Calcium sulphate biomineralisation: Artefact of sample preparation?

Calcium biomineralisation is widely documented in plants. However, crystallisation of Ca-sulphate-containing minerals is closely related to water content, and sample processing, such as drying, alters the water balance of plant tissues. We hypothesised that common sample processing practices may favour the formation of crystals, leading to spurious crystallisation not observed in unaltered plant tissues. We selected three species (Ononis tridentata, Helianthemum squamatum and Gypsophila struthium) with reported gypsum biomineralisation. We used x-ray diffractometry on fresh intact or sliced leaves, and on the same leaves processed by subsequent drying, to address whether sample processing alters crystal formation. Ca-sulphate crystals were detected in dry samples of all species but not in fresh intact samples. Ca-sulphate crystallisation occurred in some cut fresh samples, although the accumulation greatly increased after drying. In addition, G. struthium exhibited Ca-oxalate crystals in both fresh and dry treatments, with a tendency for greater accumulation in dry treatments. Our results demonstrate that the Ca-sulphate crystals observed by x-ray diffractometry in these species are artefacts caused by common sample processing practices, such as excessive drying and slicing samples. We encourage future studies on the biomineral potential of plants to avoid the use of procedures that alter the water balance of tissues.

A sustainable one-pot method to transform seashell waste calcium carbonate to osteoinductive hydroxyapatite micro-nanoparticles.

We have developed a straightforward, one-pot, low-temperature hydrothermal method to transform oyster shell waste particles (bCCP) from the species Crassostrea gigas (Mg-calcite, 5 wt% Mg) into hydroxyapatite (HA) micro/nanoparticles. The influence of the P reagents (H3PO4, KH2PO4, and K2HPO4), P/bCCP molar ratios (0.24, 0.6, and 0.96), digestion temperatures (25-200 °C), and digestion times (1 week-2 months) on the transformation process was thoroughly investigated. At 1 week, the minimum temperature to yield the full transformation significantly reduced from 160 °C to 120 °C when using K2HPO4 instead of KH2PO4 at a P/bCCP ratio of 0.6, and even to 80 °C at a P/bCCP ratio of 0.96. The transformation took place via a dissolution-reprecipitation mechanism driven by the favorable balance between HA precipitation and bCCP dissolution, due to the lower solubility product of HA than that of calcite at any of the tested temperatures. Both the bCCP and the derived HA particles were cytocompatible for MG-63 human osteosarcoma cells and m17.ASC murine mesenchymal stem cells, and additionally, they promoted the osteogenic differentiation of m17.ASC, especially the HA particles. Because of their physicochemical features and biological compatibility, both particles could be useful osteoinductive platforms for translational applications in bone tissue engineering.

Rational Coformer Selection in the Development of 6-Propyl-2-thiouracil Pharmaceutical Cocrystals.

Pharmaceutical multicomponent solids have proved to efficiently modulate the physicochemical properties of active pharmaceutical ingredients. In this context, polyphenols are interesting coformers for designing pharmaceutical cocrystals due to their wide safety profile and interesting antioxidant properties. The novel 6-propyl-2-thiouracil multicomponent solids have been obtained by mechanochemical synthesis and fully characterized by powder and single-crystal X-ray diffraction methods. The analysis of supramolecular synthons has been further performed with computational methods, with both results revealing a robust supramolecular organization influenced by the different positions of the hydroxyl groups within the polyphenolic coformers. All novel 6-propyl-2-thiouracil cocrystals show an enhanced solubility profile, but unfortunately, their thermodynamic stability in aqueous media is limited to 24 h.

Metformin-NSAIDs Molecular Salts: A Path towards Enhanced Oral Bioavailability and Stability.

According to the World Health Organization, more than 422 million people worldwide have diabetes. The most common oral treatment for type 2 diabetes is the drug metformin (MTF), which is usually formulated as a hydrochloride to achieve higher water solubility. However, this drug is also highly hygroscopic, thus showing stability problems. Another kind of worldwide prescribed drug is the non-steroidal anti-inflammatory drug (NSAID). These latter, on the contrary, show a low solubility profile; therefore, they must be administered at high doses, which increases the probability of secondary effects. In this work, novel drug-drug pharmaceutical solids combining MTF-NSAIDs have been synthesized in solution or by mechanochemical methods. The aim of this concomitant treatment is to improve the physicochemical properties of the parent active pharmaceutical ingredients. After a careful solid-state characterization along with solubility and stability studies, it can be concluded that the new molecular salt formulations enhance not only the stability of MTF but also the solubility of NSAIDs, thus giving promising results regarding the development of these novel pharmaceutical multicomponent solids.

