Selective binding of albumin to Gm1 ganglioside micelles containing paclitaxel

In our previous work we showed that taxanes (paclitaxel (Ptx) and docetaxel(Dtx)) can be spontaneously loaded into monosialoganglioside (GM1) nanomicelles, increasing their water solubility about 6,000 times, to render stable water soluble formulations that could be used as a novel strategy to deliver drugs in cancer. Here, we describe the hydrophobic interaction of Human Serum Albumin (HSA) with GM1 micelles loaded with Ptx, as a strategy that could improve tumour drug accumulation. This interaction is regulated by conditions such pH and temperature and generates ternary complexes GM1/Ptx/HSA with sizes around 19 to 24 nm and hydrodynamic radius equivalent to a globular protein of 140-180 kDa. These mixed micelles were stable in solution for at least 40 days and also upon freeze-thawing or lyophilization-solubilization cycles. The results of in vitro assays showed that the nano-structures developed are taken up by cell cultures with an antimitotic activity of Ptx on tumoral and nontumoral cell lines that was similar to that observed with the free drug in DMSO solution.Fil: Leonhard, Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia En Productos y Procesos de Córdoba. Laboratorio de Biotecnología; ArgentinaFil: Alasino, Roxana Valeria. Centro de Excelencia En Productos y Procesos de Cordoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Bianco, Ismael Dario. Universidad Nacional de La Rioja; Argentina. Centro de Excelencia En Productos y Procesos de Cordoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Beltramo, Dante Miguel. Centro de Excelencia En Productos y Procesos de Cordoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Católica de Córdoba; Argentin

Antibodies can be spontaneously loaded onto monosialoganglioside micelles containing oncological drugs

Recently, we demonstrated that GM1 micelles transport paclitaxel and doxorubicin with high efficiency. Whenthis GM1-drugs complex is incubated with whole serum, albumin was the only one protein that binds spontaneouslyto form GM1-drug-albumin complex. Here, we show that, under specific physicochemical conditions, these micellesinteract with antibodies forming GM1-IgG complexes. The load of IgG in GM1 reaches a maximum at ratios of 1/4(w / w) incubating to 4.5 and preheating the micelles of GM1 at 55-60°C. The IgG of the GM1-IgG complex obtainedunder these experimental conditions retains the biological activity against the soluble and cellular antigens and is notdisplaced from the micelles in the presence of albumin, the main competitive binding protein.Treatment of GM1-IgG with pepsin, does not show the breakage of the IgG like control of free IgG, suggesting thatIgG is deeply bound into GM1, probably via Fc. Moreover, the presence of 1 M NaCl does not prevent neither dissociatethe complex, suggesting the hydrophobic nature of the interaction. The DLS and TEM results shows that GM1-IgGcomplexes have sizes significantly higher than those of GM1 micelles; this is directly related to the amount of IgGloaded. On the other hand GM1-IgG complex also retain the ability to encapsulate oncological drugs, but, an adequatesequence must be followed during the preparation, in order to obtain efficient GM1-drug-IgG ternary complexes. Moreover, the presence of IgG into GM1-oncological drugs complex do not affect the release or the citotoxic activity ofthe encapsulated molecules such as Ptx or Doxo.Fil: Garro, Ariel Gustavo. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Alasino, Roxana Valeria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Leonhanrd, Victoria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Heredia, Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Beltramo, Dante Miguel. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

LDL-conjugated to GM1 micelles incorporating anticancer drugs to improve tumor cell uptake

Objective(s): The role of lipoproteins (LDL) as active molecules with preferential tumor interaction, but limited drug delivery capacity, has been previously reported. On the other hand, in a previous report, we demonstrated the high capacity of monosialogangliosides (GM1) micelles as drug transporters. Materials and Methods: In this work, GM1 was loaded with high doses of oncologic drugs such Paclitaxel or Doxorubicin and binded to LDL lipoproteins to form GM1-drug-LDLwater soluble complex. Evidence suggests that both, hydrophobic and electrostatic forces, participate in the interaction, regulated by conditions such as pH, temperature and ionic strength. Results: Results of DLS and TEM show that GM1-LDL complexes are considerably larger than the sum of their individual compounds, with a high charge of electronegative surface (-55.9 mV). In addition, the cytotoxic effect on cell cultures is greater when drugs are contained in GM1-LDL complexes than when loaded in GM1 micelles. Conclusion: The results suggest the participation of active energy-dependent mechanism in the uptake of GM1-LDL drug, probably linked to the LDL receptor by the tumor cells. However, we could not confirm that the transport through LDL receptors is the only one that participates in the cellular uptake of the micelles.Fil: Garro, Ariel Gustavo. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Alasino, Roxana Valeria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Leonhard, Victoria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Heredia, Valeria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Beltramo, Dante Miguel. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Universidad Católica de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentin

Polyamines in the Surface of Lipid Micelles Improve the Cellular Uptake of Antitumoral Agents

Purpose: The ability of spermidine to increase the selectivity of anticancer agents has been studied extensively. In this research we report the combination of this polyamine and GM1 ganglioside micelles and characterize their behavior for drug delivery.Methods: Dynamic light scattering and electron microscopy were used to characterize size and morphology of micelles. Zeta potential was determined using a Nano-zeta potential analyzer. Sizeexclusion chromatography was used to separate different populations. Cytotoxic effect of micelles was evaluated on Hep2 cell line.Results: Covalent conjugation of spermidine to gangliosides produces a clear reduction of the electronegative z potential of micelle surface. DLS analysis shows no significant differences between both micelles, while TEM image shows a smaller size of Spermidine-GM1.These modified micelles load hydrophilic or hydrophobic antitumor drugs and conjugation does not affect the stability of micelles/drug in solution. Spermidine-GM1/Doxo micelles show faster drug release into cells as compared with GM1/Doxo micelles; however, no evidence can be found for the participation of the polyamine transport system in the uptake of modified micelles.Conclusion: While Spermidine-GM1 and GM1 micelles show similar physical properties, spermidine modified micelles are most efficient to release drugs, making this an interesting alternative to consider for drug delivery.Fil: Garro, Ariel Gustavo. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Alasino, Roxana Valeria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Leonhard, Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Heredia, Valeria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Beltramo, Dante Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentin

An in silico analysis of Ibuprofen enantiomers in high concentrations of sodium chloride with SARS-CoV-2 main protease

2020 will be remembered worldwide for the outbreak of Coronavirus disease (COVID-19), which quickly spread until it was declared as a global pandemic. The main protease (Mpro) of SARS-CoV-2, a key enzyme in coronavirus, represents an attractive pharmacological target for inhibition of SARS-CoV-2 replication. Here, we evaluated whether the anti-inflammatory drug Ibuprofen, may act as a potential SARS-CoV-2 Mpro inhibitor, using an in silico study. From molecular dynamics (MD) simulations, we also evaluated the influence of ionic strength on the affinity and stability of the Ibuprofen–Mpro complexes. The docking analysis shows that R(−)Ibuprofen and S(+)Ibuprofen isomers can interact with multiple key residues of the main protease, through hydrophobic interactions and hydrogen bonds, with favourable binding energies (−6.2 and −5.7 kcal/mol, respectively). MM-GBSA and MM-PBSA calculations confirm the affinity of these complexes, in terms of binding energies. It also demonstrates that the ionic strength modifies significantly their binding affinities. Different structural parameters calculated from the MD simulations (120 ns) reveal that these complexes are conformational stable in the different conditions analysed. In this context, the results suggest that the condition 2 (0.25 NaCl) bind more tightly the Ibuprofen to Mpro than the others conditions. From the frustration analysis, we could characterize two important regions (Cys44-Pro52 and Linker loop) of this protein involved in the interaction with Ibuprofen. In conclusion, our findings allow us to propose that racemic mixtures of the Ibuprofen enantiomers might be a potential treatment option against SARS-CoV-2 Mpro. However, further research is necessary to determinate their possible medicinal use. Communicated by Ramaswamy H. Sarma.Fil: Clemente, Camila Mara. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentina. Universidad Nacional de Villa María. Instituto Académico Pedagógico de Ciencias Básicas y Aplicadas; ArgentinaFil: Freiberger, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Ravetti, Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones y Transferencia de Villa María. Universidad Nacional de Villa María. Centro de Investigaciones y Transferencia de Villa María; Argentina. Universidad Nacional de Villa María. Instituto Académico de Ciencias Humanas; ArgentinaFil: Beltramo, Dante Miguel. Universidad Católica de Córdoba; Argentina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Garro, Ariel Gustavo. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Universidad Nacional de Villa María. Instituto Académico de Ciencias Humanas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones y Transferencia de Villa María. Universidad Nacional de Villa María. Centro de Investigaciones y Transferencia de Villa María; Argentin

Development of a method to control the RNA extraction and reverse transcription steps for the detection of dengue virus present in human blood samples

