嗜鹽菌硫化多醣之大量生產及其於抗氧化與醫學功能之探討

The mass production of a halophilic bacteria’s sulfated polysaccharide and it’s antioxidant function with biomedical application

陳依瑩(Yi-Ying Chen)

指導教授 ： 陳志成

大同大學/工程學院/生物工程學系所/碩士(2008年)

摘要

多醣已廣泛的應用於食品、製藥與化妝品及其他工業上。多醣大部分的來源是從藻類或植物萃取所得，缺點在於萃取分離的成本過高。而利用微生物來生產胞外多醣則具有可大量生產與易分離的優點。已知極端嗜鹽菌Haloferax mediterranei所生產的胞外多醣，是一種含有硫酸根與甘露糖的多醣聚合物。因此本研究乃尋找醱酵培養H. mediterranei，生產胞外硫化多醣(exopolysaccharide，EPS)之最適培養基配方與其應用。實驗發現利用飼料級玉米粉為碳源，以溫度42℃，轉速180 rpm和碳源濃度15.5 g/L的條件下可得最佳的菌體量以及EPS產量，分別為1.84×106 cells/ml及5.9 g/L的多醣。同時發現EPS的分子量約為3.1×104 Da且含硫量約為7.5 %。而經由細胞培養試驗得知，此EPS以內差法求得對人類肺癌細胞(A549)和人類橫紋肌瘤細胞(RD)細胞達到50％毒性抑制的濃度(CC50)分別1.94及1.49 mg/mL；在抗氧化試驗可得知，EPS於43.7 mg/mL濃度下對清除DPPH自由基可達50 %的清除能力(IC50)；於91.2 mg/mL濃度下對螯合亞鐵離子可達50 %的螯合能力；在抗病毒試驗得知，EPS在0.63 mg/mL濃度之下對腺病毒41型(AD41)有16.35%的些微抗病毒效果，當EPS濃度於1.25 mg/mL時對，腸病毒71型(EV71)有較明顯的抑制效果，為23.62 %；在抗菌試驗可得知，EPS於50 mg/mL濃度下對野生型大腸桿菌可達中度的抑菌活性(抑制圈13.17±0.3 mm)，於1 mg/mL濃度下對金黃色葡萄球菌可達中度抑菌活性(抑制圈15.33±0.3 mm)。上述數據顯示，嗜鹽菌EPS具有可大量生產與醫學上的應用價值。

關鍵字

多醣 ； 嗜鹽菌

並列摘要

Polysaccharide recently has been used in some industries like food, pharmacy and cosmetics. Most polysaccharides can be extracted from algae or plants, however, the cost of extracting process is expensive. On the contrary, there is a benefit to produce and purify the mass quantity of exopolysaccharide(EPS) from microorganism. Haloferax mediterranei can produce EPS with functional qroup of sulfate and mannose units. In this stuy, we are seeking for the suitable medium and culture method to produce EPS by H. mediterranei, and also looking for the application of EPS. By the shaking-flask experiment, it is found that using 15.5 g/L field corn as the carbon source, and culture condition at 42℃ and rotational speed of 180 rpm, we could obtain the best quantity of cell concentration of 1.84×106 cells/ml and 5.9 g/L EPS. The molecular weight of EPS was measured as 3.1×104 Da, and sulfur content in the EPS was measured as 7.5%. By the cell culture test, the 50% toxicity(CC50) of EPS to the human lung A549 cell and Rhadomyosarcoma cell line were 1.94 and 1.49 mg/mL respectively by linear interpolation. In the anti-oxidation experiments, it is found that the 50% radical scavenging activity (IC50) to DPPH is 43.7 mg/mL, and the IC50 of Fe2+ chelating ability was 91.2 mg/mL. Besides, in the of anti-virus experiments, the concentrations of 0.63 mg/mL EPS shown a slight anti-virus activity (16.4%) to Adeno Virus (AD41). The anti-virus effect to Enterovirus EV71) is 23.62% when the concentrations of EPS is 1.250 mg/mL. By the antibacterial test, the concentration of EPS of 50 mg/ml shows a mild effect of inhibition (with inhibition zone of dia, 13.17±0.3 mm to E.coli). The concentration of 1 mg/ml EPS shows a mild inhibition to the growth of Staphylococcus aureus with inhibition zone of 15.33±0.3 mm). We conclude that the EPS from H. mediterranei could be mass produced and also shows the potential medical applications.

並列關鍵字

polysaccharide ； halophilic bacteria

參考文獻

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