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標題: Lactobacillus reuteri 冷凍耐受性提升的探討: 細胞微膠囊化及海藻糖添加的利用
Studies on the Enhancement of the Freeze-Tolerance of Lactobacillus reuteri: the Application of Cell Microencapsulation and Trehalose Addition
作者: 徐亞莉
Hsu, Ya-Li
關鍵字: Lactobacillus reuteri
Lactobacillus reuteri
frozen storage
出版社: 食品暨應用生物科技學系所
引用: 方繼、李根永、李清福、林建谷、林順富、范晉嘉、陳惠英、虞積凱、蔡國珍 (1999) 現代食品微生物學,pp.364-369。偉明圖書有限公司,台北。 白壽雄、陳惠蓉 (1997) 天然生物保存物質-海藻糖的特性、製造及其應用。生物產業。8(1):33-42。 朱琳、劉甯、張英華、霍貴成 (2006) 乳酸菌細胞膜的凍乾損傷。食品科學。27(2): 266-269。 李福臨 (2000) 乳酸菌分類之研究近況。食品工業。32(8): 36-42。 施坤河 (1998) 能降低甜度的糖-海藻糖。烘培工業。79:57-59。 徐進財 (1977) 實用食品加工手冊,p.184。復文書局,台南。 張為憲、李敏雄、呂政義、張永和、陳昭雄、孫璐西、陳怡宏、張基郁、顏國欽、林志城、林慶文 (2004) 食品化學,pp.64-65。華香園出版社,台北。 邱雪惠 (2004) 乳酸菌之抗癌機制。食品工業。36(3):37-33。 夏慧芬 (2000b) 海藻膠的特性及於食品上的應用。食品資訊。178: 60-63。 陳慶源、黃崇真、邱雪惠、廖啟成 (2007) 乳酸菌之保健功效與產品開發。食品生技。11: 60-68。 葉娟美、張釵如 (2004) 蛋白質分子量之決定。生物化學實習手冊。國立中興大學農資學院生化核心實驗室。 廖啟成 (1998) 乳酸菌之分類及應用。食品工業。30(2): 1-10。 廖基元、蔡啟智、余進光、黃文雄 (2008) 益生菌與益菌生。台灣兒童過敏氣喘及免疫學會學會通訊。9(1): 5-7。 楊純華 (2007) 間歇性遠紅外線加熱應用於乳酸控低溫真空乾燥及冷凍乾燥的探討:2.Lacctobacillus acidophilus BCRC 10695。碩士論文。國立中興大學食品暨應用生物科技學系。 趙紅霞、李應彪 (2007) 微膠囊包埋技術在益生菌製品中的應用。中國乳品工業。10:32-34。 潘子明 (2008) 乳酸菌的保健功效。健康世界。266: 41-66。 潘子明、謝依庭 (2003) 乳酸菌降血壓之功效。生物產業。14(4): 302-309。 蔡英傑 (1998) 乳酸菌與益生菌。生物產業。9(2): 98-104。 賴茲漢、金安兒、柯慶文 (1999) 食品加工學 (方法篇)。國立中興大學出版,台中。 Angelis, M. De and Gobbetti, M. (2004) Environmental stress responses in Lactobacillus: A review. Proteomics 4(1): 106-122. Antonov, Y. and Friedrich, C. (2007) Aqueous phase-separated biopolymer mixture compatibilized by physical interactions of the constituents. Polymer Bulletin 58: 969–978. Baek, S. H., Park, S. and Lee, H. G. (2005) Hypocholesterolemic action of fermented brown rice supplement in cholesterol-fed rats: cholesterol-lowering action of fermented brown rice. Journal of Food Science 70(8):S527-S531. Beal, C., Fonseca, F. and Corrieu, G. (2001) Resistance to freezing and frozen storage of streptococcus thermophilus is related to membrane fatty acid composition. Journal of Dairy Science 84:2347–2356. Bengmark, S. (2002) Gut microbial ecology in critical illness: is there a role for prebiotics, probiotics, and synbiotics. Current Opinion in Critical Care 8:145-151. Brennan, M., Wanismail, B., Johnson, M. C. and Ray, B. Cellular damage in dried lactobacillus acidophilus. Journal of Food Protection 49(1): 47-53. Broadbent, J.R. and Lin, C. (1999) Effect of heat shock or cold shock treatment on the resistance of lactococcus lactis to freezing and lyophilization. Cryobiology 39(1): 88–102. Carvalho, A.S., Silva, J., Ho, P., Teixeira, P., Malcata, F.X. and Gibbs, P.(2004a) Effects of various sugars added to growth and drying media upon thermotolerance and survival throughout storage of freeze-dried Lactobacillus delbrueckii ssp bulgaricus. Biotechnology Progress 20: 248–254. Casas, I. A. and Dobrogosz, W. J. (1997) Lactobacillus reuteri: overview of a new probiotic for humans and animals. Microecology and Therapy 26:221–231. Casas, I. A. and W. J. Dobrogosz. (2000) Validation of the probiotic concept: Lactobacillus reuteri confers broad-spectrum protection against disease in humans and animals. Microbial Ecology and Disease 12:247–285. Champagne, C. P., Morin, N., Couture, R., Gagnon, C., Jelen, P. and Lacroix, C. (1992) The potential of immobilized cell technology to produce freeze-dried, phage-protected cultures of Lactococcus lactis. Food Research International 25:419-427. Champagne, C. P. and Fustier, P. (2007) Microencapsulation for the improved delivery of bioactive compounds into foods. Current Opinion in Biotechnology 18:184–190. Chen, C. N., Liang, H. F., Lin, M. H. and Sung, H. W.