Thiamine Monophosphate Research Articles

Heteroatom(S) (N, S, P) Co‐Doped Reduced Graphene Oxide‐Based Binder‐Free Novel Energy Storage Electrode Material for High Energy Density Supercapacitor

AbstractThis manuscript reports a one‐pot synthesis of ternary heteroatoms (N/S/P) co‐doped reduced graphene oxide as an electrode material by simultaneous reduction/doping of/into graphene oxide employing eco‐friendly natural molecule(s) (thiamine monophosphate (TM) and thiamine pyrophosphate (TP)) under mild heating (≤60 °C) and near‐neutral pH (≤8) conditions. It produces N, S, P co‐doped rGO‐TM and N, S, P co‐doped rGO‐TP nanohybrids displaying a well‐connected porous microstructure with significantly doped atomic(%) ‐N (10.0%), S (4.1%), P (0.1%) and N (6.7%), S (2.0%), and P (0.5%), respectively. The symmetric supercapacitor designed using N, S, P‐rGO‐TM‐10 in water‐in‐salt 17 m NaClO4 electrolyte exhibits stable cell voltage of 3.0 V, superb gravimetric energy density (areal energy density) of 47.9 Wh kg−1@522.8 W g−1 (0.2363 Wh cm−2@ 2.5 W cm−2); the excellent cyclic stability (88.8 %) and Coulombic efficiency (99.2%) even after 20 000 cycles. Their energy storage capability is corroborated by the illumination of 109 white LEDs upon lighting a single SSC for 50 s each, driving a motor to run a fan, and forming the tandem device by joining three such cells in series.

Pre-analytical challenges from adsorptive losses associated with thiamine analysis

Thiamine (vitamin B1) is an essential vitamin serving in its diphosphate form as a cofactor for enzymes in the citric acid cycle and pentose-phosphate pathways. Its concentration reported in the pM and nM range in environmental and clinical analyses prompted our consideration of the components used in pre-analytical processing, including the selection of filters, filter apparatuses, and sample vials. The seemingly innocuous use of glass fiber filters, glass filter flasks, and glass vials, ubiquitous in laboratory analysis of clinical and environmental samples, led to marked thiamine losses. 19.3 nM thiamine was recovered from a 100 nM standard following storage in glass autosampler vials and only 1 nM of thiamine was obtained in the filtrate of a 100 nM thiamine stock passed through a borosilicate glass fiber filter. We further observed a significant shift towards phosphorylated derivatives of thiamine when an equimolar mixture of thiamine, thiamine monophosphate, and thiamine diphosphate was stored in glass (most notably non-silanized glass, where a reduction of 54% of the thiamine peak area was observed) versus polypropylene autosampler vials. The selective losses of thiamine could lead to errors in interpreting the distribution of phosphorylated species in samples. Further, some loss of phosphorylated thiamine derivatives selectively to amber glass vials was observed relative to other glass vials. Our results suggest the use of polymeric filters (including nylon and cellulose acetate) and storage container materials (including polycarbonate and polypropylene) for thiamine handling. Losses to cellulose nitrate and polyethersulfone filters were far less substantial than to glass fiber filters, but were still notable given the low concentrations expected in samples. Thiamine losses were negated when thiamine was stored diluted in trichloroacetic acid or as thiochrome formed in situ, both of which are common practices, but not ubiquitous, in thiamine sample preparation.

Ectopic expression of a bacterial thiamin monophosphate kinase enhances vitamin B1 biosynthesis in plants.

Plants and bacteria have distinct pathways to synthesize the bioactive vitamin B1 thiamin diphosphate (TDP). In plants, thiamin monophosphate (TMP) synthesized in the TDP biosynthetic pathway is first converted to thiamin by a phosphatase, which is then pyrophosphorylated to TDP. In contrast, bacteria use a TMP kinase encoded by ThiL to phosphorylate TMP to TDP directly. The Arabidopsis THIAMIN REQUIRING2 (TH2)-encoded phosphatase is involved in TDP biosynthesis. The chlorotic th2 mutants have high TMP and low thiamin and TDP. Ectopic expression of Escherichia coli ThiL and ThiL-GFP rescued the th2-3 mutant, suggesting that the bacterial TMP kinase could directly convert TMP into TDP in Arabidopsis. These results provide direct evidence that the chlorotic phenotype of th2-3 is caused by TDP rather than thiamin deficiency. Transgenic Arabidopsis harboring engineered ThiL-GFP targeting to the cytosol, chloroplast, mitochondrion, or nucleus accumulated higher TDP than the wild type (WT). Ectopic expression of E. coli ThiL driven by the UBIQUITIN (UBI) promoter or an endosperm-specific GLUTELIN1 (GT1) promoter also enhanced TDP biosynthesis in rice. The pUBI:ThiL transgenic rice accumulated more TDP and total vitamin B1 in the leaves, and the pGT1:ThiL transgenic lines had higher TDP and total vitamin B1 in the seeds than the WT. Total vitamin B1 only increased by approximately 25-30% in the polished and unpolished seeds of the pGT1:ThiL transgenic rice compared to the WT. Nevertheless, these results suggest that genetic engineering of a bacterial vitamin B1 biosynthetic gene downstream of TMP can enhance vitamin B1 production in rice.

