Applications of the hexanic fraction of Agave sisalana Perrine ex Engelm (Asparagaceae): control of inflammation and pain screening

The present study evaluated the anti-inflammatory and analgesic properties of Agave sisalana Perrine in classic models of inflammation and pain. The hexanic fraction of A. sisalana (HFAS) was obtained by acid hydrolysis followed by hexanic reflux. Anti-inflammatory properties were examined in three acute mouse models (xylene ear oedema, hind paw oedema and pleurisy) and a chronic mouse model (granuloma cotton pellet). The antinociceptive potential was evaluated in chemical (acetic-acid) and thermal (tail-flick and hot-plate test) models of pain. When given orally, HFAS (5, 10, 25 and 50 mg/kg) reduced ear oedema (p < 0.0001; 52%, 71%, 62% and 42%, respectively). HFAS also reduced hind paw oedema at doses of 10 mg/kg and 25 mg/kg (p < 0.05; 42% and 58%, respectively) and pleurisy at doses of 10 mg/kg and 25 mg/kg (41% and 50%, respectively). In a chronic model, HFAS reduced inflammation by 46% and 58% at doses of 10 mg/kg and 25 mg/kg, respectively. Moreover, this fraction showed analgesic properties against the abdominal writhing in an acetic acid model (at doses of 5-25 mg/kg) with inhibitory rates of 24%, 54% and 48%. The HFAS also showed an increased latency time in the hot-plate (23% and 28%) and tail-flick tests (61% and 66%) for the 25 mg/kg and 50 mg/kg doses, respectively. These results suggest that HFAS has anti-inflammatory and analgesic properties.26327

Estudo dos papeis biologicos de dois inibidores de Kunitz isolados de sementes de Dimorphandra mollis BENTH: atividades inseticida e inflamatoria

