Inactive Vibrio cholerae whole-cell vaccine-loaded biodegradable microparticles: in vitro release and oral vaccination

An approach is proposed using Vibrio cholerae (VC)-loaded microparticles as oral vaccine delivery systems for improved vaccine bioavailability and increased therapeutic efficacy. The VC-loaded microparticles were prepared with 50:50 poly(DL-lactide-co-glycolide) (PLG), 75:25 poly(DL-lactide-co-glycolide) and poly(lactide acid) (PLA)/PEG blend copolymers by the solvent evaporation method. VC was successfully entrapped in three types of microparticles with loading efficiencies and loading levels as follows: 50:50 PLG systems: 97.8% and 55.4 ± 6.9 µg/mg; 75:25 PLG systems: 89.2% and 46.5 ± 4.4?µg/mg; PLA/PEG-blended systems: 82.6% and 53.7 ± 5.8?µg/mg. The different distributions of VC in the core region and on the surface were as follows: 50:50 PLG systems 25.7 ± 1.9 and 6.2 ± 0.9?µg/mg; 75:25 PLG systems: 25.8 ± 2.2 and 3.6 ± 0.4?µg/mg; PLA/PEG-blended systems: 32.4 ± 2.1 and 5.2 ± 1.0?µg/mg, respectively. In vitro active release of VC was affected mainly by matrix type and VC-loaded location in microparticles. The therapeutic immunogenic potential of VC loaded with 50:50 PLG, 75:25 PLG and PLA/PEG-blended microparticles was evaluated in adult mice by oral immunization. Significantly higher antibody responses and serum immunoglobin Ig G, IgA and IgM responses were obtained when sera from both VC-loaded 75:25 PLG and PLA/PEG-blended microparticles immunized mice were titrated against VC. The most immunogenicity in evoking serum IgG, IgA and IgM responses was immunized by VC-loaded PLA/PEG-blended microparticles, and with VC challenge in mice, the survival rate (91.7%).     

Cellulose acetate phthalate microparticles containing Vibrio cholerae: steps toward an oral cholera vaccine

Abstract

Oral cholera vaccine (OCV) has been recommended in some endemic areas and epidemic situations since 1999. Although safe and effective vaccines are currently on the market, the burden of transport and storage remains an issue. Herein, we report an approach to develop an alternative OCV in the form of a gastro-resistant powder. Heat-killed Vibrio cholerae (VC) was encapsulated with a spray-drying technique at different temperatures. Cellulose acetate phthalate (Aquacoat® CPD) was chosen as the core polymer and the addition of alginate was studied. The microparticles (MPs) produced were characterized by surface morphology, particle size, drug loading, antigenicity and gastro resistance. The MPs obtained were 6?µm in size and had appropriate drug content, ranging from 8.16 to 8.64%. Furthermore, antigenicity was maintained, never dropping below 85%, and enteric properties were achieved for all the formulations. Next, an in vivo study was carried out with Aquacoat® CPD MP prepared at 80?°C with and without alginate. Two different doses were assayed, 30 and 60?mg, and compared to the VC suspension. The evoked immune responses showed that alginate containing MPs, especially at the 30?mg dose, displayed values that were very similar to those of VC. In conclusion, spray-dried alginate VC MPs seem to be a promising step toward a powder-form cholera vaccination.     

Fluphenazine release from biodegradable microparticles: Characterization and modelling of release

Abstract

The release of actives encapsulated in biodegradable poly-lactide-co-glycolide (PLGA)-based microparticles may be diffusion controlled, dependent on polymer degradation, or may occur by a combination of drug diffusion and polymer degradation. This report applies a model, describing combined diffusional and polymer degradation-assisted drug release, to quantify the release of fluphenazine HCl (F-HCl) from PLGA microspheres. Parameters for the release process showed that both the initial drug release phase and the polymer controlled drug release phase were dependent on the F-HCl loading of the microspheres. The percentage drug released in the burst phase and the length of the lag phase were dependent on F-HCl loading. In the degradation controlled release phase, drug release was faster the higher the loading, as shown by the decrease in t_max from 27 to 10 days, as F-HCl loadings increased from 4.2 to 16.6%w/w. The presence of F-HCl was found to catalyse the degradation of PLGA polymer during particle manufacture and during dissolution. When compared to drug free microspheres, F-HCl accelerated PLGA degradation as shown by the ～5-fold increase in both PLGA degradation rate constant (k) and reduction in t_max.     

