Anal. Chem. 1988, 60, 1645-1648
nal-to-noise characteristics ( S I N = 3) indicated a detection limit of 1pM, i.e., 0.64 ng, in the 20-pL sample. Hence, the excellent sensitivity of amperometric detection is not compromised by the electrode coating. In conclusion, amperometric detection for flowing streams is greatly improved by the multifunctional character of CoPC/CA coatings. The incorporation of CoPC into the CA domain has essentially no effect on the catalytic behavior. Such effective coupling of electrocatalysis and permselectivity provides a means for minimizing problems and extending the scope of electrochemical measurements in flowing streams. This study suggests also that cellulose acetate could be an ideal host material for the preparation of other powerful multifunctional coatings.
Registry No. CoPC, 3317-67-7; CA, 9004-35-7; carbon, 7440-44-0; hydrogen peroxide, 7722-84-1; hydrazine, 302-01-2; cysteine, 52-90-4; oxalic acid, 144-62-7.
1845
LITERATURE CITED (1) Stulik, K.; P a d k o v i , V. €/echoanalytical Measurements h Flowing Liquids; Ellis Horwood: Chichester, England, 1987. (2) Kisslnger, P. T. Anal. Chem. 1977, 49, 447A. (3) Bard, A. J. J. Chem. Educ. 1983, 60,302. (4) Murray, R. C.; Ewing, A. G.; Durst, R. A. Anal. Chem. 1987, 59,
379A.
(5) Korfhage, K. M.; Ravichandran, K.; Baldwin, R . P. Anal. Chem. 1984, 56, 1514. (6) Wang, J.; Freiha, B. Anal. Chem. 1984, 56, 2266. (7) Halbert, M. K.; Baldwin, R. P. Anal. Chem. 1985, 57, 591. (8) Marko-Varga, G.; Appelqvist, R.; Gorton, L. Anal. Chim. Acta 1988, 179, 370. (9) Wang, J.; Hutchins, Anal. Chem. 1985, 57, 1536. (10) Wang, J.; Tuzhi, P.; Golden, T. Anal. Chim. Acta 1987, 794, 129. (11) Wang, J.; Golden, T.; Tuzhi, P. Anal. Chem. 1987, 59, 740. (12) Santos, L. M.; Baldwin, R. P. Anal. Chem. 1986, 58, 848.
RECEIVED for review December 29,1987. Accepted March 28, 1988. This work was supported by the National Institutes of Health (Grant No. GM 30913-04) and Battelle Pacific Northwest Laboratory.
Thin-Layer Spectroelectrochemical Cuvette Cells with Long Optical Path Lengths Yupeng Gui,’ Steven A. Soper, and Theodore Kuwana* Center for Bioanalytical Research, University of Kansas, 2095 Constant Avenue, Lawrence, Kansas 66046 Thin-layer transmission spectroelectrochemistry has been widely used to investigate various heterogeneous and homogeneous electrochemical processes in both aqueous and nonaqueous solutions (1-6). However, the conventional thin-layer spectroelectrochemical cell, in which the light is passed through an electrode, has two major disadvantages. One is its short optical path length, which is defined by the thickness of the thin solution layer, and provides poor optical sensitivity for observing solution species. Another is the requirement of optically transparent electrodes (OTE), such as metal-coated and metal oxide coated glass of Pt and Au minigrid electrodes. Recently, a new spectroelectrochemical cell, the long optical path length thin-layer cell (LOPTLC), has been developed in this and other laboratories (7-9) to overcome the above disadvantages. In this LOPTLC configuration, the light irradiates the thin solution layer along the axis of the electrode/solution interface, thus greatly improving its optical sensitivity (about 100 times more sensitive than a conventional thin-layer cell) while all other thin-layer spectoelectrochemical behaviors are maintained. Several studies (410-1 7)have demonstrated the excellent utility of this LOPTLC to investigate various heterogeneous and homogeneous phenomena, such as adsorption, desorption, catalytic oxidation, catalytic hydrogenation, and solution chemical reactions associated with a solid electrode. However, there are a few shortcomings with this LOPTLC: (1) Fragile electrodes, such as glass-coated and Si semiconductor electrodes, are difficult to use because pressure must be applied to the body of the electrode to seal for vacuum-tight conditions. (2) It is very inconvenient and time-consuming to remove the electrode from the cell in order to pretreat the electrode. (3) The current cell does not allow simultaneous transmission spectroelectrochemistry to be conducted with other spectroscopy methods, such as luminescence and surface reflectance. (4) It is difficult to perform flowing solution experiments. ( 5 ) It is difficult to rigorously exclude oxygen within the cell which is made from Teflon or Kel-F (10,16) and the LOPTLC made from a material that is not O2 Present address: Department of Chemistry, University of Cincinnati, Cincinnati, OH 45221-0172.
