Concentrations Of Succinate Research Articles

Effect of copper stress on Phaseolus coccineus in the presence of exogenous methyl jasmonate and/or Serratia plymuthica from the Spitsbergen soil

Copper stress in the presence of exogenous methyl jasmonate and Serratia plymuthica in a complete trifactorial design with copper (0, 50 µM), methyl jasmonate (0, 1, 10 µM) and Serratia plymuthica (without and with inoculation) was studied on the physiological parameters of Phaseolus coccineus. Copper application reduced biomass and allantoin content, but increased chlorophyll and carotenoids contents as well as catalase and peroxidases activities. Jasmonate did not modify biomass and organic acids levels under copper treatment, but additional inoculation elevated biomass and content of tartrate, malate and succinate. Jasmonate used alone or in combination with bacteria increased superoxide dismutase activity in copper application. With copper, allantoin content elevated at lower jasmonate concentration, but with additional inoculation – at higher jasmonate concentration. Under copper stress, inoculation resulted in higher accumulation of tartrate, malate and citrate contents in roots, which corresponded with lower allantoin concentration in roots. Combined with copper, inoculation reduced catalase and guaiacol peroxidase activities, whereas organic acids content was higher. Under metal stress, with bacteria, jasmonate reduced phenolics content, elevated superoxide dismutase and guaiacol peroxidase activities. The data indicate that jasmonate and S. plymuthica affected most physiological parameters of P. coccineus grown with copper and revealed some effect on biomass.

Kinetic modeling of succinate production from glucose and xylose by metabolically engineered Escherichia coli KJ12201

This study proposed a kinetic model for succinate production from xylose and glucose by Escherichia coli KJ12201 based on the Monod equation in which inhibitions of xylose or glucose and succinate were simultaneously considered. Our proposed model showed the strong inhibition effect by xylose on the specific growth rate while glucose had no inhibitory effect. Additionally, maximum concentrations of succinate that completely inhibited the cell growth were 60.70 g/L for xylose and 82.71 g/L for glucose. The kinetic parameters of the proposed model were determined by minimizing the gap between experimental data and the model prediction. With estimated kinetic parameters, values solved by the proposed model aligned and suited experimental data in the range of substrate concentrations (20–80 g/L) used. The sensitivity analysis also revealed that the maximum specific growth rate (µmax) and the maximum inhibitory xylose concentration (C*xylose) provided extreme influences on the succinate production. Our proposed model was then validated and found that it precisely predicted the cell growth, substrate utilization and succinate production by the strain. The proposed model may be employed for effectively controlling and optimizing the succinate production from other substrates containing glucose and xylose in a large-scale process by E. coli KJ12201.

Metabolomic Analysis Reveals That the Moor Frog Rana arvalis Uses Both Glucose and Glycerol as Cryoprotectants.

Simple SummaryThe moor frog Rana arvalis can tolerate freezing to low temperatures, up to −16 °C. We performed metabolomic analysis of the liver and hindlimb muscles of frozen and control R. arvalis. We found that the moor frog synthesizes glucose and glycerol in similar concentrations as low molecular weight cryoprotectants. This is the first such case reported for the genus Rana, which was believed to use glucose only. We found that freezing upregulates glycolysis, with the accumulation of several end products: lactate, alanine, ethanol, and, possibly, 2,3-butanediol. To our knowledge, this is also the first report of ethanol as an end product of glycolysis in terrestrial vertebrates. We observed highly increased concentrations of nucleotide degradation products, implying high level of stress. We found almost no signs of adaptations to reoxygenation stress, with overall low levels of antioxidants. We also performed metabolomics analysis of subcutaneous ice that was found to contain glucose, glycerol, and several other substances.The moor frog Rana arvalis is one of a few amphibians that can tolerate freezing to low temperatures, up to −16 °C. In this study, we performed metabolomic analysis of the liver and hindlimb muscles of frozen and control R. arvalis. We found that the moor frog synthesizes glucose and glycerol in similar concentrations as low molecular weight cryoprotectants. This is the first such case reported for the genus Rana, which was believed to use glucose only. We found that freezing upregulates glycolysis, with the accumulation of several end products: lactate, alanine, ethanol, and, possibly, 2,3-butanediol. To our knowledge, this is also the first report of ethanol as an end product of glycolysis in terrestrial vertebrates. We observed highly increased concentrations of nucleotide degradation products, implying high level of stress. The Krebs cycle arrest resulted in high concentrations of succinate, which is common for animals. However, we found almost no signs of adaptations to reoxygenation stress, with overall low levels of antioxidants. We also performed metabolomics analysis of subcutaneous ice that was found to contain glucose, glycerol, and several other substances.

