Exploring and disentangling the production of potentially bioactive phenolic catabolites from dietary (poly)phenols, phenylalanine, tyrosine and catecholamines.

Following ingestion of fruits, vegetables and derived products, (poly)phenols that are not absorbed in the upper gastrointestinal tract pass to the colon, where they undergo microbiota-mediated ring fission resulting in the production of a diversity of low molecular weight phenolic catabolites, which appear in the circulatory system and are excreted in urine along with their phase II metabolites. There is increasing interest in these catabolites because of their potential bioactivity and their use as biomarkers of (poly)phenol intake. Investigating the fate of dietary (poly)phenolics in the colon has become confounded as a result of the recent realisation that many of the phenolics appearing in biofluids can also be derived from the aromatic amino acids, l-phenylalanine and l-tyrosine, and to a lesser extent catecholamines, in reactions that can be catalysed by both colonic microbiota and endogenous mammalian enzymes. The available evidence, albeit currently rather limited, indicates that substantial amounts of phenolic catabolites originate from phenylalanine and tyrosine, while somewhat smaller quantities are produced from dietary (poly)phenols. This review outlines information on this topic and assesses procedures that can be used to help distinguish between phenolics originating from dietary (poly)phenols, the two aromatic amino acids and catecholamines.

A critical examination of human data for the biological activity of quercetin and its phase-2 conjugates.

This critical review examines evidence for beneficial effects of quercetin phase-2 conjugates from clinical intervention studies, volunteer feeding trials, and in vitro work. Plasma concentrations of quercetin-3-O-glucuronide (Q3G) and 3'-methylquercetin-3-O-glucuronide (3'MQ3G) after supplementation may produce beneficial effects in macrophages and endothelial cells, respectively, especially if endogenous deglucuronidation occurs, and lower blood uric acid concentration via quercetin-3'-O-sulfate (Q3'S). Unsupplemented diets produce much lower concentrations (<50 nmol/l) rarely investigated in vitro. At 10 nmol/l, Q3'S and Q3G stimulate or suppress, respectively, angiogenesis in endothelial cells. Statistically significant effects have been reported at 100 nmol/l in breast cancer cells (Q3G), primary neuron cultures (Q3G), lymphocytes (Q3G and3'MQ3G) and HUVECs (QG/QS mixture), but it is unclear whether these translate to a health benefit in vivo. More sensitive and more precise methods to measure clinically significant endpoints are required before a conclusion can be drawn regarding effects at normal dietary concentrations. Future requirements include better understanding of inter-individual and temporal variation in plasma quercetin phase-2 conjugates, their mechanisms of action including deglucuronidation and desulfation both in vitro and in vivo, tissue accumulation and washout, as well as potential for synergy or antagonism with other quercetin metabolites and metabolites of other dietary phytochemicals.

A Systematic Review and Comprehensive Evaluation of Human Intervention Studies to Unravel the Bioavailability of Hydroxycinnamic Acids.

Significance: Hydroxycinnamic acids (HCAs) are the main phenolic acids in the western diet. Harmonizing the available information on the absorption, distribution, metabolism, and excretion (ADME) of HCAs is fundamental to unraveling the compounds responsible for their health effects. This work systematically assessed pharmacokinetics, including urinary recovery, and bioavailability of HCAs and their metabolites, based on literature reports. Recent Advances: Forty-seven intervention studies with coffee, berries, herbs, cereals, tomato, orange, grape products, and pure compounds, as well as other sources yielding HCA metabolites, were included. Up to 105 HCA metabolites were collected, mainly acyl-quinic and C6-C3 cinnamic acids. C6-C3 cinnamic acids, such as caffeic and ferulic acid, reached the highest blood concentrations (maximum plasma concentration [Cmax] = 423 nM), with time to reach Cmax (Tmax) values ranging from 2.7 to 4.2 h. These compounds were excreted in urine in higher amounts than their phenylpropanoic acid derivatives (4% and 1% of intake, respectively), but both in a lower percentage than hydroxybenzene catabolites (11%). Data accounted for 16 and 18 main urinary and blood HCA metabolites, which were moderately bioavailable in humans (collectively 25%). Critical Issues: A relevant variability emerged. It was not possible to unequivocally assess the bioavailability of HCAs from each ingested source, and data from some plant based-foods were absent or inconsistent. Future Directions: A comprehensive study investigating the ADME of HCAs derived from their most important dietary sources is urgently required. Eight key metabolites were identified and reached interesting plasma Cmax concentrations and urinary recoveries, opening up new perspectives to evaluate their bioactivity at physiological concentrations. Antioxid. Redox Signal. 40, 510-541.

