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聚合物膜离子选择电极作为脂肪酶和酯酶测定的便捷工具。

Polymer membrane ion-selective electrodes as a convenient tool for lipases and esterases assays.

作者信息

Cieplak Maciej, Ostaszewski Ryszard

机构信息

a Enzymes and Microorganisms in Organic Synthesis and Biotechnology , Institute of Organic Chemistry PAS , Warsaw , Poland.

出版信息

Prep Biochem Biotechnol. 2017 Aug 9;47(7):673-677. doi: 10.1080/10826068.2017.1303611. Epub 2017 Mar 9.

Abstract

We have proposed a novel assay for lipases and esterases activity determination based on potentiometry with ion-selective electrodes (ISEs). Enzyme preparations, obtained from the living cells, are complex mixtures of various proteins, short peptides, lipids, carbohydrates, and other compounds. The most commonly used quantitative methods in enzyme studies are based on spectrophotometric or spectroflourimetric protocols which has significant limitations. They are not valid for samples that are turbid or strongly colored. To overcome those drawbacks we have proposed an assay based on potentiometry with ISEs for lipases and esterases activity determination. This electrochemical methodology represents an attractive tool for enzyme analysis, because of its low detection limit, independence from sample volume and from sample turbidity. The usefulness of this assay has been proven by the determination of the activity of various raw enzymes "acetone powders" isolated from animal tissues. Moreover, activities of fractions obtained during purification of one of those raw biocatalysts were also determined that way. The reliability of determination enzyme activity with ISE assay was proven by comparison with a classical spectrophotometric method.

摘要

我们提出了一种基于离子选择性电极(ISE)电位分析法的新型脂肪酶和酯酶活性测定方法。从活细胞中获得的酶制剂是各种蛋白质、短肽、脂质、碳水化合物和其他化合物的复杂混合物。酶研究中最常用的定量方法基于分光光度法或荧光分光光度法,这些方法存在显著局限性。它们不适用于浑浊或颜色较深的样品。为了克服这些缺点,我们提出了一种基于ISE电位分析法的脂肪酶和酯酶活性测定方法。这种电化学方法是一种有吸引力的酶分析工具,因为它具有低检测限,不受样品体积和样品浊度的影响。通过测定从动物组织中分离出的各种粗酶“丙酮粉”的活性,证明了该测定方法的实用性。此外,还通过这种方法测定了其中一种粗生物催化剂纯化过程中获得的各组分的活性。通过与经典分光光度法比较,证明了ISE测定法测定酶活性的可靠性。

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