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PubMed1993

A high-performance liquid chromatographic method for the determination of hypoxanthine, xanthine, uric acid and allantoin in serum.

A method was developed for the simultaneous determination of hypoxanthine, xanthine, uric acid and allantoin based on isocratic reversed-phase chromatography. This HPLC-method additionally allows the direct determination with UV-detection of inosine-5'-phosphate, uridine, thymine, orotic acid, allopurinol and oxipurinol, besides hypoxanthine, xanthine and uric acid in the same chromatographic run. Allantoin elutes in this system near the void volume and a fraction is collected covering the retention time range for this substance. After hydrolysis allantoin is converted to glyoxylate-2,4-dinitrophenylhydrazone, rechromatographed and detected at 360 nm. The coefficient of variation for this method does not exceed 5.0% for a serum concentration of 0.3 mumol/l hypoxanthine and is not greater than 5.3% for a xanthine concentration of 0.3 mumol/l serum. Recoveries were 90-110% for both hypoxanthine and xanthine. The determination of uric acid had an imprecision and inaccuracy not exceeding 1.45% in the concentration range of 103-568 mumol/l. Due to the more complex procedure required for the determination of allantoin, the coefficient of variation between days was 13.6% for a sample containing 0.8 mumol/l allantoin and the recoveries for this analyte were in the range of 86-93%. Reference ranges (mean +/- SD) determined on 171 serum samples from healthy adults were 12.7 +/- 6.6 mumol/l for hypoxanthine, 3.3 +/- 1.4 mumol/l for xanthine, and 15.7 +/- 7.9 mumol/l for allantoin. No significant age or sex dependence was observed. Uric acid concentrations were 320 +/- 55 mumol/l serum for men and 206 +/- 55 mumol/l for women.

Adolescent↗
PubMed1977

Nucleic acid biosynthesis in rat embryo cells infected with X14 or H-1 parvovirus.

Nucleic acid biosynthesis was studied in rat embryo cell (REC) cultures 48 hours after infection with X14 or H-1 parvovirus. The incorporation of 14C-formate and [6-(14C]-orotic acid into purines and pyrimidines of various was lowered after infection with these parvoviruses. 14C-Formate incorporation into acid-soluble thymine was greatly inhibited in H-1 virus-infected cells whereas it was slightly inhibited in X14 virus-infected cells. These results suggest that X14 virus-infected cells can carry out the biosynthesis of thymidylic acid utilizing some endogenous pyrimidine nucleotide (e.g. deoxycytidylic acid, via deoxyuridylic acid). In the infected cells, the nucleoplasmic RNA polymerase activity was strongly inhibited. This results suggests an interference by the two viruses with hosts RNA synthesis.

Adenine↗
PubMed2002

Regulation of pyrimidine synthesis in Pseudomonas mendocina.

Regulation of the de novo pyrimidine biosynthetic enzymes in the bacterium Pseudomonas mendocina was examined when its cells were grown on succinate as a carbon source. When P. mendocina was grown in the presence of orotic acid or uracil, the de novo enzyme activities were depressed with dihydroorotase activity being significantly depressed after uracil addition. Following pyrimidine limitation of a uracil auxotroph of P. mendocina deficient for orotate phosphoribosyltransferase activity, the pyrimidine biosynthetic pathway enzyme activities were affected indicating possible regulation at the level of enzyme synthesis. Of the de novo pathway enzymes assayed, dihydroorotate dehydrogenase exhibited the highest increase in its activity. The regulation of the pyrimidine biosynthetic pathway by pyrimidines in P. mendocina appeared similar to what was previously observed for the taxonomically-related species P. stutzeri.

Bacterial Proteins↗
PubMed1988

Growth, liver lipid and blood amino acids in rats fed ethanol with an adequate diet.

The weight, histology and RNA, DNA, protein and lipid content of the liver and arterial and portal plasma amino acid concentrations were determined in male Sprague-Dawley rats fed a liquid diet which met AIN-76A standards with 36% of the calories supplied by ethanol. The dietary components of the dry mixture in percentages by weight included 20% casein, 22% sucrose, 43% dextrin, 5% corn oil, vitamins, minerals and other dietary factors. For feeding these were suspended in distilled water containing 2.5% xanthan gum with or without ethanol to supply 1 kcal/ml. The feeding method employed perforated neoprene discs floated on top of the suspended diet to control evaporative losses. Animals were pair fed or ad libitum fed for 8-10 weeks. Gain/feed ratios were virtually identical for ethanol-fed rats and their pair-fed controls. Ethanol intake of ad libitum fed rats averaged 14.8, 10.3 and 7.4 g/kg BW/day after 1, 5 and 10 weeks, respectively. No chemical or histological evidence of liver fat accumulation or significant differences in arterial or portal amino acid concentrations were detected in animals fed ethanol. The lack of apparent ethanol toxicity is discussed in relation to the results of others and to our earlier report of increased orotic acid excretion by ethanol-fed rats.

