Preparation and cytotoxicity of poly (DL-lactide-co-glycolide) microspheres encapsulating 2-methoxyestradiol

Injectable implant would be a potential way for 2-methoxyestradiol (2-ME) for the therapy of breast cancer because of its water-insolubility, short half-life, and low oral bioavailability. So 2-ME microspheres based on poly (DL-lactide-co-glycolide) were prepared by emulsion solvent extraction method and characterized for morphology, particle size, drug physical state, entrapment efficiency and drug release in vitro and in vivo. Their cytotoxicity on MCF-7 cells was evaluated by sulforhodamine B (SRB) method. The 2-ME was successfully entrapped in the interior of microspheres with particle size of 55.44 ± 12.21 μm and could exist in an amorphism. In vitro and in vivo release of 2-ME from the microspheres occurred in a Ritger-Peppas and zero-order manner with a slow release over 46 days, respectively, and their better correlation was found. The 2-ME even with very low concentration in the microspheres could efficiently inhibit the growth of MCF-7 cells compared to the equivalent amount of drug in free solution, which indicated that the release rate from the microspheres and local water-solubility of 2-ME could maintain effective drug concentration in target site. The above results indicated that the microspheres prepared could not only control prolonged release of 2-ME in vitro and in vivo but also maintain effective drug concentration in target site. So 2-ME microspheres are acceptable for controlled release devices for effective treatment of breast cancer.     

Assessment of biodegradability of polymeric microspheresin vivo: Poly (DL-lactic acid), poly (L-lactic acid) and poly (DL-lactide-co-glycolide) microspheres

To confirm a new evaluation technique for biodegradability of biopolymer microspheres in vivo condition, magnetic microsphere system was adopted for tracing the microspheres injected and lodged in mice. Microspheres of poly(DL-lactic acid), poly(L-lactic acid) and poly (DL-lactide-coglycolide)(PLGA) were prepared by solvent-extraction method and their organ distribution and biodegradation in mice was examined. Magnetic microspheres lodged in mice organs were recollected from the homogenates of mice organs with a constant flow magnetic separation apparatus. Recollected microspheres were observed by scanning electron microscopy and also were assayed for their magnetite content by atomic absorption spectrophotometry to evaluate the biodegradability of polymeric microspheres. This method seems to be practical and simple to estimate the biodegradability of biopolymers over the conventional methods.     

全反式维甲酸前体脂质体的制备及体外评价

目的:制备维甲酸前体脂质体,并对其体外性质进行考察。方法:采用乙醇注入结合冷冻干燥法制备前体脂质体;微柱离心-高效液相色谱法测定脂质体的包封率;并进一步对其粒径、Zeta电位、血浆释放率及乙醇残留量进行测定。结果:所制备的前体脂质体包封率为95.2%,Zeta电位为-(28.4±17.5)mV,粒径为(170±29)nm,乙醇残留量为3.98%。结论:乙醇注入结合冷冻干燥法制备的维甲酸前体脂质体包封率高,粒径均匀,稳定性好。     

Formulation and in-vitro characterization of retinoic acid loaded poly (lactic-co-glycolic acid) microspheres

Poly (lactic-co-glycolic acid) (PLGA) microspheres containing all-trans retinoic acid (atRA) were prepared by emulsion/solvent evaporation technique. PLGA (50:50) with inherent viscosities of 0.17 and 0.39 dL g(-1) was used. Polyvinyl alcohol (PVA) or PVA and sodium oleate (SO) combinations (4:1) were used to stabilize the emulsions. The effect of polymer viscosity, emulsifier type and concentration on the in vitro release of atRA from the microspheres was investigated. The stability of the microparticles was also tested at the temperatures of 4, 25 and 40 degrees C. The particle size ranged between 1-2 microm. Microspheres were smooth and spherical in shape, as determined by scanning electron microscope (SEM) photographs. The yield of microspheres ranged from 50-75% and the encapsulation efficiency was determined between 45-75%. In vitro release studies showed that atRA release from microspheres lasted for 11 days.     

