Technical Notes Anal. Chem. 1995, 67, 4010-4014
An Electrochemical Cell for EndlColumn Amperometric Detection in Capillary Electrophoresis Meieheng Chen and HsuanJung H u n g * Department of Chemistty, National Sun Yat-sen University, Kaohsiung 80424, Taiwan, ROC
With end-column amperometric detection in capillary electrophoresis, precise positioning and stabilization of the working electrode are highly important. Noises from vibration and breakage of &e microelectrode (e.%.,carbon fiber electrode) are typical problems. To overcome these drawbacks, a new electrochemical cell assembly was designed. In this assembly, alignment of the working electrode with the capillary outlet can be achieved precisely and easily. Coupling with a disk-type Pt electrode (50 pm diameter), detection limits of 3.0 and 5.2 am01 and separationefficienciesof about 70 000 and 150 000 theoretical plates for the determinationof dopamine and catechol, respectively,as test compounds can be obtained. The relative standard deviations (n = 21) of migration time and peak current obtained are 3.5, 5.1%and 2.2, 2.7%,respectively,for these two compounds. Applicability of this assembly as an electrochemical detector for capillary electrophoresiswas demonstratedby runninga synthetic sample containing dopamine, serotonin, norepinephrine, epinephrine, isoproterenol, and catechol. Results obtained are comparable with those from other end-columnor off-columndeterminations. Capillary electrophoresis (CE) was introduced about decade ago by Mikkers et al.' and Jorgenson and Lukac~~3~ as a highly efficient method for separating ionic compounds. Since then, CE has become an important technique in the area of liquid phase separation. Typically, CE is characterized by a minimal sample volume (microliters) requirement, short analysis time, and high separation efficiency. With various separation modes, e.g., capillary zone electrophoresis, micellar electrokinetic capillary electrophoresis, and gel capillary electrophoresis, CE can be used for the separation of anions, cations, neutral mole~ules,4-~ or even (1) Mikkers, F. E. P.; Everaerts, F. M.; Verheggen, Th. P. E. M. J. Chromatogr. 1979, 169, 11. (2) Jorgenson, J. W.; Lukacs, K. D. J. Chromatogr. 1981,218,209. (3) Jorgenson, J. W.; Lukacs, K. D. Anal. Chem. 1981,53, 1298. (4) Ludi, H.; Gassman, E.; Grossenbacher, H.; Marki, W. Anal. Chim. Acta 1988,213,215. (5) Cohen, A. S.; Terabe, S.; Smith, A; Karger, B. L. Anal. Chem. 1987,59, 1021. (6) Walbroehl, Y.; Jorgenson, J. W. Anal. Chem. 1986,58, 479. (7) Wallingford, R A;Ewing, A G. J. Chromatogr. 1988,441,299.
4010 Analytical Chemistry, Vol. 67, No. 27, November 7, 1995
optical isomers.8-'0 CE with a narrow-bore (2-10 pm id.) capillary offers better separation efficiency, but a very sensitive detector with miniial dead volume is needed to accommodate the extremely small sample volume (nanoliters to picoliters) injected. Although different detection schemes, such as UV absorption,laser-induced fluorescence,mass spectroscopy @IS), and electrochemical detection, have been developed for CE, not all of these detection modes are applicable to CE with a narrow-bore capillary. The UV detector commonly ~ s e d ~is~not~ sensitive - ~ ~ ~enough J ~ because of its light-path-dependent characteristic. A laser-induced fluorescence detectot4,8J1provides the required sensitivity for narrowbore CE however, it usually requires extra procedures of pre- or postcolumn derivatization of the analytes. An MS detectorI2 provides the highest sensitivity and also the structural information, but a cost problem arises as compared with other modes of detection. The electrochemical detector, which possesses high sensitivity, is inexpensive and can be coupled with the narrowbore capillary readily. It thus becomes one of the most popular detectors used for CE with narrow-bore ~apillary.'