Metabolomics analyses of cancer tissue from patients with colorectal cancer.

The alteration of metabolism is essential for the initiation and progression of numerous types of cancer, including colorectal cancer (CRC). Metabolomics has been used to study CRC. At present, the reprogramming of the metabolism in CRC remains to be fully elucidated. In the present study, comprehensive untargeted metabolomics analysis was performed on the paired CRC tissues and adjacent normal tissues from patients with CRC (n=35) using ultra‑high‑performance liquid chromatography‑mass spectrometry. Subsequently, bioinformatic analysis was performed on the differentially expressed metabolites. The changes in these differential metabolites were compared among groups of patients based on sex, anatomical tumor location, grade of tumor differentiation and stage of disease. A total of 927 metabolites were detected in the tissue samples, and 24 metabolites in the CRC tissue were significantly different compared with the adjacent normal tissue. The present study revealed that the levels of three amino acid metabolites were increased in the CRC tissue, specifically, N‑α‑acetyl‑ε‑(2‑propenal)‑Lys, cyclo(Glu‑Glu) and cyclo(Phe‑Glu). The metabolites with decreased levels in the CRC tissue included quinaldic acid (also referred to as quinoline‑2‑carboxilic acid), 17α‑ and 17β‑estradiol, which are associated with tumor suppression activities, as well as other metabolites such as, anhydro‑β‑glucose, Asp‑Arg, lysophosphatidylcholine, lysophosphatidylethanolamine (lysoPE), lysophosphatidylinositol, carnitine, 5'‑deoxy‑5'‑(methylthio) adenosine, 2'‑deoxyinosine‑5'‑monophosphate and thiamine monophosphate. There was no difference in the levels of the differential metabolites between male and female patients. The differentiation of CRC also showed no impact on the levels of the differential metabolites. The levels of lysoPE were increased in the right side of the colon compared with the left side of the colon and rectum. Analysis of the different tumor stages indicated that 2‑aminobenzenesulfonic acid, P‑sulfanilic acid and quinoline‑4‑carboxylic acid were decreased in stageI CRC tissue compared with stageII,IIIandIV CRC tissue. The levels of N‑α‑acetyl‑ε‑(2‑propenal)‑Lys, methylcysteine and 5'‑deoxy‑5'‑(methylthio) adenosine varied at different stages of tumorigenesis. These differential metabolites were implicated in multiple metabolism pathways, including carbohydrate, amino acid, lipid, nucleotide and hormone. In conclusion, the present study demonstrated that CRC tumors had altered metabolites compared with normal tissue. The data from the metabolic profile of CRC tissues in the present study provided supportive evidence to understand tumorigenesis.

Exploratory analyses on the effect of time since last meal on concentrations of amino acids, lipids, one-carbon metabolites, and vitamins in the Hordaland Health Study

PurposeDietary intake may have pronounced effects on circulating biomarker concentrations. Therefore, the aim was to provide a descriptive overview of serum metabolite concentrations in relation to time since last meal, focusing on amino acids, lipids, one-carbon metabolites, and biomarkers of vitamin status.MethodsWe used baseline data from the observational community-based Hordaland Health Study, including 2960 participants aged 46–49 years and 2874 participants aged 70–74 years. A single blood draw was taken from each participant, and time since last meal varied. Estimated marginal geometric mean metabolite concentrations were plotted as a function of time since last meal, up to 7 h, adjusted for age, sex, and BMI.ResultsWe observed a common pattern for nearly all amino acids and one-carbon metabolites with highest concentrations during the first 3 h after dietary intake. Homocysteine and cysteine were lowest the 1st hour after a meal, while no patterns were observed for glutamate and glutamic acid. The concentrations of phylloquinone and triglycerides were highest 1 h after dietary intake. Thiamine and thiamine monophosphate concentrations were highest, while flavin mononucleotide concentrations were lowest within the first 2 h after a meal. No clear patterns emerged for the other fat-soluble vitamins, blood lipids, or B-vitamin biomarkers.ConclusionOur findings suggest that distinguishing between “fasting” and “non-fasting” blood samples may be inadequate, and a more granular approach is warranted. This may have implications for how to account for dietary intake when blood sampling in both clinical and research settings.

