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Stevia rebaudiana leaf extracts contain stevioside and rebaudioside A, two steviol glycosides (SGs) used as natural sweeteners because of their non-toxic, thermally stable and non-caloric properties. Indeed, leaf extracts can be up to 300 times sweeter than sucrose. Stevioside and rebaudioside A have organoleptic differences, the first one having an undesirable bitterness and the second one a higher sweetener capacity. Selection of the S. rebaudiana varieties and the best environmental conditions that elicit higher SGs content and the appropriate composition is an important goal. In this study we quantified and compared the amount of stevioside and rebaudioside A in two of the most used S. rebaudiana cultivars, Morita II and Criolla. Our results show a strong differential ratio of stevioside and rebaudioside A accumulated in the leaf between these cultivars. The Criolla cultivar showed about 3 times more stevioside per mg of dry weight than Morita II, whereas the Morita II accumulated almost 10 times more rebaudioside A than that produced in Criolla. We observed an enhanced expression in Morita II of three genes (SrKA13H, SrUGT74G1 and SrUGT76G1) known to encode three enzymes that participate in SGs biosynthesis, likely contributing to the differences in the stevioside and rebaudioside A accumulation. Not only genetic variation can affect SGs composition, but also environmental factors and crop management. Numerous studies have shown that the light regime in which S. rebaudiana cultivars grow can affect SGs accumulation. However, the optimal light regime to increase total SGs content is currently controversial. By applying various light intensities, we detected an increase of expression of these three biosynthetic genes at higher light intensity, accompanied by higher levels of stevioside and rebaudioside A, demonstrating that light intensity influences the synthesis of SGs.

Aristotelia chilensis (Mol.) Stuntz is an evergreen species endemic to Chile. It grows in open areas or under tree canopy, and its leaves emerge in early spring and summer. The objective of this study was to determine changes in photosynthetic parameters, total phenol content (TPC), antioxidant activity, and anatomy of apical and basal leaves of A. chilensis during the year. Photosynthesis performance was determined by measuring electron transport rate (ETR), the quantum efficiency of photosystem II (Fv/Fm), photochemical quenching (qP), and non-photochemical quenching (NPQ) with a fluorimeter. Leaf extracts were analysed to determine TPC and antioxidant activity. The maximum ETR and qP were recorded in spring and summer when the photosynthetically active radiation (PAR) at midday was higher (1901 and 1968 µmol m-2 s-1, respectively) than in other parts of a year. The Fv/Fm had typical physiological values in both types of leaves (about 0.8 in all the seasons). Also the NPQ was not influenced by the kind of leaves and season of the year. In concordance, the basal spring leaves had the highest TPC values. In contrast, the highest values of antioxidant activity were recorded in basal winter leaves followed by basal spring leaves. The results suggested that an increase in PAR (spring) positively affected the antioxidant activity and TPC, which correlated with higher ETR and qP values. The apical leaves showed morphological adaptations during the year and areas of intercellular spaces and palisade parenchyma were larger than in the basal leaves.

Natural products are an alternative to control microorganisms that cause diseases in crops. This work aimed to evaluate different solvents for obtaining crude extracts from tobacco leaf exudates and to determine in vitro effect of these extracts against two phytopathogenic bacteria: Xanthomonas campestris(Xc) and Pectobacterium carotovorum(Pc). Crude extracts from ten tobacco lines using solvents with polarities between 3.1 and 6.2 (dichloromethane, n-butanol, ethyl acetate, methanol and ethanol 90%) were obtained. Ethanol 90% was selected as the best solvent for obtaining extracts from tobacco leaf exudates and as a substitute of dichloromethane due to the best yield. The chemical composition diversity of the ethanolic extracts was revealed by thin-layer chromatography. The antibacterial activity was evaluated by agar disk diffusion method recording the inhibition zones. Growth inhibition was observed for all extracts against Xc, and the better activity corresponded to the lines Nic 1061"TI 1738" and Nic 1016 "Incekara" until a minimal amount of 5 µg/ disc, with higher yield in case of the line Nic1061 . Only the extract of the line Nic 1015 was able to inhibit the growth of Pc until a minimal inhibitory concentration of 5 µg/disc. These results suggest a potential use of crude extracts from lines Nic 1061 and Nic 1015 "TI 1341" as an effective agent for the crop protection against Xc and Pc respectively.Key words: phytopathogens, Nicotiana tabacum, crop protection, solvents, leaf surface., Título en ingles: In vitro antibacterial effect of tobacco leaf exudates against two bacterial plant pathogensTítulo corto: Efecto antibacteriano in vitro de exudados foliares de tabacoResumen: Los productos naturales son una alternativa para el control de microorganismos que ocasionan enfermedades en los cultivos. Este trabajo tuvo como objetivo evaluar diferentes solventes para la obtención de extractos crudos a partir de exudados foliares de líneas de tabaco, y el efecto in vitro de estos extractos contra dos bacterias fitopatógenas: Xanthomonas campestris (Xc) y Pectobacterium carotovorum (Pc). Se evaluaron solventes con polaridades entre 3.1 y 6.2 (diclorometano, n-butanol, acetato de etilo, metanol y etanol 90 %). El etanol 90 % se seleccionó como mejor solvente y como sustituto del diclorometano por su mayor rendimiento. Los extractos etanólicos crudos se obtuvieron a partir de exudados foliares de diez líneas de tabaco seleccionadas. La diversidad de la composición química de los extractos etanólicos se reveló por cromatografía en capa delgada. La actividad antibacteriana se evaluó por el método de difusión en agar con discos de papel de filtro y la medición del diámetro del halo de inhibición. Se observó inhibición para todos los extractos contra Xc destacándose los correspondientes a las líneas Nic 1061 “TI 1738” y Nic 1016 “Incekara” hasta 5 µg de extracto crudo seco /disco, con un mayor rendimiento para la línea Nic 1061. El extracto de la línea Nic 1015 fue el único con actividad contra Pc hasta 5 µg de extracto crudo seco por disco. Estos resultados sugieren un uso potencial de los extractos crudos de las líneas Nic 1061 y Nic 1015 “TI 1341” como un agente efectivo para la protección de cultivos contra estas bacterias.Palabras clave: fitopatógenos, Nicotiana tabacum, protección de cultivos, solventes, superficie foliar.Key words: phytopathogens, Nicotiana tabacum, crop protection, solvents, leaf surface.Recibido: noviembre 18 de 2014 Aprobado: ab

