Electrospray Mass

Electrochemistry coupled on-line with mass spectrometry, i.e., electrochemical mass spectrometry (EC/MS), allows identification of electrochemically g...
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Anal. Chem. 1996, 68, 4244-4253

On-Line Probe for Fast Electrochemistry/ Electrospray Mass Spectrometry. Investigation of Polycyclic Aromatic Hydrocarbons Xiaoming Xu, Wenzhe Lu, and Richard B. Cole*

Department of Chemistry, University of New Orleans, Lakefront, New Orleans, Louisiana 70148

A newly invented probe accessory for fast electrochemistry/ electrospray mass spectrometry (EC/ESMS) is presented and evaluated. The device features a low-volume, threeelectrode electrochemical cell which has been designed with a minimum distance between the working electrode and the “Taylor cone” inherent to the electrospray process. This configuration limits the time between electrochemical generation of ions and mass spectrometric analysis to an absolute minimum. A fused-silica layer insulates the microcylinder working electrode from the sample solution until immediately prior to the electrospray region, postponing electrode processes until the last moment. The same fused-silica layer insulates the working electrode from the surrounding auxiliary electrode, a stainless steel capillary that also serves as the electrospray capillary. The performance and capabilities of the novel electrochemistry/electrospray mass spectrometry system have been evaluated using polycyclic aromatic hydrocarbons (PAHs) as test analytes. In the positive ion EC/ ESMS mode, oxidized forms (one-electron removal) of PAHs are produced in high yield. The ability to analyze reaction products appearing subsequent to the initial oxidation is also demonstrated. The coupling of electrochemistry to other analytical techniques can offer valuable information to aid in the elucidation of mechanisms of complex electrochemical reactions. Spectroscopic techniques (UV/visible, IR, Raman, EPR) and mass spectrometry have been coupled with electrochemistry to monitor and characterize intermediates and products generated by electrode processes. Surface analysis (Auger electron spectroscopy, X-ray photoelectron spectroscopy) and topographical imaging techniques (scanning electron microscopy, scanning tunneling microscopy, atomic force microscopy, etc.) have been used to study electrochemical deposition and adsorption to electrode surfaces. Among these analytical techniques, mass spectrometry offers some unique advantages in terms of sensitivity and specificity. Electrochemistry coupled on-line with mass spectrometry, i.e., electrochemical mass spectrometry (EC/MS), allows identification of electrochemically generated species. EC/MS has become a powerful tool in electroanalysis and electrochemical kinetics studies. It has found applications in many areas such as electrocatalysis, electrosynthesis, batteries, chemical sensors, and corrosion. The idea of determining electrochemical reaction products by coupling the electrochemical cell on-line with a mass spectrometer was first concretized by Bruckenstein and co-workers1-5 in the 4244 Analytical Chemistry, Vol. 68, No. 23, December 1, 1996

