This study investigated the cold gelation of native pea protein isolate (PPI) induced by transglutaminase (TG), focusing on how protein and enzyme concentrations interact to control gelation kinetics, crosslinking degree, and final gel properties. PPI dispersions (5-18% w/w) were treated with TG (0.2-1%, w/w) at 37 degrees C and characterized by tilting test, oscillatory rheology, microscopy, crosslinking degree, and response surface analysis. Gelation time decreased markedly with increasing protein concentration and, to a lesser extent, with increasing TG concentration. Microscopy revealed that concentrated systems formed more compact and aggregated networks, suggesting that non-covalent interactions increasingly governed gelation at high protein contents. A Doehlert design, to analyze the combined effects of PPI and TG concentrations, showed that the most elastic gels, characterized by the lowest tan delta, were obtained at intermediate PPI concentrations (10-14%) and relatively high TG levels (higher than 0.7%). In accordance, the degree of TG-mediated crosslinking showed a maximum at intermediate PPI levels (approximately 12%) and decreased at higher protein concentrations, despite faster gelation. In addition, maximum hardness was achieved at the highest protein concentration (18%) and intermediate TG concentration (approximately 0.6%). These results demonstrate that pea protein cold gelation is controlled by the competition between concentration-driven aggregation and TG-mediated covalent crosslinking. Optimal gel properties arise from a balance between sufficient protein concentration to promote intermolecular contacts and sufficient molecular mobility to allow efficient enzymatic crosslinking, enabling the design of cold-set gels with tailored firmness and elasticity.

Low dispersibility and colloidal instability limit the application of commercial pea protein concentrates in liquid food systems. This study investigates the effect of high-pressure homogenization (HPH) followed by pH modulation on the techno-functional properties (soluble fraction, particle size, colloidal stability, rheological behavior, and emulsifying properties) of a commercial pea protein concentrate. Initially, 3

This study investigates the potential of soy protein isolate (SPI) to enhance the bioaccessibility of oleic acid (OA), a beneficial fatty acid linked to reduced blood pressure and cardiovascular risk. Using an in vitro digestion model, we examined how the interactions between SPI-derived peptides and bile salts (BS) evolve during gastric and gastroduodenal digestion and how these interactions influence OA micellar solubility. SPI with a soluble fraction (SF) of 16% was digested for 10 and 60 min (tg10 and tg60), and the resulting digesta were evaluated for their ability to enhance OA solubilization in BS micelles. SF increased to 52.3% (tg10) and 64.7% (tg60) and reached 95.8% in the presence of BS under duodenal conditions. Consequently, OA solubilization improved by up to 82% compared to that of BS alone, highlighting the role of soy peptides in promoting lipid bioaccessibility through BS-peptide interactions during digestion.

The objective of the present work was to assess if the degree of crosslinking of pea proteins (PPI) by transglutaminase (TG) affects the in vitro gastric digestion of pea proteins in the bulk and at the oil-water interface as well as the duodenal lipolysis of the emulsions. PPI samples were prepared at protein concentrations of 10-18 % w/w and 1 % TG at 37 °C and characterized by small deformation tests and texture measurements. The degree of crosslinking linearly decreased with the increase of protein concentration and at the same time a loss of elasticity occurred. However, both the storage modulus G' and gel hardness increased with increasing protein concentration. These three crosslinked proteins were tested regarding their susceptibility to be hydrolysed by pepsin as well as the behaviour of their emulsions during in vitro gastroduodenal digestion. As the degree of crosslinking of the samples increases and they are more homogeneous (lower PPI concentration), they are more rapidly hydrolysed by pepsin in the bulk because the enzyme and the acid may easily diffuse into the gel microparticles. At the interface, lower degrees of PPI crosslinking would allow a higher structural flexibility, exposing peptide bonds that pepsin can more easily cleave. The rate of lipolysis increased as the degree of crosslinking decreased but at the end of duodenal digestion they converged to similar values. A correlation was found between the kinetics of gastric proteolysis of emulsions mediated by pepsin and the kinetics of duodenal lipolysis.

