Table 1.
Conditions and HPLC parameters for the measurement of enzyme activity assays.
| CYP | Substrate | Metabolite | Substrate concentration(µM) | pmol of CYP/ incubation volume (µl) | Quenching agent | Elution | Injection of sample (µl) | Detection | |
|---|---|---|---|---|---|---|---|---|---|
| UV (nm) | Fluorescence Ex/Em(nm) | ||||||||
| 1A1/2 | 7-Ethoxyresorufin | Resorufin | 2.6 | 35/100 | 100% methanol | Isocratic | 50 | 535/585 | |
| 2 A | Coumarin | 7-Hydroxycoumarin | 6.25 | 35/100 | 100% methanol | Isocratic | 5 | 338/458 | |
| 2 C | Diazepam | Desmethyldiazepam | 100 | 70/200 | 100% acetonitrile | Isocratic | 50 | 229 | |
| 2 C | Diclofenac | 4´-Hydroxydiclofenac | 16 | 35/200 | Acetonitrile/Acetic acid (94:6) | Binary gradient | 50 | 280 | |
| 2D | Bufuralol | 1´-Hydroxybufuralol | 25 | 67.3/200 | 70% HClO4 | Binary gradient | 5 | 252/302 | |
| 3 A | Midazolam | 1´-Hydroxymidazolam | 2.8 | 12.56/100 | 100% methanol | Isocratic | 50 | 240 | |