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Objective To describe the history, key features, recent enhancements, and common applications of the Dietary Supplement Label Database (DSLD). Background and history Although many Americans use dietary supplements, databases of dietary supplements sold in the United States have not been widely available. The DSLD, an easily accessible public-use database was created in 2008 to provide information on dietary supplement composition for use by researchers and consumers. Rationale Accessing current information easily and quickly is crucial for documenting exposures to dietary supplements because they contain nutrients and other bioactive ingredients that may have beneficial or adverse effects on human health. This manuscript details recent developments with the DSLD to achieve this goal and provides examples of how the DSLD has been used. Recent developments With periodic updates to track changes in product composition and capture new products entering the market, the DSLD currently contains more than 71,000 dietary supplement labels. Following usability testing with consumer and researcher user groups completed in 2016, improvements to the DSLD interface were made. As of 2017, both a desktop and mobile device version are now available. Since its inception in 2008, the use of the DSLD has included research, exposure monitoring, and other purposes by users in the public and private sectors. Future directions Further refinement of the user interface and search features to facilitate ease of use for stakeholders is planned. Conclusions The DSLD can be used to track changes in product composition and capture new products entering the market. With over 71,000 DS labels it is a unique resource that policymakers, researchers, clinicians, and consumers may find valuable for multiple applications.

The transsulfuration pathway (TS) acts in sulfur amino acid metabolism by contributing to the regulation of cellular homocysteine, cysteine production, and the generation of H2S for signaling functions. Regulation of TS pathway kinetics involves stimulation of cystathionine β-synthase (CBS) by S-adenosylmethionine (SAM) and oxidants such as H2O2, and by Michaelis-Menten principles whereby substrate concentrations affect reaction rates. Although pyridoxal phosphate (PLP) serves as coenzyme for both CBS and cystathionine γ-lyase (CSE), CSE exhibits much greater loss of activity than CBS during PLP insufficiency. Thus, cellular and plasma cystathionine concentrations increase in vitamin B6 deficiency mainly due to the bottleneck caused by reduced CSE activity. Because of the increase in cystathionine, the canonical production of cysteine (homocysteine → cystathionine → cysteine) is largely maintained even during vitamin B6 deficiency. Typical whole body transsulfuration flux in humans is 3-7 μmol/h per kg body weight. The in vivo kinetics of H2S production via side reactions of CBS and CSE in humans are unknown but they have been reported for cultured HepG2 cells. In these studies, cells exhibit a pronounced reduction in H2S production capacity and rates of lanthionine and homolanthionine synthesis in deficiency. In humans, plasma concentrations of lanthionine and homolanthionine exhibit little or no mean change due to 4-wk vitamin B6 restriction, nor do they respond to pyridoxine supplementation of subjects in chronically low-vitamin B6 status. Wide individual variation in responses of the H2S biomarkers to such perturbations of human vitamin B6 status suggests that the resulting modulation of H2S production may have physiological consequences in a subset of people. Supported by NIH grant DK072398. This paper refers to data from studies registered at clinicaltrials.gov as NCT01128244 and NCT00877812.

Background: Low chronic vitamin B-6 status can occur in a subset of women who use oral contraceptives (OCs) with uncertain metabolic consequences. An insufficiency of cellular pyridoxal 5′-phosphate (PLP), which is the coenzyme form of vitamin B-6, may impair many metabolic processes including one-carbon and tryptophan metabolism. Objective: We investigated the effects of vitamin B-6 supplementation on the in vivo kinetics of one-carbon metabolism and the concentration of one-carbon and tryptophan metabolites in vitamin B-6–deficient OC users. Design: A primed, constant infusion of [13C5]methionine, [3-13C]serine, and [2H3]leucine was performed on 10 OC users (20–40 y old; plasma PLP concentrations

Suboptimal vitamin B-6 status, as reflected by low plasma pyridoxal 5#-phosphate (PLP) concentration, is associated with increased risk of vascular disease. PLP plays many roles, including in one-carbon metabolism for the acquisition and transfer of carbon units and in the transsulfuration pathway. PLP also serves as a coenzyme in the catabolism of tryptophan. We hypothesize that the pattern of these metabolites can provide information reflecting the functional impact of marginal vitamin B-6 deficiency. We report here the concentration of major constituents of one-carbon metabolic processes and the tryptophan catabolic pathway in plasma from 23 healthy men and women before and after a 28-d controlled dietary vitamin B-6 restriction (

Vitamin B-6 deficiency is associated with impaired tryptophan metabolism because of the coenzyme role of pyridoxal 5′-phosphate (PLP) for kynureninase and kynurenine aminotransferase. To investigate the underlying mechanism, we developed a mathematical model of tryptophan metabolism via the kynurenine pathway. The model includes mammalian data on enzyme kinetics and tryptophan transport from the intestinal lumen to liver, muscle, and brain. Regulatory mechanisms and inhibition of relevant enzymes were included. We simulated the effects of graded reduction in cellular PLP concentration, tryptophan loads and induction of tryptophan 2,3-dioxygenase (TDO) on metabolite profiles and urinary excretion. The model predictions matched experimental data and provided clarification of the response of metabolites in various extents of vitamin B-6 deficiency. We found that moderate deficiency yielded increased 3-hydroxykynurenine and a decrease in kynurenic acid and anthranilic acid. More severe deficiency also yielded an increase in kynurenine and xanthurenic acid and more pronounced effects on the other metabolites. Tryptophan load simulations with and without vitamin B-6 deficiency showed altered metabolite concentrations consistent with published data. Induction of TDO caused an increase in all metabolites, and TDO induction together with a simulated vitamin B-6 deficiency, as has been reported in oral contraceptive users, yielded increases in kynurenine, 3-hydroxykynurenine, and xanthurenic acid and decreases in kynurenic acid and anthranilic acid. These results show that the model successfully simulated tryptophan metabolism via the kynurenine pathway and can be used to complement experimental investigations.