Short Chain Fatty Acids and Bacterial Taxa Associated with Reduced Salmonella enterica serovar I 4,[5],12:i:- Shedding in Swine Fed a Diet Supplemented with Resistant Potato Starch.

ABSTRACTSalmonella enterica serovar I 4,[5],12:i:- is a foodborne pathogen of concern because many isolates are multidrug-resistant (resistant to ≥3 antimicrobial classes) and metal tolerant. In this study, three in-feed additives were individually tested for their ability to reduce Salmonella I 4,[5],12:i:- shedding in swine: resistant potato starch (RPS), high amylose corn starch, and a fatty acid blend, compared with a standard control diet over 21 days. Only RPS-fed pigs exhibited a reduction in Salmonella fecal shedding, different bacterial community compositions, and different cecal short chain fatty acid (SCFA) profiles relative to control animals. Within the RPS treatment group, pigs shedding the least Salmonella tended to have greater cecal concentrations of butyrate, valerate, caproate, and succinate. Additionally, among RPS-fed pigs, several bacterial taxa (Prevotella_7, Olsenella, and Bifidobacterium, and others) exhibited negative relationships between their abundances of and the amount of Salmonella in the feces of their hosts. Many of these same taxa also had significant positive associations with cecal concentrations of butyrate, valerate, caproate, even though they are not known to produce these SCFAs. Together, these data suggest the RPS-associated reduction in Salmonella shedding may be dependent on the establishment of bacterial cross feeding interactions that result in the production of certain SCFAs. However, directly feeding a fatty acid mix did not replicate the effect. RPS supplementation could be an effective means to reduce multidrug-resistant (MDR) S. enterica serovar I 4,[5],12:i:- in swine, provided appropriate bacterial communities are present in the gut.IMPORTANCE Prebiotics, such as resistant potato starch (RPS), are types of food that help to support beneficial bacteria and their activities in the intestines. Salmonella enterica serovar I 4,[5],12:i:- is a foodborne pathogen that commonly resides in the intestines of pigs without disease, but can make humans sick if unintentionally consumed. Here we show that in Salmonella inoculated pigs, feeding them a diet containing RPS altered the colonization and activity of certain beneficial bacteria in a way that reduced the amount of Salmonella in their feces. Additionally, within those fed RPS, swine with higher abundance of these types of beneficial bacteria had less Salmonella I 4,[5],12:i:- in their feces. This work illustrates likely synergy between the prebiotic RPS and the presence of certain gut microorganisms to reduce the amount of Salmonella in the feces of pigs and therefore reduce the risk that humans will become ill with MDR Salmonella serovar I 4,[5],12:i:-.

Temperature-Induced Environmental Chain Reaction in Marine Sedimentation and Its Impact on Manila Clam Ruditapes philippinarum

Marine Heatwaves (MHWs) events have been increasing, causing severe impacts on marine ecosystems and aquaculture. In this study, the effects of temperature on changes in sediment dissolved oxygen (DO) and hydrogen sulfide (H2S) concentrations and their effects on the behavioral and physiological responses of a bivalve were investigated by simulating different sudden summer temperature change conditions. The results showed that temperature was an important factor affecting DO consumption and H2S release in sediments, and sediment type also played a key role. At higher temperatures, DO was consumed more rapidly and H2S release increased in sediments, and the DO was consumed and H2S released in sandy sediments was less than in silty sand sediments. The response of Manila clam’s exercise behavior under environmental pressure was also rapid. The excavation index of the Manila clam decreased with the DO consumption, indicating that the Manila clam could alter its burial depth and move toward the sediment-water interface when stressed. Compared with phenol oxidases (PO) activity being activated under experimental conditions, succinate concentrations only increased slightly at 32°C, and did not surpass the threshold indicative of anaerobic metabolism. However, when toxic substances such as H2S start to accumulate, they may damage the immune system and tissues of the Manila clam, thereby affecting its future survival. Therefore, when temperatures are high for long periods in summer, it is necessary to take timely action to prevent and guard against harm caused by DO and H2S to aquaculture organisms.

