Abstract
A more sensitive method for the determination of tissue-bound lithocholic acid (TBL) in rat livers is described in this report. The method has been applied to quantitate TBL in livers of rats treated with methylazoxymethanol (MAM). Charles River strain male rats (5–6 weeks old) received a single dose of MAM, 35 mg/kg b.wt., intravenously, while the corresponding control rats received only the saline vehicle After 11 months, the animals were sacrificed and their livers were excised rapidly. After extraction of the hepatic acetone-dry powder with ethanol-ammonia, the residue was predigested with various proteolytic enzymes before hydrolysis with cholanoyl amino acid hydrolase. The highest yield of lithocholic acid (LA) released from TBL (149μg LA/g dry tissue) was obtained by predigesting with 2 mg/ml trypsin for 2 h at 37°C pH 8.0. This technique not only gave higher yields of LA from TBL but also showed that livers of MAM-treated animals had significantly higher levels of TBL than the corresponding saline-treated controls.
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