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«“√ “√ √“™∫— ≥±‘ µ¬ ∂“π ªï ∑’Ë Û ©∫— ∫∑’Ë Ò ¡.§.-¡’ .§. ÚıÙ¯ 63 Somsak Damronglerd, et al. Reduction of different sulfonated azo dyes by whole cells of strain A5 in the presence of different concentrations of AQDS. The addition of quinoid redox mediators resulted in an increased reduction rate for azo compounds by various bacteria (26, 29, 30, 42, 43). To demonstrate the general applica- bility of this system for the treatment of textile wastewaters, various sulfonated azo dyes were incubated under anaerobic condition with whole cells of Paenibacillus sp. strain A5 in the presence of different concentra- tions of AQDS. Thus, it was found that the addition ofAQDS significant- ly increased the rate of decoloriza- tion of various sulfonated azo dyes (Fig. 3). Concentration of AQDS as low as 50 M significantly stimulated all sulfonated azo dyes reduction by cell suspensions of Paenibacillus sp. strain A5 (Fig. 3). The rate of azo dyes reduction was also dependent on the concentration of AQDS added. Comparison of the azo reductase activities in different cellular components of Paenibacillus sp. strainA5 with different industrially sulfonated azo dyes. In the present study it was attempted to determine the location of the enzyme system(s) responsible for the reduction of sulfonated azo dyes by Paenibacillus sp. strain A5 in the presence of AQDS. Therefore, azo reductase activities of cellular membrane and cytoplasm of strain A5 were compared in the presence of AQDS (0.05 mM) in reaction mixtures. Thus, it was found that the majority of AQDS-dependent azo reductases activities present in membrane fraction of strain A5 (Table 2). Up to the present, the most generally accepted hypothesis for bacterial reduction of azo dyes is that many bacterial cells possess a rather unspecific cytoplasmic azo reductases which transfers electrons via soluble or bound-flavins to the azo dyes (45, 56). The involvement of different low molecular weight redox mediators (e.g., flavins and quinones) in the bacterial reduction of azo dyes have been repeatedly suggested (14, 26, 29, 42, 43, 50). In this manuscript, it was also attempted for the first time to perform the comparison of the effects of both flavin-type (FAD) and quinoid-type mediator (AQDS) on azo reductase activities present in different cellular components. Without the addition of any redox mediators (FAD nor AQDS), the specific decolorization rates of orange, violet and black dye of whole cells were significantly lower than the specific decolorization rates of the same dyes determined with cell extracts (Table 2). This suggested that either the cell membrane limited the uptake of highly polar sulfonated azo dyes or the lack of some cofactors (e.g., free flavins) limited the reduc- tion of azo dyes by whole cells. The addition of FAD (0.05 mM) to whole cell suspensions and the cell membrane fraction of Paenibacillus sp. strain A5 did not significantly increase the azo reductase activities of whole cells and slightly increased the azo reductase activities present in membrane fraction. In contrast, it was observed that the addition of the same concentration of FAD in the cytoplasmic fraction of strain A5 resulting in dramatically increased in the specific reduction rates of all azo dyes used. On the other hand, the addition of the same concentration of AQDS, a quinoid redox mediator, was significantly enhanced azo reductase activities of whole cells and greatly stimulated the azo reductase activities present in membrane fraction of strain A5. In cytoplasmic fraction, however, this externally added quinoid mediator had rather lower stimulating effect on azo dyes reduction than the addition of FAD (Table 2). Thus the membrane-bound and cytoplasmic azo reductases of strainA5 are probably different enzyme systems which the latter may have insignificant impor- tance in the reduction of sulfonated azo dyes in vivo. Quinone reductase and flavin reductase activity in membrane and cytoplasmic fraction of strain A5. In cytoplasmic membranes of almost all prokaryotes, it has been shown that the reduction of soluble quinones is catalyzed by the membrane- bound respiratory NADH: quinone reductase (4, 5, 32, 58). Furthermore, it was previously suggested that in Sphingomonas xenophaga BN6 the (membrane-bound) NADH: quinone oxidoreductase of respiratory chain is responsible for the reduction of anthraquinone-2-sulfonate (and thus the azo reductase activity) (29). In the