以不同萃取方法的檸檬多醣評估其對於小鼠巨噬細胞RAW 264.7的抗發炎活性

檸檬 (Citrus limon) 為芸香科柑橘屬植物，為台灣主要飲料作物之一。栽培品種以優利卡檸檬 (Citrus limon (L.) Burm. f.) 為主，因果皮較厚、酸度較高、有耐儲耐運、一年可收獲多次等特性，使其經濟價值極高，但在被榨汁後剩餘大約全果重量三分之一的果皮廢棄物，通常被丟棄導致嚴重的浪費和環境污染。檸檬皮含有大量的植物多醣，而植物多醣透過對小鼠巨噬細胞刺激，可以有效對抗發炎、抗腫瘤、殺菌及其他治療作用。本研究以市售柑橘果膠 (P-industry)、酸萃取 (P-acid) 、鹼萃取 (P-alkaline) 之檸檬多醣為樣品，測定其多醣組成特徵，並研究市售果膠、酸、鹼萃取之多醣對以脂多醣 (Lipoplysaccharide, LPS) 誘導發炎的RAW264.7小鼠巨噬細胞是否有抑制的功效，並尋找可能抑制發炎信號途徑。結果顯示，鹼萃取多醣之多醣含量和酯化度皆低於酸萃取；檸檬多醣在500 μg/mL濃度以下對RAW264.7小鼠巨噬細胞無細胞毒性；測定一氧化氮 (Nitric oxide, NO) 含量，及使用ELISA kit進行分析，加入各樣品並以LPS誘導RAW264.7巨噬細胞產生IL-6、IL-10、IL-1β、TNF-α，在各樣品中濃度500 μg/mL時有顯著下降，可預防發炎因子增加，濃度500 μg/mL P-alkaline可最有效抑制發炎作用；再進一步以西方墨點法測定NF-κB之蛋白質表現，確認可由NF-κB路徑減輕發炎反應；再進行ROS及細胞週期試驗，可有效降低LPS誘導之ROS量及促使S週期進展。因此，我們認為檸檬多醣具有不錯的抗發炎活性，未來有潛力應用於預防醫學之上。

關鍵字

檸檬皮；生物廢棄物；多醣；免疫調節；發炎

並列摘要

Lemon (Citrus limon) is a citrus plant in the Rutaceae family and it is common used as beverage crops in Taiwan. Cultivated varieties are mainly Eureka lemon (Citrus limon (L.) Burm. f.), which has the characteristics of thick causal skin, high acidity, storage and transportation resistance, and can be harvested multiple times per year. Hences, it has high economy value. Approximately one-third of the lemon was processed into food products, and the peels are usually considered as bio-waste and caused environmental problems. However, lemon peel contains high amount of botanical polysaccharides. The botanical polysaccharides have several functions on immune-stimulatory, anti-tumor, bactericidal and other therapeutic. The pectin of industry (P-industry), acid-extracted polysaccharide (P-acid) and alkaline-extracted polysaccharide (P-alkaline) were used to prepare samples to determine the characteristics of polysaccharide composition. We examined the protein expression of inflammatory cytokines, LPS-activated macrophages and to elucidate possible signaling pathways by using Western blotting analysis. In our results, it demonstrated that polysaccharide content and degree of esterification (DE) of P-alkaline are less than P- acid and lemon polysaccharides were not cytotoxic to RAW264.7 cells at the concentrations of 500 μg/mL. All P- industry, P- acid and P- alkaline three samples showed the decreasing of content of nitric oxide, IL-6, IL-10, IL-1β and TNF-α, and it also presented that P- industry, P- acid, and P- alkaline can suppress NF-κB activity significantly. P-alkaline concentration at 500 μg/mL can suppress ROS significantly and rescue cell-cycle procession in RAW264.7 cells. Hences, we expected that lemon polysaccharides have good potential on anti-inflammation ability to be a preventive medicine in the future.

並列關鍵字

lemon peel ； bio-waste ； polysaccharides ； immunomodulatory ； inflammatory

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