Optimization of protein encapsulation in PLGA nanoparticles

Influences of process parameters were investigated on the efficiency of encapsulation of bovine serum albumin (BSA) in poly(dl-lactic-co-glycolic acid) (PLGA) nanoparticles produced by w₁/o/w₂ (water-in-oil-in-water) double emulsion-solvent evaporation method. According to a 5-factorial 3-level Box-Behnken type experimental design aqueous solution of BSA was emulsified in an immiscible organic phase composed of dichloromethane and various quantities of dissolved PLGA to get water-in-oil (w₁/o) emulsion. This latter was then dispersed in a second aqueous phase (w₂) containing poly-vinyl-alcohol (PVA) surfactant as an emulsifier/stabilising agent. PLGA nanoparticles with encapsulated BSA were obtained by evaporating the dichloromethane from the w₁/o droplets. Encapsulation efficiency was determined as the weight ratio of BSA remained in the PLGA nanoparticles relative to the total weight of BSA used in the process. By statistical evaluation of the experimental results an equation was proposed to predict the encapsulation efficiency as a function of five process variables. Two optimization procedures were carried out to increase the efficiency of encapsulation, with and without constraints referring to the required mean particle size. Correlation was found between the latter and the achievable maximal encapsulation efficiency under optimal process conditions.     

Preparation of biocompatible magnetite-PLGA composite nanoparticles using supercritical fluid extraction of emulsions

We report a new strategy for the production via supercritical fluid extraction of emulsion of biocompatible magnetic nanocomposite particles made of magnetite nanocrystals dispersed in a poly(lactic-co-glycolic) acid (PLGA) matrix. Ricinoleic acid-stabilized magnetic nanocrystals have been prepared via coprecipitation of two iron salts in alkaline environment, and subsequently dispersed in a solution of PLGA in dichloromethane. The obtained oil phase has then been dispersed in an aqueous solution of polyvinyl alcohol (PVA) in order to obtain a kinetically stable oil-in-water miniemulsion. The solvent was finally extracted via supercritical fluid extraction of emulsions. The continuous extraction of dichloromethane by means of supercritical CO₂ leads directly to a stable suspension of magnetite-PLGA composite nanoparticles in water. The influence of those parameters affecting the final particle size distribution and morphology, primarily emulsifier amount and magnetite content, has been investigated, so as to optimize the process. Analysis of the products, performed through light scattering and electron microscopy, indicates that narrower size distributions are obtained with larger amounts of emulsifier and lower amounts of magnetite. The morphology of the particles tends to be of Janus type, with the magnetite accumulated on one hemisphere of the particle. The proposed approach is suitable for the preparation of large quantities of high-quality magnetite-PLGA composite nanoparticles for biomedical applications.     

Preparation of magnetic poly(lactic‐co‐glycolic acid) microspheres featuring monodispersity and controllable particle size using a microchannel device

Poly(lactic‐co‐glycolic acid) (PLGA) microspheres prepared using a traditional solvent evaporation or double emulsification method are usually polydisperse with an uncontrollable particle size distribution, which brings about poor application performance. In our research, monodisperse magnetic PLGA microspheres were prepared using a microchannel device based on a water‐in‐oil‐in‐water composite emulsion. The composite emulsion was formed by injecting a dichloromethane–gelatin water‐in‐oil emulsion into a microchannel together with an external water phase, i.e. poly(vinyl alcohol) (PVA) aqueous solution. Mean particle size control of the microspheres was executed using the osmotic pressure difference between internal and external aqueous phases caused by regulating NaCl concentration in PVA aqueous phase. It is found that monodisperse magnetic PLGA microspheres with high magnetic responsiveness can be successfully prepared combining the microchannel device with composite emulsion method. Mean particle size of the microspheres with coefficient of variation value below 4.72% is controllable from 123 to 203 µm depending on the osmotic pressure. The resulting samples have pyknotic and smooth surfaces, as well as spherical appearance. These monodisperse magnetic PLGA microspheres with good superparamagnetism and magnetic mobility have potential use as drug carriers for uniform release and magnetic targeting hyperthermia in biological fields. © 2015 Society of Chemical Industry     

