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Hydroxyapatite (HA), Ca 10 (PO 4 ) 6 (OH) 2 , is a key material used to manufacture thin, bioactive coatings on metal implants for hard tissue regeneration. Technical challenges in their fabrication include control of the coating structure and chemical composition, as well as managing surface activation via ionic substitution. In this study, 15 – 216 nm-thick zinc-doped HA (ZnHA) thin films were grown via magnetron sputtering at room temperature. The effect of substituting Zn 2+ for Ca 2+ on the ultrastructures of the HA films was analyzed during several stages of film growth. Infrared vibrational scattering scanning near-field optical microscopy (s-SNOM) using the ultra-broad-band IR beam from synchrotron radiation was compared with Fourier transform infrared spectroscopy in attenuated total reflectance (FTIR/ATR) to reveal the close relationships between local topography and the chemical compositions of the film surface and the bulk of the film. The energy delivered via ionic bombardment induced the formation of complex structures: island-shaped nanoparticles of amorphous zinc-doped calcium phosphate, coalescent islands, disordered non-stoichiometric ZnHA nanoparticles, and long-range structures of nearly stoichiometric and crystalline ZnHA. Grazing incidence X-ray diffraction (GIXRD) from synchrotron radiation, X-ray photoelectron spectroscopy (XPS), focusing ion beam (FIB), and high-resolution transmission electron microscopy (HRTEM) confirmed Zn substitution into the HA lattice. Zinc was distributed homogeneously within the film structure (1.6 at%) but its concentration increased slightly on the film surface. The bulk of the film consisted of crystalline, columnar ZnHA domains oriented perpendicular to the substrate surface while the regions near the film/substrate interface were preferentially disordered and non-stoichiometric.

Many biomaterials are used for Bone Morphogenetic Proteins (BMPs) delivery in bone tissue engineering. The BMP carrier system&rsquo, s primary function is to hold these growth factors at the wound&rsquo, s site for a prolonged time and provide initial support for cells to attach and elaborate the extracellular matrix for bone regeneration. This study aimed to evaluate the nanostructured carbonated hydroxyapatite microspheres (nCHA) as an rhBMP-2 carrier on rats calvaria. A total of fifteen male Wistar rats were randomly divided into three groups (n = 5): clot (control group), rhBMP-2 associated with collagen membrane (COL/rhBMP-2) or associated with the microspheres (nCHA/rhBMP-2). After 45 days, the calvaria defect samples were evaluated through histological, histomorphometric, and SR-&micro, CT analyses to investigate new-formed bone and connective tissue volume densities. The descriptive histological analysis showed that nCHA/rhBMP-2 improved bone formation compared to other groups. These results were confirmed by histomorphometric and SR-&micro, CT analysis that showed substantially defect area filling with a higher percentage of newly formed (36.24 &plusmn, 6.68) bone than those with the COL/rhBMP-2 (0.42 &plusmn, 0.40) and Clot (3.84 &plusmn, 4.57) (p &lt, 0.05). The results showed that nCHA is an effective carrier for rhBMP-2 encouraging bone healing and an efficient alternative to collagen membrane for rhBMP-2 delivery.

The adaptation of trabecular bone microstructure to mechanical loads has been intensively investigated. However, loading-unrelated aspects of trabecular architecture remain unclear. We used synchrotron radiation-based X-ray microtomography to study the 3D microarchitecture of newly formed trabecular tissue in a defect produced in the cortical region of the rat tibia diaphysis, in the absence (7, 14, and 21 days) or the presence (21 days) of carbonated hydroxyapatite/alginate (cHA) microspheres. This work provides the first evidence that the woven bone trabecular network, formed during the healing process, displays a well-organized 3D microarchitecture consisting of nodes with 3 (3-N), 4 (4-N) and 5 (5-N) connecting trabeculae, with a mean relative abundance of (3-N)/(4-N)/(5-N) = 66/24/7, for the analyzed periods. The measured inter-trabecular angles (ITA) distribution presented a Gaussian profile, with mean value at 115° for 3-N nodes, and 105° for 4-N nodes, close to the angles of idealized 3D regular structures (120° and 109.5°, respectively). Changes in the dispersion of ITA distribution suggested that a highly symmetric trabecular fabric organized under tensegrity principles is formed early during the bone healing process. Post-implantation, cHA disaggregated into multiple fragments (~20–400 μm), stimulating osteoconduction and bone growth toward the interior of the medullary cavity. The presence of biomaterials in bone defects affected the trabecular dimensions; however, it did not interfere with the formation of geometrical motifs with topological parameters similar to those found in the sham-defects. Statement of Significance The trabecular bone microstructure enables the tissue to meet the necessary mechanical and functional demands. However, the process of trabecular microarchitecture formation during healing, in the absence or presence of a bone graft, is not yet well understood. This work demonstrated that, from the beginning of its formation in cortical bone defects, the woven-bone trabecular network is spatially organized according to the principle of tensegrity. This microarchitecture is comprised of highly symmetric geometric motifs and is an intrinsic characteristic of trabecular growth, regardless of hierarchical scale or mechanical stimulation. The addition of a biodegradable nanostructured calcium phosphate graft did not disrupt trabecular microarchitecture; however, graft biodegradation should be controlled to optimize the reproduction of intrinsic trabecular motifs throughout the defect.

The aim of this study was to fabricate and characterize dental composites containing hydroxyapatite nanoparticles (HApNPs). Four dental composites were produced from the same organic matrix (70 wt% Bis-GMA and 30 wt% TEGDMA), with partial replacement of BaBSi particles (65 wt%) by HApNPs in the following concentrations (wt%): E0 (0) – control, E10 (10), E20 (20) and E30 (30). Ca2+ and PO43− release was evaluated in solutions with different pHs (4, 5.5, and 7) using atomic emission spectroscopy with microwave-induced nitrogen plasma while the enamel remineralization potential was evaluated in caries-like enamel lesions induced by S. mutans biofilm using micro-CT. The following properties were characterized: degree of conversion (DC%), microhardness (KHN), flexural strength (FS), elastic modulus (EM) and translucency (TP). The higher the HApNPs content, the higher the Ca2+ and PO43− release. The ions release was influenced by pH (4 > 5.5 > 7) (p E10) (p

