胶原-磺化羧甲基壳聚糖/硅橡胶皮肤再生材料的制备及其对小型猪烫伤创面全层皮肤缺损的修复研究

制备了一种胶原-磺化羧甲基壳聚糖/硅橡胶皮肤再生材料,并以小型猪为模型,考察了其对烫伤全层皮肤缺损的修复性能.首先合成了磺化羧甲基壳聚糖,并对其结构进行了表征.制备了胶原-磺化羧甲基壳聚糖多孔支架,采用扫描电子显微镜(SEM)研究了磺化羧甲基壳聚糖含量对支架微结构的影响.随着磺化羧甲基壳聚糖含量的增大,胶原-磺化羧甲基壳聚糖支架从纤维结构向片状结构转化,且支架的孔径相对变大.采用体外成纤维细胞培养实验证明胶原-磺化羧甲基壳聚糖支架无明显细胞毒性.进一步将胶原-磺化羧甲基壳聚糖支架与硅橡胶膜复合,构建具有双层结构的皮肤再生材料.以小型猪为模型,评价了其对深度烫伤创面的修复性能.大体观察和组织学分析结果显示,胶原-磺化羧甲基壳聚糖/硅橡胶皮肤再生材料具有更快的血管化性能,且经该材料处理的创面能有效支持薄自体皮片的移植成活,实现深度烫伤创面的全层修复.     

复合转化生长因子-β1的磺化壳聚糖/聚赖氨酸纳米粒子的制备及其体外诱导干细胞分化性能

采用共沉淀法制备了复合转化生长因子-β1(TGF-β1)的磺化壳聚糖/聚赖氨酸纳米粒子(SCS/PLL/TGF-β1),研究了磺化壳聚糖的浓度对纳米粒子粒径和表面电位的影响.所得纳米微粒的粒径在240~290nm、表面电位在-0.4~-1.3 mV之间可调.采用透射电子显微镜表征了磺化壳聚糖/聚赖氨酸(SCS/PLL)和SCS/PLL/TGF-β1纳米粒子的形貌.通过体外骨髓间充质干细胞(BMSCs)培养实验,分别考察了SCS/PLL/TGF-β1纳米粒子对BMSCs的增殖、氨基葡聚糖(GAGs)分泌和Ⅱ型胶原基因表达的影响.结果显示,相对于等量的SCS/PLL或同浓度的自由TGF-β1,SCS/PLL/TGF-β1纳米粒子可有效保护TGF-β1的活性,促进BMSCs向软骨细胞分化,可望应用于软骨修复材料等领域的研究.     

磺化低聚壳聚糖的制备及其阻垢性能的研究

低聚壳聚糖和2,3-环氧丙磺酸钠开环接枝制得磺化低聚壳聚糖(SCS),其磺化度为0.77,用红外(FT-IR)光谱、核磁共振(1H NMR)氢谱进行了SCS产物结构表征。利用静态阻垢法对SCS阻硫酸钙和阻磷酸钙性能进行评价,当[Ca2+]为1900 mg/L,[SO42-]为4560 mg/L,SCS用量为32mg/L时,对硫酸钙的阻垢率能够达到88%;当[Ca2+]为100mg/L,[PO43-]为5 mg/L,SCS用量为16mg/L时,对磷酸钙的阻垢率可达到84%。研究表明磺化低聚壳聚糖是一种性能优异的绿色阻垢剂。     

壳聚糖固定化亚铁Schiff碱配合物的研究

利用具有多种生物医用活性的高分子壳聚糖的轴向配位作用合成了壳聚糖-N-亚水杨基氨基酸Schiff碱亚铁配合物,采用IR光谱、元素分析、荧光光谱、紫外-可见吸收光谱和热分析等分析手段对配体及高分子配合物进行了表征,推测了其结构.以核黄素光化学氧化-羟胺-偶氮比色法研究了该化合物对超氧离子自由基的清除能力.结果表明,壳聚糖固定化的N-亚水杨基氨基酸Schiff碱亚铁配合物对超氧离子自由基的清除能力有所降低,但仍具有较强的抑制作用.     

单模微波辐照下壳聚糖-左旋聚乳酸接枝共聚物的酶催化合成

以猪胰脂肪酶(PPL)代替传统的有机金属作为催化剂,在单模微波辐照下利用左旋丙交酯(L-LA)的开环聚合制备壳聚糖-左旋聚乳酸(CS-g-PLLA)接枝共聚物.考察了反应温度及酶用量对接枝率的影响.以此为基础,利用DTG、XRD和3T3成纤维细胞培养对产物的物理性能及细胞相容性进行分析.结果显示,猪胰酶可有效催化接枝反应的进行.反应温度和酶用量对产物的接枝率有较大影响.在单模微波作用下,较低的反应温度(50℃)可获得具有较高接枝率(178.8%)的接枝产物.与纯壳聚糖相比,接枝产物的结晶度和热稳定性降低,说明PLLA的引入破坏了壳聚糖的高结晶性.产物具有良好的细胞相容性,可作为优良的组织工程支架材料.     

高纯度(98.5%)夫沙那韦-d_4的合成

以硝基苯-d5为原料,经还原、保护、磺酰化、脱保护、氧化和亲核取代反应制得中间体——对硝基苯磺酰氯-d4(7)。以(2R,3S)-N-叔丁氧羰基-3-氨基-1,2-环氧-4-苯基丁烷为原料,经开环反应制得(1S,2R)-N-[1-苯甲基-2-羟基-3-(异丁胺基)丙基]氨甲酸叔丁酯(9);9与7经磺化反应合成(2R,3S)-N-[(3-氨基-2-羟基-4-苯基)丁基]-N-异丁基-4-硝基苯磺酰胺盐酸盐-d4(10);10依次经酰化、磷酰化、加氢还原和醋酸钙成盐反应合成了夫沙那韦-d4,总收率33%,纯度98.5%,其结构经1H NMR和EI-MS确证。     

O-羧甲基壳聚糖稀土配合物的抑菌活性及与DNA的相互作用

用体外抑菌法研究了O-羧甲基壳聚糖镧、O-羧甲基壳聚糖钕配合物对大肠杆菌(E.coli)、金黄色葡萄球菌(St.aureus)、阴沟肠杆菌(E.cloacae)、枯草芽孢杆菌(B.subtilis)、粪肠杆菌(E.faecalis)和肺炎克罗伯杆菌(S.pneumoniae)的抑菌活性。采用紫外光谱研究了两种O-羧甲基壳聚糖稀土配合物与小牛胸腺DNA的相互作用。结果表明:O-羧甲基壳聚糖稀土配合物均有抑菌活性,且O-羧甲基壳聚糖稀土配合物与DNA以静电作用为主,能使DNA双螺旋结构破坏。     

新型5-芳基-N-吡唑肟酯化合物的合成与杀菌活性

以水杨醛和丙酮缩合形成α,β不饱和酮,与水合肼和冰乙酸关环形成5-(2-羟基苯基)吡唑酮骨架,合成了10种未见报道的1-(5-(2-(苯氧基)苯基)-3-甲基)-4,5-二氢-N-吡唑肟酯类衍生物.通过元素分析、红外光谱及核磁共振氢谱测试技术对其结构进行了表征,初步测定了化合物的杀菌活性.结果表明,化合物对小麦赤霉病菌有较好的杀灭活性(其中化合物5f杀灭活性达到81.3%);化合物5g和5i对水稻纹枯病菌有中等杀灭活性,对稻瘟病菌的杀灭活性差.成了10种未见报道的1-(5-(2-(苯氧基)苯基)-3-甲基)-4,5-二氢-N-吡唑肟酯类衍生物.通过元素分析、红外光谱及核磁共振氢谱测试技术对其结构进行了表征,初步测定了化合物的杀菌活性.结果表明,化合物对小麦赤霉病菌有较好的杀灭活性(其中化合物5f杀灭活性达到81.3%);化合物5g和5i对水稻纹枯病菌有中等杀灭活性,对稻瘟病菌的杀灭活性差.     

