A RAPID HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC METHOD FOR DETERMINATION OF HOMOCYSTEINE IN PORCINE TISSUE

A simple sensitive method for determination of total L-homocysteine (H C) in tissues is described. The assay involves incubation of a tissue homogenate in the presence of a reducing agent to liberate homocystein e from protein followed by derivatization of thiols with ammonium 7-fl uorobenzo-2-oxa-1,3-diazole-4-sulphonate (SBD-F). Homocysteine is then quantified using fluoresence detection following high performance liq uid chromatography on a 5 mu M reverse phase column. The method was ap plied to heart tissue taken from normal pigs and the results compared to values obtained from pigs intermittently exposed to nitrous oxide ( N2O). It was found that N2O treatment resulted in a reduction in methi onine synthase activity (EC 2.1.1.13) and concomitantly in a massive a ccumulation of homocysteine in both tissue and plasma.