Ultrasound-Assisted Maillard Conjugation of Yeast Protein Hydrolysate with Polysaccharides for Encapsulating the Anthocyanins from Aronia.

Valorisation of food by-products, like spent brewer's yeast and fruit pomaces, represents an important strategy for contributing to sustainable food production. The aims of this study were to obtain Maillard conjugates based on spent yeast protein hydrolysate (SYH) with dextran (D) or maltodextrin (MD) by means of ultrasound treatment and to use them for developing encapsulation systems for the anthocyanins from aronia pomace. The ultrasound-assisted Maillard conjugation promoted the increase of antioxidant activity by about 50% compared to conventional heating and SYH, and was not dependent on the polysaccharide type. The ability of the conjugates to act as wall material for encapsulating various biologically active compounds was tested via a freeze-drying method. The retention efficiency ranged between 58.25 ± 0.38%-65.25 ± 2.21%, while encapsulation efficiency varied from 67.09 ± 2.26% to 88.72 ± 0.33%, indicating the strong effect of the carrier material used for encapsulation. The addition of the hydrolysed yeast cell wall played a positive effect on the encapsulation efficiency of anthocyanins when used in combination with the SYH:MD conjugates. On the other hand, the stability of anthocyanins during storage, as well as their bioavailability during gastrointestinal digestion, were higher when using the SYH:D conjugate. The study showed that hydrolysis combined with the ultrasound-assisted Maillard reaction has a great potential for the valorisation of spent brewer's yeast as delivery material for the encapsulation of bioactive compounds.

Lactiplantibacillus plantarum MIUG BL21 paraprobiotics: Evidences on inactivation kinetics and their potential as cytocompatible and antitumor alternatives.

Two new -biotics concepts, such as paraprobiotics and postbiotics were introduced, with beneficial effects beyond the viability of probiotic. In this study, the effect of individual (thermal, ohmic heating, high pressure, and ultrasound) and combined (ohmic, high pressure and ultrasound in combination with heating) treatments on the inactivation kinetics of Lactiplantibacillus plantarum was investigated. Different inactivation rates were obtained, up to 8.18 after 10 min at 90 °C, 2.07 after 15 min at a voltage gradient of 20 V/cm, 6.62 after 10 min at 600 MPa and 3.6 after ultrasound treatment for 10 min at 100 % amplitude. The experimental data were fitted to Weibullian model proposed by Peleg, allowing to estimate the inactivation rate coefficient (b) and the shape of the inactivation curves (n). At lower concentration, the samples showed both cytocompatibility and antiproliferative effect, stimulating the cell proliferation on both murine fibroblast and human colorectal adenocarcinoma cell lines.

Advanced interactional characterization of the inhibitory effect of anthocyanin extract from Hibiscus sabdariffa L. on apple polyphenol oxidase.

In this study, a comprehensive approach to advance the inhibitory effect of Hibiscus sabdariffa extract on apple polyphenol oxidase (PPO) was performed. PPO was extracted, purified, and characterized for optimal activity, whereas response surface methodology generated a quadratic polynomial model to fit the experimental results for hibiscus extraction. The optimum conditions allowed to predict a maximum recovery of anthocyanins (256.11 mg delphinidin-3-O-glucoside/g), with a validated value of 272.87 mg delphinidin-3-O-glucoside/g dry weight (DW). The chromatographic methods highlighted the presence of gallic acid (36,812.90 µg/g DW extract), myricetin (141,933.84 µg/g DW extract), caffeic acid (101,394.07 µg/g DW extract), sinapic acid (1157.46 µg/g DW extract), kaempferol (2136.76 µg/g DW extract), and delphinidin 3-O-ß-d-glucoside (226,367.08 µg/g DW extract). The inactivation of PPO followed a first-order kinetic model. A temperature-mediated flexible fit between PPO and anthocyanins was suggested, whereas the molecular docking tests indicated that PPO is a good receptor for cafestol, gallic acid, and catechin, involving hydrophobic and hydrogen bond interactions. PRACTICAL APPLICATION: It is well known that enzymatic browning is one of the most important challenges in the industrial minimal processing of selected fruit and vegetable products. Novel inhibitors for polyphenol oxidase are proposed in this study by using an anthocyanin-enriched extract from Hibiscus sabdariffa L. Based on our results, combining the chemical effect of phytochemicals from hibiscus extract with different functional groups with minimal heating could be an interesting approach for the development of a new strategy to inhibit apple polyphenol oxidase.

Alternative Processing Options for Improving the Proteins Functionality by Maillard Conjugation.