permeable should help to circumvent this problem. T o overcome these shortcomings, two new thin-layer spectroelectrochemicalcells with long optical path lengths were designed. These cells are called “cuvette” cells because they are simply made from commercially available quartz cuvettes. In this communication, the construction and characterization of two different types of cuvette cells are reported. The use of one of these cells in flow injection analysis (FIA) is also reported.
EXPERIMENTAL SECTION Cell Fabrication. A. A schematic diagram of the glassy carbon cuvette cell (cell A) is shown in Figure 1. Figure 1A is the front view of the cell, viewed along the optical path. Parts B and C of Figure 1are the left and right views, respectively, of the cross section S-S as marked in Figure 1A. The cell body is an ordinary quartz cuvette with a 10-mm optical path length. Its internal height and width are 23 mm and 2.0 mm, respectively. A small hole was made at the bottom of the cuvette by polishing with the tip of a fine file and finally drilling with a heated tungsten wire. At the top of the cuvette there are two small pieces of 125 wm thick Teflon film (type 500C, Du Pont, Wilmington, DE) on each of the two cuvette walls, as indicated in Figure 1B. The Teflon film was melted onto the cuvette wall at a temperature of ca. 300 “C. This film controlled the thickness of the solution layer. To eliminate any stray light caused by the light passing through the cuvette wall, both ends of the wall were coated with black water-insoluble ink. The coating was applied with an extra fine point marker while viewing the wall under a 45x microscope (Fisher Scientific Co., Fair Lawn, NJ). The working electrode is a piece of glassy carbon (GC) plate (GC-20 grade, Tokai Carbon Co., Tokyo, Japan). It was first cut to the approximate internal size of the cuvette with a glass saw, polished subsequently with fine sandpaper (grit 0000, Buehler, Ltd., Lake Bluff, IL), and finally polished on a glass polishing plate (Barnes Analytical Division, Stamford,CT) successively with 1-,0.3-, and 0.05-wm alumina slurry (Buehler,Ltd.). Prior to each polishing and after the fiial polishing, the electrode was sonicated in water. The final electrode was about 1.7 mm wide, 9.8 mm long, and 28 mm high. The geometric area of the electrode, which is exposed to solution, was 5.46 cm2. The electrical contact to the electrode was provided by clamping with a three-finger stainless steel tricep (Universal Technical Products, Inc., New York). Rigid contact to the electrode by the tricep was achieved
0003-2700/88/0360-1645$01.50/00 1988 American Chemical Society
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 15, AUGUST 1, 1988 A
B
A
C
Figure 1. Schematic diagram of the glassy carbon cuvette cell: (A) front view of the cuvette cell; (B) left-side view of the cross section S-S; (C) right-side view of the cross section S-S. Parts of the cell are the following: (a)cuvette, (b) a hole at the bottom of cuvette, (c) glassy carbon electrode, (d)three-finger tricep, (e) Teflon film, (f) wall of cuvette, (9) optical window of cuvette, (h) small holes on electrode.
