Using ion-template method to prepare chitosan nanoparticle for MMC sustained and targeted delivery

近二十年来，药物缓控释系统是国际医药界研究和开发的重点技术之一，该技术通过改变药物的剂型或给药方式，可以解决传统抗肿瘤药物用药量大、毒性大和副作用大等缺点。壳聚糖及其衍生物在药物释放体系中具有广泛的应用前景。将药物与壳聚糖及其衍生物偶联或者用它们来包裹抗肿瘤药物可以制成抗肿瘤药物的控缓释制剂。 本文的主要研究包括：(1)采用离子模板法，即将离子凝胶和化学交联相结合的方法制备粒径可控，稳定性较好的壳聚糖纳米载体，考察三聚磷酸钠与壳聚糖的质量比、pH以及分子量对纳米粒子大小的影响；使用动态光衍射法监测纳米粒子粒径随时间的变化情况，以及使用扫描电镜等对粒子的微观形态等进行表征。(2)对壳聚糖纳米载...Sustained/Controlled release drug delivery system, one of the fast-developing technologies over the past two decades in international medical community, by changing the dosage form or drug delivery methods, can overcome the shortcomings of traditional anti-cancer drugs. Chitosan and its derivatives are very promising candidates as sustained/controlled release carrier-materials for a variety of dru...学位：理学博士院系专业：材料学院生物材料系_生物医学工程学号：2452007115251

Protein fold pattern recognition based on ensemble classifiers

蛋白质折叠问题被列为“21世纪的生物物理学“的重要课题,他是分子生物学中心法则尚未解决的一个重大生物学问题,因此预测蛋白质折叠模式是一个复杂、困难、和有挑战性的工作。为了解决该问题,我们引入了分类器集成,本文所采用的是三种分类器(lMT、rAndOMfOrEST、SMO)进行集成以及188维组合理化特征来对蛋白质类别进行预测。实验证明,该方法可以有效表征蛋白质折叠模式的特性,对蛋白质序列数据实现精确分类;交叉验证和独立测试均证明本文预测准确率超过70%,比前人工作提高近10个百分点。Protein folding problem is listed as an important issue of 21st century bio-physics and it is a major unresolved biological problem for the central dogma of molecular biology,so predicting protein folding model is a complex,difficult and challenging work.It is introduced an ensemble classifier to solve such this problem.We use three classifiers(LMT,Random Forest,and SMO) and the 188-dimensional combination of characteristics algorithm for analyzing and predicting the class of protein fold pattern.Experiments show that it can predict the type of protein fold pattern effectively by using the ensemble classifier and classify the experimental data accurately.Crossvalidation and independent testing have proved that the accuracy rate can be higher than 70%,improving by close to 10 percent than previous work.国家自然科学基金(61001013;61102136);福建省自然科学基金(2011J05158

Preparation of Chitosan Nanoparticles for Targeting and Sustained Drug Delivery System by Ion-induced Combined with Chemical Crosslinking

