Anal. Chem. 1987, 5 9 , 1054-1056 Hansch, C.; Leo, A. Substituent Constants for Correlation Analysis in Chemistry and Biology; Wlley-Interscience: New York. 1979; pp 18-43. Miller, K. J.; Savchik, J. A. J. Am. Chem. SOC. 1979, 101, 7206. Draper, N.; Smith, H. Applied Regression Analysis, 2nd ed.; Wiley-lnterscience: New York. 1981: DD 307-312. Furnival, G. M.; Wilson; R. W.’,’jr. Technornetrics 1974, 16, 499. Neter, J.; Wasserman, W.; Kutner, M. H. Applied Linear Statistical Models. 2nd ed.. Richard D. Irwin: Homewood. IL. 1985: D 435. Chambers, J. M.; Cleveland, W. S.; Kleiner, B.; Tukey, P. A.’Graphicai Methods for Data Analysis; Wadsworth International: Belmont, CA, 1983; p 288. Bermejo, J.; Guillen, M. D. HRC C C . J . High Resolut. Chrornatogr.
Chromatogr. Commun. 1984, 7 , 191. (38) Bermejo, J.; Canga, J. S.; Gayol, 0. M.; Gulllen. M. D.J. Chromatogr. Sci. 1984, 22,252. (39) Bermejo, J.; Guillen, M. D.J. Chromatogr. 1985, 3 f 8 , 187.
RECEIVED for review September 22,1986. Accepted December 1, 1986. This work was supported by the National Science Foundation under Grant CHE-8202620. The PRIME 750 computer was purchased with partial financial support of the National Science Foundation.
CORRESPONDENCE Electron Impact Excitation of Ions in Fourier Transform Mass Spectrometry Sir: In 1979, we reported the use of ion cyclotron resonance (ICR) spectrometry to study electron impact induced fragmentation of ions trapped in the electron beam ( I ) . The technique, referred to as “electron impact excitation of ions from organics” (EIEIO), was used to obtain relative dissociation cross sections for product ions as the energy of the electron beam was varied. The “breakdown curves” obtained by this method were comparable to those obtained by angle-resolved mass spectrometry and low-energy collision-induced dissociation ( 2 , 3 ) .Electron impact excitation was also employed to produce doubly and triply charged argon ions from singly charged ions trapped in the electron beam ( 4 ) . At that time, we proposed that electron impact excitation of ions might become a useful alternative to collision-induced dissociation and photodissociation as a method for ion structure elucidation and mixture analysis. However, several problems were encountered with the method as originally employed, which presented obstacles to its general application. In particular, at the high currents employed to observe fragmentation, it is difficult to use double resonance techniques (5)for ejecting ions trapped in the electron beam. This is apparently due to the fact that the electron beam has a “shielding” effect against the applied rf. Thus, parent ion selection was only possible for compounds giving molecular ions exclusively a t electron energies slightly above the ionization potential of the compound. This limited the range of compounds available for study and required operation at a low signal level. One obvious solution to this problem would be to form ions by electron impact (or other ionization methods), turn off the electron beam to allow ion ejection, and then turn the electron beam on again at a lower electron energy to allow electron impact excitation of ions to occur, permitting the technique to be applied to a wider variety of problems. Unfortunately, this approach failed on the ICR spectrometer originally employed for these studies, because ions drifted out of the path of the electron beam after it was turned off and were not efficiently drawn back when the beam was turned on again. In this correspondence, we report the use of a Fourier transform mass spectrometer with a superconducting magnet to perform the experiment described above. Ion ejection is used to select a parent ion by gating the electron beam off. The beam is then gated on again, and the parent ion undergoes fragmentation following excitation by electron impact. The higher magnetic field produced by the superconducting 0003-2700/87/0359-1054$01.50/0
magnet confines the ions in the path of the electron beam, making this experiment possible. Signal averaging may be employed to improve signal-to-noise ratio, and it is possible to obtain very high resolution on the daughter ions produced.
