Search

In this contribution, we apply our newly developed ball-milling operando platform, which combines Raman spectroscopy and thermal (IR) imaging, as well as acoustic and high-speed optical video recordings, to the synthesis and transformation of citric acid-isonicotinamide (1:2) cocrystal polymorphs in transparent PMMA jars. Particularly, we demonstrate how Raman, temperature, acoustic, and video data are complementary and enable detection and connection of chemical and physical events happening during ball-milling in a time-resolved manner. Importantly, we show that the formation of the three cocrystal polymorphs can be detected through acoustic analyses solely. Even more impressively, the meticulous analyses of the sound data allowed subtle polymorphic transitions to be perceived in operando mode when the Raman spectroscopy was not conclusive enough. Such changes in sound signatures are greatly linked to the beads' motions in the milling jar. A new data analysis methodology of acoustic recordings is proposed through a combination of energetic and statistical approaches that simplifies the data analyses for potential users. The interpretation of the detected sound signals was further validated thanks to the high-speed videos recorded in synchronization with all other operando techniques. Finally, we broaden this acoustic methodology to opaque stainless-steel jars, showing the relevance of the acoustic analysis method for following polymorphic transformations of cocrystals, as well as pure substances, in any type of milling jar.

Octacalcium phosphate (OCP, Ca 8 (PO 4 ) 4 (HPO 4 ) 2 ·5H 2 O) is a notable calcium phosphate due to its biocompatibility, making it a widely studied material for bone substitution. It is known to be a precursor of bone mineral, but its role in biomineralisation remains unclear. While the structure of OCP has been the subject of thorough investigations (including using Rietveld refinements of X-ray diffraction data, and NMR crystallography studies), important questions regarding the symmetry and H-bonding network in the material remain. In this study, it is shown that OCP undergoes a lowering of symmetry below 200 K, evidenced by 1 H, 17 O, 31 P and 43 Ca solid-state NMR experiments. Using ab initio molecular-dynamics (MD) simulations and gauge including projected augmented wave (GIPAW) DFT calculations of NMR parameters, the presence of rapid motions of the water molecules in the crystal cell at room temperature is proved. This information leads to an improved description of the OCP structure at both low and ambient temperatures, and helps explain long-standing issues of symmetry. Remaining challenges related to the understanding of the structure of OCP are then discussed.

Oxalate ligands are found in many classes of materials, including energy storage materials and biominerals. Determining their local environments at the atomic scale is thus paramount to establishing the structure and properties of numerous phases. Here, we show that high-resolution 17 O solid-state NMR is a valuable asset for investigating the structure of crystalline oxalate systems. First, an efficient 17 O-enrichment procedure of oxalate ligands is demonstrated using mechanochemistry. Then, 17 O-enriched oxalates were used for the synthesis of the biologically relevant calcium oxalate monohydrate (COM) phase, enabling the analysis of its structure and heat-induced phase transitions by high-resolution 17 O NMR. Studies of the low-temperature COM form (LT-COM), using magnetic fields from 9.4 to 35.2 T, as well as 13 C- 17 O MQ/D-RINEPT and 17 O{ 1 H} MQ/REDOR experiments, enabled the 8 inequivalent oxygen sites of the oxalates to be resolved, and tentatively assigned. The structural changes upon heat treatment of COM were also followed by high-resolution 17 O NMR, providing new insight into the structures of the high-temperature form (HT-COM) and anhydrous calcium oxalate α-phase (α-COA), including the presence of structural disorder in the latter case. Overall, this work highlights the ease associated with 17 O-enrichment of oxalate oxygens, and how it enables high-resolution solid-state NMR, for "NMR crystallography" investigations.

The Zr-based Metal Organic Framework (MOF) UiO-66(Zr) is widely employed owing to its good thermal and chemical stabilities. Although the long-range structure of this MOF is preserved in the presence of water during several days, little is known about the formation of defects, which cannot be detected using diffraction techniques. We apply here 17 O solid-state NMR spectroscopy at 18.8 T to investigate the reactivity of UiO-66, through the exchange of oxygen atoms between the different sites of the MOF and water. For that purpose, we have selectively enriched in 17 O isotope the carboxylate groups of UiO-66(Zr) by using it with 17 O-labeled terephthalic acid prepared using mechanochemistry. In the presence of water at 50 °C and a following dehydration at 150 °C, we observe an overall exchange of O atoms between COO - and μ 3 -O 2- sites. Furthermore, we demonstrate that the three distinct oxygen sites, μ 3 -OH, μ 3 -O 2- and COO - , of UiO-66(Zr) MOF can be enriched in 17 O isotope by post-synthetic hydrothermal treatment in the presence of 17 O-enriched water. These results demonstrate the lability of Zr-O bonds and the reactivity of UiO-66(Zr) with water., (© 2024 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

While ball-milling is becoming one of the common tools used by synthetic chemists, an increasing number of studies highlight that it is possible to further expand the nature and number of products which can be synthesized, by heating the reaction media during mechanochemical reactions. Hence, developing set-ups enabling heating and milling to be combined is an important target, which has been looked into in both academic and industrial laboratories. Here, we report a new approach for heating up reaction media during ball-milling reactions, using induction heating (referred to as i-BM). Our set-up is attractive not only because it enables a very fast heating of the milling medium (reaching ≈80 °C in just 15 s), and that it is directly adaptable to commercially-available milling equipment, but also because it enables heating either the walls of the milling jars or the beads themselves, depending on the choice of the materials which compose them. Importantly, the possibility to heat a milling medium "from the inside" (when using for example a PMMA jar and stainless steel beads) is a unique feature compared to previously proposed systems. Through numerical simulations, we then show that it is possible to finely tune the properties of this heating system ( e.g. heating rate and maximum temperature reached), by playing with the characteristics of the milling system and/or the induction heating conditions used. Lastly, examples of applications of i-BM are given, showing how it can be used to help elucidate reaction mechanisms in ball-milling, to synthesize new molecules, and to control the physical nature of milling media.

