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Nanostructured fe2 o3 platform for the electrochemical sensing of folic acid

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Nanostructured fe2 o3 platform for the electrochemical sensing of folic acid

Nanostructured fe2 o3 platform for the electrochemical sensing of folic acid

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  • 1. Nanostructured a-Fe2O3 platform for the electrochemical sensing of folic acid† Thandavarayan Maiyalagan,‡a J. Sundaramurthy,‡bc P. Suresh Kumar,bc Palanisamy Kannan,*d Marcin Opallo*d and Seeram Ramakrishna*bc a-Fe2O3 nanofibers are synthesized by a simple and efficient electrospinning method and the selective determination of folic acid (FA) is demonstrated in the presence of an important physiological interferent, ascorbic acid (AA), using the a-Fe2O3 nanofiber modified glassy carbon (GC) electrode at physiological pH. Bare GC electrode fails to determine the concentration of FA in the presence of a higher concentration of AA due to the surface fouling caused by the oxidized products of AA and FA. However, modification with a-Fe2O3 nanofibers not only separates the voltammetric signals of AA and FA by 420 mV between AA and FA, but also enhances higher oxidation current. The amperometric current response is linearly dependent on FA concentration in the range of 60–60 000 nM, and the a-Fe2O3 nanofiber modified electrode displayed an excellent sensitivity for FA detection with an experimental detection limit of 60 nM (1.12 Â 10À10 M (S/N ¼ 3)). Furthermore, the a-Fe2O3 nanofiber modified electrode showed an admirable selectivity towards the determination of FA even in the presence of a 1000-fold excess of AA and other common interferents. This modified electrode has been successfully applied for determination of FA in human blood serum samples. 1 Introduction In recent years, the synthesis and fabrication of nanomaterials with tailoring their size, morphology, and porosity have been intensively pursued not only for fundamental scientic interest but also for many technological applications.1–3 Nanoparticles (zero-dimensional (0-D)) and nanowires/nanorods (one-dimen- sional (1-D)) with controlled size and shape are of key importance because their electrical, optical, and magnetic properties are strongly dependent on their size and shape.1–3 Currently, one- dimensional (1-D) nanomaterials such as silicon nanowires (SiNWs), carbon nanotubes (CNTs), and conducting polymer nanowires (CP NWs) have opened the possibility to fabricate electrochemical sensors and biosensors.4–7 Their high sensitivity and new sensing mechanisms are related to intrinsic properties associated with a high surface-to-volume ratio.4–7 Further, 1-D nanostructures can be used for both efficient transport of elec- trons and optical excitation, and these two factors make them critical to the function and integration of nanoscale devices, and have been the focus of intensive research for many potential applications in electronics, photonics, drug delivery, medical diagnostics, and magnetic materials.8–11 Hematite (a-Fe2O3) is the most stable iron oxide with n-type semiconducting properties (Eg ¼ 2.2 eV) under ambient condi- tions. It has been intensively investigated because of its wide applications in catalysts, pigments, magnetic materials, gas sensors, and lithium ion batteries.4,12–15 Fe2O3 was generally considered to be biologically and electrochemically inert, and its electrocatalytic functionality has been rarely realized directly in the past,16 whereas Fe2+ ions (instead of Fe3+ ) play the dominant role in the oxidation reaction.17–19 Meanwhile, Fe2O3 was also demonstrated to show both reversible reduction and reversible oxidation of Fe(III) in a basic carbonate buffer solution.17 Never- theless, in contrast with interests focusing on synthetic and catalytic applications of Fe3O4, reports on the electrochemical characterization of Fe2O3 nanoparticles are rather rare, and little attention has been paid to the detailed study of their sensing performance.20–22 In principle, Fe2O3 nanoparticles may effi- ciently mediate the nal heterogeneous chemical oxidation or reduction of the target agent, while the converted iron oxides can be continuously and simultaneously recovered by electro- chemical oxidation or reduction due to their high surface to volume ratio. From this key point, an electrocatalytic study of nanostructured Fe2O3 in biocompatible environments may not a School of Chemical and Biomedical