Role of the small intestine, colon and microbiota in determining the metabolic fate of polyphenols

TL;DR: While it is clear that the composition of the human gut microbiota can be modulated in vivo by supplementation with some (poly)phenol‐rich commodities, such modulation is definitely not an inevitable consequence of supplementation and it is not clear whether the modulation is sustained when supplementation ceases.

About: This article is published in Biochemical Pharmacology.The article was published on 2017-09-01 and is currently open access. It has received 227 citations till now.

Summary

1. (Poly)phenols covered and main classes.

• A wide variety of polyphenols are consumed as part of the normal diet [3] , and are also present at high levels in supplements [4] , and form essential components of many Chinese medicines [5] .
• The estimated total polyphenol consumption ranged from 584 mg/day by Greek women to 1786 mg/day for men in Aarhus-Denmark.
• This study defined the major contributors to be the phenolic acids (predominantly the caffeoylquinic acids (27-53%) , also called chlorogenic acids) and flavonoids (predominantly flavanols (16-29%), proanthocyanidins (5-9%) and theaflavins (14-25%)) [6] .
• It has been demonstrated that foods also contain a significant amount of non-extractable (poly)phenols which, nevertheless, are gut microbiota substrates, and these will further raise the (poly)phenol intake [13] .
• Flavanols are present in planta mostly in their free forms, but can be galloylated (as in green tea) or polymerised to form proanthocyanidins, common components of many foods such as cocoa.

2. Absorption into the bloodstream

• Chlorogenic acid is a general term for the esters of a phenolic acid (e.g. ferulic, caffeic or dimethoxycinnamic acids) with quinic acid [16] , and these classes are found at particularly high levels in coffee [17] .
• After reaching the small intestine, some hydrolysis of caffeoylquinic acid and of dimethoxycinnamoyl quinic acid occurs owing to the action of mammalian esterases, but the hydrolysis is relatively slow and only a proportion of the chlorogenic acid is hydrolysed [18] .
• Intact hesperidin is not absorbed passively, but hesperetin, after removal of the two sugars, is readily absorbed [33] .
• The consensus is that (−)-epicatechin-3 -O-glucuronide, (−)-epicatechin-3 -O-sulfate, and a 3 -Omethyl-(−)-epicatechin-5/7-sulfate are the predominant epicatechin metabolites in humans after oral consumption [47, 49] .
• Anthocyanins and proanthocyanidins are very poorly absorbed in the small intestine in their intact forms [50, 51] .

3. Conversion of polyphenols by the gut microbiota

• In the last decade there have been major advances in defining the composition of the gut microbiota with greatly increased use of methods based on the detection and sequencing of 16S rDNA and MALDI-TOF MS replacing conventional culture methods, but these diverse approaches remain complementary with more traditional methods required to determine microbiota functionality [56, 57] .
• The human gastro-intestinal tract hosts up to 100 trillion microbes [58] , which have been assigned to well over 1000 species.
• The unique catabolites include 4hydroxy-mandelic acid from p-sympatol [65] , tyrosols from oleuropein and related compounds [66] , S-equol, 5-hydroxy-equol and dihydrocinnamic acids with the aryl residue at C2 formed from isoflavones [67, 68] , urolithins and nasutins from ellagitannins [69] [70] [71] , diarylbutanes from lignans [32;35;36] , and dihydroresveratrol from piceid and resveratrol [72] .
• The predominant catabolites generated by the microbiota from (poly)phenols are the aromatic and phenolic acids with zero to three aromatic hydroxyls, or their mono-or di-methoxy analogues, possessing also a sidechain of one to five carbons which might bear an aliphatic hydroxyl [64] .
• Catechol and 1-methyl pyrogallol are not found in foods, but are products of catabolism by the gut microbiota.

4. Pharmacokinetic consequences of multiple doses

• (Poly)phenols associated with coffee, green tea and black tea are generally consumed frequently during the waking day, with the result that the gut microbiota have access to several bolus doses for a prolonged period of time.
• This would predispose to a larger and more uniform concentration of the associated catabolites in the gastrointestinal tract and in plasma after absorption, but little attention has been given to how this would affect the levels of (poly)phenols in circulation in human intervention studies.
• It is likely that a similar phenomenon will occur with proanthocyanidins which also bind strongly to proteins, and possibly thearubigins, but because their catabolites are not unique it would be more difficult to demonstrate than with the ellagitannins (see Figs 2 and 5 ).
• Volunteer studies of typical real world repeated consumption would, if available, circumvent the lack of data for intravenous dosing, but in their absence the authors performed a simple additive modelling using real data from a single dose study in order to obtain an indication of the potential concentrations that might be attained.
• This approach assumes that the rate of clearance from plasma does not change with dose, and would only be true if clearance was saturated, and thus the estimate obtained is on the high side.

5. Prebiotic effects

• There have been many studies of the potential of (poly)phenols substrates and catabolites to modulate the composition of the gut microbiota.
• It is also a gut microbiota catabolite of some dietary (poly)phenols and the greater excretion was ascribed provisionally to a greater proportion of Faecalibacterium prausnitzii, Bifidobacterium spp. and Lactobacillus spp. in the gut microbiota of the healthy metabotype both before and after red wine consumption [118] .
• Studies on volunteers must be viewed as the gold standard but they are susceptible to several confounding factors, such as faulty recall when food frequency questionnaires are used, or non-compliance when supplements are provided, and to substantial inter-individual variation.
• One must accept that human faecal samples do not truly represent the microbiota composition or the metabolic competence of the gastrointestinal tract from which it was voided.
• In view of the very large number of discrete species, and the recognised potential for subtle variations in metabolic competence of even closely related species, it is almost certain that there is more than one organism capable of effecting any particular transformation, and probably more than one route connecting substrate to any catabolite.

6. Effects of catabolites

• In vitro cell culture studies are probably the most effective way to investigate the effects of gut microflora catabolites because when effects are observed there is the potential also to investigate the underlying mechanism(s).
• The chosen biomarker(s) must be biologically relevant, but more importantly, those selected should be measurable with precision.
• Because multiple catabolites are present simultaneously, the total exposure in the plasma might realistically be some 2-3-fold higher and in the colon in excess of 1 mM.
• A simple comparison of plasma and colon concentrations with concentrations shown to be effective in vitro suggests that there is potential for protocatechuic acid, vanillic acid, gallic acid and 3 ,4 -dihydroxyphenylacetic acid to exert modest effects in vivo in at least some volunteers on real-world diets.
• There has been limited investigation of mixtures of (poly)phenolic catabolites.

7. Summary, conclusions and recommendations for future research

• The colonic microbiota transform a very complex range of (poly)phenol substrates with coffee chlorogenic acids, black tea theaflavins and thearubigins often dominating, with substantial contributions from proanthocyanidins and flavanols, flavonols, flavanones and anthocyanin glycosides, and unextractable (poly)phenols, from many fruit and vegetables.
• There is growing evidence from in vitro and intervention studies that some of these catabolites are biologically active, and that, along with untransformed substrates, may function as prebiotics capable of modulating the human gut microbiota composition.
• There is also a pressing need for investigation of free-living volunteers who regularly consume (poly)phenol-rich beverages at frequent intervals to better define the circulating catabolite profiles after consumption of multiple doses.
• Hesperetin after absorption is found as glucuronidated and sulfated conjugates, but here enzymic deconjugation was used to convert them back to hesperetin aglycone.
• HUVEC, human umbilical cord endothelial cell Figure legends.

Figures

Table 1: Comparison of plasma and colon catabolite concentrations in volunteers compared to concentrations shown to be effective in in vitro

Figure 8: Possible catabolic routes from hesperetin to hydroxyphenylhydracrylic acids.

Figure 6. Appearance of microbial metabolites in the plasma and urine from volunteers (n=22) after consumption of mixed berry fruits. Top panel: Plasma catechol-O-sulfate (ﾐ) and 1-methyl-pyrogallol-O-sulfate (ズ). Lower panel: Urinary catechol-O-sulfate. Redrawn from [87] and [86]. *, p < 0.05; **, p < 0.01 relative to time zero. Relative quantification is based on peak area corrected for the internal standard.

Figure 7. Measured concentration of 3胡-methoxy-4胡-hydroxy-phenylpropionic acid (DHFA) in plasma after consumption of one cup of coffee (ズ), or calculated after seven cups, one every 120 min (ﾐ). Data on single dose taken from [20] and is the DHFA plasma concentration after consumption of a cup of coffee (3.4 g of instant coffee in 200 ml water) containing 412 mg total chlorogenic acid. Effect of seven doses consumed every 120 min was calculated using simple addition of each time point from the single dose data, without taking into account any change in plasma clearance as a function of plasma concentration.

Figure 2: Diagrammatic illustration of the conversion of three classes of dominant dietary (poly)phenols to a series of common catabolites

Frequently asked questions

Q1. What have the authors contributed in "Role of the small intestine, colon and microbiota in determining the metabolic fate of polyphenols" ?

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Q2. What have the authors stated for future works in "Role of the small intestine, colon and microbiota in determining the metabolic fate of polyphenols" ?

