Evaluation of an in Vitro Assay of Lecithin:Cholesterol Acyl Transfer Rate in Plasma

Abstract

Crucial and previously criticized points in a slightly modified version of the Stokke-Norum assay of fractional lecithin:cholesterol acyl transfer (LCAT; rate in plasma were studied. LCAT activity in the albumin added to the assay medium and negative influence of remaining organic solvent were important sources of error that it was necessary to eliminate. The assumption of equilibration of labeled cholesterol among lipoproteins in vitro was supported experimentally. Addition of isolated chylomicrons had no influence on initial LCAT rate. Incubation time was decreased to 20 minutes to obtain a better estimate of initial LCAT rate in normals. Using gas-liquid chromatography to determine unesterified cholesterol concentration, the precision ot measurement of molar LCAT rate was 5.2 per cent (coefficient of variation). Molar LCAT rate in healthy males 20–60 years of age was 56–130 μmol. 1⁻¹ h⁻¹ (mean ± 2 S. D.) and compared well with reports from other laboratories and in vivo measurements. At present this methodology is considered to be suitable for studies of LCAT and cholesterol turnover in different metabolic situations and clinical materials.

Key Words:

• Albumins
• cholestane
• cholesterol
• gas-liquid chromatography
• lecithin
• cholesterol acyltransferase