Research Article
Immobilization of GOX on PEG/fluoresence functionalized nanographene oxide to describe fluctuation of glucose level

https://doi.org/10.1016/j.jmst.2023.05.006Get rights and content

Highlights

  • Special platforms combining fluorescent dyes, biocompatible polymer and nanographene oxide were fabricated.

  • Nontoxic enzyme-nanocomposites in which GOx could possess and amplify its enzyme activities were firstly fabricated.

  • These enzyme-nanocomposites provide wider application potential in biosensing and tracing of glucose.

Abstract

Enzyme, produced and worked in all living things, could work as macromolecular biological catalysts in diverse biochemical processes with particular specificity, like glucose oxidase (GOX). The efficient use of enzyme properties has great importance in pharmaceutics and therapeutics. In this work, we could fabricate naive and effective electrochemical biosensors in the determination of glucose levels via utilizing GOX. Graphene oxide, as a water-soluble derivative of graphene, has shown great promise in a variety of biomedical applications including biosensors. Thus, we established a new-type special platform for GOX immobilization to perform its prosperities, in which nanographene oxide (nGO) was employed as an ideal base and poly(ethylene glycol) (PEG) was conjugated on the edge of nGO sheets to enhance its biocompatibility. Additionally, preferable functional dyes (Rhodamine B/fluorescein isothiocyanate) were also introduced to the platform. Enzyme-nanocomposites were then provided by locating GOX on the platform, i.e., GOX@nGO-PEG-RhB and GOX@nGO-PEG-FITC. The microstructure and composite of platforms and enzyme-nanocomposites were confirmed by diverse characterizations. Finally, on account of corresponding cyclic voltammetric and typical ready-state amperometric curves, it was informed that GOX@nGO-PEG-RhB and GOX@nGO-PEG-FITC could effectively respond to the fluctuation of glucose level as electrochemical biosensor. The present work presents special platforms for the immobilization of enzymes like GOX and provides new-type biosensors in the detection of glucose levels.

Introduction

Metabolic diseases, especially diabetes, have recently caused serious health troubles for human beings [1,2]. When people suffered from hyperglycemia, each of their bodily organs worked in a glucose-filled environment, and then their organs would endure irrecoverable damages like retinopathy, nephropathy and cardiovascular disease, etc. [3,4]. As an important bioactive substance, glucose serves an indispensable role in most metabolic processes in the living body [5]. Timely and effective monitoring of blood glucose levels became increasingly necessary to keep the health of mankind nowadays [6], [7], [8].

As a burgeoning approach, real-time and constant biosensors may afford an optimal tool for glucose level detection. Recent advances in nanomaterials have provided multiple platforms for biosensors, and various techniques could be employed in constructing biosensors for glucose detection in physiological statuses, such as electrochemistry, chemiluminescence, fluorescence, surface plasmon resonance, and photoelectrochemistry [9], [10], [11]. Given their excellent stability, fairly durability, good selectivity, fast response, and cheapness, electrochemical glucose biosensors were picked as the most optional glucose levels monitoring approach [12]. With the recent advances in nanotechnology, tremendous nanomaterials which performed good chemical stability and high catalytic activity have attracted much attention as potential substitutes, such as transition metals (such as Ni, Co, Cu, and their oxides), noble metal nanoparticles (such as gold (AuNPs) and silver (AgNPs) nanoparticles), and G-quadruplex hydrogels [13], [14], [15]. Regarding some intrinsic limitations like poor dispersion and impaired colloidal stability in the physiological medium, these biosensor alternates were still faced with great challenges [16]. Differently, enzymes such as glucose oxidase (GOX), working as macromolecular biological catalysts in multiple biochemical processes, were relatively applicable as biosensors in physiological states [17]. Biosensors based on GOX enzymes could be compatible with the human body and selective to a specific substrate, and GOX could be activated in glucose oxidation reaction (GOR) at neutral pH and room temperature [18,19].

So far, previous reports have provided glucose electrochemical sensors with GOX, but the improvement of GOX, such as its stability and activity, still needs to be solved [20]. In this aspect, a large number of nanomaterials have been employed to modulate the activity of GOX via fabricating platforms, such as polymers, nanofibers, quantum dots, nanoparticles, two-dimensional (2D) nanomaterials, etc. [21], [22], [23], [24]. Among them, 2D nanomaterials, especially nanographene oxide (nGO), have always been attractive candidates in biocatalysts and biosensors, with their large specific surface area and unique physicochemical properties [25], [26], [27]. Prominently, nGO could be fabricated as a special multifunctional platform to meet various requirements in the biosensor application of GOX, on account of its good conductivity, favorable compatibility, and multiple oxygen-containing moieties and activity sites [28,29]. Extendedly, through modification with functional molecules or macromolecules, nGO could perform superior in numerous application conditions like biomedicine, bioimaging, energy storage, and information transmission [30], [31], [32]. Previous literatures have informed that nGO could be decorated with much more functionality by conjugating with many polymers like poly(ethylene glycol) (PEG), poly(ethylene imine) (PEI), polystyrene (PS), polyamidamine (PAMAM), and poly(N-vinyl-2-pyrrolidone) (PVP), based on massive oxygen-containing moieties such as hydroxyl, carboxyl, and epoxy groups [33], [34], [35], [36]. And it is feasible to provide an attractive platform for GOX modulation by introducing functional molecules in the nGO base.

Herein, we first fabricated an appropriate platform for GOX locating, in which nGO was modified with functional polymers (PEG) and molecules (Rhodamine B/ fluorescein isothiocyanate) via amidation. GOXwas located on the foregoing platforms to exercise its enzyme activity in glucose level detection as electrochemical biosensors (Scheme 1). Systematic results illustrated that these biosensors displayed good electrochemical activity, stability, and selectivity towards glucose.

Section snippets

Morphology and structure characterization of functionalized nGO

As shown in Scheme 1, to make GOX stably perform its enzyme activity in the electrochemical detection of glucose level, we first fabricated special platforms by modifying nGO with featured polymers and molecules. Initially, biocompatible PEG (6-armed PEG-NH2) was covalently grafted onto the edge of freshly prepared nGO via amidation, giving well-dispersed and compatible nGO-PEG [37]. Fluorescent dye molecules (Rhodamine B (RhB) or fluorescein isothiocyanate (FITC)) were then introduced on

Conclusion

To summarize, in view of new-prepared nanocomposites (nGO-PEG-RhB and nGO-PEG-FITC), GOX was located on these special platforms to perform its enzyme properties in the detection of glucose. According to rigorous morphological and structural characterization, it was clarified that nontoxic enzyme-nanocomposites (GOX@nGO-PEG-RhB and GOX@nGO-PEG-FITC) were firstly fabricated, in which GOX could possess its enzyme activities and special platforms could amplify performances of GOX with unique

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

The authors thank the National Natural Science Foundation of China (Nos. 52272283 & 52103094), Science and Technology Commission of Shanghai Municipality (Nos. 20ZR1452200 & 22S31902900), Program for Outstanding Medical Academic Leader (No. 2019LJ27), and Shanghai Medical Key Specialty (No. ZK2019B12).

Data Availability Statement

The data that support the findings of this study are available from the corresponding author upon reasonable request.

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