Investigation of Mediated Oxidation of Ascorbic Acid by

Anal. Chem. 2008, 80, 6515–6525

Investigation of Mediated Oxidation of Ascorbic Acid by Ferrocenemethanol Using Large-Amplitude Fourier Transformed ac Voltammetry under Quasi-Reversible Electron-Transfer Conditions at an Indium Tin Oxide Electrode Benchaporn Lertanantawong,† Anthony P. O’Mullane,‡ Jie Zhang,§ Werasak Surareungchai,† Mithran Somasundrum,† and Alan M. Bond*,‡ School of Bioresources and Technology, King Mongkut’s University of Technology, Thonburi, Thailand, School of Chemistry, Monash University, Clayton, Victoria 3800, Australia, and Institute of Bioengineering and Nanotechnology, 31 Biopolis Way, The Nanos, Singapore 138669 The ability of the technique of large-amplitude Fourier transformed (FT) ac voltammetry to facilitate the quantitative evaluation of electrode processes involving electron transfer and catalytically coupled chemical reactions has been evaluated. Predictions derived on the basis of detailed simulations imply that the rate of electron transfer is crucial, as confirmed by studies on the ferrocenemethanol (FcMeOH)mediated electrocatalytic oxidation of ascorbic acid. Thus, at glassy carbon, gold, and boron-doped diamond electrodes, the introduction of the coupled electrocatalytic reaction, while producing significantly enhanced dc currents, does not affect the ac harmonics. This outcome is as expected if the FcMeOH0/+ process remains fully reversible in the presence of ascorbic acid. In contrast, the ac harmonic components available from FT-ac voltammetry are predicted to be highly sensitive to the homogeneous kinetics when an electrocatalytic reaction is coupled to a quasi-reversible electron-transfer process. The required quasi-reversible scenario is available at an indium tin oxide electrode. Consequently, reversible potential, heterogeneous chargetransfer rate constant, and charge-transfer coefficient values of 0.19 V vs Ag/AgCl, 0.006 cm s-1 and 0.55, respectively, along with a second-order homogeneous chemical rate constant of 2500 M-1 s-1 for the rate-determining step in the catalytic reaction were determined by comparison of simulated responses and experimental voltammograms derived from the dc and first to fourth ac harmonic components generated at an indium tin oxide electrode. The theoretical concepts derived for large-amplitude FT ac voltammetry are believed to be applicable to a wide range of important solution-based mediated electrocatalytic reactions. L-Ascorbic acid (H2A) participates in many important biological reactions. Ascorbic acid also is a powerful antioxidant and

* To whom correspondence should be addressed. E-mail: alan.bond@ sci.monash.edu.au. † King Mongkut’s University of Technology. ‡ Monash University. § Institute of Bioengineering and Nanotechnology. 10.1021/ac702531f CCC: $40.75  2008 American Chemical Society Published on Web 07/31/2008

hence is of considerable interest in the food and pharmaceutical industries.1–15 Electrochemical detection of H2A by direct oxidation at conventional electrodes, such as Au,16 Pt,17 and glassy carbon (GC),7,8,14 is complicated by slow electrontransfer kinetics6,15,18–20 and surface fouling by oxidation products.21 Thus, sensitivity, selectivity, and reproducibility often are relatively poor. The peak potentials reported under conditions of dc voltammetry for H2A oxidation at GC electrodes vary significantly, ranging from 0 to 600 mV (vs Ag/AgCl). Peak potential values in the range of 0-250 mV are found at GC electrodes activated through heat,6,7 laser treatment,,23 or a rigorous polishing procedure.24 Commonly, peak potential values reported with use of conventionally polished GC electrodes9,13,25 lie in the 450-600 mV range. (1) Kirk, R.; Sawyer, R. Pearson’s Composition and Analysis of Food; Longman Scientific & Technical Press: Harlow, UK, 1991. (2) Aoki, A.; Matsue, T.; Uchida, I. Anal. Chem. 1992, 64, 44. (3) Hassan, H. N. A.; Barsoum, B. N.; Habib, I. H. I. J. Pharm. Biomed. Anal. 1999, 20, 315. (4) Kwakye, J. K. Talanta 2001, 51, 197. (5) Yang, Y. J. Pharm. Biomed. Anal. 1998, 18, 274. (6) Deakin, M. R.; Kovach, P. M.; Stutts, K. J.; Wightman, R. M. Anal. Chem. 1986, 58, 1474. (7) Hu, I. F.; Kuwana, T. Anal. Chem. 1986, 58, 3235. (8) Pournaghi-Azar, M. H.; Ojani, R. Talanta 1995, 42, 1839. (9) Yu, A.-M.; Zhang, H.-L.; Chen, H.-Y. Electroanalysis 1997, 9, 788. (10) Ageliki, B. F.; Mamas, I. P.; Miltiades, I. K.; Stella, M. T.-K. Anal. Chim. Acta 2000, 409, 113. (11) Zhang, G.; Wang, X.; Shi, X.; Sun, T. Talanta 2000, 51, 1019. (12) Moreno, G.; Pariente, F.; Lorenzo, E. Anal. Chim. Acta 2000, 420, 29. (13) Wang, J.; Wu, Z.; Tang, J.; Teng, R.; Wang, E. Electroanalysis 2001, 13, 1093. (14) Raj, C. R.; Ohsaka, T. J. Electroanal. Chem. 2003, 540, 69. (15) Salimi, A.; MamKhezri, H.; Hallaj, R. Talanta 2006, 70, 823. (16) Rueda, M.; Aldaz, A.; Sanchez.Burgos, F. Electrochim. Acta 1978, 23, 419. (17) Karabinas, P.; Jannakoudakis, D. J. Electroanal. Chem. 1984, 160, 159. (18) Pachla, L. A.; Renolds, D. L.; Kissinger, P. T. Anal. Chem. 1985, 68, 1. (19) Nassef, H. M.; Radi, A.-E.; O’Sullivan, C. Anal. Chim. Acta 2007, 583, 182. (20) Thangamuthu, R.; Kumar, S. M. S.; Pillai, K. C. Sens. Actuators, B 2007, 120, 745. (21) John, S. A. J. Electroanal. Chem. 2005, 579, 249. (22) Rice, R.; Allred, C.; McCreery, R. J. Electroanal. Chem. 1989, 263, 163. (23) Poon, M.; McCreery, R. L.; Engstrom, R. Anal. Chem. 1988, 60, 1725. (24) Aihara, M.; Komatsu, M. Bull. Chem. Soc. Jpn. 1987, 60, 1911. (25) Pournaghi-Azar, M. H.; Ojani, R. Talanta 1997, 297.

Analytical Chemistry, Vol. 80, No. 17, September 1, 2008

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In order to minimize surface fouling effects, increase the sensitivity, and enhance the reliability and reproducibility of analytical data, the mediated oxidation of H2A by a redox couple has been introduced.26,27 An ideal mediator needs to exhibit fast electron-transfer kinetics, have a formal reversible potential (E°′) less positive than that for the reversible potential for oxidation of ascorbic acid, and be stable in both oxidized and reduced forms. Successful mediators that meet these requirements are often based on ferrocene and its derivatives8,10,11,13,28–33 and they have been employed in the form of surface confined layers or when present in the solution phase. In this paper, details of the electrocatalytic oxidation of ascorbic acid in acetate buffer solution have been quantified when ferrocenemethanol (FcMeOH) dissolved in an acetate buffer is used as a mediator and the electrode materials employed are GC, Au, boron-doped diamond (BDD), and indium tin oxide (ITO). While qualitative studies of such an electrocatalytic process are common, quantitative studies are rare,8 probably because of difficulty in resolving H2A oxidation and FcMeOH electron-transfer reaction under catalytic conditions. Thus, in addition to undertaking conventional dc cyclic voltammetric experiments, the largeamplitude Fourier transformed alternating current (FT-ac) technique is introduced for the first time into studies of H2A oxidation. The FT-ac technique offers significant advantages relative to traditional dc or ac fundamental harmonic approaches. These include suppression of background capacitive current, as exploited in studies involving solution-phase34,35 and surface-confined processes.36–38 Furthermore, the ability to separate the underlying reversible electron-transfer process from the coupled catalytic chemical reaction can be achieved with the higher harmonics, as demonstrated in the case of mediated glucose oxidation.39 In the case of ascorbic acid, the underlying FcMeOH0/+ redox process is reversible at GC, BDD, and Au electrodes, but quasi-reversible at ITO electrodes. In this paper, it will be shown that the ability to vary the heterogeneous electron-transfer kinetics of the redox process as well as the relative insensitivity to the overlapping H2A oxidation process can be exploited to quantify all aspects of the catalytic reaction scheme by large amplitude FT-ac voltammetry at an ITO electrode. EXPERIMENTAL SECTION Reagents and Chemicals. L-Ascorbic acid (99.0%, BDH), sodium acetate (BDH), acetic acid (100%, BDH), lithium (26) (27) (28) (29) (30) (31) (32) (33) (34) (35) (36) (37) (38) (39)

Winograd, N.; Blount, H. N.; Kuwana, T. J. Phys. Chem. 1969, 73, 3456. Kitani, A.; Miller, L. L. J. Am. Chem. Soc. 1981, 103, 3595. Dautartas, M. F.; Evans, J. F. J. Electroanal. Chem. 1980, 109, 301. Petersson, M. Anal. Chim. Acta 1986, 187, 333. Scholl, H.; Socha, K. Electrochim. Acta 1991, 36, 689. Vidal, J. C.; Yague, M. A.; Castillo, J. R. Sens. Actuators B 1994, 21, 135. Raoof, J.-B.; Ojani, R.; Kiani, A. J. Electroanal. Chem. 2001, 515, 45. Fernandez, L.; Carrero, H. Electrochim. Acta 2005, 1233. Sher, A. A.; Bond, A. M.; Gavaghan, D. J.; Harriman, K.; Feldberg, S. W.; Duffy, N. W.; Guo, S.-X.; Zhang, J. Anal. Chem. 2004, 76, 6214. Bond, A. M.; Duffy, N. W.; Guo, S.-X.; Zhang, J.; Elton, D. Anal. Chem. 2005, 77, 186A. Guo, S-X. ; Zhang, J.; Elton, D. M.; Bond, A. M. Anal. Chem. 2004, 76, 166. Zhang, J.; Guo, S.-X.; Bond, A. M.; Honeychurch, M. J.; Oldham, K. B. J. Phys. Chem. B 2005, 109, 8935. Fleming, B. D.; Barlow, N. L.; Zhang, J.; Bond, A. M.; Armstrong, F. A. Anal. Chem. 2006, 78, 2948. Fleming, B. D.; Zhang, J.; Bond, A. M.; Bell, S. G.; Wong, L.-L. Anal. Chem. 2005, 77, 3502.

