基于嗜热菌强化水解的抗生素菌渣氮源回收资源化利用研究

Antibiotics bacterial residue, which is a special kind of hazardous waste, has large production and potential environmental risk. Thus the rational and effective treatment methods for antibiotics bacterial residue have become hot issue in current researches. Solubilization by thermophilic enzyme (S-TE) pretreatment is relyed on thermophilic enzyme secreted by thermophilic bacteria. It could accelerate the dissolve of bacteria which could make a large number of organic dissolution stored in the intracellular or extracellular polymer, to improve the resource recycling of nitrogen source. The main contents of this project include the followings: (1) Under the condition of high temperature, the thermophilic bacteria which have the thermal stability of enzyme were screened from domesticated sludge and cephalosporin bacterial residue. And the performance and mechanism of the thermophilic bacteria of hydrolysis of bacterial residue were studied to establish of the research system and to elucidate the mechanism of the enhanced effects of thermophiles on cephalosporin bacterial residue hydrolysis; (2) By adding metal ions as hydrolytic enzyme activator or inhibitors, the hydrolysis effect on cephalosporin bacterial residue by metal ions was be investigated to establish of the research system of the enhanced thermophiles hydrolysis effects by metal ion on cephalosporin bacterial residue hydrolysis; (3) Precipitation separation methods were used for protein separation from precipitate protein hydrolysate and the precipitation separation effect of the methods was compared. The research system of the precipitation and separation of solid protein from hydrolyzed protein thermophiles of hydrolysis cephalosporin bacterial residue for was established. This study is valuable to lay the theoretical foundation of for rapid hydrolysis and open up a new way to realize the effective resource utilization of antibiotics bacterial residue.

抗生素菌渣属于特殊危险废物，其产量大且存在潜在环境风险，如何对其进行合理、有效的处理处置成为当今研究的热点问题。嗜热酶溶解技术可快速提升菌体水解速率，使大量储存于胞内或胞外聚合物的有机物溶出，提高其中氮源的资源化回收。本项目开展如下研究：（1）高温条件下驯化剩余污泥及头孢菌素菌渣筛选具有热稳定酶的嗜热菌，对菌渣强化水解性能以及机理进行研究，建立嗜热菌强化头孢菌素菌渣水解的研究体系，阐明嗜热菌强化头孢菌素菌渣水解的机理；（2）添加金属离子作为水解特定酶的激活剂或抑制剂，考察各离子对菌渣的嗜热菌水解效果的影响，建立金属离子强化嗜热菌水解头孢菌素菌渣的研究体系；（3）采用沉淀分离方法对蛋白质水解液进行沉淀分离其中的蛋白，比较其沉淀分离效果，建立嗜热菌水解头孢菌素菌渣制取的蛋白质水解液沉淀分离固态蛋白研究体系。本项目能为抗生素菌渣的快速水解奠定理论基础，为实现其有效资源化利用开辟新的技术途径。

抗生素菌渣属于特殊危险废物，其产量大且存在潜在环境风险，如何对其进行合理、有效的处理处置成为当今研究的热点问题。本课题分离鉴定优势水解嗜热菌，开展环境因素对其生长及产酶特性的影响，并将其水解抗生素菌渣，考察不同处理方式和条件其对菌渣水解效果的影响。最后对水解液中蛋白质进行沉淀分离研究，考察不同沉淀法对蛋白质的沉淀分离效果。取得主要结果如下：.1）60℃下从中高温堆肥基质中分离纯化出8株具有蛋白酶和淀粉酶活性的嗜热菌DF1~DF8。以两酶活性最高的DF7为模式菌，经16SrDNA鉴定其属于嗜热芽孢杆菌属（Bacillus thermocopriae）。对其进行溶胞性能验证，DF7对大肠杆菌（E. coli）溶解率在14h时达到67%，是对照组的3.2倍。与此同时，在DF7上清液投加量为35%（V/V）时，其对头孢菌素菌渣的水解效果最好，水解率可达74.99%，较对照组提高31.41%。.2）嗜热菌强化头孢菌渣溶胞效能试验表明：嗜热菌DF7最佳水解温度为60℃，最佳投加比为10%（V/V），FDA水解酶活性可达到333.45 μgFDA/mL∙h，是对照组的3.2倍。SCOD、氨氮和溶解性蛋白质最大值分别为16255 mg/L、611.4mg/L和182.46 mg/L，分别是对照组的1.1、1.2和1.4倍。金属离子强化嗜热菌水解菌渣结果表明，[K+]为200μmol/L时水解效果最好，菌渣的SCOD和氨氮浓度分别达到18285 mg/L和570.42 mg/L，相比空白分别增加了12.66%和13.28%。.3）TCA沉淀法对蛋白质沉淀效果最好。通过单因素实验，研究TCA质量分数，水解液pH、沉淀时间和离心转数对蛋白质沉淀率的影响。在单因素实验的基础上通过Box-Behnken响应曲面法对反应条件进行优化，建立了基于蛋白质沉淀率的二次函数预测模型。结果表明，蛋白质分离沉淀的最佳工艺条件为：TCA质量分数20%、提取液pH 3.0、沉淀时间10 h和离心转数10000r/min，此条件下蛋白质的沉淀率为89.5%。