Ethyl carbamate regulate esters degradation by activating hydrolysis during Baijiu ripening

The presence of ethyl carbamate in traditional fermented food is a public health concern for the FDA. The effect of forced photoirradiation (0–150,000 lx) and thermal stress (4, 25, 40 °C) on ethyl carbamate (0–150 μg/L) on physiological characteristics of Baijiu, such as esters content, photochemical degradation, and hydrogen-bond interaction efficiency were monitored by ultra high performance liquid chromatography quadrupole-orbitrap and the dynamic changes by digital foodomics analysis. Furthermore, 748 trace components covering 11 subclasses were identified in Baijiu and 71 esters were screened by Spearman’s correlation, fold changes, P values and VIP values. A forward stepwise multiple regression and discriminant analysis were performed for predicting the content of esters from appearance characteristics obtained by foodomics analysis, reaching R-square values up to 0.91. A reduction of the present variation in ethyl lactate, ethyl caproate, diethyl succinate, ethyl oleate and ethyl linoleate concentration could possibly result in a better understanding of the ethyl carbamate effects. Ethyl carbamate was found to cause esters hydrolysis through inter-molecular interaction in various species of alcoholic drinks. It was demonstrated that the light exposure level and thermal intensity applied for ethyl carbamate-spiked Baijiu samples did not unambiguously influence esters concentration in Baijiu. Future research should focus on moderate light exposure level and thermal stress and should aim at reducing natural ethyl carbamate by more closely controlling the esters content of the ripening degree.

Not Just a Cycle: Three gab Genes Enable the Non-Cyclic Flux Toward Succinate via GABA Shunt in 'Candidatus Liberibacter asiaticus'-Infected Citrus.

Although the mitochondria retain all required enzymes for an intact tricarboxylic acid (TCA) cycle, plants might shift the cyclic flux from the TCA cycle to an alternative noncyclic pathway via γ-aminobutyric acid (GABA) shunt under specific physiological conditions. We hypothesize that several genes may ease this noncyclic flux and contribute to the citrus response to the phytopathogenic bacterium 'Candidatus Liberibacter asiaticus', the causal agent of Huanglongbing in citrus. To test this hypothesis, we used multiomics techniques (metabolomics, fluxomics, and transcriptomics) to investigate the potential roles of putative gab homologies from Valencia sweet orange (Citrus sinensis). Our findings showed that 'Ca. L. asiaticus' significantly increased the endogenous GABA and succinate content but decreased ketoglutarate in infected citrus plants. Citrus genome harbors three putative gab genes, including amino-acid permease (also known as GABA permease; CsgabP), GABA transaminase (CsgabT), and succinate-semialdehyde dehydrogenase (also known as GABA dehydrogenase; CsgabD). The transcript levels of CsgabP, CsgabT, and CsgabD were upregulated in citrus leaves upon the infection with 'Ca. L. asiaticus' and after the exogenous application of GABA or deuterium-labeled GABA isotope (GABA-D6). Moreover, our finding showed that exogenously applied GABA is quickly converted to succinate and fed into the TCA cycle. Likewise, the fluxomics study showed that GABA-D6 is rapidly metabolized to succinate-D4. Our work proved that GABA shunt and three predicated gab genes from citrus, support the upstream noncyclic flux toward succinate rather than an intact TCA cycle and contribute to citrus defense responses to 'Ca. L. asiaticus'.[Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Cold or Not So Cold?-Static Organ Preservation at 10 °C May Prolong Organ Preservation and Facilitate Transplant Logistics.

Cold or Not So Cold?-Static Organ Preservation at 10 °C May Prolong Organ Preservation and Facilitate Transplant Logistics.

Characteristics of gut microbiome, organic acid profiles and viral antibody indexes of healthy Japanese with live Lacticaseibacillus detected in stool.