Engineering, Nanyang Technological University, 62 Nanyang Drive, Singapore, 637459s b Centre for Nanobers and Nanotechnology, National University of Singapore, Singapore 117576. E-mail: seeram@nus.edu.sg; Fax: +65-6872 5563; Tel: +65-6516 6593 c Department of Mechanical Engineering, National University of Singapore, Singapore 117576 d Institute of Physical Chemistry, Polish Academy of Sciences, 44/52 ul. Kasprzaka, 01- 224 Warszawa, Poland. E-mail: ktpkannan@gmail.com; mopallo@ichf.edu.pl; Tel: +48-223 433 375. Fax: +48-223 433 333 † Electronic supplementary information (ESI) available. See DOI: 10.1039/c3an00070b ‡ Both authors contributed equally to this work. Cite this: Analyst, 2013, 138, 1779 Received 24th November 2012 Accepted 17th January 2013 DOI: 10.1039/c3an00070b www.rsc.org/analyst This journal is ª The Royal Society of Chemistry 2013 Analyst, 2013, 138, 1779–1786 | 1779 Analyst PAPER DownloadedbyUniversityofTexasLibrarieson22February2013 Publishedon31January2013onhttp://pubs.rsc.org|doi:10.1039/C3AN00070B View Article Online View Journal | View Issue
  • 2. only be of scientic interest, but could also produce real benets such as their substitution for noble metals/enzymes for practical enzyme-free biosensor applications. Due to its low cost, good stability, and reversibility, a-Fe2O3 has been proven to be an important semiconductor nanomaterial for electrochemical sensors.23 However, so far, there have been only a few reports on bio-sensing properties of 1-D nanostructural a-Fe2O3. Generally, the properties of a biosensor are strongly dependent on its surface area. The relatively low surface to volume ratio of conventional bulk a-Fe2O3 materials leads to their poor bio- sensing properties. Hence, developing 1-D nanostructure a-Fe2O3 with a high surface area is very important for increasing their applications in sensors.4,14 Recently, as prepared a-Fe2O3 nano- material has been proven to be a successful electrode material due to the intrinsic peroxidase-like catalytic activity.24 The a-Fe2O3 nanowire array modied glucose sensor exhibited an excellent biocatalytic performance towards the oxidation of glucose with a detection limit of 6 mM (S/N ¼ 3).24 In the present study, we have synthesized a-Fe2O3 nanober by a simple elec- trospinning method and used it to improve the bio-sensing performance towards the oxidation of FA for the rst time. Folic acid, known as a widely used water soluble vitamin, is reported to be a very signicant component for human health which relates to a series of diseases such as gigantocytic anemia, leucopoenia, mental devolution, heart attack and congenital malformation.25–28 FA is one of the important coenzymes of the haematopoietic system that controls the generation of ferro- haeme.28 The dosage of FA is associated with the treatments of hyperhomocysteinemic coronary artery disease, hypertension, depression, hypercholesterolemia, mammary tumor, vascular disease and neural tube defects of pregnant women.29–33 The important biomolecules such as AA and FA are present in human blood plasma,34,35 urine36,37 and blood serum samples.38,39 Since these biomolecules coexist in human uids, their simultaneous determination is essential to secure the human health from the above critical diseases risk. Therefore, the selective and sensitive determination of FA is very important from the clinical and health viewpoints. In this paper, we will show that the a-Fe2O3 nanober modied GC electrode exhibits an excellent electro- catalytic activity towards FA, and a detection limit of 60 nM FA has been achieved using the amperometry method. The a-Fe2O3 nanober array on the GC electrode with a nano-size and coarse surface provides a platform for FA oxidation by contributing both excess electroactive sites and strong adhesion to the GC electrode surface, which results in the enhanced sensitivity and long term stability of the a-Fe2O3 electrode. The application of the Fe2O3 modied electrode has been successfully demonstrated by measuring the concentration of FA in real samples. 2 Experimental section 2.1 Materials and methods Polyvinylpyrrolidone (PVP; MW ¼ 1 40 000) and iron(III) acety- lacetonate (Fe(acac)3) were purchased from Sigma-Aldrich and Fluka, Singapore, respectively. Ethanol (HPLC grade) and glacial acetic acid were purchased from Tedia, Singapore and used as received. The biomolecules, uric acid (UA) and folic acid (FA), were purchased from Merck chemicals and were used as received. All other chemicals used in this investigation were of analytical grade. The phosphate buffer solution (PBS; pH ¼ 7.2) was prepared using Na2HPO4 and NaH2PO4. Double distilled water was used to prepare the solutions in this investigation. 