Anal. Chem. 2000, 72, 4677-4682
Electrochemical Detectors Prepared by Electroless Deposition for Microfabricated Electrophoresis Chips Abdelkader Hilmi and John H. T. Luong*
Biotechnology Research Institute, National Research Council Canada, Montreal, Quebec, Canada H4P 2R2
Microfabricated capillary electrophoresis (CE) chips with integrated electrochemical detection have been developed on glass substrates. An electroless deposition procedure was used to deposit a gold film directly onto the capillary outlet to provide high-sensitivity electrochemical detection for catechol and several nitroaromatic explosives. Scanning electron microscopy revealed that the electroless gold film contains nanoscopic gold aggregates (100-150 nm) with an average thickness of 79 nm. The electroless deposition procedure can be easily and routinely performed in any wet-chemistry laboratory, and electroless gold can be deposited onto complex and internal surfaces. Intimate coupling of electrochemical detection and CE chips obviates the need for a coupling mechanism or tedious alignment procedures. With nitroaromatic compounds as a working model, microchip capillary electrophoresis equipped with electroless gold has proven to provide high sensitivity and fast response times for sensor applications. The CE microchip system was capable of separation and determination of explosive compounds including TNT in less than 130 s with detection limits ranging from 24 to 36 µg/L, i.e., 4-fold enhancements in detection efficiency in comparison to thick-film technology. Micromachined capillary electrophoresis (CE) chips have received considerable interest in analytical chemistry because of their compactness and fast and efficient separation capabilities. Such devices have the potential to simultaneously assay large number of samples, facilitate integration of detection systems, and consume only picoliters of sample volumes.1,2 Like conventional CE, fluorescence detection remains one of the most sensitive detection techniques for CE chips. However, when incorporating direct fluorescence detection into a CE system that does not have naturally fluorescing analytes, derivatization of the analyte is a prerequisite. Furthermore, relatively few compounds of biological interest fluoresce with high efficiency. This detection scheme also requires expensive off-chip instrumentation and often necessitates time-consuming alignment procedures. (1) Gordon, M. J.; Huang, X.; Pentoney, S. L., Jr.; Zare, R. N. Science 1988, 242, 224-228. (2) Manz, A.; Harrison, D. J.; Verpoorte, E. M. J.; Fettinger, J. C.; Paulus, A.; Ludi, H.; Widner, H. M. J. Chromatogr. 1992, 593, 253-258. 10.1021/ac000524h CCC: $19.00 Published 2000 Am. Chem. Soc. Published on Web 08/15/2000
Electrochemical devices have been very useful for various conventional capillary electrophoresis applications due to their low cost and inherent miniaturization.3,4 CE equipped with electrochemical detection has already been proven for analysis of electroactive compounds such as phenolics, neurotransmitters, explosives, and carbohydrates.5-8 Electrochemical methods offer high flexibility in comparison to fluorescence and compatibility with microfabrication to produce completely integrated microanalysis instrumentation. Woolley et al.9 first reported on capillary electrophoresis chips with integrated electrochemical detection, based on the photolithographic placement of the detecting electrode positioned outside the exit of the electrophoretic separation channel. Gavin and Ewing10 developed a thinfilm microfabricated electrochemical array detector for planar CE chips. Recently, Wang et al.11 described an on-chip electrochemical detector based on sputtering the working electrode directly onto the channel outlet for analysis of L-dopa, dopamine, and isoproterenol. In another approach, micromachined electrophoresis chips with thick-film