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Basis of pyruvate inhibition in Thiobacillus thiooxidans.

Addition of 10(-3)m pyruvic acid to cultures of Thiobacillus thiooxidans, at pH 2.3, results in its rapid intracellular accumulation and in the cessation of sulfur oxidation, CO(2) fixation, and oxygen consumption; at pH 7.0, pyruvate neither inhibits oxygen uptake nor accumulates appreciably intracellularly. Pyruvate does not affect CO(2) fixation in cell-free extracts. The data suggest that the cells of T. thiooxidans are passively permeable to pyruvic acid at low pH. Thus entry of pyruvic acid causes accumulation of pyruvate with a concomitant decrease in intracellular pH.

Carbon Dioxide↗

Observations on calcium oxalate stone formers.

Biochemical studies were performed on 22 adult patients with idiopathic recurrent calcium oxalate renal stone disease and 23 healthy controls. It was found that urinary glutamic-oxaloacetic acid transaminase (UGOT) and urinary glutamic-pyruvic acid transaminase (UGPT) activity was low and lactate dehydrogenase (LDH) and gamma-glutamyl transferase (gamma-GT) activity was high in the urine of calcium oxalate stone formers. No significant changes were observed in the activity of glutamic-oxaloacetic acid transaminase, glutamic-pyruvic acid transaminase and gamma-GT in their blood but a significant reduction was found in both LDH and alkaline phosphatase activity. It was concluded that the activity of UGOT and UGPT is reduced in patients with kidney stones.

Adult↗

The effects of glucose on plasma amino acids and pyruvate during upper abdominal surgery.

To assess the potential benefits of perioperative glucose administration, the plasma concentrations of insulin, free fatty acids, ketone bodies, amino acids, lactate, and pyruvate were measured in 20 patients undergoing partial gastrectomy. Ten patients received intravenous glucose at 10 g/h during the operation, and the other 10 patients, as control, received no glucose. Plasma glucose, insulin activity, pyruvate and alanine concentrations in the glucose-receiving group increased significantly during the operation in comparison with the control group. In contrast, in the glucose-receiving group the plasma ketone bodies and branched-chain amino acids, especially leucine and isoleucine, decreased significantly during surgery. These results suggest that administration of glucose stimulated insulin secretion and resulted in accumulation of the substrates such as pyruvate and alanine which were utilized readily in the body. Glucose administration is useful in suppressing catabolism during upper abdominal surgery.

Amino Acids↗

Effect of exercise and beer on the plasma concentration and urinary excretion of purine bases.

OBJECTIVE: To investigate the effects of exercise and beer ingestion separately and combined on the plasma concentration of purine bases. METHODS: Six healthy men aged 30-39 years participated in 3 different experiments, in which they exercised for 30 min (at 70% of maximum oxygen uptake) and ingested beer (10 ml/kg body weight), or did each activity separately, with each experiment performed at 2 week intervals. RESULTS: The plasma concentration of uric acid was increased by 12% (p < 0.05), 8% (p < 0.01), and 29% (p < 0.01) with exercise, beer ingestion, and a combination of exercise and beer ingestion, respectively, which showed that it increased synergistically in the combination experiment. The fractional excretion of uric acid was decreased by 44% (p < 0.01) and 52% (p < 0.01) with exercise alone and a combination of exercise and beer ingestion, respectively, while it was increased by 15% (p < 0.05) with beer ingestion alone. Creatinine clearance was decreased by 16% (p < 0.01) with both exercise alone and a combination of exercise and beer ingestion, while it was not changed with beer ingestion alone. The increase in the plasma concentration of xanthine during the beer ingestion experiment was 2.1-fold greater than that during the combination (p < 0.05), while the increase in urinary excretion of xanthine caused by beer ingestion was 2.5-fold greater than that caused by a combination of beer and exercise (p < 0.05). Finally, exercise alone as well as a combination of beer and exercise increased the blood concentrations of lactic acid and NH3, whereas beer alone decreased concentration of pyruvic acid. CONCLUSION: These results suggest that the production of uric acid caused by both exercise and beer ingestion, as well as the inhibition of urinary uric acid excretion from a high blood lactic acid concentration, were the main contributors to the synergistic effect on the increase in plasma uric acid concentration. A decrease in creatinine clearance also contributed to the effect. We considered that pyruvic acid and NH3, produced in the muscles following exercise, relieved the beer induced increase of the plasma concentration and urinary excretion of xanthine, which may have played a minor role in the increase in plasma uric acid concentration.

Adult↗

Glucose metabolism in insulin administered euthyroid, hypothyroid and hyperthyroid rats.