1970s. This methodology was further developed by Brockman and Anderson,6 Wolter and Heitbaum,7 and Vielstich and coworkers.8,9 In early experiments, porous working electrodes or permeable membranes were usually used as the interface between the cell and the MS ionization source. More recently, House and Anderson10 reported a new approach to coupling an electrochemical cell to a mass spectrometer by employing a cell containing an interdigitated electrode pair. In all of the above-mentioned EC/ MS devices, electron ionization (EI) was used prior to mass analysis. As a result, only gaseous, volatile, or semivolatile products of electrochemical reactions could be detected by MS. The inability to detect polar, nonvolatile products and intermediates of electrochemical reactions limited the applicability of EC/ EI-MS when the majority of electrochemical intermediates and products were not volatile. An important development in on-line electrochemistry/mass spectrometry was made by Hambitzer and Heitbaum11 when they connected an electrochemical cell to a thermospray (TS) mass spectrometer. This device was used to detect electrochemically generated products such as dimers and trimers which were formed during the electrooxidation of N,N-dimethylaniline. A response time (i.e., the time between the application of voltage to the working electrode and the appearance of a mass spectral signal response corresponding to an electrogenerated product ion) of 9 s was observed. Another electrochemistry/thermospray device was reported by Volk, Yost and Brajter-Toth.12 They used EC/TS/MS/MS to study the oxidation of uric acid and 6-thiopurine (an antitumor drug) and to study metabolism of xenobiotic compounds.12-14 A response time of 500 ms was reported. Because an aqueous ammonium acetate buffer solution was used in the thermospray process, all detected species appeared as either protonated molecules or ammonium adduct ions in the positive (1) Bruckenstein, S.; Gadde, R. R. J. Am. Chem. Soc. 1971, 93, 793-794. (2) Petek, M.; Bruckenstein, S.; Feinberg, B., Adams, R. N. J. Electroanal. Chem. 1973, 42, 397-401. (3) Petek, M.; Bruckenstein, S. J. Electroanal. Chem. 1973, 47, 329. (4) Gadde, R. R.; Bruckenstein, S. J. Electroanal. Chem. 1974, 50, 163-174. (5) Grambow, L.; Bruckenstein, S. J. Electrochim. Anal. 1977, 22, 377-383. (6) Brockman, T. J.; Anderson, L. B. Anal. Chem. 1984, 56, 207-213. (7) Wolter, O.; Heitbaum, J. Ber. Bunsenges. Phys. Chem. 1984, 88, 2-6. (8) Vielstich, W. Symposium Electrode Materials and Processes for Energy Conversion and Storage; Srinivasan, S., Wagrer, S., Wroblowa, H., Eds.; The Electrochemical Society Inc.: Pennington, NJ, 1987; p 394. (9) Wasmus, S.; Cattaneo, E.; Vielstich, W. Electrochim. Acta 1990, 35, 771. (10) House, S. D.; Anderson, L. B. Anal. Chem. 1994, 66, 193-199. (11) Hambitzer, G.; Heitbaum, J. Anal. Chem. 1986, 58, 1067-1070. (12) Volk, K. J.; Yost, R. A.; Brajter-Toth, A. Anal. Chem. 1989, 61, 1709-1717. (13) Volk, K. J.; Yost, R. A.; Brajter-Toth, A. J. Electrochem. Soc. 1990, 137, 17641771. (14) Volk, K. J.; Yost, R. A.; Brajter-Toth, A.; Freeman, J. A. Analusis Lett. 1992, 20, 421. S0003-2700(96)00362-9 CCC: $12.00

