Increased dissolution and physical stability of micronized nifedipine particles encapsulated with a biocompatible polymer and surfactants in a wet ball milling process.

Suspensions of nifedipine, a practically water-insoluble drug, were prepared in the presence of a biocompatible polymer, polyvinylpyrrolidone (PVP, K value 17), and three surfactants, sodium lauryl sulfate (SLS, anionic), cetyltrimethylammonium bromide (CETAB, cationic), polysorbate 80 (Tween 80, nonionic), by wet milling in ceramic ball mills. Nifedipine powders encapsulated with PVP and the surfactants were recovered from the suspensions after milling and evaluated for changes in particle size, morphology, sedimentation rate in aqueous suspensions, crystal form, and dissolution. Particle size analysis indicated that milling of suspensions in solutions of PVP and surfactants is an efficient method for reducing the particle size of nifedipine to below 10 microm. Furthermore, DSC and XPS analysis indicated that during milling the nifedipine crystals were coated with the PVP or surfactants and that milling with PVP stabilized the nifedipine crystal form during milling while nifedipine was gradually amorphisized when milled in a quaternary nifedipine/PVP/SLS/CETAB system. The decrease in particle size caused a significant decrease in sedimentation rate and increased the dissolution rate of nifedipine in simulated gastric fluid when compared to milled nifedipine and powder mixtures of the drug and the excipients.

Release of protein from highly cross-linked hydrogels of poly(ethylene glycol) diacrylate fabricated by UV polymerization.

Highly crosslinked hydrogel spheres were fabricated using UV photopolymerization of poly(ethylene glycol) diacrylate (PEG-DA) and pentaerythritol triacrylate (PETA) with 2,2'-dimethoxy-2-phenyl-acetophenone (DMPA) as the photoinitiator. Spheres were fabricated both with and without one of three comonomers: acrylic acid, acrylamide or allylamine. Photopolymerization rates and polymer morphology were determined using attenuated total reflectance/Fourier transform infrared spectroscopy and electron microscopy, respectively. These gels were further characterized for volume change, equilibrium water content, diffusivity of the expanding gel, molecular weight between crosslinks and polymer mesh size. Hydrogels with comonomers generally demonstrated an increase in equilibrium water content, average molecular weight between crosslinks and mesh size. Bovine serum albumin was incorporated into the hydrogel to simulate delivery of a model protein drug. The protein diffusion coefficients, based a Fickian release model, were calculated to be between 10(-10) and 10(-12) cm2/s with slight variance due to PETA concentration and the type of comonomer used.

Mammalian cell cultures on micropatterned surfaces of weak-acid, polyelectrolyte hyperbranched thin films on gold.

A four-step soft lithographic process based on micro-contact printing of organic monolayers, hyperbranched polymer grafting, and subsequent polymer functionalization results in polymer/n-alkanethiol patterns that direct the growth and migration of mammalian cells. The functional units on these surfaces are three-dimensional cell "corrals" that have walls 52+/-2 nm in height and lateral dimensions on the order of 60 microm. The corrals have hydrophobic, methyl-terminated n-alkanethiol bottoms, which promote cell adhesion, and walls consisting of hydrophilic poly(acrylic acid)/poly(ethylene glycol) layered nanocomposites that inhibit cell growth. Cell viability studies indicate that cells remain viable on the patterned surfaces for up to 21 days, and fluorescence microscopy studies of stained cells demonstrate that cell growth and spreading does not occur outside of the corral boundaries. This simple, chemically flexible micropatterning method provides spatial control over growth of IC-21 murine peritoneal macrophages, human umbilical vein endothelial cells, and murine hepatocytes.

Fabrication of poly(ethylene glycol) hydrogel microstructures using photolithography.

The fabrication of hydrogel microstructures based upon poly(ethylene glycol) diacrylates, dimethacrylates, and tetraacrylates patterned photolithographically on silicon or glass substrates is described. A silicon/silicon dioxide surface was treated with 3-(trichlorosilyl)propyl methacrylate to form a self-assembled monolayer (SAM) with pendant acrylate groups. The SAM presence on the surface was verified using ellipsometry and time-of-flight secondary ion mass spectrometry. A solution containing an acrylated or methacrylated poly(ethylene glycol) derivative and a photoinitiator (2,2-dimethoxy-2-phenylacetophenone) was spin-coated onto the treated substrate, exposed to 365 nm ultraviolet light through a photomask, and developed with either toluene, water, or supercritical CO2. As a result of this process, three-dimensional, cross-linked PEG hydrogel microstructures were immobilized on the surface. Diameters of cylindrical array members were varied from 600 to 7 micrometers by the use of different photomasks, while height varied from 3 to 12 micrometers, depending on the molecular weight of the PEG macromer. In the case of 7 micrometers diameter elements, as many as 400 elements were reproducibly generated in a 1 mm2 square pattern. The resultant hydrogel patterns were hydrated for as long as 3 weeks without delamination from the substrate. In addition, micropatterning of different molecular weights of PEG was demonstrated. Arrays of hydrogel disks containing an immobilized protein conjugated to a pH sensitive fluorophore were also prepared. The pH sensitivity of the gel-immobilized dye was similar to that in an aqueous buffer, and no leaching of the dye-labeled protein from the hydrogel microstructure was observed over a 1 week period. Changes in fluorescence were also observed for immobilized fluorophore labeled acetylcholine esterase upon the addition of acetyl acholine.

