Adsorption characteristics of bovine serum albumin onto alumina with a specific crystalline structure

Bone cement containing alumina particles with a specific crystalline structure exhibits the ability to bond with bone. These particles (AL-P) are mainly composed of delta-type alumina (δ-Al₂O₃). It is likely that some of the proteins present in the body environment are adsorbed onto the cement and influence the expression of its bioactivity. However, the effect that this adsorption of proteins has on the bone-bonding mechanism of bone cement has not yet been elucidated. In this study, we investigated the characteristics of the adsorption of bovine serum albumin (BSA) onto AL-P and compared them with those of its adsorption onto hydroxyapatite (HA), which also exhibits bone-bonding ability, as well as with those of adsorption onto alpha-type alumina (α-Al₂O₃), which does not bond with bone. The adsorption characteristics of BSA onto AL-P were very different from those onto α-Al₂O₃ but quite similar to those onto HA. It is speculated that BSA is adsorbed onto AL-P and HA by interionic interactions, while it is adsorbed onto α-Al₂O₃ by electrostatic attraction. The results suggest that the specific adsorption of albumin onto implant materials might play a role in the expression of the bone-bonding abilities of the materials.     

The effect of hot pressing on the physical properties of glass reinforced hydroxyapatite

Hydroxyapatite (HA), being of physiological importance, can be developed synthetically for implant application. A number of avenues have been explored in order to improve the physical and biological properties of a variety of hydroxyapatite composites. However, the fact remains, hydroxyapatite lacks the mechanical properties needed to sustain high loads. This study investigates the advantages of hot pressing on the physical properties of HA and glass reinforced HA (GR-HA). The results show a significant enhancement in the mechanical properties of GR-HA composites compared to HA e.g. flexural bending strength values were given at 91.75 and 88.87 M Nm(-2) for GR-HA (CP15F) and GR-HA (CP20F) respectively, compared to 78.9 M Nm(-2) for HA. The results for other properties such as elastic modulus, fracture toughness, Vicker's hardness, density and porosity also demonstrate the benefit of adding phosphate based glasses as a sintering aid. This is supported by XRD analysis, highlighting the presence of a secondary phase (beta-TCP) in GR-HA systems and the positive effect it has on the physical properties. It must be brought to attention that densification of hot pressed HA and GR-HA composites is reached at a lower temperature compared to a previous study on the same materials that have undergone pressureless sintering.     

Porous bioceramic bead prepared by calcium phosphate with sodium alginate gel and PE powder

The porous calcium phosphate beads were made by an alginate-interacting Ca ions mechanism on addition of a pore-forming polyethylene (PE) powder at 1250 °C sintering. The nature of the powders and porous beads were analyzed through X-ray diffraction (XRD), Fourier transform infrared spectrometry (FTIR) and heavy metal analysis by inductively coupled plasma-optical emission spectroscopy (ICP-OES). The porous beads size and the pore microstructure characteristics were determined using scanning electron microscopy (SEM). Beside, the porosity analysis was evaluated out using an Archimedes' principle and mercury porosimetry. Then, the sodium ampicillin was penetrated/adsorbed onto calcium-deficient hydroxyapatite porous beads, and was subsequently released in PBS. No matter whether the raw material was HAp, TCP or biphase, the Ca₉(HPO₄)(PO₄)₅OH phase (CDHA) was formed only after sintering. Porous beads of various calcium phosphates with different sizes (0.9–1.1 mm) and pore size groups (60–120 μm and lower than 10 μm) were appeared. The release kinetics of sodium ampicillin from these porous beads have indicated the possibility of using these materials as possible carriers for drug delivery.     

