Evidence for the Validity of Pyridoxic Acid (PDA) as a Plasma-Based Endogenous Probe for OAT1 and OAT3 Function in Healthy Subjects.

Plasma pyridoxic acid (PDA) and homovanillic acid (HVA) were recently identified as novel endogenous biomarkers of organic anion transporter (OAT) 1/3 function in monkeys. Consequently, this clinical study assessed the dynamic changes and utility of plasma PDA and HVA as an initial evaluation of OAT1/3 inhibition in early-phase drug development. The study was designed as a single-dose randomized, three-phase, crossover study; 14 Indian healthy volunteers received probenecid (PROB) (1000 mg orally) alone, furosemide (FSM) (40 mg orally) alone, or FSM 1 hour after receiving PROB (40 and 1000 mg orally) on days 1, 8, and 15, respectively. PDA and HVA plasma concentrations remained stable over time in the prestudy and FSM groups. Administration of PROB significantly increased the area under the plasma concentration-time curve (AUC) of PDA by 3.1-fold (dosed alone; P < 0.05), and 3.2-fold (coadministered with FSM; P < 0.01), compared with the prestudy and FSM groups, respectively. The corresponding increase in HVA AUC was 1.8-fold (P > 0.05) and 2.1-fold (P < 0.05), respectively. The increases in PDA AUC are similar to those in FSM AUC, whereas those of HVA are smaller (3.1-3.2 and 1.8-2.1 vs. 3.3, respectively). PDA and HVA renal clearance (CL R) values were decreased by PROB to smaller extents compared with FSM (0.35-0.37 and 0.67-0.73 vs. 0.23, respectively). These data demonstrate that plasma PDA is a promising endogenous biomarker for OAT1/3 function and that its plasma exposure responds in a similar fashion to FSM upon OAT1/3 inhibition by PROB. The magnitude and variability of response in PDA AUC and CL R values between subjects is more favorable relative to HVA.

Pyridoxal 5'-phosphate and related metabolites in hypophosphatasia: Effects of enzyme replacement therapy.

OBJECTIVE: To investigate the utility of serum pyridoxal 5'-phosphate (PLP), pyridoxal (PL), and 4-pyridoxic acid (PA) as a diagnostic marker of hypophosphatasia (HPP) and an indicator of the effect of, and patient compliance with, enzyme replacement therapy (ERT), we measured PLP, PL, and PA concentrations in serum samples from HPP patients with and without ERT. METHODS: Blood samples were collected from HPP patients and serum was frozen as soon as possible (mostly within one hour). PLP, PL, and PA concentrations were analyzed using high-performance liquid chromatography with fluorescence detection after pre-column derivatization by semicarbazide. We investigated which metabolites are associated with clinical phenotypes and how these metabolites change with ERT. RESULTS: Serum samples from 20 HPP patients were analyzed. The PLP-to-PL ratio and PLP concentration were elevated in all HPP patients. They correlated negatively with serum alkaline phosphatase (ALP) activity and showed higher values in more severe phenotypes (perinatal severe and infantile HPP) compared with other phenotypes. PL concentration was reduced only in perinatal severe HPP. ERT reduced the PLP-to-PL ratio to mildly reduced or low-normal levels and the PLP concentration was reduced to normal or mildly elevated levels. Urine phosphoethanolamine (PEA) concentration did not return to normal levels with ERT in most patients. CONCLUSIONS: The serum PLP-to-PL ratio is a better indicator of the effect of ERT for HPP than serum PLP and urine PEA concentrations, and a PLP-to-PL ratio of <4.0 is a good indicator of the effect of, and patient compliance with, ERT.

Discovery and Validation of Pyridoxic Acid and Homovanillic Acid as Novel Endogenous Plasma Biomarkers of Organic Anion Transporter (OAT) 1 and OAT3 in Cynomolgus Monkeys.