Use of physiologically based kinetic ( PBK ) modeling to study interindividual human variation and species differences in plasma concentrations of quercetin and its metabolites, Biochem. Food Chem. 59 ( 6 ) ( 2011 ) 2241-2247. [ 176 ] K. M. Keane, P. G. Bell, J. K. Lodge, C. L. Constantinou, S. E. Jenkinson, R. Bass, G. Howatson, Phytochemical uptake following human consumption of Montmorency tart cherry ( L. Prunus cerasus ) and influence of phenolic acids on vascular smooth muscle cells in vitro, Eur. J. Nutr. 55 ( 4 ) ( 2016 ) 1695-1705. [ 177 ] M. Hidalgo, S. Martin-Santamaria, I. Recio, C. Sanchez-Moreno, B. de Pascual-Teresa, G. Rimbach, S. de Pascual-Teresa, Potential anti-inflammatory, anti-adhesive, anti/estrogenic, and angiotensin-converting enzyme inhibitory activities of anthocyanins and their gut metabolites, Genes Nutr. 7 ( 2 ) ( 2012 ) 295-306. [ 178 ] M. Tognolini, C. Giorgio, M. Hassan, I, E. Barocelli, L. Calani, E. Reynaud, O. Dangles, G. Borges, A. Crozier, F. Brighenti, R. D. Del, Perturbation of the EphA2-EphrinA1 system in human prostate cancer cells by colonic ( poly ) phenol catabolites, J. Agric. The catabolites above might be absorbed and subject to mammalian metabolism ( glucuronidation, sulfation, methylation, hydrogenation, dehydrogenation, -oxidation ) and / or further microbiota catabolism ( hydrogenation, -oxidation, de-methoxylation, de-hydroxylation ) HO O OH OH HO O OH HO O OH OH HO O OH OH OH HO O OH HO O A selection of catabolites that are common to these substrates, found in plasma and / or urine as drawn or as conjugates ( glucuronidation, sulfation, methylation or glycination ) 0 100 200 300 400 500 0. 0 0. 5 p la s m a h e s p e re ti n ( M ) time ( min ) 0 100 200 300 400 500 0. 0 0. 5 p la s m a h e s p e re ti n ( M ) time ( min ) 0 100 200 300 400 500 600 0. 0 0. 5 1. 0 1. 5 2. 0 p la s m a h e s p e re ti n ( M ) time ( min ) 0 100 200 300 400 500 600 0. 0 0. 5 1. 0 1. 5 2. 0 p la s m a h e s p e re ti n ( M ) time ( min ) A B C D O O OH HO OHHO HO OH OHHO HO HO O OH OH OH OHO OH OH OH OH OHO OH OH OH OH OHO OH OH OH OH Specimen Proanthocyanidin Specimen Thearubigin A C B A A C C B A A C C B B 0h 0-2h 2-4h 4-8h 8-24h 0 10000 20000 30000 re la ti v e a m o u n t urine collection time * * * O OCH3 OH OOH HO HO OCH3 OH O HO HO OH O HO OCH3 OH O HO OCH3 OH O HO Ring scission Hydratase P450 acid 0. 02 µM, range 0. 006– 0. 08 µM [ 176 ] µM ely challenge d HUVECs producti on Gallic acid 1. 2 ± 1. 0 µM [ 82 ] 

Q3. How long does hesperetin take to be absorbed?

When hesperidin is consumed orally, hesperetin (conjugates of sulfate and glucuronide) appear in the plasma with a Tmax of 4-6 h. 

Q4. What is the role of phenols in the metabolisation of fatty acids?

It has also been demonstrated in vitro that the exposure to (poly)phenols modulates the ability of the microbiota to metabolise fructo-oligosaccharides and to generate short chain fatty acids [138, 139]. 

Q5. What are the likely substrates to dominate the microbial catabolites?

The catabolites most likely to dominate are the C6 phenols, C6–C1, C6– C2 and C6–C3 dihydro acids derived from chlorogenic acids/cinnamates, and most flavonoids including black tea thearubigins and theaflavins (see Figs. 2 and 5). 

Q6. What are the requirements for future investigations?

Future investigations must address the minimum effective dose of potentially prebiotic substrates, determine what percentage of the population are susceptible, whether susceptibility can be induced, and how long any associated benefits persist, especially if the supplementation is subsequently curtailed. 

Q7. What is the way to detect purely chemical transformations?

Some dietary (poly)phenols are unstable under the conditions employed for in vitro fermentations, and it is important to use an uninoculated control to detect purely chemical transformations. 

Q8. How did the consumption of dates affect the gut microbiota?

The consumption of seven dates per day by 22 volunteers for 21 days also did not alter the composition of the faecal microbiota, but stool ammonia was significantly lowered [112]. 

Q9. What is the approach to estimating the rate of clearance from plasma?

This approach assumes that the rate of clearance from plasma does not change with dose, and would only be true if clearance was saturated, and thus the estimate obtained is on the high side. 

Q10. What is the effect of coffee on the absorption of chlorogenic acids?

Having breakfast with coffee somewhat affected the timing of absorption, but not the overall amount absorbed [25], and non-dairy creamer, but not milk, has the same effect [26] and so the overall effect of food or beverages on the absorption and metabolism of chlorogenic acids appears to be minimal despite some reports to the contrary [27].