© 1996 American Chemical Society

ion mode or deprotonated molecules or acetate adducts in the negative ion mode. Radical cations and radical anions were not detected. Electrospray ionization (ES) is a so-called “soft” ionization technique for mass spectrometry. It can convert analytes in solution at atmospheric pressure, directly into gas-phase ions with minimal fragmentation. Since the pioneering work of Fenn and co-workers,15-17 ESMS has emerged as one of the major breakthroughs in the development of analytical mass spectrometry for the analysis of nonvolatile compounds.18-22 The compounds most amenable to ESMS are ionic compounds and compounds that can be readily ionized in solution by acid/base reactions. Analyte ions in ESMS are usually considered to be either “preformed” ions (i.e., they already existed in ionic form in solution) or neutral molecules attached to (solvating) monatomic ions which constitute the charge excess present in electrosprayed droplets. In general, neutral and nonpolar compounds are not well-suited for ESMS analysis. However, in ground-breaking experiments, Van Berkel et al.23 reported that radical cations of certain nonpolar analytes, such as metalloporphyrins and polycyclic aromatic hydrocarbons, were detectable by ESMS. We have reported ESMS studies of metallocenes,24 including a series of substituted ferrocenes. Intact molecular cations of these compounds were generated by electrochemical oxidation (electron removal) at the ES needle. These results and those of Van Berkel et al.23 affirmed the description of the electrospray device as a special type of electrochemical cell, as proposed by Blades et al.,20 who first established that the mass spectrometer could detect ions that were produced electrochemically during the electrospray process; but the ability of electrospray devices to generate molecular cations from neutral compounds has been rather limited. Abundant molecular ion signals can only be obtained from compounds having relatively low E1/2(ox) values (roughly, those below +1.0 V vs SCE) and compounds whose oxidized forms are stable in solution.23,24 Van Berkel and Zhou25 have analogized the electrochemical reactions taking place in a standard electrospray ionization source to those occurring in a controlled-current electrolytic cell. Several adjustable ES parameters, such as solution flow rate, applied voltage or current, and solution properties including reactivity visa`-vis electroactive species, were shown to influence the nature and efficiency of electrochemical reactions occurring at the metal/ solution interface. To exploit the inherent oxidation capacity of the ES source, Van Berkel and Zhou25 suggest increasing the ES current, eliminating extraneous (nonanalyte) readily oxidizable species from the solution, and using an electrospray capillary (15) Yamashita, M.; Fenn, J. B. J. Phys. Chem. 1984, 88, 4451-4459. (16) Whitehouse, C. M.; Dreger, R. N.; Yamashita, M.; Fenn, J. B. Anal. Chem. 1985, 57, 675-679. (17) Fenn, J. B.; Mann, M.; Meng, C. K.; Wong, S. F.; Whitehouse, C. M. Mass Spectrom. Rev. 1990, 9, 37-70. (18) Huang, E. C.; Wachs, T.; Conboy, J. J.; Henion, J. D. Anal. Chem. 1990, 62, 713A-725A. (19) Ikonomou, M. G.; Blades, A. T.; Kebarle, P. Anal. Chem. 1991, 63, 19891998. (20) Blades, A. T.; Ikonomou, M. G.; Kebarle, P. Anal. Chem. 1991, 63, 21092114. (21) Smith, R. D.; Loo, J. A.; Edmonds, C. G.; Barinaga, C. J.; Udseth, H. R. Anal. Chem. 1990, 62, 882-889. (22) Cole, R. B.; Harrata, A. K. J. Am. Soc. Mass Spectrom. 1993, 4, 546-556. (23) Van Berkel, G. J.; McLuckey, S. A.; Glish, G. L. Anal. Chem. 1992, 64, 1586-1593. (24) Xu, X.; Nolan, S. P.; Cole, R. B. Anal. Chem. 1994, 66, 119-125. (25) Van Berkel, G. J.; Zhou, F. Anal. Chem. 1995, 67, 2916-2923.

(needle) composed of electrode material that does not readily oxidize, such as gold. The current report presents a novel three-electrode electrochemical device, which has been coupled on-line with electrospray mass spectrometry (EC/ESMS) in order to enable the analysis of neutral compounds that are typically not amenable to ESMS and to offer the possibility to monitor and identify ionic intermediates and products generated by electrochemical reactions. Results stemming from the coupling of three-electrode electrochemical systems to electrospray mass spectrometry first appeared in 1995.26,27 The following difficulties must be overcome before coupling of an electrochemical cell with ESMS can be successful: (1) excessively lengthy response time between generation of products and MS detection caused by the low flow rate typically employed; (2) high (electrospray) voltage hazard to the electrochemistry hardware (e.g., potentiostat) and to the operator; and (3) analyte signal suppression caused by the presence of high concentrations of supporting electrolytes. In this paper, we describe some novel experimental approaches that have been used to largely overcome these difficulties. As an example of the utility of the EC/ESMS system, presented herein, we report results of the electrochemical oxidation of a series of polycyclic aromatic hydrocarbons (PAHs).26 The different abundances observed for radical cations of PAHs have been rationalized on the basis of their stabilities and structural features. Some intermediates and products generated from ensuing chemical reactions have been assigned and compared with previously proposed literature mechanisms. EXPERIMENTAL SECTION 1. Electrochemistry Instrumentation. For on-line EC/ ESMS experiments, a potentiostat (LC-4B Amperometric Controller, Bioanalytical Systems, Lafayette, IN) was used to control the potential applied to the electrochemical cell and to measure the resulting current. To reduce the potential hazard created by floating the cell at the electrospray high voltage (HV), the power sources of the electrochemical instruments were replaced with rechargeable batteries. The entire electrochemical system was placed in a safety box for shielding and was then floated at the electrospray HV by connecting the frame of the electrochemical instruments to the HV power supply. During operation, the electrochemical experimental parameters were selected before turning on the HV. Typically, optimized voltages fell in the range of 2-3.5 kV. To minimize the risk of electric shock, the potentiostat was switched from “standby” to “on” via 7-in.-long polyethylene “knob”. In off-line linear sweep experiments, a CV27 Voltammograph electrochemical system (Bioanalytical Systems) was used to measure basic electrochemical parameters, such as the accessible potential range of the solvent. Solution conductivity measurements were performed using a conductivity bridge (Model RC 16B2, Beckman Instruments, Cedar Grove, NJ). 2. Mass Spectrometer. A quadrupole electrospray mass spectrometer (Vestec 201, PerSeptive Biosystems, Houston, TX) was used in all EC/ESMS experiments. Reported mass spectra represent the average of at least 20 scans obtained at scan rates of ∼0.1-0.5 s/scan. (26) Xu, X.; Lu, W.; Suleiman, A. A., Cole, R. B. Proceedings of the 43rd ASMS Conference on Mass Spectroscopy and Allied Topics, Atlanta, GA, May 2126, 1995; p 254. (27) Van Berkel, G. J.; Zhou, F. Proceedings of the 43rd ASMS Conference on Mass Spectrometry and Allied Topics, Atlanta, GA, May 21-26, 1995; p 900. (28) U.S. Patent applied for.