Conjugation of the proteins with carbohydrates, occurring in the early stages of the Maillard reactions, received increased attention because of the high potential to ensure the improvement of the biological activity and functional properties of the proteins of different origins. The Maillard conjugates are conventionally formed through wet or dry heating, but the use of alternative technologies involving ultrasound, microwave, pulsed electric fields, high-pressure, or electrodynamic treatments appears to be efficient in accelerating the reaction steps and limiting the formation of toxic compounds. An overview of the mechanisms of these processing technologies, the main parameters influencing the Maillard conjugate formation, as well as their advantages and disadvantages, is provided in this paper. Different strategies employing these alternative technologies are reported in the literature: as pretreatment of the proteins, either alone or in admixture with the carbohydrates, followed by conventional heating, as a single alternative treatment step, or as a combination of heating and alternative processing. The desired functional properties of the proteins can be achieved by selecting the appropriate processing strategy and optimizing the reaction parameters. Moreover, alternative technologies can be exploited to obtain Maillard conjugates with remarkable biological activity in terms of antioxidant, antimicrobial, antihypertensive, anti-inflammatory, antimutagenic, or bifidogenic properties.

Proteases as Tools for Modulating the Antioxidant Activity and Functionality of the Spent Brewer's Yeast Proteins.

The functionality of the peptides obtained through enzymatic hydrolysis of spent brewer's yeast was investigated. Hydrolysis was carried out for 4-67 h with bromelain, neutrase and trypsin. The resulting hydrolysates were characterized in terms of physical-chemical, antioxidant and techno-functional properties. The solid residues and soluble protein contents increased with the hydrolysis time, the highest values being measured in samples hydrolyzed with neutrase. Regardless of the hydrolysis time, the maximum degree of hydrolysis was measured in the sample hydrolyzed with neutrase, while the lowest was in the sample hydrolyzed with trypsin. The protein hydrolysate obtained with neutrase exhibited the highest DPPH radical scavenging activity (116.9 ± 2.9 µM TE/g dw), followed by the sample hydrolyzed with trypsin (102.8 ± 2.7 µM TE/g dw). Upon ultrafiltration, the fraction of low molecular weight peptides (<3 kDa) released by bromelain presented the highest antioxidant activity (50.06 ± 0.39 µM TE/g dw). The enzymes influenced the foaming properties and the emulsions-forming ability of the hydrolysates. The trypsin ensured the obtaining of proteins hydrolysate with the highest foam overrun and stability. The emulsions based on hydrolysates obtained with neutrase exhibited the highest viscosity at a shear rate over 10 s-1. These results indicate that the investigated proteases are suitable for modulating the overall functionality of the yeast proteins.

Optimizing of the extraction conditions for anthocyanin's from purple corn flour (Zea mays L): Evidences on selected properties of optimized extract.

In this study, the Response Surface Methodology was used to optimize the extraction conditions for anthocyanins from purple corn flour (PCF), whereas the optimized extract was characterized in terms of phytochemicals content, inhibitory activity against metabolic associated enzymes and the potential to stimulate the metabolic activity of yeasts. The optimized conditions using a Box-Behnken design (BBD) of response surface methodology (RSM) were as follow: temperature (39 °C), extraction time (5 h), liquid/solid ratio (30 mL/g) and ethanol concentration (73 %), yielding a maximum response predicted and experimental values of 13.77 and 14.04 ± 0.02 mg cyanidin 3-O-glucoside equivalents (C3G)/g dry weight (DW). The extract showed a high content in myricetin, quercetin 3-ß-d-glucoside, kaempferol, whereas the main anthocyanin was C3G. The extract was test the effect on metabolic activity of Saccharomyces cerevisiae, with the highest multiplication rate obtained at 1.2 mg/100 mL. The extract showed potential antidiabetic, hypocholesterolemic and preventive effects against Parkinson's disease and melanoma. The findings provide the optimum conditions to obtain an anthocyanin enriched extract, for multiple applications.

Whey Proteins Isolate-Based Biopolymeric Combinations to Microencapsulate Supercritical Fluid Extracted Oleoresins from Sea Buckthorn Pomace.

