Identification and characterization of multifunctional cationic peptides from enzymatic hydrolysates of soybean proteins

1. Download : Download high-res image (204KB)
2. Download : Download full-size image

     

Identification of cationic peptides derived from low protein rice by-products and evaluation of their multifunctional activities

1. Download : Download high-res image (251KB)
2. Download : Download full-size image

     

发酵豆制品生物活性肽功能特性研究进展

发酵豆制品在亚洲许多国家被广泛食用,其也是生物活性肽的潜在来源。近年来,发酵豆制品生物活性肽的研究越来越深入,其降血压、抗氧化、抗菌、降血糖、抗癌等功能特性也不断地被揭示。该文对发酵豆制品生物活性肽的功能作用研究现状进行了综述,并对该领域未来研究方向进行了展望,旨在为发酵豆制品的广泛应用和深度开发提供参考。     

Assessment of multifunctional activity of bioactive peptides derived from fermented milk by specific Lactobacillus plantarum strains

Milk-derived bioactive peptides with a single activity (e.g., antioxidant, immunomodulatory, or antimicrobial) have been previously well documented; however, few studies describe multifunctional bioactive peptides, which may be preferred over single-activity peptides, as they can simultaneously trigger, modulate, or inhibit multiple physiological pathways. Hence, the aim of this study was to assess the anti-inflammatory, antihemolytic, antioxidant, antimutagenic, and antimicrobial activities of crude extracts (CE) and peptide fractions (<3 and 3–10 kDa) obtained from fermented milks with specific Lactobacillus plantarum strains. Overall, CE showed higher activity than both peptide fractions (<3 and 3–10 kDa) in most of the activities assessed. Furthermore, activity of <3 kDa was generally higher, or at least equal, to the 3 to 10 kDa peptide fractions. In particular, L. plantarum 55 crude extract or their fractions showed the higher anti-inflammatory (723.68–1,759.43 μg/mL of diclofenac sodium equivalents), antihemolytic (36.65–74.45% of inhibition), and antioxidant activity [282.8–362.3 µmol of Trolox (Sigma-Aldrich, St. Louis, MO) equivalents]. These results provide valuable evidence of multifunctional role of peptides derived of fermented milk by the action of specific L. plantarum strains. Thus, they may be considered for the development of biotechnological products to be used to reduce the risk of disease or to enhance a certain physiological function.     

Scavenging and anti-fatigue activity of fermented defatted soybean peptides

Many bioactive peptides possess specific biological properties that make these potential ingredients of health-promoting foods. Increasing attention is being focused on physiologically active peptides derived from soybean proteins. In this study, soybean peptides were produced by fermentation of defatted soybean meal with Bacillus subtilis SHZ and purified by ultrafiltration and gel chromatogram. Free radical scavenging property and the effect on anti-fatigue of the peptides were evaluated in vitro and in vivo, respectively. Swimming endurance tests of mice were carried out after 20 days of soybean peptides administration (0, 100, 200, 400 mg/kg BW/day for control, low-dose, middle-dose and high-dose group, respectively). And the blood lactate and hepatic glycogen of mice were determined. Results showed that the purified peptides exhibited significant (P < 0.01) scavenging potencies on superoxide (62%) and hydroxyl (96%) at a concentration of 10 mg/ml. The average swimming time of the low-, middle- and high-dose group was increased by 20.91, 45.45 and 70% compared with that of control group, respectively. And administration with soybean peptides could significantly accelerate the clearance of blood lactate after mice swimming (P < 0.01). The hepatic glycogen storage of middle and high dose groups were obviously increased (P < 0.05). It suggested that the soybean peptides produced by fermentation could significantly alleviate physical fatigue of the mice.     

