Azo Reductase

Ran J, A Stolz (2003) Oxygen-insensitive nitroreductases NfsA and NfsB of Escherichia coli function under anaerobic conditions as lawsone-dependent azo reductases. Appl Environ Microbiol 69 3448-3455. 

Reduction of the azo group in dyes and colorants is the key reaction in their decolorization. Although an azo reductase mediates the reduction of azo groups to amines, the metabolic situation is quite complex. The enzyme is apparently synthesized under both aerobic (Bliimel et al. 2002) and... 

Anaerobic azo dye reduction can be mediated by enzymes, low molecular weight redox mediators, and chemical reduction by biogenic reductants. These reactions can be located either intracellular or extracellular. Reduction of highly polar azo dyes, which cannot pass through the cell membranes, is located outside the cell. Like azo dyes, nicotinamide adenine dinucleotide phosphate, which is believed to be the main source of electrons, also cannot pass through the cell membranes. Azo reductase enzyme, which is oxygen-sensitive and released extracellularly, is found to be responsible for the reduction of azo dyes. 

Based on the previous publications, azo dye can be reduced by azoreductase-catalyzed reduction under anaerobic conditions. But still there is a speculation whether bacterial flavin reductases are responsible for the azo reductase activity observed with bacterial cell extracts. In a published report, it is reported that flavin reductases are indeed able to act as azo reductases [24]. Bacteria produce extracellular oxidative enzymes, which are relatively nonspecific enzymes catalyzing the oxidation of a variety of dyes. It was reported that so many diverse groups of bacteria play a role in decolorization. It has been also reported that mixed microbial community could reduce various azo dyes, and members of the y-proteabacteria and sulfate reducing bacteria (SRB) were found to be prominent members of mixed bacterial population by using molecular methods to determine the microbial population dynamics [1],... 

Though cycle time plays an important role in the SBR for the decolorization process, not many reports are found in the literature. The long retention times are often applied in the anaerobic phase of the reactor studies, such as 18 and 21 h. In several studies, it was reported that there is a positive correlation between the anaerobic cycle time and the color removal [30, 31]. Indeed, in combined anaerobic-aerobic SBRs, since bacteria shifted from aerobic to anaerobic conditions, or vice versa, anaerobic azo reductase enzyme can be adversely affected by aerobic conditions, which is essential for aromatic amine removal, thereby resulting in insufficient color removal rate. To investigate the effect of cycle time on biodegradation of azo dyes, inar et al. [20] operated SBR in three different total cycle times (48-, 24- and 12-h), fed with a synthetic textile wastewater. The results indicated that with a... 

Zimmermann T, Kulla H, Leisinger T (1982) Properties of purified Orange II azo reductase the enzyme initiating azo dye degradation by Pseudomonas KF46. Eur J Biochem 129 197-203... 

Under aerobic conditions, aerobic bacteria has so far been only found in studies capable of reducing azo compounds and produce aromatic amines by specific oxygen-catalyzed enzymes called azo reductases. These aerobic bacteria could grow with mostly simple azo compounds as sole source of carbon and energy and under strict aerobic conditions by using a metabolism that started with reductive cleavage of the azo linkage. 

The azo reductases in aerobic bacteria were found to be existent when azoreductases from obligate aerobic bacteria were isolated and characterized from strains K22 and KF46 and were shown to be flavin-free after purification, characterization, and comparison 364, 362,363. These intracellular azoreductases showed high specificity to dye structures. Furthermore, Blumel and Stolz cloned and characterized the genetic code of the aerobic azo reductase from Pagmentiphaga... 

As discussed earlier, Azo biological decolorization are mainly reduced in a direct reduction or mediated/indirect reduction with nonspecial azo reductase or reduced enzyme cofactors (Figs. 1 and 3). According to the direct enzymatic reduction mechanism, nonspecial azo reductase can catalyze the transfer of reducing equivalents originating from the oxidation of original electron donor in the azo dyes. In... 

Maier J, Kandelbauer A, Erlacher A et al (2004) A new alkali-themostable azo reductase from Bacillus sp. strain SF. Appl Environ Microbiol 70 837-844... 