N-(4-哌啶基)苯甲酰胺衍生物的设计、合成及其抗肝癌活性

以4-羟基苯甲酸乙酯为原料,经烃化、水解、缩合、脱保护、磺酰化反应,合成了6个含有磺酰基取代的4-苯氧基-N-(4-哌啶基)苯甲酰胺衍生物。 其结构经¹H NMR、¹³C NMR和MS谱等技术手段进行了表征,并以索拉非尼为阳性对照药对HepG2细胞株进行了初步体外抗肝癌细胞增殖活性的评价。 实验发现,4-苯氧基-N-(1-甲磺酰基哌啶-4-基)苯甲酰胺的体外抗肝癌生物活性优于索拉非尼,IC₅₀值为8.42 μmol/L。 研究结果表明,新结构4-苯氧基-N-(4-哌啶基)苯甲酰胺类化合物具有良好的抗肝癌活性。     

磺甲基化木质素的辣根过氧化物酶催化聚合研究

以碱木质素为原料,采用磺甲基化反应,得到磺甲基化木质素(SAL),并进一步采用辣根过氧化物酶(HRP)催化以提高其分子量,制备了高分子量高磺化度磺甲基化木质素(HPSAL).采用凝胶渗透色谱、红外光谱、核磁共振氢谱、紫外光谱和顶空气相色谱等研究了改性前后SAL的结构特征.结果表明,经HRP催化后,与SAL相比,HPSAL的重均分子量和磺化度显著增加,分别提高20倍和30%以上,羧基含量升高,而甲氧基和酚羟基含量降低.HRP使SAL形成酚氧自由基,活化其酚羟基的邻、对和侧链Cβ位,增加磺化反应活性,而磺化度的提高又有利于增加HRP催化SAL的聚合反应活性,其聚合方式主要为β-O-4'、β-β'、β-1'和β-5'联结.分子模拟结果表明,甲氧基含量的降低和磺酸基含量的增加能显著提高以β-O-4'连接键为主的聚合反应活性.     

Action of chitosan against Xanthomonas pathogenic bacteria isolated from Euphorbia pulcherrima

The antibacterial activity and mechanism of two kinds of chitosan were investigated against twelve Xanthomonas strains recovered from Euphorbia pulcherrima. Results indicated that both chitosans markedly inhibited bacterial growth based on OD loss. Furthermore, the release of DNA and RNA from three selected strains was increased by both chitosans. However, the release of intracellular proteins was inhibited by both chitosans at different concentration and incubation times, except chitosan A at 0.1 mg/mL for 0.5 h incubation and 0.2 mg/mL for 2.0 h incubation increased the release of proteins, indicating the complexity of the interaction and cell membranes, which was affected by incubation time, bacterial species, chitosan type and concentration. Transmission electron microscopic observations revealed that chitosan caused changes in protoplast concentration and surface morphology. In some cells, the membranes and walls were badly distorted and disrupted, while other cells were enveloped by a thick and compact ribbon-like layer. The contrary influence on cell morphology may explain the differential effect in the release of material. In addition, scanning electron microscope and biofilm formation test revealed that both chitosans removed biofilm biomass. Overall, this study showed that membrane and biofilm play an important role in the antibacterial mechanism of chitosan.     

壳多糖抑制细菌生长的构效关系

运用化学结构已清楚, 分属4大系列的29种壳多糖, 以4种不同类型的细菌(革兰氏阳性菌Ecoli K1、革兰氏阴性菌Bacillus cereus、Bacillus megaterium和Staphlylococcu aureus)为研究对象, 进行了壳多糖抑菌能力构效关系的研究. 在实验中采用96孔平板, 用计算机\|吸光值读数仪直接测定每个孔的吸光值, 获得了各个细菌在不同壳多糖浓度中的生长曲线和壳多糖抑制细菌生长的最低抑制浓度(MIC, Minimum inhibit concentration). 通过比较同一(各个)系列的壳多糖在这些相同(不同)细菌的MIC变化规律与壳多糖的化学结构的关系, 发现同一壳多糖对不同的细菌的MIC值是不相同的, 因而壳多糖抑制细菌生长的能力首先与细菌本身特点有关, 但与是否为革兰氏阳性菌或阴性菌无直接的相关性; 同一细菌对不同化学结构的壳多糖有一定的相关性, 在壳多糖的聚合程度(DP)相同的条件下, 壳多糖中氨基被乙酰化(DA)的程度越低, 壳多糖抑制细菌生长的MIC值越低, 壳多糖抑制细菌生长的能力就越强; 同样,在DA相同的情况下, 分子越小, 壳多糖抑制细菌生长的MIC值越低, 抑制细菌生长的能力越强. 根据上述实验结果, 初步推测壳多糖抑制细菌生长的机制可能与其在溶液中所带的正电荷多少有关.     