~-~~ Two electrochemical detection modes have been developed: and endcolumn.15-'8126-28 For the offcolumn mode, a conductive junction between the separation and detection Gozel, P.; Gassmann, E.; Michelsen, H.; Zare, R N.Anal. Chem. 1987, 59, 44. Guttman, A; Paulus, A; Cohen, A. S.; Grinberg, N.; Karger, B. L. J. Chromatogr. 1988,448,41. Honda, S.; Iwase, S.; Makino, A; Fujiwara, S. Anal. Biochem. 1989, 176, 72. Cheng, Y.;Dovichi, N.J. Science 1988,242,562. Smith,R D.; Wahl, J. H.; Goodlett, D. R; Hofstadler, S. A Anal. Chem. 1993,65, 574A. Wallingford, R A;Ewing, A G . Anal. Chem. 1989,61, 98. Olefirowicz, T. M.; Ewing, A G. Anal. Chem. 1990,62,1872. Huang, X.;Zare, R. N.; Sloss, S.; Ewing, A G. Anal. Chem. 1991,63,189, Sloss, S.;Ewing, A G. Anal. Chem. 1993,65, 577. Lu, W.;Cassidy, R M.; Baranski, A S. J. Chromatogr. 1993,640,433. Lu, W.; Cassidy, R M. Anal. Chem. 1993,65, w78. Kaniansky, D.; Havasi, P.; Marak, J.; Sokolik, R J. Chromatogr. 1986,366, 153. Wallingford, R A;Ewing, A G. Anal. Chem. 1987,59, 1762. Yik,Y. F.; Lee, H. IC; Li, S. F. Y.; Khoo, S. B.J. Chromatogr. 1991,585, 139. O'Shea. T. J.; GreenHagen, R. D.; Lunte, S. M.; Lunte, C. E. J. Chromatogr. 1992,593,305. O'Shea, T. J.; Lunte, S. M. Anal. Chem. 1993,65, 247. O'Shea, T. J.; Lunte, S. M. Anal. Chem. 1993,65, 948.
0003-2700/95/0367-4010$9.00/0 0 1995 American Chemical Society
capillaries is used to isolate the high voltage applied to the separation capillary from the electrochemical detection system. For the endcolumn detection, a microelectrode is placed directly at the end of the separation capillary without any conductive junction. Although satisfactory results can be obtained from both modes of electrochemical detection, these modes have not been applicable to routine analysis. This is primarily due to the difliculty of h d i n g an appropriate material for making the conductive junction and the requirement of fairly elaborate work on the construction of a reliable and sophisticated electrochemical cell. For end-column detection, in order to improve the sensitivity and to eliminate the noise from mechanical vibrations, precise alignment and stabilization of the working electrode are highly demanded. Breakage of the microelectrode occurs frequently during the handling or assembling of the electrochemical cell. In this study, a new electrochemical cell assembly similar to was designed the off-column detector presented by Tudos et alUz5 and used as an end-column detector. A piece of poly(tetraflue roethylene) (PTFE) tubing enclosing a Pt electrode was employed as a guide for the alignment of the capillary and the working electrode. Precise alignment can be achieved easily with a magnifier instead of a micropositioner and a microscope. With this electrochemical cell, detection limits of 3.0 and 5.2 am01 and separation efficiencies of about 70 OOO and 150 OOO theoretical plates are obtained for the determination of dopamine and catechol, respectively, as test compounds. The relative standard deviations (RSDs, 11 = 21) of migration time and peak current are found to be 3.5, 5.1%and 2.2, 2.7%,respectively, for these determinations in a concentration range of 5 x 10-*-5.0 x M. The effects of injection time and separation potential on the separation efficiency are studied. The feasibility of this cell assembly as an electrochemical detector for CE is demonstrated by running a synthetic sample containing dopamine, serotonin, norepinephrine, epinephrine, isoproterenol, and catechol. Results obtained are comparable with those from endcolumn or offcolumn determinations. EXPERIMENTAL SECTION Apparatus. Fused-silica capillary of 5 pm i.d., 365 pm 0.d. was obtained from Polymicro Technologies (Phoenix, AZ). A high-voltage dc powder supply (Model CZElO00 PN30R Spellman High-Voltage Electronics Corp., Plainview, NY) was used to provide the required voltage (from 0 to -+30 kv). For the CE system, the cathodic end is maintained at ground, and pieces of Pt were used as the contacts of anode and cathode to the power supply. An acrylic box with an interlock on the access door was used to enclose the high-voltage output and to protect the operators from electric shock. The electrophoretic potential, the electrokinetic injection process, and the data acquisition were controlled by a personal computer (386DW40 MHz) equipped with a PCL812 high-performance data acquisition card (B&C Microsystem, Sunnyvale, CA). ~~~~