electrochemical detectors have been developed for assays of mixtures of nitroaromatic explosives or catecholamines.12 From a practical viewpoint, screen-printed detectors are more beneficial, particularly with applications that require a frequent replacement of the working electrode due to severe electrode fouling. The coupling of a microelectrode detection system with CE chips was described recently in this laboratory for analysis of explosive compounds in soil and groundwater.13 This approach is particularly useful for applications that require frequent electrode cleaning or replacement due to severe surface poisoning. This paper describes an electroless deposition procedure for fabricating gold electrodes just outside the exit of the separation channel to serve as a working electrode. To our knowledge, the present work represents the first demonstration of using the electroless deposition technique to fabricate a nanosized electro(3) Wallingford, R. A.; Ewing, A. G. Anal. Chem. 1987, 59, 1762-1765. (4) O’Shea, T. J.; Lunte, S. M. Anal. Chem. 1994, 66, 307-312. (5) Olefirowicz, T. M.; Ewing, A. G. Anal. Chem. 1990, 62, 1872-1876. (6) Hilmi, A.; Luong, J. H. T.; Nguyen, A. L. J. Chromatogr., A 1997, 761, 259268. (7) Bratin, K.; Kissinger, P. T.; Briner, R.; Bruntlett, C. Anal. Chim. Acta 1981, 130, 295-311. (8) Hilmi, A.; Luong, J. H. T.; Nguyen, A. L. Anal. Chem. 1999, 71, 873-878. (9) Woolley, A. T.; Lao, K.; Glazer, A. N.; Mathies, R. A. Anal. Chem. 1998, 70, 684-688. (10) Gavin, P.; Ewing, A. G. Anal. Chem. 1997, 69, 3838-3845. (11) Wang, J.; Tian, B.; Sahlin, E. Anal. Chem. 1999, 71, 3901-3904. (12) Wang, J.; Tian, B.; Sahlin, E. Anal. Chem. 1999, 71, 5436-5440. (13) Hilmi, A.; Luong, J. H. T. Environ. Sci. Technol. 2000, 34, 3046-3050.
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Figure 1. Microcapillary electrophoretic system with electrochemical detection (dimensions are not to scale). (A) Glass microchip, (B) separation channel, 80-mm length, (C) anodic or buffer reservoir, (D) sample reservoir, (E) sample waste reservoir, (F) detection cell or cathodic reservoir, (G) detecting gold electrode (prepared by electroless deposition), (H) counter electrode, (I) reference electrode and (J) cathodic electrode (highvoltage separation), (K) anodic electrode (high voltage separation), and (L) and (M) Pt electrodes for sample injection.
chemical system in CE chip. This flexible and versatile plating permits the deposition of gold from solution onto glass surfaces without the requirement to apply an external electrical potential, and the surface to be coated needs not be electronically conductive.14 The procedure is easy to perform in any wet-chemistry laboratory for preparation of CE chips with integrated metal electrodes. The electroless deposited working electrode requires no time-consuming alignment procedures or decoupling mechanism. Cyclic voltammetry (CV) is applied to characterize the gold electrode prepared by the electroless deposition procedure and compare it to the gold disk electrode. Scanning electron microscopy (SEM) is used to examine the morphology as well as to estimate the gold film thickness. Micromachined CE chips equipped with the integrated gold electrode are investigated for sensitive detection of the explosive compounds. The inherent redox properties of nitroaromatic explosives make them ideal candidates for electrochemical detection.15 Improved detectability and separation efficiency are discussed and compared with the results reported in the literature. EXPERIMENTAL SECTION Materials. All explosives were purchased from ChemService (West Chester, PA). Catechol and all other chemicals (SnCl2, AgNO3, trifluoroacetic acid, Na2SO3, NH4OH, formaldehyde, (14) Menon, V. P.; Martin, C. R. Anal. Chem. 1995, 67, 1920-1928. (15) Bratin, K.; Kissinger, P. T.; Briner, R.; Bruntlett, C. Anal. Chim. Acta 1981, 130, 295-311.
