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Soybean, a protein-rich leguminous oilseed, is often unacceptable due to its taste, appearance, and smell. Enzymatic technologies offer an alternative to conventional chemical procedures for protein modification, allowing for accurate oversight and adjustment of reaction flow to desired nutritional and bioactive properties due to the high sensitivity of protease under mild reaction conditions. Herein, the feasibility of implementing a one- or two-step biotechnological process, triggered by commercial endo- and exo-peptidases, to enhance the nutritional and bioactive characteristics of soy protein concentrates (SPC), was investigated. Two nutritionally valuable fractions, the hydrolysate (liquid stream-fraction) and the ocara (solid stream-fraction), have been separated. The hydrolysates were characterized by examining the crude protein content, protein recovery, and free amino group content. The antioxidant activity was quantified by measuring superoxide radical inhibition, and metal-ion chelation. Commercial peptidases led to different reaction kinetics and protein recovery, resulting in different peptide profiles determined by dead-end ultrafiltration (3, 10, and 30 kDa) and subsequent gel-filtration chromatography using the Toyopearl HW40F resin. Enzymatic hydrolysis seemed to enhance the hydrolyzate's protein content while decreasing the ocara's protein content. The percentage of soluble protein recovered from SPC ranged between 68 and 82%, revealing that Flavourzyme was most suitable to solubilize SPC. The highest yield of hydrolyzed peptide bonds, correlated with higher antioxidant activity, was shown by Alcalase- Flavourzyme (~20%) as well as Neutrase-Flavourzyme (~21.5%), with a tendency to favor Neutrase due to more sensory-acceptable product. Each produced hydrolysate's amino acid content, sulfhydryl groups, and surface hydrophobicity have been examined, and substantial correlations with antioxidant activity have been found. Because the quantity of phytic

Soybean, a protein-rich leguminous oilseed, is often unacceptable due to its taste, appearance, and smell. Enzymatic technologies offer an alternative to conventional chemical procedures for protein modification, allowing for accurate oversight and adjustment of reaction flow to desired nutritional and bioactive properties due to the high sensitivity of protease under mild reaction conditions. Herein, the feasibility of implementing a one- or two-step biotechnological process, triggered by commercial endo- and exo-peptidases, to enhance the nutritional and bioactive characteristics of soy protein concentrates (SPC), was investigated. Two nutritionally valuable fractions, the hydrolysate (liquid stream-fraction) and the ocara (solid stream-fraction), have been separated. The hydrolysates were characterized by examining the crude protein content, protein recovery, and free amino group content. The antioxidant activity was quantified by measuring superoxide radical inhibition, and metal-ion chelation. Commercial peptidases led to different reaction kinetics and protein recovery, resulting in different peptide profiles determined by dead-end ultrafiltration (3, 10, and 30 kDa) and subsequent gel-filtration chromatography using the Toyopearl HW40F resin. Enzymatic hydrolysis seemed to enhance the hydrolyzate's protein content while decreasing the ocara's protein content. The percentage of soluble protein recovered from SPC ranged between 68 and 82%, revealing that Flavourzyme was most suitable to solubilize SPC. The highest yield of hydrolyzed peptide bonds, correlated with higher antioxidant activity, was shown by Alcalase- Flavourzyme (~20%) as well as Neutrase-Flavourzyme (~21.5%), with a tendency to favor Neutrase due to more sensory-acceptable product. Each produced hydrolysate's amino acid content, sulfhydryl groups, and surface hydrophobicity have been examined, and substantial correlations with antioxidant activity have been found. Because the quantity of phytic

Soy protein concentrate (SPC) was hydrolyzed using several commercial food-grade proteases (Alcalase, Neutrase, papain, Everlase, Umamizyme, Flavourzyme) and their combination to obtain promising ingredients in the manufacture of functional bakery products. In all cases, the hydrolysis caused nutritional, sensory, and rheological changes in SPC, as well as protein structural changes like increased surface hydrophobicity and content of exposed SH groups with the magnitude of these changes depending on enzyme specificity. The hydrolysis with the combination of Neutrase and Flavourzyme (NeuFlav) increased essential amino acid content by 9.8% and that of Lys by 32.6% compared to SPC. This hydrolysate showed also significant antioxidant activities including ABTS and superoxide anion scavenging activity and metal-chelating ability. The addition of all hydrolysates in wheat flour decreased water adsorption and increased development time to some extent due to gluten network weakening, but also decreased the rate of starch retrogradation, contributing to the increase of the shelf-life of bakery products. The NeuFlav tasted less bitter than other hydrolysates, while E-nose provided a discrimination index of 93 between control and hydrolysates. It appeared that the addition of the NeuFlav hydrolysate in a cookie formulation improved protein content and nutritional quality and directed to its higher general consumer acceptability than cookies formulated with only wheat flour.

Soy protein concentrate (SPC) was hydrolyzed using several commercial food-grade proteases (Alcalase, Neutrase, papain, Everlase, Umamizyme, Flavourzyme) and their combination to obtain promising ingredients in the manufacture of functional bakery products. In all cases, the hydrolysis caused nutritional, sensory, and rheological changes in SPC, as well as protein structural changes like increased surface hydrophobicity and content of exposed SH groups with the magnitude of these changes depending on enzyme specificity. The hydrolysis with the combination of Neutrase and Flavourzyme (NeuFlav) increased essential amino acid content by 9.8% and that of Lys by 32.6% compared to SPC. This hydrolysate showed also significant antioxidant activities including ABTS and superoxide anion scavenging activity and metal-chelating ability. The addition of all hydrolysates in wheat flour decreased water adsorption and increased development time to some extent due to gluten network weakening, but also decreased the rate of starch retrogradation, contributing to the increase of the shelf-life of bakery products. The NeuFlav tasted less bitter than other hydrolysates, while E-nose provided a discrimination index of 93 between control and hydrolysates. It appeared that the addition of the NeuFlav hydrolysate in a cookie formulation improved protein content and nutritional quality and directed to its higher general consumer acceptability than cookies formulated with only wheat flour.

Constant world population growth imposes the need for the production of more food, and as a consequence, has an increase in the organic-rich wastes. These agro-industrial wastes represent problem from the environmental viewpoint, and their utilization as the raw materials for the production of value-added products has become one of the main topics of the biorefinery. The aim of this research was to use soybean meal, co-product of dietary oil production, as a model agro-industrial by-product for optimization of phenolic compounds extraction. In this paper, ethanol influenced microwave-assisted extraction is chosen since it is recognized as eco-friendly and food-safe. Optimization of the extraction included finding the optimal liquid/solid ratio, extraction duration, and microwave power. The same method was applied for the extraction of phenolics from apple pomace, apple pomace mixed with soybean meal, ultrasound-treated soybean meal, and wheat bran. Antioxidative potential of all extracts was evaluated using a fixed reaction time 2,2-diphenyl-1-picrylhydrazyl (DPPH) method as well as a steady-state measurement DPPH method. 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) i.e., ABTS and ferric reducing antioxidant power (FRAP) methods were applied as well. Under optimized conditions (liquid/solid ratio of 15 mL/g for 70 s at a microwave power of 300 W) 3.87 mg gallic acid equivalent (GAE)/g was extracted from soybean meal, which represented a substantial increase compared to 2.46 mg GAE/g obtained before optimization. Results showed that extracts had different behavior towards different radicals depending on the type of the extracted phenolics. Apple pomace extract was the most potent towards DPPH• radical, while the most effective extract towards ABTS•+ radical was obtained from apple pomace and soybean meal mixture. The soybean meal extract showed the highest reducing power assessed via the FRAP method. Results obtained in this study show that optimization of

Constant world population growth imposes the need for the production of more food, and as a consequence, has an increase in the organic-rich wastes. These agro-industrial wastes represent problem from the environmental viewpoint, and their utilization as the raw materials for the production of value-added products has become one of the main topics of the biorefinery. The aim of this research was to use soybean meal, co-product of dietary oil production, as a model agro-industrial byproduct for optimization of phenolic compounds extraction. In this paper, ethanol influenced microwave-assisted extraction is chosen since it is recognized as eco-friendly and food-safe. In this paper, we used soybean meal, co-product of dietary oil production, as a model agro-industrial by-product for optimization of phenolic compounds extraction. Ethanol influenced microwave-assisted extraction is chosen since it is recognized as eco-friendly and food-safe. Optimization of the extraction included finding the: optimal liquid/solid ratio, extraction duration, and microwave power. The same method was applied for the extraction of phenolics from apple pomace, apple pomace mixed with soybean meal, ultrasound-treated soybean meal, and wheat bran. Antioxidative potential of all extracts was evaluated using a fixed reaction time 2,2-diphenyl-1- picrylhydrazyl (DPPH) method as well as a steady-state measurement DPPH method. 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) i.e., ABTS and ferric reducing antioxidant power (FRAP) methods were applied as well. Under optimized conditions (liquid/solid ratio of 15 mL/g for 70 s at a microwave power of 300 W) 3.87 mg gallic acid equivalent (GAE)/g was extracted from soybean meal, which represented a substantial increase compared to 2.46 mg GAE/g obtained before optimization. Results showed that extracts had different behavior towards different radicals depending on the type of the extracted phenolics. Apple pomace extract was the most potent towa

