In vitro inhibitory effects of bergenin on human liver cytochrome P450 enzymes

Figures & data

Table 1. Isoforms tested, marker reactions, incubation conditions and K_m used in the inhibition study.

CYPs	Marker reactions	Substrate concentration (μM)	Protein concentration (mg/mL)	Incubation time (min)	Estimated Km (μM)
1A2	Phenacetin O-deethylation	40	0.2	30	48
3A4	Testosterone 6β-hydroxylation	50	0.5	10	53
2A6	Coumarin 7-hydroxylation	1.0	0.1	10	1.5
2E1	Chlorzoxazone 6-hydroxylation	120	0.4	30	126
2D6	Dextromethorphan O-demethylation	25	0.25	20	4.8
2C9	Diclofenac 4'-hydroxylation	10	0.3	10	13
2C19	S-Mephenytoin 4-hydroxylation	100	0.2	40	105
2C8	Paclitaxel 6α-hydroxylation	10	0.5	30	16

Figure 1. The chemical structure of bergenin.

Figure 2. Effects of bergenin (100 μM) on the activity of CYP450 enzymes in pooled HLMs. All data represent mean ± S.D. of the triplicate incubations. *p < 0.05, significantly different from the negative control. Negative control: incubation systems without bergenin; bergenin: incubation systems with bergenin; positive control: incubation systems with their corresponding positive inhibitors.

Figure 3. Lineweaver–Burk plots (A) and the secondary plot for K_i (B) of effects of bergenin on CYP3A4 catalyzed reactions (testosterone 6β-hydroxylation) in pooled HLM. Data were obtained from 30 min incubation with testosterone (20–100 μM) in the absence or presence of bergenin (0–30 μM). All data represent mean ± S.D. of the triplicate incubations.

Figure 4. Lineweaver–Burk plots (A) and the secondary plot for K_i (B) of effects of bergenin on CYP2E1 catalyzed reactions (chlorzoxazone 6-hydroxylation) in pooled HLM. Data were obtained from 30 min incubation with diclofenac (25–250 μM) in the absence or presence of bergenin (0–50 μM). All data represent mean ± S.D. of the triplicate incubations.

Figure 5. Lineweaver–Burk plots (A) and the secondary plot for K_i (B) of effects of bergenin on CYP2C9 catalyzed reactions (diclofenac 4′-hydroxylation) in pooled HLM. Data were obtained from 30 min incubation with phenacetin (2–20 μM) in the absence or presence of bergenin (0–30 μM). All data represent mean ± S.D. of the triplicate incubations.

Figure 6. Time-dependent inhibition investigations of CYP3A4, 2E1 and 2C9 catalyzed reactions by bergenin (20 μM). All data represent mean ± S.D. of the triplicate incubations.

Figure 7. Time and concentration-inactivation of microsomal CYP3A4 activity by bergenin in the presence of NADPH. The initial rate constant of inactivation of CYP3A4 by each concentration (K_obs) was determined through linear regression analysis of the natural logarithm of the percentage of remaining activity versus pre-incubation time (A). The K_I and K_inact values were determined through non-linear analysis of the K_obs versus the bergenin concentration (B).