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Dopamine Adsorption at Surface Modified Carbon-Fiber Electrodes Bradley D. Bath,† Heidi B. Martin,† R. Mark Wightman,*,† and Mark R. Anderson*,‡ Department of Chemistry, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-3290, and Department of Chemistry, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061-0212 Received May 7, 2001. In Final Form: August 13, 2001 Fast-scan cyclic voltammetry at high repetition rates was used to characterize adsorptive properties of dopamine (DA) at native and modified carbon-fiber microelectrode surfaces. Disk electrodes were fabricated from Thornel P55 fibers, and cylindrical electrodes, from Thornel T650 fibers. Their surfaces were modified by physisorption of 2,6-anthraquinone disulfonic acid (2,6-AQDS) or chemisorption of 4-carboxyphenyl or catechols. Chemisorption was accomplished via electrochemical reduction of diazonium salts. The degree of DA adsorption and its oxidation kinetics were found to vary for the two types of native carbon fiber electrodes and with the different chemical overlayers on the carbon surfaces. 2,6-AQDS measurably increased DA adsorption and desorption kinetics at P55 disks without a significant change in the measurement sensitivity, the response exhibiting temporal characteristics similar to that for nonadsorbing species. 4-Carboxyphenyl modification accelerated the DA adsorption rate and sensitivity at P55 disks. However, neither 2,6-AQDS nor 4-carboxyphenyl altered the response at T650 cylinders. Chemisorption of catechols decreased the DA detection sensitivity at both P55 disks and T650 cylinders. The results suggest that electrostatic interactions at the electrode interface are crucial to DA adsorption and detection under these conditions.
Introduction Background subtracted, fast-scan cyclic voltammetry has been shown to be a useful technique for the detection of neurotransmitters released from biological cells.1,2 In this technique, cyclic voltammograms are obtained at carbon-fiber microelectrodes at repetition rates approaching 100 Hz. Although the background currents are relatively large, they are stable so that background subtraction is possible. This enables small changes in the concentration of electroactive solution species adjacent to the electrode to be revealed. Each cyclic voltammogram provides an identifier of the compound detected, and the amplitude of the peak current provides a measure of its local concentration. Because physiologically relevant concentration changes of neurotransmitters are quite rapid, subsecond response times are required to follow them in real time. Furthermore, because neurotransmitters are effective at their receptors at nanomolar concentrations, high sensitivity is also required. Recent improvements in the instrumentation and computer control of this technique has allowed quite sensitive detection to be accomplished.3,4 To improve sensitivity and selectivity of measurements with carbon-fiber electrodes, Nafion, a cation exchange membrane, has been coated on the electrode tips,5 but its * To whom correspondence should be addressed. † University of North Carolina at Chapel Hill. ‡ Virginia Polytechnic Institute and State University. (1) Garris, P. A.; Wightman, R. M. In Neuromethods: Voltammetric Methods in Brain Systems; Boulton, A. A., Baker, G. B., Adams, R. N., Eds.; Humana Press: Totowa, N.J., 1995; Vol. 27, Chapter 6. (2) Travis, E. R.; Wightman, R. M. Annu. Rev. Biophys. Biomol. Struct. 1998, 27, 77-103. (3) Michael, D.; Travis, E. R.; Wightman, R. M. Anal. Chem. 1998, 70, 586A-92A. (4) Michael, D. J.; Joseph, J. D.; Kilpatrick, M. R.; Travis, E. R.; Wightman, R. M. Anal. Chem. 1999, 71, 3941-47. (5) Kristensen, E. W.; Wilson, R. L.; Wightman, R. M. Anal. Chem. 1986, 58, 986-88.
use increases the electrode response time. Although this effect can be accounted for by deconvolution techniques,6 unmodified carbon-fiber electrodes have been shown to be highly sensitive themselves.7 Recently, we showed that the sensitivity and response time of unmodified carbonfiber microelectrodes both are affected by the degree of adsorption of DA to the electrode.8 Adsorption was found to be dependent on the surface state of the carbon: high sensitivity as a result of significant surface accumulation of DA was found on carbon-fiber surfaces that had been cleaned by soaking in 2-propanol purified with activated carbon (AC). The extent of adsorption was found to decrease with an increase in repetition frequency of the cyclic voltammograms, but under the same conditions, the temporal response was found to improve. An ideal sensor would exhibit both high sensitivity and short response times. The important role of adsorption in the electrochemistry of organic molecules at carbon electrodes has long been recognized. Carbon electrodes contain many chemical functionalities and structures at their interfaces that can dramatically affect heterogeneous kinetics of redox reactions as well as the adsorption of analytes.9-18 McCreery (6) Kawagoe, K. T.; Garris, P. A.; Wiedemann, D. J.; Wightman, R. M. Neuroscience 1992, 51, 55-64. (7) Cahill, P.; Walker, Q. D.; Finnegan, J. M.; Mickelson, G. E.; Wightman, R. M. Anal. Chem. 1996, 68, 3180-86. (8) Bath, B. D.; Michael, D. J.; Trafton, B. J.; Joseph, J. D.; Runnels, P. L.; Wightman, R. M. Anal. Chem. 2000, 72, 5994-6002. (9) Liu, Y. C.; McCreery, R. L. J. Am. Chem. Soc. 1995, 117, 1125459. (10) Chen, P. H.; Fryling, M. A.; McCreery, R. L. Anal. Chem. 1995, 67, 3115-22. (11) Chen, P. H.; McCreery, R. L. Anal. Chem. 1996, 68, 3958-65. (12) Liu, Y. C.; McCreery, R. L. Anal. Chem. 1997, 69, 2091-97. (13) Ray, K.; McCreery, R. L. Anal. Chem. 1997, 69, 4680-87. (14) Ranganathan, S.; Kuo, T.-C.; McCreery, R. L. Anal. Chem. 1999, 71, 3574-80. (15) Yang, H. H.; McCreery, R. L. Anal. Chem. 1999, 71, 4081-87. (16) DuVall, S. H.; McCreery, R. L. Anal. Chem. 1999, 71, 4594-602.
