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Electrochemical manipulation of localised pH: application to electroanalysis
Journal of Electroanalytical Chemistry 520 (2002) 13 – 17 www.elsevier.com/locate/jelechem
Electrochemical manipulation of localised pH: application to electroanalysis James Davis a,*, Jonathan M. Cooper b a
b
Department of Chemistry, Uni6ersity of Surrey, Guildford GU2 7XH, UK Department of Electronics and Electrical Engineering, Bioelectronics Research Centre, Uni6ersity of Glasgow, Glasgow G12 8QQ, UK Received 12 August 2001; accepted 5 December 2001
Abstract The electrochemical manipulation of the local pH at a polymer functionalised electrode has been achieved in order to enhance the electrochemical response to cationic analytes. The changes in pH have been shown to provide a method for significantly enhancing the analytical signal towards the model compounds, dopamine and p-aminophenol. The procedure was found to operate irrespective of the electrical properties of the film. The main requirement for this electroanalytical system is that the film contains acidic groups within the polymer backbone. In the carboxylic acid functionalised polypyrrole film studied here, the performance was found to be greatest when the bulk solution pH was less than the pKa of the acid groups. The mechanism attributed to the enhanced response is elucidated and the limitations of the technique are assessed. © 2002 Elsevier Science B.V. All rights reserved. Keywords: Polymer; Dopamine; Pyrrole; Sensor
1. Introduction The detection of catecholamine type neurotransmitters, such as dopamine, can provide a valuable diagnostic aid in a variety of clinical situations [1 – 4] and strategies for their quantitative determination are strongly represented in the analytical literature. The application of electrochemical techniques can offer numerous advantages over conventional chromatographic procedures particularly where real time in vivo monitoring is required [2 – 4]. However, ensuring adequate selectivity and detection limits for the direct analysis of these important compounds whether for in vivo or in vitro measurements without recourse to sample manipulation poses a significant challenge to electroanalytical techniques. To counter this problem, the electrode surface is usually modified either in the form of substrate pre-treatments [5,6], the addition of enzyme layers [7,8], self assembled monolayers [9 – 13], polymer coatings [14 – 21], zeolitic clays [22,23] or various combinations thereof. * Corresponding author. Tel.: + 44-1483-689-596; fax: +44-1483686-851. E-mail address: [email protected] (J. Davis).
Polymeric coatings have received the most attention with anionic functionalities promoting the electrostatic repulsion of common interferents whilst allowing the partitioning of the dopamine analyte within the film. Nafion® is commonly employed in such a role [4,14] but there is an increasing interest in the use of electropolymerisable films [16 –19]. These can offer spatially localised deposition, greater control over film thickness and morphological features which are important factors when considering the increasing application of microfabricated array type sensors. Polypyrrole films functionalised with acidic groups have been used for the selective determination of dopamine and their efficiency at excluding anionic interferents such as ascorbate has been evaluated [16 –18]. The research presented herein has investigated the electrochemically induced pre-concentration of dopamine within a carboxylic acid functionalised pyrrole film and the practicalities of using the technique as a means of further enhancing the electrode response to the analyte. The technique relies upon the electrochemical manipulation of local pH within the polymer such that the removal of protons will lead to the generation of an electrostatic imbalance whose restoration will require
0022-0728/02/$ - see front matter © 2002 Elsevier Science B.V. All rights reserved. PII: S 0 0 2 2 - 0 7 2 8 ( 0 1 ) 0 0 7 4 5 - 8
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J. Da6is, J.M. Cooper / Journal of Electroanalytical Chemistry 520 (2002) 13–17
an influx of cations. Dopamine will constitute a proportion of those ions and hence will add to the analyte already present in the polymer through passive partitioning. This subtle manipulation of local pH can be readily achieved under electrochemical control through the irreversible reduction of protons at the electrode substrate without significantly altering the bulk solution properties. The electroreduction of protons and consequent pH shifts have previously been shown to influence the properties of a bioluminescent protein immobilised within a thin agar film [24] facilitating the reversible modulation of the luminescent output. The intention was to extend this principle to cationic analytes and through building upon the discriminative properties of existing polymer systems it was hoped that a simple and generic method of enhancing the detection of species such as dopamine would emerge. The electrochemical investigations were carried out over a range of pH conditions but, given the potentially vast combination of buffer constituents, two principal cases (pH 3 and pH 7) have been selected in order to illustrate the merits and limitations of the technique. The majority of the work reported herein was conducted at pH 3 as the greater availability of protons provide more latitude for modifying the local pH and hence studying the behaviour of the model analytes within the film.
