Formate Excretion in Urine of Rats Fed Dimethylaminoazobenzene-rich Diets: The Possibility of Formate Formation from D-Lactate

This experiment was carried out to evaluate the possibility of degradation of d-lactate into formate and acetaldehyde. In order to induce hyperproduction of d-lactate in rats. Donryu male albino rats were fed diets containing 0.064% 3'-methyl-4-dimethylaminoazobenzene (3'-MDAB), 4'-methyl-4-dimethylaminoazobenzene (4'-MDAB) or 2-methyl-4-dimethylaminoazobenzene (2-MDAB) for 10 weeks. During the experiment, body mass, food and water intake and volume of urine were documented. Methylglyoxal, d-lactate and formate in the urine samples were determined. On the first day of the eleventh week, methylglyoxal, d-lactate, glutathione and enzymatic activities of demethylation and glyoxalase I and II in liver were measured. Methylglyoxal, d-lactate and clinical chemistry parameters of blood plasma were also measured. The levels of methylglyoxal and d-lactate in livers of rats fed 3'-MDAB were very high, while those of 2-MDAB fed-rats and the control group were the same. The fact that glyoxalase I activity and the level of glutathione, a cofactor of glyoxalase I, were high in the livers of the 3'-MDAB-fed rats can explain the elevated levels of methylglyoxal and d-lactate in the liver. The most striking results were the elevated formate levels in the urine of rats fed 3'- and 4'-MDAB in a precancerous state. The degradation of d-lactate, an end product of the methylglyoxal bypass, into acetaldehyde and formate was suggested as a possible way to explain the results.</p

Circulating anions usually associated with the Krebs cycle in patients with metabolic acidosis

Introduction: Acute metabolic acidosis of non-renal origin is usually a result of either lactic or ketoacidosis, both of which are associated with a high anion gap. There is increasing recognition, however, of a group of acidotic patients who have a large anion gap that is not explained by either keto- or lactic acidosis nor, in most cases, is inappropriate fluid resuscitation or ingestion of exogenous agents the cause. Methods: Plasma ultrafiltrate from patients with diabetic ketoacidosis, lactic acidosis, acidosis of unknown cause, normal anion gap metabolic acidosis, or acidosis as a result of base loss were examined enzymatically for the presence of low molecular weight anions including citrate, isocitrate, α-ketoglutarate, succinate, malate and d-lactate. The results obtained from the study groups were compared with those obtained from control plasma from normal volunteers. Results: In five patients with lactic acidosis, a significant increase in isocitrate (0.71 ± 0.35 mEq l-1), α-ketoglutarate (0.55 ± 0.35 mEq l-1), malate (0.59 ± 0.27 mEq l-1), and d-lactate (0.40 ± 0.51 mEq l-1) was observed. In 13 patients with diabetic ketoacidosis, significant increases in isocitrate (0.42 ± 0.35 mEq l-1), α-ketoglutarate (0.41 ± 0.16 mEq l-1), malate (0.23 ± 0.18 mEq l-1) and d-lactate (0.16 ± 0.07 mEq l-1) were seen. Neither citrate nor succinate levels were increased. Similar findings were also observed in a further five patients with high anion gap acidosis of unknown origin with increases in isocitrate (0.95 ± 0.88 mEq l-1), α-ketoglutarate (0.65 ± 0.20 mEq l-1), succinate (0.34 ± 0.13 mEq l-1), malate (0.49 ± 0.19 mEq l-1) and d-lactate (0.18 ± 0.14 mEq l-1) being observed but not in citrate concentration. In five patients with a normal anion gap acidosis, no increases were observed except a modest rise in d-lactate (0.17 ± 0.14 mEq l-1). Conclusion: The levels of certain low molecular weight anions usually associated with intermediary metabolism were found to be significantly elevated in the plasma ultrafiltrate obtained from patients with metabolic acidosis. Our results suggest that these hitherto unmeasured anions may significantly contribute to the generation of the anion gap in patients with lactic acidosis and acidosis of unknown aetiology and may be underestimated in diabetic ketoacidosis. These anions are not significantly elevated in patients with normal anion gap acidosis

A probable dual mode of action for both L- and D-lactate neuroprotection in cerebral ischemia.

