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An Effective Approach to Maintain Homogeneity of Test Item in Soil for Earthworm (Eisenia fetida) Toxicity Test Jigar R. Rana Ecotoxicology
Transcript of JRF Global | GLP Research...y &RQFHQWUDWLRQ PJ NJ $ VRLO %RG\ :HLJKW PJ 'D\ &RQFHQWUDWLRQ PJ NJ $...
An Effective Approach to Maintain Homogeneity ofTest Item in Soil for Earthworm (Eisenia fetida)Toxicity Test
Jigar R. RanaEcotoxicology
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Introduction
• Guideline suggests analysis for volatile, unstable, andreadily degradable substances
Q1. How about insoluble and soluble compounds?
Q2. How to maintain homogeneity in the absence ofanalysis?
3
Introduction (Continued) • Carbendazim
• Solubility in water is 8 mg/L
4
Introduction (Continued) • Eisenia fetida has been extensively used as a standard test
organism for the risk assessment of pesticides and is widelyused to assess its sensitivity to chemical pollution
Life Cycle of Earthworm
Cocoon ProductionCocoon Hatchling
Juvenile
Adult
Reproduction
5
Experimental Design
AcclimatisationArtificial soil preparation Mixing and Exposure of Test Item
Distribution in test vessels Addition of water to achieve moisture contentObservation
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Experimental Design (Continued)Conventional Method
Mixing of Test Item
Mixing in artificial soil in bulk
Distributed in replicates
Individual Replicate Exposure Method
Mixing of Test Item Mixing with sand
Mixing in artificial soil in replicate
Transfer into test vessels
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Results
200250300350400450500
0 0.7 1.3 2.4 4.2 7.6 13.7 24.7 44.4 80
Body
Weigh
t (mg)
Concentration (mg/kg A. soil)
Body Weight (mg) - Day 0
200250300350400450500
0 0.1 0.2 0.4 0.8 1.4 2.6 4.6 8.3 15.0
Body
Weigh
t (mg)
Concentration (mg/kg A. soil)
Body Weight (mg) - Day 0
200250300350400450500
0 0.1 0.2 0.4 0.8 1.4 2.6
Body
Weigh
t (mg)
Concentration (mg/kg A. soil)
Body Weight (mg) - Day 28
200250300350400450500
0 0.7 1.3 2.4 4.2 7.6 13.7
Body
Weigh
t (mg)
Concentration (mg/kg A. soil)
Body Weight (mg) -Day 28
Conventional Method Individual Replicate Exposure Method
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Results (Continued)Conventional Method Individual Replicate Exposure Method
Concentrations(mg/kg A. soil) Sign of Toxicity4.2 Sluggish, Shrunk7.6 Sluggish, Shrunk13.7 Sluggish, Shrunk24.7 Shrunk44.4 -80.0 -
Concentrations(mg/kg A. soil) Sign of Toxicity1.4 Sluggish2.6 Sluggish, Shrunk4.6 Shrunk8.3 -
15.0 -
0.0 0.0 0.0 2.517.5
50.065.0
97.5 100.0 100.0
0.020.040.060.080.0
100.0
0 0.7 1.3 2.4 4.2 7.6 13.7 24.7 44.4 80
% Mo
rtality
Concentration (mg/kg A. soil)
0 0 0 0 0 2.517.5
97.5 100 100
020406080
100
0 0.1 0.2 0.4 0.8 1.4 2.6 4.6 8.3 15.0
% Mo
rtality
Concentration (mg/kg A. soil)
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Results (Continued)Conventional Method Individual Replicate Exposure Method
0.00
20.00
40.00
60.00
0 0.7 1.3 2.4 4.2
Mean
N°of J
uvenile
s
Concentration (mg/kg A. soil)
-1.6 2.4923.22
74.02 90.95 91.83
-200
20406080
100
0.1 0.2 0.4 0.8 1.4 2.6Percen
t Redu
ction
Concentration (mg/kg A. soil)
8.58
49.10
93.25 98.96
0.0020.0040.0060.0080.00
100.00
0.7 1.3 2.4 4.2
Percen
t Redu
ction
Concentration (mg/kg A. soil)
020406080
100
0 0.1 0.2 0.4 0.8 1.4 2.6
Mean
N°of J
uvenile
s
Concentration (mg/kg A. soil)
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Results (Continued)
End pointsConventional Method Individual Replicate Exposure Method Comparison
mg/artificial soilLC50 for adult mortality (28 day) 8.26 2.81 2.9 timesEC50 for reproduction 1.32 0.68 1.9 timesNOEC 0.7 0.4 1.8 timesLOEC 1.3 0.8 1.6 times
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Published results
End points
Garcia, 2004
Sian R Ellis et al.,
2007R. Shanmuga-sundaram et
al., 2013Mcshane et
al. 2012 Method 1 Method 2
LC50 - 8.03 6.33 - 8.26 2.81
EC50 2.7 - 2.45 3.3 1.32 0.68
NOEC 0.1 - 1.14 - 0.7 0.4
LOEC - - 2.06 - 1.3 0.8
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Summary• Individual replicate exposure method leads to deduction of
the end points: survival, growth, and reproduction to higherlevel of accuracy, with compressed range of concentrations.
• While the LC50, EC50, and LOEC results of our study aresubstantially lower than the published data.
13
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.
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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 onearthworms. Rev. Environ. Contam. Toxicol. 125, 23–99.• Ellis, S.R., Hodson, M.E., Wege, P., 2007. The influence of different artificialsoil types on the acute toxicity of carbendazim to the earthworm Eiseniafetida in laboratory toxicity tests. Eur. J. Soil Biol. 43, 239-245.• Garcia, M., Römbke, J., de Brito, M.T., Scheffczyk, A., 2008. Effects of threepesticides on the avoidance behavior of earthworms in laboratory testsperformed under temperate and tropical conditions. Environ. Pollut.153(2),450-456.
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References (Continued)• Holmstrup, M., 2000. Field assessment of toxic affects on reproduction in theearthworm Aporrectodea longa and Aporrectodea rosea. Environmental toxicologyand 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 forStandardization, 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 andreproductive success. Sci. Total Environ. 363, 237–244.• OECD, 1984. Guidelines for Testing of Chemicals. Test 207: Earthworm AcuteToxicity Tests. Organization for Economic Co-operation and Development (OECD).• OECD, 2016. Guidelines for Testing of Chemicals. Test 222: Earthworm ReproductionTest (Eisenia fetida/ Eisenia andrei). Organization for Economic Co-operation andDevelopment (OECD).• Reinecke, S.A., Reinecke, A.J., 2007. The impact of organophosphate pesticides inorchards 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 soilclassification and assessment concepts. Ecotoxicol. Environ. Saf. 62, 249–265.
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