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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. 011509-1 <br />50 % MORTALITY TEST: Pass TEST RESULT LC50: >100% <br />% SURVIVAL IN 100% DILUTION: 95% <br />SAMPLE DATE: 1-14-09 TIME: 09:04AM CLIENT ID, Site 84 <br />ORGANISM: Ceriodaphnia dubia AGE: < 24 hours old <br />TEST START: 1-15-09 11:50AM TEST FINISH: 1-17-09 11:50AM <br />TYPE OF TEST: LC50, 48 hours static, renewal, moderately hard fresh synthetic dilution water. <br />MODERATELY HARD FRESH SYNTHETIC DILUTION WATER CONSTITUENT <br />AMMONIA. ND D O.: 7 40 mg/L <br />HARDNESS: 96.0 mg/L CHLORINE: ND <br />ALKALINITY: 52.25 mg/L TEMPERATURE: 20.0 deg C <br />pH: 7.90 <br />SURVIVAL DATA DILUTIONS (%) <br />DATE (HOURS)CONTROL 6.25 12.5 25 50 75 100 <br />9/9/2008 INITIAL 20 20 20 20 20 20 20 <br />9/10/2008 24 20 20 20 20 18 19 19 <br />9/11/2008 48 20 20 20 20 18 19 19 <br />MORTALITY: 0 0 0 0 2 1 1 <br />% SURVIVAL: 100% 100% 100% 100% 90% 95% 95% <br /> <br />DAILY CHEMISTRY - (MIN/MAX) <br />0 <br />6.25 <br />12.5 DILUTIONS (%) <br />25 <br />50 <br />75 <br />100 <br />CONDUCTIVITY (microS) 309/322 624/628 916/935 1515/1527 2840/2870 3880/3940 5000/5030 <br />D.O. (mg/L) 7.3/7.4 7.3/7.4 7.3/7.4 7.3/7.4 7.3/74 7.3/7.4 7.3/7.5 <br />pH 7.9/80 8.0/8.2 8.0/8.4 8.1/8.5 8.1/&6 8.1/8.7 8.1 /8.7 <br />TEMPERATURE (deg C) 20.0/20.2 20.0/20.2 20.0/20.2 20.0/20.2 20.0/20.2 20.0/20.2 20.0/20.2 <br />GENERAL CHEMISTRY - SAMPLE <br />AMMONIA: 1.40 mg/L D.O.. 7.45 mg/L <br />HARDNESS: 534.00 mg/1 CHLORINE: ND <br />ALKALINITY- 454.43 mg/L TEMPERATURE: 20.0 deg C <br />pH. 8 10 D.O AFTER AERATION NA <br />ANOEC CALCULATION METHOD METHOD REFERENCE <br />25% SPEARMAN-KARBER EPA -821-R-02-012 October 2002 pg.78 <br />ADDITIONAL COMMENTS: <br />Analysis <br />Conducted by: Reviewed by- <br />. 41; <br />Date- Date: L7og <br />11