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perchlorate (99%, TLC), and ferrocenemethanol (Aldrich) were used as supplied by the manufacturer. Ascorbic acid and ferrocenemethanol solutions were freshly prepared prior to use. pH 5.0 acetate buffer solutions (0.10 M), containing 0.10 M LiClO4, used as the electrolyte and also to control the pH, were prepared from acetic acid and sodium acetate. Distilled water (Millipore, 18.2 mΩ cm resistivity) was used to prepare all solutions. Instrumentation and Procedures. Conventional dc cyclic voltammetric experiments were undertaken in a standard threeelectrode cell with a BAS Epsilon electrochemical workstation. The reference electrode was Ag/AgCl (3 M KCl), and the counter electrode was a platinum wire. The working electrodes consisted of glassy carbon (0.067 cm2), gold (0.023 cm2), indium tin oxide-coated glass (Prazisions Glas and Optik GmbH), having an electrochemically active area of 0.048 or 0.390 cm2 and a sheet resistance of 10 Ω/sq (as quoted by the manufacturer), and boron-doped diamond (0.11 cm2, Windsor Scientific, boron carrier concentration of 1020 atoms cm-3). ITO electrodes were cleaned by sonicating in acetone and propan2-ol and then dried with a flow of nitrogen gas. GC, Au, and BDD electrodes were polished with 0.3-µm alumina slurry (Buehler) on Microcloths, rinsed in deionized water, and airdried, except BDD, which had to be sonicated for 5 min in deionized water after polishing. Rotating disk electrode (RDE) experiments, with a 3-mm-diameter GC working electrode (BAS), used the BAS Epsilon electrochemical workstation in combination with a BAS RDE-2 accessory. Bulk oxidative electrolysis of FcMeOH with a BAS 100A power module (PWR-3) was used to prepare ferriciniummethanol cation (FcMeOH+) solutions. The working electrode was a solid GC basket placed in a glass cylinder with a porous glass frit in the base, which was arranged symmetrically inside a larger solid GC basket (the auxiliary electrode). The reference electrode was the same as used for voltammetric experiments and placed within the working electrode compartment. The auxiliary electrode compartment was filled with pH 5 acetate buffer, while the working electrode compartment was filled with the acetate buffer containing 1 mM FcMeOH. The solution in the working electrode compartment was stirred vigorously during bulk electrolysis. Experiments were undertaken at room temperature (20 ± 2 °C), and all solutions were deaerated with purified nitrogen to remove oxygen for at least 10 min prior to the beginning of experiments. The pH of the solution was measured with a Metrohm 744 pH meter. Uncompensated resistance (Ru) values were obtained using the BAS-Epsilon potentiostat and applying a small potential step in a region where no faradaic reaction occurs. Analysis of the charging current versus time curve allows Ru to be extracted40 and gave values in the range of 150-250 Ω. A description of the FT-ac voltammetric instrumentation is available elsewhere.35 A single sine wave of frequency f ) 21.46 Hz and amplitude ∆E over the range of 80-150 mV was employed as the ac perturbation source. The dc component and the ac harmonics are obtained by sequential application of FT and inverse FT algorithms. In brief, the FT algorithm is used to convert the calculated time domain data to the frequency domain and

presented in the form of the power spectrum. The relevant region (dc or harmonic of interest) in the power spectrum is selected and the power at all other frequencies set to zero. Application of the inverse FT algorithm gives the required dc, fundamental, second, and higher harmonic components. In this study, the magnitude of the resolved ac component is compared with experimental data obtained by an exactly analogous procedure. A copy of the simulation programs, written in Fortran 77, are available on request to the authors. Simulations of dc cyclic voltammograms were carried out with DigiSim and DigiElch (Version 2) software. THEORY In the theory relevant to the electrocatalytic reaction scheme of interest in this paper, a redox mediator heterogeneous chargetransfer electron-transfer step (eq 1) is assumed to be followed by two irreversible second-order homogeneous chemical reactions (eqs 2 and 3) which lead to the regeneration of the starting electroactive species, R.

kf

R y\z O + e-

(1)

kb

k1

O + X 98 R + X+

k2

O + X+ 98 R + X2+

(2)

(3)

where O and R are the oxidized form and the reduced form of the redox mediator, respectively; kf and kb are the forward and backward heterogeneous charge-transfer rate constants associated with the electron-transfer process; X, X+, and X2+ are the reduced, one-electron-oxidized, and two-electron-oxidized forms of the substrate and k1 and k2 are the rate constants for the irreversible homogeneous bimolecular reactions. If Butler-Volmer kinetics are obeyed, then

Figure 1. Effect of a catalytic reaction on the voltammetric characteristics of (a, 1-5) a reversible electron-transfer process, (b, 1-5) a quasireversible electron-transfer process (k0′ ) 0.01 cm s-1, R ) 0.50) and (c, 1-5) a quasi-reversible electron-transfer process (k0′) 0.001 cm s-1, R ) 0.50) in simulations in the absence (black) and presence (red) of 10 mM X when k1 ) 300 M-1 s-1. For (a, 1-5), first-fourth ac harmonics are essentially identical irrespective of whether cX* ) 0 or 10 mM, so only the case where cX* ) 0 is presented. Other parameters: v ) 100 mV s-1, f ) 21 Hz, ∆E ) 150 mV, A ) 0.01 cm2, DR ) 7 × 10-6 cm2 s-1, DX ) 7 × 10-6 cm2 s-1 (D for oxidized and reduced forms assumed equal), E°′ ) 0.40 V, Ru ) 0 Ω, Cdl ) 0 µF cm-2, and T ) 293 K (k2 ) 1 × 107 M-1 s-1). In experiments considered later, R ) FcMeOH and X ) ascorbic acid. Analytical Chemistry, Vol. 80, No. 17, September 1, 2008

6517

RF (E(t) - E )] [ RT

kf ) k0′exp

0′

(4)

-(1 - R)F (E(t) - E0′) kb ) k exp RT 0′

[

]

(5)

where k0′ is the formal charge-transfer rate constant, R is the charge-transfer coefficient, E0′ is the formal potential of the redox process (eq 1), E(t) is the applied potential, t is the time, and R, T, and F have their usual meanings. The potential waveform employed is a combination of a sine wave and a ramped dc waveform, given by eq 6, E(t) ) Edc(t) + Eac(t)

(6)

For an oxidation process, 0 e t e ts : Edc(t) ) Estart + vt ts < t e 2ts : Edc(t) ) Estart + 2vts - vt

(7) (8)

0 e t e 2ts : Eac(t) ) ∆E sin(ωt)

(9)

where Edc(t) and Eac(t) are applied dc and ac waveforms, respectively; v is the scan rate of the dc ramp; Estart is the starting potential; ts is time required to finish a complete sweep of potential; ω() 2πf); and ∆E are the angular frequency (f is frequency) and amplitude of the applied sine wave. Under linear diffusion conditions, the mass transport for all diffusing species of interest follows the relationship, ∂cR ∂2cR ) DR 2 + k1cOcX + k2cOcX+ ∂t ∂x

(10)

∂cO ∂2cO ) DO 2 - k1cOcX - k2cOcX+ ∂t ∂x

(11)

