Switchable Graphene-Based Bioelectronics Interfaces

chemosensors

Review Switchable Graphene-Based Bioelectronics Interfaces

Meenakshi Choudhary 1, Sudheesh K. Shukla 2,3,* , Jagriti Narang 4, Vinod Kumar 5, Penny P. Govender 2,* , Avi Niv 1, Chaudhery Mustansar Hussain 6 , Rui Wang 3, Bindu Mangla 7 and Rajendran Suresh Babu 8 1 The Swiss Institute for Dryland Environmental and Energy Research, The Jacob Blaustein Institutes for Desert Research, Ben-Gurion University of the Negev, Midreshet Ben-Gurion 8499000, Israel; [email protected] (M.C.); [email protected] (A.N.) 2 Department of Chemical Sciences-DFC (Formely Department of Applied Chemistry), University of Johannesburg, P.O. Box 17011, Doornfontein, Johannesburg 2028, South Africa 3 School of Environmental Science and Engineering, Shandong University, Jimo, Qingdao 266237, China; [email protected] 4 Department of Biotechnology, Jamia Hamdard University, New Delhi 110062, India; [email protected] 5 Department of Materials Engineering, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel; [email protected] 6 Department of Chemistry and Environmental Science, New Jersey Institute of Technology, Newark, NJ 07102, USA; [email protected] 7 Department of Chemistry, J.C. Bose University of Science and Technology YMCA, Faridabad 121006, India; [email protected] 8 Laboratory of Experimental and Applied Physics, Centro Federal de Educação Tecnológica Celso Suckow da Fonseca, Rio de Janeiro 20271–110, Brazil; [email protected] * Correspondence: [email protected] (S.K.S.); [email protected] (P.P.G.)

Received: 28 April 2020; Accepted: 17 June 2020; Published: 26 June 2020

Abstract: Integration of materials acts as a bridge between the electronic and biological worlds, which has revolutionized the development of bioelectronic devices. This review highlights the rapidly emerging field of switchable interface and its bioelectronics applications. This review article highlights the role and importance of two-dimensional (2D) materials, especially graphene, in the field of bioelectronics. Because of the excellent electrical, optical, and mechanical properties graphene have promising application in the field of bioelectronics. The easy integration, biocompatibility, mechanical flexibility, and conformity add impact in its use for the fabrication of bioelectronic devices. In addition, the switchable behavior of this material adds an impact on the study of natural biochemical processes. In general, the behavior of the interfacial materials can be tuned with modest changes in the bioelectronics interface systems. It is also believed that switchable behavior of materials responds to a major change at the nanoscale level by regulating the behavior of the stimuli-responsive interface architecture.

Keywords: bioelectronics interface; electrochemical biosensing; bioreactor; 2-dimensional; graphene; stimuli-responsive

1. Introduction Bioelectronics is an emerging and exciting ﬁeld, which involves biological materials and biological architecture for information processing. According to the National Institute of Standard and Technology (NIST), bioelectronics involves the convergence of biology and electronics. Bioelectronics seeks to exploit biology in conjunction with electronics in a wider context encompassing, for example, information storage, bionics and biomaterials for information processing, biofuel cells, and electronic components

Chemosensors 2020, 8, 45; doi:10.3390/chemosensors8020045 www.mdpi.com/journal/chemosensors Chemosensors 2020, 8, 45 2 of 12 Chemosensors 2020, 8, x FOR PEER REVIEW 2 of 12 and actuators.encompassing, The mainfor example, aspect ofinformation bioelectronics storag ise, the bionics interface and between biomaterials biological for information materials and electronicsprocessing, [1–3]. biofuel Thus, thecells, key and concept electronic of thecomponents bioelectronics and actuators. is the transduction The main aspect of the of biologicalbioelectronics signals to electricalis the signalsinterface at between the sensing biological interface. materials and electronics [1–3]. Thus, the key concept of the bioelectronics is the transduction of the biological signals to electrical signals at the sensing interface. Carbon-based materials have the capability to form a bridge between the biological and the Carbon-based materials have the capability to form a bridge between the biological and the electronicelectronic world world and and to fundamentallyto fundamentally change change thethe fabricationfabrication of of bioelectronics bioelectronics devices devices such such as as bio-actuators,bio-actuators, biofuel biofuel cells, cells, and and biosensors biosensors providingproviding new new window window of opportunities of opportunities to the to future the futureof of bioelectronics.bioelectronics. Carbon-basedCarbon-based materials have have the the potential potential to tocrack crack past past technical technical problems problems by by communicatingcommunicating the biologicalthe biological entities entities to to the the electronic electronic system system at its its interface interface [4,5] [4,5 (Figure] (Figure 1).1 ).

