AN UPDATED LOOK AT THE ANALYSIS OF UNSATURATED C-27 STEROLS BY GAS-CHROMATOGRAPHY AND MASS-SPECTROMETRY

Gas chromatography-mass spectrometry (GCMS) and GC are commonly used m ethods for the identification and quantitation of sterols from samples of biological origin. To investigate the utility and limitations of t hese methods, we have determined gas chromatographic mobilities and ma ss spectral properties of 5 alpha-cholestan-3 beta-ol and 26 unsaturat ed C-27 sterols as their acetate and trimethylsilyl (TMS) ether deriva tives by GC and GC-MS. The GC retention data showed that numerous ster ols were essentially coeluted on capillary GC columns coated with eith er 5% phenyl-95% methyl polysiloxane or polyethylene glycol, although the peaks were more widely dispersed on the latter column. Mass spectr a of many groups of sterol isomers were also quite similar. Sterol mix tures of any complexity are likely to contain coeluting components, an d attempts to establish structures based on mass spectra that may repr esent a mixture of sterol isomers could easily lead to errors. Our res ults demonstrate that GC and GC-MS alone cannot generally be used for rigorous structure determinations of individual components in mixtures of unsaturated sterols. However, all but a few of the 26 sterols coul d be distinguished by their combined chromatographic mobilities on the two GC columns coupled with critical examination of their mass spectr a. GC-MS analysis of appropriate sterol subclasses or preferably indiv idual sterol components obtained by prior purification by other method s may provide valuable supporting evidence for the identification of s terol structures. Reliability of identification is dependent upon care ful attention to GC and MS conditions, calibration of GC and MS data w ith authentic sterol standards, and consideration of possible decompos ition under GC conditions and of the effect of overloading on GC reten tion times.