The Impacts of Slc19a3 Deletion and Intestinal SLC19A3 Insertion on Thiamine Distribution and Brain Metabolism in the Mouse.

The Thiamine Transporter 2 (THTR2) encoded by SLC19A3 plays an ill-defined role in the maintenance of tissue thiamine, thiamine monophosphate, and thiamine diphosphate (TDP) levels. To evaluate the impact of THTR2 on tissue thiamine status and metabolism, we expressed the human SLC19A3 transgene in the intestine of total body Slc19a3 knockout (KO) mice. Male and female wildtype (WT) and transgenic (TG) mice were fed either 17 mg/kg (1×) or 85 mg/kg (5×) thiamine hydrochloride diet, while KOs were only fed the 5× diet. Thiamine vitamers in plasma, red blood cells, duodenum, brain, liver, kidney, heart, and adipose tissue were measured. Untargeted metabolomics were performed on the brain tissues of groups with equivalent plasma thiamine. KO mice had ~two- and ~three-fold lower plasma and brain thiamine levels than WT on the 5× diet. Circulating vitamers were sensitive to diet and equivalent in TG and WT mice. However, TG had 60% lower thiamine but normal brain TDP levels regardless of diet, with subtle differences in the heart and liver. The loss of THTR2 reduced levels of nucleic acid and amino acid derivatives in the brain. Therefore, mutation or inhibition of THTR2 may alter the brain metabolome and reduce the thiamine reservoir for TDP biosynthesis.

1614-P: Metabolomic Profiling Reveals Thiamine-Mediated Regulation of Triglyceride Synthesis and Lipid Turnover in Brown Fat

Brown adipose tissue (BAT) functions as a metabolic sink to dispose glucose as well as fatty acids (FAs) and exerts profound effects on energy and glucose homeostasis. Intermittent fasting (IF) has become an attractive option for combating obesity and treating diabetes, yet how IF regimen impacts BAT metabolism and thermogenic capacity remains poorly understood. Using metabolomics and lipidomic assays, we show that alternate day fasting (ADF) elicited robust activation of upper glycolysis and increased production of glycerol-3-phosphate (G3P) and lactate accompanied with stimulation of glycolytic shunts, pentose phosphate pathway (PPP) and glycogenesis, in BAT but not in gonadal white adipose tissue (gWAT) in feeding state. As a result, feast BAT exhibited a notable increase in triglyceride (TG) synthesis, tissue volume, and thermogenic capacity compared to ad libitum group. Mechanistically, repeated fasting and refeeding suppressed the expression of thiamine transporter ThTr2 and decreased the levels of free thiamine, thiamine pyrophosphate (TPP) and thiamine monophosphate (TMP), in part through periodic activation of mTORC1. Genetic inhibition of mTORC1 in BAT restored expression of ThTr2 and diminished ADF-preserved lipid storage in vivo, and inactivating thiamine metabolism with specific inhibitors induced G3P production and TG synthesis in vitro. Taken together, our study reveals a new mechanism involving thiamine that reprograms BAT carbohydrate metabolism to preserve lipids and improves metabolic health. Disclosure C.Wang: None. X.Zhang: None. M.Liu: None.

Association between diabetes and thiamine status - A systematic review and meta-analysis

BackgroundThiamine (vitamin B1) is an essential cofactor in glucose metabolism, but it remains unclear whether thiamine status is lower in individuals with diabetes compared to individuals with normal glucose metabolism. AimsWe conducted a systematic review and meta-analysis to study whether the circulating concentrations of various thiamine analytes differ between people with and those without diabetes. MethodsPubMed and the Cochrane Central Register of Controlled Trials were searched according to the study protocol. The standardized mean difference (SMD) and 95 % confidence intervals (CI) of thiamine markers between individuals with and without diabetes were used as effect size (random effects model). Subgroup analysis considered albuminuria as an additional variable. ResultsOut of the 459 articles identified, 24 full-texts were eligible for the study, 20 of which qualified for the data analysis and four were evaluated for coherence. Compared to controls, individuals with diabetes showed lower concentrations of thiamine (pooled estimate SMD [95 % CI]: −0.97 [−1.89, −0.06]), thiamine monophosphate (−1.16 [−1.82, −0.50]), and total thiamine compounds (−1.01 [−1.48, −0.54]). Thiamine diphosphate (−0.72 [−1.54, 0.11] and erythrocyte transketolase activity (−0.42 [−0.90, 0.05]) tended to be lower in persons with diabetes than in controls without reaching statistical significance. Subgroup analysis showed that individuals with diabetes and albuminuria had lower thiamine levels than the controls (−2.68 [−5.34, −0.02]). ConclusionsDiabetes is associated with lower levels of various thiamine markers, suggesting that individuals with diabetes may have higher thiamine requirements than those without diabetes, but well-designed studies are required to confirm these findings.

