Effect of the Systemic Fungicide Carboxin on Electron Transport Function in Membranes of Micrococcus denitrificans

Abstract

The systemic fungicide carboxin (5,6-dihydro-2-methyl-1,4-oxathiin-3-carboxanilide) inhibited oxidation of succinate by membranes prepared from Micrococcus denitrificans , the K i being 16 μM. Oxycarboxin (5,6-dihydro-2-methyl-1,4-oxathiin-3-carboxanilide-4,4-dioxide), F831 (5,6-dihydro-2-methyl-1,4-oxathiin-3-carboxanilide-4-oxide), and another succinate oxidase inhibitor, 4,4,4-trifluoro-1-(2-thienyl)-1,3-butanedione (TTB) were less effective inhibitors of succinate oxidation by membranes of M. denitrificans . Oxidation of other substrates (nicotinamide adenine dinucleotide, reduced form, d -lactate, l -lactate, malate, and d,l -α-hydroxybutyrate) was inhibited to a lesser degree by carboxin, and formate oxidation was entirely resistant. With all substrates tested, oxycarboxin, the dioxide analogue of carboxin, was less effective than carboxin. Carboxin also inhibited dichlorophenol indophenol (DCIP) reductase activities by these membranes in a manner both qualitatively and quantitatively similar to the inhibition of oxidation of the various substrates. The inhibition of DCIP reductase activities by TTB was qualitatively similar to carboxin, but TTB was a less effective inhibitor with all substrates tested. The inhibition of DCIP reductase by carboxin could be relieved by phenazine methosulfate with all substrates except d -lactate. Only slight inhibition of d -lactate-stimulated uptake of [ 14 C]glycine by these membrane vesicles was seen with carboxin. Uptake of [ 14 C]glycine could be stimulated to varying degrees with the other substrates tested, but in no case did carboxin cause significant inhibition. Membranes isolated from M. denitrificans are a useful system for investigating the mechanism of inhibition of electron transport function by carboxin, and the use of this system for evaluations of carboxin and its metabolites is suggested.