RAPID COMMUNICATIONS IN MASS SPECTROMETRY, VOL. 9,911-915 (1995)

Diels- Alder Reactions of Mass-selected Ions in an Ion Trap Mass Spectrometer Manish Soni, Jon Amy, Vladimir Frankevich and R. Graham Cooks* Department of Chemistry, Purdue University, West Lafayette, IN 47907, USA

Dennis Taylor, Andrew Mckewan and Jae C. Schwartz Finnigan Corporation, San Jose, CA 95134, USA

This Communication describes the use of a bench-top ion trap mass spectrometer for the study of structurally diagnostic ion/molecule reactions. The power of the experiment is increased by the broad-band ion-isolation and tandem mass spectrometry Capabilities of the ion trap which are used to isolate the reactant ion and study its reactions under controlled conditions. The availability of multiple-stage mass spectrometry (MS", where n = number of stages) provides additional information on the nature of the product ions. The compound to be ionized is conveniently introduced into the ion trap from aqueous solution using membrane introduction techniques while the neutral reagent is leaked into the trap through the calibration gas inlet. Polar Diels-Alder reactions of the [4 + 2'1 type are investigated and comparisons are made between the reactions occurring in the ion trap and those in a pentaquadrupole mass spectrometer. Three-stage tandem mass spectra (MS3) are used to characterize the iodmolecule reaction products in the quadrupole ion trap.

The identification of gas-phase ions based on their selective reactivity with chosen neutral molecules in a mass spectrometer is the objective of an increasing amount of work.' A number of reactions, including simple proton transfer, charge exchange, and more complex reaction sequences involving the transfer of larger groups, have been developed for the purpose of ion characterization and implemented using a variety of mass spectrometers.',' To characterize these reactions, one requires versatile instruments that are capable of performing mass-selection, then of allowing the selected ions to undergo ion/molecule reactions and finally selecting and activating specific ionic products. Mass spectrometers that provide such capabilities tend to be complex instrument^,^ although the tandem-in- time capabilities of the ion trap make it a possible exception.

Many features of a quadrupole ion trap mass spec- trometer (ITMS) make it a suitable device for selective as well as highly sensitive examination of gas-phase ion/ molecule reactions. Several studies of such reactions4 including some of importance analytically as a means of differentiating isomeric specie^,^ of locating the positions of double bonds6 and of identifying ions containing certain functional groups7 have been reported using prototype ion traps such as the research grade Finnigan ITMS. Quadrupole ion traps allow control over the pressure and time for which reactions run and provide multi-stage (MS") features. The capa- bility for isolation of particular ions along with pulsed- gas introduction capability allows selection of both the charged and neutral precursors, allowing highly selec- tive reactions to be performed.6 The relatively high operating pressures (typically mTorr range) cause efficient deactivation and hence stabilization of adducts. The reaction products can also be isolated and dissociated via resonance excitation to obtain import- ant structural information. Endothermic ion/molecule

Author for correspondence.

reactions can also be examined if resonant excitation is used to activate the reactant ions.' All these capabilities have so far been available only on the larger ITMS systems; the recent introduction of bench-top ion traps with many of these advanced features should facilitate the study of ion/molecule chemistry. It is the objective of this study to explore this possibility and simul- taneously to characterize some of the features of this trap, especially its MY capability.

Acylium ions, which are well-characterized stable gas-phase species, have previously been shown to undergo site-selective polar Diels-Alder cycloaddition reactions of the [4 + 2+] type with neutral isoprene molecules.' The [4 + 2'1 type of cycloaddition involves four electrons from the neutral molecule (diene) and two electrons from the positive acylium ion (dienophile).' Eberlin et al. used a linear system of five quadrupoles (a penta uadrupole mass spectrometer) to

these with ab initio molecular orbital calculations to explore the reaction mechanism in detail.' The cycload- ducts produced in this reaction are structurally diagnos- tic and yield characteristic collision-induced dissocia- tion (CID) patterns, making this reaction analytically useful for identifying many ions of the acylium class.

