Differential temporal release and lipoprotein loading in B. thetaiotaomicron bacterial extracellular vesicles.

Bacterial extracellular vesicles (BEVs) contribute to stress responses, quorum sensing, biofilm formation and interspecies and interkingdom communication. However, the factors that regulate their release and heterogeneity are not well understood. We set out to investigate these factors in the common gut commensal Bacteroides thetaiotaomicron by studying BEV release throughout their growth cycle.

Investigating Antibody Reactivity to the Intestinal Microbiome in Severe Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS): A Feasibility Study.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a multisystemic disease of unknown aetiology that is characterised by disabling chronic fatigue and involves both the immune and gastrointestinal (GI) systems. Patients display alterations in GI microbiome with a significant proportion experiencing GI discomfort and pain and elevated blood biomarkers for altered intestinal permeability compared with healthy individuals. To investigate a possible GI origin of ME/CFS we designed a feasibility study to test the hypothesis that ME/CFS pathogenesis is a consequence of increased intestinal permeability that results in microbial translocation and a breakdown in immune tolerance leading to generation of antibodies reactive to indigenous intestinal microbes.

Benchtop (60 MHz) proton NMR spectroscopy for quantification of 16-O-methylcafestol in lipophilic extracts of ground roast coffee.

We present a method for analysing the lipophilic fraction extracted from ground coffee beans using 60 MHz proton (H) NMR spectroscopy. In addition to the triglycerides from coffee oil, spectral features are seen from a range of secondary metabolites, such as various diterpenes. We demonstrate quantitation of a peak attributed to one such compound, 16-O-methylcafestol (16-OMC), which is of interest as a coffee species marker.

Regulation of Enteroendocrine Cell Networks by the Major Human Gut Symbiont .

Gut microbes have critical roles in maintaining host physiology, but their effects on epithelial chemosensory enteroendocrine cells (EEC) remain unclear. We investigated the role that the ubiquitous commensal gut bacterium (Bt) and its major fermentation products, acetate, propionate, and succinate (APS) have in shaping EEC networks in the murine gastrointestinal tract (GIT). The distribution and numbers of EEC populations were assessed in tissues along the GIT by fluorescent immunohistochemistry in specific pathogen free (SPF), germfree (GF) mice, GF mice conventionalized by Bt or (Lr), and GF mice administered APS.

High-throughput screening of argan oil composition and authenticity using benchtop H NMR.

We use 60-MHz benchtop nuclear magnetic resonance (NMR) to acquire H spectra from argan oils of assured origin. We show that the low-field NMR spectrum of neat oil contains sufficient information to make estimates of compositional parameters and to inform on the presence of minor compounds. A screening method for quality and authenticity is presented based on nearest-neighbour outlier detection.

Sparse Epistatic Patterns in the Evolution of Terpene Synthases.

We explore sequence determinants of enzyme activity and specificity in a major enzyme family of terpene synthases. Most enzymes in this family catalyze reactions that produce cyclic terpenes-complex hydrocarbons widely used by plants and insects in diverse biological processes such as defense, communication, and symbiosis. To analyze the molecular mechanisms of emergence of terpene cyclization, we have carried out in-depth examination of mutational space around (E)-β-farnesene synthase, an Artemisia annua enzyme which catalyzes production of a linear hydrocarbon chain.

Transcriptional changes in prostate of men on active surveillance after a 12-mo glucoraphanin-rich broccoli intervention-results from the Effect of Sulforaphane on prostate CAncer PrEvention (ESCAPE) randomized controlled trial.

Background: Epidemiological evidence suggests that consumption of cruciferous vegetables is associated with reduced risk of prostate cancer progression, largely attributed to the biological activity of glucosinolate degradation products, such as sulforaphane derived from glucoraphanin. Because there are few therapeutic interventions for men on active surveillance for prostate cancer to reduce the risk of cancer progression, dietary approaches are an appealing option for patients.

