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Steroids in porcine follicular fluid analysis by liquid chromatography capillary gas chromatography and capillary gas chromatography mass spectroscopy after purification on sep pak c 18 and ion exchange chromatography



Steroids in porcine follicular fluid analysis by liquid chromatography capillary gas chromatography and capillary gas chromatography mass spectroscopy after purification on sep pak c 18 and ion exchange chromatography



Steroids 41(4): 549-567



Steroids in porcine follicular fluid have been concentrated by reverse phase chromatography on SEP-PAK C18 and purified further on the cation exchanger SP- Sephadex C-25. Fractionation into unconjugated neutral and phenolic steroids, glucuronides and sulfates was carried out on triethylaminohydroxypropyl Sephadex LH-20 (TEAP-LH-20). The unconjugated neutral fraction was analyzed by high pressure liquid chromatography (HPLC) on a C18 radial cartridge 5 mm I.D.; 10 .mu., or on a C18 5 .mu. RESOLVE column, and by capillary gas chromatography (GC) on a 12 M OV-1 cross linked fused silica column. Testosterone, progesterone and androstenedione were the major steroids detected by HPLC monitored at 254 nm, although 17-hydroxy-, 20.alpha.-dihydro- and 20.beta.-dihydroprogesterone were also present. Pregnenolone, pregnanediol, dehydroepiandrosterone, 17-hydroxypregnenolone and androsterone were detected by capillary GC as their O-methyloxime trimethylsilylether derivatives. Further confirmation of structure was provided by complete mass spectral data or by selective ion monitoring (SIM).

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