74
Anel. Chem. 1001, 53, 74-77
Although nine factor combinations were used in this work to establish the validity of the mathematical model given by eq 3, future work would require a minimum of only five factor combinations to fit the five parameters of eq 3, thus making the optimization procedure even more efficient.
ACKNOWLEDGMENT The authors thank W. P. Price,Jr., and R. Kong for helpful discussions. We thank the Ethyl Corp. for the aniline samples. LITERATURE CITED (1) Bakalyar. S. R. Am. Lab. (Fsttield, Conn.) 1978, IO@), 43-61. (2) Box, G. E. P.; Hunter. W. G.; Hunter, J. S. "Statlstlcs for Experknentefs"; M y : New York, 1978: p 218. (3) Engehardt, H. "wP r e f m n c e LiqM Chromatography", springerV W g : New York, 1979;p 128. (4) Magan, S. L.; Demlng, S.N. Sep. PLrtt. Methods 1978, 5, 315-380. (5) S. N.; Tvoff, M. L. H. Anel. Chem. 1978, 50, 548-548. (6) h u b , R. J.; Pvnell, J. H. J . Chometw.1975, 112, 71-79. (7) Laub, R. J.; RmeH, J. H. AM/. Chem. 1976, 48, 799-803.
m,
(8) Bidhgneyer, B. A.; Demhg, S. N.; Rlce, W. P. Jr.; Sachok, B.; Pebuw&, M. J. Chromatog. 1979, 186, 419-434. (9) Io&, J. C.; Jcnka, K. M.; H u k , J. F. K. J. CX%fni?tw 1977, . 142, 671-668. (10) Hoffman, N. E.; Lab, J. C. AM/. Chem. 1971, 49,2231-2234. (11) Havath. C.; Metandec, W.; Mdnar, I.; Molnar, P. Anel. Chem. 1977, 49,2295-2305. (12) WlWner. D. P.; Nueasle. N. 0.; Haney, W. G., Jr. Anel. Chem. 1975, 47, 1422-1423. (13) Deedler, R. S.; Limen. H. A. J.; KonlWB, A. P.; van de Venne, J. L. M. J. chrometw.1979, 185, 241-256. J. 8. "A Textbook of cokld Chedsby"; m y : New York, (14) W-, 1949;p 49. (15) Jandera. P.; E-&. H. ChKWnetwapM 1980, 13, 16-25. (16) Marth, A. J. P. Bkchem. SOC. Symp. 1949, No. 3, 4-20. (17) Mendsnhall. W. "Inlroductkn to Linear MoWa and the D&gn and Anelysk of ExpamentS"; Dudwry Press: Belmont, CA, 1968:p 200. (18)o"esl. R. ~ p p l s. t e w 1971. 20,338-345. (19) S. N. A d . Chem. 1971, 43, 17261728. (20) Y a d e n , W. J. Meter. Res. Stend. 1961, 1 , 862-667.
m,
Received for review July 11, 1980. Accepted September 8, 1980.
Differential Pulse Detection in Liquid Chromatography and Its Application to the Measurement of Organometal Cations W. A. MacCrehan Orgenic Ana!vthl Research Dlvkbn, Center for Anatytkal Chmktty, Nstbnal Bureau of Stan&&,
An examination is made of m e advantages of dMerelrtlal p u b over amperometrk electrochemical detection under certain measurement drcunstances. The awed Wecttvky, hlgh serWvHy, and b w base Ine drift ofthe dmerentlai mode are discussed. The separatlon and low level detectbn of organomercury and organotin c a m are examined. flnaity, the use of differential pulse detection for the elhnlnatlon of electrode fouYng problems is demonstrated.