Fluphenazine release from biodegradable microparticles: characterization and modelling of release

The release of actives encapsulated in biodegradable poly-lactide-co-glycolide (PLGA)-based microparticles may be diffusion controlled, dependent on polymer degradation, or may occur by a combination of drug diffusion and polymer degradation. This report applies a model, describing combined diffusional and polymer degradation-assisted drug release, to quantify the release of fluphenazine HCl (F-HCl) from PLGA microspheres. Parameters for the release process showed that both the initial drug release phase and the polymer controlled drug release phase were dependent on the F-HCl loading of the microspheres. The percentage drug released in the burst phase and the length of the lag phase were dependent on F-HCl loading. In the degradation controlled release phase, drug release was faster the higher the loading, as shown by the decrease in t(max) from 27 to 10 days, as F-HCl loadings increased from 4.2 to 16.6%w/w. The presence of F-HCl was found to catalyse the degradation of PLGA polymer during particle manufacture and during dissolution. When compared to drug free microspheres, F-HCl accelerated PLGA degradation as shown by the approximately 5-fold increase in both PLGA degradation rate constant (k) and reduction in t(max).     

口蹄疫DNA疫苗海藻酸钠微球的制备及体外释放的研究

目的：研制DNA疫苗海藻酸钠微球，并对其体外释药特性进行考察。方法：以口蹄疫DNA疫苗为DNA疫苗的模型药物，采用喷雾干燥一离子交联法制备DNA疫苗海藻酸钠微球；考察粒径大小、外观、载药量等理化特性；考察微球的体外释药特性及其影响因素。结果：微球球形圆整，分散性好，平均粒径为11．9μm，载药量为5％，产率为53．2％。微球的体外释放速率受载药量影响较小，而壳聚糖的交联固化度增高，微球的体外释放速率变慢。结论：以生物降解材料海藻酸钠、壳聚糖，用喷雾干燥法制备DNA疫苗微球，不需要超声和有机溶剂，因而有利于DNA疫苗结构和功能的稳定性；工艺简便，易于工业化生产。     

口蹄疫DNA疫苗聚氰基丙烯酸正丁酯纳米粒制备工艺及体外释药规律研究

目的优选制备口蹄疫DNA疫苗聚氰基丙烯酸正丁酯纳米粒(DNA-PBCA-NP)的工艺条件.方法以聚氰基丙烯酸正丁酯为载体,用乳液聚合二步法制备不同配方的DNA-PBCA-NP,采用正交设计实验法,以包封率为指标,综合分析实验结果确定工艺条件.结果优选出的制备工艺条件为:在pH值为3的条件下制备聚氰基丙烯酸正丁酯空白纳米粒,以2 mL·min-1的速度将1份口蹄疫DNA疫苗溶液滴入到5份空白纳米球乳液中,继续搅拌 8 h.运用优化的工艺条件制备的纳米粒平均粒径为(78.12±16.5)nm,分布范围为30～150 nm,平均包封率为51.25%±3.92%.体外缓释试验表明释药符合双相动力学规律.结论所确定的制备口蹄疫DNA疫苗聚氰基丙烯酸正丁酯纳米粒的工艺条件稳定、可行.     

Preparation and in vitro release of zein microparticles loaded with prednisolone for oral delivery

Zein has been proposed as a polymer for targeted-drug delivery via the oral route. Zein microparticles were loaded with prednisolone and evaluated as an oral delivery system. Microparticles were formulated using phase separation. Starting quantities of zein and prednisolone, along with the agitation method and temperature were found to significantly impact drug loading and loading efficiency. Vortex mixing produced the highest drug loading and loading efficiency. Drug release was measured in simulated conditions of the stomach and small intestine using the microparticles made with the method that best improved drug loading. In simulated stomach and small intestine conditions, prednisolone release reached almost 70% over 3 and 4?h, respectively. While a clinically relevant dose may be delivered using c. 100?mg of zein microparticles, prednisolone release from the microparticles indicates that they may not be suited as a controlled- or targeted-delivery system.     