2.2 Preparation of a-Fe2O3 nanobers Firstly, 1 g of PVP was dissolved in 10 mL of ethanol solution and homogeneously stirred at room temperature for 1 h for complete dissolution. Then, 0.4 g of Fe(acac)3 precursor was added to the PVP solution and continuously stirred for 6 h followed by addition of 1 mL acetic acid. Finally, 5 mL of Fe(acac)3–PVP precursor solutions were loaded in a 5 mL plastic syringe with a hypodermic needle (dia. 27 G). The hypodermic needle was then connected to a high-voltage supply capable of generating direct current (DC) voltages up to 30 kV. Electro- spinning was carried out by applying a power supply of around 16.5 kV at the needle in a controlled electrospinning set-up (Electrospunra, Singapore). Aluminum foil was used as the counter electrode, and the distance between the needle and the collector was maintained at 15 cm. The as-spun Fe(acac)3–PVP composite nanober mats were placed in an advanced vacuum oven at room temperature for 12 h to remove the solvent residuals. Finally, the nanobers were calcined at 500  C for 5 h in air at a heating rate of 5  C minÀ1 , and nally a-Fe2O3 nanobers were obtained and stored carefully. 2.3 Instrumentation The crystallographic information of the prepared a-Fe2O3 nanobers was studied using the powder X-ray diffraction technique (XRD, Shimadzu XRD-6000, Cu Ka radiation oper- ating at 30 kV/40 mA). The surface morphologies of the nano- structures were characterized using a eld emission-scanning electron microscope (FE-SEM) JEOL JSM-6301F. Transmission electron microscopy (TEM), JEM-2010, JEOL USA Inc., was employed to study the surface morphology of a-Fe2O3 nano- bers. The electron beam accelerating voltage of the microscope was at 200 kV. Electrochemical measurements were performed in a conventional two compartment three electrode cell with a mirror polished 3 mm glassy carbon (GC) as the working elec- trode, Pt wire as the counter electrode and a NaCl saturated Ag/AgCl as the reference electrode. The electrochemical measurements were carried out with a CHI Model 660C (Austin, TX, USA) electrochemical workstation. In cyclic voltammetry, the electrochemical oxidations of AA and FA were carried out at a scan rate 50 mV sÀ1 . Pulse width ¼ 0.06 s, amplitude ¼ 0.05 V, sample period ¼ 0.02 s and pulse period ¼ 0.20 s were used in differential pulse voltammetry (DPV). For chronoamperometric measurements, pulse width ¼ 0.25 s and potential step ¼ 1 mV were used. All the electrochemical measurements were carried out under a nitrogen atmosphere at room temperature (27  C). 2.4 a-Fe2O3 nanober modied electrodes The a-Fe2O3 nanostructure modied GC electrode was prepared as follows. First, the surface of the glassy carbon electrode for each experiment was mechanically polished with 600 grit 1780 | Analyst, 2013, 138, 1779–1786 This journal is ª The Royal Society of Chemistry 2013 Analyst Paper DownloadedbyUniversityofTexasLibrarieson22February2013 Publishedon31January2013onhttp://pubs.rsc.org|doi:10.1039/C3AN00070B View Article Online
  • 3. sand-paper and 0.050 mm alumina powders, which was then rinsed with acetone and double distilled water. A 3 mL aliquot of a-Fe2O3 nanobers (dispersed in water, 5 mg mLÀ1 , and pH ¼ 7.0) was dropped onto the surface and dried under atmospheric conditions. In addition, a bulk-Fe2O3 electrode (denoted as b-Fe2O3 GC electrode, fabricated by the above mentioned process) was used for comparison. 3 Results and discussion 3.1 Characterization of the as-prepared a-Fe2O3 nanostructure The morphology of as-electrospun Fe(acac)3–PVP composite nanober before and aer calcination was investigated by per- forming FESEM analysis. Fig. 1a shows the formation of a highly interconnected network of nanobers with an average ber diameter of 288 nm. Fig. 1a (inset) shows the distribution of bers upon applying a potential of 16.5 kV; the broad distribution of the bers was due to the dominancy of Coulombic repulsive forces upon applying such a higher potential. The formation of nanobers was observed aer calcination of the Fe(acac)3–PVP composite at 500  C for 5 h at a rate of 5  C minÀ1 in air (Fig. 1b). The novel morphology of nanostructures with ellipsoidal shape of a-Fe2O3 nanoparticles uniformly plaited along the ber directions was observed. This morphology was due to the combined effect of phase separation (thermodynamic) and electrospinning (electro-hydrody- namic).40 The phase separation of the polymer and the precursor induced the formation of precursor islands, and the electrospinning coerced the precursors to plait together with the result of spinning and whirling effects upon applying potential. During calcining, the polymer PVP present all over the brous structure was decomposed and yielded nanorod-like structures. The nanobers have a coarse surface due to the adsorption and assembly of small crystalline nanoparticles (Fig. 2b; inset), some of which even have a chain-like morphology. In comparison with the randomly packed particle counterpart, such arrayed nanowires provide more ordered spatial orientation and improved structural stability. As a result, higher mass transfer and permeation rate, a stable porous volume and less structural corruption can be expected during the electrochemical recycling. In addition, the coarse surface may also result in enhanced long-term stability due to a more secure attachment to the electrode surface. Further, the XRD analysis on calcined nanobers has been carried out to conrm the a-Fe2O3 phase formation. Fig. 2 shows the XRD pattern of a-Fe2O3 nanobers aer calcination of the Fe(acac)3–PVP composite at 500  C for 5 h in air. All the diffraction peaks were well indexed to the rhombohedral hexagonal phase of hematite (a-Fe2O3) (JCPDS: 33-0664). The strong and narrow-sharp diffraction peaks showed the purity and high degree of crystallization of synthesized a-Fe2O3 nanobers. 3.2 Electrochemical oxidation of FA We have examined the electrocatalytic activity of a-Fe2O3 GC, b- Fe2O3 GC and unmodied GC electrodes towards the oxidation of FA. We found that the a-Fe2O3 nanober modied GC electrode showed higher electrocatalytic activity towards AA and FA than the b-Fe2O3 GC and unmodied GC electrodes. Fig. 3A shows the cyclic voltammograms (CVs) obtained for 0.25 mM FA at bare and a-Fe2O3 nanober modied GC electrodes in a 0.20 M phosphate Fig. 1 (a) FE-SEM image of as-electrospun Fe(acac)3–PVP composite nanofibers at a power supply of 16.5 kV (inset: histogram of the nanofiber diameter and distribution) and (b) FE-SEM image of a-Fe2O3 nanofibers after calcining composites at 500  C for 5 h at a ramp rate of 5  C minÀ1 . Fig. 2 XRD pattern of Fe(acac)3–PVP nanofibers calcined at 500  C for 5 h at a heating rate of 5  C minÀ1 in air. Fig. 3 (A) CVs obtained for 0.25 mM FA at bare and a-Fe2O3 nanofiber modified GC electrodes after the 1st (a and d), 10th (b and e) and 20th (c and f) cycles in a 0.2 M PB solution at a scan rate of 50 mV sÀ1 and (g) CV obtained for the a-Fe2O3 nanofiber modified GC electrode in the absence of 0.5 mM FA in a 0.2 M PB solution. Inset: bulk-Fe2O3 modified GC electrode in the presence of 0.25 mM FA in 0.2 M PB solution. (B) Anodic peak current vs. square root of scan rates. This journal is ª The Royal Society of Chemistry 2013 Analyst, 2013, 138, 1779–1786 | 1781 Paper Analyst DownloadedbyUniversityofTexasLibrarieson22February2013 Publishedon31January2013onhttp://pubs.rsc.org|doi:10.1039/C3AN00070B View Article Online
  • 4. buffer (PB) solution (pH ¼ 7.2). At the bare GC electrode, an oxidation peak was observed for FA at 0.95 V in the rst cycle (curve a). In the subsequent cycles, the FA oxidation peak was shied to more positive potential with decreased peak current. Aer 20 cycles, the oxidation peak of FA almost disappeared (curve c), indicating that the bare GC electrode was not suitable for the stable and simultaneous determination of FA. The adsorption of the oxidized product of FA on the electrode surface is the possible reason for the decreased FA oxidation current and more positive peak shi in the oxidation potential at the bare GC electrode. On the other hand, a well-dened oxidation peak was observed at 0.81 V for FA at the a-Fe2O3 nanober modied GC electrode (curve d), which was a 140 mV less positive potential than at the bare GC electrode. It can be seen from Fig. 3A that the oxidation potential of FA remained stable even aer 20 repeated potential cycles (curve f), indicating that the oxidation of FA was highly stable at the a-Fe2O3 nanober modied GC electrode. The a-Fe2O3 nanober modied GC electrode did not show any oxidation response in the absence of FA (curve g). These results indicated that the a-Fe2O3 nanobers are excellent candidates toward the electrochemical oxidation of FA. For comparison, we have also modied the electrode with bulk Fe2O3 (b-Fe2O3 GC) as an electrocatalyst, and we