酶解鹿茸血ACE抑制活性及其与抗氧化活性关系的研究

实验采用木瓜蛋白酶和Alcalase碱性蛋白酶水解鹿茸血,在不同的时间取样,分别测定血管紧张素转化酶(ACE)抑制率、二苯代苦味肼基自由基(DPPH.)清除率和羟自由基(.OH)清除率。结果表明Alcalase蛋白酶水解产物ACE抑制活性和DPPH.清除活性较强,其水解3h的产物ACE抑制率为90.63%,DPPH.清除率为14.40%;木瓜蛋白酶水解5h的产物.OH清除率最高,为96.7%。用SPSS10.0软件分析结果,发现ACE抑制率与DPPH.清除率具有显著相关性。     

不同来源家蝇幼虫蛋白粉的水解与抗氧化性

本研究比较不同来源、不同酶解物浓度的家蝇幼虫蛋白及其抗氧化活性。以来自猪粪源和鸡粪源家蝇幼虫为原料制备蛋白粉,用胰蛋白酶水解蛋白粉制备水解产物,以·OH和O_2~-·清除率为指标评价两种蛋白酶解物的抗氧化活性。结果显示,家蝇幼虫蛋白水解物在4 h达水解度最高值,其水解物的·OH清除率在5 h达到最大为59.02%,其水解物的O_2~-·清除率在4 h达到最大为73.24%;随着两种水解物浓度增加,其抗氧化效果越强,当猪粪源和鸡粪源家蝇幼虫酶解物浓度为10 mg/mL时,两者O_2~-·清除率达到最大分别为70%和80%;·OH清除率分别为45%和52%。由此可见,家蝇幼虫酶水解物对超氧阴离子、羟自由基均有清除能力,鸡粪源家蝇幼虫酶解物优于猪粪源。     

丝素肽的制备及其抗氧化活性研究

本文以废蚕丝为原料,通过脱胶精炼、碱性蛋白酶Alcalase2.4降解后得到丝素肽溶液,并采用超氧自由基(O-·2)体系、羟基自由基(·OH)体系、二苯代苦味酰基自由基(DPPH·)体系考察了不同水解度丝素肽体外清除自由基的活性。结果表明,丝素蛋白溶液在底物浓度5%,加酶量([E]/[S])2%,反应温度60℃,p H值8.5时,通过控制反应时间可获得水解度为0~23.92%的丝素肽,在此范围内丝素肽清除自由基的能力随水解度的提高而不断增强,在水解度为23.92%时,丝素肽对超氧自由基(O-·2)的清除率达73.69%,对羟基自由基(·OH)的清除率为82.49%,对二苯代苦味酰基(DPPH·)的清除率为70.79%,抗氧化活性最好。     

米糠蛋白抗氧化活性肽的制备

以水解度(DH％)和对DPPH自由基清除率为指标,筛选出制备米糠蛋白抗氧化活性肽的最适蛋白酶.研究最适蛋白酶的酶解条件,探讨底物浓度、蛋白酶的加入量、pH值、酶解时间等因素对水解度(DH％)和DPPH自由基清除率的影响;在单因素基础上采用Box-Behnken响应曲面中心组合设计法,对酶解米糠蛋白的工艺进行优化.试验结果表明,在加酶量13970.82 U/g,时间3.05h,底物浓度4.97％的水解条件下,米糠蛋白的水解度能够达到23.67％,活性肽对DPPH自由基清除率达到64.26％.     