Made available in DSpace on 2020-12-12T02:02:28Z (GMT). No. of bitstreams: 0 Previous issue date: 2020-07-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Fundação de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ) The apoptosis-associated Speck-like protein containing a caspase-1 recruitment domain (ASC), present in inflammasomes, regulates inflammation events and is involved in osteogenic phenotype. Nevertheless, its function in bone repair induced by bone substitute biomaterials is unclear. This study aimed to unveil the role of ASC on osteoprogenitor and tissue response to stoichiometric-hydroxyapatite (HA), nanostructured carbonated-hydroxyapatite (CHA), and CHA containing 5% Strontium (SrCHA), characterized previously by XRD, uXRF-SR, and FTIR spectroscopy implants. Thereafter, conditioned media by the biomaterials were used later to treat pre-osteoblasts and an osteogenic stimulus was shown in response to the materials, with higher expression of Runx2, Osterix, ALP, and Collagen 1a1 genes, with significant involvement of inflammatory-related genes. Thus, to better address the involvement of inflammasome, primary cells obtained from both genotypes [Wild-Type (WT) and ASC Knockout (ASC-KO) mice] were subjected to conditioned media up to 7 days, and our data reinforces both HA and CHA induces lower levels of alkaline phosphatase (ALP) than SrCHA, considering both genotypes (p < 0.01), and ASC seems contribute with osteogenic stimulus promoted by SrCHA. Complimentarily, the biomaterials were implanted into both subcutaneous and bone defects in tibia. Histological analysis on 28 days after implantation of biomaterials into mice's subcutaneous tissue revealed moderate inflammatory response to them. Both histomorphometry and μCT analysis of tibias indicated that the biomaterials did not reverse the delay in bone repair of ASC KO, reinforcing the involvement of ASC on bone regeneration and bone de novo deposition. Also, the bone density in CHA was >2-fold higher in WT than ASC-KO samples. HA was virtually not resorbed throughout the experimental periods, in opposition to CHA in the WT group. CHA reduced to half-area after 28 days, and the bone deposition was higher in CHA for WT mice than HA. Taken together, our results show that biomaterials did not interfere with the healing pattern of the ASC KO, but CHA promoted higher bone deposition in the WT group, probably due to its greater biodegradability. These results reinforce the importance of ASC during bone de novo deposition and healing. Oral Surgery Department Universidade Veiga de Almeida Oral Surgery Department and Clinical Research Laboratory in Dentistry Universidade Federal Fluminense Oral Diagnosis Department Universidade Federal Fluminense Department of Applied Physics Centro Brasileiro de Pesquisas Físicas CBPF Department of Biotechnological and Applied Clinical Sciences University of L'Aquila UNESP – São Paulo State University Institute of Biosciences of Botucatu Department of Chemistry and Biochemistry Clinical Research Laboratory in Dentistry Universidade Federal Fluminense Directory of Life Sciences Applied Metrology Instituto Nacional de Metrologia Qualidade e Tecnologia (INMETRO) Oral Surgery Department Universidade Iguaçu UNESP – São Paulo State University Institute of Biosciences of Botucatu Department of Chemistry and Biochemistry CAPES: 001 FAPERJ: 2014/22689-3

X-ray photoelectron spectroscopy (XPS) is one of the main tools for hydroxyapatite (HA) surface characterization in developing materials for biomedical and heterogeneous catalysis. Despite the XPS technique's potential to correlate binding energies with existing photo-emitter sites on near-surfaces, few previous studies analyzed this aspect for HA and metal-substituted HA surfaces. In this work, we theoretically reconstructed the XPS spectra of stoichiometric HA and lead-substituted hydroxyapatite (PbCaHA, Ca10-xPbx(PO4)6(OH)2; x = 2, 10) using a first-principles linear combination of atomic orbitals embedded cluster approach and periodic supercell band structures within the framework of Density Functional Theory (DFT). We take into account photoemission lines contributions from Ca(1), Ca(2), Pb(1) and Pb(2) sites located on surface and near-surface depths (up to ∼15 A) along the (0 0 1) and (1 0 0) surfaces. The calculated DFT spectra of HA and PbCaHA were compared with high-resolution XPS spectra previously characterized by synchrotron X-ray diffraction (XRD), Fourier transforms infrared spectroscopy (FTIR), high-resolution transmission electron microscopy (HRTEM) and electron energy-loss spectroscopy (EELS). A combined theoretical and experimental approach enables decoding of the complex structure of HA and PbCaHA in XPS spectra. It was found that XPS binding energies profiles depend significantly on photo-emitters from near-surface sites and surface crystallographic orientation. The main Ca 2p3/2 envelope peak in HA is predominantly from Ca(1) and Ca(2) sites (∼347.4 eV), while the weaker peak is due to the Ca(2) site only (∼345.0 eV). Variations on HA nanoparticle morphology could be a critical factor for changes in XPS binding energies' profile.

Some biomaterials can be used to promote tissue repair process. The biological substitutes (biomaterials such as hydroxyapatite beads) can be used with some advantages and purpose of mimicking responses to on-site repair of the injured bone. The objective of this study was to evaluate the osteogenic potential of the biomaterial composed of hydroxyapatite and alginate in place of the critical defect. bioceramic samples stoichiometric hydroxyapatite was produced by the precipitation method, wet method with ion molar ratio of Ca 10 (PO 4) 6 (OH) 2, in which the Ca / P ratio was equal to 1.67. The reaction conditions were favorable to the composition of a biomaterial with crystalline phase. The synthesis of the biomaterial composed of hydroxyapatite and alginate microspheres (HAAlg5%; 200 ø 425mm) was obtained from two primary solutions with the aim of, in optimal reactive conditions, to form the precipitate. After synthesis the microspheres were implanted into the defect site. The potential effects of using HAAlg5% and the application of vibratory waves in the critical defect repair were unknown and the results described in this study are promising, considering the systemic therapy and at the site of injury. The biomaterial used promoted repair the injured tissue.

Calcium phosphate (CP) has been used as biomaterial to regenerate bone defects. In order to improve its biological properties, substitutions in the calcium phosphate chemical structure have been investigated. Zinc has stimulatory effects on in vitro and in vivo bone formation and an inhibitory effect on in vitro osteoclastic bone resorption. The aim of this systematic review (SR) was to assess the effect of zinc incorporation into calcium phosphate on bone repair. This SR was performed in accordance with the PRISMA guidelines and Preclinical Systematic Review & Meta-analysis Facility. An electronic search was conducted using the MEDLINE/PubMed, Cochrane, Scopus, and Scielo databases. Eligibility criteria included only animal studies that compared the effect of zinc incorporation into CP–based materials on bone repair. The selection process yielded 18 studies among 335 retrieved. The ARRIVE guidelines checklist was used to assess the articles’ quality. It was observed that zinc incorporation into CP did not affect its biocompatibility, tends to reduce the resorption rate and was considered to be osteoconductive. Most of the manuscripts suggested that zinc incorporation can enhance bone repair. The results indicate that zinc-doped CP has potential for increasing bone repair, being an interesting option for bone regeneration biomaterial.