Glucose metabolism was studied as evidenced by the sugar and pyruvic acid levels in blood and glycogen and pyruvic acid content of tissues in euthyroid, hypothyroid and hyperthyroid rats by giving insulin. Results show that in a normal thyroxine-excess insulin state, the rise in blood sugar was less, glycogenesis was much enhanced and glycolysis was reduced in comparison to these data in the euthyroid state. When tyroxine deficiency was associated with excess insulin, glycogenesis was enhanced further and an almost complete inhibition of glycolysis was observed. In excess thyroxine-excess insulin state glycogenesis was increased at the expense of glycolysis in comparison to the finding in the hyperthyroid state. Thus exogenous insulin in the euthyroid state altered the pattern of carbohydrate metabolism enhancing glycogenesis and inhibiting glycolysis. In a low thyroxine-excess insulin state, further enhancement of glycogenesis and inhibition of glycolysis were observed. But in an excess thyroxine-excess insulin state, the higher thyroxine activity was somewhat neutralized by higher insulin action allowing glycogenesis with glucose to proceed to some extent.

Animals↗

Biosensor system for continuous flow determination of enzyme activities. II. Simultaneous determination of plural enzyme activities.

A biosensor system for continuous flow determination of plural enzyme activities was prepared from the combination of two pyruvate sensors, a prereactor and a flow cell. This system was applied to the simultaneous determination of lactic dehydrogenase (LDH) and glutamic-pyruvic transaminase (GPT) activities in the same sample. These enzyme activities can be determined by measuring pyruvate produced by the enzyme reactions as follows. The amount of pyruvic acid can also be determined from the amount of oxygen consumed upon oxidation of pyruvic acid by pyruvate oxidase. (Formula: see text). Therefore, both of the detectors for the determination of lactic dehydrogenase and glutamic-pyruvic transaminase activities were prepared from the combination of a pyruvate oxidase membrane and an oxygen electrode. Pyruvate oxidase was covalently immobilized on a membrane prepared from cellulose triacetate. A linear relation was obtained between the output current and LDH or GPT activities in the range of 50 to 3,600 IU l-1 or 6 to 1,000 IU l-1, respectively. Each assay of these enzyme activities was completed within 15 min. The results obtained had a precision of ca. 4%. The sensor was stable for more than 25 days at 5 degrees C.

Alanine Transaminase↗

Derivatization of endogenous and exogenous compounds in plasma for high-performance liquid chromatographic analysis.

Methods are described for the analysis of endogenous (acetoacetic and pyruvic acids) and exogenous (propranolol) compounds in plasma. Acetoacetic and pyruvic acids were converted into 2,4-dinitrophenylhydrazones and identified by chromatography on a normal-phase silica gel column at 360 nm. The propranolol amino group was modified with dansyl chloride and the dansyl derivative, after extraction with a hydrophobic solvent, was analysed on a normal-phase silica gel column using fluorescence detection. The method allows the determination of 0.5 ng/ml of propranolol.

Acetoacetates↗

Structure of the extracellular polysaccharide secreted by Rhizobium leguminosarum var. phaseoli CIAT 899.

The structure of the extracellular polysaccharide secreted by Rhizobium leguminosarum var. phaseoli CIAT 899 has been studied by methylation analysis. 1H-n.m.r. spectroscopy, and partial acid hydrolysis. The repeating unit is an octasaccharide made up of D-glucose, D-galactose, pyruvic acid, and acetic acid in the molar ratios 6:2:1.5:1.5. Half of the terminal Gal groups are 4,6-substituted by pyruvic acid acetal groups and the other half by O-acetyl groups at position 3. Also, one of the 3-linked glucosyl residues carries a pyruvic acid 4,6-acetal group and one of the 4-linked glucosyl residues is acetylated at position 6.

Carbohydrate Sequence↗

Iron-regulated excretion of alpha-keto acids by Salmonella typhimurium.

Excretion of alpha-keto acids by clinical isolates and laboratory strains of Salmonella typhimurium was determined by high-performance liquid chromatography analysis of culture supernatants. The levels of excretion increased markedly with increasing iron stress imposed by the presence of alpha,alpha'-dipyridyl or conalbumin in the medium. The major product was pyruvic acid, but significant concentrations of alpha-ketoglutaric acid, alpha-ketoisovaleric acid, and alpha-ketoisocaproic acid were also observed. Maximal excretion occurred at iron stress levels that initially inhibited bacterial growth; the concentration of alpha,alpha'-dipyridyl at which this was observed differed between strains depending on their ability to secrete and utilize siderophores, suggesting that the intracellular iron status was important in determining alpha-keto acid excretion. However, prolonged incubation of the siderophore-deficient S. typhimurium strain enb-7 under conditions of high iron stress resulted in significant delayed bacterial growth, promoted by tonB-dependent uptake of iron complexed with the high accumulated levels of pyruvic acid and other alpha-keto acids. Strain RB181, a fur derivative of enb-7, excreted massive amounts of alpha-keto acids into the culture medium even in the absence of any iron chelators (the concentration of pyruvic acid, for example, was >25 mM). Moreover, RB181 was able to grow and excrete alpha-keto acids in the presence of alpha,alpha'-dipyridyl at concentrations threefold greater than that which inhibited the growth of enb-7.

