羧甲基壳聚糖-聚乙二醇作为胰岛素载体的研究

目的：制备胰岛素-羧甲基壳聚糖-聚乙二醇纳米粒。方法：利用红外光谱（FTIR）和核磁共振氢谱（¹H-NMR）对羧甲基壳聚糖-聚乙二醇的结构进行表征,用粒度分析仪测定纳米粒的粒径分布及电位,采用动态透析法考察纳米粒的释药性能,用CCK-8试剂盒检测纳米粒细胞毒性,以糖尿病小鼠为模型,研究纳米粒的降血糖作用。结果：聚乙二醇成功接枝到羧甲基壳聚糖上,包埋胰岛素的纳米粒的平均粒径为（257.5±12.1）nm,Zeta电位为（-15.2±0.3）mV,负载胰岛素的羧甲基壳聚糖-聚乙二醇纳米粒在中性释放介质中,5 h内胰岛素的释放速度较快,之后8 h趋于平稳,胰岛素的累计释放量可达到80%,CCK-8试剂盒显示纳米粒对L929细胞基本无细胞毒性,50 U·kg^-1的纳米粒溶液经灌胃给药后,血糖浓度明显降低。结论：胰岛素-羧甲基壳聚糖-聚乙二醇纳米粒基本无毒性,具有良好的生物相容性,对糖尿病小鼠有效发挥降血糖作用。     

纳米粒沉淀法制备阳离子载基因PLA-PEG纳米粒

目的建立纳米粒沉淀法制备阳离子PLA-PEG纳米粒的方法。方法采用单因素设计考察不同影响因素对纳米粒粒径大小的影响,在单因素考察的基础上采用正交设计优化处方,制备了粒径较小,正电荷适中的阳离子PLA-PEG纳米粒。并对纳米粒的物理性质如物理形态,平均粒径,粒度分布,Zeta电位,DNA结合率,体外细胞转染能力等进行了考察。结果采用纳米粒沉淀法,通过优化处方和工艺制得的纳米粒外观圆整,呈类球形,大小均匀,平均粒径为89.7 nm,粒径分布指数为0.185,表面荷较高的正电荷,Zeta电位为+28.9 mV,能够高效的结合DNA且能够成功的转染Hela细胞。结论优化确定了纳米粒沉淀法制备阳离子PLA-PEG纳米粒的处方和工艺,可以制备满足细胞转染要求的阳离子载基因纳米粒。     

载柚皮素壳聚糖纳米粒的制备及其体外细胞评价

目的：制备柚皮素壳聚糖纳米粒,初步探讨其对人肺腺癌细胞A549的细胞毒性和细胞摄取。方法：以壳聚糖和鱼精蛋白作为载体材料,采用离子胶凝法制备柚皮素壳聚糖纳米粒,透射电镜（TEM）观察其形态,马尔文激光粒度仪测定其粒径、分散度（PDI）和Zeta电位,离心法测定其包封率和载药量,采用恒温振荡水浴法对柚皮素壳聚糖纳米粒进行体外释放度研究,最后采用人肺癌细胞系A549细胞进行了细胞毒性、细胞摄取研究。结果：柚皮素壳聚糖纳米粒为球形或类球形粒子,结构完整,大小均一、球形度好,分散均匀,PDI、粒径、Zeta电位和包封率分别为0.268,139 nm、+15.7 mV和83.34%,柚皮素壳聚糖纳米粒体外释放呈缓释,24 h累积释放量达到了80%以上,体外释药过程用Higuchi方程拟合较好。MTT试验显示不同浓度的壳聚糖纳米粒和细胞作用72 h后,细胞活力均大于95%,本文所制备的壳聚糖纳米粒无细胞毒性。细胞摄取试验表明载FITC的壳聚糖纳米粒和A549细胞作用3 h后,可明显看到大量带绿色荧光的纳米粒穿过细胞膜进入细胞。结论：离子凝胶法成功制得粒径较小的柚皮素壳聚糖纳米粒,具有缓释性好,毒性小,壳聚糖纳米粒摄取率较高,可大大提高药物的利用率,具有广泛的应用前景。     

