不同蛋白酶水解棉籽蛋白制备抗氧化多肽的研究

利用6种蛋白酶对棉籽蛋白进行酶解,测定了各酶在水解过程中的水解度及其变化,对酶解产物的抗氧化活性进行了分析比较。研究表明,各蛋白酶在水解的前2h内,水解度迅速增加,2h之后水解曲线变得平缓。其中胃蛋白酶的水解能力最强,其4h水解产物水解度最大,为30.40%;胰蛋白酶的水解能力最差,最终水解产物的水解度为17.61%。中性蛋白酶水解产物的抗氧化活性较强,经测定其DPPH清除能力为54.95%,羟自由基清除能力为68.98%,超氧阴离子自由基清除能力为58.38%。     

银鲳酶解物抗氧化活性研究

选用胃蛋白酶、胰蛋白酶、碱性蛋白酶和中性蛋白酶对银鲳蛋白进行酶解以制备蛋白酶解物,以羟基自由基清除活性为指标确定银鲳最佳水解酶。结果显示,碱性蛋白酶的水解物抗氧化活性最强。实验对碱性蛋白酶水解银鲳的酶解条件(时间、温度、pH、酶添加量和固液比)进行正交实验设计,并对最佳水解条件下所获得的酶解物进行抗氧化活性测试。结果表明,银鲳蛋白碱性蛋白酶水解物对DPPH自由基和羟基自由基具有清除作用,其自由基清除效果呈现剂量依赖性,而且银鲳蛋白水解物还具有明显还原能力。所有这些体外抗氧化数据说明,银鲳蛋白水解物有明显的抗氧化效力。     

罗非鱼皮明胶酶解物及其模拟消化产物的抗氧化活性

以罗非鱼皮为原料提取罗非鱼皮明胶,选用风味蛋白酶和胰蛋白酶制备罗非鱼皮明胶酶解物,采用ABTS自由基、DPPH自由基、羟基自由基及亚油酸过氧化体系,初步评价罗非鱼皮明胶酶解物的抗氧化活性,再通过模拟体外胃肠道消化实验,结合分子质量分布测定,进一步考察罗非鱼皮明胶酶解物的抗氧化活性。结果显示,在酶解过程中,风味蛋白酶及胰蛋白酶酶解的水解度逐渐升高,在3 h时达到最高,分别达到5.8%和25.36%。在酶解60 min时其TCA可溶性肽得率最高,风味蛋白酶、胰蛋白酶酶解物分别达56.82%和54.44%。通过比较半抑制浓度（IC₅0）,确定了酶解60 min时风味蛋白酶酶解物的清除DPPH自由基及抑制亚油酸过氧化能力较胰蛋白酶酶解物强。模拟体外胃肠道消化后,酶解物羟基自由基清除活性均显著提高（p<0.05）,亚油酸脂质过氧化活性明显降低,消化前后样品分子量分布范围均主要集中于3000⁵000 Da,消化后风味蛋白酶及胰蛋白酶酶解物3000⁵000 Da组分的含量分别提高了45%及13%。以上研究结果表明,罗非鱼皮明胶酶解后制备的明胶水解物具有一定的抗氧化能力,具有潜在的开发价值。      

Antioxidant activities and functional properties of soy protein isolate hydrolysates obtained using microbial proteases

Soy protein isolate (SPI) hydrolysates were prepared using microbial proteases to produce peptides with antioxidant activity. The process parameters (substrate and enzyme concentrations), hydrolysis time, functional properties and the effects of ultrafiltration were further investigated. The results showed that the soy protein isolate exhibited a 7.0‐fold increase in antioxidant activity after hydrolysis. The hydrolysis parameters, defined by the experimental design, were a substrate concentration of 90 mg mL^?1 and the addition of 70.0 U of protease per mL of reaction. The maximum antioxidant activities were observed between 120 and 180 min of hydrolysis, where the degree of hydrolysis was approximately 20.0%. The hydrolysis increased solubility of the soy protein isolate; however, the hydrolysates exhibited a tendency to decrease in the interfacial activities and the heat stability. The SPI hydrolysates fractions obtained by ultrafiltration showed that the enzymatic hydrolysis resulted in samples with homogenous size and strong antioxidant activity.     

