The Chemistry Behind the Folin-Ciocalteu Method for the Estimation of (Poly)phenol Content in Food: Total Phenolic Intake in a Mediterranean Dietary Pattern.

The Folin-Ciocalteu assay is a reference method for the quantification of total (poly)phenols in food. This review explains the fundamental mechanism of the redox reaction on which the method is based and looks at some of the practical considerations concerning its application. To accurately estimate the antioxidant capacity of (poly)phenolic compounds, a thorough knowledge of their structural characteristics is essential, as the two are closely associated. Therefore, to help researchers interpret the results of the Folin-Ciocalteu method, this review also summarizes some of the main phenolic structural features. Finally, we have used the Folin-Ciocalteu method to estimate the total phenolic intake associated with high adherence to a Mediterranean diet, ranked as one of the healthiest dietary patterns, which is characterized by a high consumption of (poly)phenol-rich food such as wine, virgin olive oil, fruits, vegetables, whole grains, nuts, and legumes.

Moderate Consumption of Beer (with and without Ethanol) and Menopausal Symptoms: Results from a Parallel Clinical Trial in Postmenopausal Women.

The menopausal transition can be a challenging period for women's health and a trigger of uncomfortable symptoms. Beer is the main food source of isoxanthohumol, a precursor of 8-prenylnaringenin, the strongest phytoestrogen identified to date. As phytoestrogens are reported to reduce perimenopausal symptoms, we evaluated if a daily moderate consumption of beer with (AB) and without alcohol (NAB) could improve menopausal symptoms and modify cardiovascular risk factors. A total of 37 postmenopausal women were enrolled in a parallel controlled intervention trial and assigned to three study groups: 16 were administered AB (330 mL/day), 7 NAB (660 mL/day), and 14 were in the control group. After a 6-month follow-up of the 34 participants who finished the trial, both interventions (AB and NAB) significantly reduced the severity of the menopause-related symptoms (p-value AB vs. Control: 0.009; p-value NAB vs. Control: 0.033). Moreover, AB had a beneficial net effect on psychological menopausal discomforts compared to the control group. As the sex hormone profile did not differ significantly between the study groups, the effects of both types of beers (AB and NAB) are attributed to the non-alcoholic fraction of beer. Furthermore, moderate NAB consumption improved the lipid profile and decreased blood pressure in postmenopausal women.

Cuisinomics: MS-based untargeted approach reveals chemical modulation by a recipe during home cooking.

Most of daily eaten food are cooked, which helps in absorbing nutrients and phytochemicals, but at the same time it can decrease its content. Currently, the impact of cooking has been studied that could influence food health related compounds, but they have a limited view of compounds by not consider molecular structural modifications and new compounds formation. An untargeted approach using LC-ESI-LQT-Orbitrap-MS/MS and univariate/multivariate statistical analysis was applied to understand how the preparation of a recipe, varying its ingredients (olive oil, 5-10%; onion, 20-40%; and garlic, 2-4%) and cooking time, could modulate the chemical profile of a tomato sofrito sauce. The presence of unexplored compounds that may have a beneficial effect on health, such as phytoprostanes, hydroxycinnamic acid amides and compounds such as 3,4 dihydroxyphenylglycone was revealed. Moreover, cooking was able to modulate the content of compounds like aminoacids, thiosulfates or phenolics and could be used as a tool to increase these molecules. The untargeted approach on cooking allows to use a recipe as a tool to improve a chemical profile of a dish, which opens the view for new dietary recommendations by cuisine to improve our diet, habits and health.

Effects of the Non-Alcoholic Fraction of Beer on Abdominal Fat, Osteoporosis, and Body Hydration in Women.

