高压微胶囊成型装置制备用于成囊的海藻酸钙胶球

利用高压微胶囊成型装置进行了制备用于成囊的海藻酸钙胶珠的试验研究，从而为用于药物控制释放的聚精氨酸基微胶囊的制备提供基础制备参数。将海藻酸钠溶液经试验装置滴入氯化钙溶液中固定化成海藻酸钙凝胶珠，考察了海藻酸钠浓度对制得的胶珠性能的影响及推进速度、针头内径、电压等对制得的胶珠直径的影响。得到较佳的海藻酸钠浓度为1.8%(W/V),发现推进速度提高，胶珠直径增大：针头内径越大，胶珠直径越大；电压增大，胶珠的直径先减少后增大；不同针头内径制备出胶珠的平均直径最小时所对应的电压也随针头内径的增大而增大。     

乳化-凝胶化法制备药用载体海藻酸钙微球的研究

采用乳化-凝胶化法制备了海藻酸钙微球,考察了海藻酸钠浓度、氯化钙浓度、乳化速度、油水相比例、乳化剂浓度对制备海藻酸钙微球粒径、形态的影响;同时以牛血红蛋白为模型药物初步考察了海藻酸钙微球载药性能。通过对制备条件的优化,可制备粒径为1μm左右,分布相对均匀,球形度较好的海藻酸钙微球;微球的载药量不高,提高模型药物的初始浓度,微球载药量有一定程度增大,但当牛血红蛋白浓度高于10mg/mL时,随模型药物初始浓度增大,载药量增大幅度减小。     

海藻酸钠-壳聚糖-海藻酸钠(ACA)微胶囊的蛋白质通透性研究

本研究用激光共聚焦扫描显微镜CLSM观察荧光标记蛋白在微胶珠/囊上的吸附和渗透现象.考察了微胶囊类型、海藻酸钠粘度和壳聚糖分子量对微胶珠/囊通透性的影响.结果表明,蛋白质在膜上是先吸附后渗透的过程;AC膜具有不对称性;不同类型微胶珠/囊的通透性依次为海藻酸钠（A）微胶珠＞海藻酸钠-壳聚糖（AC）微胶囊＞海藻酸钠-壳聚糖-海藻酸钠（ACA）微胶囊;海藻酸钠粘度越高,壳聚糖分子量越大,微胶珠/囊的通透性越低.为制备低通透率即具有较好酶屏障功能的ACA蛋白质口服控释载体提供了直观的实验依据.     

海藻酸钙微胶珠培养神经干细胞研究

神经干细胞作为一种具备自我更新及多向分化潜能的细胞,已经引起国内外研究者的高度关注。神经干细胞的发现、研究与应用,将在治疗神经系统疾病以及神经损伤的修复中起到重要作用,但是神经干细胞的来源受限及数量不足是临床应用的最大障碍。采用三维动态培养从而实现神经干细胞大规模扩增是解决其数量不足的有效方法。但是,在三维动态搅拌条件下,剪切力对细胞的损害是非常大的。考虑到海藻酸钙微胶珠在细胞培养领域的应用,如果把神经干细胞包囊到微胶珠中进行动态培养,将会避免剪切力对干细胞的损伤,因此是一个非常值得期待的干细胞培养方式。本研究在静态条件下,采用海藻酸钙微胶珠作为神经干细胞的3D培养手段,对成囊工艺进行了探索,以确定适于培养神经干细胞的最佳胶珠制备工艺。通过微胶珠内部的扩散传质模型,研究了微胶珠制备工艺中各参数对其扩散系数(D)的影响,找出微胶珠最佳制备工艺参数为:直径2mm,1.5%(wt%)海藻酸钠滴入3.5%(wt%)CaCl2中,反应时间10min。确定上述工艺后,对小鼠神经干细胞进行了不同包囊密度条件下的培养,得到包囊神经干细胞的最佳密度为0.08×106cells.mL-1。此外,本研究还对海藻酸钙微胶珠培养的神经干细胞进行了nestin等特异性蛋白的免疫荧光化学染色,结果为阳性,表明所培养的神经干细胞能够保持其干细胞的特性。本研究结果表明,采用优化参数制备的海藻酸钙微胶珠是培养神经干细胞的有效手段。同时也为动态微囊培养神经干细胞打下了良好基础。     

APA微胶囊免疫隔离生物膜制备中的质量控制

为了优化制备条件, 简化工艺, 控制质量, 本文观测了APA(海藻酸盐-聚赖氨酸-海藻酸盐)微胶囊免疫隔离生物膜制备过程中部分物理因素和化学因素与质量的关系.结果表明: 微囊的粒径与静电微囊发生仪脉冲频率呈负相关; 与推进泵速度呈正相关; 与针头内径呈正相关.随着聚赖氨酸浓度的增加, 膜厚度显著增加.微囊的粒径不随聚赖氨酸成膜反应时间延长而改变, 但影响膜的厚度.柠檬酸钠的液化时间对微囊的粒径与厚度均无明显影响.通过优化制备条件, APA微胶囊免疫隔离生物膜将具有更好的通透性、柔韧性、生物相容性和强度.     

载间充质干细胞海藻酸钠-壳聚糖微胶囊的制备及工艺优化

目的：以壳聚糖为囊材来制备间充质干细胞（MSCs）海藻酸钠．壳聚糖体系微胶囊。方法：通过研究羧甲基壳聚糖、低聚壳聚糖等不同种类壳聚糖的成囊性，以及壳聚糖溶液浓度对间充质干细胞活性的影响，来优化微胶囊制备工艺。结果：羧甲基壳聚糖不能成膜；低聚壳聚糖成囊性差；高相对分子质量（尬）壳聚糖成囊性良好，但微囊粒径大且易破碎；Mr为100000～250000的壳聚糖具有良好的成囊性。结论：可用Mr100000～250000的壳聚糖来制备载MSCs海藻酸钠．壳聚糖微胶囊。同时应控制壳聚糖溶液覆膜时间〈10min，以及1g／L壳聚糖溶液来降低其对间充质干细胞活力的影响。     

海藻酸钠-壳聚糖微胶囊载体在组织工程研究中的应用

背景：用壳聚糖包裹海藻酸钠制备微囊,可以改善海藻酸钠水凝胶的力学性能,如何获得理想的海藻酸钠壳聚糖微囊以及该微囊体系的应用前景是这一研究的关键。 目的：探讨海藻酸钠壳聚糖微胶囊载体的制备方法、成型机制,分析影响微胶囊膜强度性能的几个重要因素及探讨海藻酸钠-壳聚糖微胶囊在固定化细胞技术、药物载体和组织工程方面的应用前景。 方法：由第一作者采用计算机检索PubMed数据库、Elsevier ScienceDirect、中国知网库、万方数据库1987至2013年有关海藻酸钠壳聚糖微囊制备方法、反应机制及应用前景的文章。 结果与结论：海藻酸盐水凝胶在药物释放和组织工程领域具有很多优势,但是凝胶溶蚀现象和力学性能缺陷限制了它的应用,壳聚糖与海藻酸钠通过静电相互作用形成聚电解质络合物,弥补了海藻酸钠凝胶的不足。通过控制壳聚糖溶液的性质-壳聚糖的分子质量、壳聚糖溶液的pH值和浓度制备膜强度高的微囊,海藻酸钠-壳聚糖微囊在固定化技术、药物释放和组织工程领域表现出了广阔的应用前景。中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程全文链接：     

胰岛移植用免疫隔离微胶囊的牢固度、生物相容性和通透性研究

目的为了探讨用于胰岛移植的海藻酸钠-聚赖氨酸-海藻酸钠（APA）生物膜的免疫隔离效果。方法采用高压静电成囊装置,制备APA微胶囊和微囊化胰岛,微囊直径为0.25～0.55mm;取空微囊,利用恒温振荡仪振荡后测定其破损率;将空微囊移植至小鼠腹腔,分别于不同时间由腹腔中灌洗出微胶囊,记数并观察其形态;取一定量空微囊分别与IgG、BSA和胰蛋白酶孵育,测量其浓度变化确定APA微胶囊的通透性。结果采用高压静电成囊技术制成的APA微胶囊呈球形,大小均匀、表面光滑,具有较好的生物相容性;粒径为0.25～0.35mm的微胶囊其牢固度大于粒径为0.45～0.55mm的微胶囊;葡萄糖、胰岛素等小分子物质能够自由通过微囊膜,胰蛋白酶也可通过,但速度较慢;大分子物质牛血清白蛋白和免疫球蛋白则不能透入APA微囊。结论采用高压静电成囊技术可制备高质量粒径为0.25～0.35mm的微胶囊;这是具有良好免疫隔离性能的APA微胶囊。     

海藻酸钙/壳聚糖磁性微胶囊的制备及研究

采用高压静电法制备磁性海藻酸钙/壳聚糖微胶囊,Minitab全因子实验考察不同制备条件对微胶囊形貌、粒径分布及破损率的影响.以超氧化物歧化酶(SOD)为模型药物,考察磁性微胶囊的包封率、载药量及体外释放性能,并初步研究磁性微胶囊的靶向性.     

