Nutrition epidemiology of flavan-3-ols: the known unknowns

Nutritional epidemiology has an important role, as it can provide long-term data from large populations and does not rely on surrogate markers for morbidity/mortality. Meaningful interpretation and applications of outcomes from epidemiological studies depend on the accurate assessment of dietary intake, which is currently mainly based on a combination of self-reporting and food composition data. Flavan-3-ols are a group of bioactives (non-essential dietary components with significant impact on health) that is a possible candidate for the development of dietary recommendations. The breadth of data available on their effect on health also provides the basis for investigating the suitability of the methods currently used in nutritional epidemiology to assess the health effects of bioactives. The outcomes of this assessment demonstrate that the limitations of currently used methods make it virtually impossible to estimate intake accurately from self-reported dietary data. This is due to the limitations of self-reporting, especially from food-frequency questionnaires, and the inability of currently used methods to deal with the high variability of food composition. Indeed, the estimated intake of flavan-3-ols, can only be interpreted as a marker of specific dietary patterns, but not as the actual intake amount. The interpretation of results from such studies are fraught with serious limitations, especially for establishing associations between intake and health and the development of dietary recommendations. Alternative assessment not affected by these limitations, such as biomarkers, are required to overcome these limitations. The development of nutritional biomarkers is therefore crucial to investigate the health effect of bioactives

Sugar intake among German adolescents: trends from 1990-2016 based on biomarker excretion in 24-h urine samples

Trend analyses based on dietary records suggest decreases in the intakes of total (TS), added (AS) and free sugar (FS) since 2005 among children and adolescents in Germany. In terms of age trends, TS intake decreased with increasing age. However, self-reported sugar intake in epidemiological studies is criticized, as it may be prone to bias due to selective underreporting. Furthermore, adolescents are more susceptible to underreporting than children. We thus analyzed time and age trends in urinary fructose excretion (FE), sucrose excretion (SE) and the sum of both (FE+SE) as biomarkers for sugar intake among 8.5-16.5-year-old adolescents. Urinary sugar excretion was measured by UPLC-MS/MS in 997 24-h urine samples collected from 239 boys and 253 girls participating in the DONALD study cohort between 1990 and 2016. Time and age trends of log-transformed FE, SE and FE+SE were analyzed using polynomial mixed-effects regression models. Between 1990 and 2016 FE as well as FE+SE decreased (linear time trend: p=0.0272 and p<0.0001, respectively). A minor increase in excretion during adolescence was confined to FE (linear age trend: p=0.0017). The present 24-h excretion measurements support a previously reported dietary-record based decline in sugar intake since 2005. However, the previous seen dietary record-based decrease in TS from childhood to late adolescence was not confirmed by our biomarker analysis, suggesting a constant sugar intake for the period of adolescence

Phytoestrogen consumption and association with breast, prostate and colorectal cancer in EPIC Norfolk

Phytoestrogens are polyphenolic secondary plant metabolites that have structural and functional similarities to 17β-oestradiol and have been associated with a protective effect against hormone-related cancers. Most foods in the UK only contain small amounts of phytoestrogens (median content 21 μg/100 g) and the highest content is found in soya and soya-containing foods. The highest phytoestrogen content in commonly consumed foods is found in breads (average content 450 μg/100 g), the main source of isoflavones in the UK diet. The phytoestrogen consumption in cases and controls was considerably lower than in Asian countries. No significant associations between phytoestrogen intake and breast cancer risk in a nested case-control study in EPIC Norfolk were found. Conversely, colorectal cancer risk was inversely associated with enterolignan intake in women but not in men. Prostate cancer risk was positively associated with enterolignan intake, however this association became non-significant when adjusting for dairy intake, suggesting that enterolignans can act as a surrogate marker for dairy or calcium intake

Impact of high drinking water nitrate levels on the endogenous formation of apparent N-nitroso compounds in combination with meat intake in healthy volunteers