Biomacromolecular affinity: Interactions between lysozyme and regioselectively sulfated chitosan

It is well known that the sulfate groups on different positions in polysaccharides play important roles in protein adsorption. However, the interactions between sulfated chitosans and lysozyme have not been clearly elucidated. In this study, the regioselectively sulfated chitosans, 6-O-sulfated chitosan (C6S), 2-N-6-O-sulfated chitosan (C26S) and 3,6-O-sulfated chitosan (C36S), were chosen to investigate the possible mechanisms determining the interaction between lysozyme and the sulfated chitosans. It has been found that the selectively sulfated products of chitosan (CS), C6S, C26S and C36S all exhibit lysozyme binding activity. However, the maximum binding ratios of lysozyme/polysaccharide are significantly different for C6S, C26S and C36S. In addition, though C6S possesses the lowest sulfur content among the three sulfated chitosans, it exhibits the highest binding activity with lysozyme. Furthermore, in the protein mixtures, C6S shows the highest selective binding activity with lysozyme among the three sulfated chitosans in the presence of γ-globulin and bovine serum albumin (BSA). The results indicate that 6-O-sulfate groups may be responsible for the high affinity and specific interaction of sulfated chitosan with lysozyme, while 2-O-sulfate and 3-O-sulfate groups are unfavorable to this interaction.     

Surface modification of poly(ethylene terephthalate) via hydrolysis and layer-by-layer assembly of chitosan and chondroitin sulfate to construct cytocompatible layer for human endothelial cells

Surface modification of poly(ethylene terephthalate) (PET) film was performed by surface hydrolysis and layer-by-layer (LBL) assembly followed a mechanism of electrostatic adsorption of oppositely charged polymers, exemplified with chitosan and chondroitin sulfate (CS). Hydrolysis of PET in concentrated alkaline solution produced a carboxyl-enriched surface. The changes of weight loss and surface chemistry, morphology and wettability were monitored and verified by UV-vis spectroscopy, atomic force microscopy (AFM) and water contact angle. Assembly of positively charged chitosan and negatively charged CS was then conducted in a LBL manner to create multilayers on the hydrolyzed PET film. The process of layer growth and oscillation of surface wettability were monitored by UV-vis spectroscopy and water contact angle measurement, respectively. In vitro cell culture revealed that the adherence of endothelial cells was significantly enhanced on the biomacromolecules-modified PET film with preserved endothelial cell function, in particular on those assembled with larger number of chitosan/CS layers. However, with regard to cell proliferation and viability properties after cultured for 4 days, minor difference was determined between the modified and the unmodified PET films.     

Preparation and characterizations of asymmetric sulfonated polysulfone membranes by wet phase inversion method

This paper presents an original approach to prepare the asymmetric sulfonated polysulfone membranes by using wet phase inversion method and their applications for dehydrating a water/ethanol mixture by pervaporation. The separation performances of sulfonated membranes were strongly affected by the degree of sulfonation and the degree of swelling of membranes. The substitution degree of sulfonic group enhanced the permselectivity of sulfonated polysulfone membranes by increasing the hydrophilicity of polymer backbone. Based on the observations of membrane morphology and light transmittance measurements, the degree of sulfonation of polysulfone presented less influence on the membrane formation pathway and the final structure of membrane in wet phase inversion process. It was also found that the sulfonated membranes showed well hydrophilic properties and facilitated water adsorption in the membranes. The sorption and permeation properties also showed that the permselectivity of asymmetric membrane was dominated by the permeate diffusion rather than the permeate sorption in the skin layer. The high separation performance of pervaporation membrane can be achieved by phase inverse method with sulfonated polysulfone.     

Morphological studies and ionic transport properties of partially sulfonated diblock copolymers

The morphological behavior of partially sulfonated polystyrene-block-poly(ethylene-alt-propylene) (PS-PEP) membranes cast from tetrahydrofuran (THF) solutions were investigated by small-angle X-ray scattering (SAXS) and atomic force microscopy (AFM).The uptakes of methanol and water increase as the sulfonation degree increases, the methanol uptake being overwhelmingly greater than the water uptake. The conductivity increases almost exponentially with increasing sulfonation degree of polystyrene units. Clusters of sulfonated units that are formed in the solution used for casting membranes persist in the solid state after evaporation. In contact with water, swelling of the membranes proceeds predominantly in these clusters. The original lamellar morphology of the diblock copolymer is progressively deformed with increasing degree of sulfonation by the presence of the clusters containing ion-rich sequences of sulfonated polystyrene blocks.     