Perturbation of organic anion transporter (OAT) 1- and OAT3-mediated transport can alter the exposure, efficacy, and safety of drugs. Although there have been reports of the endogenous biomarkers for OAT1/3, none of these have all of the characteristics required for a clinical useful biomarker. Cynomolgus monkeys were treated with intravenous probenecid (PROB) at a dose of 40 mg/kg in this study. As expected, PROB increased the area under the plasma concentration-time curve (AUC) of coadministered furosemide, a known substrate of OAT1 and OAT3, by 4.1-fold, consistent with the values reported in humans (3.1- to 3.7-fold). Of the 233 plasma metabolites analyzed using a liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based metabolomics method, 29 metabolites, including pyridoxic acid (PDA) and homovanillic acid (HVA), were significantly increased after either 1 or 3 hours in plasma from the monkeys pretreated with PROB compared with the treated animals. The plasma of animals was then subjected to targeted LC-MS/MS analysis, which confirmed that the PDA and HVA AUCs increased by approximately 2- to 3-fold by PROB pretreatments. PROB also increased the plasma concentrations of hexadecanedioic acid (HDA) and tetradecanedioic acid (TDA), although the increases were not statistically significant. Moreover, transporter profiling assessed using stable cell lines constitutively expressing transporters demonstrated that PDA and HVA are substrates for human OAT1, OAT3, OAT2 (HVA), and OAT4 (PDA), but not OCT2, MATE1, MATE2K, OATP1B1, OATP1B3, and sodium taurocholate cotransporting polypeptide. Collectively, these findings suggest that PDA and HVA might serve as blood-based endogenous probes of cynomolgus monkey OAT1 and OAT3, and investigation of PDA and HVA as circulating endogenous biomarkers of human OAT1 and OAT3 function is warranted.

Serum B6 vitamers (pyridoxal 5'-phosphate, pyridoxal, and 4-pyridoxic acid) and pancreatic cancer risk: two nested case-control studies in Asian populations.

BACKGROUND: Vitamin B6 is an important enzymatic cofactor in pathways relevant for the development of pancreatic cancer. In order to evaluate vitamin B6 as a preventive factor for pancreatic cancer, a biomarker approach is needed to overcome the limitations inherent in self-reported dietary information. METHODS: To determine whether levels of serum B6 vitamers, including pyridoxal 5'-phosphate (PLP), pyridoxal (PL), 4-pyridoxic acid (PA), and the PA/(PLP + PL) ratio (PAr), were associated with risk of pancreatic cancer, two nested case-control studies of 187 incident pancreatic cancer cases and 258 individually matched controls were conducted within two prospective cohorts of 81,501 participants in Shanghai, China, and Singapore. PLP, PL, and PA were quantified in pre-diagnostic serum samples. Odds ratios and 95% confidence intervals (CIs) were calculated using conditional logistic regression with adjustment for potential confounders. RESULTS: The median (5th-95th percentiles) concentrations of serum PLP among control subjects of the Shanghai and Singapore cohorts were 25.7 (10.0-91.7) nmol/L and 58.1 (20.8-563.0) nmol/L, respectively. In pooled analyses, high serum PLP was associated with a reduced risk of pancreatic cancer (P for trend = 0.048); the adjusted odds ratio for the highest category of PLP (>52.4 nmol/L) was 0.46 (95% CI 0.23, 0.92) compared to vitamin B6 deficiency (<20 nmol/L). No associations were found for serum PL, PA, or PAr with pancreatic cancer risk. CONCLUSIONS: Higher concentrations of PLP may protect against the development of pancreatic cancer. The protective effect may be more apparent in populations with low concentrations of circulating vitamin B6.

The value of plasma vitamin B6 profiles in early onset epileptic encephalopathies.

BACKGROUND: Recent decades have unravelled the molecular background of a number of inborn errors of metabolism (IEM) causing vitamin B6-dependent epilepsy. As these defects interfere with vitamin B6 metabolism by different mechanisms, the plasma vitamin B6 profile can give important clues for further molecular work-up. This has so far been investigated in only a small number of patients. METHODS: We evaluated the vitamin B6 vitamers pyridoxal 5'-phosphate (PLP), pyridoxal (PL), pyridoxamine (PM), pyridoxine (PN) and the catabolite pyridoxic acid (PA) in the so far largest patient cohort: reference (n = 50); pyridox(am)ine 5'-phosphate oxidase (PNPO) deficiency (n = 6); antiquitin (ATQ) deficiency (n = 21); tissue non-specific alkaline phosphatase (TNSALP) deficiency (n = 2) and epileptic encephalopathy (EE) of unknown etiology tested negative for ATQ and PNPO deficiency (n = 64). RESULTS: High plasma PM concentration was found in all patients with PNPO deficiency irrespective of vitamin B6 supplementation. Their PM concentration and the PM/PA ratio was significantly higher (p < 0.0001), compared to any other patients analysed. One patient with TNSALP deficiency and sampling prior to PN supplementation had markedly elevated plasma PLP concentration. On PN supplementation, patients with TNSALP deficiency, ATQ deficiency and patients of the EE cohort had similar plasma vitamin B6 profiles that merely reflect the intake of supra-physiological doses of vitamin B6. The interval of sampling to the last PN intake strongly affected the plasma concentrations of PN, PL and PA. CONCLUSIONS: PM concentrations and the PM/PA ratio clearly separated PNPO-deficient patients from the other cohorts. The plasma PM/PA ratio thus represents a robust biomarker for the selective screening of PNPO deficiency.

