Highly-Bioreactive Silica-Based Mesoporous Bioactive Glasses Enriched with Gallium(III).

Beneficial effects in bone cell growth and antibacterial action are currently attributed to Ga3+ ions. Thus, they can be used to upgrade mesoporous bioactive glasses (MBGs), investigated for tissue engineering, whenever they released therapeutic amounts of gallium ions to the surrounding medium. Three gallium-enriched MBGs with composition (in mol %) xSiO2-yCaO-zP2O5-5Ga2O3, being x = 70, y = 15, z = 10 for Ga_1; x = 80, y = 12, z = 3 for Ga_2; and x = 80, y = 15, z = 0 for Ga_3, were investigated and compared with the gallium-free 80SiO2-15CaO-5P2O5 MBG (B). 29Si and 31P MAS NMR analyses indicated that Ga3+ acts as network modifier in the glass regions with higher polymerization degree and as network former in the zones with high concentration of classical modifiers (Ca2+ ions). Ga_1 and Ga_2 exhibited a quick in vitro bioactive response because they were coated by an apatite-like layer after 1 and 3 days in simulated body fluid. Although we have not conducted biological tests in this paper (cells or bacteria), Ga_1 released high but non-cytotoxic amounts of Ga3+ ions in Todd Hewitt Broth culture medium that were 140 times higher than the IC90 of Pseudomonas aeruginosa bacteria, demonstrating its potential for tissue engineering applications.

SiO2-CaO-P2O5 Bioactive Glasses: A Promising Curcuminoids Delivery System.

In this paper, we report the study of the loading and the release of curcuminoids by bioactive glasses (BG) and mesoporous bioactive glasses (MBG). Through a detailed spectroscopic study, it was possible to determine the amount and the type of molecules released in water and in simulated body fluid (SBF). In particular, curcumin and K2T21 show a good ability to be released in di-keto and keto-enolic form, depending from the pH. However, after 24 h, the amount of pristine curcumin release is very low with a consequent increment of degradation products derived by curcuminoids. The presence of -OH groups on curcuminoids is a fundamental pre-requisite in order to obtain a high loading and release in polar solution such as water and SBF. The substrate on which we loaded the drugs does not seem to affect significantly the loading and the release of the drugs. The environment, instead, affects the release: for all the drugs, the release in SBF, buffered at pH of 7.4, is slightly worse than the release in water (basic pH values).

Evidence of catalase mimetic activity in Ce(3+)/Ce(4+) doped bioactive glasses.

The ability of Ce-containing bioactive glasses to inhibit oxidative stress in terms of reduction of hydrogen peroxide, by mimicking the catalase enzyme activity is demonstrated here for the first time. The antioxidant properties of three bioactive glasses containing an increasing amount of CeO2 have been evaluated by following the degradation of hydrogen peroxide with time after immersion in H2O2 aqueous solutions with different concentration. XPS and UV-vis measurements allowed us to determine the Ce(3+)/Ce(4+) ratio in the bulk and on the glass surface, and to correlate it with the ability of the samples to show catalase mimetic activity. Interestingly, we have found that the bioactive glass with composition 23.2Na2O-25.7CaO-43.4SiO2-2.4P2O5-5.3CeO2 immersed in 0.1 M H2O2 aqueous solution is able to degrade 90% of it in 1 week. The reduction in bioactivity of the glasses with increasing CeO2 content is here rationalized in terms of a lower amount of phosphate groups available for the hydroxyapatite layer formation, after binding with cerium ions. In fact, classical molecular dynamics simulations revealed that the addition of CeO2 leads to the formation of cerium phosphate rich regions. The formation of an insoluble CePO4 crystalline phase is also observed by XRD analysis after thermal treatment of the glass samples.

Conjugation of amino-bioactive glasses with 5-aminofluorescein as probe molecule for the development of pH sensitive stimuli-responsive biomaterials.

Bioceramics, such as silica-based glasses, are widely used in bone and teeth restoration. Nowadays, the association between nanotechnology and pharmacology is one of the most promising research fields in cancer therapy. The advanced processing methods and new chemical strategies allow the incorporation of drugs within them or on their functionalized surfaces. Bioceramics can act as local drug delivery systems to treat bone and teeth diseases. The present paper reports data related to the development of a pH-stimuli responsive bioactive glass. The glass conjugation with 5-aminofluorescein (5-AF), through a pH-sensitive organic spacer, allows to produce a pH-responsive bioactive biomaterial: when it is exposed to specific pH changes, it can favour the release of 5-AF directly at the target site. 5-AF has been chosen as a simple, low cost, non toxic model to simulate doxorubicin, an anticancer drug. As doxorubicin, 5-AF contains an amino group in its structure in order to form an amide bond with the carboxylic functionalities of the glass. Raman spectroscopy and thermal analysis confirm the glass conjugation of 5-AF by means of an amide bond; the amount of 5-AF loaded was very high (≈ 65 and 44 wt%). The release tests at two different pH (4.2 and 7.4) show that the amount of released 5-AF is higher at acid pH with respect to physiological one. This preliminary datum evidenced that a pH-sensitive drug delivery system has been developed. The low amount of 5-AF released (<1 wt% of the total 5-AF) is due to the very low solubility of 5-AF in aqueous medium. This disadvantage, may be overcome in a dynamic environment (physiological conditions), where it is possible to obtain a drug release system ensuring an effective therapeutic dose for long times and, at the same time, avoiding the drug toxicity.

New formulation of functionalized bioactive glasses to be used as carriers for the development of pH-stimuli responsive biomaterials for bone diseases.

The aim of the present contribution is to prepare a functionalized bioactive glass potentially useful as prosthetic material, but also able to release organic molecules in response to a change of the pH environment. By this approach it is possible to develop devices which can be used for a triggered drug release in response to specific stimuli; this is an attractive research field, in order to avoid either systemic and/or local toxic effects of drugs. In particular, in the present paper we report data related to the development of a new formulation of bioactive glasses, their functionalization with organic molecules to obtain a pH-sensitive bond, their physicochemical characterization and in vitro bioactivity in simulated biological fluids (SBF), and organic molecule delivery tests at different pH. The glass functionalization, by means of a covalent reaction, allows us to produce a model of pH-responsive bioactive biomaterial: when it is exposed to specific pH changes, it can favor the release of the organic molecules directly at the target site. Cysteamine and 5-aminofluorescein are used as model molecules to simulate a drug. The materials, before and after the different functionalization steps and in vitro release tests at different pH, have been characterized by means of different experimental techniques such as X-ray powder diffraction (XRPD), Raman, FTIR and fluorescence spectroscopies, N2 adsorption, thermogravimetric (TGA) and elemental analysis.

In vitro antibacterial capacity and cytocompatibility of SiO2-CaO-P2O5 meso-macroporous glass scaffolds enriched with ZnO.

Zn2+ ions exhibit osteogenic, angiogenic and antimicrobial properties. For this reason, they are often added in small amounts to bioceramics being investigated for bone tissue engineering. In this paper, the cytocompatibility and antibacterial properties of 80% SiO2-15% CaO-5% P2O5 (mol%) mesoporous bioactive glass (MBG) scaffolds substituted with 4.0% and 7.0% of ZnO were studied and compared with the Zn-free scaffold. Cell proliferation, morphology, differentiation and cytotoxic effects of Zn2+ ions released from the samples were examined by culturing human osteoblast-like cells (HOS) osteoblasts both in the presence of sample extracts and on the scaffold surface. The bacterial inhibition capacity of the scaffolds was explored by using Gram-positive Stapylococcus aureus bacteria, responsible for numerous infections in orthopedic surgery, to simulate a severe infection. Our results show that the Zn-MBG scaffolds possess a hierarchical meso-macropore structure suitable for osteoblast growth. Furthermore, the amount of Zn2+ released from the scaffold with 4.0% ZnO was found to be more favorable for HOS cell development than that released from the scaffold including 7.0% ZnO. Zn2+ released to the medium from both scaffolds exhibited antibacterial properties against S. aureus. Thus, the cytocompatibility and the antibacterial ability exhibited by the MBG scaffold containing 4.0% ZnO make it a suitable candidate for bone regeneration applications.

Gallium-containing phosphosilicate glasses: functionalization and in-vitro bioactivity.

A gallium containing glass 45.7SiO2·24.1Na2O·26.6CaO·2.6P2O5·1.0Ga2O3 (referred to as "Ga1.0") and a parent Ga-free glass 46.2SiO2·24.3Na2O·26.9CaO·2.6P2O5 (hereinafter represented as "H"), corresponding to Bioglass® 45S5, were functionalized with Tetraethoxysilane (TEOS) and (3-Aminopropyl)triethoxysilane (APTS) in order to improve their ability to bond with biomolecules, such as drugs, proteins, and peptides. Functionalization with TEOS and APTS promoted the increment in OH groups and formation of NH2 groups on the glass surface, respectively. The presence of OH or NH2 groups was investigated by means of IR spectroscopy and elemental analysis. Moreover, in vitro study of these functionalized glasses was performed in simulated body fluid (SBF) so as to investigate the effect of functionalization on the bioactive behavior of H and Ga1.0. The results showed that the functionalization was obtained along with maintaining their bioactivity. The surfaces of both functionalized glasses were covered by a layer of apatite within 30 days of SBF immersion. In addition, CaCO3 was also identified on the surface of APTS functionalized glasses. However, no gallium release was detected during SBF soaking.