Dissolution studies of hydroxyapatite and glass-reinforced hydroxyapatite ceramics

In the continuous agitation assays, glass-reinforced hydroxyapatite (GR-HA) was shown to form a calcium phosphate (CaP) layer, but hydroxyapatite (HA) only formed dispersed precipitates. The formation of this layer was first detected on the GR-HA with a 7.5% glass addition (7.5 GR-HA) after only 3 days of immersion in simulated body fluid (SBF). The time required for layer formation decreased as the amount of glass added to the HA increased. The dissolution rate of the materials followed a similar pattern, i.e. the dissolution rate for GR-HA was higher than for HA, and increased with the addition of glass. The immersion of 7.5 GR-HA in water showed almost linear dissolution kinetics over the immersion periods (3, 7, 15, 30 and 60 days). The concentration of calcium ions in solution and the scanning electron microscopy (SEM) analysis of the 7.5 GR-HA specimens immersed in water and in SBF revealed a clear competition between the material dissolution and the precipitation of a CaP phase. Fourier transformed infrared spectroscopy with alternated total reflectance (FTIR-ATR) analysis indicated that the CaP phase that formed during longer immersion times (30 and 60 days) could be a carbonate-substituted CaP precipitate.As expected from previous work, the GR-HA behavior in terms of its in vitro bioactivity is higher than HA because a homogeneous CaP layer is formed and the precipitation occurs faster. From the dissolution test and in accordance with the chemical composition of the samples, GR-HA was more soluble than HA.     

Porous spherical hydroxyapatite and fluorhydroxyapatite granules: processing and characterization

This study is aimed at the development of a method to fabricate porous spherical hydroxyapatite (HA)—fluorapatite (FA) granules. The method to produce porous granules is based on liquid immiscibility effect. A suspension of HA–FA powder mixtures in aqueous solution of gelatin and oil as a dispersion media were used. By stirring the mixtures of these immiscible liquids, granules of 50–200 μm diameter can easily be produced. The granules were characterized with respect to their microstructure, phase composition and specific area. In vitro testing of human plasma protein adsorption onto the granules of HA and fluorhydroxyapatite were performed. No kind of difference in the dynamic protein adsorption between pure HA and the HA up to 10 wt% FA materials has been revealed.     

Porous spherical hydroxyapatite and fluorhydroxyapatite granules: processing and characterization

This study is aimed at the development of a method to fabricate porous spherical hydroxyapatite (HA)—fluorapatite (FA) granules. The method to produce porous granules is based on liquid immiscibility effect. A suspension of HA–FA powder mixtures in aqueous solution of gelatin and oil as a dispersion media were used. By stirring the mixtures of these immiscible liquids, granules of 50—200 mm diameter can easily be produced. The granules were characterized with respect to their microstructure, phase composition and specific area. In vitro testing of human plasma protein adsorption onto the granules of HA and fluorhydroxyapatite were performed. No kind of difference in the dynamic protein adsorption between pure HA and the HA up to 10 wt% FA materials has been revealed.     

Synthesis of nanocrystalline fluorinated hydroxyapatite by microwave processing and its in vitro dissolution study

Synthetic hydroxyapatite, (Ca₁₀(PO₄)₆(OH)₂, HA), is an important material used for orthopedic and dental implant applications. The biological hydroxyapatite in the human bone and tooth is of nanosize and differs in composition from the stoichiometric HA by the presence of other ions such as carbonate, magnesium, fluoride, etc. Osseointegration is enhanced by using nanocrystalline HA. This stimulates the interest in synthesizing nanocrystalline HA by different routes and among the methods, microwave processing seems to form the fine grain size and uniform characteristic nanocrystalline materials. Fluorinated hydroxyapatite, (FHA, Ca₁₀(PO₄)₆(OH)_2−x F _x ), possesses higher corrosion resistance in biofluids than pure HA and reduces the risk of dental caries. The present work deals with the synthesis of nanocrystalline FHAs by microwave processing. The crystal size and morphology of the nanopowers were examined by X-ray powder diffraction (XRD) and transmission electron microscopy (TEM) methods. The functional groups present in FHA powders were ascertained by Fourier transform infrared spectroscopy (FT-IR) and laser Raman spectroscopy. Since the physiological stability is an important parameter while selecting the material for implantation, the in vitro dissolution studies of FHAs with different fluorine contents were carried out.     

