Raul Zamora-Ros

Estimation of Dietary Sources and Flavonoid Intake in a Spanish Adult Population (EPIC-Spain)

BACKGROUND Epidemiologic studies have suggested associations between flavonoid intake and health benefits. Traditional Mediterranean diets consist of a high consumption of plant products rich in flavonoids. OBJECTIVE This study estimates dietary flavonoid intake and main food sources in a Mediterranean population (Spanish adults). DESIGN The study included 40,683 subjects aged 35 to 64 years from northern and southern regions of Spain who were included in the European Prospective Investigation into Cancer and Nutrition study Spanish cohort. Usual food intake was assessed by personal interviews using a computerized version of a validated diet history method. Expanded US Department of Agriculture databases for the flavonoid, isoflavone, and proanthocyanidin content were used. RESULTS The median and mean of total flavonoids were 269.17 and 313.26 mg/day, respectively. The most abundant flavonoid subgroup was proanthocyanidins (60.1%), followed by flavanones (16.9%), flavan-3-ols (10.3%), flavonols (5.9%), anthocyanidins (5.8%), flavones (1.1%), and isoflavones (<0.01%). The main sources of total flavonoid intake were apples (23%), red wine (21%), unspecified fruit (12.8%), and oranges (9.3%). CONCLUSIONS These results should be very useful for evaluating the relationships between flavonoid intake and several diseases.

Rapid Folin-Ciocalteu method using microtiter 96-well plate cartridges for solid phase extraction to assess urinary total phenolic compounds, as a biomarker of total polyphenols intake

Nutritional markers have several advantages for epidemiologic and clinical assays, when compared to dietary data obtained by food frequency questionnaires. Few studies have assessed whether total polyphenol (TP) compounds provide a valid biomarker for TP intake. To date, there has been almost no literature describing methods to determine TP in complex matrices such as urine, which have many interfering substances. We report a rapid Folin-Ciocalteu method to determine TP in urine samples using Oasis((R)) MAX 96-well plate cartridges for solid phase extraction. These plates allow analysis of a high number of samples at the same time. We performed a prospective, randomized, crossover trial and one cross-sectional study with 60 volunteers from the PREDIMED trial, seeking to evaluate whether the TP in urine were correlated with polyphenol intake and could, therefore, be considered as a marker of intake of these compounds. The assay was optimized; the sensitivity and the polarity range of urine polyphenols were increased and the detection and quantification limits were significantly reduced. The metabolites in standards solution and urine samples were stable under the storage and handling conditions. In the clinical trial and the cross-sectional study, TP excreted in spot urine samples were positively correlated with TP intake, r=0.48, P<0.01 and r=0.257, P=0.04, respectively. The methodology described may be used to detect TP in urine samples, employing the high throughput of 96-well microtiter plates and reader. The method is fast and simple and it allows analysis of a large number of samples at the same time.

Concentrations of resveratrol and derivatives in foods and estimation of dietary intake in a Spanish population: European Prospective Investigation into Cancer and Nutrition (EPIC)-Spain cohort.

Resveratrol has been shown to have beneficial effects on diseases related to oxidant and/or inflammatory processes and extends the lifespan of simple organisms including rodents. The objective of the present study was to estimate the dietary intake of resveratrol and piceid (R&P) present in foods, and to identify the principal dietary sources of these compounds in the Spanish adult population. For this purpose, a food composition database (FCDB) of R&P in Spanish foods was compiled. The study included 40,685 subjects aged 35-64 years from northern and southern regions of Spain who were included in the European Prospective Investigation into Cancer and Nutrition (EPIC)-Spain cohort. Usual food intake was assessed by personal interviews using a computerised version of a validated diet history method. An FCDB with 160 items was compiled. The estimated median and mean of R&P intake were 100 and 933 microg/d respectively. Approximately, 32% of the population did not consume R&P. The most abundant of the four stilbenes studied was trans-piceid (53.6%), followed by trans-resveratrol (20.9%), cis-piceid (19.3%) and cis-resveratrol (6.2%). The most important source of R&P was wines (98.4%) and grape and grape juices (1.6%), whereas peanuts, pistachios and berries contributed to less than 0.01%. For this reason the pattern of intake of R&P was similar to the wine pattern. This is the first time that R&P intake has been estimated in a Mediterranean country.