Gold-containing bioactive glasses: a solid-state synthesis to produce alternative biomaterials for bone implantations.

A new melted bioactive system containing gold nanoparticles (AuNPs) was prepared exploiting a post-synthesis thermal treatment that allows one to modify crystal phases and nature, shape and distribution of the gold species in the glass-ceramic matrix as evidenced by UV-visible spectroscopy, transmission electron microscopy and powder X-ray diffraction analysis. In human MG-63 osteoblasts the presence of Au(n)(+) species caused an increase of lactate dehydrogenase leakage and malonyldialdehyde production, whereas Hench's Bioglass HAu-600-17 containing only AuNPs did not cause any effect. In addition, HAu-600-17 caused in vitro hydroxyapatite formation and an increase of specific surface area with a controlled release of gold species; this material is then suitable to be used as a model system for the controlled delivery of nanoparticles.

Mesoporous bioactive scaffolds prepared with cerium-, gallium- and zinc-containing glasses.

Mesoporous bioactive glass scaffolds (MBG_Scs), based on 80% SiO(2)-15% CaO-5% P(2)O(5) (in mol.%) mesoporous sol-gel glasses substituted with Ce(2)O(3), Ga(2)O(3) (both 0.2% or 1.0%) and ZnO (0.4% or 2.0%), were synthesized by combination of evaporation-induced self-assembly and rapid prototyping techniques. Cerium, gallium and zinc trace elements were selected because of their inherent beneficial biological properties. Fabricated scaffolds were characterized and compared with unsubstituted scaffold (B_Sc). All of them contained well interconnected ultralarge pores (pores >400 µm) ideal for vascular ingrowth and proliferation of cells. Macropores of size 100-400 µm were present inside the scaffolds. In addition, low-angle X-ray diffraction showed that B_Sc and scaffolds with substituent contents up to 0.4% exhibited ordered mesoporosity useful for hosting molecules with biological activity. The textural properties of B_Sc were a surface area of 398 m(2) g(-1), a pore diameter of 4.3 nm and a pore volume of 0.43 cm(3) g(-1). A slight decrease in surface area and pore volume was observed upon substitution with no distinct effect on pore diameter. In addition, all the MBG_Scs except 2.0% ZnO_Sc showed quite quick in vitro bioactive response. Hence, the present study is a positive addition to ongoing research into preparing bone tissue engineering scaffolds from bioceramics containing elements of therapeutic significance.

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, Mg(2+) and Sr(2+), when substituted into the structure of hydroxyapatite (Ca(10)(PO(4))(6)(OH)(2),HA). The substituted samples were synthesized by an aqueous precipitation method that involved the addition of Mg(2+)- and Sr(2+)-containing precursors to partially replace Ca(2+) ions in the apatite structure. Eight substituted HA samples with different concentrations of single (only Mg(2+)) or combined (Mg(2+) and Sr(2+)) 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 (N(2) 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 ß-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/ß-TCP ratio.

Evaluation of the behaviour of fluorine-containing bioactive glasses: reactivity in a simulated body fluid solution assisted by multivariate data analysis.

Potentially bioactive fluorine-containing glasses of formula 46.2SiO(2)·24.3Na(2)O·(26.9-x)CaO·2.6P(2)O(5)·xCaF(2) [x = (0), 5, 10, 15] have been studied: the study was carried out as a function of fluorine percentage, dimensions and time of soaking in SBF. The results are compared to those obtained in the same conditions for Bioglass(®) 45S5. Due to the high number and different kind of variables/conditions explored by this set of data, the results are rationalized for the first time by means of multivariate data analysis (MDA); in this way it is possible to classify the behaviour of bioglasses toward bioactivity. The presence of fluorine does not inhibit the formation of HA; in particular, for a fast bioactivity (in term of HA crystallization) it will be better to have large particle size or slabs, while for a fast dissolution fine particle sizes should be preferred.

Gallium-containing phospho-silicate glasses: synthesis and in vitro bioactivity.