The non-equilibrium electro-thermodynamic properties of magnetized plasma produced by a modified radio-frequency magnetron sputtering system were investigated in detail. The plasma was analyzed using Hall, Langmuir, and Faraday probes and optical emission spectroscopy, and numerical simulations were conducted to determine the electric and magnetic field profiles. It is found that the occurrence of lower hybrid resonance and excitation of helicon waves are satisfied locally along the plasma profile (Z). This unusual effect in magnetron sputtering systems plays an important role in the increase of the electrons and ions temperature improving the quality of the films produced. Films of complex structure as hydroxyapatite that have biomedical applications were produced along the Z to confirm our plasma results. The films were characterized by performing grazing incidence X-ray diffraction using synchrotron radiation, X-ray photoelectron spectroscopy and atomic force microscopy. The microstructure analysis of hydroxyapatite films produced at different Z revealed that films with high crystallinities and with ideal stoichiometries (Ca/P = 1.67 ± 0.05) were produced within a magnetic cusp region (26 mm

Internalization of hydroxyapatite nanoparticles in SAOS-2 osteoblasts for 2 and 24 h was investigated in vitro using 5 and 50 µg/mL nanoparticles in culture medium. No cytotoxic effects were observed in a PrestoBlue viability assay. Focused ion beam-scanning electron microscopy and transmission electron microscopy were used to study nanoparticle trafficking inside cells and to characterize the physicochemical properties of the remodeled nanoparticles. Nanoparticles were actively internalized by cells and maintained in intracellular membrane-bound compartments. Dissolution of hydroxyapatite nanoparticles was observed inside phagolysosome in all samples. After 24 h of internalization in cell culture assays, reprecipitation of calcium phosphate minerals was observed in membrane-bound compartments in 5 and 50 µg/mL samples. Compared to the original nanoparticles, the reprecipitated calcium phosphate phase presented a different morphology, structure, and chemical composition. Two sample preparation methods were used and confirmed that reprecipitation of the calcium phosphate crystallites occurred in the intracellular environment and not during electron microscopy sample preparation. Reprecipitation of calcium phosphate prevented the release of large amounts of calcium and phosphate ions inside the cells. This phenomenon may be linked to physiological processes in the cell that control the concentration and trafficking of intracellular calcium ions, which are highly controlled by cells. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 428-439, 2018.

Insulin-loaded calcium phosphate nanoparticles have been proposed as a potential drug delivery system for the oral treatment of diabetes and to stimulate bone cell proliferation and bone mineralization. The kinetics of insulin incorporation onto hydroxyapatite (HA) and Sr (SrHA)- and Zn (ZnHA)-substituted hydroxyapatite nanoparticles was investigated using X-ray photoelectron spectroscopy (XPS), Fourier transform infrared (FTIR) spectroscopy, zeta potential measurements and circular dichroism (CD) spectroscopy. The increase in insulin concentration on HA, SrHA and ZnHA was a typical physical adsorption process controlled by electrostatic forces and followed a Freundlich isotherm model. Zn substitution enhanced the capacity of the apatite surface to adsorb insulin, whereas Sr substitution inhibited insulin uptake. The surface stoichiometry and mesopore specific area induced by Zn and Sr substitution are proposed as the main causes of the difference in insulin adsorption. Despite the weak interaction between insulin and the apatite surface, the CD spectra revealed a decrease in the insulin ellipticity when the protein was adsorbed on the HA, SrHA and ZnHA nanoparticles. A reduction in alpha-helical structures and an increase in beta sheets were observed when insulin interacted with the HA surface. A less pronounced effect was found for ZnHA, for which a subtle decrease in alpha-helical structures was followed by an increase in turn structures. Interaction with the SrHA surface did not change the native insulin conformation. In vitro cell culture experiments lasting 24h using F-OST stromal cells showed that the insulin loaded on HA and ZnHA did not affect cell proliferation but the insulin loaded on SrHA improved cell proliferation. These results suggest that the stability of the native protein conformation is an important factor to consider when cells interact with insulin adsorbed on metal-substituted HA surfaces.

Drug delivery technology is a promising way to enhance the therapeutic efficacy of drugs. The purpose of this study is to evaluate the physical and chemical properties of hydroxyapatite ceramic microspheres loaded with doxycycline (HADOX), their effects on in vitro osteoblast viability, and their antimicrobial activity, and to determine the effects of DOX on the healing of rat sockets after tooth extraction. The internal microsphere porosity was sensitive to the treatment used to adsorb DOX onto microsphere surface; HA microspheres without DOX presented 26% of pores, whereas HADOX0.15 microspheres presented 52.0%. An initial drug release of 49.15 μg/ml was observed in the first 24 hr. The minimal inhibitory concentration (MIC) tested against Enterococcus faecalis demonstrated that bacterial growth was inhibited for up to 7 days. Results of cell viability and cell proliferation did not indicate statistical differences in the metabolic activity of HADOX samples relative to HA without DOX microspheres (p > .05). After 1 week, a discreet inflammation reaction was observed in the control group, and after 6 weeks, newly-formed bone was observed in the HADOX0.15 (p < .05). The HADOX did not interfere in the bone repair and controlled the early inflammatory response. HADOX could be a promising biomaterial to promote bone repair in infected sites.

Hydroxyapatite (HA) is the main constituent of bone mineral in vertebrates and is widely used as a biomaterial in biomedical studies and medical clinics. The synthesis of this calcium phosphate phase is a crucial factor in its potential use in various applications as well as in understanding the nucleation and growth processes in biological mineralization (biomineralization). In this work, a sequential analysis of the different steps of HA synthesis in aqueous media was performed by using cryo-transmission electron microscopy (Cryo-TEM), which allows preservation of the specimens at different stages of the synthesis process, preventing their modification during drying. X-ray energy dispersive spectroscopy and electron diffraction, in addition to high-resolution imaging, were performed on vitrified samples. We showed that drying a specimen from the synthesis solution induces the crystallization of nanoparticles into an HA phase after 30 min of reaction, while when analyzed by Cryo-TEM, crystalline nanoparticles were detected only after 15 h. Calcium phosphate nanoparticles presented three different morphologies: irregular shapes characterized by low contrast in TEM (5 min), isotropic shapes with well-defined contrast (10 min) and needle-like shapes (after 30 min). The long axis of the HA needle-like nanoparticles can be associated with the c-axis direction of the hexagonal unit cell, which can thus be assigned to the preferential crystallization direction. However, a diffraction signal of the nanoparticles in vitreous ice that “mimics” the original solution was detected by Cryo-TEM only after 15 h of synthesis. These apparently controversial results are discussed in this manuscript considering various aspects of the use of Cryo-TEM in relation to the current concepts of calcium phosphate nucleation and growth.

The knowledge on human tissue is very important to recognize desirable properties of biomaterials. Host cells, extracellular matrix, integrated vessels and interstitial fluids create a complex and dynamic system able to regenerate and respond to environmental stimuli. Myoglobin is a protein with most of α-helices on its secondary structure, and responsible for oxygen binding and release in muscles, by the heme group. This work investigates the Mb adsorption process onto zinc-hydroxyapatite (ZnHA) surface by spectroscopic studies. To do so, ZnHA (0.05 g) was incubated with 4mL of 2mg Mb/mL on phosphate buffer solution pH 6.0 for 24h at 37°C. The FTIR analyses of ZnHA powders before and after protein adsorption provided information concerning the protein content. UV-Vis spectrocopy in the reflectance mode suggested a mixture of MbO2 and Met Mb on lyophilized solid Mb, and the prevalence of MetMb form when Mb was adsorbed on ZnHA sample. The decrease of UV-Vis secondary bands suggests interactions through the Mb heme group and the ZnHA surfaces. Circular Dichroism (CD) spectroscopy indicated the maintenance of the Mb α-helices secondary structure after the adsorption process on ZnHA powders.

