Preparation and characterization of sterile and freeze-dried sub-200 nm nanoparticles

The feasibility of producing sterile and freeze-dried polyester nanoparticles was investigated. Various poly(D,L-lactide-co-glycolide) and poly(D,L-lactide) were selected as biodegradable polymers. Using the salting-out procedure, process parameters were optimized to obtain sub-200 nm particles. After purification, the nanoparticle suspensions containing different lyoprotectants were sterilized by filtration. Freeze-drying was performed using vials covered with 0.22 microm membrane filters in order to preserve the suspensions from bacterial contamination. Sterility was assessed on the final product according to pharmacopoeial requirements using the membrane filtration method. With all polymers tested, sub-200 nm particles could be obtained. Nanoparticles with a size as low as 102 nm were prepared with good reproducibility and narrow size distribution. Upon freeze-drying, it appeared that complete redispersion of all types of polyester nanoparticles could be obtained in presence of the lyoprotectants tested such as saccharides while aggregation was observed without lyoprotectant. Sterility testing showed no microbial contamination indicating that sterile nanoparticulate formulations have been achieved.     

Preparation and characterization of sterile sub-200 nm meso-tetra(4-hydroxylphenyl)porphyrin-loaded nanoparticles for photodynamic therapy.

A photosensitizer, meso-tetra(4-hydroxyphenyl)porphyrin, was incorporated into sub-150 nm nanoparticles using the emulsification-diffusion technique in order to perform sterilization by filtration using 0.22 microm membranes. The three selected polyesters (poly(D,L-lactide-co-glycolide), (50:50 PLGA, 75:25 PLGA) and poly(D,L-lactide (PLA)) for the nanoparticle production were all amorphous in nature and have similar molecular weights but different copolymer molar ratios. The influence of the copolymer molar ratio and the theoretical drug loading was investigated in terms of particle size, drug loading, entrapment efficiency and surface characteristics. With all the polymers used, sub-150 nm nanoparticles were produced with good reproducibility and narrow size distributions irrespective of both the polymer nature and the theoretical drug loading. After purification by cross-flow filtration, the nanoparticle suspensions were sterilized by membrane filtration and freeze-dried in the presence of a lyoprotectant (trehalose). For all types of nanoparticles, complete redispersion in various media could be obtained. All final freeze-dried products were refiltrable on a 0.22 microm membrane and were stable in terms of mean particle size and drug loading over a period up to 6 months. The effective drug loading increased at higher theoretical drug loading, the entrapment efficiency was however decreased. The same trend was observed with the three polyesters. The sterility of the final freeze-dried nanoparticles was confirmed by the results of the sterility testing which showed no bacterial contamination.     

Dispersion, fractionation and characterization of sub-100nm P25 TiO2 nanoparticles in aqueous media

Despite of the widespread use of manufactured nanomaterials and increasing concerns on their potential risk, only limited information is available on their physicochemical properties in toxicologically relavant aqueous media. Here, P25 TiO₂ (Evonik GmbH), one of the well-known and widely-used photocatalytic TiO₂, was dispersed and fractionated in aqueous media, and its physicochemical properties, especially for its sub-100 nm fraction, was carefullly studied. The colloidal properties of TiO₂ nanoparticles, such as ag]glomeration and sedimentation, were found strongly dependent on the physicochemical characteristics of nanoparticles (e.g., hydrodynamic size distribution, type of capping ligands and surface charge) as well as those of the aqueous media used (e.g., ionic strength and chemical compositions). This study has shown the importance of standardized dispersion and characterization protocol for toxicity tests, which is urgently needed for reliable, reproducible and impartial toxicity tests of manufactured nanomaterials.     

Enhanced photodynamic activity of meso-tetra(4-hydroxyphenyl)porphyrin by incorporation into sub-200 nm nanoparticles.

