Induction of CYP3A4 by Berberine CSV JSON

To investigate the induction effect of BER on HepG2 cells, 200 μL of 1 x10^6/mL HepG2 cell suspension was placed in a 6-well plate and incubated with 2 mL of culturemedium for 4 days (culture medium was replaced every 2 days). When the cell fusion was approximately 90%, the medium was discarded, and 2 mL of BER (1.0 μg̈ mĹ1 prepared in culture medium) was added into three wells, and 2 mL of drug-free culture medium was added into the other three wells. After incubation for three days, the medium was discarded, 2 mL of lovastatin (1.0 μg̈ mĹ1 prepared in culture medium) was added into each well prior to incubation for 90 min, then 100 μL of the sample were collected at 0, 30, 60 and 90 min, the sample was added with 50 μL ofice-cold sodium acetate buffer (100 mM; pH 4.5 at room temperature) prior to addition of 150 μL cool acetonitrile (containing 100 ng̈ mĹ1 warfarin as internal standard, 4 ̋C), then vortexed for 2 min and centrifuged at 13,500 x g for 15 min, the supernatant was placed into a 96-well polypropylene plate for analysis, where the sodium acetate buffer was used to protect the transformation of lovastatin and lovastatin acid [39]. The linearity for the analysis of lovastatin was determined with 100 μL of blank samples (200 μL of HepG2 cell suspension added with 2 mL culture medium) added with 50 μL of ice-cold sodium acetate buffer (100 mM; pH 4.5 at room temperature) and 150 μL cool acetonitrile (containing 100 ng̈ mĹ1 warfarin as internal standard, 4 ̋C) prepared at final concentrations levels of 10, 25, 50, 100, 250 and 500 ng̈ mĹ1, QC samples were prepared at final concentrations of 30, 75, 150, 300 ng̈ mĹ1 as the sample preparation. The UPLC-MS/MS analysis was performed as well as determination of lovastatin acid. MS/MS condition transitions (m/z) for lovastatin were 405.35Ñ285.22, the parameters of cone voltage and collision energy were 16 and 12 V, respectively.