负载维生素D3的海藻酸纳米粒抗佝偻病作用研究

目的:观察负载维生素D3海藻酸纳米粒和维生素D3的抗佝偻病作用.方法:36只SD大鼠随机分组为正常对照组、佝偻痛模型组、负载VD3的自组装海藻酸钠纳米粒(sSAN-VD3)低剂量组(1.70 mg·kg-1·d-1含VD315.0μg)、sSAN-VD3高剂量组(4.2 mg·kg-1·d-1含VD337.5μg)、VD3低剂量组(15.0μg·kg-1·d-1)、VD3高剂量组(37.5μg·kg-1·d-1)6组,每组6只.正常对照组大鼠在通光条件下用正常饲料饲养,其余各组大鼠置于暗室中,并喂养缺乏维生素D3的饲料,同时各预防给药组按剂量连续灌胃给药35天.观察记录大鼠的体重变化,检测大鼠血清中25羟基维生素D3(25-(OH)VD3)、血清骨碱性磷酸酶(Bone Alkaline Phosphatase,BAP)活性及股骨骨密度(Bone Mineral Density BMD).结果佝偻病模型组大鼠喂养5周后体重明显低于正常对照组,血清BAP活性明显增高,血清25-(OH)VD3含量及股骨BMD则显著降低(P<0.05);与佝偻病模型组大鼠比较,各预防给药组大鼠体重及股骨BMD均有所增加,sSAN-VD3高、低剂量组和VD3高剂量组大鼠血清BAP活性降低,25-(OH)VD3水平明显升高,与模型组差异显著(P<0.05);与同剂量的VD3低剂量组比较,sSAN-VD3低剂量组大鼠的血清BAP、25-(OH)VD3及股骨BMD等三项指标均显示出更明显的抗佝偻痛作用(P<0.05).结论:负载维生素D3海藻酸纳米粒和维生素D3均具有良好的抗佝偻病作用,以前者作用较强.     

层层自组装纳米粒作为非病毒基因载体

基因治疗的效果严重依赖于基因载体。与传统包封技术相比,在自组装技术基础上发展起来的以DNA为聚阴离子,与荷正电的高分子材料在溶液中形成纳米粒的方法,已成为目前最重要的非病毒基因载体制备手段,具有良好的应用前景。采用层层自组装(layer-by-layer assembly,LbL)技术可提高基因装载率,其优势还在于纳米粒表面性质的可控性:在温和的条件下实现多种材料在载体表面的固定,实现载体多功能化等。本文将对近年来国内外有关层层自组装纳米粒作为非病毒基因载体的研究进展以及本课题组在此方向的研究进行简要综述。     

壳聚糖-有机累托石/海藻酸钠复合纳米粒的制备

目的:以BSA作为模型药物,制备壳聚糖季铵盐-OREC复合物纳米微粒,建立一种安全有效的药物控释传递系统。方法:超声条件下,制备不同质量比的具有壳聚糖硅酸盐插层结构的复合物纳米微粒,观察其形态学特征、进行红外光谱分析。同时,测定OREC对BSA包封率和载药量的影响。结果:成功制备了不同质量比的OREC-HTCC纳米粒子。电镜结果显示纳米粒呈圆球形,均匀,平均粒径约为30nm。红外图谱分析证实,HTCC插入了OREC插层中,BSA成功地包裹入HTCC-ALG/OREC混合材料制备的纳米微粒。加入OREC后,纳米粒子的包封率及载药量均明显提高,但随着加入量的增加,包封率及载药量逐渐减少。结论:OREC-HTCC纳米粒子是良好的蛋白药物载体,具有粒径小、包封率高、缓释效果好等优点,为CS-OREC作为潜在的药物给药系统的进一步应用提供科学依据。     

阿霉素纳米粒在肿瘤治疗中的应用研究

目的:观察阿霉素纳米粒对多耐受糖蛋白介导的膀胱移行细胞癌多耐药的逆转.方法:MTT法观察MDR逆转,采用流式细胞仪对细胞内阿霉素浓度进行检测.结果:阿霉素纳米对敏感株的细胞毒作用与阿霉素差异无显著性意义,耐药株对阿霉素纳米较对阿霉素纳米粒敏感4.8倍,表现细胞内阿霉素显著增高.结论:阿霉素纳米可有效逆转P-gp介导的多药耐药.     