A series of Ga-containing phospho-silicate glasses based on Bioglass 45S5, having molar formula 46.2SiO2·24.3Na2O·26.9CaO·2.6P2O5·xGa2O3 (x=1.0, 1.6, 3.5), were prepared by fusion method. The reference Bioglass 45S5 without gallium was also prepared. The synthesized glasses were immersed in simulated body fluid (SBF) for 30 days in order to observe ion release and hydroxyapatite (HA) formation. All Ga-containing glasses maintain the ability of HA formation as indicated by main X-ray diffractometric peaks and/or electronic scanning microscopy results. HA layer was formed after 1 day of SBF soaking in 45S5 glass containing up to 1.6% Ga2O3 content. Moreover, gallium released by the glasses was found to be partially precipitated on the glass surface as gallium phosphate. Further increase in gallium content reduced the ion release in SBF. The maximum of Ga(3+) concentration measured in solution is ~6 ppm determined for 3.5% Ga2O3 content. This amount is about half of the toxic level (14 ppm) of gallium and the glasses release gallium till 30 days of immersion in SBF. Considering the above results, the studied materials can be proposed as bioactive glasses with additional antimicrobial effect of gallium having no toxic outcome.

Functionalization of sol gel bioactive glasses carrying Au nanoparticles: selective Au affinity for amino and thiol ligand groups.

It is demonstrated here that bioactive glasses containing Au nanoparticles (AuNPs) can be selectively functionalized with small molecules carrying either amino or thiol groups by simply varying the temperature and pH of the functionalization batch. The results evidence the following. (i) At room temperature (RT), no functionalization of Au-free glass occurs, whereas in the case of glasses containing AuNPs, stable linkages form only with amino groups, as in this condition Au does not bind with either thiol or hydroxyl groups. The RT functionalization with cysteine and cystine confirms the preferential functionalization through the amino groups, while the -SH groups are oxidized to S-S bridges. (ii) The functionalization with cysteine and cystine, compared at pH = 5, 9, and 12, is shown not to take place at pH = 5 and to be hindered by the glass matrix dissolution at pH = 12 (with consequent release of AuNPs), while the best results are obtained at pH = 9. (iii) For the effect of reaction temperature, at 4 °C it is possible to obtain a strong Au-S interaction, whereas at RT, a weak Au-N linkage is formed. These results should allow production, in a selective way, of different bonds exhibiting different strengths and, consequently, different release times in solution, with a wide range of possible applications (for instance, weak Au-N bonds in the case of drug delivery, strong Au-S bonds in protein immobilization).

Bioactive glasses containing Au nanoparticles. Effect of calcination temperature on structure, morphology, and surface properties.

Bioactive glasses containing gold nanoparticles (AuNPs) have been synthesized via the sol-gel route using HAuCl(4) x 3 H(2)O as gold precursor. The formation process of AuNPs was studied as a function of the thermal treatment, which induces nucleation of Au particles and influences their nature, optical properties, shape, size, and distribution. The physicochemical characterization indicates that the sample treated at 600 degrees C presents the best characteristics to be used as a bioactive material, namely high surface area, high amount of AuNPs located at the glass surface, presence of micropores, and abundant surface OH groups. In the case of samples either aged at 60 degrees C or calcined at 150 degrees C, AuNPs just begin their formation, and at this stage the gel is not completely polymerized and dried yet. A thermal treatment at higher temperatures (900 degrees C) causes the aggregation of AuNPs, forming "AuMPs" (i.e., Au microparticles) in a densified glass-ceramic material with low surface area, absence of pores, and low number of surface OH groups. These features induce in the glass-ceramic materials treated at high-temperatures a lower bioactivity (evidenced by SBF reaction), as compared with that exhibited by the glass samples treated at 600 degrees C.

In vitro and in vivo behaviour of zinc-doped phosphosilicate glasses.

The aim of this work was to study the behaviour of zinc-doped phosphosilicate glasses based on Bioglass 45S5. In vitro (in simulated body fluid), the reactivity was analysed by means of inductively coupled plasma spectrometry, environmental scanning electron microscopy-energy-dispersive spectroscopy (ESEM-EDS) and X-ray diffraction. In vivo (a rat implanted with glass), the reactivity and the tissue behaviour were analysed by conventional histology, histochemistry, microradiography and ESEM-EDS. The in vivo behaviour matches that in vitro perfectly; they show comparable glass degradation processes and rates, ruled by the amount of zinc in the glass. The reaction mechanism for the formation of a polymerized silica layer superimposed with a peripheral calcium phosphate layer is clearly substantiated by ESEM-EDS investigations. The crystallization of a biologically active hydroxyapatite (HA) layer is observed in both cases; the in vitro experiment shows the presence of HA after 4 days.

Crystallization kinetics of bioactive glasses in the ZnO-Na2O-CaO-SiO2 system.

