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International audience; In vivo, collagen I, the major structural protein in human body, is found assembled into fibrils. In the present work, we study a high concentrated collagen sample in its soluble, fibrillar, and denatured states using one and two dimensional \H-1\-C-13 solid-state NMR spectroscopy. We interpret C-13 chemical shift variations in terms of dihedral angle conformation changes. Our data show that fibrillogenesis increases the side chain and backbone structural complexity. Nevertheless, only three to five rotameric equilibria are found for each amino acid residue, indicating a relatively low structural heterogeneity of collagen upon fibrillogenesis. Using side chain statistical data, we calculate equilibrium constants for a great number of amino acid residues. Moreover, based on a C-13 quantitative spectrum, we estimate the percentage of residues implicated in each equilibrium. Our data indicate that fibril formation greatly affects hydroxyproline and proline prolyl pucker ring conformation. Finally, we discuss the implication of these structural data and propose a model in which the attractive force of fibrillogenesis comes from a structural reorganization of 10 to 15% of the amino acids. These results allow us to further understand the self-assembling process and fibrillar structure of collagen.

5th International Conference of theă Federation-of-European-Zeolite-Associations (FEZA), Valencia, SPAIN, JULă 03-07, 2011; International audience; We present in this communication that phenyl phosphonic acid can be efficiently loaded in mesoporous SBA-15 and aminopropyl-modified SBA powdered samples through the incipient wetness impregnation method. High amount of phosphonic acid can be reach up to 380 mg/g of sample. We use multinuclear solid state NMR as a method of choice for the indeep characterization of the samples. Thus we demonstrate that phosphonic acid molecules do not crystallize inside the pores. The molecules are highly mobile in SBA-15 because they are submitted to a confinement effect due to the mesoscopic size of the pores and consequently they exhibit a weak interaction with the silica walls. In the case of aminopropyl-modified SBA material, we show that the molecules are rigid and that they are in strong interaction with the aminopropyl groups. Moreover, a 2D double quantum 1H NMR experiment recorded at high field and high spinning speed permit to propose a model of the phosphonate-aminopropyl interaction. The increase in spectral resolution due to the combination of high magnetic field and fast MAS rate allows also the assignment of 1H resonances in aminopropyl-modified SBA matrix and notably allows the assignment of the protons resonance of the amino group.

International audience; The continuous-flow hyperpolarised (HP)-Xe-129 NMR and magic angle spinning-pulsed field gradient (MAS-PFG) NMR techniques have been used for the first time to study the distribution and the dynamics of ibuprofen encapsulated in MCM-41 with two different pore diameters.

International audience; A comparative study of the degradation in a model phosphate buffered saline (PBS) medium of mesoporous mixed silica–zirconia oxides and hybrid organosilica materials is reported, and their thermodynamic and kinetic stability are discussed. Thin film morphology was used to monitor the degradation rates of all these materials for the first time directly in liquid and undersaturated medium. The introduction of zirconium centers in mesoporous materials strongly inhibits degradation and allowed the self-limitation of soluble silica concentration in the immediate environment of the vector. This avoids renucleation of potentially toxic silica small nanoparticles with uncontrolled surface chemistry in the release medium. Chemical modifications (hybridation or zirconia doping) of mesoporous silica allow the fine-tuning of its degradation from hours to days. Methylated thin films are highly stable when the functionalization ratio is high (50%). The hybridation by mercaptopropyl and aminopropyl moieties can be successfully performed without any high thermal treatment, allowing not only slowing down silica dissolution but also opening the possibility to introduce in one-pot other thermally fragile molecules into the vector. Dissolution rates K measured for the first time at different biologically relevant temperatures follows an Arrhenius law: K = KoExp(−Ea/RT). Stabilities of these chemically modified silicas are mainly associated with a strong decrease of the pre-exponential factor K0 of modified silicas. Counterintuitively, activation energies Ea decrease with increasing the hybrid/silica or the zirconia/silica ratios.