In the present study, the same class of ions was examined using the Finnigan GCQm bench-top ion trap mass spectrometer. Alkyl-substituted and a, p- unsaturated acylium ions, generated from acetophe- none and cyclohexanone, respectively, and neutral iso- prene were used. The compounds were directly intro- duced into the ion trap from their aqueous solutions using the instrument's membrane inlet probe system. The ion-isolation and MS" capabilities of the trap allowed further examination of the reaction products. Comparisons of the MS3 ion trap data with those obtained by Eberlin et al.' using a pentaquadrupole instrument were of particular interest, since they showed characteristic differences which are ascribed to the effect of the differences in collision conditions in the two instruments.

perform MS2 and MS ?? experiments and supplemented

CCC 0951-4198/95/090911-05 0 1995 by John Wiley & Sons, Ltd

Received I June 1995 Accepted (revised) 24 June I995

912 DIELS-ALDER REACTIONS OF MASS-SELECTED IONS IN AN ITMS

EXPERIMENTAL A GCQ ion trap mass spectrometer (Finnigan Corp., San Jose, CA, USA) was fitted with a membrane introduction probe (MIMS Technology Inc., Palm Bay FL) to introduce low levels of the compounds directly from their aqueous solutions into the external ion source of the instrument. Liquid isoprene (Aldrich Chemical Co. , Milwaukee, WI, USA) was leaked into the ion trap through the calibration gas inlet of the GCQ. Pure water was continuously passed as a carrier through the capillary silicone membrane (length 2 cm; i.d. 0.635 mm; 0.d. 1.19 mm) of the membrane probe using the gas pressurized flow injector of the inlet system. One hundred parts-per-million (ppm) solutions of acetophenone and cyclohexanone (Aldrich) , were prepared in pure water and introduced as 1 mL plugs into the carrier stream. A flow rate of OSmL/min, probe temperature of 35 "C and 70 eV electron impact ionization (EI) were used for all experiments. Acetophenone and cyclohexanone selectively permeate the silicone membrane and yield characteristic acylium ions upon electron ionization (EI). To characterize the ion/molecule reaction products, the mass spectrometer was operated in the MS3 mode.

RESULTS AND DISCUSSION The various events in the MS3 experiment, viz. ioniza- tion of the neutral precursor, isolation of the parent acylium ions, reaction with isoprene, isolation of the resulting product ions, collision-induced dissociation (CID) and finally mass analysis, are shown in the form of a scan diagram in Fig. 1. The neutral precursor of the reagent ion was introduced continuously using mem- brane introduction mass spectrometry (MIMS) from an aqueous solution, and the vapor phase permeate was ionized, typically for 7ms. This was done in a mass- selective fashion, that is a waveform was applied during

ionization n

0 20 40 W 80 100 120 140 160 180 200 220 240

time (ms)

Figure 1. Scan diagram showing the steps of the MS3 experiment. After mass-selective ionization, the precursor acylium ions are further isolated in a narrow mass window using the GCQ broad-band waveform isolation technique. The mass-selected acylium ions are then reacted with the neutral reagent to form products from which the intact adduct is similarly isolated and then dissociated using resonance excitation. The product ions are mass-analyzed in the final step with simultaneous axial modulation for enhanced resolution.

the ionization period to continuously eject unwanted ions and store only those of interest. The waveform isolation window during ionization was set at a width of approximately 7 Thomson [l Thomson (Th) = 1 mass unit/charge] . lo Mass-selective storage was followed by an additional 20 ms period of mass-selection during which the acylium ions were isolated to unit mass resolution using another broadband waveform. Note that this isolation period employed a waveform similar to that used during ionization except that a relatively narrow isolation window (1 Th) and non-linear ampli- tude distribution for its frequency components were used. The use of such mass-selective storage improves sensitivity for the analyte ions."