Assessment of human inter-individual variability of phloretin metabolites in urine after apple consumption. AppleCOR study.

Purpose: This study aimed to assess the inter-individual variation in phloretin absorption and metabolism and to seek possible phloretin metabotypes following apple snack consumption. Methods: The excreted phloretin metabolites in 24 h urine samples were determined by UPLC-MS/MS in 62 volunteers after acute and sustained (6 weeks) interventions in a randomized and parallel study with a daily supplementation of 80 g of a low-phloretin (39.5 µmol) or a high-phloretin (103 µmol) freeze-dried apple snacks. Results: absorption estimated as phloridzin equivalents for 62 volunteers varied almost 70-fold ranging from 0.1% to 6.94% of phloretin glycoside intake. Volunteers were stratified into low, medium and high producers and by the balance between glucuronidation and sulphation. For 74% of the volunteers phloretin-O-glucuronide was the dominant urinary metabolite, especially at the higher phloretin glycoside intake and for higher producers. Sulphate conjugation assumed greater significance for the remaining volunteers especially for low producers. Females dominated glucuronide profile (64.1%) and males dominated the low excretion group. Analysis of plasma glucose and insulin at the start and end of the sustained study showed a trend towards modest reductions for high producers. Furthermore, plausible factors contributing to the inter-individual variation in phloretin uptake are discussed. Conclusions: extensive inter-individual variability exists in the excretion of phloretin phase-II conjugates following consumption of apple snacks, which could be related to oral microbiota phloridzin-hydrolysing activity, lactase non-persistence trait or the metabotype to which the subject belongs. There were inconsistent effects on post-prandial serum glucose concentrations but there was a tendency for decreases to be associated with higher excretion of phloretin phase-II conjugates. Trial registration: The acute and sustained studies were registered at ClinicalTrials.gov Identifier: NCT03795324.

Excretion by subjects on a low (poly)phenol diet of phenolic gut microbiota catabolites sequestered in tissues or associated with catecholamines and surplus amino acids.

Phenolic catabolites excreted by fasting subjects with a functioning colon and ileostomists on a low (poly)phenol diet have been investigated. Urine was collected over a 12 h fasting period after adherence to a low (poly)phenol diet for 36 h. UHPLC-HR-MS quantified 77 phenolics. Some were present in the urine of both groups in similar trace amounts and others were excreted in higher amounts by participants with a colon indicating the involvement of the microbiota. Most were present in sub- or low-µmol amounts, but hippuric acid dominated accounting on average for 60% of the total for both volunteer categories indicating significant production from sources other than non-nutrient dietary (poly)phenols. The potential origins of the phenolics associated with the low (poly)phenol diet, include endogenous catecholamines, surplus tyrosine and phenylalanine, and washout of catabolites derived from pre-study intakes of non-nutrient dietary (poly)phenols.

Human colonic catabolism of dietary flavan-3-ol bioactives.

Understanding the fate of ingested polyphenols is crucial in elucidating the molecular mechanisms underlying the beneficial effects of a fruit and vegetable-based diet. This review focuses on the colon microbiota-mediated transformation of the flavan-3-ols and the structurally related procyanidins found in dietary plant foods and beverages, plus the flavan-3-ol-derived theaflavins of black tea, and the post-absorption phase II metabolism of the gut microbiota catabolites. Despite significant advances in the last decade major analytical challenges remain. Strategies to address them are presented.

Metabolism of phenolics in coffee and plant-based foods by canonical pathways: an assessment of the role of fatty acid ß-oxidation to generate biologically-active and -inactive intermediates.

ω-Phenyl-alkenoic acids are abundant in coffee, fruits, and vegetables. Along with ω-phenyl-alkanoic acids, they are produced from numerous dietary (poly)phenols and aromatic amino acids in vivo. This review addresses how phenyl-ring substitution and flux modulates their gut microbiota and endogenous ß-oxidation. 3',5'-Dihydroxy-derivatives (from alkyl-resorcinols, flavanols, proanthocyanidins), and 4'-hydroxy-phenolic acids (from tyrosine, p-coumaric acid, naringenin) are ß-oxidation substrates yielding benzoic acids. In contrast, 3',4',5'-tri-substituted-derivatives, 3',4'-dihydroxy-derivatives and 3'-methoxy-4'-hydroxy-derivatives (from coffee, tea, cereals, many fruits and vegetables) are poor ß-oxidation substrates with metabolism diverted via gut microbiota dehydroxylation, phenylvalerolactone formation and phase-2 conjugation, possibly a strategy to conserve limited pools of coenzyme A. 4'-Methoxy-derivatives (citrus fruits) or 3',4'-dimethoxy-derivatives (coffee) are susceptible to hepatic "reverse" hydrogenation suggesting incompatibility with enoyl-CoA-hydratase. Gut microbiota-produced 3'-hydroxy-4'-methoxy-derivatives (citrus fruits) and 3'-hydroxy-derivatives (numerous (poly)phenols) are excreted as the phenyl-hydracrylic acid ß-oxidation intermediate suggesting incompatibility with hydroxy-acyl-CoA dehydrogenase, albeit with considerable inter-individual variation. Further investigation is required to explain inter-individual variation, factors determining the amino acid to which C6-C3 and C6-C1 metabolites are conjugated, the precise role(s) of l-carnitine, whether glycine might be limiting, and whether phenolic acid-modulation of ß-oxidation explains how phenolic acids affect key metabolic conditions, such as fatty liver, carbohydrate metabolism and insulin resistance.

LC-MS Characterization and Quantification of Known and Unknown (Poly)phenol Metabolites-Possible Pitfalls and Their Avoidance.

This review focuses on the LC-MS characterization and quantification of dietary (poly)phenols and their metabolites. It draws attention to errors, omissions, and misunderstandings that appear frequently in published papers, and suggests strategies for their avoidance. Aspects covered include the use of authentic standards and surrogate reference materials, the importance of collecting and archiving Total Ion Current MS data, the limitations of using on-line compilations of accurate mass MS data to assign unknown components when multiple isomers are possible, and the often understated magnitude of person-to-person variation that may significantly impact at population level any potential health benefit.

A Practitioner's Dilemma Mass Spectrometry-Based Annotation and Identification of Human Plasma and Urinary Polyphenol Metabolites.

The practitioner's dilemma in metabolite assignment can be described as follows: for compound and metabolite identification, strict guidelines should be followed using authentic standards only, or uncertainties in structure assignment of compounds with the certainty of consequential errors should be accepted. These uncertainties arise due to limitation of software and databases in combination with the complexity of the human body fluid samples.

In Vitro Faecal Fermentation of Monomeric and Oligomeric Flavan-3-ols: Catabolic Pathways and Stoichiometry.

SCOPE: The study evaluates the influence of flavan-3-ol structure on the production of phenolic catabolites, principally phenyl-γ-valerolactones (PVLs), and phenylvaleric acids (PVAs). METHODS AND RESULTS: A set of 12 monomeric flavan-3-ols and proanthocyanidins (degree of polymerization (DP) of 2-5), are fermented in vitro for 24 h using human faecal microbiota, and catabolism is analyzed by UHPLC-ESI-MS/MS. Up to 32 catabolites strictly related to microbial catabolism of parent compounds are detected. (+)-Catechin and (-)-epicatechin have the highest molar mass recoveries, expressed as a percentage with respect to the incubated concentration (75 µmol L-1 ) of the parent compound, for total PVLs and PVAs, both at 5 h (about 20%) and 24 h (about 40%) of faecal incubation. Only A-type dimer and B-type procyanidins underwent the ring fission step, and no differences are found in total PVL and PVA production (≃1.5% and 6.0% at 5 and 24 h faecal incubation, respectively) despite the different DPs. CONCLUSION: The flavan-3-ol structure strongly affects the colonic catabolism of the native compounds, influencing the profile of PVLs and PVAs produced in vitro. This study opens new perspectives to further elucidate the colonic fate of oligomeric flavan-3-ols and their availability in producing bioactive catabolites.

