柯拉斯那沉香中2-(2-苯乙基)色酮类化学成分研究

为研究柯拉斯那(Aquilaria crassna Pierre ex Lecomte)沉香的化学成分。实验采用多种柱色谱方法从该沉香中分离得到9个2-(2-苯乙基)色酮类化合物,通过现代波谱学技术分别鉴定为6-甲氧基-2-[2-(3′-羟基-4′-甲氧基苯基)乙基]色酮(1)、5-羟基-6-甲氧基-2-[2-(3′-羟基-4′-甲氧基苯基)乙基]色酮(2)、tetrahydrochromone F(3)、6-甲氧基-2-[2-(3′-甲氧基-4′-羟基苯基)乙基]色酮(4)、6-甲氧基-7-羟基-2-[2-(4′-甲氧基苯基)乙基]色酮(5)、6,7-二甲氧基-2-[2-(3′-羟基-4′-甲氧基苯基)乙基]色酮(6)、6,7-二甲氧基-2-[2-(4′-甲氧基苯基)乙基]色酮(7)、6-羟基-2-[2-(4′-羟基苯基)乙基]色酮(8)、5-羟基-2-[2-(2′-羟基苯基)乙基]色酮(9)。化合物2、3和5～9均为首次从柯拉斯那所得沉香中分离得到。采用MTT法对单体化合物的细胞毒活性进行测试,测试结果表明,化合物1,2和4具有微弱的细胞毒活性。     

2种新型ω-芋螺毒素的合成及作用靶点鉴定

目的:合成2个来自泡芋螺的新型ω-芋螺毒素Bu1、Bu13,测定其作用靶标,为研制新型镇痛药提供先导化合物。方法:固相合成线性肽,然后在折叠液(0.5 mol/L乙酸铵、1 mmol/L谷胱甘肽、0.1 mmol/L氧化谷胱甘肽)中于4℃折叠24～48 h,富集纯化得目标肽;利用膜片钳技术,测定其对N、P/Q及L型钙离子通道的抑制活性。结果:Bu1、Bu13对N型钙离子通道的半抑制浓度(IC50)分别为0.86、1.02μmol/L,抑制作用低于MⅦA(IC50=0.21μmol/L);10μmol/L Bu1、Bu13对P/Q型钙离子通道的抑制率分别为17.05%±1.34%、13.1%±2.69%,稍高于MⅦA(8.92%±2.12%);10μmol/L Bu1、Bu13对L型钙离子通道的抑制率分别为19.31%±6.22%、4.78%±0.77%,高于MⅦA(<5%)。结论:Bu1、Bu13选择性作用于N型钙离子通道,对P/Q、L型钙离子通道的抑制作用较低。     

解脂耶氏酵母(Yarrowia lipolytica) ATCC30162脂肪酶基因Yllip1和Yllip2的结构及表达特征

以目前报道油脂产量最高的解脂耶氏酵母菌株(Yarrowia lipolytica)ATCC 30162为对象,采用逆转录PCR扩增到脂肪酶编码基因Yllip1和Yllip2,编码产物分别为816和549个氨基酸。保守结构域预测表明,Yllip1包含Patatin类磷脂酶和功能未知的DUF3336结构域,而Yllip2包含lipase_3类脂肪酶结构域,且这两个蛋白都具有1～4个跨膜区域。与不同物种来源的脂肪酶同源蛋白的多序列比对表明Yllip1和Yllip2分别包含8和6个保守区域,这些生物信息学分析表明这两个来源于解脂耶氏酵母的脂肪酶作用底物可能分别为细胞内膜磷脂和酰基甘油酯。荧光定量PCR分析表明:培养基中添加油酸在短期内(6 h)诱导了这两个脂肪酶基因Yllip1和Yllip2的显著上调表达,表明它们可能参与了酵母分解利用油酸的生化过程。     

几种氧代丙酸化合物在诱捕白纹伊蚊中的应用

为了从氧代丙酸化合物中筛选出对白纹伊蚊Aedes albopictus具有良好引诱效果的化合物,采用陷阱诱捕法分别测定了丙酮酸（α-氧代丙酸）、3-甲氧基-2-甲基-3-氧代丙酸、3-(苄氧基)-3-氧代丙酸和3-氨基-3-氧代丙酸4种氧代丙酸化合物及丙酸对白纹伊蚊成虫的诱捕效果。结果表明,在供试的5种化合物中,α-氧代丙酸、3-甲氧基-2-甲基-3-氧代丙酸和3-(苄氧基)-3-氧代丙酸3种氧代丙酸化合物及丙酸的诱捕效果显著,同剂量的各化合物对白纹伊蚊的诱蚊效果依次是3-甲氧基-2-甲基-3-氧代丙酸>3-(苄氧基)-3-氧代丙酸>α-氧代丙酸=丙酸。相同质量α-氧代丙酸∶3-(苄氧基)-3-氧代丙酸=1∶1至1∶2的配方比,具有协同增效作用,平均累计诱捕量均显著优于单组分α-氧代丙酸、3-(苄氧基)-3-氧代丙酸和对照纯水（P<0.05）。其中最优配方比是α-氧代丙酸∶3-(苄氧基)-3-氧代丙酸=1∶2,平均累计诱捕量比单组分α-氧代丙酸增加45.45%,比单组分3-(苄氧基)-3-氧代丙酸增加32.33%和比对照纯水增加73.11%（P<0.05）。现场诱蚊试验结果表明,根据相似相溶原理将液体的氧代丙酸（α-氧代丙酸、3-甲氧基-2-甲基-3-氧代丙酸）分别与固体的氧代丙酸（3-(苄氧基)-3-氧代丙酸）混配并结合陷阱诱捕器,能够显著提高对白纹伊蚊的持效性及诱捕数量（混合物60 d的累计诱蚊总数量是单组分的1.28~2.15倍）。研究结果为进一步开发适用于家居办公环境中使用的诱蚊剂提供了研究基础。     

