Biotransformation of 7-ethoxycoumarin in isolated perfused rainbow trout liver

A reliable technique for rainbow trout liver perfusion has been developed for studies on xenobiotic biotransformation. Normal function of the perfused liver was indicated throughout the perfusion experiment by 1) a proper oxygen consumption, 2) a low leakage of intracellular enzymes, 3) a stable pH value in the effluent, and 4) a stable release of metabolites into the effluent perfusate and bile for at least 2 hr after maximal rate of metabolism was attained. The main metabolite of 7-ethoxycoumarin in effluent perfusate was identified as 7-hydroxycoumarin glucuronide. Only trace amounts were identified as sulfates. When fish were pretreated with Clophen A50 or beta-naphthoflavone, the amount of metabolites released into the effluent perfusate increased 3.4- and 6.4-fold, respectively, when compared to control livers. Furthermore, in livers from Clophen A50- or beta-naphthoflavone-treated fish, only 80 and 67%, respectively, of excreted products were conjugated. Influence of temperature on 7-ethoxycoumarin metabolism was studied in perfused liver and isolated liver microsomes. Results indicate that the Q10 for the metabolism of 7-ethoxycoumarin in perfused liver deviates from that found in isolated microsomes. The amount of metabolites excreted into the bile consisted of about 25% of the amount found in effluent perfusate. The only metabolite detected in bile from perfused liver from control as well as treated fish was 7-hydroxycoumarin glucuronide.