利用酶法—化学氧化法偶联工艺对活性艳蓝KN-R的脱色研究

发布时间：2019-04-01 06:48

【摘要】：本文以染料活性艳蓝(Remazol Brilliant Blue KN-R, RBBR)模拟染料废水,利用过水解酶原位催化制备过氧乙酸的方法对其进行脱色处理,并对脱色条件进行了优化。同时,采用交联酶聚体技术对过水解酶进行固定化,并用聚乙烯醇进行二次包埋。最后利用固定化酶进行染料RBBR脱色。主要的实验结果如下：1、过水解酶的纯化及酶学性质分析。利用亲和层析柱纯化过水解酶,SDS-PAGE电泳显示为单一条带,其分子量大小为28 kDa,比活力为9.03 U/mg。酶学性质分析表明,该过水解酶最适pH为6.0,最适温度为65℃,在pH 3.0-6.0及温度为30-65℃范围内具有较好的稳定性；其T502h为83.4℃。2、过水解酶催化的脱色反应。利用过水解酶原位催化制备过氧乙酸对染料RBBR进行脱色。在单因子试验的基础上,通过正交优化试验获得最优脱色条件：在5 mL反应体系中,最适pH5.0、酶量20 U/Reaction、乙酸乙酯与过氧化氢的摩尔比40：1、染料浓度80 mg/mL。在最优条件下处理6h后,染料RBBR的脱色率为81.11%,24 h的脱色率为91.96%。将体系放大50倍后(250 mL),24 h染料RBBR的脱色率为84.55%。3、过水解酶的固定化及应用效果评估。首先采用交联酶聚集体技术(CLEAs)对过水解酶进行固定化,然后采用聚乙烯醇对交联过水解酶聚集体进行二次包埋,进行脱色反应。结果表明,当使用60%(v/v)的有机溶剂S2为沉淀剂,且酶与BSA的比率为16：2,交联剂戊二醛的浓度为100mmol/L时,制备得到的过水解酶交联酶聚体(Per-CLEAs)酶活回收率可达79.72%。利用不同浓度的聚乙烯醇(PVA)进行二次包埋固定化,结果发现,当PVA浓度为10%(w/v)时,包埋后的Per-CLEAs形态较好、酶活回收率较高,而且洗涤三次后无明显的酶活泄露。用其对染料RBBR脱色,在维持80%的脱色率的前提下,至少可以进行6次循环。分别利用游离酶、Per-CLEAs以及包埋后的Per-CLEAs催化合成过氧乙酸原位氧化RBBR脱色,处理30 min后,脱色率分别为63.72%、95.98%、83.51%。
[Abstract]:In this paper, dye reactive brilliant blue (Remazol Brilliant Blue KN-R, RBBR (reactive brilliant blue dye) was used to simulate dye wastewater, and the decolorization conditions were optimized by in-situ catalytic preparation of peracetic acid by hyperhydrolysis enzyme. At the same time, cross-linked enzyme polymer technology was used to immobilize the hydrolase, and polyvinyl alcohol was used to encapsulate the hydrolase for two times. Finally, the dye RBBR was decolorized by immobilized enzyme. The main results are as follows: 1. Purification and characterization of hydrolase. The hydrolase was purified by affinity chromatography. SDS-PAGE electrophoresis showed that the hydrolase was a single band with a molecular weight of 28 kDa, and a specific activity of 9.03 U ~ (?) mg 路g ~ (- 1). The results showed that the optimal pH and temperature of the hydrolase were 6.0 鈩，

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