10.1021/la0106844 CCC: $20.00 © 2001 American Chemical Society Published on Web 10/04/2001
Dopamine Adsorption at Carbon-Fiber Electrodes
and co-workers have shown that the electrochemical characteristics of carbon electrodes, both glassy carbon (GC) and highly ordered pyrolytic graphite (HOPG), are highly sensitive to the type of surface treatment employed before the electrochemical measurement.11,16,18 Indeed, they first demonstrated the improved performance of carbon surfaces previously exposed to AC-purified solvents, the procedure that we later used at carbon-fiber electrodes. However, electrochemical responses at different carbon surfaces treated in the same way may differ. For example, the amine functionality appears to initiate adsorption of DA at carbon fibers treated with AC-purified propanol implicating electrostatic interactions,8 whereas at fractured GC surfaces the adsorption of the catechol portion of DA is most apparent.19 Whether this is due to differences in the surfaces or the experimental design is not presently clear. Recently, DuVall and McCreery reported that modification of carbon electrodes with different quinones, either physisorbed or chemisorbed, significantly reduced adsorption of DA to GC electrodes without altering the heterogeneous kinetics of the electrode reaction or significantly changing the sensitivity of the measurement.18 Chemisorption to carbon surfaces was accomplished via electrochemical reduction of the diazonium salt of the species to be surface attached. This report prompted us to investigate whether similar procedures would be successful in decreasing the response time of microelectrodes to DA while maintaining their sensitivity. Adsorption was previously shown to be the rate-limiting step in DA electrooxidation on subsecond time scales.8 In addition, surface modification allows an experimental test of the relative importance of electrostatic effects as opposed to other interactions that can promote adsorption and subsequent electron transfer. Experimental Section Reagents. Doubly distilled deionized water (Corning Megapure MP-3A) was used to prepare all aqueous solutions. Dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), ascorbic acid (AA-), 4-nitroaniline, 4-aminophenylacetic acid, tetra-n-butylammonium tetrafluoroborate (TBABF4), 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide metho-p-toluenesulfonate (carbodiimide coupling agent), potassium acetate, NaNO2, HBF4, NaCl, NaOH, HEPES, CaCl2, and the sodium salt of 2,6anthraquinone disulfonic acid (2,6-AQDS) were obtained from Sigma-Aldrich, acetonitrile (ACN, HPLC grade) was obtained from Allied Signal, and diethyl ether was obtained from Fisher. All solutions for flow cell experiments were prepared with 20 mM HEPES buffer containing 150 mM NaCl and 1.2 mM CaCl2 and adjusted to pH ) 7.4 using NaOH. 4-Aminophenyl acetic acid was recrystallized from ACNwater, and 2,6-AQDS was recrystallized from water prior to use. 2-Propanol (Fisher) was purified by mixing it with Norit A activated carbon (AC, ICN Biomedicals) and filtering it, a cleaning procedure previously described by McCreery and co-workers;14 this solvent will be referred to as AC-purified propanol. All other chemicals were used as received. Electrode Preparation. Glass-encased T650 cylinder and P55 disk carbon-fiber electrodes were fabricated as (17) Ray, K. R.; McCreery, R. L. J. Electroanal. Chem. 1999, 469, 150-58. (18) DuVall, S. H.; McCreery, R. L. J. Am. Chem. Soc. 2000, 122, 6759-64. (19) Allred, C. D.; McCreery, R. L. Anal. Chem. 1992, 64, 444-48.