Orientador: Maria Ligia Rodrigues MacedoTese (doutorado) - Universidade Estadual de Campinas, Instituto de BiologiaResumo: Os inibidores de proteinases desempenham as mais variadas funções biológicas. Em plantas, eles podem estar envolvidos no mecanismo de defesa vegetal contra predadores e doenças. Devido à habilidade dessas moléculas protéicas bloquearem enzimas, os inibidores também podem ser usados como ferramentas no estudo de processos bioquímicos relacionados a patologias, tais como a inflamação, hemorragia e até mesmo o câncer. Neste trabalho, a caracterização físico-química e o estudo dos papéis biológicos de dois inibidores isolados e purificados de sementes de Dimorphandra mollis, DMTI (Mace do et ai., 2000) e DMTI-II (Mello et aI., 2001) foram realizadas com o objetivo de melhor conhecer essas moléculas. DMTI (20 kDa) e DMTI-II (23kDa) são inibidores de tripsina pertencentes à família de Kunitz. A estabilidade da atividade inibitória de DMTI e DMTI-II foi analisada e comparada. DMTI e DMTI-II perde, respectivamente, cerca de 20% e 40% da atividade quando incubados a 60 °e por 20 minutos. Entretanto, a atividade de ambos não foi afetada por diferentes condições de pH, concentrações crescentes dos agentes redutores DTT e uréia. No entanto, a atividade DMTI-II sofreu um descréscimo de 50% quando incubado com cloreto de guanidina. A diferença da estabilidade dos inibidores levou a investigação da estrutura protéica de ambos; DMTI e DMTI-II foram submetidos a análises de dicroísmo circular e calorimetria. A ação inseticida de DMTI-II e DMTI sobre os lepidópteros Anagasta kuehniella e Corcyra cephalonica, foram analisadas através de ensaios in vitro e in vivo. Tanto DMTI-II quanto DMTI, quando incorporados ao nível de 2% na dieta artificial dos insetos, não se mostraram agentes inseticidas potenciais contra essas duas espécies. Estes resultados não descartam a utilização dos inibi dores de D. mollis na biotecnologia vegetal contra outras espécies de insetos, apenas ajudam a esclarecer as características como agentes inseticidas biológicos. Quando estudado o envolvimento de DMTI-II e DMTI na resposta inflamatória utilizando o edema de pata em ratos como modelo experimental, os resultados mostraram que doses crescentes de DMTI-II geram uma curva dose-dependente sendo que a resposta máxima ocorre 30 minutos após a sua injeção subplantar, o que sugeriu a ação do inibidor na liberação de histamina e serotonina, que são responsáveis pelo aumento da penneabilidade vascular. A elucidação do mecanismo de ação de DMTI-Il foi estudada através da utilização de drogas antiinflamatórias, como a mepiramina e ciproheptadina; sendo esta última a que melhor inibiu a ação de DMTI-Il na indução do edema. Concomitantemente, a sua ação na liberação de bradicinina também pode estar ocorrendo, pois o pré-tratamento com o antagonista de receptores B 2, HOE 140, reduziu cerca de 79% do edema induzido por DMTI-IIAbstract: Proteinase inhibitors have a wide range of biological functions. In plants, they¿re part of the defense mechanism against predators and diseases. As they can block enzymes, inhibitors are useful tools in the study of biochemical processes related to pathologies, such as inflammation, hemorrhage and even cancer. This paper aims at assessing the physical-chemical features and the biological functions of two inhibitors isolated and purified trom Dimorphandra mollis seeds, DMTI (Macedo et al., 2000) and DMTI-II (Mello et aI., 2001). DMTI (20 kDa) and DMTI-TI (23 kDa) are trypsin inhibitors ITom the Kunitz family. The stability of the inhibitory activity of DMTI and that ofDMTI-II were analyzed and compared to each other. DMTI and DMTI-II lost 20% and 40%, respectively, oftheir activities when incubated at 60°C for 20 minutes. However, neither inhibitory activity was sensitive to pH above the 2.0-10.0 range, reducing DTT agents and urea. On the other hand, DMTI-II activity was reduced by 50% when incubated with guanidine hydrochloride. The difference of stability between the inhibitors led to further investigation of their protein structure; DMTI and DMTI-II structures were analyzed by circular dichroism (CD) and differential scanning calorimetry (DSC). The effectiveness of DMTI-II and DMTI as insecticide in Lepidoptera insects, Anagasta kuehniella and Corcyra cephalonica, was analyzed in vitro and in vivo assays. Neither DMTI-II nor DMTI, when incorporated into artificial diet at 2%, were active as insecticide agents against these two species. Nevertheless, these results do not discard the use of inhibitors trom D. mollis in plant biotechnology against other species of insects; they just help us to understand how they work as biological agents. When studying DMTI and DMTI-II participation in inflammatory response in the rat paw edema experimental model, the results showed that increased doses of DMTI-II induced a dose dependent curve, and maximum response occurred 30 min after sub-plantar injection, which indicated that the inhibitor is active in the release of histamine and serotonin, which are responsible for the increase of vascular permeability. The elucidation of DMTI-II action mechanism was studied by using anti-inflammatory drugs, such as mepyramine and cyproheptadine, the latter the most effective against DMTI-II effects in edema induction. At the same time, it may also be active in bradykinin release as the pretreatment with B2 receptor antagonist HOE 140 reduced the DMTI-II induced edema by near1y 79%DoutoradoBioquimicaDoutor em Biologia Funcional e Molecula

Anti-inflammatory and antinociceptive effects of racemic goniothalamin, a styryl lactone

