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Avaliação do perfil antiinflamatório e antinociceptivo da casca do caule de Aspidosperma tomentosum (Apocynaceae). / Evaluation of Antiinflamatory and antinociceptive profile of the stem bark of Aspidosperma tomentosum (Apocynaceae).Aquino, Anansa Bezerra de 12 March 2010 (has links)
In this study we evaluated the antinociceptive and anti-inflammatory activities of
the stem bark of the species Aspidosperma tomentosum in models of pain and
inflammation induced in mice. A A. tomentosum belongs to the family Apocynaceae
that has been extensively studied and characterized chemically by the frequent
occurrence of alkaloids, some of therapeutic importance, such as vinblastine and
vincristine. This species is known as pau-pereiro do campo and was collected in the
Brazilian cerrado, in the city of Planaltina the state of Goiás, in May 2004. We assessed
the antinociceptive and anti-inflammatory activity of crude ethanol extract obtained by
extraction with 20 L of 90% ethanol, fractions obtained by chromatography adapted,
according to a solvent gradient of increasing polarity, and the flavonoid isorhamnetin,
obtained in the filtration of crude ethanolic extract of stem bark of A. tomentosum. We
conducted the following tests: test of writhing induced by acetic acid, hot plate,
formalin, ear edema induced by capsaicin, peritonitis induced by thioglycollate 3%.
Was also evaluated motility and / or depression of the central nervous system by the test
of catalepsy. For statistical analysis we used ANOVA followed by Dunnet test, the
tutorial Prisma ®. Values were considered significant when * p <0.05, ** p <0.01 and
*** p <0.001. The results were expressed as mean ± standard error of the mean. In the
abdominal writhing test induced by acetic acid, ethanol extract and fractions showed
higher inhibition between 50.8% to 59.7%, compared with dipyrone. In the hot plate
test, the fractions Hex: CHCl3 50%, 100% CHCl3 and CHCl3: MeOH 5% showed an
increase in latency of the animal on the hot plate, and this result was reversed in the
presence of naloxone. In the test of nociception induced by formalin, the crude ethanol
extract and its fractions presented significant results in both phases of testing, when
compared to control, and the CHCl3 100% fraction presented better performance in
both phases (average latency time of 61s and 75s the first and second phases,
respectively). In the test of ear edema induced by capsaicin only the soluble fraction of
crude ethanol extract, the fraction of CHCl3: MeOH 10% and isorhamnetin showed no
inhibition of edema, where the CHCl3: AcOEt 50% was the most active (64.2% of
inhibition of edema). In the trial of peritonitis the crude ethanol extract, the
isorhamnetin and the fractions used inhibited cell migration, except the fraction AcOEt
100%. The test result of catalepsy suggests that treatment with the fractions Hex:
CHCl3 50%, 100% CHCl3 and CHCl3: MeOH 5% of the isolated species A.
tomentosum not alter the motor skills of the animal. The results observed in this study
indicate that the species A. tomentosum is able to modulate the antinociceptive and antiinflammatory
response. / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / No presente trabalho foram avaliadas as atividades antinociceptiva e
antiinflamatória da casca do caule da espécie Aspidosperma tomentosum, em modelos
de dor e inflamação induzidas em camundongos. A A. tomentosum pertence à família
Apocynaceae que vem sendo extensivamente estudada e caracteriza-se quimicamente
pela ocorrência frequente de alcaloides, alguns de importância terapêutica, como
vimblastina e vincristina. A espécie é conhecida como pau-pereiro do campo e foi
coletada no cerrado brasileiro, na cidade de Planaltina no estado de Goiás, em maio de
2004. Foram avaliados as atividades antinociceptiva e antiinflamatória do extrato
etanólico bruto, obtido por extração com 20 L de etanol 90%, das frações, obtidas por
cromatografia rápida em funil de separação adaptado, com solventes segundo um
gradiente crescente de polaridade, e do flavonoide isorramnetina, obtido na filtração do
extrato etanólico bruto de casca do caule da espécie A. tomentosum. Foram realizados os
seguintes testes: teste das contorções abdominais induzidas por ácido acético, placa