Markers of vitamin B6 status and metabolism as predictors of incident cancer: the Hordaland Health Study.

Dietary intake and/or circulating concentrations of vitamin B6 have been associated with risk of cancer, but results are inconsistent and mechanisms uncertain. Pyridoxal 5'-phosphate (PLP) is the most commonly used marker of B6 status. We recently proposed the ratio 3-hydroxykynurenine/xanthurenic acid (HK/XA) as an indicator of functional vitamin B6 status, and the 4-pyridoxic acid (PA) /(pyridoxal (PL) +PLP) ratio (PAr) as a marker of vitamin B6 catabolism during inflammation. We compared plasma PLP, HK/XA and PAr as predictors of cancer incidence in a prospective community-based cohort in Norway. This study included 6,539 adults without known cancer at baseline (1998-99) from the Hordaland Health Study (HUSK). HR and 95% CI were calculated for the risk of overall and site-specific cancers using multivariate Cox proportional hazards regression with adjustment for potential confounders. After a median follow-up time of 11.9 years, 963 cancer cases (501 men and 462 women) were identified. Multivariate-adjusted Cox-regression showed no significant relation of plasma PLP or HK/XA with risk of incident cancer. In contrast, PAr was significantly associated with risk of cancer with HR (95% CI) = 1.31 (1.12-1.52) per two standard deviation (SD) increment (p < 0.01). Further analysis showed that PAr was a particular strong predictor of lung cancer with HR (95% CI) = 2.46 (1.49-4.05) per two SD increment (p < 0.01). The present results indicate that associations of vitamin B6 with cancer may be related to increased catabolism of vitamin B6, in particular for lung cancer where inflammation may be largely involved in carcinogenesis.

Common Variants at Putative Regulatory Sites of the Tissue Nonspecific Alkaline Phosphatase Gene Influence Circulating Pyridoxal 5'-Phosphate Concentration in Healthy Adults.

BACKGROUND: Vitamin B-6 interconversion enzymes are important for supplying pyridoxal 5'-phosphate (PLP), the co-enzyme form, to tissues. Variants in the genes for these enzymes [tissue nonspecific alkaline phosphatase (ALPL), pyridoxamine 5'-phosphate oxidase, pyridoxal kinase, and pyridoxal phosphatase] could affect enzyme function and vitamin B-6 status. OBJECTIVES: We tested whether single-nucleotide polymorphisms (SNPs) in these genes influence vitamin B-6 status markers [plasma PLP, pyridoxal (PL), and 4-pyridoxic acid (PA)], and explored potential functional effects of the SNPs. METHODS: Study subjects were young, healthy adults from Ireland (n = 2345). We measured plasma PLP, PL, and PA with liquid chromatography-tandem mass spectrometry and genotyped 66 tag SNPs in the 4 genes. We tested for associations with single SNPs in candidate genes and also performed genome-wide association study (GWAS) and gene-based analyses. RESULTS: Seventeen SNPs in ALPL were associated with altered plasma PLP in candidate gene analyses (P < 1.89 × 10(-4)). In the GWAS, 5 additional ALPL SNPs were associated with altered plasma PLP (P < 5.0 × 10(-8)). Gene-based analyses that used the functional linear model ß-spline (P = 4.04 × 10(-15)) and Fourier spline (P = 5.87 × 10(-15)) methods also showed associations between ALPL and altered plasma PLP. No SNPs in other genes were associated with plasma PLP. The association of the minor CC genotype of 1 ALPL SNP, rs1256341, with reduced ALPL expression in the HapMap Northern European ancestry population is consistent with the positive association between the CC genotype and plasma PLP in our study (P = 0.008). No SNP was associated with altered plasma PL or PA. CONCLUSIONS: In healthy adults, common variants in ALPL influence plasma PLP concentration, the most frequently used biomarker for vitamin B-6 status. Whether these associations are indicative of functional changes in vitamin B-6 status requires more investigation.