二级结构组件与蛋白质耐热性的关系研究

挑选了NCBI COG数据库中具有全基因组的单细胞微生物,选择其中三维结构已知的蛋白质作为研究对象,研究了不同类型的二级结构含量和长度对古细菌和细菌类蛋白质耐热性的影响作用。结果表明:耐热的古细菌类蛋白质中含有相当数量的短的3_(10)螺旋,而耐热的细菌蛋白质中含有较短的loop环。这不仅说明二级结构对蛋白质耐热性有重要的影响,还表明二级结构对古细菌和细菌类蛋白质耐热性的影响作用是不同的。     

正常人耵聍中马拉色菌菌种构成研究

目的研究正常人耵聍中马拉色菌菌种构成及同一宿主耵聍中菌种是否一致。方法采集45名健康志愿者双侧耵聍,0.1%曲拉通X-100溶解稀释后接种于含菜籽油培养基,生化及形态学方法鉴定到种,同时提取菌种DNA,用真菌通用引物ITS1/ITS4做PCR扩增并测序鉴定。结果有44例(97.78%)双侧耵聍中均培养出马拉色菌(共分离出88株菌),菌种构成:糠粃马拉色菌29株(32.95%)、斯洛菲马拉色菌23株(26.14%)、合轴马拉色菌18株(20.45%)、球形马拉色菌11株(12.50%)、限制性马拉色菌7株(7.95%)。44例(88株菌)中双侧耵聍菌种相同者有38例(76株菌)(一致率86.36%)。结论正常人耵聍中马拉色菌菌种分布较广,主要菌种为糠秕马拉色菌。同一宿主双侧耵聍中马拉色菌菌种具有一定的一致性。     

土曲霉致双外耳道曲霉病

目的报道1例发生在双外耳道的曲霉病。方法取双耳耵聍行真菌直接镜检,将耵聍接种到沙堡培养基培养,并对培养出的病原菌进行形态学鉴定和rDNA序列分析。结果耵聍真菌直接镜检阳性,耵聍在沙堡培养基25℃培养长出沙褐色菌落,28℃小培养后光镜及扫描电镜下见分生孢子头呈柱状,顶囊半球形,直径约10μm,小梗双层,两层小梗的长度无明显差别,平行紧密生长在顶囊的上2/3处。分生孢子呈球形,小而光滑。分生孢子梗壁光滑、透明。提取真菌总DNA用PCR方法扩增rDNA序列,测序后登录Gene Bank进行比对,该菌与土曲霉菌株ATCC1012 rDNA序列一致性达100%,鉴定为土曲霉。患者外用4%氟康唑注射液直接滴耳,2次/d,治疗1个月后痊愈。结论确诊1例双外耳道土曲霉病,局部应用氟康唑注射液治疗外耳道曲霉病有效。     

Effects of quorum sensing autoinducer degradation gene on virulence and biofilm formation of Pseudomonas aeruginosa

The aiiA gene from Bacillus thuringiensis was cloned into the Pseudomonas/E. coli shuttle vector and transformed into Pseudomonas aeruginosa strain PAO1. Western blotting showed that the AiiA protein was expressed in PAO1. After induction by IPTG for 6 h and 18 h, expression of the aiiA gene in PAO1 completely degraded the quorum sensing autoinducers N-acylhomoserine lactones (AHLs): N-oxododecanoyl-L-homoserine lactone (OdDHL) and N-butyryl-L-homoserine lactone (BHL). The reduced amount of AHLs in PAO1 was also correlated with decreased expression and production of several virulence factors such as elastase and pyocyanin. AiiA expression also influenced bacterial swarming motility. Most importantly, our studies indicated that aiiA played significant roles in P. aeruginosa biofilm formation and dispersion, as observed by the differences of the biofilm formation on liquid and solid surfaces, and biofilm structures under a scanning electron microscope. These authors contributed equally to this work Supported by the National Natural Science Foundation of China (Grant No. 30570020) and Natural Science Foundation of Hubei Province of China (Grant No. 2004ABA120)     

Con A affinity glycoproteomics of normal human liver tissue

In order to establish the novel high throughput, high efficiency and Iow cost technological platform for the research of N-glycoproteomics, to resolve the significance of characteristic expression profile of glycoprotein and to find the proteins with biological functional importance, the glycoproteins with high-mannose core and the two antennary types were purified and enriched by the Con A affinity chromatography. Con A affinity protein expression profiles of normal human liver tissue were generated by using SDS-PAGE, two-dimensional electrophoresis (2-DE) followed by fast fluorescence staining based on multiplexed proteomics (MP) technology. 301 visible protein spots on the gel were detected and 85 of glycoproteins were further successfully identified via peptide mass fingerprinting (PMF) by a matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS/MS) and annotated to IPI databases. Identified glycoproteins definitely take part in the regulation of cell cycle and metabolic processes. The glycosylation sites were predicted with NetNGlyc 1.0 and NetOGlyc 3.1 software, meanwhile they were classified according to the geneontology methods. The construction of Con A affinity glycoprotein database of normal human liver tissue would contribute to the subsequent research.