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Assay for methylmalonyl coenzyme A mutase activity based on determination of succinyl coenzyme A by ultrahigh-performance liquid chromatography tandem mass spectrometry.

Abstract
Methylmalonic acidemia (MMA) is an inherited metabolic disease. In this condition, metabolism from methylmalonyl coenzyme A (CoA) to succinyl-CoA is inhibited because of either low methylmalonyl-CoA mutase (MCM) activity or adenosylcobalamin deficiency owing to altered vitamin B12 metabolism. A high-precision assay for detecting MCM activity would facilitate not only MMA diagnosis but also the ability to determine the severity of MMA. We developed an MCM assay method based on ultrahigh-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) that involves the determination of succinyl-CoA, which is formed in an enzyme reaction, using peripheral lymphocytes. Using 0.05, 0.5, and 5 ╬╝mol/L succinyl-CoA, the intra-assay coefficient of variation (CV) was less than 5.2% and the inter-assay CV was less than 8.7%. The MCM activities of five healthy individuals and four patients were investigated with this assay. The MCM activities of the patients were very low in relation to those of healthy individuals. Together, these results show that the UPLC-MS/MS method is useful for a detailed MCM activity assay.
AuthorsKana Gotoh, Yoko Nakajima, Go Tajima, Yuji Hotta, Tomoya Kataoka, Yoshihiro Kawade, Naruji Sugiyama, Tetsuya Ito, Kazunori Kimura, Yasuhiro Maeda
JournalAnalytical and bioanalytical chemistry (Anal Bioanal Chem) Vol. 407 Issue 18 Pg. 5281-6 (Jul 2015) ISSN: 1618-2650 [Electronic] Germany
PMID26018627 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

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