quente, formalina, edema de orelha induzido por capsaicina, peritonite induzida por
tioglicolato a 3%. Foi também avaliada alteração motora e/ou depressão do sistema
nervoso central pelo ensaio de catalepsia. Para a análise estatística utilizou-se ANOVA
seguida do teste Dunnet, no tutorial Prisma®. Os valores foram considerados
significativos quando *p < 0,05, **p < 0,01 e ***p < 0,001. Os resultados foram
expressos como média ± erro padrão da média. No ensaio de contorção abdominal
induzida por ácido acético, o extrato etanólico e frações apresentaram uma taxa de
inibição entre 50,8% a 59,7%, comparados com a dipirona. No ensaio de placa quente,
as frações Hex:CHCl3 50%, CHCl3 100% e CHCl3:MeOH 5% apresentaram aumento no
tempo de latência do animal na placa quente, sendo esse resultado revertido na presença
da naloxona. No ensaio de nocicepção induzida por formalina, o extrato etanólico bruto
e suas frações apresentaram resultados significativos nas duas fases do ensaio, quando
comparados ao controle, sendo a fração CHCl3 100% com melhor desempenho nas duas
fases (tempo de latência médio de 61s e 75s na primeira e segunda fase,
respectivamente). No teste de edema de orelha induzida por capsaicina apenas a fração
solúvel do extrato etanólico bruto, a fração CHCl3:MeOH 10% e a isorramnetina não
apresentaram inibição do edema, onde o CHCl3:AcOEt 50% foi o mais ativo (64,2% de
inibição do edema). No ensaio de peritonite o extrato etanólico bruto, a isorramnetina e
as frações utilizadas inibiram a migração celular, exceto a fração AcOEt 100%. O
resultado do ensaio de catalepsia sugere que o tratamento com as frações Hex:CHCl3
50%, CHCl3 100% e CHCl3:MeOH 5% obtidas da espécie A. tomentosum não alteram a
capacidade motora do animal. Os resultados observados neste estudo indicam que
espécie A. tomentosum é capaz de modular a resposta antinociceptiva e antiinflamatória
aguda.
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Regeneration ecology of anemochorous tree species Qualea grandiflora (Mart.) and Aspidosperma tomentosum (Mart.) of the cerrado Aguara Ñu located in the Mbaracayú Nature Forest Reserve (MNFR), ParaguayDa Ponte Canova, Giovanna 14 February 2019 (has links)
Understanding of the diverse aspects affecting the regeneration ecology of species is crucial to make decisions regarding management and conservation strategies, especially in highly fragile and threatened ecosystem as the Neotropical savanna (cerrado) formations. Available knowledge of regeneration ecology of cerrado species is too limited to attain optimal or suitable management actions. The objectives of the present study were: i) analysis of the characteristic parameters of the anemochorous seed dispersal of study species Q. grandiflora, (ii) determine the spatial distribution of tree species Q. grandiflora for growth stages (seedlings to juveniles) and interrelations between the stages, and (iii) determining variables of the spatial distribution of recruitment of tree species A. tomentosum.
The present study was conducted in the cerrado Aguara Ñu of the Mbaracayú Nature Forest Reserve located in the northeast of Paraguay. The cerrado Aguara Ñu is part of the Mbaracayú Biosphere Reserve and represents one of the most important ecoregions in the world, the cerrado ecosystem. The cerrado biome encompasses areas from northeastern to southwestern Brazil, eastern Bolivia, and northern Paraguay. It is characterized by the presence of high plant and animal biodiversity and also high endemism (Myers et al., 2000). Tree species Q. grandiflora and A. tomentosum are typical species of the cerrado formation.
Based on the selected investigated regeneration cycle stages of study tree species Q. grandiflora and A. tomentosum, the present thesis describes the spatial analysis of recruitment of both study species and the anemochorous diaspore dispersal of tree species Q. grandiflora. The purpose of the present investigation is to address regeneration aspects not attained so far as certain seed dispersal aspects, such as seed densities and distances from conspecific adult trees and spatial arrangements of seedlings of species A. tomentosum. Results of the present study aim to contribute to existing information and at the same time provide new knowledge on ecological aspects so far not investigated.