In the next stage of the experiment, the isolated (acylium) ions were allowed to react with neutral re- agent molecules during a 80ms reaction period. The RF voltage level and helium gas pressure during this period control the energy of the ion/molecule collisions. l2 The resulting ion/molecule reaction pro- ducts could be identified by a mass-selective instability scan of the RF to record the reaction product (MS2) spectrum. Alternatively, since the mass-to-charge ratio of the characteristic adduct of a particular ion/molecule reaction is already known it can be isolated and disso- ciated directly, as is shown in the subsequent MS3 spectrum. In the case illustrated, isolation was carried out over a 20ms period. Characterization of the iso- lated ion is most conveniently achieved by CID, that is by recording the sequential product MS3 spectrum. Note that this procedure, in which a selected product of the ion/molecule reaction of a mass-selected ion is dissociated to reveal information on its identity, is commonly practised using other types of mass spectro- meters including ion cyclotron resonance instrument^.^ Dissociation was achieved using resonance excitation under relatively mild conditions (1.2 V, peak-to-peak) but for a period of 30ms. The final step in the MS3 experiment was mass analysis of the fragment ions (30- 300 Th) and this was performed by the standard mass- selective instability scan using axial modulation for improved resolution.

A convenient neutral precursor to acylium ions is acetophenone (PhC=OCH3; MW = 120) which yields two different acylium ions upon EI, viz. PhC-O+ mass/charge 105 Th and CH3C=O+, 43 Th. The first stage of this experiment involved isolation of 105 Th (Fig. 2(a)) and reaction with neutral isoprene. In this and all the other cases studied, three processes occur competitively when the mass-selected ion of interest is allowed to react with isoprene, viz. adduct formation, fragmentation and proton transfer between the reagent ions and neutral isoprene (and subsequent reactions which occur between the neutral and protonated iso- prene). However, only adduct formation is' diagnostic of the structure of the reactant ion and hence the intact ion/molecule adduct ions were selected and examined further. The quality of the selection of the original reactant ion as well as that of the intact adduct (173 Th) is illustrated in Fig. 2(a) and (b). The adduct corre- sponds in mass to the [4 + 2+] cycloaddition product illustrated in Scheme 1. Alternatives, including the [2 + 2+] product and acylic products, are known from calculations to be much higher in energy' but their formation cannot be completely discounted. To obtain more information on the nature of the ion/molecule

DIELS-ALDER REACTIONS OF MASS-SELECTED IONS IN AN ITMS 913

(a) 105

loo I I

.- E

3 v) C

C -

173

al > m .- +a - B

105

173

0

I t

1 105 I I7 1, . , . . , A , . , F a . , 1 , ; , , . , . , . ~

40 80 120 160 200 240 280 mlz (Thornson)

Figure 2. Reaction between mass-selected phenyl acylium ions (105 Th) generated from acetophenone (shown in (a)) and neutral isoprene yields an intact adduct of 173 Th; the mass-selected ion (shown in (b)) is dissociated by collision and the sequential product MS3 spectrum of 173 Th is shown in (c). The notation 0-0-0 is used to denote the mass-selection (0) and mass-analysis (0) events of the multiple-stage e~periment.'~ The isoprene molecule denotes a reaction step where the mass-selected ions are reacted with neutral isoprene molecules and the abbreviation CID indicates collision induced dissociation of the mass-selected ions.

reaction product, 173 Th was collisionally activated. Figure 2(c), an MS3 mass spectrum which records the fragments of 173 Th (itself generated from mass- selected 105 Th). The spectrum shows that dissociation yields ions of 131Th and 105Th. These fragments correspond to the elimination of neutrals of masses 66 and 42 and they are assigned to the retro-reaction (isoprene loss to give 105Th ions) and to loss of propene or ketene, to give 131Th. In order to seek more information on the product of 131Th, an MS4 experiment was performed using a modified Finnigan

173 105 isoprene

Scheme 1

131 I

I 0

50 70 90 110 130 150 170 190 mlz (Thornson)

Figure 3. MS4 spectrum showing fragments produced from the ion of 131 Th, itself obtained as a fragment of dissociation of the phenyl acylium ionlisoprene adduct. After isolation (a), the ions are dissociated (b) using resonance excitation to yield principally product ions at 91 Th.

