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固态培养基汽爆灭菌及发酵过程强化
赵志敏
学位类型博士
导师陈洪章
2016-07
学位授予单位中国科学院研究生院
学位授予地点北京
学位专业生物化工
关键词汽爆灭菌 固态发酵 压力脉动 热量传递 过程强化
摘要

固态发酵是一种清洁环保生产方式。固态发酵培养基是以固相为骨架支撑结构,液相与固相紧密结合,而气相作为连续相的三相体系。由于空气热导率低,基质体系不均一等本征特性,致使固态发酵存在灭菌困难,营养物质破坏严重,发酵热累积,过程监测困难,自动化水平低等系列问题。导致固态发酵虽环境友好,但工业化进展缓慢的局面。针对以上问题,论文研究汽爆作为固态培养基新型灭菌方法的可行性,并系统考察汽爆灭菌对固态培养基理化特性及发酵水平的影响。研发新型变频压力脉动强化固态发酵方法,促进基质热量传递。建立数字图像处理监测体系,对固态发酵过程参数进行快速监测。基于以上研究成果,并研发自动控制系统,进行集成创新,实现汽爆灭菌新型固态发酵产业化应用。论文取得了如下研究结果:(1) 针对固态培养基灭菌困难,营养物质破坏严重的问题,建立了高温短时汽爆灭菌新方法。首先,研究了汽爆温度、维压时间对固态培养基灭菌效果的影响,确定汽爆灭菌条件为172 °C * 2 min和128 °C * 5 min。发现汽爆灭菌动力学曲线非线性,表明汽爆灭菌对杀灭微生物具有多重作用。一方面,高压维持阶段产生热力灭菌效果;另一方面,瞬间泄压阶段对微生物细胞结构具有损伤作用,证实了汽爆灭菌快速有效。此外,考察了汽爆灭菌对培养基营养成分的影响,发现在汽爆条件172 °C * 2 min,128 °C * 5 min下,固态培养基中葡萄糖含量分别为0.035 g/g干重,0.054 g/g干重,较常规蒸汽灭菌121 °C * 20 min条件分别提高了66.7%,157.1%,表明汽爆灭菌有效改善固态培养基营养。最后,对比分析了大规模汽爆灭菌和常规蒸汽灭菌效率及能耗,由于汽爆灭菌提高灭菌温度并省去冷却阶段,较常规蒸汽灭菌缩短时间69.8%,蒸汽消耗减少1.66%。基于本研究成果,成功实现汽爆灭菌工业化。(2) 揭示汽爆灭菌对固态培养基物理特性的影响。研究了汽爆灭菌、常规蒸汽灭菌后固态培养基中水分分布状态,进而反映灭菌对培养基物理结构的影响。发现汽爆灭菌显著改变固态培养基物理结构,汽爆灭菌后培养基中出现结合水,且毛细水和腔内水低场核磁弛豫时间延长,表明汽爆灭菌破坏了培养基中大分子结构暴露出亲水基团,同时扩大孔和腔体积。通过主成分分析等进一步研究发现,孔是微生物生长的活性位点,汽爆灭菌改善了固态培养基多孔性,为微生物生长提供更多有效空间,较常规蒸汽灭菌提高固态发酵水平2.90倍。(3) 针对固态发酵过程热量累积问题,发明了新型变频压力脉动强化固态发酵方法。基于微生物在不同发酵时期动态产热规律,建立气相双动态反应器热量平衡模型,进而确定变频压力脉动策略。考察了变频脉动下热量传递性能和发酵水平,发现在同等能耗下,变频脉动较常规恒频脉动有效促进培养基散热,基质温度上升幅度降低40.5%,纤维素酶活提高9.34 %。因此,变频压力脉动是气相双动态固态发酵优化操作方式。(4) 建立固态发酵过程数字图像处理快速监测体系。基于微生物生物量与发酵体系图像灰度平均值的对应量化关系,确立了数字图像处理快速监测固态发酵生物量方法,具有快速、准确、成本较低等优点。采用组态软件MCGS实现了对气相双动态反应器的自动控制,对温度、湿度、压力等环境参数进行实时测定。研发了大规模气相双动态固态发酵自动控制方案,促进提升固态发酵生产自动化水平。 (5) 基于以上研究,实现汽爆灭菌新型固态发酵工程产业化应用。将汽爆灭菌,变频压力脉动,自动控制系统耦合,通过工艺工程设计,集成创新,在生态肥料新型固态发酵产业化中得到应用。 

