Faungchat Thammarakcharoen

Development of antibiotics impregnated nanosized silver phosphate-doped hydroxyapatite bone graft

Nanosized Ag3PO4 loaded hydroxyapatite which was prepared by a novel low temperature phosphorization of 3D printed calcium sulfate dihydrate at the nominal silver concentration of 0.001M and 0.005M was impregnated by two antibiotics including gentamicin and vancomycin. Phase composition, microstructure, antibiotics loading, silver content, antimicrobial performance, and cytotoxic potential of the prepared samples were characterized. It was found that the fabricated sample consisted of hydroxyapatite as a main phase and spherical-shaped silver phosphate nanoparticles distributing within the cluster of hydroxyapatite crystals. Antibacterial activity of the samples against two bacterial strains (gram negative P. aeruginosa and gram positive S. aureus) was carried out. It was found that the combination of antibiotics and nanosized Ag3PO4 in hydroxyapatite could enhance the antibacterial performance of the samples by increasing the duration in which the materials exhibited antibacterial property and the size of the inhibition zone depending on the type of antibiotics and bacterial strains compared to those contained antibiotics or nanosilver phosphate alone. Cytotoxic potential against osteoblasts of antibiotics impregnated nanosilver phosphate hydroxyapatite was found to depend on the combination of antibiotics content, type of antibiotics, and nanosilver phosphate content.

Enhancement of mechanical properties of 3D printed hydroxyapatite by combined low and high molecular weight polycaprolactone sequential infiltration.

A new infiltration technique using a combination of low and high molecular weight polycaprolactone (PCL) in sequence was developed as a mean to improve the mechanical properties of three dimensional printed hydroxyapatite (HA). It was observed that using either high (Mn~80,000) or low (Mn~10,000) molecular weight infiltration could only increase the flexural modulus compared to non-infiltrated HA, but did not affect strength, strain at break and energy at break. In contrast, a combination of low and high molecular infiltration in sequence increased the flexural modulus, strength and energy at break compared to those of non-infiltrated HA or infiltrated by high or low molecular weight PCL alone. This overall enhancement was found to be attributed to the densification of low molecular weight PCL and the reinforcement of high molecular PCL concurrently. The combined low and high molecular weight infiltration in sequence also maintained high osteoblast proliferation and differentiation of the composites at the similar level of the HA. Densification was a dominant mechanism for the change in modulus with porosity and density of the infiltrated HA/PCL composites. However, both densification and the reinforcing performance of the infiltration phase were crucial for strength and toughening enhancement of the composites possibly by the defect healing and stress shielding mechanisms. The sequence of using low molecular weight infiltration and followed by high molecular infiltration was seen to provide the greatest flexural properties and highest cells proliferation and differentiation capabilities.

Evaluation of PEG-PPG-PEG Copolymer Blends for Using as Resorbable Bone Wax

In this study, four types of PEG-PPG-PEG copolymers solids and pastes having different molecular mass (Mw~5800, 8400, 12000 and 14600) were selected and mixed at various compositions ranging from 40 to 60 % by weight. The textures and handling characteristics of the prepared samples were evaluated in comparison to that of the commercial non-resorbable bone wax. Thermal and mechanical properties of the blends were determined by a differential scanning calorimeter and a universal testing machine respectively. It was found that the combination of solid (Mw~8400) and paste-like (Mw~12000) copolymers at the ratio of 40:60 gave the closest smooth texture with good smearability and had similar melting temperature to the commercial bone wax, but having slightly lower compressive stiffness. Preliminary cytotoxicity of the selected formulations against osteoblasts revealed that the cell viability was greater than 70 % indicating that the prepared samples did not show any cytotoxic potential.

