酶解膜分离两步分离乳清中β-乳球蛋白的研究

为研究乳清中β-乳球蛋白环保、温和、高效的分离工艺,该文对采用蛋白酶选择性水解然后超滤分离乳清水解液中的β-乳球蛋白进行研究.试验对乳清水解物进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)分析,比较了胃蛋白酶、胰蛋白酶、木瓜蛋白酶和中性蛋白酶的选择性,并分别采用分子截留量10000的聚砜膜(PS-10)和聚醚膜(PES-10)对胃蛋白酶解液进行超滤处理.结果显示,β-乳球蛋白对胃蛋白酶耐受性较好,α-乳白蛋白对木瓜蛋白酶抗性较佳,但二者均对中性蛋白酶较敏感.较优的水解分离条件如下:乳清蛋白质量浓度8%,胃蛋白酶添加量为乳蛋白质量的0.3%,pH值2.1,温度37℃,水解2h,α-乳白蛋白近似完全水解而β-乳球蛋白几乎不被降解.水解液用PS-10膜超滤分离,β-乳球蛋白的纯度和产率达到了较高,分别为94.6%,75.6%.因此,采用选择性酶解处理乳清然后超滤分离β-乳球蛋白是可行的.     

酶解膜分离两步分离乳清中β-乳球蛋白的研究(简报)

为研究乳清中β-乳球蛋白环保、温和、高效的分离工艺,该文对采用蛋白酶选择性水解然后超滤分离乳清水解液中的β-乳球蛋白进行研究。试验对乳清水解物进行十二烷基硫酸钠－聚丙烯酰胺凝胶电泳（SDS-PAGE）分析,比较了胃蛋白酶、胰蛋白酶、木瓜蛋白酶和中性蛋白酶的选择性,并分别采用分子截留量10000的聚砜膜(PS-10)和聚醚膜(PES-10)对胃蛋白酶解液进行超滤处理。结果显示,β-乳球蛋白对胃蛋白酶耐受性较好,α-乳白蛋白对木瓜蛋白酶抗性较佳,但二者均对中性蛋白酶较敏感。较优的水解分离条件如下：乳清蛋白质量浓度8%, 胃蛋白酶添加量为乳蛋白质量的0.3%,pH值2.1,温度37℃,水解2 h,α-乳白蛋白近似完全水解而β-乳球蛋白几乎不被降解。水解液用PS-10膜超滤分离,β-乳球蛋白的纯度和产率达到了较高,分别为94.6%,75.6%。因此,采用选择性酶解处理乳清然后超滤分离β-乳球蛋白是可行的。     

超高压对含有琼脂猪肉凝胶特性影响的试验

多糖作为脂肪替代物可用于开发低脂肉制品,而超高压可对低脂肌肉凝胶产生改性作用。该研究将超高压引入含琼脂0.4%猪肉糜凝胶(PMGA)的加工,在压力100～600 MPa、保压时间10～40 min、加压介质温度10～40℃范围内,单因素试验考察各因素对PMGA持水性、色泽与硬度的影响。研究结果表明:与未受压的对照组相比,200～400 MPa压力导致PMGA的蒸煮损失率显著降低(p<0.05);100～600 MPa压力可不同程度地提高PMGA的保水性,并显著降低彩度指数b*值(p<0.05);400～600 MPa压力引起PMGA亮度L*值显著增加和彩度指数a*值明显下降(p<0.05);100～400 MPa压力可显著提高PMGA的硬度,而500～600 MPa压力则显著降低其硬度(p<0.05);保压时间与介质温度对PMGA持水性、色泽与硬度的影响相对有限。因此,200～400 MPa的超高压处理可获得较高的产品出品率;且改变工作压力可调控受压PMGA的硬度。     