~~
~
~
~
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(25) Tudos,A J.; Van Dyck, M. M. C.; Poppe, H.; Kok, W. Th.Chromatographia 1993,37,79. (26) Colon, L A;Dandoo, R; Zare, R N. Anal. Chem. 1993.65,476. (27) Ye, J.; Baldwin, R P. Anal. Chem. 1993,65,3525. (28) Doupherty, A M.;Woolley, C. L;Williams, D. L;Swaile, D. F.; Cole, R 0.; Sepaniak, M. J. 1.Lig. Chromatog. 1991,14 (5), 907.
Buffer Solution Inlet Figure I. Top view of the electrochemical cell configuration. Components in figure are not drawn to scale. Dimensions of the acrylic block are 40 x 25 x 20 cm3, and the distances a and b are about 200 pm and 3 mm, respectively. The capillary (5 pm i d . , 365 pm 0.d.) is the separation capillary. The disk working electrode is formed by enclosing the Pt wire (50 pm diameter) with a piece of capillary (75 pm i.d., 365 pm 0.d.). Details of the electrode configuration are given in the text.
The control program was written in Turbo C++. The postrun data processing programz9was written in Quick BASIC. The data processing program displays the electropherogram on the monitor and searches the peaks of the electropherogram. The data of migration time, peak height, peak area, width at half-height, and number of theoretical plates for each detected peak can be obtained readily. The electrochemical cell was made of an acrylic block (40 x 25 x 20 "9. Spaces for positioning the reference electrode, the capillary tubing, and the inlet and outlet of buffer solution were tapped with a l/4 in. thread, while space for allocation of the working electrode was tapped with a 3/16 in. thread. A channel with a diameter of '/I6 in. was drilled for the connection of the working electrode and the capillary tubing. Figure 1 shows the top view of the electrochemical cell configuration. The scheme shown is not to scale, but the dimensions of several key components are labeled. A disk-type working electrode is used in this experiment. To fabricate the disk electrode, a piece of Pt wire (50 pm diameter, 5 cm length) was inserted and passed through a piece of capillary (75 pm i.d., 365 pm o.d., 1 cm length) from which the polyimide coating was removed. Epoxy glue was then applied to both ends of the capillary to secure the Pt wire. No special effort has been made to ensure that the Pt wire was secured at the center of the capillary. The capillary in which the Pt electrode was enclosed was inserted into a piece of PTFE tube (300 pm i.d., l/16 in. o.d., 4 cm length) from the free Pt wire end and stopped when half of the capillary (-5 mm) was enclosed in the PTFEtubing. Both ends of the PTFE tubing were sealed with epoxy glue. The length of capillary protruding from the cured epoxy was kept to about 3 mm. A piece of Cu wire was soldered to the free Pt wire and served as the conducting lead. The disk electrode was carefully polished before use. To assemble the electrochemical cell, a piece of capillary (5 pm i.d., 365 pm 0.d.) of appropriate length (-50 cm) was inserted into a short piece of PTFE tubing (same specifications as above but about 20 mm length) until the capillary was just extruded from it. Though the inside diameter of the PTFE tubing is smaller (29) Gates, S.C.; Becker, J. Laboratory Automation Using nte IBMPC; PrenticeHall: Englewood Cliis, NJ, 1989;Chapter 11.
