Validation of a gas chromatography–mass spectrometry method for the analysis of sterol oxidation products in serum

A validated gas chromatography–mass spectrometry (GC–MS) detection method for the quantitative analysis of sterol oxidation products (SOPs) in serum is described. After a lipid extraction procedure with chloroform–methanol, a cold saponification and purification by solid phase extraction, oxysterols were derivatizated to form trimethyl-sylil-ethers which were subjected to GC–MS analysis. Calibration curves for cholesterol oxidation products showed determination coefficient (R2) of 1.0, with low values for the coefficient of variation of the response factors (<1%). Detection and quantification limits were below 5 ng/mL and 10 ng/mL, respectively. Recovery data were between 77.65% and 110.29% (CV < 10% for all compounds). Good results were obtained for within- and between-day repeatability, with values below 10%. In conclusion, the method performed is suitable for the determination and quantification of SOPs in serum.