Alcohol Drinking↗
PubMed1977

Synthesis of macromolecules by intestinal cells incubated with ammonia.

Mucosal cells isolated from the small intestine of chicks and rats were incubated with concentrations of ammonia normally found in the intestinal tract of mammals and birds. NH4Cl added to the incubation medium increased glucose metabolism in cells from both species. Ammonia stimulated incorporation of precursors into RNA and decarboxylation of orotic acid by cells isolated from chickens, but an increase in incorporation of precursors into DNA was not observed in cells from either species. Cultured embryonic chicken duodena showed increased incorporation of orotate into RNA with NH4Cl added to the medium. Rats immunized against jack bean urease showed lower urease activity per gram of dry intestinal content, lower intestinal weight, lower mucosal cell, and total gut protein and less protein per unit weight of DNA in the mucosal cell fraction. The results are compatible with the conclusion that ammonia PRODUCED IN THE INTESTINE BY BACTERIAL UREASES CAUSES SIGNIFICANT CHANGES IN THE CONTENT OF RNA and protein in intestine cells.

Ammonia↗
PubMed1980

Biosynthesis of uridine monophosphate in Plasmodium berghei.

High activities of the enzymes orotate phosphoribosyltransferase and orotidylate decarboxylase, that convert orotic acid to uridine monophosphate, have been demonstrated in crude supernatants obtained from lysed Plasmodium berghei. The enzymes are inhibited in vitro by 5-azaorotate, 5-azauracil and 6-azauracil. Of these, 5-azaorotate was the most effective and could serve as the prototype of a potential antimalarial.

Animals↗
PubMed1976

Increased synthesis of ribonucleic acids in mouse liver after treatment with a nonionic detergent.

A single injection of Tween 40 (polyoxyethylene(20)sorbitan monohexanoate) in the dose range of 600--800 mg/kg body weight induced an increased short-term labeling especially of poly(A)-containing mRNAs in mouse liver (using either [3H]orotic acid or [32P]orthophosphate as RNA precursors), and apparently increased the turnover rates of both rRNAs and mRNAs in this organ over a period of 24 h. In the early period (4 h) after the injection of Tween 40, there was also a significant increase in the content of 32P-labeled adenylic acid in microsomal poly(A)-mRNA fraction. The activity of DNA-dependent RNA polymerases in the nuclei of treated animals was stimulated up to 60% over that in control nuclei in the same period after detergent injection.

Animals↗
PubMedDate not supplied

[Synthesis of nucleic acids and proteins in the appendix of chickens infected with Eimeria tenella].

A comparative study was carried out of the synthesis of DNA, RNA and protein in blind appendages of the intestine of healthy and infected with E. tenella chickens. It was established that the inclusion of P32 into DNA and C14 - orotic acid into RNA increases in 2-10 days after the infection. An intensive inclusion of C14-lysine into protein of blind appendages was observed in 2-6 days after the infection of chickens. Under the effect of the parasite an activation of metabolism of DNA, RNA and protein takes place in blind appendages of the host's intestine.

Animals↗
PubMed1980

Urinary purines and pyrimidines in patients with hyperammonemia of various origins.

Excretion patterns of pyrimidines and purines in patients with various types of hyperammonemia have been investigated by 2-dimensional thin-layer chromatography and high pressure liquid chromatography (HPLC). For the quantitative analysis of pseudouridine, uracil and uridine a new procedure has been developed, consisting of pre-fractionation with Dowex 1 X 8, followed by dual column HPLC on a strong anion-exchanger and a reverse phase column. Thymine has also been analyzed in the pre-fractionated urine by a new HPLC method using the reverse phase column in combination with a strong cation-exchange column. Quantitative data for urinary pyrimidines and uric acid in hyperammonemia are given. In patients with a defect in one of the urea cycle enzymes, the level of pyrimidine excretion was found to depend on plasma ammonia concentrations. In other hyperammonemic patients, an increased excretion of orotic acid, uracil and uridine has only been found in one of the two patients with lysinuric protein intolerance, all other patients showing normal excretion patterns. Elevated uric acid excretions have been found frequently in our patients with hyperammonemia, but they did not always coincide with high plasma ammonia levels. A possible explanation for the difference in the excretion levels of the various pyrimidines is discussed.

Amino Acid Metabolism, Inborn Errors↗
PubMed1988

Dihydroorotic acid dehydrogenase activity in actinomycin-D-treated and normal chick embryos.

