1402
Anal. Chem. 1981, 53, 1492-1497
responsivenessis not a function of suppressor exhaustion but remains reasonably constant with use. CONCLUSION The replacement of the conventional ion-exchange resin bed suppressor column with the hollow fiber suppressor allows continuousoperation of an ion chromatographwithout varying interference from base line dips, ion-exclusion effects, or chemical reactions. Future work needs to be directed at reducing the band spreading of hollow fiber suppressors, at the development of aminated membrane suppressors for cation determination using ion chromatography and at easier means of fabricating hollow fiber suppressors.
LITERATURE CITED (1) Maugh 11, T. H. Science I980 208, 164. (2) Small, H.;Stevens, T. S.; Bauman, W. C. Anal. Chem. 1975, 47 1801. (3) Wheaton, R. M.; Bauman, W. C. Ind. Eng. Chem. 1953, 45, 229. (4) Koch, W. F. Anal. Chem. 1979, 57, 1571. (5) Wetzel, R. A.; Anderson, C. L.; Schleicher, H.; Crook, G. D. Anal. Chem., 1979, 57, 1571. (6) Heftman, E. “Chromatography”, 2nd ed.; Reinhold: New York, 1976; p 296.
RECEIVED for review March 2, 1981. Accepted May 8,1981. A United States Patent application has been filed on behalf of the authors covering the subject of this contribution.
Laser Ionization Mass Spectrometry of Nonvolatile Samples E. D. Hardln’ and M. L. Vestal” Department of Chemistty, Universlty of Houston, Houston, Texas 77004
A new laser Ionization mass spectrometer has been developed whlch employs a movlng stainless steel belt onto whlch the sample Is electrosprayed for continuous sample Introduction. Ionization Is produced by focusing the output of a tunable dye laser onto the movlng belt. The mass spectrometer system Is a conventlonal quadrupole system, wlth the exceptlon of a gated boxcar integrator to process the pulsed Ion beam. Mass spectra have been recorded for a number of nonvolatlle blomolecules Including saccharides, amlno aclds, peptldes, nucleosldes, and nucleotldes. Generally these spectra show intense catlonlzed molecular Ions, often Includlng multiple alkall addltlon, and little fragmentatlon. The major llmltatlon on the technlque at present Is the rather poor reproduclbllity of the spectra. Ion tlme-of-flight dlstrlbutlons have been measured whlch show that Ions produced by laser desorptlonAonlzation have broad kinetlc energy distrlbutlons wlth most probable kinetic energles of about 6 eV and wlth hlgh-energy tails extendlng beyond 25 eV. The time-of-flight dlstrlbutlons also show that most of the hlgh mass ions observed result from metastable decomposltlon of larger clusters formed lnltially at the surface.
In a number of areas of organic and biochemical research there is a growing need for high mass, high sensitivity mass spectrometry applicable to thermally labile molecules of low volatility (I). In particular, it is often very important to obtain easily identifiable ions characteristic of the intact molecule so that the molecular weight can be determined. In response to this need, several new techniques have been developed (2). These include field desorption (3), chemical ionization (4), plasma desorption (5), laser desorption (6),organic SIMS (7, 8), and very rapid sample heating with “in-beam’’ chemical (9-11) or electron ionization (12-14). In our own laboratory we have recently discovered a new soft ionization technique which employs thermal production of a charged macroscopic particle beam containing the sample. This technique employs very rapid heating of a liquid solution to form a particle beam
R. A. W e l c h F o u n d a t i o n Predoctoral Fellow. 0003-2700/81/0353-1492$01.25/0
and impact of this particle beam on a mildly heated surface (15).
