【作者】 王丽英；

【导师】 王谦；

【作者基本信息】 河北大学 ， 微生物学， 2002， 硕士

【摘要】 雅致放射毛霉AS3．2778是腐乳酿造业的一株优良菌株，毛霉菌种的优劣对于腐乳的生产起极其重要的作用，但是国内外对它的研究却一直少见报道。在液体培养条件下的生长特性、酶学性质的研究报道极少，将液体毛霉菌种应用于腐乳生产上的研究还是空白。 为了改进腐乳生产中菌种制作的传统工艺，将现代发酵工程技术应用于腐乳生产，进一步实现腐乳生产工艺的自动化、规模化，同时能够提高腐乳产品的质量，本项研究尝试了毛霉AS3．2778的液体深层培养及工业规模发酵液体毛霉菌种应用于腐乳生产的研究，并对毛霉AS3．2778在液体培养条件下的生长特性、发酵条件、蛋白酶活性等进行了较为全面的探讨。 研究结果表明：在本试验条件下，雅致放射毛霉AS3．2778在以黄豆粉为原料的液体培养基中，28℃、200rpm摇瓶培养条件下可以生长得很好。在前24小时内，菌丝体大量积累，最终菌丝平均生物量(菌丝体干重)可以达到1.02g／100ml。它的生长规律基本符合逻辑斯蒂生长模型，二级发酵几乎没有迟滞期，直接进入指数生长期，36—40小时进入稳定期。在发酵滤液中具有一定的糖化淀粉酶活和脂肪酶活，并具有较高的蛋白酶活。 单一组分影响实验及方差分析表明，黄豆粉、蔗糖、磷酸二氢钾都有利于毛霉AS3．2778液体培养时滤液中蛋白酶活的提高。正交实验及分析结果表明，在一定浓度范围内，饴糖的浓度越高，越不利于蛋白酶的产生。低碳氮比有利于蛋白酶活的产生，在碳氮比为1.2左右可以得到较高的蛋白酶活。毛霉AS3．2778蛋白酶的产生需要较高的通风量，最适起始pH为6，在36小时达到产酶高峰期。检测表明滤液中的蛋白酶活一般可以达到10—20u／ml。 工业发酵及生产应用结果表明，利用豆浆和黄浆水作为培养基，毛霉AS3．2778于28℃、通风比0—12h 1:1(vvm) 12h后1:2(vvm)条件下发酵培养，在50L通用式罐和180L外循环气升式罐内24小时生物量可以分别达到24.55g／100ml和27.69g／100ml(菌体湿重)，而且外循环气升式罐较通用式设备显示出多种优势。毛霉AS3．2778液体培养菌种在腐乳生产中可以得到很好的应用，液体种发酵周期在16—24小时。当菌体湿重在20g／100ml以上，菌液稀释2倍时，可以显示出比生产用种明显的优势，使前发酵周期缩短6—8小时，形成的菌膜更致密厚实。河北大学理学硕士学位论文 研究结果证实，以工业化液体深层发酵方式培养雅致放射毛霉液体菌种应用于腐乳生产完全可行。目前本项研究已经在腐乳工业化生产中得到应用，并表现出良好的应用效果。更多还原

【Abstract】 Ar.tinomucor elegans is an excellent stain used in the fermention of sufu. The quality of Mucor stain has an extraordinary effect on the production of sufu. The research on it, however, home and abroad, has seldom been reported. What is more, there is no account of the research on the property of its growth and protease activity on the condition of liquid submerged fermentation, on the liquid mucor spawn used in sufu production.In order to improve the traditional stain production process of sufu and apply the up-to-date fermentation technique to the manufacturing process of sufu, we made the research on liquid submerged culture of Actinomucor AS3.2778 and on the industrial application of the liquid mucor spawn. By this way, we hope to improve greatly the productive process automation and its scale, and the quality of sufu production. In addition, we studied the growing characteristics, the fermerntation conditions and the protease activity of Actinomucor AS3.2778.The results of the experiments showed that Ar.tinomucor elegans AS3.2778 can grow well in the soybean powder liquid medium at 28 on a rotary shaker with a speed of 200rpm.Within 24 hours, the mycelium greatly massed and in the end, the average biomass(dry weight of mycelium) reached 1.02g/l00ml.Its growth accorded with the Growth Model of Rogistic.The second stage fermentation had nearly no Lag period and came directly into Exponential period, and Steady period after 36-40 hours.There were certain glucoamylase activity and lipase activity,and higher protease activity in the culture fluid.Single factor experiments and the variance analyses showed that soybean powder, sucrose, KH2PO4 all can improve the protease activity in the liquid. The result indicated that in a certain range, the more concentrated the maltose, the less advantageous to outcome of the protease .The adequate C:N ratio in medium is about 1.2.The lower C:N ratio is advantageous to the production of mucor protease. Better ventilative is needed for the production of the proteinase of Actinomucor elegans AS3.2778.The initial optimal pH is 6,and the protease outcome was at its height at 36 hours.The industrial fermentation and production application process showed that the biomass of Actinomucor elegans AS3.2778 was able to reach respectively 24.55g/100ml and 27.69g/100ml (wet weight of mycelia:)at 24h in the 50L generalfermentor and 180L external airlift fermentor, with soymilk and soy whey as medium,on the condition of 28 ,air condition 1:1 vvm(0-12h),l:2vvm(12h -). Airlift fermentor showed many adventages. The liquid spawn of Actinomucor elegans AS3.2778 did well in sufu production, its fermentation period being 16-24 hours. When being diluted by 2 times, the culture fluid showed obvious adventages over the productive strain, shortening the primary fermentation period by 6-8 hours and forming thicker mycelial mat.The research proved that it is totally feasible for the liquid spawn of Actinomucor elegans AS3.2778 by the industrial submerged culture to be used in the production proscess of sufu. At present, the study has been already used successfully in the actual industrial production of sufu and displayed very good applicabitity.更多还原