Macroporous ZnO foams by high internal phase emulsion technique: synthesis and catalytic activity.

Zinc(II) oxide nanoparticles were used for the stabilization of dicyclopentadiene (DCPD)-water-based high internal phase emulsions (HIPEs), which were subsequently cured using ring-opening metathesis polymerization (ROMP). The morphology of the resulting ZnO-pDCPD nanocomposite foams was investigated in correlation to the nanoparticle loading and nanoparticle surface chemistry. While hydrophilic ZnO nanoparticles were found to be unsuitable for stabilizing the HIPE, oleic acid coated, yet hydrophobic ZnO nanoparticles were effective HIPE stabilizers, yielding polymer foams with ZnO nanoparticles located predominately at their surface. These inorganic/organic hybrid foam-materials were subsequently calcined at 550 °C for 15 min to obtain inorganic macroporous ZnO foams with a morphology reminiscent to the original hybrid foam, and a specific surface area of 1.5 m(2) g(-1). Longer calcination time (550 °C, 15 h) resulted in a sea urchin like morphology of the ZnO foams, characterized by higher specific surface area of 5.5 m(2) g(-1). The latter foam type showed an appealing catalytic performance in the catalytic wet air oxidation (CWAO) process for the destruction of bisphenol A.

Nano-extrusion: a promising tool for continuous manufacturing of solid nano-formulations.

Since more than 40% of today's drugs have low stability, poor solubility and/or limited ability to cross certain biological barriers, new platform technologies are required to address these challenges. This paper describes a novel continuous process that converts a stabilized aqueous nano-suspension into a solid oral formulation in a single step (i.e., the NANEX process) in order to improve the solubility of a model drug (phenytoin). Phenytoin nano-suspensions were prepared via media milling using different stabilizers. A stable nano-suspension was obtained using Tween(®) 80 as a stabilizer. The matrix material (Soluplus(®)) was gravimetrically fed into the hot melt extruder. The suspension was introduced through a side feeding device and mixed with the molten polymer to immediately devolatilize the water in the nano-suspension. Phenytoin nano-crystals were dispersed and embedded in the molten polymer. Investigation of the nano-extrudates via transmission electron microscopy and atomic force microscopy showed that the nano-crystals were embedded de-aggregated in the extrudates. Furthermore, no changes in the crystallinity (due to the mechanical and thermal stress) occurred. The dissolution studies confirmed that the prepared nano-extrudates increased the solubility of nano-crystalline phenytoin, regardless of the polymer. Our work demonstrates that NANEX represents a promising new platform technology in the design of novel drug delivery systems to improve drug performance.

In situ determination and imaging of physical properties of soft organic materials by analytical transmission electron microscopy.

Analytical transmission electron microscopy (ATEM) offers great flexibility in identification of the structural-chemical organization of soft materials at the level of individual macromolecules. However, the determination of mechanical characteristics such as hardness/elasticity of the amorphous and polycrystalline organic substances by ATEM has been problematic so far. Here, we show that energy filtered TEM (EFTEM) measurements enable direct identification and study of mechanical properties in complex (bio-)polymer systems of relevance for different industrial and (bio-)medical applications. We experimentally demonstrate strong correlations between hardness/elasticity of different polymers (polycaprolactone, polylactid, polyethelene, etc.) and their volume plasmon energy. Thickness and anisotropy effects, which substantially mask the material contrast in EFTEM bulk plasmon images, can be adequately removed by normalizing the latter by carbon elemental map. EFTEM data has been validated using atomic force microscopy phase images, where phase shift related to the hardness and elastic modulus of the materials.

Nanocomposite foams from iron oxide stabilized dicyclopentadiene high internal phase emulsions: preparation and bromination.

Nanocomposite polyHIPE foams with open-cellular morphology were obtained using nanoparticles (γFe2O3/Fe3O4), surfactant (Pluronic L121) or nanoparticle/surfactant stabilized dicyclopentadiene high internal phase emulsions (DCPD HIPEs). Upon curing, cavity sizes were found to vary drastically between 950 ± 360 µm down to 7 ±3 µm depending on the HIPE formulations. As-obtained nanocomposite polyHIPE foams were functionalized using elemental bromine in THF. Upon bromination the nanoparticles are moved from the cavities surfaces into the bulk phase of the polymer scaffold, which affects the inductive-heating capability of the magnetic nanocomposite foams decreasing it by the factor of 2.

