Sodium cholate extraction of rat liver nuclear xenobiotic-metabolizing enzymes.
DNA is the purported target of several carcinogenic and mutagenic agents. Nuclear enzymes which could generate or detoxify reactive metabolites are of major concern. Several such enzymes have been identified within nuclei, but obtaining samples with enriched content or activity is difficult, time-consuming, and uses harsh isolation techniques. Extraction of rat liver nuclear suspensions with cholate-containing buffer results in solubilization of 25-30% of the protein. Linear extraction was obtained for total protein and cytochromes P-450 and b5, NADPH-cytochrome P-450 reductase, NADH-cytochrome b5 reductase, DT-diaphorase, and microsomal-like epoxide hydrolase with specific activities comparable to values reported for isolated nuclear membrane, while the yield was five to ten times greater. Detergent extracts of rat liver nuclei were employed to study the comparative response of microsomal and nuclear enzymes to chemical treatment. While the responses to acute inductive (phenobarbital and 3-methylcholanthrene) and toxic (carbon tetrachloride and dibromochloropropane) treatments were qualitatively similar, an initiation-promotion protocol (diethylnitrosamine with phenobarbital promotion) resulted in divergent responses between the enzymes in the two subcellular fractions. Detergent extracts of nuclei offer an efficient means of recovering xenobiotic-metabolizing enzymes from rat liver nuclei, and have been utilized to demonstrate a differential response of nuclear enzymes during preneoplastic development.
D000818:Animals; D002251:Carbon Tetrachloride / Q000633:toxicity; D002467:Cell Nucleus / Q000201:enzymology*; D019826:Cholic Acid; D002793:Cholic Acids / Q000494:pharmacology*; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis; D003600:Cytosol / Q000201:enzymology; D008099:Liver / Q000187:drug effects / Q000201:enzymology*; D008297:Male; D008748:Methylcholanthrene / Q000494:pharmacology; D016660:NAD(P)H Dehydrogenase (Quinone); D009249:NADP / Q000494:pharmacology; D009251:NADPH-Ferrihemoprotein Reductase / Q000032:analysis; D004364:Pharmaceutical Preparations / Q000378:metabolism; D010634:Phenobarbital / Q000494:pharmacology; D011407:Propane / Q000031:analogs & derivatives / Q000633:toxicity; D011808:Quinone Reductases / Q000032:analysis; D051381:Rats; D011919:Rats, Inbred Strains
D002793:Cholic Acids; D004364:Pharmaceutical Preparations; D009249:NADP; D008748:Methylcholanthrene; D003577:Cytochrome P-450 Enzyme System; C007318:1,2-dibromo-3-chloropropane; D002251:Carbon Tetrachloride; D009251:NADPH-Ferrihemoprotein Reductase; D016660:NAD(P)H Dehydrogenase (Quinone); D011808:Quinone Reductases; D019826:Cholic Acid; D011407:Propane; D010634:Phenobarbital