Oxidation connects numerous and important human disorders. Therefore, it is tempting to simplify the action of polyphenols based on their intrinsic properties. Polyphenols are bioactive but not bio-accessible. A number of questions may be raised. Firstly, in plants, growing region, variety or processing procedures may change the in vitro antioxidant properties; secondly, and intriguingly, some have found potential artifacts in cell culture studies that are difficult to interpret; thirdly, the activity of parent molecules is not necessarily the same as that of their metabolites; fourthly, oral administration supposes modification in the gastrointestinal system and the effect of pH and/or microbiota can be hardly understated. Is it time, therefore, to complicate the issue (even more) and to focus research in actions independent of their antioxidant properties?
It is true that there are several issues in establishing polyphenols as antioxidants for humans. In vitro assays certainly can not be taken as experimental proof that these also act in vivo. The pharmacokinetics and in vivo bioactivity of these plant products is very important aspects which should be the focus of future research.
Yes of course sir. Antioxidants are defense system which can scavange free radicals. Antioxidants prevent oxidation reaction. Oxidation reaction can produce free radicals. which will start chain of reaction and lead to damage to the cell. Polyphenols have free radical scavenging ability. Therefore it shows high antioxidant activity. And I have attached some papers which will be usefull for your research.
Thank you all for your time. As Joseph accurately points, complexity is not a reason to give up. However, sometimes is difficult to keep confident. Measurements in vitro are consistent, and certainly a global antioxidant score is useful in vitro. The bioactivity of plant extracts (not necessarily polyphenols-only) is also consistent, in our hands, in humans. However, antioxidant activity in plasma, or blood,... simply, our data do not support a pre-empted increase. Simone and Vinod are also right indicating that complexity is probably connected with derived (undetectable?) metabolites. Luckily, Veerabahu is fully optimistic.
Best regards
One of the measurements that is often performed by some of my associates is the redused versus oxidised glutathione ratio. This allows them to estimate the oxidative state of their samples over a period of time. One can then see if your polyphenol was effective in controlling oxidation.
Cell line screening, and in vitro tests of antioxidant activity are not relevant to the uptake and in-body effects of polyphenolics in the diet. Look for publications like the one below that discuss human metabolism (and gut flora processing) of these compounds.
Impact of various factors on pharmacokinetics of bioactive polyphenols: An overview Rubió, L., Macià, A., Motilva, M.-J. 2014 Current Drug Metabolism
15 (1), pp. 62-76
No! In vitro results are preliminary results. Yonathan Asikin is quite right.
The important question to me is: what is the action space of the research? Is it the food and food ingredients space? The best (and often cheapest) solution is for people to each plenty of fruits, vegetables, pulses and whole grains each day. 'Superfoods' labels are marketing exercises to refresh the image of a specific fresh fruit or vegetable in the public consciousness. Valid enough, when so many processed foods of poor value are marketed so aggressively. Extracting antioxidant-enriched food ingredients to add to industrially produced foods seems a useful target, especially if value is extracted from waste streams, but highly purified extracts or single compounds cross over from food to pharmacology.
Is the the nutraceuticals space? To be cynical, those selling extracts often benefit by having the minimum known about their product's function. Sellers of 'antioxidant' extracts use in vitro assay results as a marketing tool to benchmark their product against others -and we can all live healthily without consuming another high-cost 'gojolo berry extract'.
Is it the pharmacology space? Bulk screening of foods for pharmacological function seems like a blunt instrument, a poor tool in an age when we can target and synthesize specific structures. This is a dangerous space to work in- look at the published negative interactions of some antioxidants with cancer therapies as an example.
All this research is also expensive to do, so we must assess the overall value of a project before spending others' research money, and our research time, to prove what we already know- that humans benefit from many of the compounds in a diet rich in no- or low- processed foods.
There are many substances in our blood and cells , they may interfere anitoxidant activity of polyphenols.
As an aside, this is regarding the critique above - the "nuraceutical space". Those who benefited most from the antioxidant story of vitamin E (may also be carotenes) were, in fact, giant pharmaceutical companies headquartered in Europe and operating out of North America. Just check who had the resources/manufacturing plants for producing and distributing tocopherols! One would also think that the potential profits that nutritional supplement marketing companies enjoyed/aspired for would have paled in comparison with that of the giant pharmaceutical outfits (not to talk of their “ethical” practices).
Surprised that we still ignore bioavaialbility of antioxidants when it is the one which limits how much of those compounds reach systemic circulation and thence the different site in the body. A highly bioavailable antioxidant could infact show a comparable in vitro and in vivo antioxidant activity. But in reality such a compound is surreal per se.
One cannot simply assume that flavonoids are poorly absorbed since they are found in foods as glycosides. Similarly, generalizations cannot be made about the bioavailability and biological accessibility of flavonoids. We need to understand more. Most flavonoids, except catechins, exist in nature as glycosides. Moreover, at least quercetin glucosides were absorbed better than the aglycone, quercetin (quercetin is the most prominent dietary flavonoid). Consequently, the extent of flavonoid glycosides consumed is a better indication than the quantity of aglycones consumed, thus raising the lower level estimated for the flavonoid aglycones. Supplementation of the diet should more appropriately use flavonoid glycosides instead of aglycones.
After the ingestion of diosmin (the 7-rhamnoglucoside of diosmetin, 5,7,3′-trihydroxy-4′-methoxyflavone) in humans, only diosmetin (the aglycone) could be detected in the plasma. The time course of diosmetin plasma concentrations indicated rapid initial distribution and prolonged final elimination half-life of 31.5 h. Neither diosmin nor diosmetin could be detected in urine. The metabolites in urine were primarily phenolic acids. The prolonged presence of diosmetin in blood is suggestive of an enterohepatic circulation.
Quercetin glycosides from onions were more readily absorbed than the pure aglycone; absorbed quercetin was eliminated slowly from the blood, suggesting that enterohepatic circulation may be operative. In related studies, it was concluded that quercetin-glucose conjugates were more readily absorbable; the suggestion was made that the glycosides might be absorbed via the intestinal sugar uptake route. Following ingestion of fried onions containing quercetin glycosides equivalent to 64 mg of quercetin aglycone, peak plasma levels of 196 μg/ml were achieved after 2.9 h. The distribution phase half-life was 3.8 h and the elimination phase half-life was 16.8 h. Thus, oral dietary (cooked vegetable) quercetin can be absorbed and reach tissues and plasma where antioxidant and other activities could be exerted. What is true for quercetin is very likely true also for other flavonoids in other vegetable sources. Thus, the cumulative concentration of quercetin plus other flavonoids could be substantially greater than that shown for quercetin alone.
An early evaluation of the absorption and metabolism of (+)-catechin in humans was presented by Das in 1971. Oral administration (83 mg/kg) resulted in rapid absorption, metabolism, and excretion of the flavonoid within 24 h. Eleven metabolites were detected in urine.
Isoflavonoid phytoestrogens and mammalian lignans, occurring in animal and human biological fluids and in feces, are diphenolic compounds with molecular weights similar to those of steroid estrogens; the mammalian compounds are produced from plant sources and isoflavonoids by intestinal microflora. The phytoestrogenic isoflavone, daidzein, has been detected in human urine. The isoflavonoids have been shown to bind with relatively high affinities to the estrogen receptors of human mammary tumor cells. They display affinity for type II binding sites. They may, therefore, be implicated in the inhibition of breast carcinoma cell growth mediated by estrogen. The isoflavone molecule and estrogen show structural similarities.
When human volunteers consumed soya flour, the urinary excretion of the isoflavonoids, genistein, daidzein, and glycitein, increased after 24 h as did the isoflavonoid metabolites equol and O-desmethylangolensin. The plasma concentrations of four isoflavonoids, daidzein, genistein, O-desmethylangolensin, and equol, were very high in Japanese men consuming a low fat diet with a high content of soy products; the mean plasma total and individual isoflavonoid levels were 7 to 110 times higher in the Japanese men than in the Finnish men. These phytoestrogen levels may inhibit the growth of prostate cancer in Japanese men, which may explain the low mortality from prostatic cancer in that country. Concentrations of the isoflavonoid, genistein, in urine of subjects consuming a traditional soy-rich Japanese diet were in the micromolar range, while these concentrations were 1/30th or less of these in the urine of omnivores.
