Oxygen-sensitive and -insensitive nitroreduction by Escherichia coli and rat hepatic microsomes.

F J Peterson,R P Mason,J Hovsepian,J L Holtzman

doi:10.1016/s0021-9258(18)50687-6

Abstract

Nitrofurazone is shown to undergo an initial 1-electron (oxygen-sensitive) or 2- or more electron (oxygen-insensitive) reduction by partially purified nitroreductases from Escherichia coli. Nitrofurazone (50 micronM) is reduced by the oxygen-sensitive reductase to a nitro anion free radical as indicated by ESR and visible spectroscopy. The visible spectrum of the nitro anion free radical is characterized by an increase in absorption at 406 nm. In the presence of the oxygen-sensitive reductase, nitrofurazone stimulates superoxide formation and oxygen consumption. This enzyme gives a steady state radical concentration which is proportional to the square root of the enzyme concentration, suggesting that the nitrofurazone anion radical is an obligate intermediate in the reduction and that the radical decays by a nonenzymatic second order process. The oxygen-insensitive reductase does not form the nitro anion free radical nor in the presence of nitrofurazone does it stimulate oxygen consumption. Visible spectroscopy shows that nitrofurazone is reduced by the oxygen-sensitive reductase to a species with an absorption maximum at 335 nm, which has been previously identified as the amine. The oxygen-insensitive reductase reduces nitrofurazone to a previously identified cyano derivative with an absorption maximum at 280 nm. Rat hepatic microsomes appear to metabolize nitrofurazone in a manner similar to the oxygen-sensitive E. coli reductase.

Highlights

Nitrofurazone is shown to undergo an initial l-electron or Z- or more electron reduction by partially purified nitroreductases from Escherichia coli
Visible spectroscopy shows that nitrofurazone is reduced by the oxygensensitive reductase to a species with an absorption maximum at 335 nm, which has been previously identified as the amine
Nitroreduction by E. coli and Liver Microsomes-Nitrofurazone metabolism by E. coli and microsomal nitroreductases was indicated by a decrease in optical density at 400 nm (Table I)

Summary

Introduction

Nitrofurazone is shown to undergo an initial l-electron (oxygen-sensitive) or Z- or more electron (oxygeninsensitive) reduction by partially purified nitroreductases from Escherichia coli. Nitrofurazone (50 pM) is reduced by the oxygen-sensitive reductase to a nitro anion free radical as indicated by ESR and visible spectroscopy. In the presence of the oxygen-sensitive reductase, nitrofurazone stimulates superoxide formation and oxygen consumption. The oxygeninsensitive reductase does not form the nitro anion free radical nor in the presence of nitrofurazone does it stimulate oxygen consumption. The oxygen-insensitive reductase reduces nitrofurazone to a previously identified cyano derivative with an absorption maximum at 280 nm. Escherichia coli contain two different nitrofurazone-reducing activities distinguishable primarily by their sensitivity to oxygen [2, 3]. The oxygen-sensitive enzyme (Reductase II), which consists of at least two different enzymes, is present in both the wild type and the mutant strain of E. coli [4, 5]. Many oxygen-sensitive nitroreductases have been described in mammalian systems (s-12)

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Conclusion