Antimicrobial Performance and Cytotoxicity of Antibiotics Impregnated Hydroxyapatite for Osteomyelitis Treatment

Localized antibiotic beads are often used for treating patients with osteomyelitis or infections of the bone by providing local, sustained and high concentrations of antimicrobial agents to the area of infection, without systemically exposing an individual to antibiotic levels that could result in numerous toxic side effects. In this study, antibiotic impregnated hydroxyapatite spheres were prepared aiming for a functional device of drug carrier and bone graft. Three types of antibiotics were employed including gentamicin, vancomycin and fosfomycin. Antimicrobial susceptibility of antibiotic impregnated beads were tested against two bacterial strains (Pseudomonas aeruginosa ATCC 27853 and Staphylococcus aureus ATCC 25923) by using modified agar diffusion assay. Differences in antimicrobial efficiency were observed and related with respect to differences in molecular weight, mechanism of action and spectrum of activity for each antibiotic. Cytotoxicity by serial extraction technique of all antibiotic impregnated beads were determined by MTT assay. No cytotoxic potential at all extraction periods was observed for vancomycin impregnated on beads. Gentamicin and fosfomycin impregnated beads showed cytotoxic potential only on day 1 extraction, but no cytotoxic potential on longer periods.

Biomimetically Co-Deposition of Bovine Serum Albumin and Calcium Phosphate on 3D Printed Hydroxyapatite: Influence of Time, Temperature and Concentration

Influence of bovine serum albumin (BSA) concentration, soaking times and soaking temperatures during biomimetic co-deposition process of three dimensionally printed hydroxyapatite (3D printed HA) on weight change, phase composition and microstructure were studied. At 23°C, weight change of all samples monotonically increased with increasing soaking time regardless of BSA concentration used while the weight change initially increased, but leveled off later with increasing soaking time when using solution temperature of 37°C. At 50°C, weight change initially increased for up to 4 h and remained constant for up to 16 h where the weight decreased afterward. This decrease could be related to the competition between the dissolution of 3D printed HA, the deposition of new calcium phosphate crystals and the incorporation of BSA during biomimetic process at high temperatures. From X-ray diffraction, no influence in BSA concentration, time and temperature on the phase composition was observed. Scanning electron microscopy revealed the deposition of new plate-like calcium phosphate crystals on all 3D printed HA. Hydroxyapatite and octacalcium phosphate were found to be the main phases of all biomimetically BSA.

The Use of Three Dimensional Printed Hydroxyapatite Granules in Alveolar Ridge Preservation

In this study, an alveolar ridge preservation using novel hydroxyapatite granules which was fabricated by three dimensional printing technique in post-extraction socket was carried out and evaluated. Clinical, radiographic and histology were assessed prior to dental implant placement. Five volunteered patients who needed an extraction of anterior tooth and scheduled for implant replacement were enrolled in this pilot study. No sign of infection or local of systemic immune reaction to the three dimensional printed hydroxyapatite granules in all patients was noted. At 8 weeks post-surgery, the grafted area was observed to be completely filled with woven bone and the formation of new vessels was seen. In addition, the bone quality and quantity of the grafted site when placing the implant showed efficient implant stability (ISQ values ∼ 65) without the need of additional bone graft surgery. Overall results indicated that three dimensional printed hydroxyapatite granules could be potentially employed as bone grafting material for alveolar ridge preservation.

Effect of Process Parameters on Biomimetic Deposition of Calcium Phosphate on 3D Printed Hydroxyapatite

Factors influencing calcium phosphate deposition on three dimensional printed hydroxyapatite (3D printed HA) by biomimetic process including soaking times (2, 4 and 6 hrs), solution temperatures (23, 37 and 50 °C) and solution refreshment (refreshed and non-refreshed) were studied. It was found that the weight change of the samples increased with increasing soaking times at all temperatures regardless of solution refreshment. Using refreshed solution resulted in greater increase in weight change than using non-refreshed solution. In the case of solution temperature, two opposite trends were observed depending on the solution refreshment. Increasing solution temperatures in non-refreshed solution exhibited a decrease in the weight change whereas the increase in the weight change was observed when using refreshed solution. This could be related to the competition between the dissolution of 3D printed HA and the deposition of new calcium phosphate crystals during biomimetic process. Octacalcium phosphate and HA were found to be the main phases of biomimetically deposited 3D printed HA.

