Quantification of major lipid classes in human triacylglycerol-rich lipoproteins by high-performance liquid chromatography with evaporative light-scattering detection

Abstract

Triacylglycerol-rich lipoproteins (TRL), comprising chylomicrons (CM) and very-low-density lipoproteins (VLDL), have been associated with cardiovascular disease. The lipid class content in the remnant particles of these lipoproteins is a determinant for the accumulation of lipids in macrophages and their transformation into foam cells. We have optimized a method for the simultaneous determination of cholesteryl esters (CE), triacylglycerols (TG), free cholesterol (FC), monoacylglycerols (MG), and phospholipids (PL) by HPLC coupled to a light-scattering detector (ELSD). A diol column and a ternary gradient of hexane, 2-propanol, and methanol were applied to CM and VLDL of human origin (n = 10), with excellent precision in terms of repeatability of peak areas and retention times. All peaks were baseline resolved although the resolution of CE and TG was compromised for the sake of simplicity of the solvent gradient. The ELSD response was fitted to second-order equations, with correlation coefficients (r 2) higher than 0.999 for a wide range of concentrations (0.25–10 μg of lipid injected). TG were the major lipid class detected in human TRL, accounting for 62% in CM obtained 2 h after the oil intake. In addition we recorded a depletion of TG and CE in CM obtained 2 h after the oil intake of about 60%. We conclude that the method reported here is suitable for a rapid and precise determination of lipid classes in human TRL and, therefore, may be a useful tool for investigations on the atherogenicity of these lipoproteins.