Research results on seed dispersal of tree species Q. grandiflora revealed that dispersal can be modeled by inverse modelling considering isotropy and lognormal density function presenting mean dispersal distances of 10.69 to 62.48 m. Estimations of the fruit production of a seed tree yielded a total 50671 to 70632 (DBH = 70 cm). Results of spatial arrangement of seedlings and juveniles revealed a significant distance effect to conspecific adult trees. Moreover, results also showed: (i) highest densities or intensities (m2) of seedlings (heights <50 cm) close to the conspecific adult trees and (ii) a shift of intensity of seedlings with increase of growth stage or size for tree species Q. grandiflora. Additionally, seedlings (up to 200 cm height) of study species Q. grandiflora indicated gradual decreasing clumping patterns and juveniles (200 – 500 cm height) presented clumping patterns.
Modelling results of spatial patterns of seedlings (heights ≤ 200 cm) of study tree species A. tomentosum revealed aggregation patterns. Moreover, shade effect resulted to be a statistical significant factor for the establishment of seedlings of tree species A. tomentosum (p-value = 0.0266), whereas distance effect to seed tree resulted not significant (p-value= 0.4936).
Considering the findings of seed dispersal and spatial patterns analysis of tree species Q. grandiflora and A. tomentosum some management aspects to be attained for conservation purposes are avoiding fragmentation of the ecosystem, management of the spatial and time fire frequency and maintain minimum amount of seed trees per unit area in order to guarantee successful recruitment.:1. Introduction 1
References 8
2. Materials and Methods 13
2.1 Characterization of the cerrado biome 13
2.2 Description of the study area and study sites 15
2.3 Characterization of the study tree species 23
2.3.1 Qualea grandiflora (Mart.) 23
2.3.2 Aspidosperma tomentosum (Mart.) 24
2.4 Principles and selection criteria 25
2.5 Data collection 26
2.5.1 Seed dispersal 26
2.5.2 Spatial patterns of plants 27
2.6 Data analysis 28
2.6.1 Statistical analysis of data 28
2.6.2 General statistical procedures of data analysis 30
2.6.3 Spatial point process analysis – Inverse modelling and spatial point patterns 31
2.6.4 Spatial point patterns analysis procedure 33
2.6.4.1 Descriptive statistics in spatial point patterns 36
2.6.4.1.1 Distance effect of seedlings from seed trees (rhohat function) 36
2.6.4.1.2 Pair correlation function (pcf) 36
2.6.4.2 Point process modelling 38
References 43
3. Seed dispersal of Qualea grandiflora (Mart.) 49
3.1 Introduction 49
3.2 Methodology 51
3.2.1 Data collection and seed trap design 51
3.2.1 Data analysis – inverse modelling 53
3.3 Results 58
3.3.1 Seed density 58
3.3.2 Inverse modelling results – seed production, dispersal and distances 60
3.3.2.1 Isotropic modelling 61
3.3.2.2 Anisotropic modelling 63
3.3.2.3 Statistical comparison isotropy vs. anisotropy 66
3.4 Discussion 67
3.4.1 Applied methodology for seed dispersal – trap design and inverse modelling 67
3.4.2 Seed dispersal modelling 69
3.5 Conclusion 74
References 75
4. Spatial analysis of Qualea grandiflora (Mart.) 80
4.1 Introduction 80
4.2 Methodology 82
4.2.1 Data collection – Field sampling 82
4.2.2 Data analysis 85
4.2.2.1 Spatial point pattern – Explorative analysis 85
4.2.2.2 Point process modelling (Poisson and Gibbs models) 87
4.2.3 Results 89
4.2.3.1 Spatial distribution of individuals of study species 89
4.2.3.2 Modelling distance effect of recruitment to adult trees 95
4.2.4 Discussion 102
4.2.4.1 Applied methodology for spatial analysis of study species 102
4.2.4.2 Spatial arrangement of study species 103
4.2.5 Conclusion 109
References 109
5. Spatial analysis of Aspidosperma tomentosum (Mart.)115
5.1 Introduction 115
5.2 Methodology 117
5.2.1 Data collection – Field sampling 117
5.2.2 Data analysis 120
5.2.2.1 Spatial point pattern – Explorative analysis 120
5.2.2.2 Point process modelling – Replicated point patterns 120
5.3 Results 123
