Azo and nitro reductases

Adamson RH, Dixon RL, Francis EL, Rail DP (1965) Comparative biochemistry of drug metabolism by azo and nitro reductase. Proc Natl Acad Sci USA 54 1386-1391... 

The excretion of amines is unusual in animals. Amines are highly toxic and one method employed by vertebrates to detoxify them is via monoamine oxidase, an enzyme which has been detected in H. diminuta (569). Amines can arise from the decarboxylation of the appropriate amino acid, e.g. glycine and alanine can give rise to methylamine and ethylamine, respectively. Another possible source of amines may be the reduction of azo or nitro compounds (39) and azo- and nitro-reductase activity has been reported from M. expansa (180, 181). Furthermore, the physiologically active amines octopamine, dopamine, adrenalin and serotonin (5-hydroxytryptamine) have been demonstrated in cestodes (283, 296, 435, 681, 682, 758, 859), where they probably function predominantly as neurotransmitters (see Chapter 2). 

Douch, P. G. C. (1976a). Azo- and nitro-reductases of the cestode Moniezia expansa. Substrate specificity, reaction products and the effects of flavins and other compounds. Xenobiotica, 6 339-404. 

Dixon DR, Jones IM, Harrison FL (1985) Cytogenetic evidence of inducible processes linked with metabolism of a xenobiotic chemical in adult and larval Mytilus edulis. Sci Total Environ 46 1-8 Douch PGC (1976) Azo- and nitro-reductase activities and cytochromes of Ascaris lumbricoides var. suum and Moniezia expansa. Xenobiotica 6 531-536... 

Reductions Epoxide hydroplase Azo and nitro reduction Carbonyl reductase Disulfide reduction Sulfoxide reduction Quinone reduction Reductive dehalogenation Microsomes, cytosol Gut microflora Cytosol Cytosol Cytosol Cytosol, microsomes Microsomes... 

Cytochrome P450 monooxygenase Azo and nitro group reductase Aldehyde dehydrogenase... 

The enzymes responsible for reduction may be located in both the microsomal fraction and the soluble cell fraction. Reductases in the microflora present in the gastrointestinal tract may also have an important role in the reduction of xenobiotics. There are a number of different reductases which can catalyse the reduction of azo and nitro compounds. Thus, in the microsomal fraction, cytochromes P-450 and possibly a flavoprotein are capable of reductase activity. NADPH is required, but the reaction is inhibited by oxygen. FAD alone may also catalyse reduction by acting as an electron donor. 

Reduction. Reduction, for example azo- and nitro-reduc-tion, is a less common pathway of drug metabolism. Reductase activity is found in the microsomal fraction and in the cytosol of the hepatocyte. Anaerobic intestinal bacteria in the lower gastrointestinal tract are also rich in these reductive enzymes. A historical example concerns Prontosil, a sulfonamide prodrug. It is metabolized by azo-reduction to form the active metabolite, sulfanilamide. Sulfasalazine is also cleaved by azoreduction by intestinal bacteria to form aminosalicylate, the active component, and sulfapyridine. Chloramphenicol is metabolized by... 

In addition to the oxidative systems, liver microsomes also contain enzyme systems that catalyze the reduction of azo and nitro compounds to primary amines. A number of azo compounds, such as Prontosil ahd sulfasalazine (Fig. 10.16), are converted to aromatic primary amines by azoreductase, an NADPFI-dependent enzyme system in the liver microsomes. Evidence exists for the participation of CYP450 in some reductions. Nitro compounds (e.g., chloramphenicol and nitrobenzene) are reduced to aromatic primary amines by a nitroreductase, presumably through hitrosamine and hydroxylamine intermediates. These reductases are not solely responsible for the reduction of azo ahd hitro compouhds reductioh by the bacterial flora ih the anaerobic environment of the intestine also may occur. 

B. Reduction The enzymology of reduction is not as well as characterized as for oxidation but, for example, reductive reactions can be catalyzed by cytochrome P-450 and P-450 reductase and soluble enzymes such as DT-diaphorase (EC 1.6.99.2) Many compounds including azo-and nitro-compounds, epoxides, heterocycles and halogenated hydrocarbons Sources of reducing equivalents for the reactions include NAD PH and NADH. Chemical groups modified include nitro, nitroso, tertiary amine oxide, hydroxylamine, azo, quinone, nitroso, alkylhalide... 

