DNA mediated electrocatalytic enhancement of α-Fe₂O₃–PEDOT–C-MoS₂ hybrid nanostructures for riboflavin detection on screen printed electrode

Abstract

A facile synthesis of iron oxide nanorods and PEDOT(poly(3,4-ethylenedioxythiophene)) nanospheres on carbon supported MoS₂ (C-MoS₂) is reported for riboflavin (RF) sensing. Furthermore, a novel aqueous-based DNA wrapped on an α-Fe₂O₃–PEDOT–C-MoS₂ scaffold shows high electrocatalytic activity compared to that of the α-Fe₂O₃–PEDOT–C-MoS₂ composite in biosensing. α-Fe₂O₃–PEDOT–C-MoS₂/GCE demonstrates the linear response of RF in the concentration range from 800 nM to 700 μM, with a detection limit of 79 nM (S/N = 3σ/b), whereas the α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE composite shows a wider range from 300 nM to 1 mM with a comparatively low detection limit of 5 nM. Similarly, α-Fe₂O₃–PEDOT–C-MoS₂–DNA/SPE exhibits a still wider range from 100 nM to 1 mM, with a detection limit of 12 nM. Interestingly, it is also observed that α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE reduces the oxidation potential of RF by 30 mV. Thus, the excellent behavior of the proposed biosensor can be attributed to the unique behavior of DNA, which provides a wider detection range and good electrocatalytic behavior towards RF. The fabricated sensor exhibited highly sensitive and selective detection of RF. Real sample analysis was also executed for human urine, milk powder and pharmaceutical drugs without any preliminary treatment.

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1. Introduction

Riboflavin (RF), a well-known vitamin, is the primary redox active component of flavin mononucleotide and flavin adenine dinucleotide in blood plasma; it undergoes a wide variety of cellular processes, such as metabolism of fats, ketone bodies, carbohydrates, and proteins.^1,2 Therefore, the determination of trace amounts of RF is important in humans and animals. This, in turn, requires superior methods for determining RF and the development of rapid, selective, and highly sensitive techniques. Among the various techniques, electrochemical methods are an attractive option because they are inexpensive, reliable long-term, reproducible and convenient.

Conducting polymers have attracted major attention for their applications in a variety of fields, including photovoltaics,^3,4 electronics,^5,6 sensing,^7,8 electroluminescence^9,10 and biosensors,^11,12 especially due to their optical properties, structural modification, conductivity and comparably low cost. PEDOT, a recently discovered, excellent conducting polymer, has been reported to exhibit enhanced behavior compared to its counterparts.^13,14 Recently, our group investigated the PEDOT polymer matrix for the development of ultra-sensitive label free electrochemical DNA sensors. The fabricated sensor was capable of effectively discriminating single and double base mismatch DNA and completely non-complementary DNA.¹⁵

For analogous reasons, nanostructured metal oxides are generating great interest, owing to their enhanced electron-transfer kinetics and strong adsorption, which provide suitable microenvironments for the immobilization of biomolecules and result in improved biosensing characteristics.^16–19 Maiyalagan et al.²⁰ reported that nanostructured α-Fe₂O₃ in fact exhibits noble metal behavior and is also found to possess an intrinsic enzyme-mimicking activity similar to that found in natural horseradish peroxidase. Hence, α-Fe₂O₃ could be utilized for enzyme free biosensing applications. Very recently, we reported that α-Fe₂O₃ can play the key role in achieving a minimal detection limit due to its catalytic and enhanced conductive behavior in combination with an Au–Pd system.²¹

