传统发酵酸马奶中ACE抑制成分的分离

研究了从传统发酵酸马奶中分离ACE抑制成分,即酸马奶经热处理、分离乳清、截留膜超滤和凝胶柱层析分离后得到ACE抑制成分,并逐步测定ACE抑制活性和肽质量浓度.酸马奶乳清经超滤膜截留分离得到3个组分(Ⅰ、Ⅱ和Ⅲ),其中分子量小于3 ku组分(Ⅲ)的ACE抑制效率最高((2.257±0.028)L/g),再经凝胶柱层析分离得到6个组分(A-F),其中组分C的ACE抑制效率最高((6.3852±0.0728)L/g).结果表明,传统发酵酸马奶中富舍ACE抑制成分,为酸马奶的降血压功能及其进一步研发提供科学依据和实验基础.     

大豆肽的大孔吸附树脂以及凝胶过滤色谱分离

采用DA201．C型大孔吸附树脂对Alcalase水解。DH14％的大豆肽进行脱盐和乙醇分布洗脱，不同浓度乙醇洗脱物的氨基酸组成分析结果表明：洗脱是按照疏水性递增的方式进行的。乙醇浓度为75％时洗脱下来的组分都具有最高的降血压活性，其ACE抑制率为56．52％。体外模拟实验测定结果表明：经过胃肠道酶的作用，大豆肽的ACE抑制活性仍达到49．52％。采用1M的醋酸作为洗脱液可以实现75％乙醇洗脱组分的Sephadex G-15有效分离，其中ACE抑制活性最强组分的抑制率为69．57％．IC50为0．144mg／mL。     

花生ACE抑制肽的分离纯化、结构鉴定及体内降血压功能研究

在水酶法提取花生油和水解蛋白的中试生产基础上,将中试生产得到的水解蛋白通过DA201-C大孔吸附树脂、葡聚糖凝胶Sephadex G-15色谱分离纯化制得花生ACE抑制肽Ⅰ,再经半制备和分析型RP-HPLC进一步分离纯化后.利用基质辅助激光解吸电离-飞行时间-飞行时间(MALDI-TOF-TOF)串联质谱对具有最高ACE抑制活性的组分进行结构鉴定,得到氨基酸序列为Pro-Gly-Arg-Val-Tyr的花生ACE抑制肽Ⅱ.按照该结构合成得到较大量的花生ACE抑制肽Ⅱ,与花生ACE抑制肽Ⅰ-起作为受试样品进行SHR大鼠体内降血压功能实验.实验证明,花生ACE抑制肽在体内确实具有降低血压的作用,与降血压药物相比其服用剂量较大,但安全性相对较高.     

三斑海马蛋白ACE抑制肽的制备及其二级结构的研究

以三斑海马为原料,经碱性蛋白酶酶解,获得富含血管紧张素转化酶(angiotensin I-converting enzyme,ACE)抑制肽的酶解液。酶解液经透析、Sephadex G-25凝胶过滤色谱和反相高效液相色谱得到进一步分离纯化。结果表明,透析产物的IC50值为(0.44±0.26)mg/mL,相比酶解液(0.81±0.12)mg/mL,其ACE抑制活性更强。透析产物经Sephadex G-25凝胶柱分离纯化后,得到3种组分,其中组分F2的ACE抑制活性最强,IC50值为(0.11±0.08)mg/mL。F2经反相高效液相色谱进一步分离后,得到6个具有ACE抑制活性的组分峰,其中组分F2-4的ACE抑制活性最强,IC50值为(0.005 7±0.000 9)mg/mL。经过3步分离纯化后,成功从三斑海马蛋白中分离得到一种活性较强ACE抑制肽:ProAla-Gly-Pro-Arg-Gly-Pro-Ala(PAGPRGPA),多肽分子量为721.39 Da。圆二色谱分析多肽二级结构表明其主要含无规则卷曲。因此,从三斑海马蛋白中分离得到的多肽可能成为营养保健品和抗高血压药品及相关疾病的一种有益成分,且对海马蛋白资源的开发利用具有重要意义。     

