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摘要：本论文主要研究了运用酶技术和微生物发酵技术，生产制备天然烟用香料。通过测定水解液中还原糖含量，确定了纤文素酶和果胶酶的最适酶解条件，包括酶解温度、pH值、料液比、加酶量、酶解时间五个单因素条件，确定单因素最佳条件后，将纤文素酶和果胶酶复合，确定复合酶的配比，得出果胶酶加酶量最佳值为0.01%(5u/g)、纤文素酶最佳加酶量为0.02%(80u/g)，果胶酶：纤文素酶为1:2；在此条件下，进行复合酶的四因素三水平的正交试验（因素有：温度、pH值、酶解时间、料液比）正交结果进行直观分析得出各因素对酶解效果的影响程度大小，并且确定复合酶解的最佳条件，复合酶酶解烟粉的最佳工艺条件为：在温度55℃、pH值3.5、料液比1:10的条件下酶解处理1.5h。按照纤文素酶和果胶酶复配的最佳条件水解烟草，运用生香酵母菌、乳酸菌、根霉菌发酵水解液，制备天然烟用香料并对比了单菌种发酵和混菌发酵产物挥发性成分的区别，得出混菌发酵效果优于单菌种发酵效果。6763
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关键词：烟用香料；微生物发酵；纤文素酶；果胶酶； GC-MS
Preparation of natural tobacco flavor in biological method
Abstract: In this paper, we used the enzyme and microbial fementation technology to produce the natural tobacco flavor. We confirmed the optimum enzymatic hydrolysis conditions of cellulase and pectinase by measuring the reducing sugar content of hydrolyzate, including the reaction temperature, pH, solid-liquid ratio, enzyme dosage and the reaction time. After determining the optimum conditions of the single factors above, we compounded the cellulase and pectinase to find the optimal ratio of the two enzymes and we concluded that the optimum enzyme dosage of pectinase and cellulose is 0.01%(5u/g) and 0.02%(40u/g) respectively, and pectinase: cellulose is 1:2. At these conditions, the orthogonal test of four factors at three different levels were used to see the influence degree of those factors and make sure the best conditions for the compound enzyme. We concluded that the optimum conditions of compound enzymes are: reaction temperature at 55℃，pH 3.5，solid-liquid ratio 1:10，reaction time 1.5h. Using cellulose and pectinase hydrolyzed tobacco in optimal conditions, and using aroma yeast, lactic acid bacteria Rhizopus fermented hydrolysate. By this way, we prepared natural tobacco flavor and compared with the volatile components of single strain fermentation, and we concluded that mixed fermentation better than single strain fermentation .
Keywords: Tobacco Flavors；Microbial fermentation； Cellulose；Pectinase；GC-MS
目录
1. 绪论 1
1.1. 本课题国内外研究现状概况 1
1.1.1. 烟用香精香料及生产概况 1
1.1.2. 生物技术在烟草生产方面的概况 2
1.2 酶法处理烟草 3
1.2.1. 纤文素酶、果胶酶酶解烟草的可行性 4
1.2.2. 影响酶解效果的因素 4
1.3 本课题的研究目的和意义 5
2. 实验材料与方法 6
2.1. 实验材料与试剂 6
2.2. 实验仪器及设备 6
2.3. 实验方法 7
2.3.1. 工艺流程 7
2.3.2. 实验内容及步骤 7
2.4 分析测定方法 12
2.4.1. 酶解液还原糖含量测定 12
2.4.2. 发酵产物挥发性成分萃取 14