ITS 40 bench-top instrument.13 In this experiment, the 131 Th ion, generated as just described, was used as the parent ion and its CID spectrum was recorded. The results, illustrated in Fig. 3, show that the major frag- ment is 91 Th, presumably C7H:. These observations are consistent with the occurrence of [4 + 2'1 cycloaddi- tion, although they do not exclude other possibilities including C1-benzoylation.

It is interesting to compare mass-selected ion/ molecule reactions in the ion trap with data for the same system recorded using a pentaquadrupole mass spectrometer. This choice of instruments also allows MS3 experiments to be performed in both cases to characterize the ion/molecule reaction products. In making such a comparison, it must be noted that there are significant differences between the conditions in the ion trap and those in a multiple stage quadrupole instrument such as the pentaquadrupole. First, the time scale in the ion trap is long compared to that in the pentaquadrupole instrument; second, collisional deac- tivation of product ions is expected to be more effective in the trap and third, collisional activation normally is more gentle in the ion trap, more collisions of lower energy transfer being involved in increasing the internal energy of the selected ion." All these factors will tend to favor lower energy rearrangement processes in the collision-induced dissociation spectra recorded using the ion trap and cause differences in ion/molecule product distributions. Interesting differences in the MS3 spectra obtained in the two instruments are indeed seen. The relative abundances of the CID fragment ions in the MS3 mass spectrum of the phenyl acylium ionlisoprene adduct (173 Th) observed earlier' using the pentaquadrupole tandem mass spectrometer were 105 Th (100%) and 131 Th (18%) while those obtained in this study were 105 Th (7%) and 131 Th (100%) (Fig. 2(c)). The pentaquadrupole results were reconfirmed by repeating the same MS3 experiment and the results are shown in Fig. 4. The yield of the 131 Th ions is

914 DIELS-ALDER REACTIONS OF MASS-SELECTED IONS IN AN ITMS

>>

105

I + 0 173

0

I + 0 173

0

105 100 -

173 b e I 0

*

t 131

I 4 J \ i. r _ _ C _ .

40 60 60 100 120 140 160 180 200

r _ _ C _ . JI .J\ 131

40 60 60 100 120 140 160 180 200

m h (Thomson)

Figure 4. MS3 spectrum showing collision-induced dissociation of the intact adduct (173Th) of the phenyl acylium ions (105Th) and neutral isoprene molecules recorded using a pentaquadrupole instru- ment for comparison with the MS3 spectrum recorded using the ion trap (Fig. 2(c)). Note the differennce in relative abundances of 131 to 105 Th in the two spectra.

higher than that for the retro process which yields 105Th ions in the case of the ion trap whereas the pentaquadrupole barely yields 131 Th. It is thus likely that formation of 131 Th, presumably due to a propene loss, may actually proceed via a phenyl migration with subsequent ketene loss from the intact adduct. Another possibility which we will not investigate further here is that the substantially different conditions in the quad- rupole ion trap can alter the course of the reaction of the benzoyl ion and isoprene, and lead to an acyclic or other addition product.

The reagent acetophenone, in addition to allowing the generation of the benzoyl ion, is a source of a second type of acylium ion, the acetyl ion of 43 Th. The reactions of this ion with neutral isoprene were also studied. The isolated reactant ion reacted with neutral isoprene to yield the intact adduct at 111 Th among other products. The sequential product spectrum (MS3 experiment) is reproduced in Fig. 5. This spectrum shows the retro-reaction to yield the starting material, 43Th, as well as a low abundance ion at 93Th corre- sponding to elimination of water from the adduct. In all these respects, as well as in a comparison with the reactions observed in the pentaquadrupole instrument, this ion behaves analogously to the benzoyl ion.

43

l m a I I . I11

0

93

m/z (Thornson)

Figure 5. Reaction of acetyl ions (43 Th) generated from acetophe- none and neutral isoprene yields a single cycloaddition product, 111 Th. This ion dissociates upon collision to generate the sequential (MS3) product spectrum shown.