observed that the electrochemical oxidation of FA is almost the same as the electrochemical response of the bare GC electrode (Fig. 3A; inset). Unlike bare GC and b-Fe2O3 modied GC electrodes, the FA oxidation peak is highly stable at the a-Fe2O3 nanober modied GC electrode. This indicated that a-Fe2O3 nanobers effectively prevent the fouling caused by the oxidized products of FA. The observed oxidation peak for FA in Fig. 3A is due to the two electron oxidation of FA to dehydrofolic acid,41 as shown in Scheme 1. The oxidation process can be deduced through an electrocatalytic mechanism involving the Fe(III)/Fe(II) ion centers, and the catalytic mechanism of the a-Fe2O3 to folic acid oxidation can be explained by the following scheme; the vol- tammetric response of FA at the a-Fe2O3 electrode is due to two steps, viz., an electrochemical process followed by a chemical reaction. In the rst step, Fe(II) was electrochemically oxidized to Fe(III) (eqn (1)) and in the second step FA was chemically oxidized to dehydrofolic acid by Fe(III) (eqn (2)). 2Fe(II) / 2Fe(III) + 2eÀ (1) 2Fe(III) + folic acid / 2Fe(II) + dehydrofolic acid + H2O (2) The a-Fe2O3 nanober does not show any oxidation peak in the absence of FA (curve e). Further, in order to understand the fast electron transfer reaction of FA at the a-Fe2O3 nanober modied GC electrode quantitatively, we have calculated the standard heterogeneous rate constant (ks) for FA at a-Fe2O3 nanobers and bare GC electrodes using the Velasco equation42 as given below: ks ¼ 1.11Do 1/2 (Ep À Ep/2)À1/2 n1/2 where, ks is the standard heterogeneous rate constant; Do is the apparent diffusion coefficient; Ep is the oxidation peak poten- tial; Ep/2 is the half-wave oxidation peak potential and n is the scan rate. In order to determine ks, it is necessary to nd the diffusion coefficient for FA. The apparent diffusion coefficient (Do) value was determined using a single potential chro- noamperometry technique based on the Cottrell slope obtained by plotting current versus 1/Otime. Chronoamperometry measurements were carried out for FA both at bare and a-Fe2O3 nanober modied GC electrodes aer 20 potential cycles. The Do of 1.98 Â 10À6 cm2 sÀ1 was obtained for FA. The estimated ks values for the oxidation of FA at bare and a-Fe2O3 nanober modied GC electrodes were found to be 1.43 Â 10À5 cm sÀ1 and 2.91 Â 10À4 cm sÀ1 , respectively. The obtained higher ks value for FA at the a-Fe2O3 nanober modied GC electrode indicated that the oxidation of FA was faster at the a-Fe2O3 nanober modied GC electrode than at the bare GC electrode. Further, we have investigated whether the oxidation of FA at the a-Fe2O3 nanober modied GC electrode is due to diffusion control or adsorbed species by varying the scan rates. The oxidation current of FA was increased while increasing the scan rates (Fig. 3B). A good linearity between the anodic peak current and the square root of the scan rate was obtained within the range from 100 to 1000 mV sÀ1 with a correlation coefficient of 0.995 for FA, as shown in the inset of Fig. 3B. This indicated that the electrode reaction process was controlled by the diffusion of FA. Further, we have studied the optimization of pH for the present FA sensor. ESI, Fig. S1A,† shows the DPVs obtained for 100 mM FA at the a-Fe2O3 nanober modied GC electrode from pH 5.2–10.2 PB solution. It can be clearly visualized that as the pH value increases, the Epa of FA shis towards negative potential, which conrms that during electrochemical oxida- tion of FA not only electrons but also protons are involved. The plot of Epa vs. pH shows good linearity in the pH range of 5.2– 10.2. The linear regression equation of Epa/V ($0.032 V) vs. pH was obtained with a correlation coefficient r ¼ 0.990, indicating that the number of protons and electrons involved is equal. Fig. S1B† also reveals that the Ipa increases with an increase in pH up to 7.2, and a further increase of pH results in the decrease of the anodic peak current. Since the present modied electrode shows a higher current for FA at pH 7.2 and it is also close to the physiological pH value, we have chosen pH 7.2 for the deter- mination of FA in this work. 3.3 Selective determination of FA in the presence of AA Further, we have investigated the determination of FA in the presence of very high concentrations of AA. It is well known that AA is an important interferent compound which coexists with FA in our body uids, and further its concentration is always Scheme 1 Electrochemical oxidation of FA at the a-Fe2O3 nanofiber modified GC electrode. 1782 | Analyst, 2013, 138, 1779–1786 This journal is ª The Royal Society of Chemistry 2013 Analyst Paper DownloadedbyUniversityofTexasLibrarieson22February2013 Publishedon31January2013onhttp://pubs.rsc.org|doi:10.1039/C3AN00070B View Article Online
