Made available in DSpace on 2014-08-20T14:37:57Z (GMT). No. of bitstreams: 1 tese_fabiane_grecco.pdf: 3401915 bytes, checksum: 93b93d478f426a7fe7b06afda9b90b14 (MD5) Previous issue date: 2009-06-08 / This study him to characterize liver morphological patterns of the intoxication by Senecio spp. observed in cattle in Southern Rio Grande do Sul State, Brazil between 2000-2008 and to text the acquireal resistance in sheep to Senecio brasiliensis poisoning. The patterns observed were correlated with climate changes occurred in the period. Two outbreaks Senecio spp. In sheep were described. It was tested if repeated doses of S. brasiliensis induce resistance to the intoxication in sheep. The macroscopic and histological lesions of cattle dead by Senecio spp. toxicosis submitled to Regional Diagnostic Laboratory of Pelotas University were analyzed. The lesions were classified in seven different patterns. The outbreaks prevalence and climate changes concerning accumulated rain and temperature mean in different seasons of year were analyzed. To determine if repeated doses of S. brasiliensis induce resistance to the intoxication in sheep, doses that cause acute intoxication was determined. The green plant was administered by gavage to 3 sheep in doses of 60, 80, 90 and 100 g/kg of body weight (bw). The doses (100g\kg of bw) that caused acute poisoning were divided in 2, 5 and 10 doses and administered daily in 2, 5 and 10 days to observe if chronic lesion would develop. Sheep that received 10g/kg of bw was challenged with 100g/kg of bw after 45 days the last doses of 10g. Doses of 15 g/kg of bw were administered by gavage for 30 days and 10 g/kg of bw for 10 days to three sheep. Two of them were challenged 24 hours after the last doses of 30g/kg of bw. One sheep was challenged 15 days after the last doses of 30g/kg of bw. The results of macroscopic and histological analyze of liver showed that the diffuse liver fibrosis and nodular regeneration were the most common lesions induced by Senecio spp. intoxication in cattle. Although between 2007 and 2008 the pattern with little fibrosis and histological subacute lesion had been observed. In outbreaks of spontaneous Senecio spp. poisoning in sheep jaundice, photodermatitis, progressive emaciation and death in 4 to 30 days were observed. Macroscopic lesion were characterized by jaundice, yellowish liver, mesentery edema, ascitis, alternativaly dark and firm liver with numerous small well-circumscribed nodules measuring 1-5mm in diameter in the cut and capsular surface. Microscopically the hepatic lesions were similar in all affected sheep and characterized by hepatomegalocytosis, periportal fibrosis, hepatocytes necrosis and biliary ductal proliferation. The results of experimental intoxication showed that doses up 90g/kg of bw cause acute intoxication and the doses of 100g/kg of bw divided in10 doses do not induces resistance to Senecio brasiliensis poisoning in sheep. It was demonstrated that 15g/kg of bw for 30 days and 30g/kg of bw for 10 days induce resistance to Senecio basiliensis poisoning if the animals were challenged with 100g/kg of bw immediately after last dose of 30g/kg of bw and lost resistance after 15 days. / Os objetivos do presente trabalho foram caracterizar os diferentes padrões morfológicos hepáticos da intoxicação por Senecio spp. em bovinos observados na área de influência do Laboratório Regional de Diagnóstico da Faculdade de Veterinária da UFPel (LRD) entre 2000-2008, fazendo uma relação com as variações climáticas ocorridas no período; descrever dois surtos da intoxicação por Senecio spp em ovinos; e determinar se doses repetidas e não tóxicas da planta induzem a resistência de ovinos à intoxicação por Senecio brasiliensis. Para tanto, este trabalho foi dividido em duas partes sendo que na primeira foram analisadas as lesões macroscópicas e histológicas dos fígados de bovinos mortos pela intoxicação que foram necropsiados e/ou tiveram órgãos remetidos ao LRD, as quais foram agrupadas em sete diferentes padrões morfológicos. Foram determinados os dados da prevalência dos surtos durante este período e as variações climáticas referentes à precipitação pluviométrica e temperatura média nas diferentes estações do ano. Na segunda parte do trabalho foram descritos dois surtos da intoxicação por Senecio spp. em ovinos. Para determinar se doses não tóxicas da planta verde induzem a resistência em ovinos inicialmente foi determinada a dose tóxica capaz de induzir intoxicação aguda nesta espécie animal. Foram administradas 60, 80, 90 e 100g/kg de peso corporal (pc) a três ovinos. As doses que causaram a morte dos animais foram divididas em duas, cinco e 10 doses administradas diariamente para determinar se os animais apresentavam lesões hepáticas crônicas, sendo que o ovino que recebeu 10g/kg de pc por 10 dias foi desafiado 45 dias após a última administração com a dose de 100g/kg de pc. Para induzir a resistência doses de 15g/kg de peso