乳酸预发酵抑菌保存对餐厨垃圾产丁醇的影响机制及应用研究

The objective of this study is to establish a high efficient and productive butanol production system from food waste. Firstly, lactic acid per-fermentation was carried out as an antimicrobial preservation procedure to avoid the carbon losses from rotten food waste, because partial carbon source would be converted to lactic acid and fixed in the system. On the other hand, ABE-producing clostridia have been reported to utilize exogenous lactic acid as co-substrate to produce butanol with sugar. Thus, in this study, we considered to couple lactic acid per-fermentation with ABE fermentation as a combined novel process for butanol production from food waste. Furthermore, based on the composition of food waste, optimal enzymatic hydrolysis parameters and butanol-producing clostridium were also conducted in this study. To elucidate the mechanism of carbon flux, 13C tracer experiment was introduced to analyze the metabolism in the complex food waste fermentation system. Further, quantitative analysis of carbon flux distribution during lactic acid per-fermentation, and the following co-carbon sources metabolized ABE fermentation could be achieved. To apply this novel butanol production process, pH-stat fed-batch culture mode and continuous culture mode with cell recycling (high cell density) under pH control will be utilized to develop the fermentation system. In this research project, a well-constructed waste to energy conversion system will be established to provide a reliable theoretical basis and detailed experimental data for the realization of the butanol industrialized production.

本研究以实现高效高速的餐厨垃圾发酵生产丁醇为目标。首先采用乳酸预发酵的方法对餐厨垃圾进行抑菌保藏，减少其因腐败酸化造成的碳源损失，将部分碳源固定到乳酸分子中，结合产丁醇发酵菌株可同时利用乳酸和糖类共同发酵产丁醇的特点，实现高碳转化率的高效丁醇生产。另外，本研究拟针对餐厨垃圾碳源组成的特点，优化酶解糖化工艺，优选适于高效利用乳酸产丁醇的梭状芽孢杆菌属作为生产菌。在机理研究方面，将13C同位素示踪法引入到餐厨垃圾复杂体系中，定量分析乳酸预发酵过程，以及乳酸糖类共碳源代谢发酵过程中碳流分布情况。在工艺应用开发方面，研究采用pH-stat自控式补料发酵，pH调控高细胞密度连续发酵的模式，实现高速率丁醇生产。本研究为产丁醇系统的快速、高效运行提供可靠的调控策略和科学依据。构建良好的废物－能源的转化系统，为实现餐厨垃圾产丁醇的工业化提供可靠的理论依据和翔实的实验数据。

本研究考察餐厨垃圾乳酸预发酵耦合丁醇发酵过程中几个关键性技术及科学问题，针对有机酸和糖类作为联合碳源进行丁醇发酵的特点，开发乳酸预发酵联合共碳源产丁醇发酵的工艺。最大限度实现餐厨垃圾的减量化、资源化及能源化，为实现餐厨垃圾转化为生物燃料丁醇这一高附加值产品提供可靠的理论依据和详实的数据基础。本研究建立了餐厨垃圾不经过任何酶制剂糖化处理，直接进行高效产丁醇发酵的工艺，单批次发酵产醇浓度可达12.1 g/L，碳转化率0.402 C-mol/C-mol，最大生产速率0.705 g/L/h。研究探索乳酸预发酵的方式实现餐厨垃圾保藏过程中的抑菌固碳，优选抑菌效果优异的植物乳杆菌作为同型乳酸预发酵作用菌，强化接种24 h后餐厨垃圾中乳酸浓度达13.4 g/L，较空白对照组提高2倍，接种7 h后pH值降至4.5抑制腐败菌株生长代谢，实现抑菌固碳。进一步探索餐厨垃圾淀粉与固碳乳酸共碳源发酵的可行性，发现初始乳酸浓度15 g/L时，丁醇产量达15.2 g/L；而初始乳酸浓度高于15 g/L时发酵受到明显抑制；此外，乳酸不能作为唯一碳源，但可作为共碳源与淀粉一起被菌株N1-4代谢消耗。研究表明以实际收运保藏条件下，利用植物乳杆菌强化乳酸预发酵联合直接产丁醇发酵工艺可明显抑制餐厨垃圾腐败并强化碳源向丁醇方向转化，最终丁醇产量14.8 g/L，最大体积生产速率为1.02 g/L/h。采用植物乳杆菌对餐厨垃圾进行强化乳酸预发酵的保藏方法，实现餐厨垃圾在收运过程中的抑菌固碳，减少碳源损失率达10%；初步建立餐厨垃圾乳酸预发酵耦合菌株N1-4进行共碳源ABE发酵的高效产丁醇模式，批次丁醇产量超过15 g/L。本研究开发了乳酸预发酵联合直接厌氧ABE发酵，即共碳源产丁醇新工艺，为丁醇生产找到廉价原料的同时，为餐厨垃圾资源化提供新途径。