Rapid Biomimetic Coating of Calcium Phosphate on Titanium: Effect of Soaking Time, Temperature and Solution Refreshing

The influence of employing three different solution temperatures (23, 37 and 50 °C), three soaking times (2, 4 and 6 h) and two solution refreshing methods (Refreshed or Non-refreshed) in rapid biomimetic coating process on phase composition, functional groups, coating content and microstructure of the resulted coating was studied. Increasing soaking times and temperature increased the coating content in all cases regardless of the use of refreshed or non-refreshed accelerated calcium phosphate solution. The use of non-refreshed solution resulted in the lower rate of coating than that of refreshed solution at all coating temperatures. However, all coatings similarly comprised octacalcium phosphate and hydroxyapatite as main phases and the microstructure consisted of sharp and interconnected plate-like crystals vertically grown on the surface of titanium. However, two types of crystal structure were produced. Low solution temperature resulted in isolated spheroids while uniform and distributed crystal structure was produced by using high solution temperature. This could be related to the difference in nucleation and precipitation rate formed in rapid biomimetic coating process as a result of the interplay between temperature and ionic strength of the solutions.

In Vitro Resorbability of Three Different Processed Hydroxyapatite

Hydroxyapatite has been used as bone substitutes in many applications due to its biocompatibility and osteoconductivity. Generally, it is considered to be biostable and shows limited resorption in the body. In some circumstances, resorption of bone substitutes is more desirable since it could accelerate the bone healing process. It is known that processing route is one of the crucial parameters that could affect the properties of materials. Three different processes were employed in this study to fabricate hydroxyapatite samples including low temperature transformation of three-dimensionally printed calcium sulfate (HA1), high temperature sintering of three-dimensionally printed hydroxyapatite (HA2) and high temperature sintering of mold pressed hydroxyapatite (HA3). HA1 was found to contain high porosity and low crystallinity whereas HA2 had high porosity and high crystallinity. HA3 had low porosity, but high crystallinity. In vitro resorbability of these samples was studied by submerging all the samples in simulated body fluid (SBF) for 1, 7, 14 and 28 days and determining their phase composition, density change, liquid absorption, ions release and microstructure. It was found that HA1 showed the greatest density loss and liquid absorption followed by HA2 and HA3 respectively. Calcium and phosphorus ions in SBF were observed to decrease with submerging times for HA1 and HA2, but remained constant for HA3. SEM studies showed that new calcium phosphate crystals were found to form on the surface of the HA1 and HA2 samples whereas none was found on HA3. These results suggested that HA1 had the greatest resorbability and calcium phosphate crystals forming ability on its surface followed by HA2 and HA3 respectively. Therefore, porosity and crystallinity of the samples resulting from different processing routes are important factors for in vitro resorbability of hydroxyapatite.

Preparation of 3DP Hydroxyapatite Composite by Single and Double Pass Poly(ε-caprolactone) Infiltration

Recently, porous hydroxyapatite was fabricated by three dimensional printing (3DP) in coupled with low temperature phosphorization to yield nanosized and low crystalline structure. However, brittleness was an intrinsic drawback for some foreseen applications. Polymer infiltration aiming to improve the toughness and mechanical integrity was thus carried out using biodegradable poly(ε-caprolactone) (PCL) as an infiltrant since it has shown good biocompatibility together with a high elongation and energy to failure as compared to other medical polymers. Three routes of infiltration were performed including melt infiltration of low molecular weight PCL (Mw ˜ 10,000), solution infiltration by 10 % high molecular weight PCL (Mw ˜ 80,000) and the combination of both melt and solution infiltration of low and high molecular weight PCL. The combination of low and high MW infiltration yielded the greatest increase in the mechanical properties and followed by the melt infiltration of low molecular weight PCL while the use of high MW infiltration yielded limited enhancement. After immersing in simulated body fluid (SBF), no significant changes in flexural properties were seen for both hydroxyapatite and high molecular weight infiltrated sample. However, flexural strength and strain at break of low molecular weight infiltrated sample largely dropped after 7 days of immersion to be closed to those of hydroxyapatite and high molecular weight infiltrated sample. The flexural properties of high-low infiltrated sample also decreased after immersion, but to a less degree and still maintained the greatest values amongst all samples. This could be associated to the difference in degradation of different molecular weight of PCL and the content of polymer infiltration induced by different infiltration routes. Calcium and phosphorus ions in the SBF were quantified and observed to be consumed continuously during immersion for all samples. Newly formed apatite crystals were observed to form on the surface of the infiltrated composites signifying that infiltration did not hinder the bioactivity of the composites.