Exploitation of agricultural and forestry waste: from the recovery of bioactive compounds for nutraceutical applications to the production of innovative materials

Agricultural and food industries generate substantial waste that, remaining underexploited, entails environmental, economic, and social issues. The shells of Pistacia vera, Prunus dulcis, Corylus avellana, and Juglans regia species represent the most abundant by-products and have the potential to be further processed into value-added nutraceutical ingredients and other innovative materials. In this thesis, dry fruit shells have been employed as sources of valuable bioactive compounds. Specifically, the Response Surface Methodology (RSM) coupled with Box-Behnken design (BBD) models were adopted to optimize entirely eco-sustainable microwave-assisted extraction (MAE) protocols. The optimized shell extracts exhibited the highest total phenolic content (TPC) and antioxidant/antiradical activity. Furthermore, in this thesis, MAE method was applied for the first time to recover antioxidants from pistachio and hazelnut shells. Although MAE was previously employed to extract bioactive compounds from almond and walnut shells, the herein optimized MAE conditions led to higher phenolics recovery rates. Moreover, MAE protocols were carefully designed to support their application at an industrial level, where the extraction of natural products using microwave irradiation is already a reality. The phytochemical composition of the optimized shell extracts was determined by hyphenated techniques, namely high-performance liquid chromatography/electrospray ionization ion-trap tandem mass spectrometric (HPLC/ESI-MS/MS) analyses. A thorough qualitative and quantitative analysis of mass spectrometric data allowed the identification of phenolic compounds and lipids as the predominant organic constituents in all the extracts. Remarkably, the phytochemical composition of Pistacia vera shells has never been reported before. Furthermore, the abundant presence of alkyl phenols, namely anacardic acids (AAs), in both P. vera and J. regia shells was herein uncovered for the first time, highlighting such biomasses as novel sources of bioactive AAs. Unique insights into the potential biological properties of the optimized shell extracts were provided in this thesis. Particularly, their reactive oxygen species (ROS) scavenging capacity, their anti-glycation properties, and their inhibitory behaviour towards pancreatic lipase, α-glucosidase, α-amylase, and tyrosinase, molecular targets of notable therapeutic importance, were in vitro investigated. Considering the remarkable anti-glycation and hypoglycemic properties observed for the AAs-rich extracts from P. vera and J. regia shells, AAs were screened, for the first time, as novel natural-derived drug candidates for the treatment of type 2 diabetes mellitus (T2DM). Noteworthy, AAs exhibited strong anti-glycation properties and higher hypoglycemic activity than the anti-diabetic drug acarbose, used as a positive control. In silico molecular modelling was consistent with the results from the in vitro bioassays. Moreover, in vitro kinetics of inhibition and fluorescence quenching experiments provided unique findings on AAs interaction with the targeted enzymes. MTT assays were performed to assess the biocompatibility of the optimized shell extracts on human intestinal (Caco-2 and HT29-MTX) and buccal (TR146 and HSC-3) cell lines. This represented the first study reporting the effects of compounds from dry fruit shells on the viability of intestinal and buccal cells. The metabolic fate of the optimized extracts from P. vera, P. dulcis, and C. avellana shells was estimated through in vitro simulated gastrointestinal digestion followed by untargeted LC-ESI-LTQ-Orbitrap-MS metabolomic analyses of the resulting oral, gastric and intestinal digests. The changes in the extracts’ phytochemical composition, antioxidant/antiradical activity (FRAP, DPPH•, and ABTS•+), and hypoglycemic properties (α-glucosidase and α-amylase inhibition) occurred after the gastrointestinal digestion simulation were widely explored providing novel insights into compounds’ bioaccessibility, hence contributing to expand the value of dry fruit shells into the field of food nutrition. Lastly, in vitro cell-based permeability models were adopted to assess the permeability of the gastrointestinal epithelium and buccal mucosa to the optimized shell extracts. Further green applications were investigated for walnut and pistachio shells. The optimized extract from walnut shells (OWS) was loaded into biodegradable polylactide (PLA)/poly (butylene adipate-co-terephthalate) (PBAT) polymeric blends. The resulting bio-film formulation showed improved performances in terms of UV-light resistance, antioxidant capacity, and mechanical properties. These results support the use of such PLA/PBAT-OWS blends as eco-friendly food packaging and bio-plastics. Another approach involved utilizing exhausted pistachio shells, leftover from the MAE treatment, as carbonaceous precursors for their transformation into activated carbons (ACs). Nowadays, ACs have multipurpose applications, such as electrode and battery production, water purification from heavy metals, and air pollution control. Hence, the proposed strategy introduces even exhausted pistachio shells as equally promising biomasses for the development of several clean-label materials with broad environmental and industrial relevance. All the findings shown in this thesis encourage the employment of dry fruit shells for the sustainable recovery of valuable bioactive compounds and antioxidants that, within the framework of a circular economy, can be included in nutraceutical formulations and other innovative materials.