Albert Lebugle

Synthesis and characterization of amino acid-functionalized hydroxyapatite nanorods

Synthesis of aqueous colloids of amino acid-functionalized hydroxyapatite (HAP; Ca10(PO4)6(OH)2) nanoparticles with rod-like morphology and uniform size was achieved by hydrothermal crystallization at 80 °C and pH 9 in highly supersaturated solutions with a Ca2+∶amino acid molar ratio of 1∶2. Stable suspensions of positively charged nanorods, less than 80 nm in length and ca. 5 nm in width, were obtained in the presence of glycine, alanine, serine, lysine and arginine, and together with FTIR spectroscopy data this was consistent with a general model in which the α-carboxylate of the amino acid was preferentially bound to the crystal surfaces. Variations in the size and shape of the nanoparticles with amino acid used were consistent with differences in the strength of binding at the HAP surfaces. In comparison, stable colloids could not be prepared with aspartic acid due to a relatively low surface charge; instead, the precipitate consisted of highly elongated nanorods, ca. 150 × 3.5 nm in dimension. XRD textural analysis suggested a model for nanorod formation based on the oriented aggregation and fusion of primary crystallite domains specifically along the c axis direction. Colloids consisting of HAP/alanine or HAP/arginine nanoparticles were cross-linked in situ by thermal, hydrothermal or chemical polymerisation of surface-attached and unbound amino acids to produce aggregated nanostructures with variable degrees of long-range order.

Europium-doped bioapatite: a new photostable biological probe, internalizable by human cells.

The authors prepared at low temperatures (37 degrees C) a novel inorganic bioprobe. It consisted of mineral nanoparticles of apatitic tricalcium phosphate doped with europium, of size, structure and composition close to those of the mineral part of calcified tissues. In contrast to organic probes which degrade rapidly (photobleaching), the red luminescence of the new probe is photostable. Moreover, this luminescence can be obtained under visible irradiation, which makes it suitable for prolonged examination of live cells. Human pancreatic epithelial cells in culture were incubated with these particles and their internalization was observed by laser scanning confocal microscopy. Transmission electron microscopy and electron microdiffraction analysis confirmed that the particles were internalized retaining their original apatitic structure. This probe may thus be of value for biovectorization.

Study of implantable calcium phosphate systems for the slow release of methotrexate.

The authors prepared and studied systems implantable in bone, for the slow release of an antineoplasic agent, methotrexate (MTX). The systems were made by compaction of a powdered mixture of an apatitic deficient calcium phosphate, dextran and various amounts of MTX. Used as a matrix, this calcium phosphate has outstanding adsorption and compaction properties. It is an osteoconductor and biodegradable. The in vitro study carried out on these systems showed that the release of MTX with time is slow and prolonged due to the phenomena of adsorption/desorption of MTX onto deficient apatite. The composition of the implants changed with time towards that of stoichiometric apatite. The in vivo pilot study was performed by implantation in the external femoral condyle of rabbits. A pharmacokinetic study revealed that the circulating concentration of MTX in the blood was always below toxic levels. Twenty percent of the initial MTX remained in the implants after 7 days. A study of the biocompatibility and bioreactivity showed no local necrosis at any time, while implants degraded and new bone formed simultaneously. These implantable systems seem appear suitable for use immediately.

Apatitic Calcium Orthophosphates and Related Compounds for Biomaterials Preparation

The authors show that to obtain well chemically defined apatitic bioceramics and to know the possible transformations of this material during sintering, it is necessary to prepare a good starting material. Moreover, they show that it is possible to prepare a new organic-inorganic phosphate compound. The precipitation of apatite in an aqueous medium at boiling temperature was studied using the methodology of experimental design. Independent variables were the volume of NH4OH in phosphate solution, the volume of NH4OH in calcium solution, and the time of precipitation; the response was the atomic Ca/P ratio of the obtained precipitate. A continuous variation of this ratio from 1.63 to 1.73 is observed. Implications of this result to the preparation of pure HA: Ca10(PO4)6(OH)2 is given. Moreover, when Ca/P greater than 1.67, HA reacts with Ca(OH)2 (after heating at 1000 degrees C in air for some days) to give rise to a single phase described as a modified HA (MHA), a Ca/P ratio of 1.75, an a value of 9.373 +/- 0.002 A, and a c value of 6.884 +/- 0.002 A. The reactivity (time versus temperature) of the MHA is described. If the precipitation of the calcium phosphate is realized at 37 degrees C in a water-ethanol medium in the presence of A2EP, a new apatite, chemically bonded to the organic molecule by pooling phosphate groups, is obtained.

