Frank A. Müller

The role of prenucleation clusters in surface-induced calcium phosphate crystallization

Unravelling the processes of calcium phosphate formation is important in our understanding of both bone and tooth formation, and also of pathological mineralization, for example in cardiovascular disease. Serum is a metastable solution from which calcium phosphate precipitates in the presence of calcifiable templates such as collagen, elastin and cell debris. A pathological deficiency of inhibitors leads to the uncontrolled deposition of calcium phosphate. In bone and teeth the formation of apatite crystals is preceded by an amorphous calcium phosphate (ACP) precursor phase. ACP formation is thought to proceed through prenucleation clusters--stable clusters that are present in solution already before nucleation--as was recently demonstrated for CaCO(3) (refs 15,16). However, the role of such nanometre-sized clusters as building blocks for ACP has been debated for many years. Here we demonstrate that the surface-induced formation of apatite from simulated body fluid starts with the aggregation of prenucleation clusters leading to the nucleation of ACP before the development of oriented apatite crystals.

Cellular Ti-6Al-4V structures with interconnected macro porosity for bone implants fabricated by selective electron beam melting.

Selective electron beam melting (SEBM) was successfully used to fabricate novel cellular Ti-6Al-4V structures for orthopaedic applications. Micro computer tomography (microCT) analysis demonstrated the capability to fabricate three-dimensional structures with an interconnected porosity and pore sizes suitable for tissue ingrowth and vascularization. Mechanical properties, such as compressive strength and elastic modulus, of the tested structures were similar to those of human bone. Thus, stress-shielding effects after implantation might be avoided due to a reduced stiffness mismatch between implant and bone. A chemical surface modification using HCl and NaOH induced apatite formation during in vitro bioactivity tests in simulated body fluid under dynamic conditions. The modified bioactive surface is expected to enhance the fixation of the implant in the surrounding bone as well as to improve its long-term stability.

Asymmetric pores in a silicon membrane acting as massively parallel brownian ratchets.

The brownian motion of mesoscopic particles is ubiquitous and usually random. But in systems with periodic asymmetric barriers to movement, directed or ‘rectified’ motion can arise and may even modulate some biological processes. In man-made devices, brownian ratchets and variants based on optical or quantum effects have been exploited to induce directed motion, and the dependence of the amplitude of motion on particle size has led to the size-dependent separation of biomolecules. Here we demonstrate that the one-dimensional pores of a macroporous silicon membrane, etched to exhibit a periodic asymmetric variation in pore diameter, can act as massively parallel and multiply stacked brownian ratchets that are potentially suitable for large-scale particle separations. We show that applying a periodic pressure profile with a mean value of zero to a basin separated by such a membrane induces a periodic flow of water and suspended particles through the pores, resulting in a net motion of the particles from one side of the membrane to the other without moving the liquid itself. We find that the experimentally observed pressure dependence of the particle transport, including an inversion of the transport direction, agrees with calculations of the transport properties in the type of ratchet devices used here.

Hydroxyapatite formation on alkali-treated titanium with different content of Na+ in the surface layer

Titanium can form a bone-like apatite layer on its surface in SBF when it is treated in NaOH. When pre-treated titanium is exposed to SBF, the alkali ions are released from the surface into the surrounding fluid. The Na+ ions increase the degree of supersaturation of the soaking solution with respect to apatite by increasing pH. On the other hand, the released Na+ cause an increase in external alkalinity that triggers an inflammatory response and leads to cell death. Therefore, it would be beneficial to decrease the release of Na+ into the surrounding tissue. The purpose of this study was to evaluate the hydroxyapatite formation on alkali-treated titanium with different content of Na+ in the surface layer. Using SEM, gravimetric analysis and measurement of calcium and phosphate concentration, it was found that the rate of apatite formation was not significantly influenced by a lower amount of Na+ in the surface layer. Titanium with the lowest content of Na+ could be more suitable for implantation in the human body. The amount of alkali ions released in the surrounding tissue is lower and the rate of apatite formation is identical to titanium with the highest content of Na+ in the surface layer.

