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Aqua-Tox, Inc. <br />4880 Robb Street, Suite 8 <br />Wheat Ridge, CO 80033 <br />(303) 621-4880 aqua-tox.com <br />ACUTE TOXICITY REPORT <br />Ceriodaphnia dubia <br />CLIENT NAME: Twentymile Coal Company SAMPLE I.D.: 042309-1 <br />50 % MORTALITY TEST: Pass TEST RESULT LC50: >100% <br />% SURVIVAL IN 100% DILUTION: 100% <br />SAMPLE DATE: 4-22-09 TIME: 14:15PM CLIENT ID: Site 87 = 007 vi <br />ORGANISM: Ceriodaphnia dubia AGE: < 24 hours old <br />TEST START: 4-23-09 11:20AM TEST FINISH: 4-25-09 11:20AM <br />TYPE OF TEST: LC50, 48 hours, static, renewal, moderately hard fresh synthetic dilution water. <br />AMMONIA: ND <br />HARDNESS: 92.14 mg/L <br />ALKALINITY: 57.96 mg/L <br />pH: 8.00 <br />D.O.: 7.26 mg/L <br />CHLORINE: ND <br />TEMPERATURE: 20.1 deg C <br />SURVIVAL DATA DILUTIONS (%) <br />DATE (HOURS)CONTROL 6.25 12.5 25 50 75 100 <br />4/23/2009 INITIAL <br />4/24/2009 24 <br />4/25/2009 48 <br />MORTALITY: <br />% SURVIVAL: <br />20 20 20 20 20 20 20 <br />20 20 20 20 20 20 20 <br />20 20 20 20 20 20 20 <br />0 0 0 0 0 0 0 <br />100% 100% 100% 100% 100% 100% 100% <br />- (MIN/MAX) DILUTIONS (%) <br />0 6.25 12.5 25 50 75 100 <br />CONDUCTIVITY (microS; <br />D.O. (mg/L; <br />pH <br />TEMPERATURE (deg C; <br />317/325 476/482 625/640 900/920 1385/1450 1899/2100 2710/2730 <br />7.0/7.3 7.1/7.3 7.1/7.3 7.1/7.3 7.1/7.4 7.1/7.4 7.2/7.5 <br />8.0/8.0 7.9/8.0 7.9/8.0 8.0/8.1 8.0/8.2 8.0/8.3 8.0/8.3 <br />19.6/20.2 19.6/20.2 19.6/20.2 19.6/20.2 19.6/20.2 19.6/20.2 19.6/20.2 <br />GENERAL CHEMISTRY - SAMPLE <br />AMMONIA: 0.06 mg/L D.O.: 7.45 mg/L <br />HARDNESS: 1301.95 mg/L CHLORINE: ND <br />ALKALINITY: 134.91 mg/L TEMPERATURE: 20.1 deg C <br />pH: 8.00 D.O AFTER AERATION: NA <br />ANOEC CALCULATION METHOD METHOD REFERENCE <br />100% SPEARMAN-KARBER EPA -821-R-02-012 October 2002 F <br />DITIONAL COMMENTS: