Herbicidal Composition comprising Chromone Derivatives and Method for Weed Control.
Inventores: Gustavo SOSA; M. Lucía TRAVAINI; Néstor CARRILLO; Eduardo CECCARELLI; Helmut WALTER. Submitted to the PCT. (2016) N/Ref: 639-3033 PCT.
Khellin and Visnagin, Furanochromones from Ammi visnaga (L.) Lam., as Potential Bioherbicides.
Maria L. Travaini, Gustavo M. Sosa, Eduardo A. Ceccarelli, Helmut Walter, Charles L. Cantrell, Néstor J. Carrillo, Franck E. Dayan, Kumudini M. Meepagala, and Stephen O. Duke. Journal of Agricultural and Food Chemistry, 2016, 64 (50), pp 9475–9487.
Plants constitute a source of novel phytotoxic compounds to be explored in searching for effective and environmentally safe herbicides. From a previous screening of plant extracts for their phytotoxicity, a dichloromethane extract of Ammi visnaga (L.) Lam. was selected for further study. Phytotoxicity-guided fractionation of this extract yielded two furanochromones: khellin and visnagin, whose herbicidal activity had not been described before. Khellin and visnagin were phytotoxic to model species lettuce (Lactuca sativa) and duckweed (Lemna paucicostata), with IC50 values ranging from 110 to 175 μM. These compounds, also inhibited the growth and germination of a diverse group of weeds at 0.5 and 1 mM. These weeds included five grasses [ryegrass (Lolium multiflorum), barnyardgrass (Echinocloa crus-galli), crabgrass (Digitaria sanguinalis), foxtail (Setaria italica), and millet (Panicum sp.)] and two broadleaf species [morningglory (Ipomea sp.) and velvetleaf (Abutilon theophrasti)]. During greenhouse studies visnagin was the most active and showed significant contact postemergence herbicidal activity on velvetleaf and crabgrass at 2 kg a. i. ha-1. Moreover, its effect at 4 kg a. i. ha-1 was comparable to the bioherbicide pelargonic acid at the same rate. The mode of action of khellin and visnagin was not a light-dependent process. Both compounds caused membrane destabilization, photosynthetic efficiency reduction, inhibition of cell division and cell death. These results support the potential of visnagin, and possibly khellin, as bioherbicides or lead molecules for the development of new herbicides.
Herbicidal Composition comprising Chromone Derivatives and Method for Weed Control.
Inventores: Gustavo SOSA; M. Lucía TRAVAINI; Néstor CARRILLO; Eduardo CECCARELLI; Helmut WALTER; Stephen DUKE; Charles CANTREL; Kumudini MEEPAGALA (2015) Provisional patent in USA 62/272.880. N/Ref.: 577 US PROV.
Khellin and visnagin, furanochromones from Ammi visnaga (L.) Lam., as potential bioherbicides.
M. L. Travaini, N. J. Carrillo, E. A. Ceccarelli, H. Walter, G. M. Sosa, C. L. Cantrell, K. M. Meepagala, S. O. Duke. 56th Meeting of the Weed Science Society of America (WSSA) and 69th Meeting of the Southern Weed Science Society (SWSS) (2016) San Juan, Puerto Rico.
There is an increasing demand of new molecules that serve as leads structures for the development of herbicides. Plants natural products provide an attractive alternative in finding effective and environmentally safe phytotoxic compounds, with high structural diversity and novel modes of action. Considering this situation, a systematic process of searching, evaluation and selection was developed in order to find plant extracts with promising phytotoxic activity.
As a result of this screening process involving nearly 2400 plant extracts from different regions of Argentina, a dichloromethane extract of toothpick weed (Ammi visnaga (L.) Lam.) was selected because of its significant herbicidal activity. Phytotoxicity assay-guided fractionation yielded two furanochromones: khellin and visnagin. These compounds have been previously reported in toothpick weed, but their herbicidal activity has not been described before.
Khellin and visnagin significantly inhibited the development of lettuce (Lactuca sativa) (germination IC50 = 700 and 740 μM, growth IC50 = 110 and 170 μM) and duckweed (Lemna paucicostata) (growth IC50 = 160 and 120 μM). Also, during laboratory bioassays in Petri dishes, both compounds at 1 mM strongly interfered with growth and germination of weeds: ryegrass (Lolium perenne), morningglory (Ipomea sp.), foxtail (Setaria italica) and millet (Panicum sp.). The natural compounds’ inhibitory effects were similar to those caused by acetochlor (0.54 mM) and glyphosate (0.75 mM) in the same bioassays.
Visnagin showed the most promising activity. It had significant contact post-emergence herbicidal activity on velvetleaf (Abutilon theophrasti) and crabgrass (Digitaria sanguinalis) during greenhouse tests at 2 kg ai ha-1. Moreover, its effect at 4 kg ai ha-1 against velvetleaf, crabgrass and barnyardgrass (Echinochloa crus-galli) was comparable to the bioherbicide pelargonic acid at the same rate. These results support visnagin’s potential as bioherbicide or lead for the development of a new herbicide.
