ABSTRACT
Snake venoms are widely used as laboratory tools for studies of physiological, pharmacological and toxicological mechanisms. Venoms used here are rich sources of several classes of proteases that act on factors of the coagulation cascade, fibrinogenolysis and fibrinolysis, altering the hemostatic processes, and phospholipases A2 which are involved mainly in inflammatory and clotting processes. Natural products such as essential oils are made up of active ingredients with wide application in the food, pharmaceutical and cosmetic industries. Thus, this study evaluates the essential oils from Mentha viridis and Mentha pulegium on coagulation, fibrinogenolysis and degradation of azocasein, induced by Bothrops sp and Lachesis muta muta venoms. These oils were achieved by hydrodistillation and presented, respectively, as the main constituents linalool (40.70 %), carvone (13.52 %) and α-terpinene (8.56 %); pulegone (50.01 %), menthol (31.90 %) and menthone (16.56 %). The essential oils were previously incubated with Bothrops alternatus venom, for two different times, then plasma was added and time was recorded. The M. viridis oil presented greater anticlotting potential. Meanwhile, the M. pulegium oil presented anticlotting or proclotting activity dependent on the dose tested. The incubation time also influenced the effect of the oils on the coagulation time. At azocaseinolytic assay, the oil from M. pulegium reduced the activity for all evaluated venoms. The highest inhibition was of 39.99 %; on activity induced by B. jararacussu, M. viridis reduced the activity in 57.72 %. On B. moojeni, the major inhibition observed was of 74.67 %. The fibrinogenolysis induced by B. moojeni venom was totally inhibited by both oils in the evaluated proportions. The results show the presence in oils of protease inhibitors, considering serine and metalloproteases (acting on clotting factors or with hemorrhagic activity), as well as phospholipase A2 (enzymes involved in inflammation and clotting processes) inhibitors of wide application in medical and biotechnology areas.
Los venenos de serpientes son ampliamente utilizados como herramientas de laboratorio para estudios de mecanismos fisiológicos, farmacéuticos y toxicológicos. Los venenos son recursos ricos en diferentes clases de proteasas que actúan sobre los factores de la cascada de coagulación, fibrinogenólisis y fibrinólisis, alterando los procesos homeostáticos y las fosfolipasas A2, los cuales están involucrados en procesos inflamatorios y de coagulación. Los productos naturales como los aceites esenciales se componen de ingredientes activos de amplia aplicación en las industrias alimentaria, farmacéutica y cosmética. El presente estudio evalúa los aceites esenciales de Mentha viridis y Mentha pulegium sobre la coagulación, fibrinogenólisis y degradación de azocaseína, inducida por los venenos de Bothrops sp. y Lachesis muta muta. Los aceites esenciales fueron obtenidos por hidrodestilación y sus componentes principales fueron identificados respectivamente como el linalool (40.70 %), carvona (13.52 %) y α-terpineno (8.56 %); pulegone (50.01 %), mentol (31.9 %) y mentona (15.56 %). Los aceites esenciales fueron previamente incubados con veneno de Bothrops alternatus, durante dos tiempos diferentes, luego se agregó el plasma y fue registrado el tiempo. El aceite de M. viridis presentó el más grande potencial anticoagulante. Mientras tanto, el aceite de M. pulegium presentó actividad anticoagulante o procoagulante dependiendo de la dosis evaluada. El tiempo de incubación también influyó en el efecto de los aceites sobre el tiempo de coagulación. En el ensayo azocaseínolitico, el aceite de M. pulegium redujo la actividad para todos los venenos evaluados. La inhibición más alta inducida para el veneno de B. jararacussu fue de 39.99 %, para M. viridis la actividad fue reducida en 57.72 %, mientras que para B. moojeni, la mayor inhibición observada fue de 74.67 %. La fibrinogenólisis inducida por el veneno de B. moojeni fue totalmente inhibida por ambos aceites en las proporciones evaluadas. Los resultados muestran la presencia de inhibidores de proteasas en los aceites, como las serinas y metaloproteasas (actuando sobre los factores de coagulación o sobre la actividad hemorrágica), como también fosfolipasas A2 (enzimas involucradas en los procesos de inflamación y coagulación), los cuales son inhibidores de amplia aplicación en áreas médicas y biotecnológicas.
