枯草芽孢杆菌发酵提取物对大肠杆菌的抑制作用

为探索枯草芽孢杆菌发酵提取物对大肠杆菌的抑菌效果及作用机理,利用倍半稀释法确定其对大肠杆菌的最小抑菌浓度,通过研究枯草芽孢杆菌发酵提取物对大肠杆菌生长曲线、细胞膜通透性以及细胞超微结构的影响,探讨提取物对大肠杆菌的抑制作用机理,同时对该提取物的化合物类型进行了初步分析。结果表明,该提取物对大肠杆菌具有明显的抑制作用,其最小抑菌浓度为0.614 mg·mL-1,在大肠杆菌生长的延滞期和对数期加入该提取物,比在稳定期加入能够显现出更好的抑菌效果。经提取物作用后的细胞表面粗糙,边缘模糊,细胞膜破裂,表明该提取物能够增加大肠杆菌细胞膜的通透性。化合物分析显示,提取物中抑菌成分主要是多烯类化合物和脂肽类化合物。     

儿茶素的抑菌效果及机理研究

以金黄色葡萄球菌和大肠杆菌为实验对象,采用抑菌环实验研究了儿茶素的最小抑菌浓度,并对儿茶素作为抑菌剂的稳定性作出了评价,同时还研究了儿荼素可能存在的抑菌机理.结果显示,儿茶素对金黄色葡萄球菌和大肠杆菌的最小抑菌浓度分别为0.32g/L和1.25g/L;儿茶素的抑菌效果较大程度地受温度和pH的影响,较小程度受紫外照射的影响;儿茶素会抑制金黄色葡萄球菌和大肠杆菌的对数生长期,增加细菌细胞膜的通透性,造成细菌胞内蛋白质和糖类物质的渗漏,使细菌代谢发生紊乱,儿茶素的抑菌作用不是通过抑制呼吸代谢而引起的.     

抗菌肽brevilaterin与ε-聚赖氨酸对金黄色葡萄球菌的协同抑菌机理

金黄色葡萄球菌是食品中常见的致病菌,控制食品中金黄色葡萄球菌的生长繁殖对提高食品安全性至关重要。本研究以金黄色葡萄球菌作为指示菌,考察了抗菌肽brevilaterin与ε-聚赖氨酸的协同抑菌机理。抑菌动力学结果表明抗菌肽与ε-聚赖氨酸对金黄色葡萄球菌具有协同抑菌效果;细胞膜质子动力势研究结果显示,抗菌肽对跨膜pH值梯度无明显影响,ε-聚赖氨酸会破坏跨膜pH值梯度,1/4最小抑菌浓度(minimal inhibitory concentration,MIC)抗菌肽+1/4 MICε-聚赖氨酸(联用组)对跨膜pH值梯度产生了协同破坏作用;采用流式细胞术结合荧光显微镜考察细胞膜的完整性,发现抗菌肽对细胞膜完整性具有较强的破坏作用,1/4 MIC抗菌肽即可导致36.3%的膜破损,而ε-聚赖氨酸对膜完整性损伤较小,1/4 MICε-聚赖氨酸仅破坏10.4%的细胞膜完整性,两者联用可对细胞膜完整性产生协同破坏作用,导致51.3%细胞膜完整性发生损伤;采用透射电子显微镜观察了细胞超微结构,发现抗菌肽和ε-聚赖氨酸联用较单独使用对细胞形态和细胞内容物的泄漏产生了协同破坏作用;十二烷基硫酸钠-聚丙烯酰胺凝胶电泳结果显示,ε-聚赖氨酸会抑制菌体蛋白质的合成或降解蛋白质,而抗菌肽对蛋白质合成无影响;琼脂糖凝胶阻滞电泳表明,抗菌肽对菌体DNA条带无明显变化,而ε-聚赖氨酸以及两者联用会造成DNA滞留,表明ε-聚赖氨酸可以通过与DNA结合发挥抑菌作用。上述结果表明,抗菌肽brevilaterin和ε-聚赖氨酸联用可以增强对细胞膜的破坏强度,且兼具抗菌肽对呼吸链脱氢酶活性的抑制与ε-聚赖氨酸对跨膜pH值梯度的破坏和DNA的结合作用,从而实现多靶位协同抑菌。     