Several of these techniques, such as field desorption and chemical ionization, both conventional and “in-beam”, have been relatively well characterized, and the apparatus is commercially available; however, with the exception of gas-phase chemical ionization, it appears that none are satisfactorily understood. Recent work employing these new ionization techniques has demonstrated that gas-phase ions characteristic of the intact molecule can be produced for previously intractable molecules. Despite the enormous advances of recent years many problems are not yet solved, and none of these new techniques has, as yet, achieved wide-spread acceptance. The present research was undertaken to explore the feasibility and practicality of laser desorption and ionization for mass spectrometric analyses of nonvolatile organic molecules adsorbed on, or contained in, solid surfaces. In our view, a practical technique should provide both molecular weight and structural information on a wide range of nonvolatile and/or thermally labile compounds; it should be compatible with conventional rapid scanning mass analyzers, both magnetic and quadrupole;and it should be suitable for combination with the techniques commonly used for separating and purifying mixtures of involatile compounds such as liquid (LC) amd thin-layer chromatography (TLC). Most of the earlier laser desorption studies have used some form of simultaneous ion detection, employing either timeof-flight analysis (16), electrooptical ion detection, or photoplate (17). Recently, the use of fast electrical scanning with laser desorption from a direct exposure probe in a chemical ionization source has been reported by Cotter (18). The use of repetitive pulsed laser desorption with magnetic scanning over a limited region of the spectrum has been reported by Heresch, Schmid, and Huber (19). The use of laser desorption using a moving belt inlet system with a tightly enclosed chemical ionization (CI) source and a quadrupole mass spectrometer has been reported by Hunt, Bone, and Shabanowitz (20). In the present work we have developed a new laser desorption mass spectrometer which uses a moving belt system to continuously supply fresh sample to a fully open laser vaporization/ionization region. The mass spectrometer is a 0 1961 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 53, NO. 9, AUGUST 1981 LASER STAINLESS STEEL
Table I. Relative Intensity of Molecular Ions DRIFT SPACE
sample
/
7/
ION ACCELERATION
/
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OUT
ELECTRON MULTIPLIER
QUADRUPOLE Figure 1. Schematic dlagrerm of the laser ionization mass spectrom-
eter. conventional system, with the exception of a boxcar integrator to process the pulsed ion signal generated by pulsed laser ionization. With the addition of differential pumping on the moving belt inlet system, the laser ionization mass spectrometer could provide a viable alternative approach to a combined LC-MS system.
EXPERIMENTAL SECTION The laser desorption mass spectrometer developed in this work is shown schematically in Figure 1. The apparatqs consists of a continuous stainless steel belt, ion accelerating and focusing lenses,a quadrupole mass illter, and an off-axis electron multiplier. This apparatus is housed in a vacuum bell jar pumped by a 4000 L/s diffusion pump backed by a 300 L/min mechanical pump. The beam from a tunable dye laser passes through a Pyrex lens of 10 cm focal length and then through a Pyrex window and impinges on the moving belt as indicated in Figure 1. The angle between the laser beam and the quadrupole axis is approximately 45" and the extended quadrupole axis is normal to the surface of the belt at the point of incidence of the laser beam. The dye laser is pumped by a 400-kWnitrogen laser, and for the experiments reported here, the dye laser was operated at 483 nm using a 0.01 M solution of 7,7-diethyl-4-(trifluoromethyl)coumarinin p-dioxane in the dye cell. The laser pulses are typically 5-7 ns wide full width at half-maximum and typical pulse energies are 400 pJ when the laser is operated at a repetition rate of 25 Hz. The ion optical system consists of a nearly uniform field accelerating region (0.34cml) followed by a field free region (0.68 cm) and a thin einzel lens (0.68cm). The einzel lens is normally operated in the accelerating mode with the central element negative (for positive ions) in the range 100-200 V. The back plate of the accelerating region and the moving belt are connected to the accelerating potential which is typically in the range 5-30 V off ground. The additional plates in the ion path are grounded as is the central axis of the quadrupole. The stainless steel belt (150cm in circumference, 0.1 cm wide and 0.005 cm thick) is driven by a small dc motor and gear train external to the vacuum system using a shaft coupled through a rotatable ''0"-ring sealed vacuum feedthrough. Belt speeds used in the present work are in the range 10-30 cm/min. This range of speeds corresponds to 50-150 pulses/cm for a laser frequency of 25 Hz which generally removes most of the sample after one rotation of the belt. Spectra were taken with scan times allowing for approximately 5 pulses/amu. For measurement the output of the electron multiplier is coupled to a fast current amplifier which is sampled by a boxcar integrator gated to measure the ion current during the appropriate time interval following each laser pulse. Spectra may be recorded by scanning the quadrupole in the conventionalmanner arid connectingthe output of the boxcar to a recorder. For most of' our recent work the instrument was interfaced to a Finnigan/hicos Model 2300 data system for data acquisition and reduction. For the time-of-flightmeasurements the current amplifier was replaced by a fast pulse amplifier/discriminator, and the time between laser pulse and ion arrival was determined by using a time-to-pulse height converter, and the results were stored in a
MH+
ak:li;+
a
(M H)(alkali); a
arginine 75 90 100 histidine 100 45 35 phenyl0 0 100 alanine serine 0 15 100 leucine 0 25 100 aspartic acid 0 35 100 glutamine 0 10 100 glutamic acid 0 100 75 0 20 100 glYtryptophan arg70 100 25 glutamine gly-gly-gly0 100 100 glycine guanosine 10 100 100 cytidine 0 100 0 adenosine 80 100 0 a Relative ion intensities were calculated for (M + alkali)+as the sum of MNa+ and MK+. For ( M - H)(alkali)l the sum of (M - H)Na,+,(M - H)NaK+,and (M - H)Kl. No salts were added to any of these samples. multichannel pulse height analyzer. For these measurementsthe quadrupole was set manually to the mass of interest and the . time-of-flight spectrum was accumulated until the desired statistical precision was achieved. For these measurements the signal levels were intentionally reduced by increasing the resolution of the quadrupole so that not more than one ion was detected per laser pulse. At present the moving belt and the quadrupole are housed in the same vacuum system. For applicationof sample the apparatus shown in Figure 1 is vented to atmcsphere with the vacuum pumps valved off. Several methods of applying sample to the belt have been tried, but the most reproducible results have been obtained by using the electrospray technique as described by McNeal, Macfarlane, and Thurston (21). Typically, about 15 min is required to fully coat the belt with sample and an additional 15 min is required to evacuate the system to normal operating pressure of lo4 torr or below.
RESULTS AND DISCUSSION Laser ionization mass spectra have been recorded for a number of nonvolatile biomolecules including amino acids, peptides, nucleosides, and saccharides. Some examples of these results are presented in Table I. The most intense pe& in the spectra are generally those produced by alkali addition. In most cases this is true even when no alkali salts are intentionally added. Notable exceptions are the basic amino acids, e.g., arginine and histidine, for which intense protonated molecular ions, MH+, are observed even when excess amounts of alkali salts are added to the samples. With other amino acids, such as phenylalanine, serine, leucine, glutamine, and aspartic acid, the most intense ions correspond to double alkali addition to the deprotonated molecular anion, (M - H)K2+, (M - H)NaK+, etc., and the cationized molecular ions, MNa+ and MK+, are weak or absent completely. With the nucleosides the cationized molecular ion is generally the most intense, but protonation and multiple alkali addition are also observed. Of the molecules studied, only the nucleosides yield significant fragment intensities with the protonated and cationized base fragments being observed in all cases studied. The background spectra consist mainly of K+ and Na+ and small clusters of salt adducts such as KCl.K+, NaCl.K+, etc.; at higher power densities ions characteristic of the stainless steel belt, such as Cr+, Fe', and Ni+, are observed. No doubly charged metal atoms have been observed. At higher masses several small peaks are observed which are apparently due
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ANALYTICAL CHEMISTRY, VOL. 53, NO. 9, AUGUST 1981
1
t
100
I
L
INTENSITY (bRBMCiITSI
I\ 5
I
n
15
IO TIME (mid
Figure 2. Intensity of cytidine MK+ as a function of time. The parameter indicates the sample coverage on the belt over the indicated measuring times. Data were obtalned by using 20-s scans over a 20 amu range including MNa+ ( m l e = 266) and MK ( m / e = 282). ~
~~
sample concn, ng/cm2 re1 responsea
re1 std devb
10
18
13
30 50 100 150 980
24 24 22 25 21
12 14 4
10 5
a Average response in arbitrary units divided by sample concentration. Standard deviation of 1 4 measurements divided by sample concentration.