In this study, high-value, carotenoid-rich oleoresin obtained by supercritical carbon dioxide (SFE-CO2) extraction was used to develop five variants of microencapsulated delivery system, based on whey proteins isolate (WPI), in combination with inulin (I), pectin (P) or lactose (L). The WPI:I and WPI:L variants were also obtained by conjugation via Maillard reaction. The microencapsulation of the SFE-CO2 sea buckthorn pomace oleoresin was performed by emulsion, complex coacervation and freeze-drying, which allowed for the obtaining of five powders, with different phytochemicals profile. The WPI:I conjugate showed the highest level of total carotenoids, whereas the counterpart WPI:L showed the highest content in linoleic acid (46 ± 1 mg/g) and palmitoleic acid (20.0 ± 0.5 mg/g). The ß-tocopherol and ß-sitosterol were identified in all variants, with the highest content in the conjugated WPI:L variant. Both WPI:L and WPI:I conjugate samples presented similar IC50 value for inhibitory activity against pancreatic lipase and α-amylase; the highest activity was observed for the conjugated WPI:I. The WPI:P combination allowed the highest release of carotenoids in the gastro-intestinal environment. All the powders exhibited poor flowing properties, whereas water activity (aw) ranged from 0.084 ± 0.03 to 0.241 ± 0.003, suggesting that all variants are stable during storage. In case of solubility, significant differences were noticed between non-heated and glycated samples, with the highest value for the WPI:I and the lowest for glycated WPI:I. The structural analysis revealed the presence of finer spherosomes in WPI:I and WPI:L, with a reduced clustering capacity, whereas the particles in the conjugated samples were more uniform and aggregated into a three-dimensional network.

CO2 Supercritical Fluid Extraction of Oleoresins from Sea Buckthorn Pomace: Evidence of Advanced Bioactive Profile and Selected Functionality.

The processing of sea buckthorn generates a significant amount of pomace, seeds and skin considered valuable sources of health-promoting macromolecules, such as carotenoids, pectin, flavonoids, phytosterols, polyunsaturated fatty acids and tocopherols. In this study, the bioactives from sea buckthorn pomace (SBP) were extracted using supercritical carbon dioxide (SFE-CO2), at different temperatures and pressures, allowing for obtaining four fractions according to separators (S40 and S45). The highest carotenoid content of 396.12 ± 1.02 mg/g D.W. was found in the S40 fraction, at extraction parameters of 35 °C/45 MPa, yielding an antioxidant activity of 32.10 ± 0.17 mMol TEAC/g D.W. The representative carotenoids in the extract were zeaxanthin, ß-carotene and lycopene, whereas all enriched SFE-CO2 extracts contained α-, ß- and δ-tocopherol, with α-tocopherol representing around 82% of all fractions. ß-sitosterol was the major phytosterol in the fractions derived from S45. All fractions contained significant fatty acids, with a predominance of linoleic acid. Remarkably, the enriched extracts showed a significant palmitoleic acid content, ranging from 53 to 65 µg/g. S40 extracts showed a good antibacterial activity against Staphylococcus aureus and Aeromonas hydrophila ATCC 7966, whereas S45 extracts showed a growth inhibition rate of 100% against Aspergillus niger after three days of growth. Our results are valuable, and they allow identifying the different profiles of extracts with many different applications in food, pharmaceutics, nutraceuticals and cosmeceuticals.

Encapsulation of Anthocyanins from Cornelian Cherry Fruits Using Heated or Non-Heated Soy Proteins.

In the current study, the effect of temperature on the potential of soy proteins to ensure the encapsulation and gastric stability of bioactives, such as anthocyanins from cornelian cherry fruits, was investigated. The powders obtained after freeze-drying were analyzed in relation to flow properties, encapsulation retention and efficiency, stability in simulated gastrointestinal medium, color, and morphology. Preheating the soy proteins generated a powder with low density. Powders obtained with native soy proteins allowed the highest encapsulation efficiency and the lowest was obtained when using preheated soy proteins. The heat treatment of the mixture of soy proteins and cornelian cherry fruits prior to encapsulation generated powders with the highest lightness and the lowest intensity of red shades among all samples. The in vitro experiments revealed that the highest protection in simulated gastric environment was provided when protein was heat treated either alone or in combination with bioactives to be encapsulated. The morphological analysis highlighted that powders consisted of large and rigid structures.

Cross-contamination of lettuce with Campylobacter spp. via cooking salt during handling raw poultry.

Campylobacter spp. are the most common bacterial pathogens associated with human gastroenteritis in industrialized countries. Contaminated chicken is the food vehicle associated with the majority of reported cases of campylobacteriosis, either by the consumption of undercooked meat or via cross- contamination of ready-to-eat (RTE) foods during the handling of contaminated raw chicken parts and carcasses. Our results indicate that cooking salt (used for seasoning) is a potential vehicle for Campylobacter spp. cross-contamination from raw chicken to lettuce, through unwashed hands after handling contaminated chicken. Cross-contamination events were observed even when the chicken skin was contaminated with low levels of Campylobacter spp. (ca. 1.48 Log CFU/g). The pathogen was recovered from seasoned lettuce samples when raw chicken was contaminated with levels ≥ 2.34 Log CFU/g. We also demonstrated that, once introduced into cooking salt, Campylobacter spp. are able to survive in a culturable state up to 4 hours. After six hours, although not detected following an enrichment period in culture medium, intact cells were observed by transmission electron microscopy. These findings reveal a "novel" indirect cross-contamination route of Campylobacter in domestic settings, and a putative contamination source to RTE foods that are seasoned with salt, that might occur if basic food hygiene practices are not adopted by consumers when preparing and cooking poultry dishes.

