具ACE抑制活性干酪乳杆菌LC-15发酵乳制备条件的研究

干酪乳杆菌LC-15发酵酸乳具有良好的血管紧张素Ⅰ转换酶（ACE）抑制效果，在“三高”现象越来越严重的今天，这种具有抗高血压活性的酸乳必将具有广阔的商业前景。以具有较高ACE抑制活性的干酪乳杆菌LC-15发酵制备酸乳，以发酵时间、发酵温度、接种量和稳定剂组合为考察因素，结合SAS响应面分析手段，通过感官评分，确定了干酪乳杆菌LC-15发酵酸乳的最佳发酵条件为接种量4％，发酵温度37℃，发酵时间14h，最适稳定剂组合为：果胶0．1％，明胶0．15％，变性淀粉0．15％。     

瑞士乳杆菌对契达干酪成熟期间所产ACE抑制肽的影响及其消化稳定性

为明确瑞士乳杆菌对契达干酪中血管紧张素转换酶（angiotensin-converting enzyme,ACE）抑制肽活性的影响,以蛋白质水解度和ACE抑制率为指标,与干酪乳杆菌组、鼠李糖乳杆菌组和空白组干酪进行对照,研究瑞士乳杆菌对干酪成熟期间蛋白质水解及ACE抑制活性的影响,并对ACE抑制活性最高时期的干酪进行消化稳定性研究。结果表明：成熟期间,3 组益生菌干酪的活菌数无明显差异（P＞0.05）,但均高于空白组;益生菌干酪的蛋白质水解程度和ACE抑制活性显著高于空白组（P＜0.05）,其中瑞士乳杆菌干酪的蛋白质水解程度最强,活性最高（79.71%）。模拟消化后,瑞士乳杆菌干酪活菌数降低14.30%,ACE抑制活性显著增加（P＜0.05）,达到86.06%,多肽质量浓度增加至2.81 mg/mL;研究不同分子质量超滤组分消化后的ACE抑制活性发现,其中大于10 kDa的多肽活性升高,小于10 kDa的活性下降。此外,添加瑞士乳杆菌不影响干酪的整体可接受性。因此,瑞士乳杆菌能促进干酪ACE抑制肽的产生并提高其活性,消化后活性的升高主要与大分子肽的降解有关。     

具有血管紧张素转换酶抑制活性的乳酸菌筛选及特性研究

血管紧张素转换酶（Angiotensin Converting Enzyme，ACE）是抗高血压药物的筛选靶位酶。以体外ACE抑制率为筛选指标，从传统发酵食品中筛选出12株具有ACE抑制活性的乳酸菌，研究了12株乳酸菌的ACE抑制活性和蛋白水解活性，以及模拟胃肠消化对乳酸菌发酵乳ACE抑制活性的影响和乳酸菌耐酸耐胆汁的特性。结果表明，CZ2112具有良好ACE抑制活性，经API鉴定系统鉴定为植物乳杆菌，ACE抑制类型为非竞争性抑制，24h发酵产物ACE抑制率可以达到83％，经模拟胃肠消化后抑制率为70％，邻苯二甲醛（OPA）法检测其蛋白水解活性为OPA指数0．43，对胃酸、胆汁具有较好的耐受性。     

具有ACE抑制活性乳酸菌菌株的筛选与鉴定

以ACE抑制活性和蛋白水解活性为检测指标,选择138株乳酸菌为出发菌株,筛选出具有强ACE抑制活性的乳酸菌菌株.结果表明,筛选出具有强ACE抑制活性的4株乳酸菌,其中3株菌为瑞士乳杆菌,1株菌为干酪乳杆菌.瑞士乳杆菌KLDS1.0485和干酪乳杆菌KLDS1.0486比例为1:1,制成的发酵乳ACE抑制活性可达到61.55%.因此,组合瑞士乳杆菌KLDS1.0485和干酪乳杆菌KLDS1.0486可作为制备乳源ACE抑制肽的优良菌株.     

