表面活性剂改性磁性氧化石墨烯共价固定脂肪酶CRL

doi:10.16085/j.issn.1000-6613.2019-0255

摘要/Abstract

摘要：

采用化学共沉淀法制备磁性氧化石墨烯（MGO），直接在反应体系中加入Span系列表面活性剂，一锅法制备得到表面活性剂改性磁性氧化石墨烯（SMGO）。X射线衍射仪（XRD）、扫描电镜（SEM）和傅里叶变换红外光谱（FTIR）表征结果表明SMGO制备成功，且具有良好的磁分离性能。以1,5-戊二醛（GA）为交联剂，褶皱假丝酵母脂肪酶（CRL）为模型酶，共价固定CRL于SMGO载体上。Span40 MGO固定化酶酶活回收率为116.5%±1.7%，为MGO固定化酶的6倍；比活可达32.5U/mg，为游离酶的1.6倍；k_cat/K_m也有较大的提升，高于游离酶50%。储存稳定性及热稳定性得到提高，用于水解反应6批次后仍然保留73.6%的相对酶活力。初步分析认为MGO经改性后表面从亲水性转为强疏水性，使得共价固定化过程中同时发生疏水性界面活化，这是酶活性提高的原因之一。文章所报道的改性策略可为类似载体改性提供新思路。

关键词: 磁性氧化石墨烯, 表面活性剂, 固定化, 酶, 界面活化

Abstract:

In this paper, MGO was prepared by chemical co-precipitation method, span series surfactant was directly added to the reaction system to prepare surfactant-modified magnetic graphene oxide （SMGO) through one pot process. The results of X-ray diffraction(XRD), scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy(FTIR) indicated that SMGO was successfully prepared and performed well in magnetic separation. With GA as cross-link agent and Candida rugose lipase (CRL) as model enzyme, CRL was immobilized covalently on the SMGO materials. The activity recovery of Span40 MGO immobilized enzyme was 116.5%±1.7%, which was 6 times that of MGO immobilized enzyme; the specific activity was up to 32.5U/mg, 1.6 times that of free enzyme; k_cat/K_m was about 0.5 times higher than that of free enzyme. The storage stability and thermal stability were improved, and 73.6% of the relative enzyme activity was retained after 6 cycles of hydrolysis reaction. Preliminary analysis indicated that the surface of MGO was changed from hydrophilic to strong hydrophobic after modification, which simultaneously caused the activation of hydrophobic interface during the process of covalent immobilization. It was one of the reasons that increased the enzyme activity. The modification strategy reported in the article can provide new ideas for the modification of similar carrier.

Key words: magnetic graphene oxide, surfactants, immobilization, enzyme, interface activation

中图分类号:

王艳娥,张子安,唐爱星,李青云,刘幽燕. 表面活性剂改性磁性氧化石墨烯共价固定脂肪酶CRL[J]. 化工进展, 2019, 38(9): 4255-4263.

Yan’e WANG,Zi’an ZHANG,Aixing TANG,Qingyun LI,Youyan LIU. Surfactant modified magnetic graphene oxide covalent immobilized Candida rugose lipase[J]. Chemical Industry and Engineering Progress, 2019, 38(9): 4255-4263.

图/表 9

参考文献 32

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载体	蛋白回收率 /%	酶活回收率 /%	比活 /U·mg^-1	表面活性剂 HLB值	接触角 /（°）
MGO	97.5±5.6	28.0±1.6	5.7	—	23.5±0.2
Span20 MGO	77.1±3.5	110.10±2.0	28.5	8.6	128.1±1.4
Span40 MGO	71.6±1.2	116.5±1.7	32.5	6.7	139.3±2.9
Span60 MGO	71.2±2.5	113.4±0.4	31.8	4.7	136.0±0.5
CRL	—	—	20.0	—	—

载体	蛋白回收率 /%	酶活回收率 /%	比活 /U·mg^-1	表面活性剂 HLB值	接触角 /（°）
MGO	97.5±5.6	28.0±1.6	5.7	—	23.5±0.2
Span20 MGO	77.1±3.5	110.10±2.0	28.5	8.6	128.1±1.4
Span40 MGO	71.6±1.2	116.5±1.7	32.5	6.7	139.3±2.9
Span60 MGO	71.2±2.5	113.4±0.4	31.8	4.7	136.0±0.5
CRL	—	—	20.0	—	—

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