Abstract 1010: Circulating biomarkers of vitamin B1, B2, and B3 status and their associations with clinical outcomes among non-metastatic colorectal cancer patients: results from the FOCUS Consortium

Abstract Introduction: Circulating levels of biomarkers indicating better B vitamin status appear to have a protective effect on various medical conditions, while they also contribute to the reduction in risk of some cancer entities, e.g. colorectal cancer (CRC). However, little is known about the associations of circulating vitamin B biomarkers with clinical outcomes among CRC patients, such as overall survival (OS), disease-free survival (DFS), and risk of recurrence. Recently, we showed that higher vitamin B6 status is associated with improved OS and DFS. In the present study, we further investigated associations of vitamin B1, B2, and B3 status with clinical outcomes in a prospective cohort of non-metastatic CRC patients. Materials and Methods: A total of 2,102 patients with stage I-III CRC participated in six cohorts in the international FOCUS Consortium. Circulating concentrations of vitamin B biomarkers (thiamine and thiamine monophosphate (representing vitamin B1), riboflavin (vitamin B2), and vitamin B3 markers N1-methylnicotinamide and nicotinamide were measured by liquid chromatography-tandem mass spectrometry in preoperative blood samples. Using Cox proportional hazards regression, we examined associations of vitamin B1, B2, B3 status, and related biomarkers with OS, DFS, and risk of recurrence. Models were adjusted for patient age, sex, circulating creatinine levels, tumor site, tumor stage, and study site. Results and Discussion: After a median follow-up of 3.2 years for OS, higher levels of thiamine monophosphate were associated with lower risk of all-cause mortality (HROS: 0.82; 95% CI, 0.71-0.95, p=0.008). Furthermore, higher riboflavin levels were associated with improved OS (HROS: 0.88; 95% CI, 0.78-0.98, p=0.03) and DFS (HRDFS: 0.91; 95% CI, 0.83-0.99, p=0.04). We did not observe any associations of thiamine, N1-methylnicotinamide, and nicotinamide with clinical outcomes among CRC patients. Conclusion: Findings from this multi-centre study of non-metastatic CRC patients yield important clinical information. We observed significantly improved OS among patients with higher levels of thiamine monophosphate after CRC diagnosis. Further, higher preoperative riboflavin may represent a promising predictor for OS and DFS among non-metastatic CRC patients. Citation Format: Biljana Gigic, Jennifer Ose, Stefanie Brezina, Anne JMR Geijsen, Arve Ulvik, Eline H. van Roekel, Andreana N. Holowatyj, Dieuwertje E. Kok, Fränzel JB van Duijnhoven, Victoria Damerell, Christopher I. Li, Nina Habermann, Alexis B. Ulrich, Matty P. Weijenberg, Andrea Gsur, Per M. Ueland, Ellen Kampman, Martin Schneider, Cornelia M. Ulrich. Circulating biomarkers of vitamin B1, B2, and B3 status and their associations with clinical outcomes among non-metastatic colorectal cancer patients: results from the FOCUS Consortium [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1010.

Maternal preconception circulating blood biomarker mixtures, child behavioural symptom scores and the potential mediating role of neonatal brain microstructure: the S-PRESTO cohort

Human brain development starts in the embryonic period. Maternal preconception nutrition and nutrient availability to the embryo may influence brain development at this critical period following conception and early cellular differentiation, thereby affecting offspring neurodevelopmental and behavioural disorder risk. However, studying this is challenging due to difficulties in characterizing preconception nutritional status and few studies have objective neurodevelopmental imaging measures in children. We investigated the associations of maternal preconception circulating blood nutrient-related biomarker mixtures (~15 weeks before conception) with child behavioural symptoms (Child Behaviour Checklist (CBCL), aged 3 years) within the Singapore Preconception Study of Long-Term Maternal and Child Outcomes (S-PRESTO) study. The CBCL preschool form evaluates child behaviours based on syndrome scales and Diagnostic and Statistical Manual of Mental Disorders (DSM) oriented scales. These scales consist of internalizing problems, externalizing problems, anxiety problems, pervasive developmental problems, oppositional defiant, etc. We applied data-driven clustering and a method for modelling mixtures (Bayesian kernel machine regression, BKMR) to account for complex, non-linear dependencies between 67 biomarkers. We used effect decomposition analyses to explore the potential mediating role of neonatal (week 1) brain microstructure, specifically orientation dispersion indices (ODI) of 49 cortical and subcortical grey matter regions. We found that higher levels of a nutrient cluster including thiamine, thiamine monophosphate (TMP), pyridoxal phosphate, pyridoxic acid, and pyridoxal were associated with a higher CBCL score for internalizing problems (posterior inclusion probability (PIP) = 0.768). Specifically, thiamine independently influenced CBCL (Conditional PIP = 0.775). Higher maternal preconception thiamine level was also associated with a lower right subthalamic nucleus ODI (P-value = 0.01) while a lower right subthalamic nucleus ODI was associated with higher CBCL scores for multiple domains (P-value < 0.05). One potential mechanism is the suboptimal metabolism of free thiamine to active vitamin B1, but additional follow-up and replication studies in other cohorts are needed.