Natural products are an alternative to control microorganisms that cause diseases in crops. This work aimed to evaluate different solvents for obtaining crude extracts from tobacco leaf exudates and to determine in vitro effect of these extracts against two phytopathogenic bacteria: Xanthomonas campestris(Xc) and Pectobacterium carotovorum(Pc). Crude extracts from ten tobacco lines using solvents with polarities between 3.1 and 6.2 (dichloromethane, n-butanol, ethyl acetate, methanol and ethanol 90%) were obtained. Ethanol 90% was selected as the best solvent for obtaining extracts from tobacco leaf exudates and as a substitute of dichloromethane due to the best yield. The chemical composition diversity of the ethanolic extracts was revealed by thin-layer chromatography. The antibacterial activity was evaluated by agar disk diffusion method recording the inhibition zones. Growth inhibition was observed for all extracts against Xc, and the better activity corresponded to the lines Nic 1061"TI 1738" and Nic 1016 "Incekara" until a minimal amount of 5 µg/ disc, with higher yield in case of the line Nic1061 . Only the extract of the line Nic 1015 was able to inhibit the growth of Pc until a minimal inhibitory concentration of 5 µg/disc. These results suggest a potential use of crude extracts from lines Nic 1061 and Nic 1015 "TI 1341" as an effective agent for the crop protection against Xc and Pc respectively., Los productos naturales son una alternativa para el control de microorganismos que ocasionan enfermedades en los cultivos. Este trabajo tuvo como objetivo evaluar diferentes solventes para la obtención de extractos crudos a partir de exudados foliares de líneas de tabaco, y el efecto in vitro de estos extractos contra dos bacterias fitopatógenas: Xanthomonas campestris (Xc) y Pectobacterium carotovorum (Pc). Se evaluaron solventes con polaridades entre 3.1 y 6.2 (diclorometano, n-butanol, acetato de etilo, metanol y etanol 90 %). El etanol 90 % se seleccionó como mejor solvente y como sustituto del diclorometano por su mayor rendimiento. Los extractos etanólicos crudos se obtuvieron a partir de exudados foliares de diez líneas de tabaco seleccionadas. La diversidad de la composición química de los extractos etanólicos se reveló por cromatografía en capa delgada. La actividad antibacteriana se evaluó por el método de difusión en agar con discos de papel de filtro y la medición del diámetro del halo de inhibición. Se observó inhibición para todos los extractos contra Xc destacándose los correspondientes a las líneas Nic 1061 "TI 1738" y Nic 1016 "Incekara" hasta 5 µg de extracto crudo seco /disco, con un mayor rendimiento para la línea Nic 1061. El extracto de la línea Nic 1015 fue el único con actividad contra Pc hasta 5 µg de extracto crudo seco por disco. Estos resultados sugieren un uso potencial de los extractos crudos de las líneas Nic 1061 y Nic 1015 "TI 1341" como un agente efectivo para la protección de cultivos contra estas bacterias.

