Mass, spectrometry of steroid glucuronide conjugates. II - Electron impactfragmentation of 3-keto-4-en- and 3-keto-5 alpha-steroid-17-O-beta glucuronides and 5 alpha-steroid-3 alpha,17 beta-diol 3- and 17-glucuronides

The steroid glucuronide conjugates of 16,16,17-d(3)-testosterone, epitestos terone, nandrolone (19-nortestosterone),16,16,17-d(3)-nortestosterone methy ltestosterone,metenolone,mesterolone,5 alpha -androstane-3 alpha, 17 beta - diol, 2,2,3,4,4-d(5)-5 alpha -androstane-3 alpha ,17 beta -diol, 19-nor-5 a lpha -androstane-3 alpha ,17 beta -diol, 2,2,4,4-d(4)-19-nor-5 alpha -andro stane-3 alpha ,17 beta -diol and 1 alpha -methyl-5 alpha -androstane-3 alph a/beta, 17 beta -diol were synthesized by means of the Koenigs-Knorr reacti on. Selective 3- or 17-O-conjugation of bis-hydroxylated steroids was perfo rmed either by glucuronidation of the corresponding steroid ketole and subs equent reduction of the keto group or via a four-step synthesis starting fr om a mono-hydroxylated steroid including (a) protection of the hydroxy grou p, (b) reduction of the keto group, (c) conjugation reaction and (d) remova l of protecting groups. The mass spectra and fragmentation patterns of all glucuronide conjugates were compared with those of the commercially availab le testosterone glucuronide and their characterization was performed by gas chromatography/mass spectrometry and nuclear magnetic resonance spectrosco py. For mass spectrometry the substances were derivatized. to methyl esters followed by trimethylsilylation of hydroxy groups and to pertrimethylsilyl ated products using labelled and unlabelled trimethylsilylating agents. The resulting electron ionization mass spectra obtained by GC/MS quadrupole an d ion trap instruments, full scan and selected reaction monitoring experime nts are discussed, common and individual fragment ions are described and th eir origins are proposed. Copyright (C) 2001 John Wiley & Sons, Ltd.