Bioassay-guided fractionation of anti-inflammatory components from residue of aqueous extract of Chinese olive

• Main Authors: Kuei-Ming Hsu, 許貴明
• Other Authors: Shu-Chen Hsieh
• Format: Others
• Language: zh-TW
• Published: 2016
• Online Access: http://ndltd.ncl.edu.tw/handle/61846939870770993964

碩士 === 國立臺灣大學 === 食品科技研究所 === 104 === Chronic inflammation is one of the factors causing chronic disorders. Many secondary metabolites extracted from herbs exhibit both anti-oxidative and anti-inflammatory activities, and may have potential against inflammation-induced chronic diseases. Canarium album is a common herb and food with excellent anti-oxidative and hepatoprotective effects. Its ethyl acetate fraction from methanol extract has been proved to exhibit good anti-inflammatory potential in our previous work.The objective of this research is to use different isolation techniques to screen bioactive fractions guided by bioassays to obtain anti-inflammatory compounds. The bioassay is a luciferase reporter assay built by our lab using the cell line RAW 264.7 that has been constructed with either NF-κB binding site or COX-2 promoter. First, the fraction was separated based on polarity differences, and the supernatant was found to have better anti-inflammatory action than the precipitate. It was then isolated with silica gel chromatography, and the anti-inflammatory action was concentrated in sub-fraction D, which also showed substantial cytotoxicity. When sub-fraction D was subjected to isolation again with silica gel chromatography, sub-fraction D-11 was found to have better anti-inflammatory action, but also with dose-dependent cytotoxicity.After isolation and purification by thin-layer chromatography and LH-20 column, the active compound from sub-fraction D-11 was identified to be scoparone by nuclear magnetic resonance. Quantification by HPLC showed that scoparone increases as purification steps increase. However, scoparone was unable to inhibit LPS-induced inflammation in the luciferase reporter assay. When spiking scoparone into the extract to explore whether there is synergy between the two, the result was negative. Furthermore, scoparone exhibited a weaker effect of inhibiting NO-induced inflammation than INF-γ/LPS-induced inflammation in literature, indicating scoparone was not the active anti-inflammatory factor in ethyl acetate fraction of methanol extract of C. album. Identification of other components in sub-fraction D-11 is required to reveal the active compound..