First-principles calculations of NMR parameters in solids are of crucial importance in chemistry and materials science. The acquisition of a high-resolution spectrum, however, does come with a cost, that is, the suppression of all anisotropies. The interactions that affect NMR spectra are inherently anisotropic and can be represented mathematically by second- and fourth-rank tensors. A third approach is to employ electronic structure techniques that model extended solids using periodic boundary conditions. The linear augmented plane-wave approach (LAPW) through the Wien series of codes has long been used to compute quadrupolar coupling constants. Recoupling experiments are introduced in the context of the measurement of interaction parameters, that is, internuclear distances and anisotropies. The main goal of this section is to highlight the fundamental concepts involved, and the methods required, for experimental measurement parameters. Particular emphasis is placed on important classes of systems such as organic structures and assemblies, inorganic architectures, polymers and hybrids, biomaterials, and nanomaterials.

International audience; In the context of nanomedicine, liposils (liposomes and silica) have a strong potential for drug storage and release schemes: such materials combine the intrinsic properties of liposome (encapsulation) and silica (increased rigidity, protective coating, pH degradability). In this work, an original approach combining solid state NMR, molecular dynamics, first principles geometry optimization, and NMR parameters calculation allows the building of a precise representation of the organic/inorganic interface in liposils. {(1)H-(29)Si}(1)H and {(1)H-(31)P}(1)H Double Cross-Polarization (CP) MAS NMR experiments were implemented in order to explore the proton chemical environments around the silica and the phospholipids, respectively. Using VASP (Vienna Ab Initio Simulation Package), DFT calculations including molecular dynamics, and geometry optimization lead to the determination of energetically favorable configurations of a DPPC (dipalmitoylphosphatidylcholine) headgroup adsorbed onto a hydroxylated silica surface that corresponds to a realistic model of an amorphous silica slab. These data combined with first principles NMR parameters calculations by GIPAW (Gauge Included Projected Augmented Wave) show that the phosphate moieties are not directly interacting with silanols. The stabilization of the interface is achieved through the presence of water molecules located in-between the head groups of the phospholipids and the silica surface forming an interfacial H-bonded water layer. A detailed study of the (31)P chemical shift anisotropy (CSA) parameters allows us to interpret the local dynamics of DPPC in liposils. Finally, the VASP/solid state NMR/GIPAW combined approach can be extended to a large variety of organic-inorganic hybrid interfaces.

Hybrid organic-inorganic approaches are used for the synthesis of bifunctional proton exchange membrane fuel cell (PEMFC) membranes owing to their ability to combine the properties of a functionalized inorganic network and an organic thermostable polymer. We report the synthesis of both sulfonic and phosphonic acid functionalized mesostructured silica networks into a poly(vinylidenefluoride-co-hexafluoropropylene) (poly(VDF-co-HFP) copolymer. These membranes, containing different amounts of phosphonic acid and sulfonic acid groups, have been characterized using FTIR and NMR spectroscopy, SA-XRD, SAXS, and electrochemical techniques. The proton conductivity of the bifunctional hybrid membranes depends strongly on hydration, increasing by two orders of magnitude over the relative humidity (RH) range of 20 to 100%, up to a maximum of 0.031 S cm(-1) at 60 °C and 100% RH. This value is interesting as only half of the membrane conducts protons. This approach allows the synthesis of a porous SiO(2) network with two different functions, having -SO(3)H and -PO(3)H(2) embedded in a thermostable polymer matrix.

In 2001, Pickard and Mauri implemented the gauge including projected augmented wave (GIPAW) protocol for first-principles calculations of NMR parameters using periodic boundary conditions (chemical shift anisotropy and electric field gradient tensors). In this paper, three potentially interesting perspectives in connection with PAW/GIPAW in solid-state NMR and pure nuclear quadrupole resonance (NQR) are presented: (i) the calculation of J coupling tensors in inorganic solids; (ii) the calculation of the antisymmetric part of chemical shift tensors and (iii) the prediction of (14)N and (35)Cl pure NQR resonances including dynamics. We believe that these topics should open new insights in the combination of GIPAW, NMR/NQR crystallography, temperature effects and dynamics. Points (i), (ii) and (iii) will be illustrated by selected examples: (i) chemical shift tensors and heteronuclear (2)J(P-O-Si) coupling constants in the case of silicophosphates and calcium phosphates [Si(5)O(PO(4))(6), SiP(2)O(7) polymorphs and α-Ca(PO(3))(2)]; (ii) antisymmetric chemical shift tensors in cyclopropene derivatives, C(3)X(4) (X = H, Cl, F) and (iii) (14)N and (35)Cl NQR predictions in the case of RDX (C(3)H(6)N(6)O(6)), β-HMX (C(4)H(8)N(8)O(8)), α-NTO (C(2)H(2)N(4)O(3)) and AlOPCl(6). RDX, β-HMX and α-NTO are explosive compounds.

