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A total of twenty organisms at each concentration were used for the daphnia test. The <br />animals were exposed in 30 ml plastic disposable beakers. Approximately 20 ml of the effluent <br />mixture was added to each beaker. Exposure medium was replaced after each 24-hour <br />period and organisms surviving at each concentration were counted and recorded. Routine <br />measurements were made of temperature, pH, and dissolved oxygen in each concentration. <br />Four acute tests were started on May 28, 2004 and were performed according to the <br />procedures outlined in EPA-821-R-02-012 (2002). The tests were conducted over a 48-hour <br />period. The original effluent was run with exposure concentrations of 100%, 50%, 25%, 12.5%, <br />and 1.9% effluent. Moderately hard laboratory reconstituted freshwater was used for the <br />control and for the dilutions. <br />Test Procedures for Diluted Acute Tests <br />Three additional acute tests were performed to confirm the potential for TDS to be a <br />source of toxicity in the effluent. These tests were designed to indicate the presence of other <br />contaminants that could be present. <br />The tests were accomplished by making three reconstituted test solutions. Aliquots of <br />the original effluent were diluted to stocks of 50%, 25%, and 12.5%. Each of these stock <br />solutions was then returned to its original state by bringing the alkalinity back to that of the <br />original effluent .Laboratory grade NaHCO3 (sodium bicarbonate) was added in order to raise <br />the alkalinity. These reconstituted samples were then run in toxicity tests at concentrations of <br />100%, 50%, 25%, 12.5% and 1.9%. This has the effect of maintaining a constant alkalinity <br />while diluting other contaminants that could be present in the original effluent. In essence, if <br />similar toxicity is observed in all of the tests, then the alkalinity can be recognized as the cause <br />of the problem. If the toxicity decreases with the increasing dilution of the reconstituted <br />samples, then it can be assumed that some other toxicant is present. <br />Data Analysis <br />Data from the tests were analyzed on a Pentium personal computer. Where applicable, <br />LC50's were calculated using EPA's TOXDATMnlti-Method Program according to the binomial, <br />probit, or moving average methods. Statistically significant differences between control and <br />exposed orgariisms were determined using the TOXSTAT software developed at the University <br />of Wyoming. <br />RESULTS <br />Original Sample <br />An LC50 of 77.46% was measured in this test (See Table 1). Survival was 35% in the <br />100% effluent and ranged from 75 -95% in the remaining effluent concentrations. Survival was <br />100% in the control. The results of all tests can be found in Appendix 2 of this report. <br />The SeaCrest Group 5 <br />