摘要
人类发展到21世纪,生活质量逐渐提高,石油化工产品越来越多的应用于人类的日常生活中,塑料作为一种广泛的金属的代替品而深受人们的喜爱,其应用广泛,经济耐用,成本低,使用方便等特点使其生产规模逐年扩大,然而低成本的塑料制品又很少有回收的价值,因此而导致了令人头疼的白色污染问题,为了解决白色污染,各国都在极力的寻找一种可以代替传统塑料的材料,聚乳酸(PLA)作为一种可降解塑料的原材料,由于其特有的性能而受到越来越多的关注,聚乳酸是利用乳酸为原料合成的聚酯材料,具有良好的生物降解性,现已应用到医疗卫生,纺织,包装,农业等领域。
本论文重点在于证明聚乳酸生物降解可行性。筛选出一株具有降解聚乳酸能力并可分泌聚乳酸解聚酶的细菌,探究该菌株分泌聚乳酸水解酶的最佳条件,分离、纯化出聚乳酸解聚酶,并对其酶学性质进行初步研究。实验结果如下:
1从抚顺石油厂的活性污泥中筛选纯化出菌株DS0701,经过生理生化性能比对,以及16SrDNA鉴定,该菌系属于纤维单胞菌属(Cellulomonas.sp.),且该菌具有降解PLA能力。
2对菌株DS0701降解PLA的条件进行检测,最终确定,最适培养温度为37℃,最适培养pH为7.0。培养时间为72h,最佳摇床转数为150r/ min。
3对DS0701菌株降解能力测定,检测结果表明,菌株DS0701不仅能够降解PLA,而且对其他的几种可降解材料也有降解能力,如聚羟基丁酸酯(PHB)、3-羟基丁酸与3-羟基戊酸的共聚物(PHBV),聚(3-羟基丁酸酯-co-4-羟基丁酸酯)(P(3-4HB))等。
4进行酶的分离与纯化,在对培养条件进行测定之后,对该菌株进行发酵产酶,通过离心,超滤,冻干,盐析,层析等方式对PLA解聚酶进行分离与纯化,进而研究其解聚酶的酶学性质。
5.进行聚乳酸薄膜降解,降解时间为9天,降解后膜透明度明显减弱,变为白色,强度大大降低,膜的边角明显变圆,质地变酥脆。从扫描电镜照片上可以观察到膜表面部分地方开始变化,聚乳酸膜逐步裂解,膜表面因被降解而开始变得粗糙至膜出现细微裂痕和空洞。
Polylactide or poly(lactic acid) (PLA) was formerly known as a hydrolyzable and unstable polyester with limited use as a biodegradable material in medical applications. Polymer processing technology has advanced quite remarkably, however, so PLA film and fiber with great stability and good physical properties are now available. Hence, the applications of PLA have expanded to include its use as a biodegradable plastic for solving the worldwide plastic waste problem.
The emphases of the essay are proving the feasibility of degrading polylactic acid by microorganisms, procuring a bacterium which can degrade polylactic acid in a relative short time. Such bacterium had be obtained, and its growing optimum condition had be obtained, too. Polylactic acid depolymerase has been purified and some primary analysis has been done on it.
The main results obtained from the work are as follows:
1. The bacterium numbered DS0701, which has the ability of degradating PLA, was purified from the mud collected from different places. The bacterium is identified as [Cellulomonas (Bergey et al., 1923), Clark, 1952]. Its growing optimum condition had be obtained. The bacterium growing in 37℃and pH=7.0 can excrete most polylactic acid depolymerase.
2. Some research has been done on the polylactic acid depolymerase which is obtained from DS0701, The most suitable reaction temperature of this polylactic acid depolymerase is 40℃and pH is7.0.
3. The bacteria DS0701 also have the ability to degrade PLA films. After nine days’degradation, PLA films partly lost their intensity and transparency. The photos by electronic scanning microscope show that the surface of the PLA films has been degraded partly.
4.Isolation and Purification of the enzyme, after measured of the culture conditions, the enzyme production of the strain by centrifugation, ultrafiltration, Freeze-dried, Salt precipitation, chromatography, etc. the PLA depolymerase were isolated and purified and then study the enzymatic properties of depolymerization of PLA.
5. The degradation of PLA film, degradation time is 9 days, significantly decreased of transparency after degradation, to white, strength greatly reduced, and the corner of the film was turned round the corner, change crisp of the texture.
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