Reversal of Oxythiamin Toxicity in Neurospora

Abstract

Oxythiamin acts as an antimetabolite of thiamin in the rat and in other organisms (1-5). De Caro et al.(5, also 3,4,6) investigated the original finding that oxythiamin increased urinary excretion of thiamin and level of blood pyruvate in the rat, but did not produce the characteristic neuromuscular syndrome produced by administration of pyrithiamin. While both oxythiamin and pyrithiamin could lower the level of thiamin in various tissues, pyrithiamin was much more potent in this respect than oxythiamin. These authors concluded that pyrithiamin could produce the neuromuscular syndrome since it was a more potent inhibitor of thiamin, whereas oxythiamin could not do so in view of its weaker antagonistic action.

The present paper reports our studies on metabolic effects of oxythiamin in Neurospora crassa(8a.) (wild strain) and the reversal of these effects by thiamin and by acetate.

Methods. The fungus was grown on the usual medium(8) reported by Horowitz and Beadle. Suitable additions were made before autoclaving, on reversal of oxythiamin toxicity (Table I) while volatile and heat-labile substances were added at room temperature after sterilization by filtering through Seitz filter pad. After growth period of 72 hours, the mycelia were filtered, dried at 105°C to constant weight and the weights recorded. Culture filtrates were analyzed for pyruvic and total ketoacids according to Friedmann and Haugen(9). In experiments on pyruvic decarboxylase activity, the mycelia were dried with acetone and the acetone powders kept refrigerated. Oxidation of pyruvic acid via Krebs cycle was minimized by addition of malonate and magnesium chloride. 15 mg of the acetone powder were shaken with 2 mg of pyruvic acid (as sodium salt), 2 mg of potassium malonate and 3 mg of magnesium chloride in total volume of 1.6-1.8 ml in a Warburg flask at 37°C.