The effect of actinomycin D on chick embryos cultivated in vitro by New's culturing method was studied. Exposure of chick embryos to actinomycin D (0.05 micrograms/ml) at the primitive streak stage (stage 4; Hamburger and Hamilton) for 6 h showed interference in orotic acid formation. The assay of the enzyme dihydroorotic acid dehydrogenase was carried out in both treated and control embryos. No enzymic activity was observed in actinomycin-D-treated embryos in contrast to the considerable activity in the controls. These observations suggest an interference by actinomycin D in the biogenesis of the enzyme dihydroorotic acid dehydrogenase.

Animals↗
PubMed2004

mer-Diaquabis(1H-imidazole-kappaN3)(orotato-kappa2N3,O4nickel(II).

The title mononuclear complex, [Ni(C(5)H(2)N(2)O(4))(C(3)H(4)N(2))(2)(H(2)O)(2)] or [Ni(HOr)(im)(2)(H(2)O)(2)] (im is imidazole and H(3)Or is orotic acid, or 2,6-dioxo-1,2,3,6-tetrahydropyrimidine-4-carboxylic acid), has been synthesized and the crystal structure determination is reported. The Ni(II) ion in the complex has a distorted octahedral coordination geometry comprised of one deprotonated pyrimidine N atom and the adjacent carboxylate O atom of the orotate ligand, two tertiary imidazole N atoms and two aqua ligands. An extensive three-dimensional network of OW-H...O and N-H...O hydrogen bonds, and pi-pi and pi-ring interactions are responsible for crystal stabilization.

Journal Article↗
PubMed1990

[Basic study of nephrotoxicity of antibiotics. I. Studies of the effects of antibiotics on nucleic acids and protein metabolism in rat kidneys].

In order to evaluate the nephrotoxicity by antibiotics, effects of these agents were investigated in terms of protein synthesis and nucleic acid metabolism in the kidneys of Wistar rats. Aminoglycoside antibiotics (streptomycin, kanamycin, gentamycin), tetracycline (doxytetracycline), chloramphenicol, and cephems (cephalothin, cephaloridine, ceftezol, latamoxef) were used. These antibiotics were given to the rats for 5 successive days. On the 6th day 14C-6-orotic acid and 14C-1-leucine were administered intraperitoneally. Incorporation rates of these tracers into ribonucleic acid and protein fraction in rat kidney ribosomes (polysomes) were measured. Another experiment was undertaken in vitro by separating the polysome fraction from the rat kidneys. In vitro acellular protein synthesis using these polysomes was established. Effects of antibiotics on the incorporation rates of 14C-1-leucine were examined. Marked reduction of incorporation of these tracers into nucleic acid and protein in vivo and in vitro was induced by aminoglycoside antibiotics. These facts suggest that aminoglycoside antibiotics develop nephrotoxicity by interfering with the metabolism of ribosomal RNA and protein.

Aminoglycosides↗
PubMed1960

Studies on independent synthesis of cytoplasmic ribonucleic acids in Acetabularia mediterranea.

1. The RNA content of anucleate and nucleate fragments of Acetabularia has been measured. It was found that there is a net synthesis of RNA in nucleate fragments. On the other hand, the RNA content of anucleate fragments did not change significantly after enucleation. 2. Anucleate fragments, however, can readily incorporate (14)C-labeled adenine, orotic acid, and carbon dioxide into their cytoplasmic RNA. 3. The results of experiments on (14)CO(2) incorporation into the RNA of anucleate and nucleate fragments suggest that there is a mechanism for de novo synthesis of RNA in anucleate cytoplasm. 4. In Acetabularia, 81 per cent of the cytoplasmic RNA is bound to a large granule fraction, consisting mainly of chloroplasts. Even after removal of the nucleus, RNA is synthesized in this "chloroplast" fraction. The chloroplasts are thus a major site of RNA synthesis in the cytoplasm of these algae. Synthesis of "chloroplastic" RNA, in anucleate fragments, possibly occurs at the expense of the RNA present in other fractions (microsomes and supernatant). 5. 8-Azaguanine stimulates regeneration and cap formation in anucleate fragments and does not inhibit RNA synthesis in these fragments.

Acetabularia↗
PubMed1975

Arginine deficiency and orotic aciduria in mammals.

Dietary arginine deficiency in rats causes significant increases in urinary excretion of urea, citric acid and orotic acid independently of feed intake. Urea excretion during arginine deficiency depends upon the diet, sex, age, and species. Thus urea excretion has limitations as an indicator of arginine availability. Although elevated urinary citric acid during arginine deficiency is more consistently observed, it may be influenced by citric acid in natural dietary ingredients. Orotic acid excretion, however, appears to be a reliable indicator of available dietary arginine based upon studies in rats, mice, hamsters, guinea pigs, rabbits, and dogs.