Microencapsulation of Red Grape Juice by Freeze Drying and Application in Jelly Formulation.

The aim of this work is to obtain new food products enriched with bioactive compounds from concentrated grape juice microencapsulated by freeze drying using a whey protein isolate-chitosan system. The obtained powder showed an encapsulation efficiency of (86.1±4.0) %, with an anthocyanin mass fraction (expressed as cyanidin-3-O-glucoside equivalent) of (1.4±0.2) mg/g, while the total polyphenolic (expressed as gallic acid equivalents) and flavonoid (expressed as catechin equivalents) mass fractions were (3.3±0.6) and (1.6±0.5) mg/g, respectively. The confocal laser microscopy revealed the presence of the flavonoid pigments wrapped inside the matrix, whereas the anthocyanins were grouped into large and compact clusters. The microencapsulated powder was used for jelly formulation. The new food formulations have a satisfactory anthocyanin mass fraction ranging from (0.03±0.01) to (0.12±0.02) mg/g, while no significant differences were observed in flavonoid content. All the value-added jelly showed appreciable antioxidant activity. The in vitro digestibility results confirm a slow release of anthocyanins from the food matrices during simulated gastric digestion and a significant release of the bioactive compounds into the gut. The addition of microencapsulated powder caused a significant decrease in firmness, cohesiveness and springiness, leading to the destabilization of the gel structure, while reducing the attraction forces between the matrix components. The sensorial analysis indicated that the panellists preferred the sample with grape juice the most.

Spectroscopic and Molecular Modeling Investigation on the Interaction between Folic Acid and Bovine Lactoferrin from Encapsulation Perspectives.

The impact of thermal treatment on the ability of lactoferrin (FL) to bind folic acid (FA) was investigated by employing fluorescence spectroscopy, molecular dynamics and docking tests. The structural and conformational particularities of LF upon heating at 80 °C and 100 °C were first estimated based on the intrinsic fluorescence changes in respect to the native protein. The emission spectra indicated gradual unfolding events accompanied by Trp exposure with increasing temperature. In agreement with the experimental results, molecular modeling investigations showed that the secondary and tertiary structure of LF are slightly affected by the thermal treatment. Some minor unfolding events related particularly to the α-helical regions of LF were observed when the temperature increased to 100 °C. The LF fluorescence quenching upon FA addition indicated that a static mechanism stands behind LF-FA complex formation. Regardless of the simulated temperature, the hydrogen bonds played an important role in regulating the interaction between the protein and ligand. FA binding to LF equilibrated at different temperatures occurred spontaneously, and all complexes displayed good thermodynamic stability. The obtained results support the suitability of LF as biocompatible material, for obtaining micro- and nanoparticles for delivery of dietary supplements or for enhancing the functionality of target delivery systems.

Valorizations of Sweet Cherries Skins Phytochemicals by Extraction, Microencapsulation and Development of Value-Added Food Products.

Sweet cherries are processed in various ways, leading to significant amounts of underutilized by-products that can potentially be used as a source of bioactive compounds, including antioxidants. The present study focuses on identifying ways to exploit bioactive compounds from sweet cherry skins, namely the extraction, microencapsulation, and functionalizing of some food product to obtain added value. The anthocyanins from skins were extracted and encapsulated in a combination of whey proteins isolate and chitosan by freeze-drying, with an encapsulation efficiency of 77.68 ± 2.57%. The powder showed a satisfactory content in polyphenols, of which anthocyanins content was 14.48 ± 1.17 mg cyanidin 3-glucoside/100 g dry weight (D.W.) and antioxidant activity of 85.37 ± 1.18 µM Trolox/100 g D.W. The powder was morphologically analyzed, revealing the presence of coacervates, ranging in size from 12-54 µm, forming large spheresomes (up to 200 µm). The powder was used as a functional ingredient to develop two value-added food products, namely yoghurt and marshmallows. The powder was tested for its prebiotic effect on L. casei 431® in the yoghurt samples during 21 days at 4 °C, when a decrease in viability was found, up to 6 log CFU·g-1. The anthocyanins and antioxidant activity decreased in yoghurt and increased in marshmallows during storage time. The obtained results support the potential use of extracts from underutilized sources in the development of functional ingredients and value-added food products.