m
WE
Flgure 2. Schematic illustration of the spectroelectrochemical system for the cuvette cell: (a) cuvette cell, (b) Torr Seal epoxy, (c) Teflon stopcock, (d) solution inlet, (e) solution outlet. by inserting each finger into a small hole on the top part of the GC electrode, as is shown in Figure 1C. The small holes were drilled with the two stripper (Abrasive Technology, Inc., Columbus, OH) and water was used as coolant and lubricant during the drilling process. The cuvette cell and GC electrode were assembled together with the solution filling and dumping system as shown in Figure 2. Although the quartz cuvette could be connected with the Pyrex solution filling and dumping system through a graded seal, Torr-seal resin (Varian Associates, Lexington, MA) was used to make a sealed connection because there was no contact with the solution and the resin was simple to use. The solution in the cuvette cell was removed by applying a positive N2pressure and then solution refilled by capillary action. The working electrode can be removed from the cuvette for pretreatment or surface analysis and reinserted very easily through the 3 24/40 inner joint (see Figure 2). The auxiliary electrode was a piece of Pt wire and the reference electrode was Ag/AgCl (saturated KC1). All potentials in the text will be referred to this reference. B. The construction of the second cuvette cell for a Pt working electrode (cell B) was similar to a design by Simone (18)and is shown in Figure 3. A piece of Teflon (Berghoff, Raymond, NH)
B
Figure 3. Schematic view of the Pt cuvette cell: (A) sue view of the cell, (a)to vacuum system, reference electrode, and pump, (b) solution exit line, (c)working electrode cell compartment, (d) auxiliary electrode cell compartment, (e)Teflon plug, (1) quartz cuvette; (E) front vlew of the cell, (a)to vacuum system, (b) channel for electrical connections to the working and auxillary electrcdes,(c)solution exit port, (d) working electrode, (e) solutlon entrance port. was milled to press fit into a standard 10-mm optical path length quartz cuvette with an internal volume of 3 mL. Two channels (1 cm X 0.5 cm) were milled into the Teflon on opposite sides of this Teflon plug, which defined the working electrode compartment and the auxiliary electrode compartment. The depth of these channels was approximately 5 mm. Small holes (1.5 mm diameter) were drilled in the middle of each channel and through the length of the Teflon plug. Wires to make electrical connections traversed the holes and then were soldered to the back of two Pt foils, each 1cm X 0.5 cm, and 0.1 mm thick. The Pt foils were then sealed onto the Teflon plug with TFE bonding epoxy (Norton Chemplast, Wayne, NJ). The cell thickness was defined by a piece of Teflon tape placed on top of each Pt foil, which gave a cell thickness of approximately 50 pm. Two metal plates were then placed over the tape and the Pt foils and clamped together with a C-clamp. The epoxy was allowed to dry for 48 h. Small diameter holes (0.53 mm diameter) were drilled directly below both cell compartments to serve as solution entrance ports. Another small hole (0.53 mm diameter) was drilled above the working electrode and through the length of the Teflon plug in order to serve as the solution exit port for FIA experiments. TFE tubing (0.079 cm inner diameter, Hamilton, Reno, NV) was used to connect the cell to a five-port HPLC valve (Hamilton, Reno, NV). The other ports of this valve were connected to the vacuum system (which was used to fill and empty the cell in nonflow experiments), the reference electrode (Ag/AgCl, saturated KCl), and a syringe pump. To the exit port of the cell was connected a two-way valve that could be closed for stationary experiments and opened for FIA. The working and auxiliary Pt electrodes were polished with 1-,0.3-, and 0.05-pm alumina slurry (Buehler, Lake Bluff, IL) and then concentrated nitric acid was placed dropwise on the electrodes to clean them. The Teflon plug and Pt electrodes were rinsed thoroughly with NANOpure water. Finally the Teflon assembly was placed into the cuvette and sealed with RTV silicone (Regal Plastics, Kansas City, MO) around the top of the cuvette. Instrumentation and Reagents. Electrochemistry was performed with a BAS (West Lafayette, IN) CV-1A potentiostat and the data were displayed on an X-Y recorder (Houston Instruments, Bellaire, TX). Spectral measurements were made with a Shimadzu UV-250 UV-vis spectrophotometer (Shimadzu Co., Kyoto, Japan) and a Perkin-Elmer X4B W/vis spectrophotometer (Perkin-Elmer, Norwalk, CT). The cuvette cells were mounted on an optical translator in order to align the cells with respect to the optical beam in the spectrometer. The position of the cell within the spectrometer was varied by the translator until maximum transmittance values resulted. Analytical reagent grade K,Fe(CN),, K,Fe(CN),, and KCl were from Mallinckrodt, Inc. (Paris, KY). o-Tolidine was from Paul Lewis Laboratories (Milwaukee, WI). NANOpure water (Sybron Barnstead, Boston, MA) was used to make all solutions. For FIA experiments, a
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 15, AUGUST 1, 1988
+
C
E L
3
0
,
I
I
0.5
0.4
I
0.3
I
0.2
I
I
0.1
0.0
Time
t
Figure 6. Double potential step chronocoulometric (- - -) and chronoabsorptometric (-) responses for 8.2 X M K,Fe(CN), in 1.0 M KCI. Each dlvislon in the time axis represents 38.7 s and each division on absorbance (left side) or charge (right side) axis represents either 0.0166 AU or 1.45 mC. T = 23 f 2 'C. See details in the text. h
potential v Figure 4. Cyclic voltammetric current-potential responses for 0.00 mM (- - -) and 0.10 mM (-) ferricyanide in 1.O M KCI. Scan rate was 0.43 mV/s and temperature was 23 f 2 'C.