FigureFigure 1. Schematic 1. Schematic depicting depicting the the mergingmerging of biotechnology, biotechnology, electronics electronics and and materials materials science science [6]. [6].

For successfulFor successful interfacing interfacing between between electronic electronic andand biological system system there there must must be bea precise a precise interface.interface. Interface Interface works works as as a shuttlea shuttle between between biologicalbiological and electrical electrical entities entities and and enhances enhances the the electronelectron transfer transfer rate. rate. To overcomeTo overcome these these challenges, challenges, carbon-basedcarbon-based materials materials emerges emerges as a as best-fitted a best-ﬁtted modelmodel for the for fabrication the fabrication of bioelectronic of bioelectronic interfaces; interfaces; they they playplay a a crucial crucial role role in in exploring exploring the thebasics basics of of their physicaltheir physical properties, properties, chemical chemical interaction, interaction, thermodynamics, thermodynamics, and and kinetics kinetics due due to its to itsremarkable remarkable surface area, morphological behavior, and electron transfer rate. surface area, morphological behavior, and electron transfer rate. Whether at a commercial or laboratory level of development of bioelectronic devices, there is a Whether at a commercial or laboratory level of development of bioelectronic devices, there is a need to comprehend the dynamic forces and respective component at molecular levels [7,8]. During need tothe comprehend past decade,the there dynamic have been forces remarkable and respective achievements component in material at molecular science levelsof bioelectronic [7,8]. During the pastdevices decade, but therethere haveis still beena need remarkable to explore achievementsthe challenges of in real-world material science scenario of [9,10]. bioelectronic In the recent devices but thereera, researchers is still a need are focusing to explore more the of challenges their attention of real-worldon the technical scenario issues [9 relating,10]. In the the practical recent era, researchersapplication are focusing of bioelectronic more of interfaces. their attention on the technical issues relating the practical application of bioelectronicIn this interfaces. review article, we look at graphene-based interface materials with their respective Inapplications this review in the article, fields weof biosensors look at graphene-basedand bioelectronics. interface In addition, materials we also highlight with their the respectivefuture applications in the ﬁelds of biosensors and bioelectronics. In addition, we also highlight the future prospects of graphene-based bioelectronic interfaces by focusing on recent reports on electrochemical biosensing performance. Chemosensors 2020, 8, x FOR PEER REVIEW 3 of 12

Chemosensorsprospects 2020of , 8graphene-based, 45 bioelectronic interfaces by focusing on recent reports3 ofon 12 electrochemical biosensing performance. 2. Fundamentals of 2D Materials and Graphene-Based Bioelectronic Interfaces 2. Fundamentals of 2D Materials and Graphene-Based Bioelectronic Interfaces Before highlighting on the behavior of switchable bioelectronic interfaces, first we need to Before highlighting on the behavior of switchable bioelectronic interfaces, first we need to deliberate on the unique properties of graphene that makes it such an important biosensing interface deliberate on the unique properties of graphene that makes it such an important biosensing interface material. The striking discovery of graphene opens a platform to initiate exceptional and extraordinary material. The striking discovery of graphene opens a platform to initiate exceptional and research in the field of 2D materials. The dimensionality of 2D materials plays a crucial role in tuning extraordinary research in the field of 2D materials. The dimensionality of 2D materials plays a their mechanical, electrical, optical properties at the molecular level, depending on the stereometric crucial role in tuning their mechanical, electrical, optical properties at the molecular level, arrangement of atoms in the dimension of zero (0D), one (1D), two (2D), or three (3D) crystal structure. depending on the stereometric arrangement of atoms in the dimension of zero (0D), one (1D), two 2D materials have atomically thin geometry along with intrinsic flexibility and they can easily be (2D), or three (3D) crystal structure. 2D materials have atomically thin geometry along with intrinsic integrated with other materials because of H-terminated ends which terminate the dangling bonds at flexibility and they can easily be integrated with other materials because of H-terminated ends their respective surfaces. The 2D crystals have an unusual physical phenomenon to tender a wide which terminate the dangling bonds at their respective surfaces. The 2D crystals have an unusual range of potential applications. In addition, 2D materials are super thin nanomaterials with a high physical phenomenon to tender a wide range of potential applications. In addition, 2D materials are degree of anisotropy behavior and chemical functionality. super thin nanomaterials with a high degree of anisotropy behavior and chemical functionality. The embryonic level discovery of 2D materials recommended different insights to develop The embryonic level discovery of 2D materials recommended different insights to develop flexible and thin materials-based bioelectronic interfaces for bioelectronic device fabrication. 2D-based flexible and thin materials-based bioelectronic interfaces for bioelectronic device fabrication. bioelectronic interface opens a new way to research for the field of material science including 2D-based bioelectronic interface opens a new way to research for the field of material science semiconductor and nanotechnology for different applications. These 2D atomic sheets with unique including semiconductor and nanotechnology for different applications. These 2D atomic sheets properties such as distinctive thickness, scalability, and so on, enable them to be used in the development with unique properties such as distinctive thickness, scalability, and so on, enable them to be used in of semiconductors, insulators, transparent conductors, and transducers [11,12]. The different types of the development of semiconductors, insulators, transparent conductors, and transducers [11,12]. The 2D materials are categorized based on their geometric distribution [13]. different types of 2D materials are categorized based on their geometric distribution [13]. In the present era, low cost flexible 2D materials opens a door for cheap and stable bioelectronic In the present era, low cost flexible 2D materials opens a door for cheap and stable bioelectronic devices such as radio-frequency sensors, integrated nano-systems with performance as compared devices such as radio-frequency sensors, integrated nano-systems with performance as compared with the conventional silicon devices. 2D layered materials also have remarkable potential for the with the conventional silicon devices. 2D layered materials also have remarkable potential for the development of electrochemical and optical biosensors,biosensors, nanomedicine drug delivery systems, cancer treatment, and tissue engineering [[14,15]14,15] (Figure(Figure2 2).).