  • 5. much higher than that of FA.43 For example, the concentrations of AA and FA in human blood serum are 53.8 Æ 36.6 mmol LÀ1 , and 34.4 Æ 10.4 nmol LÀ1 , respectively.43–45 Therefore, from a clinical point of view, the determination of FA in the presence of higher concentrations of AA is very important. Fig. 4 shows the DPVs obtained for the increment of 10 mM FA in the presence of 2500 mM AA. The concentration of FA was varied from 10 to 50 mM (curves b–f). A very clear signal was observed for 10 mM FA in the presence of 2500 mM AA in Fig. 4 (curve b), which revealed that detection of a very low concentration of FA is possible even in the presence of 250-fold AA. On increment of 10 mM FA to a PB solution containing 2500 mM AA, the oxidation current of FA was increased linearly with a correlation coefficient of 0.9995. However, the oxidation peak current of AA was almost unchanged in each addition of FA. These results demonstrated that the a-Fe2O3 nanober modied GC electrode is more selective towards FA even in the presence of very high concen- trations of AA. 3.4 Amperometric determination of FA along with AA The amperometric method was used to examine the sensitivity of the a-Fe2O3 nanober modied GC electrode towards the detection of FA individually and also along with AA. Fig. 5A shows the amperometric i–t curve for FA at the a-Fe2O3 nano- ber modied GC electrode in a homogeneously stirred 0.20 M PB solution by applying a potential of 0.90 V. The modied electrode shows the initial current response due to 600 nM FA. The current response increases and a steady state current is Fig. 4 DPVs obtained for the increment of 10 mM FA to 2500 mM AA in a 0.2 M PB solution at the a-Fe2O3 nanofiber modified GC electrode. Pulse width ¼ 0.06 s, amplitude ¼ 0.05 V, sample period ¼ 0.02 s and pulse period ¼ 0.2 s. Fig. 5 (A) Amperometric i–t curve for the determination of FA at the a-Fe2O3 nanofiber modified GC electrode in a 0.2 M PB solution. Each addition increases the concentration of 60 nM of FA. Eapp ¼ 0.90 V. (B) Calibration plot obtained for conc. of FA vs. amperometric current. (C) (a) 60, (b) 300, (c) 900, (d) 1500, (e) 4500, (f) 9000, (g) 15 000, (h) 20 000, (i) 30 000, (j) 40 000 and (k) 60 000 mM addition of FA at the a-Fe2O3 nanofiber modified electrode. (D) Amperometric i–t curve response obtained for the addition of 60 nM FA (a–c) and a mixture of 60 nM each of FA and AA (d–f) using the a-Fe2O3 nanofiber electrode in a 0.2 M PB solution at a regular interval of 50 s. This journal is ª The Royal Society of Chemistry 2013 Analyst, 2013, 138, 1779–1786 | 1783 Paper Analyst DownloadedbyUniversityofTexasLibrarieson22February2013 Publishedon31January2013onhttp://pubs.rsc.org|doi:10.1039/C3AN00070B View Article Online
  • 6. attained within 3 s for further addition of 60 nM FA in each step with a sample interval of 50 s. The dependence of the response current with respect to the concentration of FA was linear from 60 nM to 600 nM at the a-Fe2O3 nanober modied GC elec- trode with a correlation coefficient of 0.9991 (Fig. 5B). The current response for 60 nM FA was found to be 39.2 nA. Further, the amperometric current response was increased linearly with increasing FA concentration in the range of 60–60 000 nM (Fig. 5C) with a correlation coefficient of 0.9901, and the a-Fe2O3 nanober modied electrode displayed an excellent sensitivity for FA detection with an experimental detection limit of 60 nM (1.12 Â 10À10 M (S/N ¼ 3)). The linear range and the lowest detection limit for FA at a-Fe2O3 nanobers were compared with the recently reported chemically modied electrodes.46–51 Thus, the present modied electrode shows the lowest detection limit for FA (60 nM (1.12 Â 10À10 M (S/N ¼ 3)) when compared to the reported FA detection limits (see Table 1).46–51 As mentioned above, the normal level of FA in blood serum is 34.4 Æ 10.4 nmol LÀ1 . Therefore, the a-Fe2O3 nanober modied GC electrode is more suitable for the determination of FA in real (blood serum) samples even in the presence of 53.8 Æ 36.6 mmol LÀ1 AA. The amperometric method was also performed to determine the concentration of FA along with AA. The amperometric current response for the alternative addition of AA and