响应面试验优化裙带菜蛋白酶解工艺及酶解液抗氧化活性

运用响应面分析方法对裙带菜蛋白酶解工艺条件进行优化。经单酶筛选,在单因素试验基础上,以亚铁离子螯合率和1,1-二苯基-2-三硝基苯肼(1,1-diphenyl-2-picrylhydrazyl,DPPH)自由基清除率为主要指标,水解度为辅助指标,研究酶解时间、酶解温度、p H值、底物质量浓度、加酶量对裙带菜蛋白酶解产物抗氧化活性和水解度的影响,并比较优化条件下的酶解液与常用天然抗氧化剂抗坏血酸、合成抗氧化剂丁基羟基茴香醚(butyl hydroxyanisole,BHA)的抗氧化活性。结果表明:复合蛋白酶是裙带菜蛋白酶解的最适用酶,酶解液螯合亚铁离子能力和清除DPPH自由基的最优条件为酶解时间8.1 h、酶解温度50℃、p H 7.0、底物质量浓度15 g/L、加酶量0.2%(0.3 AU/g裙带菜粉末)。在此条件下,酶解液的亚铁离子螯合率为88.58%,DPPH自由基清除率为59.22%,水解度为29.72%。对比常用抗氧化剂,在亚铁离子螯合能力方面,酶解液显著高于0.01%抗坏血酸和0.01%BHA(P0.05),而在DPPH自由基清除能力和还原能力方面,酶解液低于0.01%抗坏血酸和0.01%BHA(P0.05)。     

响应面法优化龙须菜蛋白酶解工艺及酶解液的抗氧化活性

为获得高抗氧化活性的龙须菜蛋白酶酶解液,运用响应面(RSM)分析方法对龙须菜蛋白酶解工艺条件进行优化。经单酶筛选,在单因素实验基础上,以亚铁离子螯合率和DPPH自由基清除率为主要指标,水解度为辅助指标,研究酶解时间、p H、酶解温度、底物质量浓度、加酶量对龙须菜蛋白酶解液抗氧化活性和水解度的影响。结果表明:在所选的5种蛋白酶中,复合蛋白酶是龙须菜蛋白酶解的最适用酶;酶解液抗氧化活性的最优酶解条件为酶解时间8.1 h、酶解温度50.1℃、p H7.0、底物浓度10 g/L、加酶量0.1%(0.15 AU/g)。在此条件下,酶解液的亚铁离子螯合率为74.61%,DPPH自由基清除率为47.66%,水解度为17.58%。对比常用抗氧化剂,酶解液在亚铁离子螯合能力方面显著高于0.01%维生素C和0.01%BHA(p0.05),而在DPPH自由基清除能力和还原能力方面,酶解液低于0.01%维生素C和0.01%BHA(p0.05)。优化后的龙须菜蛋白酶酶解液具有一定的抗氧化活性。     

鳄鱼皮酶解产物功能特性及抗氧化活性

为了解鳄鱼皮酶解产物功能特性和抗氧化活性,采用2种商业蛋白酶(木瓜蛋白酶、碱性蛋白酶)在各自最适反应条件下分别酶解鳄鱼皮,研究水解度(DH)、酶种类及pH值对酶解产物功能特性及抗氧化活性的影响.结果显示:随着酶解时间延长,鳄鱼皮水解度逐渐增加,鳄鱼皮在碱性蛋白酶酶解作用下水解度较高,水解4h时可达12％;木瓜蛋白酶酶解产物与碱性蛋白酶酶解产物的溶解性差异不显著(P＞0.05).相同水解度下,碱性蛋白酶酶解产物的热稳定性在pH4时优于木瓜蛋白酶酶解产物.酶解时间在1h之内,木瓜蛋白酶酶解物亚铁离子螯合力明显增强;随着时间延长,酶解产物亚铁离子螯合能力变化不显著(P＞0.05).酶解3h后碱性蛋白酶酶解产物亚铁离子螯合能力高于木瓜蛋白酶酶解产物,但木瓜蛋白酶酶解产物具有较强的清除DPPH自由基能力.综上表明,碱性蛋白酶水解作用的鳄鱼皮水解度较高,其酶解产物乳化活性和热稳定性优于木瓜蛋白酶酶解产物;鳄鱼皮酶解产物抗氧化能力较强,有较高的开发利用价值.     

