A novel on-flow mass spectrometry-based dual enzyme assay.

Autor: Seidl C; Departamento de Química, Grupo de Cromatografia de Bioafinidade e Produtos Naturais, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, 14040-90, Brazil., Vilela AFL; Departamento de Química, Grupo de Cromatografia de Bioafinidade e Produtos Naturais, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, 14040-90, Brazil., Lima JM; Departamento de Química, Grupo de Cromatografia de Bioafinidade e Produtos Naturais, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, 14040-90, Brazil., Leme GM; SEPARARE Núcleo de Pesquisa Em Cromatografia, Departamento de Química, Universidade Federal de São Carlos, Caixa Postal 676, São Carlos, 13565-905, Brazil., Cardoso CL; Departamento de Química, Grupo de Cromatografia de Bioafinidade e Produtos Naturais, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, 14040-90, Brazil. Electronic address: ccardoso@ffclrp.usp.br.
Jazyk: angličtina
Zdroj: Analytica chimica acta [Anal Chim Acta] 2019 Sep 23; Vol. 1072, pp. 81-86. Date of Electronic Publication: 2019 Apr 26.
DOI: 10.1016/j.aca.2019.04.057
Abstrakt: This work describes a new simultaneous on-flow dual parallel enzyme assay based on immobilized enzyme reactors (ICERs) with mass spectrometry detection. The novelty of this work relies on the fact that two different enzymes can be screened at the same time with only one single sample injection and in less than 6 min. The system consisted of two immobilized capillary enzyme reactors (ICERs). More specifically, the ICERs comprised two different enzymes that were accommodated in parallel and were placed between a liquid chromatography (LC) system and a mass spectrometer (MS). The resulting system could be adapted to other types of enzyme reactors with different supports. All the elements in the system were interfaced by means of two 10-port/two-position switching valves. Different tubing dimensions allowed us to monitor the activity of each enzyme independently during the same analysis. Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) bioreactors were chosen as proof of concept. Acetylcholine (ACh) was used as substrate; the area of its protonated enzymatic hydrolysis product ion, choline, [M+H] + m/z 104.0, was monitored in the presence and absence of the standard cholinesterase inhibitor galantamine. This method proved to be an interesting tool for fast, simultaneous, and independent label-free dual enzyme inhibitor assay.
(Copyright © 2019 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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