Application of enzymatic fluorometric assays to quantify phosphatidylcholine, phosphatidylethanolamine and sphingomyelin in human plasma lipoproteins

Abstract

Phosphatidylcholine (PC), phosphatidylethanolamine (PE) and sphingomyelin (SM) are important surface components of plasma lipoproteins, including very-low-density lipoproteins (VLDL), low-density lipoproteins (LDL) and high-density lipoproteins (HDL). However, the pathophysiological roles of PC, PE and SM in lipoproteins have not been well characterized owing to the difficulties in quantifying phospholipid classes in lipoproteins. In this study, we assessed the precision and accuracy of the enzymatic fluorometric assays for measuring PC, PE and SM in VLDL, LDL and HDL, which were isolated from human plasma by ultracentrifugation. The within-run coefficients of variation (CV) for the measurements of PC, PE and SM in lipoproteins were 1.5–2.8 %, 1.1–2.4 % and 0.9–2.3 %, respectively, whereas the between-run CVs for the PC, PE and SM assays were 2.7–4.7 %, 2.1–4.5 % and 1.6–3.3 %, respectively. Excellent linearity and almost complete recovery were achieved for all assays measuring PC, PE and SM in VLDL, LDL and HDL. Our preliminary results using these enzymatic fluorometric assays suggested that the phospholipid compositions were different among VLDL, LDL and HDL. In conclusion, we established high-throughput enzymatic fluorometric assays to quantify PC, PE and SM in human plasma VLDL, LDL and HDL, which will be useful for further investigation of pathophysiological roles of phospholipids in lipoproteins.