Inhibition of nitric oxide production and COX-2 protein expression in LPS-stimulated RAW 264.7 cells by the hexane fraction of Murdannia loriformis
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Abstract
Background: Murdannia loriformis (ML) is a medicinal plant traditionally used for chronic bronchitis, cancers in the initial stage, colds, throat infections, pneumonia, the flu, and wound healing. The crude ethanolic extract of ML has been reported anti-inflammatory, analgesic, antipyretic and gastroprotective activities in various in vivo experiments.
Objectives: This study aimed to isolate the active fractions of ML and assess the effect on nitric oxide (NO) inhibition and cyclooxygenase 2 (COX-2) expression.
Materials and methods: The dry powder of ML was extracted with 80% ethanol. The crude ethanolic extract afterward was brought to partition with the various solvents base on polarity difference. Finally, accepted hexane, chloroform, ethyl acetate, and water fractions, respectively. The ML extract and its fractions were screened the cytotoxicity on RAW264.7 cells by sulforhodamine B assay. The non-toxic doses were selected for the next NO inhibition experiment. RAW264.7 cells were treated with the various non-toxic doses of the ML. Lipopolysaccharide (LPS) was added to induce cells inflammation and stimulate NO production. Additionally, the culture supernatants were collected and measured NO levels by Griess reagent. The fraction that revealed potent anti-inflammatory activity by reduced NO accumulation was selected to study COX-2 protein suppression by western blot analysis.
Results: The results showed that crude ethanolic extract and all fractions except for the water fraction significantly inhibited NO production of RAW264.7 cells. The hexane fraction demonstrated a superior on nitric oxide reduction as same as the standard drugs L-NAME and indomethacin. This fraction also reduced COX-2 protein expression in LPS-stimulated RAW264.7 cells.
Conclusion: The hexane fraction possesses an anti-inflammatory activity by reducing nitrite level and COX-2 protein expression. Suggesting that, the active ingredient of ML is the non-polar compound. Further studies should be carried out to isolate the pure compound from the hexane fraction and structure identification.
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