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previously described;20 the fibers were aspirated into glass capillaries and pulled on a commercial pipet puller (Narishige). For the cylinders, the protruding fiber was cut to an appropriate length, ∼75 µm. The disks were cut at the glass/fiber interface and epoxied.20 After curing, the disk electrodes were polished at a 45° angle on a diamond-embedded polishing wheel (Sutter Instruments), resulting in an elliptical surface. P55 disks and T650 cylinders were chosen because of their wide use in measuring in vivo neurotransmission events. Thornel P55 fibers are manufactured from a pitch precursor, whereas Thornel T650 fibers are from polyacrylonitrile, resulting in subtle changes in the surface chemistry. In addition, the P55 fibers are larger, ∼10 µm diameter, in comparison to T650 fibers, ∼7 µm diameter. Thus, P55 disks and T650 cylinders had surface areas of ∼10-6 cm2 and 10-4 cm2, respectively. Both types of electrodes were soaked in AC-purified propanol for a minimum of 20 min to yield clean, reproducible surfaces prior to experimental or chemical modification. Electrical contact was made via capacitive coupling with a wire inserted into the back of the electrode capillary, which was backfilled with a solution of 4 M potassium acetate/ 0.15 M NaCl. Prior to use, glassy carbon (GC) disk electrodes (Bioanalytical Systems, 3 mm diameter) were polished and treated with AC-purified propanol according to McCreery’s procedure.14 For XPS measurements, GC (Tokai) was attached to a contact wire, sealed into epoxy, and polished prior to use. Chemical Surface Modification. Carbon fiber surfaces were modified by physisorption of 2,6-AQDS or chemisorption of 4-carboxyphenyl or catechols via diazonium chemistry. Modification of a macroscopic GC disk was conducted in parallel, permitting easy verification of the synthesis procedure. The electrochemical responses of the modified GC electrodes were consistent with literature. 2,6-AQDS was physisorbed by cycling the carbon electrode potential for 20 min at 200 mV/s from +0.40 to -0.60 V vs Ag/AgCl in 0.1 M HClO4/0.010 M 2,6-AQDS. Following this electrochemical treatment, the carbon fiber electrodes were soaked in the 2,6-AQDS solution until testing. The surface coverage was estimated by integrating the charge under the voltammetric surface wave after adsorption onto GC or P55 fiber electrodes. Electrode modification by chemisorption of aryl radicals was accomplished by the reduction of diazonium salts.21,22 Schemes 1 and 2 summarize the overall synthesis routes. The precursor for addition of DOPAC was 4-nitrophenyldiazonium tetrafluoroborate, which was synthesized according to the method of Dunker.23 The crude synthesis product was recrystallized from acetonitrile-diethyl ether prior to electrochemical modification of carbon surfaces. The precursor for addition of 4-carboxyphenyl or DA was 4-carboxyphenyldiazonium tetrafluoroborate, which was synthesized according to the procedure of Bourdillon.21 The crude synthesis product was recrystallized from acetone-diethyl ether and stored over phosphorus pentoxide until being used. Both diazonium salts were chemisorbed by electrochemical reduction of the phenyl-diazonium bond during (20) Kawagoe, K. T.; Zimmerman, J. B.; Wightman, R. M. J. Neurosci. Methods 1993, 48, 225-40. (21) Bourdillon, C.; Delamar, M.; Demaille, C.; Hitmi, R.; Moiroux, J.; Pinson, J. J. Electroanal. Chem. 1992, 336, 113-23. (22) Allongue, P.; Delamar, M.; Desbat, B.; Fagebaume, O.; Hitmi, R.; Pinson, J.; Saveant, J. M. J. Am. Chem. Soc. 1997, 119, 201-07. (23) Dunker, M. F. W.; Starkey, E. B.; Jenkins, G. L. J. Am. Chem. Soc. 1936, 58, 2308-09.
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Bath et al. Scheme 1
Scheme 2
a 10 min electrolysis at -0.8 V vs Ag-Ag+ (EAg-Ag+ ) +0.26 V vs Ag-AgCl) of ACN containing 0.001 M diazonium salt and 0.10 M TBABF4. The product of this electrolytic reduction, an aryl radical (in this case, 4-nitrophenyl or 4-carboxyphenyl), readily chemisorbs to the carbon electrode surface. The cyclic voltammogram (CV) of these diazonium species displays a broad feature on the initial potential scan for the reduction of the phenyl-diazonium bond; this feature is absent or significantly reduced in size on subsequent cycles. This electrochemical behavior is characteristic of deposition of a chemical overlayer on the electrode surface and is consistent with literature on reduction of diazonium salts.21,22,24-26 The surface coverage for 4-nitrophenyl was confirmed by the presence of the surface wave at -1.6 V vs Ag-Ag+ due to the reduction and reoxidation of the confined nitrophenyl. The coverage of the nitrophenyl group was estimated by integrating the charge under the voltammetric reduction peak. 3,4-Dihydroxyphenyl acetic acid (DOPAC) was chemisorbed in a multistep procedure outlined in Scheme 1 and described in the following; the resulting surface group is labeled CAT1. After the electrolytic chemisorption of (24) Kariuki, J. K.; McDermott, M. T. Langmuir 1999, 15, 6534-40. (25) Downard, A. J.; Roddick, A. D.; Bond, A. M. Anal. Chim. Acta 1995, 317, 303-10. (26) Downard, A. J. Electroanalysis 2000, 12, 1085-96.