To estimate the health-promoting effects of Lacticaseibacillus paracasei (previously Lactobacillus casei) strain Shirota (LcS) that reached the lower gastrointestinal tract alive, we investigated the characteristics of gut microbiome, organic acid profiles, defecatory symptoms and serum viral antibody indexes of healthy Japanese adults between the group in whom live LcS was detected or not from stool. The β-diversity index of the gut microbiome constituted a significant difference between the live-LcS-detected-group (LLD) and the live-LcS-not-detected-group (LLnD). In the LLD, the Bifidobacteriaceae, Lactobacillaceae, and Coriobacteriaceae counts were significantly higher, and the succinate concentration was significantly lower than that in the LLnD. The serum herpes simplex virus (HSV) immunoglobulin (Ig)M antibody index in the LLD tended to be lower than that of the LLnD in HSV IgG-positive subjects. Of the LLD, those in the fermented milk products containing LcS (FML)-high-frequency-group (FML-HF) and those in the FML-low-frequency-group (FML-LF) had different gut microbiome and organic acid profiles. However, the pattern of differences between FML-HF and FML-LF was dissimilar those between LLD and LLnD. In contrast, among subjects with FML-LF, those in the group with LLD in stool (LF-LLD) and those in the LLnD in stool (LF-LLnD) showed a similar pattern of differences in their gut microbiome and organic acid profiles as those in the LLnD and LLD. The LLD and LF-LLD commonly had lower caloric and carbohydrate intakes from the diet than their respective control groups. In this study, we found that the presence of live LcS in stool is associated with a healthy gut environment and inhibition of the reactivation of latently infected viruses in the host. However, these health-promoting effects on the host were not related to the frequency of FML intake. Furthermore, dysbiosis of the gut microbiome and diet including caloric intake was related to the viability of ingested LcS in the gut.

Gastrointestinal Tract and Dietary Fiber Driven Alterations of Gut Microbiota and Metabolites in Durco × Bamei Crossbred Pigs.

Gastrointestinal tract and dietary fiber (DF) are known to influence gut microbiome composition. However, the combined effect of gut segment and long-term intake of a high fiber diet on pig gut microbiota and metabolite profiles is unclear. Here, we applied 16S rRNA gene sequencing and untargeted metabolomics to investigate the effect of broad bean silage on the composition and metabolites of the cecal and jejunal microbiome in Durco × Bamei crossbred pigs. Twenty-four pigs were allotted to four graded levels of DF chow, and the content of jejunum and cecum were collected. Our results demonstrated that cecum possessed higher α-diversity and abundance of Bacteroidetes, unidentified Ruminococcaceae compared to jejunum, while jejunum possessed higher abundance of Lactobacillus, Streptococcus. DF intake significantly altered diversity of the bacterial community. The abundance of Bacteroidetes and Turicibacter increased with the increase of DF in cecum and jejunum respectively. Higher concentrations of amino acids and conjugated bile acids were detected in the jejunum, whereas free bile acids and fatty acids were enriched in the cecum. The concentrations of fatty acids, carbohydrate metabolites, organic acids, 2-oxoadipic acid, and succinate in cecum were higher in the high DF groups. Overall, the results indicate that the composition of bacteria and the microbiota metabolites were distinct in different gut segments. DF had a significant influence on the bacterial composition and structure in the cecum and jejunum, and that the cecal metabolites may further affect host health, growth, and slaughter performance.

Complexation of Np(V) with the Dicarboxylates, Malonate, and Succinate: Complex Stoichiometry, Thermodynamic Data, and Structural Information