to alkali addition to residual pump oil or other background contaminants of the vacuum system. We have generally experienced little difficulty in obtaining spectra of nonvolatile compounds with samples in the 0.1-1 pg/cm2 range, and in most cases the detection limits are at about the 1 ng/cm2 level or below. In essentially all cases studied, the most intense ions observed contain the intact molecule with a proton or one or more alkali atoms or ions added. The major difficulty has been the lack of reproducibility, both in the relative intensities of the major ions and in their absolute intensities. A major source of these variations appears to be due to the local variations in surface coverage by sample and by impurities and background contaminants. In general, deposition of samples by direct application of a liquid solution followed by evaporation of the solvent has given very poor results. The use of the electrospray technique described by McNeal et al. (21)has given considerably better results, but significant variations persist in most cases. An example of the quantitative response of the laser desorption system for one of the more favorable cases studied, the nucleoside cytidine, is shown in Figure 2. In this experiment one-third of the belt was coated with 10 ng/cm2 of cytidine, one-third with 30 ng/cm2, and one-third with 100 ng/cm2. In each case an equimolar solution of cytidine and KI was used in the electrospray apparatus, and we assume that essentially all of the solution sprayed is deposited on the belt. The mass spectrum was scanned repetitively over a 20 amu range encompassing both the M Na+ ( m / e = 266) and the M + K+ ( m / e = 282 amu) peaks; under the conditions of this experiment the M K+ is the most intense peak in the laser desorption mass spectrum of cytidine. The reconstructed mass chromatogram for this ion is shown in Figure 2 with the corresponding surface concentrations of cytidine indicated. As can be seen from the figure, the M K+ is approximately proportional to surface concentration of sample in this range of surface coverage, but the spectrum to spectrum fluctuations are quite large. The average response and the standard deviation of the results from this experiment and another covering a wider range of sample concentrations are summarized in Table 11. It appears that, with careful use of the electrospray technique, linear response can be obtained over at
+
+
+
0
-~
Table 11. Summary of Quantitative Measurements on Cytidine
40
20
60
80
IO0
t (psec)
Figure 3. Measured time-of-flight distributions for K+ and K+.KI (solid lines). The dashed line under the K+ indicates the calculated timeof-flight distribution Corresponding to the inltlal kinetic energy dlstributions given by eq 4. For flight times shorter than 21 ps the experimental and calculated distributlonsare essentially identical. The dashed lines under the K+*KI distribution correspond to calculated distrlbutions for ions accelerated as K+.(KI),, with the value of n indicated on the figure which dissociate to K+.KI in the field free reglon. The relative contribution of each which give the best fit to the experimental data is (in percent): n = 1, 8; n = 2, 29; n = 3, 51; n = 4, 8; n = 5, 4 (not shown on the figure). least the range from 10 ng/cm2 to 1 pg/cm2. TIME-OF-FLIGHTANALYSIS In these initial studies we have experienced considerable difficulty in obtaining reproducible results from the laser desorption mass spectrometer. In an effort to achieve a better understanding of the sources of these difficulties, we have initiated some more fundamental studies of the laser desorption process. The most fruitful of these investigations has been measurements of the time-of-flight distributions of mass-selected product ion beams produced by pulsed laser desorption. The total time for an ion produced at the surface of the stainless steel belt to reach the detector is given approximately by
t=u1
2 4
+ uo
+ -d2u1
(1)
where
and Tois the initial kinetic energy, M the mass of the ion, e the charge, V1 the applied accelerating potential difference, dl the length of the accelerating field, and d2the total effective length of the drift region, including the einzel lens, the quadrupole, and the accelerating field in front of the multiplier. In the present appartus, dl = 0.34 cm and d2 = 15.7 cm; the error in approximating the TOF through the lens, the quadrupole, the multiplier accelerating field is estimated to be less than 1%, which is insignificant for the present work. If an ion dissociates in the field free region (0.68 cm long) between the accelerating field and the einzel lens, it will be observed with the quadrupole set to transmit the product ion, but its velocity distribution, and hence its time-of-flight distribution, will correspond to that of the precursor ion. Kinetic energy release in the fragmentation process may broaden the distribution somewhat, but this contribution to the time-of-flight distribution is relatively small compared to its effect on the laboratory kinetic energy distribution. The measured flight time distribution for K+ from laser desorption of K1 is shown in Figure 3. The resulb are in good agreement with the calculated time-of-flight distribution if
ANALYTICAL CHEMISTRY, VOL. 53, NO. 9, AUGUST 1981 m/e 100
200
300 400 500 600 700 800
--tv-r--l
1495
GUANOSINE t K I BHK'
+r
20
40
a0
60
t (psec) 30
40
50
60 70 t (ped
BO
YO
100
Figure 5. Measured time-of-fllght distribution for the cationized base fragment BHK' of guanosine ( m l e = 190). The dashed lines show the distributions calculated for ions initially formed and accelerated as the BHK' ion itself (190) and as MK+ (322)and MK+-KI (488).