A simple and versatile preparation of Zn(II)-poly(carboxylates) reticulated binders by the addition of Zn(II) precursors (ZnSO 4 , ZnO, or Zn(NO 3 ) 2 ) into a preoptimized poly(carboxylic acids) binder solution is proposed. These binders lead systematically to a significantly improved electrochemical performance when used for the formulation of silicon-based negative electrodes. The formation of carboxylate-Zn(II) coordination bonds formation is investigated by rheology and FTIR and NMR spectroscopies. Mechanical characterizations reveal that the coordinated binder offers a better electrode coating cohesion and adhesion to the current collector, as well as higher hardness and elastic modulus, which are even preserved in the presence of a carbonate solvent (i.e., in battery operation conditions). Ultimately, as shown from operando dilatometry experiments, the electrode expansion during lithiation is reduced, mitigating electrode mechanical failure. Such coordinatively reticulated electrodes outperform their uncoordinated counterparts with an improved capacity retention of over 30% after 60 cycles.

17 O NMR spectroscopy is a powerful technique, which can provide unique information regarding the structure and reactivity of biomolecules. However, the low natural abundance of 17 O (0.04 %) generally requires working with enriched samples, which are not easily accessible. Here, we present simple, fast and cost-efficient 17 O-enrichment strategies for amino acids and peptides by using mechanochemistry. First, five unprotected amino acids were enriched under ambient conditions, consuming only microliter amounts of costly labeled water, and producing pure molecules with enrichment levels up to ∼40 %, yields ∼60-85 %, and no loss of optical purity. Subsequently, 17 O-enriched Fmoc/tBu-protected amino acids were produced on a 1 g/day scale with high enrichment levels. Lastly, a site-selective 17 O-labeling of carboxylic functions in peptide side-chains was achieved for RGD and GRGDS peptides, with ∼28 % enrichment level. For all molecules, 17 O ssNMR spectra were recorded at 14.1 T in reasonable times, making this an important step forward for future NMR studies of biomolecules., (© 2022 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

The possibility of enriching in 17 O the water molecules within hydrated biominerals belonging to the Ca-pyrophosphate family was investigated, using liquid assisted grinding (LAG) in the presence of 17 O-labelled water. Two phases with different hydration levels, namely triclinic calcium pyrophosphate dihydrate (Ca 2 P 2 O 7 ·2H 2 O, denoted t -CPPD) and monoclinic calcium pyrophosphate tetrahydrate (Ca 2 P 2 O 7 ·4H 2 O, denoted m -CPPT β) were enriched in 17 O using a "post-enrichment" strategy, in which the non-labelled precursors were ground under gentle milling conditions in the presence of stoichiometric quantities of 17 O-enriched water (introduced here in very small volumes ∼10 μL). Using high-resolution 17 O solid-state NMR (ssNMR) analyses at multiple magnetic fields, and dynamic nuclear polarisation (DNP)-enhanced 17 O NMR, it was possible to show that the labelled water molecules are mainly located at the core of the crystal structures, but that they can enter the lattice in different ways, namely by dissolution/recrystallisation or by diffusion. Overall, this work sheds light on the importance of high-resolution 17 O NMR to help decipher the different roles that water can play as a liquid-assisted grinding agent and as a reagent for 17 O-isotopic enrichment.

Calcium oxalate minerals of the general formula CaC 2 O 4 . x H 2 O are widely present in nature and usually associated with pathological calcifications, constituting up to 70-80% of the mineral component of renal calculi. The monohydrate phase (CaC 2 O 4 . H 2 O, COM) is the most stable form, accounting for the majority of the hydrated calcium oxalates found. These mineral phases have been studied extensively via X-ray diffraction and IR spectroscopy and, to a lesser extent, using 1 H, 13 C, and 43 Ca solid-state NMR spectroscopy. However, several aspects of their structure and reactivity are still unclear, such as the evolution from low- to high-temperature COM structures (LT-COM and HT-COM, respectively) and the involvement of water molecules in this phase transition. Here, we report for the first time a 17 O and 2 H solid-state NMR investigation of the local structure and dynamics of water in the COM phase. A new procedure for the selective 17 O- and 2 H-isotopic enrichment of water molecules within the COM mineral is presented using mechanochemistry, which employs only microliter quantities of enriched water and leads to exchange yields up to ∼30%. 17 O NMR allows both crystallographically inequivalent water molecules in the LT-COM structure to be resolved, while 2 H NMR studies provide unambiguous evidence that these water molecules are undergoing different types of motions at high temperatures without exchanging with one another. Dynamics appear to be essential for water molecules in these structures, which have not been accounted for in previous structural studies on the HT-COM structure due to lack of available tools, highlighting the importance of such NMR investigations for refining the overall knowledge on biologically relevant minerals like calcium oxalates., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

We present a new operando approach for following reactions taking place in mechanochemistry, relying on the analysis of the evolution of the sound during milling. We show that differences in sound can be directly correlated to (physico)chemical changes in the reactor, making this technique highly attractive and complementary to others for monitoring mechanochemical reactions. Most notably, it can provide unique information on the actual movements of the beads within the milling jars, which opens new avenues for helping rationalize mechanochemical processes., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

The description of the formation, structure, and reactivity of coordination networks and metal-organic frameworks (MOFs) remains a real challenge in a number of cases. This is notably true for compounds composed of Zn 2+ ions and terephthalate ligands (benzene-1,4-dicarboxylate, BDC) because of the difficulties in isolating them as pure phases and/or because of the presence of structural defects. Here, using mechanochemistry in combination with operando Raman spectroscopy, the observation of the formation of various zinc terephthalate compounds was rendered possible, allowing the distinction and isolation of three intermediates during the ball-milling synthesis of Zn 3 (OH) 4 (BDC). An "NMR crystallography" approach was then used, combining solid-state NMR ( 1 H, 13 C, and 17 O) and density functional theory (DFT) calculations to refine the poorly described crystallographic structures of these phases. Particularly noteworthy are the high-resolution 17 O NMR analyses, which were made possible in a highly efficient and cost-effective way, thanks to the selective 17 O-enrichment of either hydroxyl or terephthalate groups by ball-milling. This allowed the presence of defect sites to be identified for the first time in one of the phases, and the nature of the H-bonding network of the hydroxyls to be established in another. Lastly, the possibility of using deuterated precursors ( e.g ., D 2 O and d 4 -BDC) during ball-milling is also introduced as a means for observing specific transformations during operando Raman spectroscopy studies, which would not have been possible with hydrogenated equivalents. Overall, the synthetic and spectroscopic approaches developed herein are expected to push forward the understanding of the structure and reactivity of other complex coordination networks and MOFs., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

The reaction between P2-type honeycomb layered oxides Na 2 Ni 2 TeO 6 and K 2 Ni 2 TeO 6 enables the formation of NaKNi 2 TeO 6 . The compound is characterized by X-ray diffraction and 23 Na solid-state nuclear magnetic resonance spectroscopy, and the structure is discussed through density functional theory calculations. In addition to the honeycomb Ni/Te cationic ordering, NaKNi 2 TeO 6 exhibits a unique example of alternation of sodium and potassium layers instead of a random alkali-mixed occupancy. Stacking fault simulations underline the impact of the successive position of the Ni/Te honeycomb layers and validate the presence of multiple stacking sequences within the powder material, in proportions that evolve with the synthesis conditions. In a broader context, this work contributes to a better understanding of the alkali-mixed layered compounds.

