Hydroxyapatite Discs Research Articles

Full-contour Y-TZP ceramic surface roughness effect on synthetic hydroxyapatite wear

ObjectiveTo investigate the effects of polishing techniques on the surface roughness of Y-TZP ceramic and on the wear behavior of synthetic hydroxyapatite (HA). MethodsThirty-two full-contour Y-TZP (Diazir®) sliders (φ=2mm×1.5mm in height) were manufactured using CAD/CAM, embedded in acrylic resin using brass holders, and randomly allocated into four groups (n=8): according to the finishing/polishing procedure: G1-as-machined, G2-glazed, G3-diamond bur finishing and G4−G3+OptraFine® polishing kit. Thirty-two sintered HA disks (φ=13mm×2.9mm in height) were similarly mounted in brass holders. Y-TZP sliders baseline surface roughness values (Ra and Rq, in μm) were recorded using a non-contact profilometer (Proscan 2000). A two-body pin-on-disc wear test was performed. HA height (μm) and volume (mm3) losses were measured. Y-TZP height loss was measured using a digital micrometer. One-way ANOVA was used to determine the effect of the polishing techniques on the surface roughness. Comparisons between groups for differences in antagonist height loss/volume, and slider height loss were performed using one-way ANOVA. Statistical significance was set at α=0.05. ResultsRoughness measurements showed significant differences (p=0.0001) among the surface treatments with G1 (Ra=0.84, Rq=1.13μm) and G3 (Ra=0.89, Rq=1.2μm) being the roughest, and G2 (Ra=0.42, Rq=0.63μm) the smoothest (p=0.0001). Y-TZP slider height loss was highest for the glazed group (35.39μm), and was lowest for the polished group (6.61μm) (p=0.0001). Antagonist volume and height losses for groups (G1–G3) were similar, while the polished group (1.3mm3, 14.7μm) showed significant lower values (p=0.0001). SignificanceAlthough glazed zirconia provides an initially smooth surface, significantly increased antagonist wear was observed compared to the polished Y-TZP zirconia surface.

Effects of Lactobacillus reuteri PTA 5289 and L. paracasei DSMZ16671 on the Adhesion and Biofilm Formation of Streptococcus mutans

Probiotics have decreased the counts of salivary mutans streptococci (MS) in clinical studies. The aim of this study was to compare the effects of Lactobacillus reuteri PTA 5289 and L. paracasei DSMZ16671 on the adhesion of a reference strain and a clinical isolate of Streptococcus mutans and on the counts of MS in a biofilm. The adhesion of S. mutans Ingbritt and the clinical isolate S. mutans 2366 to a smooth glass surface and saliva-coated hydroxyapatite (SHA) were studied in the presence of and without the lactobacilli. A three-species biofilm formed on saliva-coated hydroxyapatite discs was used in the biofilm experiments. The lactobacilli did not affect adhesion to the glass surface but interfered with binding to SHA. No effects of the lactobacilli were detected on the MS levels in the three-species biofilms. The results of the SHA binding experiments best reflected the results of the existing clinical studies.

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.

Pamidronate Enhances Bacterial Adhesion to Bone Hydroxyapatite. Another Puzzle in the Pathology of Bisphosphonate-Related Osteonecrosis of the Jaw?

