Mass Spectrometry
A truly interdisciplinary and versatile
analytical method
MS is used
• for the characterization of molecules ranging from small inorganic and
organic molecules to polymers and proteins. With MS we might be able to
determine the molecular weight of a molecule ion, the elemental
composition of a molecule ion, and the presence of certain functional
groups.
• for mechanistic studies of gas-phase (ion) chemistry. This is also relevant
for a better understanding of the chemistry in our atmosphere and in space.
• as a mass detector coupled to a GC, HPLC, capillary electrophoresis (CE),
and thermo-gravimetric analysis.
• as an integrated mass detector in many high vacuum systems.
UofW seminar speaker January 2002
The Gas-phase
ion chemistry
of cluster ions
Dr Paul M. Mayer
Assistant Professor
Chemistry Department
University of Ottawa
History of MS Instrumentation
http://masspec.scripps.edu/information/history/
History of MS Applications
1899
1956
Early Mass Spectrometry;
Identifying Organic Compounds with MS;
1964
1966
1974
1990
1991
GC/MS
Peptide Sequencing
Extraterrestrial Mass Spectrometry
Protein Structure
Non-Covalent Interactions with ESI
1992
1993
1993
1996
Low Level Peptide Analysis
Oligonucleotide Sequencing
Protein Mass Mapping/Fingerprinting
MS of a Virus
1999
1999
Desorption/Ionization on Silicon
Isotope-Coded Affinity Tags
MS Celebrities
Ion Chemistry
Francis William Aston
(1877 - 1945)
Cambridge University,
Great Britain; Nobel Prize
in Chemistry 1922
1st
MS
Joseph John Thomson
(1856 - 1940)
Cambridge University,
Great Britain; Nobel
Prize in Physics 1906
ESI of Biomolecules
John B. Fenn (1917)
Virginia Commonwealth
University, Richmond,
Virginia
Ion Trap Technique
Wolfgang Paul
(1913 - 1993)
University of Bonn,
Germany; Nobel Prize
in Physics 1989
Peptide Sequencing using MS
Klaus Biemann (1926)
MALDI
MIT, Cambridge,
Franz Hillenkamp (1936)
Massachusetts
University of Münster,
Germany
Mechanisms and Applications
R. Graham Cooks (1941)
Department of Chemistry,
Purdue University
West Lafayette, Indiana
Fragmentation
Mechanisms
Fred W. McLafferty
(1923)
Cornell University
Ithaca, New York
Mechanism of MALDI
& ESI
Michael Karas (1952)
University of
Frankfurt, Germany
Basic Concepts
A mass spectrometer is an instrument that produces ions and separates them in
the gas phase according to their mass-to-charge ratio (m/z). Today a wide
variety of mass spectrometers is available but all of these share the capability to
assign mass-to-charge values to ions, although the principles of operation and
the types of experiments that can be done on these instruments differ greatly.
Basically, a mass spectrometric analysis can be envisioned to be made up of the
following steps:
Sample Introduction → Ionization → Mass Analysis → Ion Detection/Data
Analysis
Samples may be introduced in gas, liquid or solid states. In the latter two cases
volatilization must be accomplished either prior to, or accompanying ionization.
Many ionization techniques are available to produce charged molecules in the
gas phase, ranging from simple electron (impact) ionization (EI) and chemical
ionization (CI) to a variety of desorption ionization techniques with acronyms
such as FAB (fast atom bombardment), PD (plasma desorption), ES
(electrospray) and MALD (matrix assisted laser desorption).
The following pages are in part from http://ms.mc.vanderbilt.edu/tutorials/ms/
Sample Introduction
Mass spectrometers are operated at reduced pressure in order to prevent
collisions of ions with residual gas molecules in the analyzer during the
flight from the ion source to the detector. The vacuum should be such that the
mean free path length of an ion, i.e., the average distance an ion travels
before colliding with another gas molecule, is longer than the distance from
the source to the detector. For example, the mean free path length of an
ion is approximately one meter at a pressure of 5x10-5 torr, i.e. about
twice the length of a quadrupole instrument.
