title: Accurate LC-MS analyses for microcystins using per-15N-labeled microcystins.
abstract: Per-(15)N-labeled microcystins were prepared for use as surrogates for accurate liquid chromatography-mass spectrometry analysis. Two strains of Microcystis aeruginosa were cultured in (15)NO(3)-containing TS-15 medium. To change from the incorporation of (14)N to (15)N into all cell components, cells of Microcystis aeruginosa were precultured in Na(15)NO(3)-containing medium for more than 6 months. After mass cultivation of the strains, cells of each strain were harvested and lyophilized. Microcystin variants were extracted from the lyophilized cells and per-(15)N-labeled microcystin variants were purified using high-performance liquid chromatography and high-performance thin-layer chromatography. The structures of per-(15)N-labeled microcystin variants were confirmed by their mass spectrometry spectra and NMR spectra. When per-(15)N-labeled microcystins were used as surrogates for quantitative analysis of these toxins in cyanobacterial cells, excellent accuracy (98-106%) was obtained, with the m/z of M(+), [M+1](+), and [M+2](+) of both microcystins and the per-(15)N-labeled microcystins as surrogates being completely separated. In conclusion, per-(15)N-labeled microcystins are excellent surrogates for microcystin analysis using liquid chromatography-mass spectrometry.
authors.affiliation: Laboratory of Intellectual Fundamentals for Environmental Studies, National Institute for Environmental Studies, Tsukuba, Ibaraki, Japan. sanotomo@nies.go.jp
authors.forename: Tomoharu
authors.initials: T
authors.lastname: Sano
authors.forename: Hiroo
authors.initials: H
authors.lastname: Takagi
authors.forename: Kimiyo
authors.initials: K
authors.lastname: Nagano
authors.forename: Masataka
authors.initials: M
authors.lastname: Nishikawa
authors.forename: Kunimitsu
authors.initials: K
authors.lastname: Kaya
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002855:Chromatography, Thin Layer / Q000379:methods; D013058:Mass Spectrometry; D052998:Microcystins / Q000737:chemistry* / Q000302:isolation & purification*; D046931:Microcystis / Q000737:chemistry* / Q000378:metabolism; D009587:Nitrogen Isotopes / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular
chemical_list: D052998:Microcystins; D009587:Nitrogen Isotopes