The complexation of Np(V) with malonate and succinate is studied by different spectroscopic techniques, namely, attenuated total reflection Fourier transform infrared (ATR FT-IR) and extended X-ray absorption fine-structure (EXAFS) spectroscopy, as well as by quantum chemistry to determine the speciation, thermodynamic data, and structural information of the formed complexes. For complex stoichiometries and the thermodynamic functions (log βn°(Θ), ΔrHn°, ΔrSn°), near infrared absorption spectroscopy (vis/NIR) is applied. The complexation reactions are investigated as a function of the total concentration of malonate ([Mal2-]total) and succinate ([Succ2-]total), ionic strength [Im = 0.5-4.0 mol kg-1 Na+(Cl-/ClO4-)], and temperature (Θ = 20-85 °C). Besides the solvated NpO2+ ion, the formation of two Np(V) species with the stoichiometry NpO2(L)n1-2n (n = 1, 2, L = Mal2-, Succ2-) is observed. With increasing temperature, the molar fractions of both complex species increase and the temperature-dependent conditional stability constants log βn'(Θ) at given ionic strengths are determined by the law of mass action. The log βn'(Θ) are extrapolated to IUPAC reference-state conditions (Im = 0) according to the specific ion interaction theory (SIT), revealing thermodynamic log βn°(Θ) values. For all formed complexes, [NpO2(Mal)-: log β1°(25 °C) = 3.36 ± 0.11, NpO2(Mal)23-: log β2°(25 °C) = 3.95 ± 0.19, NpO2(Succ)-: log β1°(25 °C) = 2.05 ± 0.45, NpO2(Succ)23-: log β2°(25 °C) = 0.75 ± 1.22], an increase of the stability constants with increasing temperature was observed. This confirmed an endothermic complexation reaction. The temperature dependence of the log βn°(T) values is described by the integrated Van't Hoff equation, and the standard reaction enthalpies and entropies for the complexation reactions are determined. Furthermore, the sum of the specific binary ion-ion interaction coefficients Δεn°(Θ) for the complexation reactions are obtained as a function of the t from the respective SIT modeling as a function of the temperature. In addition to the thermodynamic data, the structures of the complexes and the coordination modes of malonate and succinate are investigated using EXAFS spectroscopy, ATR-FT-IR spectroscopy, and quantum chemical calculations. The results show that in the case of malonate, six-membered chelate complexes are formed, whereas for succinate, seven-membered rings form. The latter ones are energetically unfavorable due to the limited space in the equatorial plane of the Np(V) ion (as NpO2+ cation).

Effects of ultra-long fermentation time on the microbial community and flavor components of light-flavor Xiaoqu Baijiu based on fermentation tanks.

Microbial structure and succession of fermented grains play a significant role in Baijiu’s flavor and quality. In this study, high-throughput sequencing (HTS) coupled with headspace solid-phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS) were used to analyze the microbial community structures and flavor components in the fermented grains at the end of fermentation from different fermentation time of light-flavor Xiaoqu Baijiu. HTS results showed that Lactobacillus acetotolerans, Lactobacillus helveticus, Lactobacillus buchneri, Wickerhamomyces, Saccharomyces, and Condenascus were identified as the dominant microbes, but Lactobacillus (96.28%) exhibited obvious advantages at the end of ultra-long fermentation time (day 98). HS-SPME-GC-MS analysis revealed that esters and alcohols had the most abundance in fermented grains of day 98, containing high concentrations of ethyl acetate, diethyl succinate, phenylethyl alcohol, isoamyl alcohol, and n-propanol, which were related to the succession of Lactobacillus and yeast communities. Interestingly, the content of n-propanol in the ultra-long fermentation time samples (day 98) was 6 times of that in normal fermented grains (day 14), which may be caused by higher abundance of Lactobacillus in day 98 samples. Monte Carlo permutation test showed residual starch, acidity, and amino nitrogen (p < 0.05) were important factors affecting the microbial community. Together, these results shed light on the physicochemical changes, microbial dynamics, and key flavor components of fermented grains at the end of fermentation from different fermentation time and provide a strategy for further improvement of Baijiu quality.

Abstract 11005: Electrophysiological Remodelling of Right Atria in Atrial Fibrillation Sheep Model and Role of Succinate in Electrophysiological Properties