The crystallization kinetics of Na(2)O.CaO.2SiO(2) (x = 0) and 0.68ZnO.Na(2)O.CaO.2SiO(2) (x = 0.68, where x is the ZnO stoichiometric coefficient in the glass formula) bioactive glasses have been studied using both nonisothermal and isothermal methods. The results obtained from isothermal XRPD analyses have showed that the first glass crystallizes into the isochemical Na(2)CaSi(2)O(6) phase, whereas the Na(2)ZnSiO(4) crystalline phase is obtained from the Zn-rich glass, in addition to Na(2)CaSi(2)O(6). The activation energy (Ea) for the crystallization of the Na(2)O.CaO.2SiO(2) glass is 193 +/- 10 and 203 +/- 5 kJ/mol from the isothermal in situ XRPD and nonisothermal DSC experiments, respectively. The Avrami exponent n determined from the isothermal method is 1 at low temperature (530 degrees C), and its value increases linearly with temperature increase up to 2 at 607 degrees C. For the crystallization of Na(2)CaSi(2)O(6) from the Zn-containing glass, higher values of both the crystallization temperature (667 and 661 degrees C) and Ea (223 +/- 10 and 211 +/- 5 kJ/mol) have been found from the isothermal and nonisothermal methods, respectively. The Na(2)ZnSiO(4) crystalline phase crystallizes at lower temperature with respect to Na(2)CaSi(2)O(6), and the Ea value is 266 +/- 20 and 245 +/- 15 kJ/mol from the isothermal and nonisothermal methods, respectively. The results of this work show that the addition of Zn favors the crystallization from the glass at lower temperature with respect to the Zn-free glass. In fact, it causes an increase of Ea for the Na diffusion process, determined using MD simulations, and consequently an overall increase of Ea for the crystallization process of Na(2)CaSi(2)O(6). Our results show good agreement between the Ea and n values obtained with the two different methods and confirm the reliability of the nonisothermal method applied to kinetic crystallization of glassy systems. This study allows the determination of the temperature stability field of the crystalline phases with the view of creating a different glass ceramic useful in the field of bioactive materials.

Cytotoxicity of zinc-containing bioactive glasses in contact with human osteoblasts.

Bioactive glasses such as Hench's 45S5 have applications to tissue engineering and bone repair: the insertion of zinc has been proposed to improve their bone-bonding ability and to slacken their dissolution in extracellular body fluids. In view of a potential clinical application, we have investigated whether zinc-containing 45S5 (HZ) glasses might be cytotoxic for human MG-63 osteoblasts. In our experimental conditions, after 24h of incubation HZ glasses released significant amounts of Zn(2+) and induced in MG-63 cells release of lactate dehydrogenase (index of cytotoxicity) and the following indexes of oxidative stress: (i) accumulation of intracellular malonyldialdehyde, (ii) increased activity of pentose phosphate pathway, (iii) increased expression of heme oxygenase-1, (iv) increased activity of Cu,Zn-superoxide dismutase, (v) decreased level of intracellular thiols. These effects were inversely related to the zinc content of glass powders, were mimicked by ZnCl(2) solutions and were prevented by either metal chelators (EDTA, NTA) or the antioxidant ascorbate, suggesting that Zn(2+) released fastly from HZ glasses can cause MG-63 cell damage via an oxidative stress. This work highlights the importance of designing Zn-containing bioactive glasses without cytotoxic effects and gives supplementary information about the prooxidant role of zinc in living systems.

Thermodynamic aspects of the adsorption of hexametaphosphate on kaolinite.

The adsorption of hexametaphosphate ion, an important deflocculant used in the ceramic industry, from aqueous solutions onto kaolinite has been studied at different temperatures. The adsorption isotherm follows the Langmuir model: the thermodynamic parameters DeltaG(ads)(0), DeltaH(ads)(0), and DeltaS(ads)(0) were calculated and found to be consistent with an interaction model involving the formation of an inner-sphere complex between HMP and aluminol groups. Also, the dependence of the adsorption behavior on the kaolinite volume fraction has been studied and discussed in term of association processes between the clay particles.

A computational tool for the prediction of crystalline phases obtained from controlled crystallization of glasses.

An automatic tool (named CLUSTER) for the prediction of the most probable crystal phases that can separate from glasses has been developed. The program analyzes the output of molecular dynamics simulations of glasses or glass ceramics, systematically sampling the ratios of the ions in different portions of the simulation box and comparing them to the stoichiometric ratio of compositionally equivalent crystalline phases retrieved from a crystal structure database. The efficacy of the similarity index elaborated has been judged by comparing the results obtained with the crystal phases identified by XRD analysis after thermal treatment in a series of multicomponent potential bioactive glasses and glass ceramics for which the advantages of rational-designed erosion-controlled release is straightforward.