A photosensitizer, meso-tetra(hydroxyphenyl)porphyrin (p-THPP) was incorporated into sterile submicronic nanoparticles of poly(D,L-lactide-co-glycolide) (50:50 and 75:25 PLGA) and poly(D,L-lactide) (PLA). With all polymers used, sub-130 nm p-THPP-loaded nanoparticles with similar drug loadings and entrapment efficiencies were produced using the emulsification-diffusion technique. The photodynamic activity (photocytotoxicity) of these nanoparticles was evaluated on EMT-6 mammary tumour cells in comparison with the free drug. The influence of drug concentration (3-10 microg/ml), incubation time (5-60 min) and light dose (6-9 J/cm(2)) on p-THPP photocytotoxic efficiency was investigated. With all p-THPP formulations tested, cell viability decreased with increasing values of these parameters. The beneficial effect of nanoencapsulation compared to free drug was highlighted at drug concentrations up to 6 microg/ml and short incubation times (15-30 min). The most important photocytotoxicity was observed with 50:50 PLGA nanoparticles allowing low drug doses and short drug administration-irradiation intervals for local photodynamic therapy.     

α-常春藤皂苷丙烯酸树脂纳米粒的制备及其体外评价

目的:制备和评价α-常春藤皂苷(saponins PD,SPD)丙烯酸树脂S100纳米粒(SPD-S100-NPs)。方法:采用改良乳化-溶剂扩散法制备纳米粒,以粒径大小、包封率(entrapment efficiency,EE)和多分散指数(polydisperse index,PI)为指标,通过单因素试验和正交试验设计优化制备工艺。以红外光谱(fouriertransform infrared spectrometer,FT-IR)、X射线衍射(X-ray diffraction,XRD)、差示扫描量热分析(differentialscanning calorimetry,DSC)、体外释放试验等对纳米粒的相关性质进行评价。结果:制备的纳米粒外观圆整,平均粒径(73.1±4.6)nm,包封率(99.0±0.58)%,PI值(0.249±0.029)。药物在纳米粒中均被载体材料有效包裹,其体外释放具有显著的pH依赖性。结论:改良乳化-溶剂扩散法制备了包封率高、大小均匀的pH依赖性α-常春藤皂苷纳米粒。     

Preparation and characterization of n-dodecyl-ferulate-loaded solid lipid nanoparticles (SLN)

Solid lipid nanoparticles (SLN) containing a novel potential sunscreen n-dodecyl-ferulate (ester of ferulic acid) were developed. The preparation and stability parameters of n-dodecyl-ferulate-loaded SLN have been investigated concerning particle size, surface electrical charge (zeta potential) and matrix crystallinity. The chemical stability of n-dodecyl-ferulate at high temperatures was also assessed by thermal gravimetry analysis. For the selection of the appropriated lipid matrix, chemically different lipids were melted with 4% (m/m) of active and lipid nanoparticles were prepared by the so-called high pressure homogenization technique. n-Dodecyl-ferulate-loaded SLN prepared with cetyl palmitate showed the lowest mean particle size and polydispersity index, as well as the highest physical stability during storage time of 21 days at 4, 20 and 40 degrees C. These colloidal dispersions containing the sunscreen also exhibited the common melting behaviour of aqueous SLN dispersions.     

N-琥珀酰壳聚糖纳米粒的制备及体外评价

目的制备N-琥珀酰壳聚糖纳米粒并对其进行体外评价。方法采用乳化溶剂挥发法制备N-琥珀酰壳聚糖纳米粒;以包封率、载药量及粒径为指标,采用正交设计法对处方进行优化;考察其理化特征及体外释药行为。结果纳米粒包封率及载药量分别为62.36%和18.98%,平均粒径及zeta电位分别为(206.6±64.7)nm和(-27.2±0.2)mV;1 h药物释放达到45%,随后药物的释药行为是一个缓释过程。结论作者采用乳化溶剂挥发法成功制得N-琥珀酰壳聚糖纳米粒。该方法制得纳米粒包封率较高,制备工艺简单。     

