Determination of α-keto acids as silylated oximes in urine and serum by combined gas chromatography-mass spectrometry

Nine α-keto acids in serum and urine were analyzed by gas chromatography as their oxime-trimethylsilylated derivatives, following oxime formation with hydroxylamine in pyridine and silylation with BSA containing 10% TMCS. The oxime-TMS derivatives were identified by both low- and high-resolution mass spectrometry. Recoveries from urine were greater than 90% and from serum greater than 80%. Loads of valine, leucine and isoleucine are shown to result in increased serum concentrations of corresponding α-keto acids in healthy adults. In a child with maple-syrup-urine disease under treatment by dietary restriction of the branchedchain amino acids, administration of leucine results in a large excretion of α-keto isocaproic acid and a less striking excretion of α-keto-β-methylvaleric and α-keto isovaleric acid. The excretion of the branched-chain α-keto acids is reported in two patients with maple-syrup-urine disease before and after institution of dietary treatment. Nine α-keto acids and at least 15 phenolic acids and related compounds can be determined in a single analysis using the method described. The ‘metabolic profile’ thus obtained may be used in screening for abnormalities in the metabolism of amino acids. The gas Chromatographic analysis alone is adequate for screening; when an abnormal quantity of a constituent is found, mass spectrometry is used for confirmation of identity.