(2001) A natural sterilant (reuterin) fermented from glycerol using lactobacillus reuteri: fermentation conditions. Journal of Medical and Biological Engineering 21(4): 205-212. Cleusix, V., Lacroix, C., Vollenweider, S., Duboux, M. and Blay, G. L. (2007) Inhibitory activity spectrum of reuterin produced by Lactobacillus reuteri against intestinal bacteria. BMC Microbiology 101:1-9. Cross, M. L. (2002) Microbes versus microbes: immune signals generated by probiotic lactobacilli and their role in protection against microbial pathogens. Immunology and Medical Microbiology 34:245-253. Crowe, J. H., Crowe, L. M., Oliver, A. E., Tsvetkova, N., Wolkers, W. and Tablin F. (2001) The trehalose myth revisited: introduction to a symposium on stabilization of cells in the dry state. Cryobiology 43: 89–105 Douglas, L. C. and Sanders, M. E. (2008) Probiotics and prebiotics in dietetics practice. Journal of the American Dietetic Association 108: 510-521. Fook, L. J. and Gibson, G. R. (2002) Probiotics as modulators of the gut flora. British Journa of Nutrition 88: S39-S49. Fukishima, Y. (2007) Review: Probiotics and natural defense function of the host. Bioscience Microflora 26(1):1-10. Gänzle, M.G.. and Vogel R. F. (2003) Contribution of reutericyclin production to the stable persistence of Lactobacillus reuteri in an industrial sourdough fermentation. International Journal of Food Microbiology 80(1)31-45. Grill, J. P., Cayuela, C., Antoine, J. M. and Schneider F. (2000) Effects of Lactobacillus amylovorus and Bifidobacterium breve on cholesterol. Letters in Applied Microbiology 31: 154-156 Havenaar, R. and Huis In’t Veld, J. M. J. (1992) Probiotics: a general view. In: Wood BJB: The Lactic Acid Bacteria, Vol. 1: The Lactic Acid Bacteria in Health and Disease. pp. 209-224. Chapman & Hall, New York. He, T., Priebe, M. G., Harmsen, H. J., Stellaard, F., Sun, X., Welling, G. W. and Vonk, R. J. (2006) Colonic fermentation may play a role in lactose intolerance in humans. Journal of Nutrition 136: 58-63. Hoerr, R. A. and Bostwick, E. F.(2000) Bioactive proteins and probiotic bacteria: modulators of nutritional health. Nutrition 16(7-8): 711-713. Homayouni, A., Ehsani, M. R., Azizi, A., Yarmand, M. S. and Razavi, S. H. (2007) Effect of lecithin and calcium chloride solution on the microencapsulation process yield of calcium alginate beads. Iranian Polymer Journal 16 (9): 597-606. Hong, S. H. and Marshall, R. T. (2001) Natural exopolysaccharides enhance survival of lactic acid bacteria in frozen dairy desserts. Journal of Dairy Science 84(6): 1367–1374. Hubálek, Z. (2003) Protectants used in the cryopreservation of microorganisms. Cryobiology 46:205-229. Jauhiainen, T. and Korpela, R. (2007) Milk peptides and blood pressure. Journal of Nutrition 137: 825S–829S. Irianto, A. and Austin, B. (2002) Probiotics in aquaculture. Journal of Fish Diseases 25: 633-642. Jaya, S., Durance, T. D. and Wang, R. (2009) Effect of alginate-pectin composition on drug release characteristics of microcapsules. Journal of Microencapsulation 26(2): 143–153. Juárez Tomás, M. S., Ocaña, V. S. and Nader-Macías, M.E. (2004) Viability of vaginal probiotic lactobacilli during refrigerated and frozen storage. Anaerobe 10(1):1-5. Kailasapathy, K. and Sureeta, B. S. (2004) Effect of storage on shelf life and viability of freeze-dried and microencapsulated Lactobacillus acidophilus and Bifidobacterium infantis cultures. Australian