Animals↗
PubMed1983

Biosynthesis of nuclear RNAs during 3'-methyl-4-dimethyl-aminoazobenzene hepatocarcinogenesis and in transplantable rat hepatomas.

The long-term administration of 3'-methyl-4-dimethylaminoazobenzene (3'-Me-DAB) to rats causes marked changes in concentrations and synthesis of nucleic acids in the liver. In the beginning, a decrease in total cellular RNA concentration can be seen. On the other hand, at the stage of tumor onset the concentration of nuclear RNAs in precancerous lesions and hepatomas is elevated. Incorporation of 3H-orotic acid into nuclear RNA after 45 min of in vivo labeling serving as a measure of biosynthesis of nuclear RNAs is markedly decreased when compared to control livers. This decrease takes place from the very beginning of the process and goes on until primary hepatomas arise in which its values are approximately at 10% of those observed in controls. In the present study the problem of the increase of proliferative activity at early stages of carcinogenesis is discussed which is not in correlation with the changes in nuclear RNA biosynthesis.

Animals↗
PubMedDate not supplied

[UMP synthase activity of rat tissue extracts in the bone marrow form of radiation sickness].

Whole-body gamma-irradiation of rats with a dose inducing bone marrow radiation syndrome caused phase organospecific changes in UMP-synthase activity. Disturbances of enzymic activity in the bone marrow and spleen well correlated with the dynamics of interphase and reproductive cell death. In brain extracts, UMP biosynthesis from orotic acid did not undergo essential changes.

Animals↗
PubMed1978

The influence of ammonia on purine and pyrimidine nucleotide biosynthesis in rat liver and brain in vitro.

1. The effect of ammonia on purine and pyrimidine nucleotide biosynthesis was studied in rat liver and brain in vitro. The incorporation of NaH(14)CO(3) into acid-soluble uridine nucleotide (UMP) in liver homogenates and minces was increased 2.5-4-fold on incubation with 10mm-NH(4)Cl plus N-acetyl-l-glutamate, but not with either compound alone. 2. The incorporation of NaH(14)CO(3) into orotic acid was increased 3-4-fold in liver homogenate with NH(4)Cl plus acetylglutamate. 3. The 5-phosphoribosyl 1-pyrophosphate content of liver homogenate was decreased by 50% after incubation for 10min with 10mm-NH(4)Cl plus acetylglutamate. 4. Concomitant with this decrease in free phosphoribosyl pyrophosphate was a 40-50% decrease in the rates of purine nucleotide synthesis, both de novo and from the preformed base. 5. Subcellular fractionation of liver indicated that the effects of NH(4)Cl plus acetylglutamate on pyrimidine and purine biosynthesis required a mitochondrial fraction. This effect of NH(4)Cl plus acetylglutamate could be duplicated in a mitochondria-free liver fraction with carbamoyl phosphate. 6. A similar series of experiments carried out with rat brain demonstrated a significant, though considerably smaller, effect on UMP synthesis de novo and purine base reutilization. 7. These data indicate that excessive amounts of ammonia may interfere with purine nucleotide biosynthesis by stimulating production of carbamoyl phosphate through the mitochondrial synthetase, with the excess carbamoyl phosphate in turn increasing pyrimidine nucleotide synthesis de novo and diminishing the phosphoribosyl pyrophosphate available for purine biosynthesis.

Ammonia↗
PubMed1994

Arginine becomes an essential amino acid after massive resection of rat small intestine.

We compared effects of feeding arginine- and/or proline- deficient diets (-Arg, -Pro, and -Arg, Pro) with those of a complete diet (Complete) in rats whose small intestine had been massively resected. After 4 weeks, the rats fed -Arg and -Arg, Pro lost weight (a mean of 28 and 32 g, respectively), whereas those fed Complete and -Pro gained 80 and 58 g, respectively. The average nitrogen balance was about 117,100, -20 and -14 mg/day for Complete, -Pro, -Arg, and -Arg, Pro diets, respectively. The concentration of arginine in skeletal muscle was about 310, 330, 91, and 65 nmol/g for Complete, -Pro, -Arg, and -Arg, Pro, respectively; while plasma arginine concentration averaged 95, 107, 56, and 46 microM, respectively. The weight loss, the negative nitrogen balance, and the markedly reduced arginine concentration in the muscle observed in rats fed -Arg and -Arg, Pro clearly indicate that arginine becomes a strictly essential amino acid in the rats with massive resection of the small intestine. However, sufficient proline can be synthesized from arginine in tissues such as the liver and kidney in the absence of the small intestine. Plasma glutamine, citrulline in the muscle and plasma, urinary excretion of orotic acid and nitrate (to assess nitric oxide formation from arginine) were also measured, and the changes in these metabolites are discussed.

Amino Acids↗