Fast analysis using pillar array columns: Quantification of branched-chain α-keto acids in human plasma samples

Branched-chain α-keto acids (BCKAs, namely, α-ketoisovaleric acid (KIV), α-ketoisocaproic acid (KIC), and α-keto-β-methylvaleric acid (KMV)) are related to many diseases such as myeloid leukemia, liver cancer, and diabetes mellitus. A rapid quantitative analytical method for BCKAs using pillar array columns was developed. α-Keto acids were labeled with 1,2-diamino-4,5-methylenedioxybenzene (DMB), followed by their separation on octadecylsilane-treated pillar array columns with MeOH/H₂O as the mobile phase. Five DMB-labelled α-keto acids including the internal standard were separated in 160 s. The lower limits of quantification for DMB-α-keto acids were 2-5 μM. The intra- and interday precisions were 2.9-6.6 % and 5.2-10.7 %, respectively. The developed method was applied to BCKA quantification in human plasma samples; KIV, KIC, and KMV concentrations were determined to be 13.8, 24.2, and 15.2 μM, respectively. The method realized rapid, sensitive, and precise analysis of BCKAs and can be applied for clinical diagnosis.