Effect of gastrointestinal protein adsorption on the in vitro release of AZT from ethylcellulose microspheres.

The purpose of this study was to assess the effect of gastrointestinal proteins on the in vitro release of zidovudine (AZT) from ethylcellulose microspheres, and to investigate protein adsorption as a possible mechanism that mediates this effect. AZT release from ethylcellulose microspheres was tested in the presence of different gastrointestinal proteins, both dietary (casein and albumin) and endogenous (pepsin, pancreatin, and mucin) in simulated gastric fluid and/or simulated intestinal fluid. The resulting release profiles were compared with those produced in the corresponding release media without the presence of proteins. Protein adsorption on AZT-loaded ethylcellulose microspheres was studied for the five proteins under investigation. The amounts of adsorbed proteins were determined by fluorescent spectrometry after the protein solution was reacted with fluoraldehyde reagent. All of the investigated proteins were found to slow the release of AZT from ethylcellulose microspheres. At gastric pH, ovalbumin and casein had the maximum effect on AZT release. Mucin exerted a more pronounced effect at gastric pH compared with that at intestinal pH. The negative effect of pancreatin on AZT release increased when its concentration was increased. The five proteins were found to adsorb on AZT-loaded ethylcellulose microspheres with varying quantities. The observed protein adsorption is believed to cause blockage of the small pores and channels in the microsphere structure, and consequently slow the release of AZT.

In vivo evaluation of zidovudine (AZT)-loaded ethylcellulose microspheres after oral administration in beagle dogs.

The purpose of this study was to evaluate the in vivo performance of sustained-release zidovudine (AZT) microspheres after oral administration in Beagle dogs, and to establish an in vitro-in vivo correlation. Two AZT microsphere formulations as well as AZT powder were administered to four Beagle dogs. Plasma samples were analyzed by HPLC. The plasma concentration-time data was analyzed by both compartmental and noncompartmental pharmacokinetic analyses. Based on the calculated pharmacokinetic parameters, in vivo release profiles were simulated and compared with in vitro release profiles in three different release media. Significantly longer mean residence time (MRT) was observed after administration of the sustained-release microspheres compared with AZT powder. Significantly lower maximum (Cmax) concentration values and longer times to Cmax (tmax) values were also observed. Formulation I showed the longest MRT (4.4 h). AZT plasma concentration was maintained above the minimum effective concentration for approximately 10 h after administration of Formulation I. The relative bioavailability of the microsphere formulations with respect to AZT powder was not significantly different from 1. The in vitro release of the three formulations was slower in simulated gastric fluid compared with simulated intestinal fluid. The addition of enzymes and mucin to the release media significantly lowered the in vitro release rate of AZT from the microspheres formulations, but not from AZT powder. A good level of in vitro-in vivo correlation (Level A correlation) was achieved with a release medium that was composed of simulated gastric fluid with pepsin and mucin for 2 h followed by simulated intestinal fluid with pancreatin and mucin for 8 h. This in vitro model may be used to predict the in vivo release of AZT, in the further development of controlled-release AZT formulations.

Biodegradable cisplatin microspheres for direct brain injection: preparation and characterization.

The objectives of the present study were to prepare cisplatin loaded-PLGA microspheres that are suitable for direct brain injection and to characterize them in terms of their physicochemical properties, in vitro drug release, and self-removal mechanism. The microspheres were prepared by emulsification/solvent evaporation method using PLGA (50:50) as the biodegradable matrix forming polymer. The physicochemical characterization encompassed the following: surface morphology, particle size, entrapment efficiency, surface area, and density. The in vitro release and in vitro degradation studies were performed in phosphate buffer and in 10% rat brain preparation. SEM micrographs revealed that the microspheres have a rough porous surface and a smooth interior. Particle size typically ranged from 180 to 250 microns with an average of 230 T microns. Entrapment efficiency was approximately 70% and was found to be dependent on the particle size. Surface area and density ranged from 0.038 to 0.025 m2/g and from 1.44 to 1.39 g/cm3, respectively. Both were also dependent on particle size. In the in vitro release study in phosphate buffer, approximately 80% of cisplatin was released over 30 days, after which the release rate plateaued. The release profile in 10% rat brain preparation was comparable in shape to that obtained in phosphate buffer. However, the release rate was lower and the total amount released by the end of the study was only 55% of the total cisplatin content. The degradation of PLGA microspheres in phosphate buffer and in rat brain homogenate correlated well with the respective release profiles. Based on the evidence of self-removal and the sustained release of cisplatin for over a month, cisplatin-loaded PLGA microspheres may be useful for local delivery to brain tumors.

Optima: a windows-based program for computer-aided optimization of controlled-release dosage forms.

