title: Analysis of oligosaccharide sequences of trace Caulophyllum robustum saponins by direct infusion multiple-stage tandem mass spectrometry.
abstract: The saponins in Caulophyllum robustum have not yet been fully characterized. Furthermore these saponins are often present in trace amounts and are structurally complex. Here, a simple direct infusion electrospray ion trap multiple-stage tandem mass spectrometry (DI-ESI-IT-MS(n)) method was described for the characterization of trace C. robustum saponins. Eight reference saponins from the C. robustum hairy root were investigated by DI-ESI-IT-MS(n) in positive ion mode. Some fragmentation approaches were proposed through analysis of the [M+Na](+) ions: (1) preferential cleavage of the C-28 ester glycosidic bond to provide complementary [Y0α+Na](+) and [Bα+Na](+) ions for bidesmosidic saponins; (2) diagnostically neutral loss of CO2 from free carboxyl groups at C-28 for monodesmosidic saponins; and (3) the ion intensity ratio between [C2β+Na](+) and [B2β+Na](+), which is sensitive to the structural differences between the two isomeric β-sugar chains (Glc → (2)Ara and Glc → (3)Ara). The DI-ESI-IT-MS(n) method was successfully used for the analysis of trace C. robustum saponins with [M+Na](+) ions at m/z 1745.6, 1729.5, 1583.7, 1567.7, 1421.7 and 1405.7. This article highlights the discovery and identification of complex α- and β-oligosaccharide moieties in Caulophyllum saponins by glycosidic product ions along with cross ring cleavage product ions. Five oligosaccharide moieties were unambiguously or tentatively identified as Rha → (4)Glc → (6)Glc → (4)Rha → (4)Glc → (6)Glc, Glc → (4)Glc → (6)Glc → (4)Rha → (4)Glc → (6)Glc, Rha → Glc → Glc (Glc) → (2,3)Ara, Glc → Glc (Glc) → (2,3)Ara and Glc (Glc) → (2,3)Ara. Accuracy of the analytical procedure was demonstrated by structural identification of two saponins isolated using 1D and 2D-NMR spectroscopy. The DI-ESI-IT-MS(n) method facilitates rapid discovery and analysis of trace Caulophyllum saponins and is a powerful and practical tool for structural characterization of complex oligosaccharide moieties in triterpene saponins.
authors.affiliation: Key Laboratory of Chinese Materia Medica (Heilongjiang University of Chinese Medicine), Ministry of Education, Harbin 150040, PR China.
authors.forename: Yong-Gang
authors.initials: YG
authors.lastname: Xia
authors.affiliation: Key Laboratory of Chinese Materia Medica (Heilongjiang University of Chinese Medicine), Ministry of Education, Harbin 150040, PR China.
authors.forename: Jun
authors.initials: J
authors.lastname: Liang
authors.affiliation: Pharmaceutical College, Harbin Medical University, Harbin 150086, PR China.
authors.forename: Guo-Yu
authors.initials: GY
authors.lastname: Li
authors.affiliation: Key Laboratory of Chinese Materia Medica (Heilongjiang University of Chinese Medicine), Ministry of Education, Harbin 150040, PR China.
authors.forename: Bing-You
authors.initials: BY
authors.lastname: Yang
authors.affiliation: Key Laboratory of Chinese Materia Medica (Heilongjiang University of Chinese Medicine), Ministry of Education, Harbin 150040, PR China. Electronic address: hxkuang@hotmail.com.
authors.forename: Hai-Xue
authors.initials: HX
authors.lastname: Kuang
mesh_terms: D028145:Caulophyllum / Q000737:chemistry*; D007536:Isomerism; D009844:Oligosaccharides / Q000737:chemistry*; D018517:Plant Roots / Q000737:chemistry; D012503:Saponins / Q000737:chemistry*; D053719:Tandem Mass Spectrometry / Q000379:methods; D014315:Triterpenes / Q000737:chemistry
chemical_list: D009844:Oligosaccharides; D012503:Saponins; D014315:Triterpenes
keywords: Berberidaceae; Caulophyllum robustum; ESI-IT-MS(n); Triterpene saponins