Complementary cardioprotective effects of flavonoid metabolites and terpenoid constituents of Ginkgo biloba extract (EGb 761) during ischemia and reperfusion

Hemodynamic and electron spin resonance (ESR) analyses were performed on is olated ischemic and reperfused rat hearts to assess the cardioprotective an d antioxidant effects of therapeutically relevant concentrations of Ginkgo biloba extract (EGb 761; 5, 50 or 200 mug/ml), its terpenoid constituents ( ginkgolide A; 0.05 mug/ml and ginkgolide B; 0.05, 0.25 or 0.50 mug/ml), and a terpene-free fraction of EGb 761 (CP 205; 5 or 50 mug/ml). Hearts underw ent 10 min of low-flow ischemia, 30 min of no-flow global ischemia, and 60 min of reperfusion. Test substances were added to the perfusion fluid durin g the last 10 min of control perfusion, low-Bow ischemia and the first 10 m in of reperfusion. A separate group of rats was treated with CP 205 (60 mg/ kg/day; p.o.) for 15 days, after which the hearts were perfused with plain buffer. In ESR experiments, the spin-trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) was added to the perfusate to determine the effects of treatments on post-ischemic myocardial free radical generation. Results showed that in v itro exposure of hearts to EGb 761 (5 or 50 mug/ml) or to ginkgolides A and B (both at 0.05 mug/ml), or in vivo pretreatment of the rats with CP 205 d elayed the onset of contracture during ischemia. The strong reperfusion-ind uced elevation of left ventricular end-diastolic pressure observed in untre ated hearts was significantly reduced by in vitro exposure to the lowest co ncentrations of EGb 761, by ginkgolide A, and to a lesser extent by ginkgol ide B, or by prior oral treatment with CP 205. Postischemic functional reco very was significantly improved by in vivo administration of CP 205, by per fusion with 5 mug/ml of EGb 761 or with both terpenoids as compared to untr eated group but in vitro CP 205 was not effective. ESR analyses revealed th at DMPO-OH (the DMPO/hydroxyl radical spin-adduct) concentrations in corona ry effluents were markedly decreased by all treatments, except for the lowe st concentration of ginkgolide B. Perfusing 5 mug/ml EGb 761 resulted in a better inhibition of baseline DMPO-OH concentration than 5 mug/ml CP 205 (- 70% and -48% vs, control, respectively), indicating that both terpenoid and flavonoid constituents of EGb 761 are required to produce this effect. CP 205 was significantly more efficient in reducing DMPO-OH concentration when administered in vivo than when applied in vitro, indicating that the antio xidant effect of flavonoid metabolites (formed in vivo) is superior to that of intact flavonol glycosides (present in vitro). Collectively these findi ngs provide the first evidence that part of the cardioprotection afforded b y EGb 761 is due to a specific action of its terpenoid constituents and tha t this effect involves a mechanism independent of direct free radical-scave nging. Thus, the terpenoid constituents of EGb 761 and the flavonoid metabo lites that are formed after in vivo administration of the extract act in a complementary manner to protect against myocardial ischemia-reperfusion inj ury.