Two-step purification of cytochrome P-450 from rat liver microsomes using high-performance liquid chromatography

Cytochrome P-450 from rat liver microsomes treated with phenobarbital (PB) was separated into six fractions, as was cytochrome P-450 treated with 3-methylcholanthrene (MC), by high-performance liquid chromatography (HPLC) with an anion-exchange column. PB and MC induced three forms and one form of cytochrome P-450, respectively. The major forms induced by PB and by MC were further purified to apparent homogeneity based on sodium dodecyl(lauryl)sulphate--polyacrylamide gel electrophoresis by HPLC using a hydroxyapatite column. These new HPLC techniques are simple, rapid and useful for the purification of major forms of cytochrome P-450 from solubilized microsomes.