6. THEORETICAL ESTIMATION OF PEPTIDE INTERNAL TEMPERATURE FOR
6.3. Methods and Theory
6.4.2. Comparison of Simulation and Experiment
When the first, preliminary simulations were performed, it was observed that product ions were formed with more internal energy than expected. This was clear from observing the b4+/a4+ ratio of YGGFL, which has been used as an indicator of the internal energy
deposited into the parent ion.23, 24 Typically for YGGFL, the b
4+/a4+ ratio is much greater
than one at low CID voltages, and makes a transition to less than one as CID voltage is raised. In the first simulations, the b4+/a4+ ratio was less than one as soon as b4+ was formed,
indicating that b4+ was being formed from [YGGFL+H]+ with an average internal energy
much higher than the critical energy for dissociation to a4+ of 1.32eV. Experimentally,
however, when product ions are formed they are no longer in resonance with the CID voltage (their mass and therefore secular frequency in the ion trap has changed) and collisions with the He bath gas have an overall cooling effect instead of an activating effect. Thus, product ions possibly are cooled significantly before they dissociate.24 A collisional cooling term,
132
rate of the product ions, as in Equation 6.9. The collisional cooling term is not subtracted from the dissociation rates of the parent ions because they are being accelerated by the excitation field, and therefore the probability of a collision resulting in a loss of internal energy is much smaller.
cc reaction
final k k
k = − (Equation 6.9)
The collisional cooling rates of peptides in a QITMS at 1 mTorr have been calculated previously.25 When the diffuse scattering method is applied to YGGFL25, the resulting k
cc is
1800 s-1 when activated to 830 K, and 1100 s-1 when activated to 545 K. A recent experimental study in our lab found this rate to be 400 s-1 for YGGFL at 1 mTorr when activated for 5 ms with a 50 W CO2 laser (an activation of 90 K or 0.9 eV).26, 27 This result
agrees with the master equation modeling25, where extrapolation of the master equation data
for an ion with the number of degrees of freedom of YGGFL yields kcc=600 s-1 for a 300 K
activation. The best results were found when kcc=500-2000 s-1. 2000 s-1 was used in the
results shown. This number, which was obtained empirically, is reasonable when compared to the above theoretical and experimental values. Figure 6.6 shows the results of comparing simulated to experimental MS/MS spectra for conventional and HASTE CID. The sum of the squared differences in relative peak intensities between experiment and calculation was used to find the temperature that gave the best fit. Differences in internal temperature
between HASTE and conventional CID, as opposed to absolute temperature values, are more informative for analyzing the results across all the ions studied, because disparities in ion structure and vibrational modes have an influence on the calculated p(E) at a given
temperature. For the sodiated peptide ions, the internal temperature of the HASTE activated parent ions was 80-90 K higher than the conventional CID activated ions, while the sodiated
133
cluster ion had a difference of 115 K. The protonated peptide ion, YGGFL, was activated 285 K higher with HASTE than conventional CID. Table 6.3 summarizes this temperature data. The above temperature differences translate to an average parent ion internal energy about 1.0 eV higher for the sodiated ions, and about 3.5 eV higher for protonated YGGFL. The resulting increase in product ion intensities for HASTE compared to conventional CID is apparent in Figure 6.6a-h. The result that the protonated ion [YGGFL+H]+ was activated to much higher internal temperature than the sodiated ions stands out. This higher calculated temperature might be explained by the complexity of the [YGGFL+H]+ dissociation pathways. The many competitive dissociations present may compound the error in determining dissociation parameters, and a Boltzmann distribution might be a worse approximation for [YGGFL+H]+ internal energies after HASTE activation than for conventional CID. It should be noted that if only a4+, b4+, and [M+H]+ are considered for
[YGGFL+H]+, then an activated temperature of 650 K results, which is in better agreement with the sodiated ion data.
Table 6.3: Effective Temperatures Achieved During Conventional CID and HASTE CID [YGAFL+Na]+ [YGGFL+Na]+ [YGGFL+H]+ [CH3COONa]6Na
+ Conventional Teff(K) 610 675 545 500 HASTE Teff(K) 690 765 830 615 ∆Teff(K) 80 90 285 115
The calculated intensities of the [b2+NaOH]+ ions and [CH3COONa]2Na+ ion did not
match the experiment very well, as is seen in Figure 6.6, even though the breakdown curves from their respective parent ions had a very good fit.(not shown) It is possible that the later generation ions are formed through multiple pathways, which could be determined through
134
double resonance experiments. Overall, there is a good visual fit between experimental and simulated CID spectra. This lends some credence to the original approximation that a thermal distribution of ion internal energies could be assumed for CID activated parent ions.
