探討溫度暨萃取發酵系統對於Saccharomyces cerevisiae連續生產苯乙醇之影響

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周筱軒(Siao-Syuan, Jhou) 查詢紙本館藏

畢業系所

化學工程與材料工程學系

論文名稱

探討溫度暨萃取發酵系統對於Saccharomyces cerevisiae連續生產苯乙醇之影響
(Enhanced biotransformation of L-phenylalanine into 2-phenylethanol via cultivation temperature control strategy and extractive fermentation)

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摘要(中)

苯乙醇因具有特殊的玫瑰香氣而被大量的應用在香水、化妝品及食品香精中，年產量在全球香料市場中佔有一席之地。利用微生物發酵法，依循Ehrlich pathway的生成途徑，可以透過Saccharomyces cerevisiae將苯丙胺酸轉化成苯乙醇，但在發酵過程中，面臨最大的限制在於產物的抑制效應，會對細胞產生毒性的影響。而且在微生物發酵中，不同培養溫度也會影響微生物的生理狀態和發酵行為，因此本研究主要的目的是在於探討「溫度」環境因子對於S. cerevisiae生產苯乙醇之影響，探討不同培養溫度對於菌體的生長及產物的變化，並結合萃取技術來改善產物抑制的問題。
研究結果發現在溫度為25℃下最適合S. cerevisiae的生長，但在溫度為35℃下則比較有助於單位菌重轉化成產物的能力，於是設計一種兩階段溫度轉換的策略同時提升菌體及產物的產量，在此操作模式下，苯乙醇的產量可以高達2.53 g/L，與在單一發酵溫度25℃跟35℃時相比下分別提升了15.48 %及22.38 %，最後再結合連續萃取發酵裝置，以Hytrel®作為吸附劑的角色，透過 in situ product removal 技術，減輕產物對於菌體生長的限制，最終苯乙醇的累積濃度可達到4.506 g/L。

摘要(英)

2-Phenylethanol (PEA) is one of the important flavor and fragrance compound in the world with a rose-like aroma. In addition, it is widely used in the cosmetics, perfume, and food industries.

PEA can be produced from L-phenylalanine (L-Phe) through Ehrlich pathway using yeasts. In the fermentation process, the growth of yeast will be limited by product inhibition, cultivation temperature will also has a strong connection with microbial growth and fermentation behavior.

In this work, we have explored the effects of temperature on bioconversion of L-Phe into PEA using Saccharomyces cerevisiae BCRC21812 as well as develop strategies to enhance PEA production. Experimental results showed that at 25°C, the optimum cells growth was achieved and favored bioconversion yield. But at 35°C was more favorable for product yield based on biomass. Therefore, two-stage batch fermentation by temperature control was conducted. PEA concentration using two-stage fermentation was 2.53 g/L which corresponds to 15.48 % and 22.38 % higher than single-stage fermentation at 25°C and 35°C, respectively.

To increase the PEA production, the extractive fermentation will be conducted using Hytrel®. In the preliminary test, it showed that Hytrel® can adsorb PEA to reduce the inhibitory effect and non-toxic effect on yeast growth. After the successive extraction and feeding fermentation, the final concentration of PEA was achieved 4.506 g/L.

關鍵字(中)

★ 酵母菌
★ 苯乙醇
★ 溫度效應

關鍵字(英)