In vivo study of the bioavailability and metabolic profile of (poly)phenols after sous-vide artichoke consumption.

Artichokes are a rich source of (poly)phenols, mainly caffeoylquinic acids, but little is known about their bioavailability from this source. This study investigated the absorption, metabolism and excretion of (poly)phenols after sous-vide artichoke consumption (5776 µmol of (poly)phenols) by healthy volunteers. Seventy-six (poly)phenol metabolites were identified by UHPLC-MS/MS using authentic standards, including acyl-quinic acids plus C6-C3, C6-C1, C6-C2, C6-C1-N, C6-C0 metabolites, and their phase-II conjugates. The major metabolites were 3'-methoxy-4'-hydroxycinnamic acid, 3'-methoxycinnamic acid-4'-sulfate, and 4'-hydroxycinnamic acid-3'-sulfate, which appeared early in plasma (Tmax < 4 h); plus 3-(3'-methoxy-4'-hydroxyphenyl)propanoic acid, 3-(4'-methoxyphenyl)propanoic acid-3'-glucuronide, 3-(3'-hydroxyphenyl) propanoic acid and hippuric acids, which appeared later (Tmax > 6 h). The 24 h urinary recovery averaged 8.9% (molar basis) of the (poly)phenols consumed. Hepatic beta-oxidation of 3',4'-dihydroxycinnamic acid and methylated conjugates occurred, but was limited (<0.04%). 3'-Methylation exceeded 4'-methylation and interindividual variability was high, especially for gut microbial metabolites (up to 168-fold).

Variation in the Methylation of Caffeoylquinic Acids and Urinary Excretion of 3'-methoxycinnamic acid-4'-Sulfate After Apple Consumption by Volunteers.

INTRODUCTION: It has been reported that the phenolic metabolite 3'-methoxycinnamic acid-4'-sulfate generated from 5-O-caffeoylquinic acid may have potential benefits in human health. However, the variation in 3'- and 4'-methylation of 3',4'-dihydroxycinnamic acid and its impact on the yield of this sulfate metabolite is unclear and has been poorly studied. METHODS AND RESULTS: To address this aim, the excreted 3'-methoxy and 4'-methoxy metabolites in urine samples (24-h) are determined in 14 volunteers after an acute intake of 80 g of red-fleshed apple (RFA) or white-fleshed apple (WFA). These methoxy metabolites are also determined in the same volunteers in a second acute intake after a 6-week sustained consumption of the same products. CONCLUSION: Seven 3'-methoxy and seven 4'-methoxy metabolites are determined, i.e., the free cinnamic and corresponding phenylpropanoic acid, plus their sulfate, glucuronide, and glycine conjugates. In only six volunteers, five females and one male, is 4'-methylation preferred over 3'-methylation, but it is observed that an individual's 3'- : 4'-methylation ratio can change over time, and that the yield of 3'-methoxycinnamic acid-4'-sulfate is extremely variable, ranging from undetectable to 71% of the total C6 -C3 metabolites excreted, and any benefit accruing from this metabolite will not necessarily be available to all consumers.

Bioavailability and metabolism of chlorogenic acids (acyl-quinic acids) in humans.

Acyl-quinic acids (chlorogenic acids) are produced by many plants, including fruits, vegetables, and herbal remedies, with coffee and maté particularly rich dietary sources. Epidemiological and intervention studies suggest that they can reduce the risk of developing type 2 diabetes and cardiovascular disease. This review addresses their metabolic handling after oral consumption to provide a mechanistic basis to explain their possible effects on health. Intact acyl-quinic acids are absorbed only to a small extent in the small intestine, but the cinnamic acids are efficiently absorbed after hydrolysis by either digestive or microbial enzymes in the colon. Metabolism results in phenolic conjugates in the blood and urine, but varying dependent on the acyl-quinic acid, and subject to significant interperson variability. The balance between hydrogenation and complete ß-oxidation of the cinnamic acids, both by liver and gut microbiota, determines the profile of metabolites. Pharmacokinetic data suggest that some metabolites are bound to human serum albumin and/or sequestered in tissues, and some exhibit biological activity in vitro, consistent with proposed protective action in vivo. Significant gaps in the literature include lack of plasma and urinary data for free-living individuals, and pharmacokinetic data for groups who consume coffee or maté at regular short intervals. Data are required for cis isomers. There is a critical need for precise urinary biomarkers of consumption of acyl-quinic acids, accounting for variability in individual metabolism and in beverage composition, thus facilitating better translation of urinary metabolite measurements into accurate coffee consumption data to improve the outcomes of future epidemiological and intervention studies.