Journal of Diary Technology 59(3):204-208. Kalfirtová, P. and Sovják, R. (2005) Improve the nutritional value of fermented dairy products by nutraceutical-producing food-grade microorganisms. Agricultura Tropica et Subtropica 38(2): 22-27. Khan, S. H. and Ansari, F. A. (2007) Review: Probiotics – the friendly bacteria with market potential in global market. Pakistan Journal of Pharmaceutical Sciences 20(1): 71-76. Khan, T., Park, J. K. and Kwon, J. H. (2007) Functional biopolymers produced by biochemical technology considering applications in food engineering. Korean Journal of Chemical Engineering 24(5): 816-826. Kourkoutasa, Y., Bekatoroua, A., Banatb, I. M., Marchantb, R. and Koutinas. A. A. (2004) Immobilization technologies and support materials suitable in alcohol beverages production: a review. Food Microbiology 21: 377–397. Krasaekoopt, W., Bhandari, B. and Deeth, H. (2003) Evaluation of encapsulation techniques of probiotics for yoghurt. International Dairy Journal 13: 3–13. Lee, H.S. and Coates G. A.(1999) Vitamin C in frozen, fresh squeezed, unpasteurized, polyethylene-bottled orange juice: a storage study. Food Chemistry 65(2): 165-168. Lee, J. S., Cha, D. S. and Park, H. J. (2004) Survival of freeze-dried Lactobacillus bulgaricus KFRI 673 in chitosan-coated calcium alginate microparticles. Journal of Agriculture and food Chemistry 56: 7300-7305. Lei, Y., Chenb, W. and Mulchandani, A. (2006) Microbial biosensors. Analytica Chimica Acta 568: 200–210. Leroy, F. and De Vuyst, L. (2004) Lactic acid bacteria as functional starter cultures for the food fermentation industry. Food Science & Technology 15: 67–78. Li, X. and Chen, X. (2009) Drying of micro-encapsulated lactic acid bacteria -Effects of trehalose and immobilization on cell survival and release properties. Journal of Ocean University of China 8(1): 39-44. Lim, I. S., Lee, H. S. and Kim, W. Y. (2009) The effect of lactic acid bacteria isolates on the urinary tract pathogens to infants in vitro. Journal of Korean Medical Science 24(1): S57-62. Lindfors, K., Blomqvist, T., Juuti-Uusitalo, K., Stenman, S., Venalainen, J., Maki, M. and Kaukinen, K. (2008) Live probiotic Bifidobacterium lactis bacteria inhibit the toxic effects induced by wheat gliadin in epithelial cell culture. Clinical & Experimental Immunology 152(3): 552-558. Ljungh, Å. and Wadström, T. (2006) Lactic acid bacteria as probiotics. Current Issues in Intestinal Microbiology 7: 73–90. Makarova, K. S. and Koonin, E. V. (2007) Evolutionary genomics of lactic acid bacteria. Journal of Bacteriology 189: 1199–1208. Marteau, P.,Seksik, P. and Jian, R. (2002) Probiotics and intestinal health effects: a clinical perspective. British Journal of Nutrition 88(l): S51–S57. Mattila-Sandholm, T., Myllärinen, P.,Crittenden, R., Mogensen, G., Fondén, R. and Saarela, M. (2002) Technological challenges for future probiotic foods. International Dairy Journal 12: 173–182. Miyake, Y., Karanis, P. and Uga S. (2004) Cryopreservation of protozoan parasitesq. Cryobiology 48: 1–7. Miyamoto-Shinohara, Y., Sukenobe, J., Imaivumi, T. and Nakahara, T. (2006) Survival curves for microbial species stored by freeze-drying. Cryobiology 52: 27-32. Moon, S., Ryu, B. Y., Choi,, J., Jo, B. W. and Farris R. J. (2009) The morphology and mechanical properties of sodium alginate based electrospun poly(ethylene oxide) nanofibers. Polymer Engineering and Science 49: 52–59. Muthukumarasamy, P., Allan-Wojtas, P. and Holley, R. A. (2006) Stability of Lactobacillus reuteri in different types of microcapsules. Journal of Food Science 71: M20-M24. Nguyen, T. D. T., Kang, J. H. and Lee M. S. (2007) Characterization of Lactobacillus plantarum PH04, a potential probiotic bacterium with cholesterol-lowering effects. International Journal of Food Microbiology 113: 358–361. Nelson, D. L. and Cox, M. M.