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Figure 1. Schematic representation of the on-line fast electrochemistry/electrospray mass spectrometry probe. A platinum microcylinder (2) serves as the working electrode. The fused-silica layer (4) postpones electrical contact between the working electrode and the sample solution until the latter reaches the electropray region. The fused-silica layer also insulates the working electrode from the auxiliary electrode (8); the latter functions as the electrospray capillary. The above electrode assembly is held in place by ferrules (9) on a PEEK “cross” (7), as is the Ag/Ag+ (0.01 M in CH3CN) reference electrode (6), which is isolated from the sample solution by a Vycor glass tip. Sample is infused in port 10, and carrier gas to pneumatically assist the electrospray process may be introduced in port 12 of a stainless steel tube (14).

3. Reagents. All solvents used were HPLC grade and were purchased either from J.T. Baker (Philipsburg, NJ) or from E.M. Science (Gibbstown, NJ). Acetonitrile was preliminarily dried with calcium hydride (Strem Chemicals Inc., Newburyport, MA) for 24 h followed by distillation over phosphorus pentoxide and storage over CaH2 in a desiccator. Methylene chloride was dried by passage through an activated alumina column (preheated for 7 h at 175 °C) followed by storage over alumina. Supporting electrolytes, lithium trifluoromethanesulfonate, and tetrabutylammonium perchlorate (TBAP; Fluka Chemical Corp., Ronkonkoma, NY) were dried in small quantities (∼100 mg) at 110 °C for 3 h before use. All PAHs were purchased either from Sigma (St. Louis, MO) or Aldrich (Milwaukee, WI) and were used without further purification. RESULTS AND DISCUSSION Construction of the Electrochemical Cell. The experimental setup for the on-line “fast electrochemistry/electrospray mass spectrometry” system28 is shown in Figure 1. An electrochemical flow cell was constructed inside the electrospray probe. The threeelectrode cell consists of a platinum microcylinder (0.127-mm diameter, 1.5-mm length wire) flame-sealed inside a cylindrical fused-silica capillary (0.170-mm i.d. 0.300-mm o.d.) as the working electrode. The fused-silica layer prevents electrical contact between the working electrode and the sample solution until immediately prior to the electrospray region. It also insulates the working electrode from the auxiliary electrode, a stainless steel capillary (0.35-mm i.d., 0.405-mm o.d.) which surrounds both the working electrode and the fused-silica layer. This auxiliary electrode also functions as the electrospray capillary. The reference electrode is an Ag/Ag+ (0.01 M in CH3CN) electrode (Bioanalytical Systems) which was isolated from the sample solution by a Vycor (Dow-Corning, Midland, MI) glass tip (Figure 1) that allowed migration of ions but prevented flow of solution. Because there is a limited space inside the electrospray probe (∼16 mm in diameter), the reference electrode compartment must be placed outside of the probe. The sample solution is delivered by a syringe pump (Model 341B Sage Instruments, Boston, MA) to the entry of a PEEK 4246 Analytical Chemistry, Vol. 68, No. 23, December 1, 1996

