Oil Structuring for Improving Healthy and Sustainable Diets: The Case Study of Extra Virgin Olive Oil Oleogelation

The aim of this PhD research project is to develop novel and sustainable strategies to structure liquid oils into pseudoplastic materials, called oleogels, to be used as saturated fat substitutes or as functional components able to modulate lipolysis during human digestion as well as deliver bioactive lipophilic molecules. In this context, the use of extra virgin olive oil (EVOO) as a target oil to be gelled would be particularly interesting due to its well- recognized health-promoting capacity. In this case study, EVOO-based oleogels were developed by using different gelation strategies. The oleogels, after structural characterization, were in vitro digested to study the impact of oil structure on the free fatty acids (FFA) release and polyphenols’ bioaccessibility. 1. Introduction In accordance with the PhD thesis project previously described (Ciuffarin, 2021), this poster reports the main results of the following activities: (A1) Study of strategies for oil structuring (e.g., methods and selection of gelators). (A3) Evaluation of the effect of oleogelation on the gastrointestinal behavior of oleogels by determining the bioaccessibility of selected bioactive molecules (e.g., polyphenols) as well as the lipolysis degree by using in vitro digestion methodologies. 2. Materials and Methods Oleogels were obtained by adding 10 % (w/w) of saturated monoglycerides (MG), rice bran waxes (RW), sunflower waxes (SW), and β-sitosterol/γ-oryzanol mixture (PS) in EVOO heated at temperatures higher than the melting temperatures of the different gelators. Additionally, a whey protein-based oleogel (WP) was prepared by mixing EVOO with a WP aerogel prepared following the methodology of Plazzotta et al. (2020). The final oil content in WP-based oleogels was 80% (w/w). Oleogels were characterized for their structure using a texture analyzer (35 mm-diameter cylindrical probe for 5 mm of distance at a crosshead speed of 1.5 mm/s, TA. XT Plus, Stable Micro Systems Ltd, Godalming, UK) and an accelerated release test by centrifuging samples (10000g for 15 min, Mikro 120, Hettich Zentrifugen, Andreas Hettich GmbH and Co, Tuttlingen, Germany). Unstructured EVOO and oleogels were then subjected to in vitro digestion according to the protocol proposed by Brodkorb et al. (2019). The FFAs released during digestion were assessed by titration (pH-stat). The bioaccessibility of tyrosol (T) and hydroxytyrosol (HT) was evaluated as the percentage ratio between the concentration of these components included in the micellar phase after intestinal in vitro digestion and their concentration in the undigested sample. The polyphenols were determined by HPLC. 3. Results and Discussion 3.1 Oleogel physical properties Table 1 shows the oil retention capacity and firmness of the considered oleogels. All the samples presented a very high oil retention capacity upon centrifugation (<99%) despite the different firmness. The MG-based oleogel was the weakest gel, followed by WP, RW, SW, and PS. These mechanical properties can be associated with the different natures of the networks structuring EVOO. In agreement with the literature, MG, RW, and SW formed a crystalline network (da Pieve et al., 2010; Doan et al., 2015), PS generated a fibrillar structure (Scharfe et al., 2019) and protein aerogels absorbed oil in the protein porous structure (Plazzotta et al., 2021). 3.2 In-vitro digestion: FFA release and bioaccessibility Figure 1 shows the FFA release as a function of the digestion time of structured into oleogels and unstructured EVOO. The typical curve of lipid hydrolysis was obtained. The unstructured oil presented FFA release % of about 68%, followed by PS, SW, RW, and MG with 59.1, 50.8, 50.7, and 42.8% respectively. A different behavior was acquired for WP- based oleogels showing the complete digestion of the oil. These results clearly show that the extent of lipid lipolysis was significantly affected by oil structure. In the case of liposoluble gelators (i.e., MG, RW, SW, PS), it can be inferred that the lipase activity was hindered by the presence of a structuring network behaving like a physical barrier to the access of the enzyme to the substrate sites. On the contrary, WP probably completely dissolved in the gastrointestinal environment thus favouring the emulsification of the oil and thus the lipase activity. In summary, the results demonstrated that the digestibility of the oil can be steered by selecting the proper oleogelator. In the next part of the study, the bioaccessibility of the major EVOO polyphenols (i.e., tyrosol and hydroxytyrosol) was assessed. Despite the higher content of hydroxytyrosol (HT) in EVOO than tyrosol (T) (HT: 248 mg/kg, T: 96 mg/kg), the bioaccessibility of T was significantly higher than that of HT. This result can be explained by considering the different susceptibility to oxidation of the two molecules during digestion (Alberdi-Cedeño et al., 2020). Moreover, differences were recorded among oleogels. Unstructured oil and WP presented the higher T bioaccessibility values, followed by SW, MG and RW, and PS. Since it is impossible to observe a direct effect of gel strength on polyphenol bioaccessibility, it can be speculated a possible interaction between the polyphenols and oleogel network structures. In fact, as well-known, polyphenols are surface-active molecules with the potentiality to interact with other food components. In conclusion, the results reported in the present study confirm that oleogelation could be a profitable strategy to modulate lipid digestion while delivering bioactive molecules. 4. References Alberdi-Cedeño, J., Ibargoitia, M. L., & Guillén, M. D. (2020). Study of the in vitro digestion of olive oil enriched or not with antioxidant phenolic compounds. Relationships between bioaccessibility of main components of different oils and their composition. Antioxidants, 9(6). Brodkorb, A., Egger, L., Alminger, M., Alvito, P., Assunção, R., Ballance, S., Bohn, T., Bourlieu-Lacanal, C., Boutrou, R., Carrière, F., Clemente, A., Corredig, M., Dupont, D., Dufour, C., Edwards, C., Golding, M., Karakaya, S., Kirkhus, B., le Feunteun, S., ... Recio, I. (2019). INFOGEST static in vitro simulation of gastrointestinal food digestion. Nat. Protoc., 14(4), 991–1014. da Pieve, S., Calligaris, S., Co, E., Nicoli, M. C., & Marangoni, A. G. (2010). Shear Nanostructuring of monoglyceride organogels. Food Biophys., 5(3), 211–217. Doan, C. D., van de Walle, D., Dewettinck, K., & Patel, A. R. (2015). Evaluating the oil-gelling properties of natural waxes in rice bran oil: Rheological, thermal, and microstructural study. JAOCS,92(6). Plazzotta, S., Calligaris, S., & Manzocco, L. (2020). Structural characterization of oleogels from whey protein aerogel particles. Int. Food Res. J., 132, 109099. Plazzotta, S., Jung, I., Schroeter, B., Subrahmanyam, R. P., Smirnova, I., Calligaris, S., Gurikov, P., & Manzocco, L. (2021). Conversion of whey protein aerogel particles into oleogels: Effect of oil type on structural features. Polym. J., 13(23). Scharfe, M., Ahmane, Y., Seilert, J., Keim, J., & Flöter, E. (2019). On the Effect of Minor Oil Components on β- Sitosterol/γ-oryzanol Oleogels. Eur. J. Lipid Sci. Technol., 121(8)