Associations of serum vitamin B6 status with the risks of cardiovascular, cancer, and all-cause mortality in the elderly.

Background: There are few studies investigating the relationship between serum vitamin B6 and mortality risk in the elderly. This study hereby evaluated the associations between biomarkers of serum vitamin B6 status and cardiovascular, cancer, and all-cause mortality risks in the elderly. Methods: Our study included a total of 4,881 participants aged 60 years or older from the National Health and Nutrition Examination Survey (NHANES) 2005-2010. Serum vitamin B6 status was estimated based on levels of pyridoxal 5'-phosphate (PLP), 4-pyridoxic acid (4-PA), and vitamin B6 turnover rate (4-PA/PLP) detected by high-performance liquid chromatography. Survival status and corresponding causes of death were matched through the National Death Index records through December 31, 2019. Multivariate Cox regression model was adopted to assess the relationships between serum vitamin B6 status and the risk of mortality. Results: During a median follow-up period of 10.33 years, 507 cardiovascular deaths, 426 cancer deaths, and 1995 all-cause deaths were recorded, respectively. In the multivariate-adjusted Cox model, the hazard ratios (HRs) and 95% confidence intervals (CIs) for the highest versus the lowest quartiles of PLP, 4-PA, and 4-PA/PLP were 0.70(0.54-0.90), 1.33(0.88-2.02), and 2.01(1.41-2.79) for cardiovascular mortality, 0.73(0.52-1.02), 1.05(0.71-1.57), and 1.95(1.25-3.05) for cancer mortality, and 0.62(0.53-0.74), 1.05(0.82-1.34), and 2.29(1.87-2.79) for all-cause mortality, respectively. Conclusion: Our study found that lower serum PLP levels were associated with increased risks of cardiovascular and all-cause mortality among the elderly population. And higher vitamin B6 turnover rate was associated with increased risks of cardiovascular, cancer, and all-cause mortality.

Vitamin B6 and the immunity in kidney transplant recipients.

OBJECTIVE: The study aimed to determine vitamin B6 status in elderly (age ≥ 60 years) and younger (age <60 years) recipients of allogeneic kidney graft and to investigate associations between vitamin B6 status and immunity markers. DESIGN: A retrospective observational study. SETTING: The study was conducted at the Medical University of Gdansk, Poland. SUBJECTS: We recruited 34 kidney allograft recipients (17 males and 17 females) and allocated them into 2 groups: patients aged ≥ 60 years (18 patients) and those aged <60 years (16 patients). Exclusion criteria included patients receiving vitamin B6 supplementation or drugs known to influence vitamin B6 metabolism. MAIN OUTCOME MEASURE: Plasma levels of pyridoxal 5'-phosphate (PLP), pyridoxal, pyridoxine, pyridoxamine, pyridoxamine 5'-phosphate, and 4 pyridoxic acid were determined by high-performance liquid chromatography. Measured immunity markers were serum cytokines (interleukin-6, interleukin-10, and transforming growth factor-ß), levels of T-lymphocyte subsets, and the proliferative ability of peripheral blood mononuclear cells. RESULTS: Concentrations of all vitamin B6 vitamers in plasma (PLP, pyridoxal, pyridoxamine 5'-phosphate, pyridoxamine, pyridoxine, 4 pyridoxic acid) were comparable in the 2 studied groups. There were no cases of PLP deficiency in the study population, but 29% of patients had PLP concentrations more than the upper reference limit. Vitamin B6 vitamer concentrations were not influenced by gender, estimated glomerular filtration rate, and circulating phosphate concentration. There was no difference in immunity markers according to age. However, the plasma concentrations of vitamin B6 vitamers were inversely associated with levels of CD28(+) lymphocyte subsets, as well as with the proliferative response of peripheral blood mononuclear cells in both groups. CONCLUSIONS: No cases of vitamin B6 deficiency were found among kidney allograft recipients, and we report inverse links between vitamin B6 vitamer concentrations and markers of cellular immunity, suggesting that bioactive vitamin B6 concentration in kidney allograft recipients merits further investigation.