“cross” (Upchurch Scientific, Oak Harbor, WA). The solution subsequently passes through the annulus between the outer wall of the fused-silica layer and the inner wall of the stainless steel auxiliary electrode until it reaches the vicinity of the exposed portion of the working electrode. The polyimide coating of the fused silica was resistant to all solvents used in this study. Immediately following electrochemical reaction at the working electrode, the sample solution undergoes electrospray directly into the mass spectrometer. A carrier gas can be employed to pneumatically assist the electrospray process. The most novel and important feature of this electrochemical probe is that it generates electrochemical intermediates (e.g., radical cations) and products in situ at the tip of the electrospray capillary (needle). This feature distinguishes it from all other EC/ ESMS systems reported to date employing either two-electrode29 or three-electrode30 arrangements. The latter three-electrode cell was decoupled from the electrospray high voltage by introducing a grounded metal connector between the electrochemical cell and the electrospray region. This arrangement resulted in a 30-cm distance between the working electrode and the electrospray capillary exit.30 By contrast, in the EC/ESMS design shown in Figure 1, electrogenerated species are produced immediately prior to delivery into the mass spectrometer, thus keeping response time to an absolute minimum. Another feature of this design is that it is a type of annular cell (average distance between working and counter electrode is 0.112 mm) wherein the iR drop was estimated to be only 0.0467 V for a 5 × 10-4 M electrolyte solution (see Appendix), Low concentrations of competing electrolytes are crucial for maximizing sensitivity in electrospray mass spectrometry.31,32 The volume of the prototype cell depicted in Figure 1 is very small (annulus volume surrounding the working electrode ∼0.125 µL), while the surface area of the working electrode is relatively large (∼0.60 mm2). These features are important to maintaining a favorable electrochemical conversion efficiency, (29) Bond, A. M.; Colton, R.; D’Agostino, A.; Downard, A. J.; Traeger, J. C. Anal. Chem 1995, 67, 1691-1695. (30) Zhou, F.; Van Berkel, G. J. Anal. Chem. 1995, 67, 3643-3649. (31) Kebarle, P.; Tang, L. Anal. Chem. 1993, 65, 972A-986A. (32) Wang, G.; Cole, R. B. Anal. Chem. 1994, 66, 3702-3708.

Table 1. Properties of Solvents Commonly Used in Electrochemistry36

a

solvent

boiling point (°C)

dielectric consta

viscosity at 15 °C (cp)

potential limits (V vs SCE)

acetonitrile N,N-dimethylformamide propylene carbonate methylene chloride nitromethane nitrobenzene methanol water

81.6 153 241.7 39.8 101 210.9 64.7 100

37.5 (25) 36.7 (25) 69 (25) 9.08 (20) 36.7 (20) 34.82 (25) 32.63 (25) 80.10 (20)

0.375 0.92 ( 20 °C)

-3.5 to +2.4 -3.5 to +1.5 -2.5 to +1.7 -1.7 to +1.8 -1.2 to +2.7

0.449 0.620 (25 °C) 2.24 0.623 1.139

-2.2 to +1.5 -2.7 to +1.5

nucleophilicity moderate moderate moderate low low high high

Numbers in parentheses, temperature (°C).