Application of different drying techniques to fresh-cut salad waste to obtain food ingredients rich in antioxidants and with high solvent loading capacity

Wastes from iceberg salad fresh-cut processing were submitted to air-drying, freeze-drying, and supercritical-CO2-drying with or without ethanol as co-solvent. Drying was combined with grinding to obtain flours. Samples were analysed for macro- and micro-appearance, particle size, dietary fibre, polyphenol content, antioxidant activity, water vapour sorption, water and oil holding capacity. Air-drying produced a collapsed brown material allowing a flour rich in fibre (>260 g/kg) and polyphenols (3.05 mg GAE/gdw) with antioxidant activity (6.04 OD 123/min/gdw) to be obtained. Freeze-drying maintained vegetable structure and colour while partly retaining polyphenols (1.23 mg GAE/gdw). Supercritical-CO2-drying with ethanol as co-solvent, produced an expanded material able to entrap huge amounts of water and oil (43.2 and 35.2 g of water and oil for g of dry sample). Air-dried salad waste derivatives could be used as functional food ingredients, while supercritical-CO2-dried ones can be exploited as bulking agents and absorbers of oil spills or edible oils. \ua9 2017 Elsevier Lt

Exploring the potentialities of photoinduced glycation to steer protein functionalities: The study case of freeze-dried egg white proteins/carbohydrates mixtures