(2000) Lehninger Principles of Biochemistry. 3rd ed, pp. 134. Worth Publishers, New York. Nikawa, H., Makihira, S., Fukushima, H., Nishimura, H., Ozaki, Y., Ishida, K., Darmawan, S., Hamada, T., Hara, K., Matsumoto, A., Takemoto, T. and Aimi, R.(2004) Lactobacillus reuteri in bovine milk fermented decreases the oral carriage of mutans streptococci. International Journal of Food Microbiology 95:219-223. Nil (2001) Health and nutritional properties of probiotics in food including powder milk with live lactic acid bacteria. Joint FAO/WHO expert consultation of evaluation of health and nutritional properties of probiotics in food including powder milk with live bacteria, 1-4 October. Córdoba, Argentina. Onsøyen, E. (1997) Aliginat. In: Imeson A, editor. Thickening and Gelling Agents for Food. 2nd ed. London UK: Blackie Academic and Professional. pp. 22-24. Otero, M.C., Espeche, M.C. and Nader-Macias, M.E. (2007) Optimization of the freeze-drying media and survival throughout storage of freeze-dried Lactobacillus gasseri and Lactobacillus delbrueckii subsp delbrueckii for veterinarian probiotic applications. Process Biochemistry 42: 1406–1411. Ouwehand, A. C., Salminen, S., Isolauri, E. (2002) Probiotics: an overview of beneficial effects. Antonie van Leeuwenhoek 82: 279-289. Parvez, S., Malik, K.A., Kang, S.A. and Kim, H.Y. (2006) Probiotics and their fermented food products are beneficial for health. Journal of Applied Microbiology 100: 1171-1185. Patist, A. and Zoerb, H. (2004) Preservation mechanisms of trehalose in food and biosystems. Colloids and Surfaces B: Biointerfaces 40: 107–113. Pereira, D. I. A. and Gibson, G.R. (2002) Cholesterol assimilation by lactic acid bacteria and bifidobacteria isolated from the human gut. Applied and Environmental Microbiology 68(9): 4689–4693. Rafter, J., Bennett, M., Caderni, G., Clune, Y., Hughes, R., Karlsson, P. C, Klinder, A., O’Riordan, M., O’Sullivan, G. C, Pool-Zobel, B., Rechkemmer,G., Roller, M., Rowland, I., Salvadori, M., Thijs, H., Loo, J. V., Watzl, B. and Collins, J. K (2007) Dietary synbiotics reduce cancer risk factors in polypectomized and colon cancer patients. American Journal of Clinical Nutrition 85: 488–96. Sabatini, N. and Marsilio, V. (2008) Volatile compounds in table olives (Olea Europaea L., Nocellara del Belice cultivar). Food Chemistry 107: 1522–1528. Santivarangkna, C., Kulozik, U. and Foerst P. (2008) Inactivation mechanisms of lactic acid starter cultures preserved by drying processes. Journal of Applied Microbiology 105:1–13. Savino, F., Pelle, E., Palumeri, E., Oggero, R. and Miniero, R. (2007) Lactobacillus reuteri (American Type Culture Collection Strain 55730) versus simethicone in the treatment of infantile colic: a prospective randomized study. Pediatrics 119(1) e124-e130. Schoug, Å., Olsson, J., Carlfors , J., Schnürer, J. and Håkansson, S. (2006) Freeze-drying of Lactobacillus coryniformis Si3—effects of sucrose concentration, cell density, and freezing rate on cell survival and thermophysical properties. Cryobiology 53(1): 119-127. Sheu, T.Y. and Marshall, R.T. (1993) Microencapsulation of lactobacilli in calcium alginate gels. Journal of Food Science 54:557-561. Singer, S. J. and Nicolson, G. L. (1972) The fluid mosaic