Thus, the introduction of a sample into a mass spectrometer usually requires
crossing of a rather large pressure drop, and several means have been
devised to accomplish this.
Gas samples may be directly connected to the instrument and metered into
the instrument via a needle valve. Liquid and solid samples can be
introduced through a septum inlet or a vacuum-lock system. However, when
connecting continuous introduction techniques like gas chromatography
(GC), high performance liquid chromatography (HPLC) or capillary
electrophoresis (CE), special interfacing becomes imperative to prevent
excessive gas load.
Organic Structural Spectroscopy by Lambert, Shurvell, Lightner
Ionization
The deciding criteria are often the following:
1. Physical state of the sample
2. Volatility and thermal stability of the sample
3. Type of information sought
Comparison
•
•
•
•
EI, CI, and DI are suitable for high resolution MS;
EI works well only for thermally stable and volatile samples;
CI, SI, and DI cause much less fragmentation (normally no radicals are formed);
DI and SI must be combined with tandem mass detection (MS-MS) to extract
more structural information from fragmentation;
Organic Structural Spectroscopy by Lambert, Shurvell, Lightner
Electron Impact Ionization (EI)
purely physical
processes!
The ionization energies of many compounds are on the order of 7-14 eV but
electron energies of 70 eV are often chosen for EI-MS to achieve higher signal
intensities and to avoid changes in the mass spectrum with small changes in
electron energy.
The energy domain: 1 J (kg m2 s-2) = 6.24145 x 1018 eV
1 J mol-1 = 1.03641 x 10-5 eV
energies of chemical bonds are typically between 100 and
600 kJ mol-1 (C-H = 435 kJ mol-1; C-O = 356 kJ mol-1, C-N
= 305 kJ mol-1), which are equivalent to 1-6 eV
The time domain: a 70 eV electron has a velocity of about 5 x 108 cm s-1 and
transits a molecule of 1 nm length in 2 x 10-16 s, while a typical
bond vibration requires >10-12 s;
thus, the molecular conformation remains unchanged as the
electronic excitations occurs (Franck-Condon principle)
The degree of fragmentation depends on the internal energy deposition in
the molecular ion and on the resistance of the molecular structure to bond
cleavage (fragmentation).
Schematic representation of an electron ionization ion source. M represents neutral
molecules; e-, electrons; M+· , the molecular ion; F+, fragment ions; Vacc,
accelerating voltage; and MS, the mass spectrometer analyzer.
Schematic representation of an electron ionization ion source.
about every 1/1000
molecule is ionized
the sample is heated
up until a sufficient
vapour pressure is
obtained
sample pressure in the ion source
is about 10-5 torr
only cations
Advantages & Disadvantages of EI-MS
Organic Structural Spectroscopy by Lambert, Shurvell, Lightner
Contrasting degrees of fragmentation depending on the chemical structure
Radical cations of aromatic and unsaturated conjugated hydrocarbons resist
fragmentation more than radical cations of saturated hydrocarbons
Lowering the electron beam energy to approximately 15 eV does not
necessarily result in the observation of a molecular ion.
CH3C=O++
•OCH2CH2CH2CH3
CH2=CHCH2CH3+
+ CH3COOH
It does, however, change
the probability of different
fragmentation pathways.
Here, the H-rearrangement
wins at the expense of
single-bond cleavage.
The Molecular Ion in EI-MS
Index of hydrogen deficiency (degree of unsaturation)
The index of hydrogen deficiency is the number of pairs of hydrogen atoms that
must be removed from the corresponding saturated formula to produce the
molecular formula of the compound of interest. The index is the sum of the
number of rings, double-bonds, and twice the number of triple bonds.
Compounds can contain C, H, N, O, halogen, and S.
Index = tetravalent (C, Si) - ½ monovalent (H, halogen) + ½ trivalent (N, P) + 1
Bivalent atoms such as O and S do not contribute.
The nitrogen rule
Molecular ions containing odd numbers of nitrogen must have an odd number m/z.
Example
The compound with the molecular formula C7H7NO has an index of ??
What are possible structures?
Note that a benzene ring counts for an index of 4, 3 double bonds and one ring!
The isotopic signature – recognition of elements