Myricetin is a flavonoid compound that is abundant in teas, red wine, berries, herbs and vegetables with a variety of pharmacological properties such as antioxidant, anti-inflammatory and anti-cancer effects. Although there are in vitro studies showing that myricetin inhibits human cytochrome P450(CYP)2D6 and CYP3A, the inhibitory mechanisms of myricetin on CYP enzymes are still unclear. The aim of this study was to evaluate the inhibitory effects of myricetin on human and rat CYPs, including CYP3A2/3A4, CYP2B1/2B6, CYP2C9/2C11 and CYP2D1/2D6. This study was performed to investigate the inhibitory effects of myricetin on human CYP3A4, CYP2B6, CYP2C9, CYP2D6 and rat CYP3A2, CYP2B1, CYP2C11, CYP2D1 through the cocktail approach using ultra-performance liquid chromatography tandem mass spectrometry. Typical probe substrates were used as follows-midazolam for CYP3A2/3A4, dextromethorphan for CYP2D1/2D6, tolbutamide for CYP2C9/2C11, and bupropion for CYP2B1/2B6. The results of this study showed that myricetin might not be a time-dependent inhibitor. Moreover, myricetin inhibited CYP3A4 in an uncompetitive way with aninhibition constant (Ki)value of 143.1μM. It was also a noncompetitive inhibitor of CYP2C9 and CYP2D6 with Ki values of 31.12and 53.44μM and a competitive inhibitor of CYP2B1 with a Ki value of 69.70μM, as well as a mixed inhibitor of CYP3A2, CYP2C11 and CYP2D1with Ki values of 37.57, 14.88and 17.39μM, respectively. In conclusion, this study indicates that myricetin inhibited CYP3A4/3A2, CYP2C9/2C11, CYP2D6/2D1 and CYP2B1 by various mechanisms with different Kivalues. Given that our experiments are established in vitro, further in vivo work is needed to confirm the interaction between myricetin and CYP enzymes, thus providing better guidance for the safe clinical use of myricetin.