BACKGROUND: Nitrate is converted to nitrite in the human body and subsequently can react with amines and amides in the gastrointestinal tract to form N-nitroso compounds (NOCs), which are known to be carcinogenic in animals. Humans can be exposed to nitrate via consumption of drinking water and diet, especially green leafy vegetables and cured meat. The contribution of nitrate from drinking water in combination with meat intake has not been investigated thoroughly. Therefore, in the present pilot study, we examined the effect of nitrate from drinking water, and its interaction with the consumption of white and processed red meat, on the endogenous formation of NOCs, taking into account the intake of vitamin C, a nitrosation inhibitor. METHODS: Twenty healthy subjects were randomly assigned to two groups consuming either 3.75 g/kg body weight (maximum 300 g per day) processed red meat or unprocessed white meat per day for two weeks. Drinking water nitrate levels were kept low during the first week (< 1.5 mg/L), whereas in week 2, nitrate levels in drinking water were adjusted to the acceptable daily intake level of 3.7 mg/kg bodyweight. At baseline, after 1 and 2 weeks, faeces and 24 h urine samples were collected for analyses of nitrate, apparent total N-nitroso compounds (ATNC), compliance markers, and genotoxic potential in human colonic Caco-2 cells. RESULTS: Urinary nitrate excretion was significantly increased during the high drinking water nitrate period for both meat types. Furthermore, levels of compliance markers for meat intake were significantly increased in urine from subjects consuming processed red meat (i.e. 1-Methylhistidine levels), or unprocessed white meat (i.e. 3-Methylhistidine). ATNC levels significantly increased during the high drinking water nitrate period, which was more pronounced in the processed red meat group. Genotoxicity in Caco-2 cells exposed to faecal water resulted in increased genotoxicity after the interventions, but results were only significant in the low drinking water nitrate period in subjects consuming processed red meat. Furthermore, a positive correlation was found between the ratio of nitrate/vitamin C intake (including drinking water) and the level of ATNC in faecal water of subjects in the processed red meat group, but this was not statistically significant. CONCLUSIONS: Drinking water nitrate significantly contributed to the endogenous formation of NOC, independent of the meat type consumed. This implies that drinking water nitrate levels should be taken into account when evaluating the effect of meat consumption on endogenous formation of NOC

Oat bran, but not its isolated bioactive β-glucans or polyphenols, have a bifidogenic effect in an in vitro fermentation model of the gut microbiota

Wholegrain oats are known to modulate the human gut microbiota and have prebiotic properties (increase the growth of some health-promoting bacterial genera within the colon). Research to date mainly attributes these effects to the fibre content; however, oat is also a rich dietary source of polyphenols, which may contribute to the positive modulation of gut microbiota. In vitro anaerobic batch-culture experiments were performed over 24 h to evaluate the impact of two different doses (1 and 3 % (w/v)) of oat bran, matched concentrations of β-glucan extract or polyphenol mix, on the human faecal microbiota composition using 16S RNA gene sequencing and SCFA analysis. Supplementation with oats increased the abundance of Proteobacteria (P <0·01) at 10 h, Bacteroidetes (P <0·05) at 24 h and concentrations of acetic and propionic acid increased at 10 and 24 h compared with the NC. Fermentation of the 1 % (w/v) oat bran resulted in significant increase in SCFA production at 24 h (86 (sd 27) v. 28 (sd 5) mm; P <0·05) and a bifidogenic effect, increasing the relative abundance of Bifidobacterium unassigned at 10 h and Bifidobacterium adolescentis (P <0·05) at 10 and 24 h compared with NC. Considering the β-glucan treatment induced an increase in the phylum Bacteroidetes at 24 h, it explains the Bacteriodetes effects of oats as a food matrix. The polyphenol mix induced an increase in Enterobacteriaceae family at 24 h. In conclusion, in this study, we found that oats increased bifidobacteria, acetic acid and propionic acid, and this is mediated by the synergy of all oat compounds within the complex food matrix, rather than its main bioactive β-glucan or polyphenols. Thus, oats as a whole food led to the greatest impact on the microbiota

Association between dietary phyto-oestrogens and bone density in men and postmenopausal women

Phyto-oestrogens have been associated with a decreased risk for osteoporosis, but results from intervention and observational studies in Western countries have been inconsistent. In the present study, we investigated the association between habitual phyto-oestrogen intake and broadband ultrasound attenuation (BUA) of the calcanaeum as a marker of bone density. We collected 7 d records of diet, medical history and demographic and anthropometric data from participants (aged 45–75 years) in the European Prospective Investigation into Cancer-Norfolk study. Phyto-oestrogen (biochanin A, daidzein, formononetin; genistein, glycitein; matairesinol; secoisolariciresinol; enterolactone; equol) intake was determined using a newly developed food composition database. Bone density was assessed using BUA of the calcanaeum. Associations between bone density and phyto-oestrogen intake were investigated in 2580 postmenopausal women who were not on hormone replacement therapy and 4973 men. Median intake of total phyto-oestrogens was 876 (interquartile range 412) μg/d in postmenopausal women and 1212 (interquartile range 604) μg/d in men. The non-soya isoflavones formononetin and biochanin A were marginally significant or significantly associated with BUA in postmenopausal women (β = 1·2; P < 0·1) and men (β = 1·2; P < 0·05), respectively; enterolignans and equol were positively associated with bone density in postmenopausal women, but this association became non-significant when dietary Ca was added to the model. In the lowest quintile of Ca intake, soya isoflavones were positively associated with bone density in postmenopausal women (β = 1·4; P < 0·1). The present results therefore suggest that non-soya isoflavones are associated with bone density independent of Ca, whereas the association with soya or soya isoflavones is affected by dietary Ca