Ion adsorption behaviour of hydroxyapatite with different crystallinities

This study aimed to correlate crystallinity of hydroxyapatite (HA) with the ion adsorption behaviour of the material. Hydroxyapatite powders of various crystallinities (X(c)) and specific surface area (SSA) were prepared by precipitation following heat treatment. Adsorption experiments were carried out by using (i) multi-component ion solutions containing a broad range of light and heavy ions to study competitive adsorption and (ii) lead and zinc solutions with concentrations up to 250 ppm to determine the adsorption isotherms of the material. While as-prepared HA powders of low crystallinity (X(c)=0%) and a high SSA of 170 m(2)/g showed quantitative removal for divalent Pb, Zn, Be, U, Bi, V, Al, Cu and Ga ions, calcined powders with higher crystallinity (X(c)=65-95%) and lower SSA between 5 and 30 m(2)/g led to a quantitative removal only for a few elements (Pb, Bi, Ga). The time and concentration dependant ion removal capacity for Pb(2+) and Zn(2+) single element solutions showed quantitative removal even after short immersion times of less than 10 min for as-prepared HA powders. XRD analysis of the powders after ion adsorption revealed the presence of pyromorphite (Pb(5)(PO(4))(3)OH) and hopeite (Zn(3)(PO(4))(2)) phases, respectively.

The Biopolymer Bacterial Nanocellulose as Drug Delivery System: Investigation of Drug Loading and Release using the Model Protein Albumin

Although bacterial nanocellulose (BNC) has reached enormous interest for biomedical applications because of its outstanding material properties, investigations about its potential as drug delivery system are very rare. In the present study, for the first time, the applicability of BNC as drug delivery system for proteins using serum albumin as model drug was systematically investigated. Additionally, never-dried BNC was compared with freeze-dried BNC. For both types of BNC, a dependency of concentration, temperature, time, and preswelling for albumin loading and release could be demonstrated. These findings indicated an overlay of diffusion- and swelling-controlled processes, which could be confirmed by Ritger-Peppas equation. Freeze-dried samples showed a lower uptake capacity for albumin than native BNC, which was found to be related to changes of the fiber network during the freeze drying process as demonstrated by electron microscopy and protein staining experiments. The integrity and biological activity of proteins could be retained during the loading and release processes, which was demonstrated by gel electrophoresis and the use of luciferase as biologically active molecule. In conclusion, hydrophilicity, high biocompatibility, and controllable drug loading and release render BNC an innovative and attractive biopolymer for controlled drug delivery.

Hydroxylation of dental zirconia surfaces: characterization and bonding potential.

Bioinert zirconia surfaces exhibit a low chemical bonding potential to resin-based luting agents. The aim was to hydroxylate dental zirconia surfaces and to examine tensile bond strength using commercial luting agents. The measured bond strength was compared with established mechanical conditioning techniques. Five acidic and one alkaline hydroxylation pretreatments were applied and compared with air abrasion and tribochemical silica coating. For the chemical characterization of hydroxyl groups and hydroxyl value, zirconia powders were used, chemically modified, and analyzed using Fourier-transformed infrared spectroscopy and a titrimetric method according to the ISO 4629 standard. All acidic pretreatment procedures exhibited increased hydroxyl values. The highest values were recorded after treatment with phosphoric acid or Piranha solution. Tensile bond strength was examined in a universal testing machine using the commercial dual-cure luting agents Multilink (Ivoclar, Liechtenstein) and Panavia F2.0 (Kuraray, Japan). Surface hydroxylation with Piranha solution in combination with the luting agent Multilink led to a bond strength of 12.47 +/- 3.25 MPa. Tribochemical silica-coated/silanized zirconia surfaces with Multilink produced the highest bond strength of 19.33 +/- 3.65 MPa. Using the luting agent Panavia F2.0, statistically homogenous values for the untreated (11.60 +/- 1.68 MPa) and for the hydroxylated surface (12.46 +/- 3,81 MPa) were measured. Bioinert zirconia surfaces were successfully hydroxylated in terms of tensile bond strength. Resin bonding with Multilink can be strongly increased by acidic treatment with Piranha solution. Bonding with Panavia F2.0 is not affected by hydroxylation, which is likely due to the incorporation of specific functional monomers.

