Branched Amino Acids Research Articles

The evolution of amino acids under asteroidal aqueous alteration

Carbonaceous chondrites contain amino acids, with variable abundances and isotope compositions between and within carbonaceous chondrites. The parent body processes, and the presence of clay minerals may explain those differences. Here, we experimentally investigate the evolution of 6 amino acids (glycine, β-alanine, α-alanine, 2-aminoisobutyric acid, γ-aminobutyric acid, and isovaline) exposed to hydrothermal conditions in the presence or absence of silicates. We determined the chemical nature and isotopic composition of the organic compounds of the soluble and solid fractions of the residues using X-ray diffraction, spectroscopy, and mass-spectrometry methods. Glycine and α-alanine exhibit a rather high stability, which is consistent with the measured abundances of α-alanine and glycine in chondrites having experienced various degrees of aqueous alteration. In the meantime, the evolution of β-alanine under hydrothermal conditions leads to the formation of a new compound, which likely results from the decarboxylation and deamination of β-alanine followed by recombination. More than 95 % of γ-ABA was transformed into 2-pyrrolidione though self-cyclization during the aqueous alteration. The solid residues of the experiments conducted in the presence of clay minerals contain organic material, with abundances varying depending on the amino acid used for the experiments (TOC isovaline > 2-aminoisobutyric acid > γ-aminobutyric acid > glycine > α-alanine > β-alanine). Clay minerals thus preferentially trap branched amino acids over chained amino acids, likely within their interlayer spaces as suggested by XRD data. The δ13C values of amino acids have not changed significantly during the experiments, even with the presence of silicates. Thus, the δ13C values of amino acids reported in CR and CM chondrites likely relate to synthetic conditions or the origin of their precursors (i.e. inherited from the pre-accretion processes).

Continuous pulsed electric field processing for intensification of aqueous extraction of protein from fresh green seaweed Ulva sp. biomass

The green marine macroalgae Ulva sp. can reach a high protein content and a high composition of essential amino acids. In this study, we have developed a novel device and investigated the parameters for a continuous Pulsed Electric Field (PEF) process, coupled with enzyme treatment followed by a spray drying, to facilitate the aqueous fractionation of Ulva protein. The process demonstrated 8.79 ± 0.58 (% w/w) protein extraction yield. Furthermore, the aqueous fractionation of protein exhibited a substantial 41.45% essential amino acid content and a branched amino acids composition of 17.58%. Additionally, it displayed an in vitro relative digestibility of 87.4 ± 1.36% compared to soy protein, along with a water holding capacity of 7.15 ± 0.17 g water/g sample and an oil holding capacity of 1.76 ± 0.11 g oil/g sample. These findings suggest that employing a continuous PEF process in conjunction with enzyme-induced cell wall degradation could position green marine macroalgae as an important potential source for food protein ingredients.

Systematic sequence characterization of enzymatic-derived soybean peptides for precision enhancement of anti-inflammatory properties

Food-derived soybean peptides possess inflammation-preventive effects beyond basic nutrition due to their structural configuration. However, the relationship between sequence characteristics and their anti-inflammatory properties remains poorly understood. The lack of targeted peptide production methods limits their use in functional foods. Hydrophobic and basic amino acids enhance peptide anti-inflammatory activity. Alcalase and Papain facilitate hydrophobic and basic amino acid preparation, while Bromelain lacks specific cleavage sites. Endopeptidases selectively break peptide bonds within proteins. We subsequently used Flavourzyme, lacking specific cleavage sites, to sequentially shorten peptide chains from their terminal ends. This sequential use of endo- and exopeptidases enables efficient peptide preparation. However, the proteases' ability to selectively produce peptides with specific sequence characteristics and their contribution to the soybean peptide mixture remain unknown. This study examined three soybean peptide groups prepared using distinct methods (Alcalase + Flavourzyme: hydrophobic amino acid-rich peptides, Papain + Flavourzyme: basic amino acid-rich peptides, and Bromelain + Flavourzyme: balanced amino acid compositions) with different sequence characterizations linked to endonuclease cleavage sites. All groups inhibited nitric oxide (NO) and inducible nitric oxide synthase (iNOS) secretion, suppressing protein phosphorylation in the Phosphatidylinositol 3-kinase (PI3K)/Protein kinase B (Akt)/Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) (PI3K/Akt/NF-κB) signaling pathway. PF peptides showed the strongest anti-inflammatory effect, targeting iNOS, inhibiting TLR4, and reducing Akt and IκB phosphorylation. Mass spectrometry revealed correlations between regulatory mechanisms and the abundance of branched amino acids at PF peptides' N-terminals and significant Arg presence at their C-terminals. This study validates optimal enzymatic processes, introduces sequence characterization analysis, and identifies key features influencing anti-inflammatory properties, providing a theoretical basis for soybean peptide applications in inflammation-preventing functional foods.