聚乙二醇单甲醚接枝壳聚糖自组装纳米球的制备*

目的:合成聚乙二醇单甲醚接枝壳聚糖(monomethoxy poly(ethylene glycol)-grafted-chitosan,mPEG-g- CS),并制备自组装纳米球。方法:利用甲醛连接法将聚乙二醇单甲醚(monomethoxy poly(ethylene glycol),mPEG)接枝干壳聚糖(ehitosan,CS)分子,得到聚乙二醇(poly(ethylene glycol),PEG)改性的壳聚糖衍生物,并通过傅立叶红外光谱仪(Fourier transform infrared spectroscopy,FT-IR),核磁共振仪(proton nuclear magnetic resonance,~1H-NMR)对产物进行结构表征;采用超声透析法制备自组装纳米球,并通过透射电镜(transmission electron microscopy,TEM),动态激光粒度分析仪(dynamic laser light scattering,DLLS)表征了纳米球的形态和粒径;以芘为荧光探针,通过荧光检测分析测定了mPEG-g-CS的临界胶束浓度(critical micellar concentration,CMC)。结果:通过FT-IR,~1H- NMR确证了接枝产物的存在;mPEG-g-CS在水溶液中能够自组装形成球状纳米胶束,平均粒径为250 nm。结论:通过甲醛连接法制备mPEG-g-CS,具有制备方法简捷、反应周期短、易操作的优点。利用该产物制备的纳米球有望成为长循环纳米药物载体。     

Preparation of poly (alkylcyanoacrylate) nanoparticles by polymerization of water-free microemulsions

Phase diagrams of the pseudoternary systems ethyloleate, polyoxyethylene 20 sorbitan mono-oleate/sorbitan monolaurate and propylene glycol with and without butanol as a co-surfactant were prepared. Areas containing optically isotropic, one-phase systems were identified and samples therein designated as droplet, bicontinuous or solution type microemulsions using conductivity, viscosity and self-diffusion NMR. Nanoparticles were prepared by polymerization of selected microemulsions with ethyl-2-cyanoacrylate and the morphology of the particles was investigated. Addition of monomer to all types of microemulsions led to the formation of nanoparticles, which had an average size of 244 +/- 25 nm, an average polydispersity index of 0.15 +/- 0.04 and a zeta-potential of -17 +/- 3 mV. The formation of particles from water-free microemulsions of different types is surprising, particularly considering that polymerization is expected to occur at a water-oil interface by base-catalysed polymerization. It would appear that propylene glycol is sufficiently nucleophilic to initiate the polymerization. The use of water-free microemulsions as templates for the preparation of poly (alkylcyanoacrylate) nanoparticles opens up interesting opportunities for the encapsulation of bioactives which do not have suitable properties for encapsulation on the basis of water-containing microemulsions.     

姜黄素聚乙二醇-聚己内酯纳米粒的制备及制剂学性质分析

目的制备具有肝脏靶向性的姜黄素聚乙二醇-聚己内酯纳米粒。方法采用乳化溶剂挥发法,通过正交实验筛选最佳处方工艺,并按处方工艺制备姜黄素聚乙二醇-聚己内酯纳米粒。结果当选择聚乙二醇-聚己内酯100 mg、泊洛沙姆188浓度1.5%、乙酸乙酯与二氯甲烷体积比2∶1、有机相与内水相体积比为1∶4时,制备的姜黄素聚乙二醇-聚己内酯纳米粒粒径为(102.84±2.68)nm、多分散系数为0.105±0.010、包封率为84.36%±1.19%、载药量为4.70%±0.27%。结论本方法成功制备了姜黄素聚乙二醇-聚己内酯纳米粒,为下一步的药动学及体外抗肿瘤活性研究奠定基础。     

5-氟尿嘧啶-N-琥珀酰壳聚糖纳米粒冻干粉针制备

目的:考察5-氟尿嘧啶-N-琥珀酰壳聚糖纳米粒(5-FU-Suc-Chi/NPs)稳定性,制备纳米粒冻干粉针.方法:采用乳化溶剂挥发法制备5-FU-Suc-Chi/NPs;考察纳米粒溶液稳定性;优化纳米粒冻干粉针处方.结果:选定12%的甘露醇为5-FU-Suc-Chi/NPs的支架剂,制备纳米粒冻干粉.结论:5-FU-...     