Several studies have shown that binge drinking of alcoholic beverages leads to non-desirable outcomes, which have become a serious threat to public health. However, the bioactive compounds in some alcohol-containing beverages might mitigate the negative effects of alcohol. In beer, the variety and concentration of bioactive compounds in the non-alcoholic fraction suggests that its consumption at moderate levels may not only be harmless but could also positively contribute to an improvement of certain physiological states and be also useful in the prevention of different chronic diseases. The present review focuses on the effects of non-alcoholic components of beer on abdominal fat, osteoporosis, and body hydration in women, conditions selected for their relevance to health and aging. Although beer drinking is commonly believed to cause abdominal fat deposition, the available literature indicates this outcome is inconsistent in women. Additionally, the non-alcoholic beer fraction might improve bone health in postmenopausal women, and the effects of beer on body hydration, although still unconfirmed seem promising. Most of the health benefits of beer are due to its bioactive compounds, mainly polyphenols, which are the most studied. As alcohol-free beer also contains these compounds, it may well offer a healthy alternative to beer consumers.

Microbial Phenolic Metabolites: Which Molecules Actually Have an Effect on Human Health?

The role of gut microbiota in human health has been investigated extensively in recent years. The association of dysbiosis, detrimental changes in the colonic population, with several health conditions has led to the development of pro-, pre- and symbiotic foods. If not absorbed in the small intestine or secreted in bile, polyphenols and other food components can reach the large intestine where they are susceptible to modification by the microbial population, resulting in molecules with potentially beneficial health effects. This review provides an overview of studies that have detected and/or quantified microbial phenolic metabolites using high-performance liquid chromatography as the separation technique, followed by detection through mass spectrometry. Both in vitro experimental studies and human clinical trials are covered. Although many of the microbial phenolic metabolites (MPM) reported in in vitro studies were identified in human samples, further research is needed to associate them with clinical health outcomes.

Bioactive Compounds of Cooked Tomato Sauce Modulate Oxidative Stress and Arachidonic Acid Cascade Induced by Oxidized LDL in Macrophage Cultures.

Sofrito is a mix of tomato, onion, garlic, and olive oil, which contains phenolic compounds and carotenoids. Consumption of tomato-based sofrito has been related to a lower risk of cardiovascular events, but the mechanisms behind such beneficial effects remain unclear. This study aimed to analyze the effects of representative sofrito compounds such as naringenin, hydroxytyrosol, lycopene, and ß-carotene on mechanisms involved in the pathogenesis of atherosclerosis. We demonstrated that both phenolic compounds and both carotenoids studied were able to inhibit low density lipoproteins (LDL) oxidation, as well as oxidative stress and eicosanoid production induced by oxidized LDL (oxLDL) in macrophage cultures. These effects were not the consequences of disturbing oxLDL uptake by macrophages. Finally, we observed an additive effect of these sofrito compounds, as well as the activity of a main naringenin metabolite, naringenin 7-O-ß-d-glucuronide on LDL oxidation and oxidative stress.

Mediterranean sofrito home-cooking technique enhances polyphenol content in tomato sauce.

BACKGROUND: Sofrito, a basic culinary technique widely used in the Mediterranean, may preserve dietary polyphenols and enhance their intake in the Mediterranean population. The aim of this study was to investigate if the sofrito technique improves the polyphenol extractability in a tomato-based sofrito sauce. RESULTS: A full factorial design was applied using mathematical models. The content of chlorogenic acid, ferulic acid hexoside and naringenin was higher in the sofrito sauce than in raw tomato. The bioaccessibility of some tomato polyphenols was enhanced by the presence of olive oil and they were protected from oxidation during the cooking process by the use of onion. CONCLUSION: The use of olive oil and onion in Mediterranean cooking as a base for sauces and dishes, with an appropriate cooking time, preserve the polyphenol content of food. Thus, Mediterranean cuisine may contribute to the health effects of the Mediterranean diet. © 2019 Society of Chemical Industry.

Acute Effect of a Single Dose of Tomato Sofrito on Plasmatic Inflammatory Biomarkers in Healthy Men.