Differences in dietary intakes, food sources and determinants of total flavonoids between Mediterranean and non-Mediterranean countries participating in the European Prospective Investigation into Cancer and Nutrition (EPIC) study

A greater adherence to the traditional Mediterranean (MED) diet is associated with a reduced risk of developing chronic diseases. This dietary pattern is based on higher consumption of plant products that are rich in flavonoids. We compared the total flavonoid dietary intakes, their food sources and various lifestyle factors between MED and non-MED countries participating in the EPIC study. Flavonoid intakes and their food sources for 35,628 subjects, aged 35-74 years and recruited between 1992 and 2000, in twenty-six study centres were estimated using standardised 24 h dietary recall software (EPIC-Soft®). An ad hoc food composition database on flavonoids was compiled using analytical data from the United States Department of Agriculture and Phenol-Explorer databases. Moreover, it was expanded to include using recipes, estimations of missing values and flavonoid retention factors. No significant differences in total flavonoid mean intake between non-MED countries (373·7 mg/d) and MED countries (370·2 mg/d) were observed. In the non-MED region, the main contributors were proanthocyanidins (48·2%) and flavan-3-ol monomers (24·9%) and the principal food sources were tea (25·7%) and fruits (32·8%). In the MED region, proanthocyanidins (59·0%) were by far the most abundant contributor and fruits (55·1%), wines (16·7%) and tea (6·8%) were the main food sources. The present study shows similar results for total dietary flavonoid intakes, but significant differences in flavonoid class intakes, food sources and some characteristics between MED and non-MED countries. These differences should be considered in studies about the relationships between flavonoid intake and chronic diseases.

Dietary intakes and food sources of phytoestrogens in the European Prospective Investigation into Cancer and Nutrition (EPIC) 24-hour dietary recall cohort

BACKGROUND/OBJECTIVES:Phytoestrogens are estradiol-like natural compounds found in plants that have been associated with protective effects against chronic diseases, including some cancers, cardiovascular diseases and osteoporosis. The purpose of this study was to estimate the dietary intake of phytoestrogens, identify their food sources and their association with lifestyle factors in the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort.SUBJECTS/METHODS:Single 24-hour dietary recalls were collected from 36 037 individuals from 10 European countries, aged 35–74 years using a standardized computerized interview programe (EPIC-Soft). An ad hoc food composition database on phytoestrogens (isoflavones, lignans, coumestans, enterolignans and equol) was compiled using data from available databases, in order to obtain and describe phytoestrogen intakes and their food sources across 27 redefined EPIC centres.RESULTS:Mean total phytoestrogen intake was the highest in the UK health-conscious group (24.9 mg/day in men and 21.1 mg/day in women) whereas lowest in Greece (1.3 mg/day) in men and Spain-Granada (1.0 mg/day) in women. Northern European countries had higher intakes than southern countries. The main phytoestrogen contributors were isoflavones in both UK centres and lignans in the other EPIC cohorts. Age, body mass index, educational level, smoking status and physical activity were related to increased intakes of lignans, enterolignans and equol, but not to total phytoestrogen, isoflavone or coumestan intakes. In the UK cohorts, the major food sources of phytoestrogens were soy products. In the other EPIC cohorts the dietary sources were more distributed, among fruits, vegetables, soy products, cereal products, non-alcoholic and alcoholic beverages.CONCLUSIONS:There was a high variability in the dietary intake of total and phytoestrogen subclasses and their food sources across European regions.

Dietary Intakes of Individual Flavanols and Flavonols Are Inversely Associated with Incident Type 2 Diabetes in European Populations

Dietary flavanols and flavonols, flavonoid subclasses, have been recently associated with a lower risk of type 2 diabetes (T2D) in Europe. Even within the same subclass, flavonoids may differ considerably in bioavailability and bioactivity. We aimed to examine the association between individual flavanol and flavonol intakes and risk of developing T2D across European countries. The European Prospective Investigation into Cancer and Nutrition (EPIC)–InterAct case-cohort study was conducted in 8 European countries across 26 study centers with 340,234 participants contributing 3.99 million person-years of follow-up, among whom 12,403 incident T2D cases were ascertained and a center-stratified subcohort of 16,154 individuals was defined. We estimated flavonoid intake at baseline from validated dietary questionnaires using a database developed from Phenol-Explorer and USDA databases. We used country-specific Prentice-weighted Cox regression models and random-effects meta-analysis methods to estimate HRs. Among the flavanol subclass, we observed significant inverse trends between intakes of all individual flavan-3-ol monomers and risk of T2D in multivariable models (all P-trend < 0.05). We also observed significant trends for the intakes of proanthocyanidin dimers (HR for the highest vs. the lowest quintile: 0.81; 95% CI: 0.71, 0.92; P-trend = 0.003) and trimers (HR: 0.91; 95% CI: 0.80, 1.04; P-trend = 0.07) but not for proanthocyanidins with a greater polymerization degree. Among the flavonol subclass, myricetin (HR: 0.77; 95% CI: 0.64, 0.93; P-trend = 0.001) was associated with a lower incidence of T2D. This large and heterogeneous European study showed inverse associations between all individual flavan-3-ol monomers, proanthocyanidins with a low polymerization degree, and the flavonol myricetin and incident T2D. These results suggest that individual flavonoids have different roles in the etiology of T2D.

Resveratrol metabolites in urine as a biomarker of wine intake in free-living subjects: The PREDIMED Study.

Several clinical and epidemiological studies have shown that moderate wine consumption may exert a protective effect against oxidative stress involved in several diseases, such as cardiovascular and neurodegenerative disorders. However, the epidemiological assessment of wine consumption has usually been obtained using self-reported questionnaires containing less reliable information for assessing total intake than nutritional biomarkers. A reliable biomarker for wine consumption is, therefore, needed. To validate urinary resveratrol metabolites (RMs) as a biomarker of wine consumption in a large cohort of free-living subjects, 1000 consecutive subjects entering a substudy of the PREDIMED trial (Prevención con Dieta Mediterránea) were evaluated. Data were collected in a validated semiquantitative food frequency questionnaire. RMs were measured in morning urine by LC-MS/MS. Urinary RM values correlated directly with reported daily amounts of wine consumed (r=0.895; p<0.001). One drink of wine per week can be detected. Using a cut-off of 411.4 nmol/g creatinine, the measurement of urinary RMs could discriminate wine consumers from non-wine consumers with a sensitivity of 93.3% (95% confidence interval (CI) 91.5-94.7%) and a specificity of 92.1% (CI 90.2-93.7%). Urinary RMs fulfill the criteria to be considered as a nutritional biomarker of wine consumption in a large sample of free-living subjects. This biomarker would provide an additional tool for investigating more precisely the relationship between wine consumption and health benefits.

Measuring exposure to the polyphenol metabolome in observational epidemiologic studies: current tools and applications and their limits

Much experimental evidence supports a protective role of dietary polyphenols against chronic diseases such as cardiovascular diseases, diabetes, and cancer. However, results from observational epidemiologic studies are still limited and are often inconsistent. This is largely explained by the difficulties encountered in the estimation of exposure to the polyphenol metabolome, which is composed of ∼500 polyphenols distributed across a wide variety of foods and characterized by diverse biological properties. Exposure to the polyphenol metabolome in epidemiologic studies can be assessed by the use of detailed dietary questionnaires or the measurement of biomarkers of polyphenol intake. The questionnaire approach has been greatly facilitated by the use of new databases on polyphenol composition but is limited by bias as a result of self-reporting. The use of polyphenol biomarkers holds much promise for objective estimation of polyphenol exposure in future metabolome-wide association studies. These approaches are reviewed and their advantages and limitations discussed by using examples of epidemiologic studies on polyphenols and cancer. The current improvement in these techniques, along with greater emphasis on the intake of individual polyphenols rather than polyphenols considered collectively, will help unravel the role of these major food bioactive constituents in disease prevention.