Bacterial colonization of the denuded bone in bisphosphonate-related osteonecrosis of the jaw suggests that bisphosphonates increase bacterial adhesion and biofilm formation. This study evaluated the adhesion of gram-positive and gram-negative bacteria on hydroxyapatite coated with pamidronate, one of the most potent bisphosphonates. Twenty-five Staphylococcus aureus and 25 Pseudomonas aeruginosa strains were cultured on pamidronate-coated and uncoated hydroxyapatite discs. After incubation, nonadherent bacteria were removed by rinsing and centrifugation. Formation of a biofilm was confirmed by confocal laser 3-dimensional and scanning electron microscopy. The number of bacterial colonies was counted using quantitative cultures and mean numbers were compared using the Mann-Whitney rank sum test (statistical significance defined as P ≤ .05). The Hartree-Fock method was used for the calculation of electron interactions between hydroxyapatite ions and pamidronate. Fold increases in the number of colonies formed by S aureus and P aeruginosa in the presence of pamidronate compared with controls were 7.19 ± 4.127 and 2.87 ± 0.622, respectively. Hartree-Fock analysis showed that the reactive NH3(+) group of pamidronate may act as a steric factor, facilitating anchoring of bacteria to the hydroxyapatite surface. Alternatively, the NH3(+) group may attract bacteria by direct electrostatic interaction. Increased bacterial adhesion in the presence of bisphosphonates can promote osteomyelitis in patients with bisphosphonate-related osteonecrosis of the jaw. There may be increased infection rates when bisphosphonates are used for stabilization of prostheses in joint arthroplasty and in osteotomies and open fractures in patients treated with bisphosphonates.

Quantitative high contrast X-ray microtomography for dental research

ObjectivesX-ray microtomography (XMT or micro-CT) is a miniaturized version of medical CT and has been used extensively for in vitro dental research. The technique allows three-dimensional analyses of both structure and density (or concentration), the latter requiring some a priori knowledge of composition. The aim of this paper is to demonstrate the capabilities of an XMT scanner specifically designed for dental research, especially that requiring high contrast resolution or accurate mineral concentration quantification. MethodsThe MuCAT scanners designed at Queen Mary, University of London, use high dynamic range CCD cameras with time-delay integration readout to provide high quality tomographic images. Accuracy in mineral concentration quantification is achieved using a modelling approach to beam hardening correction with automatic calibration following every scan. Accuracy was tested by measuring the density of a hydroxyapatite disc, and the utility of the high contrast and high accuracy capabilities are demonstrated in three applications from ongoing dental research projects. ResultsQuantification accuracy in the hydroxyapatite disc density was better than 1%. In dental applications, slight differences in mineral concentration allowed features such as dead tracts to be visualized and ion transport from a glass-ionomer cement into demineralized dentine to be observed. ConclusionThe improved accuracy and contrast sensitivity, together with the non-destructive nature of XMT in general, facilitates precise studies of dynamic processes in teeth. The ability to differentiate subtle differences in mineral concentration allows dead tracts to be traced in three dimensions, linking external pathology in teeth to reactive processes from the pulp. Clinical significanceThe MuCAT scanners have been optimized for quantification of mineral concentration and are particularly useful for in vitro studies of de- and re-mineralization, excavation and other treatment methodologies, and for gaining further insight into tooth morphology and pathology. Results from such studies will inform development of clinical treatment and management.

Effect of Precipitation Method on Properties of Hydroxyapatite Powders

2nd International Congress on Advances in Applied Physics and Materials Science (APMAS) -- APR 26-29, 2012 -- Antalya, TURKEY

Development of Novel Fluorescent Probes for the Analysis of Protein Interactions under Physiological Conditions with Medical Devices

In this article, a method to analyze protein adsorption on porous, clinically relevant samples under physiologically relevant conditions is described. The use of fluorescent probes was identified as a methodology that would facilitate analysis under a range of conditions including fully competitive conditions where a protein of interest may be labeled in isolation and then allowed to compete with unlabeled proteins on samples that require no specialized surface pretreatment. As a first step, this article describes the covalent labeling of isolated bovine serum albumin (BSA) with fluorescent fluoresceinthioureidoaminocaproic acid, FTCA, giving FTCA-BSA. The fluorescence intensity of FTCA-BSA was then used to monitor the adsorption and desorption of the protein under noncompetitive conditions with two forms of hydroxyapatite discs (silicate-substituted, SA and stoichiometric, HA) in phosphate-buffered saline (PBS) and minimum essential Eagles' medium (MEM). Noncompetitive conditions were used to facilitate the validation of the technique in which data obtained from these experiments were corroborated against data obtained using an established total protein assay method (Quant-IT kit, Invitrogen). These experiments demonstrated that the FTCA-BSA probe had several advantages including a greater sensitivity at lower concentrations and a considerably longer lifetime. The results also demonstrated that the interaction of BSA with SA and HA was also highly temperature- and media-dependent. Under the most physiologically relevant conditions of MEM at 37 °C, BSA was more readily adsorbed to SA with significant differences between biomaterials, but no differences were observed during the desorption process. The use of this method to analyze adsorption under competitive conditions will be the subject of further investigations.