5.3.1 Spatial distribution of natural regeneration of study species 123
5.3.2 Modelling shade and distance to seed tree effect on natural regeneration of study species 130
5.4 Discussion 133
5.4.1 Applied methodology for data collection and analysis 133
5.4.2 Spatial distribution of natural regeneration of study species 134
5.5 Conclusion 139
References 140
5. Concluding discussion and summary 146
6.1 Regeneration ecology of Qualea grandiflora and Aspidosperma tomentosum 146
6.1.1 Inferences on relation of seed dispersal and spatial distribution of recruitment of Qualea grandiflora 146
6.1.2 Inferences on spatial patterns of recruitment of Aspidoserma tomentosum 149
6.2 Management implications for Qualea grandiflora Aspidosperma tomentosum 150
6.3 Future research 153
6.4 Concluding summary 154
References 155
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Atividade antimicrobiana e estudo químico bioguiado de espécies de Aspidosperma / Antimicrobial activity and chemical study bioguided species AspidospermaPessini, Greisiele Lorena 25 September 2015 (has links)
Aspidosperma species belonging to the Apocynaceae family are restricted to the Americas. Ethnopharmacological studies have shown the use of these species as a potential agent against malaria and trypanosomatids, also used as antibiotics, anti-inflammatory and antitumor. The search for new chemical entities with antimicrobial activity has significant importance due to the versatility of microorganisms to acquire resistance to the therapeutic arsenal, and due to the prevalence of neglected diseases, such as leishmaniasis. These are endemic in developing countries have few therapeutic options, which are toxic and have limited effectiveness. The aim of this study was to evaluate the crude extracts and fractions from different parts of the species A. macrocarpon, A. tomentosum and A. pyrifolium, as the antimicrobial action and realize bioguided chemical study of the species with high activity against the microorganisms tested, and less toxic to mammalian cells. About antibacterial activity alkaloidal fraction from A. pyrifolium stem presented moderate minimal inhibitory concentration (MIC): 125 and 250 g/mL for Staphylococcus aureus and Bacillus subtilis, respectively. The ethyl acetate fraction of A. macrocarpon stem had a weak MIC of 250 g/mL on Candida parapsilosis. The activity of the extract of the stem bark of A. macrocapon against Leishmania amazonensis promastigotes showed inhibitory concentration (IC50) of 151.5 g/mL and selectivity index (SI) of 6.52 on LLCMK2 cells, this species were considered the most promising for the bioguided chemical study. The chloroform and ethyl acetate fractions of the A. macrocarpon stem bark showed IC50 29.00 1.65 and 29.50 0.95 g/mL, respectively on promastigotes forms of L. amazonensis. The bioguided study with chloroform fraction led to subfraction of monoacylglycerols, wich were the most active on L. amazonensis, with IC50 2.31 0.08 and 2.29 0.14 g/mL on promastigotes and intracellular amastigotes, respectively. SI showed that this fraction were 42 fold more toxic to L. amazonensis than to macrophages. Studies related to the mechanism of action of monoacylglycerols evaluated by optical microscopy, scanning electron (SEM) and transmission electron microscopy (TEM) showed abnormalities such as mitochondrial swelling, concentric membranes inside the mitochondria, presence of perforations on the cell surface of the parasite and autophagic vacuoles. These data together with results of flow cytometry showed necrosis and autophagy as a possible mechanism of cell death. Quercetin was identified as the active substance of the ethyl acetate fraction against evolutionary forms of L. amazonensis. This substance had, IC50 of 61.93 1.36 and 63.86 3.25 M/mL on promastigotes and intracellular amastigotes, respectively. Quercetin was evaluated in combination with amphotericin B (Q + A) and miltefosine (Q + M), the combinatorial effect of both was considered synergistic against