体外模拟淡水鱼小清蛋白对胃肠消化的影响

大多数食物过敏原的理化性质决定了蛋白质在消化道中的稳定性.模拟胃液(simulated gastric fluid,SGF)的稳定性是评估食物过敏的一个重要参数.为了解一种主要的鱼类过敏原小清蛋白(parvalbumin,PV)与来自鲤鱼(Cyprinus carpio)和鲢鱼(Hypophthalmichthys molhrix)肌肉中的非致敏性蛋白在SGF和模拟肠液(simulated intestinal fluid,SIF)中的稳定性差异,本研究采用3种蛋白水解酶(胃蛋白酶、胰蛋白酶和来自猪的胰凝乳蛋白酶)来模拟人(Homo sapiens)体肠道的消化蛋白酶;结合两次三氯乙酸沉淀法和凝胶过滤层析法纯化PV,通过Tricine-十二烷基硫酸钠聚丙烯酰胺凝胶电泳(Tricine-sodium dodecyl sulfate polyacrylamide gel electrophoresis,Tricine-SDS-PAGE)和Western blot对3种蛋白在模拟肠胃液中的稳定性进行了评估.结果显示,鲤鱼和鲢鱼的PV都获得了分子量约10kD的蛋白,同时在SGF中鲤鱼和鲢鱼的纯化PV也获得了相同的分子量蛋白.胃蛋白酶使原先的PV带几乎在60 min内完全降解,并观察到一些稳定的肽片段,而胰蛋白酶和胰凝乳蛋白酶在240 min后都无法有效降解PV.在SGF中的非致敏性鱼浆蛋白在很短的时间内迅速降解,而PV的消化时间延长.Western blot分析表明,抗鲢鱼PV多克隆抗体可以特异性地检测到PV及其降解产物.PV比非致敏性蛋白更能抵抗蛋白酶消化,相比胰蛋白酶和胰凝乳蛋白酶,胃蛋白酶处理能更有效地减少超敏反应.研究结果为未来低致敏性鱼产品的开发提供了理论参考.     

燕麦麸分离蛋白的酶解对其功能性质的影响

为了改善燕麦蛋白的功能性质以扩大其在食品工业中的应用,该文以燕麦麸为原料制备了燕麦麸分离蛋白(OBPI),并利用胰蛋白酶对其进行水解,得到了3种不同水解度(4.1%、6.4%、8.3%)的酶解产物。SDS-PAGE分析结果表明OBPI中的主要蛋白成分是球蛋白,其经过胰蛋白酶处理后,球蛋白酸性亚基被部分水解而碱性亚基相对保持完整。胰蛋白酶水解显著改变了OBPI的功能性质。在所考察的水解度范围内,随着水解度的升高,酶解产物的溶解性、持水性、乳化活性及起泡能力等方面均逐渐增加;但持油性、乳化及泡沫稳定性有不同程度的降低。     

免疫印迹分析试验感染肝片吸虫和大片吸虫绵羊的IgG应答

免疫印迹(Westernblot)分析试验感染肝片吸虫和大片吸虫绵羊的抗片形吸虫分泌捧泄产物(FhESP和FgESP)IgG应答，结果表明，FhESP中分子质量在17．1～21．6ku之间的4条主要蛋白带仅被肝片吸虫感染以后(0WPI以后)的血清识别且不被对照组血清识别；FgESP中分子质量在19．0～71．1ku之间的6条主要蛋白带仅被大片吸虫感染以后(0WPI以后)的血清识别且不被对照组血清识别。     

超声处理对牦牛乳酶促凝胶流变特性的影响研究

为探究超声牦牛乳的酶促凝胶流变特性,并进一步分析超声后牦牛乳的蛋白疏水性和脂肪球粒径如何影响凝胶网络的形成,本研究以牦牛乳为原料,进行超声处理(时间5 min和10 min,功率300、400和500 W)后,编号为US1(300 W,5 min)、US2(300 W,10 min)、US3(400 W,5 min)、US4(400 W,10 min)、US5(500 W,5 min)和US6(500 W,10 min),以未处理的牦牛乳作为对照(CK),通过小振幅振荡时间扫描获取凝胶形成的流变信息。结果表明,超声处理后牦牛乳的脂肪含量、蛋白质含量以及pH值无显著变化(P>0.05),而均质效率和乳液稳定性显著提高(P<0.05);超声处理能够增加凝胶的储能模量,使得牦牛乳酶促凝胶的弹性增加。US5和US6的储能模量增幅最多(分别为6 824和7 189 Pa),但凝胶时间长。与CK相比,超声牦牛乳US1、US2、US3和US4的凝胶时间分别缩短了1.78、3.72、5.75和4.74 min;超声处理在改变乳蛋白表面疏水性的同时减小了脂肪球尺寸。与CK相比,US3最大荧光强度显著增加了51.22%(P<0.05),脂肪球粒径分布范围为0.300~4.002 μm,集中在1.974 μm处。US4最大荧光强度显著降低了5.31%(P<0.05),脂肪球粒径呈双峰分布,范围为0.131~4.502 μm。综上所述,经过不同时间和不同功率的超声处理后,US1~US6乳蛋白表面疏水性和脂肪球尺寸受到影响的程度不同,使得凝胶网络体系存在差异,从而表现出不同的凝胶特性。本研究为超声技术在牦牛乳凝胶类制品加工中的应用提供了理论基础。     