Cocoa flavanol intake improves endothelial function and Framingham Risk Score in healthy men and women: a randomised, controlled, double-masked trial: the Flaviola Health Study

Cocoa flavanol (CF) intake improves endothelial function in patients with cardiovascular risk factors and disease. We investigated the effects of CF on surrogate markers of cardiovascular health in low risk, healthy, middle-aged individuals without history, signs or symptoms of CVD. In a 1-month, open-label, one-armed pilot study, bi-daily ingestion of 450 mg of CF led to a time-dependent increase in endothelial function (measured as flow-mediated vasodilation (FMD)) that plateaued after 2 weeks. Subsequently, in a randomised, controlled, double-masked, parallel-group dietary intervention trial (Clinicaltrials.gov: NCT01799005), 100 healthy, middle-aged (35–60 years) men and women consumed either the CF-containing drink (450 mg) or a nutrient-matched CF-free control bi-daily for 1 month. The primary end point was FMD. Secondary end points included plasma lipids and blood pressure, thus enabling the calculation of Framingham Risk Scores and pulse wave velocity. At 1 month, CF increased FMD over control by 1·2 % (95 % CI 1·0, 1·4 %). CF decreased systolic and diastolic blood pressure by 4·4 mmHg (95 % CI 7·9, 0·9 mmHg) and 3·9 mmHg (95 % CI 6·7, 0·9 mmHg), pulse wave velocity by 0·4 m/s (95 % CI 0·8, 0·04 m/s), total cholesterol by 0·20 mmol/l (95 % CI 0·39, 0·01 mmol/l) and LDL-cholesterol by 0·17 mmol/l (95 % CI 0·32, 0·02 mmol/l), whereas HDL-cholesterol increased by 0·10 mmol/l (95 % CI 0·04, 0·17 mmol/l). By applying the Framingham Risk Score, CF predicted a significant lowering of 10-year risk for CHD, myocardial infarction, CVD, death from CHD and CVD. In healthy individuals, regular CF intake improved accredited cardiovascular surrogates of cardiovascular risk, demonstrating that dietary flavanols have the potential to maintain cardiovascular health even in low-risk subjects

5‐(3′,4′‐Dihydroxyphenyl)‐γ‐Valerolactone is a substrate for human paraoxonase: a novel pathway in flavan‐3‐ol metabolism

Scope Dietary flavan-3-ols are known to mediate cardiovascular benefits. Currently, it is assumed that the levels of flavan-3-ol catabolites detected in humans, 5-(3ʹ,4ʹ-dihydroxyphenyl)-γ-valerolactone (γVL) and 5-(3ʹ,4ʹ-dihydroxyphenyl)-γ-valeric acid (γVA), and their corresponding phase II metabolites, are determined exclusively by the action of the gut microbiome. However, a family of human proteins, paraoxonase (PON), can theoretically hydrolyze γVL metabolites into the corresponding γVAs. This study aims to determine if PON is involved in γVL and γVA metabolism in humans. Methods and results A rapid conversion of γVL into γVA is detected in serum ex vivo (half-life = 9.8 ± 0.3 min) that is catalyzed by PON1 and PON3 isoforms. Phase II metabolites of γVL are also reacted with PON in serum. Following an intake of flavan-3-ol in healthy males (n = 13), the profile of γVA metabolites detected is consistent with that predicted from the reactivity of γVL metabolites with PON in serum. Furthermore, common PON polymorphisms are evaluated to assess the use of γVL metabolites as biomarkers of flavan-3-ol intake. Conclusion PONs are involved in flavan-3-ol metabolic pathway in humans. PON polymorphisms have a minor contribution to inter-individual differences in the levels of γVL metabolites, without affecting their use as a nutritional biomarker