Integrated metabolomics and machine learning approach to predict hypertensive disorders of pregnancy

Hypertensive disorders of pregnancy account for 3% to 10% of maternal-fetal morbidity and mortality worldwide. This condition has been considered one of the leading causes of maternal deaths in developing countries, such as India. This study aimed to discover hypertensive disorders of pregnancy-specific candidate urine metabolites as markers for hypertensive disorders of pregnancy by applying integrated metabolomics and machine learning approaches. The targeted urinary metabolomics study was conducted in 70 healthy pregnant controls and 133 pregnant patients having hypertension as cases. Hypertensive disorders of pregnancy-specific metabolites for disease prediction were further extracted using univariate and multivariate statistical analyses. For machine learning analysis, 80% of the data were used for training (79 for hypertensive disorders of pregnancy and 42 for healthy pregnancy) and validation (27 for hypertensive disorders of pregnancy and 14 for healthy pregnancy), and 20% of the data were used for test sets (27 for hypertensive disorders of pregnancy and 14 for healthy pregnancy). The statistical analysis using an unpaired t test revealed 44 differential metabolites. Pathway analysis showed mainly that purine and thiamine metabolism were altered in the group with hypertensive disorders of pregnancy compared with the healthy pregnancy group. The area under the receiver operating characteristic curves of the 5 most predominant metabolites were 0.98 (adenosine), 0.92 (adenosine monophosphate), 0.89 (deoxyadenosine), 0.81 (thiamine), and 0.81 (thiamine monophosphate). The best prediction accuracies were obtained using 2 machine learning models (95% for the gradient boost model and 98% for the decision tree) among the 5 used models. The machine learning models showed higher predictive performance for 3 metabolites (ie, thiamine monophosphate, adenosine monophosphate, and thiamine) among 5 metabolites. The combined accuracies of adenosine from all models were 98.6 in the training set and 95.6 in the test set. Moreover, the predictive performance of adenosine was higher than other metabolites. The relative feature importance of adenosine was also observed in the decision tree and the gradient boost model. Among other metabolites, adenosine and thiamine metabolites were found to differentiate participants with hypertensive disorders of pregnancy from participants with healthy pregnancies; hence, these metabolites can serve as a promising noninvasive marker for the detection of hypertensive disorders of pregnancy.

Maternal Liver and Egg Thiamine Concentrations in Walleye from Lake Oahe, South Dakota

Thiamine, vitamin B1, cannot be synthesized by fish. Therefore, developing embryos are reliant on thiamine stores deposited during oogenesis. Thiaminase, which breaks down thiamine in predatory fish, has been suggested as the probable reason for the poor reproductive success in salmonids. Similarly, walleye Sander vitreus reproduction may also be hampered by thiaminase-containing diets. This study documented maternal liver and egg thiamine concentrations of walleye populations from the Cheyenne, Moreau, and Grand rivers of Lake Oahe, South Dakota, USA in 2000, 2001, 2002, and 2004. Thiamine concentrations were generally lowest in the eggs from the Cheyenne River. Free thiamine, thiamine monophosphate, and total thiamine were all significantly lower in eggs from the Cheyenne River compared to the Moreau and Grand Rivers. Maternal liver thiamine pyrophosphate and total thiamine followed a similar pattern. Egg survival to hatch varied significantly between locations, at 30.5%, 41.4%, and 56.8% in the Cheyenne, Moreau, and Grand Rivers, respectively, but was not significantly correlated with any form of egg thiamine. Considerable year-to-year variation was observed when data from the locations were pooled; egg free thiamine, thiamine monophosphate, and total thiamine were significantly higher in 2002. The lowest mean egg total thiamine level of 2.6 nmol/g was recorded in 2000. Liver free thiamine and thiamine monophosphate were weakly, but significantly negatively correlated with egg survival to hatch to hatch. Egg survival to hatch was significantly negatively correlated with maternal length. The results were likely impacted by the unexpected catastrophic loss of walleye forage because of flooding in 2003.

