藏灵菇菌微囊化发酵剂的研制及发酵产物生化分析
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摘要
本研究是从稳定藏灵菇菌的菌相入手，开展藏灵菇菌微囊化发酵剂的研制。主要内容包括微囊制备、微囊内活菌的高密度培养、发酵条件优化和发酵产物的生化分析。
本研究确定微囊壁材为海藻酸钙，以液芯微囊法进行藏灵菇菌微囊化。选择海藻酸钠浓度、氯化钙浓度、壳聚糖浓度、搅拌速度等因素，经单因素优化及正交分析，确定藏灵菇菌微囊制备流程为：海藻酸钠浓度为0.6%，氯化钙浓度为2.0%,搅拌速度为40转/分，原菌液离心后制备为3mL菌悬液，壳聚糖的浓度为0.2%，氯化钙溶液固定时间为20-30min。将制备得到的微囊进行贮存性能检测分析，藏灵菇菌微囊化不但稳定了藏灵菇菌的菌相而且延长了藏灵菇菌的保存期限。
藏灵菇菌微囊进行高密度培养，微囊内的细胞密度达1011cfu以上，可以直接进行直投式发酵生产。藏灵菇菌发酵鲜奶的最佳发酵条件：未微囊化藏灵菇菌的发酵条件为接菌量为5%，培养温度为25℃，培养时间为25h；微囊化藏灵菇菌的发酵条件为接菌量为8%，培养温度为25℃，培养时间为25h。
未微囊化藏灵菇菌和微囊化藏灵菇菌的发酵产物的生化检测分析可知，微囊化藏灵菇菌发酵产物比未微囊化藏灵菇菌发酵产物的乳酸菌密度提高了10倍、氨基酸含量和种类增加了2-5倍。
Starting with stablizing the bacterial phase of Tibetan kefir, this study wasconducted to develop a microencapsulated Tibetan kefir starter culture.Microcapsulation of the Tibetan kefir, high-density culturation of theintra-microencapsulated living bacteria, optimization of the fermentation procedureand analysis of the biochemical characteristics of the fermentation products werecarried out.
Calcium alginate (C6H7O8Ca)n was confirmed in this study to be the wallmaterial of microencapsulation according to the physical-chemical property of wallmaterial and the Tibetan kefir were microencapsulated by liquid core microcapsulemethod. Factors influencing the microencapsulation of Tibetan kefir includingsodium alginate, CaCl2, chitosan and stirring speed were considered for thepreparation of Tibetan kefir microencapsulation. Through single factoroptimication and orthogonal experiments, the optimized preparation conditionswere: the concentration of Calcium alginate was0.6%,2.0%CaCl2, stirring speedwas of40r/min, original bacterial suspension was centrifuged for preparing the3mL bacterial suspension, chitosan concentration was0.2%, and CaCl2fixing timewas20-30min. The test of storage property on the resulting microencapsulationshowed that microencapsulation of Tibetan kefir not only stabled its bacterial phase,but also extended its storage time.
The Tibetan kefir microencapsulation was high-density cultured untilintra-microencapsulation living cell density arrived at1011cfu/g, then the direct vatset (DVS) fermentation could be carried out directly. The optimized fermentationprocedure of Tibetan kefir was: the medium inoculated with5%inoculums,fermented25h at25℃with natural Tibetan kefir as starter culture; on the otherhand, when using microencapsulated Tibetan kefir as starter culture, the inoculation ratio was8%, culture temperature was25℃and culture time was25h.
Based on the biochemical analysis about the fermentation products ofTibetan kefir with or without microencapsulation, the lactobacillus density wasincreased by ten folds, and the contents and types of amino acid were increased byfive folds in the fermentation products of microencapsulated compared tonon-microencapsulated Tibetan kefir.
本研究确定微囊壁材为海藻酸钙，以液芯微囊法进行藏灵菇菌微囊化。选择海藻酸钠浓度、氯化钙浓度、壳聚糖浓度、搅拌速度等因素，经单因素优化及正交分析，确定藏灵菇菌微囊制备流程为：海藻酸钠浓度为0.6%，氯化钙浓度为2.0%,搅拌速度为40转/分，原菌液离心后制备为3mL菌悬液，壳聚糖的浓度为0.2%，氯化钙溶液固定时间为20-30min。将制备得到的微囊进行贮存性能检测分析，藏灵菇菌微囊化不但稳定了藏灵菇菌的菌相而且延长了藏灵菇菌的保存期限。
藏灵菇菌微囊进行高密度培养，微囊内的细胞密度达1011cfu以上，可以直接进行直投式发酵生产。藏灵菇菌发酵鲜奶的最佳发酵条件：未微囊化藏灵菇菌的发酵条件为接菌量为5%，培养温度为25℃，培养时间为25h；微囊化藏灵菇菌的发酵条件为接菌量为8%，培养温度为25℃，培养时间为25h。
未微囊化藏灵菇菌和微囊化藏灵菇菌的发酵产物的生化检测分析可知，微囊化藏灵菇菌发酵产物比未微囊化藏灵菇菌发酵产物的乳酸菌密度提高了10倍、氨基酸含量和种类增加了2-5倍。
Starting with stablizing the bacterial phase of Tibetan kefir, this study wasconducted to develop a microencapsulated Tibetan kefir starter culture.Microcapsulation of the Tibetan kefir, high-density culturation of theintra-microencapsulated living bacteria, optimization of the fermentation procedureand analysis of the biochemical characteristics of the fermentation products werecarried out.
