【作者】 傅海燕；

【导师】 曾光明； 黄国和；

【作者基本信息】 湖南大学 ， 环境工程， 2007， 博士

【摘要】 本研究从堆肥过程中筛选得到产生生物表面活性剂的细菌,并将筛选得到的铜绿假单胞菌产生的鼠李糖脂进行分离提纯及表征,然后着重探讨了它对改善有机质颗粒降解微环境的作用机制,并在此基础上,进一步将其应用于堆肥中(实验室规模),旨在提高堆肥效率,最后初步考察了它对去除堆肥中石油烃和重金属离子污染物的潜在作用。首先,本实验对堆肥中产生生物表面活性剂的细菌进行筛选,发现枯草芽孢杆菌和铜绿假单胞菌为堆肥中产生生物表面活性剂的重要菌种。同时发现,在以葡萄糖为碳源的液体培养条件下,所筛选的铜绿假单胞菌产鼠李糖脂的能力较强。对所筛选的铜绿假单胞菌产生的鼠李糖脂发酵液进行一系列的分离提纯,对得到的较纯的鼠李糖脂用液质联用进行表征,发现发酵液中含有结构不同的单鼠李糖脂及二鼠李糖脂。接着本研究将实验制得的鼠李糖脂在堆肥中的作用机制进行了探讨。在分析生物表面活性剂基本物化性质和微生物效应的基础上,通过实验的方式,从鼠李糖脂对堆肥有机质颗粒水分分布、菌体在颗粒介质中传输、有机质组分的降解的影响及鼠李糖脂在堆肥中的降解情况等角度考察了鼠李糖脂在堆肥中的作用机制。结果发现鼠李糖脂对有机质颗粒介质的水分有较好的保持性能,有利于长时间保持堆肥微生物的活性,同时加速了外加水分的下渗,对水分在介质中的分布有潜在的优化作用;一定浓度的鼠李糖脂能够减弱铜绿假单胞菌菌体在颗粒介质表面的吸附,促进菌体在有机质颗粒中的传输,能够使菌体传输到有机质颗粒介质的深层,可见鼠李糖脂的加入对堆肥介质中菌体的分布有潜在作用;从有机颗粒介质的降解实验发现,鼠李糖脂的加入还起到了增强有机质在液相中的分散的作用,同时它促进了微生物的生长和有机质成分的降解,并且颗粒降解前后扫描电镜检测表明,鼠李糖脂的参与,使降解反应发生的主要位置发生了改变;最后,鼠李糖脂的降解实验表明,鼠李糖脂在堆肥中可以被堆肥微生物所降解,但是它不是微生物所优先降解的物质。这些实验结果表明,生物表面活性剂鼠李糖脂对改善堆肥的微环境,提高堆肥效率有潜在的作用,而且它在堆肥中不会产生二次污染。在理论研究的基础上,将鼠李糖脂进一步应用到城市生活垃圾堆肥(实验室规模)过程中,考察其作用效果。首先考察了鼠李糖脂对堆肥物料中易降解的蔬菜有机质降解的影响。实验结果表明鼠李糖脂发酵液能够在一定程度上加快蔬菜基质的降解过程。接着将生物表面活性剂鼠李糖脂引入含石油烃的堆肥过程中,考察了鼠李糖脂对堆肥中普通有机质和疏水性的石油烃降解过程的影响。实验结果表明生物表面活性剂的加入不仅提高了堆肥效率,缩短中温条件下堆肥过程达到稳定时间,同时它对提高堆肥中石油烃的降解效果显著,这一结果和土壤修复中生物表面活性剂促进石油烃类污染物降解的理论吻合。堆肥中重金属是影响堆肥质量的一个关键因素,而堆肥腐殖酸对重金属的迁移起着关键的作用,本研究最后考察了鼠李糖脂对腐熟段堆肥腐殖酸吸附重金属离子Pb2+的影响。实验结果发现,鼠李糖脂的加入可能改变了铅离子在堆肥腐殖酸上的吸附性状,当鼠李糖脂浓度较高时,它减弱了腐殖酸对Pb2+的吸附作用。本研究课题考察了生物表面活性剂鼠李糖脂的产生、分离提纯及其部分物化特性,同时着重对其在堆肥中的作用机制及应用进行研究,取得了一定的研究成果。本课题为较新的研究方向,对科研工作者有一定的参考价值。更多还原