In this paper we present an NMR methodology to characterize small organic molecules confined in mesoporous materials. In particular, we demonstrate that NMR techniques issued from solution state NMR are well suited to characterize benzoic acid encapsulated in hexagonally ordered mesoporous silica MCM-41 possessing two different averaged pore sizes (30 and 100 A). As evidenced by differential scanning calorimetry, entrapped benzoic acid molecules are highly mobile at room temperature due to confinement effect and possess a glass phase transition temperature around −55 °C. Thus, the 13C NMR characterization of encapsulated molecules has to be adapted to that particular behavior. In particular, the cross-polarization technique traditionally used in solid state NMR to record 13C magic angle spinning (MAS) spectra is of poor efficiency due to weak 1H−13C dipolar interaction. Nevertheless, the presence of 1H−13C cross-relaxation phenomenon (nuclear Overhauser effect, NOE) allows us to record 13C spectra through...

We present a simple one-pot crystallization method, inspired by biological conditions, for the synthesis of hydroxyapatite (Ca5(PO4)3OH) nanocrystals. The reaction proceeds via NH3 vapor diffusion into a CaCl2−NaH2PO4 mixed solution that is free of any organic additives. The advantage of relying on acidic calcium-phosphate precursors here is, first, that the reaction can be performed at room temperature within a short time and without direct pH control and, second, that it does not produce any secondary phases or byproduct. Furthermore, the addition of NaHCO3 to the salt solution or the introduction of (NH4)2CO3 instead of NH3 lead, respectively, to the precipitation of B- or A-type carbonate-apatite phases according to the FT-IR data. Multinuclear solid state NMR studies and especially 13C CP experiments allow an in-depth characterization showing the presence of A/B substitutions in carbonated samples as well and indicate a close similarity to deproteinated bovine compact bone. A precipitation mechanism ...

International audience; This article illustrates the suitability of solid-state NMR for the in-depth description of organic-inorganic interfaces in silica-based hybrid materials. The examples were selected as representatives of some of the most studied families of hybrid materials, such as functionalized silica particles and templated mesostructured silica.; Cette publication illustre l’adéquation des techniques de RMN à l’état solide pour la caractérisation en profondeur des interfaces organiques-inorganiques présentes dans les matériaux hybrides à base de silice. Les exemples décrits ont été choisis comme représentatifs de familles de matériaux hybrides parmi les plus étudiées, comme les silices fonctionnalisées et les silices mésostructurées par des templates.

Various Si/carbon/polymer composite electrodes were prepared to better understand the influence of the Si−polymer interactions on the stability of the Li−Si reaction and especially the superior performances of CMC-based (carboxy−methyl−cellulose) composites despite the large volume changes of the Si particles upon cycling. Via the modification of the composites formulation, the nature of the polymer, the nature and the amount of the substituting groups and the surface chemistry of the Si particles, together with the use of various characterization techniques (TEM, SEM, NMR−MAS, infrared spectroscopy, TGA, etc.) we could propose that the performances of the Si/Csp/CMC composite electrodes are nested in both the porous texture of the electrode and in the nature of the Si−polymer chemical bonding. A self-healing process of the rather strong Si−CMC hydrogen bonding which can accommodate textural stresses and can evolve during cycling is proposed to be critical for Si-based electrode performances. This better ...

We present in this communication the preparation and the solid state NMR characterization of phenyl phosphonic acid encapsulated both in pure and aminopropyl-modified SBA-15 mesoporous silica materials. The 31P and 1H MAS studies revealed two radically different behaviors of the confined molecules. The included phosphonic acid in SBA-15 is submitted to a confinement effect that implies a weak interaction with the SiO2 surface and a relative mobility at room temperature. On the contrary, phenyl phosphonic acid molecules in the aminopropyl modified sample possess a strong interaction with the hybrid surface of the material. This finding is supported by a two dimensional double-quantum 1H experiment that revealed the close proximity between phenyl phosphonic acid and aminopropyl surface groups.