Effect of the Source of Biofilm Bacteria, Level of Biofilm Maturation, and Type of Disinfecting Agent on the Susceptibility of Biofilm Bacteria to Antibacterial Agents

IntroductionBiofilm development is a dynamic process that begins with the initial attachment of planktonic bacteria to a surface, eventually leading through different stages to a mature, structurally complex biofilm. The aim of this study was to assess the effect of the source of biofilm bacteria, the level of biofilm maturation, and the type of disinfecting agent on the susceptibility of biofilm bacteria to antibacterial agents. MethodsMultispecies biofilms from plaque bacteria of 6 donors were grown for up to 8 weeks on collagen-coated hydroxyapatite disks. After 1, 2, 3, 4, and 8 weeks of growth the biofilms were exposed to 1% sodium hypochlorite, 0.2/0.4% iodine-potassium iodide, and 2% chlorhexidine for 1 and 3 minutes. The percentage of killed biofilm bacteria was determined by using LIVE/DEAD viability staining and confocal laser scanning microscopy. One-way analysis of variance was used for statistical analysis. ResultsOne- and 2-week-old biofilms were moderately or very sensitive to the tested disinfecting agents, which killed 20%–100% of the biofilm bacteria. After 3 weeks of growth the biofilm bacteria were more resistant to the same agents, and only 10%–30% of the bacteria were killed (P < .001). The same pattern of the effect of biofilm age (maturation) on the resistance of bacteria was observed in all 6 biofilms and with all 3 disinfecting agents. ConclusionThe change of biofilm bacteria from sensitive to resistant against disinfecting agents occurred between 2 and 3 weeks of biofilm maturation. This development took place simultaneously in all biofilms grown from plaque bacteria from 6 different donors and was independent of the type of disinfecting agent used. The results emphasize the importance of knowing the maturation timeline of each biofilm model used to test the effectiveness of endodontic disinfecting agents against biofilm bacteria.

Engineering nanocages with polyglutamate domains for coupling to hydroxyapatite biomaterials and allograft bone

Hydroxyapatite (HA) is the principal constituent of bone mineral, and synthetic HA is widely used as a biomaterial for bone repair. Previous work has shown that polyglutamate domains bind selectively to HA and that these domains can be utilized to couple bioactive peptides onto many different HA-containing materials. In the current study we have adapted this technology to engineer polyglutamate domains into cargo-loaded nanocage structures derived from the P22 bacteriophage. P22 nanocages have demonstrated significant potential as a drug delivery system due to their stability, large capacity for loading with a diversity of proteins and other types of cargo, and ability to resist degradation by proteases. Site-directed mutagenesis was used to modify the primary coding sequence of the P22 coat protein to incorporate glutamate-rich regions. Relative to wild-type P22, the polyglutamate-modified nanocages (E2-P22) exhibited increased binding to ceramic HA disks, particulate HA and allograft bone. Furthermore, E2-P22 binding was HA selective, as evidenced by negligible binding of the nanocages to non-HA materials including polystyrene, agarose, and polycaprolactone (PCL). Taken together these results establish a new mechanism for the directed coupling of nanocage drug delivery systems to a variety of HA-containing materials commonly used in diverse bone therapies.

