Harmonic analysis of surface instability patterns on colloidal particles.

Wrinkling of colloidal particles alter a wide variety of interfacial properties but quantitative topographical descriptions have been explored experimentally to a very limited extent. In this study, we present a harmonic analysis of surface wrinkles and folds on submicron colloidal particles, obtained using an aerosol flow route, with small radius (<300 nm) and high crust thickness-to-radius ratio (>0.1). The particle surface coordinates were mapped in their entirety using cryo-electron tomography and subsequently reconstructed using spherical harmonics, allowing a spectral topographical description of the instability patterns and the identification of their surface modes by lateral wavelength. Wrinkled and crumpled particles showed a similar surface roughness spectrum, wherein differences were found most noticeable in the large wavelength region. The analysis of preferred directions of harmonic frequencies indicated a possible axial or planar alignment attributed to the directionality of the surface corrugations. The employed characterization methodology can further the study of topographical influences on colloidal interactions.

Aerosolization, Drug Permeation and Cellular Interaction of Dry Powder Pulmonary Formulations of Corticosteroids with Hydroxypropyl-ß-Cyclodextrin as a Solubilizer.

PURPOSE: The purpose of this study was to assess the feasibility of hydroxypropyl-ß-cyclodextrin as a solubilizer for the corticosteroids prednisolone and fludrocortisone acetate in dry powder inhalation formulations. METHODS: The dry particles were simultaneously produced and coated with nanosized L-leucine crystals using an aerosol flow reactor method. The aerosolization performances of carrier-free powders were studied using Easyhaler® and Twister™ at 2 and 4 kPa pressure drops over the inhalers. Drug permeation properties of the formulations were tested across a Calu-3 cell monolayer. Toxicity and reactive oxygen species induction were tested against Calu-3 and A549 cell lines. RESULTS: The hydroxypropyl-ß-cyclodextrin in the powders promoted the dissolution of fludrocortisone the most, followed by that of prednisolone. Fine particle fractions were 52-70% from emitted doses which showed good repeatability with a coefficient variation of 0.9-0.17. In addition, hydroxypropyl-ß-cyclodextrin enhanced the permeation of the corticosteroids. The powders showed no statistically significant toxicity nor reactive oxygen species induction in the tested cell lines. CONCLUSIONS: This study demonstrated the preparation and function of fine powder formulations which combine improved dissolution of poorly soluble drugs with good aerosolization performance. These results are expected to promote particle engineering as a way to develop new types of therapeutic pulmonary powders.

Thermoresponsive Nanoparticles of Self-Assembled Block Copolymers as Potential Carriers for Drug Delivery and Diagnostics.

Thermally responsive hydrogel nanoparticles composed of self-assembled polystyrene-b-poly(N-isopropylacrylamide)-b-polystyrene block copolymers and fluorescent probe 1-anilinonaphthalene-8-sulfonic acid have been prepared by aerosol flow reactor method. We aimed exploring the relationship of intraparticle morphologies, that were, PS spheres and gyroids embedded in PNIPAm matrix, as well PS-PNIPAm lamellar structure, to probe release in aqueous solution below and above the cloud point temperature (CPT) of PNIPAm. The release was detected by fluorescence emission given by the probe binding to bovine serum albumin. Also, the colloidal behavior of hydrogel nanoparticles at varying temperatures were examined by scattering method. The probe release was faster below than above the CPT from all the morphologies of which gyroidal morphology showed the highest release. Colloidal behavior varied from single to moderately aggregated particles in order spheres-gyroids-lamellar. Hydrogel nanoparticles with tunable intra particle self-assembled morphologies can be utilized designing carrier systems for drug delivery and diagnostics.

Polypeptide-based aerosol nanoparticles: self-assembly and control of conformation by solvent and thermal annealing.