In recent years, there has been an unprecedented de-mand by both consumers and industry, for inexpensive plant-derived protein hydrolysates as potential dietary sources of antioxidants. This is a particular refers to the protein from agricultural crop, such as soybean flour and/or flakes, especially from the soy protein concentrate (SPC). However, insolubility of SPC in water limits its more extensive use as food-protein ingredient, but it can be overcome by applying various enzymatic and non-enzymatic treatments. In order to expand the application of SPC, the influence of the one-step enzyme modification by using the commercial food-grade en-doprotease from Bacillus licheniformis on the antiox-idant and functional properties has been considered. For this purpose, SPC under the various dry matter contents (DM) were pre-incubated at the optimal temperature and hydrolyzed in a mechanically stirred batch reactor. The bioactivity was evaluated using rad-ical-scavenging and metal-chelating assays. The ABTS (2,2’-azino-bis(3-ethylbenzothiazoline-6-sul-phonic acid)) radical-scavenging ability reached higher values at DM 8% and smaller hydrolysis degree (DH) 12.85%, then a decreasing for higher DH and the minimum reducing power was observed after hydrolysis of SPC with DM 2% and DH~18% (64.2%). It appeared that the all prepared hydrolysates had the high yields of bioactive peptides which had the ability to strong-ly chelating prooxidant ferrous metal ions ( gt 95%). The techno-functionalities of SPC hydrolysate (DM 8%) with relatively higher antioxidant activities were remarkably higher compared to the untreated SPC. Namely, this hydrolysate had excellent solubility over a pH range of 2-12 which was improved compared to untreated SPC-8%. The oil holding capacity was also enhanced, while enzyme treatment was damaging to both of emulsifying properties, activity and stability, and water holding capacity. The results obviously show that enzyme treated SPCs were superior to the

The objective of this research was to investigate the impact of high-intensity ultrasound (HIU) generated by a probe-type sonicator (frequency 20 +/- 0.2 kHz and an amplitude of 40%) for 2-20 min on the selected functional and structural properties of egg white proteins (EWPs) and their susceptibility to hydrolysis by alcalase. The protein solubility, foaming, and emulsifying properties were studied as a function of ultrasonication time and related to protein particle and structural properties. The length of ultrasonication exhibited important effect on EWP particle size, uniformity, and charge, affecting also the protein conformation and susceptibility to alcalase hydrolysis and determining functional properties. There was a linear correlation between the particle size decrease and the solubility while a two-step linear correlation between the foam capacity (FC)/foam stability (FS) and particle size was apparent. Specifically, FC and FS sharply increased with decreasing particle size for range from similar to 370 to similar to 260 nm, and below this range from 260.6 to 68.4 nm, the changes were not that substantial. Besides, the solubility, FC, and FS were directly and linearly related with the absolute value of the particle zeta potential. The overall emulsifying properties were also improved with an increase of sonication time, through both the decrease of the mean particle diameter and the increase of zeta potential, but there was no direct correlation between the emulsion activity/stability index and protein particle size and/or charge. Analysis of EWP structure by Raman spectroscopy revealed that the HIU leads to changes in the secondary structure, while heat and ultrasound generated by the ultrasound bath were not sufficient to exhibit this effect.

The objective of this research was to investigate the impact of high-intensity ultrasound (HIU) generated by a probe-type sonicator (frequency 20 +/- 0.2 kHz and an amplitude of 40%) for 2-20 min on the selected functional and structural properties of egg white proteins (EWPs) and their susceptibility to hydrolysis by alcalase. The protein solubility, foaming, and emulsifying properties were studied as a function of ultrasonication time and related to protein particle and structural properties. The length of ultrasonication exhibited important effect on EWP particle size, uniformity, and charge, affecting also the protein conformation and susceptibility to alcalase hydrolysis and determining functional properties. There was a linear correlation between the particle size decrease and the solubility while a two-step linear correlation between the foam capacity (FC)/foam stability (FS) and particle size was apparent. Specifically, FC and FS sharply increased with decreasing particle size for range from similar to 370 to similar to 260 nm, and below this range from 260.6 to 68.4 nm, the changes were not that substantial. Besides, the solubility, FC, and FS were directly and linearly related with the absolute value of the particle zeta potential. The overall emulsifying properties were also improved with an increase of sonication time, through both the decrease of the mean particle diameter and the increase of zeta potential, but there was no direct correlation between the emulsion activity/stability index and protein particle size and/or charge. Analysis of EWP structure by Raman spectroscopy revealed that the HIU leads to changes in the secondary structure, while heat and ultrasound generated by the ultrasound bath were not sufficient to exhibit this effect.

In recent years, there has been an unprecedented de-mand by both consumers and industry, for inexpensive plant-derived protein hydrolysates as potential dietary sources of antioxidants. This is a particular refers to the protein from agricultural crop, such as soybean flour and/or flakes, especially from the soy protein concentrate (SPC). However, insolubility of SPC in water limits its more extensive use as food-protein ingredient, but it can be overcome by applying various enzymatic and non-enzymatic treatments. In order to expand the application of SPC, the influence of the one-step enzyme modification by using the commercial food-grade en-doprotease from Bacillus licheniformis on the antiox-idant and functional properties has been considered. For this purpose, SPC under the various dry matter contents (DM) were pre-incubated at the optimal temperature and hydrolyzed in a mechanically stirred batch reactor. The bioactivity was evaluated using rad-ical-scavenging and metal-chelating assays. The ABTS (2,2’-azino-bis(3-ethylbenzothiazoline-6-sul-phonic acid)) radical-scavenging ability reached higher values at DM 8% and smaller hydrolysis degree (DH) 12.85%, then a decreasing for higher DH and the minimum reducing power was observed after hydrolysis of SPC with DM 2% and DH~18% (64.2%). It appeared that the all prepared hydrolysates had the high yields of bioactive peptides which had the ability to strong-ly chelating prooxidant ferrous metal ions ( gt 95%). The techno-functionalities of SPC hydrolysate (DM 8%) with relatively higher antioxidant activities were remarkably higher compared to the untreated SPC. Namely, this hydrolysate had excellent solubility over a pH range of 2-12 which was improved compared to untreated SPC-8%. The oil holding capacity was also enhanced, while enzyme treatment was damaging to both of emulsifying properties, activity and stability, and water holding capacity. The results obviously show that enzyme treated SPCs were superior to the

In recent years, there has been an unprecedented demand by both consumers and industry, for inexpensive plant-derived protein hydrolysates as potential dietary sources of antioxidants. This is a particular refers to the protein from agricultural crop, such as soybean flour and/or flakes, especially from the soy protein concentrate (SPC). However, insolubility of SPC in water limits its more extensive use as food-protein ingredient, but it can be overcome by applying various enzymatic and non-enzymatic treatments. In order to expand the application of SPC, the influence of the one-step enzyme modification by using the commercial food-grade endoprotease from Bacillus licheniformis on the antioxidant and functional properties has been considered. For this purpose, SPC under the various dry matter contents (DM) were pre-incubated at the optimal temperature and hydrolyzed in a mechanically stirred batch reactor. The bioactivity was evaluated using radical-scavenging and metal-chelating assays. The ABTS (2,2’-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)) radicalscavenging ability reached higher values at DM 8% and smaller hydrolysis degree (DH) 12.85%, then a decreasing for higher DH and the minimum reducing power was observed after hydrolysis of SPC with DM 2% and DH~18% (64.2%). It appeared that the all prepared hydrolysates had the high yields of bioactive peptides which had the ability to strongly chelating prooxidant ferrous metal ions (> 95%). The techno-functionalities of SPC hydrolysate (DM 8%) with relatively higher antioxidant activities were remarkably higher compared to the untreated SPC. Namely, this hydrolysate had excellent solubility over a pH range of 2-12 which was improved compared to untreated SPC-8%. The oil holding capacity was also enhanced, while enzyme treatment was damaging to both of emulsifying properties, activity and stability, and water holding capacity. The results obviously show that enzyme treated SPCs were superior to the untreated

In recent years, there has been an unprecedented demand by both consumers and industry, for inexpensive plant-derived protein hydrolysates as potential dietary sources of antioxidants. This is a particular refers to the protein from agricultural crop, such as soybean flour and/or flakes, especially from the soy protein concentrate (SPC). However, insolubility of SPC in water limits its more extensive use as food-protein ingredient, but it can be overcome by applying various enzymatic and non-enzymatic treatments. In order to expand the application of SPC, the influence of the one-step enzyme modification by using the commercial food-grade endoprotease from Bacillus licheniformis on the antioxidant and functional properties has been considered. For this purpose, SPC under the various dry matter contents (DM) were pre-incubated at the optimal temperature and hydrolyzed in a mechanically stirred batch reactor. The bioactivity was evaluated using radical-scavenging and metal-chelating assays. The ABTS (2,2’-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)) radicalscavenging ability reached higher values at DM 8% and smaller hydrolysis degree (DH) 12.85%, then a decreasing for higher DH and the minimum reducing power was observed after hydrolysis of SPC with DM 2% and DH~18% (64.2%). It appeared that the all prepared hydrolysates had the high yields of bioactive peptides which had the ability to strongly chelating prooxidant ferrous metal ions (> 95%). The techno-functionalities of SPC hydrolysate (DM 8%) with relatively higher antioxidant activities were remarkably higher compared to the untreated SPC. Namely, this hydrolysate had excellent solubility over a pH range of 2-12 which was improved compared to untreated SPC-8%. The oil holding capacity was also enhanced, while enzyme treatment was damaging to both of emulsifying properties, activity and stability, and water holding capacity. The results obviously show that enzyme treated SPCs were superior to the untreated