2. Experimental
2.1. Reagent and materials 3-(Pyrrol-1-yl)propanoic acid (PPA) was prepared from 3-(pyrrol-1-yl)propionitrile using established methods [25]. All reagents with the exception of dopamine hydrochloride (Sigma) were obtained from Aldrich and used without further purification.
2.2. Procedures Electrochemical investigations were conducted using a standard three electrode configuration with a platinum gauze counter electrode (CE) and an Ag AgCl 3 M NaCl reference. Working electrodes (WE) were prepared by the evaporation of gold (Goodfellow, UK) onto photolithographically patterned glass slides prefunctionalised with 3-mercaptoproplytrimethoxysilane. The WE consisted of a gold disk (100 nm thick, 0.03 cm2) which was electrochemically polished by repetitive potential cycling (−0.2–1.2 V at 50 mV s − 1) in 0.5 M perchloric acid and stored in 0.1 M NaCl prior to use. Polymer modified electrodes were prepared by the electropolymerisation of 10 mM PPA from 0.1 M tetraethyl ammonium perchlorate in acetonitrile. The polymerisation was conducted potentiostatically by stepping the potential to 1.15 V for either 60 or 120 s and then stepping down to − 0.1 V for an equal length of time. Non-conducting films were obtained either through the irreversible oxidation of the polymer backbone [16] or as a consequence of prolonged proton reduction with the properties of the resulting films being identical, irrespective of the method employed. Film thickness was measured using a DekTak 3ST surface profiling instrument with a typical thickness of 180 and 450 nm recorded for the polymerisation times of 60 and 120 s respectively. Unless stated otherwise, electrochemical measurements were carried out at pH 3 (0.1 M NaCl) in air saturated solution. Britton– Robinson buffers (equimolar mixtures of acetic, boric and phosphoric acids adjusted to the appropriate pH by the addition of NaOH) were used to investigate the influence of pH on the peak position of film and analyte redox processes. The scan rate in all instances was 50 mV s − 1.
3. Results and discussion
Fig. 1. Cyclic voltammograms detailing the response of a conducting film of poly(PPA) towards 50 mM dopamine in pH 3 solution before (dotted line) and after (dashed line) proton reduction. The voltammogram of the polymer in the absence of dopamine (solid line) is included for reference.
The voltammetric response of a gold electrode coated with a conducting film of poly(PPA) in a pH 3 (0.1 M NaCl) solution is shown in Fig. 1. The electrode was initially swept between −0.2 V and an upper limit of + 0.4 V (thereby preventing irreversible oxidation of the polymer) with the anodic redox process at 0.17 V
J. Da6is, J.M. Cooper / Journal of Electroanalytical Chemistry 520 (2002) 13–17
Fig. 2. Repetitive cyclic voltammograms detailing the relaxation of a conducting poly(PPA) film in a 50 mM dopamine (pH 3) back to its initial state after proton reduction. The voltammograms detail the response before (solid line), during (dotted line) and after (dashed line) proton reduction.
characteristic of film/anion doping processes [16]. The electrode response to 50 mM dopamine is also illustrated in Fig. 1 (dotted line) with its oxidation process observable as a poorly defined shoulder at 0.33 V. Upon extending the scan limit to − 1 V, a large increase in the cathodic current is observed due to the electroreduction of protons. When the electrode potential is cycled back (dashed line) into the positive region the film redox process was found to have moved to a less positive potential (EPPAox 0.08 V, DEPPAox − 90 mV). The dopamine oxidation peak was similarly displaced but significantly enhanced in terms of peak height and definition (the origin of which was confirmed by the addition of increasing dopamine aliquots, data not shown). The shift in the potential of the redox processes can be accounted for by the fact that the irreversible reduction of protons leads to an increase in the local pH. As with most organic species bearing an acid/base functionality, both polymer and analyte will be pH dependent and their redox processes will shift accordingly with exposure to the new environment. This was confirmed by cycling poly(PPA) films in buffer solutions (covering pH 2 to pH 7 at constant ionic strength) with the redox process found to move negatively, with increasing pH (DEPPAox −70 mV/pH unit). Repetitive potential sweeps, especially at slower scan rates where proton reduction is prolonged, resulted in an irreversible degradation in conductivity through the instability of the polypyrrole backbone to the electrogenerated alkaline environment and was found to occur irrespective of whether or not the solutions were degassed. While the repositioning of the film redox pro-
15
cess corresponds to little over one pH unit, the local pH during proton reduction will be significantly higher. The small shift in potential relates to the fact that the composition within the polymer will readjust to the bulk solution conditions upon commencement of the reverse sweep. This is demonstrated in Fig. 2 where the polymer is shown to relax back to its initial state after proton reduction. Three repetitive cyclic sweeps are shown —the polymer response before proton reduction, the enhancement in dopamine oxidation as a consequence of the extended negative sweep and lastly where the lower limit was again restricted to − 0.2 V. This third scan is intermediate between the first two and highlights the transitional nature of the film after proton reduction has ceased. Complete loss of film conductivity does not however prevent utilisation of this technique as a means of enhancing the electrode response to dopamine. This is illustrated in Fig. 3 where a non-conducting poly(PPA) electrode is cycled in the presence of 50 mM dopamine. On the first scan (solid line), the oxidation of dopamine is again observed at 0.33 V and subsequently shifts (EDOPox 0.27 V, DEDOPox − 60 mV) after proton reduction (dashed line). Poising the electrode potential at − 1 V for increasing periods of time (rather than allowing immediate scan reversal) would be expected to increase the magnitude of the potential shift as the increased proton depletion will promote a greater rise in the local pH. This feature is also shown in Fig. 3 (dotted line) where holding the potential at − 1 V for 10 s displaced the oxidation peak position by a further − 30 mV. The increase in the height of the dopamine oxidation peak occurs as a consequence of the electrostatic imbalance created through the removal of the positively
Fig. 3. Response of a non-conducting film of poly(PPA) towards 50 mM dopamine in a solution of pH 3 before (solid line) and after (dashed line) proton reduction. The effect of holding the electrode at −1 V for 10 s on peak position is also illustrated (dotted line).