Nanoconfined self-assemblies within aerosol nanoparticles and control of the secondary structures are shown here upon ionically complexing poly(L-lysine) (PLL) with dodecylbenzenesulfonic acid (DBSA) surfactant and using solvents chloroform, 1-propanol, or dimethylformamide. Different solvent volatilities and drying temperatures allowed tuning the kinetics of morphology formation. The supramolecular self-assembly and morphology were studied using cryo-TEM and SEM, and the secondary structures, using FT-IR. Highly volatile chloroform led to the major fraction of α-helical conformation of PLL(DBSA), whereas less volatile solvents or higher drying temperatures led to the increasing fraction of ß-sheets. Added drugs budesonide and ketoprofen prevented ß-sheet formation and studied PLL(DBSA)-drug nanoparticles were in the α-helical conformation. Preliminary studies showed that ketoprofen released with a slower rate than budesonide which was hypothesized to result from different localization of drugs within the PLL(DBSA) nanoparticles. These results instruct to prepare polypeptide aerosol nanoparticles with internal self-assembled structures and to control the secondary structures by aerosol solvent annealing, which we foresee to be useful, e.g., toward controlling the release of poorly soluble drug molecules.

Structure and dissolution of L-leucine-coated salbutamol sulphate aerosol particles.

L-Leucine formed different crystalline coatings on salbutamol sulphate aerosol particles depending on the saturation conditions of L-leucine. The work emphasizes a careful characterization of powders where structural compartments such as crystal size and particle coating may affect the performance of drug when administered. The sublimation of L-leucine from the aerosol particles took place 90°C lower temperature than the bulk L-leucine which was attributed to result from the sublimation of L-leucine from nano-sized crystalline domains. The dissolution slowed down and initial dissolution rate decreased with increasing L-leucine content. Decreasing crystalline domains to nano-scale improve heat and mass transfer which was observed as the lowered decomposition temperature of the drug salbutamol sulphate and the sublimation temperature of surface material L-leucine as well as the altered dissolution characteristics of the drug. The structure of the coated drug particles was studied by means of thermal analysis techniques (DSC and TG), and the dissolution of salbutamol sulphate was studied as an on-line measurement in a diffusion cell.

Intact nanoparticulate indomethacin in fast-dissolving carrier particles by combined wet milling and aerosol flow reactor methods.

PURPOSE: Drug development is often hindered by a drug's low dissolution rate. We present a method to increase dissolution rate of a drug powder by producing crystalline nanoparticles that are dispersed in carrier microparticles. METHODS: Indomethacin crystals of a few hundred nanometers are prepared by media milling using poloxamer 188 as a stabilizer. Nanoparticles are embedded into microparticles with a mannitol matrix and an L-leucine coating layer using an aerosol flow reactor method. RESULTS: Microparticles stabilize the primary nanoparticles in an intact crystalline form and release them when re-dispersed in aqueous medium. Secondary microparticle structure dissolves rapidly, resulting in a fast release and dissolution of indomethacin. In this manner, it is possible to change the surface layer of the particles from the one needed for nanoparticle production to one more suitable for process formulation of pharmaceuticals for, e.g., tablet or pulmonary products. CONCLUSIONS: Particle assemblies where nano-sized crystalline drug domains are embedded in solid microparticles are presented. The present work is a promising approach towards a "nanos-in-micros" concept as a tool for pharmaceutical nanoparticle processing.

Aerosolization behavior of carrier-free L-leucine coated salbutamol sulphate powders.

Aerosolization behavior of carrier-free l-leucine coated salbutamol sulphate inhalable powders has been studied. L-Leucine coatings were formed by physical vapour deposition (PVD) on the surface of the spherical particles in the gas phase. While depositing L-leucine formed pointy crystalline asperities whose size and density increased with the increased content of L-leucine in the gas phase. The asperity size changed from few nanometers to hundreds of nanometers. Due to the rough surface, all these coated fine powders were well-flowable and could be fed without the aid of coarser carriers. The aerosolization characteristics of the powders were studied with 'Inhalation Simulator' under ascending and fast inhalation profiles. When detected on-line by infrared light attenuation, the emission of the coated powders from an inhaler (Easyhaler) was distinctively dependent on the inhalation flow rate less than 30 l/min whereas that of micronized salbutamol sulphate powder solely depended on the studied inhalation flow rate range up to 100 l/min. Gravimetric measurements showed that emitted doses (ED) and fine particle fractions (FPF) of the coated powders were 5.1-7.1 mg/dose and 42-47%, respectively, which were 3-4 times higher than those of the micronized powder. The ED and FPF of the coated powders decreased as the surface roughness increased which is hypothesized as mechanical interlocking between the surface asperities.

Blocking the lateral film growth at the nanoscale in area-selective atomic layer deposition.

Area-selective atomic layer deposition (ALD) allows the growth of highly uniform thin inorganic films on certain parts of the substrate while preventing the film growth on other parts. Although the selective ALD growth is working well at the micron and submicron scale, it has failed at the nanoscale, especially near the interface where there is growth on one side and no-growth on the other side. The reason is that methods so far solely rely on the chemical modification of the substrate, while neglecting the occurrence of lateral ALD growth at the nanoscale. Here we present a proof-of-concept for blocking the lateral ALD growth also at the nanoscale by combining the chemical surface modification with topographical features. We demonstrate that area-selective ALD of ZnO occurs by applying the diethylzinc/water ALD process on cicada wings that contain a dense array of nanoscopic pillars. The sizes of the features in the inorganic film are down to 25 nm which is, to the best of our knowledge, the smallest obtained by area-selective ALD. Importantly, our concept allows the synthesis of such small features even though the film is multiple times thicker.

Simultaneous synthesis and coating of salbutamol sulphate nanoparticles with L-leucine in the gas phase.

Salbutamol sulphate nanoparticles have been simultaneously prepared and coated with L-leucine in the gas phase. Three different ways of coating can be separated based on the operation temperatures used in an aerosol flow reactor. Below the temperature of L-leucine sublimation, formation of the L-leucine layer on the core particle surface takes place via diffusion of L-leucine molecules on the droplet surfaces during droplet drying. At intermediate temperatures, the extent of sublimation of L-leucine depends notably on the concentration, and thus partial evaporation was expected. The L-leucine coating was solely formed via vapor deposition at high reactor temperatures when complete sublimation of L-leucine was obtained. The geometric mean diameter of the core salbutamol particles was approximately 65 nm. In general, particle size increased with the addition of L-leucine. The size distribution remained the same or broadened when the coating layer of the particles was formed via surface diffusion whereas notable narrowing of the distribution was observed when the coating was formed via vapor deposition. Upon desublimation and heterogeneous nucleation on the surfaces of smooth, spherical core particles, L-leucine formed a discontinuous coating with leafy crystals a few nanometers in size.

Pulmonary administration of a dry powder formulation of the antifibrotic drug tilorone reduces silica-induced lung fibrosis in mice.

The aim of this work was to study the antifibrotic effect of pulmonary administration of tilorone to lung fibrosis. L-leucine coated tilorone particles were prepared and their aerosolization properties were analyzed using two dry powder inhalers (Easyhaler and Twister). In addition, the biological activity and cell monolayer permeation was tested. The antifibrotic effect of tilorone delivered by oropharyngeal aspiration was studied in vivo using a silica-induced model of pulmonary fibrosis in mice in a preventive setting. When delivered from the Easyhaler in an inhalation simulator, the emitted dose and fine particle fraction were independent from the pressure applied and showed dose repeatability. However, with Twister the aerosolization was pressure-dependent indicating poor compatibility between the device and the formulation. The formulation showed more consistent permeation through a differentiated Calu-3 cell monolayer compared to pristine tilorone. Tilorone decreased the histological fibrosis score in vivo in systemic and local administration, but only systemic administration decreased the mRNA expression of type I collagen. The difference was hypothesized to result from 40-fold higher drug concentration in tissue samples in the systemic administration group. These results show that tilorone can be formulated as inhalable dry powder and has potential as an oral and inhalable antifibrotic drug.

Drug permeation and cellular interaction of amino acid-coated drug combination powders for pulmonary delivery.