AIMS: Molecular biology techniques based on the detection of genomic sequences by reverse transcription combined with polymerase chain reaction (PCR) have enabled the detection of different RNA viruses in serum or plasma samples. Since the dengue epidemic outbreak declared in Argentina in 2009, numerous patients´ samples were analyzed for the acute phase of infection. One of the main methodological drawbacks is the lack of internal control to measure the effectiveness of the viral extraction and reverse transcription process. In this article, we propose to standardize a molecular method to detect beta actin (â-Act) and glucose 6 phosphate dehydrogenase (G6PDH) complementary DNAs (cDNAs) present in patient´s plasma/serum, as a control process. RESULTS: RNA extraction, reverse transcription, and PCRs for human G6PDH, â-Act, and the dengue virus genome were performed. cDNA fragments for â-Act and G6PDH were amplified for all samples, regardless of the presence or absence of viral RNA. CONCLUSIONS: Amplification of â-Act and G6PDH cDNAs can be used as a control for the extraction and reverse transcription processes during dengue virus detection. This could also be a useful method for controlling the above steps when infections caused by other RNA viruses are studied, even if another methodology is employed, such as real-time PCR.Fil: Galván, Cristian Ariel. Universidad Católica de Córdoba. Facultad de Ciencias Químicas; ArgentinaFil: Elbarcha, Osvaldo C.. No especifíca;Fil: Fernández, Eduardo J.. No especifíca;Fil: Beltramo, Dante Miguel. Universidad Católica de Córdoba. Facultad de Ciencias Químicas; Argentina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Soria, Néstor Walter. Universidad Católica de Córdoba. Facultad de Ciencias Químicas; Argentin

Self-assembled micelles of monosialogangliosides as nanodelivery vehicles for taxanes

We demonstrate herein that taxanes (paclitaxel (Ptx) and docetaxel (Dtx)) can be spontaneously loaded into ganglioside nanomicelles. The efficiency of gangliosides to solubilize taxanes was highly dependent on their self-aggregating structure. Thus, GM3 that forms unilamellar vesicles was less efficient to solubilize taxanes than gangliosides that form micelles (i.e. GM1 and GM2). Sialic acid cyclization of GM1 by acid treatment led to an important reduction in its capacity to solubilize taxanes, as also did the replacement of the fatty acid of ceramide by a dicholoracetyl group. Water solubility of paclitaxel (Ptx) is less than 1 μg mL- 1 and increased up to 6.3 mg.mL- 1 upon its association with GM1 micelles. The incorporation of Ptx in GM1 reached an optimum at GM1/Ptx 20/1 molar ratio when performed at room temperature. An increase in the solubilization capacity of GM1 micelles was observed upon dehydration of their polar head group by pre-treatment at 55 °C. Loading of Ptx into the micelle induced a structural reorganization that led to an important protection of Ptx reducing its hydrolysis at alkaline pH. Diffusion of either GM1 or Ptx was restricted upon mixed-micelle formation indicating that they are kinetically more stable than pure ganglioside micelles. X-ray powder diffraction of lyophilized GM1 micelles with Ptx showed a change in their internal structure from a crystalline state to completely amorphous. Taxane-ganglioside mixed micelles were stable in solution for at least 4 months and also upon freeze-thawing or lyophilization-solubilization cycles. Upon mixing with human blood constituents, GM1/Ptx micelles did not induce hemolysis or platelet aggregation and were spontaneously covered with human serum albumin (HSA), which could aid in the delivery of micellar content to tumors. In vitro antimitotic activity of GM1/Ptx mixed micelles was qualitatively equivalent to that of free drug in DMSO solution. © 2012 Elsevier B.V. All rights reserved.Fil: Leonhard, Victoria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Alasino, Roxana Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Bianco, Ismael Dario. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Universidad Nacional de La Rioja; ArgentinaFil: Garro, Ariel G.. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Heredia, Valeria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Beltramo, Dante Miguel. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Católica de Córdoba. Facultad de Ciencias Químicas; Argentin

Sistemas nanoparticulados de vehiculización de fármacos: transportador vs fármaco

La eficacia terapéutica de un fármaco está condicionada por muchos factores, dentro de los cuales la solubilidad y el acceso al sitio de acción juegan un rol primordial. Se estima que aproximadamente el 40% de los medicamentos que se encuentran actualmente en el mercado y cerca del90% de las moléculas en desarrollo presentan baja solubilidad en agua. Así, las propiedades de solubilidad de los compuestos continúa siendo uno de los mayores obstáculos en el desarrollo de sistemas de administración de fármacos.Fil: Alasino, Roxana Valeria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Leonhard, Victoria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Garro, Ariel Gustavo. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Beltramo, Dante Miguel. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentin

Identification and Expression of Some Plant Cell Wall-Degrading Enzymes Present in Three Ontogenetics Stages of Thecaphora frezii, a Peanut (Arachis hypogaea L.) Pathogenic Fungus

Peanuts can be affected by the presence of pathogenic microorganisms. The fungus Thecaphora frezii (T. frezii), which belongs to the taxonomic class Ustilaginomycetes, is the causal agent of the disease known as “peanut smut”. The life cycle of this fungus includes three stages, namely teliospores, basidiospores and hyphae. In the hyphae stage, infection occurs in the peanut plant, which requires the involvement of some enzymes secreted by the fungus. These include the Plant Cell Wall-Degrading Enzymes (PCWDEs), which degrade various polysaccharides. This study aimed to identify the presence of transcript for enzymes belonging to the PCWDEs from three stages of T. frezii. For this, total RNA was extracted from the three ontogenetic stages of T. frezii. These samples were analyzed using an RNA-Seq approach and some transcripts were quantified using Real Time PCR. The analysis of the data provided by the RNA-Seq of the three T. frezii stages, it was possible to identify some transcripts that could encode enzymes compatible with polysaccharides degradation that are part of the plant cell wall. In T. frezii transcriptome, 40 deduced proteins would be enzymes with functions of PCWDEs were identified. They were divided into 27 glycoside hydrolases; two polysaccharide lyases; three carbohydrate esterases and eight enzymes with auxiliary activities. In addition, the fungal SNF1 gene was identified whose activity could be affected by high glucose level, and indirectly influence the levels of some PCWDEs. The analysis of the PCWDEs could help to understand part of the fungal infection process and possibly find substances that can control its development.Fil: Soria, Néstor Walter. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentina. Area de Ciencias Agrarias, Ingeniería, Ciencias Biológicas y de la Salud de la Universidad Católica de Córdoba; ArgentinaFil: Figueroa, Ana Cristina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Díaz, María Soledad. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Alasino, Roxana Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Yang, Pablo. Area de Ciencias Agrarias, Ingeniería, Ciencias Biológicas y de la Salud de la Universidad Católica de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Beltramo, Dante Miguel. Area de Ciencias Agrarias, Ingeniería, Ciencias Biológicas y de la Salud de la Universidad Católica de Córdoba; Argentina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentin

Monosialoganglioside GM1 reduces toxicity of Ptx and increase anti-metastasic effect in a murine mammary cancer model

Having demonstrated the ability of monosialoganglioside GM1 micelles as oncology drug transporter, this work focuses on evaluating its application in an in vivo system, studying the toxicity and antitumoral effect of GM1-Ptx micellar formulation. The maximum tolerated dose (MTD) obtained after intravenous administration of GM1-Ptx in mice was 55 mg/kg and the 50% lethal dose (LD50) was 70 mg/kg. This value is higher than those described for the commercial formulations TAXOL and ABRAXANE, with LD50 of 30 and 45 mg/kg respectively. The antitumor activity, mortality and incidence of metastasis were studied on a murine model of mammary gland cancer. The GM1-Ptx formulation was administered i.v. at different doses for 9 weeks using empty GM1 micelles and saline as treatment controls. Once the treatments were completed, biochemical markers were quantified and histological tissue tests were performed. The most promising results were obtained with the treatment at a dose of 15 mg/kg/twice a week, condition in which a longer survival and significant reduction in the incidence of animals with metastasis, since only one 25% of the mice showed presence of pulmonary micro metastases.Fil: Leonhard, Victoria. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Alasino, Roxana Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; ArgentinaFil: Pasqualini, María Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Ciencias de la Salud. Universidad Nacional de Córdoba. Instituto de Investigaciones en Ciencias de la Salud; Argentina. Universidad Nacional de Córdoba; ArgentinaFil: Cremonezzi, David César. Universidad Nacional de Córdoba; ArgentinaFil: Garcia, Nestor Horacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Ciencias de la Salud. Universidad Nacional de Córdoba. Instituto de Investigaciones en Ciencias de la Salud; ArgentinaFil: Beltramo, Dante Miguel. Provincia de Córdoba. Ministerio de Ciencia y Técnica. Centro de Excelencia en Productos y Procesos de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin