Electrochemical Sensing of Hydrogen Peroxide Using Block

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Electrochemical Sensing of Hydrogen Peroxide Using Block Copolymer Templated Iron Oxide Nanopatterns Salih Zeki Bas, Cian Cummins, Dipu Borah, Mustafa Ozmen, and Michael A. Morris Anal. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.analchem.7b03244 • Publication Date (Web): 11 Dec 2017 Downloaded from http://pubs.acs.org on December 12, 2017

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Analytical Chemistry

Electrochemical Sensing of Hydrogen Peroxide Using Block Copolymer Templated Iron Oxide Nanopatterns Salih Zeki Bas†, Cian Cummins‡, Dipu Borah‡*, Mustafa Ozmen†, and Michael A Morris‡* †

Salih Zeki Bas, Mustafa Ozmen

Department of Chemistry, Selcuk University, 42075, Konya, Turkey. ‡

Cian Cummins, Dipu Borah, Michael A Morris*

AMBER Centre and CRANN, Trinity College Dublin, Dublin 2, Ireland.

*Corresponding Authors: Email: [email protected] (DB), Email: [email protected] (MAM) Tel: +353 1 896 3089

1 ACS Paragon Plus Environment

Analytical Chemistry 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Abstract: A new enzyme-free sensor based on iron oxide (Fe3O4) nanodots fabricated on an indium tin oxide (ITO) substrate via a block copolymer (BCP) template was developed for high sensitive and selective detection of hydrogen peroxide (H2O2). The self-assembly based process described here for Fe3O4 formation is a simple, cost effective, and reproducible process. The H2O2 response of the fabricated electrodes was linear from 2.5×10-3 to 6.5 mM with a sensitivity of 191.6 µA mM−1cm−2 and a detection limit of 1.1×10-3 mM. The electrocatalytic activity of Fe3O4 nanodots toward the electroreduction of H2O2 was described by cyclic voltammetric and amperometric techniques. The sensor described here has a strong anti-interference ability to a variety of common biological and inorganic substances.

Keywords: enzyme-free sensor, hydrogen peroxide, electrocatalytic reduction, block copolymers, self-assembly, nanopatterned iron oxide.


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Introduction Selective and sensitive determination of hydrogen peroxide (H2O2) with accuracy and a fast response time has significant implications in food, mining, textile, pharmaceutical, clinical and environmental applications.1 In living organisms, H2O2 also plays an essential role as a signaling molecule in regulating various biological processes as well as its wellknown cytotoxic effects.2 For example, experimental studies establish that H2O2 is produced by mitochondria through a specialized enzyme to control cellular growth and death.3 H2O2 is also a product released during the oxidation of substrate catalyzed by enzymes such as glucose oxidase,4 cholesterol oxidase,5 lysine oxidase6 and xanthine oxidase.7 Hence, the studies related to developing new methodologies for the quantitative determination of H2O2 concentrations in solution are of significant clinical and industrial importance. Several analytical methods such as titrimetry, spectrophotometry, fluorimetry, chemiluminesence and electrochemical methods have been reported for quantitative

determination of H2O2.8,9,10,11 Compared with these, the determination of H2O2 based on electrochemical reduction and/or oxidation has some advantages such as simplicity of operation and automation, fast response time, accuracy, low cost, high sensitivity and selectivity.12 Commercially available H2O2 sensors can be enzyme-based or enzyme-free.13,14 Enzyme-based sensors involving redox active biomolecules such as horseradish peroxidase (HRP),15 cytochrome c,16 myoglobin,17,18 etc., have received extensive attention owing to the highly challenging task of bio-molecule immobilization at the electrode surface. However, these sensors suffer from intrinsic drawbacks that limit their applications in analytical field and are therefore unsuitable for mass production. These disadvantage include restricted activity and storage time, complicated preparation steps, poor reproducibility, rigorous operating conditions (pH, temperature, humidity, ionic strength, etc.) and high cost.19,20 These problems have ensured that enzyme free H2O2 sensor research for both academic and industrial purposes has been of signifcant interest. Current efforts to develop enzyme free H2O2 sensors have focused on using novel nanomaterial based techniques (mostly metal, metal oxides or hybrid materials based) around high electrocatalytic activity on transducers.2,21 These nanomaterials have attracted attention because of their superior chemical, physical, and electronic properties, in comparison to bulk materials.22 Recently, magnetic metal oxide nanoparticles, especially iron oxides (Fe3O4), have drawn considerable interest for the fabrication of enzyme-free sensors, owing to their good biocompatibility, strong superparamagnetic properties, low toxicity and simple preparation processes.23 3 ACS Paragon Plus Environment