Objective: We evaluated whether consumption of a glucoraphanin-rich broccoli soup for 1 y leads to changes in gene expression in prostate tissue of men with localized prostate cancer.

Lack of acute or chronic effects of epicatechin-rich and procyanidin-rich apple extracts on blood pressure and cardiometabolic biomarkers in adults with moderately elevated blood pressure: a randomized, placebo-controlled crossover trial.

Background: The reported effects of flavanol-rich foods such as cocoa, dark chocolate, and apples on blood pressure and endothelial function may be due to the monomeric flavanols [mainly (-)-epicatechin (EC)], the oligomeric flavanols [procyanidins (PCs)], or other components. Reports of well-controlled intervention studies that test the effects of isolated oligomeric flavanols on biomarkers of cardiovascular health are lacking.

Objective: We studied the acute and chronic effects of an EC-rich apple flavanol extract and isolated apple PCs on systolic blood pressure (BP) and other cardiometabolic biomarkers.

Complementary intestinal mucosa and microbiota responses to caloric restriction.

The intestine is key for nutrient absorption and for interactions between the microbiota and its host. Therefore, the intestinal response to caloric restriction (CR) is thought to be more complex than that of any other organ. Submitting mice to 25% CR during 14 days induced a polarization of duodenum mucosa cell gene expression characterised by upregulation, and downregulation of the metabolic and immune/inflammatory pathways, respectively.

Microbial taxonomic and metabolic alterations during faecal microbiota transplantation to treat Clostridium difficile infection.

Objectives: This study aimed to examine changes to the microbiota composition and metabolic profiles of seven patients with recurrent Clostridium difficile infection (rCDI), following treatment with faecal microbiota transplant (FMT).

Methods: 16S rDNA sequencing and H NMR were performed on faecal samples from the patients (pre-, post-FMT, and follow-up) and the associated donor samples. Sparse partial-least-square analysis was used to identify correlations between the two datasets.

16-O-methylcafestol is present in ground roast Arabica coffees: Implications for authenticity testing.

High-field and low-field proton NMR spectroscopy were used to analyse lipophilic extracts from ground roast coffees. Using a sample preparation method that produced concentrated extracts, a small marker peak at 3.16 ppm was observed in 30 Arabica coffees of assured origin.

Increased transcriptional and metabolic capacity for lipid metabolism in the peripheral zone of the prostate may underpin its increased susceptibility to cancer.

The human prostate gland comprises three distinct anatomical glandular zones, namely the peripheral, central and transitional zones. Although prostate cancer can arise throughout the prostate, it is more frequent in the peripheral zone. In contrast, hyperplasia occurs most frequently in the transitional zone.

Intestinal PPARγ signalling is required for sympathetic nervous system activation in response to caloric restriction.

Nuclear receptor PPARγ has been proven to affect metabolism in multiple tissues, and has received considerable attention for its involvement in colon cancer and inflammatory disease. However, its role in intestinal metabolism has been largely ignored. To investigate this potential aspect of PPARγ function, we submitted intestinal epithelium-specific PPARγ knockout mice (iePPARγKO) to a two-week period of 25% caloric restriction (CR), following which iePPARγKO mice retained more fat than their wild type littermates.

Low-field (1)H NMR spectroscopy for distinguishing between arabica and robusta ground roast coffees.

This work reports a new screening protocol for addressing issues of coffee authenticity using low-field (60MHz) bench-top (1)H NMR spectroscopy. Using a simple chloroform-based extraction, useful spectra were obtained from the lipophilic fraction of ground roast coffees. It was found that 16-O-methylcafestol (16-OMC, a recognized marker compound for robusta beans) gives rise to an isolated peak in the 60MHz spectrum, which can be used as an indicator of the presence of robusta beans in the sample.

Can we trust untargeted metabolomics? Results of the metabo-ring initiative, a large-scale, multi-instrument inter-laboratory study.

