Temperature effect in the inhibition of PLA2 activity of Bothrops brazili venom by Rosmarinic and Chlorogenic acids, experimental and computational approaches.

Myotoxicity caused by snakebite envenoming emerges as one of the main problems of ophidic accidents as it is not well neutralized by the current serum therapy. A promising alternative is to search for efficient small molecule inhibitors that can act against multiple venom components. Phospholipase A2 (PLA2) is frequently found in snake venom and is usually associated with myotoxicity. Thus it represents an excellent target for the search of new treatments. This work reports the effect of temperature in the inhibition of catalytic properties of PLA2 from Bothrops brazili venom by Rosmarinic (RSM) and Chlorogenic (CHL) acids through experimental and computational approaches. Three temperatures were evaluated (25, 37 and 50 °C). In the experimental section, enzymatic assays showed that RSM is a better inhibitor in all three temperatures. At 50 °C, the inhibition efficiency decayed significantly for both acids. Docking studies revealed that both ligands bind to the hydrophobic channel of the protein dimer where the phospholipid binds in the catalytic process, interacting with several functional residues. In this context, RSM presents better interaction energies due to stronger interactions with chain B of the dimer. Molecular dynamics simulations showed that RSM can establish selective interactions with ARG112B of PLA2, which is located next to residues of the putative Membrane Disruption Site in PLA2-like structures. The affinity of RSM and CHL acids towards PLA2 is mainly driven by electrostatic interactions, especially salt bridge interactions established with residues ARG33B (for CHL) and ARG112B (RSM) and hydrogen bonds with residue ASP89A. The inability of CHL to establish a stable interaction with ARG112B was identified as the reason for its lower inhibition efficiency compared to RSM at the three temperatures. Furthermore, extensive structural analysis was performed to explain the lower inhibition efficiency at 50 °C for both ligands. The analysis performed in this work provides important information for the future design of new inhibitors.Communicated by Ramaswamy H. Sarma.

Antiophidic potential of chlorogenic acid and rosmarinic acid against Bothrops leucurus snake venom.

Bothrops leucurus is responsible for most cases of snakebite in Northeast Brazil; however, this species is not included in the pool of venoms used in antivenom production in Brazil. The serotherapy has logistical and effectiveness limitations, which stimulates the search for therapeutic alternatives. Chlorogenic acid and rosmarinic acid present several biological activities, but their antiophidic potential has been poorly explored. Thus, the aim of this approach was to evaluate the potential inhibitory effects of these compounds on B. leucurus venom. Initially, the enzymatic inhibition of toxins was evaluated in vitro. Then, anti-hemorrhagic, anti-myotoxic, and anti-edematogenic assays were performed in vivo, as well analysis of several biochemical markers and hemostatic parameters. In addition, the interaction of inhibitors with SVMP and PLA2 was investigated by docking analysis. Results revealed that compounds inhibited in vitro the enzymatic activities and venom-induced edema, with a decrease in both myeloperoxidase and interleukin quantification. The inhibitors also attenuated the hemorrhagic and myotoxic actions and mitigated changes in serum biochemical and hemostatic markers, as well as decreased lipid peroxidation in liver and kidney tissues. Docking analysis revealed attractive interactions of both inhibitors with the zinc-binding site of SVMP and, in the case of PLA2, chlorogenic acid showed a similar inhibition mechanism to that described for rosmarinic acid. The results evidenced the antiophidic potential of both compounds, which showed higher efficiency than antivenom serum. Thus, both inhibitors are promising candidates for future adjuvants to be used to complement antivenom serotherapy.

The potential of phenolic acids in therapy against snakebites: A review.

Ophidism is a serious health problem worldwide and is included in the World Health Organization's (WHO's) list of Neglected Tropical Diseases. Although snakebite envenoming requires emergency treatment, currently the only treatment recommended by WHO is serotherapy, which has some disadvantages such as low access to the rural population, low effectiveness in neutralizing local effects, and high cost. In this context, new alternatives for the treatment of snakebites are required. The use of plant-derived compounds to inhibit the effects caused by snake venoms has been the object of a number of studies in recent years. This review aims to provide an up-to-date overview of the use of phenolic acids with therapeutic application against envenomation by snakes of different species. In this sense, structural analysis in silico and biological activities in vivo and in vitro were reported. The acids were subdivided into derivatives of benzoic and cinnamic acids, with derivatives of cinnamic acids being the most studied. Studies have revealed that these compounds are capable of inhibiting local and systemic effects induced by envenomation, and structural analyses indicate that the acids interact with important sites responsible for the action of toxins. Thus, it was reported that phenolic acids showed antiophidic potential, providing insights for future research to develop complementary drugs for the treatment of snakebites.