The capacity of UV-C light to induce glycation and modify functional properties of systems containing freeze-dried egg white proteins and carbohydrates with increasing molecular weight (i.e., glucose, maltose, trehalose and maltodextrin) was studied. Color changes induced by light exposure were taken as typical indicators of glycation. Samples were then analyzed for selected physical (critical concentration, particle size and viscosity), chemical (ovalbumin content) and technofunctional properties (gelling temperature and foaming capacity). The presence of sugars during exposure to UV-C light promoted intense browning and decreased ovalbumin content by circa 30%. Concomitantly, up to a 3-fold increase in critical concentration of the aqueous suspensions of the irradiated protein-carbohydrate powders and changes in particle size were detected. These modifications were consistent with the development of non-enzymatic browning reactions upon UV-C light irradiation. Photoinduced glycation was associated to a decrease in viscosity, a tendency to form gel at temperature lower by up to 8\u25e6C and a better capacity of foam stabilization. The intensity of these changes seems to be affected by the nature of the carbohydrates reacting with proteins, with longer carbohydrates able to produce systems with higher foam stability capacity

Understanding the impact of moderate-intensity pulsed electric fields (MIPEF) on structural and functional characteristics of pea, rice and gluten concentrates

Aim: The effect of moderate-intensity pulsed electric fields (MIPEF) was evaluated on vegetable protein concentrates from pea, rice, and gluten. Methods: Five percent (w/w) suspensions of protein concentrates (pH 5 and 6) were exposed to up to 60,000 MIPEF pulses at 1.65\ua0kV/cm. Both structural modifications (absorbance at 280\ua0nm, free sulfhydryl groups, FT-IR-spectra) and functional properties (solubility, water and oil holding capacity, foamability) were analyzed. Results: MIPEF was able to modify protein structure by inducing unfolding, intramolecular rearrangement, and formation of aggregates. However, these effects were strongly dependent on protein nature and pH. In the case of rice and pea samples, structural changes were associated with negligible modifications in functional properties. By contrast, noticeable changes in these properties were observed for gluten samples, especially after exposure to 20,000 pulses. In particular, at pH 6, an increase in water and oil holding capacity of gluten was detected, while at pH 5, its solubility almost doubled. Conclusion: These results suggest the potential of MIPEF to steer structure of proteins and enhance their technological functionality

The importance of redox state in liver damage.

Oxidative stress is a major pathogenetic event occurring in several liver disorders ranging from metabolic to proliferative ones, and is a major cause of liver damage due to Ischemia/Reperfusion (I/R) during liver transplantation. The main sources of ROS are represented by mitochondria and cytocrome P450 enzymes in the hepatocyte, by Kupffer cells and by neutrophils. Cells are provided with efficient molecular strategies to strictly control the intracellular ROS level and to maintain the balance between oxidant and antioxidant molecules. A cellular oxidative stress condition is determined by an imbalance between the generation of ROS and the antioxidant defense capacity of the cell and can affect major cellular components including lipids, proteins and DNA. Proteins are very important signposts of cellular redox status and through their structure/function modulation, ROS can also influence gene expression profile by affecting intracellular signal transduction pathways. While several enzymatic (such as superoxide dismutase, catalase, glutathione peroxidase) and non enzymatic (such as 4-hydroxynonenal, decrease of glutathione, vitamin E, vitamin C, malondialdehyde) markers of chronic oxidative stress in liver are well known, early protein targets of oxidative injury are yet not well defined. Identification of these markers will enable early detection of liver diseases and will allow monitoring the degree of liver damage, the re1 Department of Biomedical Sciences and Technologies, University of Udine, P.le Kolbe 4, 33100 Udine, Italy. 2 Department of Biochemistry, Biophysics and Macromolecular Chemistry, University of Trieste, via Giorgieri 1, 34127 Trieste, Italy. 3 Centro Studi Fegato, AREA Science Park Bldg Q, Campus Basovizza, ss 14, km 163.5, 34012 Trieste, Italy. Address for correspondence

Effect of different oleogelators on lipolysis and curcuminoid bioaccessibility upon in vitro digestion of sunflower oil oleogels

Sunflower oil enriched with curcuminoid compounds (CUs) was gelled by adding 5% (w/w) saturated monoglycerides (MG), rice bran waxes (RW) or a mixture of \u3b2-sitosterol and \u3b3-oryzanol (PS). The resulting oleogels differed for rheological properties and firmness due to the difference in gel network structure. PS oleogel was the firmest sample followed by RW and MG ones. Upon in vitro digestion, fatty acid release as a function of digestion time was greatly affected by oleogel structure: the extent of lipolysis decreased as oleogel strength increased (PS < RW < MG). On the other hand, the nature of the oleogelator affected CUs bioaccessibility, which was lower in oleogels containing crystalline particles (MG and RW). These findings appear interesting in the attempt to develop oleogels able to control lipid digestion as well as to deliver bioactive molecules in food systems

