Nuevos estudios sobre la mejora del aceite de oliva y el aprovechamiento de residuos del olivar

La finalidad de la investigación desarrollada fue aportar nuevos conocimientos sobre el cultivo del olivo, sobre su producto estrella, el aceite de oliva, y sobre los desechos generados en su producción, alperujo y hojas de olivo. De esta forma se cierra el ciclo formado por el cultivo, la producción de aceite y la gestión de residuos generados. Para ello, los objetivos que motivaron la realización de la tesis fueron varios, todos con el denominador común del binomio olivo/aceite de oliva y la metabolómica. (1) Conocer en profundidad los metabolitos secundarios que componen la fracción minoritaria del aceite de oliva virgen (VOO) y los métodos existentes en la bibliografía para determinarlos. (2) Desarrollar estrategias de análisis global y orientado de fracciones de interés del VOO para su implementación en programas de mejora de olivo destinados a la mejora del producto final, el VOO. Para ello, se ha utilizado equipamiento analítico de última generación basado en espectrometría de masas para identificar/cuantificar los metabolitos de interés. (3) Evaluar la aplicación de las estrategias de análisis metabolómico desarrolladas en el programa de mejora de olivo mediante el análisis de dos fracciones de gran interés en el VOO: los compuestos fenólicos y la fracción de ácidos grasos. (4) Estudiar el efecto en términos de calidad y estabilidad del enriquecimiento de aceites con extractos fenólicos procedentes de dos residuos del olivo: alperujo y hojas de olivo

Exploring the Mysteries of <em>Cannabis</em> through Gas Chromatography

In the last decades, cannabinoids, the active constituents of Cannabis sativa L., have been attracting a strong interest, regarding the health effects associated with the use of Cannabis and Cannabis-derived products. The progressive legalization of this species in several countries has prompted an increasing concern about the characterization and quantification of cannabinoids in diverse chemotypes of the plant, as well as the obtained final products. Therewith, Process and Product Quality Assurance (PPQA) becomes a mandatory practise to verify the Good Manufacturing Practices (GMP). Gas chromatography is one of the most used techniques in this sense due to its high attainable resolution. However, sample complexity and the thermal lability of cannabinoids hinder the analysis. In this chapter, a fully description of the recent advances in the Cannabis sativa L. analysis by gas chromatography will be presented, including different approaches that have come up to solve the obstacles encountered

Impact of different formulations of pharmaceutical cannabis-based extracts on the neuroprotective effect in cerebellar granule cell cultures

Preclinical research supports the benefits of pharmaceutical cannabis-based extracts for treating different medical conditions (e. g., epilepsy); however, their neuroprotective potential has not been widely investigated. In addition, there is still controversy about the impact of other factors in the beneficial effect of these extracts (e.g., the entourage effect, and oil formulations). We evaluated the neuroprotective activity of Epifractan (EPI), a cannabis-based medicinal extract containing a high level of cannabidiol (CBD), components like terpenoids and flavonoids, and trace levels of Δ9-tetrahydrocannabinol and the acid form of CBD. Using primary cultures of cerebellar granule cells, we determined the ability of EPI to counteract the rotenone-induced neurotoxicity by analyzing cell viability and morphology of neurons and astrocytes by immunocytochemical assays. The effect of EPI was compared with XALEX, a plant-derived and highly purified CBD formulation (XAL), and pure CBD crystals (CBD). The results revealed that EPI induced a significant reduction in the rotenone-induced neurotoxicity in a wide range of concentrations without causing neurotoxicity per se. EPI showed a similar effect to XAL suggesting that no additive or synergistic interactions (i.e., entourage effect) between individual substances present in EPI occurred. In contrast, CBD crystals did show a different profile to EPI and XAL since a neurotoxic effect per se was observed at the higher concentrations assayed. Medium-chain triglyceride oil used in EPI formulation could explain this difference. Our data support a neuroprotective effect of EPI which may provide neuroprotection in different neurodegenerative processes. The results highlight the role of CBD as the active component of EPI but also support the need for an appropriate formulation to dilute pharmaceutical cannabis-based productAgencia Nacional de Investigación e Innovació

Quantitative proteomic analysis of Pseudomonas pseudoalcaligenes CECT5344 in response to industrial cyanide-contain ing wastewaters using Liquid Chromatography- Mass Spectrometry/Mass Spectrometry (LC- MS/MS)