Polyion complex micelles composed of all-trans retinoic acid and poly (ethylene glycol)-grafted-chitosan

The goal of this study is to develop novel types of polyion complex micelles for the drug delivery to brain tumor. Methoxy poly(ethylene glycol) (mPEG)-grafted chitosan (CP) was synthesized in order to make polymeric micelles encapsulating all-trans retinoic acid (ATRA) based on polyion complex formation. Polyion complex micelles were found to have spherical shapes with sizes of about 50 approximately 200 nm. The loading efficiency of micelle was higher than 80% (w/w) for all formulations. 1H nuclear magnetic resonance (NMR) spectra confirmed the formation of polymeric micelles. The CP graft copolymer and ATRA have distinguishing peaks in their 1H NMR spectra. The specific peaks of ATRA disappeared in D2O or DMSO while it appeared at mixtures of D2O/DMSO, indicating that ATRA and chitosan formed ion complex inner-core. In the cell cytotoxicity study using U87MG cells in vitro, polyion complex micelles showed similar cytotoxicity to that of free ATRA. A migration test was performed to investigate the inhibition of tumor cell invasion in vitro. The results suggested that the polyion complex micelles was more effective at inhibiting tumor cell migration than free ATRA.     

Characterization of FITC-albumin encapsulated poly(DL-lactide-co-glycolide) microspheres and its release characteristics

In this study, the fluorescein isothiocyanate (FITC)-albumin distribution, the amount of FITC-albumin release and the degradation of poly (DL-lactic-co-glycolic acid) (PLGA) microspheres were evaluated. The higher the FITC-albumin concentration in the W1 phase, the higher the protein concentration and loading efficiency in the microspheres, the larger the particle size of microspheres. Confocal laser scanning microscopy (CLSM) showed that the FITC-albumin was distributed uniformly in the microspheres with a low concentration while it was aggregated at higher concentrations. The high protein concentration in the microspheres showed significantly large initial release (above 70% w/w) during the 10 days. A higher PVA concentration in the W2 phase resulted in a smaller particle size, higher protein contents and loading efficiency and decreased release rate. After a 10 day incubation period, the microspheres with a low FITC-albumin concentration had an almost spherical morphology, while microspheres with a high FITC-albumin concentration had a more distorted morphology, suggesting that rapid morphological changes and microspheres ruptures can be induced by an initial burst release.     

紫杉醇PLGA微球制备及体外性质评估

目的：以乳酸-羟基乙酸共聚物（PLGA）为材料,制备用于肿瘤内注射的紫杉醇PLGA长效缓释微球.方法：采用改良溶剂蒸发法制备,对微球的体外性质以及不同剂量（10,15,25 kGy）60Coγ射线对微球性质的影响进行考察.结果：制得的微球形态圆整,载药量、包封率、平均粒径和跨距分别为1.53%,97.29%,42.72μm和0.95.药物体外释放30 d累计释放达到56.19%,体外降解30 d后微球失去完整结构,表面粗糙.3个剂量60Coγ射线的灭菌效果均良好,且对微球的体外性质均无明显影响.微球中二氯甲烷的残留量低于药典规定的限度.结论：紫舷杉醇PLGA微球满足缓释长效的要求,对恶性肿瘤的间质化疗具有一定前景.     

Optimization of the encapsulation and release of beta-lactoglobulin entrapped poly(DL-lactide-co-glycolide) microspheres.

The goal of the present paper was to optimize the encapsulation of beta-lactoglobulin (BLG) within poly(lactide-co-glycolide) (PLGA) microparticles prepared by the multiple emulsion solvent evaporation method. The role of the pH of the external phase and the introduction of the surfactant Tween 20, in the modulation of the entrapment and release of BLG from microparticles, was studied. Reducing the solubility of BLG by decreasing the pH of the external phase to a value close to the pI of BLG resulted in a better encapsulation with, however, a larger burst release effect. By contrast, Tween 20 was shown to increase the encapsulation efficiency of BLG and reduce considerably the burst release effect. In fact, Tween 20 was shown to be responsible for removing the BLG molecules that were adsorbed on the particle surface. In addition, Tween 20 reduced the number of aqueous channels between the internal aqueous droplets as well as those communicating with the external medium. Thus, the more dense structure of BLG microspheres could explain the decrease in the burst release. These results constitute a step ahead in the improvement of an existing technology in controlling protein encapsulation and delivery from microspheres prepared by the multiple emulsion solvent evaporation method.     