Ex vivo killing of Enterococcus faecalis and mixed plaque bacteria in planktonic and biofilm culture by modified photoactivated disinfection

To compare the efficacy of conventional and modified photoactivated disinfection (PAD) against Enterococcus faecalis and mixed plaque bacteria in suspension and biofilms. Enterococcus faecalis (four strains) and mixed plaque bacteria from three adult volunteers were suspended in water, added to methylene blue (MB, 15 μmol L⁻¹), MB mixed with 0.5% hydrogen peroxide and 0.05% chlorhexidine (CHX), MB mixed with 0.5% hydrogen peroxide and 0.05% EDTA or MB mixed with 0.05% EDTA and 0.05% CHX and exposed to laser irradiation from 10 s to 5 min. After exposure, samples were taken, serially diluted and grown aerobically and anaerobically on Tryptic Soy Agar plates or on blood agar plates for 24 and 72 h, respectively. For biofilm experiments, E. faecalis and mixed plaque biofilms were grown on sterile hydroxyapatite (HA) discs coated overnight with bovine dermal collagen type I for 3 weeks. After exposure to MB or MB and low concentration of EDTA with either hydrogen peroxide or CHX, the percentage of killed bacteria by PAD was evaluated using viability staining and confocal laser scanning microscope. For statistical analysis, one-way analysis of variance was performed. Conventional PAD killed from 90.76% to 100% E. faecalis for 3 min, but failed to kill all plaque bacteria even after 5 min of laser irradiation. In modified PAD, up to 100% of suspended E. faecalis and mixed plaque bacteria were killed after 1 min and 30 s of irradiation. Up to twenty times more biofilm bacteria were killed by modified PAD than by conventional PAD with 15 μmol L⁻¹ MB (P < 0.001) and up to eight times more than 2% CHX (P < 0.001) and 1% sodium hypochlorite (P < 0.001). Modified PAD was superior to conventional PAD against planktonic and biofilm bacteria.

Influence of substrate curvature on osteoblast orientation and extracellular matrix deposition

BackgroundThe effects of microchannel diameter in hydroxyapatite (HAp) substrates on osteoblast behavior were investigated in this study. Microchannels of 100, 250 and 500 μm diameter were created on hydroxyapatite disks. The changes in osteoblast precursor growth, differentiation, extra cellular matrix (ECM) secretion and cell attachment/orientation were investigated as a function of microchannel diameter.ResultsCurvature did not impact cellular differentiation, however organized cellular orientation was achieved within the 100 and 250 μm microchannels (mc) after 6 days compared to the 12 days it took for the 500mc group, while the flat substrate remained disorganized. Moreover, the 100, 250 and 500mc groups expressed a specific shift in orientation of 17.45°, 9.05°, and 22.86° respectively in 24 days. The secreted/mineralized ECM showed the 100 and 250mc groups to have higher modulus (E) and hardness (h) (E = 42.6GPa; h = 1.6GPa) than human bone (E = 13.4-25.7GPa; h = 0.47-0.74GPa), which was significantly greater than the 500mc and control groups (p < 0.05). It was determined that substrate curvature affects the cell orientation, the time required for initial response, and the shift in orientation with time.ConclusionsThese findings demonstrate the ability of osteoblasts to organize and mineralize differentially in microchannels similar to those found in the osteons of compact bone. These investigations could lead to the development of osteon-like scaffolds to support the regeneration of organized bone.

In vitro antibacterial effect of carbamide peroxide on oral biofilm

This study compared the effects of carbamide peroxide (CP) and chlorhexidine (CHX) on oral biofilm in vitro. Collagen-coated hydroxyapatite discs were inoculated with subgingival plaque. After 3 weeks, the emergent biofilms were subjected to 1-, 3-, and 10-min exposures of a 1% CHX gel, a 5% CP gel and rinse, and a 10% CP gel and rinse. Subsequently, the biofilms were stained using a two-colour fluorescent dye kit for confocal laser scanning microscopy, and the volume ratio of dead bacteria to all bacteria was analysed. Compared to a non-treated gel control, the active agents killed bacteria on all the discs, with higher concentration and longer exposure times killing more bacteria. The rinse form disrupted the biofilm quicker than the gel form. Overall, 10% CP showed more disruption of biofilm and a greater proportion of killed bacteria than 1% CHX (p<0.05).