体外消化对玉米谷蛋白酶解物抗氧化活性的影响

以玉米蛋白粉为原料提取玉米谷蛋白,利用蛋白酶对其进行酶解,制备蛋白水解物,再用胃蛋白酶及胰蛋白酶对其进一步消化处理,以抗氧化活性为指标对消化过程中蛋白酶加量进行优化,制备富含谷氨酰胺的具有抗氧化功能的蛋白水解物。结果表明,玉米谷蛋白经复合蛋白酶水解后,蛋白收率(54.41±3.26)%,水解物的羟基自由基清除率、亚铁离子螯合能力及还原力分别达到(29.50±0.31)%、(73.33±0.62)%及0.50±0.01;二步消化后蛋白水解物羟基自由基清除率、亚铁离子螯合能力及还原力分别达到(55.88±0.94)%、(90.44±0.01)%及0.40±0.01。     

甲醇芽孢杆菌蛋白酶的水解特性及酪蛋白水解产物活性分析

首先确定甲醇芽孢杆菌LB-1蛋白酶(蛋白酶LB-1)最适水解温度和pH值,然后通过持续监测水解pH值和氧化还原电位变化、十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分析、质谱检测等探究蛋白酶LB-1的蛋白水解特性,并测定其酪蛋白水解产物的1,1-二苯基-2-三硝基苯肼(1,1-diphenyl-2-picrylhydrazyl,DPPH)自由基和2,2’-联氮双(3-乙基苯并噻唑啉-6-磺酸)(2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid),ABTS)阳离子自由基清除率、α-淀粉酶和α-葡萄糖苷酶抑制率、金属离子螯合率等生物活性。结果表明:蛋白酶LB-1的最适蛋白水解温度为52℃,最适pH 6.0～7.0;该酶对酪蛋白的水解作用显著强于乳清蛋白,水解程度为κ-酪蛋白α-酪蛋白β-酪蛋白;对κ-酪蛋白的水解位点主要位于Lys_(21)-Ile_(22)和Lys_(112)-Asn_(113);酪蛋白水解产物对DPPH自由基和ABTS阳离子自由基最高清除率分别为75.4%、48.45%,对α-淀粉酶和α-葡萄糖苷酶的最高抑制率分别为80.89%、93.12%,对钙离子和锌离子的最高螯合率分别为63.13%、35.31%;表明该酶的酪蛋白水解产物具有一定的抗氧化能力、降血糖作用以及良好的金属离子螯合能力。因此,甲醇芽孢杆菌LB-1蛋白酶在功能性干酪加工和乳源活性肽开发方面具有潜在的应用价值。     

米糠肽-铁锌螯合物制备工艺研究

目的 研究米糠蛋白制备米糠肽-铁锌螯合物的工艺。方法 以米糠蛋白为原料,优化米糠肽-铁锌螯合物制备工艺。以亚铁离子与锌离子的螯合率和水解度为评价指标,考察蛋白酶种类、反应物质量比、温度、时间及pH值对米糠酶解产物铁锌螯合率的影响。在单因素试验的基础上,结合JMP软件进行定制试验设计,确定米糠肽-铁锌螯合物的制备工艺。结果 碱性蛋白酶较适宜制备米糠肽-铁锌螯合物,其最佳螯合工艺条件为时间70 min、温度70 oC、反应物质量比3:1、pH 5.0;在此条件下,其产物亚铁离子螯合率达到38.29%、锌离子螯合率达到57.2%,与预测值接近。结论 本研究采用米糠肽同时螯合铁锌2种金属离子,不仅可以充分吸收米糠肽的营养成分,提高消化率,发挥其降血压、降血脂、抗氧化等功能特性,还可以同时补铁、补锌,对人体具有积极作用。     