Wheat gluten represents the major protein fraction present in wheat flour and as protein-rich material posses cohesive and viscoelastic properties which permit the retention of gas bubbles in the dough. It could be obtained as a by-product during the separation of starch from wheat flour and as such may be utilized like a functional protein additive in various non-bakery foodstuffs due to its desirable structure-enhancing properties. However, wheat gluten is hardly soluble in water, which limits their expanding utilization. In the regard, the aim of this research was to investigate the correlation between process parameters of wheat gluten hydrolysis and emulsification activity and stability of the prepared hydrolysates. The hydrolysates showing the greatest enhancement of the emulsifying properties which are closed correlated with solubility improvement was further separated by sequential ultrafiltration to obtain molecular weight distribution profile and peptide fraction with higher metal-ion chelating ability. The hydrolysis was performed by using commercial endopeptidase from Bacillus licheniformis and process was followed by monitoring the degree of hydrolysis, emulsifying properties and metal-ion chelating activity. The effects of some relevant process parameters such as gluten concentration (1-9% w/v), temperature (40-60 °C), pH (7-9), enzyme/substrate ratio (0.25-0.75 AU g-¹ of protein) and their interactions were investigated by the means of the four-factor Box-Behnken experimental design with 29 experimental points (5 central points). The obtained results showed that the second-order models developed for emulsification activity and stability of gluten hydrolysates were significant (p<0.05) with a high value of the coefficients of determination (0.965-0.995). The statistical analysis showed that each variable had a significant effect on the emulsifying properties of the tested system. In terms of emulsifying properties results showed that gluten concentrati

Native wheat gluten is a deluxe bread improver and may be utilized as a functional protein additive in multifarious non-bakery foodstuffs due to its desirable structure-enhancing properties. Also its utilization would be economically interesting, but lack of some desirable functional properties limited their expanding utilization in foodstuff formulations. This study was designed to examine the relationship between process parameters and functional properties of the obtained hydrolysates using Box-Benken experimental design and response surface methodology (RSM). The hydro-lysate showing the highest improvement of solubility and foaming ability was further separated by sequential ultrafiltration to obtain molecular weight distribution profile. The progress of wheat gluten hydrolysis was followed by monitoring the degree of hydrolysis (DH) using the pH-stat method and functional properties were measured by our methods already adopted. The effects of process parameters (pH, T, [S], [E]/[S] ration) and their interactions were investigated by the means of the four-factor Box-Behnken experimental design with 29 experimental points (5 central points). Experiments were carried out in triplicates and expressed as means with SD. The effects of different parameters under the significance level of p < 0.05 were examined using one-way analysis of variance (ANOVA) and Student t-test. The coefficients of the response function and their statistical significance were evaluated by the response surface regression analysis, using the Design software. Non-significant terms (p ≥ 0.05) were deleted from the second order polynomial and a new polynomial has been recalculated. The Fisher test (F-value) was used to determine whether the second-order model was adequate to describe the obtained data while the goodness of fit of the model was evaluated by the determination coefficient (R2). The obtained results showed that the second-order models developed for DH, solubility and foaming prop

The aim of the study was to investigate and compare several extraction protocols like 1) high-power ultrasound probe assisted solvent extraction; 2) microwave assisted solvent extraction; 3) direct acid hydrolysis; 4) direct alkali hydrolysis, and 5) two step extraction consisting of ultrasound or microwave assisted solvent extraction followed by alkaline and acid hydrolysis in terms of efficiency of the extraction of phenolic acids from the yellow soybean seed variety Laura. These extracts were screened for their total phenol content (TPC), and for their antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl radical scavenging (DPPH) assay, as well as on content of some individual phenolic acids. It appeared that the acetone containing mixtures gave much higher TPC than methanol containing ones, but the presence of acid in the methanol solvent significantly improved the extraction of phenolic compounds. To further improve the extraction, an ultrasound lab-scale probe at 20 kHz was used, with 15 and 30% of the maximum amplitude, and the extraction time was varied from 2 to 15 min. Microwave assisted extraction was performed varying the temperature from 55 to 85 degrees C, microwave power from 25 to 100 W and extraction time from 2 to 10 min. Changes in the content of six phenolic acids were examined: gallic, trans cinnamic, chlorogenic, caffeic, p-coumaric and ferulic acid. The separation and quantification of phenolic acids was accomplished by high-performance liquid chromatography-diode array detection (DAD) procedure. The results suggested that microwaves contributed to more efficient extraction of phenolic acids from the seed of yellow soybean. The amount of phenolic acids varied from 65.52 mu g/g of dry matter (d.m.) for caffeic acid, to 581.84 mu g/g d.m. for p-coumaric acid. Both, ultrasound and microwaves contributed to more efficient extraction of total phenol compounds and enhanced antioxidant activity of soybean seed extracts. TPC varied from 12.48 to 1

This study focuses on the influence of operating conditions on Alcalase-catalyzed egg white protein hydrolysis performed in a continuously stirred tank reactor coupled with ultrafiltration module (10 kDa). The permeate flow rate did not significantly affect the degree of hydrolysis (DH), but a significant increase in process productivity was apparent above flow rate of 1.9 cm(3) min(-1). By contrast, an increase in enzyme/substrate (E/S) ratio provided an increase in DH, but a negative correlation was observed between E/S ratio and productivity. The relationship between operating conditions and antioxidant properties of the hydrolysates, measured by three methods, was studied using Box-Behnken experimental design and response surface methodology. The statistical analysis showed that each variable (impeller speed, E/S ratio, and permeate flow rate) had a significant effect on the antioxidant capacity of all tested systems. Nevertheless, obtained response functions revealed that antioxidative activity measured by DPPH, ABTS and FRAP methods were affected differently by the same operating conditions. High impeller speeds and low permeate flow rates favor ABTS while high impeller speeds and high permeate flow rates had a positive effect on the DPPH scavenging activity. On the other hand, the best results obtained with FRAP method were achieved under moderate operating conditions. The integration of the reaction and ultrafiltration membrane separation in a continuous manner appears to be a right approach to improve and intensify the enzymatic process, enabling the production of peptides with desired antioxidant activity.

The objective of this study was to discover the relationship between the ultrasound probe treatment (UPT) on egg white proteins (EWPs) before EWPs hydrolysis by different proteases, and the functional properties of the obtained hydrolysates. To fulfill this goal, the protein solubility, foaming, and emulsifying properties were studied as a function of the UPT time and then related to the surface characteristics and structural properties. The changes in the hydrolysates microstructures and macromolecular conformation, induced by the UPT, were followed using scanning electron microscope analyzis (SEM) and Fourier transforms infrared spectroscopy (FTIR). The results showed that UPT influenced (P lt 0.05) the proteolysis of egg white proteins for all examined treatment times. Alcalase hydrolysates (AHs) and papain hydrolysates (PHs) were found to have a higher solubility, as a consequence of their relatively higher foaming, and emulsifying properties compared to the untreated hydrolysates. The changes in surface hydrophobicity, sulfhydryl content and surface charge of AHs and PHs indicated unfolding of EWPs affected by ultrasound. SEM analyzis showed that UPT destroyed the microstructures of AHs and PHs, while FTIR spectra indicated remarkable changes in the macromolecular conformation of AHs and PHs after UPT. This study revealed that by combining ultrasound pre-hydrolysis treatment under controlled conditions with thoughtful proteases selection, hydrolysates with improved functional properties could be produced, enhancing utilization of EWPs in food products.

