Sareh Boostani; Marzieh Moosavi-Nasab; Mahmoud Aminlari; Mehrdad Niakosari; Gholam Reza Mesbahi
Abstract
Introduction: Proteins play a fundamental role in biological systems and are often sensitive against organic solvents, heat and other damaging factors. Proteins are the basic component of food formulations and enhancement the functional characteristics and stability of the proteins has always been the ...
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Introduction: Proteins play a fundamental role in biological systems and are often sensitive against organic solvents, heat and other damaging factors. Proteins are the basic component of food formulations and enhancement the functional characteristics and stability of the proteins has always been the main goal of food industry engineers. One of the natural ways used for protein modifications is Maillard reaction. Maillard reaction as a result of covalent binding between the available amino groups of the proteins and carbonyl containing moiety of the polysaccharides, causes a loss in free amino group content of the mixture. Protein- polysaccharide hybrids, as a result of dry heating of two biopolymers mixture under controlled reaction conditions, cause the emergence of conjugates with novel functionalities. Much research has shown that conjugation can increase thermal stability and functional characteristics of food proteins and also reduce the allergenicity of suspected proteins. Although many studies have been conducted on the effects of conjugation on functional properties of proteins, the impacts of conjugation on proteins behavior after food processing have been less investigated. So, in this paper the influences of Maillard reaction on functional properties of soy proteins have been investigated. In addition, characteristics of conjugated proteins after pasteurization treatment was also studied. Materials and methods: Construction of protein- polysaccharide conjugates was done in several steps. First, soy proteins and dextran were mixed with phosphate buffer (0.1 M, pH: 8.5) and 1 to 4 ratio of protein to polysaccharide. After mixing and incubating at ambient temperature for some hours, solutions were frozen at –80 ℃ and freeze dried. Then the lyophilized powder was incubated at different times, at 60℃, under the 79 percent relative humidity in presence of saturated KBr. For each treatment a non conjugated sample was prepared in the exact same condition. Conjugation of proteins to polysaccharides was monitored by SDS-PAGE electrophoresis, browning intensity measurement and UV absorbance analysis. SDS-PAGE was conducted according to Laemmli procedure using a discontinuous buffer system. A vertical gel electrophoresis unit was used with 3% acrylamide stacking gel and 10% acrylamide running gel. Evaluation of the color changes as an indicator of grafting intensity was investigated by monitoring absorption at a wavelength of 420 nm. To investigate the UV absorption of conjugated proteins, the samples were diluted with SDS (Sodium dodecyl sulfate) solution and the absorption was read by a UV-visible spectrophotometer at 294 nm. The impact of modification on characteristics of soy proteins was monitored by examining the functional properties changes of protein samples. In the last stage soy drinks were prepared from conjugated and non conjugated proteins then the prepared beverages were subjected to thermal processing conditions and the influences of Maillard conjugation on the stability of soy drinks was monitored over time. Results and Discussion: SDS-PAGE electrophoresis profile showed that proteins-polysaccharide conjugates were formed. As a result of conjugation, the protein-dextran covalent binding occurs, leads to the formation of higher molecular weight components, resulting in its accumulation on the top of the separating gel. When heating time increased a wider and higher molecular weight bands appeared near the top of the running gel however they were not observed in the native soy pattern. Covalent linkage between amino group of proteins and carbonyl group of polysaccharides causes a color changes from light yellow to brown, browning intensity results showed that the even during early incubation time, a significant absorption was observed at 420 nm compared to the control samples. UV absorption results showed similar trend of changes in browning intensity measurement. Increasing UV absorption is due to the intermediate Maillard reaction products (MRP). Increasing UV absorption with increasing heating time indicates the fact that Maillard reaction products (MRP) formation are more favorable in the long incubation time. Data of UV absorption are a good evidence for SDS-PAGE and browning intensity results. Functional properties results indicated that grafted proteins had better functional properties. The storage stability of soy drinks prepared from conjugated proteins was significantly higher than the samples prepared from non conjugated proteins. Stability of beverages after thermal processing and during storage is one of the most important features of protein drinks and many efforts have been made to develop mentioned characteristics. Stability of soy drinks produced from the conjugated proteins was significantly higher than those prepared from non conjugated soy proteins. Functional characterization of proteins is dependent on several factors, the majority of soy drink composed of proteins that could be denaturated by heating applied during thermal processing, as the results showed conjugation with dextran caused an increase in denaturation temperature of soy proteins which enhance the resistance of proteins during thermal processing treatment. In addition, the solubility and emulsifying properties of soy proteins increased with conjugation which can be a good reason for improvement the relation between protein and surrounding water molecules and therefore increases the protein storage stability. It can be concluded that Maillard reaction could be applied as a means to prepare soy proteins–dextran conjugates with better functional properties and more stable during processing and storage.