Ramalho PA, Paiva S, Cavaco-Paulo A et al (2005) Azo reductase activity of intact Saccharomyces cerevisiae cells is dependent on the Frelp component of plasma membrane ferric reductase. Appl Environ Microbiol 71 3882-3888... 

From the biological point of view, the effect of anaerobiosis has been characterized in purely anaerobic, facultative anaerobic, and aerobic bacteria, in yeasts, and in tissues from higher organisms [6-12], From these studies it can be deduced that almost every azo compound can be biologically reduced under anaerobic conditions [4]. Reduced flavins are produced by cytosol flavin-dependent reductases [6, 13], while quinone reductase activity located in the plasma membrane [14] and extracellular azo reductase activities [9, 15] were also observed. 

A highly salt-tolerant bacterial strain Gracilibacillus sp. GTY was tested for the ability to decolorize the azo dye Acid Red B. It was observed that the dye was decolorized by growing and resting cells, as well as by extracted azo reductase, in optimum conditions and at a 10-15% NaCl concentration at very high and very low salt concentration, it was not possible to reach a good performance in decolori-zation [80],... 

Nitro- and Azo-compounds Nitro and azo reductases Nitroso, hydroxylamino and amino compounds... 

The endoplasmic reticulum is composed of a convoluted network of channels and so has a large surface area. Apart from cytochromes P-450, the endoplasmic reticulum has many enzymes and functions, besides the metabolism of foreign compounds. These include the synthesis of proteins and triglycerides and other aspects of lipid metabolism and fatty acid metabolism. Specific enzymes present on the endoplasmic reticulum include cholesterol esterase, azo reductase, glucuronosyl transferase, NADPH cytochromes P-450 reductase and NADH cytochrome b5 reductase and cytochrome b5. A FAD-containing monooxygenase is also found in the endoplasmic reticulum, and this is discussed later in this chapter. 

We have not pursued mechanisms but suggest that the enhancement of carcinogenesis may be related to a role for riboflavin in the activation of enzymatic processes involved with metabolic detoxification of MBN, similar to azo reductase and its role in the detoxification of 4-dimethylaminoazobenzene (32). In this case riboflavin activates azo reductase in the liver and this, in turn, is associated with decreased carcinogenicity. Conversely, when animals are deprived of riboflavin, there is less active enzyme present to detoxify the chemical and the induction of liver cancer is enhanced. A similar process may be functioning in our MBN, riboflavin deprived model but the exact nature of the mechanism requires additional research. 

Certain benzidine-derived (direct azo) dyes afford, by reduction, generally metabolic degradation in the gut or fiver, the free aryl amine. The azo cleavage may be brought about by gut bacteria and mammalian fiver azo reductases. In 1978, the US National Institute of Occupational Safety and Health (NIOSH) and the Occupational Safety and Health Administration (OSHA) reported that the benzidine-colorants Cl Direct Black 38, Cl Direct Blue 6 and Cl Direct Brown 95 were all reduced to 10a by rat liver in vivo, though trypan blue (35), Congo red (36) and Chicago sky blue were not. However, all six... 

Azo Prodrugs Amines have been incorporated into an azo linkage to form prodrugs that can be activated through azo reduction. In fact, sulfa dmgs were discovered because of prontosil (93), an inactive azo dye that was converted in vivo to the active sulfanilamide (95) (Scheme 15). Clinically useful balsalazide (23), olsalazine (25), and sulfasalazine (26) are azo prodrugs of mesalazine (27). They are converted in vivo by bacterial azo reductases in the gut to the active 5-aminosalicylic acid (5-ASA or mesalazine, 27), which is responsible for their anti-inflammatory activity in the treatment of ulcerative colitis, as discussed earlier. 

The drug is practically insoluble in water (1 g in 10,000 ml). Thus, being poorly absorbed from the small intestine, SZ reaches the colon intact, where bacterial azo-reductase enzymes cleave the compound to its components sulfapyridine (SP) and 5-aminosali-cylic acid (5-ASA). In slow acetylators, where the absorption of SP can lead to plasma levels above 50 pg/ml. It is very likely that it is the sulfonamide that is responsible for the toxic adverse effects associated with SZ. It is ironic that the drug s suppressive effect on ulcerative colitis was at first attributed to the local antibacterial effect of SP. With the dis-... 

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