负载金属离子的5A 沸石的体外抗菌实验

目的:探讨负载金属离子的5A沸石的体外抗菌作用。方法:选用金黄色葡萄球菌、铜绿假单孢菌、白色念珠菌,利用倍比稀释法对5A沸石组、磺胺嘧啶银组及载不同浓度Ag+、Zn2+5A沸石共15组进行了体外抗菌试验研究,确定最佳抗菌效果离子负载方案。结果:三种细菌的MIC,双金属离子负载在抗菌上具有协同载Ag+5A沸石分别达到了125μg/ml~500μg/ml、31.25μg/ml~500μg/ml、250μg/ml~500μg/ml;附载Zn2+5A沸石分别达到了12.5mg/ml~25mg/ml、6.25mg/ml~50mg/ml、25mg/ml;负载双离子5A沸石分别为250μg/ml~500μg/ml、62.5μg/ml~500μg/ml、500μg/ml。结论:5A沸石负载金属离子后均具有抗菌作用,且抗菌作用与附载金属离子的量正相关;负载相同质量Ag+的5A沸石较附载Zn2+者具有更强的抗菌作用;双金属离子负载在抗菌上具有协同作用;2%Ag++8%Zn2+与2%Ag++10%Zn2+及4%载银组与阳性对照组无显著性差异。     

五种重金属离子对克氏原螯虾（Procambarus clarkiai）的急性毒性作用研究

采用静水生物法、充气和恒温法研究了5种重金属离子对克氏原螯虾的急性毒性作用,并以直线内插法得出了24 h、48 h、72 h和96 h的半致死量（LD50）.研究结果表明：重金属离子的毒性顺序由大到小依次为：Hg2＋、Cd2＋、Cu2＋、Pb2＋和Zn2＋.根据毒性分级标准,5种重金属离子对克氏原螫虾均为高毒物,其安全质量浓度依次为：0.0143 mg/L、0.0322 mg/L、0.0401 mg/L、0.1995 ms/L和0.2795 mg/L.以浓度对数和死亡率进行回归分析,分别得出了5种重金属离子对螯虾24 h、48 h和96 h的回归方程.     

不同浓度尼古丁对牙周膜成纤维细胞增殖及纤维结合蛋白合成的作用

目的:探讨不同浓度尼古丁对人牙周膜成纤维细胞(PDLFs)增殖及纤维结合蛋白(Fn)合成的作用。方法:不同浓度的尼古丁(50 ng/ml,250 ng/ml,500 ng/ml,1μg/ml,2μg/ml,3μg/ml)作用于PDLFs,MTT比色法检测细胞增殖活性,流式细胞仪检测细胞周期,酶联免疫吸附测定法(ELISA)检测Fn合成含量。结果:不同浓度尼古丁作用下:人PDLFs的增殖均被抑制,且呈浓度依赖性。浓度为250 ng/ml~3 ug/ml的尼古丁有明显抑制人PDLFs增殖的作用(P0.05),3 ug/ml尼古丁显示出最强的抑制增殖作用(P0.01);人PDLFs的G0-G1期、S期、G2-M期与对照组相比,G0-G1期细胞周期分布比例逐渐增高,S期和G2-M期逐渐降低,差异均有统计学意义,呈浓度依赖性;人PDLFs合成Fn逐渐减少,呈浓度依赖性。浓度为50 ng/ml~3μg/ml的尼古丁均有明显抑制人PDLFs合成Fn的作用(P0.05),其中3μg/ml抑制作用最强。结论:尼古丁抑制牙周膜细胞的增殖及Fn的合成,呈浓度依赖性,并影响其细胞周期的进程,进而影响牙周新附着的形成,加重牙周病的病情。     