Sareh Boostani; Mahmoud Aminlari; Marzieh Moosavi-Nasab; Mehrdad Niakosari; Gholam Reza Mesbahi
Abstract
Introduction : Soybean is an excellent plant protein source with diverse applications in food systems. Despite numerous commercial applications and rich nutritional properties of soybeans, soy proteins are sensitive to heat and other damaging agents during food processing which can limit their applications ...
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Introduction : Soybean is an excellent plant protein source with diverse applications in food systems. Despite numerous commercial applications and rich nutritional properties of soybeans, soy proteins are sensitive to heat and other damaging agents during food processing which can limit their applications in food industries. Maillard reaction includes a series of chemical reactions between the free carbonyl groups of carbohydrates and the un-protonated amino groups of proteins under mild experimental conditions. This is one of the most desirable approaches for applying in food systems, because of the safety of the procedure and the independency of adding extra chemicals. Natural occurring Maillard reaction can be a relatively safe and inexpensive method in order to improve functionalities of food proteins. The production of conjugates haspositive influences on food proteins functionality such as solubility, water holding capacity, emulsion activity and stability, foaming properties, thermal stability, whipping ability, textural and gelation properties and also reduce allergenicity of proteins. Due to the positive characteristics and reasonable price of both soy proteins and maltodextrin in food industries, the aim of current study was to enhance the heat stability and functional properties of soy proteins through glycosylation with maltodextrin. In addition, assessment of changes in protein properties as a function of incubation time were evaluatedMaterials and methods: Preparation of purified soy proteins (Acid precipitated soy proteins) was done by a multistage process of washing, centrifugation, dialysis and freeze drying. The resulting powder contained pure soy globulins. Conjugation of acid precipitated soy proteins with maltodextrin was performed according to the method described as follows: protein-polysaccharide at a weight ratio of 1: 3 were dissolved in 0.01 M phosphate buffer, at pH values of 8, and were incubated at ambient temperature for 1 hr. Solutions were frozen at –80°C and then freeze dried. Lyophilized powders were incubated at 60 °C for 1, 3, 5 and 7 days, under 79% of relative humidity provided by saturated KBr. For each treatment an un-conjugated (control) sample was prepared under the same conditions. The formation of protein-polysaccharide conjugates was confirmed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration chromatography (Sephadex G-100 chromatographic system was used to separate the conjugated proteins from the un conjugated samples). Determination of protein denaturation temperature changes were carried out using METTLER TA Q100-DSC thermal analyser.The emulsifying properties of the samples including emulsifying activity and emulsion stability were assessed according to the procedure established by Pearce and Kinsella. Protein solubility was measured by calculating the amount of nitrogen in the supernatant and total nitrogen content of the samples and reported as percentage of protein solubility at pH 3, 5, 7 and 9. Foaming properties of the samples including foaming capacity and foaming stabilitywere determined using calibrated measuring cylinderDiscussion & Results: When the heating duration is increased, wider and heavier molecular weight bands emerge near the top of the running gel of SDS-PAGE, and yet these were not observed in the control. As a result of conjugation, the protein-carbohydrate covalent binding occurs, producing heavier molecular weight species, and thus leading to its accumulation on top of the separating gel. Compared with un-modified soy proteins, the conjugated soy proteins eluted in the void volume of G-100 gel permeation chromatography column, suggesting increase in the size and molecular weight of soy proteins due to the covalent attachment of maltodextrin. According to differential scanning calorimetry (DSC) analysis, thermal stability of soy proteins was remarkably increased by conjugation with maltodextrin and maximum denaturation temperature was observed for the mixture incubated for 7 days. The improved thermal stability is manifested in increase in denaturation temperature of globular proteins, hence conjugation leads to significant improvement of soy proteins stability. Increase in thermal stability is the result of inclusion of the hydrophilic carbohydrate moiety to the surface of the proteins. Compared to control sample, the solubility, foaming characteristics and emulsifying properties were significantly improved by increasing incubation time. The protein solubility of conjugate remarkably increased at all pH’s compared with the un-conjugated proteins. Covalent links between hydrophilic maltodextrin and soy proteins could enhance the reaction tendency between proteins and water molecules under unfavorable conditions. Improvements in the emulsifying properties of the conjugated samples can be explained by the fact that there is a combination among the emulsifying activity of proteins and the stabilizing impacts of polysaccharides per molecule. Foaming capacity of proteins can be affected by the solubility of proteins. Furthermore, maltodextrin is a hydrophilic carbohydrate which can improve the stability of soy proteins foams by acting as a thickener, thus increasing the strength of bubbles. It should also be considered that functionality of proteins are frequently influenced by protein solubility, and this could also serve the understanding of why improvements occur in functional properties of conjugated proteins, compared to un-conjugated ones. The results indicate that physiochemical and functional properties of soy proteins were modified and improved by conjugation with maltodextrin.