驼血蛋白酶解制备抗氧化肽工艺参数

由于骆驼特殊的生存环境,使得驼血组成及含量不同于其他动物血液,其蛋白质含量尤其丰富,具有较高的生物活性。研究通过单因素、正交试验,以水解度为评价指标,研究了复合蛋白酶、木瓜蛋白酶、风味蛋白酶和胃蛋白酶对驼血酶解的影响,确定了4种蛋白酶水解驼血的最优条件。在单因素正交试验基础上,以DPPH·清除能力、亚铁离子螯合能力、总抗氧化能力(ABTS法)为评价指标,确定了胃蛋白酶是水解驼血制备抗氧化肽的最佳蛋白酶;并在此基础上对胃蛋白酶水解产物超滤组分进行抗氧化活性检测,确定了组分Pep-1(10 ku)抗氧化活性最强,其DPPH·清除率为72.6%,铁离子螯合能力94.9%,总抗氧化能力(ABTS法)为1.433 mmol/g。     

芝麻蛋白制备金属螯合肽的酶解工艺研究

分别采用木瓜蛋白酶、胰蛋白酶和碱性蛋白酶Alcalase对芝麻蛋白进行水解,结果表明胰蛋白酶为制备金属螯合肽的最佳酶。通过优化胰蛋白酶酶解工艺条件发现最佳条件为:底物质量浓度5%(g/100mL),酶添加浓度20u/g(底物),水解时间5h,得到的酶解产物金属螯合率最强,与Fe2+的螯合率为90.9%,与Zn2+的螯合率为93.5%。通过考察水解度对酶解产物金属螯合率及抗氧化能力的影响,结果显示水解度在18.9%到22.4%之间的酶解产物金属螯合率强,且水解度在一定范围内金属螯合率和抗氧化能力均与水解度呈正相关性。     

复合蛋白酶水解玉米谷蛋白产物的抗氧化活性研究

以玉米谷蛋白为原料,利用复合蛋白酶对其进行限制水解,探讨不同水解时间所获得的谷蛋白酶解物的分子量分布和抗氧化活性。结果表明:不同水解时间获得的酶解物分子量分布差异较大。水解时间为120min的酶解物中,分子量分布为6 511.51~307.32Da的肽段占94.36%,此时获得的酶解物的抗氧化活性最高,其对DPPH自由基、O-2·、·OH的清除率分别为58.86%,82.64%,37.21%;还原力为0.236;与亚铁离子的螯合能力为29.92%。     

具有铁螯合能力的驴骨胶原肽酶解条件优化及微观形态

为充分开发利用驴骨资源,开发一种酶解驴骨蛋白从而得到驴骨胶原蛋白肽的方法,考察该酶解肽与亚铁离子的螯合能力,并将其产物进行表征。以亚铁离子螯合率为考察指标,筛选最优酶种类,通过单因素实验和考虑交互作用的正交实验优化确定驴骨的最佳酶解条件。结果表明,驴骨的最佳预处理条件为121 ℃蒸煮30 min,木瓜蛋白酶为最佳酶,优化过后的酶水解工艺参数为温度50 ℃、pH 7.0、酶底比1 000 U/mL、时间3 h,此时驴骨胶原肽和亚铁离子的螯合率为38.40%,可得到微观结构为片状的螯合产物。木瓜蛋白酶酶解驴骨蛋白可以制备有螯合亚铁离子能力的驴骨胶原肽,具有促铁吸收的潜力,对开发驴骨胶原肽相关产品具有科学参考作用。     

Generation,Fractionation, and Characterization of Iron‐Chelating Protein Hydrolysate from Palm Kernel Cake Proteins