while also minimizing band broadening in cases where a solutionbased separation (e.g., liquid chromatography or capillary electrophoresis) is performed with introduction of the effluent into the electrochemical cell. A potential drawback of this cell design is the need for cleaning and regenerating the surface of the working electrode. The level of fouling is quite analyte dependent, and for PAH measurements, signal deterioration was noticeable after 1-2 h of accumulated run time. However, the following procedure was found to be quite effective for regenerating an active working electrode surface after electrooxidation of PAHs: (1) the cell (needle) tip is dipped in acetonitrile contained in a 100-W ultrasonic bath for 2 min while a total of 200 µL of acetonitrile is pumped through the cell to dissolve salts and polar compounds; (2) the first step is repeated using toluene as the solvent to remove electro-deposited PAHs; (3) the cell is rinsed with 200 µL of CH2Cl2; (4) the applied potential is switched between +0.5 and -0.5 V (holding the voltage at each value for 10 s) while 100 µL of electrolyte solution (blank) is pumped through the cell for 2 min at a flow rate of 50 µL/min. This electrode pretreatment procedure enabled the acquisition of quite reproducible results. Optimization of the EC/ESMS System. Several factors influenced the performance of the prototype EC/ESMS system. These factors included the configuration of the electrochemical cell, the solvents employed, the supporting electrolytes added, the solution flow rate, and the probe tip temperature. The effects of these factors were investigated and optimized in order to achieve the fastest response time and the highest detection sensitivity. (1) Solvents. In selecting solvents for use in on-line EC/ ESMS, the requirements of both the electrochemical cell and the electrospray mass spectrometer must be taken into account. The dielectric constant, accessible potential range, surface tension, boiling point, viscosity, and reactivity toward radical ions are all factors that influence the success of the EC/ESMS experiment. The properties of a number of solvents commonly used in electrochemistry are listed in Table 1. Acetonitrile is probably the most widely used nonaqueous solvent for organic electrochemistry owing to its high dielectric constant (high polarity), wide accessible potential range (-3.5 to +2.4 V vs SCE), and convenient range of temperatures over which it stays liquid (-45 to +81.6 °C). The intermediate polarity, volatility, and low viscosity of acetonitrile also make it a good solvent for ESMS. Due to its nucleophilicity, however, it is not preferred for EC/ ESMS detection of radical cations of PAHs. For the same reason, neither methanol nor water should be used as a solvent in electrooxidation of PAHs, unless it is desirable to add a nucleophile

or methoxylating reagent for a particular electrochemical reaction. It has been reported that radical cations of PAHs are considerably more stable in nitrobenzene than in acetonitrile,33 but the volatility of nitrobenzene is rather low (high boiling point) for convenient use as a solvent for ESMS. For similar reasons, propylene carbonate is not preferred as a solvent for ESMS. Dimethylformamide (DMF) is less nucleophilic than acetonitrile, but its low volatility, high viscosity, and narrow anodic potential limit make it undesirable for an EC/ESMS oxidation study. Methylene chloride has useful electrospray properties (high volatility, low viscosity), and its anodic limit is wide enough to view the oxidation of most PAHs. In particular, methylene chloride offers considerable stability to radical cations of PAHs and other molecules, an important consideration in EC/ESMS studies. A drawback of methylene chloride, however, is its low dielectric constant and low solvent power; few salts dissolve well in methylene chloride. Although no single solvent is perfect for EC/ESMS, methylene chloride offers an acceptable compromise for the EC/ESMS detection of radical cations of PAHs. (2) Supporting Electrolytes. In preparation for an electrolysis, a high concentration of supporting electrolyte (∼10-100 times the concentration of analyte) is typically added to the solution to obtain sufficient conductivity and to maintain electroneutrality. High concentrations of electrolytes, however, are known to decrease the sensitivity of ESMS detection of analytes formed via proton attachment or proton removal.31,32 If these electrolytes possess a high degree of surface activity, analyte suppression will be further exacerabated presumably due to increased competition for sites on the droplet surface.31,34 In methylene chloride solvent, TBAP or hexafluorophosphate is most often used as the supporting electrolyte. Addition of 5 × 10-4 M TBAP to an analyte solution consisting of 10-4 M 9,10-dimethylanthracene (DMA) in methylene chloride, however, resulted in an almost complete suppression of analyte M•+ (m/z 206) signal (Figure 2a), even though a substantial faradaic current (300 nA) passed through the cell. Instead, the tetrabutylammonium monomer (m/z 242) and dimeric cluster with one counterion (m/z 584, at low resolution) were observed (Figure 2a). It has been reported that lithium trifluoromethanesulfonate (LiCF3SO3) exhibits only limited suppression of analyte signals,35 but this salt is sparingly soluble in pure CH2Cl2. The solubility problem was overcome by first dissolving LiCF3SO3 into purified (33) Marcoux, L. S.; Fritsch, J. M.; Adams, R. N. J. Am. Chem. Soc. 1967, 89, 5766-5769. (34) Fenn, J. B. J. Am. Soc. Mass Spectrom. 1993, 4, 524-535. (35) Zhou, F.; Van Berkel, G. J. Proceedings of 42nd ASMS Conference on Mass Spectrometry and Allied Topics, Chicago, June 1994; p 1002.