目的制备粒径相对可控的壳聚糖纳米粒,探索其在药物缓释体系中的应用。方法以多聚磷酸钠为离子诱导剂、戊二醛为化学交联剂,通过离子诱导结合化学交联法,制备壳聚糖纳米粒;在1-乙基-(3-二甲氨基丙基)碳二亚胺盐酸盐(EdC)作用下,分别进行丝裂霉素载药以及聚乙二醇(PEg),叶酸修饰;另外,对载药纳米粒子进行不同PH条件下的体外释药实验;对PEg,叶酸修饰的壳聚糖纳米粒罗丹明b荧光标记后,进行激光共聚焦以及活体成像实验。结果离子诱导交联法可以获得粒径范围在200--500 nM的壳聚糖纳米粒;丝裂霉素载药量可以达到25%,包封率为50%,体外释药呈现突释和缓释双相特征,并且随着PH的升高释药明显加快;未经修饰的、叶酸修饰的以及PEg和叶酸修饰的纳米粒都能有效的进入HElA细胞,而单独PEg修饰的却很少进入细胞内;叶酸修饰的纳米粒有明显的靶向作用,PEg修饰的纳米粒可以明显延长实验裸鼠血液中循环时间。结论采用离子诱导结合化学交联法可以获得粒径可控、稳定、适合于主动靶向给药的壳聚糖纳米粒。OBJECTIVE To prepare chitosan nanoparticle with controllable size and investigate its applications to targeting and sustained drug delivery system.METHODS The combination of an ion-induced and chemical crosslinking method was used to prepare chitosan nanoparticles,and mitomycin C,PEG and folic acid were bound covalently on the surface of chitosan nanoparticles in the presence of 1-ethyl-3-(3-dimethyllaminopropyl) carbodiimide hydrochloride(EDC),respectively.In vitro drug release test was carried out in different pH PBS solution.Confocal laser scanning microscope of Hela cell and in vivo imaging test were carried out using PEG and folic acid-modified chitosan nanoparticles labeled by Rhodamine B.RESULTS The diameter was in the range of 200 and 500 nm,the drug-loading capability and entrapment efficiency were 25% and 50%,respectively.The drug release was somewhat biphasic with an initial burst effect,followed by a subsequent slower release.Furthermore,increasing pH of the medium resulted in a faster release rate.Blank chitosan nanoparticles,folic acid modified and both folic acid and PEG-modified nanoparticles were effectively uptaken by Hela cells,while the single PEG-modified nanoparticles rarely entered the cell.By live imaging techniques,folic acid-modified nanoparticles had a clear targeting character.While,the PEG-modified chitosan nanoparticles enhanced circulation time in the bloodstream of mice.CONCLUSION By the method of ion-induced combined with chemical crosslinking,steady chitosan nanoparticles with controlable size were obtained and suitable for active targeting drug delivery.国家重大基础研究计划项目(973项目)资助(2006CB933300);厦门市科技计划项目(3502Z20093009

Research advances of electrochemical micro/nanofabrication based on confined etchant layer technique

与机械加工相比, 电化学加工技术具有无刀具磨损、无热效应、无机械损伤、加工效率高等优点, 而且适用于柔性、脆性及超硬材料,; 具备传统方法难以实现的复杂结构加工能力, 因而在航空航天、汽车、微电子等领域有着重要应用, 日益成为一种重要的工业制造技术.; 随着超大规模集成电路(ULSI)、微机电系统(MEMS)、微全分析系统(mu-TAS)、现代精密光学系统等高技术产业的迅速发展,; 功能性结构/器件的微型化和集成化的要求越来越高. 由于传统电化学只适用于金属材料, 为了应对微纳制造的时代要求, 拓展电化学加工的材料普适性,; 1992年田昭武院士提出了具有我国自主知识产权的约束刻蚀剂层技术(CELT). 一般的, 约束刻蚀包括3个步骤:; (1)通过电化学、光化学或光电化学的方法在模板电极表面生成刻蚀剂;; (2)通过后续的均相化学反应或自由基衰变反应将刻蚀剂约束在微/纳米厚度的液层内; (3)将模板电极逼近加工基底,; 当约束刻蚀剂层接触被加工基底时, 通过刻蚀反应实现微纳加工. 最近, 联合课题组通过仪器、原理和方法3个方面的努力, 引入外部物理场调制技术,; 实现一维铣削、二维抛光、三维微/纳结构加工, 大幅提升了CELT的技术水平.Compared with mechanical machining, ECM has several advantages, such as; avoiding tool wear, none thermal or mechanical stress on machining; surfaces, as well as high removal rate. Moreover, ECM is capable of; making complex three-dimensional structures and is appropriate for; flexible, fragile, or fissile materials even materials harder than the; machining tool. Thus, ECM has been widely used for various industrial; applications in the fields of aerospace, automobiles, electronics, etc.; ECM methods can be classified usually as electrolytic machining based on; anodic dissolution and electroforming based on cathodic deposition of; metallic materials. Recently, high technology industry, such as; ultralarge scale integration (ULSI) circuits, microelectromechanical; systems (MEMS), miniaturized total analysis systems (mu-TAS) and; precision optics, has developed more and more rapidly, where; miniaturization and integration of functional components are becoming; significant. Nowadays, the feature size of interconnectors in ULSI; circuits has been down to 20 nanometers, predicted by Moore's law.; Confined etchant layer technique (CELT) was proposed in 1992 to; fabricate three-dimensional micro- and nanostructures (3D-MNS) on; different metals and semiconductors, which has been developed an; effective machining method with independent intellectual property; rights. Generally, there are three procedures in CELT: (1) generating; the etchant on the surface of the tool electrode by electrochemical or; photoelectrochemical reactions; (2) confining the etchant in a depleted; layer with a thickness of micro- or nanometer scale; (3) etching process; when the tool electrode is fed to the workpiece, which applicable for 1D; milling, 2D polishing, and 3D microfabrication with an accuracy at micro; or nanometer scale. External physical-field modulations have recently; been introduced into CELT to improve its machining precision. In this; review, the advances of CELT in principles, instruments and applications; will be addressed as well as the prospects.国家自然科学基金; 机械制造系统工程国家重点实验室开放课题资助项