海藻酸钙/壳聚糖磁性微胶囊的制备及研究

采用高压静电法制备磁性海藻酸钙/壳聚糖微胶囊,Minitab全因子实验考察不同制备条件对微胶囊形貌、粒径分布及破损率的影响。以超氧化物歧化酶（SOD）为模型药物,考察磁性微胶囊的包封率、载药量及体外释放性能,并初步研究磁性微胶囊的靶向性。     

Influence of process conditions during impulsed electrostatic droplet formation on size distribution of hydrogel beads

In the medical applications of microencapsulation of living cells there are strict requirements concerning the high size uniformity and the optimal diameter, the latter dependent on the kind of therapeutic application, of manufactured gel beads. The possibility of manufacturing small size gel bead samples (diameter 300 microm and below) with a low size dispersion (less than 10%), using an impulsed voltage droplet generator, was examined in this work. The main topic was the investigation of the influence of values of electric parameters (voltage U, impulse time tau and impulse frequency f) on the quality of obtained droplets. It was concluded that, owing to the implementation of the impulse mode and regulation of tau and f values, it is possible to work in a controlled manner in the jet flow regime (U> critical voltage UC). It is also possible to obtain uniform bead samples with the average diameter, deff, significantly lower than the nozzle inner diameter dI (bead diameters 0.12-0.25 mm by dI equal to 0.3 mm, size dispersion 5-7%). Alterations of the physical parameters of the process (polymer solution physico-chemical properties, flow rate, distance between nozzle and gellifying bath) enable one to manufacture uniform gel beads in the wide range of diameters using a single nozzle.     

Microcapsules of alginate-chitosan. II. A study of capsule stability and permeability.

The stability of alginate-chitosan capsules was shown to depend strongly on the amount of chitosan bound to the capsules. When the capsules were made by dropping a solution of sodium alginate into a chitosan solution (one-stage procedure), all the chitosan was located in a thin alginate-chitosan membrane on the surface. These capsules were much weaker than the capsules made by reacting calcium alginate beads in an aqueous solution of chitosan and calcium chloride (two-stage procedure). Capsules with high mechanical strength were obtained after shorter reaction times when the number-average molecular weight of the chitosan was reduced to around 15,000, when the capsules were made more homogeneous and when the capsule diameter was reduced to around 300 microm. When these capsules were treated with calcium sequestrant such as citrate under conditions where calcium alginate gels normally dissolve, they still had a gel core indicating the presence of chitosan throughout the capsule matrix. The permeability of the two-stage capsules was reduced when the chitosan molecular weight was increased and the degree of acetylation was increased, and when the capsules were made more inhomogeneous. The addition of another several layers of alginate and chitosan resulted in capsules virtually impermeable to IgG, suggesting an average capsule pore diameter less than 90 A.     

Differential expression of proteoglycan epitopes by ovine intervertebral disc cells

The alginate bead culture system has been utilised by several groups to examine the in vitro proteoglycan (PG) metabolism of chondrocytes and intervertebral disc cells, but the nature of the PGs produced has not been examined in detail. This is largely due to the difficulty of separating the anionically charged sodium alginate support matrix from PGs which are similarly charged. In the present study ovine annulus fibrosus, transitional zone and nucleus pulposus cells were dissociated enzymatically from their respective matrices by sequential digestion with pronase/clostridial collagenase and DNAase and then cultured in alginate beads for 10 d. The beads were solubilised and subjected to DEAE Sepharose CL6B anion exchange chromatography to separate the sodium alginate bead support matrix material quantitatively from the disc cell PGs. The alginate free bead PGs were then subjected to composite agarose polyacrylamide gel electrophoresis to resolve PG populations and the PGs were transferred to nitrocellulose membranes by semidry electroblotting. The PGs were identified by probing the blots with a panel of antibodies to defined PG core protein and glycosaminoglycan side chain epitopes. Alginate beads of disc cells were also embedded in paraffin wax and 4μm sections cut to immunolocalise decorin, biglycan, versican, and the 7-D-4 PG epitope within the beads. Decorin and biglycan had similar distributions in the beads, being localised on the cell surface whereas versican and the 7-D-4 PG epitope were immunolocalised interterritoriarly. This study is the first to demonstrate that ovine disc cells synthesise versican in alginate bead culture. Furthermore the immunoblotting studies also showed that a proportion of the 7-D-4 PG epitope was colocalised with versican.     