表面修饰的灯盏花素聚乳酸纳米粒的制备和大鼠体内药动学*

目的:制备灯盏花素聚乳酸纳米粒并对其进行了表面修饰,同时考察了游离药物和纳米药物经大鼠尾静脉注射后在动物体内的药动学。方法:采用自乳化溶剂扩散法制备灯盏花素聚乳酸纳米粒,并用泊洛沙姆188对纳米粒进行表面修饰,采用反相高效液相色谱法(RP-HPLC)测定纳米粒的包封率、载药量和血浆样品中灯盏花素的含量,药时数据采用DASver 1.0药代计算程序处理。结果:载药纳米粒平均粒径为177.2和319.6 nm,多分散指数分别为(0.11±0.01)和(0.12±0.02),平均包封率及载药量分别为(86.9±0.9)%,(8.0±0.2)%和(93.1±0.6)%,(8.5±0.1)%,游离灯盏花素iv后呈二室模型,t1/2β为(0.81±0.14)m in,纳米组则呈一室模型,2种粒径的纳米粒的t1/2β分别为(8.90±0.16)m in(177.2 nm)和(13.90±0.07)m in(319.6 nm)。游离灯盏花素和2种粒径的灯盏花聚乳酸纳米粒的AUC0~t分别为(158.82±69.96),(1 476.25±51.22)和(704.95±25.39)mg.m in.L-1。经t检验,游离药物与纳米药物之间的t1/2β和AUC0~t均有统计学差异(P<0.01)。结论:灯盏花素制成纳米粒后明显增加了药物在动物体内的半衰期,延长了药物在体内的循环时间,且不同粒径的纳米粒对药动学有一定的影响。     

Preparation of biodegradable polycaprolactone/poly (ethylene glycol)/polycaprolactone (PCEC) nanoparticles

Biodegradable polyetherester copolymer (PCL/PEG/PCL, PCEC) was synthesized by ring-opening polymerization of epsilon-caprolactone initiated by poly(ethylene glycol) (PEG). The PCEC nanoparticles were prepared by solvent diffusion method or w/o/w double emulsion method. The obtained particles' morphology was observed on scanning electron microscopy, and the particle size distribution was determined using Malvern laser particle sizer. Bovine serum albumin was used as the model water-soluble protein drug, which was successfully encapsulated in PCEC nanoparticles, the drug release behavior was studied in detail. The hydrolytic degradation behavior of the PCEC nanoparticles was also studied.     

Preparation and evaluation of poly (caprolactone fumarate) nanoparticles containing doxorubicin HCI

Background and the purpose of the study

Biodegradable Poly(caprolactone fumarate) (PCLF) has been used as bioresorbable sutures. In this study, doxorubicin HCl (Dox) loaded PCLF nanoparticles were prepared and characterized.

Material and methods

PCLFs were synthesized by polycondensation of PCL diols (Mws of 530, 1250 and 2000) with fumaryl chloride. The degradation of PCLF in NaOH, water and phosphate buffer saline (PBS), was determined in terms of changes in Mw. Nanoparticles (NPs) were prepared by two methods. In microemulsion polymerization method, dichloromethane containing PCLF and photoinitiator were combined with the water containing surfactants and then the mixture was placed under light for crosslinking. In nanoprecipitation method, the organic solvent containing PCLF was poured into the stirring water. The effect of several variables including concentration of PCLF, polyvinyl alcohol (PVA), Dox and Trypan blue (Trb) and the Mw of PCLF and PVA on NP size and loading were evaluated.

Result

PCLF 530, 1250 and 2000 in PBS or water were not degraded over 28 days. Nanoprecipitaion method gave spherical (revealed by SEM images) stable NPs of about 225 with narrow size distribution and a zeta potential of −43 mV. The size of NP increased significantly by increase in Mw or concentration of PCLF. Although PVA was not necessary for formation of NPs, but it decreased with NP size. Dox loading and EE were 2.5–6.8% and 15–20%, respectively. Increasing the drug concentration increased the drug loading (DL) and NP size. The entrapment efficiency (EE) for Trb ranged from 1% for PCLF530 to 6% for PCLF2000. An increase in PCLF concentration resulted in an increase in EE. Dox and Trb release showed a burst followed by 80% and 78% release during 3 and 4 days respectively.