The feasibility of developing nanostructured lipid carriers (NLCs) based on coconut oil (CO) was analyzed by studying the crystallization behavior of bulk and emulsified CO in the presence of diacetyl tartaric acid ester of monoglycerides (DATEM) and glycerol monostearate (GM). Supercooling was almost halved compared to pure CO, and crystallization began at higher temperatures due to the seed effect of the emulsifiers. A significant difference in the crystallization enthalpy (ΔH) of CO was observed. Consequently, the solid fat content decreased to only 63-66 % when CO was emulsified. Both pure and emulsified CO crystallized in the β'-2 polymorph. The Avrami model showed reduced crystal dimensionality in CO-emulsifier blends and emulsions. NLCs derived from CO emulsions formulated with GM as emulsifier were most suitable due to the absence of flocculation, reduced CO crystallization, and melting above 37 °C.

Bile salts (BS) are biological surfactants that have important physiological functions mainly by participating in lipid digestion and intervening in the transport, absorption, and excretion of hydrophobic products. Bile salts have been shown to interact with the products of protein hydrolysis, being this interaction a key point to modulate several physiological processes.The type and size of peptides, released at different times of gastric digestion, could have different implications in these physiological processes modulated by the BS. In the present study we focused on understanding how the interactions of hydrolysates/peptides from native or commercial pea protein isolates with BS (formation of insoluble complexes or the modification of the BS micellar solubilization of poorly soluble molecules) evolve according to their residence time in a static gastric digestion model. The different residence times (10-60 min) would simulate different emptying points in which the partially hydrolyzed pea proteins by pepsin in the stomach are flushed into the duodenum and come into contact with the BS.It is observed that the longer residence times in the stomach the degree of hydrolysis of the pea isolates is greater, producing an increase in the soluble fraction. Both isolates showed a slight tendency to bind BS and form insoluble precipitates. The interaction of the soluble fraction of pea protein hydrolysates with the BS micelles, acted synergistically, increasing the solubilization capacity of oleic acid, taken as a model molecule.Likewise, it is shown that the structural changes of pea proteins due to processing have a profound influence on the interaction with BS and potentially affect the physiological reactions that they modulate.

The present work evaluated how a native pea protein isolate (PPI) affects the key roles carried out by bile salts (BS) in lipid digestion by means of the in vitro static INFOGEST protocol. Two gastric residence times were evaluated (10 and 60 min), and then the peptides obtained (GPPP) were mixed with BS at physiological concentration in simulated intestinal fluid to understand how they interact with BS both at the bulk and at the interface. Both GPPP give rise to a film with a predominant viscous character that does not constitute a barrier to the penetration of BS, but interact with BS in the bulk duodenal fluid. When the peptides flushing from the stomach after the different gastric residence times undergo duodenal digestion, it was found that for the longer gastric residence time the percentage of soluble fraction in the duodenal phase, that perform synergistically with BS micelles, was twice that of the lower residence time, leading to an increase in the solubilization of oleic acid. These results finally lead to a greater extent of lipolysis of olive oil emulsions. This work demonstrates the usefulness of in vitro models as a starting point to study the influence of gastric residence time of pea protein on its interaction with BS, affecting lipolysis. Pea proteins were shown to be effective emulsifiers that synergistically perform with BS improving the release and bioaccessibility of bioactive lipids as olive oil.

The study of lipid digestion has increased in recent years in order to elucidate how lipolysis can be controlled as this knowledge can aid to design healthier emulsified foods. Most of the works have attributed the decrease of the extent and rate of lipolysis of protein stabilized emulsions to droplet coalescence during the gastric phase causing a decrease of the interfacial area available for the reaction. Despite the crucial role of BS in lipids digestion, only few works have attributed a decrease of lipolysis to BS-emulsifiers interactions occurring both, at the interface, or in the bulk phase. The present work focuses in understanding the way in which a model milk protein as β-lactoglobulin (βlg), used as emulsifier, interacts with BS micelles under in vitro gastroduodenal conditions, modifying their capacity to solubilize the products of lipolysis and verify if this phenomenon is reflected in the kinetics of lipolysis of olive or chia oil in water emulsions.This work shows that the presence of βlg promotes the bioaccessibility of healthy oils such as olive oil or chia oil, which are sources of bioactive fatty acids. The mechanism involved is mediated by the interaction of the BS micelles with the peptides originated from the gastroduodenal proteolysis of the protein. As a result of this interaction, mixed micelles with a much higher capacity to solubilize the lipolysis products are formed. Therefore the lipolysis can proceed at the highest rate for a longer time.