Innovative Biomarkers for Obesity and Type 1 Diabetes Based on Bifidobacterium and Metabolomic Profiling.

The role of Bifidobacterium species and microbial metabolites such as short-chain fatty acids (SCFAs) and human milk oligosaccharides in controlling intestinal inflammation and the pathogenesis of obesity and type 1 diabetes (T1D) has been largely studied in recent years. This paper discusses the discovery of signature biomarkers for obesity and T1D based on data from a novel test for profiling several Bifidobacterium species, combined with metabolomic analysis. Through the NUTRISHIELD clinical study, a total of 98 children were recruited: 40 healthy controls, 40 type 1 diabetics, and 18 obese children. Bifidobacterium profiles were assessed in stool samples through an innovative test allowing high taxonomic resolution and precise quantification, while SCFAs and branched amino acids were measured in urine samples through gas chromatography-mass spectrometry (GC-MS). KIDMED questionnaires were used to evaluate the children's dietary habits and correlate them with the Bifidobacterium and metabolomic profiles. We found that B. longum subs. infantis and B. breve were higher in individuals with obesity, while B. bifidum and B. longum subs. longum were lower compared to healthy individuals. In individuals with T1D, alterations were found at the metabolic level, with an overall increase in the level of the most measured metabolites. The high taxonomic resolution of the Bifidobacterium test used meant strong correlations between the concentrations of valine and isoleucine, and the relative abundance of some Bifidobacterium species such as B. longum subs. infantis, B. breve, and B. bifidum could be observed.

Polyhydroxyalkanoate Copolymer Production by Recombinant Ralstonia eutropha Strain 1F2 from Fructose or Carbon Dioxide as Sole Carbon Source.

Ralstonia eutropha strain H16 is a chemoautotrophic bacterium that oxidizes hydrogen and accumulates poly[(R)-3-hydroxybutyrate] [P(3HB)], a prominent polyhydroxyalkanoate (PHA), within its cell. R. eutropha utilizes fructose or CO2 as its sole carbon source for this process. A PHA-negative mutant of strain H16, known as R. eutropha strain PHB-4, cannot produce PHA. Strain 1F2, derived from strain PHB-4, is a leucine analog-resistant mutant. Remarkably, the recombinant 1F2 strain exhibits the capacity to synthesize 3HB-based PHA copolymers containing 3-hydroxyvalerate (3HV) and 3-hydroxy-4-methyvalerate (3H4MV) comonomer units from fructose or CO2. This ability is conferred by the expression of a broad substrate-specific PHA synthase and tolerance to feedback inhibition of branched amino acids. However, the total amount of comonomer units incorporated into PHA was up to around 5 mol%. In this study, strain 1F2 underwent genetic engineering to augment the comonomer supply incorporated into PHA. This enhancement involved several modifications, including the additional expression of the broad substrate-specific 3-ketothiolase gene (bktB), the heterologous expression of the 2-ketoacid decarboxylase gene (kivd), and the phenylacetaldehyde dehydrogenase gene (padA). Furthermore, the genome of strain 1F2 was altered through the deletion of the 3-hydroxyacyl-CoA dehydrogenase gene (hbdH). The introduction of bktB-kivd-padA resulted in increased 3HV incorporation, reaching 13.9 mol% from fructose and 6.4 mol% from CO2. Additionally, the hbdH deletion resulted in the production of PHA copolymers containing (S)-3-hydroxy-2-methylpropionate (3H2MP). Interestingly, hbdH deletion increased the weight-average molecular weight of the PHA to over 3.0 × 106 on fructose. Thus, it demonstrates the positive effects of hbdH deletion on the copolymer composition and molecular weight of PHA.

