Preparation and Characterisation of S9 Fractions

Many xenobiotics are only mutagenic after metabolism by the drug metabolising enzymes. Attention has focused on one of these enzymes ‘cytochrome P450’ (a generic term for a group of haem containing mixed function oxidase isoenzymes). Since the profile of metabolites produced by each isoenzyme is different and the activity of each isoenzyme is affected by the species, sex, age, health and environmental exposure of the animals used and by the tissue selected it is important to identify any factors which may affect their bioactivation when added to in vitro mutagenicity tests such as the Ames test. Metabolic activation of mutagens in vitro is generally achieved by supplementing the system by the addition of a mammalian microsome preparation, usually in the form of 9000 g supernatant (S9) from rodent liver. This fraction contains the Cyt. P450 enzymes as well as cytosolic enzymes.