Vitamin B-6 vitamer levels in plasma and related symptoms in hemodialysis subjects taking low- and high-dose renal multivitamin supplements.

INTRODUCTION: The purpose of this study was to evaluate the B-6 vitamers in plasma and related symptoms in hemodialysis subjects taking high- or low-dose vitamins. METHODS: A total of 24 hemodialysis (HD) subjects were divided into two groups. Twelve subjects received a high-dose vitamin supplement [50 mg pyridoxine hydrochloride (PN-HCl) /tablet] and 12 received a low-dose vitamin supplements containing (10 mg PN-HCl/tablet) for 6+ months. Plasma B-6 vitamers were analyzed using HPLC. Other data were obtained from subjects' medical records. Subjects were assessed for vitamin B-6 related symptoms. Cluster analysis was used to form symptom groups. Student t-tests and analysis of variance were used to determine differences (p < 0.05) in group means. RESULTS: The mean ± SD plasma B-6 vitamer and 4-pyridoxic acid concentrations (nmol/L) were as follows in the 10-mg and 50-mg PN-HCl groups, respectively: pyridoxal- 5'-phosphate (PLP) 10 ± 3 and 16 ± 8 (p = 0.04); pyridoxal (PL) 50 ± 96 and 68 ± 06; pyridoxine (PN) 26 ± 50 and 191 ± 107; and 4-pyridoxic acid (4-PA) 43 ± 64 and 99 ± 361. The cluster group with a significantly higher (p = 0.04) plasma 4-PA concentration of 167 ± 697 nmol/L reported more tingling hands, tachycardia, and diarrhea. CONCLUSION: Plasma PLP levels and symptoms related to B-6 in HD subjects are impacted by dose of PN-HCl.

Untargeted metabolomics identifies enterobiome metabolites and putative uremic toxins as substrates of organic anion transporter 1 (Oat1).

Untargeted metabolomics on the plasma and urine from wild-type and organic anion transporter-1 (Oat1/Slc22a6) knockout mice identified a number of physiologically important metabolites, including several not previously linked to Oat1-mediated transport. Several, such as indoxyl sulfate, derive from Phase II metabolism of enteric gut precursors and accumulate in chronic kidney disease (CKD). Other compounds included vitamins (pantothenic acid, 4-pyridoxic acid), urate, and metabolites in the tryptophan and nucleoside pathways. Three metabolites, indoxyl sulfate, kynurenine, and xanthurenic acid, were elevated in the plasma and interacted strongly and directly with Oat1 in vitro with IC50 of 18, 12, and 50 µM, respectively. A pharmacophore model based on several identified Oat1 substrates was used to screen the NCI database and candidate compounds interacting with Oat1 were validated in an in vitro assay. Together, the data suggest a complex, previously unidentified remote communication between the gut microbiome, Phase II metabolism in the liver, and elimination via Oats of the kidney, as well as indicating the importance of Oat1 in the handling of endogenous toxins associated with renal failure and uremia. The possibility that some of the compounds identified may be part of a larger remote sensing and signaling pathway is also discussed.

Population pharmacokinetic modeling and simulation to support qualification of pyridoxic acid as endogenous biomarker of OAT1/3 renal transporters.

