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INTRODUCTION <br />Biomonitoring provides an effective method for testing the toxicity of effluents from <br />municipal and industrial discharges. Among the advantages of biomonitoring is the ability to test <br />complex effluents containing a broad range of contaminants. The biomonitoring methods <br />generate data that cannot be obtained solely from chemical analyses. At the same time, chemical <br />analyses often go hand in hand with the biomonitoring tests. <br />Biomonitoring tests were conducted for Mountain Coal Company during November, 1998. <br />In accordance with accepted Environmental Protection Agency (EPA) and State of Colorado <br />procedures, acute tests were performed using Daphnia magna and the fathead minnow <br />(Pimepha/es prome/as). This report details the results of the tests. <br />MATERIALS AND METHODS <br />Sample Collection <br />A sample of the Lone Pine Pipeline (#013) discharge was collected in one-gallon plastic <br />containers at 09:05 on November 4, 1998. The effluent was packed in an ice chest and shipped <br />to the lab the next day. The Chain of Custody form showing collection and lab arrival times is <br />included in Appendix 1. In the lab, the samples were refrigerated at 4°C until used. <br />Test Organisms <br />With permission from the Colorado Department of Public Health and Environment, <br />Ceriodaphnia dubia were replaced with Daphnia magna as the invertebrate test organism for <br />Mountain Coal discharge sites. Fathead minnows, Pimepha/es prome/as, were also used in the <br />tests. The Daphnia magna are cultured in the laboratory. The daphnids were less than 24 hours <br />old at the start of the test. Seven day old fathead minnows also came from in-house cultures. <br />All in-house organisms are tested at least monthly in a reference toxicant test with sodium <br />chloride to confirm their overall health. <br />Test Procedures <br />Upon receipt at the lab water samples were analyzed for alkalinity, hardness, conductivity, <br />dissolved oxygen, chlorine, ammonia and pH. Alkalinity and hardness were determined <br />titrimetrically according to methods described in Hach Chemical Co. (1978). Ammonia was <br />measured by an Orion ion specific electrode according to procedures in APHA/AWWANVPCF <br />(1985). <br />A total of twenty organisms at each concentration was used for the acute tests. Daphnia <br />magna were exposed in 30 ml plastic disposable beakers. Twenty ml of the effluent mixture was <br />added to each beaker. Fathead minnows were exposed in 260 ml plastic cups containing 200 <br />ml of medium. Exposure medium was replaced after each 24 hour period and organisms <br />surviving at each concentration were counted and recorded. Routine measurements were made <br />of temperature, pH, and dissolved oxygen in each concentration. <br />Acute tests were performed according to the procedures outlined in Weber (1993). <br />Daphnia magna tests were conducted over 48 hours with fathead minnows being exposed for 96 <br />hours. This test was run with exposure concentrations of 100%, 75%, 50%, 25%, and 12.5% <br />effluent. Water used for the control and as the dilution water was laboratory reconstituted <br />