三种桉叶油化学成分研究

通过水蒸汽蒸油试验,发现已贮藏半年多的风干桉叶,仍有较高的含油量。云南省弥勒产的直杆桉(Eucalyptus maidenii)得油率为4.14％,蓝桉(E.globulus)为3.5％,福建省惠安产的窿缘桉(E.exserta)为1.2％,桉叶油生产厂也可以通过贮藏的风干桉叶生产桉叶油。通过气相色谱和质谱分析,从直杆桉叶油中鉴定出44个组份,蓝桉叶油中鉴定出35个组份,窿缘桉叶油中鉴定出26个组份。三种桉叶油单萜部分的组份基本一致,只是相对含量各有不同。桉油素含量,以直杆桉最高为68.02％,其次是蓝桉为67.54％,窿缘桉最低为34.33％。直杆桉、蓝桉可以怍材、油两用树种,窿缘桉不宜作油用树种。     

Infectivity enhancement by human immunodeficiency virus type 1 Nef is independent of its association with a cellular serine/threonine kinase.

Nef proteins from human immunodeficiency virus type 1 isolate SF2 (HIV-1SF2) and simian immunodeficiency virus isolate mac239 (SIVmac239) have been found to associate with a cellular serine/threonine kinase designated NAK. We have recently shown that the association of Nef with NAK is isolate dependent. To identify the structural basis for Nef-kinase association, several chimeric molecules were constructed between SF2 Nef (binding NAK) and 233 Nef (a primary isolate not binding NAK) and stably expressed in HuT-78 human T cells via retrovirus-mediated gene transfer. The Nef 233/SF2/SF2 chimera in which the N-terminal 37 amino acids of SF2 Nef were replaced by those of 233 Nef showed the same ability as SF2 Nef to bind NAK. The Nef 233/SF2/233 chimera in which the N-terminal 37 amino acids and the C-terminal 72 amino acids of SF2 Nef were replaced by corresponding sequences from 233 Nef completely lost the ability to associate with the kinase activity. Furthermore, replacement of the C-terminal 72 amino acids of 233 Nef with the equivalent SF2 sequence (chimera 233/233/SF2) fully restored kinase association to 233 Nef. These results suggest that (i) the core of Nef is not sufficient for NAK binding, (ii) the C terminus of SF2 Nef contains structural determinants important for association with NAK, and (iii) the failure of 233 Nef to bind NAK is due to a defect in its C terminus. Taking advantage of the C terminus of 233 Nef being nonfunctional and using an infectious clone of HIV-1SF2, we show that association with NAK is not required for Nef-mediated infectivity enhancement. While the strong and reproducible association of some Nef isolates with NAK has been clearly established, the role of NAK in Nef function remains to be fully elucidated.     

Purification and characterization of a novel chlorpyrifos hydrolase from Cladosporium cladosporioides Hu-01

Chlorpyrifos is of great environmental concern due to its widespread use in the past several decades and its potential toxic effects on human health. Thus, the degradation study of chlorpyrifos has become increasing important in recent years. A fungus capable of using chlorpyrifos as the sole carbon source was isolated from organophosphate-contaminated soil and characterized as Cladosporium cladosporioides Hu-01 (collection number: CCTCC M 20711). A novel chlorpyrifos hydrolase from cell extract was purified 35.6-fold to apparent homogeneity with 38.5% overall recovery by ammoniumsulfate precipitation, gel filtration chromatography and anion-exchange chromatography. It is a monomeric structure with a molecular mass of 38.3 kDa. The pI value was estimated to be 5.2. The optimal pH and temperature of the purified enzyme were 6.5 and 40°C, respectively. No cofactors were required for the chlorpyrifos-hydrolysis activity. The enzyme was strongly inhibited by Hg2?, Fe3?, DTT, β-mercaptoethanol and SDS, whereas slight inhibitory effects (5-10% inhibition) were observed in the presence of Mn2?, Zn2?, Cu2?, Mg2?, and EDTA. The purified enzyme hydrolyzed various organophosphorus insecticides with P-O and P-S bond. Chlorpyrifos was the preferred substrate. The Km and Vmax values of the enzyme for chlorpyrifos were 6.7974 μM and 2.6473 μmol·min?1, respectively. Both NH2-terminal sequencing and matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometer (MALDI-TOF-MS) identified an amino acid sequence MEPDGELSALTQGANS, which shared no similarity with any reported organophosphate-hydrolyzing enzymes. These results suggested that the purified enzyme was a novel hydrolase and might conceivably be developed to fulfill the practical requirements to enable its use in situ for detoxification of chlorpyrifos. Finally, this is the first described chlorpyrifos hydrolase from fungus.     

长角血蜱卵泡抑素相关蛋白的定性

参照GenBank中长角血蜱致病性Okayama株卵泡抑素基因的核苷酸序列(GenBank Accession No.DQ248886)设计合成一对引物,从本实验室保藏的单克隆洁净长角血蜱饥饿成蜱中快速提取总RNA,通过RT-PCR扩增出814bp的卵泡抑素基因,序列比对结果显示:与长角血蜱致病性Okayama株的核苷酸序列及氨基酸序列一致性分别为97.8%和99%,将其亚克隆到表达载体pGEX-4T-1中进行表达,GST融合重组蛋白预期分子量为57kD。表达重组蛋白经MagneGSTTM蛋白纯化系统纯化后作为抗原分别与抗不同发育阶段长角血蜱(卵、幼蜱、若蜱、成蜱)多克隆抗体作为一抗进行免疫印迹,结果表明:与长角血蜱卵制备的多克隆抗体有很强的免疫反应,而与其他发育阶段(幼蜱、若蜱、成蜱)饥饿长角血蜱制备的多克隆抗体反应性很弱。以上结果表明:长角血蜱卵泡抑素蛋白在长角血蜱产卵及卵成熟发育时期的表达水平较其他发育阶段(幼蜱、若蜱、成蜱)的蛋白表达水平高。     

A New Approach to Segment Both Main and Peripheral Retinal Vessels Based on Gray-Voting and Gaussian Mixture Model

Vessel segmentation in retinal fundus images is a preliminary step to clinical diagnosis for some systemic diseases and some eye diseases. The performances of existing methods for segmenting small vessels which are usually of more importance than the main vessels in a clinical diagnosis are not satisfactory in clinical use. In this paper, we present a method for both main and peripheral vessel segmentation. A local gray-level change enhancement algorithm called gray-voting is used to enhance the small vessels, while a two-dimensional Gabor wavelet is used to extract the main vessels. We fuse the gray-voting results with the 2D-Gabor filter results as pre-processing outcome. A Gaussian mixture model is then used to extract vessel clusters from the pre-processing outcome, while small vessels fragments are obtained using another gray-voting process, which complements the vessel cluster extraction already performed. At the last step, we eliminate the fragments that do not belong to the vessels based on the shape of the fragments. We evaluated the approach with two publicly available DRIVE (Staal et al., 2004) and STARE (Hoover et at., 2000) datasets with manually segmented results. For the STARE dataset, when using the second manually segmented results which include much more small vessels than the first manually segmented results as the “gold standard,” this approach achieved an average sensitivity, accuracy and specificity of 65.0%, 92.1% and 97.0%, respectively. The sensitivities of this approach were much higher than those of the other existing methods, with comparable specificities; these results thus demonstrated that this approach was sensitive to detection of small vessels.