Discovery of a novel amino acid racemase through exploration of natural variation in Arabidopsis thaliana.

Plants produce diverse low-molecular-weight compounds via specialized metabolism. Discovery of the pathways underlying production of these metabolites is an important challenge for harnessing the huge chemical diversity and catalytic potential in the plant kingdom for human uses, but this effort is often encumbered by the necessity to initially identify compounds of interest or purify a catalyst involved in their synthesis. As an alternative approach, we have performed untargeted metabolite profiling and genome-wide association analysis on 440 natural accessions of Arabidopsis thaliana. This approach allowed us to establish genetic linkages between metabolites and genes. Investigation of one of the metabolite-gene associations led to the identification of N-malonyl-D-allo-isoleucine, and the discovery of a novel amino acid racemase involved in its biosynthesis. This finding provides, to our knowledge, the first functional characterization of a eukaryotic member of a large and widely conserved phenazine biosynthesis protein PhzF-like protein family. Unlike most of known eukaryotic amino acid racemases, the newly discovered enzyme does not require pyridoxal 5'-phosphate for its activity. This study thus identifies a new d-amino acid racemase gene family and advances our knowledge of plant d-amino acid metabolism that is currently largely unexplored. It also demonstrates that exploitation of natural metabolic variation by integrating metabolomics with genome-wide association is a powerful approach for functional genomics study of specialized metabolism.

Blueberry intake elevates post-exercise anti-inflammatory oxylipins: a randomized trial.

This study determined if 18 days of supplementation with blueberries (BL) compared to placebo (PL) could mitigate muscle soreness and damage and improve inflammation resolution in untrained adults (n = 49, ages 18-50 years) after engaging in a 90-min bout of "weekend warrior" eccentric exercise. The BL freeze dried supplement provided 1 cup of fresh blueberries per day equivalent with 805 mg/day total phenolics and 280 mg/day anthocyanins. Urine levels of eight BL gut-derived phenolics increased after 14- and 18-days supplementation with 83% higher concentrations in BL vs. PL (p < 0.001). The 90-min exercise bout caused significant muscle soreness and damage during 4d of recovery and a decrease in exercise performance with no significant differences between PL and BL. Plasma oxylipins were identified (n = 76) and grouped by fatty acid substrates and enzyme systems. Linoleic acid (LA) oxylipins generated from cytochrome P450 (CYP) (9,10-, 12,13-dihydroxy-9Z-octadecenoic acids) (diHOMEs) were lower in BL vs. PL (treatment effect, p = 0.051). A compositive variable of 9 plasma hydroxydocosahexaenoic acids (HDoHEs) generated from docosahexaenoic acid (DHA, 22:6) and lipoxygenase (LOX) was significantly higher in BL vs. PL (treatment effect, p = 0.008). The composite variable of plasma 14-HDoHE, 17-HDoHE, and the eicosapentaenoic acid (EPA)-derived oxylipin 18-hydroxyeicosapentaenoic acid (18-HEPE) (specialized pro-resolving lipid mediators, SPM, intermediates) was significantly higher in BL vs PL (treatment effect, p = 0.014). Pearson correlations showed positive relationships between post-exercise DHA-LOX HDoHEs and SPM intermediates with urine blueberry gut-derived phenolics (r = 0.324, p = 0.023, and r = 0.349, p = 0.015, respectively). These data indicate that 18d intake of 1 cup/day blueberries compared to PL was linked to a reduction in pro-inflammatory diHOMES and sustained elevations in DHA- and EPA-derived anti-inflammatory oxylipins in response to a 90-min bout of unaccustomed exercise by untrained adults.

Influence of 2 Weeks of Mango Ingestion on Inflammation Resolution after Vigorous Exercise.

Mangoes have a unique nutrient profile (carotenoids, polyphenols, sugars, and vitamins) that we hypothesized would mitigate post-exercise inflammation. This study examined the effects of mango ingestion on moderating exercise-induced inflammation in a randomized crossover trial with 22 cyclists. In random order with trials separated by a 2-week washout period, the cyclists ingested 330 g mango/day with 0.5 L water or 0.5 L of water alone for 2 weeks, followed by a 2.25 h cycling bout challenge. Blood and urine samples were collected pre- and post-2 weeks of supplementation, with additional blood samples collected immediately post-exercise and 1.5-h, 3-h, and 24 h post-exercise. Urine samples were analyzed for targeted mango-related metabolites. The blood samples were analyzed for 67 oxylipins, which are upstream regulators of inflammation and other physiological processes. After 2 weeks of mango ingestion, three targeted urine mango-related phenolic metabolites were significantly elevated compared to water alone (interaction effects, p ≤ 0.003). Significant post-exercise increases were measured for 49 oxylipins, but various subgroup analyses showed no differences in the pattern of change between trials (all interaction effects, p > 0.150). The 2.25 h cycling bouts induced significant inflammation, but no countermeasure effect was found after 2 weeks of mango ingestion despite the elevation of mango gut-derived phenolic metabolites.

