Wine grape pomace flour improves blood pressure, fasting glucose and protein damage in humans: a randomized controlled trial

Lifestyle changes, including healthy eating, exercise, and giving up smoking, constitute the most powerful tools to fight chronic diseases, which are the main causes of disability and death in the developed and developing world [1]. Among various conditions, patients with metabolic syndrome are at very high chronic disease risk. The increased risk associated with metabolic syndrome appears to be linked to higher levels of oxidative stress [2]. Recently, much attention has been paid to evidence that postprandial oxidative stress contributes to the development of cardiovascular disease and diabetes [3–7]. Thus, managing oxidative stress can be an important factor in managing the risk of chronic disease.

Mediterranean diets (MD) are considered as valuable preventive measures since several studies have shown that their intake is associated with lower incidence and prevalence of chronic diseases, mainly cardiovascular diseases, and longer life expectancy in the countries where a MD is consumed or in subgroups exhibiting a higher MD score [8, 9]. In 2013, the PREDIMED study, a primary prevention trial, reported the benefits of a MD in reducing the incidence of major cardiovascular events among a high-risk population [10]. A relatively large consumption of olive oil, poultry, fish, fruits, vegetables and cereals, a moderate intake of red wine, and a low consumption of red meats characterize MD.

The health effects of different diets can be explained by the positive functional properties of key nutrients. The bioactive components that apparently support the healthy properties of MD are antioxidants, fiber, mono and polyunsaturated fats, and phytochemicals [11]. Vegetables, fruits and legumes, which are rich in antioxidants and fiber, are associated with a lower incidence of metabolic syndrome and chronic diseases [12, 13]. Wine is also a source of polyphenol antioxidants and, when consumed in moderation, is associated with reduced risk of cardiovascular diseases [14].

Traditionally, dietary fiber and antioxidants are addressed separately as unrelated dietary compounds. Saura-Calixto has introduced the concept of antioxidant dietary fiber because ≈50 % of total dietary antioxidants, mainly polyphenolics, are transported along the small intestine linked to dietary fiber [15, 16]. These antioxidants are released from the fiber matrix within the colon by the action of bacterial microbiota, generating bioactive metabolites and an antioxidant environment [17]. Thus, it is proposed that the transport of dietary antioxidants through the gastrointestinal tract may be an essential function of dietary fiber [16].

Antioxidant dietary fiber is present in fruits and vegetables and derived products, such as grapes and apple pomace and mango peel powder [18]. Thus, fruit byproducts represent rich sources of dietary fiber and antioxidants with beneficial bioactive properties. In this regard, wine grape pomace flour (WGPF), made from seed and skin residues obtained from grapes during winemaking, contains high levels of phenolic compounds and dietary fiber.

In this study, we prepared flour from WGPF as a functional food ingredient to increase the daily intake of dietary fiber and bioactive antioxidant compounds. Based on the current evidence, we hypothesized that the daily intake of WGPF reduced the metabolic syndrome in adult subjects. On this basis, we conducted a longitudinal intervention study including male volunteers with at least one component of metabolic syndrome, who were randomly assigned to either the intervention group or the control group, to test the effects of dietary supplementation with this WGPF on components of metabolic syndrome, plasma antioxidants and protein damage.

Wine grape pomace flour prepared from red wine grape, Cabernet Sauvignon variety, is especially rich in antioxidants and fiber, two bioactive components of human diet with healthy properties. Our hypothesis was that WGPF consumption decreases the risk of developing metabolic syndrome, supporting its potential benefit for preventing cardiovascular disease and diabetes. We chose a group of men with at least one diagnostic criterion of metabolic syndrome in order to enhance the chance of detecting significant effects.

WGPF contains an antioxidant/fiber complex, in which polyphenols are transported along the gastrointestinal tract linked to fiber components. WGPF includes soluble or extractable polyphenols with low or intermediate molecular mass, which appear to be absorbed from the digestive tract with systemic effects [15]. Nevertheless, many WGPF polyphenols are non-absorbable/non-extractable high molecular mass proanthocyanidins [16]. After reaching the large intestine, these antioxidants are processed by colonic microbiota producing metabolites with physiological properties [19].