pH敏感纳米粒的制备和功能表征

目的:制备具有pH响应的甲氧基聚乙二醇甲基丙烯酸-2-六亚甲基亚胺乙酯聚合物,测试材料pH功能响应,以及建立聚合物纳米粒载药方法。方法:通过核磁共振氢谱鉴定ATRP(Atom Transfer Radical Polymerization)聚合反应所获得的化合物结构。滴加-搅拌挥发法制备聚乙二醇甲基丙烯酸-2-六亚甲基亚胺乙酯纳米粒,酶标仪测定其载药量和包封率。透射电镜下观察其形态,激光粒度仪分析测定其粒径,包载DiR红外荧光探针检测纳米粒pH响应功能。结果:分别成功合成得到2-溴代异丁酸聚乙二醇单甲醚和甲基丙烯酸-2-六亚甲基亚胺乙酯单体。通过ATRP聚合反应成功合成聚乙二醇甲基丙烯酸-2-六亚甲基亚胺乙酯聚合物材料,并通过核磁氢谱对聚合材料进行鉴定。通过滴加搅拌法制备包载有模型药物香豆素-6的纳米粒,并对纳米粒的形态表征及载药量进行测定。结论:试验结果表明制备得到的聚合物纳米粒尺寸均匀,具有预期的pH响应效果,可以装载模型药物。     

载药纳米粒作为脑靶向给药系统的研究

血脑屏障使大部分的活性药物很难由血液进入脑内发挥作用。载药纳米粒具有脑靶向性,可显著提高药物在脑内浓度,成为药物突破血脑屏障的有效途径。本文综述了近年来载药纳米粒透过血脑屏障的研究进展,并对纳米粒载中药入脑提出展望。     

载药纳米粒作为脑靶向给药系统的研究

血脑屏障使大部分的活性药物很难由血液进入脑内发挥作用。载药纳米粒具有脑靶向性,可显著提高药物在脑内浓度,成为药物突破血脑屏障的有效途径。本文综述了近年来载药纳米粒透过血脑屏障的研究进展,并对纳米粒载中药入脑提出展望。     

转铁蛋白修饰磁性纳米粒的制备及其体外抗肿瘤活性研究

目的:本研究旨在构建一种转铁蛋白修饰负载阿霉素(DOX)的磁纳米粒靶向递药系统,以提高阿霉素作用的靶向性。方法:采用化学共沉淀法制备转铁蛋白修饰负载阿霉素的磁性纳米粒(DOX@MNP),采用zeta电位及纳米粒度分析仪测定DOX@MNP的粒径及其zeta电位,透析法评价DOX@MNP的体外释药特征。通过MTT实验,研究DOX@MNP与游离DOX对A549细胞的细胞毒性,通过激光共聚焦显微镜和流式细胞仪观察A549细胞对DOX@MNP与游离DOX的摄取情况。结果:DOX@MNP的释药具有p H依赖性。MTT实验结果显示,DOX@MNP与游离DOX具有相当的细胞毒性;激光共聚焦显微镜和流式细胞仪检测结果显示A549细胞对DOX和DOX@MNP的摄取没有明显差异。结论:本文构建了一种转铁蛋白修饰包载阿霉素的磁纳米粒,体外结果显示其具有与游离DOX相当的细胞毒性,为进一步进行体内实验奠定了基础。     

5-氨基水杨酸纳米粒对TNBS诱导小鼠结肠炎的治疗作用

目的:评价超临界分散法制备的5-氨基水杨酸(5-ASA)结肠定位纳米粒对小鼠结肠炎的治疗作用。方法:将24只6周龄BABL/c小鼠随机分为空白对照组,结肠炎模型组,5-ASA治疗组,5-ASA纳米粒治疗组。三硝基苯磺酸(TNBS)造模后的第2-7 d分别给予0.9%氯化钠,5-ASA和5-ASA纳米粒灌胃,于第8 d处死所有小鼠,评估小鼠的疾病活动指数(DAI),组织病理形态以及结肠组织学损伤情况;检测结肠组织髓过氧化物酶(MPO)活性。结果:5-ASA和5-ASA纳米粒均能明显减轻结肠重量,缩小溃疡面积,降低MPO活性。超临界分散法制备的5-ASA纳米粒在DAI,减轻局部的炎症程度(MPO水平)方面,其疗效与5-ASA组(50 mg/Kg)相当。结论:超临界分散法制备的5-ASA纳米粒对TNBS诱导小鼠的结肠炎有明显的治疗作用。     

