title: Hyphenation of solid-phase extraction with liquid chromatography and nuclear magnetic resonance: application of HPLC-DAD-SPE-NMR to identification of constituents of Kanahia laniflora.
abstract: The introduction of on-line solid-phase extraction (SPE) in HPLC-NMR has dramatically enhanced the sensitivity of this technique by concentration of the analytes in a small-volume NMR flow cell and by increasing the amount of the analyte by multiple peak trapping. In this study, the potential of HPLC-DAD-SPE-NMR hyphenation was demonstrated by structure determination of complex constituents of flower, leaf, root, and stem extracts of an African medicinal plant Kanahia laniflora. The technique was shown to allow acquisition of high-quality homo- and heteronuclear 2D NMR data following analytical-scale HPLC separation of extract constituents. Four flavonol glycosides [kaempferol 3-O-(6-O-alpha-l-rhamnopyranosyl)-beta-d-glucopyranoside; kaempferol 3-O-(2,6-di-O-alpha-l-rhamnopyranosyl)-beta-d-glucopyranoside; quercetin 3-O-(2,6-di-O-alpha-l-rhamnopyranosyl)-beta-d-glucopyranoside (rutin); and isorhamnetin, 3-O-(6-O-alpha-l-rhamnopyranosyl)-beta-d-glucopyranoside] and three 5alpha-cardenolides [coroglaucigenin 3-O-6-deoxy-beta-d-allopyranoside; coroglaucigenin 3-O-(4-O-beta-d-glucopyranosyl)-6-deoxy-beta-d-glucopyranoside; 3'-O-acetyl-3'-epiafroside] were identified, with complete assignments of 1H and 13C resonances based on HSQC and HMBC spectra whenever required. Confirmation of the structures was provided by HPLC-MS data. The HPLC-DAD-SPE-NMR technique therefore speeds up the dereplication of complex mixtures of natural origin significantly, by characterization of individual extract components prior to preparative isolation work.
authors.affiliation: Department of Medicinal Chemistry, Danish University of Pharmaceutical Sciences, Universitetsparken 2, DK-2100 Copenhagen, Denmark.
authors.forename: Cailean
authors.initials: C
authors.lastname: Clarkson
authors.forename: Dan
authors.initials: D
authors.lastname: Staerk
authors.forename: Steen Honoré
authors.initials: SH
authors.lastname: Hansen
authors.forename: Jerzy W
authors.initials: JW
authors.lastname: Jaroszewski
mesh_terms: D029062:Apocynaceae / Q000737:chemistry*; D002298:Cardenolides / Q000032:analysis; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006027:Glycosides / Q000032:analysis*; D007202:Indicators and Reagents; D009682:Magnetic Resonance Spectroscopy / Q000379:methods* / Q000592:standards; D015394:Molecular Structure; D010936:Plant Extracts / Q000032:analysis*; D010946:Plants, Medicinal / Q000737:chemistry*; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013997:Time Factors
chemical_list: D002298:Cardenolides; D006027:Glycosides; D007202:Indicators and Reagents; D010936:Plant Extracts