- 學位論文
西餐香料中活性成分香芹酚抗發炎功效之探討
Anti-inflammatory properties of carvacrol, an active compound in western-type herbs
摘要
香芹酚 (Carvacrol) 是西餐常用香料牛至與百里香中所含的天然化合物成分，是一種具有生理活性的多酚類 (Polyphenols)。文獻指出多酚類物質可以保護細胞免受自由基的傷害，降低罹患癌症的風險、心血管疾病的發生率、免疫系統疾病和慢性代謝疾病。過去研究發現，香芹酚具有抗菌及抗病毒的特性，可作為飼料添加劑及食品防腐劑，同時亦具有抗氧化與神經保護作用。慢性發炎會增加疾病的發生率及罹癌風險，在慢性病患中發現皆有發炎反應，若能以天然植化素抑制發炎反應發生，將有助於減緩許多疾病的發生。本研究旨在探討香芹酚對脂多醣 (Lipopolysaccharide, LPS) 誘發小鼠巨噬細胞株 (RAW 264.7 cell) 發炎反應的抑制作用與相關機制。以不同濃度 (0、25、50、100、150 M) 香芹酚處理經 LPS 誘發 RAW 264.7 細胞，以 MTT Assay 測定天然物香芹酚對細胞是否具有毒殺作用，以 Gelatin Zymography 測定細胞中基質金屬蛋白酶 (Matrix metalloproteinase, MMP)-9 的活性，再以酵素連結免疫吸附分析法(Enzyme-linked immunosorbent assay, ELISA) 偵測促發炎因子白細胞介素 (Interleukin, IL) -6、腫瘤壞死因子 (Tumor necrosis factor, TNF) - 分泌量，最後利用 Western blot 評估香芹酚抑制蛋白表現量及相關分子機轉。結果顯示，香芹酚在處理濃度 150 M 以內對 RAW 264.7 細胞無毒殺作用，以低於此濃度進行以下實驗，香芹酚在濃度 25 – 150 M 時，可顯著抑制IL-6 與 TNF-細胞激素的分泌量；且細胞質內磷酸化之 ERK、JNK、p38 訊息蛋白與細胞核內之轉錄因子NF-B，亦隨香芹酚處理濃度增加而表現量減少 ; 此外，香芹酚也使細胞內 MMP-9的酵素活性受到抑制。綜合以上結果發現，香芹酚可能藉由抑制訊息傳遞 MAPKs 之路徑與降低 NF-B 活性，阻止 TNF-對促炎因子的表現， 降低 IL-6 釋放與 MMP-9 活性，達成抑制 LPS 誘發RAW 264.7小鼠巨噬細胞之發炎反應。
並列摘要
Carvacrol is a phenolic monoterpene presents in the essential oil of the family Lamiaceae that responsible for the biological activities of oregano. Carvacrol is used as feed additive and preservative because of its diverse bioactivities such as antimicrobial, antitumor, antimutagenic, antiplatelet, antihepatotoxic and hepatoprotective activities and so on. Carvacrol was reported that is effective as an analgesic compound by inhibiting peripheral mediators, and the function could be related with its strong antioxidant effect observed in vitro. The aim of the study was to evaluate the anti-inflammatory potential and molecular mechanism of carvacrol in cell model, RAW 264.7 cells. MTT method was used to assay the toxicity of carvacrol on cells. Cells were stimulated to the status of inflammation by lipopolysaccharide (LPS) and then treated with carvacrol at a non-toxic concentration. The levels of several inflammation-related factors were determined, including tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) by ELISA, matrix metalloproteinase (MMP)-9 by gelatin zymography, and molecular signaling MAPKs and transcription factor NF-B by Western blot. The results indicated that TNF-α and IL-6 in cells can be significantly induced by 100 ng/mL LPS, and the induced-TNF-α and IL-6 were suppressed by the treatment of carvacrol at a concentration of 25-150 M. The gelatinolytic activity and expression of MMP-9 were depressed by carvacrol. Additionally, MAPKs and NF-B in cells were inactivated by carvacrol. From the results, we concluded that carvacrol is a potential anti-inflammatory natural compound. The inflammation-inhibited effect may through downregulating TNF-α, IL-6, and MMP-9 via inactivating MAPKs and NF-B.