Preparation of prolonged release clarithromycin microparticles for oral use and theirin vitro evaluation

Prolonged release microparticles of clarithromycin (CL) were prepared using Eudragit RL 100 and RS 100 by spray-drying and casting-drying techniques. For the characterization of those microparticles, preparation yield, particle size distribution, X-ray diffraction, thermal behavior, active agent content and in vitro dissolution from the microparticles were performed. HPLC was used for the assay of clarithromycin and the assay method was validated. All the formulations obtained showed prolonged release when compared to pure clarithromycin. Microparticles prepared by spray-drying method had a slower release compared to those of casting-drying method. Spray-drying method seems to be a more suitable method to prepare microparticles for prolongation in release.     

Porcine insulin biodegradable polyester microspheres: stability and in vitro release characteristics

The stability of porcine insulin in biodegradable polymers, i.e., poly(DL-lactide-co-glycolide) 50:50 (50:50 DL-PLGA) and poly(L-lactide) (L-PLA) was investigated. Insulin encapsulated microspheres were fabricated from both polymers using double-emulsion-solvent evaporation and emulsion-solvent evaporation techniques and subjected to accelerated stability studies at 40 degrees C and 75% relative humidity. Porcine insulin was found to degrade in all microsphere formulations with an average of < 50% of the initial loading amount remaining intact at the end of 4 weeks. The two major degradation products observed in these formulations were determined to be A-21 desamido insulin and covalent insulin dimer with trace amounts of high molecular weight transformation products. In vitro release studies in phosphate buffered saline at 37 degrees C resulted in very slow and incomplete (< 30% in 30 days) release kinetics for all microsphere formulations. Extraction and analyses of the unreleased insulin within the microspheres revealed that an average of approximately 11% of the encapsulated insulin remained intact. The degradation products observed consisted of approximately 15% of three distinct deamidated hydrolysis products including A-21 desamido insulin, approximately 22% covalent insulin dimer, and trace amounts of high molecular weight transformation products. The degradation of porcine insulin within biodegradable polyester microspheres during stability and release studies can be attributed to the gradual decrease in the pH within the microspheres due to progressive polymer hydrolysis resulting in the production of DL-lactic and glycolic acids. The encapsulation of an acid-base indicator, bromophenol blue, in 50:50 PLGA microspheres (as a probe to estimate pH within the microspheres during accelerated stability studies) indicated that the pH decreased to approximately 3.8 after 3 weeks.     

Encapsulation and release of the hypnotic agent zolpidem from biodegradable polymer microparticles containing hydroxypropyl-beta-cyclodextrin

The goal of this study was to design a prolonged release system of the hypnotic agent zolpidem (ZP) useful for the treatment of insomnia. In this work, ZP alone or in the presence of HP-beta-CD was encapsulated in microparticles constituted by poly(DL-lactide) (PDLLA) and poly(DL-lactide-co-glycolide) (PLGA) and the drug release from these systems was evaluated. ZP alone-loaded microparticles were prepared by the classical O/W emulsion-solvent evaporation method. Conversely, ZP/HP-beta-CD containing microparticles were prepared by the W/O/W emulsion-solvent evaporation method following two different procedures (i.e. A and B). Following procedure A, the previously produced ZP/HP-beta-CD solid complex was added to the water phase of primary emulsion. In the procedure B, HP-beta-CD was added to the aqueous phase and ZP to the organic phase. The resulting microparticles were characterized about morphology, size, encapsulation efficiency and release rates. FT-IR, X-ray, and DSC results suggest the drug is in an essentially amorphous state within the microparticles. The release profiles of ZP from microparticles were in general biphasic, being characterized by an initial burst effect and a subsequent slow ZP release. It resulted that co-encapsulating ZP with or without HP-beta-CD in PDLLA and PLGA the drug release from the corresponding microparticles was protracted. Moreover, in a preliminary pharmacological screening, the ataxic activity in rats was investigated and it was found that intragastric administration of the ZP/HP-beta-CD/PLGA microparticles prepared according to procedure B produced the same ataxic induction time as the one induced by the currently used formulation Stilnox. Interestingly moreover, there was a longer ataxic lasting and a lower intensity of ataxia produced by the ZP/HP-beta-CD/PLGA-B-formulation already after 60 min following the administration. However, a need for further pharmacokinetic and pharmacodynamic studies resulted to fully evaluate the utility of this last formulation for the sustained delivery of ZP.     

