Beatrice Zonfrillo

Pomegranate peel, accounting for 35–50% of the fruit weight, is an underutilized agri-food by-product. This study applied, for the first time, fermentation with Saccharomyces cerevisiae as a simple and sustainable strategy to simultaneously obtain tannin-rich extracts and polysaccharide fractions with potential prebiotic activity. Peels from two cultivars, Wonderful and G1, differing in peel thickness, were subjected to three fermentation protocols (air- and not air-exposed) and monitored at 25 °C over 48 and 72 h. HPLC-DAD analysis showed that yeast-inoculated fermentation increased total tannin concentration in dry extracts (up to 70%) without inducing chemical modifications to tannin profiles. As determined by Dynamic Light Scattering, fermentation promoted significant depolymerization of native polysaccharides, while DOSY-¹H-NMR analyses revealed the presence of reduced molecular weight fractions down to 26 kDa. In vitro growth assays confirmed that fermented polysaccharides were more efficiently utilized as a carbon source by Bifidobacterium breve and Lactiplantibacillus plantarum compared to non-fermented controls, likely thanks to polysaccharide depolymerization induced by fermentation. The study demonstrated that air-exposed S. cerevisiae fermentation was an effective process alternative to chemical or enzymatic hydrolysis for modifying pomegranate peel pectin directly within a complex matrix, while simultaneously enhancing tannin recovery. This approach represents a possible sustainable strategy for pomegranate peel valorization into functional ingredients.

Pomegranate peel, accounting for 35-50% of the fruit's weight, is an underutilized by-product rich in ellagitannins and pectin. Although pomegranate juice has already been subjected to fermentation, this study aimed to demonstrate for the first time, that a simple fermentation process of pomegranate peel using Saccharomyces cerevisiae can yield tannin-rich extracts and enhance the potential prebiotic properties of pomegranate polysaccharides. Peels of ‘Wonderful’ and ‘G1’ cultivars, significantly differing in peel thickness, were used for fermentation applying three processes at room temperature monitored at 48 and 72 h. HPLC-DAD analysis showed that yeast-driven fermentation increased total tannin (up to 70% in dry extracts) compared with spontaneous fermentation (40-43%). A potential enhancement of prebiotic activity was associated with the reduction in the molecular weight of native polysaccharides (from a maximum of 705 kDa down to 26 kDa), as evaluated by DLS and DOSY-1H-NMR. Polysaccharides after fermentation were more effectively utilized by Bifidobacterium breve and Lactobacillus plantarum than those from control samples, as demonstrated by in vitro assays. These findings highlight a promising strategy for the valorization of pomegranate peel through the production of functional ingredients highly enriched in tannins and containing potentially prebiotic polysaccharides.

Background: Chemotherapy-induced neuropathic pain is a major side effect of antineoplastic treatment. This study investigated the neuroprotective potential of Acmella oleracea L. extracts containing the N-alkylamide spilanthol, phenolic acids, and glycosylated flavonoids. Methods: Hydroalcoholic extracts of aerial parts (AP) and roots (R) of in vitro seedlings were quali-quantitatively characterized by HPLC-DAD-MS and by ¹H-NMR. Different concentrations (15–150 µg/mL) of AP and R were tested in SH-SY5Y cells differentiated into neurons exposed to oxaliplatin (10 µM), assessing cell viability (MTT), cytotoxicity (LDH), SOD activity, and expression of ATF-3, Ire1α, and Nf-H genes. To evaluate the impact on neuropathic pain, CD-1 mice were treated intraperitoneally with oxaliplatin (2.4 mg/kg), the effect of AP and R extracts (200–1200 mg/kg) were measured by the cold plate test. Results: AP extract was rich in phenols and alkylamides, whereas R extract showed higher phenolic levels but lower alkylamides content. Both extracts significantly reduced mortality and cytotoxicity and counteracted oxidative imbalance by enhancing SOD activity. Gene expression analysis confirmed their neuroprotective effects. In vivo, oxaliplatin induced a 50% reduction in pain threshold, while acute treatment with AP and R extracts dose-dependently alleviated neuropathic pain. Despite the lower spilanthol content in R extract, its efficacy was comparable to AP, suggesting an important role of phenolic compounds. Conclusions: Extracts from both aerial parts and roots of A. oleracea show promise in alleviating chemotherapy-induced neuropathy through mechanisms not solely related to spilanthol. Further studies to elucidate the contribution of phenolic components are desirable.