4-nitrophenyl, the nitrophenyl group was subsequently converted to aniline (Scheme 1) by the 6 e-, 6 H+ reduction in the presence of 90:10 water:ethanol containing 0.10 M KCl.22 The electrode potential was scanned between 0.0 and -1.6 V vs Ag/AgCl for two cycles, with the surface wave for the conversion of the nitro group to the amine absent on the second scan. A 4 e- reduction is also possible, converting the amine to a hydroxylamine group; this incomplete reduction does not prevent the subsequent substitution of DOPAC. DOPAC was attached by soaking the aniline-modified electrode in an acetate buffer solution (pH ) 4.5) containing 0.010 M carbodiimide coupling agent and 0.001 M DOPAC for a minimum of 24 h. An amide bond is formed between the surface-confined amine and the carboxylic acid functionality of the DOPAC. DA was attached to the 4-carboxyphenyl-modified carbon surface using the method of Bourdillon; this is summarized in Scheme 2, giving the surface group labeled CAT2.21 The 4-carboxyphenyl-modified electrode was exposed to acetate buffer (pH ) 4.5) containing 0.010 M carbodiimide coupling agent for 1 h. The electrode was rinsed with acetate buffer, and then placed into another acetate buffer solution (pH ) 4.5) containing 0.001 M DA for a minimum of 24 h. X-ray Photoelectron Spectroscopy. X-ray photoelectron spectra were acquired on native and surfacemodified GC electrodes with a PHI model 550 ESCA/SAM system (Case Western Reserve University), using a monochromatic aluminum X-ray source (2 mm, Al KR, 300 W) at a base pressure below 5 × 10-8 Torr. An X-ray spot with an elliptical area of 0.71 mm2 was used to obtain survey (10 min) and high-resolution multiplex (30 min) spectra. The binding energy scale was calibrated with respect to the C1s binding energy of 284.6 ( 0.1 eV. Electrochemical Measurements. Voltammetry and electrolysis during the surface modification procedures were applied using a Pine RDE4 potentiostat. Carbon electrodes were positioned in a custom built three-electrode cell. Solutions were purged prior to electrochemical experiments with nitrogen saturated with the background solvent. The carbon-fiber microelectrodes were evaluated in a flow injection system as previously described.5 An electrode is positioned at the outlet of a six-port HPLC loop injector mounted on a two-position actuator (Rheodyne model 5041 valve and 5701 actuator). The loop injector introduces a bolus of electroactive substance to the electrode surface. Solvent flow is gravity driven through the valve into the electrochemical cell. All carbon fiber electrodes were cycled
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Langmuir, Vol. 17, No. 22, 2001 7035
electrode surface. The response to DA electrooxidation in Figure 1 has been converted to the surface coverage of DA (ΓDA) using the area under the oxidative wave, for the purpose of comparing it to an adsorption model. The rate-limiting adsorption/desorption of DA is characterized by eq 1:8 k1
-2e
k-2
DAsoln {\ } DAads {\} DOQads {\ } DOQsoln k k -1
2
(1)
In eq 1, k1, k-1, k2, and k-2 are the rate constants of adsorption and desorption and the subscripts “soln” and “ads” represent the solution and adsorbed concentrations of DA and the orthoquinone form (DOQ), respectively. To quantify the adsorption/desorption rate-limiting behavior, eq 1 can be modeled as a set of first-order reactions; this model has been presented in ref 8 and is summarized here. The corresponding rate equations are integrated and an infinite series is formed describing the dynamics of DAads (ΓDA) during DA exposure, eq 2:8
ΓDA )
Figure 1. Dynamic response of DA coverage (ΓDA) at (a) a P55 disk and (b) a T650 cylinder to 1.75 s exposures of (a) 2.5 µM DA and (b) 1.0 µM DA as a function of CV repetition rate. The waveform is applied at 300 V/s. The solid lines represent best fits of the kinetic model to the data. Fitted kinetic parameters: (a) k1 ) k2 ) 0.84 × 10-2 cm/s, k-1 ) 5.0 s-1, k-2 ) 21 s-1 and (b) k1 ) k2 ) 2.3 × 10-2 cm/s, k-1 ) 4.0 s-1, k-2 ) 23 s-1. From the voltammograms (DA oxidation and reduction), ∆Ep ) 0.73 and 0.64 V at the P55 disk and T650 cylinder, respectively. The analytes were in a pH ) 7.4 buffer solution consisting of (mM) NaCl (150); HEPES (20); and CaCl2 (1.2).