The effect of three amino acid coatings (L-leucine, L-valine and L-phenylalanine) on particle integrity, aerosolization properties, cellular interaction, cytocompatibility, and drug permeation properties of drug combination powder particles (beclomethasone dipropionate and salbutamol sulphate) for dry powder inhalation (DPI) was investigated. Particles with crystalline L-leucine coating resulted in intact separated particles, with crystalline L-valine coating in slightly sintered particles and with amorphous L-phenylalanine coating in strongly fused particles. The permeation of beclomethasone dipropionate across a Calu-3 differentiated cell monolayer was increased when compared with its physical mixture. Drug crystal formation was also observed on the Calu-3 cell monolayer. The L-leucine coated particles were further investigated for cytocompatibility in three human pulmonary (Calu-3, A549 and BEAS-2B) and one human macrophage (THP-1) cell lines, where they showed excellent tolerability. The l-leucine coated particles were also examined for their ability to elicit reactive oxygen species in pulmonary BEAS-2B and macrophage THP-1 cell lines. The study showed the influence of the amino acid coatings for particle formation and performance and their feasibility for combination therapy for pulmonary delivery.

High-Throughput Synthesis of Lignin Particles (∼30 nm to ∼2 µm) via Aerosol Flow Reactor: Size Fractionation and Utilization in Pickering Emulsions.

An aerosol flow reactor was used for the first time for high-throughput, high yield synthesis of spherical lignin particles with given inherent hydrophilicity, depending on the precursor biomolecule. In situ fractionation via Berner type impactor afforded populations with characteristic sizes ranging from ∼30 nm to 2 µm. The as-produced, dry lignin particles displayed excellent mechanical integrity, even after redispersion under high shear in either mineral oil or water. They were effective in the stabilization of oil-in-water (O/W) Pickering emulsions with tunable droplet size, depending on the dimension of the lignin particles used for emulsification. The emulsion stability correlated with particle concentration as well as the respective lignin type. For the O/W emulsions stabilized with the more hydrophilic lignin particles, negligible changes in phase separation via Ostwald ripening and coalescence were observed over a period of time of more than two months. Together with the fact that the lignin particle concentrations used in emulsification were as low as 0.1%, our results reveal a remarkable ability to endow emulsified systems with high colloidal stability. Overall, we offer a new, high-yield, scalable nanomanufacturing approach to producing dry spherical lignin particles with size control and high production capacity. A number of emerging applications for these organic particles can be envisioned and, as a proof-of-concept, we illustrate here surfactant-free emulsification.

Oral hypoglycaemic effect of GLP-1 and DPP4 inhibitor based nanocomposites in a diabetic animal model.

Glucagon-like peptide-1 (GLP-1), an incretin hormone, is used for type 2 diabetes mellitus (T2DM) treatment because of its ability to stimulate insulin secretion and release in a glucose-dependent manner. Despite of its potent insulinotropic effect, oral GLP-1 delivery is greatly limited by its instability in the gastrointestinal tract, poor absorption efficiency and rapid degradation by dipeptidylpeptidase-4 (DPP4) enzyme leading to a short half-life (~2min). Thus, a multistage dual-drug delivery nanosystem was developed to deliver GLP-1 and DPP4 inhibitor simultaneously. The system comprised of chitosan-modified porous silicon (CSUn) nanoparticles, which were coated by an enteric polymer, hydroxypropylmethylcellulose acetate succinate MF, using aerosol flow reactor technology. A non-obese T2DM rat model induced by co-administration of nicotinamide and streptozotocin was used to evaluate the in vivo efficacy of the nanosystem. The oral administration of H-CSUn nanoparticles resulted in 32% reduction in blood glucose levels and ~6.0-fold enhancement in pancreatic insulin content, as compared to the GLP-1+DPP4 inhibitor solution. Overall, these results present a promising system for oral co-delivery of GLP-1 and DPP4 inhibitor that could be further evaluated in a chronic diabetic study.

Multistage pH-responsive mucoadhesive nanocarriers prepared by aerosol flow reactor technology: A controlled dual protein-drug delivery system.