Optimization of Formulation Variables to Increase Antigen Entrapment in PLGA Particles

Efficient antigen entrapment is a key factor in preparation of poly (lactide-co-glycolide) acid (PLGA) vaccine formulations when the antigen is of short supply. This study presents a systematic approach in the testing of formulation variables with the objective to increase antigen entrapment in particles when the antigen stock concentration was low. Some of the experimental variables tested were poly (vinyl) alcohol (PVA) concentration in the inner (W₁) and outer (W₂) aqueous phase, W₁/oil (O) phase ratio and choice of organic solvent. The double emulsion solvent evaporation technique was applied to prepare PLGA particles with sonication as the emulsifying force. To measure antigen entrapment efficiency, the antigen (bovine serum albumin, BSA) was isotope labeled with ¹²⁵iodine (¹²⁵I). Our results demonstrated that a low PVA concentration in the inner aqueous (W₁) phase was beneficial to achieve a high encapsulation efficiency of antigen. On the contrary, in the outer aqueous (W₂) phase, a high PVA concentration favored antigen entrapment. We also demonstrated that decreasing the W₁ to O/polymer ratio contributed to increased entrapment efficiency. Testing different organic solvents (ethyl acetate, dichloromethane and chloroform), either alone or in combination, revealed that using chloroform as solvent resulted in the highest encapsulation of antigen and the highest production yield. Some of the results presented in this work are in disagreement with well-established formulation variables from previous studies.     

单分散性PLGA磁性微球制备及其缓释性研究

为获得单分散性PLGA磁性微球,文中以纳米四氧化三铁明胶分散液作为内水相(W1),PLGA（聚乳酸羟基乙酸共聚物）的二氯甲烷溶液作为油相(O),PVA（聚乙烯醇）水溶液作为外水相(W2),利用T型微通道并采用复合乳液法制备PLGA磁性微球,考察流速比和油相与内水相体积比对微球制备的影响。借助FTIR、SEM及VSM（振动样品磁强计）对磁性微球组分、形貌、粒径分布和磁学性能进行表征;并以阿司匹林作为药物模型进行缓释性测试。结果表明：流速比v(W2):v(W1/O)=120:1且体积比V(O):V(W1)=2:1时可均匀成球,其粒径分布系数CV值仅为4.66%,表现出良好单分散性;此时比饱和磁化强度可达1.52emu/g,兼具优异顺磁性。制得的载药微球在60h内表现出阶段性匀速释放,且有较好磁响应性,有望用于磁响应性药物载体。     

快速膜乳化法制备粒径均一的PLGA微球和微囊

以聚(乳酸-羟基乙酸)(PLGA)为膜材,采用快速膜乳化结合溶剂萃取法制备了胰高血糖素样肽-1(GLP-1)微囊,研究了PLGA分子量对药物装载率、药物活性和体外释放行为的影响. 制备均一微球的优化条件为过膜压力1000 kPa,过膜次数3次,外水相稳定剂聚乙烯醇浓度19 g/L,油水体积比1:5. 在此条件下,制备了粒径350 nm左右、多分散系数小于0.050的载GLP-1的PLGA微囊,GLP-1包埋率达65%以上,活性保留达85%以上,药物体外释药可达20 d.     

A Multioptimization Approach to Assessment of Drug Delivery of PLGA Nanoparticles: Simultaneous Control of Particle Size and Release Behavior

A multiple optimization strategy was implemented on the experimental data obtained in this study to assess drug-delivery behavior of poly(D,L-lactide-co-glycolide) (PLGA) particles. Albumin–fluorescein isothiocyanate conjugate (FITC-albumin)-loaded PLGA micro- and nanoparticles were prepared by water-in-oil-in-water double emulsion method. The experiments were carried out based on L8 Taguchi design where the effects of PLGA molecular weight, PLGA concentration in the oil phase, and sonication rate in the second emulsification step of preparation process on particle size and percentage of initial burst release of PLGA were discussed. Multioptimization toward PLGA particle size and initial burst elucidated that all aforementioned variables affect the characteristics of PLGA particles, but PLGA molecular weight was appeared as the most significant factor among studied variables. Multifarious optimizations were tested and executed varying the aforementioned factors to minimize both the size and percentage of initial burst of PLGA particles. The FTIR experiment of the optimal formulation showed the successful incorporation of the drug into the particles. These results enable careful selection and definition of optimal process parameters for the preparation of PLGA nanoparticles with sustained release properties. Furthermore, the methodology developed in this work introduces a useful tool to meet a drug delivery system with optimal release behavior.     

Ethylenediamino bridged bis(β‐cyclodextrin)/ poly(DL‐lactic‐co‐glycolic acid) nanoparticles prepared by modified double emulsion method: Effect of polyvinyl alcohol on nanoparticle properties

This study continues long‐standing efforts to develop protein delivery systems based on cyclodextrin‐conjugated polyester in our laboratory. The crude products of ethylenediamino bridged bis(β‐cyclodextrin)‐conjugated poly(DL ‐lactic‐co‐glycolic acid) were used in this study to make full use of unreacted reactant. With bovine serum albumin (BSA) as a model protein, the encapsulation effects (the encapsulation efficiency and particle size) of nanoparticles were similar to those of using pure conjugated products. Besides, a water‐in‐oil‐in‐water emulsification technique was conveniently modified. By adding polyvinyl alcohol (PVA) in the internal aqueous phase, a more stabilized emulsion was formed. Consequently, less PVA (～ 0.05%) was needed in the outer aqueous phase and less PVA (0.14 g/g nanoparticles) remained in the nanoparticles. This modification resulted in improved encapsulation efficiency (～ 89–94%) of BSA and an enlarged particle size (340–390 nm). Furthermore, the burst release of BSA at the 1st day was less pronounced (7–12% of the encapsulated amount) than that of nanoparticles with no PVA added in the internal aqueous phase. Degradation studies using transmission electron microscope and gel permeation chromatography suggested that the mechanism for protein release was mainly through nanoparticles erosion. © 2007 Wiley Periodicals, Inc. J Appl Polym Sci, 2008     

Production of oil-containing crosslinked poly(vinyl alcohol) microcapsules by phase separation: Effect of process parameters on the capsule size distribution

Oil-containing poly(vinyl alcohol) (PVA) microcapsules in the size range of 5–20 μm were prepared by the simple coacervation of PVA followed by chemical crosslinking of the coacervated PVA membrane with glutaraldehyde. Coacervation of the aqueous polymer solution was achieved by the addition of a phase separation inducer (e.g., sodium sulfate). PVA of different grades (e.g., molecular weight and degree of hydrolysis) was utilized both as stabilizer and wall-forming material. Dispersion of the oil phase in the aqueous PVA solution was effected by a homogenizer. The effects of the various process parameters, such as the agitation speed, the type and concentration of PVA, the volume ratio of the internal oil phase to the external aqueous phase, the viscosity of the oil phase as well as the electrolyte concentration in the aqueous solution, on the stability and the size distribution of the emulsion droplets and microcapsules were experimentally investigated. It was shown that high agitation rates and low interfacial tension (e.g., high PVA concentrations) resulted in a significant reduction of the size of the emulsion droplets and microcapsules. On the other hand, as the viscosity and the amount of the dispersed oil phase increased, the capsule size increased. Finally, it was found that the concentration of the electrolyte significantly affected the stability of the (o/w) emulsion, the size and concentration of coacervated PVA colloidal aggregates, as well as the morphology of the polymer wall membrane formed by the adsorption of the polymer-rich phase to the oil/water interface. © 1996 John Wiley & Sons, Inc.     

