Microbial abilities to detoxify chromate by reduction

Abstract

Hexavalent chromium [Cr(VI)] or chromate, is a toxic, water-soluble contaminant present in many soils and industrial eflluents. As a result of contaminated discharges from industrial applications, and inappropriate wastedisposal practices, significant amounts of chromate have found their way into the environment. This poses a health risk to man as well as animals and plants due to the carcinogenicity, mutagenicity, and teratogenicity of chromate. In man, acute, high level exposures to Cr(VI) can result in ulceration of the skin, eyes, and mucous membranes. Exposure of plants to Cr(VI) can result in reduced biomass production, and in extreme cases, death. Upon reduction ofCr(VI) to trivalent chromium [Cr(III], the toxic effects are significantly decreased because of a decrease in the solubility and bioavailability of Cr(III). Traditionally, Cr(VI) has been recovered from aqueous systems using processes exploiting the differential solubility properties described above. The use of chromate reducing bacteria represents a potential mechanism for the development of an efficacious, cost effective alternative to traditional chemical/physical processes for Cr(VI) recovery from the environment. Therefore, the aim of this research was to isolate and identify chromate reducing bacteria from soil, and characterise the chromate reductase enzyme in order to determine the potential of bacteria to detoxify chromate by reduction. Bacteria from soils and wastewater were examined for chromate reducing potential and identified on the basis of biochemical tests and API 20E. Organisms were isolated by the spread plate technique. Species of Pseudomonas maltophilia, Bacillus subtilis, Acinetobacter calcoaceticus, and Cellumonas cellasea were capable of catalyzing the reduction ofCr(VI) to Cr(IlI) in batch experiments. Reduction capability as high as 99% by the isolates was detected from an initial Cr(VI) concentration of 150 mg.L' in batch cultures. Chromate reduction was determined by means of the diphenylcarbazide method and total chromium was measured by atomic absorption spectroscopy. Pseudomonas maltophilia was observed to be the most suited organism for the efficient detoxification ofCr(VI) due to its wide temperature and pH requirements, low substrate utilization, and tolerance to heavy metal ions of'Cu', Cd2+,Zn2+,and Ni2+which commonly appear in industrial eflluents along with Cr(VI). Reduction rate in a batch reactor for this organism was calculated to be 1.75 mg.g+h'. Comparison of the rates of chromate reduction by Cr(VI) grown cells and cells grown without chromate indicated that the chromate reductase activity is constitutive. Reductase activity was detected by means of the lysozyme-EDTA method in aerobically grown cells, with highest specific activity in the cytoplasmic fraction of the cell. The Cr(VI)-reductase was found to be NAD(p)H-dependent and yielded an activity of 3.24 ml.I.mg' of protein in the cytoplasmic fraction. Once optimization of the parameters in the batch reactor was achieved, cells of Ps. maltophilia was immobilized into polyacrylamide gel and packed in a column. Mass balance studies indicated that ca 147 mg.L' chromate passing through the column undergoes reduction with an initial Cr(VI) concentration of 150 mg.L' resulting in a Cr(VI) reduction efficiency of98%. An amount of 0.11 mg.L' remained in the cells, 0.11 mg.L' in the cell wash water, and 1.65 mg.L' was unaccounted for in the mass balance. Chromate reduction rate in the continuous-upflow reactor system was calculated to be 5.34 mg.g'l.h', which was 3-fold higher than that calculated for the batch reactor. Chromium-contaminated industrial eflluent obtained from Sheffield, Natal, and Saayman Danks Electroplaters was pumped into the continuous-upflow reactor containing immobilized cells of Ps. maltophilia to determine the industrial applicability of the reactor to treat chromate-containing effluents. Complete Cr(VI) reduction