Biological treatments to degrade cyanide are a powerful technology for cyanide removal from industrial wastewaters. It has been previously demonstrated that the alkaliphilic bacterium Pseudomonas pseudoalcali genes CECT5344 is able to use free cyanide and several metal − cyanide complexes as the sole nitrogen source. In this work, the strain CECT5344 has been used for detoxification of the different chemical forms of cyanide that are present in alkaline wastewaters from the jewelry industry. This liquid residue also contains large concentration s of metals like iron, copper and zinc, making this wastewater even more toxic. To elucidate the molecular mechanisms involved in the bioremediation process, a quantitative proteomic anal- ysis by LC-MS/MS has been carried out in P . pseudoalcaligene s CECT5344 cells grown with the jewelry residue as sole nitrogen source. Different proteins related to cyanide and cyanate assimilation, as well as other proteins involved in transport and resistance to metals were induced by the cyanide-cont aining jewelry residue. GntR-like regulatory proteins were also induced by this industrial residue and mutational analysis revealed that GntR-like regulatory proteins may play a role in the regulation of cyanide assimilation in P . pseudoalcaligene s CECT5344. The strain CECT5344 has been used in a batch reactor to remove at pH 9 the dif- ferent forms of cyanide present in industrial wastewaters from the jewelry industry (0.3 g/L, ca . 12 mM total cyanide, including both free cyanide and metal − cyanide complexes). This is the first report describing the biological removal at alkaline pH of such as elevated concentra- tion of cyanide present in a heterogeneou s mixture from an industrial source

Regulation of Expression of Cannabinoid CB2 and Serotonin 5HT1A Receptor Complexes by Cannabinoids in Animal Models of Hypoxia and in Oxygen/Glucose-Deprived Neurons

Background: Cannabidiol (CBD) is a phytocannabinoid with potential in one of the most prevalent syndromes occurring at birth, the hypoxia of the neonate. CBD targets a variety of proteins, cannabinoid CB2 and serotonin 5HT1A receptors included. These two receptors may interact to form heteromers (CB2-5HT1A-Hets) that are also a target of CBD. Aims: We aimed to assess whether the expression and function of CB2-5HT1A-Hets is affected by CBD in animal models of hypoxia of the neonate and in glucose- and oxygen-deprived neurons. Methods: We developed a quantitation of signal transduction events in a heterologous system and in glucose/oxygen-deprived neurons. The expression of receptors was assessed by immuno-cyto and -histochemistry and, also, by using the only existing technique to visualize CB2-5HT1A-Hets fixed cultured cells and tissue sections (in situ proximity ligation PLA assay). Results: CBD and cannabigerol, which were used for comparative purposes, affected the structure of the heteromer, but in a qualitatively different way; CBD but not CBG increased the affinity of the CB2 and 5HT1A receptor-receptor interaction. Both cannabinoids regulated the effects of CB2 and 5HT1A receptor agonists. CBD was able to revert the upregulation of heteromers occurring when neurons were deprived of oxygen and glucose. CBD significantly reduced the increased expression of the CB2-5HT1A-Het in glucose/oxygen-deprived neurons. Importantly, in brain sections of a hypoxia/ischemia animal model, administration of CBD led to a significant reduction in the expression of CB2-5HT1A-Hets. Conclusions: Benefits of CBD in the hypoxia of the neonate are mediated by acting on CB2-5HT1A-Hets and by reducing the aberrant expression of the receptor-receptor complex in hypoxic-ischemic conditions. These results reinforce the potential of CBD for the therapy of the hypoxia of the neonate

Cannabigerol action at cannabinoid CB1 and CB2 receptors and at CB1-CB2 heteroreceptor complexes

Cannabigerol (CBG) is one of the major phytocannabinoids present in Cannabis sativa L. that is attracting pharmacological interest because it is non-psychotropic and is abundant in some industrial hemp varieties. The aim of this work was to investigate in parallel the binding properties of CBG to cannabinoid CB1 (CB1R) and CB2 (CB2R) receptors and the effects of the compound on agonist activation of those receptors and of CB1-CB2 heteroreceptor complexes. Using [3H]-CP-55940, CBG competed with low micromolar Ki values the binding to CB1R and CB2R. Homogeneous binding in living cells, which is only possible for the CB2R, provided a nanomolar Ki value. In contrast, CBG competed the binding of [3H]-WIN-55,212-2 to CB2R but not to CB1R (2.7 versus >30 µM). The phytocannabinoid modulated signaling mediated by receptors and receptor heteromers even at low concentrations of 0.1-1 µM. cAMP, pERK, ÿ-arrestin recruitment and label-free assays in HEK-293T cells expressing the receptors and treated with endocannabinoids or selective agonists proved that CBG is a partial agonist of CB2R. The action on cells expressing heteromers was similar to that obtained in cells expressing the CB2R. The effect of CBG on CB1R was measurable but the underlying molecular mechanisms remain uncertain. The results indicate that CBG is indeed effective as regulator of endocannabinoid signaling

Similarities and differences upon binding of naturally occurring Δ9-tetrahydrocannabinol-derivatives to cannabinoid CB1 and CB2 receptors.