The metabo-ring initiative brought together five nuclear magnetic resonance instruments (NMR) and 11 different mass spectrometers with the objective of assessing the reliability of untargeted metabolomics approaches in obtaining comparable metabolomics profiles. This was estimated by measuring the proportion of common spectral information extracted from the different LCMS and NMR platforms. Biological samples obtained from 2 different conditions were analysed by the partners using their own in-house protocols.

Emergence of terpene cyclization in Artemisia annua.

The emergence of terpene cyclization was critical to the evolutionary expansion of chemical diversity yet remains unexplored. Here we report the first discovery of an epistatic network of residues that controls the onset of terpene cyclization in Artemisia annua. We begin with amorpha-4,11-diene synthase (ADS) and (E)-β-farnesene synthase (BFS), a pair of terpene synthases that produce cyclic or linear terpenes, respectively.

How much is too much? Threshold dose distributions for 5 food allergens.

Background: Precautionary labeling is used to warn consumers of the presence of unintended allergens, but the lack of agreed allergen thresholds can result in confusion and risk taking by patients with food allergy. The lack of data on threshold doses below which subjects are unlikely to react is preventing the development of evidence-based allergen management strategies that are understood by clinician and patient alike.

Objective: We sought to define threshold dose distributions for 5 major allergenic foods in the European population.

Authentication of beef versus horse meat using 60 MHz 1H NMR spectroscopy.

This work reports a candidate screening protocol to distinguish beef from horse meat based upon comparison of triglyceride signatures obtained by 60 MHz (1)H NMR spectroscopy. Using a simple chloroform-based extraction, we obtained classic low-field triglyceride spectra from typically a 10 min acquisition time. Peak integration was sufficient to differentiate samples of fresh beef (76 extractions) and horse (62 extractions) using Naïve Bayes classification.

60 MHz H NMR spectroscopy for the analysis of edible oils.

We report the first results from a new 60 MHz H nuclear magnetic resonance (NMR) bench-top spectrometer, Pulsar, in a study simulating the adulteration of olive oil with hazelnut oil. There were qualitative differences between spectra from the two oil types. A single internal ratio of two isolated groups of peaks could detect hazelnut oil in olive oil at the level of ∼13%w/w, whereas a whole-spectrum chemometric approach brought the limit of detection down to 11.

A diet rich in high-glucoraphanin broccoli interacts with genotype to reduce discordance in plasma metabolite profiles by modulating mitochondrial function.

Background: Observational and experimental studies suggest that diets rich in cruciferous vegetables and glucosinolates may reduce the risk of cancer and cardiovascular disease (CVD).

Objective: We tested the hypothesis that a 12-wk dietary intervention with high-glucoraphanin (HG) broccoli would modify biomarkers of CVD risk and plasma metabolite profiles to a greater extent than interventions with standard broccoli or peas.

Design: Subjects were randomly assigned to consume 400 g standard broccoli, 400 g HG broccoli, or 400 g peas each week for 12 wk, with no other dietary restrictions.

Metabolomics-on-a-chip and predictive systems toxicology in microfluidic bioartificial organs.

The world faces complex challenges for chemical hazard assessment. Microfluidic bioartificial organs enable the spatial and temporal control of cell growth and biochemistry, critical for organ-specific metabolic functions and particularly relevant to testing the metabolic dose-response signatures associated with both pharmaceutical and environmental toxicity. Here we present an approach combining a microfluidic system with (1)H NMR-based metabolomic footprinting, as a high-throughput small-molecule screening approach.

Quantitative assessment of multi-laboratory reproducibility of SDS-PAGE assays: Digestion pattern of beta-casein and beta-lactoglobulin under simulated conditions.

A digestion protocol was applied in triplicate by ten laboratories, simulating in vivo gastric and duodenal conditions. The intra- and inter-laboratory variability in the kinetics of protein degradation was quantified, focussing on the digestion of beta-casein under gastric conditions, and of beta-lactoglobulin (beta-Lg) under duodenal conditions. The addition of surfactants such as phosphatidylcholine (PC) in the digestion mix was also evaluated.