In this contribution, we present a solid-state NMR approach to characterize drug-model molecules as ibuprofen, benzoic acid and lauric acid, encapsulated in MCM-41 silica and submitted to strong confinement effects. In particular, we show that by a careful choice of the solid-state NMR sequences, it is possible to efficiently characterize these highly mobile molecules and their interactions with the pore surface. Thus, we demonstrate that 13C NMR spectroscopy is a powerful tool to characterize and even quantify entrapped and non-entrapped species by using either single-pulse excitation (SPE) or cross-polarization (CP). Whereas the standard {1H}-13C CP experiment is of poor efficiency for mobile species, we show that 13C signal-to-noise (S/N) ratio can be significantly improved through 1H-13C cross-relaxation (namely, nuclear Overhauser effect, nOe) by using a 1H power-gated technique. The long transversal relaxation times [T2(1H) up to 22 ms] observed allow the set-up of J-coupling based experiments such as 2D {1H}-13C heteronuclear multiple-quantum coherence (HMQC) in order to fully characterize the encapsulated molecules. Thus, we demonstrate that the use of sequences derived from solution-state NMR such as these two latter experiments is highly efficient to characterize highly mobile organic molecules trapped in mesopores. Finally, we show that 1H spin diffusion-based experiments can give useful informations on the proximities between trapped molecules and the silica surface.

Hybrid "organic-inorganic" membranes are synthesized by growing a functionalized mesoporous "sol-gel"-derived SiO2 network into a hydrophobic fluorinated polymer. Processing guidelines are proposed to optimize the porosity at the macro-, meso- and micro-scales by templating the surface of the membrane through a modification of the thermal processing at 30-80 degrees C in air. Such a hierarchical design exploits the benefits of the facile water retention of the breath figure to produce membranes with high proton conductivity (50 mS cm(-1) at 70 degrees C) under 100% relative humidity and develops the surface area and the interfacial area between the hydrophobic and hydrophilic domains.

A large array of advanced solid state NMR (nuclear magnetic resonance) techniques is presented in the frame of the structural characterization of sol-gel-derived materials. These techniques include the pertinent detection of (17)O chemical shifts, MAS (magic angle spinning) J spectroscopy in the solid state, high-resolution (1)H spectroscopy, heteronuclear and homonuclear D (dipolar)-derived multidimensional correlation experiments, and first-principles calculations of NMR parameters. This spectroscopic approach is suitable for the in-depth description of multicomponent sol-gel derivatives, crystalline and amorphous biocompatible silicophosphates, Al-O-P clusters, and templated porous materials. It offers unique perspectives for the description of the hybrid interfaces in terms of chemical and spatial connectivities.

Various calcium phosphates and hydroxyapatite (HAp) have been fully characterized by one- and two-dimensional solid-state nuclear magnetic resonance (NMR) experiments and first principles calculations of NMR parameters, such as chemical shift anisotropy (CSA) and electric field gradient tensors for all nuclei. Such compounds act as useful biocompatible materials. The projector augmented wave (PAW) and gauge including PAW methods allowed the complete assignment of spectra, including 1H magic-angle spinning (MAS) spectra for which ultimate resolution is not attained experimentally. 1H CSA tensors and orientation of the principal axes systems have been also discussed. 17O parameters have been calculated for a large variety of oxo-bridges and terminal oxygen atoms, including P–O–Si fragments characteristic for silicophosphate phases. The (δiso, CQ) sets of values allowed the clear distinction between the various oxygen atoms in a calculated 17O 3-quantum MAS experiment. Such an approach should be of great help for the description of interfaces in complex materials, in terms of structure and chemical composition.

Ibuprofen (an anti-inflammatory drug that is a crystalline solid at ambient temperature) has been encapsulated in MCM-41 silica matrices with different pore diameters (35 and 116 A). Its behavior has been investigated by magic angle spinning (MAS) 1H, 13C, and 29Si solid-state NMR spectroscopy at ambient and low temperature. This study reveals an original physical state of the drug in such materials. At ambient temperature, ibuprofen is not in a solid state (crystalline or amorphous) and is extremely mobile inside the pores, with higher mobility in the largest pores (116 A). The interaction between ibuprofen and the silica surface is weak, which favors fast drug release from this material in a simulated intestinal or gastric fluid. The quasi-liquid behavior of ibuprofen allows the use of NMR pulse sequences issued from solution-state NMR, such as the INEPT sequence, to characterize these solid-state samples. The solid-state MAS NMR study shows that the proton of the carboxylic acid group of ibuprofen is i...