Additionally, our results suggest that the mode of action of these furanochromones is a process involving multiple targets: membranes destabilization, photosynthetic efficiency reduction, and cell division inhibition. The herbicidal activity could be related to the potential phototoxicity associated with this type of compound. However, although the membrane destabilization was intensified after an irradiation period, the phytotoxic activity of these natural compounds was not light-dependent.
In conclusion, the plant extract screening method developed in this work enabled the identification of two natural compounds whose herbicidal activity is hereby reported for the first time.
Screening of an aqueous plant extract library for antibiotic activities against PhoP/PhoQ two-component system in Salmonella enterica.
M. G. Mediavilla, G. Viarengo, M. L. Travaini, G. Sosa, E. Caccarelli& E. García-Véscovi. XI CongresoArgentino de Microbiología General (SAMIGE) (2015) Córdoba, Argentina.
The Salmonella enterica serovar Typhimurium (S. Typhimurium) PhoP/PhoQ system is an orthodox two-component system that serves as a master regulator of this bacterium virulence. The regulon governed by this system is globally regulated by Mg2+ limitation and subsets of genes respond to different signals such as mild acidic pH, cationic antimicrobial peptides or unsaturated fatty acids. Taking into consideration the relevant role of PhoP/PhoQ system in the adaptation to environmental challenges during interaction of the bacterium with the host and that it can only be found in prokaryotes, low eukaryotes and plants, the system becomes an ideal target to search for new compounds in order to prevent/control S. Typhimurium virulence in mammalian hosts. Plant extracts are sources of compounds with exceptional chemical diversity that constantly gives rise to novel therapeutic agents. In this work, we carried out a screening of aqueous extracts of native plant species from Provincia de Chaco, Argentina, collected during a dry season to identify naturally occurring molecules that would modulate PhoP/PhoQ activity. To accomplish this task, we performed b-galactosidase activity assays in a 96-multiwell plate system to quantitatively asses the action of the extracts on the expression of PhoP-dependent reporters. We tested the activity of 6 different PhoP activated genes with transcriptional fusions to lacZ when bacteria were grown overnight in LB (activating condition), LB + MgCl2 (repressing condition) and LB + plant extracts (test condition) either in 1 mL final volume in assay tubes or in 200 mL final volume in 96-multiwell plates. Of 38 initial lyophilized extracts we determined sub-inhibitory concentrations (in the 0,01 – 5 mg/mL range) and the regulatory effect on the different reporter strains. We selected 10 of these extracts to continue the characterization: 9 showed a repressing effect towards the reporter genes under study while only one (53H) unexpectedly showed repression of most reporters tested but activation of two of them (virK::lacZ and pcgO::lacZ ). There is no previously reported relationship between these particular genes. We are currently pursuing the isolation and identification of the compounds responsible for the repressing/activating effects of the extracts, starting with a bioguided sub-fractionation approach. We are particularly focusing in extract 53H, to also determine whether there is a functional relationship between the activated genes. In sum, we successfully set up a protocol to test a large library of samples in a short time in order to establish a high throughput screening method to test and identify bioactive compounds with modulatory effect on the Salmonella enterica PhoP/PhoQ regulatory system.
Phytotoxicity and antiproliferative activity on tumor cells in vitro of plant extracts.
Travaini, M. L.; García Labari I., Carrillo, N.; Ceccarelli, E. A.; Girardini, J. & Sosa, G. M. L Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB) (2014) Rosario, Argentina.
The interaction of plants with the environment and other organisms causes them to make compounds with diverse biological activities. These compounds are used as active ingredients for many applications. It is estimated that 85% of the world flora is still unexplored. Our goals are to identify plant extracts as source of active ingredients for agro and pharma industry. Therefore we evaluated the phytotoxic activity of plant extracts, as well as, their capacity to inhibit in vitro the proliferation of tumor cell lines. A total of 103 aqueous extracts were prepared from 62 plant species collected in Argentina. Germination assays were performed using all these extracts against two dicot and three monocot species. Four extracts were interesting because they significantly inhibited germination of these five species at 1 and 8 mg/mL. In addition, they showed pre- and post-emergence negative effect on plant growth at 1 mg/mL. On the other hand, four extracts showed anti-proliferative activity against the tumor cell line MDA-MB-231, from breast adenocarcinoma. Moreover, other four extracts were active against the H1299 line, from lung carcinoma. The proliferation inhibition was over than 50% when a dilution 1/2000 of each extract was applied. These results show promising possibilities for these samples and identification of the molecules responsible for the effects is in progress.
Búsqueda de inhibidores de la germinación y/o crecimiento con potencial aplicación como agroquímicos.
María Lucía Travaini; E. A. Ceccarelli; N. J. Carrillo y G. M. Sosa. XI Simposio Argentino y XIV Simposio Latinoamericano de Farmacobotánica. I Congreso Latinoamericano de Plantas Medicinales (2013) Rosario, Argentina.