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charged protons. At pH 3, carboxyl groups within the polymer will largely be protonated (pKa =4.5 [14]) and thus serve as an additional source of reducible protons. The subsequent electroreduction of solution and carboxyl protons leads to the generation of a net negative charge within the polymer. This induces an influx of cations throughout the film in order to maintain electrostatic balance. Dopamine constitutes a proportion of those ions and when the potential is swept back into the positive region there is an increase in the peak height reflecting the increased concentration of dopamine electrostatically attracted into the film. Sweeping the potential to −0.8 V (immediately before proton reduction) presents effectively no change to the initial sweep confirming that proton reduction is primarily responsible for the increase in the subsequent dopamine signal. Corroboration for the pre-concentration mechanism was provided by a number of observations of which the gradual relaxation of the films back to the initial voltammogram has already been described. Catechol (1,2-dihydroxybenzene) is electrostatically neutral at pH 3 and the response of a non-conducting poly(PPA) film to this analyte is shown in Fig. 4. Initial inspection reveals a response similar to that obtained with dopamine with a shift in the oxidation potential (ECATox 0.3 V, DE −40 mV) occurring after proton reduction. However, the peak height remains essentially unchanged. As the analyte is electrostatically neutral there will be no influx of catechol into the film. The peak position is displaced, as the catechol already residing within the film will experience the change in localised pH. Further supportive evidence arises when considering the influence of ionic strength on the peak height enhancement. As the ionic strength is increased the
Fig. 5. Influence of ionic strength on the height of the dopamine (50 mM, pH 3) oxidation peak (at a non-conducting film) after the electroreduction of protons.
Fig. 6. Response comparison of a variety of electrode configurations, conducting and non-conducting (NC), towards the oxidation of dopamine before (BHR) and after (AHR) the proton reduction sweep at pH 3.
Fig. 4. Response of a non-conducting poly(PPA) film towards 50 mM catechol (pH 3) before (solid line) and after proton reduction (dashed line).
dopamine will constitute a decreasing proportion of the matrix cations and a decrease in the height of the oxidation peak would be expected, Fig. 5. Conversely, increasing the concentration of dopamine should increase the peak enhancement upon proton reduction. This can be seen in Fig. 6 where a number of polymer systems are compared with the magnitude of the peak enhancement found to increase with increasing dopamine concentration. Film thickness will directly influence the electrode response to dopamine though there is a more immediate relevance to the conducting polymer systems as the dopamine is oxidised at the polymer rather than simply at the electrode substrate. This can be illustrated if we compare the response of various electrodes (uncoated,
J. Da6is, J.M. Cooper / Journal of Electroanalytical Chemistry 520 (2002) 13–17
thin film, thick film, conducting and non-conducting) to dopamine under identical conditions. The results are summarised in Fig. 6 with the film coated electrodes offering significant increases in the peak current over the bare electrode system. The best response to dopamine was observed with the conducting film system but problems with reproducibility as a consequence of their short lifetime severely restricts their applicability. Exploiting the electroreduction of protons at higher pH values is limited by their availability with alterations in local pH brought about through the reduction of protons attached to either buffer constituents or from water itself. In 10 mM phosphate buffer (pH 7), it is possible to adjust the local pH by holding the electrode at − 1 V in the same way as was achieved at pH 3. It was found that for every additional 15 s the electrode was held at −1 V there is an incremental shift of − 10 mV in the dopamine oxidation peak. Increasing the buffer concentration serves to supply more reducible protons and hence requires less time to promote the shift in pH. However, there is no increase in the height of the dopamine oxidation peak irrespective of the buffering concentration or time employed in the proton reduction. This can be explained by the fact that the carboxylic acid groups within the film will be deprotonated at pH 7 and already loaded with positive counterions. It can be envisaged that any influx of counterions will occur at the periphery of the film rather than at the substrate film interface. This is in contrast to the situation at pH 3 where the electrochemically promoted removal of protons from the carboxylic acid groups on the polymer backbone will necessitate the influx of ions throughout the film. The generic nature of the procedure was briefly assessed by examining the electrode response to paminophenol. This represents another physiologically relevant compound, which displays quasi-reversible electrochemistry and will be cationic at pH 3. Applying similar procedures to those used with dopamine analysis, an enhanced oxidation peak (not shown) was observed after proton reduction that was found to mirror effectively the behaviour observed with dopamine.