体外消化过程中低分子量寡肽释放量与饲料品质的关系

应用胃蛋白酶．胰蛋白酶,模拟鸡胃肠道消化环境,对4种饲料(鱼粉、豆粕、棉粕和菜粕)进行体外消化,研究体外消化过程中不同饲料的寡肽释放特点与其氨基酸组成的关系。4种饲料的体外酶水解产物分别经过截留分子量为3kD和1kD的中空纤维超滤组件处理,制备8种低分子量组分。测定各组分中总氨基酸(TAA)和游离氨基酸(FAA)含量,RP-HPLC法分析各组分中典型寡肽。结果表明,低分子量组分中寡肽含量(PAA/TAA)与饲料必需氨基酸(TEAA)含量之间存在显著正相关,TEAA含量高的优质饲料蛋白在体外消化过程中产生较多寡肽。     

传统风干牦牛肉加工中肌原纤维蛋白氧化与体外消化性变化

为了评价传统风干牦牛肉在加工过程中肌原纤维蛋白消化率的变化,并探讨蛋白质氧化影响其消化性的潜在机制。在牦牛肉自然风干加工过程中(40 d)采集样本,测定肌原纤维蛋白羰基、巯基、二硫键、二聚酪氨酸、粒径分布、体外消化率及脂质氧化等指标,并分析脂质氧化和蛋白质氧化对蛋白质消化性的影响。结果显示,脂质初级氧化产物(Primary Oxidation Value, POV)、硫代巴比妥酸反应物(Thiobarbituric Acid Reaction Substrates, TBARS)和蛋白质羰基化合物随加工时间的延长而显著增加(P0.05),并且相互间存在极显著相关性(P0.01)。此外,二硫键和二聚酪氨酸含量显著增加(P0.05),这是蛋白质交联的主要形式。蛋白质被胃蛋白酶和胰蛋白酶总的水解率在整个加工过程中降低了19.58%(P0.05),蛋白质的水解率与羰基、二硫键、二聚酪氨酸和蛋白质粒径呈现极显著负相关性(P0.01)。综上所述,在风干牛肉加工期间,脂质氧化产物促进了蛋白质氧化,蛋白质的羰基化和交联导致蛋白酶对蛋白质的水解率降低。研究结果对了解风干牦牛肉传统加工方式和进一步改进加工工艺具有积极的促进作用。     