★ Saccharomyces cerevisiae
★ 2-Phenylethanol
★ temperature

論文目次

目錄
摘要 ii
Abstract iii
誌謝 v
目錄 vi
表目錄 x
圖目錄 xi
第一章緒論 1
1-1研究動機 1
1-2研究目的 2
第二章文獻回顧 4
2-1 香料香精化學品市場趨勢及發展 4
2-2 芳香化合物-苯乙醇 7
2-2-1 苯乙醇基本性質介紹 7
2-2-2 苯乙醇特性及應用 7
2-2-3 苯乙醇之生產方式 9
2-3 釀酒酵母(Saccharomyces cerevisiae) 14
2-3-1 酵母菌基本介紹 14
2-3-2 酵母菌生殖方式 15
2-3-3 酵母菌代謝途徑 16
2-3-4 酵母菌生長條件 18
2-4 影響發酵之因素 19
2-4-1 培養基組成 19
2-4-2 pH值 20
2-4-3 通氣速率 21
2-4-4 光 21
2-4-5 溫度 21
2-4-6 產物抑制效應 22
2-5 微生物發酵製程改善-ISPR技術 22
2-6 Hytrel®性質及應用 27
第三章材料與方法 29
3-1 實驗規劃 29
3-2 實驗材料與設備 30
3-2-1 實驗菌株 30
3-2-2 實驗藥品 31
3-2-3 實驗儀器與設備 32
3-2-4 數據呈現之參數符號設定 35
3-2-5 實驗裝置 35
3-3 實驗方法 39
3-3-1 菌種保存 39
3-3-2培養基組成 39
3-4 分析方法 43
3-4-1 菌重濃度測定 43
3-4-2 葡萄糖殘糖分析 43
3-4-3 乙醇濃度分析 45
3-4-4 苯丙胺酸和苯乙醇濃度分析 47
第四章實驗結果與討論 50
4-1 發酵培養基最適化 50
4-1-1 碳源濃度對於Saccharomyces cerevisiae發酵動力曲線之影響 50
4-1-2 氮源濃度對於Saccharomyces cerevisiae發酵動力曲線之影響 53
4-2 培養溫度對於發酵實驗之探討 57
4-2-1 Saccharomyces cerevisiae溫度耐受性測試 57
4-2-2 好氧發酵之不同通氣速率比較 59
4-2-3 兩階段溫度控制之轉換時機點 63
4-2-4 兩階段溫度轉換策略之可行性 66
4-3 Hytrel®吸附萃取能力測試 68
4-3-1 Hytrel®基本吸附能力 68
4-3-2 Hytrel®添加量之吸附影響 70
4-3-3 Hytrel®不同脫附劑測試 72
4-3-4 Hytrel®細胞毒性測試 74
4-3-5 Hytrel®萃取次數之穩定度 76
4-4 兩階段溫度轉換發酵模式結合萃取系統應用 79
4-4-1 兩階段溫度轉換饋料測試 79
4-4-2 兩階段溫度轉換發酵結合連續式萃取及饋料操作 81
第五章結論及建議 84
5-1 結論 84
5-2 建議 85
參考文獻 86

參考文獻

[1] Maximize Market Research., "Global Aroma Chemicals Market – Industry analysis and Forecast (2018-2026) by Composition, by Application and by Region.", 2018.

[2] 陳麗婷, 鄔嫣珊, "食品香料市場現況與趨勢發展", 財團法人食品工業發展研究所, 2014.

[3] K. Rusanov et al., "Microsatellite analysis of Rosa damascena Mill. accessions reveals genetic similarity between genotypes used for rose oil production and old Damask rose varieties", Theor Appl Genet., vol. 111, pp. 804-809, 2005.

[4] C. Aubert, S. Baumann, and H. Arguel, "Optimization of the analysis of flavor volatile compounds by liquid− liquid microextraction (LLME). Application to the aroma analysis of melons, peaches, grapes, strawberries, and tomatoes", Journal of agricultural food chemistry, vol. 53, no. 23, pp. 8881-8895, 2005.

[5] M. Horbowicz, W. Wiczkowski, T. Sawicki, D. Szawara-Nowak, and H. M. Sytykiewicz, "Methyl jasmonate stimulates biosynthesis of 2-phenylethylamine, phenylacetic acid and 2-phenylethanol in seedlings of common buckwheat", J Acta Biochimica Polonica, vol. 62, no. 2, 2015.

[6] I. Kirm, F. Medina, J. E. Sueiras, P. Salagre, and Y. Cesteros, "Hydrogenation of styrene oxide in the presence of supported platinum catalysts to produce 2-phenylethanol", Journal of Molecular Catalysis A: Chemical, vol. 261, pp. 98-103, 2007.

[7] S. Serra, C. Fuganti, and E. Brenna, "Biocatalytic preparation of natural flavours and fragrances", Trends Biotechnol, vol. 23, no. 4, pp. 193-8, Apr 2005.

[8] 前瞻產業研究院香精香料行業研究院, "香精香料", 2014.

[9] J. Scognamiglio, L. Jones, C. S. Letizia, and A. M. Api, "Fragrance material review on phenylethyl alcohol", Food Chem Toxicol, vol. 50 Suppl 2, pp. S224-39, 2012.

