This test method describes procedures for assessing the biodegradation of chemicals in natural surface water samples.

Formerly under the jurisdiction of Committee E47 on Biological Effects and Environmental Fate, this test method was withdrawn in March 2013. This standard is being withdrawn without replacement due to its limited use by industry.

Most of the simpler methods used to screen chemicals for biodegradation potential employ measurements that are not specific to the test substance, such as loss of dissolved organic carbon, evolution of respiratory carbon dioxide, or uptake of dissolved oxygen. Such methods generally are used to evaluate the transformation of the test substance to carbon dioxide, water, oxides or mineral salts of other elements, or products associated with the normal metabolic processes of microorganisms (ultimate biodegradability), or both. These methods require the use of relatively high initial concentrations of the test substance, generally 10 mg/L or higher, unless the tests are conducted using ¹⁴C-radiolabeled test compounds. Biodegradation tests measuring ¹⁴C-CO₂ evolution, for example, can be conducted using initial concentration of test compound at parts per billion. These tests, however, require specialized equipment and the custom preparation of appropriately labeled compound is often very expensive.

Die-away biodegradation methods are simple simulation methods that employ water collected from natural water sources and follow the disappearance of an added amount of the test substance resulting from the activity of microorganisms in the water sample. The chemical-specific analytical techniques used to follow the disappearance of the test substance, typically are employed using relatively low initial concentrations of the test substance. Most environmental pollutants are present in the environment at relatively low concentrations (less than 1 mg/L) and it has been observed that biodegradation rates obtained using high test compound concentrations may be quite different from those observed at lower concentrations (6).

The transformation of the test substance to an extent sufficient to remove some characteristic property of the molecule, resulting in the loss of detection by the chemicalspecific analytical technique, is referred to as primary biodegradation. For many purposes, evidence of primary biodegradation is sufficient, especially when it is known or can be shown that toxicity, or some other undesirable feature, associated with the test compound is removed or significantly reduced as a result of the primary biodegradation. A determination of ultimate biodegradation, on the other hand, is usually required only when treatability or organic loading are issues of concern. Furthermore, many of the simpler methods, such as those measuring CO₂ evolution (see 4.1), may not detect primary biodegradation.

The use of low substrate concentration enhances the probability of observing first-order, or pseudo first-order, kinetics. Thus, a rate constant for the primary biodegradation reaction and a half-life can be derived from the test compound under defined incubation conditions. Rate constants are required in many environmental fate mathematical models.

1.1 This test method describes procedures for assessing the biodegradation of chemicals in natural surface water samples.

1.2 This test method provides an opportunity to evaluate rates of biodegradation in the presence and absence of natural sediment materials. It also may provide limited information on the abiotic degradation rate, and sorption to sediment and vessel walls.

1.3 This test method allows for the development of a first-order rate constant, based on the disappearance of the test compound with time, and a second-order rate constant, normalized for changes in microbial biomass.

1.4 This test method requires a chemical specific analytical method and the concentrations of test substance employed are dependent on the sensitivity of the analytical method.

1.5 This test method is designed to be applicable to compounds that are not inhibitory to bacteria at the concentrations used in the test method, which do not rapidly volatilize from water, that are soluble at the initial test concentration and that do not degrade rapidly by abiotic processes, such as hydrolysis.

1.6 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.