Adaptive responses of erythritol-producing Yarrowia lipolytica to thermal stress after evolution

Elucidation of the thermotolerance mechanism of erythritol-producing Yarrowia lipolytica is of great significance to breed robust industrial strains and reduce cost. This study aimed to breed thermotolerant Y. lipolytica and investigate the mechanism underlying the thermotolerant phenotype. Yarrowia lipolytica HT34, Yarrowia lipolytica HT36, and Yarrowia lipolytica HT385 that were capable of growing at 34 °C, 36 °C, and 38.5 °C, respectively, were obtained within 150 days (352 generations) by adaptive laboratory evolution (ALE) integrated with 60Co-γ radiation and ultraviolet ray radiation. Comparative genomics analysis showed that genes involved in signal transduction, transcription, and translation regulation were mutated during adaptive evolution. Further, we demonstrated that thermal stress increased the expression of genes related to DNA replication and repair, ceramide and steroid synthesis, and the degradation of branched amino acid (BCAA) and free fatty acid (FFA), while inhibiting the expression of genes involved in glycolysis and the citrate cycle. Erythritol production in thermotolerant strains was remarkably inhibited, which might result from the differential expression of genes involved in erythritol metabolism. Exogenous addition of BCAA and soybean oil promoted the growth of HT385, highlighting the importance of BCAA and FFA in thermal stress response. Additionally, overexpression of 11 out of the 18 upregulated genes individually enabled Yarrowia lipolytica CA20 to grow at 34 °C, of which genes A000121, A003183, and A005690 had a better effect. Collectively, this study provides novel insights into the adaptation mechanism of Y. lipolytica to thermal stress, which will be conducive to the construction of thermotolerant erythritol-producing strains.Key points• ALE combined with mutagenesis is efficient for breeding thermotolerant Y. lipolytica• Genes encoding global regulators are mutated during thermal adaptive evolution• Ceramide and BCAA are critical molecules for cells to tolerate thermal stress

Daily fluctuation of colonic microbiome in response to nutrient substrates in a pig model

Studies on rodents indicate the daily oscillations of the gut microbiota have biological implications for host. However, the responses of fluctuating gut microbes to the dynamic nutrient substrates are not fully clear. In the study, we found that the feed intake, nutrient substrates, microbiota and metabolites in the colon underwent asynchronous oscillation within a day. Short-chain fatty acids (SCFAs) including acetate, propionate, butyrate and valerate peaked during T24 ~ T27 (Timepoint 24, 12:00 pm, T27, 03:00 am) whereas branched SCFAs isobutyrate and isovalerate peaked during T09 ~ T12. Further extended local similarity analysis (eLSA) revealed that the fluctuation of feed intake dynamically correlated with the colonic carbon substrates which further influenced the oscillation of sugar metabolites and acetate, propionate, butyrate and valerate with a certain time shift. The relative abundance of primary degrader Ruminococcaceae taxa was highly related to the dynamics of the carbon substrates whereas the fluctuations of secondary degraders Lactobacillaceae and Streptococcaceae taxa were highly correlated with the sugar metabolites. Meanwhile, colonic nitrogen substrates were correlated with branched amino acids and the branched SCFAs. Furthermore, we validated the evolution of gut microbes under different carbohydrate and protein combinations by using an in vitro fermentation experiment. The study pictured the dynamics of the micro-ecological environment within a day which highlights the implications of the temporal dimension in studies related to the gut microbiota. Feed intake, more precisely substrate intake, is highly correlated with microbial evolution, which makes it possible to develop chronotherapies targeting the gut microbiota through nutrition intervention.

Whole-Blood Metabolomics of a Rat Model of Repetitive Concussion.