Palm kernel cake protein was hydrolyzed with different proteases namely papain, bromelain, subtilisin, flavourzyme, trypsin, chymotrypsin, and pepsin to generate different protein hydrolysates. Peptide content and iron‐chelating activity of each hydrolysate were evaluated using O‐phthaldialdehyde‐based spectrophotometric method and ferrozine‐based colorimetric assay, respectively. The results revealed a positive correlation between peptide contents and iron‐chelating activities of the protein hydrolysates. Protein hydrolysate generated by papain exhibited the highest peptide content of 10.5 mM and highest iron‐chelating activity of 64.8% compared with the other hydrolysates. Profiling of the papain‐generated hydrolysate by reverse phase high performance liquid chromatography fractionation indicated a direct association between peptide content and iron‐chelating activity in most of the fractions. Further fractionation using isoelectric focusing also revealed that protein hydrolysate with basic and neutral isoelectric point (pI) had the highest iron‐chelating activity, although a few fractions in the acidic range also exhibited good metal chelating potential. After identification and synthesis of papain‐generated peptides, GGIF and YLLLK showed among the highest iron‐chelating activities of 56% and 53%, whereas their IC₅₀ were 1.4 and 0.2 μM, respectively.     

Evaluation of Antioxidant Activities of Zein Protein Fractions

Zein protein was extracted from the by‐product corn gluten meal. The obtained zein protein was 1st hydrolyzed by 4 different proteases. The antioxidant activities of the hydrolysates or peptides were evaluated by free radical scavenging activity, metal ion chelating activity, and lipid peroxidation inhibitory capacity. Among hydrolysates produced, alkaline protease hydrolysates exhibited the highest antioxidant activity. A regression model was established by uniform design to optimize the alkaline protease hydrolysis conditions. The hydrolysates with molecular weight < 3 kDa obtained from ultrafiltration showed the highest antioxidant activities in all relevant assays. The hydrolysates with molecular weight <3 kDa were subsequently purified by gel filtration chromatography, and fraction F3 exhibited the highest antioxidant activities. Two peptides were identified from fraction F3 using LC‐ESI‐Q‐TOF MS/MS as Pro‐Phe (263.13 Da) and Leu‐Pro‐Phe (375.46 Da). These peptides exhibited good free radical scavenging activity and lipid peroxidation inhibitory effect. The results clearly indicated that zein protein fractions are good sources for the development of natural antioxidants for the food industry.     

Antioxidant properties and protein compositions of porcine haemoglobin hydrolysates

Porcine haemoglobin hydrolysates were prepared through hydrolysis by Alcalase followed by Flavourzyme, and their protein compositions were analyzed using Sephadex G-50 gel filtration chromatography. The antioxidant activities, including reducing power, ferrous ion chelating ability, and DPPH radical scavenging activity, of the hydrolysates were evaluated. The results showed that the hydrolysates of haemoglobin exhibited low reducing powers, but high ferrous ion chelating abilities and DPPH radical scavenging activities. The hydrolysate, obtained through hydrolysis by 2% Alcalase for 4 h and followed by 1% Flavourzyme for 6 h, had the highest ferrous ion chelating ability of 63.54% at a concentration of 5.0 mg/mL. The hydrolysate, obtained through hydrolysis by 2% Alcalase for 4 hrs, had the highest DPPH radical scavenging activity of 41.94% at a concentration of 5.0 mg/mL. According to the results of protein composition analysis, we divided the hydrolysates into three groups, including high molecular weight (MW) group (Group I), medium MW group (Group II), and low MW group (Group III). The reducing power and ferrous ion chelating ability of the hydrolysates were significantly and positively correlated to the relative amount of Group I, and negatively correlated to the relative amount of Group III. This study revealed that the antioxidant activities of porcine haemoglobin hydrolysates were dependent on their protein compositions. The high MW protein fraction (Group I) was responsible for the high reducing power and ferrous ion chelating ability of the hydrolysate.     

