Capillary electrophoresis in the evaluation of ischemic injury: simultaneous determination of purine compounds and glutathione

An understanding of tissue energy metabolism and antioxidant status is of major interest in the field of organ preservation for transplantation. Nucleotide and glutathione are indicators of cell damage occurring during ischemia and reperfusion. A high performance capillary electrophoresis (HPCE) method with UV detection (185 nm) for the simultaneous analysis of intracellular free ribonucleotides, nucleosides, bases and glutathione (oxidized and reduced form) in myocardial tissues is described. The method does not involve thiol derivatization. The separations were carried out in an uncoated fused-silica capillary, 60 cm long, 52.5 cm to detector, 75 μm ID, with 20 mM Na-borate buffer, pH 10.00, at 20 kV voltage and reading at 185 nm. Injection was hydrostatic for 12 s and total analysis time was 20 min. The technique enables optimum separation of all the compounds examined and has a resolution similar to that of HPLC analysis, with the advantage of fast simultaneous measurement of cell nucleotide metabolism and redox state, not possible with HPLC.