135
a) b)
c) d)
e) f)
g) h)
Figure 6.6. Blue, x-striped boxes are calculated ion abundances, green, diagonally-striped boxes are experimental ion abundances. Calculations were done at 1 mTorr with kcc = 2000
s-1. (a)(b) Conventional and HASTE CID for [YGAFL+Na]+, where theoretical effective temperatures were 610 K and 690 K, respectively. (c)(d) Conventional and HASTE CID for [YGGFL+Na]+, where theoretical effective temperatures were 675 K and 765 K, respectively. (e)(f) Conventional and HASTE CID for [CH3COONa]6Na+, where
theoretical effective temperatures were 500 K and 615 K, respectively. (g)(h)
Conventional and HASTE CID for [YGGFL+H]+, where theoretical effective temperatures were 545 K and 830 K, respectively.
136 6.5.Conclusions
The temperature of ions activated by conventional CID and HASTE CID has been estimated via a comparison of theoretical MS/MS spectra calculated at different parent ion effective temperatures with experimental MS/MS spectra. The effect of collisional cooling had to be included in the calculations to adequately fit the experimental to calculated spectra. Reaction parameters for each dissociation pathway were determined by fitting experimental and theoretical breakdown curves. The results indicate that the sodiated ions were activated to an average internal temperature of 95 K higher, and the protonated ion was activated 285 K higher using HASTE CID over conventional CID. This translates into an average internal energy of 1.0 eV higher for the sodiated ions and 3.5 eV higher for the protonated ion. As mentioned in the results section, the protonated ion result is somewhat suspect, and the difference in internal temperatures for the protonated ion may in fact be closer to 105 K, 1.0 eV higher internal energy. This study does not claim to provide exact values of the extent of ion activation for the investigated ions, rather, an estimation has been made of the advantage of HASTE CID over conventional CID in terms of internal temperature increase. Relatively simple algorithms were used describe the dissociation kinetics of the ions fairly accurately, as is apparent from Figure 6.6. The likelihood that a thermal internal energy distribution would approximate the activation of ions by HASTE CID was questionable in the onset of the study, because the HASTE CID process probably involves a smaller number of much higher energy collisions than conventional CID. Activation by HASTE may therefore be
comparable to higher collision energy CID, such as those obtained on a triple quadrupole instrument.
137 6.6.References
1. Vachet, R. W.; Bishop, B. M.; Erickson, B. W.; Glish, G. L. Novel Peptide Dissociation: Gas-Phase Intramolecular Rearrangement of Internal Amino Acid Residues. J. Am. Chem. Soc. 1997, 119, 5481-5488.
2. Goeringer, D. E.; McLuckey, S. A. Evolution of Ion Internal Energy During Collisional Excitation in the Paul Ion Trap: A Stochastic Approach. Journal of Chemical Physics 1996, 104, 2214-2221.
3. Marzluff, E. M.; Beauchamp, J. L. In Large Ions: Their Vaporization, Detection and Structural Analysis; Baer, T. N., Cheuk-Yiu; Powis, Ivan, Ed.; John Wiley and Sons Ltd: Chichester, 1996, pp 115-143.
4. Gabelica, V.; Karas, M.; De Pauw, E. Calibration of Ion Effective Temperatures Achieved by Resonant Activation in a Quadrupole Ion Trap. Anal. Chem. 2003, 75, 5152-5159.
5. Laskin, J.; Byrd, M.; Futrell, J. Internal energy distributions resulting from sustained off- resonance excitation in FTMS. I. Fragmentation of the bromobenzene radical cation. Int. J. Mass Spectrom. 2000, 195/196, 285-302.
6. Plass, W. R.; Cooks, R. G. A Model for Energy Transfer in Inelastic Molecular Collisions Applicable at Steady State or Non-Steady State and for an Arbitrary Distribution of Collision Energies. J Am Soc Mass Spectrom 2003, 14, 1348–1359.
7. Asano, K. G.; Goeringer, D. E.; McLuckey, S. A. Thermal Dissociation in the Quadrupole Ion Trap: Ions Derived From Leucine Enkephalin. Int. J. Mass Spectrom. 1999,
185/186/187, 207-219.
8. Cunningham, C., Jr.; Glish, G. L.; Burinsky, D. J. High Amplitude Short Time Excitation: A Method to Form and Detect Low Mass Product Ions in a Quadrupole Ion Trap Mass Spectrometer. J Am Soc Mass Spectrom 2006, 17, 81 - 84.
