由 Serratia marcescens 生產 prodigiosin 之研究

Abstract

Prodigiosin 為 Serratia marcescens、Pseudomonas spp.、Streptomyces spp. 等微生物所產生之紅色色素。近年來陸續有研究指出 prodigiosin 具有免疫抑制及抗癌活性，因此對於將來無論是用於機能性食品或是開發成新型免疫抑制劑及抗癌藥物，都具有很大的潛力。在本研究中欲使用 S. marcescens 大量生產 prodigiosin，結果顯示，S. marcescens BCRC 11576 以1.4% 大豆粉培養基於27℃、120 rpm培養 48 小時後，每瓶培養液 (100 mL) 產量可達 48 mg，為目前已知研究中最高者。另外，培養基組成會大幅影響 prodigiosin 之產量。添加葡萄糖使葡萄糖與氮源之比例達一定程度會抑制 prodigiosin 之生成，並且使培養液之 pH 快速下降至 3 以下。將培養液緩衝化使培養過程中 pH 維持在 7 ~ 9 之間，可使 prodigiosin 之生成不受到抑制，推論 pH 下降可能為導致 prodigiosin 生成受到抑制之因。 以1.4% 大豆粉培養基生產之培養液 1L (prodigiosin 含量為 0.46 g/L)，取菌體以甲醇萃取得粗萃色素 1.9 g，經矽膠管柱純化之乙酸乙酯區分為 prodigiosin，可得 0.18 g，純度達 94%。純化後之 prodigiosin由體外鼷鼠淋巴瘤 tk 分析法得知在 0.8 ~ 8 mM 之濃度並無顯著之致突變性，增加了 prodigiosin 使用上之安全性。

Prodigiosin is the red pigment produced by microorganisms such as Serratia marcescens, Pseudomonas spp., Streptomyces spp., etc. Recently, the immunosuppressive and apoptotic activities of prodigiosin have been described. Therefore, prodigiosin seems to be a promising compound for the usages in the functional food, new immunosuppressants and anti-cancer drugs. In this study, the yield of the prodigiosin produced by S. marcescens BCRC 11576 reached 48 mg/100mL broth after incubation in 1.4% soybean flour broth at 27℃ with vigorous shaking for 48 hours. This yield was the highest of all previously reported. Besides, it was found that the prodigiosin production of this strain was affected greatly depending on the composition of medium. When this strain was incubated in the presence of glucose with the glucose-to-nitrogen source ratio reached to a certain value, no prodigiosin was produced and the pH of the broth dropped quickly to a value below 3. Buffering the broth between pH 7 and pH 9 during incubation released the inhibition resulted from addition of glucose to the broth. Consequently, the inhibitory effect of glucose on prodigiosin production may be due to a lowering of the pH of the medium. S. marcescens BCRC 11576 was incubated in 1.4% soybean flour broth for 48 hours (the yield of the prodigiosin was 0.46 g/L). After 1 L broth was centrifuged, the cell pellets extracted with methanol and obtained 1.9 g of the crude extract pigment. The crude extract was purified by silica gel chromatography and the prodigiosin was eluted with ethyl acetate. The amount and the purity are 0.18 g and 94%, respectively. The purified prodigiosin showed no significant mutagenesis analyzed by the mouse lymphoma tk assay with the prodigiosin concentration between 0.8 and 8 mM. This result increased the safety for the use of prodigiosin.