The aim of the study was to investigate and compare several extraction protocols like 1) high-power ultrasound probe assisted solvent extraction; 2) microwave assisted solvent extraction; 3) direct acid hydrolysis; 4) direct alkali hydrolysis, and 5) two step extraction consisting of ultrasound or microwave assisted solvent extraction followed by alkaline and acid hydrolysis in terms of efficiency of the extraction of phenolic acids from the yellow soybean seed variety Laura. These extracts were screened for their total phenol content (TPC), and for their antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl radical scavenging (DPPH) assay, as well as on content of some individual phenolic acids. It appeared that the acetone containing mixtures gave much higher TPC than methanol containing ones, but the presence of acid in the methanol solvent significantly improved the extraction of phenolic compounds. To further improve the extraction, an ultrasound lab-scale probe at 20 kHz was used, with 15 and 30% of the maximum amplitude, and the extraction time was varied from 2 to 15 min. Microwave assisted extraction was performed varying the temperature from 55 to 85 degrees C, microwave power from 25 to 100 W and extraction time from 2 to 10 min. Changes in the content of six phenolic acids were examined: gallic, trans cinnamic, chlorogenic, caffeic, p-coumaric and ferulic acid. The separation and quantification of phenolic acids was accomplished by high-performance liquid chromatography-diode array detection (DAD) procedure. The results suggested that microwaves contributed to more efficient extraction of phenolic acids from the seed of yellow soybean. The amount of phenolic acids varied from 65.52 mu g/g of dry matter (d.m.) for caffeic acid, to 581.84 mu g/g d.m. for p-coumaric acid. Both, ultrasound and microwaves contributed to more efficient extraction of total phenol compounds and enhanced antioxidant activity of soybean seed extracts. TPC varied from 12.48 to 1

Introduction: Native wheat gluten is a deluxe bread improver and may be utilized as a functional protein additive in multifarious nonbakery foodstuffs due to its desirable structure-enhancing properties. Also its utilization would be economically interesting, but lack of some desirable functional properties limited their expanding utilization in foodstuff formulations [1]. This study was designed to examine the relationship between process parameters and functional properties of the obtained hydrolysates using Box-Benken experimental design and response surface methodology (RSM). The hydrolysate showing the highest improvement of solubility and foaming ability was further separated by sequential ultrafiltration to obtain molecular weight distribution profile. Materials and methods: The progress of wheat gluten hydrolysis was followed by monitoring the degree of hydrolysis (DH) using the pH-stat method and functional properties were measured by our methods already adopted [2]. The effects of process parameters (pH, T, [S], [E]/[S] ration) and their interactions were investigated by the means of the four-factor Box-Behnken experimental design with 29 experimental points (5 central points). Results: The obtained results showed that the second-order models developed for DH, solubility and foaming properties of gluten hydrolysates were significant (p<0.05) with a high value of coefficients of determination (0.944-0.981). The statistical analysis showed that each variable had a significant effect on degree of hydrolysis and the functional properties of tested system. Almost the linear increase in DH was observed with the rise in temperature at the highest substrate concentration, while on the other hand increasing the concentration of the substrate leads to a decrease in DH. In terms of foaming properties results showed that foam capacity range are in the range of 24.2- 80.3%, depending on the independent variables that were tested. Conclusions: Results are relevant to th

The use of protein hydrolysates in bakery products is gaining importance due to the presence of bioactive peptides in their composition. Soy protein hydrolysates (SPHs) have gained interest due to the lack of antinutritional factors and presence of various bioactivities (antioxidant activity, cholesterol-lowering activity or immunoregulatory properties). The aim of the study was to evaluate the dough rheology parameters of wheat flour replaced by 5% withdifferent SPHs. SPHs were obtained by enzymatic hydrolysis of soy protein concentrate with different proteases: Alcalase® 2.4L, Everlase 16.0L, Neutrase® 0.8L, Papain and Umamizyme. The rheological behavior of flour mixtures was evaluated using a Mixolab analyzer (Chopin Technologies) with the “Chopin+” protocol. The SPHaddition in the wheat flour resulted in decreases in water absorption with almost 3.9% as compared with control dough. Dough development time increased with SPH addition, with the highest value registered for umamizyme hydrolysate (5.9 min towards 1.4 min for control). Thus, SPHs prevented gluten network development.Stability and C2 value (which represents the beginning of the protein destabilization and unfolding) decreased with the SPH addition in the dough formulation, which proves the weakening of the gluten network. In the same trend, starch gelatinization (C3), stability of the starch gel (C4) as well as starch retrogradation (C5) were reduced by the addition of SPHs. These values were decreased in the order: control > alcalase ≈ everlase ≈ umamizyme > neutrase > papain. In conclusion, dough characteristics provided by Mixolab are the results of the interaction of SPHs components with the flour constituents.Papain hydrolysate could contribute to the increase of the shelf-life of bakery products.

Introduction: Soy proteins are widely used in several food products in the forms of soy flour, soy protein concentrate (SPC) and soy protein isolate (SPI) to improve water and fat binding ability and improve nutritional content. Recent research efforts in this field continue to look at novel functional properties of soy proteins through physical, chemical or enzymatic modifications [1]. Although it has been demonstrated that the enzymatic hydrolysis of SPI is a suitable route to improve its antioxidant properties, there is a general lack of knowledge about the potential for beneficial effects of protease hydrolysis on soy products other than SPI, such as soy flour or SPC. The aim of this study was to investigate the antioxidant activities, and color of spray-dried hydrolysates of SPC obtained with several proteases. Materials and methods: A commercial SPC (protein content 71.24%) was a gift from the SOJAPROTEIN, Bečej. The proteases used were: alcalase® 2.4L, neutrase® 0.8L, papain, everlase 16.0 L (Sigma) and umamizyme (Amano Enzyme Inc). The antioxidant activity was tested by using several methods: DPPH, ABTS, and linoleic acid emulsion system [2]. Results: Process parameters like substrate concentration, E/S ratio, temperature, and DH have been optimized for each selected enzyme or their combination. The five proteolytic enzymes used for hydrolysis of SPC led to different reaction kinetics resulting also in different peptide profiles determined by gel filtration chromatography using Sephadex G-50. Conclusions: The alcalase hydrolysate seemed to be the most effective antioxidant determined by DPPH, ABTS, and linoleic acid emulsion system assays. The enzymatic hydrolysis in all cases resulted in an increase in the antioxidant activity of hydrolysate and all hydrolysates showed significantly changed color compared to SPC.

The aim of the study was to investigate and compare several extraction protocols like 1) high-power ultrasound probe assisted solvent extraction; 2) microwave assisted solvent extraction; 3) direct acid hydrolysis; 4) direct alkali hydrolysis, and 5) two step extraction consisting of ultrasound or microwave assisted solvent extraction followed by alkaline and acid hydrolysis in terms of efficiency of the extraction of phenolic acids from the yellow soybean seed variety Laura. These extracts were screened for their total phenol content (TPC), and for their antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl radical scavenging (DPPH) assay, as well as on content of some individual phenolic acids. It appeared that the acetone containing mixtures gave much higher TPC than methanol containing ones, but the presence of acid in the methanol solvent significantly improved the extraction of phenolic compounds. To further improve the extraction, an ultrasound lab-scale probe at 20 kHz was used, with 15 and 30% of the maximum amplitude, and the extraction time was varied from 2 to 15 min. Microwave assisted extraction was performed varying the temperature from 55 to 85 degrees C, microwave power from 25 to 100 W and extraction time from 2 to 10 min. Changes in the content of six phenolic acids were examined: gallic, trans cinnamic, chlorogenic, caffeic, p-coumaric and ferulic acid. The separation and quantification of phenolic acids was accomplished by high-performance liquid chromatography-diode array detection (DAD) procedure. The results suggested that microwaves contributed to more efficient extraction of phenolic acids from the seed of yellow soybean. The amount of phenolic acids varied from 65.52 mu g/g of dry matter (d.m.) for caffeic acid, to 581.84 mu g/g d.m. for p-coumaric acid. Both, ultrasound and microwaves contributed to more efficient extraction of total phenol compounds and enhanced antioxidant activity of soybean seed extracts. TPC varied from 12.48 to 1

Native wheat gluten is a deluxe bread improver and may be utilized as a functional protein additive in multifarious non-bakery foodstuffs due to its desirable structure-enhancing properties. Also its utilization would be economically interesting, but lack of some desirable functional properties limited their expanding utilization in foodstuff formulations. This study was designed to examine the relationship between process parameters and functional properties of the obtained hydrolysates using Box-Benken experimental design and response surface methodology (RSM). The hydro-lysate showing the highest improvement of solubility and foaming ability was further separated by sequential ultrafiltration to obtain molecular weight distribution profile. The progress of wheat gluten hydrolysis was followed by monitoring the degree of hydrolysis (DH) using the pH-stat method and functional properties were measured by our methods already adopted. The effects of process parameters (pH, T, [S], [E]/[S] ration) and their interactions were investigated by the means of the four-factor Box-Behnken experimental design with 29 experimental points (5 central points). Experiments were carried out in triplicates and expressed as means with SD. The effects of different parameters under the significance level of p < 0.05 were examined using one-way analysis of variance (ANOVA) and Student t-test. The coefficients of the response function and their statistical significance were evaluated by the response surface regression analysis, using the Design software. Non-significant terms (p ≥ 0.05) were deleted from the second order polynomial and a new polynomial has been recalculated. The Fisher test (F-value) was used to determine whether the second-order model was adequate to describe the obtained data while the goodness of fit of the model was evaluated by the determination coefficient (R2). The obtained results showed that the second-order models developed for DH, solubility and foaming prop

This study focuses on the influence of operating conditions on Alcalase-catalyzed egg white protein hydrolysis performed in a continuously stirred tank reactor coupled with ultrafiltration module (10 kDa). The permeate flow rate did not significantly affect the degree of hydrolysis (DH), but a significant increase in process productivity was apparent above flow rate of 1.9 cm(3) min(-1). By contrast, an increase in enzyme/substrate (E/S) ratio provided an increase in DH, but a negative correlation was observed between E/S ratio and productivity. The relationship between operating conditions and antioxidant properties of the hydrolysates, measured by three methods, was studied using Box-Behnken experimental design and response surface methodology. The statistical analysis showed that each variable (impeller speed, E/S ratio, and permeate flow rate) had a significant effect on the antioxidant capacity of all tested systems. Nevertheless, obtained response functions revealed that antioxidative activity measured by DPPH, ABTS and FRAP methods were affected differently by the same operating conditions. High impeller speeds and low permeate flow rates favor ABTS while high impeller speeds and high permeate flow rates had a positive effect on the DPPH scavenging activity. On the other hand, the best results obtained with FRAP method were achieved under moderate operating conditions. The integration of the reaction and ultrafiltration membrane separation in a continuous manner appears to be a right approach to improve and intensify the enzymatic process, enabling the production of peptides with desired antioxidant activity.