其他摘要

Solid-state fermentation (SSF) is an eco-friendly process. Solid phase plays a role of supporting framework which is covered by a thin film of liquid phase while gas phase is continuous in solid state medium. Thermal conductivity of air is low and the solid state medium is not uniform, leading to difficult sterilization, nutrients degradation, fermentation heat accumulation, inadequate monitoring and automatic control in SSF. Therefore, SSF industrialization progress is slow. To solve the above problems, a novel sterilization strategy, high-temperature and short-time steam explosion sterilization, was exploited for solid state medium in this thesis. Effects of steam explosion sterilization on medium physicochemical properties and fermentation performance were further investigated. Moreover, a new variable pressure pulsation frequency method in gas double-dynamic solid-state fermentation was invented based on heat balance model to enhance heat transfer during fermentation process. Finally, digital image analysis system was established to monitor SSF process. Based on above research achievements with automatic control program development, SSF industrialization was achieved.The main research results obtained are as follows:(1) A new steam explosion sterilization method for solid state medium was developed. First, effects of steam explosion temperature and residence time on sterilization efficiency were investigated. Steam explosion conditions for complete sterilization were 172 °C for 2 min and 128 °C for 5 min. Steam explosion sterilization kinetics curve exhibited a non-linear behavior, which indicated that steam explosion has multiple targets of action on microbial cells. On one hand, thermal sterilization effect occurred during the high-temperature maintenance stage. On the other hand, microbial cells were damaged by the instant pressure release. These results indicated that steam explosion sterilization improved sterilization efficiency. Second, effects of steam explosion sterilization on nutrients contents were studied. Glucose content in medium under steam explosion sterilization conditions of 172 °C for 2 min, 128 °C for 5 min were 0.035 g/g dry medium, 0.054 g/g dry medium, which increased by 66.7% and 157.1% respectively compared with that after conventional thermal sterilization (121 °C for 20 min). Results indicated that steam explosion sterilization improved solid state medium nutrition effectively. Finally, sterilization efficiency and steam consumption were compared between steam explosion sterilization and conventional thermal sterilization from industrial scale view. Since temperature was increased and cooling stage was canceled, sterilization time was shortened by 69.8% and steam consumption was reduced by 1.66% in steam explosion sterilization. Based on above research results, steam explosion sterilization industrialization was achieved.(2) Physical structure changes of solid state medium were investigated to reveal effects of steam explosion sterilization on SSF. Water states in solid state medium after steam explosion sterilization and conventional thermal sterilization were compared to clarify medium structure variations. Steam explosion sterilization significantly changed physical structure of solid state medium. T2b fraction (bound water) generated and T21 (capillary water), T22 (lumen water) fraction relaxation times increased in steam explosion sterilized medium, which indicated that steam explosion exposed hydrophilic groups at molecular level and enlarged pores and cavities at three-dimensional structural level. It was interesting to find that pores where capillary water located were the active sites for microbial growth, due to the close relationship among capillary water relaxation time, specific surface area and viable cell number. Therefore, steam explosion sterilization increased the effective contact area for microbial cells on solid state medium, which increased SSF productivity by 2.90 times than that of conventional thermal sterilization.(3) Variable pressure pulsation frequency method in gas double-dynamic solid-state fermentation was invented to enhance heat transfer during fermentation process. Heat balance model integrating pressure pulsation frequency, microbial metabolism, evaporative cooling and heat exchange with air was established. Variable pressure pulsation frequency was then optimized based on the heat balance model. Heat transfer and fermentation performance were investigated under the variable pressure pulsation frequency. The medium temperature increase extent reduced by 40.5% while the fermentation production (cellulase activity) increased by 9.23% under the variable frequency than that under conventional constant frequency. Therefore, the variable frequency was the preferred operation mode than the conventional constant one in gas double-dynamic solid-state fermentation. (4) Digital image analysis system for rapid determination of SSF process parameters was established. Based on the relationship between microbial biomass and average gray value of the medium, microbial biomass was determined by digital image analysis. With advantages of reasonable accuracy and low cost, this on-line and non-destructive method would provide much convenience for microbial biomass rapid determination and process optimization in SSF. Automatic control software for laboratory-scale gas double-dynamic solid-state fermentation bioreactor was developed on MCGS platform. Temperature, humidity and pressure were monitored in real time. Furthermore, automatic control logic diagram for industrial-scale fermentation process was designed to promote SSF industrialization. (5) SSF industrialization was achieved based on the above research achievements. Steam explosion sterilization, variable pressure pulsation frequency strategy and automatic control in gas double-dynamic solid-state fermentation were integrated for ecological fertilizer industrial production, which promote the modern SSF progress. 

语种中文
文献类型学位论文
条目标识符http://ir.ipe.ac.cn/handle/122111/22917
专题研究所(批量导入)
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赵志敏. 固态培养基汽爆灭菌及发酵过程强化[D]. 北京. 中国科学院研究生院,2016.
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