The Effects of Plant Flavonoids on Mammalian Cells: Implications for Inflammation, Heart Disease, and Cancer. Pharmacol Rev, 2000 52:673-751.
http://pharmrev.aspetjournals.org/content/52/4/673.full.pdf+html
Dear Chithan C Kandaswami,
Almost all your examples are quite convincing. However, I still have some doubts on the one from Das, you mentioned: is this Dipak, you are talking about? If this is the case, unfortunately, he passed away too quickly to be able to "retract" more than 18 out of his 140 (?) falsified papers as he was condemned to in 2012 by his own Connecticut University. Even though they relate mainly to the stilbenoid series, which are the results on other polyphenols, from this group which are worthy of confidence?
About the hypothesized explanation to the implication of isoflavonoids in the inhibition of breast carcinoma cell growth mediated by estrogen, due to the fact that the isoflavone molecule and estrogen show structural similarities..., are you aware that the high "affinity" of some epicatechin oligomers with membrane (NOT NUCLEAR) androgen/estrogen receptors which are then inducing apoptosis rather than proliferation, could be another hypothesis? In addition, when using some derivatives (oleyl esters), we have shown (in vitro) that these procyanidols were able to fight against hormone-sensitive tumor cells as well as hormone-independant prostate cancer cells (http://jpet.aspetjournals.org)...
Dear Joseph, thank you. I t is good to visit with you, here, at RG after 1999 (Wine and Health International Conference, Volume: 1. Polyphenols, Wine and Health Communications, Bordeaux {France} April 14 – 16 1999 Bordeaux); back then, we presented at your meeting, work of grape seed extract - GSE - (the particular sample from Polyphenolics [Constellation Brands], contained monomers and oligomeric procyanidins, characterized by Canandaigua Wines R&D in Madera, CA) and EDRF activity (reported for the first time in 1998 by Dr Kappagoda, who developed the rabbit model for hypercholesterolemia.), from Dr Tissa Kappagoda’s group (Preventive Cardiology Unit, Dept of Internal Medicine, UC Davis, CA). We also presented other work related to GSE and co-entry receptor genes/cytokines.
No Joseph, it is not Deepak that I alluded to. He boarded the ship, so to say, after his article on HPLC determination of flavonoids, in a certain volume of “Methods in Enzymology” edited by Packer; Deepak, it may turn out, had not performed any flavonoid work at the time this article took shape. I am aware of his “paper mill” on resveratrol! I was referring to Professor Nagarathnam P. Das (who is, unfortunately, no more; he had a premature end), who was a pioneer. He edited the third volume of “Flavonoids in Biology & Medicine (the first two were edited by Cody V, Harborne J B and Middleton Jr, E.).
Das NP (1971) Studies on flavonoid metabolism: Absorption and metabolism of (+)-catechin in man. Biochem Pharmacol 20:3435–3445.
Flavonoids in Biology and Medicine III Current Issues in Flavonoids Research
Editors N.P. Das, Kevin H. Cheeseman, Department of Biology & Biochemistry, Brunel University. (doi: 10.3109/10715769109088944)
International Symposium on Flavonoids in Biology & Medicine (3rd : 1989 : Singapore) Edited by Das, N. P. (Nagaratnam Patmanatha). Current issues in flavonoids research. Singapore : National University of Singapore, 1989.
Thank you very much for the article. I am aware of this activity of epicatechin oligomers but had not read this. In many cases, flavonoids are weak inhibitors, weakly bind to the type II binding sites, which can displace the estrogen molecule. In the case of the epicatechin oligomers, it may be enzyme modulation. The binding might also influence membrane associated tyrosine kinase activities.