Indocyanine green-loaded biodegradable nanoparticles: preparation, physicochemical characterization and in vitro release

PURPOSE: The objective of this study is to develop indocyanine green (ICG)-loaded biodegradable nanoparticles by using biodegradable polymer, poly(DL-lactic-co-glycolic acid) (PLGA). METHOD: PLGA nanoparticles entrapping ICG were prepared by a modified spontaneous emulsification solvent diffusion method. To optimize the nanoparticle formulation, the influence of formulation parameters such as types of ICG, amount of ICG and the polymer were investigated. The ICG entrapment in nanoparticles, nanoparticle size and zeta potential were determined. The surface characterization was performed by atomic force microscopy (AFM) and the release of ICG from nanoparticles was determined. RESULTS: All PLGA nanoparticle formulations were found to have the mean diameter within the range of 300-410 nm with polydispersity index (PI) within the range of 0.01-0.06. Indocyanine green showed more efficient entrapment as compared to indocyanine green sodium iodide salt. All indocyanine green-loaded nanoparticle formulations were found to have almost similar ICG content of nanoparticles and showed increase in ICG entrapment with increase in the amount of polymer. The ICG entrapment reached 74% when ICG: PLGA weight ratio in the formulation reached 1:800. AFM images indicated that the nanoparticles were almost spherical in shape and had numerous pores on their surfaces. The release pattern consisted of two phases, with initial exponential phase releasing about 78% of ICG (within 8 h) followed by a slow phase releasing about 2% of ICG (within next 16 h). CONCLUSIONS: ICG-loaded PLGA nanoparticles were prepared and the formulation was optimized. The increase in amount of polymer in formulation leads to higher ICG entrapment. Nanoparticles formed were spherical and had porous surfaces and exhibited the characteristic release pattern of a monolithic matrix based system.     

Studies on in vitro release behaviour of indomethacin-loaded polystyrene microparticles

Indomethacin-loaded polystyrene microparticles were prepared by emulsion-solvent evaporation method from an aqueous system. The effect of different parameters like concentration of aqueous phase emulsion stabilizer, volume of the organic disperse phase and initial drug loading on drug content and release of drug were investigated. Keeping the drug-polymer ratio constant, variation in the concentration of emulsion stabilizer and volume of the organic disperse phase did not produce any significant change either in the actual drug content or in the drug release. The initial drug loading, however, greatly influenced the drug release which, as revealed by different analyses, was due to the presence of drug in different physical forms in the microparticles. Physical characterization using thin layer chromatography and infrared spectroscopy apparently revealed the absence of drug degradation and sizeable interaction between the drug and the polymer. Regardless of lack of interaction, thermodynamic properties such as solubility of the drug in the polymer and fraction of the drug present in crystalline form were determined by using differential scanning calorimetry and was further substantiated with scanning electron micrography and X-ray diffraction analysis.     

Lectin-coated PLGA microparticles: Thermoresponsive release and in vitro evidence for enhanced cell interaction

PLGA-microparticles with 4.7μm in diameter were prepared by the double emulsion technique and loaded with 1.7μg fluorescein/mg PLGA mimicking a hydrophilic API. In an effort to further elucidate the release and bioadhesive characteristics of lectin-grafted formulations in vitro, the particles were coated with wheat germ agglutinin. The microparticles exhibited thermo-responsive release since no free fluorescein was detected at 4°C or room temperature. At body temperature, however, more than 80% of the payload was released within 48h. The adhesion of lectin-grafted particles to Caco-2 monolayers, which were applied as a model for the human intestinal epithelium, exceeded that of plain ones 1.5-fold as also observed by fluorescence microscopy. Furthermore, the amount of model drug bound and taken up into the cells was 5.8-fold higher after incubation for 4h at 37°C as compared to fluorescein in solution. According to fluorescence imaging a considerable amount of the total fluorescein payload was accumulated intracellularily after incubation for 5h at 37°C. These findings not only confirm the utility of bioadhesives of the second generation for improved absorption of low molecular weight hydrophilic compounds but also indicate storage stability of such suspensions at 4°C and room temperature without any premature loss of API.     

