Search

Magnetic nanoparticles (MNPs) are widely used for immobilizing complex biomolecules such as proteins due to their high loading capacity, biocompatibility, and easy magnetic harvesting. However, there is not a universal solution to achieve the oriented immobilization of the protein of interest. One common strategy involves the selective binding of poly-histidine-tagged (His-tag) proteins to divalent transition metal centers (TM2+) chelated by organic molecules like nitrilotriacetic acid (NTA). This article describes an innovative procedure to functionalize MNPs by simply growing of a thin layer of TM2+ hydroxides on the MNP surface, which increases the number of coordination centers and enhances the surface interaction with the protein surface. The immobilization yields of two sets of MNPs coated with Co2+, Ni2+ and Cu2+ hydroxides were compared to the conventional coating using the same inorganic cores but coated and functionalized with NTA-TM2+ moieties. It is shown that the one-step Co coated MNPs showed the highest selectivity and specificity towards His-tags. Thus, this coating not only reduce the functionalization steps to drive an oriented protein binding but also achieve an increment of immobilization yields over 1.5-fold respect to NTA-Co2+ analogous for his-tagged proteins with different 3D complexity while preserving similar or even higher enzymatic activity.

[EN] Coronaviridae viruses usually cause mild respiratory disease in humans, nevertheless, a new infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread globally since December 2019, resulting in the ongoing 2019-2021 coronavirus pandemic. The present invention discloses the use of coated iron oxyhydroxide nanoparticles (lOHNPs) for use in the treatment and/or prevention of viral infections caused by Coronaviridae, especially those caused by respiratory syndrome-related coronaviruses such as SARS-CoV and SARS-CoV-2 and MERS-CoV. The iron oxyhydroxide nanoparticles are coated with biocompatible molecules and polymers which bind to the nanoparticle core via oxygen (e.g. sorbitol and sucrose). The lOHNPs were found stable, non-cytotoxic in vitro and can efficiently impair virus replication and transcription. Furthermore, the lOHNPs are suitable for oral, intranasal or parenteral administration in combination with a pharmaceutical carrier., [FR] Les virus Coronaviridae provoquent généralement une maladie respiratoire légère chez l'homme, cependant, une nouvelle maladie infectieuse provoquée par le coronavirus 2 du syndrome respiratoire aigu sévère (SARS-CoV-2) se propage dans le monde depuis décembre 2019, conduisant à la pandémie actuelle de coronavirus de 2019-2021. La présente invention divulgue l'utilisation de nanoparticules d'oxyhydroxyde de fer (lOHNP) enrobées destinées à être utilisées dans le traitement et/ou la prévention d'infections virales provoquées par des Coronaviridae, notamment celles provoquées par des coronavirus associés à des syndromes respiratoires tels que le SARS-CoV, le SARS-CoV-2 et le MERS-CoV. Les nanoparticules d'oxyhydroxyde de fer sont revêtues de molécules et de polymères biocompatibles qui se lient au noyau de nanoparticule via l'oxygène (par exemple, le sorbitol et le saccharose). Les lOHNP se sont révélées stables, non cytotoxiques in vitro et peuvent altérer efficacement la réplication et la transcription du virus. En outre, les lOHNP sont appropriées pour une administration orale, intranasale ou parentérale en combinaison avec un support pharmaceutique.

The present invention relates to a process to carry out two or more enzymatic reactions in a reaction medium, wherein the process comprises providing a system comprising at least two substantially homogeneous and colloidal populations of magnetic nanoparticles (MNPs) and applying one or more external alternating magnetic field to produce the simultaneous or sequential activation of the enzymes functionalized on the surface of each population of MNPs so that the enzymatic reactions can be thermally activated.

[EN] The present invention relates to a process to carry out two or more enzymatic reactions in a reaction medium, wherein the process comprises providing a system comprising at least two substantially homogeneous and colloidal populations of magnetic nanoparticles (MNPs) and applying one or more external alternating magnetic field to produce the simultaneous or sequential activation of the enzymes functionalized on the surface of each population of MNPs so that the enzymatic reactions can be thermally activated., [FR] La présente invention concerne un procédé pour effectuer deux ou plusieurs réactions enzymatiques dans un milieu réactionnel, le procédé comprenant la fourniture d'un système comprenant au moins deux populations sensiblement homogènes et colloïdales de nanoparticules magnétiques (MNPs) et l'application d'un ou plusieurs champs magnétiques alternatifs externes pour produire l'activation simultanée ou séquentielle des enzymes fonctionnalisées sur la surface de chaque population de MNPs de telle sorte que les réactions enzymatiques peuvent être activées thermiquement.