Hydrothermal preparation of tailored hydroxyapatite

Hydrothermal vapor treatment method was applied for preparation of ceramic biomaterials. Hydroxyapatite (Ca₁₀(PO₄)₆(OH)₂; HA) ceramics prepared by sintering with random crystal surface have already been used as bone-repairing materials which can directly bond to natural bones. If materials of HA could have the tailored specific crystal surface, they should have the advantage of adsorptive activity and osteoconductivity in comparison with the sintered HA. In the present study, porous HA sheets of about 50 μm to 1 mm in thickness and porous HA granules of about 50 μm to 1 mm in size with tailored crystal surface were prepared by the hydrothermal vapor exposure method at temperatures below 200°C. Porous sheets and porous granules of HA with controlled crystal surface should be suitable for scaffold of cultured bone, for bone graft material and for drug delivery system (DDS).     

Mesoporous hydroxyapatite by hard templating of silica and carbon foams for protein release

Calcium phosphates, particularly hydroxyapatite Ca₁₀(PO₄)₆(OH)₂ (HA), are widely used for bone regeneration due to their biocompatibility and good resorption properties. However, their performance upon implantation is improved when they are associated with bioactive molecules such as growth factors. Using mesoporous HA leads to improved protein adsorption and release kinetics because the diameter of the mesopores (2–50 nm) is in the same range as their size. We prepared this type of material by the nanocasting method using three different templates: a silica foam and two carbon templates derived from it using propylene or sucrose as carbon source. We investigated the influence of the template, the calcination temperature and of the conditions during template removal. We obtained HA materials with a surface area of up to 90 m² g^?1 and with an intergranular mesopore volume of up to 0.4 cm³ g^?1. In this paper, we show for the first time that the synthesis of mesoporous HA from a mesoporous silica foam template allows eliminating the template at lower temperatures (in an alkaline medium), thus preventing the sintering of the HA. These materials have interesting properties for drug delivery applications. The protein adsorption and release capacities of these HAs were tested with two model proteins, bovine serum albumin (BSA), and Cytochrome C. These materials are an important milestone for future bone regeneration systems based on HA associated with human growth factor proteins.     

Influence of antibiotic adsorption on biocidal activities of silver nanoparticles

Excessive use of antibiotics has posed two major challenges in public healthcare. One of them is associated with the development of multi‐drug resistance while the other one is linked to side effects. In the present investigation, the authors report an innovative approach to tackle the challenges of multi‐drug resistance and acute toxicity of antibiotics by using antibiotics adsorbed metal nanoparticles. Monodisperse silver nanoparticles (SNPs) have been synthesised by two‐step process. In the first step, SNPs were prepared by chemical reduction of AgNO₃ with oleylamine and in the second step, oleylamine capped SNPs were phase‐transferred into an aqueous medium by ligand exchange. Antibiotics – tetracycline and kanamycin were further adsorbed on the surface of SNPs. Antibacterial activities of SNPs and antibiotic adsorbed SNPs have been investigated on gram‐positive (Staphylococcus aureus, Bacillus megaterium, Bacillus subtilis), and gram‐negative (Proteus vulgaris, Shigella sonnei, Pseudomonas fluorescens) bacterial strains. Synergistic effect of SNPs on antibacterial activities of tetracycline and kanamycin has been observed. Biocidal activity of tetracycline is improved by 0–346% when adsorbed on SNPs; while for kanamycin, the improvement is 110–289%. This synergistic effect of SNPs on biocidal activities of antibiotics may be helpful in reducing their effective dosages.Inspec keywords: silver, silver compounds, nanomedicine, nanoparticles, drug delivery systems, drugs, antibacterial activity, microorganisms, nanofabrication, materials preparation, reduction (chemical), adsorption, surface chemistry, surface treatmentOther keywords: public healthcare, multi‐drug resistance, side effect, acute antibiotic toxicity, antibiotic adsorbed metal nanoparticle, monodisperse silver nanoparticle, two‐step SNP synthesis, SNP preparation, AgNO₃ chemical reduction, oleylamine capped SNP phase‐transfer, aqueous medium, ligand exchange, tetracycline, kanamycin, antibacterial activity, antibiotic adsorbed SNP, gram‐positive bacterial strain, Staphylococcus aureus, Bacillus megaterium, Bacillus subtilis, gram‐negative bacterial strain, Proteus vulgaris, Shigella sonnei, Pseudomonas fluorescens, SNP synergistic effect, effective dosage reduction, Ag, AgNO₃      

Preparation and evaluation of spherical Ca-deficient hydroxyapatite granules with controlled surface microstructure as drug carriers

Spherical Ca-deficient hydroxyapatite (HA) granules are expected to be useful drug carriers in bony sites because of their bone regeneration and adsorption ability. In order to control drug loading and release ability of the granules, a controlled surface microstructure was constructed. Spherical Ca-deficient granules composed of micron-sized rod-shaped particles were prepared by hydrothermal treatment of α-tricalcium phosphate (α-TCP) granules, and then, submicron HA particles were precipitated on the obtained granules by immersion in a supersaturated calcium phosphate (CP) solution. When bovine serum albumin was used as a drug model, precipitation of submicron particles causes the loading capability to increase and the release rate to decrease. The spherical Ca-deficient HA granules with the controlled surface microstructure are expected to be useful drug carriers that can act as scaffolds for bone repair.     

In vitro behavior of fluoridated hydroxyapatite coatings in organic-containing simulated body fluid

A dense and pure hydroxyapatite [HA, Ca₁₀(PO₄)₆(OH)₂] coating and a fluoridated HA [Ca₁₀(PO₄)₆(OH)_0.67F_1.33] are deposited on Ti6Al4V substrates by sol-gel dip coating method. Glucose and bovine serum albumin have been added in standard simulated body fluid (SBF) to form organic-containing SBF in simulation of the physiological blood plasma. The HA and the fluoridated HA coatings are immersed in the standard and modified SBF for time periods of 2, 4, 7, 14 and 28 days at 37 ± 0.1°C. After soaking, the coating surface is examined for nucleation and growth of apatite using SEM morphological observation. The post-soaking SBF solutions are analyzed via Inductively Coupled Plasma spectroscopy for calcium ion concentration. The results show that at concentration of 40 g/L, bovine serum albumin has significant retardation effect on apatite precipitation from SBF onto pure or fluoridated HA coatings; Fluorine-incorporation in HA has positive bio-activation effect in both standard SBF and organic-containing SBF. However, glucose addition in SBF does not generate significant influence on the bioactivity of HA and fluoridated HA.     

Influences of magnetized hydroxyapatite on the growth behaviors of osteoblasts and the mechanism from molecular dynamics simulation

To investigate the influence of magnetized hydroxyapatite on the growth and differentiation of osteoblasts, hydroxyapatite (HA) and magnetized hydroxyapatite (mHA) were synthesized and characterized. The cell viability, differentiation, and morphologies of osteoblasts were investigated in vitro, respectively. The results showed that compared to HA, cells cultured with mHA had better cell viability, and both HA and mHA were beneficial to the early differentiation of osteoblasts. Furthermore, the interaction mechanism between mHA and osteoblasts was elucidated using a molecular dynamics simulation. The simulation results indicated that when cultured with osteoblasts, HA adsorbed bovine serum protein onto its surface from the medium immediately, which was beneficial to the adhesion and proliferation of osteoblasts. The main driving force for the adsorption of bovine serum was the electronic properties of HA crystal faces. The (211) crystal face of HA had the highest electron density among its all crystal faces, thus mainly contributing to the protein adsorption of HA. Nevertheless, the (211) crystal face of mHA still had a relatively higher electron density than that of HA, thus possessing better protein adsorption than that of HA, and in turn promoting the biological functions of osteoblasts.     