In vitro and in vivo evaluation of sustained release chitosan-coated ketoprofen microparticles

Simple ketoprofen microspheres (MS) were prepared by the dry-in-oil method using ethylcellulose (EC) as a matrix polymer. Further, the microspheres modified by addition of polyethylene glycol (PEG) and hydroxypropyl cellulose (HPC), called MS-P and MS-H, respectively, were prepared. The in vitro release from MS, MS-P and MS-H was examined in the JP XIII second fluid, pH 6.8, at 37 degrees C and 60 rpm. Chitosan-coated ketoprofen microparticles (Chi-MP) were prepared by the precipitation of droplets of chitosan solution containing MS, and their adhesion to the rat small intestinal mucosa was tested. The plasma concentrations after duodenal administration were investigated for ketoprofen powder suspension, MS and Chi-MP. The particle size was raised with the increase in amount of ketoprofen added. The drug content and addition of PEG or HPC affected the drug release rate. The microspheres with moderate drug content, prepared by addition of modest amount of PEG, exhibited better gradual drug release. Chi-MP showed a good mucoadhesion. The maximum plasma concentration of ketoprofen for Chi-MP was less than one-third of that for ketoprofen powder suspension. Chi-MP tended to show the higher and steadier plasma level than MS.     

Studies on the in vitro release characteristics of ibuprofen-loaded polystyrene microparticles

Ibuprofen-loaded polystyrene microparticles were prepared by the emulsion-solvent evaporation process from an aqueous system. The effects of different parameters on the drug content and on the release of the drug from the microparticles were investigated. The drug content, in all the formulations, was less than the theoretical drug loading. The lower drug content was due to drug partitioning to the external aqueous phase during formulation. Statistical analysis revealed that the variation in the concentrations of the emulsion stabilizer and the organic disperse phase volume did not significantly alter the release of the drug. Although an increase in drug loading increased drug release from the microparticles, a biphasic linear relationship was observed between the time required for 50% drug release and the drug loading. The effect of size of the microparticles on drug release was more important for the low drug-loaded microparticles than that for the high drug-loaded microparticles. Such release behaviour from the microparticles was explained on the basis of the morphological structure of the microparticles.     

Studies on the in vitro release characteristics of ibuprofenloaded polystyrene microparticles

Ibuprofen-loaded polystyrene microparticles were prepared by the emulsionsolvent evaporation process from an aqueous system. The effects of different parameters on the drug content and on the release of the drug from the microparticles were investigated. The drug content, in all the formulations, was less than the theoretical drug loading. The lower drug content was due to drug partitioning to the external aqueous phase during formulation. Statistical analysis revealed that the variation in the concentrations of the emulsion stabilizer and the organic disperse phase volume did not significantly alter the release of the drug. Although an increase in drug loading increased drug release from the microparticles, a biphasic linear relationship was observed between the time required for 50% drug release and the drug loading. The effect of size of the microparticles on drug release was more important for the low drug-loaded microparticles than that for the high drug-loaded microparticles. Such release behaviour from the microparticles was explained on the basis of the morphological structure of the microparticles.     

beta-Estradiol biodegradable microspheres: effect of formulation parameters on encapsulation efficiency and in vitro release

The purpose of this work was to study the effect of organic solvent and surfactant type on the in vitro release behavior in general and on the burst release in particular of beta-estradiol from PLA/PLGA microspheres. Also the effect of these variables on the encapsulation efficiency was investigated. The microspheres were prepared by solvent evaporation technique using dichloromethane (DCM), ethyl acetate (EtAc), tetrahydrofuran (THF), chloroform (CHCl3) or acetone (AC) as organic solvent and polyvinyl alcohol (PVA), Tween 80, sodium lauryl sulfate (SLS) or benzalkonium chloride (BKCI) as surfactant. The obtained microspheres were tested for encapsulation efficiency and in vitro drug release using 50% methanol/buffer pH 7.4 as dissolution medium. EtAC and PVA formulations showed the highest encapsulation efficiency and the lowest burst release. These microspheres were further characterized for particle size distribution, SEM and zeta potential. The results suggested that these materials could be starting materials to prepare a beta-estradiol biodegradable controlled delivery system.     

Controlled release of huperzine A from biodegradable microspheres: In vitro and in vivo studies

This paper describes the effect on Sun Protection Factor (SPF) of the combination of inorganic and organic filters in sunscreen products as determined by an in vitro method. O/W emulsions containing inorganic filters, such as titanium dioxide and zinc oxide, combined with 18 EU-authorized UV-B organic filters were tested. SPF measurements were carried out using a spectrophotometer equipped with an integrating sphere.

This study observed a synergic effect when titanium dioxide was combined with either anisotriazine or octyldimethylPABA. The combination of zinc oxide with 11 UV-B organic filters also exhibited a similar synergy; however, the measured SPF was systematically lower than the protection factor achieved with titanium dioxide.