Sareh Boostani; Mahmoud Aminlari; Marzieh Moosavi-Nasab; Mehrdad Niakosari; Gholam Reza Mesbahi
Abstract
Introduction: Engineering of food proteins with improved functional properties and higher resistance to heat is the main goal of food scientists. Food technologists are always seeking for innovative, simple and effective methods to manipulate proteins, hence natural modification has had the most attention ...
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Introduction: Engineering of food proteins with improved functional properties and higher resistance to heat is the main goal of food scientists. Food technologists are always seeking for innovative, simple and effective methods to manipulate proteins, hence natural modification has had the most attention in last decades. One of the natural ways used for protein modifications isMaillard grafting reaction. Maillard reaction as a consequence of covalentbindingbetween the available amino groups of the proteins and carbonyl containing moiety of the polysaccharides, causes a loss in free amino group content of the mixture that can be measured through different methods. Protein-polysaccharide hybrids, as a result of dry heating of two biopolymers mixture under controlled reaction conditions, cause the emergence of conjugates with novel functionalities.Selecting appropriate reaction conditions has a significant impact on the properties of the final conjugates. Among the factors affecting the degree of conjugation, temperature, pH and incubation time have considerable roles in determining the degree of conjugation. There are various methods by which conjugation degree can be assessed and factors such as accuracy, cost, accessibility and etc. must be considered when selecting a measuring method. Therefoe, in this study the effect of different Maillard reaction conditions on the conjugation degree of soy proteins-dextran heated mixtures have been investigated. In addition, the glycation degree was compared and reported by both OPA assay and UV absorbance methods.Materials and methods: Soy proteins–dextran conjugates were prepared as described later. First, soy proteins and dextran were mixed with phosphate buffer (0.1 M, pH: 8.5 and 7) and 1 to 4 ratio of protein to polysaccharide. After mixing and incubating at ambient temperature for some hours, solutions were frozen at –80 ℃ and freeze dried. Then, the lyophilized powder was incubated at 40 ℃ for 0, 4, 8, 24 hours and 2, 4, 6, 8, 12 days, at 60 ℃ for 0, 1, 2, 3, 4, 6, 8 days and at 80 ℃ for 0, 1, 2, 4, 8, 16, 24, 48 hours, under the 79 percent relative humidity in presence of saturated KBr. For each treatment a non conjugated sample was prepared in the exact same condition. Conjugation of proteins to polysaccharides was monitored by two methods (OPA assay and UV absorbance). Determining degree of glycation by OPA method was done as follows, in this procedure the level of available amino groups was estimated using the ortho- phthaldialdehyde (OPA) reagent, the absorbance was measured at 340 nm and degree of glycation was measured using a formula. To investigate the UV absorption of conjugated proteins, the samples were diluted using distilled water and the absorption was read using a UV-visible spectrophotometer.Results & Discussion: Covalent linkage between amino group of proteins and carbonyl group of polysaccharides causes depletion in the amount of available amino groups. The extent of soy proteins-dextran conjugation under various Maillard reaction conditions was evaluated by the reduction of available amino groups of proteins, the more reduction in amount of amino groups, the more conjugation between protein and polysaccharide. OPA results showed that, in the samples heated at 40 °C and 80 °C (at both pH 7 and 8.5), the amount of free amino groups slightly reduced compared to 60 °C heated samples. The disappearance of available amino groups at 60°C was faster than other temperatures and formation of conjugates in this temperature was more successful. A stepwise reduction in free amino group content observed with increasing incubation time. When soy proteins were incubated at pH 8.5 for 8 days at 60 °C, a considerable decrease in available amino group contents occurred. UV absorption results showed similar trend of changes in OPA method. Increasing UV absorption is due to the intermediate Maillard reaction products (MRP). Increasing incubation time, temperature and pH cause a significant increase in UV absorbance. Increasing UV absorption with increasing heating time indicates the fact that Maillard reaction products (MRP) formation is more favorable at alkaline pH. Data of UV absorption are a proper evidence for OPA assay results. Conclusion: As a conclusion, both OPA assay and UV absorption methods are cost effective, accurate and simple methods which can represent valuable information concerning the Maillard conjugation.