Uniform chitosan hollow microspheres prepared with the sulfonated polystyrene particles templates

Biodegradable chitosan hollow microspheres have been fabricated by employing uniform sulfonated polystyrene (PS) particles as templates. The chitosan was adsorbed onto the surface of the sulfonated polystyrene templates through the electrostatic interaction between the sulfonic acid groups on the templates and the amino groups on the chitosan. Subsequently, the adsorbed chitosan was crosslinked by adding glutaraldehyde. After the removal of the sulfonated polystyrene core, chitosan hollow microspheres were obtained. The longer the sulfonation time used, the smaller the size of the hollow particles and the thicker the chitosan wall obtained. Fourier transform infrared spectrometry was used to characterize the component of the microspheres. The morphologies of the PS templates and the chitosan microspheres were observed by transmission electron microscopy and scanning electron microscopy. The controlled release behavior of the chitosan hollow microspheres was also primarily investigated.     

邻苯二甲酰化壳聚糖中酰胺酸取代度的红外测定

室温条件下从完全脱乙酰化壳聚糖出发合成了不同酰胺酸取代度的邻苯二甲酰化壳聚糖，并以此为标样，标样的取代度由X射线光电子能谱（XPS）确定，研究了以FT－IR作为工具测定此系列衍生物的总取代度的方法，以2887cm^-1的吸收峰作为参比谱带，探针谱带可用1712cm^-1或749cm^-1的吸收峰，两种探针谱带所得曲线的斜率分别为1．13和0．12，相关系数分别为0．997和0．977，此结果表明：红外法是一种既准确可靠又方便可行的测定方法。     

Improved cellular response on multiwalled carbon nanotube-incorporated electrospun polyvinyl alcohol/chitosan nanofibrous scaffolds

We report the fabrication of multiwalled carbon nanotube (MWCNT)-incorporated electrospun polyvinyl alcohol (PVA)/chitosan (CS) nanofibers with improved cellular response for potential tissue engineering applications. In this study, smooth and uniform PVA/CS and PVA/CS/MWCNTs nanofibers with water stability were formed by electrospinning, followed by crosslinking with glutaraldehyde vapor. The morphology, structure, and mechanical properties of the formed electrospun fibrous mats were characterized using scanning electron microscopy, Fourier transform infrared spectroscopy, and mechanical testing, respectively. We showed that the incorporation of MWCNTs did not appreciably affect the morphology of the PVA/CS nanofibers; importantly the protein adsorption ability of the nanofibers was significantly improved. In vitro cell culture of mouse fibroblasts (L929) seeded onto the electrospun scaffolds showed that the incorporation of MWCNTs into the PVA/CS nanofibers significantly promoted cell proliferation. Results from this study hence suggest that MWCNT-incorporated PVA/CS nanofibrous scaffolds with small diameters (around 160 nm) and high porosity can mimic the natural extracellular matrix well, and potentially provide many possibilities for applications in the fields of tissue engineering and regenerative medicine.     

Preparation and Drug Release of PVA Composite Nanofibers Loaded Chitosan Microsphere

In this paper, we reported the fabrication of poly(vinyl alcohol)-chitosan (PVA-CS) microspheres composite nanofibers by electrospinning technique. The chitosan microspheres were firstly prepared by electrospray with the solution of chitosan and combretastatin A4. The morphology and size distribution of chitosan microspheres were analyzed by scanning electron microscopy. The influencing factors including the concentrations of both PVA and CS microspheres were studied. The physical properties of the composite nanofibers were characterized by X-ray diffraction (XRD). The drug release rate, MTT toxicity test, and cell culture were also investigated in detail. Results indicate that the chitosan microsphere-loaded composite nanofibers can be prepared when the PVA concentration is 120 mg/mL. The continuity of the nanofibers was influenced by the concentration of CS microspheres. The characteristic peaks of CS or PVA were not observed in the diffractograms after the CS and PVA were processed using the high-voltage electrostatic technique. In addition, the drug release rate showed that nanofibers induce an obvious slow-release effect. Composite nanofibers were non-toxic to fibroblasts cells, and the fibroblasts cells could proliferate on the nanofiber mat.