Flgure 4. Measured time-of-flight distribution for MH+ from guanosine ( m l e = 284)and BH,+ ( m l e = 190). The dashed lines Indicate the maximum calculated fllghit time (corresponding to zero initial kinetic energy) for the base fragment, BHz', the protonated monomer, MH', and the protonated dimer, MH,', respectively.
PEAK m / e (amu) 200 300 400
600
800 I000
we assume that the initial ion kinetic energy distributor is given by
1400
322
I800
XI
f ( E ) = CEfle-E/%
(4) with n = 2 and Eo = 6 eV. The calculated time-of-flight distribution is shown by the dashed line in Figure 3. Except for a small discrepancy at longer times, the calculated distribution is in excellent agreement with the experimental results. The energy distributions produced by laser desorption appear qualitatively similar to those observed in SIMS (22); however, our results give rather lower energies than some of the earlier work at higher laser intensities (23). The time-of-flight distribution found for K21+, shown in Figure 3, is not compatible with the ions being formed primarily at the surface. Rather, it appears that most of the observed K21+ results from unimolecular decomposition of larger clusters K+.(KI),,,,n = 2-5, in the field free region. If we assume that each of these ions is formed with the same energy distribution as was determined above for K', then the dashed curves shown in Figure 3 correspond to the expected flight time distributions for these ions. Adjusting the relative contributions of the individual clusters to achieve the best fit to the experimental curve, we obtain the results summarized in Figure 3. With the inclusion of the cluster corresponding to n = 5 (not shown in Figure 3) the sum of the individual time-of-flight distributions agrees with the experimentalresult over the entire time range with a maximum deviation of about 5%, which is well within the experimental uncertainty. In the mass spectrum a very weak intensity is observed a t mass 371, corresponding to the cluster with n = 2, but the larger ions are not detected. These larger clusters have been detected, however, in SIM8 experiments on alkali halide crystals (22). It appears quite cllear that these larger clusters are being formed under the conditions of our laser desorption experimenta but that they are metastable and are dissociating before reaching the detector. It is quite likely that a significant fraction of the total ioniiaation may be lost by decomposition in the quadrupole. Similar experimenta have been conducted on the major ions in the laser desorption m a s spectrum of guanosine; the results are summarized in Figures 4-6. It appears from the results that the protonated product ions arise from a completely different set of precursor ions than do the cationized species. Time-of-flightdistributions for the MH+ ion ( m / e = 284) and
322
x2 XI
322 20
40
60
80
' IO0
t (pet)
Flgure 6. A series of consecutive measurements of the time-of-flight distrlbutions for the MK+ (322)and BHK' (190)ions in the laser desorption spectrum of guanosine with added KI. The scale at the top shows calculated position of the peak in the distribution as a function of mass assuming the energy distribution is that given in Figure 3. The inset scale shows calculated tlme-of-flight corresponding to mle = 322 with the indicated Initial kinetlc energies. The accelerating potential for these measurements was 20 V.