Whilst some enzyme-free H2O2 sensors based on nanomaterials exhibit good analytical performances, they are expensive and are difficult to mass produce.24 This has led to renewed interest in developing simple, facile, and reliable enzyme-free sensors. Bottom-up, thin-film based technology for fabricating nanopatterns offers advantages including low cost, simple synthesis, high durability, high surface area, good electric properties, and possibility of mass manufacturing of high quality sensors. 25,26 Electrically conductive indium-tin oxide (ITO) coated glass substrates are an alternative platform for electrochemical sensor applications25 owing to their low cost in comparison to other substrates, e.g. silicon. Developing precise nanopatterns on ITO substrates with surfaces of high surface area, as well as physical, chemical and thermal stability is a challenge. Various top-down lithographic approaches such as electron beam, optical interference, X-rays, nanoimprint, etc. are used to develop nanoengineered patterns in microelectronic industries, however, they are time intensive, costly and low throughput processes. Block copolymer (BCP) self-assembly offers an alternative low cost bottom-up methodology for surface nanoengineering.27 BCPs self-assemble into regular well-defined features in the nanometer range under suitable processing (vacuum annealing, solvent annealing etc). These patterns can be useful for various nanotechnology applications as scaffolds/templates and can be used for incorporating functional material.28 Generally inorganic nanodots/nanoparticles are incorporated into reactive BCP domains of solution borne micelles.29,30 This can lead to polymer swelling effects and other that can result in nanoparticle size variation or agglomeration. The ability to successfully incorporate metallic species post patterning of the BCP eliminates size distribution variations, and can be carried out using a BCP containing a reactive block. For example, polystyrene-blockpoly4vinylpyridine (PS-b-P4VP) is a BCP of particular interest, due to the electron donating nitrogen atom in P4VP. Other BCPs such as polystyrene-b-polyethylene oxide (PS-b-PEO) have also been successfully infiltrated to act as an electrochemical sensor for detection of EtOH and H2O2.25 Herein, we report the development of an enzyme-free sensor based on Fe3O4 nanodots for detection of H2O2 via incorporation of iron species into a nanoporous PS-b-P4VP template on ITO substrates. The electrocatalytic behavior of Fe3O4 nanodots toward the electroreduction of H2O2 was studied by using cyclic voltammetry (CV) and amperometry. The performance of the sensor, i.e. linear range, detection limit, sensitivity, response time, reproducibility, repeatability and stability was also described. The interference effect of a 4 ACS Paragon Plus Environment

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range of common biological and inorganic substances on the response of the sensor was evaluated.

Experimental Section Materials and Reagents: Indium tin oxide (ITO) coated glass slides, square with a surface resistivity of 8-12 Ω/sq were purchased from Sigma Aldrich. Polystyrene-blockpoly4vinylpyridine (PS-b-P4VP) was purchased from Polymer Source, Inc., Canada, with a molecular weight of Mn = 33.5 kg mol−1 (MnPS = 24 kg mol−1; MnP4VP = 9.5 kg mol−1, fPS = 0.70), a polydispersity (Mw/Mn) of 1.15 (where, Mn and Mw are number average and weight average molecular weights) and was used as received. Iron (III) nitrate nonahydrate (Fe(NO3)3.9H2O),

ethanol

(dehydrated,

200

proof),

toluene

(99.8%,

anhydrous),

tetrahydrofuran (THF) (99.8%, anhydrous), ethylene glycol (EG, CH2(OH)CH2(OH), 95.0%) were purchased from Sigma-Aldrich. Hydrogen peroxide solution (30%, extra pure) was obtained from Merck (Darmstadt, Germany). Phosphate buffer saline solution (PBS) was prepared using NaH2PO4 (Merck) and NaOH (Merck). NaCl (99.5%), KCl (99%,), MgCl2 (99%), Zn(NO3)2 (98.5%) and lactic acid (extra pure, from Merck); D-(+)-glucose (≥99.5%, from Sigma Aldrich); uric acid (99%, from Alfa Aesar); ascorbic acid (from J. T. Baker) and CaCl2 (99%, from Riedel) were purchased. Substrate Modification and Block Copolymer Film Deposition: ITO substrates were cleaned with ultrasonication treatment in ethanol for 20 minutes. Solution of ethylene glycol (EG) 5% (v/v) was prepared in ethanol and was stirred at room temperature (~20°C) for 30 minutes to ensure complete mixing. The ITO substrates were coated with EG by spin-coating (P6700 Series Spin-coater, Speciality Coating Systems, Inc., USA) at 3000 rpm for 30 seconds. Samples were then air dried at room temperature (~20°C) for 30 minutes, washed with absolute ethanol and then dried under a stream of nitrogen. The PS-b-P4VP BCP material was dissolved in a toluene: THF (80:20) mixture to yield a 0.5 wt. % solution. BCP films were prepared by spin coating the polymer solution onto the substrates at 3200 rpm for 30 seconds. The PS-b-P4VP generated thin films were exposed to a saturated THF environment at 50°C between 4-6 hours. Further details on this fabrication process can be found in our previously published work.31 Surface Reconstruction and Iron Oxide Inclusion: Nanoporous films were developed through use of a P4VP selective solvent (ethanol). Samples were immersed in ethanol for 20 minutes and then dried under nitrogen flow. A 0.4 wt. % Fe(NO3)3.9H2O solution was prepared in ethanol (dehydrated, 200 proof) and left to stir until completely dissolved. The 5 ACS Paragon Plus Environment


solution was then spin coated onto the reconstructed porous film at 3000 rpm for 30 seconds. Following this the polymer matrix template was removed using UV/ozone treatment for 3 hours. Samples were ozonated in a UV/Ozone system (PSD Pro Series Digital UV Ozone System; Novascan Technologies, Inc., USA). Characterization: Atomic Force Microscopy (AFM) (Park systems, XE-100) was operated in AC (tapping) mode under ambient conditions using silicon microcantilever probe tips with a force constant of 42 N m−1. AFM images were processed using ImageJ software to calculate dimensions. X-ray Photoelectron Spectroscopy (XPS) was performed under ultrahigh vacuum conditions (< 5×10-10 mbar) on a VG Scientific ECSAlab Mk II system using Al Kα X-rays (1486.6 eV). The analyzer pass energy was set to 100 eV for survey spectra recorded. High resolution scans were processed with pass energy set to 20 eV. The binding energy scale was referenced to the adventitious carbon 1s core-level at 284.8 eV. Electrochemical Measurements: A CHI-660C electrochemical workstation (CH Instruments Co., USA) was used to perform all the electrochemical measurements as described elsewhere32. A conventional three electrode system was used with a modified ITO electrode (surface area: 0.01 cm2) as working electrode, a platinum wire as auxiliary electrode and an Ag/AgCl (3 M KCl) electrode as reference electrode. In electrochemical measurements, the electrical connection to the working electrode was also provided directly. CVs were recorded in 0.1 M PBS (pH 7.4) between -0.1 and -0.8 V at a scan rate of 50 mV s1

. Amperometric measurements were performed in a cell (with a volume of 20 mL)

containing 0.1 M PBS (pH 7.4) at the applied potential of -0.55 V at room temperature. The buffer solution in the cell was saturated with nitrogen for 20 min to obtain a N2-saturated solution before all electrochemical measurements.

