Rapid Isolation of VLDL Subfractions: Assessment of Composition and Susceptibility to Copper-mediated Oxidation

VLDLs, synthesized and released by the liver, are a heterogeneous group of particles of varying composition and metabolic fates. A method is described for the rapid isolation of VLDL into four subfractions (A-D) and assessment of their susceptibility to oxidation. The total isolation procedure required less than 3.5 h, and was achieved by gradient ultracentrifugation. Each subfraction was assessed for triglyceride, cholesterol, and apolipoprotein B (apoB) composition and for the presence of contaminants such as albumin and urate. The oxidation potential, in the presence of copper ions, of each subfraction was also assessed. This rapid procedure produced VLDL fractions analogous to those produced by a previously reported but more prolonged isolation method. Comparison of the two procedures demonstrated that lipid and apoB were similar, while the rapid procedure produced subfractions void of albumin and urate contamination and lower in preformed hydroperoxides. Compositional changes were found between the subfractions: as the subfractions became smaller and more dense (A-->D), there was a decrease in the ratio of triglyceride to apoB and an increase in the ratio of cholesterol to apoB, also arachidonic acid was increased in subfraction D compared with subfractions A, B, and C. The smaller subfractions were more susceptible to oxidation, a trend similar to that reported previously for the oxidation of LDL subfractions.