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than the outside diameter of the capillary, due to the slight softness characteristic of the PTFE tubing, insertion of the capillary into the PTFE tubing can be achieved without difficulty. After the capillary was filled with buffer solution and the extruded end of the PTFE tubing submerged in the same buffer solution, the capillary was pulled gently back into the PTFE tubing until the capillary end was 150-250 pm inside the PTFE tubing. The channels of the electrochemical cell were filled with buffer solution, and the PTFE tubing in which the working electrode was enclosed was inserted into the cell along the I/16 in. channel and secured with a PTFEfinger tight fitting when the Pt electrode rested at the center of the channel. The PTFE tubing in which the separation capillary was enclosed was then inserted from the other end of the '/I6 in. channel and pushed gently until the capillary outlet was in close contact with the disk electrode. The intimate contact between the disk electrode and the capillary end can be examined with a magnifier (with a magnifying factor of 20x). After the alignment was done, the PTFE tubing in which the separation capillary was enclosed was secured with an O-ring and a PTFE flanged fitting. As the surface of disk electrode is much larger than the dimensions of the capillary outlet, a walljet-type thin-layer electrochemical cell is thus formed. A homemade Ag/AgCl reference electrode surround with male screw (1/ 428 thread) was screwed into the cell. A piece of Pt wire was used as the counter electrode and was sealed into the block directly. Amperometric determination and other voltammetry experiments were done with a polarographic analyzer (Model 264A, PARC). A current preamplifier (Model PA-1,BAS) was used for the measurement of very low current. To minimize the interference of external electric noise, the electrochemical cell assembly was housed in a Faraday cage (Model C2, BAS). Current was recorded with a stripchart recorder (Yokogawa Model 3025). For most of the following experiments, CE was run by employing a capillary (5 pm i.d., 365 pm o.d., 50 cm length) at a separation potential of 25 or 20 kV. The buffer solution used contained of 20 mM morpholinoethanesulfonic acid (MES) with pH adjusted to 6.0. Sample injection was performed by electromigration at 12 kV for 5 s. A constant electrochemical potential of 0.700 V vs Ag/AgCl was used for amperometric determination. Buffer solution in the electrochemical cell was refreshed by syringing after each electrophoretic run. Chemicals. MES and dopamine were obtained from Tokyo Chemical Industry Co. Serotonin, norepinephrine, epinephrine, isoproterenol, and catechol were obtained from Sigma. All chemicals were used as received. The buffer solution used was 20 mM MES with pH adjusted to 6.0. Stock solutions (0.01 M) were prepared in 0.1 M perchloric acid. Sample solutions were prepared by dilution of stock solutionsto the desired concentration with buffer solution. RESULTS AND DISCUSSION
Characteristics of the Cell Assembly. Literature for CE with small capillaries indicates that the applied separation potential would drop to an insignificant magnitude at the end of the highresistance capillaries.16 In the system currently studied, the internal resistance of the capillary (5 pm i.d., 50 cm length), Rc and the internal resistance of the PTFE tubing, R p , can be estimated from the following equations, 4012 Analytical Chemistry, Vol. 67, No. 21, November 1, 1995
where PB is the resistivity of buffer solution, IC is the length of capillary, lp is the length of the working electrode enclosed inside the PTFE tubing (Zp 200 pm), and Ac, Ap and AWare the crosssectional areas of capillary, PTFE tubing, and working electrode, respectively. From eqs 1 and 2, the ratio Rc/Rp is estimated to be lo6. As the internal resistance for the buffer-solution-filled, 5 pm i.d. capillary, RCis approximately equal to 1 x 10I2Q,16 a value of 1 x lo6 Q is estimated for the average resistance of the buffersolution-filled PTFE tubing. In the studied system, application of a 20 kV separation potential to the capillary should result in a 20 kV potential drop across the capillary and an -10 mV potential drop across the PTFE tubing. That means the electric field across the capillary is 8 V/200 pm, and that between the capillary outlet and the end of PTFE tubing is about 10 mV/200 pm. As the surface of the disk electrode is perpedicular to the vector of applied separation potential, there should be no potential drop across the surface of the disk electrode due to the separation potential. It is feasible to carry out the amperometric detection with the designed electrochemical cell assembly. To further justify the feasibility of the conclusion made, staircase voltammetry for dopamine and isoproterenol was run in the CE system with the electrochemical cell assembly. Peak potentials found at 0.40 and 0.46 V for these two compounds, respectively, match well the half-wave potentials obtained from batch-type experiments. This implies that the voltammetric behavior of the Pt disk electrode is not affected by the possible iR drop resulting from the applied separation potential. For end-column detection, precise alignment of the working electrode with the capillary outlet is a prerequisite and is usually achieved by careful manipulation of a micropositioner with the help of a micro~cope.