Results and discussion Fabrication of Nanopatterned Fe3O4 Electrode on ITO Substrate Scheme 1 shows the overall process flow for the fabrication of Fe3O4 nanodots via BCP selfassembly. Initially, the ITO substrate was modified through a simple functionalization using ethylene glycol that has been described and characterized previously.31,33 This functionalization enables enhanced coverage of the substrate while increasing order of the BCP film deposited thereon. A dilute PS-b-P4VP solution was spin coated on the modified ITO surface and the film was then subsequently solvent vapor annealed (50°C) using tetrahydrofuran to form perpendicular cylinder arrangements. The enhanced chain mobility


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during solvent treatment allows the PS-b-P4VP to microphase separate with a high degree of order.

Scheme 1. Overall process flow for development of iron oxide (Fe3O4) nanodots. (a) Initial ITO surface functionalization with ethylene glycol and spin coating of PS-b-P4VP BCP on functionalized surface. (b) Solvent vapor anneal with THF at 50 °C of spin cast film producing cylinder perpendicular (yellow cylinders, P4VP) to ITO substrate. (c) Surface reconstruction using ethanol giving nanoporous template and spin coating of iron nitrate ethanolic solution on surface reconstructed film. (d) UV/ozone treatment for 3 hours of iron nitrate film successfully removes polymer matrix and develops iron oxide nanodots pattern.

Figure 1. (a) AFM topographic image of PS-b-P4VP film after solvent vapor annealing. Inset shows FFT with high degree of order. (b) Size distribution calculation show P4VP nanocylinders with 23.8 nm diameter. (c) AFM topographic image of reconstructed film showing nanopores following ethanol treatment. (d) Pore size was calculated at 19.1 nm. (e) 7 ACS Paragon Plus Environment


AFM topographic image of iron oxide nanodots arrays after spin coating metal nitrate solution on reconstructed film (c). (f) Size distribution calculation shows nanodots having diameter of 21.6 nm. The resulting domains are well-ordered, highly-defined and over large areas as shown in the topographic AFM image in Figure 1a. The histogram analysis in Figure 1b shows the P4VP cylinder diameter to be 23.8 nm. Following self-assembly of the PS-b-P4VP film to form well-ordered arrays, a reconstruction process31 was carried out to enable metal inclusion. The solvent vapor annealed films were immersed in ethanol (a selective solvent for P4VP and a non-selective solvent for PS) for a 20 minute period. After this swelling treatment nanoporous domains are formed as seen in the topographic AFM image in Figure 1b. The features are notably different from the self-assembled pattern in Figure 1a, improved contrast etc., and suggestive of a change in structure as has been seen by us in earlier work.29 The pores were measured at 19.1 nm in diameter (Figure 1c). An ethanolic solution of the iron nitrate material was spin coated on the porous film to fabricate Fe3O4 nanodots. After this, UV/ozone exposure was carried out to remove the polymer matrix and leave the remaining high density iron oxide nanodots array as shown in Figure 1e. The inset in Figure 1e shows the FFT revealing the high degree of long range order. As shown in Figure 1f the Fe3O4 nanodots produced have a diameter of 21.6 nm close to that of the original BCP pattern. It should be noted that the roughness of ITO substrates is higher than that of silicon used for the formation of iron oxide nanodots arrays in our previous work. We believe this contributes to the variation seen in the self-assembled (Figure1a) and iron oxide nanodots array (Figure1e) seen in the AFM data.