corporal foram administradas por 30 dias e doses de 30g/kg de pc foram administradas por 10 dias a três ovinos. Dois foram desafiados no dia seguinte ao final da administração da planta e um foi desafiado 15 dias após. Os resultados da análise macroscópica e histológica dos fígados dos bovinos demonstraram que os padrões mais frequentemente encontrados são aqueles caracterizados pela presença e fibrose difusa e presença de nódulos regenerativos, embora nos casos ocorridos entre 2007 e 2008 padrões de lesão com menor quantidade de fibrose e 2 casos de lesão subaguda tenham sido observados. Nos surtos espontâneos de intoxicação por Senecio spp. em ovinos foi observada icterícia, fotossensibilização, perda de peso e morte entre quatro dias e um mês e as lesões macroscópicas caracterizaram-se por icterícia discreta e generalizada e fígado amarelado e firme ou palidez da carcaça, ascite, edema de mesentério e fígado escuro e firme com nódulos esbranquiçados de 1-5 mm de diâmetro tanto na superfície capsular como ao corte. Microscopicamente, as lesões hepáticas eram semelhantes em todos os ovinos afetados e caracterizadas por megalocitose, fibrose periportal, necrose individual e aleatória de hepatócitos e hiperplasia acentuada das células dos ductos biliares. Os resultados da intoxicação experimental demonstraram que doses acima de 90g/kg de pc são capazes de produzir intoxicação aguda e que a dose de 100g/kg de pc fracionada em 10 doses de 10g/kg de pc não induzem resistência à intoxicação nesta espécie animal. Foi demonstrado que doses de 15 g/kg de pc por 30 dias e 30g/kg de pc por 10 dias induzem resistência nos ovinos quando os mesmos são desafiados imediatamente após o término do experimento e que se o desafio é realizado 15 dias após o final do experimento os animais perdem a resistência.

Species belonging to the Asteraceae family are an important source of terpenoids with biological activity. Senecio bonariensis Hook. et Arn., popularly known as margarida-do-banhado-de-Buenos Aires is native from South America and can be found in south Brazil, blooming from October to December. This plant is known for contain furanoeremophylanes, steroids and pyrrolizidines alkaloids (PAs) with toxic activities. This work describes the isolation and identification of three compounds present in the CH2Cl2 extract of the aerial parts of Senecio bonariensis Hook at Arn. Leaves of this plant were collected in Eldorado do Sul, RS, Brazil, in April 2004 and identified by Prof. Dr. Nelson Ivo Matzenbacher. Voucher specimen Nº SMDB 9519 is preserved in the Herbarium of the Departamento de Botânica, Universidade Federal de Santa Maria, Santa Maria, RS, Brazil. The fresh leaves of S. bonariensis (2.3 kg) were extracted by maceration with CH2Cl2. The CH2Cl2 extract was evaporated under vacuum to yield an viscous residue (13.5 g), which gave a yield of 0.64%. The crude extract was fractionated by flash chromatography over silica-gel, using CH2Cl2, and CH2Cl2: EtOH mixture of increasing polarity, (stepwise, 99:1 to 80:20) to yield 15 fractions. The fraction 1 col. 1 (1 g) was chromatographed over silica-gel impregnated with AgNO3 (10%) eluting with hexane-acetone (99:1), to yield 13 news fractions. Fraction 10 col. 2 (253 mg) was chromatographed on silica-gel impregnated with AgNO3 (10%), eluting with hexane: ethyl ether (95:5) and purified to afford 104 mg of CS1 (caryophyllene) and 16 mg of CS2 (caryophyllene oxide). Fraction 11 col. 2 (83.5 mg) was also chromatographed on silica-gel impregnated with AgNO3 (10%), eluting with hexane: ethyl ether (98:2) and purified to afford 5 mg of CS3 (germacrene-D). The compounds obtained were analyzed by GC-EI-MS, IR, 1H and 13C NMR. The antimicrobial activity of CS1 and CS2 were evaluated by the microdilution method based on M27-A2 and M7-A4 (NCCLS). Both compounds sowed no antimicrobial activity at the tested concentrations. / Espécies pertencentes à família Asteraceae, são uma importante fonte de terpenóides com várias atividades biológicas já descritas. Senecio bonariensis Hook. et Arn., popularmente conhecida como margarida-do-banhado-de-Buenos Aires é um arbusto nativo da América do Sul, que pode ser encontrado no sul do Brasil e floresce de outubro a dezembro. Esta planta é conhecida por apresentar furanoeremofilanos, esteróides e alcalóides pirrolizidínicos, com atividades tóxicas. Este trabalho descreve o isolamento e identificação de três constituintes químicos presentes no extrato CH2Cl2 das partes aéreas de Senecio bonariensis Hook. et Arn. As folhas de S. bonariensis foram coletadas em abril de 2004, no município de Eldorado do Sul-RS. A espécie foi localizada e identificada pelo Prof. Dr. Nelson Ivo Matzenbacher, do Programa de Pós Graduação em Botânica da UFRGS. Material testemunha encontra-se depositado no Herbário do Departamento de Biologia da UFSM sob o registro n° SMDB 9519. As folhas frescas de S. bonariensis (2,3 kg) foram extraídas por maceração com