model of the structure of cell membranes. Science 175, 720–731. Swain, M.R., Kar, S., Sahoo A. K. and Ray, R. C. (2007) Ethanol fermentation of mahula (Madhuca latifolia L.) flowers using free and immobilized yeast Saccharomyces cerevisiae. Microbiological Research 162:93-98. Temmerman, R. ,Huys, G. and Swings, J. (2004) Identification of lactic acid bacteria: culture-dependent and cultureindependent methods. Food Science & Technology 15:348–359. Teramoto, N., Sachinvala, N. D. and Shibata, M. (2008) Trehalose and trehalose-based polymers for environmentally benign, biocompatible and bioactive materials. Molecules 13: 1773-1816. Teramura ,Y. and Iwata, H. (2009) Islet encapsulation with living cells for improvement of biocompatibility. Biomaterials 30(12): 2270-2275. Tsen, J. H., Huang, H. Y., Lin, Y. P. and King, V. A. E. (2007) Freezing resistance improvement of Lactobacillus reuteri by using cell immobilization. Journal of Microbiological Methods 70: 561–564. Twomey, D., Ross, R. P., Ryan, M., Meaney, B. and Hill C. (2002) Lantibiotics produced by lactic acid bacteria: structure, function and applications. Antonie van Leeuwenhoek 82: 165–185. Valeur , N., Engel, P., Carbajal, N., Connolly, E. and Ladefoged, K. (2004) Colonization and immunomodulation by lactobacillus reuteri atcc 55730 in the human gastrointestinal tract. Applied and Environmental Microbiology 70(2): 1176-1181. Walker, W. A. (2008) Mechanisms of Action of Probiotics. Clinical Infectious iseases 46:S87–S91. Wang, W., Liu, X., Xie, Y., Zhang, H., Yu, W., Xiong, Y., Xie, W. and Ma, X. (2006) Microencapsulation using natural polysaccharides for drug delivery and cell Implantation. Journal of Materials Chemistry 16: 3252–3267. Warrand, J. (2006) Healthy polysaccharides the next chapter in food products. Food Technology and Biotechnology 44 (3): 355–370. Wollowski, I., Rechkemmer, G. and Pool-Zobe, B. L. (2001) Protective role of probiotics and prebiotics in colon cancer. American Journal of Clinical Nutrition 73(2): 451S-455S. Yang, L., Ma, Y. and Zhang Y. (2007) Freeze-drying of live attenuated Vibrio anguillarum mutant for vaccine preparation. Biologicals 35(4): 265-269. Yu, X. L.,. Li, X. B., Xu, X.L. and Zhou, G.H. (2008) Coating with sodium alginate and its effects on the functional properties and structure of frozen pork. Journal of Muscle Foods 19: 333–351. Zayed, G. and Roos, Y.H. (2004) Influence of trehalose and moisture content on survival of Lactobacillus salivarius subjected to freeze-drying and storage. Process Biochemistry 39: 1081-1086. Zhao, G. and Zhang, G. (2005) Effect of protective agents, freezing temperature, rehydration media on viability of malolactic bacteria subjected to freeze-drying. Journal of Applied Microbiology 99(2): 333-338. Zhao, R., Sun, J., Torley, P., Wang, D., and Niu, S. (2008) Measurement of particle diameter of Lactobacillus acidophilus microcapsule by spray drying and analysis on its microstructure. World Journal of Microbiology and Biotechnology 24:1349–1354. Zhu, Y., Tang, Y., Xu, H., Zhang, J., Wei, D., Xing, L. and Li, M. (2009) Cloning, expression and functional analysis of the genes in TPS/TPP trehalose synthetic pathway of Meiothermus ruber. Chinese Journal of Biotechnology 25(3): 399-405. Ziaei-Nejad, S., Rezaei, M. H., Takami, G. A., Lovett, D. L., Mirvaghefi, A.R. and Shakouri, M. (2006) The effect of Bacillus spp. bacteria used as probiotics on digestive enzyme activity, survival and growth in the Indian white shrimp Fenneropenaeus indicus. Aquaculture 252: 516–524.
