In vitro assessment of the inhibitory effect of goreisan extract and its ingredients on the P-glycoprotein drug transporter and cytochrome P-450 metabolic enzymes

Figures & data

Table 1. The metabolic reaction for each CYP isoform.

Representativeinhibitor

CYP isoform	Probe substrate	Final concentration of substrate (µmol/L)	Incubation time (min)	Metabolite	Representative inhibitor
1A2	Phenacetin	30	10	Acetaminophen	α-NaphthoflavoneFurafylline
2B6	Bupropion	100	10	Hydroxybupropion	Ticlopidine
2C8	Paclitaxel	10	20	6α-Hydroxypaclitaxel	MontelukastGemfibrozil 1-O-β-glucuronide
2C9	Diclofenac	5	5	4′-Hydroxydiclofenac	SulfaphenazoleTienilic acid
2C19	(S)-Mephenytoin	20	30	4′-Hydroxymephenytoin	Tranylcypromine S-Fluoxetine
2D6	Bufuralol	10	20	1′-Hydroxybufuralol	QuinidineParoxetine
3A	Midazolam	5	5	1′-Hydroxymidazolam	KetoconazoleVerapamil
3A	Testosterone	50	10	6β-Hydroxytestosterone	KetoconazoleVerapamil

Table 2. Effect of goreisan extract on apparent permeability (P_app) and efflux ratio of bi-directional [³H]-digoxin (0.030 μmol/L) transport across polarised Caco-2 cell monolayers.

		Papp (×10^-6cm/s)		Efflux ratio
		AB	BA
Control		4.62 ± 0.790	14.3 ± 1.03	3.09
Verapamil (50 μmol/L)		7.49 ± 0.193	8.04 ± 0.585	1.07
Goreisan extract (μg/mL)	250	4.77 ± 0.537	13.1 ± 0.350	2.74
	500	4.81 ± 1.48	14.1 ± 1.14	2.93
	1000	5.42 ± 1.12	12.3 ± 0.131*	2.28
	2000	4.41 ± 0.351	11.1 ± 0.505***	2.51
	4000	5.44 ± 0.552	8.19 ± 0.401***	1.51

The Caco-2 monolayer incubation time was ∼120 min at 37 °C.

P_app values are presented as mean ± SD (n = 3).

P_{app, AB} is the apparent permeability of digoxin from the apical-to-basolateral direction.

P_{app, BA} is that from the basolateral-to-apical direction.

*p < 0.05, ***p < 0.001 as compared to the control group (Dunnett’s multiple comparison test).

Figure 1. Inhibitory effect of goreisan extract on the efflux ratio across the Caco-2 cell monolayer for the P-gp substrate, [³H]-digoxin. The open circle represents the observations, a regression curve is shown, and a nonlinear regression model was used to determine the IC₅₀ values.

Table 3. IC₅₀ values of alisol A for CYP3A activity in human liver microsomes.

CYP substrate	IC50 (μmol/L)
	Direct inhibition method	Time-dependent inhibition method
3A (midazolam)	2.99	2.71
3A (testosterone)	8.39	7.28

Direct inhibition method: each CYP substrate was incubated with HLMs (0.1 mg protein/mL) in the presence of each goreisan ingredient for the designated time.

Time-dependent inhibition method: after HLMs (0.1 mg protein/mL) were pre-incubated for 30 min with each goreisan ingredient in the presence of NADPH, the CYP substrates were added to the pre-incubated microsomes for the designated time.