There is a growing interest in magnetic nanocomposites in biomaterials science. In particular, nanocomposites that combine poly(lactide) (PLA) nanofibers and superparamagnetic iron oxide nanoparticles (SPIONs), which can be obtained by either electrospinning of a SPION suspension in PLA or by precipitating SPIONs at the surface of PLA, are well documented in the literature. However, these two classical processes yield nanocomposites with altered materials properties, and their long-term in vivo fate and performances have in most cases only been evaluated over short periods of time. Recently, we reported a new strategy to prepare well-defined PLA@SPION nanofibers with a quasi-monolayer of SPIONs anchored at the surface of PLA electrospun fibers. Herein, we report on a 6-month in vivo rat implantation study with the aim of evaluating the long-term magnetic resonance imaging (MRI) properties of this new class of magnetic nanocomposites, as well as their tissue integration and degradation. Using clinically relevant T 2-weighted MRI conditions, we show that the PLA@SPION nanocomposites are clearly visible up to 6 months. We also evaluate here by histological analyses the slow degradation of the PLA@SPIONs, as well as their biocompatibility. Overall, these results make these nanocomposites attractive for the development of magnetic biomaterials for biomedical applications.

Oxygen-17 and deuterium are two quadrupolar nuclei that are of interest for studying the structure and dynamics of materials by solid-state nuclear magnetic resonance (NMR). Here, 17 O and 2 H NMR analyses of crystalline ibuprofen and terephthalic acid are reported. First, improved 17 O-labelling protocols of these molecules are described using mechanochemistry. Then, dynamics occurring around the carboxylic groups of ibuprofen are studied considering variable temperature 17 O and 2 H NMR data, as well as computational modelling (including molecular dynamics simulations). More specifically, motions related to the concerted double proton jump and the 180° flip of the H-bonded (-COOH) 2 unit in the crystal structure were looked into, and it was found that the merging of the C=O and C-OH 17 O resonances at high temperatures cannot be explained by the sole presence of one of these motions. Lastly, preliminary experiments were performed with a 2 H- 17 O diplexer connected to the probe. Such configurations can allow, among others, 2 H and 17 O NMR spectra to be recorded at different temperatures without needing to tune or to change probe configurations. Overall, this work offers a few leads which could be of use in future studies of other materials using 17 O and 2 H NMR., (© 2021 The Authors. Magnetic Resonance in Chemistry published by John Wiley & Sons Ltd.)

In recent years, there has been increasing interest in developing cost-efficient, fast, and user-friendly 17 O enrichment protocols to help to understand the structure and reactivity of materials by using 17 O NMR spectroscopy. Here, we show for the first time how ball milling (BM) can be used to selectively and efficiently enrich the surface of fumed silica, which is widely used at industrial scale. Short milling times (up to 15 min) allowed modulation of the enrichment level (up to ca. 5 %) without significantly changing the nature of the material. High-precision 17 O compositions were measured at different milling times by using large-geometry secondary-ion mass spectrometry (LG-SIMS). High-resolution 17 O NMR analyses (including at 35.2 T) allowed clear identification of the signals from siloxane (Si-O-Si) and silanols (Si-OH), while DNP analyses, performed by using direct 17 O polarization and indirect 17 O{ 1 H} CP excitation, agreed with selective labeling of the surface. Information on the distribution of Si-OH environments at the surface was obtained from 2D 1 H- 17 O D-HMQC correlations. Finally, the surface-labeled silica was reacted with titania and using 17 O DNP, their common interface was probed and Si-O-Ti bonds identified., (© 2021 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

43 Ca nuclear magnetic resonance (NMR) spectroscopy has been extensively applied to the detailed study of octacalcium phosphate (OCP), Ca 8 (HPO 4 ) 2 (PO 4 ) 4 .5H 2 O, and hybrid derivatives involving intercalated metabolic acids (viz., citrate, succinate, formate, and adipate). Such phases are of importance in the development of a better understanding of bone structure. High-resolution 43 Ca magic angle spinning (MAS) experiments, including double-rotation (DOR) 43 Ca NMR, as well as 43 Ca{ 1 H} rotational echo DOR (REDOR) and 31 P{ 43 Ca} REAPDOR NMR spectra, were recorded on a 43 Ca-labeled OCP phase at very high magnetic field (20 T), and complemented by ab initio calculations of NMR parameters using the Gauge-Including Projector Augmented Wave-density functional theory (GIPAW-DFT) method. This enabled a partial assignment of the eight inequivalent Ca 2+ sites of OCP. Natural-abundance 43 Ca MAS NMR spectra were then recorded for the hybrid organic-inorganic derivatives, revealing changes in the 43 Ca lineshape. In the case of the citrate derivative, these could be interpreted on the basis of computational models of the structure. Overall, this study highlights the advantages of combining high-resolution 43 Ca NMR experiments and computational modeling for studying complex hybrid biomaterials., (© 2021 John Wiley & Sons, Ltd.)