in flowing buffer for 30 min prior to experimentation to ensure a steady-state background response. The data acquisition system hardware and software (written in LabVIEW) for fast-scan CV have been previously described.4 Computer generated waveforms were input into a patch clamp integrating amplifier (Axopatch model 200B) for application to the electrochemical cell. Signals were digitally acquired through a PCI-6110E acquisition board (National Instruments). A PC-TIO-10 timing board (National Instruments) synchronized flow injection, data acquisition, and waveform application events. Experimental data was imported into Transform (Fortner Software LLC, Sterling, VA) to perform background subtraction. Results Native Carbon Fiber Responses in the Flow Cell. Figure 1 shows the voltammetric responses at P55 disk (a) and T650 cylinder (b) electrodes to transient exposure of DA at a 300 V/s scan rate for various CV repetition frequencies. The voltammograms for DA (inset, Figure 1) displayed capacitive current previously attributed to DA adsorption. This is seen on the oxidative sweep between 0.0 and -0.4 V vs Ag/AgCl as positive current.8 In contrast, the temporal response for the nonadsorbing species, AA(shown in Figure 1 of ref 8), displays a faster, relatively square response as expected for a bolus injection to the
[
k1[DA] (ek-1τ - 1) + k-1 k2[DOQ] (ek-2ts - 1)(ek-1τ) k-2
]∑ m
e-n[k-1τ + k-2ts] (2)
n)1
where each term, n, corresponds to one complete voltammetric cycle, [DA] and [DOQ] represent bulk solution concentrations, and τ and ts represent the two time domains during which either DA (τ) or DOQ (ts) is the dominant species present at the electrode surface; zero time is set arbitrarily as the initial time of DA injection. The model assumes that DOQ is present on the electrode surface for 60% of the voltammetric scan. For example, a 300 V/s 10 Hz waveform would have ts ) 0.006 s and τ ) 0.094 s. Similarly, at the end of the DA injection, [DA] ) 0 and the adsorption/desorption rate-limiting behavior is described by eq 3:
ΓDA ) ΓFDAe-n[k-1τ + k-2ts]
(3)
where n is the cycle number and ΓFDA is the coverage of DA at the end of DA exposure (e.g., 1.75 s and the corresponding voltammograms in Figure 1).8 In Figure 1, the solid lines represent the best fit of eqs 2 and 3 to each data set; that is, one set of parameters simulates the data in Figure 1a and another set simulates the data in Figure 1b. Each fit was obtained by setting k1 ) k2 (i.e., equal rate constants for DA and DOQ adsorption) and then adjusting k-1 and k-2 (Table 1). Both fits were obtained with k-1 < k-2, i.e., with a faster rate of DOQ desorption than for DA. The model predicts the observed decrease in ΓDA that occurs with increased CV repetition frequency. The ΓDA decreases with increasing repetition rate (decreasing τ) because DOQ is generated more frequently and, thus, can desorb for a greater percentage of the time of electrode exposure to DA. The goodness of fit of the model decreases as the CV repetition frequency decreases; at 5 Hz (Figure 1), the model does not predict the experimentally observed slow rise to the maximum ΓDA. The calculated desorption rate constants (k-1 and k-2) are identical at the two types of carbon fibers. However, the adsorption rate constants (k1 and k2) are ∼3 times larger at T650 cylinders, suggesting that DA adsorption sites are either larger in number or stronger toward DA
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Bath et al.
Table 1. ∆Ep, S/N, and Adsorption/Desorption Kinetics at Carbon Fiber Microelectrodesa
a
k1 (cm/s) (calculated)
k-1 (s-1) (calculated)
k-2 (s-1) (calculated)
80 ( 20 170 ( 80 20 ( 10 190 ( 20
4.6 ( 0.3 10 ( 1 0.5 ( 0.2 3.8 ( 0.2
30 ( 8 37 ( 18 3.0 ( 0.5 31 ( 9
T650 Cylinders 230 ( 60 230 ( 20 280 ( 130 200 ( 30 100 ( 10 40 ( 20 240 ( 70 210 ( 50
4.4 ( 0.6 4.3 ( 0.2 0.9 ( 0.4 3.6 ( 1.1
30 ( 2 24 ( 3 11 ( 3 28 ( 9
∆Ep (V) (measured)
S/N (measured)
unmodified 2,6-AQDS CAT1 4-carboxyphenyl
0.72 ( 0.02 0.72 ( 0.02 0.71 ( 0.03 0.65 ( 0.03
P55 Disks 60 ( 20 40 ( 20 19 ( 9 160 ( 30
unmodified 2,6-AQDS CAT1 4-carboxyphenyl
0.65 ( 0.02 0.71 ( 0.01 0.81 ( 0.03 0.55 ( 0.3
The value of k2 was set equal to k1 in all simulations. Voltammograms were taken at 300 V/sec. n ) 5 for each entry.
Figure 2. 2,6-AQDS modified P55 disks. (a) ip response to 1.75 s injections of either 2.5 µM DA (scale ) -0.1 nA) or 200 µM AA- (scale ) -0.05 nA). The voltammogram is taken from the data over the plateau of ip for DA. (b) ΓDA as a function of CV repetition rate. The solid lines represent the best fits of the model to the data. k1 ) k2 ) 1.3 × 10-2 cm/s, k-1 ) 11 s-1, and k-2 ) 23 s-1.
