EPA Method 100.3

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EPA Method 100.3:
Bioaccumulation Test for Sediments Using Lumbriculus variegatus. Official Name: EPA Test Method 100.3: Lumbriculus variegatus Bioaccumulation Test for Sediments.

Sediment is homogenized and placed into test chambers in equal amounts, and overlying water is added one day prior to test initiation. Water is then renewed prior to introduction of organisms. Full water quality measurments for the overlying water are done. Oligochaetes are isolated prior to test initiation and mean group weight is measured on a subset of at least 100 organisms. Healthy organisms are then introduced to test chambers. All chambers are then checked daily and observations made to assess test organism behavior. At end of 28-d period, sediment is then sieved through a fine mesh screen. Wet weight is then determined of the survivors before they are put into a clean beaker of overlying (renewal) water for gut purging for a period of time not to exceed 24h. .

The objective of a sediment test is to determine whether chemicals in sediment are harmful to or are bioaccumulated by benthic organisms. The tests can be used to measure interactive toxic effects of complex chemical mixtures in sediment. Furthermore, knowledge of specific pathways of interactions among sediments and test organisms is not necessary to conduct the tests. Sediment tests can be used to (1) determine the relationship between toxic effects and bioavailability; (2) investigate interactions among chemicals; (3) compare the sensitivitiesof different organisms; (4) determine spatial and temporal distribution of contamination; (5) evaluate dredged material; (6) measure toxicity as part of product licensing or safety testing or chemical approval; (7) rank areas of cleanup, and (8) set cleanup goals and estimate the effectiveness of remediation or management practices.

Methods for Measuring the Toxicity and Bioaccumulation of Sediment-Associated Contaminants with Freshwater Invertebrates, Second Edition, March 2002. EPA 600/R-94/064.

No acclimation of organisms in test initiation; Feeding of test organisms; D.O.'s below 2.5 mg/L; indigenous species in sample sediment.

QC Requirements:
All practices dealing with (1) sediment sampling and handling, (2) source and condition of test organisms, (3) condition and operation of equipment, (4) test conditions, (5) instrument calibration, (6) replication, (7) use of reference toxicants, (8) record keeping, (9) data evaluation.

Maximum Holding Time:
2 weeks to less than 8 weeks


Chronic Toxicity

Ratio of total organic carbon in sediment to organism by dry weight should be no less than 50:1

Sample Prep:

Test precision estimated by reference toxicant. Intra- and Inter-laboratory precision are described by the mean, standard deviation, and relative standard deviation (percent coefficient of variation, or CV) of the calculated endpoints from the replicated tests.


Revision Number:
Second Edition, March 2000.