br Results and discussion The results and
Results and discussion The results and subgroup analysis of cholesterol and NCS from the both surveys are listed in Table 2 and shown in Fig. 1, A1 to E2.
Common discussion Unlikely to the other NCS assays, a target value for cholesterol could be reported in the present survey. For NCS assays the reported values could only be compared to the mean of values obtained from all participants or to a corrected mean value after removal of defined outlier. Thus, the true NCS analyte concentration could not be referred with certainty in any of these assays. Absolute reference methods have been developed for some analytes of diagnostic importance but for most analytes there is no “golden method” and we are thus restricted to compare results between different laboratories [9,10]. However, with the collected survey data on hand it is not possible to sufficiently evaluate the best chromatographic method for cholesterol and NCS determination in 87 7 or serum. It can be generally considered that the quality of the present type of cholesterol and NCS analysis is directly dependent upon the skill and experience of the analyst, the limitations of the analysis method, the sample quality and sampling itself, as well as on the quality of the calibration material and used internal standards. Within the survey six different methods were used whereby all measurements dependent upon a critical chromatographic step, either gas- or liquid chromatography. Due to higher chromatographic efficiency and the reduced influence of matrix effects on ionisation and signal suppression, one would expect less analytical problems with gas chromatography compared to liquid chromatography. As ideal standard for cholesterol and NCS analysis it can be expected that the use of the same deuterium or 13C isotope labelled molecule will result in the most precise mass spectrometric analysis. In respect to that it is to mention that the internationally accepted reference method for cholesterol is based on isotope-dilution mass-spectrometry combined with GC [8,9,, , ].
Fundings We gratefully acknowledge funding for materials, statistics, and travel expenses to the Foundation for Pathobiochemistry and Molecular Diagnostics, German Society of Clinical Chemistry and Laboratory Medicine, Germany. The work in University of Valencia was financed by CICYT-FEDER (AGL2008−02591-C02-01) and MINECO-FEDER (AGL2012−39503-C02-01). The work at Universitario Miguel Servet, Zaragoza, Spain was funded by Fondo de Investigación Sanitaria (FIS; PI15/01983). Work in Swansea was supported by the UK Biotechnology and Biological Sciences Research Council (BBSRC, grant numbers BB/I001735/1 and BB/L001942/1).
Acknowledgements The basic initiatives for the performance of these surveys were started during and the results presented at the annual meetings of the European Network for Oxysterol Research (ENOR; https://www.oxysterols.net/). Most of the survey participants are members of the ENOR group.
Introduction Cholesterol (Chol) metabolism consists of a complex set of interactive input and output fluxes with the aim to provide the large membrane bound Chol pool with sufficient Chol molecules. For this purpose, two input fluxes are active. One is the absorption of dietary Chol, the other is de novo Chol synthesis (ChS). Furthermore, Chol is also catabolized to produce steroid hormones and bile acids. Bile acid formation is quantitatively most important. Bile acids serve to keep biliary Chol dissolved in the bile and to support transit of dietary Chol and fat through the small intestine . Bile acids are also involved in the regulation of their own synthesis and that of glucose and lipid metabolism , . High serum Chol concentration, and in particular that of LDL-Chol, is considered a risk factor for atherosclerosis. The accumulation of serum LDL-Chol can be explained by a malfunctioning control of the LDL receptor-mediated uptake of Chol into peripheral tissues or an oversupply due to an excessive de novo synthesis or a high dietary intake. In the ultimately controlled situation, Chol absorption, Chol de novo synthesis and bile acid formation are interactive. Increased Chol absorption is thus compensated by reduced ChS and increased bile acid synthesis (BAS) . The fractional absorption of Chol (FrChA) from the intestine has been measured using radioactive and stable isotope techniques applying continuous dual isotope feeding and feces collection , ,  or single dual isotope administration and blood sampling , . In individual healthy subjects, values between 10% and 80% have been reported. While in textbook terminology, the mean FrChA is reported as 50%, mean values obtained in groups of healthy subjects vary between 24% and 70% . Stable isotope enrichments of Chol are low and measured with complex gas chromatography/mass spectrometry techniques. Isotope techniques were developed to study the incorporation of 13C-acetate or 2H2O into plasma cholesterol in order to express ChS activity , , , . A high fraction of newly synthesized Chol represents a high synthesis rate. However, no information is obtained on the daily ChS. Therefore, the classic way of ChS measurement is the cholesterol balance method . Based on total body Chol metabolism, Chol and bile acids excreted with the feces are compensated by ChS and daily dietary Chol intake (DICh). Measurement of daily fecal Chol (FChE) and bile acid excretion (FBAE) allows the calculation of ChS as [(FChE+FBAE)−DICh]. The measurement of Chol catabolism involves the measurement of BAS. This can be performed with the measurement of FBAE based on the steady state principle that BAS compensates for FBAE. Alternatively, stable isotope labeled bile acids are administered and the decay of isotope enrichment of bile acids is measured in blood over a period of four days , , . A direct comparison of both methods has been published  indicating that lower values are obtained with the Chol balance method.