Renal clearance of many drugs is mediated by renal organic anion transporters OAT1/3 and inhibition of these transporters may lead to drug-drug interactions (DDIs). Pyridoxic acid (PDA) and homovanillic acid (HVA) were indicated as potential biomarkers of OAT1/3. The objective of this study was to develop a population pharmacokinetic model for PDA and HVA to support biomarker qualification. Simultaneous fitting of biomarker plasma and urine data in the presence and absence of potent OAT1/3 inhibitor (probenecid, 500 mg every 6 h) was performed. The impact of study design (multiple vs. single dose of OAT1/3 inhibitor) and ability to detect interactions in the presence of weak/moderate OAT1/3 inhibitors was investigated, together with corresponding power calculations. The population models developed successfully described biomarker baseline and PDA/HVA OAT1/3-mediated interaction data. No prominent effect of circadian rhythm on PDA and HVA individual baseline levels was evident. Renal elimination contributed greater than 80% to total clearance of both endogenous biomarkers investigated. Estimated probenecid unbound in vivo OAT inhibitory constant was up to 6.4-fold lower than in vitro values obtained with PDA as a probe. The PDA model was successfully verified against independent literature reported datasets. No significant difference in power of DDI detection was found between multiple and single dose study design when using the same total daily dose of 2000 mg probenecid. Model-based simulations and power calculations confirmed sensitivity and robustness of plasma PDA data to identify weak, moderate, and strong OAT1/3 inhibitors in an adequately powered clinical study to support optimal design of prospective clinical OAT1/3 interaction studies.

Vitamin B6 metabolism in chronic kidney disease--relation to transsulfuration, advanced glycation and cardiovascular disease.

BACKGROUND: Vitamin deficiency is common in chronic kidney disease (CKD). Data on B(6) supply and possible relationships to cardiovascular events (CVE) in CKD are rare. Pyridoxamine exerts inhibitory effects on the formation of advanced glycation endproducts (AGE) implicated in the pathogenesis of CKD and atherosclerosis. METHODS: In 48 CKD patients at stage 2-4, 72 hemodialysis patients (HD), 38 renal transplant recipients (RTR) and 141 healthy controls (mean age 58 +/- 13, 61 +/- 12, 50 +/- 12 and 54 +/- 16 years, respectively), plasma and red blood cell (RBC) concentrations of pyridoxal-5'-phosphate (PLP), pyridoxal (PL), 4-pyridoxic acid (PA), pyridoxamine-5'-phosphate (PMP) and of the AGE pentosidine were measured by high-performance liquid chromatography, N(epsilon)-(carboxymethyl)lysine and imidazolone by an ELISA, and total homocysteine and cystathionine by gas chromatography-mass spectrometry. RESULTS: Despite routine low-dose vitamin supplementation in HD, plasma PLP was decreased in HD (79 +/- 69 nmol/l) compared with CKD stage 2-4 patients (497 +/- 944 nmol/l), RTR (416 +/- 604 nmol/l) and controls (159 +/- 230 nmol/l; p < 0.001). Plasma PA was significantly increased in HD (11,667 +/- 17,871 nmol/l) in comparison with CKD stage 2-4 (435 +/- 441 nmol/l), RTR (583 +/- 668 nmol/l) and controls (46 +/- 49 nmol/l; p < 0.001). B(6) forms were significantly affected by renal function (R = 0.792, p < 0.001 for CKD stage 2-4). There was no relation of vitamers with a history of CVE. Relationships between B(6) forms and AGE (RBC-PMP with pentosidine in CKD stage 2-4: R = -0.351, p < 0.05) were found. CONCLUSION: HD patients showed a deficiency in PLP in plasma but not in RBC. Prospective trials are needed to elucidate the potential role of elevated PA on cardiovascular and renal outcome in CKD. Vitamin B(6) supplementation might be successful in preventing AGE-related pathologies.

A simplified protein precipitation and filtration procedure for determining serum vitamin B6 by high-performance liquid chromatography.

Protein precipitation followed by centrifuge filtration was tested as a simplified sample preparation procedure for quantifying pyridoxal 5'-phosphate (PLP) and 4-pyridoxic acid (4PA) in serum by high-performance liquid chromatography. Serum samples (n=160) were prepared by both centrifuge filtration and an established technique using traditional supernatant extraction with manual filtration. Bland-Altman bias analysis (95% confidence levels [CLs]) of the results showed a -1.3 (-2.2, -0.5)% difference in PLP values and a -6.2 (-7.3, -5.2)% difference in 4PA values using the simplified sample preparation. These deviations were found to be well within allowable biases on the basis of biologic variation.

Plasma metabolites Xanthine, 4-Pyridoxate, and d-glutamic acid as novel potential biomarkers for pulmonary tuberculosis.