EXPERIMENTAL SECTION These experiments were performed on a Nicolet FTMS-2000 dual-cell Fourier transform mass spectrometer (6-8) operated at a magnetic field strength of 3 T. Samples were introduced through the batch inlet. Ionization was performed by electron impact in torr. the source cell at a pressure of 5 x The sequence of events for this experiment is shown in Figure 1. Following the quench events, ions are formed by electron impact (EI) in the source region. During the ionization event, the conductance limit is kept at the trapping potential (typically 1 V) to confine ions in the source region. A variable delay period follows, during which ion-molecule reactions can occur. This delay was set to zero for E1 experiments, and is typically 50-100 ms for self-CI (chemical ionization) (9) experiments. Two sweptfrequency ion ejection events may be used to remove ions above and below the mass of the parent ion. Ion ejection was accomplished by exciting in both the source and analyzer cells. During these ejection sweeps, the electron beam is gated off by a grid in front of the electron filament. Following the ejection sweeps, the conductance limit is grounded for 100 p s to allow ions to pass into the analyzer region. After the parent ion is selected, the electron beam is turned back on for a period of 100 ms at an electron energy below the ionization potential for the neutral compounds. The electron current through the cell was 24 FA. Ion-electron collisions during this time induce fragmentation of the parent ions. The daughter ions produced are then excited by a rapid swept-frequency (“chirp”) excitation, and the ion image currents are detected, amplified, digitized, and Fourier transformed to produce the mass spectrum. RESULTS AND DISCUSSION Figure 2A shows the 15-eV electron impact spectrum of a mixture of acetophenone and isophorone, added in roughly equal amounts through the batch inlet. Molecular ions and characteristic fragment ions from both compounds are present in the spectrum. For this spectrum, a “do nothing” delay of 100 ms followed the ion ejection event, in order to keep the time scale constant for comparison with the EIEIO spectra. Figure 2B shows the fragment ions from the dissociation of the acetophenone parent molecular ion, with the electron beam at a nominal electron energy of 6 eV during the 100-ms period. The background spectrum after isolating the mo1987 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 59, NO. 7, APRIL 1, 1987
1055
QUEJCH ION F O X A T I O N
t
C8Hg+ !
.
gi
EJACT
L
EJLCT I ,
~
IONS TRANSFERRED TO ANALYZER n
ELECTRON IMPACT
riEiKi3
( l o o mr) EX=€ I
~~
I
~~~~
DETECT
Figure 1. Event sequence for electron impact excitation of ions. >
t-0
2Z
z
-0 L
z
i
120
m > W
F
67
c
6
10 W
rx
-
Figure 3. Isobaric daughter ions at m l z 105 produced by electron impact excitation of the isobaric molecular ions of acetophenone and cumene. The daughter ion resolution is 271 000 (fwhh definition). ions having a nominal mass of 105 amu. Cumene was added to the mixture of acetophenone and isophorone to produce a doublet at m / z 105. With the instrument in high-resolution mode, these two isobaric fragments could be separated with a resolution of 271 000 (full width at half-height (fwhh) definition), and identified by accurate mass measurements (Figure 3). The C6H5CO+ion produced by the fragmentation of acetophenone molecular ion was observed at 105.0328 amu (in the absence of an internal calibrant), as compared to the theoretical value of 105.0335 amu. The ion C8H9+produced by the fragmentation of the isophorone molecular ion was detected at 105.0694 amu, compared to the calculated value of 105.0699 amu. Both measured masses differ by less than 1 mamu from the calculated values. The technique is not restricted to use with ions produced by electron impact. We have applied electron impact excitation to (M H)' ions produced by chemical ionization in the source cell, isolated by ion ejection, and then transferred to the analyzer cell for electron impact excitation. For example, tert-pentyl alcohol produces no molecular ion under E1 conditions, but produces protonated molecular ion at m / z 89 under self-CI (9) conditions (ion-molecule reactions involving E1 fragment ions and sample neutral molecules). As might be expected, the (M + H)' ions fragment under EIEIO conditions to produce m / z 71 (loss of H20). The fragmentation efficiencies for the pseudomolecular ions produced by CI are similar to those observed for the molecular ions produced by electron impact (Le., approximately lo%), provided the electron beam is left on at low voltage (1eV) during the variable delay during which ion-molecule reactions occur. The highest efficiencies were obtained when the electron beam was gated off for the minimum time necessary to accomplish ion ejection (1-2 ms). These preliminary experiments demonstrate the feasibility of performing an MS/MS experiment with electron impact excitation of ions. The use of a Fourier transform mass spectrometer with a superconducting