芹菜素丝素蛋白纳米粒的制备及其安全性、抗肿瘤活性研究

目的制备芹菜素丝素蛋白(API@SF)纳米粒,并评价其安全性和抗肿瘤活性。方法采用纳米沉淀法制备API@SF纳米粒,并对其形态、粒径、Zeta电位、载药量、体外释放等进行表征;采用溶血实验和HE染色法评价该纳米粒的安全性;采用MTT法评价该纳米粒对小鼠乳腺癌4T1细胞的抑制作用。结果本研究所制得的API@SF纳米粒呈类球形,粒径分布均匀,平均粒径为406.61 nm,多分散性指数为0.154,Zeta电位为-18.4 mV,平均载药量为5.20%。体外释放结果显示,该纳米粒在pH 5.0的释放介质中释放速率相对较快,在pH 7.4的释放介质中释放速率相对较慢。溶血实验和HE染色结果显示,该纳米粒具有良好的生物相容性。MTT实验结果显示,API@SF纳米粒对4T1细胞的抑制作用显著高于API原料药(P<0.05),其作用机制可能与提高细胞中活性氧水平有关。结论本研究成功制备了API@SF纳米粒,该纳米粒具有良好的安全性和抗肿瘤活性。     

索拉非尼-Eudragit RS 纳米粒的制备及理化性质研究

摘 要 目的： 制备索拉非尼-Eudragit RS 纳米粒（sorafenib-Eudragit RS nanoparticles,S-E NPs) ,优选制备处方及初步探索其理化性质。方法: 溶剂 非溶剂法制备索拉非尼 Eudragit RS 纳米粒,采用单因素试验研究溶剂、稳定剂类型、载体比例、有机相水相比例对纳米粒理化性质的影响。通过粒径和Zeta电位、形貌观察、体外释药考察对其进行质量评价。结果:优选处方制备的索拉非尼 Eudragit RS 纳米粒的平均粒径为(86.72±3.71) nm,多相分散系数（PDI）为(0.200±0.032),Zeta 电位为(36.6±0.3) mV,纳米粒呈球形且分布均匀;体外释放符合 Weibull 模型(r=0.966 9)。结论: 溶剂 非溶剂法适用于索拉非尼 Eudragit RS 纳米粒的制备,所制备的纳米粒粒径较小,分布均匀,形态规则完整,具有明显缓释作用。     

中心组合设计法优化载基因壳聚糖纳米粒的最佳转染制备区域

目的：采用中心组合设计法优化载基因壳聚糖纳米粒的最佳转染制备区域。方法采用复凝聚法制备载质粒基因的壳聚糖纳米粒,选择壳聚糖浓度和质粒基因浓度作为实验考察因素,应用两因素五水平中心组合设计优化最佳转染制备区域,优化指标选择平均粒径和基因转染率。通过透射电镜观察纳米粒的形态;通过动态光散射和电泳光散射技术分别测量纳米粒的粒径和Zeta电位;通过凝胶电泳分析考察质粒在纳米粒制备过程中的稳定性;通过倒置荧光显微镜观察质粒基因在细胞内的表达;通过流式细胞技术测定纳米粒的转染效率。结果成功优化了载基因壳聚糖纳米粒的最佳转染制备区域。优选条件下制备的纳米粒大多呈球形,纳米粒平均粒径为217．6 nm,粒径多分散系数为0．241,表明粒径分布较窄。纳米粒zeta电位为＋22．4 mV,表明纳米粒表面带有正电荷,可以增加纳米粒混悬液的稳定性。凝胶电泳分析结果表明质粒基因在纳米粒制备过程中没有遭到破坏。纳米粒的细胞转染效率比较高,能够高效地将绿色荧光蛋白质粒基因递送到细胞内,并且基因表达产生绿色荧光蛋白。结论本研究建立的数学模型具有良好的预测性。在优化的制备区域内制备的载基因壳聚糖纳米粒的转染性能比较理想。     

重组水蛭素-2鼻腔黏附纳米粒的体外评价

目的:对制备的重组水蛭素-2(rHV2)壳聚糖鼻腔黏附纳米粒进行体外评价。方法:测定纳米粒的形态、粒径大小分布及表面电位;以超速离心测定纳米粒在不同介质中的释放度;采用在体蟾蜍上腭纤毛运动试验法考察纳米粒的纤毛毒性。结果:制备的rHV2壳聚糖纳米粒呈较为均匀分散的颗粒状,平均粒径为213.2 nm,纳米粒带正电荷,Zeta电位值为+30.61 mV,体系相对稳定性较高;rHV2纳米粒在乙酸缓冲液中的累积释放百分数明显高于在磷酸缓冲液中的累积释放百分数;rHV2纳米粒溶液的纤毛毒性较小(与生理盐水组相比,P>0.05)。结论:壳聚糖鼻腔黏附纳米粒有望成为rHV2鼻腔给药的递释系统。     

卡巴他赛白蛋白纳米粒的制备及其体外生物相容性评价

目的 制备卡巴他赛白蛋白纳米粒（CBZ-BSA-Gd-NP）以降低药物毒性,并评价其体外生物相容性。方法 采用生物矿化法制备CBZ-BSA-Gd-NP,对其处方工艺进行优化,对粒径、Zeta电位、载药量等性质进行表征,并采用体外溶血试验考察其体外血液相容性。结果 制得的纳米粒包封率为63.04%,载药量为10.51%,平均粒径为（166.1±4.7） nm,粒径的多分散系数（PDI）为0.256,Zeta电位为（-18.14±1.16） mV,与卡巴他赛-吐温溶液相比,体外溶血作用显著降低。结论 该方法操作简便,制备的CBZ-BSA-Gd-NP载药量高,粒径均匀,体外血液相容性好,增加了药物使用的安全性。     

复合碳酸钙空腔纳米粒的制备及性能评价

目的 制备甘草次酸/海藻酸钠修饰碳酸钙空腔纳米粒并进行体外评价。方法 以可溶性淀粉为模板剂制备中空球状碳酸钙纳米粒（CaCO3 Nps）;在非均相体系中合成了甘草次酸/海藻酸钠聚合物（GA-ALG）;并以聚合物（GA-ALG）为壳以中空结构的碳酸钙纳米粒为核,合成了壳核结构的GA-ALG-CaCO3 Nps。采用Malvern粒度分析仪测定纳米粒子的粒度分布和Zeta电位,并通过SEM对纳米粒的形态进行表征。应用荧光分光光度计评价载盐酸阿霉素（DOX）纳米粒的载药量、包封率及体外释放特征。结果 纳米粒分布均一,平均粒径为（425.4±31.1）nm,PDI为0.289,Zeta 电位为（-17.0±0.3）mV。药物的载药量为（13.06±0.51）%,包封率为（78.35±3.08）%。;体外释放结果显示,纳米粒具有一定的缓释作用。结论 GA-ALG-CaCO3 Nps作为新型的药物载体,具有良好的pH响应性,并能显著提高载药量,还具有明显的缓释效果,为新型的纳米给药系统的深入研究提供参考。     

负载pVAX1-wapA的壳聚糖及季铵化壳聚糖纳米粒的制备研究

目的 制备负载抗龋DNA疫苗pVAX1-wapA质粒的壳聚糖和季铵化壳聚糖纳米粒,优化其制备工艺,测定其细胞转染效率。 方法 以包封率和粒径为主要指标,单因素法考察载体浓度、pH值、N/P、TPP浓度等因素的影响,Realtime-PCR检测细胞对质粒编码蛋白的转录表达水平以评价载质粒纳米粒的促转染作用。 结果 制得的载DNA疫苗纳米粒粒径均一,形态圆整。壳聚糖(CS)纳米粒粒径为(219.2±18.2) nm,Zeta电位为(24.7±3.5) mV,包封率为91.24%。季铵化壳聚糖(CSTM)纳米粒粒径为(222.5±15.6) nm,Zeta电位为(19.6±1.2) mV,包封率为87.66%。纳米粒可以促进pVAX1-wapA进入细胞,并成功被转录。 结论 制备的包载pVAX1-wapA的季铵化壳聚糖纳米粒可用于重组基因疫苗的运送。     

Preparation of BMP-2 Containing Bovine Serum Albumin (BSA) Nanoparticles Stabilized by Polymer Coating

Purpose  The purpose of this study was to investigate the preparation process of bone morphogenetic protein-2 (BMP-2) containing bovine serum albumin (BSA) nanoparticles (NPs), and to assess the bioactivity of BMP-2 encapsulated in such NPs. Methods  The NPs were prepared by a coacervation method, and the effects of process parameters on NP size and polydispersity were examined. Polymer coated NPs were characterized with respect to amount of adsorbed polymer, particle size and zeta potential. Using bone marrow stromal cells (BMSC), biocompatibility of the NPs was investigated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) Assay, and bioactivity of the encapsulated BMP-2 was investigated by alkaline phosphatase (ALP) induction and calcification. Results  The size of NPs could be controlled in the 50–400 nm range by process parameters including BSA concentration, non-solvent:solvent ratio and pH value. After coating with cationic polymers, the particle size and zeta potential were significantly increased. MTT assay indicated no toxicity of both the uncoated and coated NPs on BMSC. Based on ALP induction and calcification, full retention of BMP-2 bioactivity was retained in the polymer-coated NPs. Conclusions  This study described a preparation procedure for BSA NPs with controllable particle size, and such polymer-coated BSA NPs are promising delivery agents for local and systemic administration of BMP-2 in bone regeneration.     

甘草酸表面修饰N-己酰化壳聚糖纳米粒的制备及其稳定性研究

目的:制备N-己酰化壳聚糖纳米粒(CCS-NPs)和甘草酸表面修饰N-己酰化壳聚糖纳米粒(CCS-NPs-GL),并考察其稳定性。方法:应用离子凝胶法制备CCS-NPs,高碘酸盐氧化法制备CCS-NPs-GL;考察各纳米粒在冷冻干燥前、后和不同pH缓冲盐中粒径、电位的变化以及不同温度对CCS-NPs-GL的粒径、药物包封率和甘草酸结合率的影响。结果:CCS-NPs和CCS-NPs-GL在冷冻干燥后粒径稍有增大,缓冲盐溶液中迅速分散,电位下降不明显;在生理pH条件下,两者易于分散且粒径无明显变化;不同温度下,CCS-NPs-GL的粒径、药物包封率和甘草酸结合率无明显变化。结论:CCS-NPs-GL作为潜在的肝靶向主动传输载体,其粒子的稳定性可满足后续静脉给药的体内靶向研究和药效学评价。     

Quercetin Loaded Silica and Gold - Coated Silica Nanoparticles: Characterization,Evaluation and Comparison of Their in vitro Characteristics

Herein, silica nanoparticles (NPs) and gold-silica NPs were loaded with the anti-cancer agent quercetin (QC) to produce silica NPs-QC and gold coated silica NPs-QC, respectively. The nanosystems were characterized using transmission electron microscopy (TEM), dynamic light scattering (DLS) and Fourier transform infrared (FTIR). Drug encapsulation efficiency (EE), loading capacity (LC) and release rate were measured using UV spectrophotometer. The drug was encapsulated in silica NPs in a high percentage (71%) and reduced by about 16% after gold coating. The mean particle size increased after coating and QC loading with a polydispersity index (PDI) between (∼ 0.2 - 0.6) and negative zeta potential (-13 to - 15 mV). The intensity of FTIR peaks of silica NPs has been significantly decreased upon gold coating indicating a successful attachment of the gold thin layer. The drug release was slightly faster from coated compared to uncoated NPs but both slower than free QC. The percentages of their cell toxicity were almost the same but lower than free QC and generally were higher against HeLa cells compared to fibroblast cells. Both nanosystems could be considered as promising nanocarriers with reasonable EE, slower release rate and lower toxicity compared to the free drug.     