4. Conclusions The electrochemical manipulation of the local pH through the electroreduction of protons within an electropolymerised film has been demonstrated and shown to enhance the detection of dopamine significantly. The technique was found to operate irrespective of the electrical properties of the film (conducting or passive) with the main requirement being the possession of an
17
acid or base functionality within the film. In the carboxylic acid functionalised film studied, the performance was found to be greatest when the bulk solution pH was less than the pKa of the acid groups and under situations of moderate ionic strength (0.1 M). The more immediate limitations of the technique have been assessed and the generic nature has been probed through examining the response to an alternative electroactive cation, p-aminophenol.
Acknowledgements The authors are grateful to the European Commission (Contract No. BIO4-CT97-2112) for supporting this research.
References [1] A. Gowenlock (Ed.), Varley’s Practical Clinical Biochemistry, 6th ed., Heinemann, London, 1988, p. 878. [2] R.N. Adams, Anal. Chem. 48 (1976) 1128A. [3] J.A. Stamford, J.B. Justice, Anal. Chem. 68 (1996) 359A. [4] K. Pihel, Q.D. Walker, R.M. Wightman, Anal. Chem. 68 (1996) 2084. [5] R.S. Kelly, D.J. Weiss, S.H. Chong, T. Kuwana, Anal. Chem. 71 (1999) 413. [6] F. Crespi, T.G. England, D.G. Trist, J. Neurosci. Methods 61 (1995) 201. [7] S. Cosnier, C. Innocent, L. Allien, S. Poitry, M. Tsacopoulos, Anal. Chem. 69 (1997) 968. [8] E.S. Forzani, G.A. Rivas, V.M. Solis, J. Electroanal. Chem. 435 (1997) 77. [9] F. Malem, D. Mandler, Anal. Chem. 65 (1993) 37. [10] Z.M. Liu, J.H. Li, S.J. Dong, E.K. Wang, Anal. Chem. 68 (1996) 2432. [11] A. Dalmia, C.C. Liu, R.F. Savinell, J. Electroanal. Chem. 430 (1997) 205. [12] C.R. Raj, T. Ohsaka, J. Electroanal. Chem. 496 (2001) 44. [13] M.A. Chen, H.L. Li, Electroanalysis 10 (1998) 477. [14] D.J. Wiedemann, K.T. Kawagoe, R.T. Kennedy, E.L. Ciolkowski, R.M. Wightman, Anal. Chem. 63 (1991) 2965. [15] J.M. Cooper, D.J. Pritchard, J. Mat. Sci.-Mater. Electron. 5 (1994) 111. [16] D.W.M. Arrigan, Anal. Comm. 34 (1997) 241. [17] J. Wang, P.V.A. Pamidi, G. Cepria, S. Basak, K. Rajeshwar, Analyst 112 (1997) 981. [18] Z.Q. Gao, B.S. Chen, M.X. Zi, Analyst 119 (1994) 459. [19] J.W. Mo, B. Dgorevc, Anal. Chem. 73 (2001) 1196. [20] R.C. Tucker, I. Song, J.H. Payer, R.E. Marchant, J. Appl. Electrochem. 27 (1997) 1079. [21] J. Cruz, M. Kawasaki, W. Gorski, Anal. Chem. 72 (2000) 680. [22] J. Wang, A. Walcarius, J. Electroanal. Chem. 407 (1996) 183. [23] J.M. Zen, P.J. Chen, Anal. Chem. 69 (1997) 5087. [24] R.S. Chittock, A. Glidle, C.W. Wharton, N. Berovic, T.D. Beynon, J.M. Cooper, Anal. Chem. 70 (1998) 4170. [25] C.J. Pickett, K.S. Ryder, J. Chem. Soc. Dalton Trans. (1994) 2181.