9. Louris, J. N.; Cooks, R. G.; Syka, J. E. P.; Kelley, P. E.; Stafford, G. C.; Todd, J. F. J. Instrumentation, Applications, and Energy Deposition in Quadrupole Ion Trap Mass Spectrometry. Anal. Chem. 1987, 59, 1677-1685.
10. Douglas, D. J. Mechanism of the Collision-Induced Dissociation of Polyatomic Ions Studied by Triple Quadrupole Mass-Spectrometry. J. Phys. Chem. 1982, 86, 185-191.
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11. Baer, T.; Mayer, P. M. Statistical Rice-Ramsperger-Kassel-Marcus quasi-equilibrium theory calculations in mass spectrometry. J. Am. Soc. Mass Spectrom. 1997, 8, 103-115.
12. Baer, T.; Hase, W. L. Unimolecular Reaction Dynamics; Oxford University Press: New York, 1996.
13. Vestal, M. L. Theoretical studies on the unimolecular reactions of poly-atomic molecule ions. Journal of Chemical Physics 1965, 43, 1356-69.
14. LabVIEW [software] 7.1 National Instruments Corporation.
15. Frisch, M. J. Gaussian 03, Revision C.02 Gaussian, Inc., Wallingford CT. Gaussian 03, Revision C.02 Gaussian, Inc., Wallingford CT 2004.
16. Scott, A. P.; Radom, L. Harmonic Vibrational Frequencies: An Evaluation of Hartree- Fock, Moeller-Plesset, Quadratic Configuration Interaction, Density Functional Theory, and Semiempirical Scale Factors. J. Phys. Chem. 1996, 100, 16502-16513.
17. Foresman, J. B., Frisch Aeleen Exploring Chemistry with Electronic Structure Methods; Gaussian, Inc.: Pittsburgh, PA, 1993.
18. Laskin, J.; Denisov, E.; Futrell, J. H. Fragmentation energetics of small peptides from multiple-collision activation and surface-induced dissociation in FT-ICR MS. Int. J. Mass Spectrom. 2002, 219, 189-201.
19. Laskin, J.; Denisov, E.; Futrell, J. A comparative study of collision-induced and surface- induced dissociation. 1. Fragmentation of protonated dialanine. J. Am. Chem. Soc. 2000, 122, 9703-9714.
20. Vachet, R. W.; Ray, K. L.; Glish, G. L. Origin of Product Ions in the MS/MS Spectra of Peptides in a Quadrupole Ion Trap. J. Am. Soc. Mass Spectrom. 1998, 9, 341-344.
21. Jue, A.; Racine, A. H.; Glish, G. L. Comparison of Dissociation Pathways at Ambient and Elevated Temperatures in a Quadrupole Ion Trap. Nashville, TN, 2004.
22. Schnier, P. D.; Price, W. D.; Strittmatter, E. F.; Williams, E. R. Dissociation Energetics and Mechanisms of Leucine enkephalin (M+H)+ and (2M+X)+ Ions (X=H, Li, Na, K, and
139
Rb) Measured by Blackbody Infrared Radiative Dissociation. J. Am. Soc. Mass Spectrom. 1997, 8, 771-780.
23. Vachet, R. W.; Glish, G. L. Effects of Heavy Gases on the Tandem Mass Spectra of Peptide Ions in the Quadrupole Ion Trap. J. Am. Soc. Mass Spectrom. 1996, 7, 1194-1202.
24. Asano, K. G.; Goeringer, D. E.; Butcher, D. J.; McLuckey, S. A. Bath Gas Temperature and the Appearance of Ion Trap Tandem Mass Spectra of High-Mass Ions. Int. J. Mass Spectrom. 1999, 190/191, 281-293.
25. Goeringer, D. E.; McLuckey, S. A. Relaxation of Internally Excited High-mass Ions Simulated under Typical Quadrupole Ion Trap Storage Conditions. Int. J. Mass Spectrom. 1998, 177, 163-174.
26. Black, D. M.; Payne, A. H.; Glish, G. L. Determination of Cooling Rates in a Quadrupole Ion Trap. J Am Soc Mass Spectrom 2006, 17, 932-938.
27. Goeringer, D. E.; Asano, K. G.; McLuckey, S. A. Ion internal temperature and ion trap collisional activation: protonated leucine enkephalin. Int. J. Mass Spectrom. 1999, 182/183, 275-288.
Chapter 7
7. On the Time Scale of Internal Energy Relaxation of AP-MALDI and Nano-ESI Ions