中北大学2013届毕业设计说明书
年产3万吨红曲色素生产工艺设计
摘要：红曲色素是一种优良的色素，是红曲在菌体生长代谢过程中产生的天然色素，具有较高的营养价值和药用价值，目前广泛应用于医药、食品、饲料和化妆品等领域。
本设计以从土壤中筛选的红曲霉为出发菌株，通过紫外线超声波复合诱变选育出红曲色素高产菌株，通过摇瓶筛选和稳定性试验，得到稳定性高产红曲色素的突变菌株。在此基础上，尝试了三级发酵的发酵方式，优化了培养基，确定了发酵工艺条件：接种菌龄20h、接种量5%、初始pH5.0、培养最适温度32℃及发酵时间96h，并选择了合适下游提取工艺。设计的生产工艺稳定、简便，发酵后所得的红曲色素产量达到300g/L。
关键词：红曲霉，红曲色素，发酵，诱变
中北大学2013届毕业设计说明书
Abstract：Monascus pigment is a good natural pigment, which Monascus pigment in the
cell growth in the metabolism process produces, has high nutritional value and medicinal value, is now widely used in the fields of medicine, food, feed and cosmetics etc
The design of the screening from the soil of Monascus as starting strain, through ultraviolet ultrasonic compound mutation breeding of the monascus pigment producing strains, through shake flask screening and stability test, the stability of mutant strains obtained high yield of Monascus pigment. On this basis, try the fermentation level three fermentation, optimization of the culture medium, fermentation conditions were determined: inoculating age 20h, 5% inoculation quantity, initial pH5.0, culture optimum temperature is 32 ℃ and the fermentation time was 96h, and select the appropriate downstream extraction process. Design of production process stability, simple, after fermentation of Monascus pigment yield income reached 300g/L.
Keywords: Monascus, Monascus pigment, fermentation, mutation
中北大学2013届毕业设计说明书
目录
1引言 ......................................................................................................................................... 1 1.1 概述 ..................................................................................................................................... 1 1.2 发酵方式 ............................................................................................................................. 1 1.3 红曲色素合成机理 ............................................................................................................. 1 1.4 国内外研究现状 ................................................................................................................. 2 1.4.1 国内研究现状 .................................................................................................................. 2 1.4.2 国外研究现状 .................................................................................................................. 3 1.5 本课题的意义 ..................................................................................................................... 4 2红曲霉菌种的选育 ................................................................................................................. 5 2.1红曲色素生产菌 .................................................................................................................. 5 2.2菌种描述 .............................................................................................................................. 5 2.3色价分析方法 ...................................................................................................................... 5 2.4 诱变选育 ............................................................................................................................. 5 2.4.1复合诱变 ........................................................................................................................... 6 2.4.2摇瓶筛选 ........................................................................................................................... 6 2.4.3 稳定性试验 ...................................................................................................................... 6 2.5 菌种保藏 ............................................................................................................................. 6 3 培养基 .................................................................................................................................... 7 3.1红曲霉的培养基 .................................................................................................................. 7 3.2 培养基的设计 ..................................................................................................................... 7 3.2.1 碳源 .................................................................................................................................. 7 3.2.2 氮源 .................................................................................................................................. 7 3.2.3 无机盐及微量元素 .......................................................................................................... 8 4 灭菌 ........................................................................................................................................ 9 4.1 仪器灭菌 ............................................................................................................................. 9 4.2 培养基灭菌 ......................................................................................................................... 9 4.2.1分批灭菌 ........................................................................................................................... 9
