An Effective Approach to Maintain Homogeneity of Test Item in Soil - PowerPoint PPT Presentation

An Effective Approach to Maintain Homogeneity of Test Item in Soil for Earthworm ( Eisenia fetida) Toxicity Test Jigar R. Rana Ecotoxicology

Introduction Guideline suggests analysis for volatile, unstable, and • readily degradable substances Q1. How about insoluble and soluble compounds? Q2. How to maintain homogeneity in the absence of analysis? 2

Introduction (Continued) Carbendazim • Solubility in water is 8 mg/L • 3

Introduction (Continued) • Eisenia fetida has been extensively used as a standard test organism for the risk assessment of pesticides and is widely used to assess its sensitivity to chemical pollution Cocoon Hatchling Cocoon Production Juvenile Reproduction Adult Life Cycle of Earthworm 4

Experimental Design Mixing and Exposure of Test Item Artificial soil preparation Acclimatisation Addition of water to Distribution in test vessels Observation achieve moisture content 5

Experimental Design (Continued) Conventional Method Individual Replicate Exposure Method Mixing of Test Item Mixing with sand Mixing of Test Item Mixing in artificial soil in bulk Mixing in artificial soil in replicate Distributed in replicates Transfer into test vessels 6

Results Conventional Method Individual Replicate Exposure Method 500 500 Body Weight (mg) - Day 0 Body Weight (mg) - Day 0 450 Body Weight (mg) 450 Body Weight (mg) 400 400 350 350 300 300 250 250 200 200 0 0.7 1.3 2.4 4.2 7.6 13.7 24.7 44.4 80 0 0.1 0.2 0.4 0.8 1.4 2.6 4.6 8.3 15.0 Concentration (mg/kg A. soil) Concentration (mg/kg A. soil) 500 Body Weight (mg) - Day 28 500 Body Weight (mg) -Day 28 450 Body Weight (mg) 450 Body Weight (mg) 400 400 350 350 300 300 250 250 200 200 0 0.1 0.2 0.4 0.8 1.4 2.6 0 0.7 1.3 2.4 4.2 7.6 13.7 Concentration (mg/kg A. soil) Concentration (mg/kg A. soil) 7

Results (Continued) Conventional Method Individual Replicate Exposure Method Concentrations Concentrations Sign of Toxicity Sign of Toxicity (mg/kg A. soil) (mg/kg A. soil) 4.2 Sluggish, Shrunk 1.4 Sluggish 7.6 Sluggish, Shrunk 2.6 Sluggish, Shrunk 13.7 Sluggish, Shrunk 4.6 Shrunk 24.7 Shrunk 8.3 - 44.4 - 80.0 - 15.0 - 100.0 97.5 100.0 100.0 100 97.5 100 100 80.0 80 65.0 % Mortality % Mortality 60.0 60 50.0 40.0 40 17.5 17.5 20.0 20 2.5 2.5 0.0 0.0 0.0 0 0 0 0 0 0.0 0 0 0.7 1.3 2.4 4.2 7.6 13.7 24.7 44.4 80 0 0.1 0.2 0.4 0.8 1.4 2.6 4.6 8.3 15.0 Concentration (mg/kg A. soil) Concentration (mg/kg A. soil) 8

Results (Continued) Conventional Method Individual Replicate Exposure Method 100 60.00 Mean N° of Juveniles Mean N° of Juveniles 80 40.00 60 40 20.00 20 0.00 0 0 0.7 1.3 2.4 4.2 0 0.1 0.2 0.4 0.8 1.4 2.6 Concentration (mg/kg A. soil) Concentration (mg/kg A. soil) 100.00 93.25 98.96 Percent Reduction 100 90.95 91.83 80.00 74.02 Percent Reduction 80 60.00 49.10 60 40.00 40 23.22 20 20.00 8.58 2.49 -1.6 0 0.00 0.1 0.2 0.4 0.8 1.4 2.6 -20 0.7 1.3 2.4 4.2 Concentration (mg/kg A. soil) Concentration (mg/kg A. soil) 9