Holocarboxylase synthetase (HCS) catalyzes the covalent binding of biotin to both carboxylases and histones. Biotinylated carboxylases and biotinylated histones play crucial roles in the metabolism of fatty acids, amino acids, and glucose, and in gene regulation and genome stability, respectively. HCS null mammals are not viable whereas HCS deficiency is linked to developmental delays in humans and phenotypes such as short life span and low stress resistance in Drosophila.HCS-dependent biotinylation of the polypeptide p67 was detected and quantified in a 96-well plate format using IRDye-streptavidin and infrared spectroscopy.Biotinylation of p67 by recombinant HCS (rHCS) and HCS from human cell extracts depended on time, temperature, and substrate concentration, all consistent with enzyme catalysis rather than non-enzymatic biotinylation. The Michaelis-Menten constant of rHCS for p67 was 4.1±1.5 μmol/l. The minimal concentration of rHCS that can be detected by this assay is less than 1.08 nmol/l. Jurkat cells contained 0.14±0.02 U of HCS activity [μmol of biotinylated p67 formed/(nmol/l HCSh)] in 400 μg of total protein.We developed a 96-well plate assay for high-throughput analysis of HCS activity in biological samples and studies of synthetic and naturally occurring HCS inhibitors.

Measures of B6 status are categorized as direct biomarkers and as functional biomarkers. Direct biomarkers measure B6 vitamers in plasma/serum, urine and erythrocytes, and among these plasma pyridoxal 5′-phosphate (PLP) is most commonly used. Functional biomarkers include erythrocyte transaminase activities and, more recently, plasma levels of metabolites involved in PLP-dependent reactions, such as the kynurenine pathway, one-carbon metabolism, transsulfuration (cystathionine), and glycine decarboxylation (serine and glycine). Vitamin B6 status is best assessed by using a combination of biomarkers because of the influence of potential confounders, such as inflammation, alkaline phosphatase activity, low serum albumin, renal function, and inorganic phosphate. Ratios between substrate-products pairs have recently been investigated as a strategy to attenuate such influence. These efforts have provided promising new markers such as the PAr index, the 3-hydroxykynurenine:xanthurenic acid ratio, and the oxoglutarate:glutamate ratio. Targeted metabolic profiling or untargeted metabolomics based on mass spectrometry allow the simultaneous quantification of a large number of metabolites, which are currently evaluated as functional biomarkers, using data reduction statistics.

Previous studies suggest that histones H3 and H4 are posttranslationally modified by binding of the vitamin biotin, catalyzed by holocarboxylase synthetase (HCS). Albeit a rare epigenetic mark, biotinylated histones were repeatedly shown to be enriched in repeat regions and repressed loci, participating in the maintenance of genome stability and gene regulation. Recently, a team of investigators failed to detect biotinylated histones and proposed that biotinylation is not a natural modification of histones, but rather an assay artifact. Here, we describe the results of experiments, including the comparison of various analytical protocols, antibodies, cell lines, classes of histones, and radiotracers. These studies provide unambiguous evidence that biotinylation is a natural, albeit rare, histone modification. Less than 0.001% of human histones H3 and H4 are biotinylated, raising concerns that the abundance might too low to elicit biological effects in vivo. We integrated information from this study, previous studies, and ongoing research efforts to present a new working model in which biological effects are caused by a role of HCS in multiprotein complexes in chromatin. In this model, docking of HCS in chromatin causes the occasional binding of biotin to histones as a tracer for HCS binding sites.

Holocarboxylase synthetase (HCS) catalyzes the binding of biotin to lysine (K) residues in histones H3 and H4. Histone biotinylation marks are enriched in repressed loci, including retrotransposons. Preliminary studies suggested that K16 in histone H4 is a target for biotinylation by HCS. Here we tested the hypotheses that H4K16bio is a real histone mark in human chromatin, and that H4K16bio is overrepresented in repressed gene loci and repeat regions. Polyclonal rabbit anti-human H4K16bio was generated and affinity purified. An extensive series of testing with synthetic and natural targets confirmed that this new antibody is specific for H4K16bio. Using anti-H4K16bio and chromatin immunoprecipitation assays, we demonstrated that H4K16bio is overrepresented in repeat regions [pericentromeric alpha satellite repeats and long terminal repeats (LTR)] compared with euchromatin promoters. H4K16bio was also enriched in the repressed interleukin-2 gene promoter in human lymphoid cells; transcriptional activation of the interleukin-2 gene by mitogens and phorbol esters coincided with a depletion of the H4K16bio mark at the gene promoter. The enrichment of H4K16bio depended on biotin supply; the enrichment at LTR22 and promoter 1 of the sodium-dependent multivitamin transporter (SMVT) was greater in biotin-supplemented cells compared with biotin-normal and biotin-deficient cells. The enrichment of H4K16bio at LTR15 and SMVT promoter 1 was significantly lower in fibroblasts from an HCS-deficient patient compared with an HCS wild-type control. We conclude that H4K16bio is a real phenomenon and that this mark, like other biotinylation marks, is overrepresented in repressed loci where it marks HCS docking sites.

Modifications of histones such as methylation and acetylation are involved in gene regulation. We recently identified a diet-dependent histone modification, biotinylation of lysines, mediated by ho...