三斑海马蛋白肽ACE抑制活性的研究

本文通过三斑海马酶解液的制备、ACE抑制活性肽的分离和体外消化模型的建立研究了三斑海马蛋白肽的ACE抑制活性。利用碱性蛋白酶制备得到具有ACE抑制活性的三斑海马酶解液,经葡聚糖凝胶层析分离纯化后,得到具有较高ACE抑制活性的组分(其IC50为0.91 mg/m L);通过对该组分的氨基酸组成和体外消化模型分析,发现该组分中疏水性氨基酸的含量为50.48%,消化酶作用后,显著提高ACE抑制活性,经判断,该组分中的蛋白肽为前体型抑制剂。     

猪血红蛋白ACE抑制肽的分离和理化性质研究

以猪血红蛋白的胃蛋白酶水解液为原料,依次通过大孔吸附树脂DA201-C、葡聚糖凝胶Sephadex LH-20进行纯化,最终得到了3个降血压组分;并收集ACE抑制活性最高的组分α-Ⅱ,对其进行理化性质的研究.结果表明,酶解液经过大孔树脂DA201-C脱盐后,IC50为0.87mg/mL;继续经过葡聚糖凝胶Sephadex LH-20分离得到的猪血红蛋白ACE抑制肽(α-Ⅱ)的IC50为0.21mg/mL.分离后的猪血红蛋白ACE抑制肽(α-Ⅱ)在pH为7时,溶解度最低,为50.37％;胃蛋白酶对其活性影响不显著.温度在25～55℃之间以及中性和偏酸性的条件下时,α-Ⅱ具有较好的热稳定性;当NaC1浓度大于0.6mol/L时,其活性急剧下降.     

鱼鳞明胶ACE抑制肽的制备及其活性研究

利用Alcalase 2.4L酶解鱼鳞明胶制备具有血管紧张素转化酶(ACE)抑制活性的降血压肽,通过超滤处理富集得到具有较高活性的ACE抑制肽,并对该组分进行分子量分布、氨基酸组成分析及抗消化性研究。结果表明:水解明胶6 h所得水解物的ACE抑制活性最高,其IC50为0.56 mg/mL,水解度为15.54%;超滤膜处理分离后,分子量小于5 kDa的多肽组分Ⅱ的ACE抑制活性最高,其回收率达90%以上;多肽中对ACE抑制活性贡献大的脯氨酸、羟脯氨酸、色氨酸等疏水性氨基酸的保存率高;体外消化实验显示,该抑制肽在胃肠道消化酶作用下能保持较好的ACE抑制活性。     

酪蛋白源ACE抑制肽的分离纯化

采用碱性蛋白酶酶解酪蛋白制备ACE抑制肽.通过正交实验得到最佳酶解条件为:温度50℃,pH值7.5,酶用量5%,底物质量分数5%,在此条件下,ACE抑制活性为47.23%.采用超滤对酶解液进行初步分离,得到分子量小于4 ku的组分,其ACE抑制活性为57.34%.用凝胶柱SephadexG一25进行进一步纯化,纯化后的组分ACE抑制活性最高可达62.78%,比原酶解物的ACE抑制活性高了24.77%,其相对分子质量在1 500 u以下.     

棉籽蛋白源ACE抑制肽的制备过程中脱盐技术的研究

综合比较了001 ×7、201 ×7离子交换树脂、MWCO100透析袋和DA201-C大孔吸附树脂对棉籽蛋白水解液的脱盐效果,分析了DA201-C树脂脱盐对棉籽蛋白水解液ACE抑制活性和氨基酸组成的影响,并对Sephadex G-25分离纯化获得ACE抑制肽各组分进行活性测定.结果表明:DA201-C大孔吸附树脂的脱盐效果最好,控制流速为3 mL/min时,脱盐率可达到94.78％,氮回收率为86.32％,脱盐后疏水性氨基酸得到了有效富集,ACE抑制率显著提高.分离纯化后的ACE抑制肽各组分中,组分Ⅲ的抑制率最高,分子质量小于1000u.     