Nanotechnology based drug delivery systems are anticipated to overcome the persistent challenges in oral protein and peptide administration, and lead to the development of long awaited non-invasive therapies. Herein, an advanced single-step aerosol flow reactor based technology was used to develop a multifunctional site specific dual protein-drug delivery nanosystem. For this purpose, mucoadhesive porous silicon (PSi) nanoparticles encapsulated into a pH-responsive polymeric nanomatrix was developed for advanced oral type 2 diabetes mellitus therapy with an antidiabetic peptide, glucagon like peptide-1 (GLP-1), and the enzyme inhibitor, dipeptidyl peptidase-4 (DPP4). Chitosan surface modification inherited the mucoadhesiveness to the nanosystem which led to enhanced cellular interactions and increased cellular compatibility. An advanced aerosol flow reactor technology was used to encapsulate the chitosan modified nanoparticles into an enteric polymeric nanomatrix. The pH-sensitive polymeric matrix simultaneously prevented the gastric degradation of the encapsulated peptide and also preserved the mucoadhesive functionality of the chitosan-modified PSi nanoparticles in the harsh stomach environment. The multidrug loaded nanosystem showed augmented intestinal permeability of GLP-1, evaluated in an in vitro cell-based intestinal epithelium model, attributed to the permeation enhancer effect of chitosan and inhibition of GLP-1 degradation by the DPP4 inhibitor. The applied technology resulted in the development of a dual-drug delivery nanosystem that synergizes the antidiabetic effect of the loaded peptide and the enzyme inhibitor, thereby indicating high clinical potential of the system and preparation technique.

Functionalization of alkyne-terminated thermally hydrocarbonized porous silicon nanoparticles with targeting peptides and antifouling polymers: effect on the human plasma protein adsorption.

Porous silicon (PSi) nanomaterials combine a high drug loading capacity and tunable surface chemistry with various surface modifications to meet the requirements for biomedical applications. In this work, alkyne-terminated thermally hydrocarbonized porous silicon (THCPSi) nanoparticles were fabricated and postmodified using five bioactive molecules (targeting peptides and antifouling polymers) via a single-step click chemistry to modulate the bioactivity of the THCPSi nanoparticles, such as enhancing the cellular uptake and reducing the plasma protein association. The size of the nanoparticles after modification was increased from 176 to 180-220 nm. Dextran 40 kDa modified THCPSi nanoparticles showed the highest stability in aqueous buffer. Both peptide- and polymer-functionalized THCPSi nanoparticles showed an extensive cellular uptake which was dependent on the functionalized moieties presented on the surface of the nanoparticles. The plasma protein adsorption study showed that the surface modification with different peptides or polymers induced different protein association profiles. Dextran 40 kDa functionalized THCPSi nanoparticles presented the least protein association. Overall, these results demonstrate that the "click" conjugation of the biomolecules onto the alkyne-terminated THCPSi nanoparticles is a versatile and simple approach to modulate the surface chemistry, which has high potential for biomedical applications.

Influence of the solvent composition on the aerosol synthesis of pharmaceutical polymer nanoparticles.

Spherical, Eudragit L100 polymer nanoparticles with and without a ketoprofen drug were prepared by a novel aerosol flow reactor method. In this method, the polymer solution is sprayed to form nanosized droplets followed by the evaporation of a solvent. A purpose of the work was to explore the effect of solvent, solvent mixture, and co-solute (ketoprofen) on the formation of polymer particle, and particularly on particle morphology. The solvents used, i.e. ethanol, THF, toluene, and water, were selected according to their vapor pressure and dissolution capability for the polymer. At the polymer concentration range from 0.2 to 1.5 g/l of the starting solution, the geometric number mean diameters (GMD) of the particles increased from 75 to 130 nm and from 65 to 100 nm from the solutions of ethanol and THF, respectively. Particle morphology was observed by a scanning electron microscope (SEM). Particles changed from collapsed to irregular via spherical shape in the course of the decreasing solubility of the polymer in the medium. This is critically dependent on the solvent evaporation rate as well as the solute solubility, i.e. fast evaporative removal of solvent results in collapsed particles whereas low solubility results in irregular particles. Interplay between the vapor pressure of the solvents and the polymer solubility in the medium made possible to prepare particles with more complicated structures such as shriveled and blistery structures. The particle morphology as detected by SEM did not change when 10 wt.% of ketoprofen was added to the precursor solution.

Nanoparticles containing ketoprofen and acrylic polymers prepared by an aerosol flow reactor method.

