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1.7ourram Ccal Company CO.0038T5 SeaCre,t Number 303730 <br />INTRODUCTION <br />Biomonitoring provides an effective method for testing the toxicity of effluents from <br />municipal and industrial discharges. Among tl7e advantages of biomonitoring is the ability to <br />test 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, <br />chemical analyses often go hand in hand vaith the biomonitoring tests. <br />Biomonitoring tests were conducted for Mountain Coal Company during April, 2003. In <br />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 />(Pimephales prornelas). This report details the results of the tests. <br />MATERIALS AND METHODS <br />Sample Collection <br />A sample of the 017 discharge was collected in one-gallon plastic containers at 10:30 <br />cn .April 1.2003. The effluent was packed in an ice chest and shipped overnight to the lab <br />where it arrived at 09:40 on April 2, 2003. The Chain of Custody form, documenting sample <br />collection and lab arrival times. is included in Appendix 1. In the lab, the sample was <br />refrigerated at 4'C u^til used. <br />Test Organisms <br />Daphnia rnagna were chcsen as the invertebrate species most able to tolerate the high <br />dissolved solids levels found in Mountain Coal samples. The Daphnia magna used in the test <br />were cultured in the SeaCrest laboratory. The d;7phnids were Tess than 24 hours old at the start <br />of the test. Tt7e daphnids were fed prior to the test start-up but not during the 46 hour test. <br />Daphnids are fed a mixture of yeast, cereal leaves, and trout chow (YCT); and the green <br />microalgae, Selenas:rum capncomufum. <br />One day old fathead minnows were also used in the test and came from in-house <br />cultures. Adult fathead minnows are held in ten and twenty gallon glass aquaria where they <br />produce eggs that are collected daily and held under aeration until hatching occurs in 3-4 days. <br />Larval fish from one to fourteen days old are used in the acute tests. Tt7e larval fsh are fed <br />newly-hatched brine shrimp (Artemis sp.) every other day during the test. <br />All in-house organisms are tested at least monthly in a reference toxicant test using <br />sodium chloride to confirm their overall health. <br />Test Procedures <br />Upon receipt at the lab, the water samples were analyzed for alkalinity, hardness, <br />