Nutritional composition and antioxidant properties of protein hydrolysates prepared from echinoderm byproducts

Protein hydrolysates were prepared from echinoderm byproducts, including viscera (SCV) of Atlantic sea cucumber ( Cucumaria frondosa ) and digestive tract (UDT) and non-commercial grade gonads (UGN) of green sea urchin ( Strongylocentrotus droebachiensis ). Enzymatic hydrolysis was performed on defatted materials using Alcalase^® 2.4L (0.75% w/w) and reaction was carried out overnight (∼16 h, 55 °C, pH 8.0). Freeze-dried hydrolysates were analysed for their nutritional composition, nitrogen solubility index and antioxidant activity. Degree of hydrolysis was low, with values of 5.6%, 4.6% and 7.0% for SCV, UDT and UGN, respectively. Hydrolysates showed high protein content (∼55%), high proportion of essential amino acids (∼35% of total amino acids) and good solubility (nitrogen solubility index ≈ 68%). They contained variable concentrations of major and trace elements with a predominance of Na and K. Hydrolysates showed apparent antioxidant activities in both ORAC assay (267–421 μmol TE g⁻¹) and inhibition of lipid oxidation test (54–57%). Antioxidant activities were thought to be associated with the presence of antioxidant peptides in hydrolysates. Our results showed that hydrolysates from Atlantic sea cucumber and green sea urchin byproducts might serve as alternative sources of dietary proteins, with good nutritional composition, high solubility and interesting protection against oxidative stress.     

Antioxidant activity of hydrolysates derived from porcine plasma

BACKGROUND: In China alone, more than 400 million pigs are slaughtered each year to provide meat. Porcine blood is rich in proteins but is usually discarded, which can cause environmental contamination. Recovering porcine blood and converting it to high‐value products is therefore economically and environmentally desirable. However, very little information on antioxidant peptides from porcine blood by‐products is currently available. In this study the antioxidant properties of porcine plasma hydrolysates PPE and PPA prepared with pepsin and papain respectively were investigated. RESULTS: Both PPE and PPA showed excellent antioxidant activity in a linoleic acid system (AL) compared with α‐tocopherol (VE) at the same concentration (P < 0.01). Their activities were respectively 3.33 and 1.83 times stronger than that of VE at a concentration of 10 µg mL^?1 and 5.4 and 5.6 times stronger at 100 µg mL^?1. The 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) radical‐scavenging activity (DRSA) reached 48.4 and 43.1% for PPE and PPA respectively at 500 µg mL^?1. The ferrous ion‐chelating power (FICP) of PPE at 100 µg mL^?1 was about 1.5 times stronger than that of 10 µmol L^?1 ethylene diamine tetraacetic acid (EDTA) in a 50 µmol L^?1 Fe²⁺ system, whereas the FICP of PPA at 100 µg mL^?1 was 61% that of 10 µmol L^?1 EDTA. Furthermore, PPE was separated on Resource 15RPC and Superdex peptide 10/300GL columns, and the antioxidant activity of the peptides and its relationship to their polarity and molecular weight (MW) were analysed. The hydrolysate was divided into four groups (R1–R4) with hydrophobicities ranging from weak to strong by Resource 15RPC, while it was divided into three groups (S1, MW 7–12 kDa; S2, MW 3–7 kDa; S3, MW 1–3 kDa) by Superdex peptide 10/300GL. CONCLUSION: The results showed that AL was significantly and positively correlated with the relative amounts of R1, S2 and S3 and that DRSA was dependent on R3 and S1. The fractions of PPE were not responsible for FICP. Copyright © 2009 Society of Chemical Industry     

Antioxidant and anti‐acetylcholinesterase activities of anchovy (Coilia mystus) protein hydrolysates and their memory‐improving effects on scopolamine‐induced amnesia mice

Anchovy protein hydrolysates (APHs) were prepared through hydrolysis for 2, 4 or 8 h (APH‐2, APH‐4 and APH‐8, respectively). The chemical analyses, in vitro assessments [antioxidant activity and acetylcholinesterase (AchE) inhibitory activity] and in vivo mice tests were evaluated. Results revealed that APH‐8 exhibited the strongest reducing power and AchE inhibitory capacity (IC₅₀ = 159.76 ± 0.03 mg mL^?1), which may be due to its specific amino acid composition and newly formed peptides. In addition, AchE inhibitory kinetics of amino acids suggested that lysine was featured of both competitive and noncompetitive inhibitors. Furthermore, the results of in vivo study showed that all APHs exhibited memory‐improving action on scopolamine‐induced amnesia mice especially, APH‐8, indicating that anchovy protein is a potential source for health‐promoting peptides.     