Characterization, bioactivity and ampicillin release kinetics of TiO2 and TiO24SiO2 synthesized by sol-gel processing

Local drug delivery of antimicrobics by sustained release delivery system can be used to treat periodontal disease. Advantages of these systems may include maintaining high levels of antibiotic in the gingival crevicular fluid for a sustained period of time and ease of use with high patient acceptance. The materials used are TiO₂ and TiO₂4SiO₂, mixed with sodium ampicillin, a broad-spectrum antibiotic, have been synthesized by sol-gel method. The amorphous nature of the gels was ascertained by X-ray diffraction analysis. Release kinetics in a simulated body fluid (SBF) have been subsequently investigated. The amount of sodium ampicillin released has been detected by UV-VIS spectroscopy and SEM. The release kinetics seems to occur in more than one stage. HPLC analysis has also been taken to ensure the integrity of ampicillin after the synthetic treatment. Finally, SEM micrographs and EDS analysis showed the formation of a hydroxyapatite layer on the surface of the samples soaked in SBF. Both the materials showed good release and could be used as drug delivery bioactive systems. High antimicrobial effects of samples against Escherichia coliand Streptococcus mutants were found.     

Characterisation and photocatalytic activity of structure-controlled spherical granules of an anatase/hydroxyapatite composite

Materials that can purify the environments are desirable. Anatase (TiO₂) has received attention because it is stable and can decompose organic substances because of its photocatalytic activity. To make use of anatase effectively, we deposited nano-sized anatase particles on porous hydroxyapatite (HA) ceramics composed of rod-shaped particles. Spherical porous HA granules composed of rod-shaped HA particles were prepared using a hydrothermal process. The granules were soaked in a solution containing a water-soluble titanium complex and then hydrothermally treated. Nano-sized anatase particles were deposited on each rod-shaped HA particle. The anatase/HA granules composed of rod-shaped HA particles showed higher photocatalytic activity than those composed of globular HA particles. The granules are expected to be useful as an environment-purifying material with high manageability and photocatalytic activity.     

The effect of adsorbed vitamin D and K to hydroxyapatite on ALP activity of MC3T3-E1 cell

This study describes the adsorptive property of vitamins on HA and the effect of the HA adsorbed vitamin on the alkaline phosphatase (ALP) activity for effective use as a bone graft substitute. The vitamins used were calciferol (D₃), menaquirone (K₂) and 25-hydroxycholecalciferol (25(OH)D₃). These vitamins were adsorbed on HA at 4, 10, 20, 37 and 50 °C. The adsorption amount was constant below 20 °C, and decreased as the incubation temperature increased over 20 °C. The order of the adsorption amount was: 25(OH)D₃ > K₂ > D₃. The HA adsorbed vitamins (HA/D₃, HA/25(OH)D₃ and HA/K₂) were suspended in physiological saline for 48 h for the release test. The release ratio of all vitamins increased with incubation time. The order of the release ratio was: 25(OH)D₃ > K₂ > D₃, which was proportional to that of the adsorption amount. The ALP activity of MC3T3-E1 osteoblast-like cells on HA, HA/D₃, HA/25(OH)D₃ and HA/K₂ was also investigated. The ALP activity was higher on HA/25(OH)D₃ than on any other samples. However, HA/K₂ and HA/D₃ showed similar ALP activity to HA.     

<Emphasis Type="Italic">In vivo</Emphasis> assessment of hydroxyapatite and silicate-substituted hydroxyapatite granules using an ovine defect model

Phase pure hydroxyapatite (HA) and two silicate-substituted hydroxyapatites (0.8 and 1.5 wt% Si, or 2.6 and 4.9 wt% SiO₄) were prepared by aqueous precipitation methods. The filter-cakes of HA and silicate-substituted hydroxyapatite (SiHA) compositions were processed into granules 1.0–2.0 mm in diameter and sintered at 1200°C for 2 h. The sintered granules underwent full structural characterisation, prior to assessment in an ovine defect model by implantation for a period of 6 and 12 weeks. The results indicate that HA and SiHA implants were well accepted by the host tissue, with no evidence of inflammation. New bone formation was observed directly on the surfaces and in the spaces between the granular implants. Quantitative histomorphometry as determined by the percentage of bone ingrowth and bone coverage for both SiHA implant compositions was significantly greater than that for phase pure HA. These findings indicate that the in vivo bioactivity of hydroxyapatite was significantly improved by the incorporation of silicate ions into the HA structure, making SiHA ceramics attractive alternatives to conventional HA materials for use as bone graft substitute ceramics.     