Conclusion

PCLF possessed suitable characteristics for preparation of nanoparticulate drug delivery system such as desired NP size, stability and degradation time. Although PCLF530 NPs were the smallest, but their DL were lower than PCLF1250 and 2000 NPs.     

姜黄素PLGA纳米粒的制备及制剂学性质分析

目的制备姜黄素(Curcumin,Cur)聚乳酸羟基乙酸共聚物(PLGA)纳米粒(Cur-PLGA-NPs)并对其理化性质进行考察。方法采用改良的自乳化溶剂挥发法制备纳米粒,通过正交设计,以粒径、包封率和载药量为评价指标优化处方工艺。结果制备Cur-PLGA-NPs的优化条件为PLGA 100 mg,泊洛沙姆188浓度1.0%,丙酮与乙醇体积比3∶1,有机相体积15 m L。按优化条件所制备的Cur-PLGA-NPs粒径为(120.33±2.44)nm,多分散系数为0.10±0.02,包封率为84.50%±1.13%,载药量为4.75%±0.22%。结论采用改良的自乳化溶剂挥发法成功制备了Cur-PLGA-NPs,为后续"纳米粒-脂质体系统"的研究奠定了基础,有望实现药物在肝脏的浓集。     

Preparation and characterization of paclitaxel-loaded DSPE-PEG-liquid crystalline nanoparticles (LCNPs) for improved bioavailability

Lipid-based liquid crystalline nanoparticles (LCNPs) have attracted growing interest as a new drug nanocarrier system for improving bioavailability for both hydrophilic and hydrophobic drugs. In this study, self-assembled LCNPs based on soy phosphatidyl choline and glycerol dioleate, which was known possessing low toxicity and negligible hemolysis, were prepared using poly(ethylene glycol)-grafted 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine (DSPE-PEG) as the dispersing agent. Paclitaxel (PTX) was used as a model hydrophobic drug. The particle size of the optimized DSPE-PEG-LCNPs and PTX-loaded DSPE-PEG-LCNPs were around 70nm. Crossed polarized light microscopy was used to characterize the phase behavior of liquid crystalline (LC) matrices, which showed a fan-like birefringent texture in dark background indicating the coexistence of reversed cubic and hexagonal phase in the optimized LC matrix. Transmission electron microscopy and cryo-field emission scanning electron microscopy revealed its internal water channel and "twig-like" surface morphology. PTX-loaded DSPE-PEG-LCNPs exhibited a biphasic drug sustained release pattern with a relatively fast release at the initial stage and a sustained release afterwards. PTX-loaded DSPE-PEG-LCNPs presented higher AUC (410.942±72.522μg/Lh) when compared with commercial product Taxol (212.670±41.396μg/Lh). These results indicated that DSPE-PEG-LCNPs might serve as a potential sustained release system for poorly water-soluble agents.     

利福喷丁/聚乳酸-羟基乙酸纳米粒的制备及处方工艺优化

目的： 研制负载利福喷丁的聚乳酸-羟基乙酸共聚物[poly（lactic-co-glycolic acid）,PLGA]纳米粒,并对其处方及制备工艺进行优化。方法： 采用快速膜乳化法制备利福喷丁/PLGA纳米粒。通过单因素实验考察了乳化剂浓度、PLGA浓度、油相/水相体积比、初乳制备转速、初乳制备时间、过膜压力、过膜次数对纳米粒制备的影响。在此基础上以粒径、载药率、包封率为评价指标,使用正交实验设计对纳米粒制备的处方工艺进行优化,以TOPSIS法进行多指标综合分析。然后对最优处方工艺进行验证,并对载药纳米粒的体外释药行为进行考察。结果： 经最优处方工艺制备的载药纳米粒,粒径（428±11.4）nm,粒径分布为（0.186±0.036）,包封率为（76.89±2.6）%,载药率为（10.89±1.2）%。用透视电镜观察呈均匀分布的球形。在体外药物释放实验中,药物在72 h内累计释放了78.81%。结论： 采用快速膜乳化可以简单快捷地制备均匀圆整、包封性好、具有良好缓释性能的利福喷丁/PLGA纳米粒,并为新型抗结核精准治疗的开发提供了基础。     

Preparation of poly(DL-lactide-co-glycolide) nanoparticles by modified spontaneous emulsification solvent diffusion method.