A little perspective: The groundwork for structure-activity relationship governing antioxidant function of flavonoids, and the primary determinants of antioxidant function were laid as early as 1958 by Mehta and Seshadri, which is correct even now; further detailed elucidation was reported in 1986; all these are way and much before the powerful and pervasive UK groups boarded the bandwagon. It is sobering that these seminal papers get cursory mention, even if they do. To resort to an informal term, we adore the art of “reinventing the wheel”.
Mehta AC, Seshadri TR. Flavonoids as antioxidants. J Sci Ind Res1958;18b:24–8.
Das NP, Ratty AK (1986) Effects of flavonoids on induced non-enzymic lipid peroxidation. in Plant Flavonoids in Biology and Medicine: Biochemical, Pharmacological, and Structure-Activity Relationships, eds Cody V, Middleton E, Harborne JB (Alan R. Liss, Inc. New York), pp 243–248.
Ratty AK, Das NP (1988) Effects of flavonoids on nonenzymatic lipid peroxidation: Structure activity relationship. Biochem Med Metab Biol 39:69–79.
Ratty AK, Sunamoto J, Das NP (1988) Interaction of flavonoids with 1,1-diphenyl-2-picrylhydrazyl free radical, liposomal membranes and soybean lipoxygenase-1. Biochem Pharmacol 37:989–995.
It may be a bit ironic that we are now having this discussion on flavonoids/polyphenols and cancer prevention. It was conceived that flavonoids were secondary metabolites in higher vascular plants, angiosperms, with no primary function in the species in which they are elaborated (and considered to be secondary, nonessential dietary factors without any documented relevance to human health and/or disease prevention or amelioration). Unfortunately, vitamin P (associated with flavones) was removed from the list of vitamin (AIN/ASBCMB). They were mostly relegated (but for the concerted efforts of few resolute authors). The evaluation potential mutagenicity of food flavonoids, subsequent to the inception of the Ames mutagenicity test, and following the 1977 publication of Bjeldanes and Chang on the mutagenic activity of quercetin and related flavonoids, there were numerous studies aimed on assigning mutagenic/procarcinogenic roles for these “lowly” diet-derived molecules (based on supraphysiological/pharmacological concentrations in in vitro evaluations, particularly in vitro mutagenicity and micronucleus genotoxicity tests with mM concentrations even though flavonoids did not cause sister chromatid exchanges in these genotoxic evaluations, despite their effects on DNA strand breaks and chromosomal anomalies). As an aside, the availability of unprecedented funds in the area of procarcinogen identification, environmental toxicology and xenobiotic carcinogenesis could have contributed to this. This was a momentous interlude.
Bjeldanes LF, and Chang GW (1977) Mutagenic activity of quercetin and related compounds. Science (Wash DC) 197:577–578.
In the next 15 years or so, on and off, flavonoids seemed to have been “excoriated” as “noxious” arch villains orchestrating adversity. In this saga of changing paradigms, flavonoids, once marginalized and on the sidelines, gained enormous attention and publicity following the inception of the “The French Paradox”; this, one would imagine, would have had an effect in “fattening and proliferating” the coffers of wine industries. An awe inspiring leap from notoriety to respectability! With the emergence of the colorful (and the then promising) dietary antioxidant and disease prevention field, flavonoids experienced an elevation in their status, and were propelled to the pinnacle – secondary metabolites to suspected procarcinogens to prophylactic agents. The magnitude of interest is simply unmatched when one also considers the touted beneficial effects of cocoa consumption (by “giant” chocolate manufacturers) based on the presumptive actions of cocoa flavans (procyanidin dimers) on vasodilation and the promise of cardiovascular protection. This is the capricious state of affairs in this particular area.
Bjeldanes LF, and Chang GW (1977) Mutagenic activity of quercetin and related compounds. Science (Wash DC) 197:577–578.
Bjeldanes LF, and Chang GW (1977) Mutagenic activity of quercetin and related compounds. Science (Wash DC) 197:577–578.