adsorption (Table 1). The DA voltammograms exhibit a greater ∆Ep at P55 disks than at T650 cylinders (Table 1). It is unclear whether this is due to the increased adsorption at the cylindrical electrodes or to faster electron-transfer kinetics. Voltammetric Response of Physisorbed 2,6-AQDS. 2,6-AQDS was physisorbed onto the P55 carbon-fiber disk electrodes. The surface coverage for adsorption of 2,6AQDS was 42 (( 2) pmol/cm2 on GC and 80 (( 7) pmol/cm2 on P55 fibers. The DA adsorption/desorption rate constants were determined as in Figure 1, where the DA surface coverages (ΓDA) were calculated from voltammetric data at varying repetition rates, Figure 2b. As shown in Figure 2a, the ip temporal responses of a 2,6-AQDS-modified P55 disk to injections of AA- or DA were quite similar. Some
DA adsorption was still apparent because the responses were sensitive to CV repetition rate, although the variation was not nearly as significant as that for the unmodified disk electrodes (Figure 1). The S/N also remained nominally the same as that for the unmodified electrode. The adsorption rate constants (k1 and k2) approximately doubled (Table 1) compared to those measured at unmodified P55 disks. In addition, the DA desorption rate constant (k-1) doubled, whereas the DOQ desorption rate constant (k-2) remained unchanged. The ∆Ep for DA at 2,6-AQDS-modified P55 electrodes was also unchanged from the unmodified electrode. Thus, the 2,6-AQDSmodified disks exhibited faster adsorption/desorption kinetics compared with the unmodified P55 surfaces while maintaining similar sensitivity. Interestingly, during the 30 min conditioning period prior to DA measurements, 2,6-AQDS-modified P55 electrodes showed significant changes in background current prior to achieving a steady-state response, suggesting that some 2,6-AQDS desorbed. Yet, the DA adsorption/desorption kinetics increased at these modified electrodes, suggesting that the remaining adsorbed 2,6AQDS occupied and thus blocked surface sites that were responsible for slower kinetics at the unmodified carbon surfaces. 2,6-AQDS was also adsorbed to T650 cylinder electrodes. In contrast to P55 disks, the DA adsorption/desorption kinetics at 2,6-AQDS-modified T650 cylinders were unchanged from the responses at native surfaces, Table 1; the voltammograms and surface coverage plots are not shown. 2,6-AQDS adsorption did, however, increase the value of peak separation (∆Ep). Modification by Chemisorption: XPS Analysis. To achieve a more robust surface modification, reagents were attached to the surface via the reduction of diazonium salts. Three modifications were made. In the first, one kind of catechol functionality was attached to the surface by coupling DOPAC to the surface (labeled as CAT1, Scheme 1). In the second approach, a carboxyl group was attached (Scheme 2), and in the third approach, DA was attached to the carboxyl group to give a second type of catechol functionality (labeled as CAT2, Scheme 2). Because the coupling of DOPAC employed a nitrogencontaining reagent, the progress of Scheme 1 could be followed by XPS. The XPS spectra were obtained on GC electrodes and are assumed to correlate with the chemical changes on the surface-modified carbon fibers, because both the GC and fibers underwent identical surface modification steps and displayed comparable electrochemical responses. Polished GC displays a negligible nitrogen N1s peak at 400 eV ((0.5 eV) (N/C ∼ 0, O/C ) 0.14, not shown); the GC electrodes used for this study were prepared with epoxy and display a larger peak at 400 eV and higher baseline
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Langmuir, Vol. 17, No. 22, 2001 7037
Figure 3. N1s region of XPS spectra for surface-modified GC electrodes, consistent with the Scheme 1 modification process; N/C and O/C ratios are also indicated. The N1s peaks at 406, 402, and 400 eV change in relative intensity as (i) nitrophenyl groups are converted to (ii) aniline groups and then (iii) DOPAC is added to give CAT1. Additional C1s peaks are measured as DOPAC is added.
O/C ratio due to the epoxy (for cured epoxy, N/C ) 0.04 and O/C ) 0.35). Upon addition of the nitrophenyl group (Scheme 1), a major nitrogen peak at 406 eV and small nitrogen peak at 402 eV appear (Figure 3, spectrum i), consistent with the literature.18,27 The N/C ratio increases accordingly. In the carbon C1s region (not shown), a shoulder develops around 286 eV, typical of aromatic groups. After formation of the aniline (Scheme 1), a small N1s peak remains at 406 eV, from incomplete conversion of the nitrophenyl group (Figure 3, spectrum ii); the coupled peak at 402 eV is barely detectable. The major N1s peak is at 400 eV (Figure 1, spectrum ii), for N-H bonds of aniline.22 The O/C ratio is lowered by electroreduction of the surface, consistent with the loss of N-O bonds. The overall nitrogen level (N/C, Figure 3, spectra i and ii) appears comparable for the nitrophenyl and aniline surfaces, suggesting that negligible groups were lost from the surface during the electroreduction. After DOPAC addition, the nitrogen peaks (Figure 3, spectrum iii) remain consistent with those for aniline but with a smaller N1s peak at 400 eV, suggestive of substitution of half of the N-H groups (i.e., NH2 to NHR, R ) DOPAC). The N/C ratio (Figure 3, spectrum iii) is also nominally half as large as for aniline, because of the increased number of carbon atoms (2.3 times) by addition of DOPAC. The O/C ratio is higher with the addition of hydroxyl and carbonyl groups. For comparison, the spectrum (not shown) for attached DA (CAT2, Scheme 2), displayed a N1s peak at 400 eV, with N/C ) 0.10 and O/C ) 0.21; no peak at 406 eV was visible, because 4-carboxyphenyl was the intermediate instead of nitrophenyl. The nitrobenzene surface coverage, through voltammetric measurement, was found to be 1.6 × 10-9 mol/cm2, consistent with values reported by Saveant, although higher than values reported by McCreery.11,18,21,22 Voltammetric Responses of Chemisorbed Modified Surfaces. The ip dynamics at CAT1-modified (Scheme 1) P55 (a) and T650 (b) electrodes for injections of AAand DA shown in Figure 4, display several interesting features. First, the AA- and DA responses are slower at both P55 and T650 electrodes compared to those of the (27) Saby, C.; Ortiz, B.; Champagne, G. Y.; Belanger, D. Langmuir 1997, 13, 6805-13.