Theobroma cacao L. is a crop of major importance for their potential applications in pharmacy and agriculture. It contains metabolites that are of great interest for its natural biological functions in plant response to different types of stress, both biotic and abiotic. The presence, type and concentration of both primary and secondary metabolites may vary depending on the genotype analyzed and these compounds can accumulate in the architecture associated structures such as leaf trichomes. The aim of this study was to characterize 26 clones of the germplasm bank of Padrón, INIA-Miranda, Miranda State, Venezuela and study the relationship between biochemical indicators assessed and the number of trichomes. The content of phenolic compounds, soluble protein, free proline, total carbohydrates, lignins, the number of trichomes and the relationship between all traits was determined. The results showed that in the two dimensional space explained 46.68% of the variability, five homogeneous groups were defined: 10 clones were associated with the maximum gradient of proline, total carbohydrate and lignin content. Nine clones were associated with high content of phenols and a group of seven clones with the highest number of trichomes on leaves. It was found that the number of trichomes was not related to the concentration of phenolic compounds. The clones of Forastero cocoa type showed higher concentration of total phenols and carbohydrates, meanwhile the Trinitario type presented higher concentration of proteins and prolina., Theobroma cacao L. es un cultivo de gran importancia por sus posibles aplicaciones en la farmacia y la agricultura. Contiene metabolitos que resultan de gran interés por sus funciones biológicas naturales en la respuesta de la planta a los diferentes tipos de estrés, tanto bióticos como abióticos. La presencia, tipo y concentración de los metabolitos (primarios y secundarios) pueden variar de acuerdo al genotipo analizado y se pueden acumular en estructuras asociadas a la arquitectura de la hoja como son los tricomas. El objetivo de este trabajo fue caracterizar 26 clones del banco de germoplasma de Padrón, INIA-Miranda, Estado Miranda, Venezuela y estudiar la relación existente entre los indicadores bioquímicos y el número de tricomas. Se determinó el contenido de compuestos fenólicos, proteínas solubles, prolina libre, carbohidratos totales, ligninas, el número de tricomas y la relación existente entre ellos. Los resultados mostraron que en el espacio bidimensional que explica el 46,68% de la variabilidad total, se definieron cinco grupos homogéneos: diez clones se asociaron con el máximo gradiente de prolina, carbohidratos totales y contenido de ligninas, nueve con alto contenido de proteínas y fenoles y siete con mayor número de tricomas en hojas. Se comprobó que el número de tricomas no se relacionó con la concentración de compuestos fenólicos. Los clones de cacao del tipo forastero mostraron mayor concentración de fenoles totales y carbohidratos, en tanto que los trinitarios presentaron mayor concentración de proteínas y prolina.

Enzymes as immobilized derivatives have been widely used in Food, Agrochemical, Pharmaceutical and Biotechnological industries. Protein immobilization is probably the most used technology to improve the operational stability of these molecules. Bromelain (Ananas comosus) and papain (Carica papaya) are cystein proteases extensively used as immobilized biocatalyst with several applications in therapeutics, racemic mixtures resolution, affinity chromatography and others industrial scenarios. The aim of this work was to optimize the covalent immobilization of bromelain and papain via rational design of immobilized derivatives strategy (RDID) and RDID1.0 program. Were determined the maximum protein quantity to immobilize, the optimum immobilization pH (in terms of functional activity retention), was predicted the most probable configuration of the immobilized derivative and the probabilities of multipoint covalent attachment. As support material was used Glyoxyl-Sepharose CL 4B. The accuracy of RDID1.0 program´s prediction was demonstrated comparing with experimental results. Bromelain and papain immobilized derivatives showed desired characteristics for industrial biocatalysis, such as: elevate pH stability retaining 95% and 100% residual activity at pH 7.0 and 8.0, for bromelain and papain, respectively; high thermal stability at 30 °C retaining 90% residual activity for both immobilized enzymes; a catalytic configuration bonded by immobilization at optimal pH; and the ligand load achieve ensure the minimization of diffusional restrictions., Las enzimas inmovilizadas han sido ampliamente utilizadas en las industrias Alimentaria, Agroquímica, Farmacéutica y Biotecnológica. La inmovilización de proteínas es, probablemente, la tecnología más empleada para elevar la estabilidad operacional de estas moléculas. La bromelina (Ananas comosus) y la papaína (Carica papaya) son cisteín proteasas extensamente usadas como biocatalizadores inmovilizados con disímiles aplicaciones en la terapéutica, resolución de mezclas racémicas, cromatografía de afinidad, entre otros escenarios industriales. El objetivo del presente trabajo fue optimizar la inmovilización covalente de las enzimas bromelina y papaína a través de la estrategia de diseño racional de derivados inmovilizados (RDID) y el programa RDID1.0. Se predijo la cantidad máxima de proteína a inmovilizar, el pH óptimo de inmovilización (en términos de retención de la actividad funcional), la configuración más probable del derivado inmovilizado y la probabilidad de enlazamiento covalente multipuntual. Como soporte de inmovilización de empleó Glioxil-Sepharose CL 4B. La precisión de las predicciones llevadas a cabo con el programa RDID1.0 fue validada comparando con los resultados experimentales obtenidos. Los derivados inmovilizados de bromelina y papaína mostraron características deseadas para la biocatálisis a nivel industrial, tales como: elevada estabilidad al pH reteniendo el 95% y 100% de actividad residual a pH 7.0 y 8.0, para la bromelina y la papaína, respectivamente; una elevada estabilidad térmica con la retención del 90% de actividad residual a 30 °C para ambas enzimas; al pH de inmovilización óptimo la configuración obtenida es catalíticamente competente; y la carga de ligando alcanzada asegura la disminución de las restricciones difusionales.