Sofrito is a Mediterranean tomato-based sauce that typically also contains olive oil, onion, and garlic. The preparation of sofrito modifies the bioactive compounds (carotenoids and polyphenols) in the ingredients to more bioavailable forms, promoting cis-lycopene formation and polyphenol bioaccessibility. To evaluate the health benefits of this cooking technique, the effect of consuming an acute dose of sofrito on the inflammatory status was studied. In a clinical trial, 22 healthy male subjects consumed a single dose of sofrito (240 g/70 kg) after three days without ingesting any tomato products and following a low-antioxidant diet the day before the intervention. Plasma carotenoids and total polyphenol excretion (TPE) were evaluated, as well as the inflammatory biomarkers C-reactive protein (CRP), interleukin-6 (IL-6), interleukin 1ß (IL-1ß) and tumor necrosis factor-α (TNF-α). After the sofrito intake, a significant decrease in CRP (p = 0.010) and TNF-α (p = 0.011) was observed, but only TNF-α was inversely correlated with an increase in TPE and plasma ß-carotene (not the major carotenoid, lycopene). The positive health effects of this tomato-based product may be attributed not only to lycopene, but to the bioactive compounds of all the ingredients.

Polyphenols, food and pharma. Current knowledge and directions for future research.

Polyphenols are a large family of phytochemicals with great chemical diversity, known to be bioactive compounds of foods, species, medicinal plants and nutraceuticals. These compounds are ingested through the diet in significant amounts, around 1 g per day, an amount that be may be increased through supplements. The in vitro action of many representative polyphenols has been reported. However, their beneficial effects and their role in modulating the risk of high-prevalence diseases are difficult to demonstrate due to the wide variability of polyphenol structures and bioactive actions; the complexity of estimating the polyphenol content of food as a result of their variability in foods and cooked dishes; the potential modulation of the effects of polyphenols by food matrices; the addition of polyphenols and their synergistic interactions with each other and with other dietary bioactive components; the modulation of polyphenol bioavailability as a consequence of food composition and culinary techniques; their metabolism by the human body and the polyphenol gut microbiota metabolism in each metabotypes. Computational strategies, including virtual screening, shape-similarity-screening and molecular docking, were recently used to identify potential targets of polyphenols and thus gain a better understanding of the therapeutic effects exerted of polyphenols and modify natural polyphenol structures to potentiate specific activities. Here, we present the most relevant current knowledge and propose directions for future research in these fields, from the culinary world to the clinical setting. We hope this commentary will prompt scientists and clinicians to consider the therapeutic value of bioactive polyphenols and help shed some light on how much scientific truth lies in Hippocrates' famous quote: "Let your food be your medicine".

Identification of phenolic metabolites in human urine after the intake of a functional food made from grape extract by a high resolution LTQ-Orbitrap-MS approach.

Grape pomace (GP) is known to be a rich source of polyphenols with biological activity which may be used as functional ingredients for the development of new health-promoting products. Numerous studies have reported that bioactive compounds may act through multiple mechanisms. In order to verify the oral absorption and metabolism of grape polyphenols, we performed a prospective, randomised and cross-over acute study in 12 volunteers with two interventions: 500mL of a functional beverage enriched with 200mL of GP and 500mL of a control beverage without GP. In this work, liquid chromatography coupled with an electrospray ionization hybrid linear ion trap quadrupole-Orbitrap-mass spectrometry (LC/ESI-LTQ-Orbitrap-MS) technique has been used to accurately identify phenolics in GP and human urine. In GP, 41 phenolic compounds were identified mainly procyanidins, phenolic acids and flavonols, and in human urine over 70 metabolites of phenolic compounds including microbiota metabolites, glucuronides and sulfate derivatives were detected. Overall, high resolution mass spectrometry (HR-MS) enhances the identification of a large variety of polyphenols and their metabolites with great mass accuracies for all molecular ions.

Home cooking and ingredient synergism improve lycopene isomer production in Sofrito.

