天然抗癌藥物靈菌紅素之微生物醱酵製程開發

Development of fermentation processs for the production of a natural anti-tumor drugs-prodiogiosin by microorganism

徐巧玲(Chiao-Ling Hsu)

指導教授 ： 魏毓宏

元智大學/工程學院/生物科技與工程研究所/碩士(2008年)

摘要

本研究使用自台灣大學醫技系賴信志教授所提供之沙雷氏菌突變株Serratia macescens SMΔR，為革蘭氏陰性菌，具有生產紅色色素—靈菌紅素之能力。本篇論文主要使用S. macescens SMΔR進行其代謝靈菌紅素之定性定量及醱酵生產策略上的探討。首先研究證實，S. macescens SMΔR菌株醱酵產生之紅色色素經分離純化後，藉由1H-NMR及FAB-MS確認合成之靈菌紅素屬於Undecylprodigiosin
（UP），為靈菌紅素衍生物之一。其次，S. macescens SMΔR最適合生產靈菌紅素的碳、氮源分別為Mannitol及Soytone，其最佳產量為13.5 g/L，進一步將上述培養基利用回應曲面實驗設計法得最佳化培養基組成為：20.95 g/L Mannitol、16.33 g/L Soytone、5.34 g/L Proline和1.25 mM MnSO4，透過回應曲面實驗設計所得之最適化培養基，能將靈菌紅素產量提升至19.5 g/L，為一般文獻值的4倍，產率為0.406 g/L/h。在搖瓶饋料批次策略研究證實：在產率方面，於24小時，饋入2 %的碳源與氨基酸，靈菌紅素的產量為29 g/L，產率為0.48 g/ L/h為批次實驗的1.2倍；在產量方面，1%營養源（mannitol only） 採用每12小時連續定量進料方式可得到最佳的靈菌紅素產量（32.4 g/L），為批次實驗的1.7倍，為一般文獻值的7.6倍。在醱酵槽饋料批次研究證實，2 %營養源於24小時添加，可使產量由批次醱酵之19 g/L提升至26.5 g/L。因此，適時適量添加營養源有助於菌體生長與靈菌紅素的代謝合成。

關鍵字

靈菌紅素 ； Serratia macescens SMΔR ； 回應曲面法 ； 饋料批次

並列摘要

Serratia marcescens SMdeltaR ,a gram negative bacteria provided by Prof. H-C. Lai from Chang Gung University, Taipei, Taiwan was used to produce a prodigiosin-like pigment. This study aimed to develop better culture medium and fermentative strategy enabling more efficient production of prodigiosn from Serratia marcescens SMdeltaR. Undecylprodigiosin is one of prodigiosin-like pigment production by Serratia marcescens SMdeltaR was purified and characterized by1H-NMR and FAB-MS to determine its chemical structure and molecular weight. Preliminary tests indicated that a combination of mannitol and soytone as the carbon and nitrogen source gave a good prodigiosin production of 13.5 g/L. Furthermore, statistical experimental design ( Response surface methodology, RSM ) was applied for the purpose of identifying optimal formulation in the medium : 20.95 g/L Mannitol, 16.33 g/L Soytone, 5.34 g/L Proline and 1.25 mM MnSO4.
The optimization strategy led to the enhancement prodigiosin production of 19.5 g/L and productivity of 0.406g/L/h. The results of fed-batch in shaking flask revealed that optimum fed nutrient is 2 % mannitol (proline included ) and the optimum fed timing is 24hr , the prodigiosin production of 29 g/L and better productivity of 0.48 g/L/h which was successfully 1.2-fold higher than batch experiment. In constant fed (mannitol only), this strategy could enhance better proidgiosin concentration was 32.4 g/L, which was approximately 1.7-fold higher than that batch experiment and 7.6-fold higher than reference (4.28 g/L).Meanwhile, the results of fed-batch cultures in fermentor revealed that 2 % mannitol (including proline) fed at 24 hr, which led to the enhancement of prodigiosin from 19 g/L to 26.5 g/L.

並列關鍵字

Prodigiosin ； Serratia macescens SMΔR ； RSM ； Fed-batch

參考文獻

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