Apolar Bioactive Fraction ofMelipona scutellarisGeopropolis onStreptococcus mutansBiofilm

The aim of this study was to evaluate the influence of the bioactive nonpolar fraction of geopropolis on Streptococcus mutans biofilm. The ethanolic extract of Melipona scutellaris geopropolis was subjected to a liquid-liquid partition, thus obtaining the bioactive hexane fraction (HF) possessing antimicrobial activity. The effects of HF on S. mutans UA159 biofilms generated on saliva-coated hydroxyapatite discs were analyzed by inhibition of formation, killing assay, and glycolytic pH-drop assays. Furthermore, biofilms treated with vehicle control and HF were analyzed by scanning electron microscopy (SEM). HF at 250 μg/mL and 400 μg/mL caused 38% and 53% reduction in the biomass of biofilm, respectively, when compared to vehicle control (P < 0.05) subsequently observed at SEM images, and this reduction was noticed in the amounts of extracellular alkali-soluble glucans, intracellular iodophilic polysaccharides, and proteins. In addition, the S. mutans viability (killing assay) and acid production by glycolytic pH drop were not affected (P > 0.05). In conclusion, the bioactive HF of geopropolis was promising to control the S. mutans biofilm formation, without affecting the microbial population but interfering with its structure by reducing the biochemical content of biofilm matrix.

Validation of ATP bioluminescence as a tool to assess antimicrobial effects of mouthrinses in an in vitro subgingival-biofilm model

Objectives. The aim of this investigation was to evaluate whether the adenosine triphosphate (ATP) bioluminescence method is an appropriate tool to assess the efficacy of antiseptic mouthrinses in terms of quantitative reductions of total viable microbial counts in mixed biofilm populations in vitro. Study Design. Three mouthrinses, containing respectively, chlorhexidine and cetylpyridinium chloride (CHX/CPC), essential oils (EO) and amine fluoride/stannous fluoride (AFSF), as well as Phosphate Buffered Saline (PBS) used as control, were tested in an in vitro static biofilm model by ATP bioluminescence and compared to culture method. Biofilms were grown on saliva-coated hydroxyapatite disks for 72 hours and then exposed for 1 minute to the mouthrinse or control by immersion. The antibacterial effect of the rinses was tested by analysis of variance. The reliability of the ATP bioluminescence method was assessed by calculating the Pearson correlation coefficients when compared to the viable cell counts obtained by culture. Results. Using ATP bioluminescence, the antimicrobial activity of the tested mouthrinses was demonstrated when compared to the PBS control. The ATP bioluminescence values were significantly correlated (0.769, p<0.001) to the viable cell counts. CHX/CPC and AFSF showed similar antimicrobial activity, although AFSF had a less homogeneous effect, being both more effective than the EO rinse. Conclusion. ATP bioluminescence viability testing may be considered a useful tool to assess the in vitro efficacy of antibacterial compounds. In the proposed model, CHX/CPC and AFSF containing mouthrinses demonstrated superior antimicrobial activity, as compared to EO rinses, in a multispecies biofilm model. Key words:Biofilm, ATP bioluminescence,mouthrinse, essential oils, chlorhexidine, amine fluoride/stannous fluoride.

The Size Control of Hydroxyapatite Particles during Spray Pyrolysis

The size of hydroxyapatite particles during spray pyrolysis was successfully controlled by varying the concentrations of the starting calcium phosphate (CaP) solutions. Three different concentrations of CaP solutions were prepared by dissolving Ca (NO3)2·4H2O and (NH4)2HPO4 into deionized water and then they were sprayed at 900°C with the carrier gas flowing rate of 10 L/min. The size of the resultant hydroxyapatite particles decreased as lowering the concentration of the starting CaP solution. The smaller the particle size, the higher the relative density of the hydroxyapatite disks after sintering at 1100°C for 1 hour. The practical implication of the results is that highly sinterable hydroxyapatite particles can be synthesized during spray pyrolysis by controlling the starting concentration of CaP solution.