CH2Cl2. O extrato bruto foi evaporado sob pressão reduzida, resultando num resíduo pastoso (13,5 g), com rendimento de 0,64%. O extrato bruto foi fracionado por cromatografia em coluna do tipo flash sobre gel de sílica, usando CH2Cl2 e misturas de CH2Cl2 : EtOH, em gradiente, nas proporções de 99:1 até 80:20, sendo obtidas 15 frações (1 a 15 col. 1). A fração 1 col. 1 (1g) foi cromatografada sobre gel de sílica impregnado com AgNO3 (10%), eluída com hexano: acetona (99:1), obtendo-se 13 novas frações (1 a 13 col. 2). A fração 10 col. 2 (253 mg) foi cromatografada sobre gel de sílica impregnado com AgNO3 (10%), eluída com hexano:éter etílico (95:5), obtendo-se 104 mg de CS1(cariofileno) e 16 mg de CS2 (óxido de cariofileno). A fração 11 col. 2 (83,5 mg) também foi cromatografada sobre gel de sílica impregnado com AgNO3 (10%), eluída com hexano:éter etílico (98:2) e forneceu 5 mg de CS (germacreno-D). Os compostos isolados foram analisados por CG-EM-IE, IV, RMN 1H e RMN 13C e seus dados espectroscópicos foram comparados com os obtidos da literatura. Além disso, foi determinada a atividade antimicrobiana do cariofileno e do óxido de cariofileno, através do método de microdiluição em caldo, baseado nos documentos M27-A2/NCCLS para fungos e M7-A4/NCCLS para bactérias. Os compostos não apresentaram atividade antimicrobiana nas concentrações testadas.

Conselho Nacional de Desenvolvimento Científico e Tecnológico / During 36 months a study was conducted in beef cattle farms in Rio Grande do Sul, Brazil to survey outbreaks of Senecio spp. poisoning. The first part of this dissertation reports an outbreak of Senecio brasiliensis poisoning in cows where the main clinical sign was photosensitization. Adult cows that were placed during six months in a 205-hectare pasture heavily infested by Senecio brasiliensis were affected in September, 2013. Photosensitization was seen in 83 out 162 cows. Liver biopsy was performed in all cows under risk and three cows were necropsied. Histopathological findings in the liver related to pyrrolizidine alkaloids toxicosis consisted of fibrosis, megalocytosis and biliary ductal proliferation and were present in 119 of the biopsied cows. Six tested cows had increased serum activity of gamma glutamyl transferase. In the second part of this dissertation, a study of an outbreak of seneciosis in sheep in 2014 is reported. Morphological aspects considered important for the establishment of diagnosis in this species were stressed. Ten out of 860 adult sheep with clinical signs of seneciosis died spontaneously and eight sick ewes were euthanized and necropsied. Clinical signs included weight loss, apathy and photosensitization. Four out of seven tested sheep presented increased serum levels of gamma glutamil transferase and two had elevated alkaline phosphatase serum activity. At necropsy, three out of eight ewes presented slightly irregular toughened livers with multifocal nodules, two out of eight ewes had a whitish liver with thickened fibrotic Glisson s capsule partially adhered to the diaphragm, and three out of eight ewes had smooth and grossly normal livers. Histologically, the main hepatic findings that allowed for the establishment of the diagnosis were megalocytosis, proliferation of bile ducts and fibrosis. Spongy degeneration was observed in the brains of all eight necropsied sheep and was more severe at the cerebellar peduncles, mesencephalon, thalamus, and pons. These are suggested as the portions of election to investigate microscopic lesions of hepatic encephalopathy in sheep with chronic seneciosis. / Durante um período de 24 meses foram realizadas visitas periódicas a propriedades de bovinocultura de corte no Rio Grande do Sul, onde havia a suspeita de intoxicação por Senecio spp. Na primeira parte desta dissertação, relata-se a ocorrência de um surto de intoxicação natural por Senecio brasiliensis em bovinos em que o principal sinal clínico observado foi fotossensibilização. O surto ocorreu em setembro de 2013, acometendo vacas adultas que permaneceram durante os seis meses anteriores em um campo de 205 hectares com abundante infestação por Senecio brasiliensis. Fotossensibilização foi observada em 83 vacas de um total de 162 (51,3%). Realizou-se biópsia hepática em todas as vacas do rebanho e três vacas doentes foram necropsiadas. Os principais achados histopatológicos decorrentes de intoxicação por Senecio brasiliensis nos bovinos biopsiados foram fibrose hepática, megalocitose e proliferação de ductos biliares, observados em 119 animais. Seis vacas testadas apresentaram aumento da atividade sérica da gama glutamil transferase. Na segunda parte da dissertação, foi realizado um estudo de um surto de intoxicação por Senecio spp. em ovinos em 2014, dando-se ênfase aos aspectos morfológicos para o estabelecimento do diagnóstico da intoxicação nessa espécie. Dez ovelhas adultas de um rebanho de 860 ovinos morreram com sinais da doença e oito que estavam doentes