The objective of this study was to discover the relationship between the ultrasound probe treatment (UPT) on egg white proteins (EWPs) before EWPs hydrolysis by different proteases, and the functional properties of the obtained hydrolysates. To fulfill this goal, the protein solubility, foaming, and emulsifying properties were studied as a function of the UPT time and then related to the surface characteristics and structural properties. The changes in the hydrolysates microstructures and macromolecular conformation, induced by the UPT, were followed using scanning electron microscope analyzis (SEM) and Fourier transforms infrared spectroscopy (FTIR). The results showed that UPT influenced (P lt 0.05) the proteolysis of egg white proteins for all examined treatment times. Alcalase hydrolysates (AHs) and papain hydrolysates (PHs) were found to have a higher solubility, as a consequence of their relatively higher foaming, and emulsifying properties compared to the untreated hydrolysates. The changes in surface hydrophobicity, sulfhydryl content and surface charge of AHs and PHs indicated unfolding of EWPs affected by ultrasound. SEM analyzis showed that UPT destroyed the microstructures of AHs and PHs, while FTIR spectra indicated remarkable changes in the macromolecular conformation of AHs and PHs after UPT. This study revealed that by combining ultrasound pre-hydrolysis treatment under controlled conditions with thoughtful proteases selection, hydrolysates with improved functional properties could be produced, enhancing utilization of EWPs in food products.

The objective of this study was to discover the relationship between the ultrasound probe treatment (UPT) on egg white proteins (EWPs) before EWPs hydrolysis by different proteases, and the functional properties of the obtained hydrolysates. To fulfill this goal, the protein solubility, foaming, and emulsifying properties were studied as a function of the UPT time and then related to the surface characteristics and structural properties. The changes in the hydrolysates microstructures and macromolecular conformation, induced by the UPT, were followed using scanning electron microscope analyzis (SEM) and Fourier transforms infrared spectroscopy (FTIR). The results showed that UPT influenced (P lt 0.05) the proteolysis of egg white proteins for all examined treatment times. Alcalase hydrolysates (AHs) and papain hydrolysates (PHs) were found to have a higher solubility, as a consequence of their relatively higher foaming, and emulsifying properties compared to the untreated hydrolysates. The changes in surface hydrophobicity, sulfhydryl content and surface charge of AHs and PHs indicated unfolding of EWPs affected by ultrasound. SEM analyzis showed that UPT destroyed the microstructures of AHs and PHs, while FTIR spectra indicated remarkable changes in the macromolecular conformation of AHs and PHs after UPT. This study revealed that by combining ultrasound pre-hydrolysis treatment under controlled conditions with thoughtful proteases selection, hydrolysates with improved functional properties could be produced, enhancing utilization of EWPs in food products.

Native wheat gluten is a deluxe bread improver and may be utilized as a functional protein additive in multifarious non-bakery foodstuffs due to its desirable structure-enhancing properties. Also its utilization would be economically interesting, but lack of some desirable functional properties limited their expanding utilization in foodstuff formulations. This study was designed to examine the relationship between process parameters and functional properties of the obtained hydrolysates using Box-Benken experimental design and response surface methodology (RSM). The hydro-lysate showing the highest improvement of solubility and foaming ability was further separated by sequential ultrafiltration to obtain molecular weight distribution profile. The progress of wheat gluten hydrolysis was followed by monitoring the degree of hydrolysis (DH) using the pH-stat method and functional properties were measured by our methods already adopted. The effects of process parameters (pH, T, [S], [E]/[S] ration) and their interactions were investigated by the means of the four-factor Box-Behnken experimental design with 29 experimental points (5 central points). Experiments were carried out in triplicates and expressed as means with SD. The effects of different parameters under the significance level of p < 0.05 were examined using one-way analysis of variance (ANOVA) and Student t-test. The coefficients of the response function and their statistical significance were evaluated by the response surface regression analysis, using the Design software. Non-significant terms (p ≥ 0.05) were deleted from the second order polynomial and a new polynomial has been recalculated. The Fisher test (F-value) was used to determine whether the second-order model was adequate to describe the obtained data while the goodness of fit of the model was evaluated by the determination coefficient (R2). The obtained results showed that the second-order models developed for DH, solubility and foaming prop

The aim of the study was to investigate and compare several extraction protocols like 1) high-power ultrasound probe assisted solvent extraction; 2) microwave assisted solvent extraction; 3) direct acid hydrolysis; 4) direct alkali hydrolysis, and 5) two step extraction consisting of ultrasound or microwave assisted solvent extraction followed by alkaline and acid hydrolysis in terms of efficiency of the extraction of phenolic acids from the yellow soybean seed variety Laura. These extracts were screened for their total phenol content (TPC), and for their antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl radical scavenging (DPPH) assay, as well as on content of some individual phenolic acids. It appeared that the acetone containing mixtures gave much higher TPC than methanol containing ones, but the presence of acid in the methanol solvent significantly improved the extraction of phenolic compounds. To further improve the extraction, an ultrasound lab-scale probe at 20 kHz was used, with 15 and 30% of the maximum amplitude, and the extraction time was varied from 2 to 15 min. Microwave assisted extraction was performed varying the temperature from 55 to 85 degrees C, microwave power from 25 to 100 W and extraction time from 2 to 10 min. Changes in the content of six phenolic acids were examined: gallic, trans cinnamic, chlorogenic, caffeic, p-coumaric and ferulic acid. The separation and quantification of phenolic acids was accomplished by high-performance liquid chromatography-diode array detection (DAD) procedure. The results suggested that microwaves contributed to more efficient extraction of phenolic acids from the seed of yellow soybean. The amount of phenolic acids varied from 65.52 mu g/g of dry matter (d.m.) for caffeic acid, to 581.84 mu g/g d.m. for p-coumaric acid. Both, ultrasound and microwaves contributed to more efficient extraction of total phenol compounds and enhanced antioxidant activity of soybean seed extracts. TPC varied from 12.48 to 1

Predmet naučnog istraţivanja doktorske disertacije je razvoj novih netermičkih i enzimskih procesa za proizvodnju proteina belanceta i njihovih hidrolizata sa unapreĎenim tehnološko-funkcionalnim svojstvima kao visokokvalitetnih proteinskih dodataka hrani i napicima..., The scope of this academic doctoral research is the development of new non-thermal and enzymatic processes in order to obtain high-qualitiy egg white protein and their hydrolysates with improved technological-functional properties as high-quality protein additives for food and beverages...

Study on potential of different agro-industrial waste residues for supporting the growth of newly isolated Bacillus sp. TMF-1 strain under solid-state fermentation (SSF) was conducted aiming to produce several industrially valuable enzymes. Since the feasibility of the initial study was confirmed, further objectives included evaluation of several pretreatments of the studied agricultural by-products (soybean meal, sunflower meal, maize bran, maize pericarp, olive oil cake and wheat bran) on the microbial productivity as means of enhancing the yields of produced proteases, alpha-amylases, cellulases and pectinases. Among acid/alkaline treatment, ultrasound and microwave assisted methods, chemical treatments superiorly affected most of the studied substrates. Highest yields of produced proteases (50.5 IU g (1)) and alpha-amylases (50.75 IU g (1)) were achieved on alkaline treated corn pericarp. Alkaline treatment also promoted the secretion of cellulases on maize bran (1.19 IU g (1)). Highest yield of pectinases was obtained on untreated soybean meal (64.90 IU g (1)).

The objective of this research was to investigate the impact of high-intensity ultrasound (HIU) generated by a probe-type sonicator (frequency 20 +/- 0.2 kHz and an amplitude of 40%) for 2-20 min on the selected functional and structural properties of egg white proteins (EWPs) and their susceptibility to hydrolysis by alcalase. The protein solubility, foaming, and emulsifying properties were studied as a function of ultrasonication time and related to protein particle and structural properties. The length of ultrasonication exhibited important effect on EWP particle size, uniformity, and charge, affecting also the protein conformation and susceptibility to alcalase hydrolysis and determining functional properties. There was a linear correlation between the particle size decrease and the solubility while a two-step linear correlation between the foam capacity (FC)/foam stability (FS) and particle size was apparent. Specifically, FC and FS sharply increased with decreasing particle size for range from similar to 370 to similar to 260 nm, and below this range from 260.6 to 68.4 nm, the changes were not that substantial. Besides, the solubility, FC, and FS were directly and linearly related with the absolute value of the particle zeta potential. The overall emulsifying properties were also improved with an increase of sonication time, through both the decrease of the mean particle diameter and the increase of zeta potential, but there was no direct correlation between the emulsion activity/stability index and protein particle size and/or charge. Analysis of EWP structure by Raman spectroscopy revealed that the HIU leads to changes in the secondary structure, while heat and ultrasound generated by the ultrasound bath were not sufficient to exhibit this effect.

