title: Synthesis and bioanalytical evaluation of morphine-3-O-sulfate and morphine-6-O-sulfate in human urine and plasma using LC-MS/MS.
abstract: The aim of this work was to synthesize morphine-3-O-sulfate and morphine-6-O-sulfate for use as reference substances, and to determine the sulfate conjugates as possible heroin and morphine metabolites in plasma and urine by a validated LC-MS/MS method. Morphine-6-O-sulfate and morphine-3-O-sulfate were prepared as dihydrates from morphine hydrochloride, in overall yields of 41 and 39% with product purities of >99.5% and >98%, respectively. For bioanalysis, the chromatographic system consisted of a reversed-phase column and gradient elution. The tandem mass spectrometer was operated in the positive electrospray mode using selected reaction monitoring, of transition m/z 366.15 to 286.40. The measuring range was 5-500 ng/mL for morphine-3-O-sulfate and 4.5-454 ng/mL for morphine-6-O-sulfate in plasma. In urine, the measuring range was 50-5000 ng/mL for morphine-3-O-sulfate and 45.4-4544 ng/mL for morphine-6-O-sulfate. The intra-assay and total imprecision (coefficient of variation) was below 11% for both analytes in urine and plasma. Quantifiable levels of morphine-3-O-sulfate in authentic urine and plasma samples were found. Only one authentic urine sample contained a detectable level of morphine-6-O-sulfate, while no detectable morphine-6-O-sulfate was found in plasma samples.
authors.affiliation: Department of Laboratory Medicine, Division of Clinical Pharmacology, Karolinska University Hospital, Stockholm, Sweden. maria.ch.andersson@karolinska.se
authors.forename: Maria
authors.initials: M
authors.lastname: Andersson
authors.forename: Tomasz
authors.initials: T
authors.lastname: Janosik
authors.forename: Hamid
authors.initials: H
authors.lastname: Shirani
authors.forename: Johnny
authors.initials: J
authors.lastname: Slätt
authors.forename: Andreas
authors.initials: A
authors.lastname: Fischer
authors.forename: Olof
authors.initials: O
authors.lastname: Beck
mesh_terms: D002851:Chromatography, High Pressure Liquid; D018360:Crystallography, X-Ray; D006801:Humans; D008958:Models, Molecular; D009020:Morphine / Q000032:analysis* / Q000097:blood / Q000138:chemical synthesis* / Q000652:urine; D009022:Morphine Derivatives / Q000032:analysis* / Q000097:blood / Q000138:chemical synthesis* / Q000652:urine; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry
chemical_list: D009022:Morphine Derivatives; C572395:morphine-3-O-sulfate; C572396:morphine-6-O-sulfate; D009020:Morphine