A sensitive LC-MS/MS method to determine ginkgolide B in human plasma and urine: application in a pharmacokinetics and excretion study of healthy Chinese subjects.

1. Ginkgolide B (GB), the most active of the ginkgolides, has been developed as a new drug for the treatment of vascular insufficiency; however, the pharmacokinetics of GB remain unclear. Here, we investigated the pharmacokinetics and urine excretion properties of GB in healthy Chinese subjects administered single- and multiple-dose injectable GB based on a new LC-MS/MS method.2. GB pharmacokinetics were found to be dose-dependent from 20 to 60 mg. GB reached a steady state by day 6 with once-daily dosing at 40 mg. Systemic exposure to GB, as characterised by AUC0-∞, indicated accumulation following repeated once-daily dosing for seven consecutive days. The mean urinary cumulative excretion rate of GB in response to 20, 40, and 60 mg GB was 41.9 ± 18.5%, 32.9 ± 12.2%, and 43.9 ± 8.5%, respectively.3. Dose-proportional pharmacokinetics of GB were observed after intravenous administration in healthy subjects. A gradual reduction in the volume of distribution and slight change in mean resistance time led us to conjecture the limited accumulation of GB based on distribution equilibrium in vivo.4. This comprehensive study of the clinical pharmacokinetics of GB will provide useful information for its application and further development.

Human pharmacokinetics of ginkgo terpene lactones and impact of carboxylation in blood on their platelet-activating factor antagonistic activity.

Terpene lactones are a class of bioactive constituents of standardized preparations of Ginkgo biloba leaf extract, extensively used as add-on therapies in patients with ischemic cardiovascular and cerebrovascular diseases. This investigation evaluated human pharmacokinetics of ginkgo terpene lactones and impact of their carboxylation in blood. Human subjects received oral YinXing-TongZhi tablet or intravenous ShuXueNing, two standardized ginkgo preparations. Their plasma protein-binding and platelet-activating factor antagonistic activity were assessed in vitro. Their carboxylation was assessed in phosphate-buffered saline (pH 7.4) and in human plasma. After dosing YinXing-TongZhi tablet, ginkgolides A and B and bilobalide exhibited significantly higher systemic exposure levels than ginkgolides C and J; after dosing ShuXueNing, ginkgolides A, B, C, and J exhibited high exposure levels. The compounds' unbound fractions in plasma were 45-92%. Apparent oral bioavailability of ginkgolides A and B was mostly >100%, while that of ginkgolides C and J was 6-15%. Bilobalide's bioavailability was probably high but lower than that of ginkgolides A/B. Terminal half-lives of ginkgolides A, B, and C (4-7 h) after dosing ShuXueNing were shorter than their respective values (6-13 h) after dosing YinXing-TongZhi tablet. Half-life of bilobalide after dosing the tablet was around 5 h. Terpene lactones were roughly evenly distributed in various body fluids and tissues; glomerular-filtration-based renal excretion was the predominant elimination route for the ginkgolides and a major route for bilobalide. Terpene lactones circulated as trilactones and monocarboxylates. Carboxylation reduced platelet-activating factor antagonistic activity of ginkgolides A, B, and C. Ginkgolide J, bilobalide, and ginkgo flavonoids exhibited no such bioactivity. Collectively, differences in terpene lactones' exposure between the two preparations and influence of their carboxylation in blood should be considered in investigating the relative contributions of terpene lactones to ginkgo preparations' therapeutic effects. The results here will inform rational clinical use of ginkgo preparations.

The pharmacokinetics and conversion of the lactone to the carboxylate forms of ginkgolide B in rat plasma.