We have here assessed, using Δ9 -tetrahydrocannabinol (Δ9 -THC) for comparison, the effect of Δ9 -tetrahydrocannabinolic acid (Δ9 -THCA) and of Δ9 -tetrahydrocannabivarin (Δ9-THCV) that is mediated by human versions of CB1, CB2, and CB1-CB2 receptor functional units, expressed in a heterologous system. Binding to the CB1 and CB2 receptors was addressed in living cells by means of a homogeneous assay. A biphasic competition curve for the binding to the CB2 receptor, was obtained for Δ9 -THCV in cells expressing the two receptors. Signaling studies included cAMP level determination, activation of the mitogen-activated protein kinase pathway and ß-arrestin recruitment were performed. The signaling triggered by Δ9 -THCA and Δ9 -THCV via individual receptors or receptor heteromers disclosed differential bias, i.e. the bias observed using a given phytocannabinoid depended on the receptor (CB1, CB2 or CB1-CB2) and on the compound used as reference to calculate the bias factor (Δ9 -THC, a selective agonist or a non-selective agonist). These results are consistent with different binding modes leading to differential functional selectivity depending on the agonist structure, and the state (monomeric or heteromeric) of the cannabinoid receptor. In addition, on studying Gi-coupling we showed that Δ9 -THCV and Δ9 -THCA and Δ9 -THCV were able to revert the effect of a selective CB2 receptor agonist, but only Δ9-THCV, and not Δ9-THCA, reverted the effect of arachidonyl-2′ -chloroethylamide (ACEA 100 nM) a selective agonist of the CB1 receptor. Overall, these results indicate that cannabinoids may have a variety of binding modes that results in qualitatively different effects depending on the signaling pathway that is engaged upon cannabinoid receptor activatio

Biomarkers of Venous Thromboembolism Recurrence after Discontinuation of Low Molecular Weight Heparin Treatment for Cancer-Associated Thrombosis (HISPALIS-Study)

The most appropriate duration of anticoagulant treatment for cancer-associated venous thromboembolism (CAT) remains unclear. We have conducted a prospective multicenter study in CAT patients with more than 6 months of anticoagulant treatment to predict the risk of venous thromboembolism (VTE) recurrence after anticoagulation discontinuation. Blood samples were obtained when patients stopped the anticoagulation, at 21 days and at 90 days. In each sample we assessed different coagulation-related biomarkers: D-dimer (DD), high-sensitivity C-reactive protein (hs-CRP), P-selectin (PS), phospholipids, soluble tissue factor, factor VIII and the thrombin generation test. It was evaluated 325 CAT patients and 166 patients were included in the study, mean age 64 ± 17 years. VTE recurrence until 6 months after stopping anticoagulation treatment was 9.87% [95% confidence interval (CI): 6–15]. The biomarkers sub-distribution hazard ratios were 6.32 for ratio DD basal/DD 21 days > 2 (95% CI: 1.82–21.90), 6.36 for hs-CRP > 4.5 (95% CI: 1.73–23.40) and 5.58 for PS > 40 (95% CI: 1.46–21.30) after 21 days of stopping anticoagulation. This is the first study that has identified the DD ratio, hs-CRP and PS as potential biomarkers of VTE recurrence in cancer patients after the discontinuation of anticoagulation treatment. A risk-adapted strategy may allow the identification of the optimal time to withdraw the anticoagulation in each CAT patientmThis research was funded by Instituto de Salud Carlos III, grant number (PI15/01085; PI18/01640, PI20/00075), Sociedad Española de Trombosis y Hemostasia; Fundación Respira, grant number (140/2013), Fundación Neumosur, grant number (5/2013) and the LEO Pharma Research Foundation. Partial funding for open access charge: Universidad de Málaga