Recommendations for standardizing nomenclature for dietary (poly)phenol catabolites.

There is a lack of focus on the protective health effects of phytochemicals in dietary guidelines. Although a number of chemical libraries and databases contain dietary phytochemicals belonging to the plant metabolome, they are not entirely relevant to human health because many constituents are extensively metabolized within the body following ingestion. This is especially apparent for the highly abundant dietary (poly)phenols, for which the situation is compounded by confusion regarding their bioavailability and metabolism, partially because of the variety of nomenclatures and trivial names used to describe compounds arising from microbial catabolism in the gastrointestinal tract. This confusion, which is perpetuated in online chemical/metabolite databases, will hinder future discovery of bioactivities and affect the establishment of future dietary guidelines if steps are not taken to overcome these issues. In order to resolve this situation, a nomenclature system for phenolic catabolites and their human phase II metabolites is proposed in this article and the basis of its format outlined. Previous names used in the literature are cited along with the recommended nomenclature, International Union of Pure and Applied Chemistry terminology, and, where appropriate, Chemical Abstracts Service numbers, InChIKey, and accurate mass.

Plasma pharmacokinetics of (poly)phenol metabolites and catabolites after ingestion of orange juice by endurance trained men.

The health benefits of orange juice (OJ) consumption are attributed in part to the circulating flavanone phase II metabolites and their microbial-derived ring fission phenolic catabolites. The present study investigated these compounds in the bloodstream after acute intake of 500 mL of OJ. Plasma samples obtained at 0, 1, 2, 3, 4, 5, 6, 7, 8 and 24 h after OJ intake were analysed by HPLC-HR-MS. Eleven flavanone metabolites and 36 phenolic catabolites were identified and quantified in plasma. The main metabolites were hesperetin-3'-sulfate with a peak plasma concentration (Cmax) of 80 nmol/L, followed by hesperetin-7-glucuronide (Cmax 24 nmol/L), hesperetin-3'-glucuronide (Cmax 18 nmol/L) and naringenin-7-glucuronide (Cmax 21 nmol/L). Among the main phenolic catabolites to increase in plasma after OJ consumption were 3'-methoxycinnamic acid-4'-sulfate (Cmax 19 nmol/L), 3-hydroxy-3-(3'-hydroxy-4'-methoxyphenyl)propanoic acid (Cmax 20 nmol/L), 3-(3'-hydroxy-4'-methoxyphenyl)propanoic acid (Cmax 19 nmol/L), 3-(4'-hydroxyphenyl)propanoic acid (Cmax 25 nmol/L), and 3-(phenyl)propanoic acid (Cmax 19 nmol/L), as well as substantial amounts of phenylacetic and hippuric acids. The comprehensive plasma pharmacokinetic profiles that were obtained are of value to the design of future ex vivo cell studies, aimed at elucidating the mechanisms underlying the potential health benefits of OJ consumption. CLINICAL TRIAL REGISTRATION NUMBER: This trial was registered at clinicaltrials.gov as NCT02627547.

Terms and nomenclature used for plant-derived components in nutrition and related research: efforts toward harmonization.

Many terms for plant-derived food components are commonly used in the literature, but there is a notable lack of standardization and definition of nomenclature. The use of terms is often field-specific, leading to misunderstanding and problems with literature searches and systematic reviews, and results in isolated and divided research; this impacts not only publication quality but also innovation, regulatory compliance, and enforcement. To begin to address this issue, this narrative review describes the current use and definition of terms. The terms are either chemical and/or origin-based, such as phytochemical (chemicals from plants), or function-based, such as phytonutrient, bioactive, or nutraceutical. The ultimate goal is to establish a common harmonized, evidence-based understanding for when to use each term, thereby providing clarity and a specific scientific basis for such nomenclature. Neither the quality nor the quantity of evidence needed to allow the use of functional terms such as phytonutrient or nutraceutical is specifically discussed here; rather, it is simply noted that evidence is needed to apply these terms. The next step would be to define the evidence necessary for a compound to have a functional descriptor. The aim in this article is to establish scientific criteria for definitions that could be applied to clearly define and differentiate commonly used terms and thus ensure their consistent application in the scientific literature.

