A simple method has been developed for partial purification of cytochrome P-450 from phenobarbital-induced rabbit liver microsomes. The cytochrome was solubilized with 0.1 M phosphate buffer (pH 7.25) containing 0.4% sodium cholate, 20% glycerol, 1 mM dithiothreitol, and 1 mM EDTA. The solubilized hemoprotein was adsorbed on a column of an ω-amino-n-octyl derivative of Sepharose 4B and then eluted with 1 mM Emalgen 913 (a polyoxyethylene nonylphenyl ether) in the same buffer. This method permitted us to purify cytochrome P-450 to a specific content of 8–10 nmoles per mg of protein (40–50% pure assuming a molecular weight of 50,000 for the cytochrome) with an overall yield of about 35%. The purified preparation was free from both cytochromes b5 and P-420. Its spectral properties were consistent with those reported for the microsomal bound form of cytochrome P-450 and superior in several respects to those of the partially purified preparations of the cytohcrome so far described.
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Imai et al. (1974) studied this question.