鳄鱼肉蛋白酶解产物抗氧化性的研究

为更加充分利用优质的鳄鱼肉蛋白资源,采用木瓜蛋白酶、风味蛋白酶单酶酶解及木瓜蛋白酶与风味蛋白酶复合酶解制备了鳄鱼肉蛋白多肽,并对其抗氧化活性（DPPH自由基清除能力、亚铁离子螯合能力和还原力）进行分析。结果显示,酶解时间为Q5h、1h和2h木瓜蛋白酶和风味蛋白酶复合酶解产物的DPPH自由基清除能力显著（P〈0．05）,高于木瓜蛋白酶和风味蛋白酶单酶酶解产物的DPPH自由基清除能力;除酶解时间为0．5、1的风味蛋白酶鳄鱼肉蛋白多肽外,其它鳄鱼肉蛋白多肽的亚铁离子螯合率均大于80％;随酶解时间延长,木瓜蛋白酶和风味蛋白酶复合酶解鳄鱼肉蛋白多肽的还原力显著降低（P〉0．05）。鳄鱼肉蛋白酶解产物具备开发为功能食品的潜力。     

Sensory Characteristics and Antioxidant Activities of Maillard Reaction Products from Soy Protein Hydrolysates with Different Molecular Weight Distribution

Soy protein was hydrolyzed using two enzymes to obtain soy protein hydrolysate (SPH), the SPH was fractionated with ultrafiltration membranes to obtain peptide fractions below 1,000 Da (SP1) and 1,000–5,000 Da (SP2), and for the meantime, SPH was further completely hydrolyzed to get compound amino acids (CAA). Maillard reaction products (MRPs) were prepared from aqueous xylose–SPH/SP1/SP2/CAA model systems by heating at 120°C for 2.0 h. Compared with the original hydrolysates and other MRPs, the MRPs from SP2 exhibited a distinctly enhanced effect on flavor, including the caramel-like, soy sauce-like odors, umami and mouthful tastes and a greatly reduced bitterness in consomme′ soup. Antioxidant activities of SPH, SP1, SP2, CAA, and their MRPs were investigated through reducing power, DPPH radical-scavenging activity, and Fe²⁺ chelating activity. Before Maillard reaction, the antioxidant activities of peptide fractions with different molecular weights were quite different, and SP2 showed the highest activity; however, CAA exhibited very poor antioxidant activity. The antioxidant activities of SPH, SP1, SP2, and CAA were greatly enhanced by Maillard reaction, and the MRPs prepared from xylose–CAA model system exhibited a higher antioxidant activity than those from other model systems. Pyrazines, pyrroles, furans, and thiazoles were significantly correlated with reducing power and Fe²⁺ chelating activity by principal component analysis.     

美拉德糖基化对玉米蛋白粉双酶水解产物抗氧化活性的影响

利用复合蛋白酶和碱性蛋白酶协同作用于玉米蛋白粉,将得到的双酶水解产物与壳寡糖进行美拉德糖基化反应,研究不同蛋白浓度下的酶解物和糖基化产物的抗氧化活性。结果表明,酶解物和糖基化产物都具有良好抗氧化活性,美拉德糖基化反应可以提高酶解物的抗氧化活性。当糖基化产物蛋白浓度为5.00 mg/mL时,其·OH清除率和DPPH·清除率分别达到了90.91%和61.75%,蛋白浓度为1.00mg/mL时,Fe~(2+)螯合能力为83.42%,比酶解物·OH清除率和DPPH·清除率分别提高了48.78%和32.26%,Fe~(2+)螯合能力提高了22.63%。     

Enzyme-enhanced extraction of antioxidant ingredients from red algae Palmaria palmata