【Abstract】 In this study, the bacteria which produce biosurfactant were isolated from compost community. The biosurfactant rhamnolipid was produced by an isolate, Pseudomonas aeruginosa, and its purification and characterization were carried out. The functions for rhamnolipid to improve the microenvironment for degradation of organic granules were investigated. Based on these functions, it was further applied to the composting to determine its effect on enhancement of composting efficiency. Finally its potential effect to enhance removal of the petroleum hydrocarbon and metal from composting was primarily investigated.Firstly isolation of biosurfactant producers from composting was carried. Result indicated Bacillus subtilis and Pseudomonas aeruginosa are the important bacteria that produce biosurfactant in composting, in which the Pseudomonas aeruginosa has stronger capability of producing the biosurfactant product in liquid culture with glucose as the sole carbon source. The consequential separation and purification of the product, rhamnolipid, from the culture was performed and HPLC-MS was applied to the purified rhamnolipid for its characterization. Monorhamnolipid and dirhamnolipid with respectively one and two rhamnosyl group in the molecular structure were found to be major components.Next the potential functions of rhamnolipid in composting were investigated experimentally. Based on the analysis on physico-chemical properties and microbial effects presented by literatures, the rhamnolipid was assumed to have effect on water distribution and preservation, bacterial mobility and transportation as well as on organic matter degradation manner in compost granule matrix. The self-degradation of rhamnolipid in composting was also studied. The result showed that rhamnolipid attenuated water evaporation from the matrix which may favor preservation of microbial activity for longer time. It also enhanced water fluidity in the matrix and accelerated its downflow to deep layer of the matrix, which may potentially optimize distribution of added water in the composting. Rhamnolipid at certain concentrations could weaken adsorption of Pseudomonas aeruginosa cells on the granules and may potentially enhance transportation of the bacteria to deep layers or pores in compost matrix, that is, it may favor distribution of local or added bacteria. The organic granule degradation experiment showed that it even enhanced dispersion of organic matter to the aqueous phase in the matrix and meantime it enhanced microbial growth and degradation of organic matter. The SEM detection indicated that interference of rhamnolipid caused translocation of the degradation reaction. The experiment of rhamnolipid self-degradation showed that rhamnolipid could be degraded by compost microorganisms, but not preferentially to other readily degradable substrates. These experimental results indicate that biosurfactant have the potential to improve the microenvironment of compost and enhance composting effect, and it will not cause secondary pollution.Based on the result of the mechanism study, the rhamnolipid was finally applied to composting of municipal solid waste (laboratory scale) to consolidate its functions. First degradation of readily degradable vegetable substrate in the compost material in the presence of rhamnolipid was investigated and the result showed that rhamnolipid did accelerate the degradation to some extent. Next rhamnolipid was introduced to the composting containing petroleum to see the effect of rhamnolipid on degradation of both the compost organic components and the hydrophobic petroleum. As a result, the biosurfactant not only enhanced the composting efficiency by shortening the total time for stabilization of the compost under middle temperature, but also dramatically strengthened degradation of the petroleum hydrocarbons. This result is in consistence with the conclusion that biosurfactant is able to enhance petroleum degradation in soils. Heavy metals in compost is an important factor that influence the quality of compost product, and the compost humic acid has crucial impact on migration of the heavy metals. We finally investigated the effect of rhamnolipid interference on adsorption of Pb2+ by humic acid from the compost material of late composting stage. The result showed that rhamnolipid possibly changed the style of Pb2+ adsorption onto the humic acid and it weakened the adsorption when the rhamnolipid concentration is high enough.In the whole study, production and purification of the biosurfactant rhamnolipid as well as investigation on some of its physico-chemical properties were performed. More significantly its potential functions to improve the microenvironment of organic granule substrate and its application in composting were investigated, and some achievements were obtained. It is a new field for biosurfactant application and may be meaningful to the concerned researchers.更多还原