Stable isotope studies on the biosynthesis of lipoic acid in Escherichia coli

A method has been developed for the gas chromatographic-mass spectrometric (GC-MS) identification of lipoic acid in tissue. The method consists of acid hydrolysis of the tissue to free the bound lipoic acid, methylene chloride extraction of the lipoic acid, and the subsequent chemical derivatization of the lipoic acid as methyl 6,8-bis(benzylthio)octanoate prior to GC-MS analysis. By use of this method of analysis, the incorporation of deuterium into lipoic acid by Escherichia coli growing on [methyl-2H3]acetate has been studied. The results clearly show that the lipoic acid is biosynthesized from octanoic acid with the loss of only one deuterium-containing position at C8. The deuterium incorporated at C6 of octanoic acid from the labeled acetate is retained. Since this deuterium is incorporated in the L configuration during fatty acid biosynthesis and it is known to have the D configuration in lipoic acid, it is concluded that an inversion of configuration occurs at C6 during the sulfur insertion.