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Figure 2. Survey XPS spectra is shown of the iron oxide nanodots array on ITO substrate after UV ozone treatment. The inset shows the high resolution Fe 2p core level spectrum. XPS analysis of the iron oxide nanodots is shown in Figure 2. The survey spectra (Figure 2) of the Fe3O4 samples after UV/Ozone treatment show the presence of In, Sn, O, (all three elements expected from the substrate) and the presence of Fe and O from the sensing material at the substrate surface. C signal is attributed to adventitiously adsorbed carbon and/or residual material from the BCP. High resolution Fe 2p spectra were recorded to identify the exact phase of the iron oxide material developed. The high resolution Fe 2p core level spectrum in the inset of Figure 2 shows two broadened peaks associated with Fe 2p3/2 at 710.7 eV and Fe 2p1/2 at 723.1 eV due to the existence of both Fe2+ and Fe3+ ions. These values are in agreement with previous literature of iron oxide material34 and the concentration ratio of Fe3+/Fe2+ here is close to 2:1 confirming the formation of Fe3O4 material.

Electrochemical Studies Electrocatalytic Performance of Fe3O4 Nanopatterned Electrode The electrochemical performance of the Fe3O4 nanodots-ITO toward H2O2 detection was evaluated by comparing its CV with those of different ITO electrodes such as bare ITO and pol-ITO. Figure 3 provides typical CVs of bare ITO (curves a and b), pol-ITO (curves c and d) and Fe3O4 nanodots-ITO (curves e and f) in the absence and the presence of 0.25 mM H2O2 in N2-saturated 0.1 M PBS (pH 7.4) at a scan rate of 50 mV s-1. There little indication of any redox features observed at bare ITO and pol-ITO in the absence of H2O2 in the potential range between -0.1 and -0.8 V, and any signals from redox reactions are negligible compared with that of Fe3O4 nanodots-ITO. As shown in curve e of Figure 3, a pair of welldefined redox peaks with an anodic peak at -0.41 V and a cathodic peak at -0.57 V are observed for the nanodots coated substrates. In the presence of 0.25 mM H2O2, there are minor cathodic peak currents for both bare ITO and pol-ITO (inset of Figure 3). Compared with the CV (curve e) of Fe3O4 nanodots-ITO in the absence of H2O2, the CV (curve f) of Fe3O4 nanodots-ITO displays a significant enhancement of the cathodic peak current with the addition of 0.25 mM H2O2 as a result of an irreversible reduction of H2O2. The increase in the cathodic peak of Fe3O4 nanodots-ITO also indicates that Fe3O4 nanodots has excellent electrocatalytic activity to the electroreduction of H2O2.



Figure 3. CVs of bare ITO (a,b), pol-ITO (c,d) and Fe3O4 nanodots-ITO (e,f) in the absence (a,c,e) and presence (b,d,f) of 0.25 mM H2O2 in 0.1 M PBS (pH 7.4). Inset: CVs of bare ITO (a,b) and pol-ITO (c,d) in the absence (a,c) and presence (b,d) of 0.25 mM H2O2 in 0.1 M PBS (pH 7.4). Scan rate: 50 mV s-1. To determine the type of the controlled process, the effect of the scan rate on the peak current was also investigated by monitoring CVs of Fe3O4 nanodots-ITO electrode in the presence of 0.25 mM H2O2 in 0.1 M PBS (pH 7.4) at different scan rates. Naturally, the cathodic (Ipc) peak currents increased with increasing scan rate from 20 mVs-1 to 100 mVs-1 (Figure 4). The cathodic peak currents are linearly proportional to the square root of scan rate in the range of 20-100 mV s-1 with correlation a coefficient R of 0.9973, as shown in the inset of Figure 4, indicating that the redox behavior of Fe3O4 nanodots-ITO is a diffusion-controlled process.

Figure 4. CVs of Fe3O4 nanodots-ITO in the presence of 0.25 mM H2O2 in 0.1 M PBS (pH 7.4) at different scan rates of (a) 20, (b) 30, (c) 40, (d) 50, (e) 60, (f) 70, (g) 80, (h) 90, (i) 100 mV s-1. Inset: (a) Plot of cathodic (Ipc) peak currents vs. square root of scan rate.


