This paper is to report the study of the metabolism of forscolin in plasma and liver microsomes for guiding clinical therapy.  Forscolin was quantified by HPLC-MS/MS.  The metabolic stability of forscolin in rat, Beagle dog, monkey and human plasma and liver microsomes, mediated enzymes of forscolin and its inhibition on cytochrome P450 isoforms in human liver microsomes were studied.  Results showed that forscolin was not metabolized in plasma of the four species but metabolized in liver microsomes of the four species.  The t_1/2 of forscolin in rat, Beagle dog, monkey and human liver microsomes were (52.0 ± 15.0), (51.2 ± 5.9), (6.0 ± 0.2) and (11.9 ± 1.8) min; CL_int were (75.6 ± 18.7), (60.9 ± 6.8), (513.8 ± 14.3) and (176.2 ± 25.6) mL·min⁻¹·kg⁻¹; CL were (34.8 ± 4.5), (23.3 ± 1.0), (40.3 ± 0.5) and (17.9 ± 0.3) mL·min⁻¹·kg⁻¹, respectively.  Forscolin was metabolized by CYP3A4 in human liver microsomes.  There was definite inhibition on CYP3A4 at the concentrations of forscolin between 0.1 ng·mL⁻¹ and 5 μg·mL⁻¹.  Therefore, forscolin is rapidly excreted from liver microsomes.  Attention should be paid to the drug interaction when forscolin was used along with other drugs metabolized by CYP3A4 in clinics.