The aim of the study was to find the optimal operational and process parameters for the enzymatic hydrolysis of wheat gluten in a batch stirred bioreactor regarding both degree of hydrolysis and antioxidant capacity of the obtained hydrolysates. It appeared that impeller geometry and agitation speed influenced the mass transfer resulting in enhanced gluten hydrolysis. The highest initial reaction rate (0.83 0.02 min(-1)) and degree of hydrolysis (30.47%) were achieved with the pitched four-bladed impeller and agitation speed of 350-450 rpm, conditions which provided proper balance between requirements for adequate mass/heat transfer and low shear stress. The impact of other process conditions including gluten concentration, temperature, pH and enzyme gluten (E/S) ratio on the enzymatic reaction was investigated by applying a Box-Behnken experimental design from the viewpoint of the degree of hydrolysis (DH) and antioxidant activity. Three models obtained allowed calculation of the hydrolysis degree, and both 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ABTS (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulpfonic acid) radical scavenging activity from a given set of reaction conditions with good predictability. The statistical analysis showed that each variable had a significant effect on degree of hydrolysis and the antioxidant capacity of both tested systems. Hydrolysis up to around DH 15% improved DPPH radical scavenging activity, while excessive hydrolysis worsened it. The ABTS activity of the hydrolysates was not associated with the DPPH activity nor with the DH, revealing that it was not possible to fulfill all desirable quality requirements (maximum degree of hydrolysis and protein yield, maximum DPPH and ABTS scavenging activities) by using the Same reaction conditions. Overall, the study might contribute to approve wheat gluten, a by-product of wheat starch industry, as an accessible and cheap source of bioactive compounds for the development of novel nutraceuticals

In order to confirm the gluten potential for inclusion into functional foods, the synergistic effect of the heat treatment and controlled enzymatic hydrolysis on the functional and the antioxidant properties of alcalase-assisted wheat gluten hydrolysates (AWGHs) will be discussed. For this purpose, wheat gluten was heat-treated during 30min at 75 degrees C and intensively hydrolyzed with alcalase at degree of hydrolysis (DH) 16.1%, 22.9%, and 30.2%. All the hydrolysates had excellent solubility over a pH range of 2-12. Emulsifying activity and stability were also improved, while proteolysis was deleterious to foam capacity and stability, water-holding capacity, fat-binding capacity and did not show improvement at higher DH (22.9% and 30.2%). As well, controlled hydrolysis of heat-treated gluten resulted in a remarkable improvement in antioxidant activities. The results show that the heat-treated AWGHs were superior to the untreated hydrolysate in the functional and antioxidant properties tested. Practical applicationsThis report examines existing evidence regarding the wheat gluten proteins (WGP), which is a byproduct from wheat starch processing. It is known that enzymatic hydrolysis is frequently used to improve functional properties of protein hydrolysates and largely dependent on the degree of hydrolysis (DH), which needs to be controlled to elude redundant proteolysis that can deteriorate functionality and cause unfavorable effects. The DH is a substantial factor which affect the hydrolysates' performances and an appropriate selection of protease for WGP hydrolysis will result in maximum biological activity and improved functionalities. Heat treatment is often used to facilitate the proteolysis of proteins. Thus, functional and antioxidant properties of WGP hydrolysates, as a function of heat treatment and the DH were adequately examined in this study and results showed that by combining heat prehydrolysis treatment under controlled conditions, hydrolysates wit

A selection of best combination of adequate immobilization support and efficient immobilization method is still a key requirement for successful application of immobilized enzymes on an industrial level. Eupergit® supports exhibit good mechanical and chemical properties and allow establishment of satisfactory hydrodynamic regime in enzyme reactors. This is advantageous for their wide application in enzyme immobilization after finding the most favorable immobilization method. Methods for enzyme immobilization that have been previously reported as efficient considering the obtained activity of immobilized enzyme are presented: direct binding to polymers via their epoxy groups, binding to polymers via a spacer made from ethylene diamine/glutaraldehyde, and coupling the periodate-oxidized sugar moieties of the enzymes to the polymer beads. The modification of the conventionally immobilized enzyme with ethylenediamine via the carbodiimide route seems to be a powerful tool to improve its stability and catalytic activity.

The aim of the study was to find the optimal operational and process parameters for the enzymatic hydrolysis of wheat gluten in a batch stirred bioreactor regarding both degree of hydrolysis and antioxidant capacity of the obtained hydrolysates. It appeared that impeller geometry and agitation speed influenced the mass transfer resulting in enhanced gluten hydrolysis. The highest initial reaction rate (0.83 0.02 min(-1)) and degree of hydrolysis (30.47%) were achieved with the pitched four-bladed impeller and agitation speed of 350-450 rpm, conditions which provided proper balance between requirements for adequate mass/heat transfer and low shear stress. The impact of other process conditions including gluten concentration, temperature, pH and enzyme gluten (E/S) ratio on the enzymatic reaction was investigated by applying a Box-Behnken experimental design from the viewpoint of the degree of hydrolysis (DH) and antioxidant activity. Three models obtained allowed calculation of the hydrolysis degree, and both 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ABTS (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulpfonic acid) radical scavenging activity from a given set of reaction conditions with good predictability. The statistical analysis showed that each variable had a significant effect on degree of hydrolysis and the antioxidant capacity of both tested systems. Hydrolysis up to around DH 15% improved DPPH radical scavenging activity, while excessive hydrolysis worsened it. The ABTS activity of the hydrolysates was not associated with the DPPH activity nor with the DH, revealing that it was not possible to fulfill all desirable quality requirements (maximum degree of hydrolysis and protein yield, maximum DPPH and ABTS scavenging activities) by using the Same reaction conditions. Overall, the study might contribute to approve wheat gluten, a by-product of wheat starch industry, as an accessible and cheap source of bioactive compounds for the development of novel nutraceuticals

In order to confirm the gluten potential for inclusion into functional foods, the synergistic effect of the heat treatment and controlled enzymatic hydrolysis on the functional and the antioxidant properties of alcalase-assisted wheat gluten hydrolysates (AWGHs) will be discussed. For this purpose, wheat gluten was heat-treated during 30min at 75 degrees C and intensively hydrolyzed with alcalase at degree of hydrolysis (DH) 16.1%, 22.9%, and 30.2%. All the hydrolysates had excellent solubility over a pH range of 2-12. Emulsifying activity and stability were also improved, while proteolysis was deleterious to foam capacity and stability, water-holding capacity, fat-binding capacity and did not show improvement at higher DH (22.9% and 30.2%). As well, controlled hydrolysis of heat-treated gluten resulted in a remarkable improvement in antioxidant activities. The results show that the heat-treated AWGHs were superior to the untreated hydrolysate in the functional and antioxidant properties tested. Practical applicationsThis report examines existing evidence regarding the wheat gluten proteins (WGP), which is a byproduct from wheat starch processing. It is known that enzymatic hydrolysis is frequently used to improve functional properties of protein hydrolysates and largely dependent on the degree of hydrolysis (DH), which needs to be controlled to elude redundant proteolysis that can deteriorate functionality and cause unfavorable effects. The DH is a substantial factor which affect the hydrolysates' performances and an appropriate selection of protease for WGP hydrolysis will result in maximum biological activity and improved functionalities. Heat treatment is often used to facilitate the proteolysis of proteins. Thus, functional and antioxidant properties of WGP hydrolysates, as a function of heat treatment and the DH were adequately examined in this study and results showed that by combining heat prehydrolysis treatment under controlled conditions, hydrolysates wit

The effects of some relevant process parameters for the Alcalase-catalyzed wheat gluten hydrolysis such as gluten concentration (X1; 1-9% w/v), temperature (X2; 40-60 oC), pH (X3; 7-9) and enzyme/substrate ratio, [E]/[S] ratio (X4; 0.25– 0.75 AU/g of protein) were investigated by the means of an experimental design. The second-order models developed for the degree of hydrolysis, DH, DPPH and ABTS radical scavenging activity of gluten hydrolysates were significant (p<0.01) with a high value of coefficients of determination (0.981-0.992). The statistical analysis showed that each variable had a significant effect on degree of hydrolysis and the antioxidant capacity of both tested systems. Hydrolysis up to around DH = 15% improved DPPH radical scavenging activity, while excessive hydrolysis worsened it. It seemed there was not a correspondence between degree of hydrolysis and ABTS activity at different protein concentrations, suggesting that there were peptides with considerable size presenting a good antioxidant capacity.