The purpose of this work was to develop a computer program that assists optimization of controlled-release devices, both visually and mathematically, using response surface methodology (RSM). A Windows-based computer program, Optima, which interactively implemented a number of subroutines for the optimization procedure, was developed. Optima is an integrated, user-friendly, and graphically oriented program for pharmaceutical dosage form optimization. Central composite design is implemented in the program. First- and second-order models containing up to five variables can be fitted to the data. The user can also choose between linear and exponential individual desirability functions, and use them to construct an overall desirability function that combines all the response variables in a single response. The program can predict the optimum levels of experimental variables, with respect to individual responses and/or the overall desirability. Optima has been successfully used in the development of sustained-release AZT-loaded microspheres. During the optimization process, three experimental variables were investigated and four responses were measured. The experimental design was a central composite design that was generated by the program. The response values were used by the program to calculate the individual desirability functions, which were then combined into an overall desirability function. The individual responses as well as the overall desirability function were optimized by fitting to a second-order polynomial equation. The response surfaces were generated and optimum levels of the experimental variables were predicted. The observed responses of the optimized formulation were very close to those predicted by Optima. The program proved to be a very useful, integrated tool for optimization of the controlled-release microspheres.

Preparation and evaluation of zidovudine-loaded sustained-release microspheres. 2. Optimization of multiple response variables.

The formulation of zidovudine-loaded (AZT-loaded) sustained-release microspheres was prepared and optimized using response surface methodology. The objective was to use this statistical procedure to obtain a formulation with optimized overall properties. The effect of formulation variables (emulsifier concentration, drug to polymer ratio, and composition of the internal phase of the emulsion) on a number of response variables was systemically investigated. The response variables were t85, entrapment efficiency, yield, and percentage of loose surface crystals. A desirability function that combines these four response variables was constructed. A second-order polynomial equation was fitted to the data, and the resulting equation was used to predict the responses in the optimal region. All the investigated response variable were found to be highly dependent on the formulation variables, with strong interactions observed between the formulation variables. It was found that optimum overall desirability of AZT microspheres can be obtained at low levels of SDS and ethyl acetate concentrations and at intermediate levels of drug to polymer ratio. An optimized formulation was prepared under these experimental conditions and evaluated for individual responses and overall desirability. The experimental values of the response variables highly agreed with the predicted values.

Preparation and evaluation of sustained release AZT-loaded microspheres: optimization of the release characteristics using response surface methodology.

The purpose of the study was to prepare and optimize a sustained release formulation of zidovudine (AZT). Ethylcellulose microspheres containing AZT were prepared using an emulsification/solvent evaporation technique. The critical formulation variables were emulsifier concentration, drug to polymer ratio, and ethyl acetate concentration in the internal phase of the emulsion. The time to release 85% of the contents of the microspheres (t85) was used as a measure for the release time. A second-year polynomial equation was fitted to the release data to systemically investigate the effect of the formulation variables on the release rate. This equation was then used to predict t85 in the optimum region. The t85 was found to be dependent on the three formulation variables, with strong interactions observed between these variables. The microspheres were characterized in terms of their particle size and surface morphology. The study indicated no overall correlation between the mean diameter of the microspheres and the t85.

High-performance liquid chromatographic assay for sodium mercaptoundecahydrododecaborate in rat tissues.

Mercaptoundecahydrododecaborate (BSH) is an important agent for the boron neutron-capture therapy (BNCT). A sensitive high-performance liquid chromatographic (HPLC) method was developed for measuring BSH concentrations in rat tissues. Various tissue samples containing the drug were homogenized in a 1:1 (g/ml) mixture with phosphate buffered saline. The samples were then deproteinised with 4 volumes of acetonitrile and centrifuged. An aliquot of the supernatant was dried and reconstituted in 200 microliter of Tris-HC1 buffer. The samples were subjected to precolumn derivatization using the thiol reactive monobromobimane (mBB). The drug-mBB adduct was resolved by isocratic elution from a C18 reversed-phase column. The optimized mobile phase was methanol-0.02 M phosphate buffer (43:57, v/v) containing 0.01 M tetrabutylammonium dihydrogen phosphate as the ion-pairing agent with the final pH adjusted to 7.0. The flow-rate was set at 2.0 ml/min. The adduct was monitored by UV absorption at 373 nm. The analysis was completed in less than 15 min. The detection limit was 0.5 microgram/ml (0.25 microgram of boron). The assay method was linear over a concentration range of 0.5 to 50 micrograms/ml. This assay method could be used to evaluate the BSH concentrations in different tissues in studies on the targeted delivery of BSH.

Liquid chromatographic determination of sodium mercaptoundecahydrododecaborate in rat urine and plasma after precolumn derivatization.

A high-performance liquid chromatographic (HPLC) method was developed for the determination of disodium mercaptoundecahydrododecaborate (BSH) in biological fluids. Monobromobimane was used as a precolumn derivatizing agent. A stable derivative was obtained. The derivative was separated on a C18 column using reversed-phase ion-pairing chromatography and detected by a spectrophotometric detector at 373 nm. The detection limit was 200 ng/ml (0.1 ppm boron). Calibration curves were prepared for rat urine and plasma samples. The calibration curves were linear in the range of 1 microgram/ml to 100 micrograms/ml for urine samples and 0.2 micrograms/ml to 50 micrograms/ml for plasma samples.