the BH2+ion ( m / e = 152)from guanosine are shown in Figure 4. It appears that both of these ions are formed by metastable decomposition of a larger precursor; qualitative deconvolution of the results indicates that the major precursor is probably the protonated dimer of guanosine corresponding to m l e = 569. At most, about 10% of the MH+ is formed a t or near the surface. About 70% of the BH2+is formed by metastable decomposition of the protonated dimer and about 30% by metastable decomposition of the protonated monomer. Direct formation of the BH2+fragment at the surface appears negligible. A time-of-flight distribution for the cationized base fragment of guanosine, BHK+ ( m / e = 190), is shown in Figure 5. The resolution of this distribution into the distributions correspondingto the precursor masses is shown in the dashed lines on the figure. In this case it appears that about 40%
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ANALYTICAL CHEMISTRY, VOL. 53,
NO.9, AUGUST 1981
of the BHK+ fragment is formed directly on the surface; about 50% is formed by metastable decomposition of the cationized parent ion MK+ ( m / e = 322), and the remainder is formed by metastable decomposition of the cluster of KI with MK+ at m / e = 488. In these initial studies it was found that the observed time-of-flight distributions were quite dependent on the laser power density and the relative concentration of the sample and alkali salt on the surface. The kinds of variations which are observed are illustrated in Figure 6. In these series of experimentsthe belt was coated with a relatively heavy coating of guanosine, about 1pg/cm2,along with an equimolar amount of KI. The time-of-flight distributions shown in Figure 6 were taken consecutively, starting from the top, following this single application of sample. The accelerating potential, VI, was 20 V for this series of experiments. Each of these distributions corresponds approximately to one revolution of the belt. For runs 1-5 and 8 and 9, the quadrupole was set to transmit m / e = 322 (MK’), and for runs 6 and 7 m / e = 190 (BHK’) was monitored. For those experiments indicating an attenuation of xl, the laser was operated at the usual nominal operating conditions corresponding to a peak power density of about lo7 W/cm2. In runs 3, 5, 6, and 8 the power density was reduced by a factor of 2 by interposing a X 2 neutral density filter in the path of the laser beam. In the first run at “normal” laser power and with “freshly applied” sample, essentially no direct formation of MK+ is observed; the time-of-flight distribution corresponds to clusters in the range 600-1800 amu. The peaks in the distribution indicate that the major precursors are probably MzK+(605), M3K+ (888), M4K+.2(KI) (1337), and M5K+.(KI)2(1786). Peaks corresponding to the first two of these precursors are quite clearly resolved, but the identity of the higher mass clusters is less certain. In the second experiment the thickness of the sample layer has presumably been reduced and the higher mass precursors are no longer observed. In this case the dominant peak corresponds to the MzK+ ion and there appears to be a small contribution from M3K+. The dramatic effect of decreasing the laser power density by a factor of 2 is shown in the third run. In this case, most of the MK+ is produced directly at or near the surface. The tail to longer times is probably due to M2K” decomposition either in the accelerating field or in the drift region. Returning to full laser power by removing the X 2 attenuator in run 4 causes the peak in the time-of-flight distribution to return to that expected for MzK+as in the first two runs; however, in the latter case the peak is more tailed to shorter times. This may indicate that as the sample is depleted from the surface, a new precursor, such as MK+.KI, begins to contribute to the observed product. In run 5 the laser power density is again reduced by a factor of 2 and now almost all of the MK+ appears to be formed a t the surface with only a very small fraction of the total intensity formed by metastable decomposition of large clusters. In runs 6 and 7 the quadrupole was set to transmit m / e = 190 corresponding to the BHK+ fragment from MK+. At low laser power, run 6, essentially all of the observed BHK+ appears to be produced by metastable decompositon of MK+ while at the higher power there is a significant contribution from a larger cluster, probably MK+-KI. Neither experiment appears to show any significant amount of direct formation of BHK+ at the surface. The fact that the peak in run 6 occurs at shorter times than in the other runs implies either a significant fraction of the decomposition occurs in the accelerating field or the most probable kinetic energy of the precursor is shifted to a higher value. Since it appears, in general that the most probable kinetic energy decreases as the laser power is decreased, the former explanation seems more likely.