牦牛乳硬质干酪体外消化前后ACE抑制肽分析

以牦牛乳硬质干酪为原料,采用胃蛋白酶和胰蛋白酶模拟体外肠胃道消化的方法,运用毛细管液相色谱结合Quadrupole-Orbitrap质谱,对牦牛乳硬质干酪体外消化前后血管紧张素转化酶(angiotensin-converting enzyme,ACE)抑制肽活性和其肽特征进行了分析。结果表明:牦牛乳硬质干酪鉴定出ACE抑制肽15种,经胃蛋白酶、胰蛋白酶降解后,其ACE抑制肽为12种。ACE抑制肽主要源自β-酪蛋白,分子质量为800～1 500 Da。相同ACE抑制肽3种。18种ACE抑制肽共享了已知ACE抑制肽C-末端的三肽氨基酸结构,大多数ACE抑制肽具备了抑制ACE活性的结构特征,它们是潜在ACE抑制肽。干酪中SKVLPVPQ、QEPVLGPVRGPFP、YQEPVLGPVRGPFPI和干酪消化后LVYPFPGPIPN、VYPFPGPIPN、YQEPVLGPVR确定具有ACE抑制活性。干酪水溶性多肽与其经胃蛋白酶和胰蛋白酶降解的酶解物的ACE抑制活性间无显著性差异(P> 0.05),这为开发具有保健功能特性的牦牛乳硬质干酪提供了理论依据。     

抗ACE活性之瑞士乳杆菌的筛选

为了筛选能发酵牛乳产生较强抗高血压活性肽的乳酸菌,本研究通过发酵乳的抑制ACE活性试验,从二种干酪制品的混合样品中分离筛选出能发酵牛乳产生较强抗ACE活性的乳杆菌1002、1004和1006。经糖发酵、生长温度、精氨酸发酵产氨、产硫化氢试验及细胞壁中二氨基庚二酸(DAP)成分分析,确认它们均为瑞士乳杆菌。     

乳酸菌发酵乳血管紧张素转化酶抑制活力的比较研究

以18种共计61株乳酸菌为研究对象,比较了乳酸菌发酵脱脂乳的血管紧张素转化酶抑制活力(ACE抑制活力),并探讨了ACE抑制活力与pH值、滴定酸度、蛋白水解度的关系。结果表明,乳杆菌ACE抑制活力最强,双歧杆菌次之,链球菌、乳球菌、明串珠菌和片球菌ACE抑制活力很低。ACE抑制活力最高的是Lacto- bacillus helveticus 6024,发酵28 h后ACEI%可达到80%以上,其OPA指数可达到0.800以上,而球菌的ACE抑制活力和蛋白水解能力都很低,甚至检测不到。乳酸菌产酸能力做为一个单一因素与ACE抑制率没有明显相关性(Spearman相关系数r_3=0.526),乳酸菌的蛋白水解能力与ACE抑制率具有显著的正相关性(Spearman相关系数r_3=0.938)。通过对2株产ACE抑制活力高的L.helveticus 6024和L.helveticus T16的研究发现,其共同特点是产酸能力和蛋白水解能力都很强。     

不同干酪乳杆菌菌株对α-葡萄糖苷酶抑制作用的比较

从14株干酪乳杆菌中筛选出11株具有凝乳作用的菌株,采用以麦芽糖酶为靶向酶的α-葡萄糖苷酶抑制剂体外筛选模型,分析了不同干酪乳杆菌发酵脱脂乳上清对α-葡萄糖苷酶的抑制作用。结果表明,不同菌株发酵脱脂乳上清对麦芽糖酶的抑制活性具有明显的菌株特异性,且随着发酵时间延长,α-葡萄糖苷酶抑制活性呈现上升趋势。其中,干酪乳杆菌LC2W在37℃发酵10%(w/w)脱脂乳96 h后所得的上清液对麦芽糖酶抑制活性最高,达37.36%,明显高于其它被测试的干酪乳杆菌菌株,表明该菌株具有潜在的抗高血糖作用。     