Enhanced NSAIDs Solubility in Drug-Drug Formulations with Ciprofloxacin.

Drug-drug salts are a kind of pharmaceutical multicomponent solid in which the two co-existing components are active pharmaceutical ingredients (APIs) in their ionized forms. This novel approach has attracted great interest in the pharmaceutical industry since it not only allows concomitant formulations but also has proved potential to improve the pharmacokinetics of the involved APIs. This is especially interesting for those APIs that have relevant dose-dependent secondary effects, such as non-steroidal anti-inflammatory drugs (NSAIDs). In this work, six multidrug salts involving six different NSAIDs and the antibiotic ciprofloxacin are reported. The novel solids were synthesized using mechanochemical methods and comprehensively characterized in the solid state. Moreover, solubility and stability studies, as well as bacterial inhibition assays, were performed. Our results suggest that our drug-drug formulations enhanced the solubility of NSAIDs without affecting the antibiotic efficacy.

Physico-chemical and biological characterization of a new bovine bone mineral matrix available for human usage.

BACKGROUND: Anorganic bovine bone has been deeply studied for bone regeneration in the oral cavity. Different manufacturing processes can modify the final composition of the biomaterial and the responses that induce. AIM: To evaluate the physico-chemical characteristics of a bovine bone mineral matrix and the clinical, radiographical, histological, and mRNA results after using it for maxillary sinus floor augmentation in humans. MATERIALS AND METHODS: First, the physical-chemical characteristics of the biomaterial were evaluated by X-ray powder diffraction, X-ray fluorescence, and electron microscopy. A frequently used biomaterial with the same animal origin was used as comparator. Then, a clinical study was designed for evaluating clinical, radiographical, histological, and mRNA outcomes. Patients in need of two-stage maxillary sinus floor augmentation were included in the study. Six months after the grafting procedure, a bone biopsy was collected for evaluation. RESULTS: In terms of physico-chemical characteristics, no differences were found between both biomaterials. Clinically, 10 patients were included in the study. After 6 months, clinical and radiographical data showed adequate outcomes for allowing implant placement. Histological, immunohistochemical and mRNA analyses showed that the biomaterial in use provides biological support to induce responses similar to those of other commonly used biomaterials. CONCLUSION: Bovine bone mineral matrix (Creos™ Xenogain) used as a single material for maxillary sinus floor augmentation shows adequate biological, clinical, and radiological outcomes. In fact, the results from this study are similar to those reported in the literature for another bovine bone-derived biomaterial with whom it shares composition and micro- and nanoscale characteristics.

Mechanisms shaping the gypsum stromatolite-like structures in the Salar de Llamara (Atacama Desert, Chile).

The explanation of the origin of microbialites and specifically stromatolitic structures is a problem of high relevance for decoding past sedimentary environments and deciphering the biogenicity of the oldest plausible remnants of life. We have investigated the morphogenesis of gypsum stromatolite-like structures currently growing in shallow ponds (puquíos) in the Salar de Llamara (Atacama Desert, Northern Chile). The crystal size, aspect ratio, and orientation distributions of gypsum crystals within the structures have been quantified and show indications for episodic nucleation and competitive growth of millimetric to centimetric selenite crystals into a radial, branched, and loosely cemented aggregate. The morphogenetical process is explained by the existence of a stable vertical salinity gradient in the ponds. Due to the non-linear dependency of gypsum solubility as a function of sodium chloride concentration, the salinity gradient produces undersaturated solutions, which dissolve gypsum crystals. This dissolution happens at a certain depth, narrowing the lower part of the structures, and producing their stromatolite-like morphology. We have tested this novel mechanism experimentally, simulating the effective dissolution of gypsum crystals in stratified ponds, thus providing a purely abiotic mechanism for these stromatolite-like structures.

Luminescent Citrate-Functionalized Terbium-Substituted Carbonated Apatite Nanomaterials: Structural Aspects, Sensitized Luminescence, Cytocompatibility, and Cell Uptake Imaging.