FA in the mixture is shown in Fig. 5D. The a-Fe2O3 nanober modied GC electrode showed the initial current response due to the addi- tion of 60 nM AA (Fig. 5D; curves a–c) into a PB solution with a sample interval of 50 s, and the current response was increased. Further, the addition of a mixture of 60 nM AA and 60 nM FA to a stirred solution of 0.2 M PB showed a two-fold enhanced amperometric oxidation current at the same applied potential (Fig. 5D; curves d–f). The two-fold amperometric oxidation current obtained was due to the oxidation of both AA and FA. 3.5 Anti-interference ability of the a-Fe2O3 nanobers The anti-interference ability of the a-Fe2O3 nanobers was tested towards the detection of FA from various common ions such as Na+ , Ca2+ , and SO4 2À , and some physiological interfer- ents such as glucose, urea and oxalate using the amperometric method (Fig. 6). Furthermore, no change in the amperometric current response was observed for 60 nM FA in the presence of 60 mM of MgSO4, CaCl2, NaCl, K2CO3, NaF, ClÀ , FÀ , and NH4Cl, indicating that the present modied electrode is highly selective towards the determination of FA even in the presence of a 1000- fold excess of these interferents. 3.6 The stability and reproducibility of the a-Fe2O3 nanober modied electrode In order to investigate the stability of the a-Fe2O3 nanober modied GC electrode, the DPVs for 0.20 mM FA in a 0.20 M PB solution were recorded for every 5 min interval. It was found that the oxidation peak current remained the same with a relative standard deviation of 2.1% for 20 repetitive measure- ments, indicating that the electrode has a good reproducibility and does not undergo surface fouling. Aer voltammetric measurements, the electrode was kept in a pH ¼ 7.2 PB solution at room temperature. The current response decreased about 1.24% in one week and 5.54% in about two weeks. To ascertain the reproducibility of the results, three different GC electrodes were modied with the a-Fe2O3 nanobers and their response towards the oxidation of 0.50 mM AA and FA was tested by 20 repeated measurements. The separation between the voltam- metric peaks of AA–FA was the same at all the four electrodes. The peak current obtained in the 20 repeated measurements of three independent electrodes showed a relative standard devi- ation of 1.48%, conrming that the results are reproducible. The above results showed that the present modied electrode was very much stable and reproducible towards these analytes. It is worthy to compare the determination of FA at the a-Fe2O3 nanober modied GC electrode with other chemically modi- ed electrodes. In the reported papers, the procedures adopted for the modication of electrode surfaces are very tedious, more time consuming and further reproducible results cannot be obtained.47,48,52–54 In the case of a carbon paste electrode, rst the carbon paste was mixed with the palmitic and stearic acids Table 1 Comparison of different chemically modified electrodes for the deter- mination of FA with the a-Fe2O3 nanostructure modified electrode Modied electrodes Detection limit Ref. Single-walled carbon nanotube-ionic liquid paste electrode 1 Â 10À9 M 46 Single-walled carbon nanotube lm modied glassy carbon electrode 1 Â 10À9 M 47 Lead lm modied glassy carbon electrode 7 Â 10À10 M 49 Poly(5-amino-2-mercapto-1,3,4- thiadiazole) lm modied glassy carbon electrode 2.3 Â 10À10 M 50 3-Amino-5-mercapto-1,2,4-triazole polymerized lm modied glassy carbon electrode 2.5 Â 10À7 M 51 a-Fe2O3 nanostructure modied glassy carbon electrode 1.12 Â 10À10 M This work Fig. 6 Amperometric i–t curve for 60 nM addition of FA at the a-Fe2O3 nanofiber modified GC electrode (a–c, g–i), and the addition of 60 mM of Na+ , Ca2+ , SO4 2À (d–f), glucose, urea and oxalate (j–l), in a homogeneously stirred 0.2 M PB solution. 1784 | Analyst, 2013, 138, 1779–1786 This journal is ª The Royal Society of Chemistry 2013 Analyst Paper DownloadedbyUniversityofTexasLibrarieson22February2013 Publishedon31January2013onhttp://pubs.rsc.org|doi:10.1039/C3AN00070B View Article Online