发酵乳中ACE抑制肽生成的外部因素条件的研究

探讨了促进发酵乳中ACE抑制肽生成的外部因素条件。研究结果表明,菌株发酵凝乳(pH4.7～5.0)后,产生肽量迅速积累,ACE抑制活性大幅度增强,产生的肽量与ACE抑制活性具有正相关性。添加乳清蛋白和酪蛋白可提高发酵乳中的肽含量,添加酪蛋白组更为突出;添加乳清蛋白组肽粉ACE抑制活性减弱,而添加酪蛋白组肽粉ACE抑制活性增强。与发酵温度42℃相比,37℃发酵条件下有利于发酵乳ACE抑制肽生成;与4℃冷藏处理相比,37℃条件下保温培养对提高ACE抑制活性和肽含量更为显著(P<0.01),实验结果为发酵法生产ACE抑制肽提供了可靠的技术依据。     

瑞士乳杆菌发酵乳制备ACE 抑制肽的条件优化

本研究以脱脂乳为主要原料,对瑞士乳杆菌发酵产生血管紧张素转化酶(angiotensin I-converting enzyme,ACE)抑制肽工艺条件进行研究。通过单因素试验和响应面试验设计,确定瑞士乳杆菌发酵脱脂乳生产ACE 抑制肽的最佳工艺条件为：脱脂乳浓度9.67%(m/V)、底物灭菌时间30min、接种量3%(V/V),温度38.82℃、发酵时间6.26h,测得ACE 抑制率为92.26%。研究结果证实利用发酵法生产乳源ACE 抑制活性肽对控制高血压具有重要意义。     

产ACE抑制肽乳酸菌的筛选、益生特性评定及应用

目的:筛选出富产血管紧张素转化酶(Angiotensin converting enzyme,ACE)抑制肽的乳酸菌并评定其益生特性.方法:依据蛋白水解度及ACE抑制率对乳酸菌进行初筛,然后依据模拟胃肠消化后的ACE抑制率最终筛出2株乳酸菌ZJUIDS09和ZJUIDS11,进一步评价其耐酸、耐胆盐、抗生素耐药性和抑菌...     

Effect of simulated gastrointestinal digestion on the antihypertensive properties of synthetic beta-lactoglobulin peptide sequences

In this study, the antihypertensive activity in spontaneously hypertensive rats of two peptides isolated from beta-lactoglobulin hydrolysates with thermolysin was evaluated. These peptides, with sequences LLF [beta-lg f(103-105)] and LQKW [beta-lg f(58-61)], showed potent in vitro ACE-inhibitory activity. Two hours after administration, both sequences caused a clear and significant decrease in the blood pressure of these rats. The impact of a simulated gastrointestinal digestion on ACE-inhibitory and antihypertensive activities of these peptides was also studied. The results showed that both fragments were susceptible to proteolytic degradation after incubation with pepsin and Corolase PP. In addition, their in vitro ACE-inhibitory activity decreased after the simulated digestion. It is likely that fragment LQK was the active end product of the gastrointestinal digestion of peptide LQKW. The fragment LL, observed after digestion of peptide LLF, probably exert its antihypertensive effect through a mechanism of action different than ACE-inhibition.     