Background Coronaviruses usually cause mild respiratory disease in humans but as seen recently, some human coronaviruses can cause more severe diseases, such as the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the global spread of which has resulted in the ongoing coronavirus pandemic. Results In this study we analyzed the potential of using iron oxide nanoparticles (IONPs) coated with biocompatible molecules like dimercaptosuccinic acid (DMSA), 3-aminopropyl triethoxysilane (APS) or carboxydextran (FeraSpin™ R), as well as iron oxyhydroxide nanoparticles (IOHNPs) coated with sucrose (Venofer®), or iron salts (ferric ammonium citrate -FAC), to treat and/or prevent SARS-CoV-2 infection. At non-cytotoxic doses, IONPs and IOHNPs impaired virus replication and transcription, and the production of infectious viruses in vitro, either when the cells were treated prior to or after infection, although with different efficiencies. Moreover, our data suggest that SARS-CoV-2 infection affects the expression of genes involved in cellular iron metabolism. Furthermore, the treatment of cells with IONPs and IOHNPs affects oxidative stress and iron metabolism to different extents, likely influencing virus replication and production. Interestingly, some of the nanoparticles used in this work have already been approved for their use in humans as anti-anemic treatments, such as the IOHNP Venofer®, and as contrast agents for magnetic resonance imaging in small animals like mice, such as the FeraSpin™ R IONP. Conclusions Therefore, our results suggest that IONPs and IOHNPs may be repurposed to be used as prophylactic or therapeutic treatments in order to combat SARS-CoV-2 infection.

[EN] New doped inorganic nanocrystals (NC) consisting on iron oxide and other metal integrated into the structure have been synthesized in one-step by adapting the oxidant precipitation synthesis route for magnetite. Different metals have been chosen to confer extra and unique properties to the resulting magnetic hetero-nanostructure: Co and Gd for enhancing transversal and longitudinal relaxivities for magnetic resonance imaging and Bi and Au for achieving X-ray absorption for computed tomography imaging. Apart of that, gold optical properties are interesting for photothermal therapy and iron oxides for magnetic hyperthermia. All metals have been incorporated into the magnetite structure in different ways during the synthesis: by forming a solid solution, by modifying the surface of the NCs, or by co-crystallization with the magnetite. The nanostructure formed in each case depends on the ionic radius of the secondary metal ion and the solubility of its hydroxide that control the co-precipitation in the initial steps of the reaction. Magnetic properties and imaging capabilities of the hetero-nanostructures have been analyzed as a function of the element distribution. Due to the synergistic combination of the different element properties, these magnetic hetero-nanostructures have great potential for biomedical applications.

It is well known that magnetic nanoparticles (MNPs) absorb magnetic energy when exposed to an alternating magnetic field (AMF) and therefore dissipate heat in their local environment. So far, the heat dissipated by the MNPs has been widely used in biomedicine for cancer treatment and drug delivery. The FET-OPEN project HOTZYMES proposes instead to exploit this property for the biotechnological production of pharmaceuticals and bio-commodities, by conjugating thermophilic enzymes on the MNPs surface in order to selectively tune their activity by triggering this highly localized magnetic heating while maintaining the reaction media at low temperature., To this aim superparamagnetic iron oxide nanoparticles coated with different organic shells (dimercaptusuccinic acid, DMSA; polyacrylic acid, PAA) have been selected due to their good colloidal stability, high density of functionalizable chemical groups (-COOH) and a good combination of different heating efficiencies. They have been further functionalized with NTA-Cu2+, a chelating agent commonly used in protein purification by metal affinity interaction. Herein, we demonstrate that we can selectively bind a His-tag variant of the superfolded GFP protein, thanks to the high affinity of the immobilized copper to the 6xhistidine tag genetically fused to the protein. Furthermore, using the same strategy, we were able to bind the tetrameric recombinant His-tag variant of the B. stearothermophilus alcohol dehydrogenase (ADH) with also high specificity. Additionally, we demonstrate that the application of an AMF to the selected protein@MNPs derivatives can lead to the generation of different temperature gradients on the MNPs surface by studying both the GFP fluorescence (that strongly depends on temperature), and the effect of the triggered local heating on ADH activity. These first results suggest the feasibility of tuning remotely protein properties by combining their coupling to MNPs with the application of alternating magnetic fields.