Electrochemical manipulation of localised pH: application to electroanalysis James Davis a,*, Jonathan M. Cooper b a
b
Department of Chemistry, Uni6ersity of Surrey, Guildford GU2 7XH, UK Department of Electronics and Electrical Engineering, Bioelectronics Research Centre, Uni6ersity of Glasgow, Glasgow G12 8QQ, UK Received 12 August 2001; accepted 5 December 2001
Abstract The electrochemical manipulation of the local pH at a polymer functionalised electrode has been achieved in order to enhance the electrochemical response to cationic analytes. The changes in pH have been shown to provide a method for significantly enhancing the analytical signal towards the model compounds, dopamine and p-aminophenol. The procedure was found to operate irrespective of the electrical properties of the film. The main requirement for this electroanalytical system is that the film contains acidic groups within the polymer backbone. In the carboxylic acid functionalised polypyrrole film studied here, the performance was found to be greatest when the bulk solution pH was less than the pKa of the acid groups. The mechanism attributed to the enhanced response is elucidated and the limitations of the technique are assessed. © 2002 Elsevier Science B.V. All rights reserved. Keywords: Polymer; Dopamine; Pyrrole; Sensor
1. Introduction The detection of catecholamine type neurotransmitters, such as dopamine, can provide a valuable diagnostic aid in a variety of clinical situations [1 – 4] and strategies for their quantitative determination are strongly represented in the analytical literature. The application of electrochemical techniques can offer numerous advantages over conventional chromatographic procedures particularly where real time in vivo monitoring is required [2 – 4]. However, ensuring adequate selectivity and detection limits for the direct analysis of these important compounds whether for in vivo or in vitro measurements without recourse to sample manipulation poses a significant challenge to electroanalytical techniques. To counter this problem, the electrode surface is usually modified either in the form of substrate pre-treatments [5,6], the addition of enzyme layers [7,8], self assembled monolayers [9 – 13], polymer coatings [14 – 21], zeolitic clays [22,23] or various combinations thereof. * Corresponding author. Tel.: + 44-1483-689-596; fax: +44-1483686-851. E-mail address: [email protected] (J. Davis).
Polymeric coatings have received the most attention with anionic functionalities promoting the electrostatic repulsion of common interferents whilst allowing the partitioning of the dopamine analyte within the film. Nafion® is commonly employed in such a role [4,14] but there is an increasing interest in the use of electropolymerisable films [16 –19]. These can offer spatially localised deposition, greater control over film thickness and morphological features which are important factors when considering the increasing application of microfabricated array type sensors. Polypyrrole films functionalised with acidic groups have been used for the selective determination of dopamine and their efficiency at excluding anionic interferents such as ascorbate has been evaluated [16 –18]. The research presented herein has investigated the electrochemically induced pre-concentration of dopamine within a carboxylic acid functionalised pyrrole film and the practicalities of using the technique as a means of further enhancing the electrode response to the analyte. The technique relies upon the electrochemical manipulation of local pH within the polymer such that the removal of protons will lead to the generation of an electrostatic imbalance whose restoration will require
0022-0728/02/$ - see front matter © 2002 Elsevier Science B.V. All rights reserved. PII: S 0 0 2 2 - 0 7 2 8 ( 0 1 ) 0 0 7 4 5 - 8
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J. Da6is, J.M. Cooper / Journal of Electroanalytical Chemistry 520 (2002) 13–17
an influx of cations. Dopamine will constitute a proportion of those ions and hence will add to the analyte already present in the polymer through passive partitioning. This subtle manipulation of local pH can be readily achieved under electrochemical control through the irreversible reduction of protons at the electrode substrate without significantly altering the bulk solution properties. The electroreduction of protons and consequent pH shifts have previously been shown to influence the properties of a bioluminescent protein immobilised within a thin agar film [24] facilitating the reversible modulation of the luminescent output. The intention was to extend this principle to cationic analytes and through building upon the discriminative properties of existing polymer systems it was hoped that a simple and generic method of enhancing the detection of species such as dopamine would emerge. The electrochemical investigations were carried out over a range of pH conditions but, given the potentially vast combination of buffer constituents, two principal cases (pH 3 and pH 7) have been selected in order to illustrate the merits and limitations of the technique. The majority of the work reported herein was conducted at pH 3 as the greater availability of protons provide more latitude for modifying the local pH and hence studying the behaviour of the model analytes within the film.
2. Experimental
2.1. Reagent and materials 3-(Pyrrol-1-yl)propanoic acid (PPA) was prepared from 3-(pyrrol-1-yl)propionitrile using established methods [25]. All reagents with the exception of dopamine hydrochloride (Sigma) were obtained from Aldrich and used without further purification.
