The bioactivity‐guided isolation and structural identification of toxic cucurbitacin steroidal glucosides from stemodia kingii

A histologically validated murine model for the ovine intoxication by Stemodia kingii was used as a bioassay to guide the isolation of several groups of toxins from Stemodia kingii . Two of the toxins from one group were purified sufficiently to allow structural analysis and a determination of their median lethal doses (LD 50 ) for oral administration to mice. A combination of acid hydrolysis, elemental analysis, HPLC‐MS, 1D‐NMR ( 1 H, 13 C) and 2D‐NMR ( 1 H– 1 H COSY, 13 C– 1 H HSQC and HMBC, and gNOESY) was used to define stemodiosides B3 and B4 as cucurbitacin steroidal glucosides. Thus stemodioside B3 is (24Z)‐3 α ‐( β ‐glucopyranosyloxy)‐2 β ,20,27‐trihydroxy‐19‐(10→9 β )‐ abeo ‐10 α ‐lanost‐5,24‐diene‐11‐one and stemodioside B4 is (23E)‐3 α ‐( β ‐glucopyranosyloxy)‐2 β ,20,22,27‐tetrahydroxy‐19‐(10→9 β )‐ abeo ‐10 α ‐lanost‐5,23‐diene‐11‐one. The approximate oral LD 50 s for stemodiosides B3 and B4 in mice were estimated to be 99 and 42 mg/kg body weight, respectively. Copyright © 2006 John Wiley & Sons, Ltd.