Recently, molybdenum disulfide (MoS₂), a newly discovered bandgap-adjustable semiconductor composed of S–Mo–S triple layers, has attracted great interest in the fields of electrochemistry, catalysts, sensors, capacitors, and lithium-ion batteries.^23–25 MoS₂ has a sandwich structure similar to graphene;²² its electrocatalytic activity originates from the sulfur edges of the MoS₂ layers, while the inner planes are catalytically inert. W. H. Hu et al.²⁶ reported the highly active and stable electrocatalytic behavior of the MoS_x/GO matrix, owing to its good dispersion and conductivity, for the hydrogen evolution reaction. However, the improvement of MoS₂ electrocatalyst is still in its infancy due to challenges such as aggregation phenomena and poor conductivity. To date, several strategies have been proposed for the modification of MoS₂ composite materials. Among these, carbon supported MoS₂ (C-MoS₂) is attractive to facilitate electron transfer, resulting in greatly improved conductivity of the composite. W. H. Hu et al.²⁷ reported that the acid treated carbon nanospheres and MoS₂ composite exhibited enhanced conductivity and electrocatalytic activity for the hydrogen evolution reaction compared to MoS₂.

Moreover, various types of biomolecules, such as amino acids, nucleic acids, proteins, and peptides, have been utilized as templates for the fabrication of nanostructured materials.^28–31 Among the various biomolecules, deoxyribonucleic acid (DNA) is an attractive material to develop inorganic nanostructures because it is inexpensive, well-characterized, and shows controllable and compatible behavior. The phosphate groups and sugar molecules of DNA can bind with different metal/polymer cations through electrostatic interactions.^32–36

In this paper, mixed electronic/ionic transport in conducting polymer-metal oxide-dichalcogenide biomolecules is used as a platform to design a hybrid nanostructure, due to a host of novel devices that leverage the blend of these carriers to enrich catalytic activity for RF biosensor applications. The work discussed here addresses two aspects: (i) formation of a co-ordination bond between Fe–S of α-Fe₂O₃ and the monomer EDOT (3,4-ethylenedioxythiophene), which has a lone pair of electrons and may overlap with the vacant d-orbital of Fe; also, carbon enhances the active S edge sites for the electron transfer process; the unpaired spin density of α-Fe₂O₃ and C-MoS₂ will provide more spacing for insertion of other materials in the composite.³⁷ (ii) PEDOT/DNA and α-Fe₂O₃/DNA composites exhibit enhanced catalytic activity; however, MoS₂/DNA composite has less interaction. In order to enhance the conductive nature of MoS₂, carbon supported MoS₂ is utilized in this work. As DNA can be an effective intercalator and makes the composite less acidic than the PEDOT-PSS (poly(3,4-ethylenedioxythiophene)-polystyrene sulfonate) system, it is therefore expected that DNA doped biocomposite modified electrodes may also possess catalytic activity towards the oxidation of RF (Scheme 1).

Scheme 1 Illustration of preparation of the PEDOT–α-Fe₂O₃–C-MoS₂–DNA hybrid nanostructure and RF detection.

2. Experimental section

2.1 Reagents and materials

Sodium molybdate (Na₂MoO₄·2H₂O) and EDOT monomer were obtained from Sigma Aldrich, India. Glucose, thiourea, RF, and FeCl₃ were obtained from Sisco Research Laboratories, Mumbai. Stock 1 mM solution of RF was prepared before use and protected against light. Double-stranded herring testes DNA with an average molecular weight of 50 kbps (base pairs) is simply denoted as DNA throughout this paper. The stock DNA solution (0.06 g per 50 ml) was prepared by mixing a measured amount of DD (doubly distilled) water. Screen printed electrodes (SPE) were from Zensor Technologies, India. The aqueous solutions used throughout were prepared with ultra-pure water obtained from a Millipore system.

2.2 Instruments and measurements

The surface morphology of the samples was investigated with a scanning electron microscope (SEM, Carl Zeiss EVO 18); X-ray diffraction (X-RD) was performed using a Bruker Germany D8 Advance instrument with CuKα radiation (1.5418 Å). The Raman spectrum was recorded using an imaging spectrograph (model STR, 500 mm focal length) laser Raman spectrometer (SEKI, Japan). UV studies were performed using a Shimadzu UV-1700 spectrophotometer. Electrochemical experiments were performed using a CHI 6005D electro-chemical workstation (Austin, USA) using a GC working electrode (0.07 cm⁻²), an Ag/AgCl (3.0 M KCl) reference electrode and a platinum wire auxiliary electrode. All the measurements were carried out in PBS (phosphate buffer solution) as a supporting electrolyte under nitrogen atmosphere at room temperature.