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methanol (HPLC grade) and HNO3 were purchased from Aldrich (Milwaukee, WI). Commercial gold electroless plating solution (Oromerse Part B, Technic, Anaheim, CA) was diluted 40-fold in water prior to use. The smooth gold electrode was purchased from Bioanalytical Systems (BAS, West Lafayette, IN). Sample Preparations. Stock solutions of explosives were prepared in acetonitrile, whereas catechol was prepared in 0.1 M perchloric acid solution. Prior to CE analysis, all samples were diluted in the buffer and were passed through a 0.45-µm filter. To avoid any decomposition, all stock solutions were protected from light and kept at 4 °C. CE Chips. The glass microchannel separation chips were fabricated at Alberta Microelectronic Corp. (AMC, µchip-T 100, Edmonton, AB, Canada), using standard microphotolithographic technology, wet chemical etching, and thermal-bonding techniques. The CE glass chip consists of a glass plate (10 × 2 cm), with an 8-cm-long separation channel (between the inlet reservoir and the detection reservoir) and 1-cm-long injection channel (between the sample reservoir and the sample waste reservoir). The channel has a maximum depth of 20 µm and a width of 50 µm (at the top) (Figure 1). The original outlet reservoir was cut off by AMC, leaving the channel outlet at the highly flat end of the chip. After the cut edge was polished with an abrasive paper aided by an alumina slurry (CF-1050, BAS), the resulting chip was washed extensively with deionized water and sonicated for 30 min to remove any alumina particles which could adhere to
the separation or sample channel. Pipet tips (200 µL) were properly cut flat and placed over the drilled access holes (0.8 mm) to form the buffer (anodic or inlet), sample, and sample waste reservoirs. A platinum wire (0.5 mm) was inserted into each reservoir to provide electrical connections to the channels. Amperometric Detection and CE Chip Arrangement. The glass chip was fixed in a laboratory-built Plexiglas supporter whereas the channel outlet was inserted into a custom-built Plexiglas detection reservoir with silicone grease providing proper sealing (Figure 1). A platinum wire was inserted into this reservoir through a septum to serve as a cathode for electrophoresis whereas two other septa were installed on the side of this reservoir to allow insertion of the two electrodes. The Ag/AgCl reference electrode was prepared by electrochemical oxidation of a silver wire (0.5 mm diameter) in 0.1 M HCl, while a platinum wire (0.5 mm diameter) served as the counter electrode. Cyclic voltammetry experiments used a potentiostat (model 263 A, EG&G, Princeton Applied Research, Princeton, NJ) to operate the three-electrode amperometric detection system. During electrophoresis, a CV-1B voltammograph (Bioanalytical Systems, BAS, West Lafayette, IN) was used to apply -0.8 V to the detecting electrode. The time response data were digitized and treated by an A/D board (DP 500-AD) supplied with an interface box (Labtronics, Guelph, ON, Canada) that was installed on a PC 486 computer. The data were stored in ASCII files and converted to PRN files for treatment by a graphic program. No software filtration of the signal was used. Preparation of the Working Electrode by Electroless Deposition. The process of electroless metal deposition involves the use of formaldehyde, a chemical reducing agent to plate gold from solution onto a surface. The advantage of the electroless method compared to the electrochemical deposition is that the surface to be coated needs not be electronically conductive. The most important requirement of an electroless deposition bath of this type is to arrange the chemistry such that the kinetics of homogeneous electron transfer from the reducing agent to the metal ion is slow. Otherwise the metal ion would simply be reduced in the bulk solution. A catalyst that accelerates the rate of metal ion reduction is then applied to the surface to be coated. In this way, metal ion is reduced only at the surface of the substrate (glass). Initially, the end (5 mm) of the chip was cleaned by immersion in a solution of nitric acid/water (50/50) for 20 min followed by another immersion in Piranha solution (70% H2SO4/ 30% H2O2) for 30 min and then extensively rinsed with methanol and warm water (60 °C). The cleaning procedure was performed to remove organic contamination if present and to improve adhesion of the plated metal to glass. The procedure used in this work for electroless plating of gold was based on the work of Menon and Martin14 in which the deposition of a thin layer of titanium by evaporation on the surface of glass microscope slides was necessary to improve the adhesion between the gold and glass. However, with high index glass or PTCE membranes, the titanium underlayer was not necessary; therefore no deposition of titanium was performed in this study. The procedure used for electroless deposition of gold consisted of three principal steps. First, Sn2+ was adsorbed onto the substrate surface by immersion of the CE chip (5 mm of the channel outlet) into a solution of 26 mM SnCl2 and 70 mM