Figure 2. Possible applications of 2D-based nanomaterials [[16].16].

Graphene is one of thethe leadingleading andand primaryprimary 2D2D materials,materials, andand categorizedcategorized asas havinghaving spsp22 hybridization, and is single-layered with extensiveextensive surface areaarea structure.structure. The honeycomb shaped

Chemosensors 2020, 8, 45 4 of 12 structureChemosensors of the 2020 graphene, 8, x FOR PEER provides REVIEW an easy fabrication of bioelectronic devices. The interesting4 of 12 properties such as high surface area, excellent electrical conductivity, and mechanical strength make structure of the graphene provides an easy fabrication of bioelectronic devices. The interesting grapheneproperties a promising such as high candidate surface forarea, di excellentfferent typeelectrical of applications conductivity, such and mechanical as hydrogen strength storage, make sensor, solargraphene cells, and a biosensorpromising [candidate17,18]. Dating for different back to type 1940, of applications the early discussion such as hydrogen and theoretical storage, explanationsensor, alreadysolar proved cells, theand existence biosensor of [17,18]. graphene. Dating However, back to the 1940, interest thein early graphene discussion was sparkedand theoretical in 2010 with the Nobelexplanation prize awardalready in proved physics the to existence Prof. A. Geimof graphene. and Prof. However, K. Novoselov the interest for their in graphene ground-breaking was researchsparked on the in 2010 “graphene: with the 2DNobel materials” prize award [19 ].in The physics research to Prof. reported A. Geim by and two Prof. Nobel K. Novoselov Prize winners for in 2010,their cracked ground-breaking the mystery research wall of grapheneon the “graphene: by the simple2D materials” scotch [19]. tape The experiment research reported in 2004 by [20 two,21]. Recently,Nobel Prize a numberwinners ofin synthetic2010, cracked approaches the mystery have wall been of developed graphene by for the the simple synthesis scotch of graphene.tape experiment in 2004 [20,21]. All the developed methodologies can be divided into two main categories: top-down and bottom-up, Recently, a number of synthetic approaches have been developed for the synthesis of graphene. in a similar fashion to what is done with other nanostructured materials [22]. Because of the All the developed methodologies can be divided into two main categories: top-down and variousbottom-up, physicochemical in a similar properties, fashion to what graphene-based is done with other materials nanostructured promote amaterials functional [22]. interface Because of for the developmentthe various of physicochemical bioelectronics with properties, control graphene-b of interfacialased electrochemical materials promote properties. a functional Because interface graphene for showsthe anisotropic development behavior of bioelectronics at different with areas control of the of same interfacial surface, electrochemi it is said tocal possess properties. two idiosyncraticBecause partsgraphene (plane) on shows the sameanisotropic surface behavior i.e., basal at different and edge area planess of the (Figure same surface,3), with it diis ffsaiderent to possess electrochemical two properties.idiosyncratic Because parts of the(plane) structural on the same defect, surface the electrochemical i.e., basal and edge reactions planes at(Figure the basal 3), with plane different are slower thanelectrochemical at edge planes properties. on graphene Because interface. of the structural Meanwhile, defect, some the studieselectrochemical also report reactions the time-dependent at the basal chemicalplane activity are slower for than the basalat edge plane planes [23 on]. graphene interface. Meanwhile, some studies also report the time-dependent chemical activity for the basal plane [23].

FigureFigure 3. Schematic 3. Schematic representation representation of theof the basal basal plane plane (active (active part) part) and and the the edge edge plane plane (less(less activeactive part) of graphiticpart) of interfacegraphitic interface [24]. [24].

DirectDirect immobilization immobilization of graphene of graphene on macro on macro electrode electrode surfaces surfaces (glassy (glassy carbon carbon and gold)and gold) generates generates a heterogeneous interface. This type of heterogeneous interface initiates obstacles in a heterogeneous interface. This type of heterogeneous interface initiates obstacles in revealing prodigy revealing prodigy properties of the respective graphene. In this scenario, the electrode materials propertiesplay a ofcrucial the respectiverole and even graphene. dictate the In electr thisochemical scenario, thebehavior electrode of the materials interface [25,26]. play a crucial role and even dictateInteraction the electrochemical of graphene surface behavior with biomolecules of the interface in bioelectronics [25,26]. : Interaction of graphene surface Interactionwith biomolecules of graphene such as surface DNA, withprotein, biomolecules enzyme, and in bioelectronicscells depends: on Interaction colloidal force of grapheneand chemical surface withand biomolecules physical interactions. such as DNA, The interaction protein, enzyme, between and graphene cells dependssurface and on biomolecules colloidal force also and plays chemical a and physicalcrucial role interactions. in bio-catalytic The interactionprocesses. Investigations between graphene have surfaceproved that and biomoleculesthese interactions also are plays a crucialgenerally role in bio-catalyticcontrolled by processes.the size, shape, Investigations and surface havecharacteristic proved thatof graphene these interactions materials. There are generally are controlledthree possible by the size,types shape,from the and perspective surface characteristicof using graphene of graphene as bioelectronics, materials. these There include are (i) three possiblegraphene types fromin contact the perspective with the ofbiological using graphene substrate, as bioelectronics,(ii) surface of these graphene include defined (i) graphene by in physicochemical composition, and (iii) solid–liquid interfaces, where graphene interacts with the contact with the biological substrate, (ii) surface of graphene deﬁned by physicochemical composition, respective surroundings [27,28]. and (iii) solid–liquid interfaces, where graphene interacts with the respective surroundings [27,28].

3. Graphene in Biosensing Graphene is a 2D, zero-band gap semiconductor, an electroactive and transparent material with unique properties such as high enzyme operational stability, enzyme loading, good catalytic activity, larger surface area, fast electron transfer rate for enzyme immobilization and hence it is increasingly being used as immobilization carriers in bioelectronics (Figure4). Chemosensors 2020, 8, x FOR PEER REVIEW 5 of 12

3. Graphene in Biosensing Graphene is a 2D, zero-band gap semiconductor, an electroactive and transparent material with unique properties such as high enzyme operational stability, enzyme loading, good catalytic Chemosensorsactivity, 2020larger, 8, 45surface area, fast electron transfer rate for enzyme immobilization and hence it5 is of 12 increasingly being used as immobilization carriers in bioelectronics (Figure 4).

FigureFigure 4. 4.Enzyme Enzyme immobilization immobilization on graphene- diversity diversity in in applications. applications.

