Enhancing the Analytical Selectivity of Voltammetric Technique by the

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Enhancing the Analytical Selectivity of Voltammetric Technique by the Combination of Harmonic Analysis and “Fingerprint” Phase Angle Lock-in Detection Wenting Chen, Lishi Wang,* Xinjian Huang, and Min Wang College of Chemical Science and Engineering, South China University of Technology, Guangzhou 510641, China ABSTRACT: Traditionally, the selectivity of voltammetric analysis depends on the difference of redox potential. Unfortunately, the limit of discrimination imposed by the voltammogram itself is dozens of millivolts. This suggests that it is impossible to achieve selective detection of one chemical under the interference of chemicals which have very close redox potential with the target chemical. Herein, we provided an attractive solution to this problem, using phase angle instead of potential as the basis of selectivity. Specifically, the electrochemical system was perturbed with a large-amplitude sinusoidal potential signal and the responsive current signal was subsequently analyzed in the frequency domain. This technique was termed as sinusoidal voltammetry (SV). The selective detection can be realized by quantifying the amplitude of a certain harmonic element at the characteristic “fingerprint” phase angle of each redox couple; and their phase angle difference can be regulated to be close to 90° to eliminate interferences and optimize the selective detection. Feasibility of the proposed approach was verified with a model system consisting of two ferrocene derivatives. The underlying theoretical basis was interpreted as that there are inherently several phase angle dramatic transition regions near the redox potential, and thus a minimum redox potential difference can generate a significant phase angle difference in those regions.

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carbon electrodes. Angiogenin was selectively detected at the antiangiogenin/Au electrode surface with square-wave voltammetry recently.11 Besides modified working electrodes, adsorptive voltammetries combined with hanging mercury drop electrodes for selective detection gained renewed interest recently.12,13 Apart from efforts mentioned above, Kuhr et al. have made remarkable progress on electrochemical selective detection. They capitalized on Fourier transformed sinusoidal voltammetry (SV), in which a large-amplitude sine wave potential was applied as the excitation signal and the analysis of the responsive current signal was performed in the frequency domain. According to previous studies, when an electrochemical system was excited with a large-amplitude sine wave (i.e., > 50 mV), faradaic response, which was recognized to be nonlinear, will be distributed to the fundamental frequency as well as higher order harmonics; whereas, capacitive charging current response which was recognized to be linear and be dominate in intensity can be merely distributed at the fundamental harmonic.14−16 As a result, great enhancement of the signal-to-noise ratio was achieved at higher harmonics. Furthermore, a new concept of “fingerprint” phase angle, which was associated with each individual chemical was proposed. By locking in a certain phase angle, chemicals can be selectively and sensitively detected.14 A limit of detection of 8 nM for

lectroanalytical techniques as important one of the major classes of present-day analytical methods are currently gaining popularity due to advantages such as inexpensive instrumentation, rapid determination, and easy miniaturization for small volume samples.1,2 In dozens of widely used electroanalytical techniques, voltammetry is the most important one.3 This technique is most widely utilized to effectively explore the mechanism of the electrode process of redox active molecules, such as the structure of electrical double layer, adsorption phenomenon, as well as electrode process kinetics. Also, voltammetry can be used for rapid quantitative and qualitative determination of extremely small amounts of sample. However, shortcomings of voltammetry associated with reproducibility and selectivity greatly prevented its widespread application in the industrial routine analysis compared with chromatographic, spectroscopic, and other techniques. In the past half century, researchers have been unremittingly endeavoring to solve these two major problems to improve the stability and resolution in electrochemical detections. For improving the reproducibility, attention was largely paid on seeking patterns of preconditioning and materials of the electrode.4−6 In order to enhancing the selectivity, previous research placed a great amount of emphasis on modifying working electrodes. Several examples are as follows. Carbon nanotubes-ionic liquid gel modified glassy carbon electrode was prepared to selectively detect dopamine in the presence of ascorbic acid and uric acid.7 Similar determination was also achieved by graphene/Pt8 and polymer film9,10 modified glassy © 2012 American Chemical Society

Received: June 21, 2012 Accepted: November 27, 2012 Published: November 27, 2012 83

dx.doi.org/10.1021/ac302947w | Anal. Chem. 2013, 85, 83−90

Analytical Chemistry

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glucose was gained using a flow injection analysis apparatus with a Cu microelectrode as the working electrode.15 Kuhr and co-workers achieved selective detection of glucose over maltose by digital locking of the phase angle in the fourth harmonic. In further studies, Kuhr’s group made significant achievements in ultrasensitive and high-selective detections of purine- and pyrimidine-based nucleotides,17 underivatized oligonucleotides and DNA,18 and native amino acids and peptides.19 They even coupled SV to separation methods such as in capillary electrophoresis20 and microfluidic chips.21−23 However, it will be of great importance to clarify the theory and provide the widely applicable experimental guidance. Most favorable selectivity and sensitivity should possibly be achieved if the “fingerprint” phase angle of the interfering agent is close to ±90° out of that of the target analyte.19 Interestingly, we found this condition can be satisfied by adjusting the experiment setup. When Kuhr and co-workers put forward the innovative largeamplitude sinusoidal voltammetry, Bond’s group simultaneously performed similar research. Also, the studies of Kuhr and Bond were consistent in essence. Bond’s group achieved the first completely general simulation of alternating current linear sweep voltammetry and qualitatively as well as quantitatively evaluated the harmonics of large-amplitude ac voltammetry.24 Thereafter, studies were performed in-depth on developing the instrument and data process.25 They successfully applied the Fourier transformed ac voltammetry (ACV) to various electrochemical systems. Resolution of overlapping voltammetric signals for a mixture of uric acid and dopamine was realized by utilizing the fourth or fifth harmonic components.26 Besides, the electrochemical processes of biologically important organic compounds were compared on higher harmonics with high sensitivity.27,28 Recently, Bell et al. provided the theoretical interpretations of large-amplitude sinusoidal voltammetry for reversible redox and ideal surface-confined redox systems.29,30 They derived analytical approximations and new analytical solutions for the current response subjected to large-amplitude sinusoidal voltammetry. They also defined a new experimental protocol for estimating the system parameters, such as the electron transfer coefficient (α) and the kinetic rate constant (k0). The most attractive superiority of large-amplitude sinusoidal voltammetry is its ability of obtaining highly selective detections of redox molecules with similar electrochemical properties based on their unique “fingerprint” phase angle. Previous researchers obtained significant achievements, but the underlying essence and the origin of the “fingerprint” phase angle were discussed only briefly. Relevant work can be traced back to nearly half a century ago, when Smith and co-workers established a general simulation of second harmonic ac polarography and examined the influence of charge transfer kinetic influence on the second harmonic phase angle with the model.31 Thereafter they developed the phase sensitive second harmonic ac polarography and applied it in the Cd2+/Cd (Hg) system.32 Inspired by pioneer works done by Kuhr, Bond, and Smith, we proposed a reasonable explanation to the phenomenon of “fingerprint” phase angle, and this theory may guide us to further improve the selectivity of SV detections. Herein, a series of sine waves with step-up dc bias potentials were continuously applied to the model redox compounds, which were ferrocenecarboxylic acid (FcCOOH) and (dimethylaminomethyl)ferrocene (FcN) in phosphate buffer

solution (PBS). This novel method is named staircase sinusoidal voltammetry (SC-SV). We endeavored to provide direct insight into the nature of the “fingerprint” phase angle by investigating the relationship between the first to fifth harmonic phase angle and the dc bias potential (the center potential) of the applied sine wave in ferrocene derivatives systems. On the basis of the phase angle vs dc bias potential curves, we also aimed to develop a new sinusoidal voltammetry method with enhanced resolution through selecting the favorable experiment conditions, i.e., the optimum dc bias potentials. The phase difference of the fourth harmonic of the two ferrocene derivatives was regulated and controlled to be close to 90° to detect FcCOOH over FcN. Also, the dc bias potential was also set at the harmonic current zero point of FcCOOH to quantify FcN. Under the selected optimum SV experiment conditions, the interference can be markedly nulled out.

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EXPERIMENTAL SECTION Regents. Ferrocenecarboxylic acid (FcCOOH) and (dimethylaminomethyl)ferrocene (FcN) were purchased form Sigma−Aldrich and J&K Scientific (Beijing, China), respectively. To prepare 2.0 mM stock solutions of FcCOOH and FcN, the required amount of each species were first dissolved in 300 μL of ethanol (Guangzhou, China) respectively, and then diluted to 20 mL with 0.1 M phosphate buffer solution (PBS, pH 7.4). It must be noted that the PBS for further dilution to lower concentrations and background acquisition contained the same percentage (1.5%, v/v) of ethanol. All chemicals were of analytical grade and used without further purification. Doubly distilled water was used throughout all experiments. Instrumentation and Experimental Conditions. The whole experiment was organized to be two parts, SC-SV and SV, and performed on a home-built instrument. The instrument is mainly constituted of a data acquisition card (NI USB6251, National Instruments) and a homemade potentiostat. Sine waves of SC-SV and SV were digitally generated (4096 points per cycle) with software written with C++ Builder and applied with National Instrument hardware. The excitation waveform was filtered with a third order low pass filter before being applied to the electrode system in the potentiostat. The current response was also low pass filtered in the potentiostat before A/D conversion. The rate of data acquisition was identical to that of waveform generation, and they were synchronized. Thus, there were 4096 points in each cycle in the obtained current response. Note that special strategies may be taken to avoid spectrum leakage when performing Fourier transformation. Unless otherwise noted, the frequency and amplitude of excitation sine waves in the entire experiment was 3 Hz and 300 mV, respectively. Briefly, SC-SV is an electrochemical method that combined ac voltammetry (ACV) of Bond’s group24 and sinusoidal voltammetry of Kuhr.15 A schematic presentation of the excitation potential waveform and the method of data acquisition are illustrated in Scheme 1. By performing SC-SV experiment, the phase angle vs dc bias potential curves were obtained to reveal the nature of the “fingerprint” phase angle. The amplitude of the dc bias potential step was 2 mV (Estep = 2 mV), the quiet time before the application of sine wave was 5 s, and the number of cycles for each measurement was 8. Both the amplitude and phase angle of the first five harmonics at each dc bias potential step were obtained by fast Fourier transformation (FFT) of the time-domain total current response of the 8 cycles of the sine wave. Thus the first five harmonic phase angle vs dc 84



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three-electrode cell was employed for all electrochemical measurements, with a glassy carbon electrode (GCE) (3 mm diameter) as the working electrode, saturated calomel electrode (SCE) as the reference electrode, and a platinum wire as the auxiliary electrode. Prior to voltammetric experiments, the GCE was polished with 0.3 μm alumina slurry and 0.05 μm alumina slurry and rinsed with ethanol, acetone, and water successively. To enhance the quality of quantitive detection, the background current was subtracted from the entire data set to isolate the information associated with the given analytes. Prior to the SV experiment, a background signal for 8 periods was acquired and Fourier transformed after stabilizing in the same experimental condition for 80 periods. Simply, all the harmonics can be considered as a series of instantaneous current vectors (represented by magnitude and phase angle).19 The instantaneous harmonic currents are the sum of background vector and signal vector. The background signal was digitally subtracted from total instantaneous signal on each harmonic to get the signal only vector. The background current subtraction of CV and DPV was also undertaken to correct the baseline.

Scheme 1. Excitation Waveform and the Method of Data Acquisition Used in Staircase Sinusoidal Voltammetry (SCSV)a

a

The waveform consisted of a series of sine waves with the dc bias potential stepped up or down. At each dc bias potential step, one point of current response and another point of phase angle of each harmonic were recorded.

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RESULTS AND DISCUSSION Electrochemical Behavior of FcCOOH and FcN. Ferrocene derivatives are structurally similar to each other, and they differ only in the substituent on the ferrocene ring, which can eventually result in a slight difference on the halfwave potential (E1/2) of the molecules. Cyclic voltammetries of FcCOOH, FcN, and their equimolar mixture were performed in a solution of PBS at a concentration of 1.0 mM, as shown in Figure 1A. The two ferrocene derivatives displayed one pair of well-defined redox waves which can be attributed to the Fc/Fc+ redox center in the potential scan range of −0.2 to 0.8 V. On the basis of the data of FcCOOH (ΔEp = 68 mV, E1/2 = 0.285 V, ipa/ipc ≈ 1) and FcN (ΔEp = 65 mV, E1/2 = 0.325 V, ipa/ipc ≈ 1), the electron transfer process of the two ferrocene derivatives can be recognized to be a one electron transfer reversible process. Generally, it is better to express the redox potential by E1/2 than by Epa or Epc,33 and the difference of E1/2 between FcN and FcCOOH was only 40 mV. As shown by the CV data of a 1.0 mM equimolar mixture (ΔEp = 80 mV, E1/2 = 0.306 V, ipa/ipc ≈ 1), the current peaks of the two ferrocenes were fully

bias potential and magnitude vs dc bias potential curves of FcCOOH and FcN were obtained. In the part of SV experiments for selective detection, mixed solutions of the two ferrocenes were perturbed by the excitation sine wave with the same frequency and amplitude as that of in SC-SV, and the dc bias potential was optimized to selectively detect the two species with similar electrochemical properties. Details associated with the selection of the optimum dc bias potential are discussed in the Results and Discussion below. By performing the digital phase lock-in technique, the instantaneous fourth harmonic current of the mixture was monitored at the optimum phase angle for the signals of interest, thus the fourth harmonic signal of desired ferrocene was optimized and the interferential one was also depressed.19 In order to compare the electrochemical behavior of FcCOOH and FcN and their mixture, as well as check the stability of the system, cyclic voltammetry (CV) and differential pulse voltammetry (DPV) were performed via a CHI660B electrochemical workstation (Shanghai, China). A conventional

Figure 1. (A) Background-subtracted CVs of 1.0 mM FcCOOH (black line), 1.0 mM FcN (red line), and 1.0 mM equimolar mixture solution (blue line) at GCE in 0.1 M PBS (pH 7.4), at a scan rate of 50 mV·s−1. (B) Background-subtracted DPVs of 25 μM FcCOOH (black line), 25 μM FcN (red line), and 25 μM equimolar mixture solution (blue line) at GCE in 0.1 M PBS (pH 7.4). 85



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Figure 2. Fundamental to the fifth (A−E) harmonic current plots corresponding to the left ordinate for 1.0 mM FcCOOH (black lines) and 1.0 mM FcN (red lines) and phase angle plots corresponding to the right ordinate for 1.0 mM FcCOOH (black dotted lines) and 1.0 mM FcN (red dotted lines), and fundamental to the fifth harmonic current plots (F1) and phase angle plots (F2) of a 1.0 mM equimolar mixture solution obtained by SCSV (in 0.1 M PBS, pH 7.4); with parameters: ΔE = 300 mV; f = 3 Hz; dc bias potential window, −0.2 to 0.8 V; Estep = 2 mV; 8 cycles of sine wave and Fourier transformation were applied at each dc bias potential step.

−0.2 to 0.8 V is shown in Figure 2. Figure 2A−E presents current amplitude vs dc bias potential plots and the corresponding phase angle vs dc bias potential curves of 1.0 mM FcCOOH and 1.0 mM FcN, for the first to fifth harmonic, respectively. The harmonic current amplitude and phase angle curves of the 1.0 mM equimolar mixture solution are showed by Figure 2F1, F2. In this work, the harmonic currents are presented for convenience in an “envelope” format rather than the full current−time version.28 Also, all phase angles are shown between −180 and +180°. In order to enhance the response amplitude of the higher harmonics and obtain more favorable phase angle vs potential curve profile, a relatively high

overlapped with each other. The electrochemical oxidation of 25 μM FcCOOH, 25 μM FcN, and their equimolar mixture solution (25 μM) was investigated by differential pulse voltammetry (DPV) in pH 7.4 PBS. As shown in Figure 1B, the oxidation potential of FcCOOH (0.256 V) was close to the oxidation potential of FcN (0.296 V). Also in the equimolar mixture solution, only one completely overlapped peak (0.276 V) was obtained, indicating that it was impossible to selectively detect each individual chemical in the mixture by traditional dc voltammetry. Nature of “Fingerprint” Phase Angle. The experimental result of SC-SV scanned over the dc bias potential range of 86



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transition (upward or downward). This is the essential cause for the “fingerprint” phase angle lock-in detections. Selective Detection of FcCOOH/FcN. As shown in Figure 1, it is impossible to selectively detect FcCOOH or FcN by traditional dc voltammetry. In order to obtain high resolution and selective detection of FcCOOH and FcN in their mixture, “fingerprint” phase angle was utilized. The sine waves with the optimum dc bias potentials were applied to the mixed solution. By digital locking in the optimum phase angle of FcCOOH and FcN, the selectivity of one analyte over another was accomplished. The exact optimum dc bias potential, which was the pivotal factor, was picked out via the steps as follows. It was woth noting that the phase angle was recognized to be independent of concentration of the analytes,14,36 so that the phase angle data obtained with SC-SV of analytes on the concentration of 1.0 mM can serve as the guidance and criterion for other low concentration detection. As we want to selectively detect FcCOOH or FcN (target) over FcN or FcCOOH (interference), the optimum dc bias potential was obtained with information provided by Figure 2. The dc bias potential was optimized based on the principles of enhancing the target signal and suppressing the interference one. The enhancement of target signal was realized by avoiding the current transition points and making target signal as large as possible. The interference was suppressed through two methods: phase angle difference between the target and the interference is 90°or the dc bias potential is just corresponding to the current transition point of the interference. As shown in Figure 2B−E, the phase angles only differ from each other in the phase angle dramatic jumping regions, i.e., near the current transition points. Near the harmonic current transition points of FcN, the phase angle differences between the two ferrocenes were easily found to be 90°. However, because the phase angle of FcCOOH jumped much more sharp and straight than FcN, the phase angle difference was hardly 90° near the current transition points of FcCOOH (e.g., only could be adjusted to either 144° or 55°). The dc bias potentials, where the phase angle differences were 90°, could be selected and set for the detection of FcCOOH over FcN, but it was not good for detecting FcN in turn, because the current amplitude of FcN was always very small at these dc bias potentials. In order to detect FcN, we selected the dc bias potential where the interfering currents from FcCOOH approached zero. Although the phase angle difference largely deviated from 90°, the interfering current amplitude from FcCOOH was almost zero which might not affect the selective detection. It was worth noting that for the measurement of FcCOOH over FcN, it was very necessary to control the phase angle difference to be 90°, because as displayed by Figure 2B−E, the current of FcN at the transition points was not finished at zero and relatively large, which directly influenced the detection results. In general, the background current is mainly distributed on the first three harmonics;26 and the magnitude of the fourth harmonic current is higher than that of the fifth harmonic. For the purpose of diminishing the influence of the background current and obtaining a relatively large current signal, the work presented here utilized the fourth harmonic. In Figure 2D, when the dc bias potential was from 0.328 to 0.330 V, the phase angle difference of FcCOOH and FcN was from 109 to 88°. For selectively detecting FcCOOH, the optimum dc bias potential might be within 0.328−0.330 V. For detecting FcN over FcCOOH, when the current of FcCOOH was very close to zero and the current of FcN was as large as possible, the dc

concentration of 1.0 mM for the model compounds was chosen, and the applied sine waves had a relatively large amplitude (ΔE = 300 mV), a relatively low frequency (f = 3 Hz), and a small step height of dc bias potential (Estep = 2 mV). The phase angle is relevant with kinetics,19,31 so that it is very favorable to perturb the electrode system with a relative low frequency signal (less than 5 Hz), such as 3 Hz, to access the phase angle vs dc bias potential curve and investigate the nature of the “fingerprint” phase angle. As shown in Figure 2, the currents of the fundamental harmonic component commence and finish at a nonzero value (Figure 2A), whereas the base lines of the second to fifth harmonic currents are close to zero (Figure 2B−E). This implies that background charging current was mainly concentrated in the first harmonic and was not significant relative to the faradaic current in the second to fifth harmonic. Furthermore, the harmonic current response of FcN was larger and more symmetrical than that of FcCOOH, indicating that the level of reversibility of the electrochemical process for FcN is higher than that of FcCOOH. The research of second and higher harmonics manifested that E1/2 corresponds to the potential where the second harmonic current of ac voltammetry was zero with reversible processes.27,34,35 By checking the zero point of the second harmonic current in Figure 2B, the E1/2 of FcCOOH and FcN and their mixture were found to be 0.284, 0.328, and 0.314 V, which were very close to (in agreement with) that found by CV. Comparing the phase angle vs dc bias potential curves with their corresponding harmonic current plots in Figure 2, we can find out that the phase angles of the second to fifth harmonic change rapidly with dc bias potential as the amplitudes of harmonic current approached zero, whereas the phase angles of the fundamental harmonic of FcCOOH, FcN, and their equimolar mixture were all stable around 55°(±10°), Generally, the phase angle will change about 180° in a very narrow potential scope (less than 10 mV) and stay steady before the next phase shift region. Furthermore, the higher the order of the harmonic, the more the phase angle reversal points can be observed. For the specific electrochemical measurement, If the values of E1/2 for FcCOOH and FcN involved in the measurement differ slightly, the phase angle difference of the two redox couple in the higher harmonics will be very remarkable at a dc bias potential near their current transition points. It was interesting that, along with the dc bias potential shift, the phase angle of FcCOOH not only shifted upward but also shifted downward; but it mostly reversed upward in the case of FcN. On the basis of the previous research on second harmonic ac polarography of Smith et al.,31,32 this different phase angle jumping directions between FcCOOH and FcN resulted from the effects of different charge transfer kinetics, which were represented by the charge transfer coefficient (α) and charge transfer rate constant (k0). Although the phase angle jumping directions were different, their phase angles were only different within their dramatic phase angle transition dc bias potential regions (less than 10 mV). In summary, a small difference in E1/2 (even less than 10 mV) for two electroactive molecules can lead to great difference on the phase angle (even more than 100°) in higher harmonics on the same dc bias potential. The E1/2 decides the dc bias potential corresponding to the phase angle transition; and the charge transfer kinetics controls the direction of the phase angle 87



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optimum dc bias potential of 0.328 V for quantifying FcCOOH, the background-subtracted “fingerprint” phase angles of FcCOOH and FcN on the fourth harmonic was −73° and −161°, respectively (Table 1). By digital locking at −73° on the background-subtracted fourth harmonic, the selective detections of FcCOOH were realized. The calibration plots for the determination of FcCOOH in the presence of 25 μM FcN and the detection of 25 μM FcCOOH in the presence of 0 to 50 μM FcN are illustrated in Figure 3A and Table 2A,

bias potential was 0.284 V which was selected as the optimum one. Because of the different experimental conditions between SC-SV and SV, the phase angle obtained in SV might somewhat differ from that obtained with SC-SV (Figure 2D) even if the dc bias potential was identical. To pick out a more favorable dc bias potential for SV measurement of FcCOOH, a series of Fourier transformed SV experiment with the dc bias potential varied from 0.326 to 0.332 V were carried out with 25 μM FcCOOH and 25 μM FcN, respectively. From 0.326 to 0.332 V, the phase angle of FcCOOH already finished the rapid change stage and stayed stable around −71° (±2°), whereas the phase angle of FcN was still in the process of rapid increase. Table 1 shows the phase angle of the background subtracted

Table 2. Detection of FcCOOH/FcN in a Mixture with the Concentration of the Species of Interest Constant at 25 μM, and the Interference Varied from 0 to 50 μMa (A) FcCOOH (25 μM) + FcN (0−50 μM)

Table 1. Fourth Harmonic Phase Angle Measurements with Successive dc Bias Potentials of SV for FcCOOH and FcNa

c

5 10 15 20 30 40 50

phase angle/deg dc bias potential/V

FcCOOH

FcN

phase difference

0.326 0.327 0.328 0.329 0.330 0.331 0.332

−72 −72 −73 −71 −70 −71 −70

−165 −164 −161 −157 −154 −153 −151

93 92 88 86 84 82 81

FcN/μM

I/μA 0.175 0.178 0.186 0.187 0.188 0.189 0.188

RSDb

c

FcCOOH/μM

23.0 23.3 24.4 24.5 24.6 24.7 24.6 2.90%

(B) FcN (25 μM) + FcCOOH (0−50 μM) c

FcCOOH/μM

5 10 15 20 30 40 50

I/μA 0.473 0.477 0.474 0.470 0.468 0.464 0.460

RSDb

c

FcN/μM

22.2 22.4 22.2 22.0 21.8 21.6 21.4 1.64%

a

The data in parts A and B were obtained by Fourier transformed sinusoidal voltammetry (Edc = 0.328 V, ΔE = 300 mV, and f = 3 Hz) and Fourier transformed sinusoidal voltammetry (Edc = 0.284 V, ΔE = 300 mV, and f = 3 Hz) respectively. Columns 2 and 5 display the current signal of 25 μM FcCOOH and 25 μM FcN obtained by digital locking in −73° and 155°, respectively, on the background-subtracted fourth harmonic. Columns 3 and 6 displays the calculated concentration of FcCOOH and FcN by plugging the data in columns 2 and 5 back into the linear regression equations of parts A and B of Figure 3, respectively. bRSDs stand for the relative standard deviations of columns 3 and 6.

a

The phase angle data was background subtracted and obtained by Fourier transformed sinusoidal voltammetry (ΔE = 300 mV and f = 3 Hz) of 25 μM FcCOOH and 25 μM FcN with dc bias potentials from 0.326 to 0.332 V. Column 4 displays the phase angle difference between FcCOOH and FcN.

fourth harmonic data of SVs with successive dc bias potentials. From the table, 0.328 V was selected as the optimum dc bias potential for selective detection of FcCOOH, where the phase angle difference was 88°. The detection of FcCOOH and FcN in a mixture was investigated in two ways: the concentration of the species of interest varied, while the other one was kept constant at 25 μM; the concentration of the species of interest was constant at 25 μM, and the interfering one varied from 0 to 50 μM. At the

respectively. At the optimum dc bias potential of 0.284 V, the background-subtracted “fingerprint” phase angles of 25 μM FcCOOH and 25 μM FcN on the fourth harmonic was determined as 98° and 155°, respectively. Similarly, the detection of FcN over FcCOOH was realized through digital locking in 155° on the fourth harmonic. The linear calibration

Figure 3. (A) Selective detection of FcCOOH (0−50 μM) in the presence of 25 μM FcN in 0.1 M pH 7.4 PBS. Signal was isolated from the background-subtracted fourth harmonic at phase angle = −73°; SV conditions: ΔE = 300 mV, f = 3 Hz, dc bias potential = 0.328 V. (B) Selective detection of FcN (0−50 μM) in the presence of 25 μM FcCOOH in 0.1 M pH 7.4 PBS. Signal was isolated from the background-subtracted fourth harmonic at phase angle = 155°; SV conditions: ΔE = 300 mV, f = 3 Hz, dc bias potential = 0.284 V. 88



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plots for the determination of FcN in the presence of 25 μM FcCOOH and the detection of 25 μM FcN in the presence of 0−50 μM FcCOOH are illustrated in Figure 3B and Table 2B. The digitally locked current magnitude of species of interest was proportional to the concentration. The linear regression equation of FcCOOH was I (μA) = 8.01 × 10−3c (μM) − 8.95 × 10−3 (R = 0.9998); and the equation for FcN was I (μA) = 1.51 × 10−4c (μM) + 0.137 (R = 0.9999). When the interfering species varied from 0 to 50 μM, the signal of 25 μM target one was still constant and well in agreement with the calibration plots of Figure 3. The average value (24.2 μM) of the calculated concentration of FcCOOH by plugging the current data back into the linear regression equation of FcCOOH was very close to 25 μM. The average value (21.9 μM) of the calculated concentration of FcN was slightly smaller than 25 μM. However, the error was acceptable. The relative standard deviations (RSD) of FcCOOH and FcN were 2.90 and 1.64%, respectively. This demonstrates the feasibility and reliability of this selective detection method. By optimizing the dc bias potential for the analyte of interest, it is possible to detect selectively two species with similar electrochemical properties. The reproducibility of the measurements was checked by six repeated experiments for both FcCOOH and FcN. The maximum coefficients of variation in phase angle were ±2 and ±5° for FcCOOH and FcN. The same six tests gave maximum current magnitudes errors of 3.8 and 5.5% for FcCOOH and FcN. The current magnitudes and phase angles for several times of tests were found to be reproducible. In this work, a simple model system was utilized to demonstrate the feasibility of enhancing the selectivity with the proposed method. As for more complex redox system, one should pay attention to the factors which may result in the change of half-wave potentials and discordance between the individual signals and matrix signals, such as the interactions between two components and the adsorption, as it may be more difficult to find the exact optimum potentials. In general, this method is not suitable for the redox systems with severe interaction. Also, one can to some extent solve the problem of adsorption through utilizing suitable electrode materials1,2 and prestabilization. Also, SV is a fast scanning electrochemical technique, and the adsorption is proved to be avoided in the determination of DNA.17,18

also realized by a digital phase lock-in technique at the dc bias potential where the interference signal from FcCOOH was close to zero. Other applications utilizing this method to separate the signal of biologically important organic compounds (sugars and amino acids) transfer across the liquid/ liquid interface are currently underway in our laboratory to investigate the possibilities.

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AUTHOR INFORMATION

Corresponding Author

*Phone: +86 20 87112906. Fax: +86 20 87112906. E-mail: [email protected]. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This work was financially supported by the National Science Foundation of China (Grant No. 21175047).

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REFERENCES

(1) Kato, D.; Sekioka, N.; Ueda, A.; Kurita, R.; Hirono, S.; Suzuki, K.; Niwa, O. Angew. Chem., Int. Ed. 2008, 120, 6783−6786. (2) Kato, D.; Goto, K.; Fujii, S.; Takatsu, A.; Hirono, S.; Niwa, O. Anal. Chem. 2011, 83, 7595−7599. (3) Lubert, K. H.; Kalcher, K. Electroanalysis 2010, 22, 1937−1946. (4) Engstrom, R. C. Anal. Chem. 1982, 54, 2310−2314. (5) Hoogvliet, J. C.; Dijksma, M.; Kamp, B.; Bennekom, W. P. van Anal. Chem. 2000, 72, 2016−2021. (6) Sekioka, N.; Kato, D.; Ueda, A.; Kamata, T.; Kurita, R.; Umemura, S.; Hirono, S.; Niwa, O. Carbon 2008, 46, 1918−1926. (7) Zhao, Y.; Gao, Y.; Zhan, D.; Liu, H.; Zhao, Q.; Kou, Y.; Shao, Y.; Li, M.; Zhuang, Q.; Zhu, Z. Talanta 2005, 66, 51−57. (8) Sun, C. L.; Lee, H. H.; Yang, J. M.; Wu, C. C. Biosens. Bioelectron. 2011, 26, 3450−3455. (9) Ensafi, A. A.; Taei, M.; Khayamian, T. J. Electroanal. Chem. 2009, 633, 212−220. (10) Ensafi, A. A.; Taei, M.; Khayamian, T.; Arabzadeh, A. Sens. Actuators, B: Chem. 2010, 147, 213−221. (11) Li, L.; Zhao, H.; Chen, Z.; Mu, X.; Guo, L. Biosens. Bioelectron. 2011, 30, 261−266. (12) Oliveira, T. M. B. F.; Ribeiro, F. W. P.; Soares, J. E. S.; de LimaNeto, P.; Correia, A. N. Anal. Biochem. 2011, 413, 148−156. (13) da Silva, J. G.; e Silva, M. R. L.; de Oliveira, A. C.; SouzaDe, J. R.; Vaz, C. M. P.; de Castro, C. S. P. J. Food Compos. Anal. 2012, 25, 1−8. (14) Brazill, S. A.; Bender, S. E.; Hebert, N. E.; Cullison, J. K.; Kristensen, E. W.; Kuhr, W. G. J. Electroanal. Chem. 2002, 531, 119− 132. (15) Singhal, P.; Kawagoe, K. T.; Christian, N.; Kuhr, W. G. Anal. Chem. 1997, 69, 1662−1668. (16) Smith, D. E.; Bauer, H. H. Anal. Chem. 1971, 2, 247−343. (17) Singhal, P.; Kuhr, W. G. Anal. Chem. 1997, 69, 3552−3557. (18) Singhal, P.; Kuhr, W. G. Anal. Chem. 1997, 69, 4828−4832. (19) Brazill, S. A.; Singhal, P.; Kuhr, W. G. Anal. Chem. 2000, 72, 5542−5548. (20) Brazill, S. A.; Kuhr, W. G. Anal. Chem. 2002, 74, 3421−3428. (21) Hebert, N. E.; Kuhr, W. G.; Brazill., S. A. Electrophoresis 2002, 23, 3750−3759. (22) Hebert, N. E.; Kuhr, W. G.; Brazill, S. A. Anal. Chem. 2003, 75, 3301−3307. (23) Hebert, N. E.; Snyder, B.; McCreery, R. L.; Kuhr, W. G.; Brazill, S. A. Anal. Chem. 2003, 75, 4265−4271. (24) Gavaghan, D. J.; Bond, A. M. J. Electroanal. Chem. 2000, 480, 133−149. (25) Bond, A. M.; Duffy, N. W.; Guo, S. X.; Zhang, J.; Elton, D. Anal. Chem. 2005, 77, 186 A−195A.

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CONCLUSIONS In this article, direct insight was put to the nature of the “fingerprint” phase angle of different redox compounds in Fourier transformed sinusoidal voltammetry using a model system of FcCOOH and FcN. With dc bias potential as the only independent variable, the phase angle rapidly changes about 180° at the potential of current transition points which was well recognized to be associated with the half-wave potential. Thus, subtle difference of a half-wave potential results in a large phase difference, even up to 100°. Therefore the essential cause of the “fingerprint” phase angle is originating from the subtle difference of the half-wave potential of molecules with similar electrochemical properties and low resolution in traditional dc voltammetry. Through examining the behavior of the current vector (amplitude and phase angle) with respect to potential bias of SV, even structurally similar species can be effectively distinguished from one another. By picking out the optimum dc bias potentials of SV, the phase angle difference of FcCOOH and FcN was regulated to 88° for selectively quantifying FcCOOH; and the detection of FcN was 89



Article

(26) O’Mullane, A. P.; Zhang, J.; Brajter-Toth, A.; Bond, A. M. Anal. Chem. 2008, 80, 4614−4626. (27) Guo, S.; Zhang, J.; Elton, D. M.; Bond, A. M. Anal. Chem. 2004, 76, 166−177. (28) Lee, C. Y.; Bond, A. M. Langmuir 2010, 26, 5243−5253. (29) Bell, C. G.; Anastassiou, C. A.; O’Hare, D.; Parker, K. H.; Siggers, J. H. Electrochim. Acta 2011, 56, 8492−8508. (30) Bell, C. G.; Anastassiou, C. A.; O’Hare, D.; Parker, K. H.; Siggers, J. H. Electrochim. Acta 2011, 56, 7569−7579. (31) McCord, T. G.; Smith, D. E. Anal. Chem. 1968, 40, 289−304. (32) Glover, D. E.; Smith, D. E. Anal. Chem. 1971, 43, 775−779. (33) Batterjee, S. M.; Marzouk, M.; Aazab, M.; El-Hashash, M. Appl. Organomet. Chem. 2003, 17, 291−297. (34) Bond, A. M.; Smith, D. E. Anal. Chem. 1974, 46, 1946−1951. (35) Andrieux, C. P.; Hapiot, P.; Pinson, J.; Savéant, J. M. J. Am. Chem. Soc. 1993, 115, 7783−7788. (36) Brazill, S. A.; Kim, P. H.; Kuhr, W. G. Anal. Chem. 2001, 73, 4882−4890.

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Enhancing the Analytical Selectivity of Voltammetric Technique by the

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