A representative silicophosphate gel was synthesized, starting from orthophosphate groups and pyrophosphate species. At 136°C, a complex mixture of crystalline phases and amorphous components was obtained. A new panel of solid state NMR techniques was implemented, including dipolar based experiments (CP MAS), as well as J-derived techniques, in both homonuclear (31P INADEQUATE-MAS) and heteronuclear (31P/29Si HMQC-MAS) versions. These experiments are suitable for the fine characterization of P–O–P, P–O–Si, P–OH…linkages in silicophosphate gels and materials.

This paper illustrates the use of a 1H-29Si-1H double CP sequence to investigate different organic/SiO2 interfaces in templated porous silica. The advantage of this sequence is to selectively edit, in a one-dimensional experiment, the protons that are in close proximity with the Si surface sites. In order to maximize the polarization transfer efficiency, some experiments have been recorded at lower temperature (238 K). Examples will concern surfactant/silica interactions in mesostructured silicas and the behavior of benzoic acid and 4-methoxychlorophenol, a common pesticide, encapsulated in mesoporous silica.

It is shown that the phenylphosphinic acid C 6 H 5 PH(O)(OH) and the octavinylsilsesquioxane [SiO 1.5 (C 2 H 3 )] 8 act as good candidates for the set-up of the Hartmann–Hahn condition under MAS condition for 31 P→ 13 C and 29 Si→ 13 C CP transfer, respectively. The study of the polyoxometalate derivative [AsW 9 O 33 ( t BuSiO) 3 (VO)]( n Bu 4 N) 3 allowed us to clearly demonstrate the effect of { 51 V} decoupling during 29 Si acquisition for this type of compounds. These examples open new possibilities for the detailed study of inorganic compounds and hybrid materials, by triple resonance solid-state NMR experiments. To cite this article: C. Bonhomme et al., C. R. Chimie 9 (2006) .

J-derived (HMQC, INEPT) and D-derived (double and triple resonance) experiments were applied to the detailed characterization of crystalline and amorphous silicophosphate derivatives. 31P/29Si and 1H/31P/29Si CP MAS experiments were suitable for the description of complex silicophosphate gels, which can act as precursors for biocompatible materials. First principles calculations involving the GIPAW approach (first developed by Mauri and Pickard) were applied for the determination of CSA (29Si, 31P, 17O) and quadrupolar (17O) parameters. Excellent agreement between experimental and calculated data was obtained.

We report the results of the two-dimensional MAS-J-HMQC experiment providing scalar correlations between 29Si and 31P nuclei in solid state NMR, and we give the first evaluation of the 2JSi-O-P coupling constants (approximately 15 Hz) for a crystalline silicophosphate phase Si5O(PO4)6. The experiment is applied to the characterization of complex mixtures of SiP2O7 phases, through editing of 31P spin pairs by the heteronuclear 2JP-O-Si interaction.

1H, 13C, 17O and 31P NMR parameters, including chemical shift tensors and quadrupolar parameters for 17O, were calculated for phenylphosphonic acid, C6H5PO(OH)2, under periodic boundary conditions. The results are in very good agreement with experimental data and permit the unambiguous assignment of all the sites present in the structure. In particular, the 17O NMR parameters of the PO and P - OH environments were precisely determined, which should help in the characterization of the bonding mode of phosphonate molecules in hybrid solids. Moreover, the effect of intermolecular interactions on the NMR parameters were investigated by comparing the results of the calculations in the crystal and in an isolated molecule of phenylphosphonic acid.

The synthesis of vanadium oxide nanotubes containing phosphorus (VPOx-NT) is described. A mixture of V2O5.nH2O gels, phosphoric acid and hexadecylamine was heated under hydrothermal conditions at 180°C for 7 days. The VPOx nanotubes were characterized by X-ray diffraction, scanning and transmission electron microscopy, 51V and 31P solid state NMR. In-situ experiments were performed in order to get a better understanding of the mechanism of these nanotubes formation.

