发酵法生产1，3-丙二醇条件优化及相关策略研究
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摘要
1，3-丙二醇是一种重要的化工原料，主要作为单体生产聚酯(PTT)等高分子材料。1，3-丙二醇的生产方法有化学法和微生物发酵法两类。微生物发酵法以其环境友好、生产过程中不含醛类物质、产率高、产品质量好以及可再生资源为原料等优点，成为当前研究热点。为了进一步提高1，3-丙二醇产量、实现发酵的有效调控，本论文对克雷伯氏肺炎杆菌(Klebsiella pneumoniae)合成1，3-丙二醇的发酵条件优化、pH值对中心碳代谢的影响、控制3-羟基丙醛(3-HPA)保持持续发酵等进行了系统研究，从而为工艺放大和菌种的基因改造提供指导。
利用了中心组合设计的商业软件Design Expert 7．0．2 Trial对K. pneumoniae合成1，3-丙二醇的发酵条件，包括初始甘油浓度、搅拌速度、通风量以及pH值四因素进行了研究，获得了最佳的相关参数，使得批次发酵的生产强度提高，优化条件下流加补料发酵实验的发酵周期明显缩短。
以pH值作为考察因素，研究不同pH值对发酵过程中各代谢产物代谢的影响，结果显示，pH值对K. pneumoniae产物谱有明显的影响，低pH值下有机酸的合成受到抑制，较高pH值抑制2，3-丁二醇的合成。
3-羟基丙醛是甘油发酵生产1，3-丙二醇的一个中间产物，3-羟基丙醛的积累会造成发酵过程的异常终止。考察了初始甘油浓度、转速和氧化还原电势(ORP)三个因素对3-羟基丙醛积累的影响，实现了在不影响1，3-丙二醇生产强度的情况下，防止3-羟基丙醛的致死性积累。
研究了利用不同生物柴油副产物粗甘油对K. pneumoniae发酵生产1，3-丙二醇的影响。研究结果表明，生物柴油粗甘油浓度越高，1，3-丙二醇的得率和生产强度也越高，且生产单位1，3-丙二醇产品的经济效益也越高，证明了生物柴油和1，3-丙二醇耦联生产的可行性，为降低1，3-丙二醇工业化生产成本提供了依据。
1,3-Propanediol is an important chemicals mainly used for the synthesis ofpolytrimethylene terephthalate (PTT) and other polyester fibers. Compared with thechemical methods, the biological synthesis approach is increasingly favored due to theenvironmental friendly features, the high yield and the renewable feedstock. In orderto further enhance the 1,3-propanediol productivity and ensure the effectivefermentation regulation, the statistical optimization of culture conditions and thecontrol 3-hydroxypropionaldhyde(3-HPA) concentration below the toxic level to1,3-propanediol production were investigated, providing the substantial insights forthe subsequent pilot plant experiment and the metabolic analysis.
A central composite design was used to study the effect of glycerol, rate ofstirring, air aeration and pH on the synthesis of 1,3-propanediol by K. pneumoniae.The optimum combinations for 1,3-propanediol productivity was glycerol, rate ofstirring, air aeration, and pH of 50 g/L, 318 rpm, 0.6 vvm, 6.48, respectively. After theoptimization, the 1,3-propanediol productivity of the batch culture was increased. Thefermentation cycle for the fed batch culture was deceased.
The response of K. pneumoniae central carbon metabolism to environmentalpH has been studied. The results indicated that pH influenced the product spectrum.More organic acid was produced at low pH, while 2,3-butadiol synthesis was inhibtedat high pH condition.
3-Hydroxypropionaldhyde is a medium metabolite during production of1,3-propanediol. The 1,3-propanediol fermentation would terminated due to the3-hydroxypropionaldhyde accumulation. Three approaches including the initialglycerol, the stirring rate, and the oxidoreduction potential (ORP) regulation wereemployed to investigate the impacts on 3-hydroxypropionaldehyde accumulation. Thegoal of this work was to study three strategies' influences to 3-hydroxypropionaldehyde accumulation and to prevent the abnormal cessation. Theresults confirmed that the accumulation of 3-hydroxypropionaldhyde could beregulated.
