Toxicity Assessment Using a Dilution Series

Toxicity, or more appropriately, inhibition, is a measure of the interference of a compound or compounds with biodegradation. Since inhibition usually is expressed as the interference with bioconversion of an oxygen-consuming reaction – or gas producing reaction in the case of anoxic or anaerobic reactions – it is essential that a biodegradable substrate be present when assessing toxicity of specific compounds. A dilution series is one of the most common respirometric approaches used to assess toxicity. The test procedure usually consists of 1) adding a seed culture to a number of respirometer vessels, 2) adding biodegradable substrate, 3) adding adequate amounts of nutrients and minerals to support biological growth and a buffer to maintain pH within a suitable range, and 4) adding the suspected inhibitory chemical or wastewater to produce a range of test concentrations. The number of dilutions should be based on the anticipated range of exposure of the subject chemical or wastewater to a receiving treatment process. A dilution series also requires use of a respirometer that can be operated over the range of substrate concentrations and test conditions needed to assess inhibition effects. An example of a dilution series using four concentrations of phenol – and a measured using a Challenge AER-200 respirometer system – is shown in Figure A. The increasing lag time with oxygen uptake rates increasing to a maximum is a fingerprint for substrate inhibition reactions. An example using wastewater from a petrochemical industry is shown in Figure B. The test program involved analysis of four dilutions of wastewater plus a control. The control in this case was wastewater from the process that would receive the test industrial wastewater. The relative relationship between dilutions showed increased lag and declining oxygen uptake rates as wastewater concentration increased toward full-strength. The first plateau around 20% of COD represents the completion of the biodegradation reaction for the first of at least three groups of organic constituents. The fact that the oxygen uptake rate increased gradually to essentially the same maximum for each dilution is characteristic of substrate inhibition. The oxidation of the second group of organic constituents was complete at around 60% of the COD. Subsequent tests indicated that this wastewater could be added to a regional wastewater treatment process at volume ratios up to 25% of the total flow without contributing significantly to risk of failure.

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