trifluoroacetic acid prepared in water for 3 min (with stirring). The chip was thoroughly rinsed with warm water (60 °C) since rinsing of the glass substrate in warm water at this temperature was reported to improve the adhesion of the gold to glass.14 In the second step, the sensitized chip was immersed in an aqueous solution of ammoniacal AgNO3 (29 mM) for 2 min (with stirring). In this step, a redox reaction led to the oxidation of Sn2+ to Sn4+ and the reduction of Ag+ to Ag; i.e., the surface became coated with nanoscopic silver particles. After Ag deposition, the chip was then rinsed thoroughly in warm water (60 °C) several times. Finally, the Ag-coated chip was immersed in a gold plating solution (22-23 °C) consisting of 7.9 mM Na3Au(SO3)2 (commercial plating solution diluted 40 times), 127 mM Na2SO3, and 625 mM formaldehyde (with stirring). The chip was kept in the gold solution for 5 h to result in a good deposition. In this step, the Ag particles are galvanically displaced by Au since gold is a more noble metal and the resulting Au particles are excellent sites for the oxidation of formaldehyde and the concurrent reduction of Au(I) to Au(0). Consequently, gold plating continues on the gold particles and the reaction can be described as14
2Au(I) + HCHO + 3OH- f HCOO- + H2O + 2Au (0)
After plating the chip was rinsed thoroughly with water and the following treatments were performed to clean the gold deposited film. The chip was immersed in a solution of 25% HNO3 for 1 h with stirring to dissolve any residual Sn or Ag that could be strongly adsorbed on the surface. After this step, the chip was annealed at 180 °C for 6 h and then electrochemically cleaned in 0.1 M HClO4 by cycling between -0.6 and 1.2 V vs Ag/AgCl until the stability of the voltammograms was attained. A silver wire (0.125 mm) was attached to the gold film by silver epoxy and covered by an insulated layer to ensure rigid connection; only a small disk around the outlet of the separation channel was exposed to the solution in the cathodic reservoir (detection reservoir). During the gold plating process, the outlet of the separation channel was not protected. A microscopic inspection showed a thin film of gold deposited inside the channel (40 µm). Scanning Electron Microscopy. SEM images were obtained using a Hitachi scanning electron microscope (model S-4700, Tokyo, Japan) with a magnification of 20-500000-fold. The system is equipped with a built-in anticontaminator that allows highresolution microscopy at low operating voltages (2.5- and 1.5-nm resolution at 1 and 15 kV, respectively). Electrophoresis Procedure. The separation potential was effected by a high-voltage power supply with an adjustable voltage range between 0 and 50 kV (model EH, Glassman High Voltage Inc., Whitehouse Station, NJ). Prior to use, the CE channels were flushed with methanol for 2 min to remove any organic contamination and rinsed with deionized water for another 2 min. The CE channels were then filled with 0.1 M NaOH for 1 h, followed by a rinsing sequence with 0.1 M HClO4 for 15 min and deionized water for 5 min. All flushing and rinsing sequences were performed by applying a low pressure (20 psi). To begin the experiment, all the channels were filled with the running buffer (15 mM borate, pH 8.7, containing 25 mM sodium dodecyl sulfate (SDS) or 25 mM 4-morpholineethanesulfonic acid (MES), pH 6.2) by applying a low pressure (20 psi) to the inlet buffer reservoir Analytical Chemistry, Vol. 72, No. 19, October 1, 2000
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for 1 min. The buffer in the sample reservoir was emptied and replaced by the sample (the stock sample, 50 µL, diluted in the separation buffer). The injection channel between the sample reservoir and the sample waste reservoir was filled by applying +1100 V to the sample reservoir for 15 s with the sample waste reservoir grounded whereas both the inlet buffer and detection reservoirs were floating. To establish an initial stable baseline, +1500 V was applied to the inlet buffer reservoir with the detection reservoir grounded and the sample and sample waste reservoirs floating. During analysis, the sample was then injected to the separation channel by applying +1100 V to the sample reservoir for 4 s with the sample waste reservoir