Determination of cholesterol in erythrocyte membrane

Abstract

Quantification of cholesterol in biological membranes is an important step toward understanding of metabolism of intracellular cholesterol, composition of cell membrane and plasma lipid profile. The aim of our study was to optimize the method for determination of cholesterol in erythrocyte membrane and then to use this method in the determination of cholesterol concentration in erythrocyte membrane in the studied group of blood samples. Cholesterol in erythrocyte membrane was determined in dry lipid extract of erythrocyte membrane by the enzymatic manual CHOD-PAP method. Cholesterol in erythrocyte membrane and plasma lipid parameters (total cholesterol HDL-cholesterol, LDL-cholesterol and triglycerides) were determined in blood samples of 58 females, obtained by routine health control. Lipid parameters were determined by standard biochemical methods. We examined the relationship between cholesterol in erythrocyte membrane and other plasma lipid parameters as well as other atherogenic risk factors (BMI, blood pressure). Optimization of the method for determination of cholesterol in erythrocyte membrane was based on the observation that primary cholesterol standards prepared by dissolving crystal cholesterol cannot be used due to the interference of the dissolved dry extract in organic solvents with the enzymatic reagent. Commercial standard solutions of cholesterol were used for calibration because they contain detergents for solubilisation of the dry extract in enzymatic reagent. The obtained mean value for cholesterol in erythrocyte membrane, as mmol/L erythrocyte is 4.44 ? 1.019; median 4.65 5-th percentile is 2.70, and 95-th percentile is 6.26. In the examined female group we tested cholesterol concentration in erythrocyte membrane according to age. Two groups were formed (females below and above 50 years) using nonparametric t-test no statistically significant difference was found between these two age groups (p>0.05), while plasma lipid parameters of total cholesterol, triglycerides and LDL-cholesterol were different in the examined groups (p<0.05). By Spearmen nonparametric correlation method we found no statistically significant correlation between cholesterol in erythrocyte membrane and other atherogenic risk factors.