2.2. Procedures Electrochemical investigations were conducted using a standard three electrode configuration with a platinum gauze counter electrode (CE) and an Ag AgCl 3 M NaCl reference. Working electrodes (WE) were prepared by the evaporation of gold (Goodfellow, UK) onto photolithographically patterned glass slides prefunctionalised with 3-mercaptoproplytrimethoxysilane. The WE consisted of a gold disk (100 nm thick, 0.03 cm2) which was electrochemically polished by repetitive potential cycling (−0.2–1.2 V at 50 mV s − 1) in 0.5 M perchloric acid and stored in 0.1 M NaCl prior to use. Polymer modified electrodes were prepared by the electropolymerisation of 10 mM PPA from 0.1 M tetraethyl ammonium perchlorate in acetonitrile. The polymerisation was conducted potentiostatically by stepping the potential to 1.15 V for either 60 or 120 s and then stepping down to − 0.1 V for an equal length of time. Non-conducting films were obtained either through the irreversible oxidation of the polymer backbone [16] or as a consequence of prolonged proton reduction with the properties of the resulting films being identical, irrespective of the method employed. Film thickness was measured using a DekTak 3ST surface profiling instrument with a typical thickness of 180 and 450 nm recorded for the polymerisation times of 60 and 120 s respectively. Unless stated otherwise, electrochemical measurements were carried out at pH 3 (0.1 M NaCl) in air saturated solution. Britton– Robinson buffers (equimolar mixtures of acetic, boric and phosphoric acids adjusted to the appropriate pH by the addition of NaOH) were used to investigate the influence of pH on the peak position of film and analyte redox processes. The scan rate in all instances was 50 mV s − 1.
3. Results and discussion
Fig. 1. Cyclic voltammograms detailing the response of a conducting film of poly(PPA) towards 50 mM dopamine in pH 3 solution before (dotted line) and after (dashed line) proton reduction. The voltammogram of the polymer in the absence of dopamine (solid line) is included for reference.
The voltammetric response of a gold electrode coated with a conducting film of poly(PPA) in a pH 3 (0.1 M NaCl) solution is shown in Fig. 1. The electrode was initially swept between −0.2 V and an upper limit of + 0.4 V (thereby preventing irreversible oxidation of the polymer) with the anodic redox process at 0.17 V
J. Da6is, J.M. Cooper / Journal of Electroanalytical Chemistry 520 (2002) 13–17
Fig. 2. Repetitive cyclic voltammograms detailing the relaxation of a conducting poly(PPA) film in a 50 mM dopamine (pH 3) back to its initial state after proton reduction. The voltammograms detail the response before (solid line), during (dotted line) and after (dashed line) proton reduction.
characteristic of film/anion doping processes [16]. The electrode response to 50 mM dopamine is also illustrated in Fig. 1 (dotted line) with its oxidation process observable as a poorly defined shoulder at 0.33 V. Upon extending the scan limit to − 1 V, a large increase in the cathodic current is observed due to the electroreduction of protons. When the electrode potential is cycled back (dashed line) into the positive region the film redox process was found to have moved to a less positive potential (EPPAox 0.08 V, DEPPAox − 90 mV). The dopamine oxidation peak was similarly displaced but significantly enhanced in terms of peak height and definition (the origin of which was confirmed by the addition of increasing dopamine aliquots, data not shown). The shift in the potential of the redox processes can be accounted for by the fact that the irreversible reduction of protons leads to an increase in the local pH. As with most organic species bearing an acid/base functionality, both polymer and analyte will be pH dependent and their redox processes will shift accordingly with exposure to the new environment. This was confirmed by cycling poly(PPA) films in buffer solutions (covering pH 2 to pH 7 at constant ionic strength) with the redox process found to move negatively, with increasing pH (DEPPAox −70 mV/pH unit). Repetitive potential sweeps, especially at slower scan rates where proton reduction is prolonged, resulted in an irreversible degradation in conductivity through the instability of the polypyrrole backbone to the electrogenerated alkaline environment and was found to occur irrespective of whether or not the solutions were degassed. While the repositioning of the film redox pro-
15
cess corresponds to little over one pH unit, the local pH during proton reduction will be significantly higher. The small shift in potential relates to the fact that the composition within the polymer will readjust to the bulk solution conditions upon commencement of the reverse sweep. This is demonstrated in Fig. 2 where the polymer is shown to relax back to its initial state after proton reduction. Three repetitive cyclic sweeps are shown —the polymer response before proton reduction, the enhancement in dopamine oxidation as a consequence of the extended negative sweep and lastly where the lower limit was again restricted to − 0.2 V. This third scan is intermediate between the first two and highlights the transitional nature of the film after proton reduction has ceased. Complete loss of film conductivity does not however prevent utilisation of this technique as a means of enhancing the electrode response to dopamine. This is illustrated in Fig. 3 where a non-conducting poly(PPA) electrode is cycled in the presence of 50 mM dopamine. On the first scan (solid line), the oxidation of dopamine is again observed at 0.33 V and subsequently shifts (EDOPox 0.27 V, DEDOPox − 60 mV) after proton reduction (dashed line). Poising the electrode potential at − 1 V for increasing periods of time (rather than allowing immediate scan reversal) would be expected to increase the magnitude of the potential shift as the increased proton depletion will promote a greater rise in the local pH. This feature is also shown in Fig. 3 (dotted line) where holding the potential at − 1 V for 10 s displaced the oxidation peak position by a further − 30 mV. The increase in the height of the dopamine oxidation peak occurs as a consequence of the electrostatic imbalance created through the removal of the positively
Fig. 3. Response of a non-conducting film of poly(PPA) towards 50 mM dopamine in a solution of pH 3 before (solid line) and after (dashed line) proton reduction. The effect of holding the electrode at −1 V for 10 s on peak position is also illustrated (dotted line).