Mild traumatic brain injury (mTBI) and repetitive mTBI (RmTBI) are silent epidemics, and so far, there is no objective diagnosis. The severity of the injury is solely based on the Glasgow Coma Score (GCS) scale. Most patients suffer from one or more behavioral abnormalities, such as headache, amnesia, cognitive decline, disturbed sleep pattern, anxiety, depression, and vision abnormalities. Additionally, most neuroimaging modalities are insensitive to capture structural and functional alterations in the brain, leading to inefficient patient management. Metabolomics is one of the established omics technologies to identify metabolic alterations, mostly in biofluids. NMR-based metabolomics provides quantitative metabolic information with non-destructive and minimal sample preparation. We employed whole-blood NMR analysis to identify metabolic markers using a high-field NMR spectrometer (800MHz). Our approach involves chemical-free sample pretreatment and minimal sample preparation to obtain a robust whole-blood metabolic profile from a rat model of concussion. A single head injury was given to the mTBI group, and three head injuries to the RmTBI group. We found significant alterations in blood metabolites in both mTBI and RmTBI groups compared with the control, such as alanine, branched amino acid (BAA), adenosine diphosphate/adenosine try phosphate (ADP/ATP), creatine, glucose, pyruvate, and glycerphosphocholine (GPC). Choline was significantly altered only in the mTBI group and formate in the RmTBI group compared with the control. These metabolites corroborate previous findings in clinical and preclinical cohorts. Comprehensive whole-blood metabolomics can provide a robust metabolic marker for more accurate diagnosis and treatment intervention for a disease population.

Serum branched amino acids and the risk of all-cause mortality: a meta-analysis and systematic review.

Recently, the serum levels of branched-chain amino acids (BCAAs) have been considered as an indicator to evaluate health status and predict chronic diseases risk. This systematic review and meta-analysis aimed to assess the relationship between Serum BCAAs and the risk of all-cause mortality. We carried out a comprehensive and systematic search in various important databases, including PubMed, Scopus, and Web of Science databases to find the relevant studies published up to October 2022 with no language, design, or time limitation. We extracted the reported hazard ratio (HR) with 95% confidence interval (CI) and odds ratio (OR) with 95%CI in cohorts and case-control studies, respectively, and computed the log HR or OR and its standard error. Then, we used the random-effects model with inverse variance weighting method for the present meta-analysis, to calculate the pooled effect size. Ten observational studies, including nine cohort studies and one case-control study, were included in the present meta-analysis. The number of participants ranges from 53 to 26,711, with an age range of 18-99years. During 6months to 24years of follow-up, 3599 deaths were ascertained. The pooled results indicated that there was no significant association between serum BCAAs (RR: 1.17; 95% CI 0.85-1.60), isoleucine (RR: 1.41; 95%CI 0.92-2.17), leucine (RR: 1.13; 95% CI 0.94-1.36), and valine (RR: 1.02; 95%CI 0.86-1.22) and all-cause mortality. Also, there was significant heterogeneity between studies for serum BCAAs (I2 = 74.1% and P-heterogeneity = 0.021), isoleucine (I2 = 89.4% and P-heterogeneity < 0.001), leucine (I2 = 87.8% and P-heterogeneity < 0.001), and valine (I2 = 86.6% and P-heterogeneity < 0.001). Our results suggested that the serum BCAAs and its components, including isoleucine, leucine, and valine, were not associated with the risk of all-cause mortality.

Branched Chain Amino Acid Content and Antioxidant Activity of Mung Bean Tempeh Powder for Developing Oral Nutrition Supplements.

Mung bean (Vigna radiata), a non-soybean legume, is known as one of the vegetable protein sources with 27% protein content. Mung bean also has a high content of essential amino acids, including branched chain amino acids (BCAAs). The use of mung bean for tempeh production presumably increases its nutritional value and functional efficacy, most significantly in protein and BCAA content. This tempeh is further applied for developing modern functional foods such as oral nutrition supplements (ONS). ONS can be used as a substitute for food and emergency food due to its complete nutritional content, as well as to treat malnourished patients. This study was aimed to produce mung bean tempeh powder, to formulate a mung bean tempeh one shot ONS high in BCAA content, and to determine its proximate analysis, antioxidant activity, and sensory characterization. Mung bean tempeh powder was successfully obtained with a yield of 37.50%, protein 39.19%, total amino acids 286.21 mg/g, essential amino acids 117.97 mg/g, and BCAAs 54.14 mg/g. There were 6 ONS formulas that were made with the combination of mung bean tempeh powder, palm sugar or honey, olive oil, and addition of an emulsifier. The selected formulas (F1 and F4) as well as commercial mung bean juice were sensory analyzed by applying an appropriate hedonic test. The results showed that the panelists both liked ONS F1 and F4 (p > 0.05). In addition, both ONS F1 and F4 at 5% demonstrated a significant antioxidant capacity, 92.79% and 82.57% of ascorbic acid, respectively. These data suggest that mung bean tempeh containing high branched amino acids could be recommended as a functional ingredient that gives health promotion for ONS development.