foram eutanasiadas e necropsiadas. Os sinais clínicos incluíam emagrecimento, apatia e fotossensibilização. Quatro ovelhas, de um total de sete que foram testadas, apresentaram aumento da atividade sérica da gama glutamil transferase e duas apresentaram fosfatase alcalina sérica elevada. Na necropsia, em três dos oito ovinos necropsiados, o fígado estava levemente mais firme, com superfície capsular ligeiramente irregular e com nódulos pálidos na superfície de corte. Em outros três ovinos o fígado era macroscopicamente normal. Em dois dos oito ovinos necropsiados a cápsula de Glisson era brancacenta devido à fibrose, e parcialmente aderida ao diafragma. Histologicamente, as principais alterações observadas, que contribuíram para o estabelecimento do diagnóstico de intoxicação crônica por alcaloides pirrolizidínicos nas ovelhas deste surto foram hepatomegalocitose, proliferação de ductos biliares e fibrose. Todas as ovelhas apresentavam degeneração esponjosa grave no cerebelo e pedúnculos cerebelares, ponte, mesencéfalo e tálamo. Sugere-se que esses sejam os sítios anatômicos de eleição no encéfalo para a observação dessa lesão em ovinos com intoxicação por Senecio spp.

Thesis (Ph. D.)--Oregon State University, 2008. / Printout. Includes bibliographical references. Also available on the World Wide Web.

Tese (doutorado) - Universidade Federal de Santa Catarina, Centro de Ciências Físicas e Matemáticas, Programa de Pós-Graduação em Química, Florianópolis, 2014. / Made available in DSpace on 2015-02-05T20:35:53Z (GMT). No. of bitstreams: 1 331098.pdf: 3343639 bytes, checksum: 1aa3dc70f554a210df4d2516b5bbdff1 (MD5) Previous issue date: 2014 / O presente trabalho visou realizar um estudo fitoquímico analítico da espécie vegetal Senecio brasiliensis. Após o preparo do extrato bruto de diferentes partes da planta, foi desenvolvido um procedimento sistemático utilizando eletroforese capilar como guia para o subseqüente isolamento dos constituintes químicos. O procedimento aplicado permitiu verificar a presença de alcalóides, flavonóides e ácidos dicafeoilquinicos. Com o uso de técnicas espectroscópicas RMN, IV, EI-MS e ESI-MS foi possível identificar os alcalóides como senecionna, senecioina N-oxido, os ácidos x,y-dicafeoilquínicos 3,5, 3,4, 4,5 e 1,4, além dos flavonóides rutina e quercetrina. Para o estudo da distribuição dos alcalóides na planta, foi desenvolvido um método utilizando eletroforese capilar. A determinação de log P para os alcalóides foi efetuada através da EC, utilizando um mecanismo de separação em MEKC (cromatografia micelar capilar). O modelo obtido para os padrões foi log P = xlogk - y (n=6, R2 = 0.91) com log P variando de 0,4 ? 4,08. O valor de log P para a N-oxido senecionina foi determinado como < 0,4 e para a senecionina 0,7. Para a determinação da interação dos alcalóides pirrolizidinicos com albumina humana foi utilizado eletroforese capilar no modo PF-ACE. Ambos os compostos não apresentaram interação com a proteína humana. Para a determinação da permeabilidade em membrana paralela artificial (PAMPA) foi utilizada a técnica eletroforese capilar para as análises de permeabilidade. Com a finalidade de determinar alcalóides pirrolizidinicos em amostras de mel com diluição simples foi desenvolvido método utilizando HPLC-MS/MS.<br> / Abstract: This study aimed to carry out a phytochemical study of Senecio brasiliensis species under an analytical perspective. After preparation of the crude extract from different parts of plant, a systematic procedure using capillary electrophoresis (CE) as a guide for the subsequent isolation of the chemical constituents was developed. The procedure demonstrated the presence of alkaloids, flavonoids and dicaffeoylquinic acids. Using spectroscopic techniques (NMR, IR, EI-MS and ESI-MS) it was possible to identify the alkaloids senecionine[1] and N-oxide senecionine[2]; the x,y-dicaffeolylquinic acids (3,5)[3], (3,4)[4], (4,5)[5] and (1,4)[6] and also the flavonoids rutin[7] and quercetin[8]. To study the distribution of alkaloids in the plant, a novel method using CE was developed. For determining the log P for alkaloid, CE technique was used in MEKC (Micellar ElectrokineticChromatography) mode. The model obtained for the standards was logP=xlogk -y (n= 6, R2=0.91) with log P ranging from 0.4 to 4.08. The k values for senecionine and N-oxide senecionine were determined as0.7 and <0.4, respectively. To evaluate the interaction between pyrrolizidine alkaloids and human albumin (HSA), CE in PF-ACE (Partial-Filling Affinity Capillary Electrophoresis) mode was applied. The results suggest that both compounds do not present interaction with the studied protein. To determine the permeability of these alkaloids, experiments were carried out using parallel artificial membrane (PAMPA) and CE was used to analyze all solutions. In order to determine pyrrolizidine alkaloids in honey samples with simple dilution, a method using HPLC-MS/MS was developed.