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Figure 3. (a) Solution conductivity and (b) abundance of DMA radical cation, each as a function of electrolyte LiCF3SO3 concentration for solutions containing 10-4 M DMA.

Figure 2. Electrochemical/electrospray mass spectra of 10-4 M 9,10-dimethylanthracene in 5% CH3CN/95% CH2Cl2, acquired in the presence of the following supporting electrolytes: (a) 5 × 10-4 M tetrabutylammonium perchlorate; (b) 10-3 M lithium trifluoromethanesulfonate.

acetonitrile at a concentration up to 0.1 M; the resulting solution was then mixed with CH2Cl2. It was found that a preferred mixed solvent consisting of 5% CH3CN/95% CH2Cl2 (v/v) can dissolve up to 10-3 M LiCF3SO3, while 10% CH3CN/90% CH2Cl2 can dissolve up to 10-2 M. The proportion of acetonitrile was kept at a minimum to maintain the stability of radical cations. The DMA radical cation (M•+, m/z 206) was detected in high abundance by employing 5% CH3CN/95% CH2Cl2 containing 10-3 M LiCF3SO3, as shown in Figure 2b. Also detected was the [M + H2O]•+ adduct, presumably formed from reaction of M•+ with trace amounts of water present as an impurity. The conductivity of the solution and the abundance of the DMA radical cation, each as a function of electrolyte (LiCF3SO3) concentration, are shown in panels a and b of Figure 3, respectively. For an analyte (DMA) concentration of 10-4 M, the optimal concentration of supporting electrolyte was found to lie between 1 × 10-4 and 2 × 10-4 M. Below this level, the conductivity of the solution was apparently too low, resulting in a poor electrochemical conversion efficiency. At very low concentrations of supporting electrolyte (e.g., 5 × 10-4 M), severe suppression of analyte signal was systematically observed. 4248 Analytical Chemistry, Vol. 68, No. 23, December 1, 1996

(3) Temperature. In ESMS experiments not using pneumatic assistance, probe temperatures in the range of 50-70 °C are often preferred to facilitate the evaporation of solvents and to increase sensitivity. However, in our EC/ESMS experiments employing 5% CH3CN/95% CH2Cl2, the optimal probe temperature for detection of products of PAH oxidation fell in the range of 40-43 °C (source heating block temperature 186 ( 2 °C), which is consistent with previous results for signal optimization of stable anions in methylene chloride in the negative ion mode.22 Higher temperatures disrupt the electrospray process when the boiling point of the solution is exceeded (boiling point of the employed solution was slightly higher than that of pure CH2Cl2 (39.7 °C)). (4) Position of the Working Electrode. The position of the working electrode influenced both the response time and the detection sensitivity. The fastest response time and the highest sensitivity were obtained when the working electrode protruded approximately 0-0.3 mm out of the tubular auxiliary electrode; i.e., the tip of the working electrode penetrated the base of the “Taylor cone”. Further extension of the working electrode rendered the electrospray unstable or even caused discharge, especially if the boundaries of the stable Taylor cone were “pierced”. Withdrawing the working electrode inside the auxiliary electrode increased the response time, and it could also lower the detection sensitivity. Placement of the working electrode tip about 1-2 mm back from the base of the Taylor cone, however, did produce a more stable electrospray current at the cost of a longer response time. Further withdrawl of the working electrode produced no advantages, while response time increased. Off-Line Linear Sweep Experiment. To validate the performance of the electrochemical system, an off-line linear sweep experiment was conducted employing 10-4 M 9,10-diphenylanthracene (DPA) in 5% CH3CN/95% CH2Cl2 containing 10-4 M LiCF3SO3. The accessible anodic potential range was ∼+1.7 V vs Ag/Ag+ (0.01 M), i.e., ∼+2.0 V vs SCE. The E1/2,ox of DPA was measured to be 0.89 V vs Ag/Ag+ (0.01 M), close to the