2.3 C-MoS₂ synthesis

In a typical synthesis, 0.15 g of Na₂MoO₄·2H₂O, 0.50 g of glucose and 0.20 g NH₂CSNH₂ were dispersed in 30 ml DD water. After stirring for 30 minutes, the reaction solution was transferred into a 50 ml Teflon-lined stainless steel autoclave and heated at 240 °C for 24 h. The autoclave was left to cool naturally. The black precipitate was collected by centrifugation, washed with DD water and ethanol, and dried at 80 °C for 24 h. Afterwards, the as-prepared samples were annealed to 800 °C for 2 h to improve the conductivity of MoS₂ under the protection of argon.

2.4 α-Fe₂O₃ nanorods synthesis

Ferric chloride (1.0 g) and sodium hydroxide (NaOH) pellets (0.2 g) were sequentially added to a beaker of 100 ml DD water. Then, this solution was stirred and heated in the autoclave to 160 °C for 18 h. Finally, the heated mixture was centrifuged, washed and dried as earlier.

2.5 DNA stock solution preparation

In a typical synthesis, 10 ml of stock DNA solution (0.06 g per 50 ml) was prepared by mixing a measured amount of DD water. The solution containing DNA was stirred using a magnetic stirrer overnight, which is useful to obtain homogeneous DNA solution without any pop-up of adenine and guanine bases in the DNA.

2.6 PEDOT–α-Fe₂O₃–C-MoS₂–DNA hybrid nanostructure

200 μl EDOT monomer was dispersed in 50 ml distilled water and stirred for 30 minutes. Next, the oxidizing agent FeCl₃ was added to the above solution. Then, C-MOS₂ (0.05 g) and α-Fe₂O₃ (0.05 g) were also mixed into the above solution, which was stirred for 1 h. Finally, 2 ml of DNA from the stock solution was added and heated to 60 °C for 45 min. Then, the sample was used for various electrochemical studies.

3. Results and discussion

3.1 Structural analysis

3.1.1 Raman studies. The as-formed film exhibits a strong vibrational band centered at 1478 cm⁻¹, which corresponds to the symmetric stretching mode of the aromatic C–C band; the major bands in the range of 1350 to 1600 cm⁻¹ can be attributed to stretching vibrations of the thiophene ring, as shown in Fig. 1A. They are broad and not well-resolved, which is characteristic of polymeric films. The peaks ascribed to the C–H stretching vibrations of aromatic and methylene moieties³⁸ are usually positioned between 2850 and 3100 cm⁻¹. The C-MoS₂ spectrum exhibits two broad bands at 1270 cm⁻¹ (D-band) and 1525 cm⁻¹ (G-band), with some additional peaks of hexagonal structured MoS₂. The G and D-bands depict the in-plane vibration of sp² bonded carbon atoms and the vibrational modes from sp³ bonded carbon atoms in amorphous carbon.³⁹ The peaks at 454, 630, and 3044 cm⁻¹ confirm the presence of single-crystalline rod-shaped α-Fe₂O₃. Further, it is confirmed from the α-Fe₂O₃–PEDOT–C-MoS₂ spectra that the strong peaks of the G and D-bands of C-MoS₂ are unchanged, indicating that there is no structural change in the composite; furthermore, the new peaks confirm the composite formation. It can be concluded that the changes in peak intensity, the new peak formation and the morphological re-arrangements lead to enhancement in the catalytic activity of RF sensing.

Fig. 1 (A) Raman spectra, (B) XRD patterns of the α-Fe₂O₃–PEDOT–C-MoS₂ composite. (C) UV patterns of the α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid nanostructure.

3.1.2 UV studies. In Fig. 1B, the peak at 300 nm can be attributed to the π–π* transitions of the thiophene ring of PEDOT. The sharp peak at 217 nm obviously confirms the presence of carbon in the MoS₂.⁴⁰ The inter-band transition of hematite above approximately 640 nm confirms the presence of α-Fe₂O₃.^41,42 In the UV absorption spectrum of the α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid film, as a result of the strong π–π stacking interaction between PEDOT and DNA, a strong peak at 257 nm confirms the p–p* transition of the electrons of the C@C bond of the DNA bases.