[10] M. Carlquist et al., "Process engineering for bioflavour production with metabolically active yeast – a minireview", Yeast, vol. 32, pp. 123-143, 2015.

[11] N. Dudareva, F. Negre, D. A. Nagegowda, and I. Orlova, "Plant Volatiles: Recent Advances and Future Perspectives", Critical Reviews in Plant Sciences, vol. 25, no. 5, pp. 417-440, 2006.

[12] D. M. Tieman, H. M. Loucas, J. Y. Kim, D. G. Clark, and H. J. Klee, "Tomato phenylacetaldehyde reductases catalyze the last step in the synthesis of the aroma volatile 2-phenylethanol", Phytochemistry, vol. 68, no. 21, pp. 2660-2669, 2007.

[13] S. Silver and L. Wendt, "Mechanism of action of phenethyl alcohol: breakdown of the cellular permeability barrier", Journal of bacteriology, vol. 93, no. 2, pp. 560-566, 1967.

[14] S.-P. You, J. Zhao, L. Ma, M. Tudimat, S.-L. Zhang, and T. Liu, "Preventive effects of phenylethanol glycosides from Cistanche tubulosa on bovine serum albumin-induced hepatic fibrosis in rats", Daru : journal of Faculty of Pharmacy, Tehran University of Medical Sciences, vol. 23, pp. 52-52, 2015.

[15] C. h. M. Gang Xue, Yujuan Chen, "Progress of Phenylethanol Glycosides in Plants.", Journal of Organic Chemistry Research, vol. 5(2), pp. 114-119, 2017.

[16] 趙修報，唐育岐，劉天明, "β-苯乙醇的研究進展", 中國釀造, vol. 233, 2011.

[17] B. D. Mookherjee and R. A. Wilson, "Benzyl Alcohol and β‐Phenethyl Alcohol", Kirk‐Othmer Encyclopedia of Chemical Technology, 2000.

[18] M. I. N.-M. F. White, "Recovery of 2-phenylethanol."

[19] A. L. Carroll, S. H. Desai, and S. Atsumi, "Microbial production of scent and flavor compounds", Curr Opin Biotechnol, vol. 37, pp. 8-15, 2016.

[20] M. Etschmann, D. Sell, and J. Schrader, "Medium optimization for the production of the aroma compound 2-phenylethanol using a genetic algorithm", Journal of Molecular Catalysis A: Chemical, vol. 29, no. 1-6, pp. 187-193, 2004.

[21] E. Celińska, P. Kubiak, W. Białas, M. Dziadas, and W. Grajek, "Yarrowia lipolytica: the novel and promising 2-phenylethanol producer", Journal of industrial microbiology biotechnology, vol. 40, pp. 389-392, 2013.

[22] N. Eshkol, M. Sendovski, M. Bahalul, T. Katz‐Ezov, Y. a. Kashi, and A. Fishman, "Production of 2‐phenylethanol from L‐phenylalanine by a stress tolerant Saccharomyces cerevisiae strain", Journal of applied microbiology, vol. 106, no. 2, pp. 534-542, 2009.

[23] D. Hua and P. Xu, "Recent advances in biotechnological production of 2-phenylethanol", Biotechnol Adv, vol. 29, no. 6, pp. 654-60, 2011.

[24] M. M. Etschmann, W. Bluemke, D. Sell, and J. Schrader, "Biotechnological production of 2-phenylethanol", Appl Microbiol Biotechnol, vol. 59, no. 1, pp. 1-8, 2002.

[25] M. Etschmann, D. Sell, and J. Schrader, "Screening of yeasts for the production of the aroma compound 2-phenylethanol in a molasses-based medium", Biotechnology Letters, vol. 25, no. 7, pp. 531-536, 2003.

[26] D. Stark, T. Münch, B. Sonnleitner, I. Marison, and U. v. Stockar, "Extractive Bioconversion of 2‐Phenylethanol from l‐Phenylalanine by Saccharomycescerevisiae", Biotechnology Progress, vol. 18, no. 3, pp. 514-523, 2002.

[27] D. Hua et al., "Enhanced 2-phenylethanol production from L-phenylalanine via in situ product adsorption", Biocatalysis Biotransformation, vol. 28, no. 4, pp. 259-266, 2010.