Lactate has been shown to offer neuroprotection in several pathologic conditions. This beneficial effect has been attributed to its use as an alternative energy substrate. However, recent description of the expression of the HCA1 receptor for lactate in the central nervous system calls for reassessment of the mechanism by which lactate exerts its neuroprotective effects. Here, we show that HCA1 receptor expression is enhanced 24 hours after reperfusion in an middle cerebral artery occlusion stroke model, in the ischemic cortex. Interestingly, intravenous injection of L-lactate at reperfusion led to further enhancement of HCA1 receptor expression in the cortex and striatum. Using an in vitro oxygen-glucose deprivation model, we show that the HCA1 receptor agonist 3,5-dihydroxybenzoic acid reduces cell death. We also observed that D-lactate, a reputedly non-metabolizable substrate but partial HCA1 receptor agonist, also provided neuroprotection in both in vitro and in vivo ischemia models. Quite unexpectedly, we show D-lactate to be partly extracted and oxidized by the rodent brain. Finally, pyruvate offered neuroprotection in vitro whereas acetate was ineffective. Our data suggest that L- and D-lactate offer neuroprotection in ischemia most likely by acting as both an HCA1 receptor agonist for non-astrocytic (most likely neuronal) cells as well as an energy substrate

Plasmodium falciparum glyoxalase II: Theorell-Chance product inhibition patterns, rate-limiting substrate binding via Arg(257)/Lys(260), and unmasking of acid-base catalysis

Glyoxalase II (GloII) is a ubiquitous thioester hydrolase catalyzing the last step of the glutathione-dependent conversion of 2-oxoaldehydes to 2-hydroxycarboxylic acids. Here, we present a detailed structure-function analysis of cGloII from the malaria parasite Plasmodium falciparum. The activity of the enzyme was salt-sensitive and pH-log k(cat) and pH-log k(cat)/K-m profiles revealed acid-base catalysis. An acidic pK(a)(app) value of approximately 6 probably reflects hydroxide formation at the metal center. The glutathione-binding site was analyzed by site-directed mutagenesis. Substitution of residue Arg(154) caused a 2.5-fold increase of K-m(app), whereas replacements of Arg(257) or Lys(260) were far more detrimental. Although the glutathione-binding site and the catalytic center are separated, six of six single mutations at the substrate-binding site decreased the k(cat)(app) value. Furthermore, product inhibition studies support a Theorell-Chance Bi Bi mechanism with glutathione as the second product. We conclude that the substrate is predominantly bound via ionic interactions with the conserved residues Arg(257) and Lys(260), and that correct substrate binding is a pH-and salt-dependent rate-limiting step for catalysis. The presented mechanistic model is presumably also valid for GloII from many other organisms. Our study could be valuable for drug development strategies and enhances the understanding of the chemistry of binuclear metallohydrolases

The preparation of crystalline lactic acid

On account of its importance in intermediary metabolism, lactic acid was among the first compounds chosen in our plan, which we have described in a previous communication (1), to augment the available data on the free energies of formation of substances significant in biological chemistry. It was necessary for this purpose to obtain pure crystalline lactic acid, free of water, anhydride, and lactide. The only description in the literature of the preparation of crystalline lactic acid is that of Krafft and Dÿes (2). Table I shows that the product obtained by their method contains relatively large quantities of anhydro impurities. The subject of the present communication is the description of a method which yields the active isomers of lactic acid in a crystalline state, free of water, anhydride, and lactide, supplemented by the description of two methods of separating the active forms from the commercial syrup (1). Lactic acid commercially available at present either is in the form of the U.S.P. syrup, which usually exhibits a low optical activity corresponding to the excess it happens to contain, which is variable, of one or the other optical isomer, or is the expensive zinc sarcolactate. The methods described below now make it possible to obtain easily and quickly and at low cost large quantities of both active isomers in a relatively high degree of purity