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Figure 5. CVs of Fe3O4 nanodots-ITO in the presence of 0.25 mM (a), 0.50 mM (b), 0.75 mM (c), 1 mM (d) H2O2 at scan rate of 50 mV s-1. Inset: Effect of applied potential on response current of Fe3O4 nanodots-ITO to 0.25 mM H2O2 in 0.1 M PBS (pH 7.4). The electrocatalytical activity of the Fe3O4 nanodots-ITO electrode toward H2O2 was investigated by CV in the presence of different concentrations of H2O2 in 0.1 M PBS (pH 7.4) at a scan rate of 50 mV s-1 (Figure 5). It is notable that the cathodic current of Fe3O4 nanodots-ITO increased gradually with increasing the concentration of H2O2 from 0.25 mM to 1 mM due to the electroreduction of H2O2 on the surface of Fe3O4 nanodots-ITO electrode. The dependence of the response current of Fe3O4 nanodots-ITO to 0.25 mM H2O2 in 0.1 M PBS (pH 7.4) at different potentials between -0.40 and -0.70 V was examined amperometrically. As shown in the inset of Figure 5, the amperometric responses showed that the current signal increased gradually with increasing potential applied up to -0.55 V, and then decreased for potential higher than -0.55 V. Thus, a potential of -0.55 V was selected as the applied potential for amperometric detection of H2O2 in the following experiments.

Amperometric Performance of Fe3O4 Nanopatterned Electrode In order to develop an amperometric method for H2O2 determination using Fe3O4 nanodotsITO, current signals of the proposed sensor were measured in the bulk solution. Figure 6 shows the typical amperometric current of Fe3O4 nanodots-ITO for successive injection of different concentrations of H2O2 (0.05, 0.01 and 0.1 M) into 0.1 M PBS (pH 7.4) at an applied potential of -0.55 V. During the analysis performed with Fe3O4 nanodots-ITO, a gradual increase in the reduction current of H2O2 was observed with stepped increasing of H2O2 concentration in PBS, indicating that Fe3O4 nanodots in the structure of sensor can catalyze electrochemically the 11 ACS Paragon Plus Environment


Figure 6. Amperometric response of Fe3O4 nanodots-ITO for successive injection of different concentrations of H2O2 into 0.1 M PBS (pH 7.4) at the applied potential of -0.55 V. Inset: Calibration curve of Fe3O4 nanodots-ITO for H2O2 detection. reduction of H2O2 to H2O. Inset of Figure 6 shows the change in the response currents of Fe3O4 nanodots-ITO as a function of H2O2 concentration. The statistical analysis of the calibration curve of Fe3O4 nanodots-ITO gave the following equation y = 7.48x + 1.69 (with a correlation coefficient of 0.998), where y is the current (µA) and x is H2O2 concentration (mM). The response current of Fe3O4 nanodots-ITO electrode showed a linear relationship with H2O2 concentration from 2.5×10-3 to 6.54 mM. Fe3O4 nanodots-ITO reached 95% of the steady-state signal within 5-8 s, indicating a fast amperometric response to the reduction of H2O2. The detection limit of Fe3O4 nanodots-ITO was calculated to be 1.1×10-3 mM, according to the 3SD/m criterion where m is the slope of the calibration curve of sensor and SD is the standard deviation for the amperometric response to 5×10-3 mM H2O2 for 5 times.35 The comparison of the analytical performance of Fe3O4 nanodots-ITO for H2O2 detection with other H2O2 sensors reported recently in the literature is shown in Table 1 in terms of linear range, detection limit (LOD) and sensitivity. The present sensor has better or comparable characteristics than most of the H2O2 sensors given in Table 1. As can be seen from Table 1, the linear range of Fe3O4 nanodots-ITO is only lower than that of Cu2O/Cu nanocomposite modified electrode,36 Ag-SiO2 modified GCE37 and AuPd-GR modified ITO electrode,38 and wider than those of the other sensors. The current sensitivity was found to be 191.57 µA mM-1 cm-2 to H2O2 for Fe3O4 nanodots-ITO, indicating that the present sensor has high sensitivity compared with those of other hydrogen peroxide sensors.37,38,39,40,41,42 12 ACS Paragon Plus Environment

