Determination of carbonyl oxygen exchange rates in α-ketoacids by gas chromatography-mass spectrometry

✍ Scribed by Tenkasi S. Viswanathan; Charles E. Hignite; Harvey F. Fisher

• Publisher: Elsevier Science
• Year: 1982
• Tongue: English
• Weight: 883 KB
• Volume: 123
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(82)90449-3

✦ Synopsis

A method for determining the rate of exchange of the carbonyl oxygen in ~-ketoglutarate using gas chromatography-mass spectrometry is described. The method is based on a new procedure for quenching the exchange reaction by rapid oxidative decarboxylation of the LYketoacid to the next lower homologous carboxylic acid using concentrated hydrogen peroxide. Using this method the rate constant for carbonyl oxygen exchange in ol-ketoglutarate in 0.1 M imidazole buffer, pH 7.5, was found to be (2.8 + 0.2) X lo-' s-' at 25'C. Data obtained using this technique suggest that hydration is the mechanism for carbonyl oxygen exchange in ru-ketoacids. The method is also applicable to the measurement of oxygen exchange rates of other Lu-ketoacids free in solution and bound in enzyme complexes.

' Abbreviation used: CC-MS, gas chromatographymass spectrometry.