The effect of histatin 5, adsorbed on PMMA and hydroxyapatite, on Candida albicans colonization

The limited number of treatments for oral candidiasis resulted in the emergence of azole-resistant Candida albicans strains, thus enforcing the need for novel antifungal treatments. Although histatin 5 (H5) demonstrates antifungal activity, its inhibitory effect when adhered to hydroxyapatite and Polymetylmethacrylate (PMMA) surfaces, resembling conditions of the in vivo pellicle, remains unexplored. The objective of this in vitro study was to determine whether surface-adhered H5 inhibits the colonization of C. albicans on hydroxyapatite and/or PMMA. The C. albicans assay involved developing a mono-protein pellicle (either H5 or albumin) on hydroxyapatite and PMMA discs, introducing C. albicans and counting the number of adhered cells, throughout time, using scanning electron microscopy. A negative binomial statistical model and the Tukey-Kramer test were used for statistical analysis, with p < 0.01 indicating significance. H5-coated PMMA had significantly reduced number of cells compared to albumin-coated PMMA at 30, 90 and 1440 min (p < 0.0001), with the number of cells decreasing significantly in 90 and 1440 min (p < 0.0001). Similarly, H5-coated hydroxyapatite had significantly fewer cells compared to the albumin-coated surface at 90 and 1440 min (p < 0.0001), with the number of cells decreasing significantly at 30, 90 and 1440 min (p < 0.0001). In conclusion, C. albicans colonization was most inhibited by PMMA and hydroxyapatite-adhered H5 after 1440 min, illustrating the time-dependent effect of H5. In addition, yeast cells colonized albumin-coated PMMA, while dense hyphal networks formed on albumin-coated hydroxyapatite, suggesting that C. albicans morphology is influenced by the surface available for albumin adhesion.

DNA-lipid-apatite composite layers enhance gene expression of mesenchymal stem cells

DNA-apatite composite layer (D-Ap layer) and DNA-lipid-apatite composite layer (DLp-Ap layer) were prepared on ceramic hydroxyapatite disk and scaffold using supersaturated calcium phosphate solutions supplemented with 0.5–5μg/mL plasmid and 0–10μL/mL lipid transfection reagent FuGENE®. Both in vitro and in vivo studies were carried out using mesenchymal stem cells (MSCs) and two kinds of gene (luciferase and bone morphogenetic protein (BMP)-2) for demonstrating potential application of the gene transfer system using the D-Ap and DLp-Ap layers in bone tissue engineering. In the in vitro study using luciferase gene, the DLp-Ap layers showed 1–2 orders of magnitudes higher gene transfer efficiency to MSCs than the D-Ap layer. In the in vivo study using BMP-2 gene, DLp-Ap layer slightly increased BMP-2 protein concentration than D-Ap layer, thereby enhancing their osteogenic differentiation than D-Ap layer. The present gene transfer system using the DLp-Ap layers, with the advantages of good biocompatibility, bone-bonding ability, and efficacy in in vitro and in vivo gene transfer to MSCs, would be useful in bone tissue engineering.

