The effect of phenothiazine on the metabolism of liver mitochondria

Abstract

Phenothiazine at a concentration of 3.3 mM inhibited the oxidation by liver mitochondria of the Krebs' cycle substrates, citrate, α-oxoglutarate and l-malate, the fatty acid, octanoate, and the amino acid, l-glutamate, all of which reactions require NAD coenzymes. NAD-dependent systems of mitochondria were inhibited by quite low concentrations of phenothiazine; for example, 10 μM produced appreciable inhibition of l-glutamate oxidation. The oxidation of succinate, which does not require NAD coenzymes, was not inhibited by 3.3 mM phenothiazine. Phenothiazine also inhibited oxidative phosphorylation by liver mitochondria, regardless of whether an NAD-dependent substrate or succinate was being oxidized. Inhibition of the oxidation of NAD-dependent substrates by phenothiazine was largely prevented by including NAD, GSH and nicotinamide in the respiratory reaction mixture, and it was also slightly prevented by EDTA. The inhibition was shown to be due to the loss of NAD coenzymes from mitochondria during incubation with phenothiazine. Approximately 10 μM phenothiazone or its leuco equivalent was formed when 3-3 mM phenothiazine was incubated for 40 min with respiring mitochondria.