The spectroscopic study of pathological calcifications (including kidney stones) is extremely rich and helps to improve the understanding of the physical and chemical processes associated with their formation. While Fourier transform infrared (FTIR) imaging and optical/electron microscopies are routine techniques in hospitals, there has been a dearth of solid-state NMR studies introduced into this area of medical research, probably due to the scarcity of this analytical technique in hospital facilities. This work introduces effective multinuclear and multidimensional solid-state NMR methodologies to study the complex chemical and structural properties characterizing kidney stone composition. As a basis for comparison, three hydrates ( n = 1 , 2 and 3) of calcium oxalate are examined along with nine representative kidney stones. The multinuclear magic angle spinning (MAS) NMR approach adopted investigates the 1 H , 13 C , 31 P and 31 P nuclei, with the 1 H and 13 C MAS NMR data able to be readily deconvoluted into the constituent elements associated with the different oxalates and organics present. For the first time, the full interpretation of highly resolved 1 H NMR spectra is presented for the three hydrates, based on the structure and local dynamics. The corresponding 31 P MAS NMR data indicates the presence of low-level inorganic phosphate species; however, the complexity of these data make the precise identification of the phases difficult to assign. This work provides physicians, urologists and nephrologists with additional avenues of spectroscopic investigation to interrogate this complex medical dilemma that requires real, multitechnique approaches to generate effective outcomes., Competing Interests: The authors declare that they have no conflict of interest., (Copyright: © 2021 César Leroy et al.)

Two mechanochemical procedures for 17O/18O-isotope labeling of fatty acids are reported: a carboxylic acid activation/hydrolysis approach and a saponification approach. The latter route allowed first-time enrichment of important polyunsaturated fatty acids (PUFAs) including docosahexaenoic acid (DHA). Overall, a total of 9 pure labeled products were isolated in high yields (≥80%) and with high enrichment levels (≥37% average labeling of C=O and C-OH carboxylic oxygen atoms), under mild conditions, and in short time (

Fatty acids are ubiquitous in biological systems and widely used in materials science, including for the formulation of drugs and the surface-functionalization of nanoparticles. However, important questions regarding the structure and reactivity of these molecules are still to be elucidated, including their mode of binding to certain metal cations or materials surfaces. In this context, we have developed novel, efficient, user-friendly, and cost-effective synthetic protocols based on ball-milling, for the 17 O and 18 O isotopic labeling of two key fatty acids which are widely used in (nano)materials science, namely stearic and oleic acid. Labeled molecules were analyzed by 1 H and 13 C solution NMR, IR spectroscopy, and mass spectrometry (ESI-TOF and LC-MS), as well as 17 O solid state NMR (for the 17 O labeled species). In both cases, the labeling procedures were scaled-up to produce up to gram quantities of 17 O- or 18 O-enriched molecules in just half-a-day, with very good synthetic yields (all ≥84%) and enrichment levels (up to an average of 46% per carboxylic oxygen). The 17 O-labeled oleic acid was then used for the synthesis of a metal soap (Zn-oleate) and the surface-functionalization of ZnO nanoparticles (NPs), which were characterized for the first time by high-resolution 17 O NMR (at 14.1 and 35.2 T). This allowed very detailed insight into (i) the coordination mode of the oleate ligand in Zn-oleate to be achieved (including information on Zn···O distances) and (ii) the mode of attachment of oleic-acid at the surface of ZnO (including novel information on its photoreactivity upon UV-irradiation). Overall, this work demonstrates the high interest of these fatty acid-enrichment protocols for understanding the structure and reactivity of a variety of functional (nano)materials systems using high resolution analyses like 17 O NMR.

While 17 O NMR is increasingly being used for elucidating the structure and reactivity of complex molecular and materials systems, much effort is still required for it to become a routine analytical technique. One of the main difficulties for its development comes from the very low natural abundance of 17 O (0.04%), which implies that isotopic labeling is generally needed prior to NMR analyses. However, 17 O-enrichment protocols are often unattractive in terms of cost, safety, and/or practicality, even for compounds as simple as metal oxides. Here, we demonstrate how mechanochemistry can be used in a highly efficient way for the direct 17 O isotopic labeling of a variety of s-, p-, and d-block oxides, which are of major interest for the preparation of functional ceramics and glasses: Li 2 O, CaO, Al 2 O 3 , SiO 2 , TiO 2 , and ZrO 2 . For each oxide, the enrichment step was performed under ambient conditions in less than 1 h and at low cost, which makes these synthetic approaches highly appealing in comparison to the existing literature. Using high-resolution solid-state 17 O NMR and dynamic nuclear polarization, atomic-level insight into the enrichment process is achieved, especially for titania and alumina. Indeed, it was possible to demonstrate that enriched oxygen sites are present not only at the surface but also within the oxide particles. Moreover, information on the actual reactions occurring during the milling step could be obtained by 17 O NMR, in terms of both their kinetics and the nature of the reactive species. Finally, it was demonstrated how high-resolution 17 O NMR can be used for studying the reactivity at the interfaces between different oxide particles during ball-milling, especially in cases when X-ray diffraction techniques are uninformative. More generally, such investigations will be useful not only for producing 17 O-enriched precursors efficiently but also for understanding better mechanisms of mechanochemical processes themselves.

Janus bis(N-heterocyclic carbenes) composed of a porphyrin core with two N-heterocyclic carbene (NHC) heads fused to opposite pyrroles were used as bridging ligands for the preparation of metal complexes. We first focused our attention on the synthesis of gold(i) chloride complexes [(NHC)AuCl] and investigated the substitution of the chloride ligand by acetylides to obtain the corresponding [(NHC)AuC[triple bond, length as m-dash]CR] complexes. Polyacetylides were then used to obtain molecular multiporphyrinic systems with porphyrins fused to only one NHC ligand, while main-chain organometallic polymers (MCOPs) were obtained when using Janus porphyrin bis(NHCs). Interestingly, MCOPs incorporating zinc(ii) porphyrins proved to be efficient as heterogeneous photocatalysts for the generation of singlet oxygen upon visible light irradiation.

Materials containing a calcium phosphate component have been the subject of much interest to NMR spectroscopists, especially in view of understanding the structure and properties of mineralized tissues like bone and teeth, and of developing synthetic biomaterials for bone regeneration. Here, we present a selection of recent developments in their structural characterization using advanced solid state NMR experiments, highlighting the level of insight which can now be accessed., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The development of amorphous phosphate-based materials is of major interest in the field of biomaterials science, and especially for bone substitution applications. In this context, we herein report the synthesis of gel-derived hydrated amorphous calcium/sodium ortho/pyrophosphate materials at ambient temperature and in water. For the first time, such materials have been obtained in a large range of tunable orthophosphate/pyrophosphate molar ratios. Multi-scale characterization was carried out thanks to various techniques, including advanced multinuclear solid state NMR. It allowed the quantification of each ionic/molecular species leading to a general formula for these materials: [(Ca 2+ y Na + z H + 3+ x -2y-z )(PO 4 3- ) 1-x (P 2 O 7 4- ) x ](H 2 O) u . Beyond this formula, the analyses suggest that these amorphous solids are formed by the aggregation of colloids and that surface water and sodium could play a role in the cohesion of the whole material. Although the full comprehension of mechanisms of formation and structure is still to be investigated in detail, the straightforward synthesis of these new amorphous materials opens up many perspectives in the field of materials for bone substitution and regeneration. STATEMENT OF SIGNIFICANCE: The metastability of amorphous phosphate-based materials with various chain length often improves their (bio)chemical reactivity. However, the control of the ratio of the different phosphate entities has not been yet described especially for small ions (pyrophosphate/orthophosphate) and using soft chemistry, whereas it opens the way for the tuning of enzyme- and/or pH-driven degradation and biological properties. Our study focuses on elaboration of amorphous gel-derived hydrated calcium/sodium ortho/pyrophosphate solids at 70 °C with a large range of orthophosphate/pyrophosphate ratios. Multi-scale characterization was carried out using various techniques such as advanced multinuclear SSNMR ( 31 P, 23 Na, 1 H, 43 Ca). Analyses suggest that these solids are formed by colloids aggregation and that the location of mobile water and sodium could play a role in the material cohesion., (Copyright © 2019 Acta Materialia Inc. All rights reserved.)

The synthesis, characterization, and biological evaluation of a series of compounds incorporating two or three benzoxaborole moieties is reported. Three different synthetic strategies were used to explore within this series as much chemical space as possible, all starting from the 6-aminobenzoxaborole reagent: amide coupling, imine bond formation, and squarate coupling. Eleven new compounds were isolated in pure form, and single crystals were obtained for two of them. These compounds were then evaluated as carbonic anhydrase inhibitors against the cytosolic hCA I and II and the transmembrane hCA IV, IX, and XII isoforms. While the benzoxaborole scaffold has been recently introduced as a new chemotype for carbonic anhydrase inhibition, these new multivalent derivatives exhibited superior inhibitory activity against the tumor-associated isoform hCA IX. In particular, compared to monovalent 6-aminobenzoxaborole ( K I = 813 nM) and 6-carboxybenzoxaborole ( K I = 400 nM), derivative 2h characterized by a glutamic acid structural core and two benzoxaborole moieties was found to be more potent ( K I = 64 nM) and more selective over human hCA II., Competing Interests: The authors declare no competing financial interest.

Composites combining superparamagnetic iron oxide nanoparticles (SPIONs) and polymers are largely present in modern (bio)materials. However, although SPIONs embedded in polymer matrices are classically reported, the mechanical and degradation properties of the polymer scaffold are impacted by the SPIONs. Therefore, the controlled anchoring of SPIONs onto polymer surfaces is still a major challenge. Herein, we propose an efficient strategy for the direct and uniform anchoring of SPIONs on the surface of functionalized-polylactide (PLA) nanofibers via a simple free ligand exchange procedure to design PLA@SPIONs core@shell nanocomposites. The resulting PLA@SPIONs hybrid biomaterials are characterized by electron microscopy (scanning electron microscopy and transmission electron microscopy) and energy-dispersive X-ray spectroscopy analysis to probe the morphology and detect elements present at the organic-inorganic interface, respectively. A monolayer of SPIONs with a complete and homogeneous coverage is observed on the surface of PLA nanofibers. Magnetization experiments show that magnetic properties of the nanoparticles are well preserved after their grafting on the PLA fibers and that the size of the nanoparticles does not change. The absence of cytotoxicity, combined with a high sensitivity of detection in magnetic resonance imaging both in vitro and in vivo, makes these hybrid nanocomposites attractive for the development of magnetic biomaterials for biomedical applications.

A new layered hybrid polythiophene-silica material was obtained directly by hydrolysis and polycondensation (sol-gel) of a silylated-thiophene bifunctional precursor, and its subsequent oxidative polymerization by FeCl₃. This precursor was judiciously designed to guarantee its self-assembly and the formation of a lamellar polymer-silica structure, exploiting the cooperative effect between the hydrogen bonding interactions, originating from the ureido groups and the π-stacking interactions between the thiophene units. The lamellar structure of the polythiophene-silica composite was confirmed by X-ray powder diffraction (XRD) and transmission electron microscopy (TEM) analyses. The solid-state nuclear magnetic resonance (NMR), UV-Vis, and photoluminescence spectra unambiguously indicate the incorporation of polythiophene into the silica matrix. Our work demonstrates that using a polymerizable silylated-thiophene precursor is an efficient approach towards the formation of nanostructured conjugated polymer-based hybrid materials.

Natural abundance 43Ca solid state NMR experiments are reported for the first time at ultra-high magnetic field (35.2 T) on a series of Ca-(pyro)phosphate and Ca-oxalate materials, which are of biological relevance in relation to biomineralization processes and the formation of pathological calcifications. The significant gain in both sensitivity and resolution at 35.2 T leads to unprecedented insight into the structure of both crystalline and amorphous phases.

17 O NMR spectroscopy has been the subject of vivid interest in recent years, because there is increasing evidence that it can provide unique insight into the structure and reactivity of many molecules and materials. However, due to the very poor natural abundance of oxygen-17, 17 O labeling is generally a prerequisite. This is a real obstacle for most research groups, because of the high costs and/or strong experimental constraints of the most frequently used 17 O-labeling schemes. Here, we show for the first time that mechanosynthesis offers unique opportunities for enriching in 17 O a variety of organic and inorganic precursors of synthetic interest. The protocols are fast, user-friendly, and low-cost, which makes them highly attractive for a broad research community, and their suitability for 17 O solid-state NMR applications is demonstrated., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

With the increasing number of organocalcium and organomagnesium complexes under development, there is a real need to be able to characterize in detail their local environment in order to fully rationalize their reactivity. For crystalline structures, in cases when diffraction techniques are insufficient, additional local spectroscopies like 25 Mg and 43 Ca solid-state NMR may provide valuable information to help fully establish the local environment of the metal ions. In this current work, a prospective DFT investigation on crystalline magnesium and calcium complexes involving low-coordination numbers and N-bearing organic ligands was carried out, in which the 25 Mg and 43 Ca NMR parameters [isotropic chemical shift, chemical shift anisotropy (CSA) and quadrupolar parameters] were calculated for each structure. The analysis of the calculated parameters in relation to the local environment of the metal ions revealed that they are highly sensitive to very small changes in geometry/distances, and hence that they could be used to assist in the refinement of crystal structures. Moreover, such calculations provide a guideline as to how the NMR measurements will need to be performed, revealing that these will be very challenging.

The interfaces within bones, teeth and other hybrid biomaterials are of paramount importance but remain particularly difficult to characterize at the molecular level because both sensitive and selective techniques are mandatory. Here, it is demonstrated that unprecedented insights into calcium environments, for example the differentiation of surface and core species of hydroxyapatite nanoparticles, can be obtained using solid-state NMR, when combined with dynamic nuclear polarization. Although calcium represents an ideal NMR target here (and de facto for a large variety of calcium-derived materials), its stable NMR-active isotope, calcium-43, is a highly unreceptive probe. Using the sensitivity gains from dynamic nuclear polarization, not only could calcium-43 NMR spectra be obtained easily, but natural isotopic abundance 2D correlation experiments could be recorded for calcium-43 in short experimental time. This opens perspectives for the detailed study of interfaces in nanostructured materials of the highest biological interest as well as calcium-based nanosystems in general., Competing Interests: The authors declare no competing financial interests.

Unlabelled: Benzoxaboroles are a family of molecules that are finding an increasing number of applications in the biomedical field, particularly as a "privileged scaffold" for the design of new drugs. Here, for the first time, we determine the interaction of these molecules with hydroxyapatites, in view of establishing (i) how benzoxaborole drugs may adsorb onto biological apatites, as this could impact on their bioavailability, and (ii) how apatite-based materials can be used for their formulation. Studies on the adsorption of the benzoxaborole motif (C7H7BO2, referred to as BBzx) on two different apatite phases were thus performed, using a ceramic hydroxyapatite (HAceram) and a nanocrystalline hydroxyapatite (HAnano), the latter having a structure and composition more similar to the one found in bone mineral. In both cases, the grafting kinetics and mechanism were studied, and demonstration of the surface attachment of the benzoxaborole under the form of a tetrahedral benzoxaborolate anion was established using (11)B solid state NMR (including (11)B-(31)P correlation experiments). Irrespective of the apatite used, the grafting density of the benzoxaborolates was found to be low, and more generally, these anions demonstrated a poor affinity for apatite surfaces, notably in comparison with other anions commonly found in biological media, such as carboxylates and (organo)phosphates. The study was then extended to the adsorption of a molecule with antimicrobial and antifungal properties (3-piperazine-bis(benzoxaborole)), showing, on a more general perspective, how hydroxyapatites can be used for the development of novel formulations of benzoxaborole drugs., Statement of Significance: Benzoxaboroles are an emerging family of molecules which have attracted much attention in the biomedical field, notably for the design of new drugs. However, the way in which these molecules, once introduced in the body, may interact with bone mineral is still unknown, and the possibility of associating benzoxaboroles to calcium phosphates for drug-formulation purposes has not been looked into. Here, we describe the first study of the adsorption of benzoxaboroles on hydroxyapatite, which is the main mineral phase present in bone. We describe the mode of grafting of benzoxaboroles on this material, and show that they only weakly bind to its surface, especially in comparison to other ionic species commonly found in physiological media, such as phosphates and carboxylates. This demonstrates that administered benzoxaborole drugs are unlikely to remain adsorbed on hydroxyapatite surfaces for long periods of time, which means that their biodistribution will not be affected by such phenomena. Moreover, this work shows that the formulation of benzoxaborole drugs by association to calcium phosphates like hydroxyapatite will lead to a rapid release of the molecules., (Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Unlabelled: The development of bioactive phosphate-based glasses is essential in biomaterials science, and especially for bone substitution applications. In this context, the preparation of amorphous calcium-phosphorus hydroxide/oxide monoliths at low temperature is a key challenge for being able to develop novel hybrid materials for these applications. We herein report for the first time the synthesis and physical chemical characterisation of a novel family of pyrophosphate-based glasses (with the formula: {[(Ca(2+))1-x(H(+)/K(+))2x]2[(P2O7(4-))1-y(PO4(3-))4y/3]} n(H2O)), which were prepared by soft chemistry using low temperatures (T<70°C) and water as a solvent. The effect of the initial Ca/Pyrophosphate ratio on the structure and morphology of these pyrophosphate glasses was investigated in detail. Depending on this ratio, a glass (mixed calcium pyro- and orthophosphate) or a glass-ceramic (Ca10K4(P2O7)6·9H2O crystals embedded in the amorphous phase) was obtained. The proportion of the crystalline phase increased with an increase in the Ca/Pyrophosphate ratio in the batch solution. As expected for a glass, the formation of the glassy material was demonstrated not to be thermodynamically but rather kinetically driven, and the washing step was found to be crucial to prevent crystallisation. The stability of the amorphous phase was discussed considering the structural degrees of freedom of pyrophosphate entities, ionic strength of the initial solution and the inhibitory effect of orthophosphate ions. Overall, this new strategy of preparation of monolithic calcium-(pyro)phosphate based glasses using soft chemistry in water is highly promising in view of preparing new functional organic-inorganic hybrids for bone substitution applications., Statement of Significance: Phosphate-based glasses have gradually emerged as a potential alternative to silicate bioactive glasses in order to induce different biological mechanisms of degradation. The synthesis of such monolithic glasses at low temperature is a key step to allow new inorganic glass compositions to be reached and hybrid materials to be prepared. Although sol-gel and coacervate methods (respectively orthophosphate and metaphosphate precursors) have already been described to prepare such glasses, the use of toxic solvents and/or the final temperature treatment associated to these processes could limit the use of these materials for biomedical applications and/or the further development of hybrids. It is shown here that pyrophosphate precursors are an alternative strategy to obtain monolithic calcium (pyro)phosphate glasses under soft conditions (water solvent, 70°C)., (Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

