Studies of Baker's yeast metabolism: I. Pyridine nucleotide reduction and oxygen utilization during alcohol oxidation

Abstract

The time course of oxygen utilization and pyridine nucleotide reduction during the oxidation of various alcohols by baker's yeast has been investigated. Four distinct phases of metabolism are defined. The fluorometric measurement of pyridine nucleotide reduction was found to reflect qualitatively the state of reduced diphosphopyridine nucleotide produced by the action of ethanol in the cell. This has been assessed by comparative spectrophotometric and direct chemical analysis of pyridine nucleotides. These studies also indicate a fourfold fluorescence enhancement for reduced pyridine nucleotide presumably associated with its binding to some cellular constituent. Differences in the steady-state pattern of pyridine nucleotide reduction with saturated and unsaturated alcohols are attributed in part to the subsequent oxidation of aldehyde. In general, the rate of oxygen utilization parallels the extent of pyridine nucleotide reduced during the aerobic steady state. A concentration of 1.2 mm ethanol is necessary to give a half-maximal stimulation of respiration and pyridine nucleotide reduction. Measurements of CO₂ evolution during alcohol oxidation indicate a delay in the production of CO₂. The measurement of aldehyde formation with the unsaturated alcohol, cinnamyl alcohol, shows a stoichiometric formation of aldehyde with oxygen utilized. Determination of aldehyde concentration during ethanol oxidation, in short-term experiments (less than 5 minutes), suggests that 70% of the oxygen utilized is attributable to oxidation of alcohol to acetaldehyde, and the remaining 30% associated with the oxidation of alcohol, to acid. Ethanol appears to be completely oxidized, however, in long-term experiments.