3.1.3 X-RD studies. As expected for PEDOT, the pattern does not yield any characteristic peaks except the low angle peak at ∼25, indicating the amorphous nature of the polymeric material, as shown in Fig. 1B.^43,44 The rods of α-Fe₂O₃ have a rhombohedral structure that matches well to iron oxide; no other hydroxide, maghemite, or magnetite peaks are observed in the hematite phase.⁴⁵ From the C-MoS₂ pattern, the peak shift confirms the role of carbon in inducing interlayer distances in its structure compared with bulk MoS₂.⁴⁶ As shown in the α-Fe₂O₃–PEDOT–C-MoS₂ sample, a sharpened intensity of the peaks can be observed due to the interaction and composite formation. It is also confirmed that the strong interaction between PEDOT and C-MoS₂ is probably due to hydrogen bonding; PEDOT interleaves between the layers of MoS₂ using the soft chemistry route of intercalation, which in turn significantly enhances the conductivity. Similarly, the presence of ferrite particles in the hybrid matrix is also confirmed.

3.2 Morphology analysis

The SEM image in Fig. 2A reveals the well dispersed regular and desirable rod shapes of Fe₂O₃ with crystalline morphology; the average particle size remains 96.5 nm. Fig. 2B shows the C-MoS₂ nanosheets with slightly folded dentations on the edges which considerably enhance the exposure of the active edge sites.⁴⁷ This is due to the presence of carbon, and it is used to decentralize the MoS₂ nanosheets. Fig. 2C depicts the nanospheres of PEDOT; their size is quite uniform (∼36.5 nm). The presence of α-Fe₂O₃ and PEDOT embedded on the C-MoS₂ sheets is seen in Fig. 2D. The DNA wrapped on the α-Fe₂O₃–PEDOT–C-MoS₂ composite is exhibited in Fig. 2E. The size enhancement after coating the α-Fe₂O₃–PEDOT–C-MoS₂ composite with DNA further confirms the presence of DNA; the sizes of the individual particles increased to approximately 222 nm. The EDS measurement also reports their presence in the hybrid nanostructure (Fig. 2F).

Fig. 2 SEM images of (A) α-Fe₂O₃ nanorods, (B) C-MoS₂ nanosheets, (C) PEDOT nanospheres, (D) α-Fe₂O₃–PEDOT–C-MoS₂ composite, (E) α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid nanostructure, (F) EDS spectrum of hybrid nanostructure.

3.3 Electrochemical characterization

3.3.1 CV behavior of the α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid structure in [Fe(CN)₆]^3−/4−. The formation of the α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid structure was characterized by CV in the presence of 1 mM [Fe(CN)₆]^3−/4− in 0.1 M KCl at a scan rate of 50 mV s⁻¹, as shown in Fig. 3. As can be seen in Fig. 3a, at the bare GCE, a pair of well-defined redox peaks is observed, with a peak to peak separation of 72 mV. Deposition of the C-MoS₂ onto the electrode surface decreases the reversibility of [Fe(CN)₆]^3−/4− due to the its barrier properties, whereas the α-Fe₂O₃ deposited electrode (curve c) increases the reversibility. After being coated with PEDOT, the electrode displays a strongly enhanced current value. Interestingly, α-Fe₂O₃–PEDOT–C-MoS₂ composite remarkably enhances the reversible nature due to the electron transfer kinetics towards [Fe(CN)₆]^3−/4−]. This is supported by earlier reports which state that PEDOT/MoS₂ composites have faster charge transfer rates in which C-MoS₂ is favorable for rapid diffusion of electrons by providing a lower resistance pathway than pristine MoS₂.⁴⁸ However, the α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid coated electrode, surprisingly, exhibited a decreased current value which is close to curve b. This is due to electrostatic repulsion between the negatively charged DNA modified composite and [Fe(CN)₆]^3−/4−.