Preparation and characterization of chitosan-treated alginate microparticles incorporating all-trans retinoic acid

Chitosan treated alginate microparticles were prepared with the purpose of incorporating all-trans retinoic acid (ATRA) using an inexpensive, simple and fast method, enhancing dermal localization and sustaining the release of ATRA into the skin. Microparticles characterization, drug-polymer interaction, release profile and in vitro skin retention were investigated. Microparticles presented spherical shape and drug loading capacity of 47%. The drug content of these microparticles was affected by ATRA concentration and by the solvent used and it was more weakly affected by chitosan concentration. The release of ATRA was also affected by chitosan concentration. Microparticles prepared with 0.4% chitosan (w/w) resulted in drug release with a more sustained profile. The results of in vitro retention studies showed that chitosan treated alginate microparticles decreased the drug retention in the stratum corneum (SC), where occur the skin irritation, but maintained the ATRA concentration in the deeper skin layers, where occur the pathologies treated with ATRA. Then, the microparticles developed in this work can be a good candidate to improve the topical therapy with retinoid.     

Preparation and characterization of chitosan-treated alginate microparticles incorporating all-trans retinoic acid

Chitosan treated alginate microparticles were prepared with the purpose of incorporating all-trans retinoic acid (ATRA) using an inexpensive, simple and fast method, enhancing dermal localization and sustaining the release of ATRA into the skin. Microparticles characterization, drug–polymer interaction, release profile and in vitro skin retention were investigated. Microparticles presented spherical shape and drug loading capacity of 47%. The drug content of these microparticles was affected by ATRA concentration and by the solvent used and it was more weakly affected by chitosan concentration. The release of ATRA was also affected by chitosan concentration. Microparticles prepared with 0.4% chitosan (w/w) resulted in drug release with a more sustained profile. The results of in vitro retention studies showed that chitosan treated alginate microparticles decreased the drug retention in the stratum corneum (SC), where occur the skin irritation, but maintained the ATRA concentration in the deeper skin layers, where occur the pathologies treated with ATRA. Then, the microparticles developed in this work can be a good candidate to improve the topical therapy with retinoid.     

紫杉醇PLGA微球制备及工艺优化

目的：以乳酸-羟基醋酸共聚物[-poly（lactic-co-glycolic acid），PLGA3为材料，制备用于肿瘤内注射的紫杉醇PLGA长效缓释微球。方法：采用改良溶剂蒸发法制备，使用星点设计一效应面法优化处方，并对药物体外释放进行评估。结果：以投药比（％）和聚乙烯醇（PVA）浓度为自变量，载药量、包封率、粒径和跨距为因变量进行多元线形回归和二次多项式拟合，结果表明二次多项式拟合的效果较好，较优工艺条件为投药比1．6％、PVA浓度2％。照优化工艺制得的微球形态圆整，载药量、包封率、平均粒径和跨距分别为1．53％，97．29％，42．72胛和0．95，与方程预测值的偏差分别为4．08％，1．91％，6．48％和-4．9HD％。微球体外释放30d累计释放率达56．19％。结论：本实验所得的紫杉醇PLGA微球满足缓释长效的要求，所建立的模型预测性良好。     

超氧化物歧化酶乳酸-羟乙酸共聚物微球的制备及其性质

利用复乳溶剂挥发法制备了超氧化物歧化酶（SOD）的乳酸－羟乙酸共聚物（PLGA）微球，考察了各工艺因素对微球粒径、包封率等的影响，通过扫描电子显微镜（SEM）、差示扫描量热分析（DSC）初步研究了其性质，结果表明，通过调整内水相的体积及浓度，分散相体积及PH值，可得到较高包封率，粒径在20－30μm，形态圆整，表面多孔的SOD微球，DSC表明SOD被有效地包入了PLGA微球中。     

Preparation, characterization, cytotoxicity and transfection efficiency of poly(DL-lactide-co-glycolide) and poly(DL-lactic acid) cationic nanoparticles for controlled delivery of plasmid DNA

The objective of this study was to investigate the effect of formulation parameters (i.e. polymer molecular weight and homogenization speed) on various physicochemical and biological properties of cationic nanoparticles. Cationic nanoparticles were prepared using different molecular weights of poly(DL-lactide-co-glycolide) (PLGA) and poly(DL-lactic acid) (PLA) by double emulsion solvent evaporation at two different homogenization speeds, and were characterized in terms of size, surface charge, morphology, loading efficiency, plasmid release, plasmid integrity, cytotoxicity, and transfection efficiency. Cationic surfactant, cetyltrimethylammonium bromide (CTAB), was used to provide positive charge on the surface of nanoparticles. Reporter plasmid gWIZ Beta-gal was loaded on the surface of nanoparticles by incubation. Use of higher homogenization speed and lower molecular weight polymer led to a decrease in mean particle size, increase in zeta potential, increase in plasmid loading efficiency, and a decrease in burst release. The nanoparticles displayed good morphology as evident from scanning electron micrographs. In vitro cytotoxicity study by MTT assay showed a low toxicity. Structural integrity of the pDNA released from nanoparticles was maintained. Transfecting human embryonic kidney (HEK293) cells with nanoparticles prepared from low molecular weight PLGA and PLA resulted in an increased expression of beta-galactosidase as compared to those prepared from high molecular weight polymer. Our results demonstrate that the PLGA and PLA cationic nanoparticles can be used to achieve prolonged release of pDNA, and the plasmid release rate and transfection efficiency are dependent on the formulation variables.     