BACKGROUND: The lack of rapid and efficient diagnostic methods has been one of the most frustrating challenges in controlling the pulmonary tuberculosis (TB) epidemic. This study was aimed to identify novel non-invasive biomarkers for pulmonary TB. METHODS: The subjects in this study were divided into four groups: the pulmonary TB group, the community-acquired pneumonia (CAP) group, the lung cancer (LC) group, and the normal control (NC) group. Plasma small molecule metabolites were investigated in each group by using ultra-high performance liquid chromatography coupled with Q Exactive mass spectrometry. Multivariate statistical methods and bioinformatics were used to analyze the data. RESULTS: We identified three differential plasma metabolites such as, Xanthine, 4-Pyridoxate and d-glutamic acid in the pulmonary TB group, compared to the other groups (CAP, LC and NC). The pathway enrichment analysis indicated that the energy source in pulmonary TB was multi-center, which might be involved in maintaining the reproductive ability and virulence of Mycobacterium tuberculosis. CONCLUSION: The results suggested that Xanthine, 4-Pyridoxate, and d-glutamic acid may serve as potential biomarkers for pulmonary TB. The present study provides experimental basis for developing potential biomarkers of pulmonary TB.

[Food Poisoning by Ginkgo Seeds through Vitamin B6 Depletion].

Overconsumption of Ginkgo biloba seeds induces food poisoning characterized by tonic-clonic convulsions and vomiting. The primary toxic component, 4'-O-methylpyridoxine (MPN), was purified from the seeds in 1985. This review includes the following aspects of ginkgo seed poisoning: 1) toxicity related to the content of MPN and MPN glucoside in G. biloba seeds; 2) the effect of MPN on vitamin B6 analogs, including an increase in pyridoxal and pyridoxic acid and decrease in pyridoxal-5'-phosphate plasma concentrations; 3) case reports of ginkgo seed poisoning in Asia, North America, and Europe, and their effective treatment via vitamin B6 administration. Considering the increase in the use of G. biloba seeds, it is essential to raise global awareness of their potential toxicity.

The PAr index, an indicator reflecting altered vitamin B-6 homeostasis, is associated with long-term risk of stroke in the general population: the Hordaland Health Study (HUSK).

Background: Vitamin B-6 homeostasis is altered during inflammation and immune activation. It is unknown whether altered vitamin B-6 homeostasis is associated with the risk of stroke. Objective: We investigated the relation between the ratio plasma 4-pyridoxic acid: (pyridoxal + pyridoxal-5'-phosphate) (PAr) as an indicator of altered vitamin B-6 homeostasis and the risk of stroke in the general population. Design: We conducted a prospective analysis of the community-based Hordaland Health Study (HUSK) in 6891 adults (born during 1925-1927 and 1950-1951) without known stroke at baseline (1998-1999). Participants were followed via linkage to the CVDNOR (Cardiovascular Disease in Norway) project and the Cause of Death Registry. HRs and 95% CIs were calculated using Cox proportional hazards analyses. Results: A total of 390 participants (193 men and 197 women) developed stroke over a median follow-up period of 11 y. Study participants with elevated PAr experienced a higher risk of incident stroke in an essentially linear dose-response fashion. The HR (95% CI) for the highest compared with the lowest quartile of PAr was 1.97 (1.42, 2.73; P-trend <0.001) for total stroke and 2.09 (1.42, 3.09; P-trend <0.001) for ischemic stroke after adjustment for age, sex, body mass index (BMI), smoking, education, physical activity, estimated glomerular filtration rate, hypertension, diabetes, total cholesterol, and statin use. PAr had greater predictive strength than did C-reactive protein, current smoking, diabetes, hypertension, estimated glomerular filtration rate, and physical activity. The associations were similar in subgroups stratified by age group, sex, BMI, current smoking, hypertension, diabetes, and statin use at baseline. Conclusions: Higher plasma PAr was independently associated with increased risk of incident stroke in all participants and across all subgroups stratified by conventional risk predictors. Our novel findings point to and expand the range of inflammation and immune activation processes that may be relevant for the pathogenesis and prevention of stroke. This trial was registered at clinicaltrials.gov as NCT03013725.

A Non-Targeted LC-MS Profiling Reveals Elevated Levels of Carnitine Precursors and Trimethylated Compounds in the Cord Plasma of Pre-Eclamptic Infants.