There has been increasing interest in tomato products rich in lycopene Z-isomers since these carotenoids present greater bioavailability and antioxidant capacity than the all-E lycopene form. Intrinsic food properties as well as processing and the interaction between dietary components can all influence the content, type and bioavailability of carotenoids. The aim of this study was to evaluate whether carotenoid content and isomerization in tomato-based Mediterranean sofrito is affected by the process of home cooking and the presence of other ingredients such as extra virgin olive oil, onion and garlic. We used a full factorial design to clarify the contribution of each ingredient to the carotenoid composition of sofrito and to determine whether this can be improved by the cooking time and ingredient synergism. Cooking time and onion content were associated with a higher production of 5-Z-lycopene, 9-Z-lycopene and 13-Z-lycopene in sofrito. Onion proved to be the most interesting ingredient in the sofrito formulation due to their enhancing effect on lycopene isomerization. The use of onion combined with an adequate processing time may improve the bioavailability of lycopene in tomato products.

trans-Lycopene from tomato juice attenuates inflammatory biomarkers in human plasma samples: An intervention trial.

SCOPE: The effect of carotenoids from tomato juice (TJ) on inflammatory biomarkers was evaluated by performing a 4-week dose-response nutritional trial in a population at high cardiovascular risk. METHODS AND RESULTS: An open, prospective, randomized, cross-over, and controlled clinical trial was carried out with 28 volunteers (mean age 69.7 ± 3.1 years; mean BMI 31.5 ± 3.6 kg/m2 ) at high cardiovascular risk, which were assigned to consume daily for 4 weeks in random order: 200 mL (LD) or 400 mL (HD) of TJ, or water as a control (C), with a 21-day wash-out period between each intervention. Blood samples were collected at baseline (B) and after each intervention. Endpoints included significant changes in plasmatic carotenoids, and adhesion molecules ICAM-1, and VCAM-1, as well as a tendency to decrease the chemokine IL-8. Compared to C, concentration of ICAM-1, and VCAM-1 were significantly lower (p ˂ 0.001), after each TJ intervention. Decreases were correlated remarkably with the trans-lycopene, while the other carotenoids present in TJ have presented a minor association or no association with changes in these molecules. CONCLUSION: trans-Lycopene from TJ may attenuate the risk of cardiovascular disease by reducing the concentration of important inflammatory molecules related to atherosclerosis.

Targeted filtering reduces the complexity of UHPLC-Orbitrap-HRMS data to decipher polyphenol polymerization.

UHPLC-LTQ-Orbitrap-high resolution mass spectrometry (HRMS) was applied to investigate complex polymeric polyphenols, before and after acid-catalysed depolymerisation in the presence of a nucleophile (phloroglucinol). Reaction products of (-)-epicatechin with acetaldehyde formed in model solution were selected for a proof-of concept experiment. The complexity of the UHPLC-HRMS dataset obtained after 4h incubation was reduced with petroleomics-inspired strategies using Van Krevelen diagrams and modified Kendrick mass defect filtering targeting ethyl-epicatechin (C17H16O6) units. Combining these approaches with mass fragmentation and phloroglucinolysis allowed us to describe reaction of epicatechin and acetaldehyde. More than 65 compounds were found, including the homogeneous bridged derivatives (up to the undecamer), vinyl and ethanol adducts, and xanthene and xanthylium salt derivatives which were identified for the first time.

Absorption and disposition of naringenin and quercetin after simultaneous administration via intestinal perfusion in mice.