Antioxidant dietary fiber and extractable and non-extractable polyphenol content in pomace obtained during winemaking usually ranges from 50 to 75 %, 1 to 9 % and 15 to 30 % by dry matter, respectively [15]. Our WGPF contains 52 % dietary fiber (7 % soluble and 93 % insoluble), and 4.4 % of extractable polyphenols with an antioxidant capacity of 362.9 ORAC (µmol TE/g dry matter). A similar product obtained from Cencibel wine grape in Spain showed an antioxidant capacity of 214.2 ORAC units [20]. Thus, flour derived from Cabernet Sauvignon WGPF exhibits a composition profile that suggests potential health benefits.

Indeed, dietary fiber (or non-starch polysaccharides) is a collective term for a variety of plant substances resistant to digestion by human gastrointestinal enzymes. Structural fiber (cellulose, lignin, and hemicelluloses) is insoluble, whereas gel-forming fiber (pectins, gums, and mucilages) is water-soluble [21]. Intervention trials have shown that soluble dietary fiber lowers blood cholesterol concentrations, reduces blood pressure, promotes body weight loss, and improves insulin sensitivity, eventually leading to reduced risk of cardiovascular disease [21, 22].

We observed a statistically significant lowering of blood pressure after WGPF intake. Previous studies have shown that soluble dietary fiber reduces blood pressure [21, 22]. Furthermore, fiber, especially soluble types, also improves intestinal potassium and magnesium absorption, which may have an indirect favorable effect on blood pressure [22]. However, the WGPF used in this study contains only 7 % of soluble fiber. Consequently, it is difficult to attribute the hypotensive effect exclusively to this component. Nevertheless, extracts of polyphenols from grape seeds, grape pomace, and/or wine have shown positive effects on blood pressure. Polyphenols activate endothelial nitric oxide synthase (eNOS) and this effect has been suggested as the mechanism that lowers blood pressure [24]. The bioavailability of single and 2- or 3-polymer unit polyphenols also support this mechanism [25].

A relatively small fraction (1–9 %) of these polyphenols associated with dietary fiber are extractable from WGPF [15], so only a minor amount of this form of antioxidants are absorbed by the intestine. However, plasma antioxidant capacity increases postprandially, becoming significant 8 h after consuming grape pomace [26]. This delayed increase is explained by the presence of more complex polyphenols (high polymerization proantocyanidins) linked to fiber in grape pomace. These proantocyanidins reach the colon, where they are partially fermented by bacterial microflora along with indigestible carbohydrates. Indeed, colonic fermentation of grape pomace generates metabolites that are detectable in human plasma [17]. Fiber-associated antioxidant products released by fermentation may explain the reduction in blood pressure after WGPF supplementation. Additional studies are required to test this hypothesis.

Moreover, we observed a significant reduction in the plasma concentrations of fasting glucose in the WGPF supplemented group. There is evidence that dietary fiber consumption is associated with lower fasting glucose levels in diabetic individuals [27]. The mechanisms associated with this beneficial effect have not yet been elucidated [28].

We also found, a statistically significant difference, when comparing the change in postprandial insulin concentrations along the study, among control and WGPF groups. There have been few intervention studies on the effects of dietary fiber on postprandial glucose and insulin. Recently, an intervention trial reported that a whole-grain cereal-based diet significantly lowers postprandial plasma insulin concentration in individuals with metabolic syndrome [29]. It is likely that bioactive compounds in this fiber-enriched ingredient, such as trace minerals, phenolic antioxidants, vitamin E or phytoestrogens, operating by different unknown pathways may contribute to this positive postprandial metabolic effect [30].

In prospective observational studies, normal postprandial increases in blood glucose and triglyceride levels have been linked to the risk of cardiovascular disease and diabetes [3–7]. The evidence suggests that postprandial hyperglycemia and hypertriglyceridemia cause oxidative stress. Postprandial oxidative stress occurs within minutes to hours following consumption of a meal rich in fat, carbohydrates and protein [31, 32]. Over the long-term, oxidative stress leads to insulin resistance, pancreatic β-cell dysfunction, abnormal serum lipids, coagulopathy, systemic inflammation, and endothelial dysfunction [33]. Exaggerated postprandial spikes in glucose and lipids can also trigger inflammation, endothelial dysfunction, and sympathetic hyperactivity [33]. With multiple repetition on a daily basis, these postprandial changes that lead to atherosclerosis and diabetes. Dietary strategies aimed at attenuating these pathogenic alterations provide new opportunities to reduce the risk of chronic disease. In fact, consumption of red wine polyphenols and grape seed extract with a meal rich in fat, carbohydrates and protein significantly reduces postprandial oxidative stress [34–36]. We hypothesize that dietary fiber rich in antioxidant in our WGPF reduces postprandial oxidative stress and plasma oxidative damage when consumed with meals. In accord, we observed a significant decrease in protein oxidation in WGPF supplemented group. Oxidative damage reduction may partially explain its beneficial effect on blood pressure and glucose homeostasis, with important implications for the prevention and management of metabolic syndrome [24]. Further investigation is needed to validate this proposal.