The purpose of this study was to outline the effects of interactions between a model drug and various acrylic polymers on the physical properties of nanoparticles prepared by an aerosol flow reactor method. The amount of model drug, ketoprofen, in the nanoparticles was varied, and the nanoparticles were analyzed for particle size distribution, particle morphology, thermal properties, IR spectroscopy, and drug release. The nanoparticles produced were spherical, amorphous, and had a matrix-type structure. Ketoprofen crystallization was observed when the amount of drug in Eudragit L nanoparticles was more than 33% (wt/wt). For Eudragit E and Eudragit RS nanoparticles, the drug acted as an effective plasticizer resulting in lowering of the glass transition of the polymer. Two factors affected the preparation of nanoparticles by the aerosol flow reactor method, namely, the solubility of the drug in the polymer matrix and the thermal properties of the resulting drug-polymer matrix.

Injected nanoparticles: the combination of experimental systems to assess cardiovascular adverse effects.

When nanocarriers are used for drug delivery they can often achieve superior therapeutic outcomes over standard drug formulations. However, concerns about their adverse effects are growing due to the association between exposure to certain nanosized particles and cardiovascular events. Here we examine the impact of intravenously injected drug-free nanocarriers on the cardiovasculature at both the systemic and organ levels. We combine in vivo and in vitro methods to enable monitoring of hemodynamic parameters in conscious rats, assessments of the function of the vessels after sub-chronic systemic exposure to nanocarriers and evaluation of the direct effect of nanocarriers on vascular tone. We demonstrate that nanocarriers can decrease blood pressure and increase heart rate in vivo via various mechanisms. Depending on the type, nanocarriers induce the dilation of the resistance arteries and/or change the responses induced by vasoconstrictor or vasodilator drugs. No direct correlation between physicochemical properties and cardiovascular effects of nanoparticles was observed. The proposed combination of methods empowers the studies of cardiovascular adverse effects of the nanocarriers.

Gas-phase synthesis of solid state DNA nanoparticles stabilized by l-leucine.

Aerosol flow reactor is used to generate solid-state nanoparticles in a one-step process that is based on drying of aerosol droplets in continuous flow. We investigated the applicability of aerosol flow reactor method to prepare solid state DNA nanoparticles. Precursor solutions of plasmid DNA with or without complexing agent (polyethylenimine), coating material (l-leucine) and mannitol (bulking material) were dispersed to nanosized droplets and instantly dried in laminar heat flow. Particle morphology, integrity and stability were studied by scanning electron microscopy. The stability of DNA was studied by gel electrophoresis. Plasmid DNA as such degraded in the aerosol flow process. Complexing agent protected DNA from degradation and coating material enabled production of dispersed, non-aggregated, nanoparticles. The resulting nanoparticles were spherical and their mean diameter ranged from 65 to 125nm. The nanoparticles were structurally stable at room temperature and their DNA content was about 10%. We present herein the proof of principle for the production of dispersed solid state nanoparticles with relevant size and intact plasmid DNA.

Coated particle assemblies for the concomitant pulmonary administration of budesonide and salbutamol sulphate.

The aims were to prepare stable and well-dispersible pulmonary fine powders composed of combination drugs with different water solubility, to facilitate concomitant release of corticosteroid budesonide and short acting ß-agonist salbutamol sulphate and to improve the dissolution of the budesonide. The budesonide nanosuspensions were prepared by a wet milling which were mixed then with salbutamol sulphate, mannitol (bulking material) and leucine (coating material) for the preparation of micron-sized particles by an aerosol flow reactor wherein leucine formed a rough coating layer on particle surface. The stable and intact particle assemblies showed excellent aerosolization performance. The emitted doses from the inhaler, Easyhaler(®), were ~3 mg/dose with a coefficient variation of 0.1, and the fine particle fractions were ~50%. Complete dissolution of budesonide nanocrystals from the particles took place within 20 min with the same rate as salbutamol sulphate. Combining the two formulation technologies enabled the encapsulation of drugs with different solubility into a single, intact particle. The leucine coating provided excellent aerosolization properties which allowed fine powder delivery from the inhaler without carrier particles. This study showed the feasibility of preparing powders for combination therapy that are utilized, for instance, in inhalation therapy.