Enzymatic Reactions in a Lab-on-Valve System: Cholesterol Evaluations.

Jucineide S Barbosa,M Das Graças A Korn,Marieta L.C Passos,M Lúcia M.F.S Saraiva

doi:10.3390/molecules24162890

Abstract

The micro sequential injection analysis / lab-on-valve (µSIA-LOV) system is a miniaturized SIA system resulting from the implementation of a lab-on-valve (LOV) atop of the selection valve. It integrates the detection cell and the sample processing channels into the same device, promoting the reduction of reagent consumption and waste generation, the improvement of the versatility, and the reduction of the time of analysis. All of these characteristics are really relevant to the implementation of enzymatic reactions. Additionally, the evaluation of cholesterol in serum samples is widely relevant in clinical diagnosis, since higher values of cholesterol in human blood are actually an important risk factor for cardiovascular problems. An automatic methodology was developed based on the µSIA-LOV system in order to evaluate its advantages in the implementation of enzymatic reactions performed by cholesterol esterase, cholesterol oxidase and peroxidase. Considering these reactions, the developed methodology was also used for the evaluation of cholesterol in human serum samples, showing reliable and accurate results. The developed methodology presented detection and quantification limits of 1.36 and 4.53 mg dL−1 and a linear range up to 40 mg dL−1. This work confirmed that this µSIA-LOV system is a simple, rapid, versatile, and robust analytical tool for the automatic implementation of enzymatic reactions performed by cholesterol esterase, cholesterol oxidase, and peroxidase. It is also a useful alternative methodology for the routine determinations of cholesterol in real samples, even when compared with other automatic methodologies.

Highlights

The popularity of biocatalysis has been increasing in the last decades, due to the high specificity and selectivity of enzymes
The objective to implement enzymatic reactions promoted by cholesterol esterase, cholesterol oxidase andisperoxidase, in asome μSIA-LOV
It was verified that the that the lowest concentration of Triton X-100 that allows the solubilization of cholesterol was 1.5%

Summary

Introduction

The popularity of biocatalysis has been increasing in the last decades, due to the high specificity and selectivity of enzymes. Their use can present some drawbacks, such as their dependence on some factors such as temperature, ionic strength, pH and cofactors. They can suffer inhibition by some compounds, and their lifetime is limited [1]. In order to overcome these limitations, enzyme has been used in flow-based methodologies. Flow systems allow efficient control of the reaction conditions, maximizing the enzyme activity [1,2], and increasing the reproducibility

Methods

Results

Conclusion