The histidine tag (His-tag) is one of the most used affinity-tag for protein purification due to its small size and versatility. Agarose and sepharose beads containing nitriloacetic acid (NTA) transition metal derivatives are widely used for the purification of His-tagged proteins, thanks to their high affinity to the His-tag genetically fused to the protein of interest [1]. The same chemistry can be used to conjugate enzymes to magnetic nanoparticles (MNPs) with the aim of tuning their activity by magnetic heating [2, 3]. Within the frame of the FET-OPEN project HOTZYMES (https://www.hotzymes.eu), different MNPs have been synthetized and coated with polyacrylic acid and dimercaptosuccinic acid, and then were further functionalized with NTA-Cu2+ as His-tag chelating agent. Different proteins were expressed as His-tag variants and immobilized on the MNPs, including monomeric (superfolded GFP), dimeric (C. violaceum transaminase, CvTA; C. uda cellobiose phosphorylase, CuCbP), and tetrameric (B. stearothermophilus alcohol dehydrogenase, ADH) variants. While for the monomeric protein selected as model no difficulties in the bioconjugation processes were observed, when using dimeric or tetrameric enzymes the aggregation of the MNPs occurs very easily due to crosslinking between the nanoparticles. This colloidal destabilization of the MNPs is favored due to its high surface area and the presence of several tags per enzyme molecule. To avoid this situation, different strategies have been developed: saturation of the binding site of the nanoparticles, presence of a small percentage of imidazole in reaction, changes in the incubation conditions (pH, ionic strength, …). First positive results confirm that by playing with different factors it is possible to conjugate different His-tagged enzymes to very different MNPs in terms of size, shape, surface area, and colloidal stability. Actually, it is possible to avoid protein and MNPs aggregation while obtaining good activity yiel

The contactless heating capacity of magnetic nanoparticles (MNPs) has been exploited in fields such as hyperthermia cancer therapy, catalysis, and enzymatic thermal regulation. Herein, we propose an advanced technology to generate multiple local temperatures in a single-pot reactor by exploiting the unique nanoheating features of iron oxide MNPs exposed to alternating magnetic fields (AMFs). The heating power of the MNPs depends on their magnetic features but also on the intensity and frequency conditions of the AMF. Using a mixture of diluted colloids of MNPs we were able to generate a multi-hot-spot reactor in which each population of MNPs can be selectively activated by adjusting the AMF conditions. The maximum temperature reached at the surface of each MNP was registered using independent fluorescent thermometers that mimic the molecular link between enzymes and MNPs. This technology paves the path for the implementation of a selective regulation of multienzymatic reactions.

The scientific community has made great efforts in advancing magnetic hyperthermia for the last two decades after going through a sizeable research lapse from its establishment. All the progress made in various topics ranging from nanoparticle synthesis to biocompatibilization and in vivo testing have been seeking to push the forefront towards some new clinical trials. As many, they did not go at the expected pace. Today, fruitful international cooperation and the wisdom gain after a careful analysis of the lessons learned from seminal clinical trials allow us to have a future with better guarantees for a more definitive takeoff of this genuine nanotherapy against cancer. Deliberately giving prominence to a number of critical aspects, this opinion review offers a blend of state-of-the-art hints and glimpses into the future of the therapy, considering the expected evolution of science and technology behind magnetic hyperthermia.