Factors influencing alginate gel biocompatibility

Alginate remains the most popular polymer used for cell encapsulation, yet its biocompatibility is inconsistent. Two commercially available alginates were compared, one with 71% guluronate (HiG), and the other with 44% (IntG). Both alginates were purified, and their purities were verified. After 2 days in the peritoneal cavity of C57BL/6J mice, barium (Ba)-gel and calcium (Ca)-gel beads of IntG alginate were clean, while host cells were adhered to beads of HiG alginate. IntG gel beads, however, showed fragmentation in vivo while HiG gel beads stayed firm. The physicochemical properties of the sodium alginates and their gels were thoroughly characterized. The intrinsic viscosity of IntG alginate was 2.5-fold higher than that of HiG alginate, suggesting a greater molecular mass. X-ray photoelectron spectroscopy indicated that both alginates were similar in elemental composition, including low levels of counterions in all gels. The wettabilities of the alginates and gels were also identical, as measured by contact angles of water on dry films. Ba-gel beads of HiG alginate resisted swelling and degradation when immersed in water, much more than the other gel beads. These results suggest that the main factors contributing to the biocompatibility of gels of purified alginate are the mannuronate/guluronate content and/or intrinsic viscosity.     

Ionic liquid as a potential solvent for preparation of collagen-alginate-hydroxyapatite beads as bone filler

In this study, collagen/alginate/hydroxyapatite beads having different proportions were prepared as bone fillers for the restoration of osteological defects. Ionic liquid was used to dissolve the collagen and subsequently the solution was mixed with sodium alginate solution. Hydroxyapatite was added in different proportions, with the rationale to enhance mechanical as well as biological properties. The prepared solutions were given characteristic bead shapes by dropwise addition into calcium chloride solution. The prepared beads were characterized using FTIR, XRD, TGA and SEM analysis. Microhardness testing was used to evaluate the mechanical properties. The prepared beads were investigated for water adsorption behavior to ascertain its ability for body fluid uptake and adjusted accordingly to the bone cavity. Drug loading and subsequently the antibacterial activity was investigated for the prepared beads. The biocompatibility was assessed using the hemolysis testing and cell proliferation assay. The prepared collagen-alginate-HA beads, having biocompatibility and good mechanical properties, have showed an option of promising biologically active bone fillers for bone regeneration.     

两种万古霉素藻酸盐微缓释球释放曲线的对比研究

目的制备两种不同的万古霉素藻酸盐微球(Vanco-AB),对比其体外缓释性能和抗菌活性,探讨其在骨科抗感染治疗中的可能应用。方法应用滴注法制备万古霉素藻酸盐微球,通过分别改变万古霉素和藻酸盐溶液的浓度,优选万古霉素含量高的微球,然后用不同浓度纤维蛋白凝胶包裹,通过检测各组万古霉素有效的释放时间来筛选适当包裹微球。结果随着万古霉素和藻酸盐溶液浓度的升高,微球中的万古霉素含量增高,16%藻酸盐和50 mg/mL万古霉素等体积混合液制备的微球中万古霉素含量最高,达(27.36±0.90)%。在前述微球的基础上,75 mg/mL纤维蛋白包裹的微球释放万古霉素的浓度可有效杀灭金黄色葡萄球菌19 d。结论优选后的FG-Vanco-AB可达到有效杀菌浓度的释放天数显著延长,增加了临床应用的可能。     

Alginate-controlled formation of nanoscale calcium carbonate and hydroxyapatite mineral phase within hydrogel networks

A one-step method was used to make nanostructured composites from alginate and calcium carbonate or calcium phosphate. Nanometer-scale mineral phase was successfully formed within the gel network of alginate gel beads, and the composites were characterized. It was found that calcite was the dominating polymorph in the calcium carbonate mineralized beads, while stoichiometric hydroxyapatite was formed in the calcium phosphate mineralized beads. A combination of electron microscopy, Fourier-transform infrared spectroscopy, thermogravimetric analysis and powder X-ray diffraction showed that alginate played an active role in controlling mineral size, morphology and polymorphy. For the calcium phosphate mineralized beads, alginate was shown to modulate stoichiometric hydroxyapatite with low crystallinity at room temperature, which may have important applications in tissue engineering. The results presented in this work demonstrate important aspects of alginate-controlled crystallization, which contributes to the understanding of composite material design.     