The effect of various protease and carbohydrase treatments on the extraction of polyphenols and other antioxidant ingredients from the red algae Palmaria palmata (dulse) was investigated. In addition, the relative contribution of different fractions to the overall antioxidant capacity of the hydrolysate was evaluated. Considerable differences were observed both in total phenolic content (TPC) and antioxidant activities of the hydrolysates evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity, oxygen radical absorbance capacity (ORAC) and ferrous ion-chelating ability assays. All the proteases tested had significant enhancing effect on the extraction of polyphenols and other active components compared to carbohydrases and cold water extraction (control). The Umamizyme extract had the highest TPC and consequently exhibited the strongest scavenging capacity against DPPH and peroxyl radicals. Further fractionation of the Umamizyme extract revealed that the crude polyphenol fraction possessed the highest peroxyl radical scavenging activity, whereas the crude polysaccharide fraction was more effective for chelating ferrous ions. The data from this study suggest the potential of protease treatment to improve value-added utilization of dulse extracts as antioxidants in functional foods and nutraceuticals.     

Effect of ultrasonic pretreatment on the antioxidant properties of porcine liver protein hydrolysates

In this study, the effect of ultrasonic pretreatment on the antioxidant activity of porcine liver protein hydrolysates (PLPHs) was investigated. The results showed that the degree of hydrolysis (DH) and peptide contents of the PLPHs increased as the time of ultrasonication increased. The hydrolysate pretreated with ultrasonication for 60 s exhibited the highest DH and peptide contents. The hydrolysate pretreated with ultrasonication for 45 s exhibited the highest ferrous ion chelating ability and reducing power. The hydrolysate pretreated with ultrasonication for 30 s exhibited the highest 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) radical scavenging activity and the higher inhibitory activity in the linoleic acid autoxidation system. The molecular weight of peptides in the hydrolysates was less than 6.2 kDa. The results clearly demonstrated that ultrasonic pretreatment enhances the antioxidant activities of the PLPHs in a short period of time (15–30 s).     

Sweet potato protein hydrolysates: antioxidant activity and protective effects on oxidative DNA damage

In this study, sweet potato protein (SPP) hydrolysates were prepared by six enzymes (alcalase, proleather FG‐F, AS1.398, neutrase, papain and pepsin). The antioxidant activities and protective effect against oxidative DNA damage of SPP hydrolysates were investigated. Alcalase hydrolysates exhibited the highest hydroxyl radical‐scavenging activity (IC₅₀ 1.74 mg mL^?1) and Fe²⁺‐chelating ability (IC₅₀ 1.54 mg mL^?1) (P < 0.05). Compared with other five hydrolysates, the hydrolysates obtained by alcalase had the most abundant <3‐kDa fractions. In addition, below 3‐kDa fractions of alcalase hydrolysates showed the highest antioxidant activities and protective effects against DNA damage through both scavenging hydroxyl radicals and chelating Fe²⁺, which was probably because of the increase in several antioxidant amino acids, such as His, Met, Cys, Tyr and Phe, as well as the hydrophobic amino acids. The results suggested that enzymatic hydrolysis could be used as an effective technique to produce high value‐added peptides products from SPP.     

太平洋鳕鱼排酶解产物的制备及其钙螯合活性研究

目的以太平洋鳕鱼排为原料,制备具有良好钙螯合活性的酶解产物,并对其促钙吸收转运功效进行评价。方法采用动物蛋白水解酶、中性蛋白酶和菠萝蛋白酶分别进行水解,比较3种酶解产物的水解度与蛋白回收率,确定最佳水解酶,并研究酶解产物的钙螯合活性与促钙吸收转运活性。结果经动物蛋白水解酶制备的酶解产物水解度与蛋白回收率最高。当酶解时间为320 min时,该酶解产物的钙螯合活性最高。氨基酸分析表明谷氨酸、甘氨酸、天冬氨酸、丝氨酸和苏氨酸等特定氨基酸所占比例较高。该酶解产物对Caco-2细胞无毒性作用,能够明显促进钙转运率。结论太平洋鳕鱼排酶解产物具有较好的钙螯合活性与促钙转运活性。