UseUse of biosensorsof biosensors approach approach is paramount is paramount for improvingfor improving the quality the quality of human of human life. This life. approach This is ableapproach to detect is able a wideto detect range a wide of compounds, range of comp andounds, it has and been it employedhas been employed to determine to determine a variety a of parametersvariety of of parameters biological of systems biological by translatingsystems by atranslating bio-recognition a bio-recognition event into aevent measurable into a measurable electronic or lightelectronic signal. or In light the last signal. century, In the several last century, researchers several have researchers reported have the reported interface the between interface the between biological andthe electronic biological world and electronic as bioelectronics world as [ 29bioelectronics–31]. Bioelectronics [29–31]. Bioelectro also assistsnics researchers also assists to researchers study the basicto study the basic relation between living systems and the artificial world. Because of breakthrough relation between living systems and the artificial world. Because of breakthrough experiments in experiments in the field of biosensors and basic research, functional bio-device developments can the field of biosensors and basic research, functional bio-device developments can build a platform build a platform whereby electronics can be integrated with biotechnology [32–34]. These types of whereby electronics can be integrated with biotechnology [32–34]. These types of advancement and advancement and developments in the field of biomaterials with various functionalities have developments in the field of biomaterials with various functionalities have increased the interests in increased the interests in bioelectronics [35]. Irrespective of the tremendous development in bioelectronicsbioelectronics, [35 there]. Irrespective is a lack of ofregulation the tremendous and control development of biological in systems bioelectronics, at devices there level. is a lack of regulationThe andremarkable control ofadvantage biological of systemsgraphene at over devices single-walled level. carbon nanotubes (SWCNTs) is its respectiveThe remarkable active surface advantage area of(2630 graphene m2 g−1) over[36]. single-walledGraphene-based carbon materials nanotubes have well-known (SWCNTs) is its respective active surface area (2630 m2 g 1)[36]. Graphene-based materials have well-known characteristics of metallic impurities, which· can− be elude by using the CNT. Graphene has been used characteristicsas a transducer of metallic in bio-field-effect impurities, whichtransistors can beand elude different by using biosensors the CNT. like Graphene electrochemical, has been usedimpedance, as a transducer and fluorescence. in bio-field-e Recentffect reports transistors of graphene-related and different materials biosensors have like been electrochemical, focused on impedance,(bio) sensor and development fluorescence. [17,37 Recent]. Functionalization reports of graphene-related of graphene materials by polymers have been enhances focused its on (bio)application sensor development in different [fields.17,37]. Because Functionalization of better biocompatibility of graphene by polymersand electro-catalytic enhances its behaviors, application inShan different et fields. al. Becausereported of betterin biocompatibility2009, a novel and electro-catalyticpolyvinyl pyrrolidone-protected behaviors, Shan et al. reportedgraphene/polyethylenimine-functionalized in 2009, a novel polyvinyl pyrrolidone-protected ionic liquid/glucose graphene/polyethylenimine-functionalized oxidase (GOD)-based ionicelectrochemical liquid/glucose biosensor oxidase with (GOD)-based an impressive electrochemical response for 14 biosensor mM of glucose with anconcentration. impressive Shan response et foral. 14 immobilized mM of glucose the enzyme concentration. onto pre-prepared Shan et al.graphene/AuNPs/chitosan immobilized the enzyme nanocomposite onto pre-prepared matrix grapheneand the/ AuNPsresultant/chitosan electrode nanocomposite showed a great matrix response and against the resultant glucose electrode with a linear showed range a great from response2 mM againstto 10 glucosemM, at with−0.2 V a linearand showed range fromgreat 2electro-catalytic mM to 10 mM, activity at 0.2 Vtoward and showed H2O2 and great O2 electro-catalytic[38]. Another − activity toward H2O2 and O2 [38]. Another study confirmed that the immobilization of graphene-Pt nanoparticles (NP) improved the detection performance with a wider linear range and lower detection limit for H2O2 and trinitrotoluene (TNT) [39]. The modification of reduced graphene with biomolecules to detect glucose was also reported by Wang et al. [40]. The authors reported a nontoxic direct electrochemical method that was used to functionalize and reduce graphene oxide (rGO) (Figure5) Chemosensors 2020, 8, x FOR PEER REVIEW 6 of 12

study confirmed that the immobilization of graphene-Pt nanoparticles (NP) improved the detection performance with a wider linear range and lower detection limit for H2O2 and trinitrotoluene (TNT) Chemosensors[39]. The2020 modification, 8, 45 of reduced graphene with biomolecules to detect glucose was also reported6 of 12 by Wang et al. [40]. The authors reported a nontoxic direct electrochemical method that was used to functionalize and reduce graphene oxide (rGO) (Figure 5) by using by using 3-aminopropyltriethoxysilane-modiﬁed conductive electrodes, and the modiﬁcation of the 3-aminopropyltriethoxysilane-modified conductive electrodes, and the modification of the reduced reduced GO-glassy carbon electrode (GCE) with glucose oxidase (GOx). GO-glassy carbon electrode (GCE) with glucose oxidase (GOx).

FigureFigure 5. 5.A A schematic schematic diagram diagram forfor electrochemicalelectrochemical reduction of of GO GO and and the the subsequent subsequent modification modiﬁcation ofof rGO rGO with with GOx GOx [40 [40].].

