扫频超声处理对玉米醇溶蛋白酶解特性的影响

为了探索扫频超声技术促进蛋白质酶解反应的效果,提高酶解产物的血管紧张素转换酶(ACE)相对抑制活性,利用扫频超声处理玉米醇溶蛋白,进行单因素考察来寻找最佳扫频超声工作模式、超声预处理参数和最佳酶解条件。在最佳的超声预处理及最佳的酶解条件下,ACE抑制率为48.48%、水解度为11.20%,酶解产物的IC50值为3.77mg/mL,比常规酶解(无超声处理)降低了31.20%,产品得率为66.15%,比常规酶解提高了14.39%。这表明扫频超声预处理能够有效地促进玉米醇溶蛋白的酶解反应,提高产物的ACE相对抑制活性。      

The use of ultrasound for enzymatic preparation of ACE-inhibitory peptides from wheat germ protein

The effects of ultrasonic treatment during proteolysis on kinetic characterisation of the hydrolysis of defatted wheat germ protein (DWGP), and on ACE-inhibitory activity of the hydrolysate, were investigated. The effects of ultrasonic pretreatment on the release of peptides with ACE-inhibitory were also studied. The results showed that the value of k_A for DWGP hydrolysis under ultrasonic irradiation increased by about 22.2%, and K_M decreased about 13.0%, compared with that obtained without ultrasound. Analysis of ACE-inhibitory activity indicated that ultrasound during enzyme treatments had less effect on the ACE-inhibitory activity, while ultrasonic pretreatment caused a 21.0–40.7% increase in ACE-inhibitory activity of DWGP hydrolysate. Analyses of hydrophobicity, microstructure, and amino acid composition revealed that ultrasonic pretreatment could accelerate the release of hydrophobic amino acids from DWGP during enzymatic hydrolysis. In conclusion, ultrasonic treatment during proteolysis could facilitate the enzymatic hydrolysis of DWGP, whereas ultrasonic pretreatment could promote the release of ACE-inhibitory peptides from DWGP during enzymatic hydrolysis.     

双酶法制备沙丁鱼ACE抑制肽的工艺研究

本实验以广东大宗低值沙丁鱼为原料,采用不同蛋白酶降解沙丁鱼以筛选合适的酶组合,然后对原料的酶解制备血管紧张素转化酶(ACE)抑制肽的工艺进行研究。结果表明,制备沙丁鱼ACE抑制肽的酶组合为:木瓜蛋白酶与碱性蛋白酶;酶解的最佳工艺参数为:第一步采用木瓜蛋白酶进行酶解,酶解温度70℃、p H7.5、反应时间2.25 h、酶添加量4000 U/g、料液比30%(w/v);采用碱性蛋白酶进行第二步酶解,酶解温度70℃、p H10.0、反应时间2.0 h、酶添加量5000 U/g。通过工艺优化,沙丁鱼蛋白质的水解度提高到28.44%,并且最终水解蛋白肽的ACE抑制率为73.44%。      

双酶水解玉米醇溶蛋白制备抗氧化肽的工艺优化

为了制备抗氧化活性肽,利用Alcalase碱性蛋白酶和中性蛋白酶分步酶解玉米醇溶蛋白。在单因素的基础上,以1.1-二苯基苦基苯肼（DPPH·）自由基清除率、羟基自由基清除率和水解度（DH）为响应值,采用响应面（RSM）中心组合实验,选取Alcalase碱性蛋白酶加酶量、中性蛋白酶与Alcalase碱性蛋白酶活力之比、底物浓度为自变量,探讨最佳酶解工艺条件。采用Design-Expert软件,通过响应面优化确定修正后各因素的最佳工艺条件为:Alcalase碱性蛋白酶加酶量12880 U/（g底物）、中性蛋白酶与Alcalase碱性蛋白酶活力之比为1∶4,底物浓度为3.4%。在此修正条件下,DPPH·自由基清除率为42.98%,水解度为32.18%,与预测值的相对误差为1.04%。浓度为20 mg/m L时,玉米醇溶蛋白的DPPH·自由基清除率和羟基自由基清除率分别为同浓度VC的85.8%和67.0%。      

扫频脉冲超声辅助提取慈姑淀粉的试验研究

利用扫频脉冲超声提取慈姑淀粉,分别以淀粉提取率和纯度为评价指标,对超声时间、超声间歇比、扫频范围及超声频率等超声提取条件进行单因素实验,并在此基础上进行了提取工艺参数优化试验,得出优化的超声提取条件为:提取时间10min,频率33kHz,扫频范围±0.5kHz,间歇比30s/5s,在此条件下慈姑淀粉提取率为96%,纯度87.5%。     

