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A Spectrophotometric Determination of α-Dicarbonyl Compounds and Its Application to the Enzymatic Formation of α-Ketobutyrate

✍ Scribed by R.S. Li; G.L. Kenyon

Publisher
Elsevier Science
Year
1995
Tongue
English
Weight
304 KB
Volume
230
Category
Article
ISSN
0003-2697

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✦ Synopsis

A simple, rapid, and sensitive spectrophotometric method has been developed for the determination of alpha-keto acids, alpha-diketones, and alpha-ketoaldehydes using o-phenylenediamine as a derivatizing reagent in acidic media to yield chromophoric quinoxaline derivatives. Application to the determination of alpha-ketobutyrate and the rate of its formation from isomerization of vinylglycolate catalyzed by mandelate racemase is described. Under optimal conditions 1 to 4 mM o-phenylenediamine in 80% acetic acid was incubated at 50 degrees C for 10 min with the alpha-dicarbonyl compounds. Solutions of alpha-ketobutyrate as low as 5 microM in the buffer solution can be measured quantitatively. This method has also been used for determining the rate of formation of the phenylglyoxalate from (S)-mandelate catalyzed by (S)-mandelate dehydrogenase.

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