HuangHuang et al.et inal. 2011in 2011 studied studied the synthesis the synthesis procedure procedure of graphene-based of graphene-based biosensors. biosensors. The graphene The ﬁlmgraphene was prepared film was by prepared chemical by vapor chemical deposition, vapor deposition, and then functionalizedand then functionalized with anti- withE. coli anti-antibodiesE. coli viaantibodies the passivation via the passivation technique (Figuretechnique6). (Figure The nano-electronic 6). The nano-electronic sensor wassensor generated was generated based based on an antibody-modiﬁedon an antibody-modified chemical-vapor chemical-v depositionapor deposition grown grown large-sized large-size graphened graphene to detect to detect bacteria bacteria (E. coli ) with(E. greatcoli) with sensitivity great sensitivity (10 cfu/mL) (10 [cfu/mL)41]. [41]. Chemosensors 2020, 8, x FOR PEER REVIEW 7 of 12

FigureFigure 6.6. SchematicSchematic diagram diagram of anti-of E.anti- coliE.antibody coli antibody functionalized functionalized graphene-ﬁeld-e graphene-field-effectiveﬀective transistor transistor(FET), inset: (FET), SEM in imageset: SEM [41 image]. [41].

4. Switchable Graphene Bioelectronics Switchable biosensing system of bioelectronics is one of the recent and most promising applications. In the present era, switchable bioelectronics aims to develop new platforms for controlling and regulating the physiological mechanism in response to real life chemical and physical stimuli [42]. Switchable bioelectronics explores the natural biochemical interaction and mimics the biochemical reactions along with electron transfer phenomenon while controlling the environment under the influence of external stimuli (physical, chemical, electrical, and magnetic) [43]. A recent study in the field of switchable bioelectronics on graphene interface is at the stage of graphene-stimuli-responsive polymer hybrids capable of regulating and controlling the enzyme based bio-molecular reaction under the influence of temperature, pH, and light. Recently in 2018, Wang et al. fabricated highly sensitive electrochemical phenol biosensor by using a graphene oxide-modified tyrosinase interface. This fabricated phenol biosensor was based on the covalent bonding of tyrosinase (TYR) onto a GO modified glassy carbon electrode (GCE) via glutarldehyde (GA) (Figure 7). The -OH group of GO played an important role for covalent bonding between graphene interface and TYR for the constructing electrochemical phenol biosensors [44].

Figure 7. Schematic presentation of the development of TYR/GA/GO/GCE biosensor for the electrochemical determination of phenol derivatives; (a) GO casting on GCE, (b) covalent bonding of GO and GA, (c) covalent bonding of GA and TYR, and (d) electrochemical detection of phenol derivatives [44].

Chemosensors 2020, 8, x FOR PEER REVIEW 7 of 12

ChemosensorsFigure 20206. ,Schematic8, 45 diagram of anti-E. coli antibody functionalized graphene-field-effective7 of 12 transistor (FET), inset: SEM image [41].

4.4. Switchable Switchable Graphene Graphene Bioelectronics Bioelectronics SwitchableSwitchable biosensingbiosensing system system of bioelectronicsof bioelectronics is one is of one the recentof the and recent most promisingand most applications. promising applications.In the present era,In the switchable present bioelectronics era, switchable aims tobioelectronics develop new platformsaims to develop for controlling new andplatforms regulating for controllingthe physiological and regulating mechanism the in responsephysiological to real mechanism life chemical in andresponse physical to stimulireal life [42 chemical]. Switchable and physicalbioelectronics stimuli explores [42]. Switchable the natural biochemicalbioelectronics interaction explores and the mimics natural the biochemical biochemical interaction reactions along and mimicswith electron the biochemi transfercal phenomenon reactions along while with controlling electron the tran environmentsfer phenomenon under the while influence controlling of external the environmentstimuli (physical, under chemical, the influence electrical, of external and magnetic) stimuli [(physical,43]. A recent chemical, study electrical, in the field and of switchablemagnetic) [43].bioelectronics A recent study on graphene in the field interface of switchable is at thestage bioelectronics of graphene-stimuli-responsive on graphene interface polymeris at the stage hybrids of graphene-stimuli-responsivecapable of regulating and controlling polymer the hybrids enzyme capable based bio-molecularof regulating reactionand controlling under the the influence enzyme of basedtemperature, bio-molecular pH, and reaction light. under the influence of temperature, pH, and light. RecentlyRecently in in 2018, Wang et al. fabricated highly sensitive electrochemical phenol phenol biosensor biosensor by by usingusing a graphene oxide-modified oxide-modified tyrosinase interface. This This fabricated fabricated phenol phenol biosensor was based onon the covalent bonding of tyrosi tyrosinasenase (TYR) onto a GO modified modified glassy carbon electrode (GCE) via glutarldehydeglutarldehyde (GA) (Figure 77).). The -OH group ofof GOGO played an important rolerole forfor covalentcovalent bondingbonding betweenbetween graphene interface and TYR for the constructing electrochemical phenol biosensors [[44].44].

