Liquid chromatography-mass spectrometry (LC-MS) analysis of
methanol extracts of Oenanthe crocata roots revealed that
oenanthotoxin co-eluted with another major polyalkyne, 2,3-dihydro-oenanthotoxin, using the existing high performance liquid chromatography (HPLC) method (isocratic elution from C18 with aqueous
methanol) for investigating Oenanthe
poisoning. Positive ES or APCI gave [(M+H)-H(2)O](+) and its
methanol adduct as major ion species for
oenanthotoxin, whereas 2,3-dihydro-oenanthotoxin formed [M+H](+) and its
methanol adduct. The two polyalkynes could be chromatographically resolved on C18 by gradient elution with aqueous
acetonitrile. This provides superior analysis for
oenanthotoxin using HPLC with photodiode array (PDA) detection alone, but for LC-MS/MS aqueous
acetonitrile was less suitable due to poor ionisation and, with APCI, an increase in the relative abundance of a [M-1](+) species, which could confuse compound assignment. HPLC-PDA and LC-MS/MS methods using an aqueous
acetonitrile or aqueous
methanol mobile phase, respectively, were successful when applied to the analysis of the stomach contents of a pony suspected to have eaten O. crocata. Relevant product ion spectra, by ion trap MS/MS, accurate mass data and complete sets of (1)H and (13)C NMR spectral assignments are given for the two compounds.