We present the structure and a multinuclear solid-state NMR study of a new cyclic aluminophosphinate. The crystallographic structure of [Al(2)(HC(6)H(5)PO(2))(2)(C(4)H(9)OH)(8)]Cl(4) (compound 1) was obtained at low temperature (a = 11.830(7) A, b = 14.216(6) A, c = 17.790(6) A, beta = 91.25(4) degrees, monoclinic, P21/c, Z = 2). (13)C IRCP (inversion recovery cross polarization) and NQS (non quaternary suppression) NMR experiments allowed the complete assignment of the quaternary carbon atom of the phenyl ring and the precise determination of the isotropic /(1)J(P-C)/ coupling constant. (31)P CP MAS dynamics was carefully studied by varying the contact time. Dipolar oscillations even at slow MAS were observed. Up to 11 kHz, these oscillations were more pronounced, and the P-H distance was easily extracted. (27)Al NMR quadrupolar parameters for 1 were obtained with very good accuracy, and unusual satellite transition splitting was observed. Furthermore, the isotropic lines of the inner and outer transitions were clearly observable, leading to the unambiguous determination of the quadrupolar parameters.

New Al-O-P clusters are presented. They are characterized by single-crystal X-ray diffraction (XRD) and solid-state NMR. High-resolution techniques such as magic angle spinning (MAS) and multiple quantum magic angle spinning (MQ-MAS) are used. The latest developments of high-resolution 1H solid-state NMR, including very high field and very high speed MAS as well as homonuclear off-resonance Lee-Goldburg (LG) decoupling, are presented. All the experiments are described in the frame of a new pictorial representation of interactions involving quadrics and 4th degree cartesian surfaces.

The synthesis and characterisation (by single crystal X-ray diffraction and solid state 13C, 27Al, 31P NMR) of a new Al–O–P cluster related to the 41 Secondary Building Unit (SBU) of zeolites are presented.

The mother-of-pearl found in the shell of certain mollusks is a fascinating material with remarkable physical (mechanical and optical) properties. This biological material of organic and mineral composite nature presents a complex architecture whose structure and mechanisms of formation are not yet perfectly understood. In this work, taking as a model study the European abalone Haliotis tuberculata, we sought to contribute to the knowledge around this material by focusing our study on three aspects in particular: i) the ultrastructural study of the phase mineral and more particularly disordered mineral environments, ii) the study of the organo-minerale interface in the adult nacre in formation as well as in the larval shell and iii) the study of the first biominerals deposited in the larval shell at very young stages of development. To carry out this study, we used a coupling of advanced spectroscopic techniques (Solid-State Nuclear Magnetic Resonance, Scanning transmission X-ray microscopy and Electron energy loss spectroscopy) coupled to advanced electron microscopy techniques (High-Resolution Transmission Electronic Microscopies and Field Emission Gun Scanning Electronic Microscopy) and preparation of sample (Focused Ion beam) and molecular biology (immunohistochemistry, Correlative Light and Electronic Microscopy).; La nacre présente dans la coquille de certains mollusques est un matériau fascinant aux propriétés physiques (mécaniques et optiques) remarquable. Ce matériau biologique de nature composite organique et minérale présente un architecture complexe dont la structure et les mécanismes de formation ne sont pas encore parfaitement compris. Dans ces travaux, en prenant comme modèle d’étude comme modèle l’ormeau Européen Haliotis tuberculata, nous avons cherché à contribuer au savoir autour de ce matériau en axant notre étude sur trois aspects en particulier : i) l’étude ultrastructurale de la phase minérale et plus particulièrement des environnements minéraux désordonnés, ii) l’étude de l’interface organominérale dans la nacre adulte en formation ainsi que dans la coquille larvaire et iii) l’étude des premiers biominéraux déposés dans la coquille larvaire aux très jeune stades de développement. Pour réaliser cette étude nous avons eu recours à un couplage de techniques avancées spectroscopiques (Résonnance Magnétique Nucléaire à l’Etat Solide, Microscopie en transmission à balayage de rayons X et Spectroscopie des pertes d'énergie) couplé à des techniques de pointes de microscopie électronique (Microscopie électronique à transmission à haute résolution et Microscope électronique à balayage à effet de champ) et de préparation d’échantillon (Abrasion ionique focalisée) et de biologie moléculaire (Immunohistochimie, Microscopie optique et électronique corrélative).