T
i '-
b Q
Figure 5. Cyclic voltammogram of 0.60 mM ferrocyanide in 1 M KCI at a scan rate of 2 mV/s with the corresponding absorbance vs time spectra, monitored at 420 nm. The small letters on the electrochemical and spectral curves correspond to the same time after the start of the potential scan.
Rheodyne Model 7126 injector (Cotati, CA, 20-pL loop) was used. The pump was a Sage Instruments syringe pump (Model 249-2, White Plains, NY). The flow rate used throughout the FIA experiments was 0.80 mL/min.
RESULTS AND DISCUSSION Cyclic Voltammetry. Typical thin-layer cyclic voltammetric responses of the GC cuvette cell (cell A) are shown in Figure 4. The solid and dashed curves were obtained in the presence and absence of 0.1 mM K,Fe(CN),, respectively. The integrated cathodic and anodic charges, determined by integrating the currents of the CV waves, were found to be proportional to the K3Fe(CN), concentration. The cell volume calculated from the charge was 88.5 p L (f2.0 KL,seven trials). Briefly polishing the electrode with 0.05-pm alumina on a polishing cloth (Buehler, Ltd., Lake Bluff, IL) or heat-treating the electrode in a Pyrex container under a N2 atmosphere at temperatures around 300 'C ( 1 7 ) did not change the cell volume within experimental error. Therefore, any changes in the geometric dimensions of the GC electrode caused by the above polishing and heating procedures are negligible. The average solution thickness, as calculated from the cell volume and electrode geometric area, was 162 pm. The formal potential of the ferricyanide/ferrocyanide redox reaction, determined by taking an average from the anodic and cathodic peak potentials, was 0.268 V, which is in close agreement to the literature value (19). Figure 5 shows the cyclic voltammogram of 0.60 mM Fe(CN)64-at a scan rate of 2 mV/s for the Pt cuvette cell (cell
T 48rec
Figure 7. FIA detection of o-tolidine in 1 M H,S04/0.5 M CH,COOH: (A) electrochemical detection at ,Eappl = 800 mV at concentration of (a) 0.16 mM, (b) 0.12 mM, and (c) 0.079 mM of o-tolidine; (B) optical transmission detection at X = 250 nm of o-tolidine at a concentration of 0.16 mM.