超声波对球等鞭金藻脂肪酸组成的效应研究

国家自然科学基金!39870565;;福建省自然科学基金!C9700

Preparation and Characterization of Folic Acid and PEG Conjugated Chitosan Nanoparticles

目的利用离子交联和化学交联相结合的方法制备壳聚糖纳米粒子(nPS),并对nPS分别进行了叶酸(fA)和聚乙二醇(PEg)的修饰。方法通过红外光谱进行结构验证;用扫描电镜和粒度分析仪对粒子的微观形态、粒径、电位等进行了表征;通过与HElA细胞摄取实验对其靶向作用进行验证。结果离子交联和化学交联相结合的方法制备壳聚糖纳米粒子粒径在200 nM左右并且粒径分布窄,修饰后的nPS(fA-nPS、PEg-nPS及fA+PEg-nPS)粒径不受功能基团修饰的影响。激光共聚焦试验证明fA-nPS及fA+PEg-nPS能显著提高细胞对粒子的摄取,而PEg-nPS则明显降低其对粒子的摄取。结论 fA+PEg-nPS有望成为一种新型的药物载体,用于抗癌药物对癌细胞的主动靶向。OBJECTIVE An ionic gelation combined with chemical crosslinking method was developed to prepare chitosan nanoparticles,followed by conjugation with folate(FA) and polyethylene glycol(PEG).METHODS The structures were verified by infrared spectroscopy.The morphology,diameter and Zeta electric potential of the nanoparticles were assayed by environmental scanning electron microscope and scattering particle analyzer.The specificity of the FA+PEG-NPs targeting cancer cells was demonstrated by human adenocarcinoma Hela cells.RESULTS The chitosan NPs presented a narrow size distribution with an average diameter about 200 nm regardless of the type of functional group.Laser confocal scanning imaging proved that both FA+PEG-NPs and FA-NPs could greatly enhance uptake by Hela cells.However,the PEG-NPs showed contrary results.CONCLUSION FA+PEG-NPs can be applied as a new vehicle to actively deliver anticancer drugs to tumor cells.厦门市科学技术计划资助项目(3502Z20114007

Construction and identification of interference plasmid targeting on TNFAIP8

目的:构建并筛选出干扰效率最佳的TnfAIP8-SHrnA-P SIrEn-rETrO Q干扰质粒。方法:通过生物软件选择3个TnfAIP8基因干扰位点,构建干扰质粒并测序验证,将干扰质粒及对照质粒分别转染至A549细胞,通过rT-PCr、WESTErn blOT检测干扰效率。结果:经rT-PCr和WESTErn-blOT证实TnfAIP8-SHrnA-P SIrEn-rETrO Q干扰质粒能有效干扰并抑制细胞内TnfAIP8基因的表达,通过流式检测发现降低TnVAIP8表达可以提高细胞对A dr5SC fV诱导凋亡的敏感性。结论:成功构建和设计了对TnfAIP8基因具有显著干扰效率的干扰质粒,为进一步研究TnfAIP8基因的功能奠定了基础。Objective: To construct and screen the high efficiency interference plasmid of TFAIP8-shRNA-p SIRENRetro Q.Methods: Selected and synthesized three Target Sequence of TNFAIP8 shRNA1,TNFAIP8 shRNA2,TNFAIP8 shRNA3,and construct the TNFAIP8 interference plasmid.Transfection TNFAIP8-shRNA-p SIREN-Retro Q interference plasmid to A549 cells.Filter out the highest interference efficiency plasmid by detecting the mRNA and protein levels using RT-PCR and Western blot methods.Results: We successfully design and built three TNFAIP8-shRNA-p SIREN-Retro Q interference plasmids,and screen out the highest efficiency interference plasmid.Conclusion: Three interference plasmids targeting the TNFAIP8 gene have been constructed successfully and provide a useful tool for studying the function of TNFAIP8.国家自然科学基金项目(81272720); 福建省卫计委医学创新课题(2014-CXB-43); 厦门市科技计划项目(3502Z2083008)资