聚乳酸纳米粒穿透血脑屏障的分析电镜研究

观察以聚乳酸 (D ,L-polylacticacid,PLA)为材料制备、经吐温-80(T-80)表面改性的纳米粒对血脑屏障的穿透效果并探讨其机制 ,分别将FITC-Dextran、叶绿素铜作为PLA纳米粒的示踪标记 ,应用荧光显微镜、透射电镜及分析电镜观察经静脉注射入小鼠体内的PLA纳米粒在脑组织中的分布、穿透血脑屏障的特性。荧光显微镜观察到小鼠脑组织中散在及沿毛细血管壁分布的荧光颗粒 ,透射电镜可观察到小鼠脑毛细血管内皮细胞及周围脑组织中圆形或类圆形的外源性纳米粒 ;进一步采用分析电镜对颗粒处组织进行能谱分析证实其为叶绿素铜标记的PLA纳米粒。证实了T-80修饰的PLA纳米粒具有穿透血脑屏障的特性 ,机制可能是毛细血管内皮细胞的胞吞转运作用 ,同时 ,为研究纳米粒在组织内的定位提供了新的标记方法.     

Optimization of covalent immobilization of pectinase on sodium alginate support

Pectinase was immobilized on a sodium alginate support using glutaraldehyde and retained 66% activity. The optimal pH for activity shifted from 3.0 to 3.5 after immobilization; however, the optimum temperature remained unchanged at 40°C. The immobilized enzyme also had a higher thermal stability and reusability than the free enzyme, and retained 80% of initial activity after 11 batch reactions.     

Immobilization of the proteins in the natural rubber with dialdehyde sodium alginate

The biodegradable dialdehyde sodium alginate (DASA) was exploited to immobilize the proteins in the natural rubber latex (NRL) and the variations of the properties for the NRL films were estimated in detail. As demonstrated, the proteins were distributed more uniformly in the NRL films with DASA and the extractable protein (EP) content was effectively decreased. Particularly, the EP content was lowered to a value about 46 μg/g with 0.40% DASA, which could meet with the demands of the allergy protein threshold limit of 50 μg/g as described in ASTM D 5712 standard. Furthermore, there was some improve on the burial degradability of the NRL films modified with DASA. The mechanical properties, however, had no evident variation in the presence of DASA. In conclusion, the immobilization of the proteins with DASA should be a potential alternative to tackle the protein allergy problem for the NRL and its products.     

An amperometric biosensor fabricated from electro-co-deposition of sodium alginate and horseradish peroxidase

A convenient and effective strategy for fabrication of hydrogen peroxide biosensor based on sodium alginate (SA) and polyvinyl butyral (PVB) as matrices was reported in this paper. The horseradish peroxidase (HRP) and SA were electro-co-deposited onto the surface of gold electrode, and the HRP–SA/Au electrode was further coated with PVB. The interaction between HRP and SA was characterized by UV–vis absorption spectroscopy, and the fabricating process of biosensor was characterized by electrochemical impedance spectroscopy (EIS). The electrochemical characteristics of the biosensor were studied by cyclic voltammetry and chronoamperometry. Experimental conditions were investigated which influence the performance of the biosensor, such as pH, and applied potential. The biosensor showed a linear response to H₂O₂ over a concentration range from 7.0 × 10⁻⁶ to 4.1 × 10⁻³ M with a detection limit of 1.8 × 10⁻⁶ M based on a signal-to-noise ratio of 3 under optimum conditions. The value of HRP in the composite was evaluated to be 1.38 mM. The biosensor obtained from this study possesses high sensitivity, good reproducibility, and long-term stability.     

海藻酸钠包埋法制备固定化菠萝蛋白酶

以海藻酸钠为载体,包埋法固定菠萝蛋白酶,对固定化奈件进行优化,同时探讨固定化菠萝蛋白酶的部分酶学性能。结果表明：固定化菠萝蛋白酶的质量受海藻酸钠质量分数、固定化酶量、固定化时间以及CaCl2质量分数的影响,其最佳固定化条件为：海藻酸钠质量分数1．0％,CaCl2质量分数3％,固定化酶液量与海藻酸钠体积之比1：2,固定化时间60min,在此条件下,制备的固定化菠萝蛋白酶的比活力为211．8U／g（湿质量载体）,由此制得的固定化酶的最适pH为7．6,与游离酶相比,升高了0．8个pH单位,同时显示固定化菠萝蛋白酶能耐受较高的碱性环境,固定化酶最适温度与游离酶相同,均为50℃,固定化酶在较高温度范围内,仍能保持较高的相对活力。     

Glutamate decarboxylase-derived IDDM autoantigens displayed on self-assembled protein nanoparticles