Genetic Tool Development for a New Host for Biotechnology, the Thermotolerant Bacterium Bacillus coagulans

Bacillus coagulans has good potential as an industrial production organism for platform chemicals from renewable resources but has limited genetic tools available. Here, we present a targeted gene disruption system using the Cre-lox system, development of a LacZ reporter assay for monitoring gene transcription, and heterologous D-lactate dehydrogenase expression

Current status of laboratory and imaging diagnosis of neonatal necrotizing enterocolitis

Necrotizing enterocolitis continues to be a devastating disease process for very low birth weight infants in Neonatal Intensive Care Units. The aetiology and pathogenesis of necrotizing enterocolitis are not definitively understood. It is known that necrotizing enterocolitis is secondary to a complex interaction of multiple factors that results in mucosal damage, which leads to intestinal ischemia and necrosis. Advances in neonatal care, including resuscitation and ventilation support technology, have seen increased survival rates among premature neonates and a concomitant detection in the incidence of this intestinal disease.Diagnosis can be difficult, and identifying infants at the onset of disease remains a challenge. Early diagnosis, which relies on imaging findings, and initiation of prompt therapy are essential to limit morbidity and mortality. Moreover, early management is critical and life-saving.This review summarizes what is known on the laboratory and instrumental diagnostic strategies needed to improve neonatal outcomes and, possibily, to prevent the onset of an overt necrotizing enterocolitis

Modelling and Optimization of the (R)-(+)-3,4-dihydroxyphenyllactic Acid Production Catalyzed with D-lactate Dehydrogenase from Lactobacillus leishmannii Using Genetic Algorithm

A mathematical model for the enzymatic kinetics of the synthesis of (R)–(+)–3,4-dihydroxyphenyllactic acid (DHPL) was developed. The synthesis was catalyzed by D-lactate dehydrogenase from Lactobacillus leishmannii. Since this enzyme requires NADH as a coenzyme, formate dehydrogenase system was used for NADH regeneration. Kinetic constants of both enzymes were estimated independently from initial reaction rate experiments. The developed mathematical model was verified by the batch reactor experiment (volumetric productivity in this experiment was 4.76 g dm–3 d–1). Optimization of initial reaction conditions for DHPL synthesis was performed using the genetic algorithm (GA). The genetic algorithm as a flexible optimization tool had been used to obtain the experimental conditions where maximal volumetric productivity could be achieved. The optimal initial conditions were found in the investigated parameter area: c3,4-dihydroxyphenylpyruvic acid = 4.69 mmol dm–3, cNAD+= 4.95 mmol dm–3, cformate = 36.85 mmol dm–3, D-lactate dehydrogenase = 3 mg cm–3, formate dehydrogenase = 2.94mg cm–3 and the reaction time 8.5 min. At these conditions volumetric productivity of 93.06 g dm–3 d–1 can be achieved

The Explanatory Indispensability of Memory Traces

During the first half of the twentieth century, many philosophers of memory opposed the postulation of memory traces based on the claim that a satisfactory account of remembering need not include references to causal processes involved in recollection. However, in 1966, an influential paper by Martin and Deutscher showed that causal claims are indeed necessary for a proper account of remembering. This, however, did not settle the issue, as in 1977 Malcolm argued that even if one were to buy Martin and Deutscher’s argument for causal claims, we still don’t need to postulate the existence of memory traces. This paper reconstructs the dialectic between realists and anti-realists about memory traces, suggesting that ultimately realists’ arguments amount to inferences to the best explanation. I then argue that Malcolm’s anti-realist strategy consists in the suggestion that causal explanations that do not invoke memory traces are at least as good as those that do. But then, Malcolm, I argue that there are a large number of memory phenomena for which explanations that do not postulate the existence of memory traces are definitively worse than explanations that do postulate them. Next, I offer a causal model based on an interventionist framework to illustrate when memory traces can help to explain memory phenomena and proceed to substantiate the model with details coming from extant findings in the neuroscience of memory