球孢白僵菌不同菌株清除自由基活性评价

选择不同来源的12株球孢白僵菌,分别测定菌丝体2种溶剂提取物在不同抗氧化模型中的抗氧化作用。结果表明,不同溶剂提取物对二苯基苦味酰基苯肼自由基(DPPH)和羟自由基(.OH)均有清除作用,但是清除自由基的活性不同,乙酸乙酯提取物对.OH的清除作用最为显著,活性较高,最高清除率达到71.54%。甲醇提取物活性均较低,30 min时甲醇提取物对DPPH和.OH清除率最高分别为36%和38.96%,12株白僵菌间的清除自由基活性有较大的差异。     

马鹿茸血免疫活性肽的制备及其活性研究

本文以免疫活性和DPPH·的清除能力为指标,研究了用木瓜蛋白酶与中性蛋白酶水解马鹿茸血制备活性肽的条件,并初步探讨了此活性肽的免疫活性与对DPPH·清除能力之间的关系.实验结果表明:当[E/S]为9000 U/g时,中性蛋白酶和木瓜蛋白酶的水解温度各为50和45 ℃,pH各为7.1和6.8,分别水解2和1.5 h可以获得最佳的酶解效果.在相同蛋白含量的情况下,两酶复合水解物的淋巴细胞增殖率比单酶水解增加了20.8%.免疫活性与DPPH·清除能力之间存在一定的相关性(r=0.957,P<0.01).     

壳聚糖季铵盐合成及其抗氧化性能研究

通过Eschweiler Clarke反应得到N,N-二甲基壳聚糖中间体。然后与碘甲烷反应生成水溶性的壳聚糖季铵盐:N,N,N-三甲基壳聚糖碘化铵(TMCI)。检测三种不同取代度的季铵盐产品对羟基自由基(.OH)、超氧自由基(O2-.)的清除率。结果发现:取代度为33.9%时,对.OH的最大清除率为40.2%,对O2-.的最大抑制率为65.6%。并且季铵盐产品对两种自由基的清除率都随着取代度的增大而减小,取代度为80.3%的产品对这两种自由基几乎无清除作用。     

固态发酵苦荞制备多肽菌种的筛选

【目的】筛选固态发酵苦荞高产多肽及发酵产物液具有抗菌、抗氧化活性的菌株。【方法】采用米曲霉、酱油曲霉、雅致放射毛霉和少孢根霉分别对苦荞进行固态发酵,以蛋白酶活力、水解度、可溶性肽得率、抑菌率和体外自由基清除率作为筛菌指标。【结果】米曲霉固态发酵苦荞的可溶性肽得率最高达38.83%±1.18%,发酵产物液对大肠杆菌和金黄色葡萄球菌的抑菌率分别为96.62%±1.66%和97.54%±0.54%,同时羟自由基(·OH)清除率和二苯基苦味酰基苯肼自由基(DPPH·)清除率分别为55.65%±1.25%和10.84%±1.03%。对米曲霉发酵2 d发酵产物液的不同分子量分布及活性分析表明,分子量大小对抗菌及抗氧化活性有一定的影响。【结论】米曲霉可作为固态发酵苦荞制备多肽且发酵产物液具有抗菌及抗氧化活性的最佳菌株,并在多肽产量提升及抗菌、抗氧化活性的研究上具有巨大空间。     

金银花尺蠖幼虫粪便乙醇提取物体外抗氧化活性

自由基过量累积会影响生物体正常机能,筛选具有自由基清除功能的昆虫代谢产物对于开发虫茶至关重要。为了解金银花尺蠖幼虫粪便提取物的抗氧化活性,在室内测定了金银花尺蠖幼虫粪便乙醇提取物的体外抗氧化活性。结果表明,金银花尺蠖幼虫粪便乙醇提取物具有一定的体外抗氧化活性:对二苯代苦味酰自由基(DPPH)、羟基自由基(·OH)、超氧阴离子自由基(O-·2)、ABTS+·自由基均具有一定的清除能力,在0.05mg/m L时的清除率分别为37.78%、25.31%、21.41%和35.82%;随着提取物浓度的增大,对自由基的清除率逐渐增加,当浓度上升到0.65 mg/m L时的清除率分别为95.52%、69.07%、63.27%和72.46%,分别增加了57.74%、43.76%、41.86%和36.64%,差异达显著水平;对卵黄脂蛋白脂质过氧化也具有一定的抑制作用,在0.05 mg/m L时的抑制率为26.78%,随着提取物浓度的增大,对脂质过氧化的抑制率也逐渐增加,当浓度上升到0.65 mg/m L时的抑制率为50.63%,增加了23.85%,差异达显著水平。在0.05~0.65 mg/m L的浓度范围内,对自由基的清除率、对脂质过氧化的抑制率与浓度之间呈显著的线性关系。综上,金银花尺蠖幼虫粪便提取物具有开发为虫茶的潜力。     