~~J~ Poor alignment between the electrode and the capillary outlet will result in a sigdicant loss of sensitivity and the deterioration of the detection limit.17 For a system with a carbon fiber microelectrode, once optimized alignment is achieved, minor displacement of the electrode due to room vibrations or other unavoidable phenomena produces marked changes in the detection currents.17 In this system, the capillary and the electrode are secured with the PTFE tubing and are guided within the same channel. Additionally, the cross-sectional area of capillary outlet to the surface area of the working electrode is in a ratio of 1:100, so proper positioning and good alignment for the capillary and working electrode can be easily attained. As the working electrode and capillary are secured inside the cell assembly, the noises that arise from mechanical vibrations and air drafts are thus eliminated. This would lower the noise level and improve the sensitivity of the electrochemical detector; a similar result has been illustrated in Baldwin's w0rk.2~It takes -20 min to assemble the electrochemical cell. Once assembled, it takes only about 5 min to replace either the working electrode or the capillary tubing. The reproducibility of the alignment operations is evaluated by measuring the detection current of dopamine after each operation cycle of disassembling and assembling the working electrode and separation capillary. The RSDs (n = 8) for the variations of migration time, peak height, and peak area obtained in alignment operations were found to be 3.0, 6.0, and 3.2%,respectively. The
RSDs obtained are better than the value of 10-15% for the walljet arrangement with a normal size electrode and the value of 65% for the conventional endcolumn scheme using a carbon fiber mi~roelectrode.2~ Performance of the Cell Assembly. The characteristics of the designed cell assembly were demonstrated by running the electrophoresis with a solution containing dopamine (5.0 pM), isoproterenol (5.0 pM), and catechol (10 pM). The reproducibility of this system was tested by 21 determinations. The injected amounts were estimated to be 0.45, 0.41, and 0.54 fmol, respectively. Fairly sharp peak were found, but peaks with minute tailing for the cationic solutes (e.g., dopamine and isoproterenol) were also found in the electropherograms. The separation efficiencies, represented by the theoretical plate number N, were estimated to be about 70 OOO, 82 OOO, and 150 OOO, respectively. The RSDs (n = 21) of migration time were 3.5,2.5, and 226, respectively. A gradual increment in the migration time for the successive determinations of the three analytes was also found. This can be attributed to the gradual decrease of the electroosmotic flow, which is caused by the adsorption of cations at the inner wall of the capillary.28~30The RSDs (n = 21) for peak current were found to be 5.1, 6.1, and 2.7%,respectively. The slight decrement of peak current may be due to fouling of the Pt disk electrode. Due to possible fouling on the electrode surface, Pt is not the customarily chosen material for the analysis of catecholamines. In this experiment, as the concentration of catecholamines used was small, fouling of the Pt disk electrode was found to be insignificant. The performance of the Pt disk electrode should be taken as normal and is compatiblewith that of the conventional carbon fiber electrode. The effect of injection time on the sensitivity of the electrochemical cell assembly was studied. It was found that the peak current increased rapidly as the injection time increased from a very short period of time but leveled off when injection time was larger than 10 s; the peak width increased linearly with the increment of injection time. The larger the amount of solute injected, the higher the sensitivity and the poorer the resolution would be. As the migration rate of analyte in the capillary was enhanced by increasing the separation potential, the wall-jet effect on amperometric detection was also enhanced,31and a larger peak current thus resulted. The linear relationship found from plots of the reciprocal of migration time versus the separation potential applied for the analytes studied agrees with theory very well. Although the analysis time can be shortened by applying a larger separation potential, the larger amount of Joule heat dissipated may offset this advantage. The response and separation efficiency are affected by the gap distance between