grounded and the inlet buffer and detection reservoirs floating. Unless otherwise stated, the solutions were not deaerated and all separations were performed at room temperature (21-23 °C). The electropherogram peaks were identified by spiking with individual explosives. For each component, a series of runs at different concentrations was performed to obtain a peak height vs concentration plot. Separation efficiency, defined as the number of theoretical plates (N) for each peak in the electropherogram, was calculated as 5.54(tR/w1/2)2, where tR and w1/2 represent the migration time of the analyte and the full peak width at half-maximum. The resolution (Rs) of one peak from the preceding peak was calculated as 1.18(t2 - t1)/(w1/2,1 + w1/2,2), where ti and w1/2,i are defined above for the two peaks.16 Safety Considerations. TNT and other explosive compounds cause headache, weakness, anemia, and liver injury, and the vapor of such chemicals is very dangerous. Stock solutions of the explosives must be prepared and handled in a ventilated hood. Skin contact must be avoided since such explosives may be absorbed through the skin. Special care must also be taken to dispose of waste solutions. During the course of experiments, to avoid electrical shock the high-voltage power supply must be handled with extreme care. RESULTS AND DISCUSSION Morphology of the Gold Film. The electron microscopic image of the gold film, prepared by electroless deposition around the outlet of the separation channel, is illustrated in Figure 2. It should be noted that ∼5 mm of the channel outlet was immersed into the gold solution so the gold film was also expected to form onto the internal surface of the separation channel, which could not be observed by electron microscopy. The gold film consisted of 100-150-nm-sized gold aggregates on the surface of the separation channel outlet. Gold aggregates were also well connected to form a thin-film layer with an average thickness of 77 nm. This result was in agreement with that of Hou et al.,17 who reported that the average thickness of the films of electroless gold deposited on glass substrates was ∼100 nm. Completeness of the gold film deposition was not attained on the entire edge of the CE chip, which indicated that adhesion between the electroless gold film and glass was still not very strong even though thermal annealing was applied to improve their adhesion. However, this should not be a problem here since a small gold film attained around the separation channel was considered sufficient to serve as the detecting electrode to provide electrical contact with high sensitivity as demonstrated below. (16) Jorgenson, J. W.; Lukas, K. D. Anal. Chem. 1981, 53, 1298-1302. (17) Hou, Z.; Abbott, N. L.; Stroeve, P. Langmuir 1998, 14, 3287-3297.
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Figure 2. Scanning electron microscopic image of the gold film prepared by electroless deposition. The image was obtained at 5 kV in view of the capability of the system to possess high resolution at low operating voltages (2.5 nm at 1 kV and 1.5 nm at 15 kV, respectively).
Figure 3. Cyclic voltammograms of gold electrode in HClO4 (0.1 M) at 50 mV/s (scan rate) and 21-23 °C. Dotted line: CV obtained with a smooth gold electrode. Solid line: CV obtained with a microdisk electrode prepared by electroless deposition.
Cyclic Voltammetry of the Gold Film. Cyclic voltammetry was used to estimate the surface of the gold electrochemically active and to evaluate the purity of the surface. Figure 3 shows the cyclic voltammograms recorded with a smooth BAS gold electrode (dotted line) and the film of electroless gold deposited on the chip (working electrode). Both cyclic voltammograms presented similar anodic oxidation peaks around 0.75-1 V; however, the potential and the shape of the reduction peaks were noticeably different. In the case of the smooth gold electrode, the peak is very defined and appears at +0.6 V, whereas with the gold film, the reduction starts at +0.7 V with a maximum at +0.45 V. Furthermore, CV of the gold film displayed a shoulder around +0.4 V and this could be explained by the presence of some highly oxygenated gold species formed during the annealing treatment. The reduction of such species needs lower potential than that obtained with the smooth gold electrode, which could explain the difference of 150 mV. The activity of the gold film was estimated by studying the electrochemical reduction of TNT and 2,3-DNT (Figure 4). The
Figure 4. Cyclic voltammograms of gold electrode prepared by electroless deposition in 15 mM borate, pH 8.7, containing 25 mM SDS at 50 mV/s and 21-23 °C in the presence of 1 mg/L TNT (dotted line) and 1 mg/L 2,3-DNT (solid line).
cyclic voltammogram recorded with the latter (solid line) displayed two peaks around -0.74 and -1.02 V; however, with the former, only one reduction peak at -0.9 V was observed. These results were in good agreement with those obtained previously with a smooth gold electrode (disk), but with higher activity in the case of the electrode prepared by the electroless deposition. As reported by Menon and Martin14 and Hou et al.,17 electrodes or films prepared by the electroless procedure contain nanoscopic gold particles (∼10-30 nm) with a high dispersion. In this study, the size of gold aggregates ranged from 100 to 150 nm. In contrast, the smallest electrodes that can be obtained with electrochemical deposition are of the order of 200 nm.18 Consequently, cyclic voltammetric detection limits for electroactive species at ensembles containing such nanoscopic gold particles were appreciably lower than those at large (3.17 mm) gold or smooth disk electrodes. Characterization of the CE Chip Equipped with the Integrated Gold Electrode. A primary advantage of the on-chip electroless deposition is the simplicity of the procedure to form a metal electrode at the capillary outlet as an effective end-column detection. Therefore, with this integrated system, no timeconsuming adjustment is required to optimize the space between the capillary outlet and the detecting electrode, one of the drawbacks of CE chips equipped with an external detecting electrode. A series of experiments was first conducted to examine the performance of the CE chip for analyzing catechol at different separation potentials. The separation efficiency (plate number), current signals, baseline slope, and separation time were affected by the separation potential. Increasing the separation potential from +1100 to +1800 V drastically reduced the migration time of catechol from 120 to 40 s (Figure 5A). Although there was a noticeable increase in the baseline current and its slope (-680 nA at +1400 V to -950 nA at +2300 V as shown in Figure 5B), very low noise levels were observed in all cases. The amperometric signal decreased noticeably upon increasing the separation potential from +1100 to +1500 V but remained almost unchanged upon further increasing the separation potential from +1500 to (18) Penner, R. M.; Martin, C. R. Anal. Chem. 1989, 61, 762-766.
Figure 5. Influence of the separation potential on the detection of catechol [(A) (a) +1100, (b) +1500, and (c) +1800 V] and TNT [(B) (a) +1400, (b) +1850, and (c) +2300 V]. The separation buffer consisted of 15 mM borate, pH 8.7, and 25 mM SDS for TNT and 25 mM MES, pH 6.2, for catechol, respectively. Sample injection at +1100 V for 4 s; detection potential -0.8 V vs Ag/AgCl for TNT and + 0.8 V for catechol.
+1800 V. However, a surprisingly different trend in separation efficiency was observed since the highest theoretical plates resulted at +1500 V. At this separation potential, the number of theoretical plates for catechol was highest and estimated to be 143 000 plates/m. Such a result was favorably compared to that of Wang et al.11 (490-4100 plates for a 7.7-cm-long separation channel, i.e., 6400-53 000 plates/m). During the course of separation and measurement, the activity of the working electrode annealed at 180 °C was stable and peak height variation for 10 repeated analyses was ∼4%. In the case of the electrode treated only by acidic solution (25% HNO3), there was a variation higher than 12% for 10 analyses, which could be explained by a loss of gold particles. Notice that the treatment at high temperature improves the adhesion of gold particles on glass substrate. The detection limits for each compound were estimated as the concentration at which the peak area was equal to 3 times the standard deviation of the peak area vs concentration plot. The limit of detection was estimated to be 26.4 µg/L (0.24 µM) for catechol., i.e., 3-fold lower than the value obtained with thick-film electrochemical detectors (0.78 µM).12 The electrochemical activity of the gold film, i.e., the amperometric signal, was evaluated for analyzing TNT at different separation potentials (Figure 5B). As expected,9,12 increasing Analytical Chemistry, Vol. 72, No. 19, October 1, 2000
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Table 1. Detection Limit and Theoretical Plates of the Nitroaromatic Compounds detection limit (µg/L) compds
electroless gold film
disk electrodea
theor plates (N)
TNT 2,4-DNT 2,6-DNT 2,3-DNT
24 33 36 35
110 150 160 150
53 200 88 100 80 100 93 700
aValues obtained from ref 8.