  • 7. in the presence of carbon tetrachloride and then dried over- night at room temperature.52 Similarly, for the fabrication of a multi-walled carbon nanotube coated Au electrode, the multi- walled carbon nanotubes (MWNTs) were reuxed in the mixture of concentrated H2SO4 and HNO3 for 4–5 h, then washed with water and dried in vacuum at room temperature.53 When compared to the reported procedure for the electrode modi- cations, the procedure for the deposition of a-Fe2O3 nanobers on the GC electrode in the present study is very easy, less time consuming (<12 min), highly stable and reproducible. 3.7 Determination of FA in human blood serum samples The practical application of the a-Fe2O3 nanober modied GC electrode was tested by measuring the concentration of FA in human blood serum samples. The human blood serum samples were collected from a local hospital (Muthu clinic and X-rays, Dindigul district, India). The standard addition technique was used for the determination of FA in serum samples. The DPV of blood serum in a PB solution (pH ¼ 7.2) shows two oxidation peaks at 0.35 and 0.81 V as shown in Fig. 7, green line, and these peaks may be due to the oxidation of AA and FA, respectively. To conrm the observed oxidation peak at 0.81 V for FA in Fig. 7, green line, we have added a known concentration of FA into the same blood serum solution, the oxidation current at 0.81 V was further enhanced (Fig. 7; blue line) and recovery results are given in Table 2. The enhanced oxidation peak current at 0.81 V indicated that the peak corresponds to the oxidation of FA. The proposed method shows a better recovery of spiked FA in serum samples, indicating that the adopted method could be effi- ciently used for the determination of FA in real samples in the presence of possible interferents. 4 Conclusions We have demonstrated the synthesis of a-Fe2O3 nanobers by a simple electrospinning method and their application in vol- tammetric determination of FA in the presence AA (pH 7.2). The a-Fe2O3 nanober modied electrode not only separates the voltammetric signals of AA and FA with a potential difference of 420 mV between AA and FA, but also shows a higher oxidation current than the bulk-Fe2O3 and unmodied electrodes. The amperometric current response is linearly dependent on FA concentration in the range of 60–60 000 nM, and the a-Fe2O3 nanober modied GC electrode displayed an excellent sensi- tivity for FA detection with an experimental detection limit of 60 nM (1.12 Â 10À10 M (S/N ¼ 3)). The practical application of the present modied electrode was successfully demonstrated by determining the concentration of FA in human blood serum samples. The excellent analytical performance and low cost nanomaterials are not only scientically signicant for the development of effective biosensors, but also could produce real benets such as energy and cost savings in comparison with other noble metals or enzymes for a wide range of potential applications in medicine, catalysis, and biosensing. Acknowledgements The authors thank the National University of Singapore and Nanyang Technological University for providing excellent research facilities to carry out this work. Palanisamy Kannan and Marcin Opallo thank NanOtechnology, Biomaterials and aLternative Energy Source for the ERA Integration [FP7- REGPOT-CT-2011-285949-NOBLESSE] Project from the Euro- pean Union. References 1 J. Kong, N. R. Franklin, C. Zhou, M. G. Chapline, S. Peng, K. Cho and H. Dai, Science, 2000, 287, 622–625. 2 P. Poizot, S. Laruelle, S. Grugeon, L. Dupont and J. M. Tarascon, Nature, 2000, 407, 496–499. 3 D. Yu and V. W.-W. Yam, J. Am. Chem. Soc., 2004, 126, 13200– 13201. 4 G. Neri, A. Bonavita, S. Galvagno, P. Siciliano and S. Capone, Sens. Actuators, B, 2002, 82, 40–47. 5 Y. Cui, Q. Wei, H. Park and C. M. Lieber, Science, 2001, 293, 1289–1292. 6 K. Ramanathan, M. A. Bangar, M. Yun, W. Chen, N. V. Myung and A. Mulchandani, J. Am. Chem. Soc., 2004, 127, 496–497. 7 R. J. Chen, S. Bangsaruntip, K. A. Drouvalakis, N. Wong Shi Kam, M. Shim, Y. Li, W. Kim, P. J. Utz and H. Dai, Proc. Natl. Acad. Sci. U. S. A., 2003, 100, 4984–4989. 8 Y. Ding, P. X. Gao and Z. L. Wang, J. Am. Chem. Soc., 2004, 126, 2066–2072. Fig. 7 DPVs obtained for blood serum (green line) and after the addition of 10 mM commercial FA (blue line) to blood serum at the a-Fe2O3 nanofiber modified GC electrode in a 0.2 M PB solution. Pulse width ¼ 0.06 s, amplitude ¼ 0.05 V, sample period ¼ 0.02 s and pulse period ¼ 0.2 s. Table 2 Determination of FA in human blood serum samples Human blood serum Original (mM) Added (mM) Found (mM) Recovery (%) Sample 1 50.10 10 59.80 99.5 Sample 2 25.40 10 35.16 99.3 This journal is ª The Royal Society of Chemistry 2013 Analyst, 2013, 138, 1779–1786 | 1785 Paper Analyst DownloadedbyUniversityofTexasLibrarieson22February2013 Publishedon31January2013onhttp://pubs.rsc.org|doi:10.1039/C3AN00070B View Article Online
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