Antihypertensive activity of casein-enriched milk fermented by Lactobacillus helveticus

Milk proteins contain peptidic angiotensin I-converting enzyme (ACE) inhibitors, which can be released by proteolysis during milk fermentation by some strains of Lactobacillus helveticus. Reconstituted milk media containing skim milk powder (12%), skim milk powder (10%) with added sodium caseinate (5%) or whey protein isolate (5%) were fermented by L. helveticus strains R211 and R389, and further tested for bacterial growth, proteolysis (free NH₃ groups) and ACE-inhibitory activity. The antihypertensive activity in spontaneously hypertensive rats (SHR) was also investigated with caseinate-enriched milk unfermented (UFM) and fermented by the two strains of L. helveticus. Caseinate-enriched milk fermented by both strains showed higher proteolysis and ACE-inhibitory activity, indicating that ACE-inhibitory peptides are probably released from caseins during milk fermentation. Significant decreases in mean arterial blood pressure in SHR rats were measured following oral administration of UFM milk at doses of 1.0 and 2.5 g kg⁻¹ of body weight, and milk fermented by R211 or R389 strains at doses of 0.5, 1.0 and 2.5 g kg⁻¹ of body weight. The antihypertensive activity of UFM could be explained by the release of ACE-inhibitory peptides from caseins during the digestion process.     

Initial proteolysis of milk proteins and its effect on formation of ACE-inhibitory peptides during gastrointestinal proteolysis: a bioinformatic, in silico, approach

Angiotensin-I-converting enzyme (ACE)-inhibitory peptides are encrypted in milk protein sequences. They may express an antihypertensive effect if they are released by proteolysis in foods and/or during gastrointestinal digestion. A bioinformatic, in silico, approach was developed to evaluate how systematic initial proteolysis, i.e. cleavage after one specific type of amino acid (C-end) at a time in milk proteins, influence the formation of ACE-inhibitory peptides by subsequent gastrointestinal proteolysis. Computer simulation was done and a peptide QSAR model was used to estimate the combined ACE inhibition by digested proteins. Initial proteolytic cleavage at the C-end of amino acids isoleucine and proline gave, based on calculations, increased effect of ACE-inhibitory peptides after gastrointestinal proteolysis of milk proteins. Cleavage after most other amino acid residues had little or no effect. Results indicate that initial proteolysis in foods have to be specific in order to increase formation of bioavailable ACE-inhibitory peptides during gastrointestinal digestion.     

Angiotensin-converting enzyme inhibitory activity of peptides derived from caprine kefir

In this study, a potent angiotensin-converting enzyme (ACE)-inhibitory activity was found in a commercial kefir made from caprine milk. The low molecular mass peptides released from caseins during fermentation were mainly responsible for this activity. Sixteen peptides were identified by HPLC-tandem mass spectrometry. Two of these peptides, with sequences PYVRYL and LVYPFTGPIPN, showed potent ACE-inhibitory properties. The impact of gastrointestinal digestion on ACE-inhibitory activity of kefir peptides was also evaluated. Some of these peptides were resistant to the incubation with pepsin followed by hydrolysis with Corolase PP. The ACE-inhibitory activity after simulated digestion was similar to or slightly lower than unhydrolyzed peptides, except for peptide β-casein f(47-52) (DKIHPF), which exhibited an activity 8 times greater after hydrolysis.     

Angiotensin I-converting enzyme inhibitory activity in milk fermented by wild-type and peptidase-deletion derivatives of Lactobacillus helveticus CNRZ32

Development of molecular methods has enabled detailed studies of the proteolytic system of Lactobacillus helveticus, which has a central role in the release of bioactive peptides during the fermentation of milk. The impact of general aminopeptidase (PepN) and X-prolyl dipeptidyl aminopeptidase (PepX) activity of L. helveticus on the level of angiotensin I-converting enzyme (ACE) inhibitory activity in fermented milk was elucidated by taking advantage of peptidase-negative derivatives of L. helveticus CNRZ32. According to the results, increased level of ACE-inhibitory activity was attained in milk fermented by the peptidase-deficient mutants. The ACE-inhibitory activity determined with the PepN-deficient strain decreased towards the end of fermentation, suggesting that PepN gives rise to a provisional accumulation of ACE-inhibitory peptides before their hydrolysis to shorter peptides or free amino acids. The ACE-inhibitory activity determined with the PepX-deficient strain increased throughout the fermentation, indicating specific blocking of the further hydrolysis of ACE-inhibitory peptides.     