Aromatic nitro and azo compounds are reduced by hepatic enzymes of mammals, birds, reptiles, fish, and bacteria. Azo-reductase and nitro-reductase are similar in that they have low affinities for their substrates in addition, they are both flavoproteins having FAD as their prosthetic group. Fouxs et al. [57]... 

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

Major reduction reactions are azo reduction and nitro reduction. The enzymes (reductases) are found in gut flora but also mammalian tissues. Reduction catalyzed by cytochrome P-450 can occur (e.g., dehalogenation). DT diaphorase carries out two-electron reductions. 

Biorcduction of nitro compounds is carried out by NADPH-dependent microsomal and soluble nitro reductases present in the liver. A multicomponent hepatic microsomal reductase system requiring NADPH appears to be responsible for azo reduction. " " In addition, bacterial reductases present in the intestine can reduce nitro and azo compounds, especially those that are absorbed poorly or excreted mainly in the bile. - ... 

The ej. can perform yet other reactions such as dechlorination of chlorinated aliphatic hydrocarbons. This can be either oxidative, in which case the products are ketones, or reductive, as with carbon tetrachloride which produces the highly toxic free radical CH2 (Salmon, Jones and Mackrodt, 1981). Moreover, the ej. carry at least two reducing enzymes a nitro-reductase and an azo-reductase, both of which produce primary amines. 

Yamashina and co-workers (94, 95) showed that azoxy-, azo- and hydrazo-com-pounds were not reduced by bacterial nitroreductase. Their results supported the observations of Saz and coworkers (78, 78, 80, 81) on the occurrence of individual nitro andnitroso reductase activities, and inhibitor studies indicated the involvement of metals in enzymatic activity. Furthermore, these studies contradicted the earlier work of Egami et al. (24) which indicated that nitroreductase activity could be attributed to nitrite reductase. 

While the cytochrome P-450 monooxygenase reaction described in Eq. (1) often involves hydroxylation of carbon, many other reactions are catalyzed by these enzyme systems. These reactions include oxidation of nitrogen and sulfur, epoxidation, dehalogenation, oxidative deamination and desulfuration, oxidative N-, O-, and S-dealkylation, and peroxidative reactions (56). Under anaerobic conditions, the enzyme system will also catalyze reduction of azo, nitro, N-oxide, and epoxide functional groups, and these reductive reactions have been recently reviewed (56, 57). Furthermore, the NADPH-cytochrome P-450 reductase is capable of catalyzing reduction of quinones, quinonimines, nitro-aromatics, azoaromatics, bipyridyliums, and tetrazoliums (58). 

A number of functional groups, such as nitro, diazo, carbonyls, disulfides, sulfoxides, and alkenes, are susceptible to reduction. In many cases it is difficult to determine whether these reactions proceed nonenzymatically by the action of biological reducing agents such as NADPFI, NADH, and FAD or through the mediation of functional enzyme systems. As noted above, the molybdenum hydroxylases can carry out, in vitro, a number of reduction reactions, including nitro, azo, A-oxidc, and sulfoxide reduction. Although the in vivo consequences of this are not yet clear, much of the distribution of reductases described below may be, in whole or in part, the distribution of molybdenum hydroxylases. 

Although insects contain reductases that catalyze the reduction of xenobiotics, reduction is less common than oxidation. Three types of reduction reactions, i.e., nitro reduction, azo reduction, and aldehyde or ketone reduction, are known to occur in insects. 

Microsomal reduction of many compounds is mediated by NADPH-cytochrome P-450 reductase. Quinones, nitro compounds, and azo compounds are just a few examples of materials that will accept an electron to form the corresponding radical anion. Many such radical anions have been fully characterized and identified on the basis of their hyperfine structure and g values. The primary radicals in each case are tumbling freely in solution, although prolonged reaction sometimes leads to immobilized species. 

Reductive biotransformations of several compounds such as polyhalogenated, keto, nitro and azo derivatives, are catalysed by a variety of enzymes which differ according to the substrates and the species. The liver cytochrome P-450-dependent drug metabolizing system is capable of reducing Af-oxide, nitro and azo bonds, whereas the cytosolic nitrobenzene reductase activity is mainly due to cytochrome P-450 reductase, which transforms nitrobenzene into its hydroxylamino derivative. NADPH cytochrome c reductase is also able to catalyse the reduction of nitro compounds. These metabolic conversions may also be brought about by gastrointestinal anaerobic bacteria. 

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