Identification of antioxidant and ACE‐inhibitory peptides in fermented milk

The angiotensin‐converting enzyme (ACE)‐inhibitory activity and antioxidant properties of a commercial fermented milk from Europe were evaluated. This dairy product showed moderate ACE‐inhibitory activity and ABTS^?+ radical‐scavenging capacity. The peptides from most active fractions collected by reverse phase high‐performance liquid chromatography (RP‐HPLC) were sequenced by RP‐HPLC–tandem mass spectrometry. This technique allowed rapid identification of peptides included in the most active fractions, and various potentially active peptides were recognised according to previous studies of structure–activity relationship. Three of the identified sequences had previously been described as potent ACE inhibitors. The structure of some sequences substantiated the presence of peptides with ACE‐inhibitory, antioxidant and immunomodulatory activities. Copyright © 2005 Society of Chemical Industry     

锡盟蒙古族传统发酵马乳中功能性特征肽段的研究

乳基中的小分子活性肽的相关研究,近几年被重点关注,它可以提高机体免疫能力,是调节生理代谢的功能成分。发酵马乳已被证实具有较高的营养价值和医疗保健作用。该研究针对锡盟蒙古族传统发酵马乳,用高效凝胶过滤色谱与超高效液相色谱-电喷雾串联三重四极杆质谱分析发酵马乳样品中蛋白水解物的分子质量分布,并对主要的功能二肽进行定性定量检测。研究结果表明,发酵后马乳的分子质量在1 000 Da以下的蛋白水解物比例有显著提升,检测出24条具有潜在血管紧张素转化酶(angiotensin-converting enzyme,ACE)抑制功能的二肽,标记含量较高且有功能活性的肽段VP、AF为该发酵马乳样品的特征肽段,并对以上功能肽段的构效关系进行探讨。该研究为进一步开发来源于蒙古族传统发酵食品中的功能乳肽提供参考。     

传统发酵豆酱中产蛋白酶菌株的筛选

该研究从东三省各地采集家庭自制豆酱254份,通过分离培养,获得纯培养真菌菌株389株。采用脱脂牛奶平板初筛获得具有蛋白酶活性的菌株31株,其中曲霉属真菌12株、青霉5株、毛霉4株、镰孢菌3株、帚霉1株、芽枝孢3株、犁头霉1株、红曲2株。通过发酵复筛提取其粗酶液,用福林酚法测定粗酶液的蛋白酶活性,筛选出了两株具有较高蛋白酶活性的米曲霉。     

嗜酸乳杆菌发酵干酪成熟过程中蛋白质降解产生的肽类的性质分析

对嗜酸乳杆菌和嗜热链球菌发酵生产的具有功能性的低脂和全脂干酪4℃成熟90d时蛋白质的降解程度进行了研究。通过电泳分析表明,两种干酪成熟后蛋白质的降解程度比市售的普通低脂和全脂干酪要充分,且低脂干酪比全脂干酪蛋白降解程度要深,降解物含量要多;反相高效液相色谱(RP-HPLC)显示的干酪中肽图谱表明,成熟后,虽然低脂干酪和全脂干酪中仍然存在和牛奶中相同的蛋白物质,但部分发生了降解并产生了新的肽类,且两种干酪所生成的肽组分各不相同。     

鹰嘴豆肽、大豆肽功能性质的研究

研究对比了鹰嘴豆肽和大豆肽的水解度、抗氧化性、吸油性、吸湿及保湿性等功能特性的差异,以及蛋白酶种类对肽产物功能性质的影响。结果表明,①酶Ⅱ(Protease from Bacillus sp.)制备的蛋白肽抗氧化性和吸油能力最好,且在低湿度下的吸湿性和高湿度下的保湿性也最好;酶Ⅲ(Papain from papaya latex)制备的蛋白肽水解度最高;酶Ⅰ(Protease fromAspergillus melleus)制备的蛋白肽在不同湿度条件下都有较好的吸湿性能。②大豆肽的水解度和抗氧化能力比鹰嘴豆肽好。③Desi肽吸油能力最强,在抗氧化性上仅次于大豆肽,高低湿度环境下都有很好的保湿能力,其中Desi肽Ⅱ这些特征最为明显。④Kabuli肽在不同湿度条件下的吸湿和保湿能力都较好。     

传统发酵豆酱中产脂肪酶菌株的筛选

脂肪酶属于羧基酯水解酶类,能将甘油三酯逐步地水解成甘油和脂肪酸。该研究从东北三省传统的家庭自制豆酱上获得纯培养真菌389株,通过三丁酸甘油酯平板法初筛获得具有脂肪酶活性的菌株41株,其中青霉属的真菌27株、曲霉5株、毛霉3株、犁头霉2株、酵母菌1株、镰孢菌2株、帚霉菌1株。结果表明,青霉属的真菌无论从数量还是酶活都高于其他属的真菌,其中橘灰青霉（Penicillium aurantiogriseum）数量最多。在分离到的菌株中,毛霉属的卷枝毛霉（Mucor circinelloides）酶活最高,是具有生物催化潜力的功能菌株。     