This work explores the preparation of luminescent and biomimetic Tb3+-doped citrate-functionalized carbonated apatite nanoparticles. These nanoparticles were synthesized employing a citrate-based thermal decomplexing precipitation method, testing a nominal Tb3+ doping concentration between 0.001 M to 0.020 M, and a maturation time from 4 h to 7 days. This approach allowed to prepare apatite nanoparticles as a single hydroxyapatite phase when the used Tb3+ concentrations were (i) ≤ 0.005 M at all maturation times or (ii) = 0.010 M with 4 h of maturation. At higher Tb3+ concentrations, amorphous TbPO4·nH2O formed at short maturation times, while materials consisting of a mixture of carbonated apatite prisms, TbPO4·H2O (rhabdophane) nanocrystals, and an amorphous phase formed at longer times. The Tb3+ content of the samples reached a maximum of 21.71 wt%. The relative luminescence intensity revealed an almost linear dependence with Tb3+ up to a maximum of 850 units. Neither pH, nor ionic strength, nor temperature significantly affected the luminescence properties. All precipitates were cytocompatible against A375, MCF7, and HeLa carcinogenic cells, and also against healthy fibroblast cells. Moreover, the luminescence properties of these nanoparticles allowed to visualize their intracellular cytoplasmic uptake at 12 h of treatment through flow cytometry and fluorescence confocal microscopy (green fluorescence) when incubated with A375 cells. This demonstrates for the first time the potential of these materials as nanophosphors for living cell imaging compatible with flow cytometry and fluorescence confocal microscopy without the need to introduce an additional fluorescence dye. Overall, our results demonstrated that Tb3+-doped citrate-functionalized apatite nanoparticles are excellent candidates for bioimaging applications.

A Comprehensive Methodology for Monitoring Evaporitic Mineral Precipitation and Hydrochemical Evolution of Saline Lakes: The Case of Lake Magadi Soda Brine (East African Rift Valley, Kenya).

Lake Magadi, East African Rift Valley, is a hyperalkaline and saline soda lake highly enriched in Na+, K+, CO3 2-, Cl-, HCO3 -, and SiO2 and depleted in Ca2+ and Mg2+, where thick evaporite deposits and siliceous sediments have been forming for 100 000 years. The hydrogeochemistry and the evaporite deposits of soda lakes are subjects of growing interest in paleoclimatology, astrobiology, and planetary sciences. In Lake Magadi, different hydrates of sodium carbonate/bicarbonate and other saline minerals precipitate. The precipitation sequence of these minerals is a key for understanding the hydrochemical evolution, the paleoenvironmental conditions of ancient evaporite deposits, and industrial crystallization. However, accurate determination of the precipitation sequence of these minerals was challenging due to the dependency of the different hydrates on temperature, water activity, pH and pCO2, which could induce phase transformation and secondary mineral precipitation during sample handling. Here, we report a comprehensive methodology applied for monitoring the evaporitic mineral precipitation and hydrochemical evolution of Lake Magadi. Evaporation and mineral precipitations were monitored by using in situ video microscopy and synchrotron X-ray diffraction of acoustically levitated droplets. The mineral patterns were characterized by ex situ Raman spectroscopy, X-ray diffraction, and scanning electron microscopy. Experiments were coupled with thermodynamic models to understand the evaporation and precipitation-driven hydrochemical evolution of brines. Our results closely reproduced the mineral assemblages, patterns, and textural relations observed in the natural setting. Alkaline earth carbonates and fluorite were predicted to precipitate first followed by siliceous sediments. Among the salts, dendritic and acicular trona precipitate first via fractional crystallization-reminiscent of grasslike trona layers of Lake Magadi. Halite/villiaumite, thermonatrite, and sylvite precipitate sequentially after trona from residual brines depleted in HCO3 -. The precipitation of these minerals between trona crystals resembles the precipitation process observed in the interstitial brines of the trona layers. Thermonatrite precipitation began after trona equilibrated with the residual brines due to the absence of excess CO2 input. We have shown that evaporation and mineral precipitation are the major drivers for the formation of hyperalkaline, saline, and SiO2-rich brines. The discrepancy between predicted and actual sulfate and phosphate ion concentrations implies the biological cycling of these ions. The combination of different in situ and ex situ methods and modeling is key to understanding the mineral phases, precipitation sequences, and textural relations of modern and ancient evaporite deposits. The synergy of these methods could be applicable in industrial crystallization and natural brines to reconstruct the hydrogeochemical and hydroclimatic conditions of soda lakes, evaporite settings, and potentially soda oceans of early Earth and extraterrestrial planets.

Production of Cross-Linked Lipase Crystals at a Preparative Scale.