Electrochemical detection has become a frequently employed alternative to optical detectors in liquid chromatography, by virtue of the selectivity and sensitivity of the approach. Usually a constant potential is applied to the detector electrode and the current monitored. This has been found to be the best approach (I) for easily oxidized or reduced analytes because the sensitivity is very high and the chance of coeluting, detectable interference is small. However, for compounds that require the application of larger potentials, the selectivity of the amperometric approach is frequently inadequate. One method of improving the selectivity in such cases is to employ differential pulse detection (DPD) (2,3). This advantage and some others have come to light in the use of DPD and will be discussed in this paper. EXPERIMENTAL SECTION Apparatus. A liquid chromatograph for reductive electrochemical detection was constructed by enclosing a solvent reservoir, delivery system, and a sampling valve in an inert atmosphere glovebox (2). External controls were added for convenient manipulation of gas flow, pump speed, sampling, and replacement of the column and detector. The solvent reservoir was continuously purged with oxygen-free argon, which was transferred entirely through stainless steel tubing. The glovebox was maintained at an oxygen level of less than 0.2% by a flow of nitrogen and monitored with an oxygen analyzer.
Washington, D.C. 20234
The electrochemical detector cell is pictured in Figure 1. Nylon was c h w n as the body material by virtue of its easy machining, resistance to all LC solvents, and oxygen impermeability. The cell spacer gasket material was a 4-mil (0.1-mm) polyethylene sheet, cut to define a 3 mm wide cell. The electrodes were positioned in the thin-layer cell to minimize the electrical resistance (and hence the time constant of the cell). The values of the reaistance were determined with a capacitanecompenaated ac bridge to be 1.83 kR for the working and reference electrode pair, with a total cell value of 8.3 kR, for a 0.1 mol/L NH40Ac, 50% MeOH/H20 solvent. The cell is connected to the column with stainless steel tubing, with a drilled-out Omnifit Teflon 1/18 in. to in. 28 end fitting. Frequently, a W/visible absorbance detector was connected to the outlet of this low dead volume cell. A PARC Model 174 A polarographic analyzer was used for both amperometric and differential pulse detection modea. Undistorted chromatograms were obtained with a 3-s added time constant in the amperometric mode and a 0.3-s added time constant (0.5s pulse time) in the differential pulse mode. Reagents. Methanol (MeOH) was obtained from Burdick and Jackson Laboratories. The aqueous NH,OAc was made from reagent grade acetic acid by adding sufficient ammonia to adjust the pH. This solution was then purified for at least 1 week by continuous electrolysisat a mercury cathode. The organometal reagents were obtained from Alfa Products and were used as received.
RESULTS AND DISCUSSION Selectivity. A significant increase in the selectivity of electrochemical detection can be obtained with the use of DPD, when compared to simple amperometry. Figure 2 shows a hypothetical pulse voltammogram of three coeluting species. A base potential (either E b l or Eb2)for DPD is chosen near the half-wave potential of the analyte, periodically a potential pulse (hEl or hE2)is superimposed, and the difference between the current before and a t the end of the pulse is measured. Clearly, only the compound that has its voltammetric response within the pulse width will be detected. The
Thls article not s u b p to U.S. Copyright. PUMlshed 1980 by the American chemical Sodely
ANALYTICAL CHEMISTRY, VOL. 53, NO. 1, JANUARY 1981
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Flgurr 3. Comparison of the selecthrky of a m p e r m t r y and DPD for a 40 ng/g “spUte” of oyster tissue workup. The first chromatogram lsdone with amperometricdetectbnat -0.8 V. The second USB% DPD with A € = 100 mV, E, = -0.70 V. The third is done with A€ = 25 mV with Eb-0.725 V. The final chromatogam is done with A € = 5 mV and Eb= -0.73 V. Chromatogaphic conditions: column, Akex Sphertsorb ODs (5 pm) 0.46 X 25 cm; mobile phase, 40% M#H/ H2O. 0.1 W L NH‘OAC, pH 5.2, 0.01 % V/V PWCaptOethanOl: ROW rate, 1.0 mUmin. Flgwo 1. Electrochemical detector cell. (1) Reference electrode Ag/AgCI, 3 m l / L C r ; (2) threaded Teflon hdder; (3) R auxilary electrode; (4) assembly screw; (5) solution outlet; (6) Nykn body; (7) porous Vycor frit; (8) polyethylene cell gasket: (9) solutbn Inlet; (10) working electrode: gold amalgamated with mercury electrode (GAME), 1.0 mm diameter.