摘要: Lactobacillus reuteri 為常用於保健產品製造的格蘭氏陽性菌,對於人體生理具有多項保健的功效,能產生抗菌物質(reuterin),故視為動物腸道中的重要菌種之ㄧ,在宿主健康的維護上扮演重要角色。ㄧ般利用冷凍貯藏的方式,保存食品中的活菌數,然而冷凍可能會對菌體造成傷害,因此可結合細胞微膠囊化技術及添加保護劑以減少菌體在冷凍貯藏時之傷害。本研究利用褐藻酸鈣來對L. reuteri 進行細胞微膠囊化處理,同時分別添加不同濃度(1%、2%、3%)的海藻糖作為冷凍保護劑,再放置於不同凍結溫度下(-20℃、-40℃、-60℃、-80℃、-196℃)來進行貯藏,探討其對L. reuteri在冷凍貯藏中的保護效果,以提升菌體之存活率、活性與貯藏安定性,並比較不同處理 間的差異。結果顯示,細胞經微膠囊化處理後能有效提高菌體冷凍貯藏的存活率,保護效果較佳。在凍結貯藏溫度方面,溫度對菌體的存活率的影響並無ㄧ致性,但菌體於-60℃貯藏有得到一較低的死滅速率常數,即添加2%海藻糖並經微膠囊化處理之菌體在-60℃冷凍貯藏下有較好的存活率,保護效果較佳。不同濃度海藻糖的添加,對於菌體的死滅皆有減緩的效果,且於2%的添加濃度可以獲得較小的死滅速率,亦即最高存活率。综合微膠囊化技術、不同冷凍貯藏溫度及不同濃度冷凍保護劑的添加對菌株存活率所得之結果,發現經微膠囊化處理於冷凍溫度-60℃貯藏並添加2%海藻糖,可以獲得最小死滅速率常數,此即為菌體較佳之貯藏條件。在β-半乳糖苷酶酵素活性方面,發現此酵素活性會因為菌體的死滅而減少。由細胞膜脂肪酸組成分析結果,可發現各處理組之不飽和脂肪酸含量皆增加且不飽和脂肪酸/飽和脂肪酸(U/S)比例亦增加。而蛋白質分析結果顯示各處理組皆無新的胞內蛋白質生成。利用掃描式電子顯微鏡觀察發現L. reuteri呈短桿狀(約1 μm), 且有時幾乎接近球狀,而細胞表面也出現有皺縮的現象,推測可能是菌體在經冷凍乾燥或臨界點乾燥時受到壓迫。
Lactobacillus reuteri is a gram-positive bacterium that commonly used in manufacturing health products, and possesses lots of important physiological functions beneficial to human body, and produces antibacterial material (reuterin). L. reuteri is considered a major organism of the human gastrointestinal tract and plays very important roles in the maintenance of host's health. In order to achieve the effective viable cell numbers in food, freezing storage is the most frequently used technique to preserve the cells. However, freezing will cause damage to the cell, so the cell microencapsulation technology and the addition of protectants are frequently used to reduce the cell damage during frozen storage. In this study, Ca-alginate was applied to L. reuteri for cell microencapsulation, and different concentrations (1%, 2%, 3%) of trehalose were added as cryoprotectants at the same time for the storage test at different frozen temperatures (-20℃, -40℃, -60℃, -80℃, -196℃) in order to study their protection effects on the enhancement of cell survival, activity and storage stability of L. reuteri during frozen storage and effects of different treatments were also compared. Results indicated that cell microencapsulation could effectively increase the cell survival during frozen storage, and the good protection effect was obtained. Wth respect to frozen temperatures, results indicated that there is no consistency between the temperatures and the cell survival existed. But -60℃ frozen storage could obtain a lower death rate constant among various frozen temperatures used, and microencapsulated L. reuteri with 2% trehalose addition during storage at -60℃ could obtain the highest survival. Regarding trehalose addition, various concentrations of trehalose addition could reduce cell death. 2% addition of trehalose could obtain the lowest death rate constant and the highest survival. It was found that 2% trehalose addition combined with cell microencapsulation and -60℃ frozen temperature could obtain the lowest death rate constant and were found to be the best conditions among all the treatments. Concerning β-galactosidase activity, results indicated that freezing kills many of the cells and causing loss of enzymatic activity. Results of the cellular membrane fatty acid composition analysis showed that the concentration of unsaturated fatty acids and the ratio between unsaturated and saturated fatty acids (U/S) all increased in all treatments. In SDS-PAGE analysis, no new intracellular protein was found. The scanning electron microscopic images of L. reuteri were short rod-shaped (approximately 1 μm in length), and sometimes nearly spherical, and then the cell surfaces were also appeared to be shrinked and wrinkled. Extrapolating from this trend, it''s probable that cells were subjected to the suppression of freeze drying or critical point drying.
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