 50-mL aliquots were placed into four replicate test chambers. Approximately 10,000 cells per mL were added to the test chamber and placed in random order under highintensity 24-hour light for four days. Temperature, conductivity, pH, hardness, and alkalinity was measured at test initiation. Temperature and pH was measured on days 1 to 4. The test chambers were shaken twice a day and randomized once a day. At the end of the test period, chambers were analyzed for chlorophyll. Test acceptability was determined by evaluating the Weston Solutions, Inc. 5
Toxicity Identification Evaluation of Sweetwater River Stormwater Using Selenastrum capricornutum August 2006 response of the control organisms. The test was considered invalid if the criterion of a cell density of 200,000 cells per mL in the control was not met. Variability between the control replicates should not have exceeded 20%. A reference toxicant test was conducted using copper sulfate with concentrations of 7, 14, 28, 56, and 112 µg Cu 2+ /L to establish the sensitivity of test organisms used in the evaluation of the Sweetwater stormwater. 3.2 TEST SOLUTION PREPARATION Control and dilution water for the S. capricornutum tests was synthetic moderately-hard water prepared with Epure to achieve a hardness of 80-100 mg/L as CaCO3. This water source has been used successfully on numerous similar bioassay testing programs conducted by Weston and others. Extensive testing with a variety of species and biannual chemical analysis of this water type has shown that this water source provides for good survival in laboratory controls with little to no measurable levels of contaminants. 3.3 WATER QUALITY Water quality was monitored daily as appropriate for each test, and data were recorded on data sheets. Dissolved oxygen and temperature was measured using Orion Model 840 oxygen meters and probes; pH was measured using Orion Model 230A pH meters and probes. Conductivity was measured with Orion Models 142 conductivity/salinity meters. Ammonia was analyzed using an Orion 720 digital ion analyzer with a three-point calibration curve (1, 10, and 100 mg/L). Hardness and alkalinity were measured utilizing LaMotte titration kits. 3.4 SAMPLE RECEIPT The stormwater samples were composited at the laboratory and stored at 4°C. A chain-ofcustody was completed for all samples received. Before samples were used in the tests, initial water quality measurements were taken. These measurements included temperature, total chlorine, total ammonia, pH, dissolved oxygen, and salinity. 3.5 PHASE I TIE METHODS 3.5.1 Protocol Modifications Measures to conserve time and resources required to conduct TIE testing have been developed and approved by the USEPA (USEPA 1992). This study incorporated modifications allowed for reduction in number of test concentrations, and replicates for all TIE tests conducted, relative to test conditions used in standard toxicity tests. The concentrations of stormwater samples used for the TIE test in this investigation included the 50, 75, and 100% dilutions of the stormwater samples. Three replicates were used in each stormwater sample treatment. Treatment blanks were created for each TIE test to determine the effects of the manipulation on laboratory dilution water. The results of these blanks were used to determine if any changes in toxicity of the control (dilution water) were impacted by the chemical or physical manipulation of the sample. 3.5.2 Baseline Tests The baseline test assesses the toxicity of the unmanipulated sample run concurrently with the TIE tests. This test would confirm the presence of toxicity in the stormwater sample, and toxicity in TIE treatments are compared to toxicity in the baseline test. 3.5.3 Ethylenediaminetetraacetic Acid (EDTA) Tests Two series of EDTA tests were conducted with concentrations of 3.0 and 8.0 milligrams per liter (mg/L). A stock solution of 2.5 grams (g) EDTA/L was prepared. EDTA treatments were prepared by diluting the EDTA stock solution to final concentrations of 3.0 and 8.0 mg/L in stormwater (50, 75, and 100% sample concentrations) and dilution water. The EDTA was Weston Solutions, Inc. 6
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