Introduction: Atrial fibrillation (AF) is the most common arrhythmia and is associated with deleterious remodelling processes. In AF patients, the plasmatic succinate content increases, indicating a potential role for metabolic remodelling in AF progression. Objective: To assess the impact of succinate on the electrophysiological properties of right atria (RA) in a persistent AF sheep model. Methods: Optical mapping experiments were performed in ex vivo RA from a burst pacing AF sheep model (SHAM=5; AF=5). The right coronary artery was perfused with Tyrode solution initially containing glucose (5.6mM), subsequently replaced by succinate (5mM). Action potential durations at 80% of repolarization (APD80) and mean conduction velocities (CV) were assessed from 2 to 5Hz pacing frequency and during sinus rhythm (SR). An S1S2 protocol was used to determine effective refractory period (ERP) and a burst pacing procotol (20Hz) to assess ex vivo AF vulnerability. Finally, an organ donation program allowed us to investigate these properties in a human RA from an AF patient. Results: SR frequency was higher in AF than SHAM sheep (2.0 vs. 1.6 Hz; p=0.19). Succinate perfusion decreased SR frequency by 18% in SHAM sheep (1.7 vs. 1.4 Hz; p=0.12) and by 43% in AF sheep (2.0 vs. 1.2 Hz; p=0.009). The ERP was lower in AF than SHAM sheep (184 vs. 210 ms; p=0.5) and succinate induced a significant increase of 136% in SHAM sheep (210 vs. 286 ms; p=0.06) and of 188% in AF sheep (184 vs. 346 ms; p=0.0008). Epicardial APD80 was lower when pacing at 2.5Hz in AF than SHAM sheep (165 vs. 197 ms ; p=0.3). Succinate increased APD80 in both SHAM and AF RA (191 to 224 ms; 165 to 231 ms) but its effect was more important in AF than SHAM sheep (148 vs. 117 %; p=0.1). Mean CV was slower in AF sheep than SHAM sheep (110 vs. 130 cm/s; p=0.47). Succinate slowed mean CV on AF sheep (110 vs. 76; p=0.4) but did not affect SHAM RA. In the human RA, APD80 (257 vs; 309 ms) and total activation time were increased by succinate. Conclusions: Our results indicate that succinate alters atrial electrophysiological properties, most likely through the associated ROS overproduction. Importantly, AF sheep showed an increased sensitivity to succinate perfusion. Preliminary data in a human RA are in line with these observations.

Biochemical Response to Freezing in the Siberian Salamander Salamandrella keyserlingii.

Simple SummaryThe Siberian salamander is a unique amphibian that is capable to survive long-term freezing at −55 °C. We used 1H-NMR analysis to study quantitative changes of multiple metabolites in liver and hindlimb muscle of the Siberian salamander in response to freezing. For the majority of molecules we observed significant changes in concentrations. Glycerol content in frozen organs was as high as 2% w/w, which confirms its role as a cryoprotectant. No other putative cryoprotectants were detected. Freezing resulted in increased concentrations of glycolysis products: lactate and alanine. Unexpectedly, we detected no increase in concentrations of succinate, which accumulates under ischemia in various tetrapods. Freezing proved to be a dramatic stress with high levels of nucleotide degradation products. There was also significant increase in the concentrations of choline and glycerophosphocholine, which may be interpreted as the degradation of biomembranes. Thus, we found that freezing results not only in macroscopical damage due to ice formation, but also to degradation of DNA and biomembranes.The Siberian salamander Salamandrella keyserlingii Dybowski, 1870 is a unique amphibian that is capable to survive long-term freezing at −55 °C. Nothing is known on the biochemical basis of this remarkable freezing tolerance, except for the fact that it uses glycerol as a low molecular weight cryoprotectant. We used 1H-NMR analysis to study quantitative changes of multiple metabolites in liver and hindlimb muscle of S. keyserlingii in response to freezing. For the majority of molecules we observed significant changes in concentrations. Glycerol content in frozen organs was as high as 2% w/w, which confirms its role as a cryoprotectant. No other putative cryoprotectants were detected. Freezing resulted in ischemia manifested as increased concentrations of glycolysis products: lactate and alanine. Unexpectedly, we detected no increase in concentrations of succinate, which accumulates under ischemia in various tetrapods. Freezing proved to be a dramatic stress with reduced adenosine phosphate pool and high levels of nucleotide degradation products (hypoxanthine, β-alanine, and β-aminoisobutyrate). There was also significant increase in the concentrations of choline and glycerophosphocholine, which may be interpreted as the degradation of biomembranes. Thus, we found that freezing results not only in macroscopical damage due to ice formation, but also to degradation of DNA and biomembranes.