BACKGROUND The application of chia mucilage still remains restricted due to the difficulty in achieving high extraction yields. The effect of ultrasonic-assisted extraction (UAE) conditions (temperature, seed:water ratio and time) on the rheological properties of chia mucilage extracts and the relation to the proportion of translucent phase (TP) and opaque phase (OP) of the mucilage in the extract were evaluated. RESULTS UAE allowed the efficient extraction of chia mucilage from chia seeds. The desired overall optimal combination to maximize both yield and apparent viscosity was achieved at a seed:water ratio 1:10, a temperature of 25.3 degrees C and 53.7 min extraction time; the optimal conditions to obtain the maximum yield and minimum apparent viscosity were a seed:water ratio close to 1:20, temperature of 48.8 degrees C and 208.4 min extraction time. CONCLUSION The results obtained in the present work demonstrated that the differences in rheological properties of chia mucilage extracts are due to the extraction methods used. Therefore, it is possible to modulate the extraction conditions in order to obtain different characteristics of the mucilage, maintaining a high extraction yield. (c) 2022 Society of Chemical Industry.

Protein-based carriers are important tools for the delivery of bioactive molecules. The objective of this work is to assess gelatin as folic acid carrier, a water-soluble vitamin with numerous fundamental biological functions. In particular, the aim of this research is to assess how this interaction affects some important properties of gelatin, to what extent it protects folic acid from degradation caused by UV light exposure and how bioaccessible the vitamin is after in vitro gastro-duodenal digestion. Firstly, it was determined that the interaction with gelatin (2.5%) at pH 3 was more efficient to avoid its degradation. In this system, it was verified that the vitamin was released from the protein carrier during the duodenal stage and would be bioaccessible in this site of absorption. Finally, the effect of folic acid on the gelling properties of this protein was analyzed, given the importance when used as food ingredient. No relevant differences were observed in the rheological and functional parameters due to the presence of the vitamin, which would allow a wide application of the developed carrier.

The caseinomacropeptide (CMP) is a bioactive peptide produced during cheese making. It is found in abundance in whey. CMP aqueous solutions allow the incorporation of large amounts of CaCl2 but the mechanism of calcium-CMP interactions are unknown. In order to evaluate its calcium binding capacity, the following techniques were performed: Dynamic Light Scattering (DLS), Fourier Transform Infrared spectroscopy (FTIR), dialysis, conductivity, precipitation of CaCl2/CMP complex by ethanol, electrochemical Ca2+ binding isotherms, and inhibition of calcium phosphate precipitation. One mole of CMP can bind 9 mol of calcium, and the CMP self-assembles as a hexameric form. A model is proposed to explain the CMP self-association in presence of CaCl2.

Consumer demands for healthier foods are constantly on the rise and results in the diversification of sources such as vegetable oils. Chia oil is a relevant source of alpha-linolenic acid. One of the targeted applications is the inclusion of chia oil in emulsified form in beverages where it is necessary to obtain stable submicron emulsions, acting as carrier of nutrients such as lipids. In the present work, high-pressure homogenization was employed to produce emulsions (10% chia oil) with small oil droplets (300 nm), formulated with different emulsifiers (beta-lacto-globulin (BLG), lecithin and Tween 80). These emulsions were submitted to a dynamic in vitro digestion (DIDGI) to better understand the main mechanisms involved in the process of lipolysis, in order to design systems that may allow the modulation of the digestive action of lipases, thus ensuring the maximum bioaccessibility of the lipids. This device mimics part of the transient biochemical conditions as well as the flow of foods through the different compartments occurring in the real digestion. Significant differences were observed in the lipolysis during the gastric phase. BLG emulsions reached a lipolysis degree of 30%. Lecithin-stabilized emulsions behave similarly to BLG ones during the first 90 min but lipolysis levelled off further similar to 20%. Finally, Tween-80-stabilized emulsions remained almost undigested (around 2%). No significant differences were noticed in duodenum and distal small intestine, irrespective of the emulsifier employed. After digestion, the fatty acid bioaccessibility was higher with BLG as emulsifier.