all forms of L. amazonensis. The combinations Q+A and Q+M showed antagonistic action on macrophages and human erythrocytes. Evaluation of the combination using SEM and TEM associated with literature data indicate apoptotic cell death after treatment with both combinations. These findings are relevant as a first step in the search for new therapies for the treatment of cutaneous leishmaniasis. / Conselho Nacional de Desenvolvimento Científico e Tecnológico / As espécies do gênero Aspidosperma pertencentes à família Apocynaceae são restritas às Américas. Estudos etnofarmacológicos revelaram a utilização destas espécies como agente potencial contra a malária e tripanosomatídeos, como antimicrobianos, anti-inflamatórios e antitumorais. A pesquisa em busca de novas entidades químicas com ação antimicrobiana apresenta significativa importância devido à versatilidade dos micro-organismos em adquirir resistência ao arsenal terapêutico, e devido à prevalência de doenças consideradas negligenciadas, como as leishmanioses. Estas são endêmicas nos países em desenvolvimento e possuem poucas opções terapêuticas, sendo estas tóxicas e de eficácia limitada. O objetivo do estudo consistiu em avaliar os extratos brutos e frações de diferentes partes das espécies de A. macrocarpon, A. tomentosum e A. pyrifolium, quanto à ação antimicrobiana e realizar o estudo químico bioguiado da espécie mais ativa sobre os micro-organismos testados e menos tóxica para células de mamíferos. Sobre a atividade antibacteriana a fração alcaloídica da casca de A. pyrifolium apresentou concentração inibitória mínima (CIM) moderada de 125 e 250 μg/mL para Staphylococcus aureus e Bacillus subtilis, respectivamente. A fração acetato de etila do caule de A. macrocarpon apresentou CIM fraca de 250 μg/mL sobre Candida parapsilosis. A atividade do extrato da casca do caule de A. macrocapon contra formas promastigotas de Leishmania amazonensis apresentou concentração inibitória (CI50) de 151,5 g/mL e índice de seletividade (IS) de 6,52 sobre células LLCMK2, sendo a espécie mais promissora para a realização do estudo químico bioguiado. As frações de clorofórmio e acetato de etila obtidas da casca do caule de A. macrocarpon demonstraram CI50 de 29,00 1,65 e 29,50 0,95 g/mL respectivamente, sobre as formas promastigotas de L. amazonensis. O estudo bioguiado do fracionamento da fração clorofórmio conduziu à obtenção da subfração com monoacilglicerois (MAG) como a mais ativa sobre as formas evolutivas de L. amazonensis, com CI50 2,31 0,08 e 2,29 0,14 g/mL sobre promastigotas e amastigotas intracelulares, respectivamente e IS sobre macrófagos J774-A1 de 42, para ambas as formas evolutivas. Estudos relacionados ao mecanismo de ação da fração MAG avaliados por microscopia óptica, eletrônica de varredura (MEV), e transmissão (MET) evidenciaram alterações como inchaço mitocondrial, membranas concêntricas dentro da mitocôndria, presença de perfurações na superfície celular do parasito e vacúolos autofágicos. Estes dados quando analisados em conjunto com resultados da citometria de fluxo indicaram a autofagia e necrose como provável mecanismo de morte celular. Após o fracionamento da fração acetato de etila foi identificada a quercetina como substância ativa sobre as formas evolutivas de L. amazonensis, com CI50 de 61,93 1,36 e 63,86 3,25 M/mL sobre promastigotas e amastigotas intracelulares, respectivamente. A quercetina foi avaliada em combinação com anfotericina B (Q+A) e miltefosina (Q+M), o efeito combinatório de ambas foi considerado sinérgico sobre as formas evolutivas de L. amazonensis. As combinações Q+A e Q+M demonstraram ação antagônica sobre macrófagos e eritrócitos humanos. A avaliação da combinação por MEV e MET associada aos dados da literatura indicaram provável morte celular por apoptose de ambas as combinações. Estes achados nos ensaios in vitro são relevantes como primeiro passo na busca por novas terapias para o tratamento da leishmaniose cutânea.
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