Fig. 3 (A) CV and (B) EIS profiles: (a) bare, (b) C-MoS₂, (c) α-Fe₂O₃, (d) PEDOT, (e) α-Fe₂O₃–PEDOT–C-MoS₂, (f) α-Fe₂O₃–PEDOT–C-MoS₂–DNA modified electrodes at 50 mV s⁻¹ in 0.1 M KCl and 1 mM [Fe(CN)₆]^3−/4−.

The impedance behavior of the modified electrode recorded in the frequency region of 100 kHz to 1 Hz at an applied DC potential of 230 mV and an amplitude of ±5 mV is shown in Fig. 3B. The values of the charge transfer resistances (R_CT) were determined from the Randles equivalent circuit. The R_CT values for the α-Fe₂O₃–PEDOT–C-MoS₂ composite, α-Fe₂O₃ bare GC, C-MoS₂, PEDOT and α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid modified electrodes have been estimated as 117, 193, 229, 5341, and 8986 Ω, respectively. The obtained impedance plots were in good agreement with the CV measurements (Fig. 3A).

3.3.2 Electrocatalytic oxidation of RF. Cyclic voltammetry (CV) was performed in the potential range of −0.7 to −0.2 V in GCE in the presence of 0.1 M PBS at pH 7.500 μM RF at 50 mV s⁻¹ was injected into PBS. Then, the GC electrode was separately modified with (a) bare (b) C-MoS₂ (c) α-Fe₂O₃ (d) PEDOT (e) α-Fe₂O₃–PEDOT–C-MoS₂ (f) α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid nanostructured samples by a drop casting technique; the electrodes were allowed to dry for 1 h and then placed into the electrochemical cell to study their current values (Fig. 4A), which is widely used for the generation of electrochemical signals. The CV of a bare GCE exhibited cathodic peak current I_pc is 5.8 × 10⁻⁶ A, indicating a reversible one electron redox process of RF. With the MoS₂ modified GCE, an increased cathodic peak current is seen, with a small potential shift from −0.388 V to −0.417 V. The attachment of α-Fe₂O₃ resulted in 7.7 × 10⁻⁶ A due to its catalytic behavior (E_pc = −0.402 V), while the PEDOT modified electrode shows 1.7 × 10⁻⁵ A. Interestingly, the α-Fe₂O₃–PEDOT–MoS₂ modified electrode depicts a maximum current of 5.6 × 10⁻⁵ A due to the electrocatalytic activity of materials present in the composite, whereas the α-Fe₂O₃–PEDOT–MoS₂–DNA hybrid nanostructure modified electrode reduces the current to 5.5 × 10⁻⁶ A owing to electrostatic repulsion between C-MoS₂, DNA and RF. However, the lowest oxidation potential at −0.384 V confirms the catalytic behavior of the modified electrode.

Fig. 4 CV: (A) (a) bare, (b) C-MoS₂, (c) α-Fe₂O₃, (d) PEDOT, (e) α-Fe₂O₃–PEDOT–C-MoS₂ composite, (f) α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid in 500 μM RF at 50 mV s⁻¹, (B) pH versus current.

Since electrons/protons participate in the redox chemistry of biomolecules, the redox peak current and potential are pH dependent. As shown in Fig. 4B, the α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid nanostructure system generated the maximal catalytic current at pH 7 and gradually decreased its catalytic activity when the pH was increased to neutral and basic values.

In comparison, a linear relationship of I_pc with scan rate^1/2 was observed (Fig. 5A–C), suggesting that RF was electrocatalytically oxidized at α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE, α-Fe₂O₃–PEDOT–C-MoS₂/GCE and α-Fe₂O₃–PEDOT–C-MoS₂–DNA/SPE, respectively. This report confirms that all these electrode processes are diffusion-controlled.

Fig. 5 (A) CVs obtained for RF 100 μM: (A) α-Fe₂O₃–PEDOT–C-MoS₂/GCE, (B) α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE, (C) α-Fe₂O₃–PEDOT–C-MoS₂–DNA/SPE modified electrode recorded in PBS (pH 7.0) at different scan rates of 10 to 100 mV s⁻¹.