Oridonin impedes breast cancer growth by blocking cells in S phase and inhibiting the PI3K/AKT/mTOR signaling pathway

Breast cancer is one of the most common cancers. Oridonin, a traditional Chinese medicine, is believed to inhibit tumor growth, but its particular effects on breast cancer remain unknown. In this study, we examined oridonin's effects on 4T1, MCF-7, and MDAMB-231 cellular activity using CCK8. Scratch assays were used to detect oridonin's effects on cellular migration. Oridonin's effects on the breast cancer cell cycle were studied using flow cytometry, and expression of cell cycle related proteins p53, CDK2, and p21 was detected using Western blot assays. Metabolomics assays were used to detect changes in small molecule metabolites and metabolic pathways in breast cancer cells after treatment with oridonin. Oridonin's effects on breast cancer growth were also studied using xenograft mice. Metabolomics assays were used to detect changes in metabolites and metabolic pathways in xenograft mouse plasma in a control group, model group, and drug administration group. Experimental results showed that oridonin could significantly inhibit breast cancer growth both in vivo and in vitro. Scratch experiments showed that oridonin could inhibit breast cancer cell migration. Oridonin was also able to arrest cells in S phase by affecting several cell cycle-related proteins, including p53, CDK2, and p21. Metabolomic analysis of 4T1 cells identified a total of 33 differential metabolites, including multiple amino acids (such as l-Glutamic acid, l-Asparagine, l-Histidine, l-Valine, and l-Isoleucine). KEGG pathway enrichment analysis showed significant changes in aminoacyl-tRNA biosynthesis, and in multiple amino acid metabolic pathways. Plasma metabolomic analyses of xenograft mice revealed 28 differentially-expressed metabolites between the different animal model groups, including multiple amino acids. KEGG pathway analysis showed significant alterations in multiple amino acid metabolic pathways in oridonin-treated mice. Additionally, changes in the expression of PI3K, AKT and mTOR proteins, as well as in branched amino acids, suggest that oridonin affects the PI3K/AKT/mTOR signaling pathway by inhibiting the biosynthesis of valine, leucine and isoleucine. Taken together, our results suggest that oridonin has strong anti-tumor activity in vitro and in vivo, and has potential as an adjuvant to breast cancer treatment regimens.

Proteins and protein components for sportspeople: quality control of dietary supplements

Quality control of protein supplements intended for a large audience of consumers such as sportspeople is particularly important. A case study on quality control of dietary supplements containing protein and protein components is presented. The objective of the study was to evaluate the conformity of the quantities of amino acids, essential and branched-chain amino acids, declared on the label through measurements with chromatographic analytical tools. 16 sportspeople supplements from different European countries were tested. Analysis of concentrated whey protein highlighted some differences between the label and what was experimentally determined; in these samples some amino acids (6 amino acids out of 19) exceeded the maximum tolerance (>20%) regulated by the European Commission. To a lesser extent, analysis of the other classes revealed amino acid concentrations that exceeded the maximum analytical tolerance percentage. As regards the essential and branched amino acid supplements, it was seen that the declared quantity conforms with that determined experimentally.

Integrative transcriptome and proteome revealed high-yielding mechanisms of epsilon-poly-L-lysine by Streptomyces albulus.