Analytical electron microscopy for characterization of fluid or semi-solid multiphase systems containing nanoparticulate material.

The analysis of nanomaterials in pharmaceutical or cosmetic preparations is an important aspect both in formulation development and quality control of marketed products. Despite the increased popularity of nanoparticulate compounds especially in dermal preparations such as emulsions, methods and protocols of analysis for the characterization of such systems are scarce. This work combines an original sample preparation procedure along with different methods of analytical electron microscopy for the comprehensive analysis of fluid or semi-solid dermal preparations containing nanoparticulate material. Energy-filtered transmission electron microscopy, energy-dispersive X-ray spectroscopy, electron energy loss spectroscopy and high resolution imaging were performed on model emulsions and a marketed product to reveal different structural aspects of both the emulsion bulk phase and incorporated nanosized material. An innovative analytical approach for the determination of the physical stability of the emulsion under investigation is presented. Advantages and limitations of the employed analytical imaging techniques are highlighted.

An analytical technique to extract surface information of negatively stained or heavy-metal shadowed organic materials within the TEM.

Using a series of uranyl acetate stained or platinum-palladium shadowed organic samples, an empirical analytical method to extract surface information from energy-filtered transmission electron microscopy (EFTEM) images is described. The distribution of uranium or platinum-palladium atoms, which replicate the sample surface topography, have been mathematically extracted by dividing the image acquired in the valence bulk plasmon energy region (between 20 and 30 eV) by the image acquired at the carbon K ionization edge (between 284 and 300 eV). The resulting plasmon-to-carbon ratio (PCR) image may be interpreted as a precise metal replica of the sample surface. In contrast to conventional EFTEM elemental mapping, including an absolute quantification approach, this technique can be applied to 200-600 nm thick organic samples. A combination of conventional TEM and PCR imaging allows one to detect complementary transmission and topographical information with nanometer precision of the same area of carbon-based samples. The advantages and limitations of PCR imaging are highlighted.

Electron microscopy of pharmaceutical systems.

During the last decades, the focus of research in pharmaceutical technology has steadily shifted towards the development and optimisation of nano-scale drug delivery systems. As a result, electron microscopic methods are increasingly employed for the characterisation of pharmaceutical systems such as nanoparticles and microparticles, nanoemulsions, microemulsions, solid lipid nanoparticles, different types of vesicles, nanofibres and many more. Knowledge of the basic properties of these systems is essential for an adequate microscopic analysis. Classical transmission and scanning electron microscopic techniques frequently have to be adapted for an accurate analysis of formulation morphology, especially in case of hydrated colloidal systems. Specific techniques such as environmental scanning microscopy or cryo preparation are required for their investigation. Analytical electron microscopic techniques such as electron energy-loss spectroscopy or energy-dispersive X-ray spectroscopy are additional assets to determine the elemental composition of the systems, but are not yet standard tools in pharmaceutical research. This review provides an overview of pharmaceutical systems of interest in current research and strategies for their successful electron microscopic analysis. Advantages and limitations of the different methodological approaches are discussed and recent findings of interest are presented.

Formation of a physical stable delivery system by simply autoclaving nanostructured lipid carriers (NLC).

A unique long term stable carrier system for poorly soluble drugs was obtained by simply autoclaving nanostructured lipid carriers (NLC). NLC composed of Precirol ATO, oleic acid, glycerol 85%, Eumulgin SML 20 and water as well as NLC loaded with the poorly soluble drug Itraconazole formed a special organized matrix containing lamellar bilayers similar to biological membranes after the sterilization process. The tendency to expel the poorly soluble drug from the matrix was thereby avoided.

Microscopic analysis of the surface functionalization of polymer particles and subsequent grafting of polymer chains from the surface.

The characterization of the surface functionalization of polymer particles and subsequent grafting of hydrated polymer chains from their surface by microscopic techniques are essential to obtain reliable data about the actual morphology of the system. Since the size range of morphological features of functionalized polymer surfaces has long ago reached the lower end of the nanometer scale, classical light microscopy and dynamic light scattering have been replaced by electron and atomic force microscopy techniques which provide sufficient resolution for the visualization of nano-sized structures. Moreover, only polymer particle aggregates and fine organization of hydrated polymer chains which are not efficiently characterized by particle size measurements can be detected accurately with microscopy methods. Both solid and hydrated systems can be characterized by transmission electron microscopy and scanning electron microscopy (inc. cryo-electron microscopy (EM)) after appropriate sample preparation. Moreover, analytical EM methods allow not only for the size, shape and internal structure characterization, but also for the chemical composition with high spatial resolution.