magnet has overcome the major obstacles to the development of a useful mixture analysis or structure determination method utilizing electron impact excitation of ions. In order to develop routine methods, it will be necessary to learn more about the nature of electron impact excitation. We must verify that the method is generally applicable to all compound types and determine whether the dissociation efficiency and cross section are dependent upon the nature of the parent ion. Unlike collision-induced dissociation (CID) in FTMS, the parent ion excitation in EIEIO is not mass selective. In other words, in a CID experiment, it is possible to selectively accelerate a given mass by irradiating it at its resonant frequency. Only that mass will have enough kinetic energy to undergo fragmentation upon collision with a neutral molecule. It is usually not necessary to have a clean parent ion selection in a CID experiment, since the parent ion acceleration can
+
z
10 W CY
2 2 10
40
60
80 100 MASS I N A M U
120
140
Figure 2. (A) 15-eV electron impact spectrum of mixture of acetophenone and isophorone. (B) Fragment ions produced by electron impact excitation of acetophenone molecular ion. (C) Fragment Ions produced by electron impact excitation of the isophorone molecular ion. lecular ion by ion ejection has been subtracted out to obtain this figure. Since the abundance of the molecular ion is reduced by the dissociation process, background subtraction (i.e., subtraction of the spectrum without dissociation from the spectrum with dissociation) causes the molecular ion peak to go negative, and hence, the molecular ion does not appear in the subtracted spectrum. As expected, loss of the methyl group to produce a peak at m / z 105 is predominant, followed by loss of CO to produce a peak at m / z 77. A further loss of ethylene produces a small peak at m / z 51. Figure 2C shows the fragments produced by electron impact excitation of the molecular ion of isophorone. The fragment at m / z 82 corresponds to C6H50+,which is the base peak in the 70-eV electron impact spectrum of isophorone. Further loss of a methyl group leads to the peak at m / z 67. Daughter ions produced by electron impact excitation may be detected with high resolution. As an example, both cumene and acetophenone have a molecular weight of 120 amu, and both molecular ions undergo fragmentation to produce isobaric
1056
Anal. Chem. 1987, 59. 1056-1059
be done selectively, and any ions that are not completely ejected can be removed by background subtraction. This is not true for electron impact excitation, since any ions trapped in the electron beam may undergo fragmentation, regardless of mass. Hence, electron impact excitation will require a more careful use of ion ejection for parent ion selection. For this reason, the use of a “notch ejection” ( I O ) , or preferably a tailored excitation ( I I ) , is to be preferred over successive swept-frequency ejection for the EIEIO experiment. The fact that ions must be trapped in the electron beam in order to perform the excitation means that the method will be restricted to positive ions. We believe that this limitation will not prevent the technique from being applied to solving a broad range of chemical problems. It should be possible to substitute a positive ion beam (e.g., a Cs+ beam) for the electron beam to produce fragmentation of negative ions. Electron impact excitation offers some characteristics which might make it a desirable alternative to coIlision-induced dissociation in some cases. For example, with the dual-cell geometry, ions may be transferred to the analyzer cell for CID experiments (7). This isolates the ions from reactive neutrals, prevents daughter ions from undergoing ion-molecule reactions, and permits higher resolution to be obtained. In order to perform a CID experiment, ions must be excited to a larger orbital radius. Once their orbital radius has been increased, they cannot be transferred through the conductance limit. This limitation does not apply in an EIEIO experiment, since the ions remain on-axis throughout the experiment until detected. This is an important capability for making full use of the dual-cell geometry for ion-molecule reaction experiments with different reactant of collision gases present in the source and analyzer cells, permitting multiple MS/MS experiments. For CID experiments in the dual-cell instrument, it is necessary to move ions into the analyzer cell (isolated from any reactive neutral species) and then introduce a collision gas through a pulsed valve (12). This requires several hundred milliseconds for accelerated ions to collide with the collision gas and for the gas to be pumped out of the analyzer region
to obtain high resolution. Since no collision gas is required for the electron impact experiment, less time is required for the experiment. This may make EIEIO a more desirable choice for experiments where time is a factor, such as GC-MS. Registry No. Acetophenone, 98-86-2; isophorone, 78-59-1; cumene, 98-82-8.