PURPOSE: The objectives of this study were to establish a new preparation method for poly(DL-lactide-co-glycolide) (PLGA) nanoparticles by modifying the spontaneous emulsification solvent diffusion (SESD) method and to elucidate the mechanism of nanoparticle formation on the basis of the phase separation principle of PLGA and poly(vinyl alcohol) (PVA) in the preparation system. METHODS: PLGA nanoparticles were prepared by the modified-SESD method using various solvent systems consisting of two water-miscible organic solvents, in which one solvent has more affinity to PLGA than to PVA and the other has more affinity to PVA than to PLGA. The yield, particle size, size distribution and PVA content of the PLGA nanoparticles were evaluated, and the phase separation behaviors of the polymers were elucidated. RESULTS: The modified-SESD method provided a good yield of PLGA nanoparticles over a wide range of composition ratios in the binary mixture of organic solvents. Several process parameters, including the fed amount of PLGA, PLGA concentration and PVA concentration were examined to achieve the optimum preparation conditions. The discrete powder of PLGA nanoparticles was obtained by freeze-drying. No change in the PVA content of PLGA nanoparticles was observed even after several times of washing treatment by ultrafiltration, suggesting a strong surface adsorption. It was found that the appropriate selections of binary solvent mixtures and polymeric concentrations in both organic and aqueous phases could provide excellent yield and favorable physical properties of PLGA nanoparticles. CONCLUSION: The proposed modified-SESD method can be used to provide PLGA nanoparticles of satisfactory quality at an acceptable yield for industrial purposes.     

Preparation of avidin-labelled gelatin nanoparticles as carriers for biotinylated peptide nucleic acid (PNA)

The possibility of preparing uniform nanoparticles consisting of proteins such as gelatin followed by covalent linkage of avidin was investigated. Gelatin nanoparticles were prepared by two step desolvation. Functional groups at the surface of the particulate system were quantified with site-specific reagents. The surface of the nanoparticles was thiolated and avidin was covalently attached to the nanoparticles via a bifunctional spacer at high levels. Biotinylated peptide nucleic acid (PNA) was effectively complexed by the avidin-conjugated nanoparticles. Avidin-conjugated protein nanoparticles should prove as potential carrier system for biotinylated drug derivatives in antisense therapy.     

雷公藤甲素聚乳酸纳米粒的制备及毒性

目的探索可生物降解聚乳酸［poly(D,L-lactic acid)，PLA］纳米粒口服给药后降低毒性的可能性。方法 采用改良的自乳化溶剂蒸发法制备雷公藤甲素聚乳酸纳米粒；透射电子显微镜(TEM)观察纳米粒的形态；动态激光粒度分析仪测定其平均粒径大小和分布；采用反相高效液相色谱法(RP-HPLC)测定纳米粒的包封率及载药量；X-射线粉末衍射(X-ray)初步研究纳米粒中药物的物理状态；考察雷公藤甲素的体外释放特性；评价口服给予纳米粒对大鼠的降毒性作用。 结果确定适合处方的工艺为：水相-有机相为40∶15(v/v)，表面活性剂浓度为1% (w/v)，药物在有机相中的浓度为0.3% (w/w)，TP-PLA为1∶15 (w/w)。处方条件下制备的纳米粒平均粒径为149.7 nm，多分散指数为0.088，平均包封率及载药量分别为74.27% 和1.36%；雷公藤甲素的体外释放分为两相；纳米粒非常显著降低肝的毒性(P<0.01)，显著降低肾的毒性(P<0.05)。结论聚乳酸纳米粒可能成为雷公藤甲素口服给药的新型载体。