title: Structural evaluation of the glucuronides of morphine and formoterol using chemical derivatization with 1,2-dimethylimidazole-4-sulfonyl chloride and liquid chromatography/ion trap mass spectrometry.
abstract: For the first time chemical derivatization of isomeric drug glucuronides with 1,2-dimethylimidazole-4-sulfonyl chloride (DMISC) has been successfully applied as a tool for determining the site of conjugation. This provides a way to differentiate between glucuronide isomers containing aliphatic and phenolic hydroxyl groups. The analyses were performed with liquid chromatography/electrospray ion trap mass spectrometry (LC/ESI-MSn). DMISC has previously been shown to react selectively with phenols in estrogens, thus improving sensitivity in ESI-MS. The model compounds selected for this study were commercially available standards of formoterol, morphine, morphine-3-glucuronide (M3G), and morphine-6-glucuronide (M6G). Formoterol glucuronides were produced with an enzymatic method in house. Both formoterol and morphine possess one phenolic and one aliphatic hydroxyl group where glucuronidation could take place. The product ion mass spectra of the native morphine glucuronides were indistinguishable due to the initial neutral loss of monodehydrated glucuronic acid (176 u). However, a significant difference between the isomers was observed with DMISC derivatization, as only the form with a free phenol, M6G, gave a detectable reaction product. Formoterol formed two detectable glucuronide isomers in the enzymatic reaction. Their respective sites of conjugation could not be directly determined from the product ion spectra. Reaction with DMISC, however, gave a detectable product with only one of the isomers. Based on previous experience of the preferred DMISC reactions with phenols, and interpretation of the fragmentation pattern of the derivative, it was concluded that the reactive isomer had a free phenol, and was thus conjugated on the aliphatic chain.
authors.affiliation: Division of Analytical Pharmaceutical Chemistry, Department of Medicinal Chemistry, Uppsala University, Biomedical Centre, P.O. Box 574, SE-751 23 Uppsala, Sweden.
authors.forename: Matilda Lampinen
authors.initials: ML
authors.lastname: Salomonsson
authors.forename: Ulf
authors.initials: U
authors.lastname: Bondesson
authors.forename: Mikael
authors.initials: M
authors.lastname: Hedeland
mesh_terms: D000701:Analgesics, Opioid / Q000737:chemistry / Q000378:metabolism*; D001993:Bronchodilator Agents / Q000737:chemistry* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004983:Ethanolamines / Q000737:chemistry* / Q000378:metabolism; D000068759:Formoterol Fumarate; D020719:Glucuronides / Q000737:chemistry* / Q000378:metabolism; D015394:Molecular Structure; D009020:Morphine / Q000737:chemistry* / Q000378:metabolism; D009022:Morphine Derivatives; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
chemical_list: D000701:Analgesics, Opioid; D001993:Bronchodilator Agents; D004983:Ethanolamines; D020719:Glucuronides; D009022:Morphine Derivatives; D009020:Morphine; D000068759:Formoterol Fumarate