The mother-of-pearl found in the shell of certain mollusks is a fascinating material with remarkable physical (mechanical and optical) properties. This biological material of organic and mineral composite nature presents a complex architecture whose structure and mechanisms of formation are not yet perfectly understood. In this work, taking as a model study the European abalone Haliotis tuberculata, we sought to contribute to the knowledge around this material by focusing our study on three aspects in particular: i) the ultrastructural study of the phase mineral and more particularly disordered mineral environments, ii) the study of the organo-minerale interface in the adult nacre in formation as well as in the larval shell and iii) the study of the first biominerals deposited in the larval shell at very young stages of development. To carry out this study, we used a coupling of advanced spectroscopic techniques (Solid-State Nuclear Magnetic Resonance, Scanning transmission X-ray microscopy and Electron energy loss spectroscopy) coupled to advanced electron microscopy techniques (High-Resolution Transmission Electronic Microscopies and Field Emission Gun Scanning Electronic Microscopy) and preparation of sample (Focused Ion beam) and molecular biology (immunohistochemistry, Correlative Light and Electronic Microscopy).; La nacre présente dans la coquille de certains mollusques est un matériau fascinant aux propriétés physiques (mécaniques et optiques) remarquable. Ce matériau biologique de nature composite organique et minérale présente un architecture complexe dont la structure et les mécanismes de formation ne sont pas encore parfaitement compris. Dans ces travaux, en prenant comme modèle d’étude comme modèle l’ormeau Européen Haliotis tuberculata, nous avons cherché à contribuer au savoir autour de ce matériau en axant notre étude sur trois aspects en particulier : i) l’étude ultrastructurale de la phase minérale et plus particulièrement des environnements minéraux désordonnés, ii) l’étude de l’interface organominérale dans la nacre adulte en formation ainsi que dans la coquille larvaire et iii) l’étude des premiers biominéraux déposés dans la coquille larvaire aux très jeune stades de développement. Pour réaliser cette étude nous avons eu recours à un couplage de techniques avancées spectroscopiques (Résonnance Magnétique Nucléaire à l’Etat Solide, Microscopie en transmission à balayage de rayons X et Spectroscopie des pertes d'énergie) couplé à des techniques de pointes de microscopie électronique (Microscopie électronique à transmission à haute résolution et Microscope électronique à balayage à effet de champ) et de préparation d’échantillon (Abrasion ionique focalisée) et de biologie moléculaire (Immunohistochimie, Microscopie optique et électronique corrélative).

This work aims to understand: How is reached the plywood architecture in bone? How is bone apatite formed and what is the effect of the environment on apatite formation? Thus, histological bone thin sections were investigated focusing on the interface between the osteoid and mature bone tissues. Our results show that this interface is acidic and collagenic but not in the form of fibrils. Thus, we propose a new mechanism for bone formation in bone remodelling where osteoclasts dissolution and new fibrils formation from osteoblasts lead to the formation of an acidic collagen mesophase in which apatite is then formed. In such mechanism, apatite forms from an acidic solution in interaction with an organic matrix (collagen molecules, citrate and non-collagenous proteins). Acidic biomimetic models have been set and the apatite formation has been followed in vitro using in situ Raman and ssNMR spectroscopies. Without organic molecules, biomimetic apatite is formed through the precipitation of an amorphous phase (ACP) that transforms into OCP which then turns into apatite. With pAsp, the same scenario is observed but the nucleation is delayed and residual OCP is stabilized. With a high concentration of citrate, only ACP is observed whereas with a lower concentration formation of OCP is inhibited. The same sequence is also observed with collagen but the final product is a more disorganized apatite. Apatite formation in dense and organized collagen solutions leads to the formation of a liquid ionic solution stable at basic pH during the first 72 hours explaining the resulting apatite/collagen co-alignement.; Les travaux réalisés ont pour but de répondre à deux questions : comment le collagène s'organise-t-il dans l'os mature ? Comment la phase minérale de l'os se forme-t-elle et quel est l'effet de son environnement sur sa formation ? Nous avons étudié d'un point de vue structural des coupes d'os en nous concentrant sur l'interface tissu ostéoïde/os mature, impliquant le front de minéralisation. Nous avons conclu qu'un domaine acide fait de collagène non fibrillaire existe à cette interface. Nous proposons ainsi un nouveau mécanisme pour la formation osseuse lors du remodelage osseux impliquant une mésophase acide de collagène au sein de laquelle l'apatite se forme. Nous avons donc étudié l'effet de cet environnement acide sur la formation d'apatite in vitro par Raman in situ et RMN du solide. Nous avons observé que la séquence de précipitation de l'apatite seule en solution passe par la formation d'une phase amorphe (ACP) qui se transforme en OCP puis en apatite. En présence de pAsp, la nucléation est ralentie et de l'OCP est stabilisé. Une grande concentration en citrate inhibe la formation de toute autre phase que l'ACP tandis qu'une plus faible concentration entraine la formation d'apatite directement depuis l'ACP. Cette séquence ACP/apatite est également observée lorsque la minéralisation est réalisée en présence de collagène quelle que soit sa concentration. Le collagène entraine la formation d'une apatite beaucoup plus désorganisée de cristallinité proche de celle de l'apatite osseuse. Enfin, une concentration en collagène supérieure à 80 mg/mL mène à la stabilisation d'une phase ionique stable à pH basique expliquant le co-alignement apatite/collagène.

