Studies on the reduction of nitroblue tetrazolium chloride mediated through the action of NADH and phenazine methosulphate

Abstract

The general features of the reduction of nitroblue tetrazolium chloride (NBT) by NADH and phenazine methosulphate (PMS) have been studied under aerobic and anaerobic conditions. Under aerobic conditions the reduction appears to be mediated through the intermediate formation of the superoxide anion radical O 2 −; this reaction is strongly inhibited by superoxide dismutase and by a number of O 2 − scavengers such as propyl gallate, (+)-catechin, manganous ions, reduced glutathione and benzoquinone. Cupric ions inhibited the overall reaction by reoxidising reduced PMS. Under anaerobic conditions, superoxide dismutase had only a small inhibitory action and, with the exception of cupric ions, the other substances mentioned above were ineffective as inhibitors. The data presented show that the use of NBT to detect the presence of O 2 − is fraught with difficulties due to an equally rapid reduction of NBT by NADH and PMS under anaerobic conditions.