Electron microscopy of nanoemulsions: an essential tool for characterisation and stability assessment.

The characterisation of pharmaceutical formulations by microscopic techniques is essential to obtain reliable data about the actual morphology of the system. Since the size range of colloidal drug delivery systems has long ago reached the lower end of the nanometer scale, classical light microscopy has been replaced by electron microscopy techniques which provide sufficient resolution for the visualisation of nano-sized structures. Indeed, the superior resolution and methodological versatility of electron microscopy has rendered this technique an indispensable tool for the analysis of nanoemulsions. Microscopic analysis of these lipid-based drug delivery systems with particle sizes in the lower submicron range provides critical information about the size, shape and internal structure of the emulsion droplets. Moreover, surfactant aggregates such as liposomes or multilamellar structures which remain unnoticed during particle size measurements can be detected in this fashion. This review provides a brief overview about both transmission electron microscopy (TEM) and scanning electron microscopy (SEM) techniques which have been employed to characterise nanoemulsions. Of special interest are sophisticated cryo techniques of sample preparation for both TEM and SEM which deliver high-quality images of nanoemulsions in their natural state. An overview about the instrumentation and sample preparation for all presented methods is given. Important practical aspects, sources of error and common artefacts as well as recent methodological advances are discussed. Selected examples of electron microscopic studies of nanoemulsions are presented to illustrate the potential of this technique to reveal detailed and specific information.

Development of sucrose stearate-based nanoemulsions and optimisation through γ-cyclodextrin.

Nanoemulsions aimed at dermal drug delivery are usually stabilised by natural lecithins. However, lecithin has a high tendency towards self-aggregation and is prone to chemical degradation. Therefore, the aim of this study was to develop nanoemulsions with improved structure and long-term stability by employing a natural sucrose ester mixture as sole surfactant. A thorough comparison between the novel sucrose stearate-based nanoemulsions and corresponding lecithin-based nanoemulsions revealed that the sucrose ester is superior in terms of emulsifying efficiency, droplet formation as well as physical and chemical stability. The novel formulations exhibited a remarkably homogeneous structure in cryo TEM investigations, as opposed to the variable structure observed for lecithin-based systems. The in vitro skin permeation rates of lipophilic drugs from sucrose stearate nanoemulsions were comparable to those obtained with their lecithin-based counterparts. Furthermore, it was observed that addition of γ-cyclodextrin led to enhanced skin permeation of the steroidal drug fludrocortisone acetate from 9.99±0.46 to 55.10±3.67 µg cm(-2) after 24 h in the case of sucrose stearate-based systems and from 9.98±0.64 to 98.62±24.89 µg cm(-2) after 24 h in the case of lecithin-based systems. This enhancement effect was significantly stronger in formulations based on lecithin (P<0.05), which indicates that synergistic mechanisms between the surfactant and the cyclodextrin are involved. Cryo TEM images suggest that the cyclodextrin is incorporated into the interfacial film, which might alter drug release rates and improve the droplet microstructure.

Decrease of liposomal size and retarding effect on fluconazole skin permeation by lysine derivatives.

Liposomes are ideal dermal drug delivery systems because of their ability to alter the biodistribution profile of incorporated drugs. In a novel approach to optimize the liposomal microstructure, lysine derivatives were employed. The effect of the oligopeptides Lys-5 and Lys-7 on the structure as well as on the skin permeation of the antimycotic drug fluconazole in 1,2-dipalmitoyl-sn-glycero-3-phosphocholine vesicles was studied using a variety of techniques. It was demonstrated by addition of the shift reagent praseodymium(III)chloride and subsequent (31)P NMR measurements that the liposomes produced consisted mainly of unilamellar vesicles. This was confirmed by cryo-transmission electron microscopy. The addition of Lys-5 and Lys-7 induced a structural change resulting in a decrease in particle size between 10% and 40% and a retarding effect on fluconazole skin permeation.

Silicon: The key element in early stages of biocalcification.