Preparation of Highly Active CuO-CeO2 Catalysts and Their Catalytic Performance for Benzene Oxidation

挥发性有机物是环境污染源中一类重要的污染源，它不仅会造成环境污染，而且对人类健康危害也很大，因此消除挥发性有机物引起了人们的重视。在所有挥发性有机物中，苯是一种工业生产中非常常见的污染物，由于其毒性大，因此相关法律法规对其排放标准非常严格。如何消除空气中低浓度的苯是目前的研究热点之一。 催化氧化技术是消除工业源排放挥发性有机物的主要技术之一，催化剂的性能是决定这一技术效能的重要因素。在众多催化剂中，CuO-CeO2混合氧化物被认为是一类非常具有应用前景的催化剂。但CuO与CeO2形成固溶体的范围非常小，且CuO耐热性差、易迁移、高温失活严重，因此CuO-CeO2催化剂中CuO的含量通常都较低，导致CuO-CeO2混合氧化物催化剂对挥发性有机物的催化氧化活性难提高。另外，CuO-CeO2热稳定性差，催化剂在高温连续反应下失活非常严重，因此提高CuO-CeO2催化剂的热稳定性也是一个非常重要的方面。 为了解决上述问题，本论文开展了以下几个方面的工作： （1）通过合成具有高比表面积的介孔CuO-CeO2催化剂来提高其对苯的催化氧化活性。这是因为高的比表面积能提供更多的活性中心，而介孔有利于消除催化反应过程中大分子有机物造成的扩散控制效应的影响。与传统模板法不同，本研究中开发的合成介孔的方法主要基于硝酸铵的分解，该方法不仅具有廉价、快速等特点，而且具有一定的通用性，也适用于制备其它介孔二元氧化物体系，如CeO2-ZrO2和Y2O3-ZrO2体系。实验结果表明，本研究中合成的介孔CuO-CeO2催化剂对苯的催化完全氧化反应中表现出较高的活性，脉冲实验表明苯在CuO-CeO2催化剂上的反应行为符合氧化-还原机制。但该介孔CuO-CeO2催化剂在260oC下连续反应10h后，比表面积严重下降，对苯的氧化性能也随之降低。 （2）为了降低比表面积的影响，通过探索合成了CuO-CeO2催化剂新的纳米结构来提高其对苯的催化氧化活性。对这种片状纳米结构的形成机理研究表明，纳米片的形成主要是由纳米颗粒之间的团聚造成的，并且这种方法同样也适用于合成MnO-CeO2二元体系纳米片状结构。虽然CuO-CeO2纳米片催化剂比表面积较小，但由于在这种结构中CuO能充分暴露在催化剂的表面，因此其对苯表现出良好的催化性能。但CuO-CeO2片状催化剂的热稳定性也较差，在240oC连续反应30h后即出现明显失活现象。高度分散的CuO形成块状晶相CuO可能是其催化性能降低的重要原因。 （3）为了提高CuO-CeO2催化剂的热稳定性，主要通过掺杂ZrO2对CeO2进行改性来提高催化剂的稳定性。采用超临界干燥制备的CuO-ZrxCe1-xOy（0<x≤0.5）经过800oC高温老化之后形成了固溶体，不仅具有晶粒小、比表面积高等特点，而且由于CuO-CeO2-ZrO2形成了三元固溶体，产生了氧缺陷，提高了晶格氧的移动性，因此对苯的氧化也表现较好的催化性能。同时，由于CuO渗透进入到CeO2晶格中，提高了CuO和其它氧化物之间的相互作用力，因此降低了其表面迁移能力，使得CuO-ZrxCe1-xOy催化剂对苯的氧化表现出良好的热稳定性。经过800oC高温老化后的CuO-Zr0.5Ce0.5Oy催化剂在260oC下连续反应50 h，几乎没有失活。 （4）由于燃烧反应器的设计通常需要动力学相关数据，因此进一步考察了苯在CuO-Ce0.5Zr0.5Oy催化剂上的动力学行为。采用Power-Rate law模型和Mars-Van Krevelen模型对实验所获得的动力学数据进行了拟合。结果表明，Mars-van Krevelen模型对苯在CuO-Ce0.5Zr0.5Oy催化剂上的动力学数据拟合效果要优于Power-Rate law模型