In contrast to earlier studies with pomace with animal models and humans [16, 20, 37, 38], our WGPF did not influence serum cholesterol levels. In a human trial, supplementation with grape pomace flour obtained from Cencibel wine grapes significantly decreased total cholesterol in normo- and hypercholesterolemic subjects [20]. Furthermore, LDL cholesterol and triglycerides diminished significantly only in hypercholesterolemic subjects, who represented 38 % of study participants and were mostly women (65 %) [20]. Although the fiber and antioxidant composition and dose were similar to those in our WGPF, we did not observe any effect on lipid profile. However, our study design included only male subjects with 40 % hypercholesterolemic (total cholesterol >200 mg/dL) at baseline. Alternatively, the lack of effect in our study may be attributed to differential functional properties of wine grape types (e.g., Cabernet Sauvignon versus Cencibel). In hamsters, different effects were observed of a diverse variety of grape pomace in plasma total cholesterol and LDL-cholesterol concentrations [39].

In terms of antioxidant capacity, measured as DPPH, and vitamin C, we observed similar changes in both group. They consumed same food at the cafeteria of work place, so these variations could be explained by the diet.

Interestingly we found a significant increase in plasma concentrations of γ-tocopherol and δ-tocopherol in the WGPF supplemented group. Also, a statistically significant difference, when comparing the change in γ-tocopherol concentrations along the study, among control and WGPF groups was detected. The raise in plasma concentrations of γ- and δ-tocopherols in the WGPF supplemented group could be explained by the antioxidant effect of WGPF polyphenols in the gastrointestinal tract [40]. This effect would depend on the quantity of tocopherols in the entire diet. Volunteers consumed soybean oil that is a good source of γ-tocopherol (80 mg/100 g) and δ-tocopherol (27 mg/100 g) [41]. WGPF by its antioxidant capacity would protect these vitamins from its oxidation, improving its bioavailability and increasing the antioxidant defenses of the volunteers in terms of plasma concentration of vitamin E.

The significance of plasma increase of these forms of vitamin E on health is matter of discussion [41]. Most research of vitamin E has focused on α-tocopherol, because it is the predominant form of vitamin E in tissues. Despite well-documented antioxidant and other beneficial effects, supplementation with α-tocopherol has failed to offer consistent benefits to prevention of chronic diseases, as cancer and cardiovascular diseases, in clinical intervention studies in people with adequate nutrient status. Current studies show that other forms of vitamin E, such as γ-tocopherol and δ-tocopherol have unique antioxidant and anti-inflammatory properties in prevention and therapy against chronic diseases. These vitamin E forms scavenge reactive nitrogen species, inhibit cyclooxygenase- and 5-lipoxy-catalyzed eicosanoids, and suppress proinflammatory signaling. It has been suggested that γ-tocopherol and δ-tocopherol may be useful against inflammation-associated diseases [41].

Strength of the current study was that workers had a similar diet during weekdays, because they were offered same lunch menus at the cafeteria of the company. Also we supervised daily the intake of WGPF during lunch as well as similar consumption of regular bread in control subjects. However, one limitation of the study was that the number of participants who completed the protocol in each group was relatively small, because six and three participants quit the study in WGPF and control groups, respectively. Second, this study was an open-labelled trial that may have led to potential biases in the subjects as well as in the researchers. Third, we did not measured differences in caloric intake at the end of the interventions, but no changes in abdominal obesity or body weight were detected during the study in both groups suggesting that no major differences in energy balance occurred during the study. In addition, both groups were encouraged to maintain their regular eating habits and lifestyle.

The consumption of WGPF as a food supplement to a regular diet, improves some components of metabolic syndrome, as blood pressure and glycaemia. Also, postprandial insulin levels, measured in an OGTT, were reduced. Moreover, fiber and polyphenols-containing WGPF increased antioxidant defenses, rising γ- and δ-tocopherols and diminishing oxidative protein damage, contributing to a better handling of oxidative stress. WGPF seems to be a promising functional ingredient that can be incorporated into different food matrices to increase fiber and antioxidant consumption and eventually contribute to the management of chronic disease burden.