Fig. S2† shows the linear sweep voltammogram (LSV) obtained for RF sensing with different concentration ranges at α-Fe₂O₃–PEDOT–C-MoS₂/GCE and α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE. The oxidation current of RF increased with a slight potential shift with increasing scan rate upon each increment of analyte. The oxidation currents had linear relationships with the concentration of RF, with correlation coefficients of 0.9849 and 9890, respectively (inset of Fig. S2†).

3.4 Square wave voltammetry on RF determination

On α-Fe₂O₃–PEDOT–C-MoS₂/GCE, the SWV (Square Wave Voltammetry) oxidation peak potential of RF was at −0.486 V, and the oxidation peak currents of the analyte increased linearly from 2.890 × 10⁻⁴ A to 4.476 × 10⁻⁴ A with increasing concentration. Linear responses for RF determination were observed in the concentration range of 800 nM to 700 μM with a detection limit of 79 nM (S/N = signal/noise = 3σ/b, σ – standard deviation and b slope of the linear fit, Fig. 6). The greater gap of the interlayer distance in C-MoS₂ compared with bulk MoS₂ can provide desirable channels for electron transfer with reduced diffusion barriers.⁴⁹ Meanwhile, on α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE, the peak RF potential was at −0.458 V and the oxidation peak currents were from 2.841 × 10⁻⁶ A to 4.482 × 10⁻⁵ A with increased concentration. The linear responses for this modified electrode were in the range of 300 nM to 1 mM, with a detection limit of 5 nM (Fig. 7). Two points have been noted from the above discussion: (i) the DNA modified electrode reduces the oxidation potential of RF by 30 mV, clearly showing electrocatalytic behavior. (ii) However, the oxidation peak current values decreased in the DNA modified electrode by 2 orders on the lower concentration side and one order in the higher concentration side in comparison with the modified electrode without DNA. Interestingly, the interaction between ferrites and phosphate present in DNA in the composite provides excellent electrocatalytic activity towards the oxidation of RF.⁵⁰ On α-Fe₂O₃–PEDOT–C-MoS₂–DNA/SPE, the SWV oxidation peak potential of RF is at −0.588 V and linear responses for the determination of RF were observed in the concentration range of 100 nM to 1 mM, with a detection limit of 12 nM (Fig. 8). The oxidation peak current values were from 2.267 × 10⁻⁶ A to 6.201 × 10⁻⁵ A. It is clear from the above 3 SWV results that (i) the DNA modified electrode detects a wider range than the other electrodes; (ii) even though the electroactive surface area (8.15 ×10⁻⁶ cm⁻²) of the α-Fe₂O₃–PEDOT–C-MoS₂/GCE is higher, the sensitivity is not satisfactory, whereas the α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE (2.11 × 10⁻⁵ cm⁻²) and α-Fe₂O₃–PEDOT–C-MoS₂–DNA/SPE (5.27 × 10⁻⁵ cm⁻²) possess comparatively less area, but exhibit more sensitivity and wide detection ranges. It is believed that the DNA in the hybrid nanostructure oxidizes the RF by a lower potential of 30 mV, which is responsible for the enhanced detection range. The corresponding detection limits and response ranges of different modified electrodes are listed (Table S4†) for comparison. Therefore, this hybrid sample, as an advanced electrode material, may have promising applications in electrochemical detection. The following statements suggest reasons for the reduced conductivity of the Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid material:

Fig. 6 SWV profiles: (A) 800 nM to 700 μM RF. (B) Plot of the oxidation peak current against the concentration of RF at α-Fe₂O₃–PEDOT–C-MoS₂ composite/GCE in 0.1 M PBS (pH 7.0).

Fig. 7 SWV profiles: (A) 300 nM to 1 mM RF. (B) Plot of the oxidation peak current against the concentration of RF at α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid nanostructure/GCE in 0.1 M PBS (pH 7.0).

Fig. 8 SWV profiles: (A) 100 nM to 1 mM RF. (B) Plot of the oxidation peak current against the concentration of RF at α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid nanostructure/SPE in 0.1 M PBS (pH 7.0).