益生菌发酵乳模拟胃肠液环境下抗氧化活性

为间接评价益生菌发酵乳在人体内的抗氧化活性,分别采用嗜热链球菌（Streptococcus thermophilus）、德氏乳杆菌保加利亚亚种（Lactobacillus delbrueckii subsp. bulgaricus）、动物双歧杆菌（Bifidobacterium）BB12及嗜酸乳杆菌（Lactobacillus acidophilus）制备SL、SL+BB12、SL+LA三种发酵乳,并对其冷藏过程中模拟胃肠液环境下对ABTS、DPPH自由基的清除能力进行了测定。结果表明,三种发酵乳随冷藏时间的延长其抗氧化活性都逐渐降低,添加菌种BB12和LA的发酵乳抗氧化活性下降的幅度小于SL发酵乳,在相同的冷藏时间下模拟胃肠液环境后抗氧化活性都较原发酵乳降低,但添加益生菌BB12和LA发酵乳在模拟胃肠液环境下抗氧化活性高于SL发酵乳,其中BB12高于LA。SL、SL+BB、SL+LA发酵乳冷藏7 d时对ABTS自由基最大清除率分别为46.21%、58.54%、51.99%,对DPPH自由基最大清除率分别为42.17%、53.34%、49.48%。     

Angiotensin I converting enzyme-inhibitory activity of bovine, ovine, and caprine kappa-casein macropeptides and their tryptic hydrolysates

This work evaluated the angiotensin-converting enzyme (ACE)-inhibitory activities of bovine, ovine, and caprine kappa-casein macropeptides (CMPs) and their tryptic hydrolysates. The results obtained indicate that bovine, ovine, and caprine CMPs exhibited moderate in vitro ACE-inhibitory activities that increased considerably after digestion under simulated gastrointestinal conditions. Active peptides could also be produced from CMPs via proteolysis with trypsin, with tryptic hydrolysates exhibiting a more extensive ACE-inhibitory activity than intact CMPs during simulated gastrointestinal digestion. Two active fractions were chromatographically separated from the tryptic hydrolysate of the bovine CMP, but their complexity hampered the assignment of the ACE-inhibitory activity to specific peptide sequences. Evidence for the release of the strong ACE-inhibitory tripeptide IPP was found upon simulation of the gastrointestinal digestion of peptides released by trypsin from the CMP sequence. These findings might help to promote further exploitation of cheese whey in the preparation of nutraceuticals for inclusion in the composition of functional food products with high added values.     

超高压处理对消减乳源酪蛋白抗原性影响的体外模拟研究

目的:研究超高压结合不同模拟胃肠消化对于消减乳源酪蛋白抗原性的影响。方法:以乳源酪蛋白为研究对象,采用不同的压力、温度、保压时间、超高压次数来处理酪蛋白,并对处理后的酪蛋白进行模拟成人、婴儿胃肠消化,用间接酶联免疫吸附法来检测经过处理后酪蛋白过敏原消减效果,并且使用正交设计确定消减酪蛋白过敏原最优处理条件组合。结果:在相同超高压处理条件下,对酪蛋白进行模拟成人、婴儿胃肠消化环境,其结果显示成人比婴儿胃肠消化后酪蛋白过敏原消减的效果更好;高压结合模拟成人胃肠消化的最佳条件组合是:350 MPa、温度30℃、保压时间15 min、高压3次,抑制率为22.67%;高压结合模拟婴儿胃肠消化的最佳条件组合是:350 MPa、温度30℃、保压时间20 min、高压1次,抑制率为39.95%。结论:高压结合不同胃肠消化后都可以消减酪蛋白过敏反应,而且结合成人胃肠消化比结合婴儿胃肠消化对过敏原的消减效果更好。