La pérdida de actividad de algunos herbicidas convencionales y la evolución de resistencia hacia muchos de ellos ha generado la necesidad de encontrar nuevos principios activos con mecanismos de acción diferentes a los conocidos. En este sentido, los productos naturales de las plantas ofrecen una fuente de moléculas diversas. Particularmente, la presencia de ciertos metabolitos fitotóxicos se ha asociado a especies colonizadoras o invasivas y estaría controlada por factores genéticos así como medioambientales. En este contexto, planteamos una búsqueda de inhibidores de la germinación, a partir de extractos vegetales, que puedan ser utilizados en el desarrollo de agroquímicos. Para ello se recolectaron especies que exhibían un comportamiento invasivo, en regiones de Argentina que presentan condiciones climáticas y geográficas adversas. Utilizando los distintos órganos de las mismas se prepararon extractos acuosos y se evaluó su fitotoxicidad contra lechuga (Lactuca sativa). De esta manera se seleccionaron 27 extractos que inhibieron la germinación de dicha especie al 100%. Usando diferentes concentraciones de cada uno de éstos se realizaron nuevos ensayos de germinación de lechuga y especies consideradas modelo de malezas: moha (Setaria itálica), ryegrass anual (Lolium multiflorum) y avena (Avena sativa). Se destacaron particularmente los extractos de hoja 118 y 1429, y el 1503 de fruto. El primero proveniente de una especie recolectada en el Sureste de la provincia de Santiago del Estero; y los dos siguientes, en el Noroeste de Santa Fe. No pueden detallarse las especies puesto que existe un acuerdo de confidencialidad. Estos extractos mostraron el CI50 más bajo para inhibir la germinación de lechuga, en un rango de 1,1-3,2 mg/mL de extracto seco, y generaron porcentajes de inhibición de la germinación de malezas superiores al 80% cuando fueron aplicados a una concentración del 25%. Al realizar curvas de crecimiento de lechuga y ryegrass anual en presencia de los extractos destacados, éstos inhibieron significativamente el crecimiento y desarrollo de ambas especies luego de una semana de tratamiento. Los resultados expuestos hasta aquí han permitido identificar al menos tres extractos vegetales atractivos como potencial fuente de compuestos con actividad herbicida.
Aqueous extracts from plants with phytotoxic activity.
Travaini, M. L.; Carrillo, N. J.; Ceccarelli, E. A.; Sosa G. XLVIII Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB) (2012) Mendoza, Argentina.
The extensive use of synthetic herbicides directed against a limited number of targets to control weeds has accelerated the rate of appearance of resistance among them. The use of new substances with different sites of action would reduce that problem. In that sense, we propose the use of phytotoxic compounds isolated from plants. Several known vegetal species release germination and/or growth inhibitors as a strategy to colonize or invade grounds occupied by other plants. We collected samples (organs and surrounding soil) of plants that exhibited an invasive behaviour in different regions of Argentina. We also harvested plants popularly known for their medicinal use. A total of 2103 samples were processed. Aqueous extracts were obtained from each sample and their inhibitory activity was tested in assays of lettuce and agronomic seeds germination. We detected that 251 extracts inhibited lettuce germination at 100%. Moreover, 4 extracts were selected because they showed the lowest IC50 (below 3 mg/mL) when tested for lettuce germination inhibition and produced also a post germination effect. Longer exposures of the seeds to these extracts produced an increase on inhibition on lettuce germination and growth. Particularly, the effect on germination was irreversible: after an exposition of seeds to these 4 extracts normal levels of germination were never recovered.
Screening of germination inhibitors from plant extracts.
Travaini, M. L.; Ceccarelli, E. A.; Carrillo, N. J.; Sosa G. XLVII Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB) (2011) Potrero de los Funes, Argentina.
Several plant species release germination and/or growth inhibitors as a strategy to colonize and invade grounds occupied by other plants. As a result, these plants populate successfully the bare ground or displace other native plant communities. It is known that the production of these substances increases if plants grow in a harsh environment under stressful conditions. We have collected plant species that exhibited an invasive behavior in regions of Argentina with hostile geography and harsh environments. We have also harvest those plants popularly known for their medicinal use. Using parts and soil of these plants, aqueous extracts were obtained (8 g material/70 mL water). These extracts were tested for their inhibition activity in germination assays of lettuce. Extracts that produced inhibition were then tested on the germination of seeds of agronomic interest. 804 extracts were prepared and 184 of them inhibited lettuce germination at 100%: 98 were derived from Chaco and Santiago del Estero plants, 32 from Cordoba and San Luis and 54 from medicinal species. Regarding the distribution of extracts with inhibition activity, most of them were obtained from leaves (52%), followed by stems (14%) and flowers (12%). Among the extracts used in the germination of agriculturally important species, 23 were noted for their selectivity or aggressiveness in general.