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 neurotensin analogue-containing poly(dl-lactic acid) microspheres formed by oil-in-water solvent evaporation.

This report describes the preparation of injectable microspheres containing a neurotensin analogue (NA), which is a hexapeptide with neurotensin activity. NA, a hydrophilic drug, was successfully entrapped into poly(dl-lactic acid) microspheres prepared by a novel oil-in-water solvent evaporation method. The preparation method was investigated with regard to the partition of NA into the oily phase and the rapid phase separation of the polymer. Successful entrapment was achieved with the following conditions: (1) an alkaline water phase, (2) addition of fatty acid salt in the oily phase, and (3) addition of a water-miscible solvent in the oily phase. Under these conditions, NA was completely entrapped into the microspheres at poly(dl-lactic acid):NA molar ratios of greater than 3.     

全反式维A酸醇质体的处方优化及评价

目的：制备全反式维A酸(all-trans retinoic acid,ATRA)醇质体并对其稳定性及体外经皮渗透等进行考察。方法：采用注入法制备ATRA醇质体,通过正交设计优化制备工艺;同时测定其Zeta电位及粒径;以改进的Franz扩散池法,进行体外小鼠经皮渗透实验,测定药物累积渗透量及透皮速率。结果：优选处方组成为20%乙醇（w/w）,4%磷脂（w/w）,磷脂[DK]∶胆固醇（w/w）2[DK]∶1,磷脂[DK]∶维A酸（w/w）10[DK]∶1,检测平均粒径为237.3nm,Zeta电位为-36.31mV;ATRA醇质体放置1,10,20d后,7h累积透皮量分别为（210.6±1.7）,（196.2±3.8）和（181.1±4.2）μg[DK]?cm-2,而水醇混合物累积透皮量仅为（120.4±5.4）μg[DK]?cm-2。结论：ATRA醇质体制备工艺简单可行,所得传递体粒径较小且均匀,室温放置较稳定,透皮效果较好并可以促进维A酸经皮转运。     

Preparation and characterization of poly(lactic-co-glycolic acid) microspheres loaded with a labile antiparkinson prodrug

L-dopa-α-lipoic acid (LD-LA) is a new multifunctional prodrug for the treatment of Parkinson's disease. In human plasma, LD-LA catechol esters and amide bonds are chemically and enzymatically cleaved, respectively, resulting in a half-life time of about fifty minutes. In the present work, the unstable LD-LA was entrapped into biodegradable poly(lactic-co-glycolic acid) (PLGA) microspheres designed as depot systems to protect this prodrug against degradation and to obtain a sustained release of the intact compound. The microspheres were prepared by an oil-in-water emulsion/solvent evaporation technique and the effect of formulation and processing parameters (polymer concentration in the organic solvent, volumes ratio of the phases, rate of the organic solvent evaporation) on microspheres characteristics (size, loading, morphology, release) was investigated. Also emphasis was given on the stability of the drug before and after release as well as on the underlying mass transport mechanisms controlling LD-LA release. Interestingly, when encapsulated in appropriate conditions into PLGA microspheres, the labile prodrug was stabilized and released via Fickian diffusion up to more than one week.     

乳酸-羟基乙酸共聚物微球的研究进展

通过整理和归纳国内外文献,介绍乳酸-羟基乙酸(PLGA)载药微球的制备方法和作为药物载体的应用。     

Granulomatous tubulointerstitial nephritis induced by all-trans retinoic acid

We report the first case of granulomatous tubulointerstitial nephritis induced by all-trans retinoic acid (ATRA) in a patient with acute promyelocytic leukemia (APL). Acute renal failure during treatment with ATRA has been previously reported as a part of an ATRA syndrome or a thrombotic complication of a hypercoagulable state. This case indicates an alternative mechanism of acute renal failure occurring during ATRA therapy.