医院内鼠伤寒沙门氏菌流行株耐药质粒的分析

采用质粒及其酶切片段的指纹分析,配合以生化反应、血清学鉴定、最小抑菌浓度(MIC)测定,就我院一次鼠伤寒沙门氏菌的暴发流行中任选的7个菌株进行了分子分析。所有流行株均有相同的生化反应谱,血清学鉴定均属0．Hi型。按Mic结果及质粒指纹分析,可将这七个株分成两组,甲组：5个株耐青霉素(PC>500mg/ml),氨苄青霉素(AP,>500μg/ml),头孢唑啉(Cz,16μg/ml),红霉素(Er,>500μg／ml),氯霉素(Cm,250μg／ml),链霉素(Sm,>500μg／ml),卡那霉素(Km,>500μg/ml)及四环素(Tc,>500μg／ml)8种抗生素。     

胡萝卜苷抑制HepG-2细胞增殖和迁移及拓扑异构酶表达

目的探讨胡萝卜苷对HepG2细胞增殖、迁移及DNA拓扑异构酶(topoisomerase,TOPO)Ⅰ和Ⅱ表达的影响。方法体外培养人肝癌HepG2细胞。CCK-8实验观察不同浓度(200μg/ml、150μg/ml、100μg/ml和50μg/ml)胡萝卜苷对HepG2细胞增殖的影响。qRT-PCR实验观察200μg/ml胡萝卜苷对HepG2细胞TOPOⅠ和TOPOⅡmRNA表达的影响。Transwell小室模型检测200μg/ml胡萝卜苷对HepG2细胞迁移的影响。结果胡萝卜苷对HepG2细胞增殖有明显抑制作用,并在一定范围内(0μg/ml~200μg/ml)呈现浓度依赖性。与未用胡萝卜苷处理的HepG2比较,经200μg/ml胡萝卜苷作用的HepG2细胞穿越基膜的数量显著减少,TOPOⅠ和TOPOⅡmRNA相对表达量显著降低。结论胡萝卜苷可能通过下调TOPOⅠ、TOPOⅡ基因表达而抑制HepG2细胞增殖,并具有抑制HepG2细胞迁移的能力。     

树状多节孢发酵生产紫杉醇工艺条件的初步研究

研究了树状多节孢HQD33内生真菌融合子TPF－1瓶发酵工艺条件,进行了2．8L和10L通用式机械搅拌罐的发酵试验。结果表明,HQD33适宜发酵工艺条件是：发酵时间16－18d,培养基中蔗糖、苯丙氨酸、醋酸钠、酪氨酸,2,4-氯苯氧乙酸和亚油酸的加量分别为180g/L、1mg/L,1.5g/L,15mg/L,5mg/L和15mg/L,摇瓶装置为150ml/500ml,在此条件下摇瓶发酵液中紫杉醇平均含量为448．52μg/L;2.8L和10L罐发酵液中紫杉醇含量达406．95和395．12μg/L（平均值）。     

Ⅰ型胶原蛋白在云母表面自组装的AFM研究

目的：利用原子力显微镜(AFM)观察不同浓度、pH值的I型胶原溶液在云母表面的自组装情况,从而研究样品浓度和酸度对胶原蛋白自组装的影响。方法：配制0．8μg/ml、8μg/ml、0．5mg/ml、1．0mg/ml、4mg/ml的胶原醋酸溶液(pH值2．7—3.0)和0.2mg/ml、0．5mg/ml、1．0mg/ml的胶原PBS溶液(pH值7．2—7．4)。取5μl的胶原蛋白溶液,滴于新剥离的云母表面,空气中在25℃室温下自然干燥后用AFM观察。结果：酸性条件下,低浓度时胶原蛋白在云母表面只是简单的吸附,随着浓度的增加,胶原分子在云母表面开始组装形成网状的结构,当浓度达到饱和浓度时,胶原形成无规线团结构。在中性条件下,胶原更容易行成纤维,随着浓度的增加,纤维的直径和长度都随着增加。结论：胶原蛋白分子有在固体表面进行自组装的特性,自组装的结果与样品的浓度、样品溶液的pH值有着很大的关系。     

