In vitro evaluation of the antiviral properties of Arachis hypogaea L. extracts against dengue virus serotype 1

Abstract

Dengue disease affects 50-100 million people every year worldwide. In Argentina, three serotypes circulate, with a major prevalence of DENV-1 (72%). Effective antiviral drugs are not currently available. Medicinal plants may contain bioactive principles capable of inhibiting the replication of the dengue virus, with Arachis hypogaea L. (peanut) standing out for its biological properties. The aim of the present work was to evaluate the antiviral activity of Tegument ethanolic extract (TEE) and Seed ethanolic extract (SEE) of peanuts against DENV-1 on Vero cells.

TEE and SEE were obtained by simple alcoholic extraction. The 50% cytotoxic concentration (CC50) on Vero cells was determined by Neutral Red Uptake (NRU) and MTT assays at 7 days. Antiviral activity was then assessed throughout the replication cycle using the lysis plate reduction technique. Cell monolayers were infected with 50 PFU/well of DENV-1, and treated with TEE (30μg/mL) and SEE (300μg/mL) during adsorption and penetration and, in intracellular stages of replication, incubated 7 days at 37°C. Subsequently, the cells were fixed with formaldehyde solution (10%) and stained with violet crystal (1%). Lysis plates were counted and viral inhibition was determined. Antiviral action was then evaluated at different stages of the replication cycle using the same technique. Statistical analysis (t-test and ANOVA, p<0.05) was performed using Graph Pad Prism 6.0.

The CC50 values were 169μg/mL and 65μg/mL for TEE and >1400μg/mL and 636μg/mL for SEE by NRU and MTT, respectively. The study of antiviral activity throughout the cycle of replication showed that TEE inhibited DENV-1 (92.5±0.9%) (p<0.001). The study of the target of action indicated that TEE acted in stages of viral adsorption and penetration (92.5±0.9%) but did not act in stages post-penetration (15.7±6.6%). Also, it exerted virucidal action (98±5.0.7%) and inhibited the virus (78±3.04%) in cellular pre-treatment (p<0.001). On the other hand, the SEE did not show inhibitory capacity against DENV-1 (23.1±2.9%).

In conclusion, the data obtained demonstrates the potential application of tegument extract from peanut in the development of an antiviral phytotherapeutic against the dengue virus.