the disk electrode and the capillary outlet in the electrochemical cell. It is difficult to control or measure the gap distance, but an effective gap distance can be obtained by pushing the € W E tubing in which the separation capillary was enclosed gently to the working electrode until a soft contact is attained. The optimal gap distance obtained for the electrode and the capillary is evidenced by the sharp and large peak current that results. It is found that when the gap distance is larger than the optimal value, decrement of the peak current, broadening of the peak width, and thus lowering of the separation efficiency will result. If the gap distance set is too small, a back pressure in the (30) Towns,J. IC; Regnier, F.E. Anal. Chem. 1992,64, 2473. (31) Yamada, J.; Matsuda, H. /. Electrounal. Chem. 1973,44, 189.
53.71
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Tlme (sec) Fl@um2. Electropherogram obtained for the separation of six different neurotransmitters. The concentrations used for (1) dopamine, (2) serotonin, (3) norepinephrine, and (4) epinephrine are 25 pM and for (5) isoproterenol and (6) catechol are 50 pM each. Conditions: separation Capillary, 5 pm i.d., 44 cm length; separation potential, 25 kV; injection potential, 12 kV for 3 s; buffer, 30 mM MES, 20% (vh) 2-propanol at pH 6.0.
capillary result and will subsequently induce the diminution or even the disappearance of peak current. Whenever the electrode and capillary outlet are too close, the effective gap distance can be attained by releasing the flanged fitting in a backward direction with an angle of about 5". It is justified from the very good reproducibility obtained for alignment operation that an optimal contact between the disk electrode and the capillary outlet can be easily attained by following the assembly procedures described above. With this electrode configuration, electrodes of smaller surface area and of different electrode material can be fabricated as long as thin wire materials are available with appropriate dimensions, i.e., instead of enclosing a piece of 50 pm diameter Pt wire with a segment of 75 pm i.d. capillary, Pt, Au wire, or carbon fiber with a diameter smaller than 25 or 10 pm can be enclosed and secured with a segment of capillary which has a specified inside diameter of 25 or 10pm and 375 pm 0.d. Although an advantage with respect to the enhancement of S/N ratio can be obtained by employing a smaller disk electrode, a problem inherent with the center alignment between the electrode surface and the capillary outlet is induced. The deterioration of electrode response due to off alignment may offset the advantage gained by using an electrode of smaller surface area. As long as the disk electrode surface is kept relatively larger than the capillary bore size, center alignment should not be a real problem. Fabrication of a smaller disk electrode may deserve further effort. Applicability of this cell assembly is demonstrated by running a synthetic sample containing six different neurotransmitters.The neurotransmitters used in sample solution include 25 pM each of dopamine, serotonin,norepinephrine, and epinephrine and 50 pM each of isoproterenol and catechol. Figure 2 shows the electropherogram obtained by using a 5 pm i.d., 44 cm long capillary at a separation potential of 25 kV. The buffer solution used contains 30 mM MES and 20%(v/v) of 2-propanol and has its pH adjusted to 6.0. The electropherogram shown in Figure 2 is similar to that obtained by Ewing et al.13 Dopamine and serotonin are well resolved, but a rather small peak of catechol is also found. Tailings present in the peaks corresponding to the cationic solutes are also similar to those found when a 5 pm diameter carbon fiber microelectrode was employed.13 The much smaller peak of Analytical Chemistry, Vol. 67, No. 21, November 1, 1995
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Table 1. Comparison of the Electrode Configuration and Detection Limits among Different End-Column and the Optlmized End-Column Amperometric Detectors
working electrode electrode surface area hmz) detection limitd for dopamine (amol) apparent injection volume (pL) detection limir‘ for catechol (amol) apparent injection volume (pL)
end-columno
end-columnb
optimized end-columnc
F‘t disk 50 pm diam 2000 2.0 133 3.5 80
carbon fiber 10 pm diam x 200 pm 6400 64 > 160 56 160
carbon fiber 11pm diam x 200 pm 7000 18 45 19 27
Data obtained from this work. * Data quoted or estimated from Ewing’s Conditions: separation potential, 20 kV; injection, 20 kV for 5 s; elecfrochemical detetion at 0.8 V; separation capillary, 5 pm id., 56.6 cm length; buffer, 20 mM MES at pH 6.0. Data quoted or estimated kom Ewng’s results.22 Conditions: separation potential, 20 kV; injection, 10 kV for 10 s; electrochemical detection at 0.8 V; separation capillary, 2 pm i.d., 58 cm length; buffer, 25 mM MES at pH 5.65. The detection limit obtained is based on S/N = 2 criterion.