化学合成抗菌肽抑制致病性大肠杆菌F41的分子作用机理

本文以化学合成抗菌肽为研究对象,对合成抗菌肽的抑菌活力、抑菌动力学进行研究;通过圆二色谱法评价抗菌肽二级结构的变化;电镜观察抗菌肽对细菌微观结构影响;K+和紫外吸收物质泄漏实验分析抗菌肽对细胞膜通透性影响;DNA凝胶阻滞实验分析抗菌肽与细菌DNA相互作用关系。结果表明:不同抗菌肽的抗菌活力、抑菌动力学存在差异;抗菌活力与抗菌肽浓度、作用时间有关;抗菌肽Tac、Tac W、Tac V明显改变细胞微观结构,使胞内K+和紫外吸收物质泄漏,细胞膜通透性改变,造成细胞坍塌破裂,进入细胞与DNA结合,产生凝胶阻滞现象。得出结论:抗菌肽作用机理与抗菌肽结构中碱性氨基酸、两亲性氨基酸的比例密切相关,其分子作用机理包括使细胞膜通透性发生变化、改变细菌细胞微观结构、结合基因组DNA,抑制复制、转录等,形成多个作用靶点,是一种多效协同的作用机制。     

表没食子儿茶素没食子酸酯对嗜水气单胞菌抑菌机制的研究

以嗜水气单胞菌为实验对象,研究了表没食子儿茶素没食子酸酯(EGCG)对其的抑菌活性及抑菌机制。结果表明,EGCG对嗜水气单胞菌的最小抑菌浓度均为25～50μg/mL。菌体在EGCG作用下,改变细胞膜的通透性,电导率增大,细胞内物质渗漏,从而影响细胞代谢,最终导致细胞死亡。     

香芹酚对阴沟肠杆菌的抑菌作用及机理

香芹酚具有较好的抑菌效果,符合人们对食品防腐剂安全无毒、无副作用的要求。本研究通过最小抑菌浓度和杀菌实验分析了香芹酚对阴沟肠杆菌（Enterobacter cloacae C4）的抑菌活性,并检测其对阴沟肠杆菌细胞膜电势、胞外ATP化学发光值的影响,使用激光共聚焦显微镜观察细胞渗透性,并通过扫描电子显微镜观察香芹酚对阴沟肠杆菌细胞形态的影响,从而探究香芹酚对阴沟肠杆菌的抑菌机理。结果表明：香芹酚对阴沟肠杆菌的最小抑菌浓度为0.5 μL/mL;阴沟肠杆菌细胞膜通透性随着香芹酚含量（0.25～2 μL/mL）的增加而逐渐增加。经香芹酚处理的阴沟肠杆菌细胞膜内外电势改变,ATP大量漏出。通过扫描电子显微镜观察,发现经香芹酚处理的菌体细胞形态干瘪且明显被破坏,含量高于1 μL/mL的香芹酚几乎能够溶解菌体。推测香芹酚通过作用于细胞膜改变膜内外电势,造成细胞膜通透性改变,引起细胞ATP等内容物泄露,从而影响细胞功能,达到抑菌的作用。     

卵黏蛋白抑菌活性及其作用机制

研究了卵黏蛋白对导致禽蛋腐败的常见微生物——金黄色葡萄球菌、腐生葡萄球菌、枯草芽孢杆菌、大肠杆菌、沙门氏菌以及对致龋齿的变异链球菌的抑菌活性及作用机制。采用最小抑菌浓度(MIC)和牛津杯法测定卵黏蛋白的抑菌活性,分析卵黏蛋白对其中3种菌生长动力曲线的影响;利用流式细胞仪与扫描电镜分析其抑菌机制。结果表明:卵黏蛋白对金黄色葡萄球菌、腐生葡萄球菌、变异链球菌的MIC分别为62.5,125,1 000μg/m L,并有明显的抑菌圈产生;对枯草芽孢杆菌、大肠杆菌、沙门氏菌的抑制作用较差,无明显抑菌圈。卵黏蛋白对菌体生长起到延缓对数期或缩短稳定期的作用,并使菌体细胞膜的通透性发生改变。扫描电镜结果显示,卵黏蛋白使菌体细胞形态发生变化,细胞膜损伤或者破坏。卵黏蛋白通过对菌体细胞膜造成损害而发挥其抑菌活性。     

