**1. Introduction**

Numerous novel and cutting-edge technologies and materials are necessary to satisfy the new trends and requisites of analytical systems as the needs for environmental, biomedical, food and beverage analysis are progressing very quickly. The development of biosensors has evolved as one of the most promising research directions to overcome these challenges. Therefore, biosensor-based techniques have recently started being applied for the determination of different clinically, environmentally and biologically active materials [1–3]. In this regard, the design of biosensors in nanoscience/nanotechnology, environmental, medicine and food monitoring has been significantly increased during the past decade for their extensive applications. These advanced technologies have assisted the construction of highly sensitive, selective, customizable, and portable sensors for the determination of various clinically significant materials such as glucose, etc. [4]. The progress of such glucose biosensors has an inordinate significance in diagnosing and controlling diabetes mellitus, which is considered a worldwide public health problem. Diabetes mellitus would increase the risk of heart disease, kidney failure, blindness, postoperative and wound infections [5,6].

Diabetes mellitus has increased worldwide over the past five decades. Diabetes is a medical condition in which patients experience glucose concentration diverging from the normal range of 80–120 mg/dL (4.4–6.6 mM) [7]. In 2019, the International Diabetes Federation (IDF) assessments indicated that approximately 463 million adults have diabetes, and it might rise to 700 million by 2045 [8]. Diabetic patients are required to perform glucose testing several times a day to maintain normal glucose levels. Hence, the rapid quantification of glucose concentration in bodily fluids is vital for diagnosing and treating

**Citation:** Settu, K.; Chiu, P.-T.; Huang, Y.-M. Laser-Induced Graphene-Based Enzymatic Biosensor for Glucose Detection. *Polymers* **2021**, *13*, 2795. https://doi.org/10.3390/ polym13162795

Academic Editors: José Miguel Ferri and Claudio Gerbaldi

Received: 5 July 2021 Accepted: 17 August 2021 Published: 20 August 2021

**Publisher's Note:** MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

**Copyright:** © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

diabetic patients. For this purpose, the design of an easy, rapid and low-cost technology for the determination of glucose is essential in clinical diagnosis [9,10].

Glucose biosensors have significantly contributed to the detection of glucose levels in diabetic patients [11,12]. Studies have indicated that among various biosensors, glucose oxidase (GOx) enzyme-based electrochemical biosensors were considered to offer good selectivity and sensitivity for glucose detection [13,14]. Amperometry is the widely used electrochemical technique for glucose detection. Amperometric sensors could provide several advantages, such as ease of use, short analysis time, high sensitivity, and higher signal-to-noise ratio compared to other sensors [14–16]. The common idea applied for the development of amperometric biosensors is the efficiency of charge transfer, which can be better enhanced. Additionally, the biocompatibility issues of the sensors could be resolved by modifying electrodes with polymers such as chitosan or hydrogels [17]. In addition, various features of the electrodes could easily be altered by selecting the optimal chemical and electrochemical parameters during the effective electrode modifications [18,19].

The amperometric glucose biosensor generally uses an enzyme glucose oxidase (GOx), which catalyzes glucose oxidation at the electrode and provides high selectivity in glucose detection. Most enzymatic amperometric biosensors are based on disposable screen-printed enzyme electrode strips [20–22]. However, the wastage of materials might occur during the screen-printing process, limiting the applications of screen-printed electrodes.

Graphene, a carbon-based nanomaterial, has gained substantial attention in many areas. In terms of electrochemical properties, graphene could provide high conductivity with a remarkable heterogeneous electron transfer rate [23,24]. In 2014, it was found that polymers such as polyimide (PI) could be directly converted into porous graphene using a CO<sup>2</sup> laser machine with a 10.6 µm wavelength [25]. In addition to infrared CO<sup>2</sup> (10.6 µm) laser, visible laser [26–31] and ultraviolet laser [32] have also been successfully used to synthesize laser-induced graphene (LIG). The laser-irradiation of the PI film caused the photo-thermal generation of the graphene due to the local heating of the film. Upon heating the film, the carbon atoms bonded with oxygen (C–O, C=O) and nitrogen (C–N) atoms via sp<sup>3</sup> and sp<sup>2</sup> hybridization breakdown and rearranged to form several layers of sp<sup>2</sup> hybridized carbon atoms of graphene [25,33]. The laser induction of graphene has been performed in ambient conditions without any material wastage. In addition, the shape/pattern of LIG could also be easily customized by computer design, which holds great promise toward developing glucose biosensors.

Recently, Pereira et al. demonstrated the electrochemical response of GOx adsorbed on a CO<sup>2</sup> laser-scribed LIG [34]. The GOx enzyme adsorbed on LIG remained catalytically active even after running the cyclic voltammetry up to +1.0 V for glucose detection. The LIG electrodes facilitated the direct electron transfer between the GOx and the electrode surface without mediators.

In this study, we fabricated a laser-induced graphene electrode (LIGE) by simple direct laser engraving with the UV laser on polyimide tape. The LIGE surface was immobilized with GOx/Chitosan composite for selective detection on glucose. Amperometric measurement was used to quantify the glucose concentration with the developed LIGE enzymatic biosensor. The novelty of the present work lies in the detection of glucose with enhanced sensitivity using a simple, low-cost LIGE-based biosensor.