ε-poly-L-lysine (ε-PL) is a high value, widely used natural antimicrobial peptide additive for foods and cosmetic products that is mainly produced by Streptomyces albulus. In previous work, we developed the high-yield industrial strain S. albulus WG-608 through successive rounds of engineering. Here, we use integrated physiological, transcriptomic, and proteomics association analysis to resolve the complex mechanisms underlying high ε-PL production by comparing WG-608 with the progenitor strain M-Z18. Our results show that key genes in the glycolysis, pentose phosphate pathway, glyoxylate pathway, oxidative phosphorylation, and L-lysine biosynthesis pathways are differentially upregulated in WG-608, while genes in the biosynthetic pathways for fatty acids, various branched amino acids, and secondary metabolite by-products are downregulated. This regulatory pattern results in the introduction of more carbon atoms into L-lysine biosynthesis and ε-PL production. In addition, significant changes in the regulation of DNA replication, transcription, and translation, two component systems, and quorum sensing may facilitate the adaptability to environmental pressure and the biosynthesis of ε-PL. Overexpression of ppk gene and addition of polyP6 further enhanced the ε-PL production. This study enables comprehensive understanding of the biosynthetic mechanisms of ε-PL in S. albulus WG-608, while providing some genetic modification and fermentation strategies to further improve the ε-PL production.

Hepatic triglyceride content is intricately associated with numerous metabolites and biochemical pathways.

Non-alcoholic fatty liver disease (NAFLD) is characterized by the pathological accumulation of triglycerides in hepatocytes and is associated with insulin resistance, atherogenic dyslipidaemia and cardiometabolic diseases. Thus far, the extent of metabolic dysregulation associated with hepatic triglyceride accumulation has not been fully addressed. In this study, we aimed to identify metabolites associated with hepatic triglyceride content (HTGC) and map these associations using network analysis. To gain insight in the spectrum of metabolites associated with hepatic triglyceride accumulation, we performed a comprehensive plasma metabolomics screening of 1363 metabolites in apparently healthy middle aged (age 45-65) individuals (N = 496) in whom HTGC was measured by proton magnetic resonance spectroscopy. An atlas of metabolite-HTGC associations, based on univariate results, was created using correlation-based Gaussian graphical model (GGM) and genome scale metabolic model network analyses. Pathways associated with the clinical prognosis marker fibrosis 4 (FIB-4) index were tested using a closed global test. Our analyses revealed that 118 metabolites were univariately associated with HTGC (p-value <6.59 × 10-5 ), including 106 endogenous, 1 xenobiotic and 11 partially characterized/uncharacterized metabolites. These associations were mapped to several biological pathways including branched amino acids (BCAA), diglycerols, sphingomyelin, glucosyl-ceramide and lactosyl-ceramide. We also identified a novel possible HTGC-related pathway connecting glutamate, metabolonic lactone sulphate and X-15245 using the GGM network. These pathways were confirmed to be associated with the FIB-4 index as well. The full interactive metabolite-HTGC atlas is provided online: https://tofaquih.github.io/AtlasLiver/. The combined network and pathway analyses indicated extensive associations between BCAA and the lipids pathways with HTGC and the FIB-4 index. Moreover, we report a novel pathway glutamate-metabolonic lactone sulphate-X-15245 with a potential strong association with HTGC. These findings can aid elucidating HTGC metabolomic profiles and provide insight into novel drug targets for fibrosis-related outcomes.

Glucose supplementation improves intestinal amino acid transport and muscle amino acid pool in pigs during chronic cold exposure.

