4. Discussion
WAHA
In this sub-study of the WAHA trial, we showed that the daily ingestion of walnuts for 2 years significantly increased the total dietary polyphenols and the subclasses of flavonoids, flavanols and phenolic acids in healthy elderly participants. To our knowledge, this is the first study to show that the inclusion of a single food (i.e., walnuts), with no other changes made to the usual diet, could significantly increase the total polyphenol intake. As expected, those who ate walnuts daily also showed higher intakes of energy, fiber, total fat, and unsaturated fatty acids. The results of this trial also show that participants in the walnut group consumed significantly higher amounts of total polyphenols and flavonoids (flavones, flavonols, and anthocyanidins), flavanols, and phenolic acids from nuts compared to those in the control group. This finding demonstrates that a single food such as walnuts can increase the intakes of total polyphenols and the polyphenol subclasses except for lignans. The walnut group had a higher intake of total polyphenols from fruits, and the flavanols and lignans from vegetables. The median daily intake of the total polyphenols of the control and walnut groups at 1897 mg/d and 2480 mg/d, respectively, of this elderly cohort residing in California was higher compared to that reported in adults in the U.S. by the National Health and Nutrition Examination Survey (NHANES) at 884 mg per 1000 kcal per day [While the walnut group showed a higher daily intake of polyphenols, this was not reflected in the urinary excretion of polyphenols tested in the spot urine samples obtained at the baseline, and at the end of either year 1 or year 2. Increased polyphenol metabolites have been identified in urine following the consumption of plant-based foods, suggesting that selected urinary polyphenols could be useful biomarkers to assess the intakes of polyphenol-rich foods and diets [In relation to the total polyphenol concentrations in the spot urine samples, our analyses did not show associations either with the dietary total polyphenols or flavanols and phenolic acid subclasses, but disclosed an inverse association with an intake of the flavonoid (flavanones + flavones) category. Studies in which the total urine polyphenols were measured with the F–C assay have shown weak to moderate associations with dietary polyphenol intakes in the 24-h [The quality of phenolic compounds present in walnuts is diverse, ranging from simple phenolic acids and flavonoids to highly polymerized molecules such as tannins. Walnut phenolics are usually found at the highest concentration in the seed coat (also called the pellicle) surrounding the edible kernel and may be bound to other plant components such as carbohydrates and proteins. Consequently, some polyphenolic compounds might not be released in compositional testing studies. Walnuts are distinguished by the predominance of the hydrolysable gallotannins, glansrins (ellagitanins), and ellagic acid and the condensed tannins that are polymers of flavan-3-ol (flavanol) catechin and epicatechin subunits. The Polyphenol-Explorer 3.6 database reports an average amount of total polyphenol assayed by the F–C reagent as 1575 mg/100 g of kernel. Average amounts per 100 g of the total flavonoids, flavanols, and phenolic acids in walnuts are reported as 65 mg, 60 mg, and 449 mg, respectively [Aside from the wide diversity and complexity of the phenolic substances found in walnuts, a number of other factors complicate efforts to obtain the exact accounts of their polyphenol composition. The concentration of phenolic compounds from different genomic walnut species and cultivars have been found to vary widely, with mean coefficients of variation of 25% or greater [It is important to note that the concentration of polyphenol in urine is determined by factors beyond the walnut phenol content and its structural matrix, but is related to human physiology, mainly sex and age, along with factors such as digestive and metabolic efficiency and the gut microbiota [Our study has many strengths. The WAHA study has a significantly long duration of intervention (2 years). The study also includes a relatively large number of participants who demonstrated excellent compliance, with a retention rate of 90%. Moreover, the dietary intake data were extensive, having been acquired using multiple 24-h recalls (up to five recalls) obtained throughout the 2 year period and carefully matched with Phenol-Explorer values to obtain a profile of its polyphenol content. The main limitation was that the urine samples were obtained from fasting participants following the first morning void, and as such, may not have captured a large enough quantity and diversity of phenolic metabolites excreted in a 24-h urine or in a longer collection period. It is well-known that the half-life of most polyphenol metabolites is relatively short and typically appear in urine within 1 to 24 h following ingestion [