酶法与非酶法磷酸化改性食品蛋白质的研究进展

磷酸化是一种重要的蛋白质修饰改性手段,可以有效改善食物蛋白质的功能性质,如乳化性、溶解性、凝胶性、热稳定性等。蛋白质磷酸化改性方法主要可分为酶法与非酶法两种,其中酶法磷酸化常使用蛋白激酶作为磷酸基团的供体,对蛋白质进行修饰。主要的蛋白激酶包括环磷酸腺苷依赖蛋白激酶(CAMPdPK)和酪蛋白激酶Ⅱ(CK-Ⅱ)。非酶法磷酸化常见的改性试剂主要包括三氯氧磷、焦磷酸钠、三聚磷酸钠和葡萄糖-6-磷酸等。本文对近年来蛋白质磷酸化的有关报道进行了总结归纳,并通过介绍磷酸化蛋白的磷酸键性质、磷酸化肽段及位点的鉴定,阐述了不同磷酸化试剂的反应机制及其对蛋白质构效关系的影响,为通过定向的磷酸化修饰而改善蛋白质特定的功能性质提供了理论依据与参考。     

Improvement of functional properties of whey soy protein phosphorylated by dry-heating in the presence of pyrophosphate

The physicochemical and functional properties of a whey soyprotein (WSP), which was phosphorylated by dry heating, were investigated. The phosphorus content of WSP was 0.85 g/100 g by dry-heating at pH 4.0 for 5 days in the presence of pyrophosphate. ³¹P NMR spectral data suggested that the serine and sugar of WSP were phosphorylated. Although changes in the random coil structure of WSP measured by circular dichroism were mild, increased surface hydrophobicity and decreased tryptophan fluorescence intensity were observed following phosphorylation. Phosphorylation improved the stability against heat-induced insolubility, emulsifying properties, foaming properties, water absorption capacity, and oil absorption capacity of WSP. These results suggested that dry-heating phosphorylation is a useful method for improving the functional properties of WSP.     

白酒糟醇溶蛋白的磷酸化改性及功能性质

酒糟是酿酒产业的最大副产物。酒糟富含醇溶蛋白,然而由于水溶性差,限制其进一步应用。采用醇碱法从清香型白酒糟中提取醇溶蛋白(prolamin from distiller’s grains, PDG),用三聚磷酸钠对PDG进行磷酸化改性,通过溶解度、持水性、持油性、乳化性等方面的分析,研究磷酸化改性对PDG功能性质的影响。结果表明,醇碱法提取的白酒糟PDG主要由4种不同分子质量的蛋白组成,原材料酒糟粉粉碎度影响PDG的提取率,粒度小时有利于提高蛋白提取率,以200目细粉为原料时PDG的提取率可达22.69%±0.26%。差示扫描量热实验结果显示PDG的热变性温度为123.47℃,PDG具有良好的热稳定性。傅里叶红外光谱结果表明与三聚磷酸钠反应后磷酸基团修饰到PDG上。磷酸化改性改善了PDG在水中的溶解度,30℃时磷酸化后PDG的溶解度较PDG提高了3.53倍,达到943.08±16.04μg/mL。由于引入磷酸基团,PDG的功能性质显著改善,磷酸化改性后PDG较PDG的持水性和持油性分别增加了44.9%和33.8%(30℃),乳化性及乳化稳定性分别提高了54.9%和3.6倍(pH10)。...     

Physicochemical,nutritional and functional properties of marama bean flour

Marama bean is a protein-rich, underutilised, drought-tolerant legume in sub-Saharan Africa. Its utilisation may be increased through value addition into protein-rich flours. Defatted flour from unheated and dry-heated (150 °C/20 min) whole marama beans (UMF and HMF) were analysed for physicochemical and protein-based functional properties. Heating significantly increased in vitro protein digestibility of marama bean flour. Of functional importance are the high levels of tyrosine present in marama bean flours. Heating significantly reduced protein solubility and emulsifying capacity of marama bean flours whilst water absorption capacity was improved. Foaming capacity was not affected by heating. UMF could be useful in food systems requiring high emulsifying capacity, but would not be suitable for applications requiring high water absorption and foaming capacity. Due to its high protein contents, marama bean flour could be used to increase the protein quality of cereal-based foods to help alleviate protein-energy nutrition in the region.     