FigureFigure 7. SchematicSchematic presentation presentation of of the the developm developmentent of of TYR/GA/GO/GCE TYR/GA/GO/GCE biosensor biosensor for for the electrochemicalelectrochemical determination determination of of phenol phenol derivatives; derivatives; ( (aa)) GO GO casting casting on on GCE, GCE, ( (b)) covalent bonding ofof GO and GA, ( c) covalent bonding of of GA and TYR, and ( d) electrochemical detection of of phenol derivativesderivatives [44]. [44].

In the series of the development of graphene-based bioelectronics interface, in 2019, Sun et al. proposed a label-free and sandwiched-type immunosensor for the early diagnosis of primary liver cancer. In this study, an electrochemical immunosensor, based on label-free and sandwiched strategies was used where AuPt-vertical linked with graphene/GCE (AuPt-VG/GCE) for alpha-fetoprotein (AFP) detection. Because of noble biocompatibility and excellent conductivity, AuPt nanoparticles were used to immobilize the primary antibody (Ab1), while vertical graphene sheet helped to accelerate the electron transfer at the liquid–solid and solid–solid interface (Figure 8). The outcome of this investigation highlights that the sensing electrode played a crucial role in signal amplification and sensitivity of the sandwiched-type sensor, which is higher than that of label-free sensor. The fabricated immunosensor (AuPt-VG/GCE) has excellent performance for the detection of AFP in human serum along with the promising stability, recommending its potential application in clinical monitoring of AFP and other tumor marker [45]. The electrochemical measurements have shown that the sensing properties can be tuned by varying the external stimuli such as pH, temperature, ionic strength, and magnetic field. The fundamental design leads to a self-switching bio-catalysis by utilizing simple to use, cost-effective, and stable biodevices. In this respect, in 2014 Song et al. fabricated a PH-responsive electrochemical sensing platform based on the chitosan-reduced graphene oxide/concanavalin A (CS-rGO/ConA) for bi-analytical detection of glucose and urea. The strategy for the glucose and urea sensing involved the in situ pH-switchable enzyme catalyzed sensing behavior. The net charge of pH-dependent behavior of CS-rGO/ConA interface exhibited the pH-switchable responses against the negatively charged Fe(CN)63−. In this graphene-based dual bioreactor, oxidation of glucose catalyzed by glucose oxidase or the hydrolyzation of urea catalyzed by urease enzyme resulted in pH changes of the electrolyte solution to give a set of electrochemical response (Figure 9). This investigation reveals that sensing performance of graphene-based bioelectronic systems have potential for simultaneous assay of dual analytes (glucose and urea) [46]. This work has proven that two analytes can be detect in one sample and they can be discriminated based on their pH responsive environment of analytes solutions. In addition, this report also provides a strategy for the detection of non-electroactive species based on the enzyme catalyzed reaction. The summary of pioneered graphene-based Chemosensorsswitchable2020, 8, bioelectronic 45 is presented in Table 1. The table shows some of the pioneered works on the 8 of 12 fabrication of graphene-based devices that have been reported during the past decades.

FigureFigure 8. Schematic 8. Schematic illustration illustration ofof th thee development development label-free label-free and sandwiched-type and sandwiched-type approach for approach AuPt-VG/GCE-based bioelectronic interfaces for the detection of liver cancer marker i.e., for AuPt-VG/GCE-based bioelectronic interfaces for the detection of liver cancer marker i.e., alpha-fetoprotein [45]. alpha-fetoprotein [45].

The electrochemical measurements have shown that the sensing properties can be tuned by varying the external stimuli such as pH, temperature, ionic strength, and magnetic ﬁeld. The fundamental design leads to a self-switching bio-catalysis by utilizing simple to use, cost-eﬀective, and stable biodevices. In this respect, in 2014 Song et al. fabricated a PH-responsive electrochemical sensing platform based on the chitosan-reduced graphene oxide/concanavalin A (CS-rGO/ConA) for bi-analytical detection of glucose and urea. The strategy for the glucose and urea sensing involved the in situ pH-switchable enzyme catalyzed sensing behavior. The net charge of pH-dependent behavior of CS-rGO/ConA 3 interface exhibited the pH-switchable responses against the negatively charged Fe(CN)6 −. In this graphene-based dual bioreactor, oxidation of glucose catalyzed by glucose oxidase or the hydrolyzation of urea catalyzed by urease enzyme resulted in pH changes of the electrolyte solution to give a set of electrochemical response (Figure9). This investigation reveals that sensing performance of graphene-based bioelectronic systems have potential for simultaneous assay of dual analytes (glucose and urea) [46]. This work has proven that two analytes can be detect in one sample and they can be discriminated based on their pH responsive environment of analytes solutions. In addition, this report also provides a strategy for the detection of non-electroactive species based on the enzyme catalyzed reaction. The summary of pioneered graphene-based switchable bioelectronic is presented in Table1. The table shows some of the pioneered works on the fabrication of graphene-based devices that have been reported during the past decades.