The recombinant ferritin heavy chain (FTN-H) formed self-assembled spherical nanoparticles with the size comparable to native one. We tried to express the GAD65 COOH-terminal fragments, i.e., 448-585 (GAD65(448-585)), 487-585 (GAD65(487-585)), and 512-585 (GAD65(512-585)) amino acid fragments, using FTN-H as N-terminus fusion expression partner in Escherichia coli. All of recombinant fusion proteins (FTN-H::GAD65(448-585), FTN-H::GAD65(487-585), and FTN-H::GAD65(512-585)) also formed spherical nanoparticles due probably to the self-assembly function of the fused ferritin heavy chain. The antigenic epitopes within GAD65(448-585), GAD65(487-585), and GAD65(512-585) against insulin-dependent diabetes mellitus (IDDM) marker (autoantibodies against GAD65) were localized at the surface of the spherical protein nanoparticles so that anti-GAD65 Ab could recognize them. Protein nanoparticles like FTN-H seem to provide distinct advantages over other inorganic nanoparticles (e.g., Au, Ag, CdSe, etc.) in that through the bacterial synthesis, the active capture probes can be located at the nanoparticle surface with constant orientation/conformation via covalent cross-linking without complex chemistry. Also it is possible for the protein nanoparticles to have uniform particle size, which is rarely achieved in the chemical synthesis of inorganic nanoparticles. Thus, the recombinant ferritin particles can be used as a three-dimensional (spherical) and nanometer-scale probe structure that is a key component in ultra-sensitive protein chip for detecting protein-small molecule interactions and protein-protein interactions.     

Biodistribution studies of protein cage nanoparticles demonstrate broad tissue distribution and rapid clearance in vivo

Protein cage nanoparticles have the potential to serve as multifunctional cell targeted, imaging and therapeutic platforms for broad applications in medicine. However, before they find applications in medicine, their biocompatibility in vivo needs to be demonstrated. We provide here baseline biodistribution information of two different spherical protein cage nanoplatforms, the 28 nm viral Cowpea chlorotic mottle virus (CCMV) and the 12 nm heat shock protein (Hsp) cage. In naive and immunized mice both nanoplatforms show similar broad distribution and movement throughout most tissues and organs, rapid excretion, the absence of long-term persistence within mice tissue and organs, and no overt toxicity after a single injection. These results suggest that protein cage based nanoparticles may serve as safe, biocompatible, nanoplatforms for applications in medicine.     

Phase diagram of the system sodium alginate/water: a model for biofilms

Sodium alginate is a polyelectrolyte consisting of the monomer units β-d-mannuronate and -l-guluronate. Mainly based on the theory of Khokhlow et al., the state diagram of the binary system alginate/water has been calculated using different sets of parameters like degree of ionization, degree of polymerization and interaction function. The calculations comprise miscibility gaps, liquidus curves, eutectic points and the behaviour at temperatures below the melting point of water. Also gel and swelling curves have been treated, where gels are physically crosslinked. The DSC diagram of a 0.5 by wt.% polymer sol shows a double melting peak, which is explained by a heterogeneity above 0 °C. The crystallization of water seems to concentrate the gelled system irreversibly.     

Disposable biosensor and biocatalysis of horseradish peroxidase based on sodium alginate film and room temperature ionic liquid

Composite material based on room temperature ionic liquid (RTIL) N-butylpyridinium hexafluorophosphate (BPPF₆), sodium alginate (SA), and graphite was used to construct a novel horseradish peroxidase (HRP) biosensor for the determination of H₂O₂. The morphology of the as-prepared biosensor was characterized by scanning electron microscopy (SEM). Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to characterize the process of the performance of the biosensor. Parameters affecting the performance of the biosensor, including the concentrations of o-aminophenol (OAP) and HRP and the pH value of substrate solution, were optimized. Under the optimized experimental conditions, H₂O₂ could be detected in a linear calibration range of 1.0 to 6.0 μM with a correlation coefficient of 0.9847 (n = 7) and a detection limit of 0.5 μM at a signal/noise ratio of 3. The prepared biosensor not only had economic and disposable property but also showed good detection precision, bioactivity, storage stability, and reproducibility.     

Microwave assisted rapid method for hydrolysis of sodium alginate for M/G ratio determination

A rapid one-pot method for hydrolysis of sodium alginate for determining M/G ratio has been developed under mild conditions, using microwave irradiation. Poly-mannuronic acid (PMA) and poly-guluronic acid (PGA) ratio was determined (M/G 0.38), which was similar to that (M/G 0.39) obtained by the conventional method, using sodium alginate of Sigma as reference sample for bench marking. For validation of the method PMA and PGA were characterized by density, porosity, viscosity, optical rotation measurements, ¹³C NMR, FT-IR, thermogravimetric analysis (TGA), X-ray diffraction, circular dichroism (CD), molar mass distribution (GPC) and scanning electron microscopy (SEM).