Development of simultaneous quantitative analysis of tricarboxylic acid cycle metabolites to identify specific metabolites in cancer cells by targeted metabolomic approach

The tricarboxylic acid (TCA) cycle is one of the most important pathways of energy metabolism, and the profiles of its components are influenced by factors such as diseases and diets. Therefore, the differences in metabolic profile of TCA cycle between healthy and cancer cells have been the focus of studies to understand pathological conditions. In this study, we developed a quantitative method to measure TCA cycle metabolites using LC-MS/MS to obtain useful metabolic profiles for development of diagnostic and therapeutic methods for cancer. We successfully analyzed 11 TCA cycle metabolites by LC MS/MS with high reproducibility by using a PFP column with 0.5% formic acid as a mobile phase. Next, we analyzed the concentration of TCA cycle metabolites in human cell lines (HaCaT: normal skin keratinocytes; A431: skin squamous carcinoma cells; SW480: colorectal cancer cells). We observed reduced concentration of succinate and increased concentration of citrate, 2-hydroxyglutarate, and glutamine in A431 cells as compared with HaCaT cells. On the other hand, decreased concentration of isocitrate, fumarate, and α-ketoglutarate and increased concentration of malate, glutamine, and glutamate in A431 cells were observed in comparison with SW480 cells. These findings suggested the possibility of identifying disease-specific metabolites and/or organ-specific metabolites by using this targeted metabolomic analysis.

Study of Wine Volatile Composition of Tempranillo versus Tempranillo Blanco, a New White Grape Variety

The aim of this work was to analyze and compare the concentration of higher alcohols, esters, and acids in wines from Tempranillo and Tempranillo Blanco. Tempranillo Blanco is a new and little-studied white variety that originated from Tempranillo by a natural mutation. During three seasons, grapevines of both varieties were harvested, and nine wines were made from each. The volatile composition of the wines was determined by GC-MS. In the wines of both varieties, the content of higher alcohols was higher than those of esters and acids. Wines from Tempranillo Blanco had lower content of 2-phenylethanol, methionol, 1-hexanol, benzyl alcohol, and total higher alcohols, but higher hexyl acetate and ethyl decanoate than Tempranillo wines. Total ethyl esters and total esters were higher in Tempranillo wines due to the higher ethyl lactate and ethyl succinate content derivate from the malolactic fermentation that was not made in Tempranillo Blanco. The content of hexanoic and octanoic acids and total acids was also higher in Tempranillo Blanco wines than in Tempranillo. This is one of the first studies carried out on the wine volatile composition of Tempranillo Blanco and therefore contributes to a better understanding of the oenological characteristics of this white variety.

Predicting chemical shelf life of mozzarella cheese submitted to irregular refrigeration practices by Nuclear Magnetic Resonance spectroscopy and statistical analysis

The complex mixture found in mozzarella cheese favors microorganism proliferation that causes spoilage. Appropriate refrigeration during storage is one step in the supply chain that is critical to guarantee food quality. However, the impact of the absence of night-time refrigeration, practiced by some commercial establishments, in mozzarella chemical profile has not yet been investigated. To this end, during the evening the simulation of improper refrigeration was carried out using an uncovered Styrofoam box containing ice, allowing the system to reach the thermal balance with the environment. The key degradation compounds produced in mozzarella cheese from simulated inadequate refrigeration were identified and quantified using NMR (Nuclear Magnetic Resonance). While the composition and physicochemical analyses were unable to distinguish refrigeration practices, the NMR data revealed that the succinate contents changed significantly between different refrigeration treatments. At the labelled expiry date, inadequate refrigeration had led to an increase in the succinate content of more than 600%. The results suggested a reduction of 4 weeks of the chemical shelf life. Together, NMR and Principal Response Curve analysis are powerful tools to identify mozzarella cheese submitted to different refrigeration conditions.

Untargeted metabolomics identifies succinate as a biomarker and therapeutic target in aortic aneurysm and dissection.