The olive oil-production sector engages with the environment on multiple levels, and the valorization of olive pomace (OP) has emerged as a key strategy to improve the entire system’s sustainability. Numerous studies have investigated the biological effects of OP phenolic fraction for nutraceutical applications, highlighting its antioxidant properties. This study aimed to assess the effect of an OP extract (OPE) and its phenolic content on ferroptosis induced by RAS-selective lethal 3 (RSL3), an inhibitor of glutathione peroxidase 4. After characterization of OPE phenolic composition, its antioxidant properties were confirmed through the Fenton reaction assay. Subsequently, we examined the effect of OPE on ter-butyl hydroperoxide-induced ROS generation and lipid peroxidation in TPH-1 and HIECs cells and found that OPE reduced ROS and lipid peroxidation. RSL3 decreased the number of vital cells, which was associated with an elevation in ROS and lipid peroxidation, and a reduction in GSH. Interestingly, all these detrimental effects were reversed by OPE. Furthermore, OPE was also found to significantly increase GSH and the GSH/GSSG ratio per se. In conclusion, the fact that OPE decreases ROS and lipid peroxidation induced by RSL3 and augments GSH and cell viability suggests that OPE has potential as a ferroptosis inhibitor.

Fruits and vegetables processing produces significant amounts of by-products rich in valuable bioactive compounds such as polyphenols and dietary fiber. Food by-product re-use promotes the eco-sustainability of several crops. This study aimed to apply green extractions of bioactive compounds from by-products of basil, tomato, and red bell pepper production. Tests were performed by applying extraction procedures both at laboratory scale and using the Timatic^® extractor. Water and ethanol 10% and 20% were used for extraction of red bell pepper and tomato, testing different temperatures (30, 50, and 90 °C; water at 90 °C and ethanol 20% were applied for basil. The obtained phenolic extracts were analyzed by HPLC-DAD-MS. Polysaccharides of tomato and red bell pepper were extracted at laboratory scale and chemically characterized using ¹H-NMR to define the methylation and acylation degree, and DLS to estimate the hydrodynamic volume. Laboratory extraction tests allowed efficient scaling-up of the process on the Timatic^® extractor. Phenolic content in the dried extracts (DE) ranged 8.0–11.2 mg/g for tomato and red bell pepper and reached 240 mg/g for basil extracts. Polysaccharide yields (w/w on DM) reached 6.0 and 10.4% for dried tomato and red bell pepper, respectively. Dry extracts obtained using the Timatic^® extractor and water can be useful sources of bioactive phenols. The study provided new data on tomato and red bell pepper polysaccharides that may be useful for future applications.

Chlorogenic acids, the esters of caffeic and quinic acids, are the main phenolic acids detected in Acmella oleracea extracts and have gained increasing interest in recent years due to their important biological activities. Given their structural similarity and instability, the correct analysis and identification of these compounds in plants is challenging. This study aimed to propose a simple and rapid determination of the A. oleracea caffeoylquinic isomers, applying an HPLC-MS/MS method supported by a mathematical algorithm (Linear Equation of Deconvolution Analysis (LEDA)). The three mono- and the three di-caffeoylquinic acids in roots of Acmella plants were studied by an ion trap MS analyzer. A separation by a conventional chromatographic method was firstly performed and an MS/MS characterization by energetic dimension of collision-induced dissociation mechanism was carried out. The analyses were then replicated using a short HPLC column and a fast elution gradient (ten minutes). Each acquired MS/MS data were processed by LEDA algorithm which allowed to assign a relative abundance in the reference ion signal to each isomer present. Quantitative results showed no significant differences between the two chromatographic systems proposed, proving that the use of LEDA algorithm allowed the distinction of the six isomers in a quarter of the time.

The effects of the phenolic compounds of extra virgin olive oil (EVOO) on AGS cells have never been studied so far, which is the aim of this study. The profiles of the main phenolic components in EVOOs, mainly secoiridoid compounds derived from the transformation of oleuropein during the olive milling process, were evaluated and compared. Oils of different origins were evaluated aiming at verifying whether chemical differences in the phenolic composition of the dry extracts played a role in the metabolism and in maintaining the cellular redox state of AGS cells. The following key enzymes of some metabolic pathways were studied: lactate dehydrogenase, enolase, pyruvate kinase, glucose 6-phosphate dehydrogenase, citrate synthase, 3-Hydroxyacyl-CoA dehydrogenase and hexokinase. As confirmed through PCA analysis, pretreatments with the dry extracts of EVOOs at different concentrations appeared to be able to counteract the enzymatic activity alterations due to oxidative stress induced by H₂O₂ 1 mM and 2 mM. The studied phytocomplexes showed the ability to protect AGS cells from oxidative damage and the secoiridoid derivatives from both oleuropein and ligstroside contributed to the observed effects. The results suggested that EVOOs with medium to high concentrations of phenols can exert this protection.