Figure 5. Effect of flow rate of a 10-4 M solution of DPA on (a) response time and (b) signal response, both pertaining to detection of M•+ (m/z 330). Figure 4. Potential-step (+1.5 V vs Ag/Ag+) experiment for determination of response time employing 10-4 M DPA in 5% CH3CN/95% CH2Cl2 containing 10-4 M LiCF3SO3: (a) Applied potential vs time; (b) m/z 330 ion current vs time; (c) mass spectrum showing DPA M•+ (m/z 330).

reported value of 0.92 V also vs Ag+/Ag,36 which corroborates the estimation (Appendix) that iR drop contributes little error to the applied potential at the working electrode. Diffusion-limited current was attained at +1.3 to +1.5 V. Response Time of the EC/ESMS System. The response time, i.e., the time delay between the application of potential to the working electrode, and the appearance of the selected signal, should be as short as possible in order to detect species that are short-lived due to reactions in the solution. The response time of the prototype fast EC/ESMS system was determined by a potential-step experiment with DPA (10-4 M) under the preferred conditions described above; results are shown in Figure 4. The moment that +1.5 V (vs Ag/Ag+) was applied to the working electrode (Figure 4a) served to define the “start” time. In response to the applied potential, the signal for the molecular radical cation of DPA (m/z 330, Figure 4b) increased sharply. The “end” time was defined as the time required for the data system (scanning every 0.1 s) to register a signal above an arbitrarily defined threshold value. By this method, a response time of 1.7 s was measured at a flow rate of 3.8 µL/min. The mass spectrum obtained after the potential step is shown in Figure 4c. The response time is a function (roughly the sum) of the time delays of several different processes including diffusion of the analyte to the electrode surface, electron transfer, diffusion of the (36) Weissberger, A.; Proskaner, E. S.; Riddick, J. A.; Tops, E. E., Jr. Organic Solvents, 2nd ed.; Wiley (Interscience): New York, 1955.

electrochemically generated species away from the electrode surface, transportation of these species to the tip of the Taylor cone, charged droplet formation, droplet evaporation, desorption into the gas-phase, transportation of ions through the mass spectrometer to the detector, and acquisition of data. The transportation of electrochemically generated species to the tip of the Taylor cone, is perhaps the slowest step. If this assumption is correct, then the flow rate of the sample solution will influence the response time. The effects of flow rate on the response time and on the sensitivity of EC/ESMS have been investigated using the same 10-4 M DPA solution employed to obtain Figure 4b and c, in the absence of pneumatic assistance. The results are shown in Figure 5. A priori, the response time should decrease with increasing flow rate because the electrochemically generated species will move out of the cell more rapidly. Experimental results confirmed this expectation when the flow rate was increased from 1.1 to 3.8 µL/min. When the flow rate was increased above 3.8 µL/min, instrument sensitivity and stability were reduced, as was the electrochemical conversion efficiency. The measured response time (which was linked to an arbitrary threshold signal level) thus appeared to increase slightly (Figure 5a). It should be noted, however, that over the range of flow rates tested, the overall influence of flow rate on response time was not very dramatic. Even at the lowest flow rate tested (1.1 µL/min), the response time was