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Therefore, we can conclude that the Fe3O4 nanodots-ITO can be used as an enzyme-free platform for the effective determination of H2O2 with wide linear range, low detection limit, high sensitivity and selectivity. In biosensing systems based on the electroreduction of H2O2 formed during the enzymatic reaction, many compounds such as ferric hexacyanoferrate, Prussian blue which has very attractive redox properties have been widely used as redox mediators.48,49 Alternatively, Fe3O4 nanodots-ITO may exhibit the effective electron-transfer ability towards the reduction of H2O2 produced between an enzyme and its substrate. Fe3O4 nanodots-ITO can provide also many advantages such as large surface-to-volume ratio, high surface activity, and strong adsorption ability for the immobilization of biomolecule in the construction of a biosensor.50,51,52 Table 1. Comparison of the analytical characteristics of various modified electrodes toward H2O2 detection. Eapp

Linear range

LOD

Sensitivity

(V)

(mM)

(mM)

(µA mM-1cm-2)

Ag@BSA/Au

-0.5

5×10-3 – 1.5

1.6×10-4

101.3

39

Pt NP-Ni foam

0

5×10-3 – 0.85

3×10-4

829

43

Cu2O/Cu NCs

-0.2

4×10-4 – 10

2×10-4

870

36

Electrodea

PEDOT-PtNPs/SPC CuS/CS/GCE MnO2-Ag nanowire/GCE

-0.6 -0.1

up to 6.0

1.6×10

-3

1×10 – 0.1

3×10

-4

-3

-4

Reference

19.29

40

b

36.4

41

2.4×10

-4

-

44

-3

-

45

-0.5

2.4×10 – 4

PDDA-AuNSs/GCE

-0.4

-2

2×10 – 2.5

9.7×10

Ag@SiO2@Ag/GCE

-0.2

5×10-3 – 24

1.7×10-3

56.07

37

Pt0.5Au0.5@C

0.3

7×10-3 – 6.5

2.4×10-3

210.3

46

CoSn(OH)6 Nanocube

-0.6

4×10-3 – 0.4

1×10-3

-0.6

-3

AuPd@GR3:15/ITO

5×10 – 11.5 -2

-

47

1×10

-3

186.86

38

-3

119.35

42

191.57

Present work

Grass-like CuO/GCE

-0.6

1×10 – 0.3

5×10

Fe3O4 nanodots-ITO

-0.55

2.5×10-3 – 6.54

1.1×10-3

a

BSA: bovine serum albumin, PEDOT: poly(3,4-ethylenedioxythiophene), SPC: screen-

printed carbon, CS: chitosan, PDDA: poly(diallyldimethylammonium chloride), AuNSs: gold nanostars, TeO2-NWs: TeO2 nanowires, GR: graphene. b

µA mM-1. 13 ACS Paragon Plus Environment


Reproducibility, Repeatability, Interference and Stability of Fe3O4 Nanopatterned Electrode The reproducibility and repeatability are important parameters for the evaluation of the sensor performance. The reproducibility of Fe3O4 nanodots-ITO electrode was estimated from the response to 0.25 mM H2O2 using five different electrodes. The relative standard deviation was found to be 2.93%, indicating a good reproducibility of the sensor preparation. The repeatability of the sensor was also examined by monitoring the current response to 0.25 mM H2O2 for five times using the same electrode. The relative standard deviation was 2.14% for Fe3O4 nanodots-ITO.