Botanist Kjell-Arne Olsson inventoried the plant fen ragwort Senecio paludosus 1983 in Scania andfound a big number of locations within Biosphere Reserve Kristianstad Vattenrike. In 2012, a reinventoryof the 1983 survey was carried out to study whether there has been any change in the fenragwort population in Kristianstad Vattenrike since the last inventory. The results show that therehas been no overall change in the number of fen ragwort in Kristianstad Vattenrike, on certainpremises they have increased in number while in others they have fallen. However, the inventoryshows that almost half of the locations with fen ragwort have disappeared during the same period.The main reason for this is that fen ragwort did not manage to competitive with other vegetation,taking in consideration this has been smaller premises with young plants. In addition to competitionother factors such as prolonged high water, grazing and caterpillars from Tyria jacobaeaecontributed to the decrease. Despite this the future looks stable for fen ragwort in KristianstadVattenrike where several very strong premises exist in the region.

En este trabajo de tesis se seleccionaron tres especies entre nueve recolectadas en las Sierras Australes Bonaerenses, cuyos extractos mostraron inhibición de la enzima acetilcolinesterasa (ACE): Senecio ventanensis, Grindelia argentina y Grindelia ventanensis. No se hallaron antecedentes fitoquímicos o de actividades biológicas en literatura de estas asteráceas endémicas. Mediante el fraccionamiento bioguiado de los extractos activos se buscó aislar los metabolitos secundarios responsables de la actividad inhibitoria de ACE. Los alcaloides pirrolizidínicos N-óxido de usaramina (1), N-óxido de retrorsina (2) y N-óxido de integerrimina (3) fueron aislados a partir del extracto etanólico de S. ventanensis y un par de diastereoisómeros del 3,6-epidioxi-1,10-bisaboladieno (4 y 5) a partir de su sub-extracto diclorometánico. Por otra parte, distintos ácidos clorogénicos fueron detectados en fracciones activas del sub-extracto butanólico de G. argentina: ácido 5-Ocafeoilquínico (6), ácido 5-O-p-cumaroilquínico (7), ácido 5-O-p-feruloilquínico (8) y ácido-3,5- dicafeoil-epi-quínico (9). Por último, el fraccionamiento bioguiado del extracto etanólico de G. ventanensis, el más activo, condujo al aislamiento de un diterpenoide labdano de la serie normal identificado como ácido 17-hidroxicatívico (10). Considerando la actividad anticolinesterasa de 10 y su extracción sencilla y en buen rendimiento, se prepararó una serie de veinte derivados nuevos del ácido catívico (16-19) a través de transformaciones sobre el grupo carboxilo (C15) de 10, introduciendo un espaciador de 2 a 6 carbonos y una amina terciaria. La mayoría de los derivados fueron más activos que el compuesto natural 10. El derivado con un anillo de pirrolidina unido al diterpenoide por un espaciador de cuatro carbonos (16c) fue el inhibidor más potente tanto de ACE como de butirilcolinesterasa; asimismo mostró inhibición significativa de ACE en células de neuroblastoma humanas SH-SY5Y, sin efecto citotóxico. El estudio cinético enzimático y el modelado molecular revelaron que 16c se une tanto al sitio activo catalítico como al sitio aniónico periférico de ACE. Por otro lado, los derivados 11-14 se obtuvieron mediante transformaciones químicas sencillas de 10, pero su actividad inhibitoria de ACE no fue mejorada por las derivatizaciones realizadas. La inhibición de la producción de óxido nítrico (NO) observada para el extracto etanólico de G. argentina llevó a su fraccionamiento bioguiado, permitiendo el aislamiento de tres saponinas tipo oleanano nuevas, grindeliósidos A-C (20-22), y una flavona conocida, hispidulina (23). Los metabolitos fueron evaluados por su actividad inhibitoria de la producción de NO inducida por LPSIFN- g en macrófagos RAW264.7 y por su actividad citotóxica contra la línea celular de leucemia humana CCRF-CEM y de fibroblastos de pulmón MRC-5. Hispidulina redujo marcadamente la producción de NO inducida por LPS-IFN-g, mientras que grindeliósidos A-C mostraron citotoxicidad, siendo grindeliósido C el más activo contra ambas líneas celulares. / In this work three of nine species collected in the Sierras Australes Bonaerenses were selected because of the acetylcholinesterase (AChE) inhibition showed by their extracts: Senecio ventanensis, Grindelia argentina y Grindelia ventanensis. There is no phytochemical or biological activities previous record in the literature for these endemic species of the Asteraceae family. The isolation of the