金属离子抑制细菌整合子捕获耐药基因盒的机制研究

【目的】探索金属离子对整合子捕获耐药基因盒的影响及其相关机制。【方法】在大肠埃希菌中构建一个1类整合子捕获耐药基因盒的体内模型。通过实时荧光定量PCR(real-time fluorescence quantitative PCR,RT-qPCR)测定不同浓度银(0.3、0.9、1.5μg/mL的Ag+)和铜离子(5、50、100、150、210μg/mL的Cu²⁺)干预后实验组和无金属离子干预对照组整合子整合频率,并用表型筛选法验证。利用质谱法测量细菌吸收的金属离子浓度,并进一步通过转录组测序方法分析银离子抑制细菌整合子捕获耐药基因盒的分子机制。【结果】qPCR和表型筛选法的结果表明,0.9μg/mL银离子组整合频率为9.42×10^-5 (6.49×10^-5,1.44×10^-4),1.5μg/mL银离子组整合频率为7.29×10^-5 (4.45×10^-5,9.03×10^-5),与对照组2.59×10^-4 (2....     

金属离子对酵母胞内核黄素产量的影响

研究了金属离子对脆壁酵母 (Saccharomycesefragilis)RY 5胞内核黄素积累的影响 ,运用均匀设计方法进行了培养基优化。结果表明 :Mg2 + ,Zn2 + ,Fe2 + 对RY 5的核黄素产量有着显著影响 ,培养基中金属离子的最佳浓度为 :MgSO4·7H2 O 1 .1g/L ;CoSO4·7H2 O 2 8mg/L ;CuSO4·5H2 O 0 .0 1mg/L ;MnCl2 0 .0 2mg/L ;ZnSO4·7H2 O 3 4mg/L ;FeSO4·7H2 O 1 4mg/L ,RY 5的核黄素产量可达 1 40mg/kg。     

不同补料发酵方式对发夫酵母产虾青素的影响

对发夫酵母的不同补料发酵方式进行了研究,期望提高发酵产率,以便对工业化生产提供一定的指导作用。实验结果表明：采用恒pH葡萄糖-氨水流加培养,色素量和生物量均具有最大值,分别为54.3μg/ml和49.5μg/ml;其次是指数流加培养,色素量和生物量分别为49.9μg/ml和47.4mg/ml;恒pO2流加方式下色素量和生物量分别为34.04ug/ml和35.4mg/ml;恒pH葡萄糖流加方式所得的色素量和生物量最小,分别为32.3μg/ml和31.5mg/ml。不同的补料发酵方式对发夫酵母生长和色素形成的影响很大。     

Rapid reduction of Legionella pneumophila on stainless steel with zeolite coatings containing silver and zinc ions

AIMS: To determine the rate of reduction of Legionella pneumophila by stainless steel surfaces with zeolite ceramic coatings containing 2.5% (w/w) silver (Ag) and 14% zinc (Zn) ions. METHODS AND RESULTS: Stainless steel pans with and without Ag/Zn coatings were inoculated with solutions of Leg. pneumophila ATCC 33155 and incubated at 37 degrees C. Survival was monitored using the spread-plate technique on selective buffered charcoal yeast extract agar. Significant reductions of Leg. pneumophila were effected by the Ag/Zn zeolite coatings within 2 h of exposure. CONCLUSIONS, Significance and Impact of the Study: Zeolite ceramic Ag/Zn coatings impart significant anti-Legionella properties to stainless steel surfaces. Coated stainless steel could be used in the manufacture of air ducts, condensation pans and intake and exhaust vents. These products have the potential to reduce numbers of Legionella in air-handling systems.     