可生物降解聚乳酸纳米粒的制备及表征

采用乳化 -溶剂蒸发法 ( O/W)制备聚乳酸 ( PLA)纳米粒 ,用透射电子显微镜和激光粒度仪对纳米粒进行了表征 ,纳米粒具有规整的球形且正态分布 ,同时从有机相和水相的体积比、聚合物的浓度、表面活性剂的选择及表面活性剂的浓度等几个因素对纳米粒粒径大小的影响作了较详细地讨论。有机相与水相的体积比从 1∶ 3减小到 1∶ 30 ,纳米粒的粒径从 ( 30 6.2 + 1 1 ) nm减小到( 1 87.1 + 2 .4) nm;聚合物在有机相的浓度从 1 %(质量分数 )增加到 5 %(质量分数 ) ,纳米粒的粒径从 1 94nm增加到 32 1 nm;随着水相中表面活性剂浓度从 0 .5 (质量体积分数 )增大到 3.5 %(质量体积分数 ) ,纳米粒粒径从 2 0 2 nm减小 1 78nm且有一个低的多分散指数。而且还比较了搅拌蒸发法和减压抽提法对纳米粒表面形态的影响     

奥曲肽长效生物可降解微球的制备工艺研究

运用复乳法制备奥曲肽PLGA长效生物可降解微球,并用正交法优化微球制备工艺。利用HPLC、显微镜、激光粒度仪等对微球进行综合质量研究。结果表明,复乳法制备奥曲肽微球的最佳工艺参数为:内水相药物与中油相PLGA的质量比为1∶5,中油相PLGA的浓度为10%,外水相乳化剂为1%的22 000分子量聚乙烯(PVA)水溶液,中油相与外水相的体积比不小于1∶50,复乳化采用机械搅拌法,搅拌速度为1 200 r/min。在该工艺条件下制得的微球,包封率为35.1%,载药量为2.98%,平均粒径为26.3μm,微球外观圆整,形态良好。     

Production of double‐walled nanoparticles containing meloxicam

Double‐walled nanospheres, containing meloxicam, were fabricated with poly‐(D,L ‐lactide‐co‐glycolide) (PLGA) and poly(L ‐lactide) (PLLA) using the solvent evaporation technique. This article discusses the effect of formulation variables [sonication power, sonication time, concentration of poly(vinyl alcohol), organic/aqueous volume ratio in the first emulsion] on the production of double‐walled nanospheres. The involved phase separation of these two polymers was investigated using differential scanning calorimetry. Double‐walled microspheres containing meloxicam were also produced to determine the composition of the shell and core polymer, based on different solubilities of polymers in ethyl acetate, and to examine the inner morphology and drug distribution using optical and fluorescence microscopy. The produced microparticles have shown a double‐walled structure with meloxicam solubilized in the PLGA‐rich phase. Therefore, adjusting the selected formulation variables and using a mass ratio of 1:1 PLLA/PLGA, double‐walled nanospheres where meloxicam is dispersed within the core can be produced. POLYM. ENG. SCI., 2013. © 2012 Society of Plastics Engineers     

Preparation of magnetic poly(lactic‐co‐glycolic acid) microspheres with a controllable particle size based on a composite emulsion and their release properties for curcumin loading

Because of their unique magnetic features and good biocompatibility, magnetic poly(lactic‐co‐glycolic) acid (PLGA) microspheres have great application potential in magnetic targeted drug‐delivery systems. In this research, magnetic PLGA microspheres with controllable particle sizes were successfully prepared from a composite emulsion with a T‐shaped microchannel reactor. A water‐in‐oil‐in‐water composite emulsion was generated by the injection of a dichloromethane/gelatin water‐in‐oil initial emulsion into the microchannel together with a coating aqueous phase, that is, the aqueous solution of glucose and poly(vinyl alcohol). The mean particle size of the microspheres could be controlled by the manipulation of the osmotic pressure difference between the internal and external aqueous phases via changes in the glucose concentration. Curcumin, a drug with an inhibitory effect on tumor cells, was used to exemplify the release properties of the magnetic PLGA microspheres. We found that the mean particle size of the microspheres ranged from 16 to 207 μm with glucose concentrations from 0 to 20 wt %. The resulting microspheres showed a rapid magnetic response, good superparamagnetism, and a considerable magnetocaloric effect, with a maximum magnetic entropy of 0.061 J·kg^?1·K^?1 at 325 K. An encapsulation efficiency of up to 77.9% was achieved at a loading ratio of 3.2% curcumin. A release ratio of 72.4% curcumin from the magnetic PLGA microspheres was achieved within 120 h in a phosphate‐buffered solution. The magnetic PLGA microspheres showed potential to be used as drug carriers for magnetic targeted tumor therapy. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2016 , 133, 43317.     

