Uric acid degradation studies

Uric acid is one of the principal products of purine metabolism in man 12 13). However, in many other organisms further oxidative degradation of the purine molecule occurs. One of the most important enzymes involved in uric acid oxidation is uricase, which has been studied to some extent in vitro. 

Hypoxanthine, on the other hand, which accounts for only a fifth or so of the urinary uric acid is an active intermediate. It is degraded to xanthine and then to uric add by xanthine oxidase. This enzyme is found mainly in liver, kidney, and bowel, while guanase is widely distributed and would quickly deaminate any guanine formed. The product xanthine is a poor substrate for hypoxanthine phosphoribosyltrans-ferase (HPRT). Most of the hypoxanthine formed is reutiliiced by conversion to inosinic acid. Similar conclusions were reached by Ayvazian and Skupp in 1965 when they administered C-labeled purines to patients (A2). Furthermore, these studies and those earlier studies show that the xanthine is converted to hypoxanthine, presumably at the nucleotide level, and on the basis of what we know about microorganisms, we would assume it to be via guanine nucleotides (M2). Since label was found in urinary 7-methylguanine as early as 4 hours after administration of C-labeled purines, and since methylation of RNA occurs at the macromolecular level (B13), interconversion must be rapid and incorporation of some of these products into nucleic acids must also occur quickly. 

Early studies with labeled compoimds were conducted simply to determine if adminbtered isotopes could be found in urinary uric acid or in the purines of tissue nucleic acids. More information was to be gained, however, by localizing the label in specific positions in the purine ring, and methods were therefore developed by which the uric acid molecule could be chemically degraded and each ring atom separately isolated. 

Early studies by Lieber and MacLachlan presented evidence suggesting that ethanol led to hyperlactic acidemia and decreased the excretion of uric acid l>2 Delbarre indicated the possibility of increased urate production and a study by Grunst, et al. suggested that ethanol infusion lead to increased release of uric acid from the liver. We have reexamined whether ethanol-induced hyperuricemia may be related to decreased renal excretion of uric acid, increased urate production by either accelerated degradation of nucleotides or increased novo synthesis or both mechanisms together. 

Further studies were aimed at the elucidation of the mechanisms whereby the hepatic adenine nucleotide pool is preserved in anoxic conditions, especially in the fasted state. In hepatocytes from fed rats, indeed, this protection is mainly due to a better better maintenance of the ATP concentration by anaerobic glycolysis. In hepatocytes from fasted animals, however, it is chiefly caused by a restriction of the degradation of AMP, as evidenced by the more marked accumulation of this nucleotide (Fig. 1) and the lower production of uric acid (Fig. 2) in comparison with the fed state. 

Based on our understanding of fructose metabolism, we have examined four potential mechanisms to account for fructose-induced hyperuricemia in man. 1) Shift in the uric acid pool, 2) decreased renal clearance of uric acid, 3) increased purine synthesis novo by stimulating PP-ribose-P production,and 4) accelerated degradation of purine ribonucleotides. Our studies were designed to distinguish which of these mechanisms in man could account for the hyperuricemia observed after fructose infusion. 

Our observations are most consistent with the hypothesis that the infusion of fructose leads to a rapid degradation of purine nucleotides with the consequent formation of inosine, hypoxanthine, xanthine, and finally uric acid (Fig. 6. Recent studies in the rat in vivo (Raivio, Kekomaki, and Maenpaa,... 

Studies with labeled lactic acid demonstrated that the carboxyl and a-carbons of lactate were converted into carbons 4 and 5 of uric acid, respectively. Since CO2 is the specific precursor of Ce, it was thought that lactic acid was converted into a 2-carbon compound prior to its utilization for uric acid formation. In view of the demonstration by Shemin of the conversion of serine to glycine, it was postulated that lactate was converted to serine which was then degraded to glycine, the latter being the more immediate purine precursor. This was confirmed by experimental evidence when high isotope concentrations were found in C4 after administration of carboxyl-labeled glycine. 

The purine-containing nucleotides are degraded stepwise to uric acid. Not all of the enzymes concerned with purine breakdown have been studied extensively. Those that have been investigated were found in many tissues of higher animals, in especially high concentrations in cell nuclei... 

Urine is also a very commonly studied biological matrix. It is much less complex than blood, and contains only a small amount of macromolecules. It has a high salt content and both organic and inorganic constituents. Its main components include NaCl (10 g/1), K (1.5 g/1), sulfate (0.8 g/1), phosphate (0.8 g/1), Ca and Mg (0.15 g/1), urea (20 g/1), creatinine (1 g/1), and uric acid (0.5 g/1). Urine should be protected from bacterial degradation, which is mostly accomplished by freezing the samples until analysis. 

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