The reactivity of benzimidazol-2-ylidenes with respect to gold nanoparticles (AuNPs) has been investigated using a combined experimental and computational approach. First, the grafting of benzimidazol-2-ylidenes bearing benzyl groups on the nitrogen atoms is described, and comparisons are made with structurally similar N-heterocyclic carbenes (NHCs) bearing other N-groups. Similar reactivity was observed for all NHCs, with 1) the erosion of the AuNPs under the effect of the NHC and 2) the formation of bis(NHC) gold complexes. DFT calculations were performed to investigate the modes of grafting of such ligands, to determine adsorption energies, and to rationalize the spectroscopic data. Two types of computational models were developed to describe the grafting onto large or small AuNPs, with either periodic or cluster-type DFT calculations. Calculations of NMR parameters were performed on some of these models, and discussed in light of the experimental data., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Hydrated calcium pyrophosphates (CPP, Ca2P2O7·nH2O) are a fundamental family of materials among osteoarticular pathologic calcifications. In this contribution, a comprehensive multinuclear NMR (Nuclear Magnetic Resonance) study of four crystalline and two amorphous phases of this family is presented. (1)H, (31)P and (43)Ca MAS (Magic Angle Spinning) NMR spectra were recorded, leading to informative fingerprints characterizing each compound. In particular, different (1)H and (43)Ca solid state NMR signatures were observed for the amorphous phases, depending on the synthetic procedure used. The NMR parameters of the crystalline phases were determined using the GIPAW (Gauge Including Projected Augmented Wave) DFT approach, based on first-principles calculations. In some cases, relaxed structures were found to improve the agreement between experimental and calculated values, demonstrating the importance of proton positions and pyrophosphate local geometry in this particular NMR crystallography approach. Such calculations serve as a basis for the future ab initio modeling of the amorphous CPP phases., Statement of Significance: The general concept of NMR crystallography is applied to the detailed study of calcium pyrophosphates (CPP), whether hydrated or not, and whether crystalline or amorphous. CPP are a fundamental family of materials among osteoarticular pathologic calcifications. Their prevalence increases with age, impacting on 17.5% of the population after the age of 80. They are frequently involved or associated with acute articular arthritis such as pseudogout. Current treatments are mainly directed at relieving the symptoms of joint inflammation but not at inhibiting CPP formation nor at dissolving these crystals. The combination of advanced NMR techniques, modeling and DFT based calculation of NMR parameters allows new original insights in the detailed structural description of this important class of biomaterials., (Copyright © 2016. Published by Elsevier Ltd.)

Benzoxaboroles are a family of organoboron molecules, which have been finding over the past few years an increasing number of biological applications, notably for the design of new drugs. Given that these molecules are still relatively new in the biomedical context, very few investigations regarding their formulation have been reported to date. Here, a complete study on the formulation of benzoxaboroles in a biopolymer, poly-l-lactic acid (PLLA), is reported. The incorporation of two small benzoxaboroles, namely the simplest benzoxaborole molecule (BBzx) and the antifungal drug tavaborole (AN2690), inside PLLA films was investigated. Different variations in the film composition and texture were looked into, by performing a heat-treatment on the PLLA films, or by preparing PLLA-PEO (polyethylene oxide) blends or PLLA-LDH (layered double hydroxide) composites. In each case, the impact of these changes in formulation on the local environment of the benzoxaboroles in the material (as determined by multinuclear solid state NMR), and on the kinetics of release in physiological media were analyzed, showing that a variety of release profiles could be achieved. Finally, cellular assays were carried out looking at the migration of MDA-MB-231 cancer cells. These tests revealed for the first time that benzoxaboroles like AN2690 and BBzx inhibited the migration of these cells. Moreover, the molecules incorporated in the films were found to remain active, and their effect on cancer cells was directly related to the release kinetics from the films. All in all, PLLA-based materials appear as highly versatile and attractive matrices for formulating benzoxaborole-based drugs.

Using aminoglycoside antibiotics as drug models, it was shown that electrostatic complexes between hydrophilic drugs and oppositely charged double-hydrophilic block copolymers can form ordered mesophases. This phase behavior was evidenced by using poly(acrylic acid)-block-poly(ethylene oxide) block copolymers in the presence of silica precursors, and this allowed preparing drug-loaded mesoporous silica directly from the drug-polymer complexes. The novel synthetic strategy of the hybrid materials is highly efficient, avoiding waste and multistep processes; it also ensures optimal drug loading and provides pH-dependence of the drug release from the materials.

The surface of aligned, electrospun poly-L-lactic acid (PLLA) fibers was chemically modified to determine if surface chemistry and hydrophilicity could improve neurite extension from chick dorsal root ganglia. Specifically, diethylenetriamine (DTA, for amine functionalization), 2-(2-aminoethoxy)ethanol (AEO, for alcohol functionalization), or GRGDS (cell adhesion peptide) were covalently attached to the surface of electrospun fibers. Water contact angle measurements revealed that surface modification of electrospun fibers significantly improved fiber hydrophilicity compared to unmodified fibers (p < 0.05). Scanning electron microscopy (SEM) of fibers revealed that surface modification changed fiber topography modestly, with DTA modified fibers displaying the roughest surface structure. Degradation of chemically modified fibers revealed no change in fiber diameter in any group over a period of seven days. Unexpectedly, neurites from chick DRG were longest on fibers without surface modification (1651 ± 488 μm) and fibers containing GRGDS (1560 ± 107 μm). Fibers modified with oxygen plasma (1240 ± 143 μm) or DTA (1118 ± 82 μm) produced shorter neurites than the GRGDS or unmodified fibers, but were not statistically shorter than unmodified and GRGDS modified fibers. Fibers modified with AEO (844 ± 151 μm) were significantly shorter than unmodified and GRGDS modified fibers (p<0.05). Based on these results, we conclude that fiber hydrophilic enhancement alone on electrospun PLLA fibers does not enhance neurite outgrowth. Further work must be conducted to better understand why neurite extension was not improved on more hydrophilic fibers, but the results presented here do not recommend hydrophilic surface modification for the purpose of improving neurite extension unless a bioactive ligand is used.