metabolites responsible of the activity was conducted by a bioassay guided fractionation. The pyrrolizidine alkaloids usaramine N-oxide (1), retrorsine N-oxide (2) and integerrimine N-oxide (3) were isolated from the ethanolic extract of S. ventanensis and a pair of distereoisomers of 3,6-epidioxy-1,10-bisaboladiene (4 and 5) from its dichloromethane sub-extract. Additionally, different chlorogenic acids were detected in the active fractions of G. argentina butanolic sub-extract: 5-O-caffeoylquinic acid (6), 5-O-p-coumaroylquinic acid (7), 5-O-feruloylquinic acid (8), and 3,5-dicaffeoyl-epi-quinic acid (9). Finally, the bioactivity guided fractionation of the ethanolic extract of G. ventanensis, the most active one, led us to isolate a normal labdane diterpenoid identified as 17-hydroxycativic acid (10). Taking into account the anticholinesterase activity of 10 and its easy extraction in good yield, a set of twenty new cativic acid derivatives (16-19) was prepared from 10 through transformations on the carboxylic group at C15, introducing a C2-C6 linker and a tertiary amine group. Most of the tested compounds were better AChE and butyrylcholinesterase (BChE) inhibitors than the natural diterpenoid 10. The derivative with a four carbon spacer and a pyrrolidine moiety (16c) was the most potent AChE and BChE inhibitor; in addition it exhibited significant inhibition of AChE activity in SHSY5Y human neuroblastoma cells and was non-cytotoxic. Enzyme kinetic studies and molecular modeling revealed that 16c targeted both the catalytic active site and the peripheral anionic site of AChE. On the other hand, derivatives 11-14 were obtained through simple structural modifications of 10, but their AChE inhibitory activity was not improved by the derivatizations. The inhibitory activity of nitric oxide (NO) production of the ethanolic extract of G. argentina prompted us to isolate three new oleanane-type saponins, grindeliosides A-C (20-22), and a known flavone, hispidulin (23). The isolated compounds were evaluated for their inhibitory activities against LPS/IFN-g induced NO production in RAW 264.7 macrophages and for their cytotoxic activities against the human leukemic cell line CCRF-CEM and MRC-5 lung fibroblasts. Hispidulin markedly reduced LPS/IFN- g induced NO production while grindeliosides A–C were found to be cytotoxic, with grindelioside C being the most active against both cell lines.

Senecio L. es el género que presenta mayor número de especies dentro de la familia Asteraceae. Exhibe importantes conflictos sistemáticos, sus límites no están claramente definidos y la identificación de sus especies se ve dificultada por el solapamiento que presentan los caracteres exomorfológicos diagnósticos; que sumado a la gran variabilidad que presentan algunos taxones, lleva a descripciones y claves poco precisas y ambiguas. Los estudios anatómicos foliares y palinológicos son escasos para las especies argentinas del género en tanto que se cuenta con mayor conocimiento de su citogenética, en particular de la serie Corymbosi. En esta tesis se evaluó la utilidad taxonómica de la anatomía foliar, micromorfología de estambres y estilos, morfología polínica y número cromosómico (microcaracteres) en el género Senecio, en particular, de especies argentinas que presentan conflictos taxonómicos. Parte de la información obtenida permite profundizar el conocimiento del género tanto a nivel local como general y contribuye a esclarecer problemas taxonómicos. Los estudios se realizaron, principalmente en Senecio sect. Senecio serie Xerosencio y los resultados fueron comparados con miembros de Senecio sect. Senecio ser. Corymbosi; Senecio sect. Senecio serie Culcitium y Senecio sect. Senecio serie Otopteri. Se efectuó una recopilación y revisión exhaustiva del material bibliográfico relacionado con la diversidad y taxonomía del género, se examinaron especímenes Resumen ii provenientes de tres herbarios de la Argentina y se estudiaron plantas vivas en su ambiente natural. Se observaron caracteres vegetativos y reproductivos de los ejemplares de herbario y de plantas provenientes de su entorno natural. Se elaboraron descripciones ampliadas y detalladas de cada una de las especies consideradas, incluyendo aspectos micromorfológicos vegetativos (anatomía foliar) y reproductivos (micromorfología de estambres y estilos, palinología y número cromosómico). Para todas las especies, se señalaron sus respectivas áreas de distribución geográfica con comentarios sobre su hábitat. Los microcaracteres que se enuncian a continuación mostraron importancia