We report results of CaCO3 crystallization experiments by counter diffusion in agar gel with two different solid contents (0.5 and 2 wt %) and two solute concentrations (0.1 M CaCl2, 0.1 M Na2CO3; 0.5 M CaCl2, 0.5 M Na2CO3). Solute concentration and hydrogel strength influence the characteristics of the gel–mineral composite formation. High reagent solution concentrations give rise to high supersaturation and high growth rates. When combined with a light gel, single crystal composites form; in a dense gel, the aggregates are mosaic crystal composites. Low reagent solution concentrations result in low supersaturation and low growth rates; when combined with a light gel, single crystal composites form; in a dense gel, the precipitate is a co-oriented polycrystal composite. Gel occlusion within the mineral increases with gel density. Gel distribution inside the mineral is homogeneous for high growth rates. For low growth rates, the gel accumulates locally in the precipitates. Light gels are pushed ahead by the growing crystals, and gel occlusion into the mineral is decreased; at low reagent solution concentrations, slightly more gel gets occluded. In conclusion, agar gel solid content determines the amount of gel occlusion and calcite orientation organization; reagent solution concentration influences the mode of gel distribution inside the mineral/gel composite aggregates.

In vitro experiments have shown the great potential of magnetic nanocarriers for multimodal imaging diagnosis and non-invasive therapies. However, their extensive clinical application is still jeopardized by a fast retention in the reticuloendothelial system (RES). The other issue that restrains their potential performance is slow degradation and excretion, which increases their risks of toxicity. We report a promising case in which multicore iron oxide nanoparticles coated with a poly(4-vinylpyridine) polyethylene glycol copolymer show low RES retention and high urinary excretion, as confirmed by single photon emission computerized tomography (SPECT), gamma counting, magnetic resonance imaging (MRI) and electron microscopy (EM) biodistribution studies. These iron oxide-copolymer nanoparticles have a high PEG density in their coating which may be responsible for this effect. Moreover, they show a clear negative contrast in the MR imaging of the kidneys. These nanoparticles with an average hydrodynamic diameter of approximately 20 nm were nevertheless able to cross the glomerulus wall which has an effective pore size of approximately 6 nm. A transmission electron microscopy inspection of kidney tissue revealed the presence of iron containing nanoparticle clusters in proximal tubule cells. This therefore makes them exceptionally useful as magnetic nanocarriers and as new MRI contrast agents for the kidneys.

[EN] Continuous processes are always preferred over batch ones when reproducible and scalable industrial procedures are needed. This work illustrates the production of magnetite nanoparticles by oxidative precipitation in aqueous media, following a continuous approach that offers additional advantages. Particularly, the developed reaction setup succeeds (i) the complete separation of the green rust’s precipitation from Fe3O4 nucleation, (ii) the achievement of constant concentrations in all ionic and solid forms throughout the production line when steady-state is reached, what means constant supersaturation from both the formation of green rust and Fe3O4, and (iii) the possibility to control critical parameters, such as OH- excess over the initial stoichiometric Fe(OH)2 precipitation, through online regulation of synthesis parameters such as the reactor’s pH and redox potential. Importantly, continuous flow synthesis of Fe3O4 nanoparticles enables high production capacities, low energy consumption and proportional scale-up at any volume. As a proof of concept, obtained nanoparticles were evaluated according to their magnetic response as potential magnetic hyperthermia agents indicating significant improvement of heating efficiency that goes up to 1.5-2 kW/gFe3O4 for both smaller (~40 nm) and larger (~200 nm) particles.

[EN] Thermally activated relaxation over energy barriers concurrently related to local properties and interparticle interactions constitutes a major contribution to both the coercivity and the applied field frequency dependence of that quantity. We have measured the slow magnetic relaxation of magnetite nanoparticles (NPs), synthetized by using the oxidative precipitation technique, having spherical and octahedral shapes, monodispersed size distributions and similar transverse dimensions. From our relaxation data we evaluated the temperature dependencies of a) the irreversible demagnetization susceptibility, b) the fluctuation field (associated to the thermally induced demagnetization occurring during the measuring time range) and c) the activation volume (corresponding to the demagnetization produced by the fluctuation field). We conclude that i) the irreversible susceptibility peaks in both samples at ca. 135 K (Verwey transition temperature) and ii) the monotonically increasing temperature variation of the activation volume shows the same values in both samples for temperatures below ca. 135 K and at 290 K reaches values corresponding to 10 and 30 times the average particle volume of the spherical and octahedral NPs, respectively. Those large increases of the activation volume are compatible with a transition from local to collective of the thermally activated processes.