2,2'-Dipyridyl↗

Effect of glucose and pyruvate in acidic and non-acidic peritoneal dialysis fluids on leukocytes cell functions.

A new peritoneal dialysate containing pyruvate anions has been tested for its effects on cell functions and compared with conventional lactate and bicarbonate based solutions. The dialysate has a final pH of 5.4 to 5.6 and is composed of 1.36 to 3.86% glucose-monohydrate, 132 mmol/liter sodium, 1.75 mmol/liter calcium, 0.75 mmol/liter magnesium, 102 mmol/liter chloride and 35 mmol/liter pyruvate. For cytotoxicity testing peritoneal macrophages and peripheral blood mononuclear cells (PBMC) were exposed to conventional lactate dialysate, pyruvate dialysate, bicarbonate dialysate and a control medium RPMI 1640 (Biochrom KG, Berlin, Germany), followed by activation with different bacterial stimuli. In addition, the study further investigated the effect of varying glucose concentration in the different dialysates ranging from 0 to 3.86% and pH changes between 5.2 and 7.4 on the cytotoxicity effect on the selected cells. Mononuclear cells exposed to pyruvate-based dialysate before stimulation with endotoxin exhibited a tumor necrosis factor (TNF)-mRNA signal comparable to those of cells exposed to RPMI. In contrast, exposure to lactate-based dialysate completely inhibited TNF-mRNA synthesis. In addition, cytokine synthesis in macrophages and PBMCs after exposure to pyruvate was less inhibited when compared to the corresponding levels measured after exposure to lactate. The chemotactic response of polymorphonuclear cells and O-2 generation in all tested cell types after exposure to pyruvate was found not to be inhibited, whereas a complete inhibition was observed after exposure to lactate. The results demonstrate that cytotoxicity effects of peritoneal dialysate on cell lines can be minimized by using a new dialysate formulation containing pyruvate anions instead of lactate.

Cell Line↗

The effects of the interactions between amino acids on pyruvate kinase activity from the brain cortex of young rats.

Considering that pyruvate kinase activity, a crucial enzyme for glucose metabolism and energy liberation in brain, may be regulated by some amino acids, it is possible that diminution of this enzyme activity may contribute to the brain damage caused by amino acids accumulated in metabolic diseases, such as phenylalanine, tryptophan and cystine. Therefore, the present study was undertaken to investigate the effect of these amino acids on pyruvate kinase activity in the brain cortex of rats. We also investigated the effect of serine and alanine on pyruvate kinase activity in the same tissue. The results suggested that phenylalanine, tryptophan, cystine, alanine, and serine act at the same site on the enzyme, phenylalanine, tryptophan, and cystine causing inhibition, and alanine and serine preventing this effect. Cystine also inhibited the enzyme activity through a different mechanism, possibly acting on the enzyme thiol groups. Considering that this enzyme is a target for amino acids accumulated in some metabolic diseases of amino acid metabolism, it is possible that its inhibition may contribute to the brain damage found in these diseases.

Amino Acid Metabolism, Inborn Errors↗

[Effects of ultraviolet irradiation of blood and sodium hypochlorite on aromatic amino acid metabolism in phlegmons of the maxillofacial region].

The investigation was performed in 39 patients with the severe course of phlegmons of the maxillofacial area. The concentration of the phenyl-pyruvic and para-hydroxyphenyl-pyruvic acids in the diurnal urine was found to be elevated which suggested the disturbed functional state of the liver. Treatment including the ultraviolet irradiation of blood and sodium hypochlorite resulted in rapid normalization of the level of para-hydroxyphenyl-pyruvic acid and in lower concentration of phenyl-pyruvic acid.

Adult↗

Branched-chain keto-acids and pyruvate in blood: measurement by HPLC with fluorimetric detection and changes in older subjects.

BACKGROUND: Measurement of keto-acids is important in various clinical situations. The aim of the present work was to develop a rapid HPLC method for the determination of keto-acids in human serum and to assess the concentrations of these acids in young adults and institutionalized elderly adults. This method was applied to the determination of blood keto-acid concentrations of young adults and institutionalized elderly people, divided into age groups METHODS: Four keto-acids (alpha-ketoisocaproate, alpha-ketoisovalerate, alpha-keto-beta-methylvalerate, and pyruvate) were derivatized with o-phenylenediamine to give fluorescent derivatives. After the sample preparation step (75 min to prepare 20 samples), the derivatives were separated chromatographically on a reversed-phase column using a binary gradient. RESULTS: The fluorometric detection of the four keto-acids was rapid, <12 min. The method is repeatable and reproducible: the CVs were <6% and <11%, respectively, for each of the keto-acids. We found no significant difference between males and females. Concentrations of the branched-chain keto-acids decreased after age 60 years, especially alpha-ketoisocaproate, which decreased approximately 40%. CONCLUSIONS: The proposed method allows rapid and reliable measurement of keto-acids. The data demonstrate that changes in branched-chain keto-acids concentrations in serum occur with age.

Adult↗