Polyphosphate Kinases Phosphorylate Thiamine Phosphates

Polyphosphate kinases (PPKs) catalyze the reversible transfer of the γ-phosphate moiety of ATP (or of another nucleoside triphosphate) to a growing chain of polyphosphate (polyP). In this study, we describe that PPKs of various sources are additionally able to phosphorylate thiamine diphosphate (ThP2) to produce thiamine triphosphate (ThP3) and even thiamine tetraphosphate in vitro using polyP as phosphate donor. Furthermore, all tested PPK2s, but not PPK1s, were able to phosphorylate thiamine monophosphate (ThP1) to ThP2 and ThP3 although at low efficiency. The predicted masses and identities of the mono- and oligo-phosphorylated thiamine metabolites were identified by high-performance liquid chromatography tandem mass spectrometry. Moreover, the biological activity of ThP2, that was synthesized by phosphorylation of ThP1 with polyP and PPK, as a cofactor of ThP2-dependent enzymes (here transketolase TktA from Escherichia coli) was confirmed in a coupled enzyme assay. Our study shows that PPKs are promiscuous enzymes in vitro that could be involved in the formation of a variety of phosphorylated metabolites in vivo.

THIAMIN REQUIRING2 is involved in thiamin diphosphate biosynthesis and homeostasis.

The THIAMIN REQUIRING2 (TH2) protein comprising a mitochondrial targeting peptide followed by a transcription enhancement A and a haloacid dehalogenase domain is a thiamin monophosphate (TMP) phosphatase in the vitamin B1 biosynthetic pathway. The Arabidopsis th2-3 T-DNA insertion mutant was chlorotic and deficient in thiamin diphosphate (TDP). Complementation assays confirmed that haloacid dehalogenase domain alone was sufficient to rescue the th2-3 mutant. In pTH2:TH2-GFP/th2-3 complemented plants, the TH2-GFP was localized to the cytosol, mitochondrion, and nucleus, indicating that the vitamin B1 biosynthetic pathway extended across multi-subcellular compartments. Engineered TH2-GFP localized to the cytosol, mitochondrion, nucleus, and chloroplast, could complement the th2 mutant. Together, these results highlight the importance of intracellular TMP and thiamin trafficking in vitamin B1 biosynthesis. In an attempt to enhance the production of thiamin, we created various constructs to overexpress TH2-GFP in the cytosol, mitochondrion, chloroplast, and nucleus. Unexpectedly, overexpressing TH2-GFP resulted in an increase rather than a decrease in TMP. While studies on th2 mutants support TH2 as a TMP phosphatase, analyses of TH2-GFP overexpression lines implicating TH2 may also function as a TDP phosphatase in planta. We propose a working model that the TMP/TDP phosphatase activity of TH2 connects TMP, thiamin, and TDP into a metabolic cycle. The TMP phosphatase activity of TH2 is required for TDP biosynthesis, and the TDP phosphatase activity of TH2 may modulate TDP homeostasis in Arabidopsis.

Thiazolium Salts as Chalcogen Bond Donors

The nitrogen atom is commonly considered to be the positive site in thiazolium cations. Counterintuitively, the shortest cation···anion contacts in thiazolium salts and their benzoderivatives involve the sulfur atom. The geometric features of these contacts allow for their rationalization as charge assisted chalcogen bonds (ChBs). Analyses of the Cambridge Structural Database and calculation of the surface electrostatic potential of benzo[d]thiazolium cations support this rationalization. The relevance and impact of the general tendency of thiazolium derivatives to function as ChB donors are exemplified by the ChBs as short as 87% of the sum of van der Waals radii of involved atoms in thiamine monophosphate, a key compound in human metabolism, and in two commonly used thiazolium organocatalysts.

Vitamin B1 THIAMIN REQUIRING1 synthase mediates the maintenance of chloroplast function by regulating sugar and fatty acid metabolism in rice.

