Determination of cholesterol and its six metabolism markers in serum by high performance liquid chromatography-quadrupole/electrostatic field orbitrap high resolution mass spectrometry

Abstract

A method for the determination of cholesterol and its six metabolism markers (desmosterol, lathosterol, campesterol, stigmasterol, β-sitosterol, squalene) in serum by high performance liquid chromatography-quadrupole/electrostatic field orbitrap high resolution mass spectrometry (HPLC-Q/Orbitrap HRMS) was developed. The sample was ultrasonically extracted with acetonitrile. The separation was performed by using 95%(v/v) methanol-acetonitrile (2:8, v/v)-5%(v/v) water as the mobile phase. Isocratic elution was performed at a flow rate of 0.4 mL/min on an Acquity UPLC BEH C18 column (100 mm×2.1 mm, 1.7 μm) in 15 min. The atmospheric pressure chemical ionization (APCI)/positive mode was used in the MS detection. Cholesterol and its six metabolism markers were qualitatively and quantitatively determined by using the full MS scan/date dependent MS2 (Full MS/dd MS2) mode. The resolution of the precursor mass was 70000 FWHM, while the resolution of the product mass was 17500 FWHM. The correlation coefficients (r2) of all the seven target compounds were no less than 0.992 in the linear range. The limits of detection were between 0.8 μg/L and 62.1 μg/L, the recoveries were in the range of 82.1%-97.5% with the relative standard deviations in range of 1.6%-7.4%. The method is accurate, simple, and rapid, and can be used for the determination of cholesterol and its six metabolism markers in serum.