In addition to evidence regarding their potential for inclusion into functional foods, the effect of the high-intensity ultrasound extraction (HIUE) on the functional properties of soy proteins will be discussed. The aim of this study was to examine the effects of HIUE as a function of the treatment time and ultrasound amplitude, in order to obtain better understanding of physicochemical effects of HIUE on soy protein which may lead to improving its applications in the food industry. For this purpose, samples of the yellow defatted soybean (Glycine max) variety Laura were treated with ultrasound probe system (20±0.2 kHz) by varying the treatment time (30 s, 2, 5, 10, 15 min) and an amplitude (15% and 30%). Functional properties of the obtained samples were characterized in terms of solubility and sulfhydryl content. HIUE resulted in a slight, but gradual decline in solubility with a treatment time, but significant increase (p < 0.05) in acidic environment was observed with the increase of ultrasound amplitude. Sulfhydryl content has been greater influenced with a treatment time for both amplitude applied. The results show that the ultrasound extracted soy proteins were superior to the original (untreated) in the functional properties tested and can be concluded that by combining the treatment time and the ultrasonic power, the samples with enhanced functionalities can be produced enhancing utilization of soy proteins in food products.

In order to confirm the gluten potential for inclusion into functional foods, the synergistic effect of the heat treatment and controlled enzymatic hydrolysis on the functional and the antioxidant properties of alcalase-assisted wheat gluten hydrolysates (AWGHs) will be discussed. For this purpose, wheat gluten was heat-treated during 30min at 75 degrees C and intensively hydrolyzed with alcalase at degree of hydrolysis (DH) 16.1%, 22.9%, and 30.2%. All the hydrolysates had excellent solubility over a pH range of 2-12. Emulsifying activity and stability were also improved, while proteolysis was deleterious to foam capacity and stability, water-holding capacity, fat-binding capacity and did not show improvement at higher DH (22.9% and 30.2%). As well, controlled hydrolysis of heat-treated gluten resulted in a remarkable improvement in antioxidant activities. The results show that the heat-treated AWGHs were superior to the untreated hydrolysate in the functional and antioxidant properties tested. Practical applicationsThis report examines existing evidence regarding the wheat gluten proteins (WGP), which is a byproduct from wheat starch processing. It is known that enzymatic hydrolysis is frequently used to improve functional properties of protein hydrolysates and largely dependent on the degree of hydrolysis (DH), which needs to be controlled to elude redundant proteolysis that can deteriorate functionality and cause unfavorable effects. The DH is a substantial factor which affect the hydrolysates' performances and an appropriate selection of protease for WGP hydrolysis will result in maximum biological activity and improved functionalities. Heat treatment is often used to facilitate the proteolysis of proteins. Thus, functional and antioxidant properties of WGP hydrolysates, as a function of heat treatment and the DH were adequately examined in this study and results showed that by combining heat prehydrolysis treatment under controlled conditions, hydrolysates wit

The objective of this research was to investigate the impact of high-intensity ultrasound (HIU) generated by a probe-type sonicator (frequency 20 +/- 0.2 kHz and an amplitude of 40%) for 2-20 min on the selected functional and structural properties of egg white proteins (EWPs) and their susceptibility to hydrolysis by alcalase. The protein solubility, foaming, and emulsifying properties were studied as a function of ultrasonication time and related to protein particle and structural properties. The length of ultrasonication exhibited important effect on EWP particle size, uniformity, and charge, affecting also the protein conformation and susceptibility to alcalase hydrolysis and determining functional properties. There was a linear correlation between the particle size decrease and the solubility while a two-step linear correlation between the foam capacity (FC)/foam stability (FS) and particle size was apparent. Specifically, FC and FS sharply increased with decreasing particle size for range from similar to 370 to similar to 260 nm, and below this range from 260.6 to 68.4 nm, the changes were not that substantial. Besides, the solubility, FC, and FS were directly and linearly related with the absolute value of the particle zeta potential. The overall emulsifying properties were also improved with an increase of sonication time, through both the decrease of the mean particle diameter and the increase of zeta potential, but there was no direct correlation between the emulsion activity/stability index and protein particle size and/or charge. Analysis of EWP structure by Raman spectroscopy revealed that the HIU leads to changes in the secondary structure, while heat and ultrasound generated by the ultrasound bath were not sufficient to exhibit this effect.

Study on potential of different agro-industrial waste residues for supporting the growth of newly isolated Bacillus sp. TMF-1 strain under solid-state fermentation (SSF) was conducted aiming to produce several industrially valuable enzymes. Since the feasibility of the initial study was confirmed, further objectives included evaluation of several pretreatments of the studied agricultural by-products (soybean meal, sunflower meal, maize bran, maize pericarp, olive oil cake and wheat bran) on the microbial productivity as means of enhancing the yields of produced proteases, alpha-amylases, cellulases and pectinases. Among acid/alkaline treatment, ultrasound and microwave assisted methods, chemical treatments superiorly affected most of the studied substrates. Highest yields of produced proteases (50.5 IU g (1)) and alpha-amylases (50.75 IU g (1)) were achieved on alkaline treated corn pericarp. Alkaline treatment also promoted the secretion of cellulases on maize bran (1.19 IU g (1)). Highest yield of pectinases was obtained on untreated soybean meal (64.90 IU g (1)).

A selection of best combination of adequate immobilization support and efficient immobilization method is still a key requirement for successful application of immobilized enzymes on an industrial level. Eupergit® supports exhibit good mechanical and chemical properties and allow establishment of satisfactory hydrodynamic regime in enzyme reactors. This is advantageous for their wide application in enzyme immobilization after finding the most favorable immobilization method. Methods for enzyme immobilization that have been previously reported as efficient considering the obtained activity of immobilized enzyme are presented: direct binding to polymers via their epoxy groups, binding to polymers via a spacer made from ethylene diamine/glutaraldehyde, and coupling the periodate-oxidized sugar moieties of the enzymes to the polymer beads. The modification of the conventionally immobilized enzyme with ethylenediamine via the carbodiimide route seems to be a powerful tool to improve its stability and catalytic activity.

The aim of the study was to find the optimal operational and process parameters for the enzymatic hydrolysis of wheat gluten in a batch stirred bioreactor regarding both degree of hydrolysis and antioxidant capacity of the obtained hydrolysates. It appeared that impeller geometry and agitation speed influenced the mass transfer resulting in enhanced gluten hydrolysis. The highest initial reaction rate (0.83 0.02 min(-1)) and degree of hydrolysis (30.47%) were achieved with the pitched four-bladed impeller and agitation speed of 350-450 rpm, conditions which provided proper balance between requirements for adequate mass/heat transfer and low shear stress. The impact of other process conditions including gluten concentration, temperature, pH and enzyme gluten (E/S) ratio on the enzymatic reaction was investigated by applying a Box-Behnken experimental design from the viewpoint of the degree of hydrolysis (DH) and antioxidant activity. Three models obtained allowed calculation of the hydrolysis degree, and both 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ABTS (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulpfonic acid) radical scavenging activity from a given set of reaction conditions with good predictability. The statistical analysis showed that each variable had a significant effect on degree of hydrolysis and the antioxidant capacity of both tested systems. Hydrolysis up to around DH 15% improved DPPH radical scavenging activity, while excessive hydrolysis worsened it. The ABTS activity of the hydrolysates was not associated with the DPPH activity nor with the DH, revealing that it was not possible to fulfill all desirable quality requirements (maximum degree of hydrolysis and protein yield, maximum DPPH and ABTS scavenging activities) by using the Same reaction conditions. Overall, the study might contribute to approve wheat gluten, a by-product of wheat starch industry, as an accessible and cheap source of bioactive compounds for the development of novel nutraceuticals

Study on potential of different agro-industrial waste residues for supporting the growth of newly isolated Bacillus sp. TMF-1 strain under solid-state fermentation (SSF) was conducted aiming to produce several industrially valuable enzymes. Since the feasibility of the initial study was confirmed, further objectives included evaluation of several pretreatments of the studied agricultural by-products (soybean meal, sunflower meal, maize bran, maize pericarp, olive oil cake and wheat bran) on the microbial productivity as means of enhancing the yields of produced proteases, alpha-amylases, cellulases and pectinases. Among acid/alkaline treatment, ultrasound and microwave assisted methods, chemical treatments superiorly affected most of the studied substrates. Highest yields of produced proteases (50.5 IU g (1)) and alpha-amylases (50.75 IU g (1)) were achieved on alkaline treated corn pericarp. Alkaline treatment also promoted the secretion of cellulases on maize bran (1.19 IU g (1)). Highest yield of pectinases was obtained on untreated soybean meal (64.90 IU g (1)).

Kaolin showed as a very perspective carrier for the enzyme immobilization and it was used for the adsorption of horseradish peroxidase (HRP). The effects of the enzyme concentration and pH on the immobilization efficiency were studied in the reaction with pyrogallol and anthraquinone dye C.I. Acid Violet 109 (AV 109). In addition, Fourier transform infrared spectroscopy, scanning electron microscopy and analysis by Brunauer-EmmettTeller were performed for kaolin, thermally activated kaolin and the immobilized enzyme. It has been shown that 0.1 IU of HRP-kaolin decolorized 87 % of dye solution, under the optimal conditions (pH 5.0, temperature 24 degrees C, dye concentration 40 mg/L and 0.2 mM of H2O2) within 40 min. The immobilized HRP decolorization follows the Ping Pong Bi-Bi mechanism with dead-end inhibition by the dye. The biocatalyst retained 35 +/- 0.9 % of the initial activity after seven cycles of reuse in the decolorization reaction of AV 109 under optimal conditions in a batch reactor. The obtained kinetic parameters and reusability study confirmed improvement in performances of k-HRP compared to free, indicating that k-HRP has a great potential for environmental purposes.