In the last few decades, there has been a growing interest in understanding the mechanisms involved in lipid digestion with the purpose of developing strategies to control this complex physiological process. Bile salts (BS) are natural bio-surfactants that play crucial functions in this process and may represent a key strategy to modulate the lipolysis. One of the main functions is the removal of lipolysis products present at the interface of lipid droplets by solubilizing them in BS micelles, thus avoiding the reaction inhibition. However, there are few studies that analyzed the effects that could have the emulsion components on the solubilization capacity of BS micelles. Thus, the main purpose of the present work was to evaluate the impact of a typical food emulsifier (Tween 80) on the fatty acid (FA) solubilization capacity of BS micelles by using a method recently developed, involving a combination of turbidity and dynamic light scattering (DLS) determinations. As FA solubilization into BS micelles may strongly affect the kinetics of lipolysis, the lipolysis of T80-stabilized oil-in-water (O/W) emulsions was also studied. The results showed that a higher concentration of the emulsifier in the duodenal medium causes the lipolysis rate to be maximum for a longer time. The mechanism involved could be the contribution of T80 to increase the solubilization of FA (and possibly other products of the lipolysis) in the duodenal medium, consequently affecting the potential bioavailability of fatty acids.

This review is focused at discussing the existing literature on proteins/peptides ? bile salts (BS) interactions regarding of the physiological processes they can modulate, as well as to analyze the in vitro assays mostly used to assess protein/peptides-bile salt interactions. When partially hydrolyzed proteins by pepsin in the stomach are flushed into the duodenum, they come into contact with the enzymes secreted by the pancreas (trypsin, chymotrypsin and lipase/colipase) and with the BS secreted by the gallbladder. It is at this stage where different types of interaction between the peptides present and the BS can take place, which can have potential health implications affecting (a) bile excretion, (b) the transport and absorption of hydrophobic molecules (cholesterol, fatty acids, vitamins and fat-soluble bioactive compounds) and (c) the degree and rate of lipolysis of dietary lipids. The molecular mechanisms underlying the modulation of processes (a) has to do with the ability of peptides to bind and precipitate BS, producing their elimination in the feces, whereas the modulation of process (b) and (c) are related to the role that BS have in the duodenal transport and absorption of lipids, fatty acids or any other hydrophobic molecule. In spite of a general evidence supporting that all food proteins or their hydrolysates/peptides have a certain degree of interaction with bile salts when testing their in vitro binding capacity or their influence on the micellar solubility of hydrophobic molecules, more studies are needed at a molecular level and using in vitro models that better simulate gastrointestinal digestion, to elucidate the mechanism underlying those effects.

• Competitive adsorption controls the degree of lipolysis. • The interfacial rheology was a valuable tool to analyze the competitive adsorption. • HPMC dominated the interface, controlling the emulsions size and lipolysis degree.

Complexing iron with organic compounds has been considered an alternative strategy to mitigate the problems associated with the level of bioavailable iron and the acceptability of products supplemented with this mineral. CMP contains specific amino acids associated with iron binding. The present study aims to optimize the conditions of Fe/CMP complex formation and understand the molecular basis of interactions between CMP and iron ions. Results showed that CMP can bind ferrous iron in a 1:1.5 M ratio, forming a stable peptide-iron complex, where CMP assembles in a tetrameric form. FTIR spectra indicated that iron binding altered the secondary structures of CMP. The iron-binding sites of CMP corresponded primarily to acid residues of Glu, Asp and sialic acid. Moreover, Fe/CMP complex remained stable in a wide pH range (2.0-6.5), suggesting the adequacy to be efficiently added in food or beverages and to keeping complexed in the digestion environment. Finally, Fe/CMP complex was added to a commercial beverage (2 mg of Fe per serving of beverage) and no changes were observed in their colour during storage. A model to explain the binding between CMP and iron is proposed. These results suggest a potential application of this peptide for iron fortification.