Nanocrystalline calcium phosphate apatites are biomimetic compounds analogous to bone mineral and are at the origin of the bioactivity of most biomaterials used as bone substitutes. Their unique surface reactivity originates from the presence of a hydrated layer containing labile ions (mostly divalent ones). So the setup of 3D biocompatible apatite-based bioceramics exhibiting a high reactivity requests the development of «low» temperature consolidation processes such as spark plasma sintering (SPS), in order to preserve the characteristics of the hydrated nanocrystals. However, mechanical performances may still need to be improved for such nanocrystalline apatite bioceramics, especially in view of load-bearing applications. The reinforcement by association with biopolymers represents an appealing approach, while preserving the advantageous biological properties of biomimetic apatites. Herein, we report the preparation of composites based on biomimetic apatite associated with various quantities of microcrystalline cellulose (MCC, 1-20 wt%), a natural fibrous polymer. The SPS-consolidated composites were analyzed from both physicochemical (X-ray diffraction, Fourier transform infrared, solid state NMR) and mechanical (Brazilian test) viewpoints. The preservation of the physicochemical characteristics of apatite and cellulose in the final material was observed. Mechanical properties of the composite materials were found to be directly related to the polymer/apatite ratios and a maximum crushing strength was reached for 10 wt% of MCC.

Ultrasmall gold nanoparticles (Au UNPs) represent a unique class of nanomaterials making them very attractive for certain applications. Herein, we developed an organometallic approach to the synthesis of Au UNPs stabilized with the C18H37-NHC ligand by the solvent free thermolysis of [RMIM][Au(C6F5)2] () or [Au(C6F5)(RNHC)] () (with R = C18H37-), by controlling the reactivity of pentafluorophenyl ligands as deprotonating or reductive elimination agents; Au UNPs can be achieved by solvent free thermolysis. Pentafluorophenyl Au(i) complexes and are synthesized from the corresponding ionic and neutral precursors. The presence of long alkyl chain imidazolium or carbene species in the complexes makes them to behave as isotropic liquids at moderate temperatures. The use of multinuclear NMR allows the description of the mechanism of formation of the UNPs as well as the surface state of the UNPs.

TiO2/conjugated polymers are promising materials in solar energy conversion where efficient photoinduced charge transfers are required. Here, a "grafting-from" approach for the synthesis of TiO2 nanoparticles supported with conjugated polymer brushes is presented. Poly(3-hexylthiophene) (P3HT), a benchmark material for organic electronics, was selectively grown from TiO2 nanoparticles by surface-initiated Kumada catalyst-transfer polycondensation. The grafting of the polymer onto the surface of the TiO2 nanoparticles by this method was demonstrated by (1)H and (13)C solid-state NMR, X-ray photoelectron spectrometry, thermogravimetric analysis, transmission electron microscopy, and UV-visible spectroscopy. Sedimentation tests in tetrahydrofuran revealed improved dispersion stability for the TiO2@P3HT hybrid material. Films were produced by solvent casting, and the quality of the dispersion of the modified TiO2 nanoparticles was evaluated by atomic force microscopy. The dispersion of the P3HT-coated TiO2 NPs in the P3HT matrix was found to be homogeneous, and the fibrillar structure of the P3HT matrix was maintained which is favorable for charge transport. Fluorescence quenching measurements on these hybrid materials in CHCl3 indicated improved photoinduced electron-transfer efficiency. All in all, better physicochemical properties for P3HT/TiO2 hybrid material were reached via the surface-initiated "grafted-from" approach compared to the "grafting-onto" approach.

A new method for the functionalization of detonation nanodiamonds (DNDs) is proposed, on the basis of surface modification with phosphonic dichloride derivatives. DNDs were first modified by phenylphosphonic dichloride, and the grafting modes and hydrolytic stability under neutral conditions were investigated using (1)H, (13)C, and (31)P solid state NMR spectroscopy, Fourier transform infrared spectroscopy, as well as elemental analysis. Then, in order to illustrate the possibilities offered by this method, DNDs functionalized by mesityl imidazolium groups were obtained by postmodification of DNDs modified by 12-bromododecylphosphonic dichloride. The oxidative thermal stability of the functionalized DNDs was investigated using thermogravimetric analysis.

The reaction of gold nanoparticles with benzimididazol-2-ylidene ligands leads to the formation of well-defined bis-carbene gold(i) complexes, as shown by characterization techniques such as powder XRD and solid state NMR.

In this contribution, the latest developments in solid state NMR are presented in the field of organic-inorganic (O/I) materials (or hybrid materials). Such materials involve mineral and organic (including polymeric and biological) components, and can exhibit complex O/I interfaces. Hybrids are currently a major topic of research in nanoscience, and solid state NMR is obviously a pertinent spectroscopic tool of investigation. Its versatility allows the detailed description of the structure and texture of such complex materials. The article is divided in two main parts: in the first one, recent NMR methodological/instrumental developments are presented in connection with hybrid materials. In the second part, an exhaustive overview of the major classes of O/I materials and their NMR characterization is presented., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Mechanical resilience of bone tissue decreases with age. The ability to comprehensively probe and understand bone properties could help alleviate this problem. One important aspect of bone quality that has recently been made evident is the presence of dilatational bands formed by osteocalcin (OC) and osteopontin (OPN), which contribute to fracture toughness. However, experimental evidence of the structural role of these two proteins at the organic-mineral interface in bone is still needed. Solid state nuclear magnetic resonance (SSNMR) is emerging as a useful technique in probing molecular level aspects of bone. Here, we present the first SSNMR study of bone tissue from genetically modified mice lacking OC and/or OPN. Probing the mineral phase, the organic matrix and their interface revealed that, despite the absence of OC and OPN, the organic matrix and mineral were well preserved, and the overall exposure of collagen to hydroxyapatite (HA) nanoparticles was hardly affected. However, the proximity to the HA surface was slightly increased for a number of bone components including less abundant amino acids like lysine, suggesting that this is how the tissue compensates for the lack of OC and OPN. Taken together, the NMR data supports the recently proposed model, in which the contribution of OC-OPN to fracture toughness is related to their presence at the extrafibrillar organic-mineral interfaces, where they reinforce the network of mineralized fibrils and form dilatational bands. In an effort toward further understanding the structural role of individual amino acids of low abundance in bone, we then explored the possibility of specific (13)C enrichment of mouse bone, and report the first SSNMR spectra of 97% (13)C lysine-enriched tissue. Results show that such isotopic enrichment allows valuable molecular-level structural information to be extracted, and sheds light on post-translational modifications undergone by specific amino acids in vivo.