大豆分离蛋白不同酶解方式水解度与乳化性和起泡性关系

研究了大豆分离蛋白酶解产物水解度与乳化和起泡功能特性的关系。试验结果表明:大豆分离蛋白酶解产物起泡性随着水解度的增加而增加,不同酶处理液的起泡性以AS1.398酶解液最好,水解度为20%时起泡性达到了360±2.46%。研究还发现,乳化性随着水解度增加而逐渐下降,以双酶复合酶解液最差(DH=25%时乳化性最差,17.60±0.80m l.g-1)。     

Protein hydrolysates from Bluefin Tuna (Thunnus thynnus) heads as influenced by the extent of enzymatic hydrolysis

Functional properties and antioxidant activities of protein hydrolysates from tuna (Thunnus thynnus) heads (THPHs), with different degrees of hydrolysis, obtained by treatment with Bacillus mojavensis A21 alkaline proteases and Alcalase, were investigated. Protein content of all freeze-dried THPHs ranged from 73.74 ± 0.5 to 78.56 ± 1.2%. The THPHs had excellent solubility, compared to untreated tuna head proteins and possessed interfacial properties, which were governed by their concentrations. Similarly, at a degree of hydrolysis (DH) of 12 and 15%, > 90% nitrogen solubility was observed at all experimental pH values tested. The emulsifying activity index (EAI) and emulsion stability index (ESI) of both hydrolysates at different DHs decreased (p < 0.05) with increasing DH. At low DH (5%), hydrolysates exhibited strong emulsifying properties. All THPHs produced by the A21 proteases generally showed higher antioxidative activity than that of the Alcalase protein hydrolysates. The highest DPPH radical-scavenging activity (78 ± 2.1% at 3 mg/mL) was obtained with a DH of 15%. The IC₅₀ value for the β-carotene bleaching assay was 0.5 ± 0.03 mg/mL. Alcalase (DH = 12%) and A21 (DH = 15%) protein hydrolysates contained glutamic acid/glutamine and arginine as the major amino acids, followed by lysine, aspartic acid/ asparagine, histidine, valine, phenylalanine, and leucine. In addition, the THPHs had a high percentage of essential amino acids, which made up 50.52 and 50.47%, of the protein hydrolysates obtained by the Alcalase and A21 proteases, respectively. Therefore, THPHs can be used as a promising source of functional peptides with antioxidant properties.     

Optimization of Enzymatic Hydrolysis Conditions of Caspian kutum (Rutilus frisii kutum)ˮ By-product for Production of Bioactive Peptides with Antioxidative Properties

The enzymatic hydrolysis was performed by Alcalase to recover the fish protein hydrolysate from Caspian kutum by-product (CB). The degree of hydrolysis (DH) was applied for monitoring the hydrolysis reaction of CB. The response surface methodology was applied based on a D-optimal design to perform the optimization process for obtaining the high yield of CB protein hydrolysate. The effect of four independent variables including pH (7.5–8.5), temperature (45–55 °C), time (1–3 h), and enzyme concentration (0.5–1.5% w/w) on DH was studied. The results indicated that the predicted and actual values of the optimum condition had no significant difference. The optimum enzymatic hydrolysis conditions were achieved at pH 8.5, temperature of 55 °C, enzyme concentration of 1.5% w/w, and time of 3 h, which resulted in the maximum value of DH (19.08%). Antioxidant assays including DPPH scavenging and metal chelating activities showed that Caspian kutum protein hydrolysates had antioxidant properties.