CNPQ – CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICOAims: The present study aimed to further investigate the anti-inflammatory activity of goniothalamin (GTN), a styryl lactone, as well as its antinociceptive effects. Main methods: The anti-inflammatory activity was evaluated in models of paw edema induced by different mediators in mice and carrageenan-induced peritonitis. Evaluation of the antinociceptive effect was performed through acetic acid-induced writhing test and formalin test. Activity of GTN on gene expression levels of interleukin-1beta (IL-1 beta), induced nitric oxidase synthase (iNOS) and cyclooxygenase-2 (COX-2) were evaluated in vitro in lipopolysaccharide (LPS)-stimulated macrophage (RAW 264.7), as well as gene expression and protein levels of tumor necrosis factor-alpha (TNF-alpha). Key findings: Pretreatment with GIN (300 mg/kg) significantly reduced paw edema induced by compound 48/80, prostaglandin E-2, phospholipase A(2) and bradyldnin. GIN (10,30 and 100 mg/kg) inhibited leukocyte migration in the peritonitis model and gene expression levels of IL-1 beta, iNOS and TNF-alpha, as well as TNF-alpha protein levels, in LPS-stimulated macrophages, without affecting COX-2 gene expression levels. GIN inhibited nociception induced by acetic acid in the writhing model and in the formalin test, when both neurogenic and inflammatory phases were inhibited. Significance: For the first time the acute anti-inflammatory profile of GTN is characterized and its antinociceptive activity reported. The current study shows that GTN inhibits both vascular and cellular phases of inflammation, with bradykinin and PLA(2) induced inflammation being the most affected by GIN. Its anti-inflammatory effects also involved the in vitro inhibition of gene expression of alarm cytokines and mediators as IL-1 beta, iNOS and TNF-alpha. (C) 2015 Elsevier Inc. All rights reserved.The present study aimed to further investigate the anti-inflammatory activity of goniothalamin (GTN), a styryl lactone, as well as its antinociceptive effects. Main methods: The anti-inflammatory activity was evaluated in models of paw edema induced by different mediators in mice and carrageenan-induced peritonitis. Evaluation of the antinociceptive effect was performed through acetic acid-induced writhing test and formalin test. Activity of GTN on gene expression levels of interleukin-1beta (IL-1 beta), induced nitric oxidase synthase (iNOS) and cyclooxygenase-2 (COX-2) were evaluated in vitro in lipopolysaccharide (LPS)-stimulated macrophage (RAW 264.7), as well as gene expression and protein levels of tumor necrosis factor-alpha (TNF-alpha). Key findings: Pretreatment with GIN (300 mg/kg) significantly reduced paw edema induced by compound 48/80, prostaglandin E-2, phospholipase A(2) and bradyldnin. GIN (10,30 and 100 mg/kg) inhibited leukocyte migration in the peritonitis model and gene expression levels of IL-1 beta, iNOS and TNF-alpha, as well as TNF-alpha protein levels, in LPS-stimulated macrophages, without affecting COX-2 gene expression levels. GIN inhibited nociception induced by acetic acid in the writhing model and in the formalin test, when both neurogenic and inflammatory phases were inhibited. Significance: For the first time the acute anti-inflammatory profile of GTN is characterized and its antinociceptive activity reported. The current study shows that GTN inhibits both vascular and cellular phases of inflammation, with bradykinin and PLA(2) induced inflammation being the most affected by GIN. Its anti-inflammatory effects also involved the in vitro inhibition of gene expression of alarm cytokines and mediators as IL-1 beta, iNOS and TNF-alpha1398390CNPQ – CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICOCNPQ – CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICOFAPESP [07/58665-7, 09/51602-5, 2012/18281-3]CNPq [307365/2013-1]sem informaçã

Applications of the hexanic fraction of Agave sisalana Perrine ex Engelm (Asparagaceae): control of inflammation and pain screening

The present study evaluated the anti-inflammatory and analgesic properties of Agave sisalana Perrine in classic models of inflammation and pain. The hexanic fraction of A. sisalana (HFAS) was obtained by acid hydrolysis followed by hexanic reflux. Anti-inflammatory properties were examined in three acute mouse models (xylene ear oedema, hind paw oedema and pleurisy) and a chronic mouse model (granuloma cotton pellet). The antinociceptive potential was evaluated in chemical (acetic-acid) and thermal (tail-flick and hot-plate test) models of pain. When given orally, HFAS (5, 10, 25 and 50 mg/kg) reduced ear oedema (p < 0.0001; 52%, 71%, 62% and 42%, respectively). HFAS also reduced hind paw oedema at doses of 10 mg/kg and 25 mg/kg (p < 0.05; 42% and 58%, respectively) and pleurisy at doses of 10 mg/kg and 25 mg/kg (41% and 50%, respectively). In a chronic model, HFAS reduced inflammation by 46% and 58% at doses of 10 mg/kg and 25 mg/kg, respectively. Moreover, this fraction showed analgesic properties against the abdominal writhing in an acetic acid model (at doses of 5-25 mg/kg) with inhibitory rates of 24%, 54% and 48%. The HFAS also showed an increased latency time in the hot-plate (23% and 28%) and tail-flick tests (61% and 66%) for the 25 mg/kg and 50 mg/kg doses, respectively. These results suggest that HFAS has anti-inflammatory and analgesic properties