Olive oil boasts numerous health benefits due to the high content of the monounsaturated fatty acid (MUFA) and functional bioactives including tocopherols, carotenoids, phospholipids, and polyphenolics with multiple biological activities. Polyphenolic components present antioxidant properties by scavenging free radicals and eliminating metabolic byproducts of metabolism. The objective of this research project was to recover the biologically active components rich in polyphenols, which include treatment of olive oil mills wastewater, and, at the same time, to remove the pollutant waste component resulting from the olive oil manufacturing processes. With specific focus on using technologies based on the application of ultra and nanofiltration membranes, the polyphenols fraction was extracted after an initial flocculation step. The nano-filtration permeate showed a reduction of about 95% of the organic load. The polyphenols recovery after two filtration steps was about 65% w/v. The nanofiltration retentate, dried using the spray dryer technique, was tested for cell viability after oxidative stress induction on human keratinocytes model in vitro and an improved cell reparation in the presence of this polyphenolic compound was demonstrated in scratch assays assisted through time lapse video-microscopy. The polyphenols recovered from these treatments may be suitable ingredients in cosmeceuticals and possibly nutraceutical preparations or functional foods.

Second cheese whey (SCW) derived from buffalo milk is the main by-product of the mozzarella cheese dairy industry. The objective of this study was to develop a membrane-based purification procedure to obtain specific fractions from SCW and assess their biological and applicative potential. Special interest was paid to the proteins and newly identified health-promoting compounds that could be recovered and used as value-added products in different sectors of food and pharmaceutical industries. SCW has been treated, sequentially, with microfiltration (MF), ultrafiltration (UF), and nanofiltration (NF) membranes giving the possibility to obtain three different fractions, namely retentates recovered on 100 and 10 kDa (R100 and R10) and a nanofiltration retentate (RNF). These retentates were compared for their ability to preserve human keratinocytes from dehydration, to form protein-based films by casting, and finally they were used for probiotic cultivations as the main substrate. Results showed that Lactobacillus rhamnosus could grow without any further additional nutrient up to 2.2 ± 0.3 × 10⁹ CFU/mL in the RNF medium. Dehydration tests on HaCat cells proved R100 as the most efficient fraction in preserving cell viability from this specific stress. R10, after diafiltrations, formed transparent films with improved features when glycerol was added as a plasticizer.

Pharma-grade extractive chondroitin sulfate (CS) is widely used for osteoarthritis (OA) treatment. Recently, unsulfated biofermentative chondroitin (BC) proved positive effects in OA in vitro model. This study, based on primary pathological human synoviocytes, aimed to analyze, by a multiplex assay, a panel of OA-related biomarkers in response to short-term treatments with bovine (CS_b), pig (CS_p) and fish (CS_f) chondroitins, in comparison to BC. As expected, all samples had anti-inflammatory properties, however CS_b, CS_f and especially BC affected more cytokines and chemokines. Based on these results and molecular weight similarity, CS_f and BC were selected to further explore the synoviocytes’ response. In fact, Western blot analyses showed CS_f and BC were comparable, downregulating OA-related biomarkers such as the proteins mTOR, NF-kB, PTX-3 and COMP-2. Proteomic analyses, performed by applying a nano-LC-MS/MS TMT isobaric labelling-based approach, displayed the modulation of both common and distinct molecules to chondroitin treatments. Thus, CS_f and BC modulated the biological mediators involved in the inflammation cascade, matrix degradation/remodeling, glycosaminoglycans’ synthesis and cellular homeostasis. This study helps in shedding light on different molecular mechanisms related to OA disease that may be potentially affected not only by animal-source chondroitin sulfate but also by unsulfated biofermentative chondroitin.

Currently, chondroitin sulfate (CS) and hyaluronic acid (HA) pharma-grade forms are used for osteoarthritis (OA) management, CS as an oral formulations component, and HA as intra-articular injective medical devices. Recently, unsulfated chondroitin, obtained through biofermentative (BC) manufacturing, has been proposed for thermally stabilized injective preparation with HA. This study aimed to highlight the specific properties of two commercial injective medical devices, one based on HA/BC complexes and the other containing HA, extractive CS, and cyclodextrins, in order to provide valuable information for joint disease treatments. Their biophysical and biomechanical features were assayed; in addition, biological tests were performed on human pathological chondrocytes. Rheological measurements displayed similar behavior, with a slightly higher G′ for HA/BC, which also proved superior stability to the hyaluronidase attack. Both samples reduced the expression of specific OA-related biomarkers such as NF-kB, interleukin 6 (IL-6), and metalloprotease-13 (MMP-13). Moreover, HA/BC better ensured chondrocyte phenotype maintenance by up-regulating collagen type 2A1 (COLII) and aggrecan (AGN). Notwithstanding, the similarity of biomolecule components, the manufacturing process, raw materials characteristics, and specific concentration resulted in affecting the biomechanical and, more interestingly, the biochemical properties, suggesting potential better performances of HA/BC in joint disease treatment.

Lactobacillus paracasei IMC502^® is a commercially successful probiotic strain. However, there are no reports that investigate growth medium composition in relation to improved biomass production for this strain. The major outcome of the present study is the design and optimization of a growth medium based on vegan components to be used in the cultivation of Lactobacillus paracasei IMC502^®, by using Design of Experiments. Besides comparing different carbon sources, the use of plant-based peptones as nitrogen sources was considered. In particular, the use of guar peptone as the main nitrogen source, in the optimization of fermentation media for the production of probiotics, could replace other plant peptones (e.g. potato, rice, wheat, and soy) which are part of the human diet, thereby avoiding an increase in product and process prices. A model with R² and adjusted R² values higher than 95% was obtained. Model accuracy was equal to 94.11%. The vegan-optimized culture medium described in this study increased biomass production by about 65% compared to growth on De Man–Rogosa–Sharpe (MRS) medium. Moreover, this approach showed that most of the salts and trace elements generally present in MRS are not affecting biomass production, thus a simplified medium preparation can be proposed with higher probiotic biomass yield and titer. The possibility to obtain viable lactic acid bacteria at high density from vegetable derived nutrients will be of great interest to specific consumer communities, opening the way to follow this approach with other probiotics of impact for human health.

There is growing interest in newly isolated lactic acid bacteria from traditional sources. In this study, a Lactococcus lactis strain identified and isolated from natural whey starter cultures of cow milk for the production of artisanal cheeses was cultivated in optimized vegan grade medium to assess its growing ability and metabolic fingerprint. In fact, traditionally fermented dairy products are considered nutritionally complete and possibly functional to human health with a number of benefits not only to the gastro-intestinal tract but also in a systemic manner. In fed batch experiments, we achieved 14 g/L dry cell weight and 1.9∙1010 viable colony forming unit increasing the values of experiment threefold and twofold respect to the batch processes, respectively. Additionally, the lactic acid (LA) production was quantified, and a maximal concentration of 78 g/L was achieved, which is approximately five-fold corresponding to the batch experiment results. This is in agreement with kinetic modeling of LA inhibition studies that highlighted a halved growth rate (µ) at 35 ± 5 g/L of LA whilst the growth blockage occurred at about 80 g/L. The samples obtaining after ultrafiltration processes and tested on three different pathogens, Enterococcus faecalis, Salmonella enterica subsp. enterica serovar Typhimurium and enteroinvasive Escherichia coli, gave a reduction on viability by 6–9 log on average. This paper focused its attention on optimization of fermentation conditions (in particular a vegan grade media using Design of Experiment approach) using fed-batch and microfiltration processes increasing the production of biomass and bioactive molecules, with respect to the batch processes. At the end of the fed-batch fermentation, a downstream process based on membranes was performed in order to obtain bioactive molecules that proved antimicrobial activity against intestinal/food spoilage pathogens.