Vitamin B1 (VB1), including thiamin, thiamin monophosphate (TMP), and thiamin pyrophosphate (TPP), is an essential micronutrient for all living organisms. Nevertheless, the precise function of VB1 in rice remains unclear. Here, we described a VB1 auxotrophic mutant, chlorotic lethal seedling (cles) from the mutation of OsTH1, which displayed collapsed chloroplast membrane system and decreased pigment content. OsTH1 encoded a phosphomethylpyrimidine kinase/thiamin-phosphate pyrophosphorylase, and was expressed in various tissues, especially in seedlings, leaves, and young panicles. The VB1 content in cles was markedly reduced, despite an increase in the expression of VB1 synthesis genes. The decreased TPP content affected the tricarboxylic acid cycle, pentose phosphate pathway, and de novo fatty acid synthesis, leading to a reduction in fatty acids (C16:0 and C18:0) and sugars (sucrose and glucose) of cles. Additionally, irregular expression of chloroplast membrane synthesis genes led to membrane collapse. We also found that alternative splicing and translation allowed OsTH1 to be localized to both chloroplast and cytosol. Our study revealed that OsTH1 was an essential enzyme in VB1 biosynthesis and played crucial roles in seedling growth and development by participating in fatty acid and sugar metabolism, providing new perspectives on VB1 function in rice.

Integrating Metabolomics and Network Analysis for Exploring the Mechanism Underlying the Antidepressant Activity of Paeoniflorin in Rats With CUMS-Induced Depression.

Background: Paeoniflorin (PF) represents the major bioactive constituent of the traditional Chinese medicine plant Paeonia suffruticosa (Ranunculaceae), which has a long history as a folk medicine in Asian. Paeoniflorin, a bitter pinene monoterpene glycoside, has antidepressant effects, but its potential therapeutic mechanism has not been thoroughly explored. Methods: Experimental depression in rats was established by the chronic unpredictable mild stress (CUMS) combined with orphan method, and the efficacy of paeoniflorin on depression was evaluated by the sucrose preference test and open field test. The antidepressant mechanism of paeoniflorin was investigated by metabolomic and network pharmacology. The relevant pathways of biomarkers highlighted in metabolomics were explored, and the possible targets of paeoniflorin in the treatment of depression were further revealed through network analysis. The binding activity of paeoniflorin to key targets was verified by molecular docking. Results: Metabolomics showed that rats with CUMS-induced depression had urine metabolic disorders, which were reversed by paeoniflorin through the regulation of metabolic pathways. Metabolites that play a key role in the function of paeoniflorin include citric acid, thiamine monophosphate, gluconolactone, 5-hydroxyindoleacetic acid and stachyose. Key predicted targets are SLC6A4, TNF, IL6 and SLC6A3. An important metabolic pathway is the Citrate cycle (TCA cycle). Conclusion: Network integrative analysis in this study showed that paeoniflorin could improve depressive-like symptoms in model rats with CUMS-induced depression and overall correct the disordered metabolic profile through multiple metabolic pathways.

VITAMIN B1 BIOLOGICAL FUNCTION IMPLEMENTATION IN THE BLOOD OF PATIENTS WITH STOMACH CANCER UNDER SURGICAL INTERVENTION

Background. The probability of undergoing surgery always predetermines the state of stress in a person; therefore, it is advisable to search for ways to optimize and (or) reduce this unfavourable effect. Objective. To find out the mechanism of vitamin B1 antistress activity during surgery. Material and methods. Metabolism intensity was investigated on donors’ blood lysates (n = 19) and those of patients with stage III stomach cancer (n = 64), referred to an elective surgery, aged 51-70. The blood was taken from the cubital vein three days before the operation, after premedication, during the most traumatic moment of the operation, after extubation, as well as on the first and third days of the postoperative period. The surgery was performed under combined multicomponent anesthesia using nitric oxide, sodium hydroxybutyrate, and epidural block. Thiamine and thiamine diphosphate kinase activities were assessed by the concentration of the formed thiamine di- and triphosphates of the vitamin. The activities of thiamine mono-, di- and triphosphatases were determined by the release of inorganic phosphate. The concentration of inorganic phosphate was recorded colorimetrically. The content of B1 and its derivatives in the blood was determined by the method of ion-pair reversed-phase HPLC. Results. There has been observed an increased content of thiamine monophosphate and that of free thiamine in the blood of donors and patients with stomach cancer. The registered rate of the hydrolytic thiamine monophosphatase reaction is not high. At the stages of premedication and maximum trauma of surgical exposure, the concentration of monophosphoric ester rapidly decreases alongside with monophosphatase activation. Therefore, the thiamine monophosphate hydrolysis is the rate-limiting link of vitamin B1 metabolism. The level of free thiamine remains persistently increased at all stages of surgical treatment. Thiamine monophosphatase activity is manifested at two pH optima – of 6.0 and 9.0. Thiamine monophosphate hydrolysis at pH of 9.0 is catalyzed by alkaline phosphatase. At pH of 6.0, in addition to thiamine monophosphoric ester, the enzyme hydrolyzes only p-nitrophenyl phosphate, flavin mononucleotide and phosphotyrosine, that allows it to be classified as hepatic acid phosphatase. The noted changes in B1 metabolism under stress concern mainly non-coenzyme forms - thiamine mono-, triphosphate, and free thiamine, which are used at the stages of thiol reduction as important components of insulin synthesis. Conclusions. The use of vitamin B1 allows to optimize the development of the stress response at all stages of surgical treatment. Its protective effect is achieved through the activation of the insulin-synthetic function of the pancreas, which increases the level of immunoreactive insulin in the blood. The formation of the most favorable physiological conditions for insulin synthesis provides an increased background of free thiamine, which is created due to the hydrolysis of noncoenzyme forms of the vitamin. The relationship between thiamine metabolism and B2 exchange and regulation of intracellular signaling pathways has been traced.