Figure 6. Analysis of a mixture of explosives (0.8-1 mg/L) using a gold electrode prepared by electroless deposition. The separation buffer consisted of 15 mM borate, pH 8.7, and 25 mM SDS. The separation voltage was applied at +1500 V and sample injection at +1100 V for 4 s; detection potential -0.8 V vs Ag/AgCl.
separation voltage from +1400 to +2300 V drastically decreased the migration time of the analyte and resulted in a sloping baseline with only minor increases in the background noise. The same order of improvement (4-5-fold) was obtained with explosive compounds (Table 1). This improvement in sensitivity could be explained by the presence of a highly active surface generated by the presence of nanoparticles of gold. Such favorable detectability is useful for on-site screening and environmental monitoring. Separation of TNT and Other Explosives. Figure 6 demonstrates the applicability of the CE/electrochemical system for analyzing a mixture of TNT, 2,4-dinitrotoluene (2,4-DNT), 2,6DNT, and 2,3-DNT. A separation buffer consisting of 15 mM borate, 25 mM SDS, pH 8.7, was used to resolve TNT from 2,4DNT, 2,6-DNT, and 2,3-DNT. The applicability of this buffer for separating several nitroaromatic explosives has been documented earlier;8 therefore, there was no attempt to modify the separation buffer.19 Electrophoretic separations of TNT and three other nitroaromatic explosives in under 130 s demonstrated the good resolution as well as separation efficiency of the CE chip. Despite (19) Grossman, P. D.; Colburn, J. C. Capillary Electrophoresis Theory and Practice; Academic Press: San Diego, CA, 1992.
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the negative potential (-0.8 V) essential for reduction of the nitro moiety, the electrochemical detector displays a low background noise and sharp peaks. For a 80-mm-long separation channel, the separation efficiency, represented by the plate number (N), corresponds to 53 200 for TNT, 88 100 for 2,4-DNT, 80 100 for 2,6-DNT, and 93 700 for 2,3-DNT. The detection limit of the four explosives was estimated to be 24 µg/L for TNT, 33 µg/L for 2,4DNT, 35 µg/L for 2,3-DNT, and 36 µg/L for 2,6-DNT. It should be noted that such detection limits are significantly lower than the values obtained with the disk electrode8 (110 µg/L for TNT, 150 µg/L for 2,4-DNT, 150 µg/L for 2,3-DNT, and 160 µg/L for 2,6-DNT). The reusability and reproducibility of a single CE chip were verified with repetitions during 80 days of operation. In this period of testing, there were no appreciable variations (more than 3%) in retention time and peak area. However, to obtain such reproducibility after use the channel was flushed with methanol for 2 min and then followed a rinsing sequence of 0.1 M NaOH for 4 min, water for 2 min, and the separation buffer for 2 min. When a significant change was noticed in the sensitivity, the working electrode was simply treated electrochemically by recycling the potential between -0.8 and +1 V at 60 mV/s for 5 min. It should be noted that, due to the sample dispersion into the separation channel during the injection procedure, it was somewhat difficult to estimate exactly the injected sample volume. If this effect is neglected and the electrophoretic mobility of the analyte (neutral) is zero, the injected volume is estimated as µeofVπr2t/L, where µeof (electroosmotic flow) is ∼6.1 × 10-8 m2/ V‚s, V is the applied potential, L is the separation channel length, r is the channel radius, and t is the injection time.19 In this study, the injected volume is estimated to be 7.5 nL, i.e., ∼5.5% of the separation channel volume. In brief, thin films of gold prepared by electroless deposition can be used as an electrochemical detector in CE chips. The procedure can be easily performed, and electroless gold films can be deposited onto complex and internal surfaces. With nitroaromatic compounds as an electroactive model, microchip CE equipped with electroless gold can provide high sensitivity and fast response times for sensor applications. The integrated CE chip/amperometric detection system requires no coupling mechanisms or alignment procedures to optimize the space between the capillary outlet and the working electrode. ACKNOWLEDGMENT Scanning electron microscopic images of the gold film were performed by Jeff Fraser of the Institute of Microfabrication Sciences, National Research Council, Ottawa, Canada. The authors also thank Dr. H. John Crabtree of the Alberta Microfabrication Incorporation of Edmonton, Alberta, Canada, for several useful discussions related to chip fabrication and modification. Received for review May 11, 2000. Accepted July 6, 2000. AC000524H