Active wound dressings based on bacterial nanocellulose as drug delivery system for octenidine

Although bacterial nanocellulose (BNC) may serve as an ideal wound dressing, it exhibits no antibacterial properties by itself. Therefore, in the present study BNC was functionalized with the antiseptic drug octenidine. Drug loading and release, mechanical characteristics, biocompatibility, and antimicrobial efficacy were investigated. Octenidine release was based on diffusion and swelling according to the Ritger-Peppas equation and characterized by a time dependent biphasic release profile, with a rapid release in the first 8h, followed by a slower release rate up to 96 h. The comparison between lab-scale and up-scale BNC identified thickness, water content, and the surface area to volume ratio as parameters which have an impact on the control of the release characteristics. Compression and tensile strength remained unchanged upon incorporation of octenidine in BNC. In biological assays, drug-loaded BNC demonstrated high biocompatibility in human keratinocytes and antimicrobial activity against Staphylococcus aureus. In a long-term storage test, the octenidine loaded in BNC was found to be stable, releasable, and biologically active over a period of 6 months without changes. In conclusion, octenidine loaded BNC presents a ready-to-use wound dressing for the treatment of infected wounds that can be stored over 6 months without losing its antibacterial activity.

Zirconia nanoparticles prepared by laser vaporization as fillers for dental adhesives.

Zirconia nanoparticles prepared by laser vaporization were incorporated into the primer or into the adhesive of a commercial adhesive system in order to evaluate its effect on bond strength to dentin. Zirconia nanoparticles (20-50nm) were prepared using a particular laser vaporization technique and incorporated into the primer (P) or into the adhesive (A) of the Adper Scotchbond Multi-Purpose (SBMP) system at 5, 10, 15 and 20wt.% by means of mechanical mixing (stirring) and ultrasonication. Control (unfilled) and experimental groups (filled) were applied, according to the manufacturers instructions, onto flat mid-coronal human dentin. Composite crowns were built up, stored in distilled water for 24h at 37°C and cut into 0.65±0.05mm² beams following a non-trimming microtensile technique. Specimens were fractured in tension using a universal testing machine (Zwick) and examined by scanning electron microscopy for fractographic analysis. Microtensile bond strength (μTBS) data were analyzed using a two-way ANOVA and modified LSD test at α=0.05. Analysis of the nanofiller distribution and ultramorphological characterization of the interface were performed by transmission electron microscopy (TEM). Zirconia nanoparticle incorporation into the primer or into the adhesive of SBMP significantly increased μTBS to dentin. Filler concentration only affected μTBS significantly in the P group. Statistically significant differences between groups P and A occurred only at 20wt.% filler content, with a significantly higher μTBS in group P. TEM micrographs revealed nanoparticle deposition on top of a hybrid layer when incorporated into the primer, whereas they remained dispersed through the adhesive layer in group A. Zirconia nanoparticles incorporation into SBMP increased bond strength to dentin by reinforcing the interface adhesive layer. Nanofiller incorporation into the primer solution showed a tendency of increasing bond strength with increasing concentration. At high concentrations (20wt.%) nanofiller incorporation was more efficient in increasing bond strength if incorporated in the primer solution. Adding nanofillers to the primer and to the adhesive solutions resulted in different particle distributions at the interface.

Quantitative analysis of the grain morphology in self-assembled hexagonal lattices.

We present a methodology for the analysis of the grain morphology of self-ordered hexagonal lattices and for the quantitative comparison of the quality of their grain ordering based on the distances between nearest neighbors and their angular order. Two approaches to grain identification and evaluation are introduced: (i) color coding the relative angular orientation of hexagons containing a central entity and its six nearest neighbors, and (ii) incorporating triangles comprising three nearest neighbors into grains or repelling them from grains based on deviations of the side lengths and the internal angles of the triangles from those of an ideal equilateral triangle. A spreading algorithm with tolerance parameters allows single grains to be identified, which can thus be ranked according to their size. Hence, grain size distributions are accessible. For the practical evaluation of micrographs displaying self-ordered structures, we suggest using the size of the largest identified grain as a quality measure. Quantitative analyses of grain morphologies are key to the systematic and rational optimization of the fabrication of self-assembled materials.