Identification of angiotensin I-converting enzyme inhibitory peptides from koumiss, a traditional fermented mare's milk

Angiotensin I-converting enzyme (ACE) inhibitory activities in untreated koumiss and koumiss digested with ACE, pepsin, trypsinase, and chymotrypsin were compared and analyzed. Four novel ACE inhibitory peptides (P_I, P_K, P_M, and P_P) were purified using ultrafiltration and high performance liquid chromatography (HPLC). The classification study showed that these 4 peptides were of the true inhibitor type. The amino acid sequences of these peptides are YQDPRLGPTGELDPATQPIVAVHNPVIV, PKDLREN, LLLAHLL, and NHRNRMMDHVH, respectively. Their individual IC₅₀ (50% inhibitory concentration) values were as follows: 14.53 ± 0.21 μM, 9.82 ± 0.37 μM, 5.19 ± 0.18 μM, and 13.42 ± 0.17 μM. From sequence analysis, we determined that P_I was part of β-casein in mare's milk. The 3 peptides P_K, P_M, and P_P did not correspond with any known milk protein. The results suggest that koumiss is rich in ACE inhibitory peptides, and the ACE inhibitors in koumiss are of the pro-drug type or a mixture of the pro-drug type and the true inhibitor type. These results may provide evidence about the beneficial effects of koumiss, especially on cardiovascular health.     

PCR-DGGE分析吉林市传统发酵豆酱中细菌的多样性

该研究以吉林市农家传统发酵豆酱为试材,利用PCR-DGGE技术分析豆酱不同发酵阶段细菌的生物多样性及变化规律。结果表明,豆酱发酵过程中细菌菌群种类较为丰富,主要有戊糖片球菌（Pediococcus pentosaceu）、乳明串珠菌（Leuconostoc lactis）、葡萄球菌属（Staphylococcus sp.）、屎肠球菌（Enterococcus faecium）、解淀粉芽孢杆菌（Bacillus amyloliquefaciens）、弗氏柠檬酸杆菌（Citrobacter freundii）、霍氏肠杆菌（Enterobacter hormaechei）、产酸克雷伯氏菌（Klebsiella oxytoca）、不可培养杆菌（uncultured bacterium）。其中乳酸菌是发酵过程中的优势菌,对豆酱的风味起着主要作用。传统豆酱发酵过程中菌群有一定的亲缘性,菌落结构变化较小,为传统发酵豆酱的进一步开发和研究提供理论基础。     

Technological characterization of lactococci isolated from traditional Chinese fermented milks

To screen lactic acid bacteria for starter cultures in cheese production, 21 Lactococcus strains previously isolated from natural fermented milk and koumiss made by herdsman families in the Xinjiang, Gansu, and Qinghai provinces of China were evaluated for optimal growth temperature, acidification activity, proteolytic activity, aminopeptidase activity, and autolytic activity. All isolates presented low acidification rates, and the pH value did not reach 5.3 after 6 h of inoculation in sterile reconstituted skim milk at 30°C. Strains X9C2 and T7C showed the highest proteolytic activity of 24.67 and 23.58 mg of glycine/L of milk, respectively. For aminopeptidase activity, strains X9C2 and T1C2 displayed the highest activities of 30.56 and 27.70 U/mg of protein using l-leucine-p-nitroanilide as substrate, respectively. Autolytic activity in simulated cheese-like buffer ranged from 7.45 to 34.76%, and strains Q14C2 and Q16C showed the highest values of 34.76 and 34.20%, respectively. Collectively, one main finding is that some technological characteristics of Lactococcus isolates from Chinese traditional fermented products varied greatly. Some isolates with potentially important properties could be valuable for application as starter cultures of cheese or could constitute a mixed culture.     