超声波辅助酶解制备猪肩胛骨降血压肽的工艺及酶解效果比较

采用超声波辅助风味蛋白酶酶解制备猪肩胛骨降血压肽并与常规酶解(未经超声处理的酶解)所得酶解液做比较。以酶解液的血管紧张素转化酶(ACE)抑制率为主要指标,考察超声功率、超声时间、超声温度、超声工作间隙、超声后酶解时间对酶解液ACE抑制率的影响,并在此基础上进行响应面优化实验。通过实验,获得的最佳超声条件为:超声时间25min,超声功率717W,超声温度40℃,超声工作间隙比1∶1.5(s/s),酶解时间3h,在该条件下,得到的酶解液ACE抑制率理论值为75.29%。此条件下制备出ACE抑制率为75.58%的猪肩胛骨降血压肽,比常规酶解提高了10.27%。半数抑制浓度(IC50)值下降32.8%,酶解时间缩短1.5h。     

超声波辅助酶解制备猪肩胛骨降血压肽的工艺及酶解效果比较

采用超声波辅助风味蛋白酶酶解制备猪肩胛骨降血压肽并与常规酶解(未经超声处理的酶解)所得酶解液做比较。以酶解液的血管紧张素转化酶(ACE)抑制率为主要指标,考察超声功率、超声时间、超声温度、超声工作间隙、超声后酶解时间对酶解液ACE抑制率的影响,并在此基础上进行响应面优化实验。通过实验,获得的最佳超声条件为:超声时间25min,超声功率717W,超声温度40℃,超声工作间隙比1∶1.5(s/s),酶解时间3h,在该条件下,得到的酶解液ACE抑制率理论值为75.29%。此条件下制备出ACE抑制率为75.58%的猪肩胛骨降血压肽,比常规酶解提高了10.27%。半数抑制浓度(IC50)值下降32.8%,酶解时间缩短1.5h。      

Effect of pretreatment on enzymatic hydrolysis of bovine collagen and formation of ACE-inhibitory peptides

Bovine collagen was pre-treated (boiled or high pressure (HP)-treated) and then hydrolysed by 6 proteases. The degree of hydrolysis (DH) and the angiotensin-converting enzyme (ACE)-inhibitory activity of hydrolysates were measured. All enzymes used were able to partly degrade collagen and release ACE-inhibitory peptides. The highest ACE-inhibitory activity was obtained with Alcalase. Pretreatment significantly influenced the DH and ACE-inhibition. For most enzymes, boiling for 5 min resulted in a significantly higher DH and ACE-inhibitory activity. With Alcalase and collagenase, hydrolysis and release of ACE-inhibitory peptides occurred without any pretreatment, but HP-treatment significantly improved the DH and ACE-inhibitory activity. HP did not markedly affect the hydrolysis with the other enzymes. The major peptides obtained with Alcalase were identified; all were released from the triple helix structure of collagen. Many of these peptides had C-terminal sequences similar to known ACE-inhibitory peptides. The present results suggest that collagen-rich food materials are good substrates for the release of potent ACE-inhibitory peptides, when proper pre-treatment and enzymatic treatment is applied.     

四角蛤蜊废弃肉渣制备ACE抑制肽的工艺研究

采用胰蛋白酶水解四角蛤蜊废弃肉渣制备ACE抑制肽。以体外ACE抑制率为指标,在单因素实验的基础上,通过响应面法优化酶解工艺。结果表明,胰蛋白酶水解四角蛤蜊肉渣制备ACE抑制肽的最佳工艺条件为:酶解温度48℃,pH为8.50,胰蛋白酶加入量为0.60%,酶解时间为2h,酶解产物的平均ACE抑制率为77.08%,与预测值77.22%的相对误差为0.18%,证明响应面法对于酶解肽工艺优化具有指导意义。      

超声预处理对脱脂小麦胚芽酶解制备ACE抑制肽的影响

为了研究不同工作模式超声预处理对脱脂小麦胚芽水解度和ACE抑制率的影响,在相同能耗条件,利用聚能逆流单频、聚能逆流双频、脉冲扫频多频、发散三频、对振双频五种工作模式超声,对脱脂小麦胚芽进行预处理。同时进行单因素实验来寻找最佳超声预处理参数。结果表明,超声波预处理对水解度没有显著的影响,但可以显著提高酶解产物的ACE抑制活性,最佳的超声波工作模式为脉冲平板式40 k Hz/28 k Hz双频超声;在此模式下超声预处理单位体积超声功率60 W/L、超声时间70 min、超声初始温度60℃、底物浓度7%时得到的酶解产物IC₅₀值低至2.483 mg/m L,为较优结果,和未超声相比,其产物IC₅₀值降低了5.8%。      