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charged protons. At pH 3, carboxyl groups within the polymer will largely be protonated (pKa =4.5 [14]) and thus serve as an additional source of reducible protons. The subsequent electroreduction of solution and carboxyl protons leads to the generation of a net negative charge within the polymer. This induces an influx of cations throughout the film in order to maintain electrostatic balance. Dopamine constitutes a proportion of those ions and when the potential is swept back into the positive region there is an increase in the peak height reflecting the increased concentration of dopamine electrostatically attracted into the film. Sweeping the potential to −0.8 V (immediately before proton reduction) presents effectively no change to the initial sweep confirming that proton reduction is primarily responsible for the increase in the subsequent dopamine signal. Corroboration for the pre-concentration mechanism was provided by a number of observations of which the gradual relaxation of the films back to the initial voltammogram has already been described. Catechol (1,2-dihydroxybenzene) is electrostatically neutral at pH 3 and the response of a non-conducting poly(PPA) film to this analyte is shown in Fig. 4. Initial inspection reveals a response similar to that obtained with dopamine with a shift in the oxidation potential (ECATox 0.3 V, DE −40 mV) occurring after proton reduction. However, the peak height remains essentially unchanged. As the analyte is electrostatically neutral there will be no influx of catechol into the film. The peak position is displaced, as the catechol already residing within the film will experience the change in localised pH. Further supportive evidence arises when considering the influence of ionic strength on the peak height enhancement. As the ionic strength is increased the
Fig. 5. Influence of ionic strength on the height of the dopamine (50 mM, pH 3) oxidation peak (at a non-conducting film) after the electroreduction of protons.
Fig. 6. Response comparison of a variety of electrode configurations, conducting and non-conducting (NC), towards the oxidation of dopamine before (BHR) and after (AHR) the proton reduction sweep at pH 3.
Fig. 4. Response of a non-conducting poly(PPA) film towards 50 mM catechol (pH 3) before (solid line) and after proton reduction (dashed line).
dopamine will constitute a decreasing proportion of the matrix cations and a decrease in the height of the oxidation peak would be expected, Fig. 5. Conversely, increasing the concentration of dopamine should increase the peak enhancement upon proton reduction. This can be seen in Fig. 6 where a number of polymer systems are compared with the magnitude of the peak enhancement found to increase with increasing dopamine concentration. Film thickness will directly influence the electrode response to dopamine though there is a more immediate relevance to the conducting polymer systems as the dopamine is oxidised at the polymer rather than simply at the electrode substrate. This can be illustrated if we compare the response of various electrodes (uncoated,
J. Da6is, J.M. Cooper / Journal of Electroanalytical Chemistry 520 (2002) 13–17
thin film, thick film, conducting and non-conducting) to dopamine under identical conditions. The results are summarised in Fig. 6 with the film coated electrodes offering significant increases in the peak current over the bare electrode system. The best response to dopamine was observed with the conducting film system but problems with reproducibility as a consequence of their short lifetime severely restricts their applicability. Exploiting the electroreduction of protons at higher pH values is limited by their availability with alterations in local pH brought about through the reduction of protons attached to either buffer constituents or from water itself. In 10 mM phosphate buffer (pH 7), it is possible to adjust the local pH by holding the electrode at − 1 V in the same way as was achieved at pH 3. It was found that for every additional 15 s the electrode was held at −1 V there is an incremental shift of − 10 mV in the dopamine oxidation peak. Increasing the buffer concentration serves to supply more reducible protons and hence requires less time to promote the shift in pH. However, there is no increase in the height of the dopamine oxidation peak irrespective of the buffering concentration or time employed in the proton reduction. This can be explained by the fact that the carboxylic acid groups within the film will be deprotonated at pH 7 and already loaded with positive counterions. It can be envisaged that any influx of counterions will occur at the periphery of the film rather than at the substrate film interface. This is in contrast to the situation at pH 3 where the electrochemically promoted removal of protons from the carboxylic acid groups on the polymer backbone will necessitate the influx of ions throughout the film. The generic nature of the procedure was briefly assessed by examining the electrode response to paminophenol. This represents another physiologically relevant compound, which displays quasi-reversible electrochemistry and will be cationic at pH 3. Applying similar procedures to those used with dopamine analysis, an enhanced oxidation peak (not shown) was observed after proton reduction that was found to mirror effectively the behaviour observed with dopamine.
