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Progress in the field of biocompatible SERS nanoparticles has promising prospects for biomedical applications. In this work, we have developed a biocompatible Raman probe by combining anisotropic silver nanoparticles with the dye rhodamine 6G followed by subsequent coating with bovine serum albumin. This nanosystem presents strong SERS capabilities in the near infrared (NIR) with a very high (2.7 × 107) analytical enhancement factor. Theoretical calculations reveal the effects of the electromagnetic and chemical mechanisms in the observed SERS effect for this nanosystem. Finite element method (FEM) calculations showed a considerable near field enhancement in NIR. Using density functional quantum chemical calculations, the chemical enhancement mechanism of rhodamine 6G by interaction with the nanoparticles was probed, allowing us to calculate spectra that closely reproduce the experimental results. The nanosystem was tested in cell culture experiments, showing cell internalization and also proving to be completely biocompatible, as no cell death was observed. Using a NIR laser, SERS signals could be detected even from inside cells, proving the applicability of this nanosystem as a biocompatible SERS probe.

In this work a cheap detector of organic molecules is developed. It comprises a cellulose fiber doped with a mixture of magnetite and reduced silver nanoparticles, the latter ones synthesized anew. The nanoparticles and the fiber were characterized with well-established spectroscopic, microscopic and magnetic techniques, namely infrared, UV-vis spectroscopies, vibrating sample magnetometry and electronic microscopies. The so-obtained doped-fibers were tested as surface enhanced Raman spectroscopy detector in aqueous samples with a diluted mixture of two pollutant models (rhodamine 6G and picric acid), being able to differentiate between both organic compounds. Hence, the nanoparticle-impregnated fiber is proposed as a reliable preliminary qualitative and semiquantitative test of the presence of specific organic molecules in solutions. Moreover, the magnetite nanoparticles provide the detector with a saturation magnetization value that enables the separation of the fiber from the solution with the aid of a commercial magnet.

Tannic acid (TA) has multiple effects against cancer, being especially promising in those types that overexpress the epidermal growth factor receptor (EGFR), as TA modulates its activation and downstream signaling pathways, triggering apoptosis. Nonetheless, despite the important role of this receptor in the pathogenesis and progression of a wide variety of tumors, no TA systems targeted to this receptor have been described yet. In this work, we synthesize, characterize by physico-chemical techniques and study the cytotoxic effect and cell uptake of TA nanoparticles targeted to EGFR in both tumoral and normal human skin cells. Our nanoparticles exhibited an extremely high entrapment efficiency, and were only toxic for the tumoral cells. The uptake assay demonstrated that nanoparticles are able to enter the cells through a receptor-mediated mechanism. Furthermore, we have included fluorescent markers in these nanoparticles to combine imaging and therapeutic applications, thus building effectively a multifunctional tool for biomedicine.

La presente invención se refiere a un procedimiento post-síntesis de modificación de la superficie de nanopartículas superparamagnéticas de óxidos de hierro con grupos hidroxilo, sin espaciador, a las nanopartículas de superficie modificada obtenidas por el mismo, así como el uso de estas nanopartículas de superficie modificada en terapia, diagnóstico y en técnicas de concentración y separación de muestras químicas y biológicas.

Camptothecin (CPT; (S)-(+)-4-ethyl-4-hydroxy-1H-pyrano[3',4':6,7]indolizino[1,2-b]quinoline-3,14-(4H,12H)-dione) is a highly cytotoxic natural alkaloid that has not yet found use as chemotherapeutic agent due to its poor water-solubility and chemical instability and, as a consequence, no effective administration means have been designed. In this work, camptothecin has been successfully loaded into iron oxide superparamagnetic nanoparticles with an average size of 14 nm. It was found that surface modification of the nanoparticles by polyethylene glycol enables loading a large amount of camptothecin. While the unloaded nanoparticles do not induce apoptosis in the H460 lung cancer cell line, the camptothecin-loaded nanoparticle formulations exhibit remarkable proapoptotic activity. These results indicate that camptothecin retains its biological activity after loading onto the magnetic nanoparticles. The proposed materials represent novel materials based on naturally occurring bioactive molecules loaded onto nanoparticles to be used as chemotherapeutic formulations. The procedure seems apt to be extended to other active molecules extracted from natural products. In addition, these materials offer the potential of being further implemented for combined imaging and therapeutics, as magnetic nanoparticles are known to be multifunctional tools for biomedicine.