Ionically crosslinked alginate hydrogels as scaffolds for tissue engineering: part 1. Structure, gelation rate and mechanical properties

Alginate gels have been used in both drug delivery and cell encapsulation applications in the bead form usually produced by dripping alginate solution into a CaCl2 bath. The major disadvantages to these systems are that the gelation rate is hard to control; the resulting structure is not uniform; and mechanically strong and complex-shaped 3-D structures are difficult to achieve. In this work controlled gelation rate was achieved with CaCO3-GDL and CaSO4-CaCO3-GDL systems, and homogeneous alginate gels were formulated as scaffolds with defined dimensions for tissue engineering applications. Gelation rate increased with increasing total calcium content, increasing proportion of CaSO4, increasing temperature and decreasing alginate concentration. Mechanical properties of the alginate gels were controlled by the compositional variables. Slower gelation systems generate more uniform and mechanically stronger gels than faster gelation systems. The compressive modulus and strength increased with alginate concentration, total calcium content, molecular weight and guluronic acid (G) content of the alginate. MC3T3-E1 osteoblastic cells were uniformly incorporated in the alginate gels and cultured in vitro. These results demonstrated how alginate gel and gel/cell systems could be formulated with controlled structure, gelation rate, and mechanical properties for tissue engineering and other biomedical applications.     

Preparation of novel silica-coated alginate gel beads for efficient encapsulation of yeast alcohol dehydrogenase

Biomimetic formation has undoubtedly inspired the preparation of novel organic-inorganic hybrid composites. In this study, silica-coated alginate gel beads were prepared by coating the surface of alginate gel beads with silica film derived from tetramethoxysilane (TMOS). The composition and structure of the silica film were characterized by FT-IR and SEM equipped with EDX. The swelling behavior of silica-coated alginate gel beads was studied to be more stable against swelling than that of alginate gel beads. The results showed that silica-coated alginate gel beads exhibited appropriate diffusion property. The effective diffusion coefficient (D(e)) of NADH in silica-coated alginate beads was 1.76 x 10(-10) m2/s, while the effective diffusion coefficient in alginate beads was 1.84 x 10(-10) m2/s. The model enzyme yeast alcohol dehydrogenase (YADH) was encapsulated in silica-coated alginate and pure alginate beads, respectively. Enzyme leakage of YADH in alginate gel beads was determined to be 32%, while the enzyme leakage in silica-coated alginate gel beads was as low as 11%. Furthermore, the relative activity of YADH in alginate gel beads decreased almost to zero after 10 recycles, while the relative activity of YADH in silica-coated alginate gel beads was 81.3%. The recycling stability of YADH in silica-coated alginate gel beads was found to be increased significantly mainly due to the effective inhibition of enzyme leakage by compact silica film.     

Preparation of novel silica-coated alginate gel beads for efficient encapsulation of yeast alcohol dehydrogenase

Biomimetic formation has undoubtedly inspired the preparation of novel organic–inorganic hybrid composites. In this study, silica-coated alginate gel beads were prepared by coating the surface of alginate gel beads with silica film derived from tetramethoxysilane (TMOS). The composition and structure of the silica film were characterized by FT-IR and SEM equipped with EDX. The swelling behavior of silica-coated alginate gel beads was studied to be more stable against swelling than that of alginate gel beads. The results showed that silica-coated alginate gel beads exhibited appropriate diffusion property. The effective diffusion coefficient (D _e) of NADH in silica-coated alginate beads was 1.76 × 10^?10 m²/s, while the effective diffusion coefficient in alginate beads was 1.84 × 10^?10 m²/s. The model enzyme yeast alcohol dehydrogenase (YADH) was encapsulated in silica-coated alginate and pure alginate beads, respectively. Enzyme leakage of YADH in alginate gel beads was determined to be 32%, while the enzyme leakage in silica-coated alginate gel beads was as low as 11%. Furthermore, the relative activity of YADH in alginate gel beads decreased almost to zero after 10 recycles, while the relative activity of YADH in silica-coated alginate gel beads was 81.3%. The recycling stability of YADH in silica-coated alginate gel beads was found to be increased significantly mainly due to the effective inhibition of enzyme leakage by compact silica film.