艾塞那肽聚乳酸-羟基乙酸共聚物纳米粒的制备及其分析方法研究

目的以聚乳酸-羟基乙酸共聚物(PLGA)为载体,用乳化复乳法制备包载艾塞那肽(EX)的PLGA纳米粒(EX-PLGANPs),并对其分析方法进行研究。方法2018年10月至2019年8月,采用Box-Behnken Design(BBD)响应面分析法对纳米粒制备的处方工艺进行优化,动态光散射技术检测EX-PLGA NPs粒径和Zeta电位;通过高效液相色谱法(HPLC)测定EX-PLGANPs中艾塞那肽含量并进行方法学验证。结果制备的EX-PLGA NPs粒径为(157.2±3.1)nm,Zeta电位为(¹19.5±2.6)mV;载药量和包封率分别为(4.41±0.28)%和(73.43±0.59)%,透射电镜图显示纳米粒外观圆整,分布均匀;EX-PLGA NPs体外稳定性良好,透析袋法释放结果显示其具有缓释效果。结论制备的EX-PLGA NPs粒径分布均一,包封率和载药量高,稳定性好,艾塞那肽含量分析方法科学有效,为艾塞那肽抗糖尿病口服缓释制剂的分析和开发提供了实验基础。     

Enhanced drug encapsulation and extended release profiles of calcium–alginate nanoparticles by using tannic acid as a bridging cross-linking agent

Abstract

Calcium alginate nanoparticles (NPs) suffer from sub-optimal stability in bio-relevant media leading to low drug encapsulation efficiency and uncontrolled release profiles. To sort out these drawbacks, a novel approach is proposed herein based on introducing tannic acid into these NPs to act as a bridging cross-linking aid agent. Calcium–alginate NPs were prepared by the ionotropic gelation method and loaded with diltiazem hydrochloride as a model drug. These NPs were characterized in terms of particle size, zeta potential, and morphology, and results were explained in accordance with Fourier-transform infrared (FTIR) spectroscopy and differential scanning calorimetry (DSC). The incorporation of tannic acid led to more than four folds increase in drug encapsulation efficiency (i.e. from 15.3% to 69.5%) and reduced burst drug release from 44% to around 10% within the first 30?min. These findings suggest the possibility of improving the properties of Ca–alginate NPs by incorporating cross-linking aid agents under mild conditions.     

Preparation and characterization of pH-sensitive calcium carbonate-chlorin e6 nanoparticles for photodynamic therapy

In the present study, we combined CaCO₃ NPs and Ce6 to construct CaCO₃-Ce6 nanoparticles (NPs). CaCO₃-Ce6 NPs were characterized in terms of particle size, zeta potential, UV-Vis absorption spectrum, fluorescence spectrum, FTIR spectrum, and pH-responsive behavior. The reactive oxygen species (ROS) generation in vitro was measured in 4T1 cells. The results showed that CaCO₃-Ce6 NPs were uniform-sized NPs with excellent fluorescence properties and pH-responsive behavior. The ability of ROS generation by CaCO₃-Ce6 NPs was stronger compared with Ce6 in 4T1 cells because Ca²⁺ could enhance the ROS generation, which could contribute to a stronger anti-tumor effect.     

紫杉醇PLGA 纳米粒的处方优化与评价

目的采用生物可降解材料乳酸-羟基乙酸共聚物(PLGA)为载体,比较不同的制备方法和工艺对紫杉醇(PTX)PLGA纳米粒(PTX-PLGA NPs)粒径的影响,筛选出最优制备工艺,并考察所制备纳米粒的体外表征以及对人源胃癌细胞SGC-7901的抗肿瘤效果,为紫杉醇缓释制剂在胃癌中的开发提供一定的实验基础。方法采用单因素实验法优化PTXPLGA NPs的制备处方及工艺,对粒径、Zeta电位、形态、稳定性、包封率和载药量及体外释放进行表征,在SGC-7901细胞中考察PTX-PLGA NPs的体外抗肿瘤效果。结果制得的PTX-PLGA NPs平均粒径为(219.9±4.5)nm,表面电位为(-21.7±1.81)mV,载药量为(4.42±1.33)%,包封率为(80.73±2.66)%,外观圆整,在37℃下PTX-PLGA NPs能够在含有血清的PBS中保持稳定72 h。24 h时,PTX-PLGA NPs的体外释放率仅24%。PTX-PLGA NPs对人胃癌细胞SGC-7901的IC₅₀为1.56×10^-10mol·L^-1,是游离PTX IC₅₀的44.4%,具有较好的抗肿瘤效果。结论所制备的PTX-PLGA NPs粒径分布均一、稳定性好、载药量高、具有缓释效果,且对胃癌细胞SGC-7901有明显的抗肿瘤作用。     

