能以3-羥基丁酸作為生長單一碳源的海洋細菌之分離與特性探討

Isolation and characterization of the marine bacteria which utilize β-hydroxybutyrate as a sole carbon source

林姵伶(Pei Ling Lin)

指導教授 ： 謝文陽

國立臺灣大學/理學院/海洋研究所/碩士(2013年)

https://doi.org/10.6342/NTU.2013.11047

摘要

聚羥基烷酸酯 (polyhydroxyalkanoates, PHAs)為一類聚酯，可由多種微生物合成。由於其生物可降解性、生物相容性及熱塑性等特性，可做為生物塑膠的材料，近年來相關的研究及應用吸引許多學者與產業的注意。聚羥基丁酸酯 (polyhydroxybutyrate, PHB)為一 PHAs，由3-羥基丁酸 (3-hydroxybutyrate, 3-HB)聚合而成，目前僅發現原核生物具有合成 PHB之能力。3-HB可被許多類群的細菌作為單一碳源供生長所需。本研究旨在篩選與探討能以 3-HB作為單一碳源的細菌，並進一步了解其是否可蓄積 PHB。樣本取自台灣東北角沿岸，經連續稀釋後接種至 glucose mineral medium (GM medium) 及 3-HB mineral medium (HBM medium) 中進行增菌培養，以最大可能計數法 (most- probable-number, MPN)估算可利用葡萄糖或 3-HB作為單一碳源的細菌豐度。GM medium中所測得之生菌數約介於 2.4×102至 1.3×104 cells/ml，HBM medium中所測得之生菌數約介於 3.3×102至 1.1×104 cells/ml。共得出 45株能以 3-HB作為單一碳源的分離株，皆為可運動的革蘭氏陰性菌，均屬好鹽性，生長溫度範圍在 4-45 ℃之間。經 16S rRNA基因定序並與已知菌株比對，多數歸類為 Pseudoalteromonas及 Alteromonas兩屬，其餘分屬Marinobacterium, Vibrio及 Ruegeria。經 Sudan Black B染色法檢測，Vibrio屬的 A3與 A23具有蓄積 PHB之能力。

關鍵字

聚羥基烷酸酯 ； 聚羥基丁酸酯 ； 3-羥基丁酸 ； 弧菌屬

並列摘要

Polyhydroxyalkanoates (PHAs) are polyesters which can be produced by various microorganisms. Because of the properties of biodegradability, biocompatibility and thermoplasticity, PAHs have become a bio-plastics material applied in many industrial fields in recent years. They have also gained a great attention of scientists in basic researches and industrial applications. Polyhydroxybutyrate (PHB), a PHA, is a polymer of 3-hydroxybutyrate (3-HB). It was found only synthesized by prokaryotes. 3-HB could be used for growth of many bacteria of different taxa as the sole carbon source. The aims of this study were to screen and explore the bacteria which could utilize 3-HB as a sole carbon source and those which could further accumulate PHB. The samples were obtained from Taiwan north coastal seawater and cultured in glucose mineral medium (GM medium) and 3-HB mineral medium (HBM medium) after serial dilution. The abundance of bacteria, which could utilize glucose or 3-HB as a sole carbon source, was calculated by most-probable-number method (MPN). The amounts of culturable bacteria were found ranged from 2.4×102 to 1.3×104 cells/ml in GM and 3.3×102 to 1.1×104 cells/ml in HBM. Forty-five isolates were found able to utilize 3-HB. They were motile and Gram-negative. All isolates required NaCl for growth and grew at 4-45 °C. According to 16S rRNA gene sequence analysis, most isolates belonged to Pseudoalteromonas and Alteromonas, and others were Marinobacterium, Vibrio and Ruegeria. Isolates A3 and A23, belonging to Vibrio genus, were found able to accumulate PHB as revealed by Sudan Black B staining.

並列關鍵字

polyhydroxyalkanoates ； polyhydroxybutyrate ； β-hydroxybutyrate ； Vibrio

參考文獻

邱秀慧 (2005)。台灣沿岸海洋弧菌之收集、分離與分類。台灣大學海洋研究所博士論文。

Chimetto, L. A., Cleenwerck, I., Brocchi, M., Willems, A., De Vos, P., & Thompson, F. L. (2011). Marinobacterium coralli sp. nov., isolated from mucus of coral (Mussismilia hispida). International Journal of Systematic and Evolutionary Microbiology, 61(Pt 1), 60-64.

Ivanova, E. P., Bowman, J. P., Lysenko, A. M., Zhukova, N. V., Gorshkova, N. M., Sergeev, A. F., & Mikhailov, V. V. (2005). Alteromonas addita sp. nov. International Journal of Systematic and Evolutionary Microbiology, 55(Pt 3), 1065-1068.

Antongiovanni, M., Buccioni, A., Petacchi, F., Leeson, S., Minieri, S., Martini, A., & Cecchi, R. (2007). Butyric acid glycerides in the diet of broiler chickens: effects on gut histology and carcass composition. Italian Journal of Animal Science, 6(1), 19-25.

Arun, A., Arthi, R., Shanmugabalaji, V., & Eyini, M. (2009). Microbial production of poly-beta-hydroxybutyrate by marine microbes isolated from various marine environments. Bioresource Technology, 100(7), 2320-2323.