Anti-oxidative capacity of enzymatically released peptides from soybean protein isolate

Shelf-life of dietary products and the need for health supporting functional ingredients often require components with high anti-oxidative properties. The study aimed at the characterisation of peptides with antioxidant properties from enzymatically hydrolysed soybean protein isolate. Soybean protein isolate was hydrolysed with a food grade pancreatic trypsin/chymotrypsin for industrial applications, ultrafiltered and the permeate was fractionated by size exclusion chromatography. Anti-oxidative properties of these fractions were tested in a Trolox^®-equivalent-anti-oxidative capacity assay, which is based on the absorbance suppression of 2,2′-azinobis(3-ethyl benzothiazoline-6-sulfonate) radical cations by anti-oxidative substances at λ 414 nm. The amino acid compositions and sequences of the peptide fractions were determined by liquid chromatography and mass spectrometry analysis. Compared to the intact soybean protein isolate a peptide fraction with an anti-oxidative capacity of 113 mg TEAC/g was obtained. This fraction was shown to be enriched in peptides smaller than 1 kDa with prevailing aromatic amino acid residues. Nine peptides derived from glycinin and β-conglycinin were sequenced by tandem mass spectrometry. The particular sequences were synthesized and assayed. The synthetic peptide analogue TTYY (β-conglycinin fragment 291–294) revealed significant radical scavenging properties of 13.6 up to 59.6% with 0.18–18 mM in a concentration dependent manner. Peptides from soybean protein isolate were shown to possess anti-oxidative capacities which might be linked to the presence of carboxy terminal tyrosine. These findings offer new functional application possibilities in nutrition, cosmetics and pharmaceuticals.     

Identification and functional characterization of Lactobacillus strains isolated from milk and Gioddu,a traditional Sardinian fermented milk

Raw milk from Sardinian cows and sheep, and sheep's milk yoghurt (Gioddu), are food products that are poorly characterized in terms of the presence of mesophilic Lactobacillus strains, and may be good sources from which to isolate novel bacterial strains. Sixteen Lactobacillus strains were identified by genotypic and phenotypic methods from samples of sheep's milk (n=7), fermented sheep's milk (n=7) and one sample of cows’ milk. These strains were further characterized for the presence of functional traits useful for probiotic applications, such as resistance to simulated gastric acid, resistance to bile salts, and adhesion of two different cell lines (Caco-2 and MIM/PPK). Moreover, selected aspects correlated with safety (antimicrobial susceptibility and presence of plasmids) and ecological roles (ability to produce biofilms and bacteriocin-like substances) were tested. All but 1 of the 16 strains were unambiguously allotted to three species, L. paracasei (5 strains), L. plantarum (8 strains), and L. reuteri (2 strains). Many of the strains exhibited characteristics suggesting that they would survive in the gastrointestinal tract and also had the capability to attach to various cells. Greater variability was observed for the other traits analyzed. These data suggest that Sardinian artisan dairy products are interesting sources for the isolation of bacterial strains with characteristic and differentiated functional traits.     

大豆多肽特性及其应用研究现状

本文主要综述了大豆多肽加工工艺流程、大豆多肽加工特性和功能特性及当前研究现状,并对今后发展提出几点展望。     

酶解小黑豆乳清多肽抗肿瘤作用的研究

研究以小黑豆乳清为原料,比较木瓜蛋白酶、537酸性蛋白酶、ProteAX蛋白酶三种蛋白酶水解多肽产物的分子量分布及其对小鼠H22肿瘤细胞生长抑制作用。结果表明,ProteAX酶解多肽的分子量分布明显不同于另外两种多肽,且对小鼠H22肿瘤细胞生长抑制率最高,当ProteAX酶解多肽浓度为40mg/mL时,可达51.36%。用ProteAX酶解多肽喂食用氨基比林-亚硝酸钠（AP-NaNO2）诱导的肝损伤模型小鼠,结果显示,与模型组相比,该多肽可以显著降低肝脏MDA和血清ALT、AST活性（p<0.05）,并提高肝脏GSH-Px活性（p<0.05）。肝脏组织切片结果显示,模型组小鼠大部分肝细胞形态不正常,细胞损伤严重,出现明显的癌前期病变;而小黑豆乳清多肽高剂量组动物的肝细胞形态基本正常,接近正常肝组织;提示ProteAX蛋白酶解多肽可以抑制AP-NaNO2诱导的肝细胞损伤和癌前期病变。