Mammals in northern regions chronically suffer from low temperatures during autumn-winter seasons. The aim of this study was to investigate the response of intestinal amino acid transport and the amino acid pool in muscle to chronic cold exposure via Min pig models (cold adaptation) and Yorkshire pig models (non-cold adaptation). Furthermore, this study explored the beneficial effects of glucose supplementation on small intestinal amino acid transport and amino acid pool in muscle of cold-exposed Yorkshire pigs. Min pigs (Exp. 1) and Yorkshire pigs (Exp. 2) were divided into a control group (17°C, n=6) and chronic cold exposure group (7°C, n=6), respectively. Twelve Yorkshire pigs (Exp. 3) were divided into a cold control group and cold glucose supplementation group (8°C). The results showed that chronic cold exposure inhibited peptide transporter protein 1 (PepT1) and excitatory amino acid transporter 3 (EAAT3) expression in ileal mucosa and cationic amino acid transporter-1 (CAT-1) in the jejunal mucosa of Yorkshire pigs (P<0.05). In contrast, CAT-1, PepT1 and EAAT3 expression was enhanced in the duodenal mucosa of Min pigs (P<0.05). Branched amino acids (BCAA) in the muscle of Yorkshire pigs were consumed by chronic cold exposure, accompanied by increased muscle RING-finger protein-1 (MuRF1) and muscle atrophy F-box (atrogin-1) expression (P<0.05). More importantly, reduced concentrations of dystrophin were detected in the muscle of Yorkshire pigs (P<0.05). However, glycine concentration in the muscle of Min pigs was raised (P<0.05). In the absence of interaction between chronic cold exposure and glucose supplementation, glucose supplementation improved CAT-1 expression in the jejunal mucosa and PepT1 expression in the ileal mucosa of cold-exposed Yorkshire pigs (P<0.05). It also improved BCAA and inhibited MuRF1 and atrogin-1 expression in muscle (P<0.05). Moreover, dystrophin concentration was improved by glucose supplementation (P<0.05). In summary, chronic cold exposure inhibits amino acid absorption in the small intestine, depletes BCAA and promotes protein degradation in muscle. Glucose supplementation ameliorates the negative effects of chronic cold exposure on amino acid transport and the amino acid pool in muscle.

Targeted metabolomics detects a putatively diagnostic signature in plasma and dried blood spots from head and neck paraganglioma patients

Head and neck paragangliomas (HNPGLs), rare chemoresistant tumors curable only with surgery, are strongly influenced by genetic predisposition, hence patients and relatives require lifetime follow-up with MRI and/or PET-CT because of de novo disease risk. This entails exposure to electromagnetic/ionizing radiation, costs, and organizational challenges, because patients and relatives are scattered far from reference centers. Simplified first-line screening strategies are needed. We employed flow injection analysis tandem mass spectrometry, as used in newborn metabolic screening, to compare the plasma metabolic profile of HNPGL patients (59 samples, 56 cases) and healthy controls (24 samples, 24 cases). Principal Component Analysis (PCA) and Partial Least Discriminant Analysis (PLS-DA) highlighted a distinctive HNPGL signature, likely reflecting the anaplerotic conversion of the TCA cycle to glutaminolysis and catabolism of branched amino acids, DNA damage and deoxyadenosine (dAdo) accumulation, impairment of fatty acid oxidation, switch towards the Warburg effect and proinflammatory lysophosphatidylcholines (LPCs) signaling. Statistical analysis of the metabolites that most impacted on PLS-DA was extended to 10 acoustic neuroma and 2 cholesteatoma patients, confirming significant differences relative to the HNPGL plasma metabolomic profile. The best confusion matrix from the ROC curve built on 2 metabolites, dAdo and C26:0-LPC, provided specificity of 94.29% and sensitivity of 89.29%, with positive and negative predictive values of 96.2% and 84.6%, respectively. Analysis of dAdo and C26:0-LPC levels in dried venous and capillary blood confirmed that dAdo, likely deriving from 2′-deoxy-ATP accumulated in HNPGL cells following endogenous genotoxic damage, efficiently discriminated HNPGL patients from healthy controls and acoustic neuroma/cholesteatoma patients on easily manageable dried blood spots.

Metabolic and proteomic indications of diabetes progression in human aqueous humor.

Diabetes mellitus is a multiorgan systemic disease impacting numerous ocular structures that results in significant ocular morbidity and often results in more frequent corneal and glaucoma surgeries for affected individuals. We hypothesize that the systemic metabolic and proteomic derangement observed in the progression of diabetes influences the composition of the aqueous humor (AH), which ultimately impacts the anterior segment health of the eye. To identify changes associated with diabetes progression, we mapped the metabolite profile and proteome of AH samples from patients with varying severities of type II diabetes (T2DM). Patients were classified as nondiabetic (ND or control), non-insulin-dependent diabetic without advanced features of disease (NAD-ni), insulin-dependent diabetic without advanced features (NAD-i), or diabetic with advanced features (AD). AH samples collected from the anterior chamber during elective ophthalmic surgery were evaluated for metabolite and protein expression changes associated with diabetic severity via gas chromatography/mass spectrometry and ultra-high performance liquid chromatography tandem mass spectrometry, respectively. Metabolic and proteomic pathway analyses were conducted utilizing MetaboAnalyst 4.0 and Ingenuity Pathway Analysis. A total of 14 control, 12 NAD-ni, 4 NAD-I, and 14 AD samples were included for analysis. Elevated levels of several branched amino acids (e.g., valine, leucine, isoleucine), and lipid metabolites (e.g., palmitate) were found only with increasing diabetic severity (i.e., the AD group). Similar proteomic trends were noted in amino acid and fatty acid metabolism and the unfolded protein/stress response. These results represent the first report of both metabolomic and proteomic evaluation of aqueous humor. Diabetes results in metabolic and proteomic perturbations detectable in the AH, and unique changes become manifest as T2DM severity worsens. Changes in AH composition may serve as an indicator of disease severity, risk assessment of anterior segment cells and structures, and potential future therapies.

Next-generation sequencing of the athletic gut microbiota: a systematic review.

Aim: There is growing evidence that physical activity modulates gut microbiota composition through complex interactions between diet and microbial species. On the other hand, next-generation sequencing techniques include shotgun metagenomics and 16S amplicon sequencing. These methodologies allow a comprehensive characterisation of microbial communities of athletes from different disciplines as well as non-professional players and sedentary adults exposed to training. This systematic review summarises recent applications of next-generation sequencing to characterise the athletic gut microbiome. Methods: A systematic review of microbiome research was performed to determine the association of microbiota composition profiles with sports performance. Results: Bibliographic analysis revealed the importance of a novel research trend aiming at deciphering the associations between individual microbial species and sports performance. In addition, literature review highlighted the role of butyrate-producing bacteria such as Anaerostipes hadrus, Clostridium bolteae, Faecalibacterium prausnitzii, Roseburia hominis and unidentified species belonging to Clostridiales, Lachnospiraceae and Subdoligranulum species in gut health and sports performance across several disciplines. Interestingly, metabolic activities of Prevotella copri and Veillonella atypica involved in branched amino acid and lactate metabolism may contribute to reducing muscular fatigue. Other microbial metabolic pathways of interest involved in carbohydrate metabolism showed increased proportions in athletes´ metagenomes. Conclusion: Future research will aim at developing personalised nutrition interventions to modulate key species associated with certain components of exercise.

Profiling of lipids in Thermus thermophilus HB8 grown under various conditions.

The membrane lipids of Thermus species have unique structures. Only four polar lipid species have so far been identified in Thermus thermophilus HB8; namely, are two phosphoglycolipids and two glycolipids, both of which have three branched fatty acid chains. Other lipid molecules may be present; however, they have not been identified so far. To clarify the whole lipid profile of T. thermophilus HB8, we cultured this organism under four different growth (temperature and/or nutrition) conditions and analyzed the compositions of polar lipids and fatty acids by high-performance thin-layer chromatography (HPTLC) and gas chromatograph-mass spectrometry (GCｰMS), respectively. Thirty-one lipid spots were detected on HPTLC plates and profiled in terms of the presence or absence of phosphate, amino, and sugar groups. Then, we allocated ID numbers to all the spots. Comparative analyses of these polar lipids showed that the diversity of lipid molecules increased under high temperature and minimal medium conditions. In particular, aminolipid species increased under high temperature conditions. As for the fatty acid comparison by GC-MS, iso-branched even-numbered carbon atoms, which are unusual in this organism, significantly increased under the minimal medium condition, suggesting that kinds of branched amino acids at the fatty acid terminus varies under different nutrition conditions. In this study, several unidentified lipids were detected, and elucidation of the lipid structures will provide important information on the environmental adaptation of bacteria.

Analysis of Branched Amino Acids by UPLC as an Alternative Method for the Management of Patients with MUSD

Analysis of Branched Amino Acids by UPLC as an Alternative Method for the Management of Patients with MUSD