The work of this thesis is focused on the characterization of bone apatite crystals, mainly by solid-state nuclear magnetic resonance. The originality of this work was to study a fresh bone sample, analyzed within two hours after its extraction from the animal (i.e. a two years old sheep). This approach avoids any alteration of the bone sample, and keeps its natural state of hydration. Such experimental rigor led to the evidence that the apatite crystals of bone own a high hydrophilic character. Furthermore, a analogous hydrophilic character has also been reported for two samples of biomimetic apatites, both studied in a artificial wet state. It is shown here that this hydrophilicity is given by the presence of a particular mineral domain located at the surface of the bone apatite (and biomimetic apatite) crystals, the so called non-apatitic domain. The NMR signatures (1H and 31P nuclei) and the hydrophilic properties of the non-apatitic domain were compared with those of a synthetic amorphous calcium phosphate, and all were found to be similar. The chemical composition of the non-apatitic domain was also studied, sometimes performed on a bone sample and sometimes on labeled (13C and 43C) biomimetic apatites. It has been shown that this non-apatitic domain is mainly composed of divalent species : Ca2+ , CO32- and HPO42-. Finally, some X-ray diffraction and cryo-electron transmission microscopy experiments were done to study the behavior of apatite crystals in aqueous media. It has been shown, surprisingly, that the water molecules strongly adsorbed on the non-apatitic domain at the surface of apatite crystals, can promote the adhesion between crystals and their 3D organization.; Les travaux de cette thèse portent sur la caractérisation fine des cristaux d’apatite osseuse, principalement par résonance magnétique nucléaire à l’état solide. Leur originalité majeure a été de se tourner vers l’étude d’un échantillon d’os frais et intact, analysé dans les deux heures après son extraction de chez l’animal (i.e. une brebis âgée de deux ans). Cette démarche évite toute altération de l’échantillon d’os, lequel se trouve proche de son état d’hydratation naturel. Une telle rigueur expérimentale a permis de mettre en lumière le fort caractère hydrophile des cristaux d’apatite du minéral osseux. Un même caractère hydrophile a également été rapporté pour des analogues biomimétiques de synthèse étudiés dans un état humide artificiel. Il est prouvé ici que ce caractère hydrophile est fourni par la présence d’un domaine minéral particulier se trouvant en surface des cristaux d’apatite osseuse et biomimétique, appelé domaine non-apatitique. Les signatures RMN (des noyaux 31P et 1H) ainsi que les propriétés d’hydrophilie de ce domaine non-apatitique ont été comparées à celles d’un échantillon de phosphate de calcium amorphe de synthèse, et se sont avérées similaires. Une étude sur la composition chimique de ce domaine non-apatitique a également été entreprise, réalisée parfois sur un échantillon d’os intact et parfois sur des analogues biomimétiques de synthèse enrichis en certains isotope (i.e. 13C et 43Ca). Il a de cette manière été montré que ce domaine non-apatitique est principalement composé d’espèces divalentes : Ca2+, HPO42- et CO32-. Enfin, des expériences de diffraction des rayons X et de cryo-microscopie électronique en transmission ont permis d’étudier le comportement en solution des cristaux d’apatite. Il a de cette manière été prouvé, de manière inattendue, que les molécules d’eau rigidement adsorbées sur le domaine de surface des cristaux d’apatite peuvent favoriser l’adhésion entre les cristaux ainsi que leur organisation 3D.