(i) In DNA, the nucleobases are buried between the densely negatively charged phosphate backbones, and the DNA stayed away from the basal plane of C-MoS₂, resulting in weak interaction.⁵¹

(ii) The affinity of MoS₂ toward ds-DNA is lower than toward ss-DNA.⁵²

(iii) Strong interactions between DNA and RF could be responsible for possible damage or shielding of the oxidizable groups of guanine and adenine bases present in DNA.⁵³

However, a strong interaction/fast formation of PEDOT and DNA in the hybrid due to electrostatic attraction between the phosphate groups of DNA and PEDOT may result in the enhanced electrocatalytic activity.⁵⁴ Moreover, it is believed that in the PEDOT–DNA system, the increased conductivity is due to the modulation ability of the DNA conformation in the redox process, which also supports the enhanced catalytic behavior. Hence, the design of this scaffold with DNA could be used for the fabrication of a nanostructured catalyst for biosensing applications.

3.5 Reproducibility and stability of sensors

The reproducibility of the α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE was investigated by the SWV responses of RF. The relative standard deviation (RSD) responses for six independent electrodes were 5.4%. The fabricated sensors with α-Fe₂O₃–PEDOT–C-MoS₂/GCE, α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE and α-Fe₂O₃–PEDOT–C-MoS₂–DNA/SPE were subjected to 100 continuous cycles in the presence of 1 mM RF (Fig. S3†). The long-term stability of the electrode stored at 4 °C was measured after one month of storage in dry conditions. The electrode retained 88% of its initial response. These results demonstrate that the modified electrodes possessed satisfactory reproducibility and tremendous stability for analytical applications.

3.6 Practical applications

The anti-interference ability of the α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE was examined by the addition of several types of ions and other small molecules to PBS solution containing 100 μM RF (Fig. S4 and Table S1†). No interference was observed in the presence of 10 fold excesses of ascorbic acid, KCl, FeCl₃, folic acid, MgCl₃, uric acid and tyrosine. It is clear that the proposed sensor exhibited good selectivity for the determination of RF. The utilization of the α-Fe₂O₃–PEDOT–C-MoS₂–DNA/GCE sensor in real samples was also studied by the standard addition method in human urine, milk powder and pharmaceutical drug samples. All the details of the real sample analysis results are summarized in Tables S2 and S3.†

The urine sample was diluted 10 times with 0.1 M PBS solution (pH 7.0) without any other treatment before measurement. While adding the diluted sample, a sharp peak appeared for RF at 0.42 V for 10 μM concentration. For different concentrations (10 to 80 μM) of urine sample, the corresponding RF peaks were recorded and are shown in Fig. S5(A).† The same experiment was performed with milk powder, and a peak appeared at 0.48 V for 100 μM. The responses for different concentrations (100 μM to 1 mM) are shown in Fig. S5(B).† Finally, the RF oxidation of the pharmaceutical drug was analyzed at 0.41 V for 100 μM. The square wave studies for different concentrations of the drug (10 μM to 100 μM) are depicted in Fig. S5(C).†

4. Conclusions

A simple chemical route has been reported for the synthesis of α-Fe₂O₃–PEDOT–C-MoS₂–DNA hybrid nanostructures. The fabricated α-Fe₂O₃–PEDOT–C-MoS₂–DNA modified electrodes, which integrate the unique behavior of the components (α-Fe₂O₃ has enzymatic activity, C-MoS₂ mimics the activity of graphene, and DNA has catalytic activity), provided superior currents for the analyte RF compared to those of α-Fe₂O₃–PEDOT–C-MoS₂/GCE. It is demonstrated that the DNA modified composite exhibits a significantly low detection limit and high electrocatalytic activity. Additionally, based on its electrochemical performance, the DNA modified hybrid structure may be a platform with high potential in biomedical and non-corrosive device fabrication.

Acknowledgements

The author J. W. gratefully acknowledges the University Grant Commission (MRP-MAJOR-ELEC-2013-37628) for financial assistance.

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Footnote

† Electronic supplementary information (ESI) available. See DOI: 10.1039/c6ra16279g

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