Ginkgolide B consists of three lactone groups, which may undergo hydrolysis, and lead to the rings opening in aqueous solution with different pHs. From mechanisms of pharmacological activity in vivo, the lactone appears to be the active form of the drug. Pharmacokinetics of lactone form (GB-lac) and the total of the lactone and carboxylate form (GB-tot) of ginkgolide B were investigated after intravenous administration of a dose of 4 mg/kg ginkgolide B. The rate of lactone hydrolysis was also studied in plasma in vitro. After intravenous administration, ginkgolide B in the original form was converted to its carboxylate form under simulated physiological conditions. The AUC0 - ∞ of GB-lac constituted 63.5 ± 17.4% of the AUC0 - ∞ of GB-tot. The ratio of average cumulation of excretion of lactone to carboxylate reached approximately 1 to 1 in urine. From the equilibrium of lactone hydrolysis in rat plasma in vitro, the k obs was - 0.0176 min(- 1) and t 1/2 was 39.38 min. In conclusion, the equilibrium existed between lactone of ginkgolide B and its carboxylate form in vivo at physiological pH, which suggested that more attention should be focused on the original and the ionization forms of ginkgolide B and the conversion of the lactone into carboxylate in vivo.

Identification of ginkgolide B metabolites in urine and rat liver cytochrome P450 enzymes responsible for their formation in vitro.

AIM: To identify metabolites of ginkgolide B in rat urine, the predominant metabolism of ginkgolide B and the major cytochrome (CYP) P450 enzymes responsible for the metabolism of ginkgolide B in rat liver microsomes. METHODS: A liquid chromatography quadrupole mass spectrometer and liquid chromatography ion-trap-time-of-flight mass spectrometer with electrospray ionization in negative-ion mode were used for the structure elucidation of metabolites in rat urine and liver microsome incubation. Various selective CYP450 inhibitors were applied to investigate their effects on the metabolism of ginkgolide B and the formation of the major metabolite in rat liver microsomes. RESULTS: Three metabolites were identified in rat urine. One hydroxyl metabolite of ginkgolide B were identified in rat liver microsomes, and quinidine uncompetitively inhibited the formation of the metabolite; its inhibitor constant (Ki) value for the inhibition of hydroxyl metabolite was estimated to be 8 micromol/L, while alpha-naphthoflavone, ketoconazole, sulfaphenazole, and diethyldithiocarbamate had no inhibitory effects. CONCLUSION: Ginkgolide B was metabolized to its hydroxyl metabolite in rats, and CYP2D6 was the major rat CYP isoform responsible for the ginkgolide B metabolism in rat liver microsomes.

Pharmacokinetics of the ginkgo B following intravenous administration of ginkgo B emulsion in rats.

Ginkgo B (GB) is an extract from the leaves of Ginkgo biloba, used in the treatment of dementia, cerebral insufficiency or related cognitive decline. In this paper, the main features of the pharmacokinetics of GB emulsion in rats were reviewed and the binding rate of GB to rat plasma and human plasma protein were investigated meanwhile. The concentrations of GB in plasma, tissue, and excretion of rats after i.v. administration of GB were measured using HPLC-ESI-MS. The metabolite was qualitated by LC-MS/MS. Intravenously administered GB was eliminated in a biphasic manner with a prominent initial phase (half-life of 0.3 h) followed by a slower terminal phase (half-life of 1.5 h). After i.v. 4, 12 and 36 mg/kg GB emulsion, the pharmacokinetic parameters from a two compartment model analysis of plasma samples were AUC(0-tau) (microg x min/ml): 53.7, 165.5 and 649.7; CL (l/min/kg): 0.07, 0.07 and 0.05; V(C) (l/kg): 2.27, 3.27 and 2.76, respectively. Peak concentrations generally occurred at 10 min except brain and fat. Tissue concentration then declined by several-fold during 6 h although still present in most tissues at 6 h. Single intravenous dose was mainly excreted in the urine (40-50%), feces contained less than 30%. The binding rate to rat plasma was little higher than to human plasma, but the difference was negligible. Some metabolites were found in urine and bile through qualitative analysis on the urine and bile by LC-MS/MS.