Metabolic Clearance in Liver S9 Fractions
Assay System:
  • Mammalian liver S9 fractions.
  • Human S9 fractions are pooled from 10 donors (mixed gender).  All other species are male.
  • Metabolic clearance determined by measuring percent remaining of test compound at multiple time points during incubation with liver S9 fractions.

Assay Conditions:

Test Compound Concentration           1 µM
S9 Protein Concentration           1.25 mg/mL
Cofactors           NADPH, UDPGA, and PAPS
Percent Organic Solvent in Incubation           0.5% acetonitrile
Incubation Time           0, 5, 10, 15, 30, and 45 minutes
pH           7.4
Temperature           37 °C
Positive Control           Testosterone and 7-hydroxycoumarin
Analysis           LC-MS (LC-MS/MS if necessary)
Number of Repeats           2

Intrinsic clearance, t1/2, and predicted blood extraction ratio (EH).

Test Compound Requirement: 1-2 mg dry powder or 100-200 µL of 10 mM DMSO stock solution.