1. Ginkgo biloba is one of the most popular herbal medicines worldwide due to its memory-enhancing and cognition-improving effects. The current study was designed to investigate the effects of five major constituents (bilobalide, ginkgolide A, B, quercetin, and kaempferol) in the standardized G. biloba extract EGb 761 on various cytochrome P450s (CYPs) in rats. 2. The activity of CYP450 was measured by the quantification of six metabolites from multiple cytochrome P450 probe substrates using a validated liquid chromatography coupled with tandem mass spectrometry detection (LC-MS/MS) method. The levels of messenger RNA (mRNA) and protein of various CYPs were determined by reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting analysis, respectively. 3. Bilobalide significantly induced the activity, protein, and mRNA expression of CYP3A1 and 1A2, and increased CYP2E1 activity and CYP2B1/2 protein expression in a dose-dependent manner. 4. Ginkgolide A, B, quercetin, and kaempferol did not affect CYP3A1, but induced CYP1A2 in a dose-dependent manner. EGb 761 and the five individual constituents had no effects on rat CYP2D2, 2C11 and 2C7. 5. The results indicate that bilobalide, and to a lesser extent ginkgolide A, B, quercetin, and kaempferol, play a key role in the effects of EGb 761 on CYP induction. Further study is needed to elucidate the mechanism of CYP3A induction by EGb 761 and bilobalide.