喷射蒸煮技术在制备易消化玉米浓缩蛋白中的应用

为了提供一种具有良好消化性的玉米蛋白,该研究以玉米黄粉(corn gluten meal,CGM)为原料,利用亚临界脱脂及酶解超滤技术制备了一种蛋白纯度较高的玉米浓缩蛋白(corn protein concentrates,CPC),并重点考察了制备过程中喷射蒸煮对玉米浓缩蛋白功能性质及消化性的影响。研究结果表明,经喷射蒸煮处理后的玉米浓缩蛋白(jet cooking corn protein concentrates,JC-CPC),普通高温处理的玉米浓缩蛋白(heat treatment corn protein concentrates,HT-CPC)以及未经高温处理的玉米浓缩蛋白(CPC),三者在蛋白质质量分数及氨基酸组成上无明显差异(P0.05)。但JC-CPC的溶解性及功能性质(持水性、起泡性及泡沫稳定性)均显著高于CPC和HT-CPC(P0.05)。体外模拟消化试验结果表明,JC-CPC的水解度(24.02%±0.49%)明显高于CPC和HT-CPC(分别为9.23%±0.45%和14.52%±1.26%)(P0.05),其可溶性氮释放量(62.05%±0.75%)亦高于HT-CPC(40.25%±0.19%)、JC-CPC(21.02%±0.72%)(P0.05)。同时,JC-CPC消化产物的抗氧化试验结果表明,其消化产物具有较高的还原力及1,1-二苯基-2-苦基肼(1,1-Diphenyl-2-picrylhydrazyl,DPPH)自由基的清除能力。因此,利用喷射蒸煮技术,结合亚临界及超滤除杂技术能够为食品工业提供一种具有良好消化性的玉米浓缩蛋白,有望为玉米黄粉的利用提供一条有效途径。     

Cross-linking and rheological changes of whey proteins treated with microbial transglutaminase

Modification of the functionality of whey proteins using microbial transglutaminase (TGase) has been the subject of recent studies. However, changes in rheological properties of whey proteins as affected by extensive cross-linking with TGase are not well studied. The factors affecting cross-linking of whey protein isolate (WPI) using both soluble and immobilized TGase were examined, and the rheological properties of the modified proteins were characterized. The enzyme was immobilized on aminopropyl glass beads (CPG-3000) by selective adsorption of the biotinylated enzyme on avidin that had been previously immobilized. WPI (4 and 8% w/w) in deionized water, pH 7.5, containing 10 mM dithiothreitol was cross-linked using enzyme/substrate ratios of 0.12-10 units of activity/g WPI. The reaction was carried out in a jacketed bioreactor for 8 h at 40 degrees C with continuous circulation. The gel point temperature of WPI solutions treated with 0.12 unit of immobilized TGase/g was slightly decreased, but the gel strength was unaffected. However, increasing the enzyme/substrate ratio resulted in extensive cross-linking of WPI that was manifested by increases in apparent viscosity and changes in the gelation properties. For example, using 10 units of soluble TGase/g resulted in extensive cross-linking of alpha-lactalbumin and beta-lactoglobulin in WPI, as evidenced by SDS-PAGE and Western blotting results. Interestingly, the gelling point of WPI solutions increased from 68 to 94 degrees C after a 4-h reaction, and the gel strength was drastically decreased (lower storage modulus, G'). Thus, extensive intra- and interchain cross-linking probably caused formation of polymers that were too large for effective network development. These results suggest that a process could be developed to produce heat-stable whey proteins for various food applications.     

Protein-stabilized nanoemulsions and emulsions: comparison of physicochemical stability, lipid oxidation, and lipase digestibility

The properties of whey protein isolate (WPI) stabilized oil-in-water (O/W) nanoemulsions (d(43) ≈ 66 nm; 0.5% oil, 0.9% WPI) and emulsions (d(43) ≈ 325 nm; 0.5% oil, 0.045% WPI) were compared. Emulsions were prepared by high-pressure homogenization, while nanoemulsions were prepared by high-pressure homogenization and solvent (ethyl acetate) evaporation. The effects of pH, ionic strength (0-500 mM NaCl), thermal treatment (30-90 °C), and freezing/thawing on the stability and properties of the nanoemulsions and emulsions were compared. In general, nanoemulsions had better stability to droplet aggregation and creaming than emulsions. The nanoemulsions were unstable to droplet flocculation near the isoelectric point of WPI but remained stable at higher or lower pH values. In addition, the nanoemulsions were stable to salt addition, thermal treatment, and freezing/thawing (pH 7). Lipid oxidation was faster in nanoemulsions than emulsions, which was attributed to the increased surface area. Lipase digestibility of lipids was slower in nanoemulsions than emulsions, which was attributed to changes in interfacial structure and protein content. These results have important consequences for the design and utilization of food-grade nanoemulsions.     

Uniaxial compression of thermal gels based on microfluidized blends of WPI and heat-denatured WPI.

