title: LC/ESI-MS/MS analysis of urinary 3β-sulfooxy-7β-N-acetylglucosaminyl-5-cholen-24-oic acid and its amides: new biomarkers for the detection of Niemann-Pick type C disease.
abstract: We developed a sensitive, reliable, and accurate LC/ESI-MS/MS method for measurement of 3β-sulfooxy-7β-N-acetylglucosaminyl-5-cholen-24-oic acid and its glycine and taurine amides in urine. This atypical C24 bile acid has been reported previously to be present in the urine of patients with Niemann-Pick Type C (NPC) disease. In the method, targeted analytes are concentrated at the front edge of a trapping column, Shim-pack MAYI-C8, which permits elimination of contaminating molecules in the urinary matrix. The trapped analytes are then eluted, separated on a YMC-Pack Pro C18, and quantified with MS/MS using selected reaction monitoring. The method could detect (as amount injected) 2pg of nonamidated 3β-sulfooxy-7β-N-acetylglucosaminyl-5-cholen-24-oic acid, 2pg of its glycine-amide, and 0.6pg of its taurine-amide, and is linear up to 300pg. The method was then used to measure the three analytes in the urine of NPC patients (N=2), 3β-hydroxysteroid dehydrogenase deficiency patients (N=2), and healthy volunteers (N=8). Measurable concentrations of all three analytes were present in all subjects. The urinary concentration of the sum of all three analytes was four hundred times greater in the 3month NPC patient and 40times greater in the adult patient than that of healthy volunteers. The markedly elevated urinary concentration of 3β-sulfooxy-7β-N-acetylglucosaminyl-5-cholen-24-oic acid and its amides in NPC patients suggests that these compounds may be valuable biomarkers for detection of NPC disease.
authors.affiliation: Department of Pharmaceutical Sciences, Tohoku University Hospital, 1-1 Seiryo-machi, Aoba-ku, Sendai 980-8574, Japan.
authors.forename: Masamitsu
authors.initials: M
authors.lastname: Maekawa
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Misawa
authors.forename: Ayako
authors.initials: A
authors.lastname: Sotoura
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Yamaguchi
authors.forename: Masami
authors.initials: M
authors.lastname: Togawa
authors.forename: Kousaku
authors.initials: K
authors.lastname: Ohno
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Nittono
authors.forename: Genta
authors.initials: G
authors.lastname: Kakiyama
authors.forename: Takashi
authors.initials: T
authors.lastname: Iida
authors.forename: Alan F
authors.initials: AF
authors.lastname: Hofmann
authors.forename: Junichi
authors.initials: J
authors.lastname: Goto
authors.forename: Miki
authors.initials: M
authors.lastname: Shimada
authors.forename: Nariyasu
authors.initials: N
authors.lastname: Mano
mesh_terms: D000293:Adolescent; D000328:Adult; D015415:Biomarkers / Q000652:urine; D016022:Case-Control Studies; D005260:Female; D006801:Humans; D007223:Infant; D008297:Male; D052556:Niemann-Pick Disease, Type C / Q000175:diagnosis / Q000652:urine*; D012503:Saponins / Q000652:urine*; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry; D055815:Young Adult
chemical_list: C587858:3-sulfooxy-7-N-acetylglucosaminyl-5-cholen-24-oic acid; D015415:Biomarkers; D012503:Saponins
keywords: 3β-HSD; 3β-hydroxy-Δ(5)-C(27)-steroid dehydrogenase/isomerase; Biomarker; C.V.; CE; ESI; GlcNAc; IS; LC/ESI-MS/MS; LC/MS/MS; Multiple conjugate; N-acetylglucosamine; NPC; Niemann–Pick disease type C; Niemann–Pick type C disease; R.E.; S/N; SNAG-Δ(5)-CA; SNAG-Δ(5)-CG; SNAG-Δ(5)-CT; SRM; coefficient of variation; collision voltage; electrospray ionization; glycine-amidated 3β-sulfooxy-7β-N-acetylglucosaminyl-5-cholen-24-oic acid; internal standard; liquid chromatography/tandem mass spectrometry; nonamidated 3β-sulfooxy-7β-N-acetylglucosaminyl-5-cholen-24-oic acid; relative error; selected reaction monitoring; signal-to-noise ratio; taurine-amidated 3β-sulfooxy-7β-N-acetylglucosaminyl-5-cholen-24-oic acid