I

55 I I

0

123

0 55

I I

123

0

95 I

rnlz (Thornson)

Figure 6. The a, j3-unsaturated acylium ions of cyclohexanone also undergo cycloaddition with isoprene to yield a complex MS2 spec- trum also containing the adduct ions at 123 Th (a). Collision-induced dissociation of the mass-selected intact adduct yields products at 95, 81 and 55 Th which are the result of losses of CO, C3H, and isoprene, respectively, as shown in the MS3 spectrum in (b).

In another experiment of the same type, cyclo- hexanone was introduced into the trap using MIMS and the resulting a, P-unsaturated acylium ions CH,=CHC=O+ (55 Th) were isolated for iodmolecule reaction. Upon reaction with isoprene under identical conditions as before, adduct ions at 123Th are obtained. Figure 6(a) is a MS2 spectrum recorded after the ion/molecule reaction period without any isolation. It is complex and contains in addition to the diagnostic adduct ions, species that are the result of various different reaction channels which are difficult to inter- pret. Ions at 67Th and 69Th are characteristic of charge exchange and/or proton transfer reactions of isoprene while 95 and 81Th are fragments of the adduct (123 Th), but this is only confirmed by the MS3 experiment. It should also be noted that without mass- selection (isolation) the mass spectrum shows poor resolution and sensitivity because excess ion abundance and high operating pressures cause space-charge induced distortion. l1 The intact adducts were then iso- lated and then fragmented to yield the MS3 mass spectrum (Fig. 6(b)). Fragmentation yielded major products at 95, 81 and 55Th and additional minor fragments at 121, 107, and 93 Th. Of the major prod- ucts, 55 Th is the retro product, and 81 and 95 are most readily explained as the products of ketene and carbon monoxide loss from cycloaddition at the activated C=C bond of the reactant ion. The fact that loss of 28 mass units occurs exclusively for the a, P-unsaturated acy- lium ion adduct but not from adducts derived from the acetyl or benzoyl ion is an indication that site-selective

DIELS-ALDER REACTIONS OF MASS-SELECTED IONS IN AN ITMS 915

isoprene 55 123

Scheme 2

cycloaddition takes place at the C=C bond in this case (Scheme 2). The same reaction has also been proposed to occur in the pentaquadrupole mass spectrometer on the basis of MS2 and MS3 spectra and its occurrence is attributed to activation of the C=C double bond by the electron-withdrawing effect of the C=O+ group.' As in the case of the benzoyl ion reaction, there are signifi- cant differences between the MS3 spectra recorded using the two instruments. Again, fragment ions (at 81 Th) which involve significant rearrangement are higher in abundance than the retro fragmentation pro- duct ions (at 55 Th). Note that unlike 173 Th (benzoyl ion adduct), the abundance of 123 Th (a , @-unsaturated ion adduct) was so low in the pentaquadrupole study that an MS3 experiment could not be performed for comparison.

CONCLUSION Polar Diels-Alder cycloaddition reactions of the [4+2+] type were carried out using acylium ions and neutral isoprene on a bench-top ion trap mass spectro- meter. The quality of the data agree well with that obtained in a previous pentaquadrupole study, even though the reaction conditions differ sufficiently to produce differences in ion abundances, and possibly even differences in reaction mechanism. The selective nature of this reaction makes it diagnostic for identify- ing acylium ions in the gas phase. It is shown that the broad-band ion-isolation and MS" features of the GCQ allow such complex but analytically useful experiments to be performed with relative ease and simplicity.

Acknowledgements This work was supported by the Division of Chemical Sciences, Offiice of Basic Energy Sciences, Office of Energy Research, U.S. Department of Energy (DE-FG02-94ER14470) and Finnigan

Corporation through the Chemistry Department Industrial Associates Program. We thank Eric Johnson, George Stafford and Mark Bier for their assistance. Special thanks to Sheng Sheng Yang for obtaining the MS3 data recorded using the pentaquadrupole mass spectrometer.