Enhanced photoluminescence of porous silicon nanoparticles coated by bioresorbable polymers

A significant enhancement of the photoluminescence (PL) efficiency is observed for aqueous suspensions of porous silicon nanoparticles (PSiNPs) coated by bioresorbable polymers, i.e., polylactic-co-glycolic acid (PLGA) and polyvinyl alcohol (PVA). PSiNPs with average size about 100 nm prepared by mechanical grinding of electrochemically etched porous silicon were dispersed in water to prepare the stable suspension. The inner hydrophobic PLGA layer prevents the PSiNPs from the dissolution in water, while the outer PVA layer makes the PSiNPs hydrophilic. The PL quantum yield of PLGA/PVA-coated PSiNPs was found to increase by three times for 2 weeks of the storage in water. The observed effect is explained by taking into account both suppression of the dissolution of PSiNPs in water and a process of the passivation of nonradiative defects in PSiNPs. The obtained results are interesting in view of the potential applications of PSiNPs in bioimaging.     

PLGA Biodegradable Nanoparticles Containing Perphenazine or Chlorpromazine Hydrochloride: Effect of Formulation and Release

In our study, poly(dl-lactide-co-glycolide) (PLGA) nanoparticles loaded with perphenazine (PPH) and chlorpromazine hydrochloride (CPZ-HCl) were formulated by emulsion solvent evaporation technique. The effect of various processing variables, including PLGA concentration, theoretical drug loading, poly(vinyl alcohol) (PVA) concentration and the power of sonication were assessed systematically to obtain higher encapsulation efficiency and to minimize the nanoparticles size. By the optimization formulation process, the nanoparticles were obtained in submicron size from 325.5 ± 32.4 to 374.3 ± 10.1 nm for nanoparticles loaded with PPH and CPZ-HCl, respectively. Nanoparticles observed by scanning electron microscopy (SEM) presented smooth surface and spherical shape. The encapsulation efficiency of nanoparticles loaded with PPH and CPZ-HCl were 83.9% and 71.0%, respectively. The drug loading were 51.1% and 39.4% for PPH and CPZ-HCl, respectively. Lyophilized nanoparticles with different PLGA concentration 0.8%, 1.3% and 1.6% (w/v) in formulation process were evaluated for in vitro release in phosphate buffered saline (pH = 7.4) by using dialysis bags. The release profile for both drugs have shown that the rate of PPH and CPZ-HCl release were dependent on a size and amount of drugs in the nanoparticles.     

快速膜乳化法制备载生长激素释放肽-6的PLGA微球

采用快速膜乳化法制备了聚(乳酸-羟基乙酸)(PLGA)微球,得到制备PLGA微球的优化条件为：过膜压力5 kPa,水相中PVA浓度19 g/L,油/水相体积比1:10,该条件下所制空白微球的平均粒径约为24 mm,粒径分布系数Span<0.7. 在此基础上制备载生长激素释放肽-6(GHRP-6)微球,油相乳化剂浓度2.5 g/L、外水相中NaCl浓度10 g/L条件下所制载GHRP-6微球包埋率最高可达85%,初乳制备方式对药物包埋率及体外释放行为均有较大影响,超声法制备的初乳所得微球内部结构紧密,药物包埋率较高(85%),但释药缓慢;而均质法制备的初乳所得微球内部结构疏松,药物包埋率较低(76.8%),但在体外释放更完全.     