The Association Between Dietary Flavonoid and Lignan Intakes and Incident Type 2 Diabetes in European Populations: The EPIC-InterAct study

OBJECTIVE To study the association between dietary flavonoid and lignan intakes, and the risk of development of type 2 diabetes among European populations. RESEARCH DESIGN AND METHODS The European Prospective Investigation into Cancer and Nutrition-InterAct case-cohort study included 12,403 incident type 2 diabetes cases and a stratified subcohort of 16,154 participants from among 340,234 participants with 3.99 million person-years of follow-up in eight European countries. At baseline, country-specific validated dietary questionnaires were used. A flavonoid and lignan food composition database was developed from the Phenol-Explorer, the U.K. Food Standards Agency, and the U.S. Department of Agriculture databases. Hazard ratios (HRs) from country-specific Prentice-weighted Cox regression models were pooled using random-effects meta-analysis. RESULTS In multivariable models, a trend for an inverse association between total flavonoid intake and type 2 diabetes was observed (HR for the highest vs. the lowest quintile, 0.90 [95% CI 0.77–1.04]; P value trend = 0.040), but not with lignans (HR 0.88 [95% CI 0.72–1.07]; P value trend = 0.119). Among flavonoid subclasses, flavonols (HR 0.81 [95% CI 0.69–0.95]; P value trend = 0.020) and flavanols (HR 0.82 [95% CI 0.68–0.99]; P value trend = 0.012), including flavan-3-ol monomers (HR 0.73 [95% CI 0.57–0.93]; P value trend = 0.029), were associated with a significantly reduced hazard of diabetes. CONCLUSIONS Prospective findings in this large European cohort demonstrate inverse associations between flavonoids, particularly flavanols and flavonols, and incident type 2 diabetes. This suggests a potential protective role of eating a diet rich in flavonoids, a dietary pattern based on plant-based foods, in the prevention of type 2 diabetes.

Dietary Protein Intake and Incidence of Type 2 Diabetes in Europe: The EPIC-INTERACT Case-Cohort Study

OBJECTIVE The long-term association between dietary protein and type 2 diabetes incidence is uncertain. We aimed to investigate the association between total, animal, and plant protein intake and the incidence of type 2 diabetes. RESEARCH DESIGN AND METHODS The prospective European Prospective Investigation into Cancer and Nutrition (EPIC)-InterAct case-cohort study consists of 12,403 incident type 2 diabetes cases and a stratified subcohort of 16,154 individuals from eight European countries, with an average follow-up time of 12.0 years. Pooled country-specific hazard ratios (HRs) and 95% CI of prentice-weighted Cox regression analyses were used to estimate type 2 diabetes incidence according to protein intake. RESULTS After adjustment for important diabetes risk factors and dietary factors, the incidence of type 2 diabetes was higher in those with high intake of total protein (per 10 g: HR 1.06 [95% CI 1.02–1.09], Ptrend < 0.001) and animal protein (per 10 g: 1.05 [1.02–1.08], Ptrend = 0.001). Effect modification by sex (P < 0.001) and BMI among women (P < 0.001) was observed. Compared with the overall analyses, associations were stronger in women, more specifically obese women with a BMI >30 kg/m2 (per 10 g animal protein: 1.19 [1.09–1.32]), and nonsignificant in men. Plant protein intake was not associated with type 2 diabetes (per 10 g: 1.04 [0.93–1.16], Ptrend = 0.098). CONCLUSIONS High total and animal protein intake was associated with a modest elevated risk of type 2 diabetes in a large cohort of European adults. In view of the rapidly increasing prevalence of type 2 diabetes, limiting iso-energetic diets high in dietary proteins, particularly from animal sources, should be considered.