Ultrasound-assisted generation of ACE-inhibitory peptides from casein hydrolyzed with nanoencapsulated protease

BACKGROUND: Bioactive peptides generated from milk proteins are eminent ingredients for functional foods and nutraceuticals. Amongst several approaches to release these peptides, hydrolysis of milk proteins with proteolytic enzymes is a promising choice. It is, however, required to inactivate the enzyme after a predetermined time, which leads to impurity of the final product. Immobilization of enzyme molecules can overcome this problem as it simplifies enzyme separation from the reaction mixture. A fungal protease from Aspergillus oryzea was encapsulated within nanoparticles yielded via silicification of polyamidoamine dendrimer template generation 0. It was used to hydrolyze the dominant milk protein (casein) in the absence or presence of sonication. The production of angiotensin converting enzyme (ACE)‐inhibitory peptides was monitored during hydrolysis. RESULTS: Sonication did not affect maximum ACE‐inhibitory activity but shortened the process sixfold. Ultrafiltration permeate of the centrifugal supernatant of casein solution hydrolyzed during sonication inhibited ACE activity as efficiently as the supernatant obtained from it. CONCLUSION: The protease from Aspergillus oryzea encapsulated within nanospheres is suitable for generation of ACE‐inhibitory peptides from casein. The nanoncapsulation procedure is simple, rapid and efficient. This may enable the industrial production of functional products from milk. Copyright © 2011 Society of Chemical Industry     

Quantitative structure-activity relationship modelling of ACE-inhibitory peptides derived from milk proteins

The potential of Lactobacillus plantarum, Pediococcus acidilactici, Pediococcus pentocaceus, Lactobacillus cellobiosus, different mixtures of these lactic acid bacteria and backslop starter cultures to acidify and form flavour compounds in uji was investigated. The bacteria chosen are the most prevalent species in fermented uji. Flavour compounds were analysed using GC-MS and GC-FID with HP5 non-polar column and DB-Wax polar columns respectively. Use of pure single or mixed cultures did not improve the flavour profile of fermented uji. On the basis of peak areas of unfermented and fermented uji aromagrams, pentanal, hexanal and hexadecanoic, 9,12-octadecadienoic, oleic and octadecanoic acids were found to be native to the flours, while 3-methyl-1-butanol, octanoate, nonanoate, hexadecanoate, linoleate, oleate and hexanoic, heptanoic, octanoic and nonanoic acids were synthesised during submerged culture fermentation. Ethanol, 1-pentanol, 1-hexanol, lactic acid and ethylacetate were synthesised prior to fermentation and synthesis of these compounds continued during fermentation.     

Primary and secondary structure of novel ACE-inhibitory peptides from egg white protein

The primary structure of novel angiotensin converting enzyme (ACE) inhibitory peptide from egg white protein was investigated, and secondary structure of the peptide was explored for the first time. The potential effects of bioactive peptides were submitted to bioactivity screening with ACE inhibitory activity, antioxidant property, and anticoagulation activity. Bioactive peptides from egg white protein were characterized by LC tandem mass spectrometric, and secondary structures of those peptides were investigated by FT-IR. Our results showed that total 11 bioactive peptides with three new and eight known structures were identified with LC/MS/MS, which then were synthesized by Fmoc solid phase method. Peptide Thr-Asn-Gly-Ile-Ile-Arg (TNGIIR) exhibited higher activity against ACE to other two new peptides. The concentration of the peptide TNGIIR, necessary to inhibit 50% the activity of ACE was 70 μM. Results also suggested that the secondary structural differences between peptides could also influence the ACE inhibition capacity. Thus, it appears that primary and secondary structure of peptide plays the potential role inhibiting the ACE activity.     

超声预处理辅助酶解玉米胚芽ACE抑制肽的研究

旨在研究不同工作模式的超声预处理对玉米胚芽蛋白酶解制备血管紧张素转换酶(Angiotensin-I Converting Enzyme,ACE)抑制肽的影响。以蛋白转化率和高活性肽占比为指标,利用聚能逆流双频、发散三频和对振双频的超声设备,对玉米胚芽粕进行预处理,得到最优的超声预处理模式;采用单因素逐级优化法来确定最佳超声预处理参数;在最优超声处理条件下,优化酶解反应条件。结果表明分子量在300~1000 Da的多肽ACE抑制活性最高,IC₅₀值为0.78 mg/mL;最优的超声模式为20/40 kHz交替双频,最佳超声预处理参数为功率密度100 W/L、底物浓度为8%、超声时间20 min、超声温度30 ℃,酶解条件为加酶量2000 U/g蛋白、酶解时间2.5 h。在最优条件下,蛋白转化率为85.00%,相比于未超声组的73.01%提高了16.42%;高活性肽占比为29.63%,相比于未超声组的26.00%提高了13.96%。因此,逆流双频超声波辅助酶解法能有效提高蛋白转化率和产物ACE抑制活性,有利于ACE抑制肽的制备。     