B). The corresponding spectrum is also shown in Figure 5, which was taken a t 420 nm, the A, of Fe(CN)63-. The peak to peak separation was found to be 0.057 V and decreased with decreasing scan rate CAEp = 0.042 V at a scan rate of 1mV/s). The peak currents were linearly related to the scan rate, which is typical for thin-layer electrochemistry. Integration of the area under the current peaks gave a cell volume of 19.5 pL. The cell thickness, again calculated from the electrode geometric area and the cell volume, was 39.0 pm. Double Potential Step Chronocoulometry ( 8 - t ) a n d Chronoabsorptometry ( A-t ). The results of double potential Q-t and A-t experiments are presented in Figure 6 for cell A. The initial applied potential was 0.500 V. It was then stepped to 0.000 V a t tl and stepped back to 0.500 V a t t 2 . The A-t curve (solid line) was obtained by continuously monitoring the absorbance at X = 420 nm with time during the double potential step experiment. The decrease in absorbance after tl is due to the reduction of ferricyanide to ferrocyanide, which is essentially transparent at X = 420 nm. The increase in absorbance after t 2 is due to ferricyanide generation. The Q-t curve (dashed line) was obtained by drawing a smooth curve through the background-corrected charges (the solid circles). These background-corrected charges are obtained by integrating the area between the sample and background chronoamperometric current-time
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 15, AUGUST 1, 1988
responses a t certain time intervals. A typical sample current-time response, as recorded simultaneously with the A-t response, is shown in Figure 6. The background current-time responses were recorded under the same conditions as above but with a pure electrolyte solution. From both the A-t and Q-t curves in Figure 6, it can be seen that full conversion of ferricyanide to ferrocyanide, or vice versa, can be achieved within 130 s. Furthermore, from the experimental data in Figure 6, the value of the electron stoichiometry, n = 1.02, can be calculated without any knowledge of the ferricyanide concentration from the following equation:
n = ebQ/FVA where Q is the total electrolysis charge a t t 2 and A is the absorbance difference between tl and t2. The other parameters have their usual meanings. Multipotential Step, Steady-State Chronoabsorptometry. In this experiment, the potential was stepped successively from 0.500 to 0.000 V in 0.050 V incremental steps. At each potential step, the absorbance at X = 420 nm was monitored as a function of time. The potential was stepped to a new value only after the absorbance reached a steady-state value. From the absorbance a t each potential, the concentration ratio of ferricyanide to ferrocyanide, [Fe(CN)63-]/ [Fe(CN)64-], can easily be calculated with the following equation: [Fe(CN)63-] [Fe(CN)64-]
A
=-
-
Af
Ai - A
where Ai is the initial absorbance a t 0.500 V (Ei),where Ei >> E"', and Af is the final absorbance at 0.000 V (Ef). A is the steady-state absorbance at any applied potential. Based on the above calculations, a plot of applied potential vs log [Fe(CN)63-]/[Fe(CN),4-] was obtained. According to the Nernst equation, the intercept and slope of the plot give E"' = 0.265 V and n = 1.03, respectively. These values are in good agreement with the E"' value obtained in the cyclic voltammetric experiment and the n value obtained from the double potential Q-t and A-t experiments as described previously. Similar results were also obtained for the Pt cuvette cell (cell
B). F I A Experiments. o-Tolidine was used for FIA experiments since it has a relatively high molar extinction coefficient for the reduced form at X = 250 nm ( 6 = 180o0 M-l cm-') while the oxidized form is transparent at this wavelength but does absorb at X = 438 nm. o-Tolidine also exhibits a two electron transfer redox reaction in acidic medium (E"' = 0.657 V). Figure 7 shows the response for the electrochemical (E,,, = 0.800 V) and the optical transmission (A = 250 nm) detection of o-tolidine at a linear flow rate of 0.80 mL/min for the solution. In this experiment the electrochemical response was monitored while simultaneously monitoring the optical response. Calibration plots were then constructed for the electrochemical and optical responses. Both responses were linear over the concentration range of 0.16-0.01 mM and with excellent correlation coefficients ( r = 0.9932 for the electrochemical response and r = 0.9992 for the optical response). The estimated limits of detection for o-tolidine at a signal to noise (S/N) ratio of 2 was found to be 0.3 p M for the electrochemical response and 6.7 pM for the optical response. CONCLUSION The design and construction of two types of thin-layer cuvette cells with long optical path lengths have been described. Thin-layer spectroelectrochemical behaviors of the cells were confirmed by using ferri-/ferrocyanide as a test case. The inertness of the cell material and the simplicity of operation made these cells advantageous compared to earlier cell