The effect of heat-denatured whey protein isolate (dWPI)/whey protein isolate (WPI) ratio (0-0.6), microfluidization pressure (0-1000 bar), and number of passes (1-10) on the uniaxial shear stress at 10% (sigma(10)) and 80% (sigma(80)) relative deformation of dWPI/WPI heat-induced gels (14% total protein, w/w) was studied. No correlation between the average diameter of aggregates and the dWPI/WPI ratio, microfluidization pressure, or number of passes was found. However, increasing the microfluidization pressure or the number of passes resulted in a narrower size distribution of aggregates. Increasing the dWPI/WPI ratio and the number of passes resulted in a decrease and an increase of gel hardness, respectively. The results were interpreted in terms of more random aggregation/gelation of proteins in the presence of aggregates that could result in localized heterogeneities into gels and more dissipation of the deformation energy during compression. The positive effect of the number of passes on the gel hardness was also considered to be due to a more homogeneous aggregation/gelation of proteins in the presence of smaller aggregates.     

Formation of whey protein isolate (WPI)-dextran conjugates in aqueous solutions

The conjugation reaction between whey protein isolate (WPI) and dextran in aqueous solutions via the initial stage of the Maillard reaction was studied. The covalent attachment of dextran to WPI was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with both protein and carbohydrate staining. The formation of WPI-dextran conjugates was monitored by a maximum absorbance peak at approximately 304 nm using difference UV spectroscopy. The impact of various processing conditions on the formation of WPI-dextran conjugates was investigated. The conjugation reaction was promoted by raising the temperature from 40 to 60 degrees C, the WPI concentration from 2.5 to 10%, and the dextran concentration from 10 to 30% and lowering the pH from 8.5 to 6.5. The optimal conjugation conditions chosen from the experiments were 10% WPI-30% dextran and pH 6.5 at 60 degrees C for 24 h. WPI-dextran conjugates were stable under the conditions studied.     

自由基引起的氧化对牛乳清蛋白凝胶特性的影响

为了深入了解蛋白氧化对凝胶特性的影响,以此探讨乳清蛋白氧化对其功能性质的影响机制,该文主要研究了氧化对乳清蛋白凝胶质地、流变学特性和微观结构变化的影响。试验采用羟基自由基氧化体系,在不同H2O2浓度(1～20mmol/L)及不同FeCl3浓度(0.1～1mmol/L)对乳清蛋白分别氧化3h,通过质构仪、流变仪和扫描电镜对凝胶特性和微观结构进行研究。结果显示:同未氧化乳清蛋白相比,在所有氧化条件下,凝胶硬度降低了90%以上,贮藏模量(G')值降低了17%以上,复合模量(G*)值降低了20%以上;高浓度氧化条件下,弹性降低了20%以上。氧化明显改变了凝胶的微观结构,随着氧化剂的加入,导致了疏松多孔且不规则凝胶的形成。上述结果表明,氧化对蛋白凝胶质地和凝胶形成能力起着很大的破坏作用,并影响着其微观结构。     

Heat-induced soy-whey proteins interactions: formation of soluble and insoluble protein complexes

The aggregation behavior during heating of a solution containing soy protein and whey protein isolate (WPI) was studied using rheology, confocal microscopy, gel filtration, and electrophoresis. Soy/WPI mixtures formed gels at 6% total protein concentration with a high elastic modulus (G') and no apparent phase separation. The ratio of soy to WPI was fundamental in determining the type of network formed. Systems containing a high soy to WPI ratio (>70% soy protein) showed a different evolution of the elastic modulus during heat treatment, with two apparent stages of network development. Whey proteins formed disulfide bridges with soy proteins during heating, and at low ratios of soy/WPI, the aggregates seemed to be predominantly formed by 7S, the basic subunits of 11S and beta-lactoglobulin. Size exclusion chromatography indicated the presence of high molecular weight soluble complexes in mixtures containing high soy/WPI ratios. Results presented are the first evidence of interactions between soy proteins and whey proteins and show the potential for the creation of a new group of functional ingredients.     