Esta invención se refiere a un procedimiento de obtención de nanoestructuras que comprende las siguientes etapas: a) preparación de una suspensión que comprende al menos un óxido metálico semiconductor, carbón activo y al menos un compuesto de un metal noble; y b) adición de un agente reductor a la suspensión obtenida en la etapa (a).

Apoptotic microtubule network (AMN) is organized during apoptosis, forming a cortical structure beneath the plasma membrane which plays a critical role in preserving cell morphology and plasma membrane integrity. The aim of this study was to examine the effect of cold/warming exposure on apoptotic microtubules and plasma membrane integrity during the execution phase of apoptosis. We demonstrated in camptothecin-induced apoptotic H460 cells that cold/warming exposure disorganized apoptotic microtubules and allowed the access of active caspases to the cellular cortex and the cleavage of essential proteins in the preservation of plasma membrane permeability. Cleavage of cellular cortex and plasma membrane proteins, such as ¿-spectrin, paxilin, focal adhesion kinase and calcium ATPase pump (PMCA-4) involved in cell calcium extrusion resulted in increased plasma permeability and calcium overload leading apoptotic cells to secondary necrosis. The essential role of caspase-mediated cleavage in this process was demonstrated because the addition of the pan-caspase inhibitor z-VAD during cold/warming exposure that induces AMN depolymerization avoided the cleavage of cortical and plasma membrane proteins and prevented apoptotic cells to undergo secondary necrosis. Likewise, apoptotic microtubules stabilization by taxol during cold/warming exposure also prevented cellular cortex and plasma membrane protein cleavage and secondary necrosis. Furthermore, microtubules stabilization or caspase inhibition during cold/warming exposure was also critical for proper phosphatidylserine externalization and apoptotic cell clearance by macrophages. These results indicate that cold/warming exposure of apoptotic cells induces secondary necrosis which can be prevented by both, microtubule stabilization or caspase inhibition.

Apoptosis is characterized by degradation of cell components but plasma membrane remains intact. Apoptotic microtubule network (AMN) is organized during apoptosis forming a cortical structure beneath plasma membrane that maintains plasma membrane integrity. Apoptotic cells are also characterized by high reactive oxygen species (ROS) production that can be potentially harmful for the cell. The aim of this study was to develop a method that allows stabilizing apoptotic cells for diagnostic and therapeutic applications. By using a cocktail composed of taxol (a microtubule stabilizer), Zn2+ (a caspase inhibitor) and coenzyme Q10 (a lipid antioxidant), we were able to stabilize H460 apoptotic cells in cell cultures for at least 72 h, preventing secondary necrosis. Stabilized apoptotic cells maintain many apoptotic cell characteristics such as the presence of apoptotic microtubules, plasma membrane integrity, low intracellular calcium levels and mitochondrial polarization. Apoptotic cell stabilization may open new avenues in apoptosis detection and therapy.

Procedimiento de obtención de nanopartículas metálicas funcionalizadas con moléculas orgánicas fluorescentes. La presente invención se refiere a un procedimiento de obtención de nanopartículas metálicas funcionalizadas con moléculas orgánicas fluorescentes mediante el tratamiento de una o más sales metálicas con un agente reductor en presencia de una molécula orgánica fluorescente. La invención se refiere también a las nanopartículas metálicas obtenibles por dicho procedimiento y a su uso para la detección de moléculas y analitos.

[EN] The present invention relates to a method for producing metallic nanoparticles functionalized with fluorescent organic molecuIes by means of the treatment of one or more metal salts with a reducing agent in the presence of a fluorescent organic molecule. The invention likewise relates to the metallic nanoparticles that can be obtained by means of said method and to the use thereof for detecting molecuIes and analytes., [ES] La presente invención se refiere a un procedimiento de obtención de nanopartículas metálicas funcionalizadas con moléculas orgánicas fluorescentes mediante el tratamiento de una o más sales metálicas con un agente reductor en presencia de una molécula orgánica fluorescente. La invención se refiere también a las nanopartículas metálicas obtenibles por dicho procedimiento y a su uso para la detección de moléculas yanalitos.

Guía didáctica del alumno curso 2011-2012.