LITERATURE CITED Cody, R. 8.; Freiser, B. S. Anal. Chem. 1979, 51, 547-551. Fedor, D. M.; Cody, R. B.;Burinsky, D. J.; Freiser, B. S.; Cooks, R . G. Int. J . Mass Spectrom. Ion Phys. 1981, 3 9 , 55-64. McLuckey, S. A.; Salkns. L.; Cody, R. B.; Burnler, R. C.; Verma, S.; Freiser, B. S.; Cooks, R. G. Int. J . Mass Spectrom Ion Phys. 1982, 4 4 , 215-229. Freiser, 0. S. Int. J . Mass Spectrom Ion Phys. 1980, 3 3 , 263-267. Cornisarow, M. B.; Grassi, V.; Parisod, G. Chem. Phys. Left. 1978, 5 7 , 413-416. U S . Patent Application Serial No. 610502. Ghaderi, S.; Littlejohn, D. P. Collected Abstracts of the 33rd Annual Conference on Mass Spectrometry and Allied Topics ; ASMS: San Diego, CA, 1985; 727-728. Cody, R. B.; Kinsinger, J. A.; Ghaderi, S.; Amster, I. J.; McLafferty, F. W.; Brown, C. E. Anal. Chim. Acta 1988, 178, 43-66. Ghaderi, S.; Kulkarni, P. S.; Ledford, E. B., Jr.; Wilkins, C. L.; Gross, M. L. Anal. Chem. 1981, 5 3 , 428-437. Noest, A. J.; Kort, C. W. F. Comput. Chem. 1983, 7 , 81-86. Marshall, A. G.; Wang, T . G . L.; Ricca, T. L. Collected Abstracts of the 33rd Annual Conference on Mass Spectrometry and Allied Topics; ASMS: San Diego, CA, 1985; 717-710. Carlin, T. J.; Freiser, B. S. Anal. Chem. 1983, 5 5 , 571-574.
Robert B. Cody* Nicolet Analytical Instruments 5225-1 Verona Road Madison, Wisconsin 53711
Ben S. Freiser Department of Chemistry Purdue University West Lafayette, Indiana 47917 RECEIVED for review February 13, 1986. Resubmitted November 7, 1986. Accepted November 7, 1986.
Fluorescence Detection of Alkylphosphonic Acids Using p -( 9-Anthroy1oxy)phenacyl Bromide Sir: Organophosphonates have found wide use as herbicides, insecticides, and antibiotics ( I ) . Given their biocidal potency, sensitive means for monitoring trace amounts of organophosphonates in the environment are needed. Laserbased detection methods have provided excellent sensitivities in the determination of a variety of compounds (2-7). This report elaborates an analysis technique for methylphosphonic acid (la),a residue resulting from complete hydrolysis of sarin. 0
II
H3C-
P
I
fluorescence is used in tandem with microcolumn high-pressure liquid chromatography. Central to the methodology is derivatization with a fluorescent labeling agent, p ( 9 anthroy1oxy)phenacyl bromide (5). Laser-induced fluorescence analysis allows detection of these derivatives in the femtomole range.
0
-F
II R-P-OH
I OH
l a R=CH,
sarin
2a R=CH,CH, 3a R (CH,),CH 4a R = CH3(CH,),
The analysis methodology is applicable to other alkylphosphonic acids such as Za, 3a, and 4a. Laser-induced
5
To date, organophosphonates have been detected via a variety of techniques. Conversion of organophosphonates to
0003-2700/87/0359-1056$01.50/00 1987 American Chemical Society