     

Opposite Contributions of Glycinin- and β-Conglycinin-Derived Peptides to the Aggregation Behavior of Soy Protein Isolate Hydrolysates

The aggregation behavior as a function of pH was studied for hydrolysates obtained by hydrolysis of soy protein isolate (SPI) and glycinin- and β-conglycinin-rich protein fractions with subtilisin Carlsberg. The substrates were hydrolyzed up to degrees of hydrolysis (DH) of 2.2% and 6.5%. Compared with nonhydrolyzed SPI, a decrease in solubility was observed for the hydrolysates of SPI [0.8% (w/v) protein, I = 0.03 M] around neutral pH. At pH 8.0, glycinin hydrolysates had a much lower solubility (∼43% and 60%, respectively, for DH 2.2% and 6.5%) than SPI and β-conglycinin-derived hydrolysates, which were almost completely soluble. Peptides that aggregated were all larger than 5 kDa, and as estimated by size-exclusion chromatography their composition was almost independent of the aggregation pH. The solubility of hydrolysates of SPIs with a varying glycinin and β-conglycinin composition showed that glycinin-derived peptides are the driving force for the lower solubility of SPI hydrolysates. The solubility of SPI hydrolysates at pH 8.0 was shown not to be the sum of that of glycinin and β-conglycinin hydrolysates. Assuming that the separate hydrolysis of glycinin and β-conglycinin did not differ from that in the mixture (SPI), this indicates that β-conglycinin-derived peptides have the ability to inhibit glycinin-derived peptide aggregation.     

Structural characteristics of hydrolysates of proteins from extracted sunflower flour at the air-water interface

The structural and topographical characteristics of a sunflower protein isolate (SPI) and its hydrolysates at different degrees of hydrolysis (DH = 5.62%, 23.5%, and 46.3%) spread at the air-water interface at pH 7 and 20 degrees C were determined from pi-A isotherms coupled with Brewster angle microscopy (BAM). The structural characteristics of SP hydrolysate spread monolayers depend on the degree of hydrolysis. We observed a significant shift of the pi-A(APPARENT) isotherms toward lower molecular areas as the degree of hydrolysis (DH) increased. This phenomenon was attributed to spreading of the protein at the interface, especially at DH 46.3%. A change in the monolayer structure was observed at a surface pressure of 12-15 mN/m. At a microscopic level, the heterogeneous monolayer structures visualized near the monolayer collapse and during the monolayer expansion proved the existence of large regions of protein aggregates. Reflectivity increased with surface pressure and was a maximum at the monolayer collapse. The monolayer thickness decreased as the degree of hydrolysis increased. These phenomena explain the poor functional properties for the formation and stabilization of a dispersion (emulsion or foam) of protein hydrolysates at high degrees of hydrolysis.     

Influence of the rapeseed protein hydrolysis process on CHO cell growth

Different protein hydrolysates were prepared from enzymatic hydrolyses of a rapeseed isolate (>90% protein content) using different commercial enzymes of non-animal origin. The extent of hydrolysis was controlled to produce hydrolysates corresponding to various degrees of hydrolysis (DH) from 5 to 30. These hydrolysates were characterized according to their solubility and size peptide pattern. Different growth behaviours of Chinese Hamster Ovary cells were observed when these various hydrolysates were added in serum-free medium containing transferrin, albumin and insulin. Hydrolysates from low degree of hydrolysis generally did not exhibit significant positive effect on cell growth; conversely hydrolysates from extensive hydrolysis, corresponding to a major low molecular size peptides content, usually allowed an increase of the maximal cell density. However, depending on the enzyme used, the supplementation with hydrolysates corresponding to a high degree of hydrolysis and composed of at least 70% peptides with a molecular size under 1kDa, led to different maximal cell density values, indicating the importance of enzyme specificity and consequently the nature of the released peptides. This result showed that the positive influence of the rapeseed hydrolysates on cell growth was not only due to a nutritional support tied to the addition of small peptides but may be related to the presence of peptides exhibiting growth or survival factor effects. Furthermore, total substitution of proteins (transferrin, albumin and insulin) in the cell culture medium by some rapeseed hydrolysates appeared to be a promising alternative to improve the cell growth in protein-free media.     