相分离-溶胀法制备生物可降解PELA多孔纳米微球

采用相分离-溶剂去除法制备纳米尺度的单甲氧基聚乙二醇-聚乳酸共聚物(PELA)微球,分析了纳米微球在溶液中的形成机理;用有机溶剂对纳米微球进行溶胀制孔,制备出具有不同孔道特征的纳米微球. 结果表明,以乙醇+丙酮为油相、去离子水为水相,油相中PELA含量6.5 g/L、水相中SDS含量1%、油与水相体积比1:6、油相中乙醇含量50%(j)条件下,所制微球粒径为78.48 nm. 溶胀时间为0.5 h时,以甲苯为溶胀剂所制PELA微球具有中空单孔结构,以二氯甲烷为溶胀剂所制PELA微球具有多孔结构. 用相同方法制备了具有孔结构的聚乳酸、聚(乳酸-羟基乙酸)共聚物纳米微球,其与PELA的成孔趋势相同. 以模拟体液考察多孔PELA纳米微球的降解性能,30 d可充分降解.     

双乳化-凝胶法制备单分散海藻酸钙微球及其载BSA研究

采用双乳化-凝胶法制备了单分散的海藻酸钙凝胶微球,并通过正交试验系统考察了海藻酸钠浓度、氯化钙浓度、表面活性剂浓度、搅拌速度和油水比对海藻酸钙凝胶微球粒径及形貌的影响。在优化的条件下,制备出了平均粒径为4μm、单分散和球形度好的海藻酸钙凝胶微球。包埋模型药物牛血清白蛋白(BSA)的过程中,以去离子水作为洗涤液洗涤海藻酸钙微球时,BSA的包封率仅为13%左右;当水洗液的pH值为3.2时,BSA的包封率提高到66%左右,载药率可达16%,这是海藻酸钙pH值响应溶胀和BSA与海藻酸盐之间静电作用的结果。微球中BSA的体外释放曲线表明,该系统具有在模拟胃液中释药速率慢、释药量低、模拟肠液中释药迅速的特性。因此,双乳化-凝胶法制备海藻酸钙微球有望成为制备蛋白类药物控释制剂的一种新方法,以达到靶向快速给药的目的。     

Synthesis strategies for optimizing sizes of PLGA nanoparticles containing recombinant Chenopodium album (rChe a 3) allergen

The size of antigen delivery systems impact on the immune induction. The purpose of this study was to obtain poly(lactide-co-glycolide) (PLGA) nanoparticles (NPs) containing rChe a 3 allergen in different sizes (about 200, 400, and 800?nm). The optimal conditions were obtained by adjusting different parameters comprising molecular weight/concentration of polyvinyl alcohol, solvent type (dichloromethane or dichloromethane/acetone), sonication time, and addition of cryoprotectant (sucrose). The optimized formulations were found to be stable for 60 days at 4°C. In conclusion, the preparation conditions need to be adjusted to obtain allergen-PLGA NPs with specific aforementioned sizes.     

Low-temperature synthesis of redispersible iron oxide nanoparticles under atmospheric pressure and ultradense reagent concentration

Highly dispersed α-Fe₂O₃ nanoparticles ca. 3 to 8 nm in diameter were prepared at atmospheric pressure, low temperature, and at an ultradense reagent concentration by titrating an aqueous ammonia solution into a dense iron oleate/toluene mixture. A transparent suspension was obtained by redispersing the prepared particles in nonpolar solvents since they were redispersible to primary particles without aggregate formations. The prepared particles were characterized by TEM, XRD, and FT-IR, and their dispersion stability in organic solvents was determined by dynamic light scattering (DLS) and viscosity measurements. In order to analyze the formation process of the highly dispersed α-Fe₂O₃ nanoparticles, time-course measurements of DLS and viscosity during the nanoparticle synthesis in toluene were carried out. A significant increase in the suspension viscosity and the formation of an aggregated structure were observed as soon as the titration of the aqueous ammonia solution. The suspension viscosity and aggregated particle size gradually reduced with continuous vigorous stirring; finally, α-Fe₂O₃ nanoparticles that were completely redispersible in nonpolar solvents were obtained after ca. 24 h. The particle size could be controlled by the synthesis temperature, and such redispersible α-Fe₂O₃ nanoparticles were obtained even when the reagent concentration was increased to 2.8 mol/L.