Improving the stability of an unstable lipase by applying different immobilization strategies for the selective hydrolysis of fish oil

IF 1.9 4区 农林科学 Q3 CHEMISTRY, APPLIED Journal of the American Oil Chemists Society Pub Date : 2024-03-12 DOI:10.1002/aocs.12833
Faezeh Ahrari, Mitra Pourmohammadi Lish, Maryam Yousefi, Mehdi Mohammadi
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Abstract

Rhizopus oryzae lipase (ROL) is known to present high selectivity in chemical reactions. However, the poor stability of ROL effectively limits its industrial applications. In this study, several immobilization protocols, such as hydrophobic adsorption, covalent immobilization, multi-point covalent attachment, ionic adsorption/cross-linking, and ionic interaction, were applied to improve the stability of ROL. Heterogeneous modification of aspartic and glutamic acid residues on the surface of ROL was carried out by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) to introduce new amine groups with lower pKb. The highest immobilization yield of 89% was achieved for octyl-agarose, producing specific activity of 45 U/mg, which is 15 folds higher than the specific activity of the soluble enzyme. Improved stability of ROL was observed, in particular for those derivatives obtained by multi-point covalent attachment of ROL on glyoxyl-agarose (Gx-ROL) and aminated ROL on glyoxyl-agarose (Gx-NH2-ROL) by retaining 28%–36% of their initial activities after 24 h incubation at 60°C. Immobilization also altered the co-solvent stability profile of the immobilized derivatives producing biocatalysts with varied co-solvent stabilities. Furthermore, utilization of the immobilized preparations in fish oil hydrolysis revealed the selective release of cis-5,8,11,14,17-eicosapentaenoic acid (EPA) and cis-4,7,10,13,16,19-docosahexaenoic acid (DHA) in favor of EPA. The highest EPA/DHA selectivity of 33 was observed for the hydrophobically immobilized ROL on octyl-sepharose.

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在选择性水解鱼油过程中采用不同的固定化策略提高不稳定脂肪酶的稳定性
众所周知,根瘤菌脂肪酶(ROL)在化学反应中具有高选择性。然而,ROL 的稳定性较差,这有效地限制了其在工业上的应用。本研究采用疏水吸附、共价固定、多点共价连接、离子吸附/交联和离子相互作用等几种固定化方案来提高 ROL 的稳定性。利用 1-乙基-3-(3-二甲基氨基丙基)碳二亚胺(EDC)对 ROL 表面的天冬氨酸和谷氨酸残基进行异构修饰,引入 pKb 值较低的新胺基。辛基琼脂糖的固定化率最高,达到 89%,产生的比活度为 45 U/mg,比可溶性酶的比活度高 15 倍。通过观察发现,ROL 的稳定性有所提高,尤其是那些通过多点共价连接将 ROL 固定在乙醛基-琼脂糖上(Gx-ROL)和将 ROL 氨基化后固定在乙醛基-琼脂糖上(Gx-NH2-ROL)而得到的衍生物,它们在 60°C 孵育 24 小时后仍能保持 28%-36% 的初始活性。固定化还改变了固定化衍生物的共溶剂稳定性曲线,从而产生了具有不同共溶剂稳定性的生物催化剂。此外,利用固定化制剂水解鱼油发现,顺式-5,8,11,14,17-二十碳五烯酸(EPA)和顺式-4,7,10,13,16,19-二十二碳六烯酸(DHA)的选择性释放有利于 EPA。在辛基-sepharose 上亲水固定的 ROL 的 EPA/DHA 选择性最高,达到 33。
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来源期刊
CiteScore
4.10
自引率
5.00%
发文量
95
审稿时长
2.4 months
期刊介绍: The Journal of the American Oil Chemists’ Society (JAOCS) is an international peer-reviewed journal that publishes significant original scientific research and technological advances on fats, oils, oilseed proteins, and related materials through original research articles, invited reviews, short communications, and letters to the editor. We seek to publish reports that will significantly advance scientific understanding through hypothesis driven research, innovations, and important new information pertaining to analysis, properties, processing, products, and applications of these food and industrial resources. Breakthroughs in food science and technology, biotechnology (including genomics, biomechanisms, biocatalysis and bioprocessing), and industrial products and applications are particularly appropriate. JAOCS also considers reports on the lipid composition of new, unique, and traditional sources of lipids that definitively address a research hypothesis and advances scientific understanding. However, the genus and species of the source must be verified by appropriate means of classification. In addition, the GPS location of the harvested materials and seed or vegetative samples should be deposited in an accredited germplasm repository. Compositional data suitable for Original Research Articles must embody replicated estimate of tissue constituents, such as oil, protein, carbohydrate, fatty acid, phospholipid, tocopherol, sterol, and carotenoid compositions. Other components unique to the specific plant or animal source may be reported. Furthermore, lipid composition papers should incorporate elements of year­to­year, environmental, and/ or cultivar variations through use of appropriate statistical analyses.
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Issue Information Issue Information JAOCS special issue on advancement in plant protein-based emulsions Issue Information Issue Information
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