具ACE抑制活性的油茶饼粕酶解物的制备及分离表征

以油茶饼粕为原料,研究通过5种蛋白酶水解制备的茶粕酶解物ACE抑制率随酶解时间的变化规律,结果表明除蛋白酶C外,其余4种蛋白酶水解的茶粕酶解物ACE抑制率随酶解时间呈现先增加后减小的趋势。比较不同蛋白酶水解的茶粕酶解物ACE抑制活性得出经蛋白酶A水解后的茶粕酶解物具有较高的ACE抑制率(94.4%);为获得高纯度并具有高ACE抑制活性的酶解组分,文章采用CM-Sephadex C-25阳离子交换色谱和RP-HPLC纯化经蛋白酶A酶解的茶粕酶解物。由CM-Sephadex C-25阳离子交换色谱分离得到一个组分A,通过RP-HPLC进一步分离组分A得到6个组分。对ACE抑制率最高的组分A-2进行RP-HPLC二次纯化,得出该分离组分达到色谱纯,其半数抑制浓度(IC_(50))为98μg.m L~(-1)。     

Purification and identification of angiotensin converting enzyme inhibitory peptides from beef hydrolysates

Sarcoplasmic protein extracts from beef rump (biceps femoris) were hydrolyzed (for 0, 4, 8, 12, and 24 h) with three enzymes or their paired combinations. Ultrafiltration, gel-filtration, and RP-HPLC were used to separate angiotensin converting enzyme (ACE) inhibitory peptides from the hydrolysates. The highest ACE inhibitory activity of enzyme hydrolysates resulted from 4 h incubation with enzymes or their paired combinations. The activities of gel filtrated fractions from these hydrolysates were assayed in vitro, demonstrating that the 3rd peak of enzyme thermolysin + proteinase A hydrolysate had the highest ACE inhibition activity (52.8%). The 3rd peak of this hydrolysate was separated by RP-HPLC into five peaks, of which peak 3 showed 30.1% ACE inhibition activity. Its peptide sequence was determined to be Val-Leu-Ala-Gln-Tyr-Lys. The results suggested that this peptide may be a potent ACE inhibitor which might perhaps be used to develop beef with a bioactive peptide to lower blood pressure.     

Fractionation and identification of a novel hypocholesterolemic peptide derived from soy protein Alcalase hydrolysates

A novel hypocholesterolemic peptide was fractionated by gradient ethanol elution from a macroporous adsorption resin (MAR DA201-C), and then separated on Sephadex G-15 and RP-HPLC from a soy protein hydrolysate (SAPH DH 18%). Identification of the hypocholesterolemic peptide structure was accomplished with HPLC–MS. The peptide with the highest hypocholesterolemic activity was found in 75% ethanol fraction among the four fractions from gradient ethanol elution with MAR DA201-C. The calculated average hydrophobicity by amino acid composition of each ethanol eluted fraction suggested that the peptides could be separated in terms of hydrophobicity with MAR DA201-C. Four peaks were obtained with further fractionation on Sephadex G-15, the highest cholesterol micellar solubility inhibition rate, 81.3%, was obtained in Peak 2, corresponding to the molecular weight fraction of 300–800 Da. Fifteen main peaks were obtained with RP-HPLC fractionation, the highest cholesterol micellar solubility inhibition rate (94.3%) was in Peak 7. The amino acid sequence of this peptide was identified as WGAPSL with LC–MS and amino acid composition analysis.     

谷朊粉酶解物的制备及其ACE抑制肽的分离鉴定

以谷朊粉为原料,以血管紧张素转换酶（ACE）抑制率为指标,比较4种蛋白酶的酶解效果。采用超滤、葡聚糖凝胶色谱、反相高效液相色谱（RP-HPLC）的方法分离纯化ACE抑制肽并用电喷雾飞行时间质谱联用(ESI-TOF-MS)鉴定其结构。结果表明：碱性蛋白酶酶解3 h的谷朊粉酶解物具有较高的ACE抑制活性;超滤后,分子质量Mr<1 ku的组分具有较高的ACE抑制率;分离纯化后得到小肽的ACE抑制率高达（81.03±1.20）%;由ESI-TOF-MS质谱图得出该小肽的m/z为630.89,氨基酸序列为Trp-Phe-Gln-Pro（WFQP）。     

Purification and identification of an ACE inhibitory peptide from the peptic hydrolysate of Acetes chinensis and its antihypertensive effects in spontaneously hypertensive rats

