Isolation and structure elucidation of a highly haemolytic saponin from the Merck saponin extract using high-field gradient-enhanced NMR techniques

Saponins SAPO50 and SAPO30, of which SAPO50 is highly haemolytic, have been isolated from the commercial Merck Saponin. Their structures have been determined exclusively by high-field gradient-enhanced NMR methods. The H-1 and C-13 NMR spectra of these saponins in pyridine-deuterium oxide have been assigned by homonuclear and heteronuclear correlation experiments. Anomeric configurations were obtained by combined use of (1)J(CH), (3)J(H-1,H-2), and 1D-NOESY data. Sugar residues were identified by use of (3)J(HH) values obtained from their subspectra recorded using an optimized 1D-z-TOCSY sequence. Linkage assignments were made using the ge-HMBC and 1D-NOESY spectra. This study shows that SAPO50 represents a hitherto undescribed saponin with the following structure: 3-O-beta-D-xylopyranosyl-(1 --> 3)-[beta-D-galactopyranosyl-(1 --> 2)]-beta-D-glucuronopyranosyl gypsogenin 28-O-(6-deoxy-beta-D-glucopyranosyl)-(1 --> 4)-[beta-D-xylopyranosyl-(1 --> 3)-beta-D-xylopyranosyl-(1 --> 4)]-alpha-L-rhamnopyranosyl-(1 --> 2)-beta-D-fucopyranoside. SAPO30, however, corresponds to a saponin previously described [D. Frechet, B. Christ, B. Monegier du Sorbier, H. Fischer, and M. Vuilhorgne, Phytochemistry, 30 (1991) 927-931]. (C) 1997 Elsevier Science Ltd.