Javad Tavakoli; Mohammad Hossein Hadad Khodaparast; Reza Esmaeilzadeh kenari; Mahmoud Aminlari; Ali Sharif
Abstract
In this study, antioxidant activity of kolkhung skin oil, baneh skin oil and seasame oil were compared. The ratio between polyunsaturated to saturated fatty acids (PUFA/SFA) of kolkhung skin oil, bane skin oil and seasame seed oil were 0.52, 0.26 and 3.06, respectively. Total phenolic contents of kolkhung ...
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In this study, antioxidant activity of kolkhung skin oil, baneh skin oil and seasame oil were compared. The ratio between polyunsaturated to saturated fatty acids (PUFA/SFA) of kolkhung skin oil, bane skin oil and seasame seed oil were 0.52, 0.26 and 3.06, respectively. Total phenolic contents of kolkhung skin oil, bane skin oil and seasame seed oil were 99.67, 645.73 and 81.72 mg/kg, respectively. Total tocopherol content of kolkhung skin oil (2154.3mg/kg) was significantly higher than those of the other investigated oils, followed by seasame seed oil and bane skin oil ( 993.69 and 648.91 mg/kg, respectively). Antioxidant activity of the three mentioned oils were determined by DPPH, FRAP and rancimat tests. Antioxidant activity of kolkhung skin oil was significantly higher than those of the other investigated oils, followed seasame seed oil and bane skin oil, respectively. Also it was found a correlation between amount of antioxidant compounds of oils and their antioxidant activity.
Nazanin Darabzadeh; Sahar Sadat Mousavi Nasab; Mahmoud Aminlari; Roghaieh Ramezani
Abstract
In recent years soybean derived foods (such as soymilk) have received considerable attention due to their high nutritional value and functional properties. The purpose of this study was to investigate the changes in protein solubility and electrophoretic behavior of soymilk stored at refrigerator and ...
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In recent years soybean derived foods (such as soymilk) have received considerable attention due to their high nutritional value and functional properties. The purpose of this study was to investigate the changes in protein solubility and electrophoretic behavior of soymilk stored at refrigerator and freezer conditions. A 12% total solid content soymilk was produced and stored at 4 and -18ºC. At different time intervals samples were removed an analyzed for protein solubility and SDS-PAGE. A gradual decrease in solubility of proteins stored at -18ºC with time was observed such that after 3 weeks solubility was dropped from 75% to 45% (30% decrease) and remained at 41% thereafter up to 6 weeks. These changes appeared to follow a first order kinetics with a rate constant (k) of 0.014(day)-1 and half life (t1/2) of 62 days. Similar pattern was seen in the refrigerated sample except that the decrease in solubility occurred slowly and at final stage, solubility decreased by 17%. At this temperature the decrease in protein solubility followed a zero order kinetics with k of 0.625 (day)-1 and t1/2 of 144 days These results indicate that long term refrigeration and freezing of soymilk must be avoided.Keywords: Soymilk, solubility, SDS-PAGE