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中北大学2013届毕业设计说明书
4.2.2连续灭菌 ......................................................................................................................... 10 4.3 空气除菌 ........................................................................................................................... 10 4.4 发酵罐灭菌 ....................................................................................................................... 10 5 种子扩大培养和装料发酵 .................................................................................................. 11 5.1 斜面培养 ........................................................................................................................... 11 5.2 一级种子培养 ................................................................................................................... 11 5.3 二级种子培养 ................................................................................................................... 11 5.4 装料发酵 ........................................................................................................................... 12 6 发酵控制 .............................................................................................................................. 13 6.1 pH值的控制 ...................................................................................................................... 13 6.3 溶氧的控制 ....................................................................................................................... 13 6.4 补料工艺的控制 ............................................................................................................... 13 6.4.1磷酸与氨水 ..................................................................................................................... 14 6.4.2 葡萄糖与蛋白胨 ............................................................................................................ 14 7 下游加工 .............................................................................................................................. 16 7.1下游加工工艺的一般流程 ................................................................................................ 16 7.2发酵液的预处理与固液分离 ............................................................................................ 16 7.3提取 .................................................................................................................................... 16 7.31 胞外色素 ......................................................................................................................... 16 7.3.2 胞内色素 ........................................................................................................................ 16 7.4 成品加工与包装 ............................................................................................................... 16 8 物料衡算 .............................................................................................................................. 18 8.1 发酵罐数量衡算 ............................................................................................................... 18 8.2 发酵液组成衡算 ............................................................................................................... 19 8.3 发酵罐工艺设计 ............................................................................................................... 19 9 结果与讨论 .......................................................................................................................... 21 致谢 .......................................................................................................................................... 25
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中北大学2013届毕业设计说明书
1引言
1.1 概述
红曲色素，是以大米、大豆为主要原料，经红曲霉菌液体发酵培养、提取、浓缩、精制而成及以红曲米为原料，经萃取、浓缩、精制而成的天然红色色素。作为天然色素，红曲色素一直以来被认为是安全性较高的食用色素，试验已证明不含黄曲霉毒素。动物性试验表明，使用红曲色素及其制品的食物均未发现急、慢性中毒现象，也无致突变作用。此外，还具有降低血脂、血压，抗突变，防腐、保鲜等生理活性，因此红曲色素具有“天然、营养、多功能”的多重优点[1]。《本草纲目》中记载，红曲消食活血，健脾燥胃，治赤白痢，下水谷，治打扑伤损，治女人血气痛及产后恶血不尽。近年来，随着国内外学者对红曲色素研究的深入，其应用范围也在不断扩大。 1.2 发酵方式
目前红曲生产主要有两种工艺，一种是我国的传统工艺，即固体发酵工艺，固态发酵是指没有或几乎没有自由水存在下，在有一定湿度的水下溶性固态基质中，用一种或多种微生物的一个生物反应过程。从生物反应过程中的本质考虑，固态发酵是以气相为连续相的生物反应过程。固体发酵具有操作简便、能耗低、发酵过程容易控制、对无菌要求相对较低、不易发生大面积的污染等优点。其生产环境卫生条件差，种子纯度低，生产周期长，机械化程度低，产品色阶低，产品次品率高，出口率低。无法满足功能性红曲全程无菌的工艺条件[2]。
另一种是液体深层发酵生产红曲色素，是指在生化反应器中，模仿自然界将食药用菌在生育过程中所必需的糖类、有机和无机含有氮素的化合物、无机盐等一些微量元素以及其它营养物质溶解在水中作为培养基，灭菌后接入菌种，通入无菌空气并加以搅拌，提供食用菌菌体呼吸代谢所需要的氧气，并控制适宜的外界条件，进行菌丝大量培养繁殖的过程。工业化大规模的发酵培养即为发酵生产，亦称深层培养或沉没培养[25]。此工艺自动化程度高，污染小，色素粉色价高，但设备投资大，生产成本高，且规模小，另外该工艺只能生产红曲红色素。 1.3 红曲色素合成机理
红曲色素一直是国内外学者研究的焦点，但是其合成机理尚未研究清楚。不同氨基酸和短链脂肪酸等对色素合成的影响，结果表明缬氨酸、异亮氨酸、蛋氨酸、谷氨酸以
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