[28] J. Sikkema, J. A. de Bont, and B. Poolman, "Mechanisms of membrane toxicity of hydrocarbons", Microbiol. Mol. Biol. Rev., vol. 59, no. 2, pp. 201-222, 1995.

[29] H. Wang, Q. Dong, A. Guan, C. Meng, X. a. Shi, and Y. Guo, "Synergistic inhibition effect of 2-phenylethanol and ethanol on bioproduction of natural 2-phenylethanol by Saccharomyces cerevisiae and process enhancement", Journal of bioscience bioengineering, vol. 112, no. 1, pp. 26-31, 2011.

[30] A. Alba Pérez, "Enhanced microbial production of natural flavors via in-situ product adsorption", PhD thesis, Swiss Federal Institute of Technology Zurich (ETHZ), 2001.

[31] S.-O. Suh, J. V. McHugh, D. D. Pollock, and M. Blackwell, "The beetle gut: a hyperdiverse source of novel yeasts", Mycological research, vol. 109, no. Pt 3, pp. 261-265, 2005.

[32] 許瑞純, "探討以PDMS海綿萃取發酵系統連續生產乙醇之研究", 碩士, 中央大學化材所, 2016.

[33] I. Herskowitz, "Life cycle of the budding yeast Saccharomyces cerevisiae", Microbiological reviews, vol. 52, no. 4, p. 536, 1988.

[34] 孫萬儒, "酵母菌", 生物學通報, 第42卷第11期, pp. 5-10, 2007.

[35] L. Gethins et al., "Influence of carbon and nitrogen source on production of volatile fragrance and flavour metabolites by the yeast Kluyveromyces marxianus", Yeast, vol. 32, no. 1, pp. 67-76, 2015.

[36] Z. Cui, X. Yang, Q. Shen, K. Wang, and T. Zhu, "Optimisation of biotransformation conditions for production of 2-phenylethanol by a Saccharomyces cerevisiae CWY132 mutant", Natural product research, vol. 25, no. 7, pp. 754-759, 2011.

[37] M. Jianfeng, "Breeding of yeast strain for production of 2-phenylethanol by biotransformation", FOOD FERMENTATION INDUSTRIES, vol. 33, no. 5, p. 22, 2007.

[38] Z. Ciesarova, D. Šmogrovičová, and Z. Dömény, "Enhancement of yeast ethanol tolerance by calcium and magnesium", Folia microbiologica, vol. 41, no. 6, pp. 485-488, 1996.

[39] M. Jianfeng, M. Hang, and L. Zhenmei, "Biocatalytic synthesis of 2-phenylethanol by yeast cells", Chinese Journal of Catalysis, vol. 28, no. 11, pp. 993-998, 2007.

[40] S. Ghosh, B. W. Kebaara, A. L. Atkin, and K. W. Nickerson, "Regulation of aromatic alcohol production in Candida albicans", Appl. Environ. Microbiol., vol. 74, no. 23, pp. 7211-7218, 2008.

[41] 陳怡君, "探討藉PDMS海綿萃取發酵系統提升Saccharomyces cerevisiae生產苯乙醇之研究", 碩士, 中央大學化材所, 2017.

[42] T. W. Nagodawithana, C. Castellano, and K. H. Steinkraus, "Effect of dissolved oxygen, temperature, initial cell count, and sugar concentration on the viability of Saccharomyces cerevisiae in rapid fermentations", Appl. Environ. Microbiol., vol. 28, no. 3, pp. 383-391, 1974.

[43] 黃俊凱, "探討光照對Saccharomycescerevisiae生產乙醇之影響", 碩士, 中央大學化材所, 2008.

[44] C.-J. Huang, S.-L. Lee, and C.-C. Chou, "Production of 2-phenylethanol, a flavor ingredient, by Pichia fermentans L-5 under various culture conditions", Food research international, vol. 34, no. 4, pp. 277-282, 2001.

[45] F. Gao and A. J. Daugulis, "Bioproduction of the aroma compound 2‐phenylethanol in a solid–liquid two‐phase partitioning bioreactor system by Kluyveromyces marxianus", Biotechnology bioengineering, vol. 104, no. 2, pp. 332-339, 2009.

[46] K. Chreptowicz, M. Wielechowska, J. Główczyk-Zubek, E. Rybak, and J. Mierzejewska, "Production of natural 2-phenylethanol: From biotransformation to purified product", Food bioproducts processing, vol. 100, pp. 275-281, 2016.

[47] M. M. Etschmann, D. Sell, and J. Schrader, "Production of 2‐phenylethanol and 2‐phenylethylacetate from l‐phenylalanine by coupling whole‐cell biocatalysis with organophilic pervaporation", Biotechnology bioengineering, vol. 92, no. 5, pp. 624-634, 2005.

[48] M. Herrero, J. A. Mendiola, A. Cifuentes, and E. Ibáñez, "Supercritical fluid extraction: Recent advances and applications", Journal of Chromatography a, vol. 1217, no. 6, pp. 2495-2511, 2010.

[49] C. E. Fabre, J. S. Condoret, and A. Marty, "Extractive fermentation of aroma with supercritical CO2", Biotechnology bioengineering, vol. 64, no. 4, pp. 392-400, 1999.

[50] D.-l. Hua et al., "Extractive Bioconversion of L-Phenylalanine to 2-Phenylethanol Using Polypropylene Glycol 1500", Asian Journal of Chemistry, vol. 25, no. 11, pp. 5951-5955, 2013.

[51] D. Serp, U. Von Stockar, and I. Marison, "Enhancement of 2‐phenylethanol productivity by Saccharomyces cerevisiae in two‐phase fed‐batch fermentations using solvent immobilization", Biotechnology bioengineering, vol. 82, no. 1, pp. 103-110, 2003.

[52] X. Qian et al., "Current status and perspectives of 2-phenylethanol production through biological processes", Critical reviews in biotechnology, vol. 39, no. 2, pp. 235-248, 2019.

[53] R. G. Berger, "Flavours and fragrances: chemistry, bioprocessing and sustainability", Springer Science & Business Media, 2007.

[54] D. Stark, D. Zala, T. Münch, B. Sonnleitner, I. Marison, and U. Von Stockar, "Inhibition aspects of the bioconversion of L-phenylalanine to 2-phenylethanol by Saccharomyces cerevisiae", Enzyme Microbial Technology, vol. 32, no. 2, pp. 212-223, 2003.

[55] H. Wang, Q. Dong, C. Meng, X. ai Shi, and Y. Guo, "A continuous and adsorptive bioprocess for efficient production of the natural aroma chemical 2-phenylethanol with yeast", Enzyme microbial technology, vol. 48, no. 4-5, pp. 404-407, 2011.

[56] J. Mei, H. Min, and Z. Lü, "Enhanced biotransformation of L-phenylalanine to 2-phenylethanol using an in situ product adsorption technique", Process Biochemistry, vol. 44, no. 8, pp. 886-890, 2009.

[57] C. Fabre, "Extraction of 2-phenylethyl alcohol: by techniques such as adsorption, inclusion, supercritical CO2, liquid-liquid and membrane separations", Perfum. Flavor., vol. 21, pp. 27-40, 1996.

[58] S.-Y. Kwak and N. Nakajima, "Morphology formation in mixing of copolyester thermoplastic elastomer (Hytrel) with poly (vinyl chloride) and nuclear magnetic resonance relaxation study on solid structures of the mixture", Macromolecules, vol. 29, no. 10, pp. 3521-3524, 1996.

[59] M. Montes, A. J. Daugulis, M. C. Veiga, and C. Kennes, "Characterization of absorbent polymers for the removal of volatile hydrophobic pollutants from air", Journal of Chemical Technology Biotechnology, vol. 86, no. 1, pp. 47-53, 2011.

[60] G. P. Prpich and A. J. Daugulis, "Polymer development for enhanced delivery of phenol in a solid‐liquid two‐phase partitioning bioreactor", Biotechnology progress, vol. 20, no. 6, pp. 1725-1732, 2004.

[61] G. L. Miller, "Use of dinitrosalicylic acid reagent for determination of reducing sugar", Analytical chemistry, vol. 31, no. 3, pp. 426-428, 1959.

[62] S. Saerens, P. Verbelen, N. Vanbeneden, J. Thevelein, and F. Delvaux, "Monitoring the influence of high-gravity brewing and fermentation temperature on flavour formation by analysis of gene expression levels in brewing yeast", Applied microbiology biotechnology, vol. 80, no. 6, pp. 1039-1051, 2008.

指導教授

徐敬衡(Chin-Hang Shu)

審核日期

2019-8-8

推文