Characterization of amphiphilic dendrimer modified PEG-PLA nanoparticles prepared by a double emulsion-solvent evaporation method

Drug delivery by nanocarriers requires characterizations of suitable particle size, high drug loading and safety. In this work, we prepared an amphiphilic dendrimer modified PEG-PLA mixed nanoparticles (NPs) by a double emulsion-solvent evaporation (DESE) method. The particle size and drug encapsulation efficacy (EE) were compared to evaluate and optimize the preparation parameters. The mixed NPs had average size ranging from (102±1) nm to (137±5) nm, and the zeta potential turned to positive with incorporation of the amphiphilic dendrimer. The NPs showed different EE of docetaxel (DTX) and paclitaxel (PTX) with higher affinity to more lipophilic PTX. The blank mixed NPs showed little cytotoxicity, and the DTX-loaded NPs could effectively facilitate the antiproliferation activity on PC-3 cells. The NPs could be used as an effective drug delivery system, and its anti-tumor effect is worthy of further study.     

Improved anticancer delivery of paclitaxel by albumin surface modification of PLGA nanoparticles

Background

Nanoparticles (NPs) play an important role in anticancer delivery systems. Surface modified NPs with hydrophilic polymers such as human serum albumin (HSA) have long half-life in the blood circulation system.

Methods

The method of modified nanoprecipitation was utilized for encapsulation of paclitaxel (PTX) in poly (lactic-co-glycolic acid) (PLGA). Para-maleimide benzoic hydrazide was conjugated to PLGA for the surface modifications of PLGA NPs, and then HSA was attached on the surface of prepared NPs by maleimide attachment to thiol groups (cysteines) of albumin. The application of HSA provides for the longer blood circulation of stealth NPs due to their escape from reticuloendothelial system (RES). Then the physicochemical properties of NPs like surface morphology, size, zeta potential, and in-vitro drug release were analyzed.

Results

The particle size of NPs ranged from 170 to 190 nm and increased about 20–30 nm after HSA conjugation. The zeta potential was about -6 mV and it decreased further after HSA conjugation. The HSA conjugation in prepared NPs was proved by Fourier transform infrared (FT-IR) spectroscopy, faster degradation of HSA in Differential scanning calorimetry (DSC) characterization, and other evidences such as the increasing in size and the decreasing in zeta potential. The PTX released in a biphasic mode for all colloidal suspensions. A sustained release profile for approximately 33 days was detected after a burst effect of the loaded drug. The in vitro cytotoxicity evaluation also indicated that the HSA NPs are more cytotoxic than plain NPs.

Conclusions

HSA decoration of PLGA NPs may be a suitable method for longer blood circulation of NPs.     

Hyaluronic Acid Gel Incorporating Curcumin-Phospholipid Complex Nanoparticles Prevents Postoperative Peritoneal Adhesion

Peritoneal adhesions following abdominal procedures remain a major surgical issue. This study aimed to evaluate the preventive effect of curcumin-soybean phosphatidylcholine (CUR-SPC) complex-loaded nanoparticles (NPs) embedded in hyaluronic acid (HA) gel. CUR-SPC complex, CUR-SPC NPs, and CUR-SPC-HA gel were fully characterized regarding particle size, zeta potential, morphology, XRD, FTIR, in-vitro release, and rheology. The in vivo efficacy was investigated in a rat model of cecum abrasion and sidewall defect. The optimized spherical-shaped CUR-SPC NPs had a high CUR loading with nanometric particle size (93 nm). CUR-SPC-HA gel showed a pseudoplastic behavior with a homogeneous distribution of CUR-SPC NPs and a sustained release profile over 3 weeks. The inhibitory effect of CUR-SPC-NP-HA gel on fibroblast proliferation and collagen synthesis was evidenced by histopathological (H&E and Masson trichrome staining) and immunohistochemical analyses (vimentin and collagen I expression). Thus, the proposed NP-gel hybrid system could be a promising postoperative anti-adhesion candidate, employing the advantages of CUR, SPC, and HA.