鳙鱼酶解物体外抗氧化的研究

利用碱性蛋白酶Alcalase对鳙鱼进行控制酶解,制备出具有清除自由基活性的酶解物并比较其还原能力和对脂肪氧合酶活性的影响,并考察其对红细胞溶血、肝线粒体肿胀的影响,结合不同处理后肝脑组织中MDA浓度的测定以对鳙鱼酶解物在模拟体系中的抗氧化活性进行综合评价。结果表明三种鳙鱼酶解物的肽提取率均在70%以上且主要由寡肽(含5～6个氨基酸残基)组成,三种酶解物均具有一定的还原能力,在较高浓度时能抑制脂肪氧合酶的活性并可在体外抑制因H2O2引发的新西兰家兔红细胞溶血和由FeSO4和Vc诱导的肝线粒体肿胀,同时降低肝脑组织中MDA的产生。说明鳙鱼酶解物在一定浓度范围内具有抗氧化作用,比较三种酶解物的抗氧化特性可知酶解物C具有较强的综合抗氧化能力,可作为后续研究的材料。     

虾夷扇贝生殖腺酶解物-核糖美拉德反应产物抗氧化特性研究

本文选用碱性蛋白酶、中性蛋白酶、风味蛋白酶及木瓜蛋白酶制备虾夷扇贝雌性生殖腺酶解物,再与核糖进行美拉德反应获得虾夷扇贝雌性生殖腺酶解物-核糖美拉德反应产物。采用DPPH自由基和亚油酸过氧化体系,评价美拉德产物的抗氧化能力。结果显示,碱性蛋白酶酶解效果较好,在酶解3 h时其水解度及TCA可溶性寡肽得率最高,分别达24.16%和20.79%。以DPPH自由基清除活性及亚油酸过氧化抑制活性为指标,确定了pH7条件下制备的虾夷扇贝雌性生殖腺酶解物-核糖美拉德产物抗氧化能力相对较强。通过比较半抑制浓度（IC50）,美拉德产物的抗氧化能力较酶解物及核糖本身均显著提高（p<0.05）,但弱于抗氧化剂2,6-二叔丁基-4-甲基苯酚（BHT）。以上研究结果说明,虾夷扇贝生殖腺酶解物-核糖美拉德反应产物具有一定的抗氧化能力,具有潜在的开发价值。      

罗非鱼鱼肉及组分蛋白酶解产物的抗氧化特性

采用胃蛋白酶、中性蛋白酶、Alcalase2.4L碱性蛋白酶分别酶解罗非鱼肉蛋白,研究比较不同蛋白酶在1、2、4、6、8h对罗非鱼肉蛋白的酶解效果及产物的抗氧化活性,结果表明:3种酶的酶解产物都具有一定的抗氧化活性,中性蛋白酶酶解产物的水解度和蛋白回收率最高,胃蛋白酶酶解产物对DPPH自由基和羟自由基的清除效果最好;综合各项指标选择中性蛋白酶对罗非鱼肉的组分蛋白(肌原纤维蛋白、肌浆蛋白、基质蛋白)进行4h酶解,研究比较不同组分蛋白的酶解效果及产物的抗氧化活性,结果表明:组分蛋白中基质蛋白酶解产物的蛋白回收率最高,为89.8%,肌原纤维蛋白酶解产物的水解度最高,为10.1%,肌浆蛋白酶解产物的抗氧化活性最好,羟自由基和DPPH自由基的IC50值分别为12.352mg/m L和3.554mg/m L。      

Valorisation of blood protein from livestock to produce haem iron-fortified hydrolysates with antioxidant activity

Blood is the main by-product from slaughterhouses bearing high levels of suspended solids (18% w/w) and BOD₅ (250 000–375 000 mg O₂/L), which makes difficult its handling and disposal. This study proposes the valorisation of blood protein to produce enzymatic hydrolysates rich in haemic iron and antioxidant peptides. Haemic iron presents higher bioavailability compared to its inorganic form, but its incorporation into foodstuffs is restrained by its high tendency to oxidation. Six commercial proteases of animal, plant and bacterial origin were employed to produce the enzymatic hydrolysates. Subtilisin and trypsin treatments were able to recover 70% of haem iron as soluble peptides, in contrast with plant enzymes where the proteolysis was reduced. Moreover, some enzymatic treatments led to hydrolysates with high levels of some in vitro antioxidant activities such as radical scavenging (Protamex, IC₅₀ 3.52 mg/mL) or metal chelating activity (trypsin, IC₅₀ 0.27 mg/mL). We conclude the enzymatic valorisation of blood protein increases haemic iron bioavailability and produces antioxidant peptides. Both properties are of interest for their use as iron fortification supplements.     

