Analysis of Ginkgo biloba for the presence of ginkgotoxin and ginkgotoxin 5 '-glucoside

Hot water extracts of Ginkgo biloba seeds were analyzed for the presence of ginkgotoxin (4'-O-methylpyridoxine) by reversed-phase liquid chromatograph y (LC) using methanol-0.05M KH2PO4(1 + 9, v/v) adjusted to pH 3 as mobile p hase. Detection was by fluorescence (excitation 280 nm, emission 370 nm). A straight line calibration curve was obtained for the 10-100 ng injected. A fter addition of beta -glucosidase (37 degreesC/h), an earlier eluting peak disappeared and the ginkgotoxin peak increased. The identity of the ginkgo toxin was confirmed by LC/MS and LC/MS/MS. LC/MS/MS also confirmed the 5'-g lucoside by comparison with the 3-glucoside. This is the first identificati on of a glucoside of ginkgotoxin in Ginkgo biloba. An unknown compound of M W 267 also observed in the Ginkgo biloba seed extract was shown not to be 3 ,5'-diacetylginkgotoxin by its different LC retention time. Extraction of g round Ginkgo biloba seeds with boiling water in a Soxhlet for 2 x 2 h yield ed a total of 179 mug/g of free ginkgotoxin. The concentration in powder fr om Ginkgo biloba capsules was several times lower than this (17-64 mug/g) i n 3 samples but higher in another (457 mug/g). Canned ginkgo seeds (white n uts) contained no detectable free ginkgotoxin but the glucoside was present . Different extraction times were studied: 0.5 h gave only 52 mug/g free gi nkgotoxin in the ginkgo seeds. However, boiling an extract for 4 h showed a bout 15% loss of ginkgotoxin and its glucoside.