Functional properties and structural characteristics of phosphorylated pea protein isolate

In order to expand the application of pea proteins in the food industry, pea protein isolate (PPI) was chemically phosphorylated to improve its functional properties. Based on the comprehensive membership value (CMV) degree, the optimal condition for PPI phosphorylation modification was as follows: pH 12, modification temperature at 70 °C and an addition of 7.0% (w/v) sodium tripolyphosphate (STP). Under this condition, the solubility, emulsifying property, emulsifying stability, foaming property and oil absorption capacity of the modified PPI were improved substantially. In addition, the structure of modified PPI was characterised by scanning electron microscope (SEM) and Fourier transform infrared (FTIR) spectroscopy. The results showed that the modified PPI had a smooth and uniform lamellar structure, where the content of α-helix and β-sheet structure increased, but the content of β-corner and random coil structure decreased. The thermodynamic properties were analysed by differential scanning calorimetry (DSC) and the results showed that ∆H of the modified PPI increased significantly. Finally, the optimum phosphorylated PPI was mixed with 0.4% (w/v) xanthan gum to form PPI fat mimics (PPI-FM). PPI-FM was added into mango mousse cake to reduce the amount of light cream, and the result showed up to 20% of light cream could be substituted.     

The phosphorylation and characterisation of soybean isolate and its β‐conglycinin component by casein kinase II

A commercial soybean isolate was phosphorylated using casein kinase II purified from the yeast Yarrowia lipolytica. Both major reserve proteins, β‐conglycinin and glycinin, were phosphorylated in a sequential way. The soybean isolate incorporated up to 0.7 mol phosphate per mole in 2 h. It was found that the phosphoester bonds were stable over time. The solubility of the phosphorylated isolate with respect to pH was not dramatically increased in comparison with the native one. However, counting the radioactivity of ³²P incorporated into the proteins (only the solubility of the phosphorylated proteins was measured in this case) showed that the solubility of the proteins was dramatically improved (up to 90% solubility for phosphorylated β‐conglycinin at pH 4). β‐Conglycinin became more soluble in the presence of CaCl₂ upon phosphorylation; this was not the case for the isolate. The iron‐binding capacity of the soy isolate and β‐conglycinin was significantly improved after phosphorylation (two and six times respectively). © 1999 Society of Chemical Industry     

Effect of pH on phosphorylation of potato protein isolate

Potato protein isolate (PPI) was phosphorylated with sodium trimetaphosphate (STMP) at ambient temperature and various reaction pH (5.2, 6.2, 8.0 and 10.5) to improve the functional properties without impairing the nutritional availability. Changes in chemical composition (total and coagulable protein content, ash and minerals content and amino acid composition), functional properties (protein solubility index, emulsifying activity and foaming capacity, water and oil absorption capacity) and phosphorus were determined. The chemical composition and functional properties of phosphorylated potato protein isolate (PP-PPI) were significantly different (p < 0.05). The PP-PPI at pH 5.2 was characterised by the highest content of all amino acids, whereas, PP-PPI under alkaline conditions (pH 10.5) caused decrease in these compounds. PP-PPI at pH 8.0 had the highest oil absorption capacity, emulsion activity and foam capacity, whereas, PP-PPI at pH 10.5 had the highest WAC.     

Improvement of functional properties of ovalbumin phosphorylated by dry-heating in the presence of pyrophosphate

The structural and functional properties of a ovalbumin (OVA), which was phosphorylated by dry-heating, were investigated. The phosphorus content of OVA increased to 0.86 % by dry-heating at 85 °C and at pH 4.0 for 3 days in the presence of pyrophosphate. ³¹P NMR spectral data indicated that phosphoesters, polyphosphate bonds and/or phosphodiester were introduced into phosphorylated ovalbumin (PP-OVA). The CD spectrum of native ovalbumin (N-OVA) was not changed by dry-heating. However, the α-helix content decreased in the presence of pyrophosphate, which was down from 31.2 % in DH-OVA to 27.8 % in PP-OVA. Increased surface hydrophobicity and decreased tryptophan fluorescence intensity were observed following phosphorylation. Phosphorylation improved heat-induced insolubility, emulsifying properties, water- and oil-binding capacity of OVA. Furthermore, the calcium phosphate-solubilizing ability of OVA was enhanced. Thus, PP-OVA may be expected to enhance the absorption of calcium. These results indicated that dry-heating phosphorylation is a useful method for improving the functional properties of OVA.     