The hydroxyapatite (HA) is a biocompatible and bioactive biomaterial used as bone substitute, however, the high crystallinity of HA and consequently its low solubility may be a limitation for its clinical use. In order to improve the biosorption of HA, the partial substitutions in the chemical structure and doping with small amounts of impurities have been study. The objective of this study was to evaluate the biocompatibility of 3% Zinc-containing nanostructured carbonated hydroxyapatite (ZncHA) compared with the carbonated hydroxyapatite (cHA), both synthesized at 37°C and non-sintered, using as control the stoichiometric HA microspheres in subcutaneous of mice. The X-ray Diffraction (XRD) and Vibrational Spectroscopy in Infra Red Fourier Transform (FTIR) were used to characterize the biomaterials. In vivo test was performed in BALB/c mice by implanting of HA, cHA and ZncHA spheres in the subcutaneous tissue for 1, and 9 weeks (n=5). The negative control consisted in incision without material implantation (Sham group). The samples were histological processed to descriptive analysis of biological effect. The microscopic analysis showed a similar granulation reaction between groups at the first experimental period. In 9 weeks there was a time dependent biosorption of cHA compared with other groups. In conclusion, the biomaterials tested were biocompatible and cHA group showed a significant biosorption in comparison with HA and ZncHA groups. The doping of zinc did not influence the biocompatibility of biomaterial, however, change the biosorption response

Nanostructured carbonated hydroxyapatite (nCHA) is a promising biomaterial for bone tissue engineering due to its chemical properties, similar to those of the bone mineral phase and its enhanced in vivo bioresorption. However, the biological effects of nCHA nanoparticles on cells and tissues are not sufficiently known. This study assessed the impact of exposing pre-osteoblasts to suspensions with high doses of nCHA nanoparticles with high or low crystallinity. MC3T3-E1 pre-osteoblasts were cultured for 1 or 7 days in a culture medium previously exposed to CHA nanoparticles for 1 day. Control groups were produced by centrifugation for removal of bigger nCHA aggregates before exposure. Interaction of nanoparticles with the culture medium drastically changed medium composition, promoting Ca, P, and protein adsorption. Transmission Electron microscopy revealed that exposed cells were able to internalize both materials, which seemed concentrated inside endosomes. No cytotoxicity was observed for both materials, regardless of centrifugation, and the exposure did not induce alterations in the release of pro-and anti-inflammatory cytokines. Morphological analysis revealed strong interactions of nCHA aggregates with cell surfaces, however without marked alterations in morphological features and cytoskeleton ultrastructure. The overall in vitro biocompatibility of nCHA materials, regardless of physicochemical characteristics such as crystallinity, encourages further studies on their clinical applications.

A new study about corrosion behavior of CoCrMo alloy used as implant material exposed to the physiological serum and Hank's solution in different temperatures, electrolyte compositions and immersion time were simulated, using open circuit potential (OCP), potentiodynamic polarization curves (PPC) and electrochemical impedance spectroscopy (EIS.) From the results, it can be concluded that CoCrMo biomaterial is influenced by electrolyte composition and temperature, and the passive films were more protective when formed in presence of physiological serum. Comparing the Ecorr for both solutions, it is possible to verify that the Hank's solution presented values more negative for the two studied temperatures (25 and 37 °C), indicating a greater susceptibility to the corrosion process. Regarding the results of potentiodynamic polarization, the ipass, decreased as function of time, indicating a more protective passive film at 25 °C. In regards to the physiological serum, the highest ipass values at 37 ºC evidenced the formation of a less stable passive film. The different behavior of ipass in Hank's solution may be related to the competitiveness between a possible film of D-glucose and oxides of the alloy metals. EIS studies suggest that the film formed in presence of physiological serum is nobler than that with Hank's solution.

In the present work, we investigated the nanostructure of calcium phosphate films formed during the initial stages of deposition (between 30 and 240 s; deposition rate of 3.7 nm/min) by radio-frequency magnetron sputtering. The sputtering deposition parameters and the plasma energy were previously adjusted to produce nearly stoichiometric nanocrystalline hydroxyapatite films after 180 min. Grazing-incidence X-ray diffraction and X-ray photoelectron spectroscopy analyses revealed that the films formed in the initial stages were composed of an amorphous calcium phosphate phase with a Ca/P ratio Ca/P = 1.60 ± 0.12, which is close to that of hydroxyapatite. Using high-resolution transmission electron microscopy, we demonstrated that the films were not a unique amorphous phase. An additional phase composed of dense calcium phosphate nanospheres was found in all periods and investigated. The nanospheres grew in size as the deposition time increased, and small nanospheres were always observed. This observation suggests the continuous formation of nanospheres at the surface of the film. Nanocrystalline regions were revealed by electron diffraction and high-resolution electron microscopy after a deposition of 60 s with a corresponding film thickness of 3.7 nm. Electron energy-loss spectroscopy analysis comparing individual nanospheres to the amorphous calcium phosphate layer showed differences in the oxygen K edge, suggesting a difference in the chemical bonding of the oxygen species. Nanospheres were observed in association with isolated crystals, suggesting that the nanospheres may participate in the crystallization of hydroxyapatite according to a heterogeneous nucleation process.

Various synthetic bone substitutes have been developed to reconstruct bone defects. One of the most prevalent ceramics in bone treatment is hydroxyapatite (HA) that is a useful material as bone substitute, however, with a low rate of biodegradation. Its structure allows isomorphic cationic and anionic substitutions to be easily introduced, which can alter the crystallinity, morphology, biocompatibility, and osteoconductivity. The objective of this study was to investigate the in vitro and in vivo biological responses to strontium-containing nanostructured carbonated HA/sodium alginate (SrCHA) spheres (425

The use of nanotechnology - especially in the area of human health - is increasing every day, with the application of various materials such as hydroxyapatite being amongst the most studied. Consequently, the affinity of hydroxyapatite compatible with so many applications in the human body is evolving cutting edge technology - the result of which is nanoparticles. However, despite these important

A radio-frequency magnetron sputtering technique operating in right-angle geometry (RAMS) with high plasma confinement was revised to produce thin films (15–570 nm) of fluorine-substituted hydroxyapatite, FHA, adapted to be used as nano-coatings for biomedical implants. An electron temperature of T eff ≈ 9.0 eV and a plasma electron density of 1.2 × 10 15 m − 3 assured the nucleation of an amorphous fluorine-substituted hydroxyapatite phase on Si and Ti surfaces. With the aid of a Langmuir probe, the RAMS plasma energy was tuned to control the coating stoichiometry and the ratio between the crystalline and amorphous phases. The energy delivered over time from the bombardment of ions and electrons transformed the amorphous calcium phosphate phase into crystalline fluorine-substituted hydroxyapatite. The crystalline films were obtained at room temperature. The partial substitution of OH − for F − in the HA structure was confirmed by X-ray diffraction using synchrotron radiation in grazing-incidence mode, X-ray photoelectron spectroscopy and attenuated total reflection Fourier transform infrared spectroscopy. High-resolution transmission electron microscopy carried out on cross-section film samples prepared by a focused ion beam (FIB) technique revealed that the film ultrastructure was composed of columnar crystals oriented perpendicularly to the substrate surface. The crystals were connected to the substrate surface by ordered nanolayers, indicating the existence of a continuous binding between the two materials. This work demonstrates that the RAMS technique is able to produce FHA nano-coatings with controlled chemical compositions and structures on metallic implants for clinical applications.