The autoimmobilization of enzymes via cross-linked enzyme crystals (CLECs) has regained interest in recent years, boosted by the extensive knowledge gained in protein crystallization, the decrease of cost and laboriousness of the process, and the development of potential applications. In this work, we present the crystallization and preparative-scale production of reinforced cross-linked lipase crystals (RCLLCs) using a commercial detergent additive as a raw material. Bulk crystallization was carried out in 500 mL of agarose media using the batch technique. Agarose facilitates the homogeneous production of crystals, their cross-linking treatment, and their extraction. RCLLCs were active in an aqueous solution and in hexane, as shown by the hydrolysis of p-nitrophenol butyrate and α-methylbenzyl acetate, respectively. RCLLCs presented both high thermal and robust operational stability, allowing the preparation of a packed-bed chromatographic column to work in a continuous flow. Finally, we determined the three-dimensional (3D) models of this commercial lipase crystallized with and without phosphate at 2.0 and 1.7 Å resolutions, respectively.

Crystallization, Luminescence and Cytocompatibility of Hexagonal Calcium Doped Terbium Phosphate Hydrate Nanoparticles.

Luminescent lanthanide-containing biocompatible nanosystems represent promising candidates as nanoplatforms for bioimaging applications. Herein, citrate-functionalized calcium-doped terbium phosphate hydrate nanophosphors of the rhabdophane type were prepared at different synthesis times and different Ca2+/Tb3+ ratios by a bioinspired crystallization method consisting of thermal decomplexing of Ca2+/Tb3+/citrate/phosphate/carbonate solutions. Nanoparticles were characterized by XRD, TEM, SEM, HR-TEM, FTIR, Raman, Thermogravimetry, inductively coupled plasma spectroscopy, thermoanalysis, dynamic light scattering, electrophoretic mobility, and fluorescence spectroscopy. They displayed ill-defined isometric morphologies with sizes ≤50 nm, hydration number n ~ 0.9, tailored Ca2+ content (0.42-8.11 wt%), and long luminescent lifetimes (800-2600 µs). Their relative luminescence intensities in solid state are neither affected by Ca2+, citrate content, nor by maturation time for Ca2+ doping concentration in solution below 0.07 M Ca2+. Only at this doping concentration does the maturation time strongly affect this property, decreasing it. In aqueous suspensions, neither pH nor ionic strength nor temperature affect their luminescence properties. All the nanoparticles displayed high cytocompatibility on two human carcinoma cell lines and cell viability correlated positively with the amount of doping Ca2+. Thus, these nanocrystals represent promising new luminescent nanoprobes for potential biomedical applications and, if coupled with targeting and therapeutic moieties, they could be effective tools for theranostics.

Bioinspired crystallization, sensitized luminescence and cytocompatibility of citrate-functionalized Ca-substituted europium phosphate monohydrate nanophosphors.

Biocompatible nanosystems exhibiting long-lifetime (∼millisecond) luminescence features are particularly relevant in the field of bioimaging. In this study, citrate-functionalized calcium-doped europium phosphates nanophosphors of the rhabdophane type were prepared at different synthesis times by a bioinspired crystallization route, consisting in thermal decomplexing of Ca2+/Eu3+ /citrate/phosphate/carbonate solutions. The general formula of this material is CaαEu1-α(PO4)1-α(HPO4)α·nH2O, with α ranging from 0 to 0.58 and n ∼ 1. A thorough characterization of the nanoparticles has been carried out by XRD (including data processing with Topas 6.0), HR-TEM, TEM, FTIR, TG/DTA, ICP, dynamic light scattering (DLS), electrophoretic mobility, and fluorescence spectroscopy. Based on these results a crystallization mechanism involving the filling of cationic sites with Ca2+ions associated to a concomitant adjustment of the PO4/HPO4 ratio was proposed. Upon calcium doping, the aspect ratio of the nanoparticles as well as of the crystalline domains decreased and the relative luminescence intensity (R.L.I.) could be modulated. Neither the pH nor the ionic strength, nor the temperature (from 25 to 37 °C) affected significantly the R.L.I. of particles after resuspension in water, leading to rather steady luminescence features usable in a large domain of conditions. This new class of luminescent compounds has been proved to be fully cytocompatible relative to GTL-16 human carcinoma cells and showed an improved cytocompatibility as the Ca2+ content increased when contacted with the more sensitive m17. ASC murine mesenchymal stem cells. These biocompatible nanoparticles thus appear as promising new tailorable tools for biomedical applications as luminescent nanoprobes.

Thermal assisted self-organization of calcium carbonate.