脂肽和茶多酚对副溶血弧菌的协同抑菌效应和机理

研究脂肽和茶多酚对副溶血弧菌的协同抑菌效应和机理,为脂肽协同茶多酚应用于副溶血弧菌的控制提供理论依据。利用"棋盘法"研究脂肽和茶多酚对副溶血弧菌的协同抑菌效应;通过研究脂肽和茶多酚独立作用及协同作用对细胞膜完整性、细胞蛋白质合成及磷代谢方面影响来研究二者的协同抑菌机理。结果表明:脂肽和茶多酚对副溶血弧菌存在强烈的协同抑菌效应,部分抑菌浓度指数达到0.19;与1/16最小抑菌浓度(minimum inhibitory concentration,MIC)脂肽或1/8 MIC茶多酚各自独立作用相比,在1/16 MIC脂肽和1/8 MIC茶多酚协同作用下,副溶血弧菌的细胞膜通透性显著增强,细胞部分蛋白质合成的抑制作用得到进一步加强,并且细胞的磷代谢完全受到抑制。结果表明,脂肽和茶多酚对副溶血弧菌具有强烈的协同抑菌效应,其协同抑菌作用主要通过二者协同破坏细胞膜的完整性,从而影响细胞部分蛋白质的合成与正常代谢实现的。     

枯草芽孢杆菌抗菌脂肽对嗜水气单胞菌抑菌效果

为了研究Bacillus subtilis fmbJ 产生的抗菌脂肽对嗜水气单胞菌的抑菌效果,采用抑菌圈法测定抗菌脂肽对嗜水气单胞菌的最小抑菌质量浓度(MIC)和最小杀菌质量浓度(MBC),并从其对嗜水气单胞菌形态结构、膜通透性来探讨其抗菌作用。结果表明,嗜水气单胞菌对枯草芽孢杆菌抗菌脂肽具有较高的敏感性,最小抑菌质量浓度(MIC)为20μg/mL,最小杀菌质量浓度(MBC)为32μg/mL;该抗菌脂肽导致嗜水气单胞菌细胞膜通透性增加,使细胞内一些离子以及大分子的蛋白质和核酸泄漏到细胞外,从而引起细胞的死亡;显微结构影响的研究表明,该抗菌脂肽还可以造成嗜水气单胞菌细胞壁缺失、鞭毛散失、胞内物质模糊不清等现象,抑菌效果较明显,可用于嗜水气单胞菌感染引起的水生动物疾病预防和控制。     

石竹烯对热杀索丝菌的抑菌机理

本实验研究石竹烯对热杀索丝菌的抑菌效果及其作用机理。通过石竹烯对热杀索丝菌的最小抑菌浓度(minimum inhibition concentration,MIC)和生长曲线的测定考察其抑制活性;通过观察扫描电子显微镜和碱性磷酸酶的泄漏判定石竹烯对细胞形态和细胞壁结构的破坏程度;通过钾离子、二乙酸荧光素分子和蛋白质的泄漏考察其对细胞膜的影响;通过苹果酸脱氢酶以及丙酮酸激酶活力的测定探究其对细胞代谢的影响;通过荧光光谱扫描法研究石竹烯与细菌DNA的结合能力。结果表明,石竹烯对热杀索丝菌的MIC值为4.51 mg/mL,1×MIC和2×MIC石竹烯能够破坏细胞的形态、结构以及细胞膜的通透性,影响细胞代谢,导致苹果酸脱氢酶、丙酮酸激酶活力显著降低(P0.05),同时对基因组DNA构象和结构造成破坏。     

儿茶素及其衍生物合成研究进展

儿茶素具有抗肿瘤、抗氧化、抗病菌以及保护心脑器官等多种药理作用,这些药理活性主要由其多羟基结 构决定,然而,多羟基结构也使儿茶素在中性和碱性条件下结构不稳定。儿茶素被人体摄入后会很快在生物酶作用 下发生甲基化或糖苷化,研究发现,在儿茶素结构中引入其他基团可有效提高其稳定性和生物利用率,并特异性增 强儿茶素药理作用。其中,甲基儿茶素抗过敏和逆转肿瘤细胞耐药性表现突出,糖苷儿茶素水溶性显著提高并可有 效防止褐变,而酰化儿茶素由于脂溶性明显增强,其抗氧化和抗癌活性也随之明显增强。在不同位置进行取代达到 的效果也不相同,通过研究儿茶素及其不同衍生物的合成途径和药理作用,将有助于进一步研究发现儿茶素新的药 理功能并加快促进儿茶素药物的临床应用,本文详细介绍了儿茶素生物合成和化学合成途径,重点综述儿茶素不同 衍生物的合成途径并简单说明其药理作用。     