taxonómica: ornamentación de la cutícula, tipos de tricomas, contorno de las células epidérmicas, características de las células oclusivas de los estomas, espesor de la pared tangencial externa de las células epidérmicas, presencia de células epidérmicas papilosas y estructura del mesofilo (caracteres anatómicos foliares), presencia de margo y de mesoapertura en los granos de polen, forma del collar del filamento, de la base y del apéndice estéril de la antera. Se realizaron estudios citogenéticos de las especies estudiadas, estableciéndose por primera vez el número cromosómico para cuatro taxones argentinos de Senecio, se confirma el contabilizado previamente por otros autores para S. vira-vira y se registra un nuevo nivel de ploidía para S. filaginoides. Se presentan claves dicotómicas que permiten diferenciar especies afines y variedades considerando el uso de estos microcaracteres. Se presenta un análisis sobre la validez de las series y subseries en que tradicionalmente se agruparon las especies estudiadas mediante la realización de un Análisis de componentes principales. Resumen iii La nueva información obtenida complementa la previamente existente y permite apoyar la sinonimia de las entidades infraespecíficas de S. perezii y los tratamientos infraespecíficos propuestos para S. subumbellatus y S. microcephalus. Por otra parte, no se apoya la sinonimia propuesta para S. subulatus var. erectus y S. subulatus var. salsus. Se proveen nuevos caracteres que justifican la exclusión de S. otites de Senecio sensu lato. / Senecio L., with more than 1250 species, is the largest genus of the family Asteraceae. It exhibits important systematic conflicts, its boundaries are not clearly defined and species identification is hampered by the overlapping of diagnostic exomorphological characters, which added to the large variability of some taxa, leads to ambiguous descriptions and makes keys difficult to use. Leaf anatomy and palynological studies are scarce for the Argentinean species of the genus, while there is a better knowledge of its cytogenetics, in particular for the series Corymbosi. The taxonomic utility of leaf anatomy, micromorphology of stamens and styles, pollen morphology and chromosome number (microcharacters) in the genus Senecio, particularly, of Argentinean species that present taxonomic conflicts, were evaluated. These studies were carried out mainly in Senecio sect. Senecio series Xerosenecio and their results were compared with members of Senecio sect. Senecio ser. Corymbosi; Senecio sect. Senecio series Culcitium and Senecio sect. Senecio series Otopteri. A comprehensive compilation and revision of bibliography related to the diversity and taxonomy of the genus was carried out. Vegetative (leaf anatomy) and reproductive (micromorphology of stamens and styles, palynology and chromosome number) characteristics of herbarium specimens and plants in their natural environments were observed and detailed descriptions were made of each species. The following microcharacters showed taxonomic importance: ornamentation of the cuticle, type of trichomes, type of contour of the epidermal cells and characteristics of the stomatal occlusive cells, thickness of the external wall of the epidermal cells, presence of papillary epidermal cells, and mesophyll structure, the presence of margo and mesoaperture in pollen grains, and shape of the filament collar, anthers base and sterile anther appendix. Chromosome numbers for four Argentinean taxa of Senecio are established for the first time, previous counts for S. vira-vira are confirmed, and a new level of ploidy for S. filaginoides is reported. However, this character has little value in the clustering of Senecio species. Dichotomous keys to differentiate related species and varieties considering microcharacters are presented. The validity of the series and subseries in which the species studied were traditionally grouped is tested using Principal Component Analysis. New morphological support is provided for the synonymy of the infraspecific entities of S. perezii which complements the infraspecific treatment given to S. subumbellatus and S. microcephalus. On the other hand, the synonymy of S. subulatus var. erectus and S. subulatus var. salsus, is not supported by the new data. The consideration of these microcharacters supports the exclusion of S. otites from Senecio sensu lato. Some of the information obtained throughout this thesis contributes to the knowledge of the genus Senecio and are an aid for the resolution of its internal systematics at the local and global levels.

