Valorization of grape by-products through extraction and encapsulation of bioactive compounds

There is a current trend for reutilization of food industry by-products, focused on the extraction of bioactive compounds and application in the development of new feed, food, cosmetic or nutraceutical ingredients. The wine industry is no exception and generates up to 7 million tons of wasted biomass in the form of grape bagasse, which is reported to be rich in high added value compounds such as neutral polysaccharides, structural proteins and phenolic compounds. This project aimed to valorize a major wine production by-product, grape pomace, through the production and characterization of a high valued extract for food industry, improvement of intestinal bioavailability through a gastrointestinal delivery system, and application in the development of a functional beverage as a proof-of-concept. Optimization of grape pomace extract (GPE) was performed through the optimization of xylooligosaccharides (XOS) production, using conventional (acid and alkaline extraction) or enzymatic processes. Conventional methods recovered 21.8 to 74.6% and 5.2 to 96.3% of total XOS, for acid and alkaline processes, respectively. Enzymatic process extracted up to 88.7 ± 0.12% of total. Taking into account the extraction yield, the quality of the extract and the fact of being considered a green process, the enzymatically produced extract with A. niger xylanases (GPE) was selected to continue this study. GPE was characterized for its chemical composition and biological activities, and submitted to the simulation of gastrointestinal digestion to understand how these properties are affected by the gastrointestinal environment. The extract presented high content of dietary soluble fiber (26.1 ± 1.59 g.100g-1) and other carbohydrates, including XOS, minerals and phenolics. In vitro simulated digestion allowed to conclude that XOS were resistant to gastric conditions but phenolics were not. The use of 2% (w/v) of GPE proved to be a potential carbon source that could be fermented by different probiotics, even after digestion. GPE also exhibited strong antioxidant and antimicrobial activity against different pathogens, however, after digestion, these bioactivities were strongly reduced. These results allow to conclude that GPE had low intestinal bioavailability due to action of digestion enzymes and acidic pH. Aiming at improving its bioavailabiliy, GPE was then encapsulated into chitosan or alginate microparticles (MPs) , in order to improve its bioavailability. MPs were characterized for their size, polydispersity, zeta potential and total phenolics (TPC) association efficiency, and the simulation of gastrointestinal digestion was performed to evaluate the release profile of polyphenols, the antioxidant and antimicrobial activities, and permeability of phenolics and XOS across Caco-2/ HT29-MTX cell layer. Also, chitosan was modified with a fluorescent probe for cellular uptake studies. GPE-loaded alginate MPs presented size of 523 nm, polydispersity of 0.112, zeta potential of -15.0 mV and 68% of association efficiency of polyhenols. GPE-loaded chitosan MPs presented size of 853 nm, polydispersity of 0.358, zeta potential of 14.9 mV and 65% of association efficiency of polyphenols. Both systems allowed the delivery of GPE in the intestine, increasing the bioavailability of different polyphenols, the antioxidant and antimicrobial activities. Permeability of XOS across the intestinal cell layer decreased from 45% to 7.9 and 15.7%, after encapsulation in alginate or chitosan MPs, respectively. Modification of chitosan MPs with Cyanine5.5 for cellular uptake studies did not affect the biocompatibility with intestinal cells, and confocal microscopy analysis confirmed the integrity of these cells tight junctions. As a proof of concept, it was developed a coconut beverage through the incorporation 2.5% (w/v) of the encapsulated GPE. Physicochemical stability under two storage conditions (freeze-dried at room temperature or liquid at 4 ˚C) was evaluated along 60 days. The antimicrobial and prebiotic potential after the digestion of the beverage were assessed using different microbial strains. Room-temperature storage accelerated the rate of degradation of phenolics, comparing to storage at 4 ˚C. Alginate and chitosan functional beverages decreased the growth of different pathogens and promoted the growth of different probiotics. Sensory analysis allowed to conclude that the incorporation of particles did not promoted significant differences in most of evaluated attributes. The results from this work will contribute for the sustainability of wine industry in circular economy context, through the development of value-added ingredients with positive biological impact.