Granular Properties of Different Starch Phosphate Monoesters

Different starch types (corn, rice, and potato starch, corn amylose and corn amylopectin) were phosphorylated by reaction with a mixture of mono and disodium phosphate at different molar ratios (mol phosphate/mol anhydrous glucose) under heat and vacuum. The starch granules of the modified and the native starches were microscopically examined for their sizes and morphology. The correlation between the variation in granular size of the modified starches with the extent of phosphorylation and some other physicochemical properties was studied. The granular size was generally increased while the iodine absorption capacity was decreased by phosphorylation. There were strong correlations between the variation in the starch granular size in dependence on phosphorylation and the corresponding changes in some physicochemical parameter of starch, e.g. solubility, swelling and paste clarity. This relationship was most evident in the case of phosphorylated corn amylopectin. Starch granular size can be taken as a quick indicator of the physicochemical properties of the native and modified starches.     

Improvement of antioxidant activity of egg white protein by phosphorylation and conjugation of epigallocatechin gallate

Egg white protein (EWP) was phosphorylated by dry heating in the presence of pyrophosphate. The antioxidant activity of phosphorylated EWP (PP-EWP) was then investigated before and after conjugation of epigallocatechin gallate (EGCG). The binding interaction between EWPs and EGCG was investigated by UV and FTIR, which indicated that more EGCG bound to PP-EWP than to native EWP and dry-heated EWP. The antioxidant activities including ABTS⁺ free-radical scavenging capacity, oxygen radical antioxidant capacity, reducing power, chelating capacity, and superoxide anion scavenging activity of EWP were remarkably improved by phosphorylation before and after conjugation of EGCG. The improved antioxidant property of PP-EWP can be attributed to the introduced phosphate group, more exposed hydrophobic group, increased surface thiol group of protein, and more EGCG bound to EWP. The secondary structure of PP-EWP decreased with more bound EGCG. This study is the first to describe an improvement in the antioxidant property of EWP by phosphorylation and conjugation of EGCG, thereby providing insight into the design of food protein antioxidant.     

Functional properties of protein isolates,globulin and albumin extracted from Ginkgo biloba seeds

In this study, the functional properties of Ginkgo seed protein isolate (GPI), Ginkgo seed globulin protein (GGP) and Ginkgo seed albumin protein (GAP) extracted from Ginkgo biloba seeds were investigated. The protein contents of GPI, GGP and GAP were 91.0%, 93.4% and 87.8%, respectively in the samples in which the sugar, polyphenol and crude fibre were removed by the preparation procedure. For functional properties of Ginkgo seed proteins in the natural state, GAP showed the highest oil-absorption capacity (9.3 ml/g), foaming capacity (67.8%), emulsifying capacity (65.4%) and emulsion stability (90.6%); while GPI showed the highest water absorption capacity (1.93 ml/g), and GGP showed the highest foam stability (55.5%). The differences of the chemical components, surface hydrophobicity, disulphide bond (SS) and sulfhydryl group (SH) contents of GPI, GGP and GAP, which were correlated significantly with functional properties of Ginkgo seed proteins, were also investigated. The improved functional properties, such as water absorption capacity, solubility, foaming properties and emulsifying properties of Ginkgo seed proteins were observed in a pH range of 8.0–10.0 or sodium chloride concentration of 0.5–0.75 M.     

High-pressure homogenization treatment of red seaweed Bangia fusco-purpurea affects the physicochemical,functional properties and enhances in vitro anti-glycation activity of its dietary fibers

In this study, high-pressure homogenization (HPH) technique was applied for the pretreatment of Bangia fusco-purpurea, and the effect of HPH on the composition, physiochemical and functional properties, and in vitro anti-glycation activity of dietary fiber from this seaweed (B. fusco-purpurea dietary fiber, BDF) was studied. Results showed that HPH significantly increased water-soluble dietary fiber (SDF) content in BDF. Water-holding capacity, oil-holding capacity, and glucose delay dialysis index of BDF were significantly enhanced after HPH treatment. Additionally, HPH significantly improved the in vitro anti-glycation activity of BDF by inhibiting the formation of advanced glycation end products (AGEs) and mitigating damage induced by AGEs on intestinal cells. These improvements could be attributed to the formation of the coarse and porous structure and greater exposure of hydroxyl groups of BDF caused by HPH treatment. These results implied the potential of HPH in seaweed processing and provided a scientific basis for the in-depth, comprehensive utilization of B. fusco-purpurea.Industrial relevanceHPH is an emerging non-thermal food processing technique with promising application potential in food industry. In this study, we found that HPH technique could significantly change the composition, improve physiochemical and functional properties and enhance anti-glycation activity of dietary fiber from seaweed B. fusco-purpurea. Our results validated the efficiency of HPH on dietary fiber modification, implying the potential of HPH in food industry and healthy industry.     