nBu,Sn+
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Potential F l g m 2. The principle of differential pulse detectkn. A€ is the pulse height; k the base potential.
selectivity can be controlled by the selection of the magnitude of the pulse height. Of the two pulse heights shown, AE2 will provide greater selectivity than A&. Another important feature becomes quite apparent, the DPD signal will increase with increasing pulse height up to a point, with an upper bound determined by the S shape of the current-voltage curve. AEl will provide the maximum signal intensity. However, as will be shown later, the baseline noise increases dramatically with increasing pulse height. Thus, in order to achieve the highest signal-to-noise ratio, a pulse height is chosen that is a compromise between signal intensity and base-line noise. Figure 3 shows a comparison of amperometric and DPD with three pulse heights (100, 25, and 5 mV) for a dilute “workup” of NBS SRM 1% oyster tissue (sample preparation as in ref 2). The methylmercury content is not evident with amperometric detection because other reducible, coeluting species in the sample are detected and obscure the methylmercury signal. However, by choosing DPD and carefully controlling the pulse height, the methylmercury is easily measured without interference. The methylmercury content of this SRM could only be estimated, because the “workup” was very close to the limit of detection (LOD). Reaults of the standard addition procedure using a 10-mV pulse height indicated approximately 50 ng/g, which agrees quite closely with the total mercury value certified to be 57 f 15 ng/g. Although DPD greatly increases the selectivity of electrochemical detection, it does not preclude simultaneous mul-
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8 10 12 14 16 18 20 22 24 Minutes
w e d . Separatbnofsomeorgenothcetkns. condttkns: dum, Whatman PXS catbn exchange, (10 pm) 0.46 X 25 an:roWe phase, 6 0 % MeOH/H20, 0.042 mdR FW,OAc, pH 5.3, Row rate 1.0 mUrnh: sample, 1 X I O 4 md/L (approximately 30 pglg) of each; A € = 100 mV, E, = -1.10 V.
ticomponent analyses for compounds of similar behavior, such as species with the same electroactive functional group or for a homologous series. Figure 4 shows the simultaneous d e tection of tributyl-, triethyl-, and trimethyltin cations by use of a 100-mV (the least selective) DPD pulse height. The separation of these organotin species was first accomplished by Jewett and Brinckman at NBS (4, 5) using a graphite furnace atomic absorption LC detector approach. Methyl-, ethyl-, and phenylmercury can also be simultaneously detected with DPD (2). Sensitivity of DPD Compared to Amperometry. It has been stated, in earlier work on DPD in LC by Swartzfager (31, that the LOD was usually a factor of 10-20 times poorer than for amperometric detection, resulting from the unwanted capacitive current encountered by using the pulsed potential. However, we have observed that in many cases a t mercury electrodes, the LOD is quite similar (6).The limiting base-line noise with amperometric detection, in many c a w , is caused by variations in the residual faradaic current, resulting from flow rate variations. The noise is a function of the magnitude of the residual current and of the ratio of the flow variation to the total flow rate (3). In order to minimize the baseline noise, it is necessary to maintain a constant and very low faradaic current and also to provide pulse-free flow. Most LC pumps providing high-preasure solvent delivery employ pistons
76
ANALYTICAL CHEMISTAY,
VOL. 53, NO. 1, JANUARY 1981
and flow check valves. Although pressure feedback systems have been added to many pumps to lower the pulsations, no such pump can provide completely pulse-free flow. Pulse dampeners may be used (6) but are often cumbersome for routine work, as a result of their large holdup volumes. I t should be stressed that the flow noise will be the determinant noise only under conditions when the residual current is a result of flowsensitive faradaic reactions. Reactions that are not mans-transfer controlled, such as proton reduction in acidic media at mercury electrodes, can result in detector residual current that is independent of flow (7). The base-line noise in DPD will arise not only from faradaic processes but also from the flow of capacitive current at the pulsed electrode. In addition, the noise introduced by the measurement instrumentation is more significant in DPD than in amperometry. One contribution to the noise is the much shorter time that the signal is actually being measured. In amperometry the signal is measured continuously and optimal filtering can be applied. However, in DPD the electronic measurement time is much less, in this case 16 ms/500 ma or 3.2% of the time. For random noise, the signal-to-noise ratio is expected to increase with the square root of the signal observation time; thus a lower signal-to-noise ratio is expected for the sampled DPD experiment. However, the DPD measurement provides some signal-tonoise advantages that help offset the increased capacitive and