Kozak序列对金黄色葡萄球菌黏附因子FnBPA-A DNA疫苗诱导小鼠免疫应答的影响

黏附因子FnbpA(纤连蛋白结合蛋白A,Fibronectin binding protein A)是金黄色葡萄球菌表面的蛋白质成分,是该菌感染早期最重要的致病因子,可促进其对寄主组织的侵入,也是一个有潜力的免疫靶标。将牛乳源金黄色葡萄球菌中FnBPA基因的A功能区克隆至真核表达载体中,分别构建含Kozak序列和不含Kozak序列的FnBPA-A基因真核表达载体,重组质粒经鉴定测序正确后,免疫C57BL/6小鼠检测其抗体水平和淋巴细胞增殖情况,并对各组小鼠进行攻毒实验。检测的结果表明Kozak修饰的重组DNA在血清抗体效价(P<0.05)和免疫保护率方面均优于不含Kozak序列的重组DNA,在刺激淋巴细胞增殖方面Kozak修饰的重组DNA的刺激效果虽然也高于不经修饰的重组DNA,但是差异不显著(P>0.05)。根据总体的免疫效果来看,Kozak序列对增强FnBPA的重组DNA疫苗诱导的免疫应答起了不容忽视的作用。     

马鹿茸血酶解肽体内免疫功能及抗氧化功能关系的研究

本文分别选用由木瓜蛋白酶水解天山马鹿茸血获得的肽Ⅰ、肽Ⅱ以及原血为受试样品,研究天山马鹿茸血及其酶解肽对小鼠免疫功能和抗氧化功能的影响,其中肽Ⅰ、肽ⅡDH分别为18．1％、12．2％时,清除OH&#183;能力最强,清除率为50．8％。分别测定了脏器指数,脾细胞增殖实验,血清中溶血素和抗体生成细胞的含量以及小鼠血清总抗氧化能力、SOD活力和MDA含量。结果表明：与对照组相比较,肽Ⅱ组能极显著提高小鼠体液和细胞免疫功能,加强小鼠的抗氧化功能（P〈0．01或P〈0．05）。此外,具有较强抗氧化活性的肽显示出了很强的免疫活性（P〈0．05）。     

Hydrolysis of rapeseed protein isolates: Kinetics,characterization and functional properties of hydrolysates

Two rapeseed protein isolates corresponding to albumins and globulins, respectively, were produced from an industrial defatted rapeseed meal. A pilot-scale process of protein extraction has been developed to remove major anti-nutritional compounds, easy to scale-up and using recyclable solvents. The kinetics of the hydrolyses of these two protein substrates using Alcalase 2.4L^® were compared by the measurement of the degree of hydrolysis (DH) when varying the initial proteins and enzyme concentrations. The globulins isolate was hydrolysed more efficiently than the albumins isolate mainly due to the compact and globular conformation of albumin (napin). Kinetic parameters have been determined for both substrates and a log-linear relation has been established between the DH values at a definite time and the initial enzyme/substrate ratio. Such relationships allow an effective monitoring of hydrolysis process since the hydrolysates analysis using reverse-phase chromatography coupled with mass spectrometry revealed that peptide maps corresponding to peptides of molecular weight inferior to 1 kDa are similar at a specific DH, independently of the reaction temperature and initial concentrations of substrate and enzyme. Thus, it is demonstrated that the DH is the sole parameter needed to control the physico-chemical properties and consequently the functionalities (solubility, foaming and emulsifying properties) which depend on the nature of peptides present in the hydrolysates.