By using N-methyl-N-trimethylsilyl trifluoroacetamide (MSTFA) or N-methyl-N-trimethylsilyl hepta-fluorobutyramide (MSHFBA) in the presence of 5% trimethylchlorosilane (TMCS) as catalyst, the trimethylsilyl ether esters of the following steroid glucuronides (G) were formed: oestrone 3-G, oestradiol-17β 3-G, oestriol 3-G, 2-hydroxyoestradiol-17β5 2-G, 2-methoxyoestradiol-17β 3-G, 2-hydroxyoes-tradiol-17β 3-methyl ether 2-G, oestradiol-17β 17-G, oestriol 16α-G, 17-epi-oestriol 16α-G, oestriol 17β-G, testosterone 17-G, dehydroepiandrosterone 3-G, and pregnanediol 3-G. These derivatives proved to be most suitable for direct mass spectrometry. All steroid glucuronide derivatives investigated exhibited characteristic mass spectra. The spectra showed molecular ion peaks with different intensities. Relatively high intensities were observed, when an aliphatic hydroxyl group was substituted by the glucuronyl residue. Lower intensities were found in steroid glucuronides, in which an aromatic hydroxyl group was conjugated. The main fragment of the trimethylsilylated glucuronic acid moiety of “aliphatic” glucuronides was solm e 465 and that of “aromatic” glucuronides solm e 464. Furthermore, “aromatic” glucuronides yielded a fragment of M-392 which represents the aglycon moiety and one additional TMS group from glucuronic acid. In contrast, “aliphatic” glucuronides showed a fragment of M-481, due to the loss of the glucuronic acid moiety together with the glycosidic oxygen. The fragment M-322 which results from the loss of a part of the glucuronic acid moiety was found only in “aliphatic” glucuronides.