4. Conclusions The electrochemical manipulation of the local pH through the electroreduction of protons within an electropolymerised film has been demonstrated and shown to enhance the detection of dopamine significantly. The technique was found to operate irrespective of the electrical properties of the film (conducting or passive) with the main requirement being the possession of an
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acid or base functionality within the film. In the carboxylic acid functionalised film studied, the performance was found to be greatest when the bulk solution pH was less than the pKa of the acid groups and under situations of moderate ionic strength (0.1 M). The more immediate limitations of the technique have been assessed and the generic nature has been probed through examining the response to an alternative electroactive cation, p-aminophenol.
Acknowledgements The authors are grateful to the European Commission (Contract No. BIO4-CT97-2112) for supporting this research.
References [1] A. Gowenlock (Ed.), Varley’s Practical Clinical Biochemistry, 6th ed., Heinemann, London, 1988, p. 878. [2] R.N. Adams, Anal. Chem. 48 (1976) 1128A. [3] J.A. Stamford, J.B. Justice, Anal. Chem. 68 (1996) 359A. [4] K. Pihel, Q.D. Walker, R.M. Wightman, Anal. Chem. 68 (1996) 2084. [5] R.S. Kelly, D.J. Weiss, S.H. Chong, T. Kuwana, Anal. Chem. 71 (1999) 413. [6] F. Crespi, T.G. England, D.G. Trist, J. Neurosci. Methods 61 (1995) 201. [7] S. Cosnier, C. Innocent, L. Allien, S. Poitry, M. Tsacopoulos, Anal. Chem. 69 (1997) 968. [8] E.S. Forzani, G.A. Rivas, V.M. Solis, J. Electroanal. Chem. 435 (1997) 77. [9] F. Malem, D. Mandler, Anal. Chem. 65 (1993) 37. [10] Z.M. Liu, J.H. Li, S.J. Dong, E.K. Wang, Anal. Chem. 68 (1996) 2432. [11] A. Dalmia, C.C. Liu, R.F. Savinell, J. Electroanal. Chem. 430 (1997) 205. [12] C.R. Raj, T. Ohsaka, J. Electroanal. Chem. 496 (2001) 44. [13] M.A. Chen, H.L. Li, Electroanalysis 10 (1998) 477. [14] D.J. Wiedemann, K.T. Kawagoe, R.T. Kennedy, E.L. Ciolkowski, R.M. Wightman, Anal. Chem. 63 (1991) 2965. [15] J.M. Cooper, D.J. Pritchard, J. Mat. Sci.-Mater. Electron. 5 (1994) 111. [16] D.W.M. Arrigan, Anal. Comm. 34 (1997) 241. [17] J. Wang, P.V.A. Pamidi, G. Cepria, S. Basak, K. Rajeshwar, Analyst 112 (1997) 981. [18] Z.Q. Gao, B.S. Chen, M.X. Zi, Analyst 119 (1994) 459. [19] J.W. Mo, B. Dgorevc, Anal. Chem. 73 (2001) 1196. [20] R.C. Tucker, I. Song, J.H. Payer, R.E. Marchant, J. Appl. Electrochem. 27 (1997) 1079. [21] J. Cruz, M. Kawasaki, W. Gorski, Anal. Chem. 72 (2000) 680. [22] J. Wang, A. Walcarius, J. Electroanal. Chem. 407 (1996) 183. [23] J.M. Zen, P.J. Chen, Anal. Chem. 69 (1997) 5087. [24] R.S. Chittock, A. Glidle, C.W. Wharton, N. Berovic, T.D. Beynon, J.M. Cooper, Anal. Chem. 70 (1998) 4170. [25] C.J. Pickett, K.S. Ryder, J. Chem. Soc. Dalton Trans. (1994) 2181.