Structural characteristics of a globular protein investigated by X-ray photoelectron spectroscopy: comparison between a legumin film and a powdered legumin

Films of legumin, a pea protein, were deposited onto a glass support using the Langmuir-Blodgett method, at various surface pressures. XPS study of these films show that their thickness increases with the deposition pressure. At the pressure limits of films stability, the thickness values (respectively 73 and 110 A) are close to the protein dimensions. Layered at low pressure, the oblate protein stands up when pressure increases. Furthermore, XPS study shows that the orientation of the external flexible loops depends on the obtention conditions. Thus, in the case of Langmuir-Blodgett films, hydrophobic residues are turned towards the external surface, and the hydrophilic ones towards the glass substrate. But, in the opposite, when protein is obtained by lyophilization, the hydrophilic residues are orientated outsides. It seems possible to determine by XPS the nature of the residues which give to the protein its reactivity, since they are located at its external surface.

Adsorption d'acides aminés sur des phosphates de calcium carbonatés analogues aux tissus calcifiés

Abstract The adsorption of alanine and phenylalanine onto apatites, containing simultaneously HPO 4 2− and CO 3 2− ions in various amount, was studied. These apatites are similar to the mineral part of calcified tissues at different stages of its maturation. The adsorption isotherms are Langmuirian in all cases whatever the amounts of HPO 4 2 or CO 3 2− . However, these amounts play a role in the adsorption properties of these apatites. The largest amounts of aminoacids were adsorbed by the compounds with the highest HPO 4 2− content. The influence of the presence of PO 4 3− ions in the incubation media was studied. These ions hinder the adsorption process by competing with the aminoacid carboxylate groups for the calcium ions at the surface of the solid. The nature of the aminoacid lateral chain, especially its size, plays a role in the adsorption. The amount of aminoacid bound at saturation decreases as the size of the side chain increases.

Organization of a globular protein film on solid surfaces: an X-ray photoelectron spectroscopy study

Abstract The organization of legumin films deposited on different solid substrates using the Langmuir-Blodgett technique has been studied by X-ray photoelectron spectroscopy (XPS). The XPS data provide information on the identity of the chemical elements present as well as on chemical bonding. From the decomposed XPS C 1s spectra, the different types of carbon (peptidic, aliphatic etc.) have been identified. The variation in atomic ratios as a function of the take-off angle shows a structural orientation of the protein films in which the hydrophobic aliphatic amino acids are at the external surface whereas the hydrophilic amino acids are oriented towards the solid substrate. Moreover, the attenuation in the S 2p signal intensity with decreasing take-off angle due to the protein layer suggests that the adsorbed protein forms a continuous overlayer on a glass substrate whereas the weak dependence of the F 1s signal intensity with the take-off angle suggests an incomplete coverage of the polytetrafluoroethylene substrate.

Influence du séchage sur la composition de phosphates de calcium carbonatés, analogues osseux

Abstract Carbonated calcium phosphates were prepared at 37 °C by coprecipitation, in partially aqueous medium (lowered dielectric constant). Then they were either freeze dried, or oven dried at 40, 60 or 80 °C. The study of these samples by XR diffraction and IR spectroscopy showed that their crystallisation into apatite depends both on their composition, and on their drying temperature. Crystallisation decreases when the drying temperature decreases or carbonate content increases in these compounds and the samples finally become amorphous. In addition, chemical analysis showed that their carbonate content, expressed by the CO3/PO4 atomic ratio, decreased when the drying temperature was raised. The decrease was due to reaction of CO3 ions with HPO4 during drying. Although this reaction has already been observed near 200–500 °C in apatites, it is the first time that it was noted at temperatures as low as 40 °C. Apatites obtained in that way present a potential for use as bone substitutes, due to their analogy with bone. Nevertheless before any applications can be done, their synthesis must be fully controlled especially during drying, in order to get materials of well defined composition.