南瓜籽蛋白酶解液的超滤分离及其抗氧化活性研究

采用超滤技术分离南瓜籽蛋白酶解液,通过研究操作压力、料液浓度、pH和时间四个因素对膜通量和膜效能的影响,依次得到截留分子量分别为10、4ku两次超滤的最佳工艺条件,并比较超滤前后酶解液的抗氧化活性。结果表明,截留分子量为10ku的超滤膜的操作压力为0.25MPa、料液浓度为3.0%、料液pH为7,操作时间为60min;截留分子量为4ku的超滤膜的操作压力为0.20MPa、料液浓度为2.0%、料液pH为7,操作时间为60min。经过超滤以后分子量小于4ku的组分具有更强的清除自由基的能力,其中清除50%DPPH自由基所需的多肽浓度即半抑制浓度IC50值为2.76mg/mL、清除羟基自由基（·OH）的IC50值为2.91mg/mL、清除超氧阴离子自由基（O2-·）的IC50值为23.58mg/mL,同时对铁离子还具有显著的还原能力,吸光度为0.5时所需的多肽浓度为9.43mg/mL。      

Antioxidant activities and functional properties of grass carp (Ctenopharyngodon idellus) protein hydrolysates

BACKGROUND: Grass carp, with an annual production exceeding 4 × 10⁶ t in China in 2009, has not been developed into a high‐value product. In this study the antioxidant activities and functional properties of grass carp protein hydrolysates prepared with Alcalase 2.4L (HA) and papain (HP) were investigated. The hydrolysate with strongest radical‐scavenging activity and reducing power was assessed further for changes in its antioxidant activity during simulated gastrointestinal digestion. RESULTS: As the degree of hydrolysis (DH) increased, the metal‐chelating activity of both HA and HP increased while their reducing power and 1,1‐diphenyl‐2‐picrylhydrazyl radical (DPPH^?)‐scavenging activity decreased (P < 0.05). At the same DH, HP possessed higher DPPH^?‐scavenging activity and reducing power than HA (P < 0.05). The metal‐chelating activity of HP with 10% DH was significantly increased after in vitro gastrointestinal metabolism (P < 0.05). Regarding their functional properties, all hydrolysates were more than 81% soluble over a wide range of pH (3–8). At the same DH, HP showed higher emulsion activity index but lower solubility and foaming capacity than HA. CONCLUSION: Grass carp protein hydrolysates showed high solubility over a wide pH range and could be used as natural antioxidants in food systems. Copyright © 2011 Society of Chemical Industry     

贻贝酶解物体外抗氧化试验研究

为了鉴定世界食品产品中受到很高评价的海洋生物的抗氧化性,研究了贻贝的抗氧化性。通过测定酶解物的抗氧化性,从胰蛋白酶、木瓜蛋白酶和胃蛋白酶3种酶中,筛选出木瓜蛋白酶和胰蛋白酶作为酶解贻贝制备具有较高抗氧化性酶解物的理想水解酶,用正交试验L9(34)对2种酶的水解条件进行了优化。结果表明,木瓜蛋白酶在温度60℃、酶解时间15min、pH7.5、酶质量分数1.00%的水解条件下,酶解物的抗氧化性最好;胰蛋白酶在温度50℃、酶解时间60min、pH8.5、酶质量分数0.75%的水解条件下,酶解物的抗氧化效果最佳。     

Enzymatic production of bioactive protein hydrolysates from tuna liver: effects of enzymes and molecular weight on bioactivity

Tuna liver is fish by‐product, which is normally discarded and/or used as fish meal and animal feed because of poor functionality. In this study, we attempt to recover functional peptides from tuna liver protein by enzymatic hydrolysis using various proteases, and further hydrolysates were fractionated into four categories base on their molecular weight (MW) by ultrafiltration membranes. All fractionated hydrolysates produced by Alcalase, Neutrase and Protamex following Flavourzyme hydrolysis showed excellent antioxidant activities against DPPH, hydrogen peroxide and hydroxyl radical scavenging, and reducing power. Their bioactivity was dependent on treated enzymes, and antioxidant activities of fractions dependent on employing bioassay. Moreover, they confirmed antioxidant ability toward the protection effects on hydroxyl radical‐induced DNA damage by the measuring the conversion of supercoiled pBR322 plasmid DNA to the open circular form. In addition, all fractionated hydrolysates inhibited acetylcholinesterase activity, which is involved in Alzheimer’s diseases, and high MW fractions showed high AChE inhibition activity than those of low MW fractions.     

