On the nature of the 'nothing dehydrogenase' reaction.
The biochemical mechanism underlying the 'nothing dehydrogenase' reaction during the histochemical demonstration of dehydrogenases using tetranitro BT as the final electron acceptor has been investigated in unfixed, frozen rat liver sections. The reaction is stronger with NAD+ than either with NADP+ or in the absence of coenzyme. As much as 50% of the reaction is due to lactate dehydrogenase converting endogenous lactate and is largely inhibited by pyruvate. No NAD+-dependent alcohol dehydrogenase activity was detected at pH 7.45, the pH used for the incubations. The coenzyme-independent activity may be caused by SH-groups present in proteins and compounds like glutathione and cysteine and can be inhibited by N-ethylmaleimide and p-chloromercuribenzoic acid. It was also found that the 'nothing dehydrogenase' reaction mainly occurs during the first few minutes of incubation, levelling off quickly to a slow rate. When studying the kinetics of dehydrogenase reactions with tetrazolium salts, it should be realized that the 'nothing dehydrogenase' reaction, which as a whole is nonlinear with time, can interfere seriously with the dehydrogenase reaction to be analysed and may yield initial reaction rates that are too high. The findings of the present study reveal the nature of the reactions used for detection of necrosis in tissues with tetrazolium salts.
D000426:Alcohol Dehydrogenase; D000429:Alcohol Oxidoreductases / Q000378:metabolism; D000818:Animals; D002729:Chloromercuribenzoates / Q000494:pharmacology; D005954:Glucosephosphate Dehydrogenase / Q000032:analysis; D006651:Histocytochemistry; D066298:In Vitro Techniques; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000201:enzymology; D008297:Male; D009580:Nitroblue Tetrazolium; D010088:Oxidoreductases / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D020245:p-Chloromercuribenzoic Acid
D002729:Chloromercuribenzoates; D009580:Nitroblue Tetrazolium; D020245:p-Chloromercuribenzoic Acid; D010088:Oxidoreductases; D000429:Alcohol Oxidoreductases; D000426:Alcohol Dehydrogenase; D007770:L-Lactate Dehydrogenase; D005954:Glucosephosphate Dehydrogenase