catechol shown in Figure 2 is due to the effect of 2-propanol on the electroosmotic behavior of MES buffer solution. The addition of 2-propanol to the MES buffer solution results in a decrement of the electroosmoticflow in the capillary that influences favorably the separation of serotonin and dopamine in solution. The decrement of electroosmoticflow influences further the sensitivity of catechol in the studied solution. As catechol is an almost neutral molecule in the pH 6 buffer solution and the sample is injected by the electrokinetic method, the amount of catechol injected into the capillary is thus much smaller than the amount of other neurotransmitters in the sample solution. A much lower sensitivity for catechol thus results. Standard calibration graphs based on the peak current for dopamine, isoproterenol, and catechol are plotted. With injection amounts ranging from 6.3 to 3500 amol (0.05-25 pM) for dopamine and isoproterenol and from 7.9 to 3400 amol (0.10-50 pM) for catechol, calibration graphs with linear correlation coefficients better than 0.999 are obtained. The detection limits based on the criterion of S/N = 3 are estimated to be 3.0 (23 nM), 3.6 (28 nM), and 5.2 amol (66 nM) for dopamine, isoproterenol, and catechol, respectively. Though it can be improved by proper optimization, no further effort has been made to pursue a lower detection limit. A comparison of the electrode configuration and the detection limit of the designed electrochemical cell assembly with those of carbon fiber end-column and the optimized end-column detector for the determination of the two neurotransmitters is shown in Table 1. From Table 1, the detection limit obtained from this work is about one order lower than that obtained from the optimized endcolumn detector and the carbon fiber end-column detector. The better performance of this work
4014 Analytical Chemistry, Vol. 67, No. 27, November I, 7995
may be attributed to the smaller electrode surface adopted in the electrochemical cell, which enhances the S/N ratio and the larger sample volume injected. CONCLUSIONS The primary advantage of this electrochemical cell assembly is its easier construction compared with other endcolumn electrochemical detectors, in which a micropositioner is needed.21-23 With the help of a piece of guide tubing, alignment between the capillary outlet and the working electrode can be achieved easily and reliably. The noise from mechanical vibrations and drafts is eliminated. With the proposed fabrication procedures, electrodes of various materials such as Au, Cu, Ni, or other metal wire and even carbon fiber, can be fabricated,thus extending the applicability of this cell assembly. The easy construction of this electrochemical cell assembly, the flexibility of adopting different electrode material, the high reproducibility inherent with the assembling processes, and the feasibility of reconditioning the disk electrode should make this electrochemical detector more acceptable to other CE researchers and the electrochemical detection method more useful for CE routine analysis. ACKNOWLEWMENT The authors thank the National Science Council of the ROC for financial support of this work (Contract No. NSC 83-0421-M110-0252). Received for review May 2, 1995. Accepted August 18,
1995.a AC950428U @
Abstract published in Advance ACS Abstracts, September 15, 1995.
