Abstract
Increasing numbers of laboratories develop new methods based on gas-liquid and high-performance liquid chromatography to determine serum concentrations of oxygenated cholesterol metabolites such as 7α-, 24(S)-, and 27-hydroxycholesterol. We initiated a first international descriptive oxycholesterol (OCS) survey in 2013 and a second interventional survey 2014 in order to compare levels of OCS reported by different laboratories and to define possible sources of analytical errors. In 2013 a set of two lyophilized serum pools (A and B) was sent to nine laboratories in different countries for OCS measurement utilizing their own standard stock solutions. In 2014 eleven laboratories were requested to determine OCS concentrations in lyophilized pooled sera (C and D) utilizing the same provided standard stock solutions of OCS. The participating laboratories submitted results obtained after capillary gas-liquid chromatography-mass selective detection with either epicoprostanol or deuterium labelled sterols as internal standards and high-performance liquid chromatography with mass selective detection and deuterated OCS as internal standard. Each participant received a clear overview of the results in form of Youden-Plots and basic statistical evaluation in its used unit. The coefficients of variation of the concentrations obtained by all laboratories using their individual methods were 58.5–73.3% (survey 1), 56.8–60.3% (survey 2); 36.2–35.8% (survey 1), 56.6–59.8, (survey 2); 61.1–197.7% (survey 1), 47.2–74.2% (survey 2) for 24(S)-, 27-, and 7α-hydroxycholesterol, respectively. We are surprised by the very great differences between the laboratories, even under conditions when the same standards were used. The values of OCS's must be evaluated in relation to the analytical technique used, the efficiency of the ample separation and the nature of the internal standard used. Quantification of the calibration solution and inappropriate internal standards could be identified as major causes for the high variance in the reported results from the different laboratories. A harmonisation of analytical standard methods is highly needed.
Highlights
Serum or plasma concentrations of the cholesterol precursors lanosterol, lathosterol, and desmosterol are widely used as surrogate markers of endogenous cholesterol synthesis [1]
These non-cholesterol sterols (NCS) show even stronger correlations with cholesterol absorption and synthesis when expressed as ratios to total cholesterol, which standardizes for variations in sterol transport protein concentrations [4]
The first part of the cholesterol and NCS survey was initiated in the year 2013 under the expertise of the German Reference Institute for Bioanalytics (RfB) and the Laboratory for Specialized Lipid Diagnostic of the University Hospital, both located in Bonn, Germany
Summary
Serum or plasma concentrations of the cholesterol precursors lanosterol, lathosterol, and desmosterol are widely used as surrogate markers of endogenous cholesterol synthesis [1]. The cholesterol metabolite 5α-cholestanol and the plant sterols campesterol and sitosterol are used as markers of cholesterol absorption [2,3] These non-cholesterol sterols (NCS) show even stronger correlations with cholesterol absorption and synthesis when expressed as ratios to total cholesterol, which standardizes for variations in sterol transport protein concentrations [4]. The interest in surrogate markers of cholesterol metabolism is recently increasing and more laboratories develop new methods for the determination of cholesterol and NCS using different chromatographic separation and mass spectrometric detection methods. We reflect the comparability of cholesterol and NCS concentrations determined by different separation and detection methods and discuss the suitability of different compounds used as internal standards for sterol quantification. The focus was on the influence of the utilized calibration solutions and the participants were requested to use provided stock solutions for the quantification of the sample material
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