Effects of a Folic Acid 5 Mg Daily Intervention on Markers of Vitamin B Status and Kynurenine Pathway

BackgroundIn Brazil, most folic acid (FA) containing supplements contain 5 mg FA. This dose is usually prescribed for patients with chronic hemolytic anemia and for women planning pregnancy. The effects of high FA doses on vitamin B1, B2 and B6 status including the kynurenine pathway are still unclear.AimsTo assess the effects of a daily intervention with 5 mg FA in healthy subjects on vitamins B1, B2 and B6 serum marker status and serum concentration ok kynurenines.Material and MethodsThirty health subjects participated in a 5 mg/day FA intervention. Blood samples were obtained at baseline (D0) and after 45 (D45) and 90 days (D90) of intervention. We assessed serum concentrations of vitamin B6 (pyridoxal-5'-phosphate (PLP), pyridoxal (PL) and 4-pyridoxic acid (PA)), vitamin B2 (riboflavin and flavin mononucleotide (FMN)) and vitamin B1 vitamers (thiamine and thiamine monophosphate (TMP)), as well as tryptophan, kynurenine and several metabolites of tryptophan catabolism in the kynurenine pathway (3-hydroxykynurenine (HK), kynurenic acid (KA), anthranilic acid (AA), xanthurenic acid (XA), 3-hydroxyanthranilic acid (HAA), picolinic acid (Pic) and quinolinic acid (QA)) using LC-MS/MS. PA ratio (PAr) was calculated by PA/(PLP + PL). All variables were normalized by Box-Cox transformation. Comparisons were performed by repeated-measures ANOVA.ResultsConcentrations of all vitamin B6 vitamers (PLP, PL and PA), as well as PAr, were similar at baseline and D45 and D90 of FA intervention. No differences were found among serum vitamin B2 vitamers (riboflavin and FMN) after FA intervention. However, a trend for a decrease in serum TMP concentrations was observed at D90 (P = 0.054). No changes in serum tryptophan and all serum kynurenine pathway metabolites were observed after intervention with FA compared with baseline, except for AA, which was higher at D45 and D90 than baseline (P < 0.001).ConclusionOur data suggest that daily use of 5 mg FA by healthy subjects may interfere with vitamin B1 status markers. In addition, serum AA increased with FA intervention, which suggest that the use of high dose FA supplements might affect the kynurenine pathway.Financing: FAPESP (2016/18854-4) and CNPq (401586/2014-6) DisclosuresNo relevant conflicts of interest to declare.

The ThiL enzyme is a valid antibacterial target essential for both thiamine biosynthesis and salvage pathways in Pseudomonas aeruginosa

Thiamine pyrophosphate (TPP) is an essential cofactor for various pivotal cellular processes in all living organisms, including bacteria. Thiamine biosynthesis occurs in bacteria but not in humans, so the enzymes in this pathway are attractive targets for antibiotic development. Among these enzymes, thiamine monophosphate kinase (ThiL) catalyzes the final step of this pathway, phosphorylating thiamine monophosphate (TMP) to produce TPP. Here, we extensively investigated ThiL in Pseudomonas aeruginosa, a major pathogen responsible for hospital-acquired infections. We demonstrate that thiL deletion abolishes not only thiamine biosynthesis, but also thiamine salvage capability, and results in growth defects of the ΔthiL strain even in the presence of thiamine derivatives, except for TPP. Most importantly, the pathogenesis of the ΔthiL strain was markedly attenuated compared with wild-type cells, with lower inflammatory cytokine induction and 103–104-fold decreased bacterial load in an in vivo infection model where the intracellular TPP level was in the submicromolar range. To validate P. aeruginosa ThiL (PaThiL) as a drug target, we further characterized its biochemical properties, determining a Vmax of 4.0 ± 0.2 nmol·min-1 and KM values of 111 ± 8 and 8.0 ± 3.5 μM for ATP and TMP, respectively. These comprehensive biological and biochemical results indicate that PaThiL represents a potential drug target for the development of an augmented repertoire of antibiotics against P. aeruginosa.