Mass spectrometry and synchrotron-FTIR microspectroscopy reveal the anti-inflammatory activity of Bua Bok extracts

Abstract

Introduction: Bua Bok or Centella asiatica (CA) is an Asian vegetable with anti-inflammatory benefits. Asiaticoside, asiatic acid, madecassoside and madecassic have been characterised as major active ingredients with a wide range of pharmacological advantages. In manufacturing processes, high-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LCMS) are used to routinely determine the active compounds in raw materials.

Objectives: This research aims to explore anti-inflammatory properties, characterise metabolites and observe the biochemical changes of the inflammatory induced macrophages after pretreatment with the potential extracted fractions.

Methods: Bua Bok leaf extracts were prepared. Macrophages were pretreated with non-toxic fractions to determine the anti-inflammatory action. Tentative metabolites of effective fractions were identified by LC-MS. Synchrotron fourier-transform infrared (S-FTIR) microspectroscopy was utilised to observe the biochemical change of the lipopolysaccharide (LPS)-induced cells after pretreatment with potential fractions.

Results: Fractions of ethyl acetate, 30% and 100% ethanol highly increased the nitrile scavenging and suppressed the function of phospholipase A₂ . Fractions of 70% and 100% ethanol strongly decreased nitric oxide production. The comparison of 39 chemical compounds was presented. The change of proteins was improved after pretreatment of macrophages with fraction 70% ethanol. Fraction of 100% ethanol revealed the lipid accumulation was lower than 70% ethanol and diclofenac.

Conclusion: While the anti-inflammatory actions of 70% and 100% ethanol were similar. S-FTIR expressed they inhibited inflammatory response with the distinct features of biomolecules. The S-FTIR, LC-MS and biological assay confidently provided the efficient strategies to inform the advantage of herbal extract on cellular organisation instead of a single compound.

Keywords: Bua Bok; Centella asiatica; anti-inflammatory; mass spectrometry; synchrotron-FTIR.