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Allelopathy occurs when one organism releases a compound into the environment that affects the functioning of another organism. Scientists have long suspected that alleopathic plant compounds could offer novel, softer chemistries to the ongoing battle of controlling pests in agricultural fields. Strawberry growers rely on toxic fumigants to kill soilborne fungal pests, weeds, nematodes, and insects. Increased regulations have reduced the use of fumigants (including methyl bromide), and strawberry growers need new sustainable pest control solutions. We selected four putative allelochemicals with known fungicidal and herbicidal activity (ferulic acid, gallic acid, juglone, and p-Coumaric acid). We assessed the pesticidal activity of these plant compounds both in agar and in soil on two emerging soilborne fungal pathogens (Macrophomina phaseolina and Fusarium oxysporum f.sp. fragariae), and four annual weeds commonly found in strawberry production fields (Malva parviflora, Melilotus officinalis, Poa annua, and Senecio vulgaris). We also assayed lettuce (Lactuca sativa ‘Inferno’), which served as a positive control plant species due to its sensitivity to phytotoxic compounds. Fitted sigmoidal dose-response curves predicted EC50 and EC75 values for each combination of plant compound and pest. All plant compounds inhibited the in vitro radial mycelial growth of the two soilborne fungal pathogens in a dose-dependent manner. Fusarium oxysporum f.sp. fragariae was more sensitive to the plant compounds than Macrophomina phaseolina. Average EC50 values for the radial mycelial growth of two F. oxysporum f.sp. fragariae isolates were 75.1 parts per million by weight (ppmw) juglone, 469 ppmw p-Coumaric acid, and 687 ppmw ferulic acid. Average EC50 values for the radial mycelial growth of two M. phaseolina isolates were 196 ppmw juglone, 2869 ppmw p-Coumaric acid, and 5716 ppmw ferulic acid. The three compounds we assayed in vitro also reduced M. phaseolina colony forming unit counts in soil and the EC50 values were 476 ppmw ferulic acid, 612 ppmw juglone, and 827 ppmw p-Coumaric acid. Metconazole, the conventional fungicide control, did not inhibit M. phaseolina colony forming unit counts in soil at its label high rate. The plant compounds required similar or lower rates to inhibit colony forming units that grew from M. phaseolina overwintering structures (microsclerotia) in soil as to inhibit radial mycelial growth in vitro. Based on the EC50 value in soil assays, ferulic acid was the least expensive plant compound to apply on a per acre basis to inhibit M. phaseolina ($74,226). In F.oxysporum f.sp. fragariae soil assays, the compounds induced hormesis at lower rates and may be germination stimulant candidates. Metconazole and the high rates of every compound effectively or completely inhibited F. oxysporum f.sp. fragariae colony forming units in soil. The plant compounds were more herbicidal than fungicidal in vitro. When combining the in vitro seedling length results for L. sativa, M. parviflora, P. annua, and S. vulgaris the EC50 values differed significantly (p < .0001) and were: 47 ppmw juglone, 120 ppmw p-Coumaric acid, 189 ppmw ferulic acid, and 297 ppmw gallic acid. At least one rate of ferulic acid, juglone, and p-Coumaric acid inhibited the germination of all plant species, while gallic acid only inhibited the germination of P. annua at 1000 ppmw (p < .05). In soil, visible microbial contamination in individual wells of 24-well plates and seed dormancy made it difficult to fit curves to weed seedling length data. The soil assay L. sativa seedling length EC50 values 11 days after initial treatment were slightly higher than in vitro, although plant compounds were in the same order of phytotoxicity: 129 ppmw juglone, 616 ppmw p-Coumaric acid, 644 ppmw ferulic acid, and 1584 ppmw gallic acid. Based on the EC50 value in soil assays, the least expensive compound to inhibit L. sativa seedling length on a per acre basis was gallic acid ($21,676). Germination 26 days after initial soil treatment generally declined in a dose-dependent manner for each compound. There was a direct relationship between plant compound rate and seedling damage in soil with the higher rates of all compounds, except p-Coumaric acid, inducing damage comparable to a conventional herbicide (pendimethalin or oxyfluorfen). Contaminated treatments appeared to be due to an interaction between plant compounds and microorganisms because herbicide and water controls had almost no microbial growth 11 days after initial treatment. Further, there was a significant positive linear relationship between level of contamination in phenolic acid-treated wells (ferulic acid, gallic acid, and p-Coumaric acid, p < .0001) and the in soil rate. This relationship was slightly negative in juglone soil treatments (p = .0167), which may be due to its greater antimicrobial activity than the phenolic acids. We propose that herbicidal effects in soil were due to the joint effect of the plant compounds themselves, and the microbial growth in wells. Microbial growth was either antagonistic or additive to the inhibitory action of the plant compounds. The plant compounds we assayed were inhibitory of emerging fungal pathogens in strawberry production and common annual strawberry field weeds. Evidence presented in this thesis correlates well with past research that not only found plant compounds to be herbicidal and fungicidal, but also described their modes-of-action (such as the production of reactive oxygen species that causes necrotic lesions on roots, and inhibition of glycolytic enzyme activity that prevents germination), and implicate plant compounds as carbon sources for a variety of microorganisms. Compound prices are currently exorbitant, but may decline as demand increases. Whether or not they provide effective pest control may depend on soil texture, organic matter, microbial diversity, and other edaphic factors.