Figure 7. (a) Amperometric response of Fe3O4 nanodots-ITO with addition of 2 mM D(+)glucose (Glu), 2 mM uric acid (UA), 2 mM lactic acid (LA), 2 mM ascorbic acid (AA) and 1 mM H2O2 in 0.1 M PBS (pH 7.4) at the applied potential of -0.55 V. (b) Amperometric response of Fe3O4 nanodots-ITO with addition of 2 mM NaCl, 2 mM KCl, 2 mM CaCl2, 2 mM MgCl2, 2 mM Zn(NO3)2 and 1 mM H2O2 in 0.1 M PBS (pH 7.4) at the applied potential of -0.55 V.


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The interference effect of some common biological substances such as glucose, uric acid, lactic acid, ascorbic acid on the sensor was investigated at -0.55 V in 0.1 M PBS (pH 7.4). The current responses of Fe3O4 nanodots-ITO in the presence of 2 mM D(+)-glucose, 2 mM uric acid, 2 mM lactic acid, 2 mM ascorbic acid were compared with the current signal obtained for 1 mM H2O2 solution. As seen from Figure 7a, no appreciable signals were observed with the addition of these substances, revealing a high selectivity of the sensor for the amperometric detection of H2O2. The effect of various inorganic substances such as NaCl, KCl, CaCl2, MgCl2, Zn(NO3)2 on the sensor was also studied, and the amperometric responses were shown in Figure 7b. No noticeable changes in the current responses of Fe3O4 nanodots-ITO were detected in 2 mM CaCl2, 2 mM MgCl2 and 2 mM Zn(NO3)2 solutions. This may be due to precipitation with phosphate ions in the electrolyte. The phosphate ions tend to precipitate with Ca2+, Mg2+ and other polyvalent cations. The concentration of divalent cations decreases with precipitation formation and no noticeable signal is observed. However, it was found that the presence of 2 mM NaCl and 2 mM KCl have the current values of 1.52% and 1.38% for Fe3O4 nanodots-ITO when the response of 1 mM H2O2 is taken as 100%. The effects of interferences from NaCl and KCl can be negligible for the H2O2 sensing. These results indicate that the proposed sensor has a strong anti-interference ability. The storage stability of the sensor was determined by measuring steady-state response current of 0.25 mM H2O2 over a period of 20 days. The sensor was stored under dry conditions at room temperature when it was not in use. The results obtained for Fe3O4 nanodots-ITO were shown in Figure 8, where i0 is the initial response current of the sensor freshly fabricated, i is the response current at any storage time, i-i0 is the change in the response current at any storage time. Fe3O4 nanodots-ITO exhibited good stability during 5 days and the response current remained about 90% of its initial response. After that, an activity loss of 28% was observed after 12 days.



Figure 8. Stability of Fe3O4 nanodots-ITO (Each data point of graph is based on measuring the amperometric response of 0.25 mM H2O2 in 0.1 M PBS (pH 7.4) at the applied potential of -0.55 V). Conclusions The fabrication and development of a new enzyme-free sensor for monitoring H2O2 is described in this paper. The prepared sensors where patterned with Fe3O4 nanodots using a BCP nanofabrication strategy. The simplicity of the BCP approach was used to include functional material (Fe3O4) as a viable nanosensor to detect H2O2. The results of the electrochemical experiments exhibited that Fe3O4 nanodots has an excellent electrocatalytic activity toward the reduction of H2O2. Both voltammetric and amperometric measurements showed that Fe3O4 nanodots can be used as an enzyme-free H2O2 sensor with a low detection limit, high selectivity and sensitivity. The linear range of the detection of H2O2 was from 2.5×10-3 to 6.54 mM with a detection limit of 1.1×10-3 mM. Furthermore, the proposed sensor has a strong anti-interference ability to some common biological and inorganic substances. The feasibility of employing BCP templated Fe3O4 nanodimensioned H2O2 sensor as demonstrated in our work may enable the application of such a low cost platform for detecting various (bio)electrochemical species.

Acknowledgements Financial support for this work is provided by the EU FP7 NMP project, LAMAND (grant number 245565) project and the Science Foundation Ireland (grant number 09/IN.1/602), and gratefully acknowledged.


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