Bioactive peptides derived from bovine whey proteins: opioid and ace-inhibitory peptides

Regulatory peptides can be released by enzymatic proteolysis of food proteins and may act as potential physiological modulators of metabolism during the intestinal digestion of the diet. The possible regulatory effects of peptides relate to nutrient uptake, immune defence, opioid and antihypertensive activities. Milk proteins, especially caseins, are an important source of these bioactive peptides. During recent years, major whey protein components, α-lactalbumin and β-lactoglobulin, were also shown to contain bioactive sequences. Peptides showing opioid and angiotensin I-converting enzyme (ACE) inhibitory activity were found in α-lactalbumin and β-lactoglobulin. Opioid peptides, α-lactorphin and β-lactorphin, were liberated during in vitro proteolysis of bovine whey proteins, and pharmacological activity was observed at micromolar concentrations. Whey hydrolysates showed ACE-inhibitory activity after proteolysis with different digestive enzymes, and several active peptides were identified. The results demonstrated the existence of several biologically active whey-derived peptides and hydrolysates. The findings of the study can be exploited in the development of foods with special health claims (e.g. treatment of hypertension) as well as in identifying new applications in food.     

Enhancement of Functional Properties of Rice Bran Proteins by High Pressure Treatment and Their Correlation with Surface Hydrophobicity

Changes in functional properties of rice bran proteins as influenced by high-pressure (HP) treatment (100–500 MPa, 10 min) were studied. Properties evaluated were protein solubility, water absorption capacity, oil absorption capacity, foaming capacity, foam stability, emulsifying activity, emulsion stability, least gelation concentration, and surface hydrophobicity. HP treatment at 100 and 200 MPa significantly improved the solubility and oil absorption capacity, while water absorption and foaming capacities increased further reaching the maximum at 500 MPa. Compared with the untreated control sample, the emulsifying activity and foam stability of treated samples were significantly higher and least gelation concentration was lower, but none of them showed any specific trend with pressure level. Emulsion stability and surface hydrophobicity increased with the pressure level until 400 MPa and decreased slightly at 500 MPa. Pearson correlation coefficients clearly showed that surface hydrophobicity was positively correlated with water absorption capacity, foaming capacity, emulsifying activity index, and emulsion stability index, but negatively correlated with least gelation concentration. The pressure treated rice bran protein possessed good functional properties for use as a food ingredient in the formulations.     

Bioactivity of β-lactoglobulin and α-lactalbumin—Technological implications for processing

The dairy industry faces new technological challenges in order to exploit and maintain some of the bioactive properties of dairy components throughout processing. This review outlines these issues with respect to the two major whey proteins β-lactoglobulin (β-lg) and α-lactalbumin (α-la). Biological activities of both the intact proteins, and peptides derived from the proteins, are discussed, e.g. inhibition of angiotensin-converting enzyme (ACE), anti-microbial activity, anti-carcinogenic activity, hypocholesterolemic effect, metabolic and physiological effects. The levels necessary to provide beneficial effects and, if available, evidence from clinical trials are reported. Developments in the purification and enrichment of the proteins are discussed, and the technological implications of industrial processing on the bio-activity of the proteins are examined. The supplementation of infant formulas with α-la enriched whey proteins is also discussed in light of its potentially improved bioactive properties.     

Improvement of functional properties of bovine serum albumin through phosphorylation by dry-heating in the presence of pyrophosphate

ABSTRACT:  Bovine serum albumin (BSA) was phosphorylated by 2 methods. One is dry-heating in the presence of pyrophosphate, and the other is conjugation with maltopentaose through the Maillard reaction and subsequent dry-heating in the presence of pyrophosphate. The phosphorus content of BSA was increased to approximately 0.45% by dry-heating at pH 4.0 and 85 °C for 5 d in the presence of pyrophosphate, and approximately 0.91% by glycation and subsequent phosphorylation. The circular dichroism spectra showed that the change of secondary structure in the BSA molecule by phosphorylation was mild. However, tryptophan fluorescence intensity of BSA decreased by phosphorylation. The differential scanning calorimetry thermograms of BSA showed a disappearing of the 1st peak and a lowering of the 2nd peak denaturation temperature by phosphorylation. These results indicated molten (partially unfolded) conformations of BSA formed by both phosphorylation methods. The functional properties of BSA such as heat stability and calcium phosphate solubilizing ability were improved by phosphorylation alone and further by phosphorylation after glycation. Transparent gels of BSA with relatively high water-holding capacity were obtained by phosphorylation alone, and the immunogenicity of BSA was reduced significantly by glycation and phosphorylation, respectively.     