Transdermal extraction of interstitial fluid by low-frequency ultrasound quantified with 3H2O as a tracer molecule.

The transdermal extraction of interstitial fluid by low-frequency ultrasound offers a potential minimally invasive method of obtaining a fluid sample for at-home blood glucose monitoring. Here we show that the application of low-frequency ultrasound (20 kHz) enhances the transdermal transport of interstitial fluid across hairless rat skin. Using 3H2O as a tracer injected intravenously, a measurable amount of water (>1 microL) was extracted without producing any histologic evidence of injury, even after repeated exposures. Piezoelectric transducers were imbedded in the extraction chamber and used to correlate ultrasound spectral properties to the amount of fluid extracted. Results indicate that the highest amount of water extracted occurs when the acoustic coupling media on the surface of the skin is cavitating, resulting in mild ablation of the stratum corneum and a reduction in its resistance to water mass transfer.

Glucose and lactate biosensors based on redox polymer/oxidoreductase nanocomposite thin films.

Glucose and lactate enzyme electrodes have been fabricated through the deposition of an anionic self-assembled monolayer and subsequent redox polymer/enzyme electrostatic complexation on gold substrates. These surfaces were functionalized with a negative charge using 11-mercaptoundecanoic acid (MUA), followed by alternating immersions in cationic redox polymer solutions and anionic glucose oxidase (GOX) or lactate oxidase (LAX) solutions to build the nanocomposite structure. The presence of the multilayer structure was verified by ellipsometry and sensor function characterized electrochemically. Reproducible analyte response curves from 2 to 20 mM (GOX) and 2-10 mM (LAX) were generated with the standard deviation between multiple sensors between 12 and 17%, a direct result of the reproducibility of the fabrication technique. In the case of glucose enzyme electrodes, the multilayer structure was further stabilized through the introduction of covalent bonds within and between the layers. Chemical cross-linking was accomplished by exposing the thin film to glutaraldehyde vapors, inducing linkage formation between lysine and arginine residues present on the enzyme periphery with amine groups present on a novel redox polymer, poly[vinylpyridine Os(bisbipyridine)2Cl]-co-allylamine. Finally, an initial demonstration of thin-film patterning was performed as a precursor to the development of redundant sensor arrays. Microcontact printing was used to functionalize portions of a gold surface with a blocking agent, typically 1-hexadecanethiol. This was followed by immersion in MUA to functionalize the remaining portions of gold with negative charges. The multilayer deposition process was then followed, resulting in growth only on the regions containing MUA, resulting in a "positive"-type pattern. This technique may be used for fabrication of thin-film redundant sensor arrays, with thickness under 100 angstrom and lateral dimensions on a micrometer scale.

Monte Carlo modeling for implantable fluorescent analyte sensors.

A Monte Carlo simulation of photon propagation through human skin and interaction with a subcutaneous fluorescent sensing layer is presented. The algorithm will facilitate design of an optical probe for an implantable fluorescent sensor, which holds potential for monitoring many parameters of biomedical interest. Results are analyzed with respect to output light intensity as a function of radial distance from source, angle of exit for escaping photons, and sensor fluorescence (SF) relative to tissue autofluorescence (AF). A sensitivity study was performed to elucidate the effects on the output due to changes in optical properties, thickness of tissue layers, thickness of the sensor layer, and both tissue and sensor quantum yields. The optical properties as well as the thickness of the stratum corneum, epidermis, (tissue layers through which photons must pass to reach the sensor) and the papillary dermis (tissue distal to sensor) are highly influential. The spatial emission profile of the SF is broad compared that of the tissue fluorescence and the ratio of sensor to tissue fluorescence increases with distance from the source. The angular distribution of escaping photons is more concentrated around the normal for SF than for tissue AF. The information gained from these simulations will be helpful in designing appropriate optics for collection of the signal of interest.

Active growth factor delivery from poly(D,L-lactide-co-glycolide) foams prepared in supercritical CO(2).

A method for the production of microporous poly(D, L-lactide-co-glycolide) foams containing encapsulated proteins using supercritical carbon dioxide is described. Foams generated as aqueous protein emulsions in a polymer-solvent solution were saturated with carbon dioxide at supercritical conditions, and then suddenly supersaturated at ambient conditions causing bubble nucleation and precipitation of the polymer. Proteins contained in the water phase of the emulsion were encapsulated within the foams, including basic fibroblast growth factor (bFGF), an angiogenic factor of interest in tissue engineering applications. The release and activity of bFGF from these foams was determined in vitro and compared with similar porous scaffolds prepared by traditional solvent casting-salt leaching techniques. Total protein release rate was greater from structures made in CO(2) than those made by the salt leaching technique, however a large initial burst of bFGF was released from the salt leached structures. This initial burst was not observed from the polymer foams processed in CO(2) and active bFGF was released at a relatively constant rate. Residual methylene chloride levels were measured in the foams made with CO(2) and were found to be above the limits imposed by the US Pharmacopoeia implying that further solvent removal would be required prior to in vivo use.