QUANTITATION OF TOTAL HOMOCYSTEINE IN HUMAN PLASMA BY DERIVATIZATION TO ITS N(O,S)-PROPOXYCARBONYL PROPYL ESTER AND GAS-CHROMATOGRAPHY MASS-SPECTROMETRY ANALYSIS

Much evidence supports the hypothesis that mild or moderate hyperhomoc ysteinaemia represents an important and independent risk factor for oc clusive vascular diseases. Therefore, the accurate and reliable determ ination of total plasma homocysteine has gained major importance for r isk assessment. Furthermore, it can help in the detection of folate an d vitamin B-12 deficiency. This has prompted us to develop a sensitive gas chromatography-mass spectrometry (GC-MS) method in order to quant ify total homocysteine in human plasma. Prior to chromatography, reduc ed homocysteine was released from disulfide bonds by incubation with e xcess dithiothreitol and converted into its N(O,S)-propoxycarbonyl pro pyl ester by derivatization with II-propyl chloroformate. Aminoethylcy steine served as internal standard. The method proved to be highly lin ear over the entire concentration range examined (corresponding to 0-2 66 mu M homocysteine) and showed intra-assay and inter-assay variation (relative standard deviations) of approximately 5 and 5-10%, respecti vely. External quality control by comparison with duplicate analyses p erformed on a HPLC-based system revealed satisfactory correlation. The newly developed GC-MS based method provides simple, reliable and fast quantitation of total homocysteine and requires only inexpensive chem icals, which are easy to obtain. (C) 1997 Elsevier Science B.V.