Analytical methods for characterizing magnetic resonance probes.

The efficiency of Gd(III) contrast agents in magnetic resonance image enhancement is governed by a set of tunable structural parameters. Understanding and measuring these parameters requires specific analytical techniques. This Feature describes strategies to optimize each of the critical Gd(III) relaxation parameters for molecular imaging applications and the methods employed for their evaluation.

Reporter protein-targeted probes for magnetic resonance imaging.

Contrast agents for magnetic resonance imaging are frequently employed as experimental and clinical probes. Drawbacks include low signal sensitivity, fast clearance, and nonspecificity that limit efficacy in experimental imaging. In order to create a bioresponsive MR contrast agent, a series of four Gd(III) complexes targeted to the HaloTag reporter were designed and synthesized. HaloTag is unique among reporter proteins for its specificity, versatility, and the covalent interaction between substrate and protein. In similar systems, these properties produce prolonged in vivo lifetimes and extended imaging opportunities for contrast agents, longer rotational correlation times, and increases in relaxivity (r(1)) upon binding to the HaloTag protein. In this work we report a new MR contrast probe, 2CHTGd, which forms a covalent bond with its target protein and results in a dramatic increase in sensitivity. A 6-fold increase in r(1), from 3.8 to 22 mM(-1) s(-1), is observed upon 2CHTGd binding to the target protein. This probe was designed for use with the HaloTag protein system which allows for a variety of substrates (specific for MRI, florescence, or protein purification applications) to be used with the same reporter.

The berry health tool chest - an evidence map and interactive resource.

Berry consumption is linked to diverse health benefits, but numerous questions remain regarding mechanism of action, dose efficacy, and optimal duration and frequency of intake. Addressing these outstanding questions requires an organized assessment of current research, to inform future study designs and fill critical knowledge gaps. Tools that organize such information will also facilitate consumer messaging, targeted nutritional health initiatives, and dietary intake guidelines. This review aimed to describe the development and utility of the "Berry Health Tool Chest," an evidence map summarizing trial design features of studies characterizing the impact of berry consumption upon human health biomarkers. A systematic search strategy identified relevant high-quality human feeding studies, whose study design parameters were collected and compiled into an evidence map that is freely available as an interactive online interface enabling tabulated data to be interrogated, filtered, and exported. Of the 231 included studies, approximately 70% were of less than 3 months' duration and/or fewer than 50 participants, illustrating research gaps that could potentially inform the design of future studies.

Variation in anthocyanin profiles of 27 genotypes of red cabbage over two growing seasons.

Acylated anthocyanins, such as those found in red cabbage, are more heat-, light-, and alkaline pH-stable than non-acylated anthocyanins, making them attractive for a variety of commercial applications. A UPLC-DAD-MSE method with an optimized chromatographic strategy was used to identify 29 red cabbage anthocyanins, predominantly acylated and glucosylated cyanidin derivatives. Anthocyanin profiles of 27 red cabbage genotypes harvested in consecutive growing seasons were measured and assessed for variation. Three unique anthocyanin profile fingerprints were identified through hierarchical clustering analysis. PCA analysis identified anthocyanin accumulation traits and genotypes with high diversity which can be utilized in future investigations into the genetic and molecular basis for anthocyanin production, acylation, and diversity.

Increased Plasma Levels of Gut-Derived Phenolics Linked to Walking and Running Following Two Weeks of Flavonoid Supplementation.

Using a randomized, double-blinded, placebo-controlled, parallel group design, this investigation determined if the combination of two weeks of flavonoid supplementation (329 mg/day, quercetin, anthocyanins, flavan-3-ols mixture) and a 45-minute walking bout (62.2 ± 0.9% VO2max (maximal oxygen consumption rate)) enhanced the translocation of gut-derived phenolics into circulation in a group of walkers (n = 77). The walkers (flavonoid, placebo groups) were randomized to either sit or walk briskly on treadmills for 45 min (thus, four groups: placebo⁻sit, placebo⁻walk, flavonoid⁻sit, flavonoid⁻walk). A comparator group of runners (n = 19) ingested a double flavonoid dose for two weeks (658 mg/day) and ran for 2.5 h (69.2 ± 1.2% VO2max). Four blood samples were collected (pre- and post-supplementation, immediately post- and 24 h post-exercise/rest). Of the 76 metabolites detected in this targeted analysis, 15 increased after the 2.5 h run, and when grouped were also elevated post-exercise (versus placebo⁻sit) for the placebo⁻ and flavonoid⁻walking groups (p < 0.05). A secondary analysis showed that pre-study plasma concentrations of gut-derived phenolics in the runners were 40% higher compared to walkers (p = 0.031). These data indicate that acute exercise bouts (brisk walking, intensive running) are linked to an increased translocation of gut-derived phenolics into circulation, an effect that is amplified when combined with a two-week period of increased flavonoid intake or chronic training as a runner.