Synthetic hydroxyapatite (HA) is a widely used ceramic biomaterial due to its well described biocompatibility. Some modifications in HA surface can be made to increase surface porosity. Likewise, HA can be modified by the coating with proteins, which may impact on biocompatibility. In this work, we aimed to evaluate the impact of two surface modifications – coating with albumin, a major serum protein, and augmented porosity - over osteoblast adhesion on stoichiometric HA discs. Dense HA discs were obtained by pressing HA powder at 30 KN and sinterization at 1000°C, while porous HA was molded after the addition of alginate (15:1), followed by thermal treatment. Protein adsorption was attained by incubation on 0.5mg/mL bovine serum albumin (BSA) for 24 h at 37°C. MC3T3 mouse preosteoblasts were seeded over both protein-coated and uncoated dense or porous tablets, and cell viability after 24 h was estimated by XTT and Neutral Red assays. Cell density was quantified by fluorescence microscopy. While both dense and porous discs presented altered surfaces after protein treatment, as observed by scanning electron microscopy, porous HA tablets presented significantly higher levels of adsorbed protein. There was a decrease in the concentration of calcium ions in all samples analyzed. Porous HA treated with protein presented significant higher mitochondrial dehydrogenase activity (XTT) than non treated tablets (p

Hydroxyapatite is the main ceramic material that has being used in bone repair, although its physico-chemical and in vivo behavior should be better understood. A method to improve the biocompatibility of HA is the substitution of calcium with divalent cations which enhance mechanic resistance and can modulate inflammatory response against implanted material. In this study we analyzed the biocompatibility of HA doped with one per cent of Zn2+ or Pb2+. The first one has being described as an inflammation modulator and the second would be a model for chronic toxicity assay. Biocompatibility of the both materials was studied in vivo following the ISO 10993-6 standard. HA cylinders (ZnHA, PbHA and stoichiometric HA as positive control) were implanted into subcutaneous tissue of 45 Balb-c mice and after 1, 3 and 9 weeks the animals were euthanized (5 for each experimental condition). Necropsies of the skin containing reactional tissue were removed, fixed in 10% formaldehyde and followed the histological processing for paraffin embedding and staining with Hematoxylin-Eosine and Picrosirius red. Microscopic analysis showed for all groups moderate inflammatory response, decreasing throughout the experimental periods, with ZnHA group showing more intense response. Similar presence of macrophages, fibrosis and angiogenesis were observed among the groups. Thereby, we can conclude that ZnHA and PbHA are biocompatible and not bioresorbable, being the ZnHA potentially indicated as bone graft. Detailed studies are required to better understand the role of PbHA as chronic model for lead toxicity.

Bone tissue in ideal conditions morphofunctional remodeling properly. The bone can be affected by fractures, tumors, hormonal dysfunction, senescence, genetic modifications, among others. In such circumstances, the proper diet, drug use, exercise and other factors are important to the prevention of bone mineral loss. The effect of kinesiotherapy obtained through the application of vibratory waves administered through the vibrating platform, Juvent1000 ® already been established in the prevention of bone mineral density, muscular trophism, among other systems in humans. The response by analyzing bone tissue of bone repair in critical defect is not known in experimental animals and in human clinical. This research evaluated the osteogenic potential critical defect in the calvaria of rats subjected to the application of vibratory waves obtained by vibrating platform and implant in the critical defect of rat calvaria. The bone tissue response was evaluated showed satisfactory results obtained in biological points 15, 45 and 120 days.

It was recently shown that the crystallinity of extracted bone mineral samples from the fin bones of zebrafish could increase over storage time. This would have implications in many studies in which the samples need to be stored until analysis. The aim of this study was to further evaluate if the crystallinity of extracted bone mineral increases over storage time. The extracted mineral was a biological bone-like apatite produced in osteoblast cell cultures. The overall characterization of the mineral was done by energy dispersive X-ray spectroscopy, Fourier transform infrared spectroscopy, and high-resolution transmission electron microscopy. In order to evaluate possible changes in crystallinity over storage time, the same sample was analyzed by X-ray diffraction immediately after mineral extraction and after 18 months of storage. In conclusion, no statistically relevant changes were observed over storage time, although the occurrence of a slight increase in crystallinity could be discussed in the stored mineral sample.

This study investigates the long-term biocompatibility of 0.5 % zinc-containing hydroxyapatite compared with hydroxyapatite. Spheres (425 < ∅ < 550) of both materials were produced by extrusion of ceramic slurry in calcium chloride and characterized by FTIR, XRD, XRF and SEM. Fifteen White New Zealand rabbits were submitted to general anesthesia, and an perforation (2 mm), was made in each tibia, one for zinc-containing hydroxyapatite sphere implantation and one for hydroxyapatite sphere implantation. After 26, 52 and 78 weeks, the animals were euthanized, and the fragment containing the biomaterial was harvested. A 30–50 μm section was obtained for histological analysis in bright field and polarized light. SEM images revealed similar morphologies between the tested biomaterials. Histological analysis showed that there was no difference between the test groups. The morphometric analysis, however, indicates that there was a greater absorption. The materials are biocompatible, promote osteogenesis and that the zinc-containing hydroxyapatite microspheres were absorbed more quickly.

In this work, the synthetic hydroxyapatite (HAP) was studied using different preparation routes to decrease the crystal size and to study the temperature effect on the HAP nano-sized hydroxyapatite crystallization. X-ray diffraction (XRD) analysis indicated that all samples were composed by crystalline and amorphous phases . The sample with greater quantity of amorphous phase (40% of total mass) was studied. The nano-sized hydroxyapatite powder was heated and studied at 300, 500, 700, 900 and 1150 °C. All samples were characterized by XRD and their XRD patterns refined using the Rietveld method. The crystallites presented an anisotropic form, being larger in the (001) direction. It was observed that the crystallite size increased continuously with the heating temperature and the eccentricity of the ellipsoidal shape changed from 2.75 at 300 °C to 1.94, 1.43, 1.04 and 1.00 respectively at 500, 700, 900 and 1150 °C. In order to better characterize the morphology of the HAP the samples were also examined using atomic force microscopy (AFM), infrared spectrometry (IR) and thermogravimetric analysis (TGA).

We report the ultrastructure of regenerated bone surrounding two types of biomaterials: hydroxyapatite–alginate composite and sintered hydroxyapatite. Critical defects in the calvaria of Wistar rats were filled with micrometer-sized spherical biomaterials and analyzed after 90 and 120 days of implantation by high-resolution transmission electron microscopy and Fourier transform infrared attenuated total reflectance microscopy, respectively. Infrared spectroscopy showed that hydroxyapatite of both biomaterials became more disordered after implantation in the rat calvaria, indicating that the biological environment induced modifications in biomaterials structure. We observed that the regenerated bone surrounding both biomaterials had a lamellar structure with type I collagen fibers alternating in adjacent lamella with angles of approximately 90°. In each lamella, plate-like apatite crystals were aligned in the c-axis direction, although a rotation around the c-axis could be present. Bone plate-like crystal dimensions were similar in regenerated bone around biomaterials and pre-existing bone in the rat calvaria. No epitaxial growth was observed around any of the biomaterials. A distinct mineralized layer was observed between new bone and hydroxyapatite–alginate biomaterial. This region presented a particular ultrastructure with crystallites smaller than those of the bulk of the biomaterial, and was possibly formed during the synthesis of alginate-containing composite or in the biological environment after implantation. Round nanoparticles were observed in regions of newly formed bone. The findings of this work contribute to a better understanding of the role of hydroxyapatite based biomaterials in bone regeneration processes at the nanoscale.

