Spectrophotometric enzymatic assay for S-3-hydroxyisobutyrate

✍ Scribed by Paul M. Rougraff; Ralph Paxton; Gary W. Goodwin; Reid G. Gibson; Robert A. Harris

• Publisher: Elsevier Science
• Year: 1990
• Tongue: English
• Weight: 399 KB
• Volume: 184
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(90)90687-5

✦ Synopsis

An enzymatic spectrophotometric end-point assay has been developed for determination of S-3-hydroxyisobutyrate in biological fluids. The assay measures NADH production at 340 nm after initiation of the reaction with rabbit liver 3-hydroxyisobutyrate dehydrogenase (EC 1.1.1.31). The assay is not affected by R-3-hydroxyisobutyrate, lactate, malate, S-hydroxybutyrate, 2-methyl-3-hydroxybutyrate, 3-hydroxyisovalerate, 3-hydroxy-n-valerate, 2-methyl-3-hydroxyvalerate, and 3-hydroxypropionate.

The assay does measure 2-ethyl-3-hydroxypropionate, a minor metabolite produced by catabolism of alloisoleucine. Application of the method to measure S-3-hydroxyisobutyrate in plasma obtained from normal, 48-h starved, and mildly and severely diabetic rats gave levels of 28,42, 112, and 156 PM, respectively.