抄録
Biosynthesis of guanidinosuccinic acid (GSA) increases in renal failure, and the mechanisms of its synthesis has been thought to be closely related to the urea cycle. To clarify the pathway of GSA synthesis and the mechanisms of its increased synthesis in renal failure, the effect of urea cycle members and D, L-norvaline, an inhibitor of urea cycle enzymes on GSA synthesis was studied in isolated hepatocytes prepared from normal rats. GSA was separated by high pressure liquid chromatog-raphy, using a cation exchange resin column for amino acids analysis. It was measured fluorometrically after reacting with 9, 10-phenanthrequinone which selectively reacts with mono-substituted guanidine derivatives. We previously reported that isolated hepatocytes (7×106 cells) which were incubated in 6 ml of Krebs-Henseleit bicarbonate buffer (+3% bovine serum albumin and 10 mM sodium lactate) synthesized GSA at a rate of 40 nmol/108 cells/4 h in the presence of 36 mM urea, but a detectable amount of GSA was not synthesized in the presence of 10 mM NH4CI. The effect of other urea cycle members on GSA synthesis was also examined. In the presence of 1 mM carbamyl-phosphate, 10 mM citrulline, 10 mM argininosuccinate, 5 mM ornithine or 10 mM NH4CI + 5 mM ornithine, hepatocytes did not synthesize a detectable amount of GSA. In the presence of 10 mM NH4Cl and 36 mM urea, 0.5 mM of ornithine or arginine inhibited GSA synthesis by 63%. When the ammonium chloride was omitted, GSA synthesis increased with 36 mM urea, was strongly inhibited by 1 mM arginine, but was little inhibited by 1 mM ornithine. In the presence of NH4CI, 1mM and 5 mM citrulline inhibited GSA synthesis by 33% and 85%, respectively. D, L-Norvaline (5 mM), an inhibitor of urea cycle enzymes (ornithine transcarbamylase, arginin-osuccinate synthetase and arginase), strongly inhibited GSA synthesis stimulated by urea with or without 10 mM NH4Cl. These results suggest that GSA synthesis stimulated by urea is competitive with urea synthesis and that the synthesis of the former is partly catalyzed by the urea cycle enzymes.