Effects of sugars on whey protein isolate gelation

Whey protein isolate (WPI) gels were prepared from solutions containing ribose or lactose at pH values ranging from 6 to 9. The gels with added lactose had no color development, whereas the gels with added ribose were orange/brown. Lactose stabilized the WPI to denaturation, which increased the time and temperature required for gelation, thus decreasing the fracture modulus of the gel compared to the gels with added ribose and the gels with no sugar added. Ribose, however, favored the Maillard reaction and covalent cross-linking of proteins, which increased gel fracture modulus. The decreased pH caused by the Maillard reaction in the gels containing ribose occurred after protein denaturation and gelation, thus having little if any effect on the gelation process.     

Effects of pH and the gel state on the mechanical properties, moisture contents, and glass transition temperatures of whey protein films.

The mechanical properties, moisture contents (MC), and glass transition temperature (T(g)) of whey protein isolate (WPI) films were studied at various pH values using sorbitol (S) as a plasticizer. The films were cast from heated aqueous solutions and dried in a climate chamber at 23 degrees C and 50% relative humidity (RH) for 16 h. The critical gel concentrations (c(g)) for the cooled aqueous solutions were found to be 11.7, 12.1, and 11.3% (w/w) WPI for pH 7, 8, and 9, respectively. The cooling rate influenced the c(g), in that a lower amount of WPI was needed for gelation when a slower cooling rate was applied. Both cooling rates used in this study showed a maximum in the c(g) at pH 8. The influence of the polymer network on the film properties was elucidated by varying the concentration of WPI over and under the c(g). Strain at break (epsilon(b)) showed a maximum at the c(g) for all pH values, thus implying that the most favorable structure regarding the ability of the films to stretch is formed at this concentration. Young's modulus (E) and stress at break (sigma(b)) showed a maximum at c(g) for pH 7 and 8. The MC and epsilon(b) increased when pH increased from 7 to 9, whereas T(g) decreased. Hence, T(g) values were -17, -18, and -21 degrees C for pH 7, 8, and 9, respectively. E and sigma(b) decreased and epsilon(b) and thickness increased when the surrounding RH increased. The thickness of the WPI films also increased with the concentration of WPI.     

Enzymatic hydrolysis of heat-induced aggregates of whey protein isolate

The effects of heat-induced denaturation and subsequent aggregation of whey protein isolate (WPI) solutions on the rate of enzymatic hydrolysis was investigated. Both heated (60 °C, 15 min; 65 °C, 5 and 15 min; 70 °C, 5 and 15 min, 75 °C, 5 and 15 min; 80 °C, 10 min) and unheated WPI solutions (100 g L(-1) protein) were incubated with a commercial proteolytic enzyme preparation, Corolase PP, until they reached a target degree of hydrolysis (DH) of 5%. WPI solutions on heating were characterized by large aggregate formation, higher viscosity, and surface hydrophobicity and hydrolyzed more rapidly (P < 0.001) than the unheated. The whey proteins exhibited differences in their susceptibility to hydrolysis. Both viscosity and surface hydrophobicity along with insolubility declined as hydrolysis progressed. However, microstructural changes observed by light and confocal laser scanning microscopy (CLSM) provided insights to suggest that aggregate size and porosity may be complementary to denaturation in promoting faster enzymatic hydrolysis. This could be clearly observed in the course of aggregate disintegration, gel network breakdown, and improved solution clarification.     

Physical properties, molecular structures, and protein quality of texturized whey protein isolate: effect of extrusion temperature

Although extrusion technology has contributed much to increasing the effective utilization of whey, the effect of extrusion conditions on the functional properties of the proteins is not well understood. In this work, the impact of extrusion temperature on the physical and chemical properties, molecular structures, and protein quality of texturized whey protein isolate (WPI) was investigated at a constant moisture content and compared with WPI treated with simple heat only. The Bradford assay methods, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and reversed-phase high-performance liquid chromatography techniques were used to determine protein solubility and to analyze compositional changes in the two major whey proteins, α-lactalbumin and β-lactoglobulin. Circular dichroism and intrinsic tryptophan fluorescence spectroscopic techniques were applied to study the secondary and tertiary structures of the proteins. This study demonstrated that extrusion temperature is a critical but not the sole determining factor in affecting the functional properties of extruded WPI.