∂cX ∂2cX ) DX 2 - k1cOcX ∂t ∂x

(12)

∂cX+ ∂2cX+ ) DX+ 2 + k1cOcX - k2cOcX+ ∂t ∂x

(13)

Figure 2. Effect of a catalytic reaction on the voltammetric characteristics of (a1-5) a reversible electron-transfer process (k0′) 1 × 104 cm s-1, R ) 0.50), (b, 1-5) a quasi-reversible electron-transfer process (k0′) 0.01 cm s-1), and (c, 1-5) a quasi-reversible electron-transfer process (k0′ ) 0.001 cm s-1) used in simulations in the absence (black) and presence (red) of 10 mM X when k1 ) 3000 M-1 s-1. For (a, 1-5), first-fourth ac harmonics are essentially identical irrespective of whether cX* ) 0 or 10 mM, so only the case where cX* ) 0 is presented. All other conditions and parameters are as stated in Figure 1. 6518


Figure 3. Cyclic voltammograms obtained at a scan rate of 15 mV s-1 at GC (---), BDD (-•-), Au (ss), and ITO (•••) electrodes for the oxidation of (a) 1 mM H2A (as HA-) and (b) 10 mM H2A (as HA-) in 0.1 M acetate buffer (pH 5.0) containing 0.1 M LiClO4.

where ci and Di (i ) R, O, X, and X+) are the concentration and diffusion coefficient of the species i, respectively, and x is the distance from electrode surface. This equation can be solved numerically using a fully implicit finite difference method with the Richtmeyer modification,41 application of expanding space grid in order to enhance numerical efficiency and accuracy,42 and application of the following initial and boundary conditions: t ) 0, all x: cO ) 0, cR ) c/R, cX ) c/X and cX+ ) 0

(14)

t > 0, x ) 0 (electrode/solution interface): -DO

∂cO ∂cR ∂cX ∂cX+ ) DR ) kfcR - kbcO and DX ) DX+ )0 ∂x ∂x ∂x ∂x (15)

t > 0, x ) ∞: cO ) 0, cR ) c/R, cX ) c/X and cX+ ) 0

(16)

where cR* and cX* are the bulk concentrations of the reduced forms of electroactive species and substrate, respectively. Current, I, can be calculated according to eq 17 once the space and time dependent concentration is known,

( )

I ) FADR

∂cR ∂x

x)0

(17)

where A is the electrode area. The effects of double layer capacitance (Cdl) and uncompensated resistance (Ru) are also taken into account in the simulation using widely adopted procedures.43 The theoretical model outlined above involves key parameters such as Cdl, Ru, f, ∆E, v, k0′, R, k1, and k2. However, in this paper, the influence of cX*, k0′, and k1 on the voltammetric characteristics is of particular interest. Furthermore, we are only interested in the case where the homogeneous reaction in eq 2 is the ratelimiting step. Consequently, k2 . k1, so the exact value of k2 used in simulations has no influence on the characteristics of the voltammograms although the simulation code is generally applicable to all scenarios. Simulated currents, I, are calculated at 293 K, and the results given immediately below refer to use of the following parameters: v ) 100 mV s-1, f ) 21 Hz, ∆E ) 150 mV, A ) 0.01 cm2, cR* ) 1 mM, cX*) 10 mM, D (R, O, X, X+, and X2+) = 7 × 10-6 cm2 s-1, k1 ) 300 (Figure 1) or 3000 (Figure 2) s-1 M-1 and k2 ) 1 × 107 s-1 M-1. Other parameters used in the simulations are provided in the figure captions. The parameters used in initial simulations encompass the range of values expected for the electrocatalytic oxidation of ascorbic acid using FcMeOH as a mediator. A large ∆E ) 150 mV was required to study the fourth harmonic under conditions where the R h O + e- process is quasi-reversible. This large amplitude also results in splitting of the dc component when the FcMeOH0/+ process is fully reversible (Figure 1, a1) and when k0′ ) 0.01 cm s-1 (Figure 1, b1). Electrode capacitance and uncompensated resistance are ignored in the following discussion of simulated voltammograms, but will be taken into account in the theory versus experiment comparisons. The simulated voltammograms in Figure 1 were obtained with k1 ) 300 s-1 M-1 and for various rates of electron transfer and cX* ) 10 mM. In the case of reversible electron transfer (Figure 1, a2-5), symmetrical ac components are obtained for all harmonics. Furthermore, the ac components are independent of the presence or absence of the catalytic reaction. In contrast, the characteristics of the dc components are altered, and the magnitude of the dc current is enhanced significantly by the presence of the catalytic reaction (Figure 1, a1, b1, c1). The magnitude of ac peak current but not dc limiting current becomes diminished by the onset of quasi-reversibility (e.g., k0′) 0.01 cm s-1 in Figure 1, b2-5) as expected.44 Nevertheless, the presence of the catalytic reaction still does not significantly alter the voltammetric characteristics of the ac components under conditions relative to Figure 1, b2-5. If the kinetics of heterogeneous electron transfer are lowered to 0.001 cm s-1 (Figure 1 c1-5), the faradic currents of the ac components are of course further diminished relative to the reversible case, but slightly enhanced by the presence of catalytic reaction. Results of simulations with faster coupled homogeneous kinetics (k1 ) 3000 s-1 M-1) were also examined. Again (Figure 2, a1-5) for reversible electron transfer, even though the magnitude of the dc current is further enhanced by the presence of an even faster catalytic reaction, the ac components remain virtually unaffected. In (40) Bard, A. J.; Faulkner, L. R. Electrochemical Methods, 2nd ed.; John Wiley and Sons: New York, 2001. (41) Mocak, J.; Feldberg, S. W. J. Electroanal. Chem. 1994, 378, 31. (42) Feldberg, S. W. J. Electroanal. Chem. 1981, 127, 1. (43) Rudolph, M.; Reddy, D. P.; Feldberg, S. W. Anal. Chem. 1994, 66, 589A. (44) Zhang, J.; Guo, S.-X.; Bond, A. M.; Marken, F. Anal. Chem. 2004, 76, 3619.


