Proceedings of the 5th Annual GRASP Symposium, Wichita State University, 2009
What Does That Molecule Look Like?
Using Tandem Mass Spectrometry, Computational Chemistry and
Vibrational Spectroscopy to Determine Molecular Structure
Ryan P. Dain* and Michael J. Van Stipdonk
Department of Chemistry, Fairmount College of Liberal Arts and Sciences
Abstract: Scientists wanting to determine the structure of a molecule have many tools at their disposal. Tandem mass
spectrometry (MS/MS) allows one to study the fragmentation pathways of molecules, examining how a molecule will fall apart
when energy is added to it through a process known as collision induced dissociation (CID). By measuring the mass and
abundance of these fragments, one can make determinations about the original, or parent, species. Computational chemistry
allows one to model a molecule with many different structures, determining which represents the most likely one by looking at
the relative energies and theoretical infrared (IR) vibrational spectra. Vibrational spectroscopy is used because each molecule, in
principle, has a different IR spectrum that depends on its structure, much like a fingerprint. The theoretical IR spectra for various
structures can then be compared to an experimental IR spectrum, to establish the true conformation. Therefore, using these three
tools a scientist can confidently determine the structure of a molecule, and a better understanding about the innate chemistry of
that molecule.
1. Introduction
Determining molecular structure is important because it allows for the investigation of the fundamental
properties of the systems of interest. Mass spectrometry and vibrational spectroscopy have been used for many years
to investigate chemical structures. With the rapid increase of availability and quality of computational resources,
computational chemical modeling has bridged the gap in the understanding of experimental results. These three
methods provide an excellent way for scientists to investigate the intrinsic chemistry of discrete gas-phase ions by
determining their molecular structure. Our research group at Wichita State has used these methods extensively over
the past years and what follows will give a better understanding of our methods and what we can do with them.
2. Experiment, Results, Discussion, and Significance
Mass spectrometry experiments are conducted in our lab at Wichita State University. We use aThermoFinnigan LCQ-DECA quadrapole mass spectrometer. The species of interest are generated by a process known as
electrospray ionization (ESI), where a liquid sample is sprayed through a fine needle that has a voltage running
across it. This process causes the liquid sample to go into the gas phase and the voltage creates a charge on the
molecule, turning them into ions. Mass spectrometry measures these discrete, gas-phase ions and displays them
according to their mass to charge (m/z) ratio. Ions with a certain m/z can be isolated and reacted with energized
helium atoms to break apart the bond in the ion through a process known as collision induced dissociation (CID).
The fragments will be displayed at a lower m/z ratio and information about the structure of the original, or parent,
ion can be deduced from the m/z and abundance of these fragments. This is a good way to get basic structural
information about a molecular system.
Since we work in the gas-phase, we can study discrete molecules. This means that when we want to
examine a certain system, we can look at just one single molecule without worrying about other factors affecting the
molecule. Luckily the best computational chemistry programs are set up to model single molecule systems, making
them an important tool in the effort to determine molecular structure. We use the Gaussian 03 series of programs,
developed by John Pople in the 70’s and 80’s. The program allows us to build a model of the molecule, starting with
the basic structure derived from the CID spectrum, and then using complex quantum mechanical equations to
theoretically predict the behavior of the electrons in that molecule. By modeling the interactions of the electrons, the
program can predict the bonding behavior of the molecule. When the program models, or optimizes, the geometry of
the molecule, it also provides intrinsic information about the system, such as energy and vibrational modes. By
comparing relative energies, you can make deductions about which conformation is more likely to be the true
conformation. The best way to tell is to compare the theoretical vibrational spectrum to an actually experimental
benchmark.
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Proceedings of the 5th Annual GRASP Symposium, Wichita State University, 2009
Infrared multiple photon dissociation (IRMPD) spectroscopy provides this benchmark for our systems. This
process involves creating ions as described above, but then bombarding them with energized photons at different
wavelengths, generated by a free electron laser. The photons add energy to the molecule, making it vibrate to the
point where the bonds break. The fragments are then measured at each different wavelength and graphed as
photodissociation as a function of wavelength. This provides an infrared (IR) spectrum that can be used as the
benchmark for the theoretical to experimental comparison. By matching the IRMPD spectrum to the predicted IR
spectrum generated by the Gaussian program, you can make a definitive determination about the structure of your
species of interest. Since free electron lasers are very rare and very expensive, we work in collaboration with the
FOM Institute for Plasma Physics, located in Nieuwegein, The Netherlands. They run the Free Electron Laser for
Infrared eXperiments (FELIX) facility where the IRMPD spectra for our work are collected.
This method has been used again and again by our research group at Wichita State to study the fundamental
chemistry of many interesting systems, both organic and inorganic in nature [1-5]. This method has become a vital
tool for scientists in this field to learn as much as they can about the systems they are studying. Below is an example
of these three tools in use, to determine the structure of the b2+ product from the peptide Trialanine, AAA.
IR spectroscopy of b2+ from AAA
Photodissociation
yield
Formation of b2+ from protonated AAA
143
100
b2+
R. I. (%)
80
60
O
H2N
b2+
a2+
N
H
H
N
CID (MS/MS) of (M+H)+
O
OH
O
IRMPD
1
0
-89 (Ala)
40
2
(M+H)+
6
DFT*
oxazolone
232
20
3
115
R. I. (%)
80
Intensity
0
100
CID (MS3) of b2+
a2+
0
9
6
60
-28 (CO)
DFT
diketopiperazine
3
40
0
20
0
25
600
50
75
100
125
150
175
200
225
250
*B3LYP/6-311+g(d,p)
m/z
800
1000
1200
1400
1600
1800
2000
-1
Frequency (cm )
J. Oomens … M. J. Van Stipdonk, J. Am. Soc. Mass Spectrom., 20, 334-339 (2008).
This shows the CID of AAA, forming the b2+ product ion. The two possible structures of the b2+ product ion, either
featuring an oxazolone or a diketopiperazine ring structure, were modeled and compared to the IRMPD spectrum. It
can be seen that the b2+ product ion most likely features the oxazolone ring.
3. Conclusions
So, it can be seen that these different methods can be used to determine the structure of ions of interest that
we want to study. While any one technique will give you part of the picture, all three are needed to give you the
whole story.
4. Acknowledgements
I would like to thank all responsible for the ongoing support of this work, including WSU, NSF, US
Department of Energy, Nederlandse Organisatie voor Wetenschappelijk Onderzoek, all members of the Van
Stipdonk research group, past and present, and our talented collaborators around the globe.
[1] Groenewold, G. S.; Oomens, J.; de Jong, W. A.; Gresham, G. L.; McIlwain, M. E.; Van Stipdonk, M. J. Vibrational Spectroscopy of Anionic
Nitrate Complexes of UO22+ and Eu3+ Isolated in the Gas Phase. Phys. Chem-Chem. Phys. 2008, 10, 1192-1202.
[2] Oomens, J.; Myers, L.; Dain, R.; Leavitt, C.; Pham, V.; Gresham, G.; Groenewold, G.; Van Stipdonk, M. Infrared Multiple-Photon
Photodissociation of Gas-Phase Group II Metal-Nitrate Anions. Int. J. Mass Spectrom. 2008, 273, 24-30.
[3] Van Stipdonk, M. J.; Kerstetter, D.K.; Leavitt, C.M.; Groenewold, G.S.; Steill, J; Oomens, J. Spectroscopic Investigation of H Atom Transfer
in a Gas-Phase Dissociation Reaction: McLafferty Rearrangement of Model Gas-Phase Peptide Ions. Phys. Chem-Chem. Phys. 2008,
10, 3209 – 3221.
[4] Groenewold, G. S.; Gianotto, A. K.; Cossel, K. C.; Van Stipdonk, M. J.; Moore, D. T.; Polfer, N.; Oomens, J.; de Jong, W. A.; Visscher, L.
Vibrational Spectoscopy of Mass-Selected [UO2(ligand)n]2+ Complexes in the Gas Phase: Comparision with Theory. J. Am. Chem.
Soc. 2006, 128, 4802-4813.
[5] Leavitt, CM; Oomens, J; Dain, RP; Steill, J; Groenewold, GS; Van Stipdonk, MJ. IRMPD Spectroscopy of Anionic Group II Metal Nitrate
Cluster Ions. J. Amer. Soc. Mass. Spect. Article in Press.
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