Fabrication of mineral multi-textured architectures by self-organization is a formidable challenge for engineering. Current approaches follow a biomimetic route for hybrid materials based on the coupling of carbonate and organic compounds. We explore here the chemical coupling of silica and carbonate, leading to fabrication of inorganic-inorganic biomimetic structures known as silica-carbonate biomorphs. So far, biomorphic structures were restricted to orthorhombic barium, strontium, and calcium carbonate. We demonstrate that, monohydrocalcite a hydrous form of calcium carbonate with trigonal structure can also form biomorphic structures, thus showing biomorphic growth is not dictated by the carbonate crystal structure. We show that it is possible to control the growth regime, and therefore the texture and overall shape, by tuning the growth temperature, thereby shifting the textural pattern within the production of a given architecture. This finding opens a promising route to the fabrication of complex multi-textured self-organized material made of silica and chalk.

Luminescent biomimetic citrate-coated europium-doped carbonated apatite nanoparticles for use in bioimaging: physico-chemistry and cytocompatibility.

Nanomedicine covers the application of nanotechnologies in medicine. Of particular interest is the setup of highly-cytocompatible nanoparticles for use as drug carriers and/or for medical imaging. In this context, luminescent nanoparticles are appealing nanodevices with great potential for imaging of tumor or other targetable cells, and several strategies are under investigation. Biomimetic apatite nanoparticles represent candidates of choice in nanomedicine due to their high intrinsic biocompatibility and to the highly accommodative properties of the apatite structure, allowing many ionic substitutions. In this work, the preparation of biomimetic (bone-like) citrate-coated carbonated apatite nanoparticles doped with europium ions is explored using the citrate-based thermal decomplexing approach. The technique allows the preparation of the single apatitic phase with nanosized dimensions only at Eu3+ doping concentrations ≤0.01 M at some timepoints. The presence of the citrate coating on the particle surface (as found in bone nanoapatites) and Eu3+ substituting Ca2+ is beneficial for the preparation of stable suspensions at physiological pH, as witnessed by the ζ-potential versus pH characterizations. The sensitized luminescence features of the solid particles, as a function of the Eu3+ doping concentrations and the maturation times, have been thoroughly investigated, while those of particles in suspensions have been investigated at different pHs, ionic strengths and temperatures. Their cytocompatibility is illustrated in vitro on two selected cell types, the GTL-16 human carcinoma cells and the m17.ASC murine mesenchymal stem cells. This contribution shows the potentiality of the thermal decomplexing method for the setup of luminescent biomimetic apatite nanoprobes with controlled features for use in bioimaging.

Switchable Surface Hydrophobicity-Hydrophilicity of a Metal-Organic Framework.

Materials with surfaces that can be switched from high/superhydrophobicity to superhydrophilicity are useful for myriad applications. Herein, we report a metal-organic framework (MOF) assembled from ZnII ions, 1,4-benzenedicarboxylate, and a hydrophobic carborane-based linker. The MOF crystal-surface can be switched between hydrophobic and superhydrophilic through a chemical treatment to remove some of the building blocks.

Continuous Sensing Photonic Lab-on-a-Chip Platform Based on Cross-Linked Enzyme Crystals.

Microfluidics or lab-on-a-chip technology offer clear advantages over conventional systems such as a dramatic reduction of reagent consumption or a shorter analysis time, which are translated into cost-effective systems. In this work, we present a photonic enzymatic lab-on-a-chip reactor based on cross-linked enzyme crystals (CLECs), able to work in continuous flow, as a highly sensitive, robust, reusable, and stable platform for continuous sensing with superior performance as compared to the state of the art. The microreactor is designed to facilitate the in situ crystallization and crystal cross-linking generating enzymatically active material that can be stored for months/years. Thus, and by means of monolithically integrated micro-optics elements, continuous enzymatic reactions can be spectrophotometrically monitored. Lipase, an enzyme with industrial significance for catalyzed transesterification, hydrolysis, and esterification reactions, is used to demonstrate the potential of the microplatforms as both a continuous biosensor and a microreactor for the synthesis of high value compounds.

Engineering Homochiral Metal-Organic Frameworks by Spatially Separating 1D Chiral Metal-Peptide Ladders: Tuning the Pore Size for Enantioselective Adsorption.

The reaction of the chiral dipeptide glycyl-L(S)-glutamate with Co(II) ions produces chiral ladders that can be used as rigid 1D building units. Spatial separation of these building units with linkers of different lengths allows the engineering of homochiral porous MOFs with enhanced pore sizes, pore volumes, and surface areas. This strategy enables the synthesis of a family of isoreticular MOFs, in which the pore size dictates the enantioselective adsorption of chiral molecules (in terms of their size and enantiomeric excess).