Acetes chinensis is a marine shrimp found in the coastal waters of China. The shrimp was hydrolysed by pepsin to prepare hydrolysates with angiotensin I‐converting enzyme (ACE) inhibitory activity. The hydrolysate with the highest ACE inhibitory activity resulted from a 3–5 h incubation at 45 °C and pH 2.5 with pepsin. Gel filtration and RP‐HPLC were used to separate ACE inhibitory peptides from the hydrolysate. The gel filtration fraction of the hydrolysate with a molecular weight range from 1320 Da to 311 Da exerted the highest ACE inhibition activity. This fraction was separated by RP‐HPLC into fifteen fractions, of which fraction F9 showed 92.7% of the ACE inhibition activity. Its peptide sequence was determined to be Leu–His–Pro. It showed a potent antihypertensive activity in spontaneously hypertensive rats. The results suggested that this peptide may be a potent ACE inhibitor which might be developed into a healthy food to lower blood pressure.     

利用对虾消化腺丝氨酸蛋白酶制备鲍鱼外套膜ACE抑制肽

利用鲍鱼加工副产物外套膜制备具有抑制血管紧张素转移酶(angiotensin converting enzyme,ACE)活性的生物活性肽,为鲍鱼加工副产物的高值化利用提供新思路.采用从凡纳滨对虾消化腺中制备的丝氨酸蛋白酶,酶解皱纹盘鲍外套膜蛋白,以酶解物ACE抑制活性为评价指标优化条件.酶解液通过3 kDa超滤膜后,...     

大豆肽的离子交换色谱分离及其活性评价

采用Sephadex C-25阳离子交换剂对经过DA201-C型大孔吸附树脂脱盐、乙醇梯度洗脱(75%乙醇洗脱组分)的Alcalase水解大豆肽(DH14%)进行进一步的分离纯化,并对分离得到的各组分ACE抑制活性进行了评价。试管试验确定的SephadexC-25色谱分离条件为:上样量0.004g/mLIE,起始缓冲液为1M醋酸,上样吸附率为61.19%。经SP-Sephadex C-25分离得到6个组分,其中未吸附的3个组分ACE抑制活性低,但NaCl梯度洗脱的3个组分ACE活性均为60%左右,从纯化ACE抑制活性肽的角度考虑,分离效果比SephadexG-15差。     

马氏珍珠贝肉蛋白水解特征及其ACE抑制肽的筛选

为探索马氏珍珠贝肉的水解规律及快速鉴定血管紧张素转化酶(angiotensin-converting enzyme,ACE)抑制活性肽,本研究以不同时间(2、4、6、8、10h)的酶解物为对象,采用三羟甲基甘氨酸-十二烷基硫酸钠-聚丙烯酰胺凝胶电泳、分子质量分布、反相高效液相色谱(reversed-phase high...     

HPLC-ESI-MS对珠蛋白酶解液中的血管紧张素I转换酶抑制肽的分离鉴定

本实验利用ESI-MS/MS对反相高效液相色谱分离的具有血管紧张素I转换酶(ACE)抑制活性的珠蛋白小肽的结构进行鉴定。结果表明:此肽的序列为Val-Val-Tyr-Pro-Trp-Thr(VVYPWT),位于猪的血红蛋白β链的34-39氨基酸序列片断,它对ACE有很好的抑制活性,其IC50为6.02μmol/L。     

Studies on purification and the molecular mechanism of a novel ACE inhibitory peptide from whey protein hydrolysate

This study sought to purify and identify a novel angiotensin I-converting enzyme (ACE) inhibitory peptide from whey protein hydrolysed by trypsin. The peptide’s amino acid sequence, as well as the molecular mechanism of the interactions between the peptide and the ACE, were also studied. Using ultrafiltration, the hydrolysate was separated into three fractions. The fraction with molecular weight of <6 kDa had the greatest ACE inhibitory activity and was further separated by size exclusion chromatography on Sephadex G-25 and G-10 columns. Reverse-phase high performance liquid chromatography (RP-HPLC) was used to separate the most active fraction. The amino acid sequence of the peptide with the greatest ACE inhibitory characteristics was confirmed as Leu–Leu (LL). The molecular mechanisms, position, type, and energy of the LL/ACE interaction were investigated by using flexible molecule docking technology.