Improvement of Quality Properties and Shelf Life Stability of New Formulated Muffins Based on Black Rice.

Effects of partial (50%) and total replacement of wheat flour with black rice flour on the phytochemical, physico-chemical, sensorial, and textural properties of muffins were studied. Partial or total replacement of wheat flour with black rice flour in muffins improved their nutritional and antioxidative properties with a positive effect on microbiological and color stability during the storage period in accelerated conditions. The low gluten muffins had an anthocyanin content of 27.54 ± 2.22 mg cyanidin-3-glucoside (C3G)/100 g dry weight (DW), whereas the gluten free muffins had 46.11 ± 3.91 mg C3G/100 g DW, with significant antioxidant values. Retention of 60% and 64% for anthocyanins and 72% and 80% for antioxidant activity after baking was found. The fracturability and hardness scores increased with the addition of black rice flour, whereas firmness and chewiness increased for gluten free muffins. The confocal analysis revealed a tendency of glucidic components to aggregate, with gathers of small bunches of black rice starch granules comprising anthocyanin. The results allowed designing two new value added bakery products, low and free gluten muffins, with significant high amounts of bioactive compounds, suggesting the functional potential of black rice flour.

New insights into xanthine oxidase behavior upon heating using spectroscopy and in silico approach.

Thermal dependent conformational changes of xanthine oxidase (XOD) were studied using sensitive and non-destructive methods like fluorescence spectroscopy and molecular modeling in the temperature range of 25-85°C. Intrinsic fluorescence studies showed that the microenvironment of tryptophan and tyrosine residues becomes more exposed to solvent as the temperature increased up to 85°C, whereas in case of flavin cofactor is rather conserved. At higher temperatures, the flavin adenine dinucleotide is displaced from the core of the protein, but is not fully released as shown by the Stern Volmer quenching constant and accessible fraction of the cofactor. Anyway, no significant changes in the structure of XOD monomer were identified after running molecular dynamics simulations at temperatures 25°C, 65°C and 85°C. Therefore, we can conclude that the most important changes in the protein structure at thermal treatment mainly consist on molecular aggregation and dissociation events.

Exploring the heat-induced structural changes of ß-lactoglobulin -linoleic acid complex by fluorescence spectroscopy and molecular modeling techniques.

Linoleic acid (LA) is the precursor of bioactive oxidized linoleic acid metabolites and arachidonic acid, therefore is essential for human growth and plays an important role in good health in general. Because of the low water solubility and sensitivity to oxidation, new ways of LA delivery without compromising the sensory attributes of the enriched products are to be identified. The major whey protein, ß-lactoglobulin (ß-Lg), is a natural carrier for hydrophobic molecules. The thermal induced changes of the ß-Lg-LA complex were investigated in the temperature range from 25 to 85 °C using fluorescence spectroscopy techniques in combination with molecular modeling study and the results were compared with those obtained for ß-Lg. Experimental results indicated that, regardless of LA binding, the polypeptide chain rearrangements at temperatures higher than 75 °C lead to higher exposure of hydrophobic residues causing the increase of fluorescence intensity. Phase diagram indicated an all or none transition between two conformations. The LA surface involved in the interaction with ß-Lg was about 497 Çº(2), indicating a good affinity between those two components even at high temperatures. Results obtained in this study provide important details about heat-induced changes in the conformation of ß-Lg-LA complex. The thermal treatment at high temperature does not affect the LA binding and carrier functions of ß-Lg.

Monitoring the heat-induced structural changes of alkaline phosphatase by molecular modeling, fluorescence spectroscopy and inactivation kinetics investigations.

The heat induced conformational changes of calf alkaline phosphatase (ALP) were analyzed using different methods, based on fluorescence spectroscopy, molecular modeling and inactivation studies. Experimental studies were conducted in buffer solution in the temperature range between 25 and 70 °C. Molecular dynamic (MD) simulation provided details on thermally induced changes in ALP structure, highlighting that heating favored the hydrophobic exposure and important alteration of the catalytic site above 60 °C. Additional information to MD data were obtained by using different fluorescence spectroscopy methods, which revealed a complex mechanism of thermal denaturation. Therefore, the emissive properties indicated an unfolding of ALP at temperatures below 60 °C, whereas at higher temperatures, the polypeptides chains fold leading to a higher exposure of Trp residues. In order to establish a structure-function relationship, the results were correlated with inactivation studies of ALP in buffer at pH 9.0. The inactivation data were fitted using a first-order kinetic model, resulting in an activation energy value of 207.26 ± 21.68 kJ · mol(-1).