Reactivation of apo horse liver alcohol dehydrogenase with the monovalent metal ion Ag(I)

Each subunit of the liver alcohol dehydrogenase dimer contains one catalytic and one structural Zn(II) atom. Enzyme with the catalytic metal atoms selectively removed is inactive but can be partly reactivated in the presence of Ag(I) ions. Reactivation results from Ag(1) ions entering the empty metal-binding site in the catalytic center. The specific activity of this silver enzyme reached 24% of the native enzyme. Atomic absorption analysis gave equal amounts of Ag(I) and Zn(II), corresponding to one mole of each metal per monomer. Metal-directed affinity labelling using bromo-imidazolyl propionate showed that the properties of the silver-reactivated enzyme were distinct from those of the native enzyme.     

Preparation of chitosan-nylon-6 blended membranes containing silver ions as antibacterial materials

Chitosan-nylon-6 blended membranes were prepared by combining solvent evaporation and a phase inversion technique, and then used to chelate silver ions. Gram-positive bacteria (Staphylococcus aureus) and Gram-negative bacteria (Escherichia coli) were used to study the antibacterial properties of the membranes. Fourier-transform infrared spectroscopy (FTIR) and differential scanning calorimetry (DSC) indicated hydrogen-bond interactions between chitosan and nylon-6. From the scanning electron microscopy (SEM) pictures, it was observed that with the increase of nylon-6 content, the blended membrane gradually became a material with porous morphology. After chelating silver ions, the tensile strength of the membranes increased. The antibacterial activity with the variation of chitosan content, the pH value and the concentration of the silver nitrate solution used to prepare Ag(+)-loaded membranes were investigated systematically. The results indicated that the chitosan-nylon-6 blended membranes with Ag(+) were antibacterial to both Gram-positive bacteria and Gram-negative bacteria. The antibacterial activity improved with the increased chitosan content due to the larger amount of silver ions loaded. The antibacterial property of the chitosan-nylon-6 blended membranes could be primarily attributed to the content of chitosan and silver ions as well as the surface morphology of the membranes.     

一个外切菊粉酶基因的克隆表达及酶学性质

从马克斯克鲁维酵母（Kluyveromycesmarxianus）DSM5418中克隆出外切菊粉酶（INU）的成熟肽编码区域,在毕赤酵母（Pichiapastoris）GS115中实现了高效分泌表达,体积酶活力达到15．27U／mL,进一步对重组酶进行了纯化与表征。经过（NH4）2SO4沉淀、透析和分子筛过滤后,得到了纯度大于95％的纯化重组酶,SDS-PAGE分析发现INU的表观相对分子质量为9．0×10^4,大于理论预测值6．0×10^4。纯化酶液的表征结果表明,INU的最适温度和最适pH分别为55℃和5．0,在此条件下INU对菊粉的K。值和比酶活分别为1．90mmol／L和433．86U／mg,对蔗糖的K。值和比酶活分别为27．81mmol/L和1249．49U／mg,I/S值为0．34;HPLC分析表明,INU酶解菊粉的产物由果糖和葡萄糖组成;金属离子Mn2＋、Fe3｜、K｜和Co2＋对酶有促进作用,而Zn2＋、Cu2＋、Ni2＋、SDS和EDTA对酶活力有不同程度的抑制作用。     

表达条件对金属激活酶NAOase离子依赖的活性分析

N-乙酰鸟氨酸脱酰基酶为一种新型手性拆分酶制剂,酶活性依赖于Mg2＋、Mn2＋、Zn2＋及Co2＋ 中的某种金属离子。以高表达NAOase的重组菌DH10B/argE-pHsh为研究对象,考察不同培养条件下Mg2＋、Mn2＋、Zn2＋及Co2＋ 4种离子对重组菌的生长、酶的表达活性及表达量的影响。结果发现：同种离子在不同条件下对重组菌的生长影响不大,但对酶活性影响显著。4种离子在合适浓度时皆能提高酶活性,促进强度从高到低依次为Mn2＋、Mg2＋、Co2＋和Zn2＋。在TB培养基中,Mn2＋为15 mmol/L时：NAOase比酶活达到1 272.7 U/mL,是未添加时的4.67倍,激活作用显著高于其他离子。SDS-PAGE电泳实验表明4种离子的蛋白表达量基本相同。     