The 1,3-propanediol production with the crude glycerol from biodiesel processwas investigated. The results revealed that higher concentration crude glycerolexhibited higher productivity and yield of 1,3-propanediol, The cost analysis showedhigher concentration crude glycerol could reduce the cost of production. This workwas expected to provide the basis to effectively improve utilization ratio of crudeglycerol from biodiesel process, produce biodiesel and 1,3-propanediol, and reducethe cost of production of 1,3-propanediol.
利用了中心组合设计的商业软件Design Expert 7．0．2 Trial对K. pneumoniae合成1，3-丙二醇的发酵条件，包括初始甘油浓度、搅拌速度、通风量以及pH值四因素进行了研究，获得了最佳的相关参数，使得批次发酵的生产强度提高，优化条件下流加补料发酵实验的发酵周期明显缩短。
以pH值作为考察因素，研究不同pH值对发酵过程中各代谢产物代谢的影响，结果显示，pH值对K. pneumoniae产物谱有明显的影响，低pH值下有机酸的合成受到抑制，较高pH值抑制2，3-丁二醇的合成。
3-羟基丙醛是甘油发酵生产1，3-丙二醇的一个中间产物，3-羟基丙醛的积累会造成发酵过程的异常终止。考察了初始甘油浓度、转速和氧化还原电势(ORP)三个因素对3-羟基丙醛积累的影响，实现了在不影响1，3-丙二醇生产强度的情况下，防止3-羟基丙醛的致死性积累。
研究了利用不同生物柴油副产物粗甘油对K. pneumoniae发酵生产1，3-丙二醇的影响。研究结果表明，生物柴油粗甘油浓度越高，1，3-丙二醇的得率和生产强度也越高，且生产单位1，3-丙二醇产品的经济效益也越高，证明了生物柴油和1，3-丙二醇耦联生产的可行性，为降低1，3-丙二醇工业化生产成本提供了依据。
1,3-Propanediol is an important chemicals mainly used for the synthesis ofpolytrimethylene terephthalate (PTT) and other polyester fibers. Compared with thechemical methods, the biological synthesis approach is increasingly favored due to theenvironmental friendly features, the high yield and the renewable feedstock. In orderto further enhance the 1,3-propanediol productivity and ensure the effectivefermentation regulation, the statistical optimization of culture conditions and thecontrol 3-hydroxypropionaldhyde(3-HPA) concentration below the toxic level to1,3-propanediol production were investigated, providing the substantial insights forthe subsequent pilot plant experiment and the metabolic analysis.
A central composite design was used to study the effect of glycerol, rate ofstirring, air aeration and pH on the synthesis of 1,3-propanediol by K. pneumoniae.The optimum combinations for 1,3-propanediol productivity was glycerol, rate ofstirring, air aeration, and pH of 50 g/L, 318 rpm, 0.6 vvm, 6.48, respectively. After theoptimization, the 1,3-propanediol productivity of the batch culture was increased. Thefermentation cycle for the fed batch culture was deceased.
The response of K. pneumoniae central carbon metabolism to environmentalpH has been studied. The results indicated that pH influenced the product spectrum.More organic acid was produced at low pH, while 2,3-butadiol synthesis was inhibtedat high pH condition.
3-Hydroxypropionaldhyde is a medium metabolite during production of1,3-propanediol. The 1,3-propanediol fermentation would terminated due to the3-hydroxypropionaldhyde accumulation. Three approaches including the initialglycerol, the stirring rate, and the oxidoreduction potential (ORP) regulation wereemployed to investigate the impacts on 3-hydroxypropionaldehyde accumulation. Thegoal of this work was to study three strategies' influences to 3-hydroxypropionaldehyde accumulation and to prevent the abnormal cessation. Theresults confirmed that the accumulation of 3-hydroxypropionaldhyde could beregulated.
The 1,3-propanediol production with the crude glycerol from biodiesel processwas investigated. The results revealed that higher concentration crude glycerolexhibited higher productivity and yield of 1,3-propanediol, The cost analysis showedhigher concentration crude glycerol could reduce the cost of production. This workwas expected to provide the basis to effectively improve utilization ratio of crudeglycerol from biodiesel process, produce biodiesel and 1,3-propanediol, and reducethe cost of production of 1,3-propanediol.
引文
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