Effects of enzymatic treatment on anthocyanic pigments from grapes skin from chilean wine

The preparation of pectic enzymes are used for a more efficient extraction of desirable red grape pigments and other compounds which are bound in plant cells and can be faster released by the action of pectic enzymes; shorten the time of maceration, setting, and filtration. The main objective of the present study was to investigate the enzymatic extraction of anthocyanic pigments from the residue remaining after the vinification process of three varieties of Vitis vinifera from Central Chile. The best results of extraction of anthocyanins can be obtained with Vinozym EC using skin grape Ribier after 2 h of treatment.     

油茶籽粉碎程度对水酶法提油效果的影响

研究油茶籽破碎程度对水酶法提取油茶籽油的得率及提油过程形成的乳状液稳定性的影响。研究结果表明,当利用刀片式粉碎机将油茶籽平均粒径减小至37.92μm时,水酶法提取油茶籽油的总油得率、清油得率分别达到96.85%和91.83%。当对油茶籽粉继续粉碎时,茶籽粉的平均粒径基本保持不变,但水酶法提油过程中乳状液稳定性增加且乳化现象加重,导致总油得率降低,使后续破乳工艺难度加大。干法粉碎和湿法粉碎对油茶籽水酶法提取所得的毛油品质无明显区别,基本理化指标符合油茶籽油国家标准,重金属含量均在0.005mg/kg的检出限水平以下,表明刀片式粉碎方式是一种安全可行的油茶籽粉碎方式。     

超声作用下蛋白酶解产生大豆肽的比较研究

以大豆蛋白粉和碱性2709蛋白酶为原料,研究超声功率以及有无超声作用下水解温度、pH、底物浓度、加酶量对大豆蛋白水解度及水解度提高百分率的影响。结果表明,超声作用效果与超声功率有关,40kHz、128W的超声对大豆蛋白酶解产生大豆肽具有明显的促进作用;与没有超声作用的大豆蛋白酶解相比,超声作用没有改变DH与水解温度、pH、底物浓度、加酶量之间关系曲线的变化趋势,没有改变酶反应的最适温度和最适pH,两者均为温度55℃,pH10.5,但使DH明显提高。     

Production and characterisation of antioxidant peptides from sweet potato protein by enzymatic hydrolysis with radio frequency pretreatment

Effect of radio frequency (RF at 70, 80 and 90 °C) pretreatment on production and characterisation of sweet potato protein hydrolysates (SPPH) prepared using Alcalase (ALC), Protease (PRO) and the combination of ALC + PRO was investigated. RF highly improved degree of hydrolysis (DH) and enhanced antioxidant activity of all SPPH produced by ALC, PRO and the combination as compared to traditional water bath (WB) heating pretreatment (P < 0.05). RF significantly increased molecular weight (MW) <3 kDa peptide fraction from SPPH produced by ALC, and MW <3 kDa peptide fraction from RF80- and RF90-pretreated samples has higher antioxidant activity. Diverse peptides in MW <3 kDa fractions with RF80 and RF90 pretreatments were identified using LC–QTOF–MS/MS, which matched the sequences of sporamins and contained antioxidant amino acids Trp, Tyr, Met, Phe and/or His. There is a great potential application of using SPPH in functional foods as a novel ingredient.     

Potential ACE-inhibitory activity and nanoLC-MS/MS sequencing of peptides derived from aflatoxin contaminated peanut meal

Our lab has developed a process for sequestering aflatoxin from contaminated peanut meal (PM) using commercial bentonite clays while protein is simultaneously extracted and hydrolyzed by a commercial protease. The objectives of this study were to sequence generated peptides and evaluate their potential ACE-inhibitory properties. Aflatoxin in the unprocessed PM was 610 μg kg⁻¹ compared to 9.7 μg kg⁻¹ on a dry weight basis in the 120 min hydrolysate. This hydrolysate displayed significant ACE-inhibitory activity with an IC₅₀ of 295.1 μg mL⁻¹. Ultrafiltration and size exclusion chromatography (SEC) improved the ACE-inhibitory properties, with the SEC fraction containing the smallest peptides having an IC₅₀ = 44.4 μg mL⁻¹. Additionally, 271 unique peptides were identified by nanoLC-MS/MS, of which 147 belonged to major seed storage proteins. This advanced characterization data will ultimately allow for more efficient production of hydrolysates with ACE-inhibitory activity or other bioactivities of interest from PM.