High-resolution transmission electron microscopy (HRTEM) and ab initio quantum-mechanical calculations of electronic structure were combined to investigate the structure of the hydroxyapatite (HA) (0 1 0) surface, which plays an important role in HA interactions with biological media. HA was synthesized by in vitro precipitation at 37 °C. HRTEM images revealed thin elongated rod nanoparticles with preferential growth along the [0 0 1] direction and terminations parallel to the (0 1 0) plane. The focal series reconstruction (FSR) technique was applied to develop an atomic-scale structural model of the high-resolution images. The HRTEM simulations identified the coexistence of two structurally distinct terminations for (0 1 0) surfaces: a rather flat Ca(II)-terminated surface and a zig-zag structure with open OH channels. Density functional theory (DFT) was applied in a periodic slab plane-wave pseudopotential approach to refine details of atomic coordination and bond lengths of Ca(I) and Ca(II) sites in hydrated HA (0 1 0) surfaces, starting from the HRTEM model.

This work evaluates the suitability of biphasic calcium phosphate (BCP) granules (β-TCP/HA 70:30) as potential carriers for cell-guided bone therapy. The BCP granules were obtained by synthesis in the presence of wax, thermal treatment, crushing and sieving and characterized by scanning electron microscopy (SEM), X-ray diffraction and Fourier transform infrared spectroscopy. The cytocompatibility of the BCP granules was confirmed by a multiparametric cytotoxicity assay. SEM analysis showed human bone cell adhesion and migration after seeding onto the material. Rat subcutaneous xenogeneic grafting of granules associated to human bone cells revealed a more accentuated moderate chronic inflammatory infiltrate, without signs of a strong xenoreactivity. Histomorphometrical analysis of bone repair of defects in rat skulls (∅ = 5 mm) has shown that bone cell associated-BCP and autograft promoted a two- and threefold increase, respectively, on new bone formation after 45 days, as compared to BCP alone and blood clot. The increase in bone repair supports the suitability the biocompatible (70:30) BCP granules as injectable and mouldable scaffolds for human cells in bone bioengineering.

Hydroxyapatite (HA) is largely used as bone graft; it seems to be the most promising synthetic implant material, mainly because of its excellent biocompatibility. The crystallinity, particle and pore size of HA are important characteristics and can be modified by decreasing basic structural form below 100 nm and have evoked a great amount of attention for improving prevention, diagnosis, and disease treatment, besides improving bone repair through the biodegradation of the material. The aim of this study was to investigate bone mineral content in bone samples with nanohydroxyapatite and HA spheres, specially its spatial distribution on bone microarchitecture. Circular bone defects were made in both tibiae of 12 White New Zeland adult rabbits (Oryctolagus cuniculus) and were divided randomly into five groups – blood clot (control group), sintered HA, non-sintered HA, sintered nanoHA and non-sintered nanoHA – all materials in spherical shape, to smooth handling and accommodation of the surgical bed, and to minimize inflammatory response. The rabbits were euthanatized according to the experimental period of 1 and 4 weeks after surgery. The samples were evaluated by polarized microscopy as well as X-ray microfluorescence in order to account the bone mineral content bone-implant interfaces, through synchrotron radiation. Our results revealed greater newly formed bone area in the non-sintered materials and control groups, and the used technique showed that the amount of calcium of new bone was consistent with both mature bone and HA spheres. In conclusion, the present findings suggest that HA-based biomaterials are biocompatible, promote osteoconduction and favored bone repair. Copyright © 2011 John Wiley & Sons, Ltd.

Carbonated apatite (CHA) is commonly considered a promising synthetic material for biomedical applications in orthopedic and dental surgery due to its biocompatibility, bioresorption and bioactivity. CHA5, CHA37 and CHA90 powders were synthesized from wet method and the DRX patterns showed that the crystallinity and particle size of CHA samples increased proportionally with the synthesis temperature. Powder extracts medium were obtained from each sample to interact with MC3T3-E1 osteoblastics cells. It was evaluated morphology, citotoxicity, pH and Ca2+ concentration. Citotoxicity assays showed high metabolic activity on all samples when compared to control. The polygonal shaped and the confluent monolayer observed in control cells progressively changed according to the crystallinity increase of samples. Cells under mitosis and spindle-like shaped where the main alterations observed. In addition the cell viability could be sensitive to the acid reactivity and crystallinity of carbonated apatite samples.

Cell adhesion, proliferation and differentiation are important specific parameters to be evaluated on biocompatibility studies of candidate biomaterials for clinical applications. Several different methodologies have been employed to study, both qualitative and quantitatively, the direct interactions of ceramic materials with cultured mammal and human cells. However, while quantitatively evaluating cell density, viability and metabolic responses to test materials, several methodological challenges may arise, either by impairing the use of some widely applied techniques, or by generating false or conflicting results. In this work, we tested the inherent interference of different representative calcium phosphate ceramic surfaces (stoichiometric dense and porous hydroxyapatite (HA) and cation-substituted apatite tablets) on different tests for quantitative evaluation of osteoblast adhesion and metabolism, either based on direct cell counting after trypsinization, colorimetric assays (XTT, Neutral Red and Crystal Violet) and fluorescence microscopy. Cell adhesion estimation after trypsinization was highly dependent on the time of treatment, and the group with the highest level of estimated adhesion was inverted from 5 to 20 minutes of exposition to trypsin. Both dense and porous HA samples presented high levels of background adsorption of the Crystal Violet dye, impairing cell detection. HA surfaces also were able to adsorb high levels of fluorescent dyes (DAPI and phalloidin-TRITC), generating backgrounds which, in the case of porous HA, impaired cell detection and counting by image processing software (Image Pro Plus 6.0). We conclude that the choice for the most suitable method for cell detection and estimation is highly dependent on very specific characteristics of the studied material, and methodological adaptations on well established protocols must always be carefully taken on consideration.

In this work myoglobin (Mb) adsorption was carried out in a batch system using hydroxyapatite (HA) powder during 24 hours at 37°C. The HA samples were analyzed after protein adsorption by Fourier Transformed Infrared (FTIR) and UV-Vis Spectroscopy in reflectance mode. UV-Vis analyses showed that Soret and Q bands shifted to lower wavelengths when Mb is associated with HA surface. This result suggests that Mb Heme group is sensitive to the protein adsorption onto HA surface. HA disks coated with myoglobin and cultured with human osteoblastic cells during 7 days showed that cell adhesion and proliferation were not inhibited by the protein coating after 7 days in cell culture.

The corrosion resistance of Ti and Ti–6Al–4V was investigated through electrochemical impedance spectroscopy, EIS, potentiodynamic polarisation curves and UV–Vis spectrophotometry. The tests were done in Hank solution at 25 °C and 37 °C. The EIS measurements were done at the open circuit potential at specific immersion times. An increase of the resistance as a function of the immersion time was observed, for Ti (at 25 °C and 37 °C), and for Ti–6Al–4V (at 25 °C), which was interpreted as the formation and growth of a passive film on the metallic surfaces.