Magnesium- and strontium-co-substituted hydroxyapatite: the effects of doped-ions on the structure and chemico-physical properties

The present study is aimed at investigating the contribution of two biologically important cations, Mg²⁺ and Sr²⁺, when substituted into the structure of hydroxyapatite (Ca₁₀(PO₄)₆(OH)₂,HA). The substituted samples were synthesized by an aqueous precipitation method that involved the addition of Mg²⁺- and Sr²⁺-containing precursors to partially replace Ca²⁺ ions in the apatite structure. Eight substituted HA samples with different concentrations of single (only Mg²⁺) or combined (Mg²⁺ and Sr²⁺) substitution of cations have been investigated and the results compared with those of pure HA. The obtained materials were characterized by X-ray powder diffraction, specific surface area and porosity measurements (N₂ adsorption at 77?K), FT-IR and Raman spectroscopies and scanning electron microscopy. The results indicate that the co-substitution gives rise to the formation of HA and ??-TCP structure types, with a variation of their cell parameters and of the crystallinity degree of HA with varying levels of substitution. An evaluation of the amount of substituents allows us to design and prepare BCP composite materials with a desired HA/??-TCP ratio.     

Silicon-substituted hydroxyapatite composite coating by using vacuum-plasma spraying and its interaction with human serum albumin

The incorporation of silicon can improve the bioactivity of hydroxyapatite (HA). Silicon-substituted HA (Ca₁₀(PO₄)_6−x (SiO₄) _x (OH)_2−x , Si-HA) composite coatings on a bioactive titanium substrate were prepared by using a vacuum-plasma spraying method. The surface structure was characterized by using XRD, SEM, XRF, EDS and FTIR. The bond strength of the coating was investigated and XRD patterns showed that Ti/Si-HA coatings were similar to patterns seen for HA. The only different XRD pattern was a slight trend toward a smaller angle direction with an increase in the molar ratio of silicon. FTIR spectra showed that the most notable effect of silicon substitution was that –OH group decreased as the silicon content increased. XRD and EDS elemental analysis indicated that the content of silicon in the coating was consistent with the silicon-substituted hydroxyapatite used in spraying. A bioactive TiO₂ coating was formed on an etched surface of Ti, and the etching might improve the bond strength of the coatings. The interaction of the Ti/Si-HA coating with human serum albumin (HSA) was much greater than that of the Ti/HA coating. This might suggest that the incorporation of silicon in HA can lead to significant improvements in the bioactive performance of HA.     

The impact of the RGD peptide on osteoblast adhesion and spreading on zinc-substituted hydroxyapatite surface

The incorporation of zinc into the hydroxyapatite structure (ZnHA) has been proposed to stimulate osteoblast proliferation and differentiation. Another approach to improve cell adhesion and hydroxyapatite (HA) performance is coating HA with adhesive proteins or peptides such as RGD (arginine–glycine–aspartic acid). The present study investigated the adhesion of murine osteoblastic cells to non-sintered zinc-substituted HA disks before and after the adsorption of RGD. The incorporation of zinc into the HA structure simultaneously changed the topography of disk’s surface on the nanoscale and the disk’s surface chemistry. Fluorescence microscopy analyses using RGD conjugated to a fluorescein derivative demonstrated that ZnHA adsorbed higher amounts of RGD than non-substituted HA. Zinc incorporation into HA promoted cell adhesion and spreading, but no differences in the cell density, adhesion and spreading were detected when RGD was adsorbed onto ZnHA. The pre-treatment of disks with fetal bovine serum (FBS) greatly increased the cell density and cell surface area for all RGD-free groups, overcoming the positive contribution of zinc to cell adhesion. The presence of RGD on the ZnHA surface impaired the effects of FBS pre-treatment possibly due to competition between FBS proteins and RGD for surface binding sites.