TABLE 1.
Incubation conditions of the CYP probe substrates in human liver microsomes
| CYP substrate | CYP form | Substrate concn (μM) | Protein contenta (mg/ml) | Incubation
|
Analytical methodb | |
|---|---|---|---|---|---|---|
| Time (min) | Vol (ml) | |||||
| Caffeine | CYP1A2 | 10 | 1 | 30 | 0.5 | Radio-HPLC |
| Coumarin | CYP2A6 | 50 | 0.05-0.1 | 10 | 1 | Fluorimetry |
| Cyclosporine | CYP3A4 | 5 | 1 | 30 | 1 | Radio-HPLC |
| Testosterone | CYP3A4 | 100 | 0.5 | 25 | 0.5 | LC-MS |
| Midazolam | CYP3A4 and -5 | 50 | 0.5 | 10 | 1 | HPLC-UV |
| Phenytoin | CYP2C8 and -9 | 150 | 0.5 | 120 | 1 | Radio-HPLC |
| Tolbutamide | CYP2C8 and -9 | 1,200 | 0.5 | 90 | 1 | HPLC-UV |
| S-Mephenytoin | CYP2C19 | 100 | 0.5 | 30 | 0.5 | Radio-HPLC |
| Debrisoquine | CYP2D6 | 49.3 | 2 | 30 | 0.15 | Radio-HPLC |
| Chlorzoxazone | CYP2E1 | 500 | 0.5 | 20 | 1 | HPLC-UV |
a
Protein content was measured by the Lowry method, as modified by Miller (13), using bovine serum albumin as a standard.
b
Radio-HPLC, HPLC with online radioactivity detection; LC, liquid chromatography; MS, mass spectrometry.