Nonenzymatic acetolactate oxidation to diacetyl by flavin, nicotinamide and quinone coenzymes

• Published in: Biochimica et biophysica acta Vol. 1245; no. 3; pp. 366 - 370
• Main Authors: Park, Sang Ho, Xing, Ruye, Whitman, William B.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 14.12.1995
• Subjects: Acetohydroxy acid synthase; Acetolactate; Benzoquinones - metabolism; Chromatography, Thin Layer; Diacetyl - metabolism; Flavin reduction; Flavins - metabolism; Lactates - metabolism; Methanococcus; NAD reduction; Niacinamide - metabolism; Oxidation-Reduction; Ubiquinone reduction; NAD reduction; Flavin reduction; Tricine; Methanococcus; Acetolactate; AHAS; Ubiquinone reduction; Acetohydroxy acid synthase
• ISSN: 0304-4165; 0006-3002; 1872-8006
• DOI: 10.1016/0304-4165(95)00103-4

Acetolactate nonenzymatically reduced flavins, quinones and nicotinamide coenzymes in a time-dependent manner at physiological pH and moderate temperature. In the presence of excess acetolactate, the reduction of FAD and NAD + followed pseudo-first-order kinetics. The rate of reduction was proportional to the concentration of acetolactate, and the rate constants at 37°C and pH 7.5 were 4.8 · 10 −2 M −1 s −1 and 7.4 · 10 −3 M −1 s −1 for FAD and NAD +, respectively. In contrast, ubiquinone reduction followed pseudo-zero-order kinetics in the presence of excess acetolactate. At 37°C and pH 7.5, the rate of reduction was proportional to the acetolactate concentration, and the apparent rate constant was 8.3 · 10 −6 s −1. In contrast to FAD, the rate of reduction of ubiquinone was higher at low pH. The kinetics of ubiquinone reduction suggested that the rate-limiting step was acetolactate decarboxylation and formation of the enolate anion, whereas the rate of FAD reduction was governed by the second-order reaction of the enolate anion. Following the oxidation, acetolactate was converted to diacetyl. Reduced FAD formed by the reaction with acetolactate generated a low rate of O 2 consumption during assays of the oxygenase activity of acetohydroxy acid synthase. The reaction of acetolactate with quinones may provide a mechanism for the nonenzymatic formation diacetyl in whole milk.