Results (Continued) Conventional Individual Replicate Method Exposure Method End points Comparison mg/artificial soil LC 50 for adult 8.26 2.81 2.9 times mortality (28 day) EC 50 for reproduction 1.32 0.68 1.9 times NOEC 0.7 0.4 1.8 times LOEC 1.3 0.8 1.6 times 10

Published results Sian R R. Shanmuga- End Garcia, Mcshane et Ellis et al., sundaram et Method 1 Method 2 points 2004 al. 2012 2007 al., 2013 LC 50 - 8.03 - 6.33 8.26 2.81 EC 50 2.7 - 3.3 2.45 1.32 0.68 NOEC 0.1 - - 1.14 0.7 0.4 - - - 2.06 1.3 0.8 LOEC 11

Summary Individual replicate exposure method leads to deduction of • the end points: survival, growth, and reproduction to higher level of accuracy, with compressed range of concentrations. While the LC 50 , EC 50 , and LOEC results of our study are • substantially lower than the published data. 12

Conclusion The Individual Replicate Exposure Method : Helps in maintaining the homogeneity of the test item in  soil Fulfills the objective of test to get enhanced accuracy for  the end points. 13

References Allen, H.E., 2002. Bioavailability of Metalsin Terrestrial Ecosystems: • Importance of Partitioning for Bioavailability to Invertebrates, Microbes, and Plants. SETAC, NewYork. Boudina, C. E., Baaliauamer, A., Grenier-Loustalot, M.F., Choven, J.M., 2003. • Photochemical behaviour of carbendazim in aquatic solution. Chemosphere. 50, 649-655. Edwards, C.A., 1998. Earthworm Ecology. St. Lucie Press, New York. • Edwards, C.A., Bohlen, P.J., 1992. The effects of toxic chemicals on • earthworms. Rev. Environ. Contam. Toxicol. 125, 23–99. Ellis, S.R., Hodson, M.E., Wege, P., 2007. The influence of different artificial • soil types on the acute toxicity of carbendazim to the earthworm Eisenia fetida in laboratory toxicity tests. Eur. J. Soil Biol. 43, 239-245. Garcia, M., Römbke, J., de Brito, M.T., Scheffczyk, A., 2008. Effects of three • pesticides on the avoidance behavior of earthworms in laboratory tests performed under temperate and tropical conditions. Environ. Pollut.153(2), 450-456. 14

References (Continued) • Holmstrup, M., 2000. Field assessment of toxic affects on reproduction in the earthworm Aporrectodea longa and Aporrectodea rosea . Environmental toxicology and chemistry.19,1781-1787. ISO, 1998. Soil quality Effects of Pollutants on Earthworms ( Eisenia fetida ). Part2: • Determination of Effects on Reproduction. ISO - The International Organization for Standardization, Genève, Switzerland. Landrum, M., Cañas, J.E., Coimbatore, G., Cobb, G.P., Jackson, W.A., Zhang, B., derson, • T.A., 2006. Effects of perchlorate on earthworm ( Eisenia fetida ) survival and reproductive success. Sci. Total Environ. 363, 237–244. OECD, 1984. Guidelines for Testing of Chemicals. Test 207: Earthworm Acute • Toxicity Tests. Organization for Economic Co-operation and Development (OECD). OECD, 2016. Guidelines for Testing of Chemicals. Test 222: Earthworm Reproduction • Test ( Eisenia fetida / Eisenia andrei ). Organization for Economic Co-operation and Development (OECD). Reinecke, S.A., Reinecke, A.J., 2007. The impact of organophosphate pesticides in • orchards on earthworms in the Western Cape, South Africa. Ecotoxicol. Environ. Saf. 66(2), 244–251. Römbke, J., Jänsch, S., Didden, W., 2005. The use of earthworms in ecological soil • classification and assessment concepts. Ecotoxicol. Environ. Saf. 62, 249–265. 15

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