Anti-inflammatory activity of standardized dichloromethane extract of Salvia connivens on macrophages stimulated by LPS

Abstract

Context: A previous study demonstrated that the chloroform extract of Salvia connivens Epling (Lamiaceae) has anti-inflammatory activity.

Objective: Identification of the active components in the dicholorometane extract (DESC), and, standardization of the extract based in ursolic acid.

Material and methods: DESC was prepared by percolation with dichlromethane and after washed with hot hexane, its composition was determined by CG-MS and NMR, and standardized by HPLC. The anti-inflammatory activity was tested on acute TPA-induced mouse ear oedema at doses of 2.0 mg/ear. The cell viability of macrophages was evaluated by MTT method, and pro- and anti-inflammatory interleukin levels were measured using an ELISA kit.

Results: Ursolic acid, oleanolic acid, dihydroursolic acid and eupatorin were identified in DESC, which was standardized based on the ursolic acid concentration (126 mg/g). The anti-inflammatory activities of DESC, the acid mixture, and eupatorin (2 mg/ear) were 60.55, 57.20 and 56.40% inhibition, respectively, on TPA-induced ear oedema. The IC₅₀ of DESC on macrophages was 149.4 μg/mL. DESC (25 μg/mL) significantly reduced TNF-α (2.0-fold), IL-1β (2.2-fold) and IL-6 (2.0-fold) in macrophages stimulated with LPS and increased the production of IL-10 (1.9-fold).

Discussion: Inflammation is a basic response to injuries, and macrophages are involved in triggering inflammation. Macrophage cells exhibit a response to LPS, inducing inflammatory mediators, and DESC inhibits the biosynthesis of the pro-inflammatory and promote anti-inflammatory cytokines.

Conclusion: DESC has an anti-inflammatory effect; reduced the levels of IL-1β, Il-6 and TNF-α; and increases IL-10 in macrophages stimulated with LPS. Ursolic acid is a good phytochemical marker.

Keywords:

• Ursolic acid
• HPLC
• cellular viability
• cytokine determination
• eupatorin

Disclosure statement

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of this article.

Additional information

Funding

We acknowledge the financial support of the CONACYT for Consejo Nacional de Ciencia y Tecnologia (project number 129903) and PRODE for Programa Para el Desarrollo Profesional Docente.