Binding affinity of tea catechins for HSA: Characterization by high‐performance affinity chromatography with immobilized albumin column

Catechins are the major polyphenols in green tea leaves. Recent studies have suggested that the catechins form complexes with HSA for transport in human blood, and their binding affinity for albumin is believed to modulate their bioavailability. In this study, the binding affinities of catechins and their analogs were evaluated and the relationship between the chemical structure of each catechin and its binding property were investigated. Comparing these catechins by HPLC analysis with the HSA column, we showed that galloylated catechins have higher binding affinities with HSA than non‐galloylated catechins. In addition, pyrogallol‐type catechins have a high affinity compared to catechol‐type catechins. Furthermore, the binding affinity of the catechin with 2,3‐trans structure was higher than those of the catechin with 2,3‐cis structure. The importance of the hydroxyl group on the galloyl group and B‐ring was confirmed using methylated catechins. These results indicate that the most important structural element contributing to HSA binding of tea catechins is the galloyl group, followed by the number of hydroxyl groups on the B‐ring and the galloyl group or the configuration at C‐2. Our findings provide fundamental information on the relationship between the chemical structure of tea catechins and its biological activity.     

Scavenging effect of tea catechins and their epimers on superoxide anion radicals generated by a hypoxanthine and xanthine oxidase system

Catechins are a major group of polyphenolic compounds contained in abundance in green tea. Using electron spin resonance spectroscopy along with a spin‐trapping agent, the scavenging effect of tea catechins and their corresponding epimers against superoxide anion radicals generated by a hypoxanthine and xanthine oxidase reaction system was evaluated. The presence of 3′,4′,5′‐trihydroxyl groups attached to the B‐ring of the flavan skeleton of tea catechins elevated their radical‐scavenging efficiency in comparison to those with 3′,4′‐dihydroxyl groups. There were no significant differences between the four dominant tea catechins and their corresponding epimers with regard to radical‐scavenging ability. Under the different spin‐trapping agent concentrations, the sigmoid curves of reducing spin‐trapping adducts produced by tea catechins were shifted leftward, suggesting that a likely possible action of tea catechins is to scavenge superoxide anion radicals directly, not to inhibit the function of xanthine oxidase. Although caffeine is also known as a major ingredient of tea, its superoxide anion radical‐scavenging effect was much weaker than that of the catechin family. It is concluded that tea catechins and their epimers serve as powerful antioxidants for directly eliminating superoxide anion radicals, and may be useful in the prevention of diseases relating to in vivo oxidative stress. © 2000 Society of Chemical Industry     

儿茶素改性的研究进展

儿茶素是茶叶中的主要活性物质,但由于较低的溶解性、稳定性及生物利用率使其在食品、医药、化工等领域的应用受到限制,因此通过改性可以提高儿茶素的应用特性。综述儿茶素改性的主要物理、生化方法以及改性后儿茶素的理化性质和生物活性,并展望儿茶素改性的发展方向。     

儿茶素单体分离制备方法

茶叶中含有丰富的儿茶素类物质,占茶叶干重12%～24%,是茶叶中最具生理活性和保健功效的物质。文中对目前国内外纯化制备儿茶素的常见方法进行了综述。常见的儿茶素制备方法包括纤维素柱层析、葡聚糖凝胶(Sephadex LH—20)、硅胶柱层析、吸附树脂柱层析和高速逆流色谱等方法。通过这些方法制备的儿茶素单体纯度不高,仍需结合结晶、制备型液相色谱技术或膜分离技术进行进一步纯化,从而才能获得高纯度的儿茶素单体。在儿茶素单体的制备方法中,每种方法都有相应的优缺点。Sephadex LH—20柱层析分离的量较大,但是分离时间较长,而且Sephadex LH—20柱填料昂贵;高速逆流色谱分离效果较好,得到的儿茶素单体的纯度较高,分离时间相对要短,但是目前市场上只有分析型,分离的量相对少;硅胶材料廉价,但是分离的效果不佳,且分离过程相对繁琐,单体制备量较小;吸附树脂柱层析分离已实现EGCG单体的工业化生产,但在其他儿茶素单体的分离制备上尚不成熟。     

Epimerisation and hydrolysis of catechins under ultrasonic treatment

This study investigated the influence of ultrasonic treatments on the chemical conversions of catechins. Ultrasound treatments significantly enhanced the epimerisation and the hydrolysis of catechins in both the catechins solution and the tea extract solution. Catechins are more stable in the tea extract solution compared to the catechins solution. The conversion of catechins induced by ultrasound can be controlled by temperature and pH. Regard to the pyrogallol-type catechins, both the epimerisation and the hydrolysis were enhanced at the higher temperatures (60 and 70 °C) and the acidic condition (pH 6). The epimerisation and the hydrolysis of (-)-epigallocatechin gallate (EGCG) improved by 47% and 18%, respectively, when the temperature rose from 50 to 70 °C. Compared to pH 8, the epimerisation and the hydrolysis of EGCG at pH 6 improved by 27% and 39%, respectively. This study can be utilised to develop appropriate ultrasound-assisted method to produce particular catechin monomers.     

Taste characterisation of green tea catechins

There has been interest in biological activities of green tea catechins. However, little is known about the taste characteristics of catechins. To assess the taste characteristics of catechins ((?)–epicatechin (EC), (?)–epicatechin gallate (ECg), (?)–epigallocatechin (EGC) and (?)–epigallocatechin gallate (EGCg)), sensory evaluations were performed. The taste intensity increased with increased catechins concentration. Among them, ECg showed the strongest taste. Catechins had tastes that consisted primarily of astringency and bitterness. Therefore, taste palatability decreased with increasing catechin concentrations. In line with taste intensity, taste palatability of ECg was the lowest. Taste sensor analysis and mouse behavioural assays showed same results. EC and ECg were more stable in Ringer’s solution than EGC and EGCg. Furthermore, taste cell responses to ECg that had the strong taste and better stability among catechins used were recorded. Some taste cells responded to ECg. This result suggests that ECg might be recognised by taste cells.     

Using chromatography and mass spectrometry to monitor isomerization of catechin in alkaline aqueous with thermal processing

Polyphenols are associated with a wide range of physiological properties. Catechins are a class of polyphenolic flavonoids predominantly found in foods and beverages that are chemically unstable. This study investigated the effects of thermal processing and pH on the structural changes to catechins and examined how these effects impact the antioxidant capacity of these molecules. The results indicated the total phenolic contents of catechin were increased after alkalifying and thermal processing. The catechin is modified by thermal processing in an alkaline environment. The chromatographic analysis demonstrated that basification and thermal processing reduced catechin, resulting in cleavage of the heterocyclic ring of the flavan‐3‐ols via oxidation cleavage of the ether bond. A novel reaction intermediate for this transformation is proposed.

Practical applications

Heating process and pH adjustment are used to meet hygienic requirements and maintain flavor during tea infusion processing in tea beverage manufacturing. Catechins account for the largest sum of polyphenols found in green tea. Basification and heating could catalyze the isomerization of catechin through an opened heterocyclic ring pathway. Catechin remains stable in acidic solutions during thermal processing, thus, avoiding changes of quality.     

In vitro human skin permeation and cutaneous metabolism of catechins from green tea extract and green tea extract-loaded chitosan microparticles

Catechins are major antioxidants in green tea (Camellia sinensis or Camellia assamica), but because they do not permeate the skin well, the application of green tea in cosmetic products has so far been limited. This study aims to evaluate the cutaneous absorption of catechins from an extract of green tea and from a green tea extract-loaded chitosan microparticle. The catechin skin metabolism was also examined. The results suggest that chitosan microparticles significantly improve the ability of catechins to permeate skin. The cutaneous metabolism of the catechins significantly affected their permeation profiles. Epicatechin (EC) and epigallocatechin (EGC) penetrated the skin more than epigallocatechin gallate (EGCG) and epicatechin gallate (ECG). The galloyl groups in EGCG and ECG were enzymatically hydrolysed to EGC and EC, respectively. Dehydroxylation of catechins was also observed. Chitosan microparticles effectively prevented enzymatic changes of the catechins; therefore, chitosan microparticles are here found to be the promising carriers for enhancing the skin permeation.