The impact of ultrasound waves generated by probe-type sonicator and ultrasound cleaning bath on egg white protein susceptibility to hydrolysis by alcalase compared to both thermal pretreatment and conventional enzymatic hydrolysis was quantitatively investigated. A series of hydrolytic reactions was carried out in a stirred tank reactor at different substrate concentrations, enzyme concentrations, and temperatures using untreated, and pretreated egg white proteins (EWPs). The kinetic model based on substrate inhibition and second-order enzyme deactivation successfully predicts the experimental behavior providing an effective tool for comparison and optimization. The ultrasound pretreatments appear to greatly improve the enzymatic hydrolysis of EWPs under different conditions when compare to other methods. The apparent reaction rate constants for proteolysis (k(2)) are 0.009, 0.011, 0.053, and 0.045 min(-1) for untreated EWPs, and those pretreated with heat, probe-type sonicator, and ultrasound cleaning bath technologies, respectively. The ultrasound pretreatment also decreases hydrolysis activation (E-a) and enzyme deactivation (E-d) energy, enthalpy (Delta H), and entropy (Delta S) of activation and for the probe-type sonication this decrease is 61.7%, 61.6%, 63.6%, and 32.2%, respectively, but ultrasound has little change in Gibbs free energy value in the temperature range of 318 to 338 K. The content of sulfhydryl groups and zeta potential show a significant increase (P lt 0.05) for both applied ultrasound pretreatments and the reduction of particle size distribution are achieved, providing some evidence that the ultrasound causes EWP structural changes affecting the proteolysis rate.

The problem of environmental pollution becomes more worrisome day by day, primarily due to the large amounts of wastewater contaminated with various harmful organic compounds, discharged untreated or partially clean into the environment. Feasibility of use of horseradish peroxidase (Amoracia rusticana) in the synthetic dyes decolorization was approved by many researchers. Among a number of supports used for the immobilization, it was found that natural clay, kaolin, has excellent features which are a precondition for obtaining biocatalysts with the excellent performances. For this reason, a horseradish peroxidase was immobilized onto kaolin using glutaraldehyde as a cross-linking agent. Obtained biocatalyst was applied in the decolorization of anthraquinone dye C.I. acid violet 109. Under determined optimal conditions (pH 4.0, hydrogen peroxide concentration 0.6 mM, dye concentration 30 mg L-1, temperature 24 degrees C) around 76% of dye decolorization was achieved. Reusability study showed that resulting biocatalyst was possible to apply four times in the desired reaction with relatively high decolorization percentage.

The production of bioactive peptides from egg white proteins (EWPs) and their separation are emerging areas with many new applications. The objective of this study was to compare antioxidant activity of three distinct EWP hydrolysates and their peptide fractions prepared by membrane ultrafiltration using membranes with 30, 10 and 1 kDa molecular weight cut-off. The hydrolysates were obtained by thermal and ultrasound pretreated EWPs hydrolyzed with a bacterial protease, Alcalase. It appeared that the pretreatment significantly affected peptide profiles and antioxidant activity of the hydrolysates measured by ABTS, DPPH and FRAP methods. The hydrolysate prepared using alcalase and ultrasound pretreatment at 40 kHz - 15 min has shown to be the most effective in scavenging both DPPH and ABTS radicals (28.10 +/- 1.38 and 79.44 +/- 2.31%, respectively). It has been noticed that this hydrolysate had a nutritionally more adequate peptide profile than the other hydrolysates with a much lower amount of peptides lt 1 kDa (11.19 +/- 0.53%) and the greatest content of the peptide fraction in the molecular weight (MW) range of 1-10 kDa (28.80 +/- 0.07%). This peptide fraction has shown the highest DPPH and ABTS antioxidant activity compared to all other fractions having a potential to be used as a functional food ingredient.

Finding a sustainable and ecofriendly methods for recalcitrant synthetic dyes removal is a researchers major challenge. A carrier-free technique for commercial HRP immobilization is a focus of the present study. The immobilized biocatalyst, HRP-CLEAs with 580 U g-1 of the activity was obtained under the following immobilization conditions: precipitation reagent 80% ammonium sulphate, cross-linking reagent 1% of glutaraldehyde and protein-fedder, bovine serum albumin (BSA) concentration 5 mg ml-1. The obtained HRP-CLEAs showed great affinity towards model anthraquinone dye, C. I. Acid Violet 109. 88.4% of the dye was oxidized under the reaction conditions: pH 4, dye concentration 40 mg l-1, H2O2 concentration 1 mM and 0.1 U of HRP-CLEAs. The possibility of the immobilized biocatalyst application in five and eight oxidation cycles of the dye and pyrogallol (retained activity 80%), respectively, indicates that HRP-CLEAs is an efficient and environmentally friendly biocatalyst with great potential in aromatic compounds removal from wastewater. This paper is a continuation of our earlier research related to HRP from horseradish extract immobilization in the form of CLEAs and the application in the wastewater colored with a synthetic anthraquinone dye treatment., Pronalazak održivih i ekološki prihvatljivih metoda za uklanjanje sintetičkih boja je jedan od vodećih izazova za istraživače. U ovom radu akcenat je na metodi imobilizacije bez primene nosača. Imobilisani preparat peroksidaze iz rena, aktivnosti 580 U g-1 dobijen je pod sledećim uslovima imobilizacije: taložni reagens 80% amonijum-sulfat, umrežavajući reagens 1% glutaraldehid i goveđi serum albumin koncentracije 5 mg ml-1. Dobijeni umreženi agregat peroksidaze iz rena pokazao je veliki afinitet prema model antrahinonskoj boji, C. I. AV 109. Pod optimalnim uslovima: pH 4, koncentracja boje 40 mg-1, koncentracija H2O2 i 0,1 U enzimske aktivnosti oksidovano je 88,4% ispitivane boje. Mogućnost primene imobilisanog enzima kroz pet i osam uzastopnih ciklusa oksidacije boje i pirogalola (zadržana aktivnost 80%) pokazuje da je imobilisani enzim efikasan i ekološki prihvatljiv biokatalizator koji se može koristiti u oksidaciji aromatičnih jedinjenja iz otpadnih voda. Ovaj rad predstavlja nastavak istraživanja vezanog za umrežavanje peroksidaze iz svežeg ekstrakta rena, koja se takođe pokazala kao imobilisani biokatalizator sa velikim potencijalom primene u tretmanu obojenih otpadnih voda.

Distillery stillage is abundant industrial waste with a great potential for utilization as a substrate in production of valuable bio-based products. Processing of distillery stillage on cost effective way can significantly decrease the price of bioethanol as alternative fuel and provide a considerable benefit to the environment. In this paper, combined potato stillage and sugar beet molasses substrate is evaluated for integrated lactic acid and probiotic biomass production by Lactobacillus rhamnosus ATCC 7469. Waste substrate based on potato stillage and sugar beet molasses enabled lactic acid and biomass production with maximal volumetric lactic acid productivity of 1.11 g L-1 h-1 and maximal number of viable L. rhamnosus ATCC 7469 cells of 1.1×109 CFU mL-1. Residue after removal of lactic acid could be recommended as probiotics and betaine-enriched animal feed., Destilerijska džibra je lako dostupan industrijski otpad koji nastaje u proizvodnji bioetanola, a zbog svog kompleksnog sastava može biti potencijalno dobar supstrat u mnogim biotehnološkim procesima. Imajući u vidu potrebu za povećanjem postojećih i izgradnjom novih kapaciteta za proizvodnju bioetanola, iskorišćavanje otpadne džibre može biti značajno sa aspekta povećanja konkurentnosti bioetanola kao alternativnog goriva, a može imati i pozitivan uticaj na životnu sredinu. U radu je ispitivana mogućnost iskorišćavanja krompirove džibre i melase šećerne repe kao supstrata za paralelnu proizvodnju mlečne kiseline i bakterijske biomase, pomoću probiotskog soja Lactobacillus rhamnosus ATCC 7469. U šaržnoj fermentaciji na ovom kombinovanom supstratu bez dodatnog obogaćivanja vitaminima, mineralnim materijama ili izvorima azota je postignuta maksimalna produktivnost od 1,11 g L-1 h-1, maksimalan broj vijabilnih ćelija od 1,1×109 CFU mL-1 i koncentracija mlečne kiseline od 18,08 g L-1. Krompirova džibra se može koristiti kao supstrat za rast probiotske biomase i može biti adekvatna zamena za skupe, najčešće korišćene organske izvore azota, kao što su kvaščev ekstrakt ili pepton. Usled nemogućnosti L. rhamnosus ATCC 7469 da podjednako dobro metaboliše sve šećere u melasi šećerne repe za efikasniju proizvodnju mlečne kiseline treba razmotriti druge izvore ugljenika ili izvršiti selekciju Lactobacillus sp. koji efikasno fermentiše saharozu iz melase. Zbog značajnog rasta bakterijske biomase tokom procesa, fermentisani medijum koji zaostaje nakon ekstrakcije mlečne kiseline predstavlja vredan sporedni proizvod fermentacije bogat probiotskom biomasom i betainom koji se može koristiti kao visoko kvalitetna hrana za životinje.