designs. For the glassy carbon cell, the accessibility of the electrode will make this cell useful for various in situ and ex situ surface analysis techniques. Evaluation of such cells for the simultaneous measurement of transmission spectroelectrochemistry and fluorescence emission is in progress. It was shown that it was possible to perform stationary as well as flowing solution experiments with the Pt cell. For FIA, the cell can be used to simultaneously monitor the electrochemical as well as the optical responses. Preliminary data indicate that this cell may also be used to monitor the fluorescence emission of fluorescing species, some of which may be the product of the electrode reaction. The theoretical aspects of utilizing a spectroelectrochemical thin-layer flow cell with a long optical path length was investigated by Fosdick and Anderson (20,21). One of their conclusions was that the optimal arrangement for absorbance measurements was one in which the optical beam was parallel to the electrode surface and to the flow of the solution when the optical beam fills the cell volume, which was the arrangement employed in the present work. However, as noted by Fosdick and Anderson (20,21),poorer S/N ratios may result in the optical responses when utilizing a spectroelectrochemical flow cell as compared to a conventional absorbance detector because of concentration inhomogeneity within the cell caused by depletion of the absorber within the diffusion layer of the electrode. The versatility of this cell is realized by the fact that a number of different input/output signals can be utilized (stationary or flowing, electrochemical, optical transmission or fluorescence detection). The cross correlation of information from the multitude of simultaneously applied detection methods makes it possible to make direct comparisons of selectivity, sensitivity, and reliability of responses for a particular analyte in LC or FIA experiments without the need for changing the detector cell (20-22). Also, analytes that are not electroactive but may absorb in the UV-vis region or are electroactive but UV-vis transparent may be detected without the need for switching to a different detector cell or employing two detector cells in series. ACKNOWLEDGMENT The authors appreciate the free samples of Teflon film from Du Pont Co. The authors also express their appreciation to one of the reviewers (J.L.A.) for the helpful comments given. LITERATURE C I T E D (1) Kuwana, T. Ber. Bunsen-Ges. Phys. Chem. 1973, 77, 858. (2) Kuwana, T.; Winograd, N. I n Electroanalytical Chemistry; Bard, A. J., Ed.; Marcel Dekker: New York, 1974; Voi. 7. (3) Heineman, W. R. Anal. Chem. 1977, 50, 390A. (4) Heineman, W. R.; Hawkridge, F. M.; Blount, H. N. I n Necfroanalyficai Chemistry; Bard, A. J., Ed.; Marcel Dekker: New York, 1983; Vol. 13. (5) McCreery, R. L. I n Physical Methods of Chemistry; Rossiter, 8. W., Hamilton, J. F., Eds.; Wiley: New York, 1986; Voi. 2. (6) Winograd, N.; Kuwana, T. Nectroanal. Chem. 1973, 77, 858. (7) Zak, J.; Porter, M. D.; Kuwana, T. Anal. Chem. 1983, 55, 2219. (8) Gui, Y.; Kuwana, T. Langmuir 1986, 2, 471. (9) Brewster, J. D.: Anderson, J. L. Anal. Chem. 1982, 54,2560. (10) Gui, Y.; Porter, M. D.; Kuwana, T. Anal. Chem. 1985, 57, 1474. (11) Gui, Y.; Kuwana, T. Chem. Lett. 1987, 231. (12) Gui, Y.; Kuwana, T. J. Electroanal. Chem. 1987, 222, 321. (13) Gui, Y.; Kuwana. T. J. Electroanal. Chem. 1987, 226, 199. (14) Gui, Y. Ph.D. Thesis, The Ohio State University, 1987. (15) Hance, G.; Kusu, F., private communications. (16) Myers, A.; Tammela, V.; Stannett, V.; Szwarc, M. Mod. flast. 1960, 37, 139. (17) Hance, G.; Kuwana, T. Anal. Chem. 1987, 59, 131. (18) Simone, M.; Heineman, W.; Kreishman, G. Anal. Chem. 1982, 54, 2382. (19) Kolthoff. I.M.; Tomsicek, W. J. J. Phys. Chem. 1935, 39, 945 (20) Fosdick, L. E.; Anderson, J. L. Anal. Chem. 1988, 60, 156. (21) Fosdick, L. E.: Anderson, J. L. Anal. Chem. 1988, 60, 163. (22) Clark, G. J.; Goodin, R. R.; Smiley, J. W. Anal. Chem. 1985, 57, 2223.
RECENEDfor review December 29, 1987. Accepted March 29, 1988. This work was supported by the National Science Foundation through Grant CH3-8515663.
