RESUMEN
Enzymic formations of catechol- and guaiacol-estrogens by rat brains have been investigated using classical- and catechol-estrogens as substrates, respectively. The incubation mixtures were pretreated with liquid-liquid and/or solid-phase extraction, and the products were identified by comparison with authentic samples using liquid chromatography-mass spectrometry (-mass spectrometry) [LC-MS (-MS)]. The enzymic activities were also determined by measuring the formed products with LC-MS.
Asunto(s)
Encéfalo/metabolismo , Catecol O-Metiltransferasa/metabolismo , Cromatografía Liquida/métodos , Citocromo P-450 CYP1A1 , Sistema Enzimático del Citocromo P-450/metabolismo , Estrógenos/análisis , Espectrometría de Masas/métodos , Esteroide Hidroxilasas/metabolismo , Animales , Catecoles/química , Estrógenos/biosíntesis , Estrógenos/química , Guayacol/química , Ratas , Ratas WistarRESUMEN
The existence of catechol estrogens in rat brains was clarified using liquid chromatography-atmospheric pressure chemical ionization-ion trap-mass spectrometry-mass spectrometry (LC-APCI-MS2). The catechol estrogens were extracted in the presence of ascorbic acid and then derivatized to acetates with acetic anhydride and pyridine. After a successive purification with silica gel mini-column chromatography, reversed-phase solid-phase extraction and preparative HPLC, the obtained fractions containing the catechol estrogen acetates were subjected to LC-APCI-MS2. 2-Hydroxyestrone, 2-hydroxyestradiol and their 4-hydroxy isomers were identified as acetates by comparison with authentic samples based on their chromatographic behavior and mass spectral data. The derivatization to acetate was useful for the treatment of labile catechol estrogens.