Effects of dielectric barrier discharge plasma on the physicochemical and functional properties of myofibrillar proteins

The present study was designed to investigate the effects of dielectric barrier discharge (DBD) plasma on myofibrillar proteins extracted from beef. For this purpose, myofibrillar proteins were extracted from Longissimus dorsi muscle and treated with DBD plasma for 0, 5, 10, 15, and 20 min to evaluate free sulfhydryl and carbonyl, FTIR, and DSC profiles as well as pH, color, foaming capacity and stability, emulsion capacity and stability, and water binding capacity of myofibrillar proteins. Evaluation of the chemical changes revealed that protein oxidation was the main event occurring during exposure to DBD plasma. Moreover, mild oxidation during the first 10 min of exposure was found to improve the functional properties of myofibrillar proteins such that significant (p < 0.05) increases were observed not only in foaming capacity but also in emulsion capacity and stability. However, the detrimental effects of severe oxidation on functional properties due to long-time exposure cannot be ignored. This is while long-time exposure was observed only to lead to significantly (p < 0.05) improved water binding capacity (WBC) (rising it from ~25% in the control treatment to ~92% in the 20-min treatment). The findings of this study can be beneficially exploited by the food industry toward appropriate treatment of myofibrillar proteins with non-thermal plasma.     

Improvement of functional properties of whey protein isolate through glycation and phosphorylation by dry heating

Whey protein isolate (WPI) was glycated with maltopentaose (MP) through the Maillard reaction, and the MP-conjugated WPI (MP-WPI) was then phosphorylated by dry heating in the presence of pyrophosphate. Glycation occurred efficiently, and the sugar content of WPI increased approximately 19.9% through the Maillard reaction. The phosphorylation of MP-WPI was enhanced with an increase in the dry-heating time from 1 to 5 d, and the phosphorus content of WPI increased approximately 1.05% by dry heating at pH 4.0 and 85°C for 5 d in the presence of pyrophosphate. The electrophoretic mobility of WPI increased with an increase in the phosphorylation level. The stability of WPI against heat-induced insolubility at pH 7.0 was improved by conjugation with MP alone, and further improved by phosphorylation. Although the emulsifying activity of WPI was barely affected by glycation and phosphorylation, the emulsifying stability of phosphorylated MP-WPI (5 d), was 2.2 times higher than that of MP-WPI. Gelling properties such as hardness, resiliency, and water-holding capacity of heat-induced WPI gel were markedly improved, and the gel was rendered transparent by phosphorylation. The calcium phosphate-solubilizing ability of WPI was enhanced by phosphorylation. These results suggested that phosphorylation by dry heating in the presence of pyrophosphate after conjugation with MP is a useful method for improving the functional properties of WPI.     

Functional properties of isolated porcine blood proteins modified by Maillard’s reaction

Plasma proteins (albumin and immunoglobulins) and haemoglobin from porcine blood can be recovered from slaughterhouse waste. These proteins are employed as ingredients in food products on account of their functional properties. There are different methods to improve these properties, Maillard’s reaction probably being the most promising technique for food purposes. In the present study, 10 kDa dextran was employed to produce the conjugates. Three reaction temperatures were assayed, 80 °C for 60 min being found to be the most suitable to produce conjugates with enhanced functional properties. Both thermal stability and emulsification capacity were improved; gelling temperature was increased 15 °C; and gel strength was lowered 50% compared to native proteins. However, solubility decreased slightly. It has been demonstrated that the functional properties of blood proteins are enhanced through conjugation, showing a improvement in the application of the blood proteins in food products.     

Phosphorylation of starch and dextrin by dry‐heating in the presence of phosphate,and their calcium phosphate‐solubilizing ability

Starch and dextrin were phosphorylated by dry‐heating in the presence of phosphate, and their properties were examined. The phosphorylation of starch was accelerated with decreasing moisture, rising incubation temperature and prolongation of the incubation period. However, a rise in incubation temperature, and lengthened incubation period caused increased browning and degradation. A decrease in the pH from 5.5 to 3.0 resulted in a modest increase of phosphorylation, but also marked browning and degradation. When potato starch was phosphorylated at 140°C and pH 5.5 for 24 h, its phosphorus content was increased up to 3.47%, with fewer side reactions. Phosphorlylated starch and dextrin had calcium phosphate‐solubilizing abitity. Phosphorylated dextrin with 2.42% phosphorus had about half the calcium phosphate‐solubilizing ability of casein phosphopeptide (CPP). The possibility of substituting of phosphorylated starch and dextrin for CPP as a calcium phosphate‐absorption enhancing material is discussed herein.