REFERENCES 1. (a) H. I. Kenttamaa, Org. MassSpectrom. 29,1(1994). (b) H.-F.

Grutzmacher, Org. Mass Spectrom. 28, 1375 (1993). (c) P. Longevialle, Mass Spectrom. Reu. 11, 157 (1992). (d) H. Schwarz, Chem. Reu. 91, 1121 (1991). (e) N. M. M. Nibbering, Acc. Chem. Res. 23, 279 (1990). (f) C. Dass, Mass Spectrom. Reu. 9,1(1990). (g) M. Vairamani, U. A. Mirza and R. Srinivas, Mass Spectrom. Reu. 9,235 (1990); (h) E. G. Eichman and J. S. Brodbelt, J . Am. SOC. Mass Spectrom. 4, 97 (1993); (i) J. Einhorn, H. I. Kenttamaa and R. G. Cooks, J . Am. SOC. Mass Spectrom. 2, 305 (1991).

2. (a) M. N. Eberlin, N. H. Morgon, S. S. Yang, B. J. Shay and R. G. Cooks, J . Am. SOC. Mass Spectrom. 6 , l (1995). (b) R. D. Guettler, G. C. Jones Jr., L. A. Posey and R. N. Zare, Science, 266, 259 (1994). (c) G. Bouchoux and J.-Y. Salpin, Rapid Commun. Mass Spectrom. 8, 325 (1994). (d) M. J. Cole and C. G. Enke, J . Am. SOC. Mass Spectrom. 2,470 (1991).

3. Techniques for the Study of Ion-Molecule Reactions, J. M. Farrar and W. H. Saunders Jr. (Eds), John Wiley, New York 1988.

4. (a) M. Moini, J . Am. SOC. Mass Spectrom. 3, 631 (1992). (b) T. Donovan, C.-C. Liou and J. Brodbelt, J. Am. SOC. Mass Spectrom. 3, 39 (1992). (c) B. D. Nourse, K. A. Cox and R. G. Cooks, Org. Mass Spectrom. 27,453 (1992). (d) J. Brodbelt and R. G. Cooks, Anal. Chim. Acta 206,239 (1988).

5. (a) G. L. Glish, Analyst 119, 533 (1994). (b) K. J. Hart, S. A. McLuckey and G. L. Glish, J. Am. SOC. Mass Spectrom. 3, 680 (1992). (c) C. Kascheres and R. G. Cooks, Anal. Chim. Acta. 215,223 (1988).

6. J. Einhorn, H. I. Kenttamaa and R. G. Cooks, J . Am. SOC. Mass Spectrom. 2, 305 (1991).

7. T. D. McCarley and J. Brodbelt, Anal. Chem. 65, 2380 (1993). 8. H.-F. Wu and J. S. Brodbelt, Int. J . Mass Spectrom. Ion

Processes, 124, 175 (1993). 9. M. N. Eberlin and R. G. Cooks, J . Am. Chem. SOC. 115, 9226

(1993). 10. R. G. Cooks and A. L. Rockwood, Rapid Commun. Mass

Spectrom. 5, 93 (1991). 11. M. H. Soni, S. Bauer, J. Amy, P. Wong and R. G. Cooks, Anal.

Chem. 67, 1409 (1995). 12. (a) K. J. Hart and S. A. McLuckey, J . Am. Chem. SOC. 5,250

(1994). (b) J. Gronowska, C. Paradisi, P. Traldi and U. Vettori, Rapid Commun. Mass Spectrom. 4,306 (1990). (c) J. N. Louris, R. G. Cooks, J. E. P. Syka, P. E. Kelley, G. C. Stafford and J. F. J. Todd, Anal. Chem. 59,1677 (1987).

13. M. H. Soni, P. Wong and R. G. Cooks, Anal. Chim. Acta, 303, 149 (1995).

14. J. C. Schwartz, A. P. Wade, C. G. Enke and R. G. Cooks, Anal. Chem. 62, 1809 (1990).