Synthesizing iron oxide nanoparticles for positive contrast in magnetic resonance imaging is the most promising approach to bring this nanomaterial back to the clinical field. The success of this approach depends on several aspects: the longitudinal relaxivity values, the complexity of the synthetic protocol, and the reproducibility of the synthesis. Here, we show our latest results on this goal. We have studied the effect of Cu doping on the physicochemical, magnetic, and relaxometric properties of iron oxide nanoparticles designed to provide positive contrast in magnetic resonance imaging. We have used a one-step, 10 min synthesis to produce nanoparticles with excellent colloidal stability. We have synthesized three different Cu-doped iron oxide nanoparticles showing modest to very large longitudinal relaxivity values. Finally, we have demonstrated the in vivo use of these kinds of nanoparticles both in angiography and targeted molecular imaging. ©

The assessment of diagenetic overprint on microstructural and geochemical data gained from fossil archives is of fundamental importance for understanding palaeoenvironments. The correct reconstruction of past environmental dynamics is only possible when pristine skeletons are unequivocally distinguished from altered skeletal elements. Our previous studies show (i) that replacement of biogenic carbonate by inorganic calcite occurs via an interface-coupled dissolution–reprecipitation mechanism. (ii) A comprehensive understanding of alteration of the biogenic skeleton is only given when structural changes are assessed on both, the micrometre as well as on the nanometre scale. In the present contribution we investigate experimental hydrothermal alteration of six different modern biogenic carbonate materials to (i) assess their potential for withstanding diagenetic overprint and to (ii) find characteristics for the preservation of their microstructure in the fossil record. Experiments were performed at 175°C with a 100 mM NaCl + 10 mM MgCl2 alteration solution and lasted for up to 35 days. For each type of microstructure we (i) examine the evolution of biogenic carbonate replacement by inorganic calcite, (ii) highlight different stages of inorganic carbonate formation, (iii) explore microstructural changes at different degrees of alteration, and (iv) perform a statistical evaluation of microstructural data to highlight changes in crystallite size between the pristine and the altered skeletons. We find that alteration from biogenic aragonite to inorganic calcite proceeds along pathways where the fluid enters the material. It is fastest in hard tissues with an existing primary porosity and a biopolymer fabric within the skeleton that consists of a network of fibrils. The slowest alteration kinetics occurs when biogenic nacreous aragonite is replaced by inorganic calcite, irrespective of the mode of assembly of nacre tablets. For all investigated biogenic carbonates we dist

We present results of bioaragonite to apatite conversion in bivalve, coral and cuttlebone skeletons, biological hard materials distinguished by specific microstructures, skeletal densities, original porosities and biopolymer contents. The most profound conversion occurs in the cuttlebone of the cephalopod Sepia officinalis, the least effect is observed for the nacreous shell portion of the bivalve Hyriopsis cumingii. The shell of the bivalve Arctica islandica consists of cross-lamellar aragonite, is dense at its innermost and porous at the seaward pointing shell layers. Increased porosity facilitates infiltration of the reaction fluid and renders large surface areas for the dissolution of aragonite and conversion to apatite. Skeletal microstructures of the coral Porites sp. and prismatic H. cumingii allow considerable conversion to apatite. Even though the surface area in Porites sp. is significantly larger in comparison to that of prismatic H. cumingii, the coral skeleton consists of clusters of dense, acicular aragonite. Conversion in the latter is sluggish at first as most apatite precipitates only onto its surface area. However, the process is accelerated when, in addition, fluids enter the hard tissue at centers of calcification. The prismatic shell portion of H. cumingii is readily transformed to apatite as we find here an increased porosity between prisms as well as within the membranes encasing the prisms. In conclusion, we observe distinct differences in bioaragonite to apatite conversion rates and kinetics depending on the feasibility of the reaction fluid to access aragonite crystallites. The latter is dependent on the content of biopolymers within the hard tissue, their feasibility to be decomposed, the extent of newly formed mineral surface area and the specific biogenic ultra- and microstructures.