Polycrystalline solid solution of phosphate and vanadate lead calcium apatites Pb 10 -x Ca x( VO 4)1( PO 4)5( OH )2 apatites [x = 0.0, 2.0, 4.0, 6.0, 8.0, and 10.0] was reduced with hydrogen gas at the temperature of 300 and 400°C for 1 h. All these compounds were investigated by XRD and ESR spectroscopy. Only one reduced oxidation state V +4 was detected. Two types of species like isolated vanadium (+4) and clusters of V 4+ had been observed. The stability of the apatites increases and the particle size of the crystal decreases with calcium doping concentrations. As the concentrations of Ca increase the intensity of ESR lines deviates due to the spin–lattice relaxation interactions. The deviation in the intensity of ESR spectra can be attributed as the coexistence of paramagnetic and antiferromagnetic phases in the apatites.

Tissue engineering is a multidisciplinary research area that aims to develop new techniques and/or biomaterials for medical applications. The objective of the present study was to evaluate the osteogenic potential of a composite of hydroxyapatite and alginate in bone defects with critical sizes, surgically made in the calvaria region of rats. The rats (48 adult males), Rattus norvegicus Wistar, were divided into two groups: control (without composite implantation) and experimental (with composite implantation) and analyzed by optical microscopy at the biological time points 15, 45, 90 and 120 d, and transmission electron microscopy 120 d after the implantation of the biomaterial. It was observed that the biomaterial presented a high degree of fragmentation since the first experimental points studied, and that the fragments were surrounded by new bone after the duration of the project. These areas were studied by analytical transmission electron microscopy using an energy dispersive X-ray spectrometer. Three regions could be distinguished: (1) the biomaterial rich in hydroxyapatite; (2) a thin contiguous region containing phosphorus but without calcium; (3) a region of initial ossification containing mineralizing collagen fibrils with a calcium/phosphorus ratio smaller than the particles of the composite. The intermediate region (without calcium or containing very low amounts of calcium), which just surrounded the composite had not been described in the literature yet, and is probably associated specifically to the biocomposite used. The high performance of the biomaterial observed may be related to the fact that alginate molecules form highly anionic complexes and are capable of adsorbing important factors recognized by integrins from osteoblasts. Regions of fibrotic tissue were also observed mainly in the initial experimental points analyzed. However, it did not significantly influence the final result. In conclusion, the biomaterial presents a great potential for application as bone grafts in the clinical area.

Biomaterials for treatment of bone defects have been studied for a long time. Alloplastic materials, mainly hydroxyapatite (HA), are under intense investigation due to its biocompatibility and osteoconductive properties. The HA can be modified by the incorporation of bivalent cations as Zn2+ known as a positive effectors for bone repair. The purpose of this study was to evaluate comparatively the effectiveness of 5% zinc-containing hydroxyapatite (ZnHA) in the treatment of critical size defect (CSD) in rat’s calvaria. CSD (8mm diameter) created in the skull of forty-five Wistar rats were filled with autogenous bone, HA and ZnHA. Skulls harvested after 30, 90 and 180 days were submitted to histological processing for paraffin embedding. Sections of 5 µm-thick stained with hematoxylin and eosin (HE) allowed histomorphometric analysis. The area of neoformed bone increased (p

In this work we applied a new route to synthesize magnetic iron oxide nanoparticles into alginate polymer for future application as drug delivery system activated by magnetic external stimuli. Calcium-alginate was used to encapsulate iron oxide nanoparticles, and as scaffold for particle nucleation and its influence on particles size and magnetic properties were studied. The iron oxide mean sizes were between 4.3 and 9.5 nm. Iron is dispersed throughout the polymer matrix mainly as iron oxide particles, and a small fraction as iron (III) occupying calcium sites in the polymer network. The temperature dependence of the Mossbauer spectra is typical of superparamagnetic particles in agreement with the magnetic susceptibility data.

Millimetric spherical beads of a biocompatible composite were produced from sodium alginate, a natural polysaccharide, and nanostructured hydroxyapatite (HA). It was shown that the composite was effective in the removal of lead ions and lead phosphate nanoparticles from high-contaminated simulated gastric fluid. X-ray diffraction spectroscopy and scanning electron microscopy analyses performed on HA–alginate beads after the Pb2+ uptake showed that nanocrystals of a lead phosphate, [Pb10–xCax(PO4)6Cl2], were precipitated on the bead surface. The cross-linked polymer chain had a double role: (i) keep Pb2+ ions and lead phosphate nanoparticles bounded to the bead surface, preventing their bioavailability in stomach fluid; and (ii) delay HA dissolution in the acidic conditions of the stomach, assuring that an excess of Ca2+ will not be released to simulated gastric fluid. Desorption experiments in simulated enteric fluid revealed that lead remained immobilized in the calcium phosphate phase in the intestinal tract. These results indicate HA–alginate composite as a potential system for heavy metals removal from contaminated gastric and enteric human fluids, minimizing its adsorption by the human body.

Hydroxyapatite (HA), widely used as bone graft, can be modified by the incorporation of bivalent cations (Mg2+ and Zn2+) and its gradual release could favor the bone repair. The purpose of this research was to evaluate the effect of the HA and zinc-containing hydroxyapatite (Zn-HA) in the bone repair in rat calvaria in comparison to autogenous bone. Critical size defect in the calvaria was filled with the graft material and the samples were harvested at the 30, 90 and 180 days. The light microcopy observations showed the biocompatibility of the graft materials. In the Zn-HA group the area of neoformed bone was larger than in the HA group, but smaller than in the autograft. A fibrous connective tissue was more evident around HA granules. It could be conclude that the presence of zinc ions in HA crystal accelerated the osteogenesis and increased the area of newly formed bone in relation to HA.

Crystalline hydroxyapatite thin coatings have been prepared using a novel opposing RF magnetron sputtering approach at room temperature. X-ray diffraction (XRD) analysis shows that all the principal peaks are attributable to HA, and the as-deposited HA coatings are made up of crystallites in the size range of 50-100nm. Fourier transform infrared spectroscopy (FTIR) studies reveal the existence of phosphate, carbonate and hydroxyl groups, suggesting that HA coatings are highly crystalline. To study the biocompatibility of these coatings, murine osteoblast cells were seeded onto various substrates. Cell density counts using fluorescence microscopy show that the best osteoblast proliferation is achieved on an HA RAMS-coated titanium substrate. These experiments demonstrate that RAMS is a promising coating technique for biomedical applications.

A solid solution of phosphate and vanadate lead apatites Pb 10( PO 4)6-x( VO 4)x( OH )2 (x = 0.5, 1.0, 2.0, 3.0, and 6.0) were reduced with hydrogen gas at temperature 300°C for one hour. All these compounds were investigated by XRD and ESR spectroscopy. Only one reduced oxidation state, V 4+, was detected. Two types of species-like isolated vanadium (+ 4) and clusters of V 4+ were observed. The EPR powder spectrum computer simulations were performed using the program Simfonia from Bruker. Spin Hamiltonian parameters were also calculated. As the concentrations of vanadium ions increase to 2.0, the intensity of the ESR lines increases, and x = 3.0 the intensity of the ESR line decreases sharply. After this concentration, i.e. x > 3.0, there is again an increase in the intensity. The variation of the line shape and intensity of the ESR spectra can be attributed to increases in the hopping rate of the charge carriers (polarons) in different apatite compositions.

