Antinociceptive activity of 3ß-6ß-16ß-trihydroxylup-20 (29)-ene triterpene isolated from Combretum leprosum leaves in adult zebrafish (Danio rerio).

Drugs used to treat pain are associated with adverse effects, increasing the search for new drugs as an alternative treatment for pain. Therefore, we evaluated the antinociceptive behavior and possible neuromodulation mechanisms of triterpene 3ß, 6ß, 16ß-trihydroxylup-20(29)-ene (CLF-1) isolated from Combretum leprosum leaves in zebrafish. Zebrafish (n = 6/group) were pretreated with CLF-1 (0.1 or 0.3 or 1.0 mg/mL; i.p.) and underwent nociception behavior tests. The antinociceptive effect of CFL-1 was tested for modulation by opioid (naloxone), nitrergic (L-NAME), nitric oxide and guanylate cyclase synthesis inhibitor (methylene blue), NMDA (Ketamine), TRPV1 (ruthenium red), TRPA1 (camphor), or ASIC (amiloride) antagonists. The corneal antinociceptive effect of CFL-1 was tested for modulation by TRPV1 (capsazepine). The effect of CFL-1 on zebrafish locomotor behavior was evaluated with the open field test. The acute toxicity study was conducted. CLF-1 reduced nociceptive behavior and corneal in zebrafish without mortalities and without altering the animals' locomotion. Thus, CFL-1 presenting pharmacological potential for the treatment of acute pain and corneal pain, and this effect is modulated by the opioids, nitrergic system, NMDA receptors and TRP and ASIC channels.

Antimicrobial activity of the lupane triterpene 3ß,6ß,16ß-trihydroxylup-20(29)-ene isolated from Combretum leprosum Mart.

Introduction. Combretum leprosum (Combretaceae) is commonly found in the Northeast Region of Brazil and is known for several bioactivities, including antimicrobial ones. Because of increasing bacterial antibiotic resistance, natural products from several plants have been studied as putative adjuvants to antibiotic activity, including products from C. leprosum. Aims. This study was carried out to investigate the structural properties, bactericidal activity and antibiotic modifying action of the lupane triterpene 3ß,6ß,16ß-trihydroxylup-20(29)-ene (CLF1) isolated from C. leprosum Mart. leaves.Methods. The CLF1 was evaluated by the Fourier transform infrared spectroscopy method and the antibacterial activity of this compound was assayed alone and in association with antibiotics by microdilution assay.Results. Spectroscopic studies confirmed the molecular structure of the CLF1 and permitted assignment of the main infrared bands of this natural product. Microbiological assays showed that this lupane triterpene possesses antibacterial action with clinical relevance against Staphylococcus aureus. The CLF1 triterpene increased antimicrobial activity against the multidrug-resistant Escherichia coli 06 strain when associated with the antibiotics gentamicin and amikacin. Synergistic effects were observed against the S. aureus 10 strain in the presence of the CLF1 triterpene with the antibiotic gentamicin.Conclusion. In conclusion, the CLF1 compound may be useful in the development of antibacterial drugs against the aforementioned bacteria.

A lupane-triterpene isolated from Combretum leprosum Mart. fruit extracts that interferes with the intracellular development of Leishmania (L.) amazonensis in vitro.

BACKGROUND: 3beta,6beta,16beta-trihydroxylup-20(29)-ene is a lupane triterpene isolated from Combretum leprosum fruit. The lupane group has been extensively used in studies on anticancer effects; however, its possible activity against protozoa parasites is yet poorly known. The high toxicity of the compounds currently used in leishmaniasis chemotherapy stimulates the investigation of new molecules and drug targets for antileishmanial therapy. METHODS: The activity of 3beta,6beta,16beta-trihydroxylup-20(29)-ene was evaluated against Leishmania (L.) amazonensis by determining the cytotoxicity of the compound on murine peritoneal macrophages, as well as its effects on parasite survival inside host cells. To evaluate the effect of this compound on intracellular amastigotes, cultures of infected macrophages were treated for 24, 48 and 96 h and the percentage of infected macrophages and the number of intracellular parasites was scored using light microscopy. RESULTS: Lupane showed significant activity against the intracellular amastigotes of L. (L.) amazonensis. The treatment with 109 µM for 96 h reduced in 80 % the survival index of parasites in BALB/c peritoneal macrophages. At this concentration, the triterpene caused no cytotoxic effects against mouse peritoneal macrophages. Ultrastructural analyses of L. (L.) amazonensis intracellular amastigotes showed that lupane induced some morphological changes in parasites, such as cytosolic vacuolization, lipid body formation and mitochondrial swelling. Bioinformatic analyses through molecular docking suggest that this lupane has high-affinity binding with DNA topoisomerase. CONCLUSION: Taken together, our results have showed that the lupane triterpene from C. leprosum interferes with L. (L.) amazonensis amastigote replication and survival inside vertebrate host cells and bioinformatics analyses strongly indicate that this molecule may be a potential inhibitor of topoisomerase IB. Moreover, this study opens major prospects for the development of novel chemotherapeutic agents with leishmanicidal activity.

Counteraction of Bothrops snake venoms by Combretum leprosum root extract and arjunolic acid.

ETHNOPHARMACOLOGICAL RELEVANCE: Serotherapy against snakebite is often unavailable in some regions over Brazil, where people make use of plants from folk medicine to deal with ophidic accidents. About 10% of Combretum species have some ethnopharmacological use, including treatment of snakebites. MATERIALS AND METHODS: We evaluated the ability of the extract of Combretum leprosum and its component arjunolic acid to reduce some in vivo and in vitro effects of Bothrops jararacussu and Bothrops jararaca venoms. The protocols investigated include phospholipase, proteolytic, collagenase, hyaluronidase, procoagulant, hemorrhagic, edematogenic, myotoxic and lethal activities induced by these venoms in Swiss mice. RESULTS: Oral pre-treatment with arjunolic acid reduced the Bothrops jararacussu lethality in up to 75%, while preincubation prevented the death of all the animals. Hemoconcentration effect of Bothrops jararacussu venom was confirmed two hours after i.p. injection, while preincubation with arjunolic acid preserved the hematocrit levels. Both Combretum leprosum extract and arjunolic acid abolished the myotoxic action of Bothrops jararacussu venom. Preincubation of Bothrops jararacussu venom with the extract or arjunolic acid prevented the increase of plasma creatine kinase activity in mice. The hemorrhagic activity of Bothrops jararaca crude venom was reduced down to about 90% and completely inhibited by preincubation with 10 mg/kg or 100 mg/kg Combretum leprosum extract, respectively, while the preincubation and the pretreatment with 30 mg/kg of arjunolic acid reduced the venom hemorrhagic activity down to about 12% and 58%, respectively. The preincubation of the venom with both extract and 30 mg/kg arjunolic acid significantly reduced the bleeding amount induced by Bothrops jararacussu venom. The extract of Combretum leprosum decreased the edema formation induced by Bothrops jararacussu venom both in preincubation and pretreatment, but not in posttreatment. Similarly, arjunolic acid preincubated with the venom abolished edema formation, while pre- and posttreatment have been partially effective. Some enzymatic activities of Bothrops jararacussu and Bothrops jararaca venoms, i.e. phospholipase A2, collagenase, proteolytic and hyaluronidase activities, were to some extent inhibited by the extract and arjunolic acid in a concentration-dependent manner. CONCLUSIONS: Altogether, our results show that Combretum leprosum extract can inhibit different activities of two important Brazilian snake venoms, giving support for its popular use in folk medicine in the management of venomous snakebites.

Optimization of subcritical fluid extraction of bioactive compounds using Hansen solubility parameters.

Process engineering operations in food and nutraceutical industries pertaining to the design of extraction of value-added products from biomass using pressurized liquids involve a careful selection of the solvent and optimal temperature conditions to achieve maximum yield. Complex molecular structure and limited physical property data in the literature of biological solutes extracted from biomass compounds have necessitated the process modeling of such operations. In this study, we have applied the Hansen 3-dimensional solubility parameter concept to optimize the extraction of molecularly complex solutes using subcritical fluid solvents. Hansen solubility spheres characterized by the relative energy differences (RED) have been used to characterize and quantify the solute-subcritical solvent interactions as a function of temperature. The solvent power of subcritical water and compressed hydroethanolic mixtures above their boiling points has been characterized using the above-mentioned method. The use of group contribution methods in collaboration with computerized algorithms to plot the Hansen spheres provides a quantitative prediction tool for optimizing the design of extraction conditions. The method can be used to estimate conditions for solute-solvent miscibility, an optimum temperature range for conducting extractions under pressurized conditions, and approximate extraction conditions of solutes from natural matrices.