US EPA

2204909 

Journal Article 

Measurement of cytochrome P-450 dependent dealkylation of alkoxyphenoxazones in hepatic S9s and hepatocyte homogenates: Effects of dicumarol 

Lubet, RA; Nims, RW; Mayer, RT; Cameron, JW; Schechtman, LM 

1985 

1 

Mutation Research
ISSN: 0027-5107
EISSN: 1873-135X 

142 

3 

127-131 

English 

2579331 

The effects of dicumarol (66762) on cytochrome-P-450 dependent dealkylation of alkoxyphenoxazones were investigated in rat and hamster hepatocytes. Male Fischer-rats and Syrian-golden-hamsters were treated with one intraperitoneal (ip) injection of 500 milligrams per kilogram (mg/kg) Aroclor-1254 (11097691). Additional groups of three rats were treated with either three daily ip injections of 25mg/kg 3-methylcholanthrene (56495) (MC), four daily ip injections of 60mg/kg phenobarbital, or three daily ip injections of 150mg/kg isosafrole (120581). Animals were killed on day 4 of the MC and isosafrole regimens or on day 5 of the PB regimen and 3 or 5 days after Aroclor-1254. Hepatic tissue from each treatment group was pooled for ethoxyresorufin-O-deethylase and pentoxyresorufin-O-dealkylase assays. Microsomal enzyme activities were highly induced by pretreatment with MC or Aroclor-1254. Hepatic tissue from PB pretreated rats yielded similar results. Values obtained for enzyme activities in Aroclor-1254 induced hepatocyte homogenates were more than 2 times greater when dicumarol was included in the reaction mixtures. When enzyme activities were measured in hepatocyte homogenates, they were more than twice those measured in supernatant fraction S9 prepared from homogenates. Reconstitution of homogenates by combining S9 supernatant and pellet yielded activity similar to that obtained with the initial homogenate. The authors conclude that the addition of dicumarol to a reaction mixture containing isolated rat and hamster hepatocytes enables the direct assay of O-dealkylation activities in hepatic S9 and cell homogenates.