重组textlinin-1抑制活性的性质研究

目的对毕赤酵母表达的textlinin-1(Q8008)进行分离纯化,并对其抑制plasmin活性的性质进行研究。方法构建表达Q8008pachia pastoris工程菌,经发酵、纯化得到纯度达95%以上的Q8008。考察不同的温度、pH缓冲液和金属离子等条件对Q8008抑制plasmin活性的影响。结果 Q8008在中性条件下比较稳定,其最适pH值为7.0,最适温度为40℃,在30~50℃相对酶活力在90.0%以上;有2个金属离子对Q8008活性测定方法的影响产生了显著性差异,Zn2+起激活作用,Co2+起抑制作用,其余金属离子均无显著性差异。结论 Q8008在中性pH下抑制活性稳定;在50℃以下,受温度变化影响较小;金属离子对其抑制活性测定有一定影响。     

毛果杨HMA基因家族的生物信息学分析

重金属转运ATP酶（heavy metal transporting ATPase,HMA）是一种通过水解ATP跨膜运送重金属阳离子的转运蛋白,属于P-ATPase家族中一个亚类。不同HMA蛋白对重金属离子的转运具有选择性,在植物修复重金属污染土壤方面起着重要作用。依据毛果杨全基因组测序的结果,以及HMA基因蛋白的序列和功能特征,从毛果杨基因组中鉴定了13个HMA基因家族成员,分属于Zn亚类（Zn2＋/Co2＋/Cd2＋/Pb2＋P1B-ATPase）和Cu亚类（Cu＋/Ag＋P1B-ATPase）两个亚家族,主要分布于1、3号染色体上。生物信息学分析表明,毛果杨HMA基因的氨基酸序列一致性介于21.3%~89.3%,且具有保守的基序CPC、HP、DKTGT、TGEx、GDG、PxD和CxxC等。蛋白理化特征分析显示,多数毛果杨HMA蛋白稍偏酸性,结构稳定性较好,蛋白脂溶指数高,稍具疏水性。密码子偏好性分析显示,毛果杨HMA蛋白14个氨基酸中存在16个高频密码子,另有1个终止密码子为高频密码子,显示出毛果杨的种属特征。研究结果展示了毛果杨HMA基因家族的基本信息和特点,为深入研究毛果杨HMA基因的功能搭建了基础平台。     

The antibacterial activity of biogenic silver and its mode of action

In a previous study, biogenic silver nanoparticles were produced by Lactobacillus fermentum which served as a matrix preventing aggregation. In this study the antibacterial activity of this biogenic silver was compared to ionic silver and chemically produced nanosilver. The minimal inhibitory concentration (MIC) was tested on Gram-positive and Gram-negative bacteria and was comparable for biogenic silver and ionic silver ranging from 12.5 to 50 mg/L. In contrast, chemically produced nanosilver had a much higher MIC of at least 500 mg/L, due to aggregation upon application. The minimal bactericidal concentration (MBC) in drinking water varied from 0.1 to 0.5 mg/L for biogenic silver and ionic silver, but for chemically produced nanosilver concentrations, up to 12.5 mg/L was needed. The presence of salts and organic matter decreased the antimicrobial activity of all types of silver resulting in a higher MBC and a slower inactivation of the bacteria. The mode of action of biogenic silver was mainly attributed to the release of silver ions due to the high concentration of free silver ions measured and the resemblance in performance between biogenic silver and ionic silver. Radical formation by biogenic silver and direct contact were found to contribute little to the antibacterial activity. In conclusion, biogenic nanosilver exhibited equal antimicrobial activity compared to ionic silver and can be a valuable alternative for chemically produced nanosilver.