As stated above, following intial reports on the mutagenic activity of flavonoids, these molecules underwent ongoing intense scrutiny as promutagenic and procarcinogenic agents. In the next 15 years or so, on and off, flavonoids seemed to have been “excoriated” as “noxious” arch villains orchestrating adversity. In this saga of changing paradigms, flavonoids, once marginalized and on the sidelines, gained enormous attention and publicity following the inception of the “The French Paradox”; this, one would imagine, would have had an effect in greatly adding to the ting” the coffers of wine industries. An awe inspiring leap from notoriety to respectability! With the emergence of the colorful (and the then promising) dietary antioxidant and disease prevention field, flavonoids experienced an elevation in their status, and were propelled to the pinnacle – secondary metabolites to suspected procarcinogens to prophylactic agents. The magnitude of interest is simply unmatched when one also considers the touted beneficial effects of cocoa consumption (by “giant” chocolate manufacturers) based on the presumptive actions of cocoa flavans (procyanidin dimers) on vasodilation and the promise of cardiovascular protection. This is the capricious state of science in this particular area.
Well, it would certainly depend on your definition of “in humans”. One can expect to have antioxidant activity of polyphenols in stomach and the rest of the digestive tract. The main leitmotiv here is that polyphenols are still capable to scavenge for free radicals introduced with food. But if you are looking for polyphenols in the blood samples you may be disappointed.
Q: Polyphenols scavenge free radicals in vitro: does it imply an antioxidant activity in humans?
Animal experiments had been done to support the anti-oxidant activity of polyphenols in scavenging free radicals. In Human, drinking Green Teas or taking their health products, we experienced their anti-oxidant effects of lesser oxidation stresses or damages. I think the polyphenols ingredients in these Green Teas or their Health products do have in vivo anti-oxidant effects in Human. Do you agree?
Thank you, Dr Zhou!
Population (epidemiological) studies have concluded that the daily ingestion of five or more cups of green tea might have beneficial effects.
Two controlled clinical trials reported that the daily ingestion of 4 to 5 cups of black tea during a four-week duration improved endothelium dependent vasodilation in humans with coronary artery disease or mildly elevated plasma cholesterol results. A number of small investigations have indicaed that green tea or its constituent catechins may have similar vascular effects.
Anecdotal evidence suggest the health promoting potential of green tea consumption. However, it has not been unequivocally established that the consumption of green tea (or of its catechins) in humans evokes in vivo antioxidant effects. A cup (8 ounces) of green tea contains 10-50 mg of caffeine.
Dear Dr. Joven,
This has already happened over the past several years. Redox (reduction and oxidation maintenance) balance, membrane and capillary stabilization, endocrine function, gene modulation, cell signaling function, cell surface signal transduction, anti-inflammatory and antiallergic effects and promotion of immune cell function, interaction with proteins and effects on protein conformation maintenance, cell differentiation, modulation of ATP binding effects, dihedral angles of flavonoids and their significance, receptor function, modulation of growth factor effects, modulation of stress response, stabilization of thiols, etc., are non-antioxidant functions of flavonoids. Antioxidant function of flavonoids is only one facet of their biologic effects.
The Effects of Plant Flavonoids on Mammalian Cells: Implications for Inflammation, Heart Disease, and Cancer
Elliott Middleton, Jr., Chithan Kandaswami, and Theoharis C. Theoharides
The Effects of Plant Flavonoids on Mammalian Cells:Implications for Inflammation, Heart Disease, and Cancer Pharmacol Rev December, 2000 52:673-751
Oral administration supposes modification in the gastrointestinal system may have many free radical implications. Free radicals may effect membrane function, transport of ions and fluids and energy molecules, These may disturb and capillary stabilization may effect endocrine function, gene activity cell signaling signal transduction. Modulation of ATP binding effects, receptor function, modulation of growth factor effects, modulation of stress response, stabilization of thiols, or are important antioxidant functions of flavonoids. Hence, antioxidant function of flavonoids is only one facet of their biologic effects on Mammalian Cells.
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