Microwave-assisted derivatization: application to steroid profiling by gas chromatography/mass spectrometry.
Gas chromatography-mass spectrometry (GC-MS) remains as the gold-standard technique for the study of the steroid metabolome. A main limitation is the need of performing a derivatization step since incubation with strong silylations agents for long periods of time (usually 16 h) is required for the derivatization of hindered hydroxyls present in some steroids of interest. In the present work, a rapid, simple and reproducible microwave-assisted derivatization method was developed. In the method, 36 steroids already treated with methoxyamine (2% in pyridine) were silylated with 50 μl of N-trimethylsilylimidazole by using microwave irradiation, and the formed methyloxime-trimethylsilyl derivatives were analyzed by GC-MS. Microwave power and derivatization time silylation conditions were optimized being the optimum conditions 600 W and 3 min respectively. In order to evaluate the usefulness of this technique, the urine steroid profiles for 20 healthy individuals were analyzed. The results of a comparison of microwave irradiation with the classical heating protocol showed similar derivatization yields, thus suggesting that microwave-assisted silylation is a valid tool for the rapid steroid metabolome study.
Biochemistry and Molecular Genetics, Hospital Clínic of Barcelona, University of Barcelona, Spain. Electronic address: casals@clinic.cat.
Gregori
G
Casals
Bioanalysis Research Group, IMIM (Institut Hospital del Mar d'Investigacions Mèdiques), Barcelona, Spain; Department of Experimental and Health Sciences, Universitat Pompeu Fabra, Barcelona, Spain.
Josep
J
Marcos
Bioanalysis Research Group, IMIM (Institut Hospital del Mar d'Investigacions Mèdiques), Barcelona, Spain.
Oscar J
OJ
Pozo
Biochemistry and Molecular Genetics, Hospital Clínic of Barcelona, University of Barcelona, Spain.
José
J
Alcaraz
Biochemistry and Molecular Genetics, Hospital Clínic of Barcelona, University of Barcelona, Spain.
María Jesús
MJ
Martínez de Osaba
Biochemistry and Molecular Genetics, Hospital Clínic of Barcelona, University of Barcelona, Spain.
Wladimiro
W
Jiménez
D000328:Adult; D000368:Aged; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D057230:Limit of Detection; D008297:Male; D055442:Metabolome; D008875:Middle Aged; D015203:Reproducibility of Results; D013256:Steroids / Q000737:chemistry* / Q000652:urine*
D013256:Steroids
Gas chromatography/mass spectrometry; Human urine; Metabolome; Microwave assisted derivatization; Steroids