Aortic aneurysm and dissection (AAD) are high-risk cardiovascular diseases with no effective cure. Macrophages play an important role in the development of AAD. As succinate triggers inflammatory changes in macrophages, we investigated the significance of succinate in the pathogenesis of AAD and its clinical relevance. We used untargeted metabolomics and mass spectrometry to determine plasma succinate concentrations in 40 and 1665 individuals of the discovery and validation cohorts, respectively. Three different murine AAD models were used to determine the role of succinate in AAD development. We further examined the role of oxoglutarate dehydrogenase (OGDH) and its transcription factor cyclic adenosine monophosphate-responsive element-binding protein 1 (CREB) in the context of macrophage-mediated inflammation and established p38αMKOApoe-/- mice. Succinate was the most upregulated metabolite in the discovery cohort; this was confirmed in the validation cohort. Plasma succinate concentrations were higher in patients with AAD compared with those in healthy controls, patients with acute myocardial infarction (AMI), and patients with pulmonary embolism (PE). Moreover, succinate administration aggravated angiotensin II-induced AAD and vascular inflammation in mice. In contrast, knockdown of OGDH reduced the expression of inflammatory factors in macrophages. The conditional deletion of p38α decreased CREB phosphorylation, OGDH expression, and succinate concentrations. Conditional deletion of p38α in macrophages reduced angiotensin II-induced AAD. Plasma succinate concentrations allow to distinguish patients with AAD from both healthy controls and patients with AMI or PE. Succinate concentrations are regulated by the p38α-CREB-OGDH axis in macrophages.

Perturbed Brain Glucose Metabolism Caused by Absent SIRT3 Activity.

Acetylation is a post-translational modification that regulates the activity of enzymes fundamentally involved in cellular and mitochondrial bioenergetic metabolism. NAD+ dependent deacetylase sirtuin 3 (SIRT3) is localized to mitochondria where it plays a key role in regulating acetylation of TCA cycle enzymes and the mitochondrial respiratory complexes. Although the SIRT3 target proteins in mitochondria have been identified, the effect of SIRT3 activity on mitochondrial glucose metabolism in the brain remains elusive. The impact of abolished SIRT3 activity on glucose metabolism was determined in SIRT3 knockout (KO) and wild type (WT) mice injected with [1,6-13C]glucose using ex vivo 13C-NMR spectroscopy. The 1H-NMR spectra and amino acid analysis showed no differences in the concentration of lactate, glutamate, alanine, succinate, or aspartate between SIRT3 KO and WT mice. However, glutamine, total creatine (Cr), and GABA were lower in SIRT3 KO brain. Incorporation of label from [1,6-13C]glucose metabolism into lactate or alanine was not affected in SIRT3 KO brain. However, the incorporation of the label into all isotopomers of glutamate, glutamine, GABA and aspartate was lower in SIRT3 KO brain, reflecting decreased activity of mitochondrial and TCA cycle metabolism in both neurons and astrocytes. This is most likely due to hyperacetylation of mitochondrial enzymes due to suppressed SIRT3 activity in the brain of SIRT3 KO mice. Thus, the absence of Sirt3 results in impaired mitochondrial oxidative energy metabolism and neurotransmitter synthesis in the brain. Since the SIRT3 activity is NAD+ dependent, these results might parallel changes in glucose metabolism under pathologic reduction in mitochondrial NAD+ pools.

Recombinant cyanobacteria cultured in CO2 and seawater as feedstock for coproduction of acetoin and succinate by engineered Enterobacter cloacae

In this work, engineered freshwater cyanobacteria (Synechococcus elongatus PCC7942-hspA) were cultivated with CO2 and seawater to harvest biomass, optimize biomass hydrolysis conditions to increase the sugar concentration in the hydrolysate and use it as a feedstock for the coproduction of acetoin and succinate. Since the hydrolysate contains glucose and xylose, to improve the utilization efficiency of the strain mixed sugar, Enterobacter cloacae (strain CL80) in which the budC, ldhA, and ptsG genes were knocked out was constructed. When using cyanobacteria biomass hydrolysate as the fermentation feedstock of CL80, the concentrations of acetoin and succinate were 10.55 and 1.8 g L−1, respectively. This work is the first attempt to use freshwater cyanobacteria biomass hydrolysate obtained by culture with CO2 and seawater as the feedstock for the coproduction of acetoin and succinate, and it will provide a practical strategy for reducing carbon dioxide emissions and producing biobased value-added products by the biorefinery method.