A reproducible oral microcosm biofilm model for testing dental materials

Most studies of biofilm effects on dental materials use single-species biofilms, or consortia. Microcosm biofilms grown directly from saliva or plaque are much more diverse, but difficult to characterize. We used the Human Oral Microbial Identification Microarray (HOMIM) to validate a reproducible oral microcosm model. Saliva and dental plaque were collected from adults and children. Hydroxyapatite and dental composite discs were inoculated with either saliva or plaque, and microcosm biofilms were grown in a CDC biofilm reactor. In later experiments, the reactor was pulsed with sucrose. DNA from inoculums and microcosms was analysed by HOMIM for 272 species. Microcosms included about 60% of species from the original inoculum. Biofilms grown on hydroxyapatite and composites were extremely similar. Sucrose pulsing decreased diversity and pH, but increased the abundance of Streptococcus and Veillonella. Biofilms from the same donor, grown at different times, clustered together. This model produced reproducible microcosm biofilms that were representative of the oral microbiota. Sucrose induced changes associated with dental caries. This is the first use of HOMIM to validate an oral microcosm model that can be used to study the effects of complex biofilms on dental materials.

Formation mechanism of nano-sized hydroxyapatite powders through spray pyrolysis of a calcium phosphate solution containing polyethylene glycol

Abstract Nano-sized hydroxyapatite powders were synthesized by spray pyrolysis of a calcium phosphate solution containing polyethylene glycol (PEG) and a subsequent dewaxing process. Calcium phosphate solutions (Ca/P ratio of 1.67) were prepared by dissolving calcium nitrate tetrahydrate and diammonium hydrogen phosphate in deionized water with and without PEG and subsequently adding nitric acid. When the calcium phosphate solution was sprayed at 1100 °C at a carrier gas flow rate of 40 L/min, micron-sized hollow hydroxyapatite particles with dense or collapsed shells were obtained. However, when the calcium phosphate solution containing PEG was sprayed under the same conditions, micron-sized porous hydroxyapatite particles consisting of loosely packed hydroxyapatite crystals containing randomly placed carbons were obtained. When these hydroxyapatite particles were dewaxed at 650 °C for 3 h, they were completely disintegrated into nano-sized hydroxyapatite particles, whereas the micron-sized hollow hydroxyapatite particles without carbons remained intact. After sintering at 1100 °C for 3 h, the relative densities of hydroxyapatite disks made from micron- and nano-sized powders were 78.5 ± 1.7% and 99.8 ± 0.3%, respectively. The practical implication of these results is that highly sinterable nano-sized hydroxyapatite powders can be synthesized through spray pyrolysis of a calcium phosphate solution containing PEG.

Quantitative Proteomic Analysis of the Effect of Fluoride on the Acquired Enamel Pellicle

The acquired enamel pellicle (AEP) is a thin film formed by the selective adsorption of salivary proteins onto the enamel surface of teeth. The AEP forms a critical interface between the mineral phase of teeth (hydroxyapatite) and the oral microbial biofilm. This biofilm is the key feature responsible for the development of dental caries. Fluoride on enamel surface is well known to reduce caries by reducing the solubility of enamel to acid. Information on the effects of fluoride on AEP formation is limited. This study aimed to investigate the effects of fluoride treatment on hydroxyapatite on the subsequent formation of AEP. In addition, this study pioneered the use of label-free quantitative proteomics to better understand the composition of AEP proteins. Hydroxyapatite discs were randomly divided in 4 groups (n = 10 per group). Each disc was exposed to distilled water (control) or sodium fluoride solution (1, 2 or 5%) for 2 hours. Discs were then washed and immersed in human saliva for an additional 2 hours. AEP from each disc was collected and subjected to liquid chromatography electrospray ionization mass spectrometry for protein identification, characterization and quantification. A total of 45 proteins were present in all four groups, 12 proteins were exclusively present in the control group and another 19 proteins were only present in the discs treated with 5% sodium fluoride. Relative proteomic quantification was carried out for the 45 proteins observed in all four groups. Notably, the concentration of important salivary proteins, such as statherin and histatin 1, decrease with increasing levels of fluoride. It suggests that these proteins are repulsed when hydroxyapatite surface is coated with fluoride. Our data demonstrated that treatment of hydroxyapatite with fluoride (at high concentration) qualitatively and quantitatively modulates AEP formation, effects which in turn will likely impact the formation of oral biofilms.