Nanopartículas metálicas funcionalizadas con el neuropéptido VIP y procedimiento de preparación. Constituye el objeto de la presente invención nanopartículas metálicas funcionalizadas con el neuropéptido VIP, así como el procedimiento de preparación de dichas nanopartículas. Las nanopartículas objeto de la presente invención presentan uniones selectivas de nanopartículas a péptidos en dos orientaciones posibles, grupo NH2 o grupo COOH. En esta última orientación, los péptidos si son reconocidos por los receptores de membranas celulares, lo que proporciona una herramienta que permite discernir efectos dependientes e independientes de receptor. El péptido empleado ha sido el VIP, mediante el cual se obtiene un amplio espectro de funciones biológicas, incluida inmunomodulación, actuando predominantemente como un potente anti-inflamatorio y un agente inhibidor de la respuesta del Th1 en el sistema inmunitario y emergiendo como un importante factor terapéutico para el tratamiento de enfermedades con componentes inflamatorias y autoinmunes.

The invention relates to metal nanoparticles functionalised with neuropeptide VIP as well as to the method for preparing the nanoparticles. Said nanopartcles have selective bonds to peptides in two possible orientations, group NH2 or group COOH. In the latter orientation, the peptides are recognised by the cell membrane receptors, providing a tool that can be used to distinguish dependent and independent receptor effects. Peptide VIP has been used, enabling a wide range of biological functions, including immunomodulation, to be obtained, acting predominantly as a powerful anti-inflammatory and an agent inhibiting the Th1 response in the immune system and emerging as an important therapeutic factor for the treatment of diseases with inflammatory and autoimmune components., La invención relaciona a las nanopartículas del metal funcionalizadas con el neuropeptide VIP así como al método para preparar las nanopartículas. Los nanopartcles dichos tienen enlaces selectivos a los péptidos en dos posibles orientaciones, grupos NH2 o grupos COOH. En la orientación más última, el son de los péptidos reconocido por los receptores de la membrana celular, proporcionando una herramienta que el bote sea utilizado distinguir efectos dependientes y independientes del receptor. El péptido VIP ha sido utilizado, posibilitando una amplia gama de funciones biológicas, incluyendo el immunomodulation, para ser obtenido, actuando predominante como un antiinflamatorio potente y agente que inhibían la respuesta Th1 en el sistema inmune y que emergían como factor terapéutico importante para el tratamiento de enfermedades con los componentes inflamatorios y autoinmunes.

We characterized a method to conjugate functional silver nanoparticles with vasoactive intestinal peptide (VIP), which could be used as a working model for further tailor-made applications based on VIP surface functionality. Despite sustained interest in the therapeutic applications of VIP, and the fact that its drugability could be largely improved by the attachament to functionalized metal nanoparticles, no methods have been described so far to obtain them. Materials & methods: VIP was conjugated to tiopronin-capped silver nanoparticles of a narrow size distribution, by means of proper linkers, to obtain VIP functionalized silver nanoparticles with two different VIP orientations (Ag-tiopronin-PEG-succinic-[His]VIP and Ag-tiopronin-PEG-VIP[His]). VIP intermediate nanoparticles were characterized by transmission-electron microscopy and Fourier transform infrared spectroscopy. VIP functionalized silver nanoparticles cytotoxicity was determined by lactate dehydrogenase release from mixed glial cultures prepared from cerebral cortices of 1-3 days-old C57/Bl mice. Cells were used for lipopolysaccharide stimulation at day 18-22 of culture. Results: Two different types of VIP-functionalized silver nanoparticles were obtained; both expose the C-terminal part of the neuropeptide, but in the first type VIP is attached to silver nanoparticle through its free amine terminus (Ag-tiopronin-PEG-succinic-[His]VIP), while in the second type, VIP N-terminus remains free (Ag-tiopronin-PEG-VIP[His]). VIP-functionalized silver nanoparticles did not compromise cellular viability and inhibited microglia-induced stimulation under inflammatory conditions. Conclusion: The chemical synthesis procedure developed to obtain VIP-functionalized silver nanoparticles rendered functional products, in terms of biological activity. The two alternative orientations designed, reduced the constraints for chemical synthesis that depends on the nanosurface to be functionalized. Our study provides, for the

The molecular dynamics of hydantoin, 5,5-dimethylhydantoin, 5,5-diphenylhydantoin, barbituric acid and 5-ethyl-5-phenylbarbituric acid in solution are determined to investigate the molecular basis for the activity of these neuroactive drugs. From dipolar 13C and 15N spin-lattice relaxation times the correlation times for the rotational dynamics of specific bond vectors are evaluated. Conclusions can be drawn concerning the anisotropy of the overall molecular rotational motion and the internal rotations of molecular segments. The translational molecular diffusion coefficients are obtained from self-diffusion measurements by the pulsed gradients method. The observed differences in the dynamics as well as structural differences may account for the variation in the neuroactivity of the studied compounds.