Hydroxyapatite (HA) films have been widely recognized for their biocompatibility and utility in promoting biointegration of implants in both osseous and soft tissue. Conventional sputtering techniques have shown some advantages over the commercially available plasma spraying method; however, the as-sputtered films are usually amorphous which can cause some serious adhesion problems when post-deposition heat treatment is necessitated. In this paper we present an opposing radio frequency (RF) magnetron sputtering approach for the preparation of HA thin films on various substrates at low power levels. Using this alternative RF magnetron geometry, as-sputtered HA films are nearly stoichiometric, highly crystalline, and strongly bound to the substrate. Post-deposition heat treatment under 800 °C did not result in a marked improvement in the degree of crystallinity of the films. In addition, dissolution experiments show that as-sputtered films are more stable than annealed ones. As-sputtered films grown on amorphous silica substrates exhibit X-ray diffraction (XRD) patterns similar to those of randomly orientated HA powder. On the other hand, films deposited on oriented substrates such as Si(100) and Si(111) show a polycrystalline HA XRD pattern but with some strongly preferred orientations, indicating that HA crystallization is sensitive to the nature of the substrate. The results suggest that the opposing RF magnetron sputtering approach has some potential to produce high quality HA films on metallic implants.

Hydroxyapatite (HA) has been investigated as a delivery system for antimicrobial and antibacterial agents to simultaneously stimulate bone regeneration and prevent infection. Despite evidence supporting the bactericidal efficiency of these HA carriers, few studies have focused on the effect of this association on bone regeneration. In this work, we evaluated the physico-chemical properties of hydroxyapatite microspheres loaded with chlorhexidine (CHX) at two different concentrations, 0.9 and 9.1 $$\upmu {\text{g}}_{\text{CHX}} /{\text{cm}}^{ 2}_{\text{HA}}$$ , and characterized their effects on in vitro osteoblast viability and bone regeneration. Ultraviolet–visible spectroscopy, scanning and transmission electron microscopy associated with energy-dispersive X-ray spectroscopy and electron energy loss spectroscopy were used to characterize the association of CHX and HA nanoparticles. The high CHX loading dose induced formation of organic CHX plate-like aggregates on the HA surface, whereas a Langmuir film was formed at the low CHX surface concentration. Quantitative evaluation of murine osteoblast viability parameters, including adhesion, mitochondrial activity and membrane integrity of cells exposed to HA/CHX extracts, revealed a cytotoxic effect for both loading concentrations. Histomorphological analysis upon implantation into the dorsal connective tissues and calvaria of rats for 7 and 42 days showed that the high CHX concentration induced the infiltration of inflammatory cells, resulting in retarded bone growth. Despite a strong decrease in in vitro cell viability, the low CHX loading dose did not impair the biocompatibility and osteoconductivity of HA during bone repair. These results indicate that high antimicrobial doses may activate a strong local inflammatory response and disrupt the long-term osteoconductive properties of CHX-HA delivery systems.

In this work, the in vitro behavior of human osteoblast cells on the undulated surfaces of biphasic calcium phosphate tablets was investigated. The tablets were produced by uniaxial pressing with convex cylindrical undulations occupying only half of the surface area; the other half was flat. Chemical and physical characterization was performed by scanning electron microscopy (SEM), atomic force microscopy (AFM), X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR). XRD and FTIR analyses revealed the presence of hydroxyapatite (HA) and alpha-tricalcium phosphate (alpha-TCP) in a well-defined ratio. Moreover, microtopography, evaluated by SEM and AFM, was similar on the flat region and on that with undulations. However, surface undulations induced different cellular arrangements, confirming the influence of the macrotopography on the cells orientation.

Isolated iron ions and iron oxide nanoparticles entrapped in alginate microspheres were studied by 57 Fe Mfssbauer spectroscopy (MS), electron paramagnetic resonance (EPR) and magnetization measurements. Ferric alginate beads were prepared by dropping sodium alginate solution into ferric chloride solution (0.1 and 0.5 M) with stirring under Ar atmosphere. MS and EPR spectroscopy show that iron is absorbed in alginate as Fe(III). The EPR suggests that isolated Fe(III) substitutes sodium in the polymeric structure (narrow line paramagnetic species at g=4.23). This mechanism induces the linking of the alginate units and the formation of the alginate microspheres. During this process, superparamagnetic iron hydroxide particles (broad line species at g=2.22) are precipitated inside the polymer network. MS measurements performed in diluted and concentrated Fe(III) samples displays a paramagnetic doublet at 300 K, and two components at 4.2 K due to a paramagnetic site and blocked superparamagnetic particles. D 2004 Published by Elsevier B.V.

Immobilization of toxic metals by calcium phosphates is a promising technology for treating contaminated soil, water and wastes. A detailed study on the mechanisms of lead immobilization by hydroxyapatite has been carried out using scanning electron microscopy (SEM), transmission electron microscopy (TEM) and X-ray diffraction (XRD). For this, synthetic hydroxyapatite powder were submitted to a sorption process through exposure to an aqueous solution containing 917 mg L−1 of lead for times that varied from 3 min to 54 h. The results obtained reinforce the hypothesis that hydroxypyromorphite formation is the end of a kinetic process in which the hydroxyapatite crystals are continuously dissolved and recrystallized in order to form more stable structures with higher lead content. Consequently, the use of calcium phosphates to immobilize lead ions seems to be technically viable.

Archaeological artefacts made of obsidian are all the more precious for archaeologists as they are witnesses of cultural, social or 'economic' relationships between prehistoric populations. The main obsidian sources reachable in these times have been identified and characterized by chemical analysis and fission track dating. Because of iron presence (from 1% to 10% expressed in Fe 2 O 3 ) it is possible to use Mossbauer spectroscopy and electron spin resonance (ESR) on geological samples to characterize different sources. We studied by electron spin resonance about seventy obsidians coming from six Mediterranean volcanic islands. The complex spectra are mainly due to iron in different states and site location (and sometimes isolated Mn 2+ ions). X-band (9 GHz) spectra exhibit a signal at g = 4.3 with a shoulder at 9.8 ascribed to isolated Fe 3+ in the glassy matrix with a rhombic environment (C 2y ). Condensed clusters of Fe 3+ ions give rise to a resonance line at g = 2.0 whose position and width do not depend on temperature and in addition to this signal broad resonance lines are ascribed to micro-crystallites of mixed iron oxides (hematite, magnetite and related spinels, titanates, silicates) as inclusions into the amorphous matrix of obsidian. When temperature decreases from 300 to 5 K these signals shift towards weak fields. The contribution of every signal to ESR spectrum is different from one source to the other according to the different thermodynamic conditions that occurred during obsidian formation. It is thus possible to discriminate a priori one geological source using ESR spectra. At room temperature the first differences appear at X-band, low temperatures experiments may allow to remove some ambiguities.