Comparative anti-inflammatory efficiencies of the leaf extracts of Chromolaena odorata, Praxelis clematidea, Ageratum conyzoides and Ageratina Adenophora

Abstract

Chromolaena odorata (L.) R.M. King & H. Rob., Praxelis clematidea R.M. King & H. Rob., Ageratum conyzoides L. and Ageratina adenophora R.M. King & H. Rob. are Asteraceae plants that widespread in Thailand and many countries. They have been used as hemostatic, anti-inflammatory, and wound dressing agents. C. odorata and its main active component, scutellarein tetramethyl ether (scu) has been intensively reported for their anti-inflammatory molecular mechanisms at the molecular level. However, other three plants have not been studied. This study investigated the anti- inflammatory activity of these three plants compared with C. odorata extract and identified their active compounds. All plants leaves were extracted with 70% ethanol and analyzed main components by Thin layer chromatography (TLC). The anti- inflammatory activity was investigated in gene and protein levels in RAW 264.7 macrophage cell line. The main active compound was high-performance liquid chromatography (HPLC). From our results, the four extracts contained flavonoids i.e.scu and quercetin, phenolic compounds i.e. eugenol and gallic acid, stilbene i.e. lupeol, and terpenoids i.e. oxyresveratrol. Afterward, the extracts and all compounds were examined on cytotoxicity using MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5- Diphenyltetrazolium Bromide) assay. The expressions of inflammatory COX-2 and iNOS proteins and genes including COX-2, iNOS, IL-6, TNF- α, PGE-2(EP2) and EP4 were studied using western blot and quantitative reverse transcription polymerase chain reaction (qRT-PCR), respectively. All the extracts exhibited no cytotoxicity in high doses. C. odorata extract at a concentration of less than 600 µg/ml, P. clematidea, A. conyzoides and A. adenophora extracts at a concentrations of less than 800 µg/ml, scu at 20-200 µg/ml, gallic acid and quercitin at 1-500 µg/ml, eugenol at 1-100 µg/ml and oxyresveratrol at 2.5-25 µg/ml, and lupeol at 2.5-10 µg/ml had no cytotoxicity, providing over 70% of cell viability. All plant extracts and their compounds could dramatically reduce COX-2 and iNOS proteins and all genes expressions in a dose- dependent manner. Among all extracts, P. clematidea was the most effective extract. However, the HPLC presented that scu is not the only one main active component. P. clematidea did not have highest amount of scu according to highest activity. A. adenophora (10.11 %w/w), P. clematidea (8.64 %w/w), C. odorata (7.64 %w/w) and A. conyzoides (6.94 %w/w) contained the highest to lowest amount of scu in order. This study supported that all active compounds had synergistic effect. This research supports the anti-inflammatory efficacy of these four medicinal plants and their active compounds through NF-kB pathway.