Capillary electrophoresis to monitor the oxidative modification of low density lipoproteins

Abstract

A procedure has been developed that uses high performance capillary electrophoresis to monitor the changes in the electrophoretic mobility of low density lipoproteins (LDL) resulting from Cu2+-catalyzed lipid peroxidation. Using uncoated fused silica capillaries, methylglucamine-Tricine, pH 9.0, as electrophoresis buffer and a field strength of 350 V/cm, separation of native LDL and oxidized LDL could be achieved in 8-10 min. The electrophoretic mobility of native LDL under these conditions was 1.32 x 10(-4) cm2 x V(-1) x s(-1), and the migration time could be measured with a coefficient of variation of 0.44%. The increase in the electronegativity of LDLs during incubation with 10 microM Cu2+ for 0.25-2.0 h resulted in a progressive increase in migration time. Monitoring the absorbance of the migrating LDL particles at a wavelength of 234 nm showed a progressive increase in peak area, which paralleled that in diene conjugation measured spectrophotometrically. Electronegative LDL particles formed by modification with malondialdehyde could also be separated from native LDL particles under these conditions. This new procedure should be useful in studies of factors influencing low density lipoprotein oxidation in vitro and in vivo.