As common constituents of tomatoes and other fruits and/or vegetables, naringenin and quercetin are usually ingested together, so for a clearer understanding of their bioavailability, metabolic fates and health benefits, it is more insightful to study them together. The purpose of the present work was to study how co-administration of naringenin and quercetin at realistic doses (3.5 µg ml(-1) and 2.36 µg ml(-1), respectively) influences their absorption and intestinal first-pass metabolism. A single-pass intestinal perfusion model in mice (n = 4-6) was used. Perfusate (every 10 minutes), blood (at 60 min) and bile samples were analysed by an UPLC-ESI-MS/MS method to evaluate the presence of the aglycones and their metabolites. Both naringenin and quercetin showed high permeability coefficients when administered separately (7.71 ± 0.82 × 10(-4) cm s(-1)vs. 7.30 ± 1.95 × 10(-4) cm s(-1), respectively), but these values decreased by 50% with co-administration (4.09 ± 0.89 × 10(-4) cm s(-1) for naringenin and 3.18 ± 0.45 × 10(-4) cm s(-1) for quercetin). Moreover, the level of phase II metabolites in perfusion, plasma and bile samples increased when naringenin and quercetin were administered together. The higher biliary excretion of these metabolites could thus favour the entero-hepatic recycling of the aglycones and metabolites. The results of this study may have several useful applications: to know and consider the possible interactions between polyphenols and drugs that use the same mechanism of absorption and elimination; when polyphenol-rich nutritional supplements are supplied, and in our regular diets to optimize the health benefits afforded by the biological activities of such aglycones and/or metabolites.

Use of metabolomics and lipidomics to evaluate the hypocholestreolemic effect of Proanthocyanidins from grape seed in a pig model.

SCOPE: This work aims to evaluate changes in the fecal metabolomic profile due to grape seed extract (GSE) intake by untargeted and targeted analysis using high resolution mass spectrometry in conjunction with multivariate statistics. METHODS AND RESULTS: An intervention study with six crossbred female pigs was performed. The pigs followed a standard diet for 3 days, then they were fed with a supplemented diet containing 1% (w/w) of MegaNatural® Gold grape seed extract for 6 days. Fresh pig fecal samples were collected daily. A combination of untargeted high resolution mass spectrometry, multivariate analysis (PLS-DA), data-dependent MS/MS scan, and accurate mass database matching was used to measure the effect of the treatment on fecal composition. The resultant PLS-DA models showed a good discrimination among classes with great robustness and predictability. A total of 14 metabolites related to the GSE consumption were identified including biliary acid, dicarboxylic fatty acid, cholesterol metabolites, purine metabolites, and eicosanoid metabolites among others. Moreover, targeted metabolomics using GC-MS showed that cholesterol and its metabolites fecal excretion was increased due to the proanthocyanidins from grape seed extract. CONCLUSION: The results show that oligomeric procyanidins from GSE modifies bile acid and steroid excretion, which could exert a hypocholesterolemic effect.

Phenolic metabolites and substantial microbiome changes in pig feces by ingesting grape seed proanthocyanidins.

Proanthocyanidin (PAC) consumption has been linked to better colonic health, but PACs are poorly absorbed, making them a target for colonic metabolism. The resulting metabolites are low molecular weight and could potentially be absorbed. To understand the effects of dietary PACs it would be important to resolve the metabolic issue and link these changes to microbial population changes in a suitable model for human digestion. Here, six crossbred female pigs were fed a diet containing 1% (w/w) of MegaNatural® Gold grape seed extract (GSE) daily for 6 days. Fecal samples were analyzed by normal phase LC coupled to fluorescence detection and LC-MS/ToF. DNA was extracted from pig fecal samples and the V3/V4 region of the 16S rRNA gene was sequenced using an Illumina MiSeq. Intact parent PACs (dimer-pentamer) were observed in the feces on days 3 and 6 at similar high levels (∼400 mg kg(-1) total) during ingestion of GSE but were absent 48 h post-feeding. The major phenolic metabolites were 4-hydroxyphenylvaleric acid and 3-hydroxybenzoic acid which increased by ∼30 and 3 mg kg(-1) respectively. The GSE diet also caused an ecological shift in the microbiome, dramatically increasing Lachnospiraceae, Clostridales, Lactobacillus and Ruminococcacceae. The relationship between dietary PACs and colon health may be attributable to the altered bacterial populations or phenolic compounds in the colon.

Olive oil intake and risk of cardiovascular disease and mortality in the PREDIMED Study.

