Localization of the Enzyme System Involved in Anaerobic Reduction of Azo Dyes by Sphingomonas sp. Strain BN6 and Effect of Artificial Redox Mediators on the Rate of Azo Dye Reduction

Abstract

The effect of different artificial redox mediators on the anaerobic reduction of azo dyes by Sphingomonas sp. strain BN6 or activated sludge was investigated. Reduction rates were greatly enhanced in the presence of sulfonated anthraquinones. For strain BN6, the presence of both cytoplasmic and membrane-bound azo reductase activities was shown. Various bacterial strains reduce azo dyes under anaerobic conditions. The most generally accepted hypothesis for this phenomenon is that many bacterial cells possess a rather unspecific cytoplasmic azo reductase which transfers electrons under anaerobic conditions via (soluble) flavins to the azo dyes (7, 12, 30, 32). Recently, it was shown that the naphthalenesulfonate-degrading Sphingomonas sp. strain BN6 converted 2-naphthalenesulfonate to some kind of redox mediator (19). It was suggested that these redox mediators enabled the strain under anaerobic conditions to transfer redox equivalents to the azo dyes. In the present study, the location of the enzyme system which is responsible for the reduction of azo dyes by whole cells was determined. The effect of artificial redox mediators on the anaerobic reduction of azo dyes by whole cells from different bacterial strains was also studied. Bacterial strains and media. The isolation and characterization of Sphingomonas sp. strain BN6 (DSM 6383) and the composition of the minimal media have been described before (15, 27). Reduction of amaranth by resting cells. Strain BN6 was grown in a mineral medium with glucose (10 mM). Cells were harvested by centrifugation and resuspended in 50 mM Na-Kphosphate (pH 7.4) buffer to an optical density at 546 nm of 11. This corresponded to a protein concentration of approximately 1.1 g/liter. The reaction mixture contained, in a final volume of 7.2 ml, 80 mol of glucose, 400 mol of Na-K-phosphate buffer (pH 7.4), and different concentrations of the redox mediators. This cell suspension was transferred to a rubber-stoppered serum bottle (30 ml). Oxygen was removed from the medium by at least 15 2-min cycles of evacuation and flushing with nitrogen gas. The reaction was started by the injection of 0.8 ml from an anaerobic stock solution of amaranth (5 or 10 mM). The reduction of the azo dye was determined spectrophotometrically at a of 520 nm. Experiments with activated sludge. The activated sludge was obtained from the aerobic part of the sewage treatment plant of the University of Stuttgart (Büsnau). Particulate material was collected by centrifugation (8,500 ϫ g, 30 min), resuspended in Na-K-phosphate buffer (50 mM; pH 7.4), and passed through a sieve with a mesh size of 1.0 mm to eliminate

Chemical stress-responsive genes from the lignin-degrading fungus Phanerochaete chrysosporium exposed to dibenzo-p-dioxin

“…More recently, NUO was reported to couple the reduction of 2,4,6-trinitrotoluene with the respiratory system in Pseudomonas sp. JLR11 [17]. Thus, two possible roles of NUO are proposed; one is as ATP generator and the other is as a membrane-bound reduction system for the xenobiotic metabolism.…”

Azo Dye Reduction by Mesophilic and Thermophilic Anaerobic Consortia

“…Initially nonspecific enzymes reduce the mediators, which are subsequently chemically reoxidized by the reduction of the azo dyes (16). The reduction of the mediator in whole cells, which can occur either intracellularly (13) or extracellularly by a membrane-bound enzyme (17), will depend on many factors such as its size, polarity, standard redox potential (E 0 ′) and other physical-chemical characteristics (14).…”

Structure–activity Relationship Study on the Bioreduction of Azo Dyes by Clostridium Paraputrificum

“…Redox mediators are reduced by bacterial enzymes, and those reduced mediators can effect the reduction of azo dyes in the absence of bacteria. The intermediacy of redox mediators in azo dye reduction has been supported in a study by Kudlich et al [20], in which the rate of reduction of amaranth was significantly enhanced in the presence of anthroquinone. Brown [4] had suggested that a mediated reduction process would mean minimal structure-activity relationships, but this is not necessarily so.…”

“…Recently, Bragger et al [18] have correlated the half-wave potential of some azo compounds with their rates of reduction, with faster reductions occurring in dyes with less negative half-wave potentials. A number of reports have suggested the intermediacy of low molecular-weight electron carriers, which would act as the de facto reducing agents before being re-reduced themselves by an enzyme [4,19,20].…”