In run number 8 the quadrupole was again set to transmit mass 322 and the laser beam attenuator was in place, repeating run 5. Again, it appears at low laser power density that the MK+ ions are primarily formed directly on the surface. Also, this run clearly shows that the most probable kinetic energy of the ions is significantly lower as the sample coverage is depleted. In the final run, the attenuator is removed and at the higher laser power most of the MK+ ions are formed by metastable decomposition of larger clusters. In this final run, it appears that the precursors correspond to about equal amounts of MK+.KI and MzK+. It should be noted in connection with these experiments that the absolute ion intensity is a very steep function of the laser power density. Under the conditions of the experiments summarized in Figure 6 reducing the laser power by a factor of 2 may reduce the absolute ion intensity by as much as an order of magnitude. Furthermore, the absolute ion intensity in the final run of Figure 6 is about an order of magnitude less than the first run. It should be noted that the results presented here were all obtained at laser power densities of about lo7 W/cm2 or less. With the laser and lens system used in this work it is quite straightforward to focus the beam spot on the surface so that the power density is 1-2 orders of magnitude larger. Some experiments were conducted early in this work at power densities up to lo8 W/cm2 but the results were quite disappointing. The pseudomolecular ions were smaller in intensity at these high-power levels and a rather large mass independent background signal was observed. Time-of-flight measurements with the quadrupole set to reject ions of all masses indicate that this background signal is mainly due to neutrals with very high kinetic energy, perhaps (depending on their mass which is presently unknown) with energies of several hundred electronvolts. In the present apparatus the ions spend approximately the first 4% of their flight time in the accelerating field and the next 4% in the field free region from which metastable decompositions are observed. Most of the remainder (about 90%) of the total flight time is spent in the quadrupole mass filter. Ions dissociating in the quadrupole are not detected at any mass. As discussed above, it appears that most of the high mass sample ions observed in the laser ionization instrument are the result of metasable decomposition in the field-free region. This statement appears to be particularly true for experiments at high laser power density and high surface concentration of sample. Furthermore, since the drift region is quite short (0.68 cm) compared to the quadrupole (14 cm), it seems quite likely that a large fraction of the total ions produced may be lost due to unimolecular decomposition during transit through the quadrupole. If this is true, substantial improvements in performance may be achieved by providing an extended drift space between the desorbing surface and the quadrupole mass filter. Incorporation of an rf excited strong focusing quadrupole lens might provide a practical solution. The major difficulty presently limiting the utility of the technique is the poor reproducibility. The ion time-of-flight measurements have provided some new insight into the ion formation mechanism. Work presently in progress is aimed at achieving a better understanding of the sources of the observed variability of the spectra; if this work is successful, the laser ionization technique may provide a practical method for routine mass spectral analysis of nonvolatile samples.
ACKNOWLEDGMENT The authors are grateful to J. W. Rabalais for lending the time-to-pulseheight converter and multichannelanalyzer used in the time-of-flight measurements. We also acknowledge C. R. Hsieh for assisting with the calculations, L. M. Marks for
Anal. Chem. 1981, 53, 1497-1504
preparing the figures, and C. R. Blakley for many helpful discussions during the course of this work.
LITERATURE CITED ( I ) Burlingame, A. L.; Baillie. T. A.; Derrick, P. J.; Chizov, 0. S. Anal. (2) (3) (4) (5) (6) (7)
(8) (9) (10) (11) (12)
(13) (14)
Chem. 1980, 52, 214,R. Daves, G. D., Jr. Acc. Chem. Res. 1979, 12, 359. Beckey, H. D. Int. J. Mass Spectrom. Ion Phys. 1969, 2 , 500. Fiekl, F. H. Acc. Chem. Res. 1968, 1 , 42. Macfarlane. R. D.; Torgerson, D. F. Science 1976, 191, 920. Mumma, R. 0.; Vastola, F. J. Org. Mass Spectrom. 1972, 6 , 1373. Benninghoven, A.; Sichterman, W. K. Anal. Chem. 1978, 50, 1160. Day, R. J.; Unger, S. I!.; Cooks, R. F. Anal. Chem. 1980, 52, 557A. Baldwln, M. A.; McLafferty, F. W. Org. Mass Spectrom. 1973, 7 , 1141, 1353. Hunt, D. F.; Shabanowitz, J.; Botz, F. K.; Brent, D. A. Anal. Chem. 1977. 49. 1160. Hansen, G.; Munson, 13. Anal. Chem. 1980, 52, 245. Dell, A.; Willlams, D. ti.; Morris, H. R.; Smith, G. A.; Feeney, J.; Roberts, G. C. K. J. Am. Chem. SOC. 1975, 97, 2497. Ohashi, M.; Tsujimoto, K.; Yasuda, A. Chem. Lett. 1976, 439. Anderson, W. R., Jr.; hick, W.; Daves, G. D., Jr. J. Am. Chem. Soc. 1978, 100 1974.
1497
Blakely, C. R.; Carmody, J. J.; Vestal, M. L. J. Am. Chem. SOC. 1980. 102. 5931. Kauf&nn.’R.; Hillenkamp, F.; Nitsche, R.; Schurmann, M.; Wechsung, R. Microsc. Acta 1978, 2 , 297. Posthumus, M. A.; Kistemaker, P. G.; Meuzelaar, H. L. C.; Ten Noever de Brauw, M. C. Anal. Chem. 1978, 50, 985. Cotter, R. J. Anal. Chem. 1980, 52, 1767. Heresch, F.; Schmid, E. R.; Huber, J. F. K. Anal. Chem. 1980, 52, 1803. Hunt, D. F.; Bone, W. M.; Shabanowitz, J. Paper RPMP18, 28th Annual Conference on Mass Spectrometry and Allied Topics, May 25-30; ASMS: New York, 1980. McNeal, C. J.; Macfarlane, R. D.; Thurston, E. L. Anal. Chem. 1979, 51, 2036. Honda, F.; Fukuda, Y.; Rabalais, J. W. J. Chem. Phys. 1979, 70, 4834.