Calcium alginate (C6H7O8Ca)n was confirmed in this study to be the wallmaterial of microencapsulation according to the physical-chemical property of wallmaterial and the Tibetan kefir were microencapsulated by liquid core microcapsulemethod. Factors influencing the microencapsulation of Tibetan kefir includingsodium alginate, CaCl2, chitosan and stirring speed were considered for thepreparation of Tibetan kefir microencapsulation. Through single factoroptimication and orthogonal experiments, the optimized preparation conditionswere: the concentration of Calcium alginate was0.6%,2.0%CaCl2, stirring speedwas of40r/min, original bacterial suspension was centrifuged for preparing the3mL bacterial suspension, chitosan concentration was0.2%, and CaCl2fixing timewas20-30min. The test of storage property on the resulting microencapsulationshowed that microencapsulation of Tibetan kefir not only stabled its bacterial phase,but also extended its storage time.
The Tibetan kefir microencapsulation was high-density cultured untilintra-microencapsulation living cell density arrived at1011cfu/g, then the direct vatset (DVS) fermentation could be carried out directly. The optimized fermentationprocedure of Tibetan kefir was: the medium inoculated with5%inoculums,fermented25h at25℃with natural Tibetan kefir as starter culture; on the otherhand, when using microencapsulated Tibetan kefir as starter culture, the inoculation ratio was8%, culture temperature was25℃and culture time was25h.
Based on the biochemical analysis about the fermentation products ofTibetan kefir with or without microencapsulation, the lactobacillus density wasincreased by ten folds, and the contents and types of amino acid were increased byfive folds in the fermentation products of microencapsulated compared tonon-microencapsulated Tibetan kefir.
引文
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[2]肖仔君,杨汝德,黄国清.西藏雪莲中乳酸菌的分离鉴定及抑菌活性的研究[J]. ModernFood Science and Technology,2009,25(3):249-251.
[3]刘变芳,孔庆学,郭蔼光.自然发酵剂“西藏雪莲”的初步研究与菌种鉴定[J].中国酿造,2004,11(11):11-15.
[4]王和平,田瑞华,乌云,等.类开菲尔粒发酵特性的研究[J].乳品科学与技术,2004,109(4):153-155.
[5] Gareiela, L., Garrote, Analia, G., Abrham, Graciela, L. Chemical and microbiologicalcharacterization of kefir grains. Journal of Dairy Research,2001,68:639-652.
[6] Simova, E., Beshkova, D., Angelov, A. et al. Lactic acid bacteria and yeasts in kefir grainsand kefir made from them. Journal of Industrial Microbiology&Biotechnology,2002，28:1-6.
[7]周剑忠,黄开红,董明盛,等.开菲尔粒和西藏灵菇中细菌和酵母群落结构的比较[J].江苏农业学报,2008,24(5):634-639.
[8]周剑忠.藏灵菇微生物分子生态分析及微囊化发酵剂的研究[D].南京农业大学,2005:3-5,11-27.
[9]周剑忠,董明盛,江汉湖. PCR-DGGE指纹技术与分离技术结合筛选藏灵菇发酵过程的优势菌[J].中国农业科学,2006,39(8):1632-1638.
[10]薄芯,李京霞.西藏灵菇奶一般毒性和生殖毒性的初步探索[J].中国乳品工业,2007,35(8):28-30.
[11]刘宇峰,王金英,曲晓军,等.西藏灵菇菌的菌相菌学的研究[J].中国乳品工业,2005,33(9):35-39.
[12]薄芯,李京霞.西藏灵菇发酵奶对小鼠消化系统的保健作用探索[J].中国乳品工业,2010,7(38):29-31.
[13]李丽秋,王春敏,宋淑贤,等.西藏灵菇奶对抗生素相关性腹泻小鼠肠道菌群的调整作用[J].中国微生态学杂志,2004,12(4):187-200.
[14]李永奇.藏灵菇奶对慢性肠道疾病功能调节作用的临床观察[J].河南中医,2005,25(3):41-42.
[15]肖琳琳.乳酸菌降胆固醇作用及机理研究[D].南京农业大学，2003:20-41.
[16]陶应龙,陈青,张月明,等.食品级藏灵菇培养液对高脂血症患者血脂血压的影响[J].中国临床营养杂志,2005,13(5):273-275.
[17]冉冉,王世平,刘慧.藏灵菇源酸奶复合菌发酵剂对大鼠降胆固醇的研究[J].营养学报2009,31(1):59-62.
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