Effect of different biopolymer-based structured systems on the survival of probiotic strains during storage and in vitro digestion

BACKGROUND: This study aimed to evaluate the protective effect of different biopolymer systems on the viability of two probiotics (Lactobacillus rhamnosus and Streptococcus thermophilus) during storage and in vitro digestion. Methylcellulose (MC), sodium alginate (SA), and whey protein (WP)-based structures were designed and characterized in terms of pH, rheological properties, and visual appearance. RESULTS: The results highlighted that the WP-system ensured probiotic protection during both storage and in vitro digestion. This result was attributed to a combined effect of the physical barrier offered by the protein gel network and whey proteins as a nutrient for microbes. On the other hand, surprisingly, the viscous methylcellulose-based system was able to guarantee good microbial viability during storage. However, this was not confirmed during in vitro digestion. The opposite results were obtained for sodium alginate beads. CONCLUSION: The results suggest that the capacity of a polymeric structure to protect probiotic bacteria is a combination of structural organization and system formulation. \ua9 2020 Society of Chemical Industry

Impact of high pressure homogenization on physical properties, extraction yield and biopolymer structure of soybean okara

The effect of high pressure homogenization (HPH) on soy okara was studied. To this purpose, okara dispersions (10 g/100 g) were subjected to 1 pass at 50, 100 and 150 MPa and to 5 passes at 150 MPa. Samples were analyzed for stability, particle size, microstructure, and viscosity. Results highlighted that the increase of HPH intensity was associated with the structural disruption of okara particles, leading to physically stable homogenates having increasing viscosity. This was mainly attributed to an increase in okara solubility, due to fibre and protein release. The latter resulted almost complete, reaching values up to 90% of the protein originally entrapped in okara matrix. Absorbance at 280 nm, SH groups and dimension of proteins revealed that HPH treatments favoured the extraction of the main protein fractions even if, at the higher intensity level, extracted proteins probably underwent conformational changes and reassembling phenomena

Modeling the effect of the oxidation status of the ingredient oil on stability and shelf life of low-moisture bakery products: The case study of crackers

In packed low-moisture foods such as crackers, oxidation is generally the main cause of quality depletion during storage. It is commonly believed, but scarcely investigated, that product shelf life depends on the oxidative status of the lipid ingredients. In this study, the influence of oxidation degree of the ingredient sunflower oil on cracker oxidative stability and hence shelf life was investigated. To this aim, oil with increasing peroxide values (PVs) (5, 11, and 25 mEqO2/kgoil) was used to prepare crackers. Just after production, crackers presented similar peroxide and rancid odor intensity, probably due to the interactive pathways of oxidative and Maillard reactions. Crackers were packed and analyzed for PV and rancid odor during storage at 20, 40, and 60 \u25e6C. Rancid odor well discriminated cracker oxidative status. Relevant oxidation rates were used to develop a shelf life predictive model based on the peroxide value of the ingredient oil. It was estimated that an oil PV from 5 to 15 mEqO2/kgoil shortens cracker Shelf Life (SL) by 50%, independently of storage temperature. These results demonstrate the critical impact of ingredient quality on product performance on the market

Modulation of Extra Virgin Olive Oil Digestibility through Oleogelation

Background. Extra virgin olive oil (EVOO) represents a key player in the Mediterranean diet for its health-promoting capacity. Although its use as a functional ingredient would be particularly interesting, the direct addition of EVOO to food is challenging due to its liquid state. EVOO conversion into a solid-like material through by oleogelation could enlarge its possible applications. Methods. EVOO was gelled by adding 10% (w/w) of saturated monoglycerides (MG), rice bran waxes (RW), sunflower waxes (SW) or a β-sitosterol/γ-oryzanol mixture (PS). Oleogels were characterised for their structure and subjected to static in vitro digestion. The fatty acid release and destructuring behavior were assessed. Results. The resulting oleogels differed for rheological properties and firmness due to the differences in gel network structure. PS oleogel was the firmest sample followed by SW, RW and MG ones. During in vitro digestion, the fatty acid release was significantly lower for all oleogels compared to unstructured oil. The different network provided by the four oleogelators not only influenced FA release, but also the intestinal micellar size. Conclusion. Acquired results could open new horizons for EVOO application through oleogelation to obtain novel EVOO-based fat replacers and better deliver the EVOO health functionalities