Table 1. Graphene-based switchable bioelectronics.

Switchable S. No. Material System Preparation Method References Mechanism pH-induced on/oﬀ 1. Graphene-glucose oxidase pH based [47] bioelectronic interface Glucose oxidase/Tannic Direct electrochemistry pH and 2. [48] acid-reduced graphene oxide based interface temperature based 3. PANI-Graphene-AuNP Zipper-like interface Temperature based [49] Light-switchable 4. Graphene-polymer Light based [50] graphene interface Graphene/poly(NIPAM- Zipper like graphene pH and 5. [51] co-DEAEMA) based interface temperature based Flexibly switchable 6. Graphene/Fe O –AuNPs sDNA based [52] 3 4 peroxidase-like activity Chemosensors 2020, 8, x FOR PEER REVIEW 9 of 12

Table 1. Graphene-based switchable bioelectronics.

Switchable S. No. Material System Preparation Method References Mechanism pH-induced on/off 1. Graphene-glucose oxidase pH based [47] bioelectronic interface Glucose oxidase/Tannic Direct electrochemistry pH and temperature 2. acid-reduced graphene [48] based interface based oxide 3. PANI-Graphene-AuNP Zipper-like interface Temperature based [49] Light-switchable 4. Graphene-polymer Light based [50] graphene interface Graphene/poly(NIPAM-co Zipper like graphene pH and temperature 5. [51] -DEAEMA) based interface based Chemosensors 2020, 8, 45 9 of 12 Flexibly switchable 6. Graphene/Fe3O4–AuNPs sDNA based [52] peroxidase-like activity

Figure 9. Schematic illustration of pH-switchable behavior of Fe(CN)63−3 on the ConA/CS-rGO/GCE Figure 9. Schematic illustration of pH-switchable behavior of Fe(CN)6 − on the ConA/CS-rGO/GCE triggeredtriggered by by the the enzyme-catalyzed enzyme-catalyzed reactions reactions [[46].46].

5.5. Conclusions Conclusions and and Future Future Perspective Perspective InIn conclusion, conclusion, a widea wide range range of grapheneof graphene with with diff differenterent morphology, morphology, composition, composition, size, size, and and shape haveshape been have explored been forexplored the electrical for the communicationelectrical communication between electronic between andelectronic biological and counterparts.biological Thesecounterparts. electrical communicationsThese electrical co explainsmmunications the understanding explains the of understa the electronnding transfer of the andelectron charge transfer transfer pathwaysand charge in biologicaltransfer pathways systems. in It biological is very di ffisystems.cult to It predict is very the difficult impact to of predict this emergent the impact technology. of this Althoughemergent potential technology. applications Although of potential this technology applications heightens of this thetechnology interest heightens for further the research interest works for forfurther practical research and fundamentalworks for practical solutions and realfundamenta problems.l solutions This review real problems article focused. This review on switchable article focused on switchable bioelectronics technology based on stimuli-responsive interface. However, bioelectronics technology based on stimuli-responsive interface. However, this area is still at the this area is still at the zygotic level and needs more exploration. Meanwhile, the first report on zygotic level and needs more exploration. Meanwhile, the first report on graphene opened consequence graphene opened consequence advancement in the area of bioelectronics with large-scale, low-cost, advancement in the area of bioelectronics with large-scale, low-cost, high-quality methods for the high-quality methods for the identification, detection, and quantification of biomolecules. identification, detection, and quantification of biomolecules. Nevertheless, the present approach for Nevertheless, the present approach for fabricating bioelectronic devices is not well established for fabricating bioelectronic devices is not well established for practical application and commercialization. In conclusion, the device and materials biocompatibility along with new technological approaches for wearable and implantable bioelectronic devices needs further investigation.

Author Contributions: M.C., S.K.S., J.N. and V.K. planned to designed and prepare the draft; P.P.G., A.N., C.M.H., R.W., B.M. and R.S.B. put their contribution to scientific and English proof editing. All authors have read and agreed to the published version of the manuscript. Funding: This research received no external funding. Conflicts of Interest: There is no conflict of interest.

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