Toxicity-directed fractionation of tannery wastewater using solid-phase extraction and luminescence inhibition in microtiter plates

A four-step solid-phase extraction (SPE) method is presented for toxicity-directed fractionation of industrial wastewater. This fractionation procedure serves as a key step for identifying unknown organic toxicants in complex samples. Toxicity was determined as luminescence inhibition of Vibrio fischeri using microtiter plates. This method was compared to standard tests in glass cuvettes using both 37 standard chemicals and 24 wastewater fractions with EC₅₀ values covering five orders of magnitude. Results of both methods correlated well. 22 tannery wastewater samples were sequentially extracted using C₁₈e and polystyrene-divinylbenzene phases in combination with pH-changes. Final solid-phase filtrates showed low inhibition, so toxicity of inorganics could be neglected. Using 1/EC₅₀ values, the SPE eluates showed clearly different toxicity patterns. Even in eluates of the fourth extraction step, high toxic effects could be observed. In several cases, luminescence inhibition was increasing at the anaerobic treatment step compared to the corresponding untreated samples. After aerobic treatment, toxicity of most wastewater fractions was greatly diminished. HPLC/DAD analyses of the wastewater fractions showed a fair separation concerning compound polarity. However, the samples were still too complex to identify single compounds responsible for the detected toxicity. Therefore, a further clean-up step accompanied with toxicity testing is needed.