Biocalcification is a widespread process of forming hard tissues like bone and teeth in vertebrates. It is also a topic connecting life sciences and earth sciences: calcified skeletons and shells deposited as sediments represent the earth's fossil record and are of paramount interest for biogeochemists trying to get an insight into the past of our planet. This study reports on the role of silicon in the early biocalcification steps, where silicon and calcium were detected on the surface of cyanobacteria (initial stage of lacustrine calcite precipitation) and in crustacean cuticles. By using innovative methodological approaches of correlative microscopy (AFM in combination with analytical TEM: EFTEM, EELS) the chemical form of silicon in biocalcifying matrices and organic-inorganic particles is determined. Previously, silicon was reported to be localized in active growth areas in the young bone of vertebrates. We have found evidence that biocalcification in evolutionarily distant organisms involves very similar initial phases with silicon as a key element at the organic-inorganic interface.

Semi-solid Sucrose Stearate-Based Emulsions as Dermal Drug Delivery Systems.

Mild non-ionic sucrose ester surfactants can be employed to produce lipid-based drug delivery systems for dermal application. Moreover, sucrose esters of intermediate lipophilicity such as sucrose stearate S-970 possess a peculiar rheological behavior which can be employed to create highly viscous semi-solid formulations without any further additives. Interestingly, it was possible to develop both viscous macroemulsions and fluid nanoemulsions with the same chemical composition merely by slight alteration of the production process. Optical light microscopy and cryo transmission electron microscopy (TEM) revealed that the sucrose ester led to the formation of an astonishing hydrophilic network at a concentration of only 5% w/w in the macroemulsion system. A small number of more finely structured aggregates composed of surplus surfactant were likewise detected in the nanoemulsions. These discoveries offer interesting possibilities to adapt the low viscosity of fluid O/W nanoemulsions for a more convenient application. Moreover, a simple and rapid production method for skin-friendly creamy O/W emulsions with excellent visual long-term stability is presented. It could be shown by franz-cell diffusion studies and in vitro tape stripping that the microviscosity within the semi-solid formulations was apparently not influenced by their increased macroviscosity: the release of three model drugs was not impaired by the complex network-like internal structure of the macroemulsions. These results indicate that the developed semi-solid emulsions with advantageous application properties are highly suitable for the unhindered delivery of lipophilic drugs despite their comparatively large particle size and high viscosity.

Enhancement of stability and skin permeation by sucrose stearate and cyclodextrins in progesterone nanoemulsions.

Lecithin-based nanoemulsions are colloidal drug delivery systems which offer fundamental advantages in topical therapy, such as excellent skin permeation of lipophilic drugs; however, their physicochemical long-term stability is usually rather poor without the use of additional synthetic surfactants such as polysorbates. In a novel approach negatively and positively charged formulations were developed without the use of conventional synthetic surfactants. Natural substances such as sucrose esters and different cyclodextrins were additionally used as stabilising agents. Emphasis was laid on optimisation of the homogenisation process and formulation properties. The optimised formulations were tested for their potential as drug delivery systems for progesterone. Furthermore, crucial formulation parameters such as particle size and zeta potential were monitored for more than a year. In this context, the effect of the natural excipients sucrose stearate and cyclodextrins alpha, beta and gamma on in vitro skin permeation was investigated; the influence of the positive particle surface charge induced by incorporation of the cationic phytosphingosine was evaluated as well. The results showed that in particular the cyclodextrins seemed to induce fundamental changes in formulation microstructure as confirmed by cryo TEM, thus leading to remarkably increased skin permeation rates of progesterone compared to the control.

Atomic force microscopy applied to study macromolecular content of embedded biological material.

We demonstrate that atomic force microscopy represents a powerful tool for the estimation of structural preservation of biological samples embedded in epoxy resin, in terms of their macromolecular distribution and architecture. The comparison of atomic force microscopy (AFM) and transmission electron microscopy (TEM) images of a biosample (Caenorhabditis elegans) prepared following to different types of freeze-substitution protocols (conventional OsO4 fixation, epoxy fixation) led to the conclusion that high TEM stainability of the sample results from a low macromolecular density of the cellular matrix. We propose a novel procedure aimed to obtain AFM and TEM images of the same particular organelle, which strongly facilitates AFM image interpretation and reveals new ultrastructural aspects (mainly protein arrangement) of a biosample in addition to TEM data.