C o k m i u s , R. J.; Capellen, J. M. Int. J . Mass Spectrom. Ion Phys. 1980, 34, 197.
RECEIVED for review February 13, 1981. Accepted May 12, lg8l.This work was supported by the NIGMSunder Grant GM24031 and by the Robert A. Welch Foundation.
Peak Resolution by Semiderivative Voltammetry Jeffrey J. Toman and Steven D. Brown”’ Department of Chemistry, University of California, Berkeley, California 94720, and Materials and Molecular Research Division, Lawrence Berkeley Laboratory, 1 Cyclotron Road, Berkeley, California 94720
A technique based on semidifferentialvoltammetry has been developed for rapld electrochemical peak resolution. The approach was tested by resolutlon of synthetic fused peak systems. Subsequently, the approach was applied to semldifferentiated h e a r scan voltammograms of Cd2+, Pb2+,and Ina+ and to semidifferentiated llnear scan anodlc stripplng voltammograms of Cd”, Ina+, and TI’. Resolutlons were directly characterlred by peak height, potential, and halfwidth, as well as by the, coefficient of determination between experlmental and fit peaks. Studies of individual peak systems and of multlple peak systems showed that the method achieved excellent resolution accuracy, failing only as charge transfers became irrewerslble. Synthetic data were totally resolved with peak separatlons as small as 25 mV, while real systems were deconvoluted with separatlons as small as 40 mV. Peak parameters obtained from these resolutlons allow observations of electrode processes, even in systems contalnlng overlapped peaks.
A principal difficulty in the use of dynamic electrochemistry as a quantitative analytical technique is the problem of resolution of overlapping waves. This is of particular importance in linear sweep voltammetry (lsv) and anodic stripping voltammetry (asv) at a stationary mercury electrode because of the very broad, asymmetric nature of the peaks. Another difficulty is that the current-potential relationship is not describable by an anallytic function. These problems are particularly unfortunate since lsv instrumentation is easy to use and is relatively inexpensive. There have been two general approaches to electrochemical peak resolution, a “hardware” approach and a “software” approach. The hardware approach is typified by the methods Present address: D e tartment of Chemistry, Washington State University, Pullman, W x 99164.
of Martin and Shain (I) and Bond and Grabaric (2). In both methods a sample is run along with a blank solution containing a suitable background electrolyte and one of the overlapped components. The concentration of this component is varied in the blank until, upon subtraction of the blank voltammogram from the sample voltammogram (whether by analog or digital means), no trace of the overlapped component is left. Besides being very time intensive, the making of a suitable blank may not even be feasible for cases in which the supporting electrolyte is complex. The software approach, that of resolution after data taking, is perhaps more promising. Amino (3) divides software resolution techniques into two types: frequency domain and time domain (curve fitting). Peak resolution in the frequency domain can be accomplished by the method of Kirmse and Westerburg (4). In this method the Fourier transform of an overlapped spectrum is divided by the transform of a single component, resulting in a sharpened spectrum. In electrochemistry Binkley and Dessey (5) have recently done work on the least-squares fitting of FT transformed square-wave voltammograms. In this procedure the transform of the overlapped peak is fit with the transforms of single peak data in a least-squares manner by varying what would be the peak height in the time domain. This procedure is advantageous in two ways. The fit is reduced in complexity by the reduction of points in going to the frequency domain, and fits may be done on both real and imaginary parts of the transform in order to obtain an averaged, improved fit. The procedure does rely on utilization of empirical single-peak parameters, however. Curve fitting of electrochemical data has been hampered by the lack of analytic current-potential functions. Some work has been done in this area, principally by Perone and Coworkers (6-8). Boudreau and Perone (8) describe the application of an empirical peak shape function (the sum of skewed Gaussian and Cauchy functions) in the use of curve fitting for the resolution of square-wave voltammetric peaks.
0003-2700/81/0353-1497$01.25/00 1961 American Chemlcal Society