Innovamos, jugamos y aprendemos combinando conocimientos de diferentes áreas

El objetivo principal de este proyecto de innovación docente es elaborar nuevas herramientas docentes para la enseñanza de la Bioquímica y la Fisiología, así como facilitar su aprendizaje y la relación de los conocimientos adquiridos entre ambas asignaturas en diferentes Grados tanto del Campus de Melilla como del Campus de Granada. Para ello nos planteamos los siguientes objetivos específicos: 1. Identificar una historia para hilar el contenido de la asignatura de Bioquímica. 2. Crear ligas o torneos para retar a los alumnos en su aprendizaje con estos juegos. 3. Relacionar contenidos entre las asignaturas de Bioquímica y Fisiología. 4. Evaluar el material didáctico al final del semestre mediante cuestionarios para comprobar su evolución en el aprendizaje de los conocimientos adquiridos. Por otro lado, también pretendemos que los alumnos identifiquen los conocimientos y competencias adquiridos y la utilidad y aplicación de esas tareas y habilidades en el ámbito empresarial, como un primer paso para el autoconocimiento profesional y la búsqueda de empleo. La dinámica de juego empleada será la realización de un Escape Room Educativo Virtual. Para alcanzar el primer objetivo en clase, identificaremos una historia o juego de interés entre los alumnos de primero. Si existen varias, se elegirá la que sea propuesta por el mayor número de alumnos. Tras la presentación de la historia, crearemos una liga o torneo para retar a los alumnos entre ellos. En el caso de que haya algún alumno/a con necesidades especiales, se nombrar un alumno colaborador, el cual le facilitará su participación durante la realización del juego. Además, el contenido será evaluado de modo que todos los alumnos tengan acceso a él, esto es, aumento del tamaño de letra, grabación de voz en off en caso necesario, aumento en el tiempo de realización de la actividad, etc. Algunas de las preguntas introducidas en el juego permitirán la relación de contenidos entre las asignaturas de Bioquímica y Fisiología y su aplicabilidad en su futuro profesional. Por último, al finalizar la actividad se les pasará a los alumnos un cuestionario para evaluar su satisfacción con este proyecto de innovación docente. Dicho proyecto docente ha sido aplicado al final del semestre en los Grados de Enfermería, Fisioterapia y el Doble Grado en Educación Primaria y Ciencias de la Actividad Física y del Deporte del Campus de Melilla y en los Grado de Fisioterapia y el Grado en Ciencias de la Actividad Física y del Deporte del Campus de Granada en la Universidad de Granada.The main objective of this teaching innovation project is to develop new teaching tools for the teaching of Biochemistry and Physiology, as well as to facilitate their learning and the relationship of the knowledge acquired between both subjects in different Degrees in both Melilla and Granada Campus at the University of Granada. To this end, we set ourselves the following specific objectives: 1. Identify a story to string together the content of the Biochemistry subject. 2. Create leagues or tournaments to resume students in their learning with these games. 3. Relate content between the subjects of Biochemistry and Physiology. 4. Evaluate the didactic material at the end of the semester by means of questionnaires to verify its evolution in the learning of the acquired knowledge. On the other hand, we also intend that students identify the knowledge and skills acquired and the usefulness and application of these tasks and skills in the business field, as a first step for professional self-knowledge and job search. The game dynamics used will be the realization of a Virtual Educational Escape Room. To achieve the first objective in class, we will identify a story or game of interest among the first graders. If there are several, the one proposed by the largest number of students will be chosen. After the presentation of the story, we will create a league or tournament to take the students back to each other. In the event that there is a student with special needs, a collaborating student will be appointed, who will facilitate their participation during the game. In addition, the content will be evaluated so that all students have access to it, that is, increased font size, voiceover recording if necessary, increased time to complete the activity, etc. Some of the questions presented in the game will allow the content relationship between the subjects of Biochemistry and Physiology and its applicability in their professional future. Finally, at the end of the activity, students will be given a questionnaire to assess their satisfaction with this teaching innovation project. This teaching project has been applied at the end of the semester in the Degrees of Nursing, Physiotherapy and the Double Degree in Primary Education and Physical Activity and Sports Sciences of the Melilla Campus and in the Degrees of Physiotherapy and the Degree in Sciences of the Physical Activity and Sports of the Granada Campus at the University of Granada

Cannabigerol Action at Cannabinoid CB1 and CB2 Receptors and at CB1–CB2 Heteroreceptor Complexes

Cannabigerol (CBG) is one of the major phytocannabinoids present in Cannabis sativa L. that is attracting pharmacological interest because it is non-psychotropic and is abundant in some industrial hemp varieties. The aim of this work was to investigate in parallel the binding properties of CBG to cannabinoid CB1 (CB1R) and CB2 (CB2R) receptors and the effects of the compound on agonist activation of those receptors and of CB1–CB2 heteroreceptor complexes. Using [3H]-CP-55940, CBG competed with low micromolar Ki values the binding to CB1R and CB2R. Homogeneous binding in living cells, which is only technically possible for the CB2R, provided a 152 nM Ki value. Also interesting, CBG competed the binding of [3H]-WIN-55,212-2 to CB2R but not to CB1R (Ki: 2.7 versus &gt;30 μM). The phytocannabinoid modulated signaling mediated by receptors and receptor heteromers even at low concentrations of 0.1–1 μM. cAMP, pERK, β-arrestin recruitment and label-free assays in HEK-293T cells expressing the receptors and treated with endocannabinoids or selective agonists proved that CBG is a partial agonist of CB2R. The action on cells expressing heteromers was similar to that obtained in cells expressing the CB2R. The effect of CBG on CB1R was measurable but the underlying molecular mechanisms remain uncertain. The results indicate that CBG is indeed effective as regulator of endocannabinoid signaling