Figure 4. CAT1-modified carbon fiber electrodes. (a) ip response at a P55 disk to injections of either 200 µM AA- (scale ) -0.05 nA) or 10 µM DA (scale ) -0.2 nA). (b) ip response at a T650 cylinder to injections of either 100 µM AA- (scale ) -0.8 nA) or 1.0 µM DA (scale ) -1.0 nA). The voltammograms are taken at 1.75 s in each plot.
unmodified surfaces. In Table 1, adsorption/desorption rate constants (k1, k-1, k2, and k-2) are considerably smaller at both CAT1-modified electrodes, as obtained from fitting eqs 2 and 3 to the electrode responses at varying CV repetition rates, similar to Figure 1. ∆Ep is nominally unchanged at CAT1-modified P55 electrodes but was larger at CAT1-modified T650 electrodes. These results are consistent with an interfacial chemical layer on the electrode blocking access to the electrode surface for the solution species. The shape of the DA voltammogram also indicates that far less DA adsorption occurs; no capacitive current from DA adsorption is observed between 0.0 and -0.4 V, as was seen in Figure 1 for the unmodified electrode and as reported in ref 8. The sensitivity also is much lower. The second modification of P55 disk electrodes with 4-carboxyphenyl (Scheme 2) accelerated the DA adsorption kinetics without altering the desorption kinetics (Table 1 and Figure 5), giving a net increase in adsorption. The increased adsorption translated into an increased S/N ratio compared to unmodified electrodes. At 4-carboxyphenylmodified T650 cylindrical electrodes, the rate constants were not changed. However, the modification did decrease ∆Ep at both P55 and T650 electrodes. No detection of DA or AA- was evident on 4-carboxyphenyl-modified P55 disks and cylinders that were further modified with DA to form CAT2, Scheme 2 (data not shown). The surface appeared completely blocked with respect to the approach of the test analytes. The background surface waves corresponding to the oxidation of the bound DA were not observed at high scan rates. This
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Figure 5. 4-Carboxyphenyl-modified carbon fiber electrodes. Dynamic response of the oxidative peak current (ip) at (a) a P55 disk and (b) a T650 cylinder to 1.75 s exposures of (a) 200 µM AA- (scale ) -0.125 nA) and 2.5 µM DA (scale ) -0.5 nA) or (b) 100 µM AA- (scale ) -1.25 nA) and 1.0 µM DA (scale ) -5.0 nA) in the flow cell. A 300 V/s voltammetric waveform is applied at 10 Hz. The voltammograms shown (DA oxidation and reduction) are taken at the end of the injection (1.75 s). ∆Ep ) 0.64 and 0.59 V at the P55 disk and T650 cylinder, respectively. The analytes were in a pH ) 7.4 buffer solution consisting of (mM) NaCl (150); HEPES (20); and CaCl2 (1.2).
may be due to inhibited electron transfer across the CAT2 monolayer. Discussion It has long been recognized that electrochemical phenomena at carbon electrode surfaces are dependent on the handling and preparation of the electrode prior to the electrochemical measurement.9-18,28-30 This behavior is due to both the electrode cleanliness and the introduction of specific chemical features at the interface by different electrode treatments. By following the cleaning procedures presented by McCreery,14 we assume that our electrodes are reasonably clean and that differences in behavior following other surface treatments arise from interfacial chemical differences. Indeed, McCreery and co-workers showed that the specific molecular features present at the carbon surface have a significant role in determining the kinetics of heterogeneous reactions.11,12,15,17,30 Selective chemical modification of these individual sites clearly demonstrated the heterogeneity of the carbon surface and the role that these individual features have in the electrochemical response of different classes of electro(28) Kagan, M. R.; McCreery, R. L. Langmuir 1995, 11, 4041-47. (29) Fryling, M. A.; Zhao, J.; McCreery, R. L. Anal. Chem. 1995, 67, 967-75. (30) McCreery, R. L.; Cline, K. K.; McDermott, C. A.; McDermott, M. T. Colloids Surf. A 1994, 93, 211-19.
Bath et al.
chemical reactions.10,11 Given this background, it is not unreasonable to expect that the electrochemical behavior of carbon-fiber microelectrodes will depend on the method of manufacture of the fiber and the electrode treatment prior to experimentation. The expected heterogeneity of surface sites at the interface of the carbon-fiber electrode and the electrolyte leads to complex adsorption phenomena, involving a distribution of interaction strengths between each type of surface site and adsorbed DA. The behavior of the 2,6AQDS-modified P55 electrode during preconditioning, when the background current changed dramatically, supports this view. We suggest that, during this cycling, the physisorbed 2,6-AQDS achieves a new, lower equilibrium coverage, related to the switching of electrolytes (that is, 2,6-AQDS was initially adsorbed from 0.10 M HClO4, whereas DA measurements were made in pH 7.4 buffer containing 0.10 M NaCl). The 2,6-AQDS most likely to desorb is that more weakly held to the carbon surface. Thus, once equilibrium is established (i.e., when the background current stabilizes), the strongly held 2,6AQDS remains on the surface, and only the weakeradsorbing carbon surface sites remains exposed for DA adsorption. These data appear consistent with those of Ta et al., who observed that 2,6-AQDS preferentially binds to edge plane sites on HOPG electrodes, with the edges being the more electroactive sites.31 In addition, they reported that 2,6-AQDS multilayers are adsorbed from solutions of concentrations exceeding 10 µM. Our physisorption was accomplished from a 0.01 M solution, 1000 times higher than this limit; therefore, some of the 2,6AQDS may have desorbed because of such a multilayer structure. Chemical modification of the electrode may generate a more homogeneous interface, with improved behavior. Indeed, at the 2,6-AQDS-modified P55 disks, DA/DOQ adsorption equilibrium was achieved much faster than was measured at the unmodified electrode, Figure 2 and Table 1. The DA temporal response qualitatively approached the faster response found for AA-, a species that does not adsorb at carbon surfaces, because both the adsorption and desorption rates were accelerated. Thus, the surface sites blocked by 2,6-AQDS appear to be responsible for the slower adsorption kinetics on unmodified electrodes. However, despite displaying the faster kinetics typical of nonadsorbing species, DA adsorption is still apparent: capacitive current for DA adsorption is visible in the CV (Figure 2) and the DA sensitivity remains nominally the same as that for the unmodified surface. The enhancement of adsorption kinetics on P55 disk electrodes by electrostatic interactions, as with the sulfonate anion functionality of 2,6-AQDS, is consistent with our previous finding that the cationic nitrogen on DA is primarily responsible for adsorption to carbon-fiber electrodes at low DAsoln concentrations.8 In contrast to the 2,6-AQDS-modified surface, the 4-carboxyphenyl-modified surface also exhibited an increase in sensitivity because desorption was not accelerated. The nature of the carboxyl group on the modified surface appears to promote stronger adsorption than for the naturally occurring, negative functionalities of disk-polished surfaces. Indeed, the thermodynamic strength of adsorption can be estimated from the slope in the linearized portion of a Langmuir isotherm, previously shown equivalent to the ratio k1/ k-1.8 Thus, the data in Table 1 show that the strength of DA adsorption increases by a factor of ∼2-3 times by (31) Ta, T. C.; Kanda, V.; McDermott, M. T. J. Phys. Chem. B 1999, 103, 1295-302.
Dopamine Adsorption at Carbon-Fiber Electrodes
attaching 4-carboxyphenyl to the P55 disk electrode surface. Consistent with the view that electrostatic attraction enhances DA adsorption, both the CAT1 and the CAT2-modified electrodes, “uncharged” surfaces, displayed less adsorption. Saby et al. have previously shown that electrostatic interactions of 4-carboxyphenyl and 4-nitrophenyl groups on GC affect the heterogeneous kinetics for several electroactive redox species.27 However, in contrast to the P55 disks, physisorbed 2,6AQDS and chemisorbed 4-carboxyphenyl do not impact the DA adsorption/desorption kinetics at T650 cylindrical electrodes. This result is attributed to the differences in the surface chemistry of the carbon structures. Even prior to surface modification, the observed adsorption kinetics of P55 and T650 surfaces are quite different, and modification of the P55 disks increases the adsorption kinetics so that they are only equal to those of the T650 cylinders. These differences cannot be linked solely to inherent differences in surface chemistry or electrode geometry but may also be affected by the fabrication procedure. As part of electrode fabrication, the native P55 disks are physically abraded (polished) prior to soaking in the AC-purified propanol; the native T650 cylinder electrodes are simply treated with the AC-purified propanol after the fiber is sealed in glass. Polishing the P55 disk contributes to cleaning the electrode surface and exposes different chemical functionalities as compared to unpolished carbon surfaces (e.g., the walls and tip of the T650 cylinder). Further studies involving surface abrasion of the T650 cylinders or use of unpolished P55 electrodes are necessary for any further comparison. DuVall and McCreery show that modification of macroscopic carbon electrodes with quinones (e.g., anthraquinone) or catechols eliminates DA adsorption without significantly impacting the heterogeneous kinetics of DA oxidation.18 That does not appear to occur with the 2,6AQDS-modified P55 disk microelectrode, as the shape of the CV (Figure 2) is consistent with oxidation of an adsorbed material: the peak is Gaussian in shape, and a change in the capacitive current from DA adsorption is visible.8 Yet, these different findings may be due to the vast differences in our respective experimental designs; the time scale for our adsorption was