In food formulations, lipids are normally incorporated as emulsions stabilized by different types of emulsifiers. The emulsifiers can affect fatty acid (FA) solubilization as they can interact with FA. The main purpose of the present work is the development of a methodology to evaluate the FA solubilization in an aqueous medium in the absence and presence of exogenous emulsifiers. To this end, a combination of turbidimetry, oiling off and dynamic light scattering (DLS) was used. The FA solubility, as well as its supramolecular assemblies, were determined by analyzing the changes in the turbidity profile and the corresponding size of particles obtained by DLS. Oleic acid (OA) was used as a model FA and a simulated intestinal fluid (SIF) as the aqueous phase. Emulsifiers of low (Tween 80) and high (protein and polysaccharide) molecular weight were tested. Tween 80 was the only emulsifier that improved OA solubilization, whereas the macromolecules only affected the supramolecular structure that OA adopted, being the structure of these assemblies governed by the emulsifier nature.

The objective of this work is to assess bioaccessibility of folic acid in egg white (EW) nanocarriers after digestion of solutions and emulsions containing the vitamin-protein complexes. Furthermore, the influence of folic acid binding on the peptides profile after digestion is also evaluated. To this end an in vitro gastrointestinal digestion was performed and the pattern of digestion products was determined by high performance liquid chromatography. When folic acid was bound to nanocarriers, the vitamin peak area was approximately 80% lower than the one for folic acid alone. In emulsion, even less amount was detected. After digestion, no significant differences on folic acid peak areas were observed. Folic acid was released from nanocarriers after digestion, thus indicating that the vitamin would be bioaccessible in the site of absorption. Moreover, it was demonstrated by tricine-SDS-PAGE that folic acid binding induced a slight aggregation of proteins, although it did not alter the profile of peptides obtained after digestion. Binding of folic acid to nanocarriers proved to cause no negative effect on EW proteins digestibility. On the contrary, it seemed to slightly enhance their digestion. These nanocarriers would constitute an adequate system for the preservation, transport and target delivery of this bioactive.

The purpose of this research was to evaluate the influence of particle size distribution, pH (5.0-9.0), concentration (l-15g/100g.) and temperature (5-60 degrees C) on the steady shear flow properties of caseinomacropeptide (CMP) aqueous solutions. These measurements were carried out by using a controlled stress rheometer. Flow curves were satisfactorily fitted by the Herschel-Bulkley model. CMP solutions exhibited Newtonian flow dependence, particularly at pH values 5.0-6.0. Non-Newtonian shear thinning behaviour was observed at pH 7.0-9.0. The concentration dependence on viscosity showed two different regimes of viscosity increase (dilute and concentrated). The overlap concentration was 8 g/100 g. The temperature dependent behaviour of CMP solutions fitted to the Arrhenius model regardless pH and concentration, and the calculated activation energy was 20 kJ/mol. The flow behaviour of CMP is explained in terms of peptide-peptide and peptide-water interactions. Based on these results, CMP molecules would form spontaneously micelles at pH > 4.5 in ultrapure water.

During the last decade a special interest has been focused on studying the relationship between the composition and structure of emulsions and the extent of lipolysis, driven by the necessity of modulate lipid digestion to decrease or delay fats absorption or increase healthy fat nutrients bioavailability. Because bile salts (BS) play a crucial role in lipids metabolism, understanding how typical food emulsifiers affect the structures of BS under duodenal conditions, can aid to further understand how to control lipids digestion. In the present work the BS-binding capacity of three emulsifiers (Lecithin, Tween 80 and β-lactoglobulin) was studied under duodenal conditions. The combination of several techniques (DLS, TEM, ζ-potential and conductivity) allowed the characterization of molecular assemblies resulting from the interactions, as modulated by the relative amounts of BS and emulsifiers in solution.