Phenyl-γ-valerolactones and phenylvaleric acids, the main colonic metabolites of flavan-3-ols: synthesis, analysis, bioavailability, and bioactivity.

Covering: 1958 to June 2018 Phenyl-γ-valerolactones (PVLs) and their related phenylvaleric acids (PVAs) are the main metabolites of flavan-3-ols, the major class of flavonoids in the human diet. Despite their presumed importance, these gut microbiota-derived compounds have, to date, in terms of biological activity, been considered subordinate to their parent dietary compounds, the flavan-3-ol monomers and proanthocyanidins. In this review, the role and prospects of PVLs and PVAs as key metabolites in the understanding of the health features of flavan-3-ols have been critically assessed. Among the topics covered, are proposals for a standardised nomenclature for PVLs and PVAs. The formation, bioavailability and pharmacokinetics of PVLs and PVAs from different types of flavan-3-ols are discussed, taking into account in vitro and animal studies, as well as inter-individual differences and the existence of putative flavan-3-ol metabotypes. Synthetic strategies used for the preparation of PVLs are considered and the methodologies for their identification and quantification assessed. Metabolomic approaches unravelling the role of PVLs and PVAs as biomarkers of intake are also described. Finally, the biological activity of these microbial catabolites in different experimental models is summarised. Knowledge gaps and future research are considered in this key area of dietary (poly)phenol research.

Postprandial glycaemic and lipaemic responses to chronic coffee consumption may be modulated by CYP1A2 polymorphisms.

There is much epidemiological evidence suggesting a reduced risk of development of type 2 diabetes (T2D) in habitual coffee drinkers, however to date there have been few longer-term interventions, directly examining the effects of coffee intake on glucose and lipid metabolism. Previous studies may be confounded by inter-individual variation in caffeine metabolism. Specifically, the rs762551 SNP in the CYP1A2 gene has been demonstrated to influence caffeine metabolism, with carriers of the C allele considered to be of a 'slow' metaboliser phenotype. This study investigated the effects of regular coffee intake on markers of glucose and lipid metabolism in coffee-naïve individuals, with novel analysis by rs762551 genotype. Participants were randomised to either a coffee group (n 19) who consumed four cups/d instant coffee for 12 weeks or a control group (n 8) who remained coffee/caffeine free. Venous blood samples were taken pre- and post-intervention. Primary analysis revealed no significant differences between groups. Analysis of the coffee group by genotype revealed several differences. Before coffee intake, the AC genotype ('slow' caffeine metabolisers, n 9) displayed higher baseline glucose and NEFA than the AA genotype ('fast' caffeine metabolisers, n 10, P<0·05). Post-intervention, reduced postprandial glycaemia and reduced NEFA suppression were observed in the AC genotype, with the opposite result observed in the AA genotype (P<0·05). These observed differences between genotypes warrant further investigation and indicate there may be no one-size-fits-all recommendation with regard to coffee drinking and T2D risk.

Chlorogenic acids and the acyl-quinic acids: discovery, biosynthesis, bioavailability and bioactivity.

Covering: 2000 up to late 2017This review is focussed upon the acyl-quinic acids, the most studied group within the ca. 400 chlorogenic acids so far reported. The acyl-quinic acids, the first of which was characterised in 1846, are a diverse group of plant-derived compounds produced principally through esterification of an hydroxycinnamic acid and 1l-(-)-quinic acid. Topics addressed in this review include the confusing nomenclature, quantification and characterisation by NMR and MS, biosynthesis and role in planta, and the occurrence of acyl-quinic acids in coffee, their transformation during roasting and delivery to the beverage. Coffee is the major human dietary source world-wide of acyl-quinic acids and consideration is given to their absorption and metabolism in the upper gastrointestinal tract, and the colon where the microbiota play a key role in the formation of catabolites. Evidence on the potential of the in vivo metabolites and catabolites of acyl-quinic acids to promote the consumer's health is evaluated.