electronically added noise. First, under the conditions used in this work, the thickness of the diffusion layer is controlled predominently by the pulse not by the hydrodynamic flow (3). Since the thickness of the diffusion layer is smaller in DPD than amperometry, some signal enhancement is achieved over steady-state (nonpulsed) conditions. Another result of the pulsed experiment is that flow pulsation will have a greatly reduced effect on the faradaic components of the background noise, eliminating the most frequent limiting noise in amperometric LC detection. The second benefit of using DPD in measurementa of high sensitivity is one that is common to all differential approaches: linear changes in the background intensity become a constant. The base-line drifts resulting from changing levels of detectable impurities in the solvent or from electrode surface changes frequently make the practical measurement of small faradaic signals very difficult. This is especially true in reductive LC detection, where days are often required to reach the lowest residual currents (resulting from the oxygen reaction) and drifting base lines may remain a severe problem. Since DPD measures only the change in slope, a linear base-line drift is eliminated, by virtue of the differential measurement. The optimum signal-to-noise ratio in DPD will also be dependent on two controllable experimental parameters: the base potential E b and the pulse height AE. The optimum E b for a given AE can be chosen by examination of the voltammogram of Figure 2. Pulses that are symmetrical about the El12of the compound of interest will provide the maximum signal for the chosen a.The optimum criterium will be &, = E I I z- A E / 2 for reducible analytes and both (MI, Ebl) and (Up, Eb2)fulfill this relationship. As will be shown later, asymmetric (M,Eb) pairs may provide detection advantages that overcome electrochemical difficulties, that in some casea are worth the attendant loss in signal compared to symmetric (AE,Eb) pairs. The signal-to-noise ratio as a function of symmetric (a,, Eb) pairs was studied for MeHg+. The signal increases quite sharply as a function of AE from 5 to 25 mV but begins to plateau as AE approaches 100 mV. In contrast, the residual current noise rises exponentially with increasing AE. Thus, as Figure 5 shows, an optimum AE for methylmercury de-
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by use of c h r m t o g a p h k conditions of Figure 3. Noise was measued p e a k - t v k , wlth an added tlm constant of 0.3 s. Values of (A€, €Jused w e (5mV,-0.77V), (10mV,-0.77V), (25mV,-0.76
V), (50 mV, -0.74 V), and (100mV, -0.72V).
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2(4 6 81012 Minutes 6. Low-level detectkn of organomtals with DPD. condltkns: (upper figue) separation as in Figue 3; A € = 100 mV, Eb= -0.73 V; sample, 1 .O x lo4 m o ( (2.2 ~ ng/g o( 40 p ~ ) (lower me) as Figue 4 except concentration, 1.0 X 10 mol/L (approxlmately 250 pg/g or 6 ng), A € = 100 mV, Eb = -1.10 V. Inject
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tection exists and is near 25 mV. Figure 6 shows two chromatograms near the LOD for two cases employing DPD using AE of 100 mV. The upper chromatogram for methylmercury is obtained a t Eb = -0.73 V and where the residual current is relatively low. The lower chromatogram, for tributyl- and triethyltin is obtained under much less favorable conditions of capacitive and faradaic current a t Eb = -1.10 v. I t is clear that the detection limit is degraded in this latter case. When the LOD of DPD is compared to amperometric detection, the results are about the same for methylmercury, but alkyltin cations cannot be reliably measured by amperometry, for reasons given in the following section. Elimination of Electrode Fouling Reactione. A serious problem in the electrochemical detection of certain analytes is the passivation of the electrode surface by the products of the detection reaction. Such a problem exists with the oxidation of phenols (8 and references therein) which form electroinactive phenoxy polymers. We have encountered a similar problem with the amperometric detection of the organotin cations. The upper portion of Figure 7 shows the
ANALYTICAL CHEMISTRY, VOL. 53, NO. 1, JANUARY 1981
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current-potential wave is chosen, such as Ebl in Figure 2. Thus, during about 87% of the reductive “duty cycle” (444 ms), the Ph3Sn+ is not being electrolyzed. When the 56-ms pulse is superimposed (representing about a 13% active duty cycle) the Ph&n+ is converted to Ph&n.. The lowering of the duty cycle means that the concentration of P h a n . available for the dimerization reaction will be much less. Furthermore, because of the electaochemical reversibility of reaction 1, much of the Ph& is reoxidized when the electrode is returned to the base potential. The formation of the electrode fouling product P h S n z is suppressed so that a completely pctsitive response is obtained (in the lower half of Figure 7),that varies linearly with concentration in the 104-10” mol/L range.