BACKGROUND: It is unknown whether individuals at high cardiovascular risk sustain a benefit in cardiovascular disease from increased olive oil consumption. The aim was to assess the association between total olive oil intake, its varieties (extra virgin and common olive oil) and the risk of cardiovascular disease and mortality in a Mediterranean population at high cardiovascular risk. METHODS: We included 7,216 men and women at high cardiovascular risk, aged 55 to 80 years, from the PREvención con DIeta MEDiterránea (PREDIMED) study, a multicenter, randomized, controlled, clinical trial. Participants were randomized to one of three interventions: Mediterranean Diets supplemented with nuts or extra-virgin olive oil, or a control low-fat diet. The present analysis was conducted as an observational prospective cohort study. The median follow-up was 4.8 years. Cardiovascular disease (stroke, myocardial infarction and cardiovascular death) and mortality were ascertained by medical records and National Death Index. Olive oil consumption was evaluated with validated food frequency questionnaires. Multivariate Cox proportional hazards and generalized estimating equations were used to assess the association between baseline and yearly repeated measurements of olive oil intake, cardiovascular disease and mortality. RESULTS: During follow-up, 277 cardiovascular events and 323 deaths occurred. Participants in the highest energy-adjusted tertile of baseline total olive oil and extra-virgin olive oil consumption had 35% (HR: 0.65; 95% CI: 0.47 to 0.89) and 39% (HR: 0.61; 95% CI: 0.44 to 0.85) cardiovascular disease risk reduction, respectively, compared to the reference. Higher baseline total olive oil consumption was associated with 48% (HR: 0.52; 95% CI: 0.29 to 0.93) reduced risk of cardiovascular mortality. For each 10 g/d increase in extra-virgin olive oil consumption, cardiovascular disease and mortality risk decreased by 10% and 7%, respectively. No significant associations were found for cancer and all-cause mortality. The associations between cardiovascular events and extra virgin olive oil intake were significant in the Mediterranean diet intervention groups and not in the control group. CONCLUSIONS: Olive oil consumption, specifically the extra-virgin variety, is associated with reduced risks of cardiovascular disease and mortality in individuals at high cardiovascular risk. TRIAL REGISTRATION: This study was registered at controlled-trials.com (http://www.controlled-trials.com/ISRCTN35739639). International Standard Randomized Controlled Trial Number (ISRCTN): 35739639. Registration date: 5 October 2005.

Extravirgin olive oil consumption reduces risk of atrial fibrillation: the PREDIMED (Prevención con Dieta Mediterránea) trial.

BACKGROUND: The PREDIMED (Prevención con Dieta Mediterránea) randomized primary prevention trial showed that a Mediterranean diet enriched with either extravirgin olive oil or mixed nuts reduces the incidence of stroke, myocardial infarction, and cardiovascular mortality. We assessed the effect of these diets on the incidence of atrial fibrillation in the PREDIMED trial. METHODS AND RESULTS: Participants were randomly assigned to 1 of 3 diets: Mediterranean diet supplemented with extravirgin olive oil, Mediterranean diet supplemented with mixed nuts, or advice to follow a low-fat diet (control group). Incident atrial fibrillation was adjudicated during follow-up by an events committee blinded to dietary group allocation. Among 6705 participants without prevalent atrial fibrillation at randomization, we observed 72 new cases of atrial fibrillation in the Mediterranean diet with extravirgin olive oil group, 82 in the Mediterranean diet with mixed nuts group, and 92 in the control group after median follow-up of 4.7 years. The Mediterranean diet with extravirgin olive oil significantly reduced the risk of atrial fibrillation (hazard ratio, 0.62; 95% confidence interval, 0.45-0.85 compared with the control group). No effect was found for the Mediterranean diet with nuts (hazard ratio, 0.89; 95% confidence interval, 0.65-1.20). CONCLUSIONS: In the absence of proven interventions for the primary prevention of atrial fibrillation, this post hoc analysis of the PREDIMED trial suggests that extravirgin olive oil in the context of a Mediterranean dietary pattern may reduce the risk of atrial fibrillation. CLINICAL TRIAL REGISTRATION URL: http://www.controlled-trials.com. Unique identifier: ISRCTN35739639.