ABSTRACT
Most of the snakebites recorded in Brazil are caused by the Bothrops genus. Given that the local tissue damage caused by this genus cannot be treated by antivenom therapy, numerous studies are focusing on supplementary alternatives, such as the use of medicinal plants. Serjania erecta has already demonstrated anti-inflammatory, antiseptic and healing properties. In the current study, the aerial parts of S. erecta were extracted with methanol, then submitted to chromatographic fractionation on a Sephadex LH20 column and eluted with methanol, which resulted in four main fractions. The crude extract and fractions neutralized the toxic activities of Bothrops jararacussu snake venom and isolated myotoxins (BthTX-I and II). Results showed that phospholipase A2, fibrinogenolytic, myotoxic and hemorrhagic activities were inhibited by the extract. Moreover, the myotoxic and edematous activities induced by BthTX-I, and phospholipase A2 activity induced by BthTX-II, were inhibited by the extract of S. erecta and its fraction. The clotting time on bovine plasma was significantly prolonged by the inhibitory action of fractions SF3 and SF4. This extract is a promising source of natural inhibitors, such as flavonoids and tannins, which act by forming complexes with metal ions and proteins, inhibiting the action of serineproteases, metalloproteases and phospholipases A2.
Subject(s)
Animals , Male , Mice , Bothrops , Plant Extracts/antagonists & inhibitors , Plants, Medicinal , Crotalid Venoms/toxicity , AntiveninsABSTRACT
Los extractos vegetales constituyen una fuente rica de moléculas farmacológicamente activas, cuya aplicación en medicina tradicional permite un acercamiento a potenciales actividades biológicas. En este trabajo se evalúa la capacidad inhibitoria de extractos etanólicos de hojas, raicillas y fracciones obtenidas por cromatografía en columna de Renealmia alpinia (Rottb) Mass, cultivada in vitro, sobre los efectos hemolítico indirecto, proteolítico y coagulante inducidos por el veneno de Bothrops asper. La actividad hemolítica indirecta es inhibida en mayor medida por la fracción 7-8 (47,3 ± 2,20%), seguida en su orden por los extractos de raicillas (32,6 #177; 6,90%) y hojas (24,2 ± 4,43%) de origen in vitro y hojas ex vitro (16,2 ± 3,88%). La actividad proteolítica se inhibe ampliamente por los extractos de hojas tanto in vitro como ex vitro sin diferencias significativas. Contra la actividad coagulante se registra una mayor neutralización por parte de las raicillas in vitro (81,73 ± 9,94s). Se descarta un potencial mecanismo de acción proteolítico de Renealmia alpinia sobre el veneno de Bothrops asper dado que no se producen cambios en los patrones electroforéticos del veneno. Los resultados viabilizan la aplicación de Renealmia alpinia como coadyuvante en el tratamiento del accidente ofídico y sustentan la utilidad de la micropropagación para la producción masiva de componentes activos.
Plant extracts are a rich source of pharmacologically active molecules. Their application allows a tradicional medicine approach to potential biological acitivities. This paper evaluates the inhibitory capacity of ethanolic extracts of leaves and little roots and also fractions chromatographically fractions obtained from Renealmia alpinia (Rottb) Mass, cultured in vitro, on the indirect hemolytic activity, proteolytic activity and coagulant activity induced by the Bothrops asper venom. Indirect hemolytic activity is inhibited to a greater extent by the fraction 7-8 (47.3 ± 2.20%) followed in order by the extracts from little roots (32.6 ± 6.90%) and leaves (24.2 ± 4.43%). They came from in vitro and ex vitro leaves (16.2 ± 3.88%). The proteolytic activity is largely inhibited by the leaves extracts in vitro and ex vitro without significant differences between them. Little roots in vitro showed the highest neutralization effect on coagulant activity (81.73 ± 9.949s). Proteolytic activity from Renealmia alpinia extracts on Bothrops asper venom is ruled out since there are not changes in the electrophoretic pattern of the venom. The results make possible the implementation of Renealmia alpinia as adjuvant for the treatment of ophidic accidents and sustain the value of micropropagation for mass production of active components.