I
CONCLUSIONS DPD can provide a useful adjunct to amperometric electrochemical detection in LC in cases where the selectivity of the latter is inadequate. Although the LOD of DPD is equal to or poorer than that of amperometry, the enhanced selectivity may make it possible to measure smaller amounts of analyte without interference in complex matrices.
Minutes
Flgm 7. Triphenytth detection by amperomeby and DPD. COnBtbns: as Flgwe 4 except 20 pL of 1.0 X lo4 molL &%+; (upper figtre) amperomtry at -1.00 V; (bwer m e ) DPD with A € = 100 mV, E b = -0.85 v (asymmetric c h o h of Eb).
amperometric response of Ph3Sn+. Initially a reductive response is obtained for the reaction Ph3Sn+
+ e- + Ph3Sn.
(1)
which is an electrochemically reversible reaction but which is followed by a chemically irreversible dimerization (9). Ph3Sn. Ph&. Ph&n2 (2)
+
77
-
The hexaphenylditin is electroinactive and adsorbed on the electrode surface. This product passivates the electrode toward the solvent discharge reaction, decreasing the faradaic current, and causing a “dip” in the base line. The electrode remains passivated only as long as it takes the flowing solvent to “elute” the adsorbed product from the electrode. A similar problem and reaction sequence also exist for the alkyltin cations, even at injected concentrations as low as IO4 mol/L. This problem can be handily overcome by the use of DPD. Rather than choosing the symmetrical base potential and pulse height pair, a base potential at the “foot” of the voltammetric
ACKNOWLEDGMENT The author wishes to thank L. Doane, R. Durst, and Bertocci for helpful discussions.
U.
LITERATURE CITED (1) (2) (3) (4)
(5) (6)
(7) (8)
(9)
KLsehger, P. T. Anal. chem.1977, 49, 445A-458A. MecCrehan, W. A,; Dwst, R. A. Anel. Chem. 1878. 50. 2106-2112. Swartzfager, D. Q Anel. chem.1976, 48,2189-2192. Jewett. K. L.; Brlndtman, F. E.; Btat. W. R. “Abstracts of Papers”. 178th Natbnel Meet@ of the Americen -1 society. Washlng ton. DC, Sept 1979: Amedan chemlcel Sockty: Waehlngton, DC, 1979; ANAL 128. Jewett, K. L.; Blat, W. R.; Brhdtman. F. E. 6th Intematknal S y m p skm on the W d e d Release of Bkactjve Materials; Ne~wOrleene, LA, 1979. MecCfehan, W. A.; W e t , R. A. EPA-806/7-7Q-211; E n v t m t a l Rotectbn Agency: Washhgton, DC, 1979. Metson. W.; Dnk. E.; Wevltch, R. Am. Lab. ( F e w , ccw?n.) 1877, &My,59-73. Kale, R. C.; Johnson. D. C. A d . Chem. 1878, 51, 741-744. Booth, M. D.; Fleet, B. Anal. Chem. 1970, 42, 825-831.
RECEIVED for review September 12,1980. Accepted October 20, 1980. The author wishes to thank the Environmental Protection Agency for partial support of this work through the Interagency Energy/Environment Program (EPA-IGA05E684). The specification of commercial products does not imply endorsement by the National Bureau of Standards.
