Mean Ki values (micromolar) for inhibition of diclofenac 4′-hydroxylase activity by baicalein, galangin, and 6-hydroxyflavone in the CYP2C9 RECO system, in human liver microsomes, and in S9 fraction from wild-type CYP2C9.1 and two variants transiently expressed in COS-7 cells
Rate of diclofenac 4′-hydroxylation by each enzyme system was 11.5, 18.8, 28.6, 25.9, and 26.7 nmol/min/nmol CYP2C9. Incubations performed using 0.3 pmol of RECO CYP2C9, S9 fraction containing 0.3 pmol of CYP2C9, or 0.2 μl of pooled human liver microsomes, 2.5 to 100 μM diclofenac, and a test compound with a concentration in the range 0 to approximately 6 times Ki. 4′-Hydroxydiclofenac/diclofenac <1/20 in all the assays. Global standard error for data fitting was less than 30% and γ2 > 0.90 for each effector.
RECO CYP2C9 | Human Liver Microsomes | CYP2C9 Expressed in COS-7 Cells | |||||
|---|---|---|---|---|---|---|---|
| CYP2C9.1 | CYP2C9 Phe100Asp | CYP2C9 Phe100Trp | |||||
| Flavonoid | |||||||
| Baicalein | 0.91 | 4.0 | 1.0 | ||||
| Galangin | 0.15 | 0.73 | 0.50 | ||||
| 6-Hydroxyflavone | 2.2a | 11a | 17a | 25 | 24 | ||
| Diclofenac 4′-hydroxylation kinetic parameters | |||||||
| Km (μM) | 14 | 4.0 | 2.3 | 2.5 | 2.6 | ||
| Vmax (nmol/min/nmol CYP2C9) | 13.1 | 19.6 | 29.3 | 26.5 | 27.4 | ||
↵ a Noncompetitive inhibition; in all other situations, competitive inhibition.