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Figure 4. Cyclic voltammograms obtained with a scan rate of 15 mV s-1 for oxidation of 1 mM FcMeOH in 0.1 M acetate buffer (pH 5.0) in the absence (s), the presence of 1 mM H2A (---), and 10 mM H2A (as HA-) (-•-) at (a) GC, (b) Au, (c) BDD, and (d) ITO electrodes.

the quasi-reversible case when k0′) 0.01 cm s-1 (Figure 2, b1-5), the presence of the catalytic reaction now clearly starts to influence the voltammetric shapes and potentials of the ac components, even though the magnitudes of the faradic ac harmonic current remain similar. When the level of irreversibility is increased so that k0′ ) 0.001 cm s-1 (Figure 2, c1-5), even the magnitudes of faradic ac components are enhanced significantly by the presence of the catalytic reaction. An important conclusion derived from analysis of simulation is that the underlying heterogeneous electron-transfer process should exhibit quasi-reversible kinetics in order that the homogeneous aspects of the process may be quantified by FT-ac voltammetry. These simulated results imply that the magnitude of ac components is dominated by the process which has the most significant influence on the concentration of the redox couple at the interfacial region. In the case of reversible electron transfer, the concentration of the redox species at the interfacial region is not strongly influenced by the catalytic step and remains predominantly governed by the Nernst relationship. However, for a quasi-reversible electron-transfer process, the concentrations of the redox-active species at the interfacial region become more significantly affected by the homogeneous chemical reaction process. Under these circumstances, the coupled homogeneous reaction may significantly influence the ac wave shape and current magnitude. The level to which this influence occurs is increased when the rate of electron transfer is decreased or the rate of the coupled catalytic homogeneous reaction is increased. Consequently, it is concluded that complete quantification of the catalytic scheme elucidated in this work by FT-ac voltammetry requires slow electron transfer for the FcMeOH0/+ process, a condition it 6520


will emerge that is obtained at an ITO electrode, but not at GC, Au, or BDD surfaces, where the process is essentially reversible. RESULTS AND DISCUSSION dc Cyclic Voltammetry for the Oxidation of Ascorbic acid at GC, Au, BDD, and ITO Electrodes. Ascorbic acid has two acidic protons (pKa1 ) 4.04 and pKa2 ) 11.34).45 Consequently, at pH 5.0, the condition employed in this study, the monoascorbate anion is the dominant species. The oxidation of the monoascorbate anion has been demonstrated to proceed via two consecutive oneelectron processes, involving the participation of a radical anion intermediate, to form dehydro-L-ascorbic acid. The latter species subsequently undergoes a hydration reaction to form the final electroinactive product.6,16,46 Accordingly, a chemically irreversible oxidation process (no reduction process on reverse scans of cyclic voltammograms) is observed. The reaction pathway is summarized by eqs 18–20, where the reaction described by eq 18 is fast and nonlimiting H2ATHA- + H+

(18)

HA-TA•- + e- + H+

(19)

dc cyclic voltammograms for the oxidation of 1 mM ascorbic acid at GC, BDD, Au, and ITO electrodes in acetate buffer (pH

Figure 5. Comparison of the experimental (s) cyclic voltammograms obtained at (a) an ITO electrode with a sweep rate of 126.66 mV s-1 for (i) 1 mM FcMeOH and (ii) 1 mM FcMeOH and 10 mM H2A (as HA-) in 0.1 M acetate buffer (pH 5.0) containing 0.1 M LiClO4 and simulated data (O). Simulation parameters: v ) 126.66 mV s-1, A ) 0.048 cm2, DFcMeOH ) 7.6 × 10-6 cm2 s-1, DHA- ) 7 × 10-6 cm2 s-1 (D for oxidized and reduced forms assumed equal), E°′ ) 0.190 V, Ru ) 250 Ω, Cdl ) 20 µF cm-2, T ) 293 K, k0′ ) 0.006 cm s-1, and R ) 0.55. In case ii, as for (i) but in addition k1 ) 2500 M-1 s-1, (k2 ) 1 × 107 M-1 s-1), and Ru ) 150 Ω. (b) Cyclic voltammograms obtained with a sweep rate of 15 mV s-1 for 1 mM FcMeOH and 10 mM H2A (as HA-). All simulation parameters as in (ii) except v ) 15 mV s-1, A ) 0.39 cm2, and Ru ) 150 Ω.

5.0) are shown in Figure 3. These voltammograms are presented in the current density versus potential format in order to facilitate comparison of data as a function of electrode material. Clearly, the oxidation of the ascorbate anion (Figure 3a) is highly dependent on the electrode material. The most positive oxidation peak potential is detected at the ITO electrode with the order being ITO > BDD > Au > GC under conditions of Figure 3. This electrode-dependent behavior is also observed at higher ascorbic acid concentrations (10 mM, Figure 3b). The peak current density at Au is lower than at GC or BDD electrodes and the shape of the voltammogram is different, which may be associated with surface fouling.47 Interestingly, the dc voltammogram at the ITO electrode is similar to that commonly reported at nonactivated GC electrodes in the sense that it exhibits a broad oxidation process. The voltammograms obtained at GC in this study are similar (45) Williams, N. H.; Yandell, J. K. Aust. J. Chem. 1982, 35, 1133. (46) Hu, I. F.; Kuwana, T. Anal. Chem. 1986, 58, 3235. (47) Hinoue, T.; Kuwamoto, N.; Watanabe, I. J. Electroanal. Chem. 1999, 466, 31.

to those reported at electrodes activated by heating under vacuum6 or electrochemical pretreatment.48 A response similar to that shown in Figure 3 was observed at several different GC electrodes, all polished in a conventional manner as outlined in the Experimental Section. This electrode-dependent response will emerge to be significant when analyzing data obtained when ferrocenemethanol mediator is present in solution. dc Cyclic Voltammetry for the Oxidation of Ascorbic Acid in the Presence of Ferrocenemethanol. The FcMeOH0/+ process, FcMeOH h FcMeOH+ + e-, has a formal reversible potential of 190 mV versus Ag/AgCl. This value is less positive than that reported for irreversible ascorbic acid oxidation at GC in previous studies9,13,25 and at ITO in this work. However, under conditions employed in this study, the reversible potential for the FcMeOH0/+ process is close to that found for oxidation of ascorbic acid at Au, BDD, and GC electrodes. FcMeOH, as required, is a stable species in aqueous solution and can be oxidized reversibly to FcMeOH+ at GC, metallic electrodes,49,50 and quasi-reversibly at ITO electrodes.51 According to theory, presented above, under large-amplitude ac conditions, only in the case of the ITO electrode should the ac harmonics be sensitive to the process of the catalytic reaction. Cyclic voltammograms obtained at different electrodes for the electrocatalytic oxidation of 1 and 10 mM ascorbic acid in the presence of 1 mM ferrocenemethanol at pH 5.0 are shown in Figure 4a-d. In all cases, the peak potential associated with ascorbic acid oxidation is shifted negatively in the presence of FcMeOH, and concomitantly, the magnitude of the reduction component of the FcMeOH0/+ couple obtained on reversing the scan direction is substantially decreased. Increasing the concentration of ascorbic acid to 10 mM completely eliminates the reduction response associated with the FcMeOH0/+ couple. The proximity of the FcMeOH0/+ and ascorbic acid oxidation processes means that care must be taken to ascertain the magnitude of the catalytic current. Figure S1 (SI) provides data obtained at a GC electrode by simple addition of the current magnitudes of the individual cyclic voltammograms for the FcMeOH0/+ and ascorbic acid oxidation processes compared to the response recorded when both species are present in solution. This experiment shows that the magnitude of the oxidation current is not greatly enhanced by catalysis. Thus, the extent of electrocatalysis may not be as significant as previously reported.8 However, it is clear that electrocatalysis does occur by noting the decrease in the magnitude of the reduction component of the FcMeOH0/+ process on addition of ascorbic acid. Mechanism. The electrocatalytic oxidation of ascorbic acid by FcMeOH8 and other ferrocene (Fc) species 8,11,13 has been extensively reported. The mechanism proposed at pH 5.0 is outlined in eqs 21–23. Fc h Fc+ + e-

(21)

k1

Fc+ + HA- 98 A•- + Fc + H+

(22)

In this scheme, k1 is rate determining, so the scheme used in simulations will be appropriate to determine k1. Also it should be Analytical Chemistry, Vol. 80, No. 17, September 1, 2008

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Figure 6. Large-amplitude dc and fundamental to fifth (a-f) harmonic Fourier transformed ac cyclic voltammograms obtained over a potential range of -250 to 650 mV for the oxidation of 1 mM FcMeOH (red), 1 mM H2A (as HA-) (blue), and 1 mM FcMeOH and 1 mM HA- (black) in 0.1 M acetate buffer (pH 5.0) containing 0.1 M LiClO4. Conditions employed: f ) 21.46 Hz, ∆E ) 80 mV, and ν ) 107.29 mV s-1.

noted that while the generic term ascorbic acid is commonly used for convenience in this paper and others, the species present at pH 5.0 and assumed to be mechanistically important is the ascorbate anion, HA-. To confirm the overall stoichiometry is correct and that 2 mol of FcMeOH is required to oxidize 1 mol of ascorbic acid, a bulk electrolysis experiment was carried out in which a 2 mM FcMeOH solution was quantitatively oxidized to 2 mM FcMeOH+ (confirmed by coulometry and measuring the FcMeOH+ + e- h FcMeOH steady-state limiting current change from fully oxidative to fully reductive current at a RDE). Addition of 0.5 mM ascorbic acid to the solution resulted in a 50% decrease in the magnitude of the reductive limiting current measured at the RDE, confirming that the stoichiometry of the reaction is 2FcMeOH+:1HA-. Initially, the electron-transfer step (eq 21) was simulated separately as a function of sweep rate (15-200 mV s-1) and FcMeOH concentrations (0.1 to 5 mM) at a GC electrode. The diffusion coefficient for FcMeOH was experimentally determined (48) Premkumar, J.; Khoo, S. B. J. Electroanal. Chem. 2005, 576, 105. (49) Bourdillon, C.; Demaille, C.; Moiroux, J.; Saveant, J.-M. J. Am. Chem. Soc. 1995, 117, 11499. (50) Miao, W.; Ding, Z.; Bard, A. J. J. Phys. Chem. B 2002, 106, 1392. (51) Neufeld, A. K.; O’Mullane, A. P. J. Solid State Electrochem. 2006, 10, 808.

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from the Randles-Sevcik relationship40 and found to have a value of 7.6 × 10-6 cm2 s-1, which is in close agreement with the literature.52 An uncompensated resistance value of 150 Ω was experimentally determined (see Experimental Section) and is the origin of the slight increase in the peak-to-peak separation in cyclic voltammograms of the FcMeOH0/+ couple when the FcMeOH concentration was increased from 0.5 to 5 mM. Comparison of simulated and experimental data revealed that the FcMeOH0/+ process was reversible at a GC electrode (k0′ g 1 cm s-1). Other studies at a range of electrode surfaces have reported k0′ values between 0.2 and 2 cm s-1 for the FcMeOH0/+ process in aqueous solution.49,50 The diffusion coefficient of ascorbate (HA-) was experimentally determined using RDE voltammetry at a GC electrode and found to be 7.0 × 10-6 cm2 s-1. This value is in close agreement to literature values53–55 and was used as an input value in simulations of the catalytic reaction scheme outlined in eqs 21–23. To ensure that the ascorbic acid oxidation response does not contribute significantly to the FcMeOH-induced catalytic current, ITO was chosen as an electrode material for evaluation of the full catalytic scheme. As noted above, and in contrast to the situation at other electrode surfaces, the ascorbic acid oxidation response

Figure 7. Large-amplitude dc and fundamental to fourth (a-e) harmonic Fourier transformed ac cyclic voltammograms obtained over a potential range of -200 to 650 mV for the oxidation of 1 mM FcMeOH (---) and 1 mM FcMeOH and 1 mM HA- (s) in 0.1 M acetate buffer (pH 5.0) containing 0.1 M LiClO4. Conditions employed: f ) 21.46 Hz, ∆E ) 150 mV, and ν ) 126.66 mV s-1.

detected at ITO (Epox ) 0.870 V) is well removed from the FcMeOH0/+ process (E°′ ) 0.198 V). Simulation of the FcMeOH0/+ electron-transfer reaction (eq 21) and comparison with experimental data (Figure 5a) resulted in a k0′ value of 0.006 cm s-1 being assigned to the FcMeOH0/+ process at ITO. The dc method is insensitive to R in the range 0.30-0.70, but R ) 0.55 was used in the simulation (value derived from ac method, see later). Simulation of the complete reaction scheme (eqs 21–23) using a k1 value of 2500 M-1 s-1 and other parameters given in the figure caption provided excellent agreement with experimental data derived from 1 mM FcMeOH in the presence of 10 mM ascorbic acid using an ITO electrode of area 0.048 cm2 with a scan rate of 100 mV s-1 (Figure 5a) and electrode area 0.39 cm2 with a scan rate of 15 mV s-1 (Figure 5b). The value for k1 is 2 orders of magnitude lower than that reported previously.8 In that study, the mediated oxidation of ascorbic acid by FcMeOH in a glycine buffer (pH 4) at a GC electrode was described by the following scheme: kf

R - e- h O Z + O 98 R

(24)

A pseudo-first-order rate constant, evaluated from the dc limiting catalytic current, was determined and plotted versus (52) Cannes, C.; Kanoufi, F.; Bard, A. J. J. Electroanal. Chem. 2003, 547, 83. (53) Roy, P. R.; Saha, M. S.; Okajima, T.; Ohsaka, T. Electrochim. Acta 2006, 51, 4447. (54) Manzanares, M. I.; Solis, V.; de.Rossi, R. H. J. Electroanal. Chem. 1996, 407, 141.

ascorbic acid concentration to yield the second-order homogeneous rate constant value of k ) 7.36 × 105 M-1 s-1. The higher value than found in this study for k1 by a full simulation may in part be due to the different form of data analysis, mechanism, and pH used. In particular, correction for the contribution to the measured total current from the FcMeOH0/+ process, or direct oxidation of ascorbic acid, represents a difficulty under conditions used in ref 8, but not in simulations relevant to data obtained at an ITO electrode. Large-Amplitude Fourier Transformed ac Voltammetry for the FcMeOH-Mediated Oxidation of Ascorbic Acid. As outlined in the theoretical section, the presence of quasi-reversibility for the FcMeOH0/+ process as found in the ITO electrode case is predicted to have a significant impact on ac components of a catalytic experiment. To test this hypothesis, theory versus experiment comparisons were carried out at electrode surfaces where the FcMeOH0/+ process was fully reversible (GC) and quasi-reversible (ITO). Figure 6 shows dc and ac harmonics obtained from a FT largeamplitude ac voltammetric experiment for the FcMeOH0/+ redox response in the absence and presence of 1 mM ascorbic acid at a GC electrode (dc scan rate 107.29 mV s-1, f ) 21.46 Hz, and ∆E ) 80 mV for the sine wave perturbation). Clearly, the magnitude of the dc component is increased on addition of ascorbic acid. However, as predicted theoretically, the ac harmonic components remain almost completely unaffected by (55) Marian, I. O.; Sandulescu, R.; Bonciocat, N. J. Pharm. Biomed. Anal. 2000, 23, 227.


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Figure 8. Large-amplitude dc and fundamental to fourth (a-e) harmonic Fourier transformed ac cyclic voltammograms from the oxidation of 1 mM FcMeOH in 0.1 M acetate buffer (pH 5.0) containing 0.1 M LiClO4 over the potential range of -200 to 650 mV. Conditions employed: f ) 21.46 Hz, ∆E ) 150 mV, and ν ) 126.66 mV s-1. Also shown are simulated harmonic components (O) with parameters: v ) 126.66 mV s-1, f ) 21.46 Hz, ∆E ) 150 mV, A ) 0.048 cm2, cFcMeOH ) 1.0 mM, cHA ) 10 mM, DFcMeOH ) 7.6 × 10-6 cm2 s-1, DHA ) 7 × 10-6 cm2 s-1 (D for oxidized and reduced forms assumed equal), R ) 0.55, k0′ ) 0.006 cm s-1, E°′ ) 0.19 V, Ru ) 250 Ω, Cdl ) 20 µF cm-2, and T ) 293 K.

the presence of ascorbic acid (Figure 6). The small difference observed for the oxidation process in the fundamental and second harmonic ac cyclic voltammograms may be due to a residual response derived from direct oxidation of ascorbic acid itself (Figure 6). Ac cyclic voltammograms for fourth and higher harmonics exhibit no measurable current for oxidation of ascorbic acid. This observation emphasizes the effectiveness of the higher harmonics derived from the FT technique in achieving kinetic discrimination between overlapping reversible and irreversible processes. Analogous resolution was achieved previously for the reversible [Ru(NH3)6]3+/2+ process, which was unperturbed by the presence of the irreversible oxygen reduction reaction in the higher ac harmonics.44 With an ITO electrode, no significant ac response is detected in any harmonic for ascorbic acid oxidation in the potential range of the FcMeOH0/+ process. Consequently, the mediated electrocatalytic oxidation of ascorbic acid by ferrocenemethanol can be studied at this electrode surface by ac methods without the need to correct for the background contribution from residual ascorbic acid oxidation. Figure 7 provides experimental results for the FcMeOH0/+ redox response in the absence and presence of 10 mM ascorbic acid at an ITO electrode (dc sweep rate 126.66 mV s-1, ∆E ) 150 mV, and f ) 21.46 Hz for the sine wave perturbation). The larger amplitude was needed in this case to enable the fourth harmonic component to be detected for the quasi-reversible FcMeOH0/+ process at the ITO electrode. For these slow electron-transfer conditions, the expected enhancement in the dc current is found when 10 mM 6524


ascorbic acid is present. Importantly, the fundamental to fourth ac harmonics now exhibit current enhancement in the presence of ascorbic acid as predicted by theory if the underlying redox process is quasi-reversible (k0′ in the range of 0.001-0.01 cm s-1 (Figure 2)). Comparison of simulated and experimental data for the FcMeOH0/+ process (Figure 8) under FT-ac conditions gave a value of k0′ ) 0.006 cm s-1 and R ) 0.55 at an ITO electrode. The k0′value is consistent with the value determined by simulating dc voltammetric data (Figure 5a), but R is far more precisely measured by the ac technique (dc method is insensitive to R; see above). The data attained at an ITO electrode for the mediated oxidation of 10 mM ascorbic by FcMeOH were compared with simulations (Figure 9), and given the complexity of the mechanism and number of parameters that needed to be included in the simulation, excellent agreement was found by employing the following input parameters: v ) 126.66 mV s-1, f ) 21.46 Hz, ∆E ) 150 mV, A ) 0.048 cm2, cFcMeOH ) 1.0 mM, cHA- ) 10 mM, DFcMeOH = 7.6 × 10-6 cm2 s-1, DHA- = 7 × 10-6 cm2 s-1 (D values for oxidized and reduced forms assumed equal), R ) 0.55, k0′ ) 0.006 cm s-1, E°′ ) 0.19 V, Ru ) 150 Ω, Cdl ) 20 µF cm-2, T ) 293 K, k1 ) 2500 M-1 s-1, and k2 ) 1 × 107 M-1 s-1 (not quantitatively meaningful; see above), where cFcMeOH is the concentration of FcMeOH, cHA- is the concentration of ascorbate anion, and DFcMeOH and DHA- are the diffusion coefficients of FcMeOH and ascorbate anion. Thus, simulation of both dc and ac voltammograms with a rate constant for the irreversible homogeneous

Figure 9. Large-amplitude dc and fundamental to fourth (a--e) harmonic Fourier transformed ac cyclic voltammograms obtained over a potential range of -200 to 650 mV for the oxidation of 1 mM FcMeOH and 1 mM FcMeOH in 0.1 M acetate buffer (pH 5.0) containing 0.1 M LiClO4. Conditions employed: f ) 21.46 Hz, ∆E ) 150 mV, and ν ) 126.66 mV s-1. Also shown are ac harmonic components (red) simulated with parameters: v ) 126.66 mV s-1, f ) 21.46 Hz, ∆E ) 150 mV, A ) 0.048 cm2, cFcMeOH ) 1.0 mM, cHA- ) 10 mM, DFcMeOH ) 7.6 × 10-6 cm2 s-1, DHA- ) 7 × 10-6 cm2 s-1 (D for oxidized and reduced forms assumed equal), k0′ ) 0.006 cm s-1, R ) 0.55, E°′ ) 0.190 V, Ru ) 150 Ω, Cdl ) 20 µF cm-2, T ) 293 K, and k1 ) 2500 M-1 s-1 (and k2 ) 1 × 107 M-1 s-1).

bimolecular reaction k1 of 2500 M-1 s-1 provides excellent agreement with experimental data. CONCLUSIONS The mediated electrocatalytic oxidation of ascorbic acid by ferrocenemethanol at a range of electrode materials has been investigated by dc and large-amplitude Fourier transformed ac voltammetry. Experimental and simulated results are compared to calculate the heterogeneous electron-transfer rate (k0′), chargetransfer coefficient (R), and homogeneous catalytic rate constant (k1). The choice of the electrode material can significantly impact on the reliability of the evaluation of the catalytic reaction scheme determined by dc voltammetry. At pH 5.0 and at GC, Au, and BDD electrodes, ascorbic acid oxidation overlaps significantly with that for the FcMeOH0/+ process, which inhibits facile and accurate determination of the homogeneous second-order chemical rate constant. Under FT-ac conditions, and at these electrode materials, the FcMeOH0/+ process is reversible. Consequently, and in accordance with simulations, the presence of a coupled electrocatalytic reaction does not affect the response detected in the higher ac harmonics. Furthermore, the kinetic discrimination between the overlapping reversible (FcMeOH0/+) and irreversible ascorbic acid oxidation also was demonstrated in the higher harmonics. Employing ITO as an electrode material allowed the second-order homogeneous chemical rate constant to be accurately determined as 2500 M-1 s-1 by dc and FT-ac methods. In the dc case, this was possible at the ITO electrode because

ascorbic acid oxidation occurs at potentials much more positive than for the FcMeOH0/+ process. In the FT-ac case, the quasireversible nature of the FcMeOH0/+ couple at ITO allowed the reaction between ascorbic acid and oxidized form of FcMeOH to be quantitatively evaluated. The Ft-ac technique, when an electrocatalytic reaction is coupled to a quasi-reversible electrontransfer process, gives rise to significant current enhancement and change in shape of the higher ac harmonic components. This form of data analysis may be extended to other solution-based mediated electrocatalytic reactions. ACKNOWLEDGMENT Funding from the Australian Research Council in support of this project is gratefully acknowledged. B.L. expresses her appreciation to the National Center for Genetic Engineering and Biotechnology and King Mongkut’s University of Technology Thonburi in Thailand for provision of a grant that enabled her to work in Australia on this project. SUPPORTING INFORMATION AVAILABLE Figure S1 illustrates the addition of the current magnitudes of individual cyclic voltammograms for the FcMeOH0/+ and ascorbic acid oxidation processes relative to the data obtained when both species are present simultaneously in solution. This material is available free of charge via the Internet at http://pubs.acs.org. Received for review December 13, 2007. Accepted June 14, 2008. AC702531F Analytical Chemistry, Vol. 80, No. 17, September 1, 2008

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Investigation of Mediated Oxidation of Ascorbic Acid by

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