Preeclampsia (PE) is a complex pregnancy disorder. It is not extensively known how the metabolic alterations of PE women contribute to the metabolism of newborn. We applied liquid chromatography-mass spectrometry (LC-MS) based non-targeted metabolomics to determine whether the metabolic profile of plasma from umbilical cord differs between infants born to PE and non-PE pregnancies in the FINNPEC study. Cord plasma was available from 42 newborns born from PE and 53 from non-PE pregnancies. 133 molecular features differed between PE and non-PE newborns after correction for multiple testing. Decreased levels of 4-pyridoxic acid were observed in the cord plasma samples of PE newborns when compared to non-PE newborns. Compounds representing following areas of metabolism were increased in the cord plasma of PE newborns: urea and creatine metabolism; carnitine biosynthesis and acylcarnitines; putrescine metabolites; tryptophan metabolism and phosphatidylcholines. To our knowledge, this study is the first one to apply LC-MS based metabolomics in cord plasma of PE newborns. We demonstrate that this strategy provides a global picture of the widespread metabolic alterations associated with PE and particularly the elevated levels of carnitine precursors and trimethylated compounds appear to be associated with PE at birth.

Plasma vitamin B-6 forms and their relation to transsulfuration metabolites in a large, population-based study.

BACKGROUND: Vitamin B-6 exists in different forms; one of those forms, pyridoxal 5'-phosphate (PLP), serves a cofactor in many enzyme reactions, including the transsulfuration pathway, in which homocysteine is converted to cystathionine and then to cysteine. Data on the relations between indexes of vitamin B-6 status and transsulfuration metabolites in plasma are sparse and conflicting. OBJECTIVE: We investigated the distribution and associations of various vitamin B-6 species in plasma and their relation to plasma concentrations of transsulfuration metabolites. DESIGN: Nonfasting blood samples from 10 601 healthy subjects with a mean age of 56.4 y were analyzed for all known vitamin B-6 vitamers, folate, cobalamin, riboflavin, total homocysteine, cystathionine, total cysteine, methionine, and creatinine. All subjects were genotyped for the methylenetetrahydrofolate reductase (MTHFR) 677C-->T polymorphism. RESULTS: Plasma concentrations of the main vitamin B-6 vitamers--PLP, pyridoxal, and 4-pyridoxic acid--were strongly correlated. Among the vitamin B-6 vitamers, PLP showed the strongest and most consistent inverse relation to total homocysteine and cystathionine, but the dose response was different for the 2 metabolites. The PLP-total homocysteine relation was significant only in the lowest quartile of the vitamin B-6 distribution and was strongest in subjects with the MTHFR 677TT genotype, whereas cystathionine showed a graded response throughout the range of vitamin B-6 vitamer concentrations, and the effect was not modified by the MTHFR 677C-->T genotype. CONCLUSION: This large population-based study provided precise estimates of the relation between plasma concentrations of vitamin B-6 forms and transsulfuration metabolites as modified by the MTHFR 677C-->T genotype.

Pyridoxal 5'-phosphate, pyridoxal, and 4-pyridoxic acid in the paired serum and cerebrospinal fluid of children.

BACKGROUND: We quantified pyridoxal 5'-phosphate (PLP), pyridoxal (PL), and 4-pyridoxic acid (PA) in paired serum and cerebrospinal fluid (CSF) samples from children and investigated the effect of age on the concentrations and CSF-to-serum ratios of these vitamers. METHODS: Serum and CSF samples prospectively collected from 49 pediatric patients were analyzed. PLP, PL, and PA were measured using high-performance liquid chromatography with fluorescence detection, using pre-column derivatization by semicarbazide. Effects of age on these vitamers, the PLP-to-PL ratio, CSF-to-serum PLP ratio, and CSF-to-serum PL ratio were evaluated using correlation analysis. RESULTS: The PLP, PL, and PA concentrations in the serum and CSF were higher at younger ages, except for CSF PA concentrations that were mostly below the limit of detection (<1.2nmol/l). The PLP-to-PL ratios in the serum and CSF correlated positively with age. The CSF-to-serum PLP ratio and CSF-to-serum PL ratio were independent of age. CONCLUSIONS: Age-related changes in PLP, PL, and PA in serum and in CSF from pediatric patients and CSF-to-serum ratios of PLP and PL demonstrated in this study will provide valuable information for evaluating PLP supply to the central nervous system from the peripheral blood.