美拉德反应对泥鳅蛋白酶解产物抗氧化性能的强化研究

为提高泥鳅肉水解产物的抗氧化活性,本研究利用木瓜蛋白酶和风味蛋白酶对泥鳅蛋白进行酶解,在此基础上,加入单种糖或者混合糖与酶解产物以质量比1:1进行美拉德反应,并以产物对羟基自由基、超氧阴离子、DPPH自由基的清除率为指标评价其抗氧化性能。结果表明,加入单种糖进行反应的产物抗氧化性能低于混合糖,其中又以木糖和乳糖质量比1:3混合效果最佳,对羟基自由基、超氧阴离子自由基、DPPH自由基的清除率分别为47.13%±2.24%、58.27%±2.19%、68.09%±1.33%。再通过正交实验得到在pH7、80 ℃下反应45 min的产物对羟基自由基的清除率效果最佳,为58.86%;在pH6、100 ℃下反应45 min的产物对超氧阴离子自由基的清除率最佳,为70.63%;在pH8,温度90 ℃下反应60 min的产物对DPPH自由基的清除率最佳,为85.40%。研究表明,采用混合糖参与美拉德反应可显著提高泥鳅蛋白酶解产物的抗氧化活性,为泥鳅蛋白的工业化生产提供科学的理论依据。     

Nutritional and functional characterisation of hydrolysates from quinoa flour (Chenopodium quinoa) using two proteases

The objective was to study the nutritional and functional properties of hydrolysates from quinoa (Chenopodium quinoa) obtained by enzymatic hydrolysis of defatted quinoa flour (DQF). The commercial enzymes alcalase and flavourzyme were used to obtain the hydrolysates defatted quinoa flour hydrolysate with alcalase (DQFA) and defatted quinoa flour hydrolysate with flavourzyme (DQFF), respectively, after 3 h of digestion at 50°C and pH 8. The degree of hydrolysis (47.32%), yield (31.05%) and protein recovery (88.80%) values were higher in DQFA; however, its protein content (48.34%) was lower compared to that of DQFF (55.06%). Also, DQFA had a solubility greater than 57% over a wide pH range (2–10) and good foam stability (70–90%). On the other hand, DQFF presented adequate emulsifying activity (61.30 m²/g), emulsifying stability (158.62 min) and foaming capacity (131%). Due to the high content of macro- and micronutrients and adequate emulsifying and foaming properties, DQFA and DQFF could be used as ingredients in various processed food products and protein supplements.     

鹅血红蛋白抗氧化活性肽的制备及工艺研究

以新鲜鹅血为原料提取血红蛋白,利用碱性蛋白酶、中性蛋白酶及风味蛋白酶来酶解鹅血红蛋白,采用甲醛滴定法测定鹅血红蛋白水解液的水解度,通过研究水解液对DPPH自由基清除能力来检测水解产物的抗氧化活性,并分析了酶解温度、时间、pH及酶与底物比对鹅血红蛋白水解产物的影响。结果表明,采用中性蛋白酶获得的酶解产物抗氧化活性最高。在中性蛋白酶的作用下,通过单因素及正交实验,优化出产物抗氧化活性较高的酶解工艺条件:温度50℃,酶解时间8h,pH7.0,酶与底物比8000U/g,在最佳酶解条件下,产物对DPPH自由基的清除率为89.6%。      

Antioxidant and chelating activity of Jatropha curcas L. protein hydrolysates

BACKGROUND: Antioxidant and chelating activities were determined in protein hydrolysates that were produced by treating a protein isolate of a non‐toxic genotype of Jatropha curcas with the protease preparation alcalase. RESULTS: 50 min protein hydrolysate with a degree of hydrolysis of 31.7% showed highest antioxidant and chelating activity. These activities were also determined in six peptidic fractions that were separated by gel filtration chromatography of the 50 min hydrolysate. The lower‐molecular‐weight peptidic fractions had the highest antioxidant and chelating activities, which correlated with a higher content in antioxidant and chelating amino acids such as tyrosine and histidine. CONCLUSION: Results show that J. curcas represents a good source of bioactive peptides. This may be important for the revalorization of defatted J. curcas flour, a by‐product resulting form oil extraction for biodiesel production. This is especially important in Third World and developing countries such as Mexico. Copyright © 2011 Society of Chemical Industry