Mass Spectrometry E-Book

Edmond de Hoffmann

• Herausgeber: John Wiley & Sons
• Kategorie: Wissenschaft und neue Technologien
• Sprache: Englisch

Seitenzahl: 768

Veröffentlichungsjahr: 2013

Beschreibung

The latest edition of a highly successful textbook, Mass Spectrometry, Third Edition provides students with a complete overview of the principles, theories and key applications of modern mass spectrometry. All instrumental aspects of mass spectrometry are clearly and concisely described: sources, analysers and detectors. Tandem mass spectrometry is introduced early on and then developed in more detail in a later chapter. Emphasis is placed throughout the text on optimal utilisation conditions. Various fragmentation patterns are described together with analytical information that derives from the mass spectra. This new edition has been thoroughly revised and updated and has been redesigned to give the book a more contemporary look. As with previous editions it contains numerous examples, references and a series of exercises of increasing difficulty to encourage student understanding. Updates include: Increased coverage of MALDI and ESI, more detailed description of time of flight spectrometers, new material on isotope ratio mass spectrometry, and an expanded range of applications. Mass Spectrometry, Third Edition is an invaluable resource for all undergraduate and postgraduate students using this technique in departments of chemistry, biochemistry, medicine, pharmacology, agriculture, material science and food science. It is also of interest for researchers looking for an overview of the latest techniques and developments.

Leseprobe

Contents

Preface

Introduction

Principles

Diagram of a Mass Spectrometer

History

Ion Free Path

1 Ion Sources

1.1 Electron Ionization

1.2 Chemical Ionization

1.3 Field Ionization

1.4 Fast Atom Bombardment and Liquid Secondary Ion Mass Spectrometry

1.5 Field Desorption

1.6 Plasma Desorption

1.7 Laser Desorption

1.8 Matrix-Assisted Laser Desorption Ionization

1.9 Thermospray

1.10 Atmospheric Pressure Ionization

1.11 Electrospray

1.12 Atmospheric Pressure Chemical Ionization

1.13 Atmospheric Pressure Photoionization

1.14 Atmospheric Pressure Secondary Ion Mass Spectrometry

1.15 Inorganic Ionization Sources

1.16 Gas-Phase Ion-Molecule Reactions

1.17 Formation and Fragmentation of Ions: Basic Rules

2 Mass Analysers

2.1 Quadrupole Analysers

2.2 Ion Trap Analysers

2.3 The Electrostatic Trap or ‘Orbitrap’

2.4 Time-of-Flight Analysers

2.5 Magnetic and Electromagnetic Analysers

2.6 Ion Cyclotron Resonance and Fourier Transform Mass Spectrometry

2.7 Hybrid Instruments

3 Detectors and Computers

3.1 Detectors

3.2 Computers

4 Tandem Mass Spectrometry

4.1 Tandem Mass Spectrometry in Space or in Time

4.2 Tandem Mass Spectrometry Scan Modes

4.3 Collision-Activated Decomposition or Collision-Induced Dissociation

4.4 Other Methods of Ion Activation

4.5 Reactions Studied in MS/MS

4.6 Tandem Mass Spectrometry Applications

5 Mass Spectrometry/Chromatography Coupling

5.1 Elution Chromatography Coupling Techniques

5.2 Chromatography Data Acquisition Modes

5.3 Data Recording and Treatment

6 Analytical Information

6.1 Mass Spectrometry Spectral Collections

6.2 High Resolution

6.3 Isotopic Abundances

6.4 Low-mass Fragments and Lost Neutrals

6.5 Number of Rings or Unsaturations

6.6 Mass and Electron Parities, Closed-shell Ions and Open-shell Ions

6.7 Quantitative Data

7 Fragmentation Reactions

7.1 Electron Ionization and Fragmentation Rates

7.2 Quasi-Equilibrium and RRKM Theory

7.3 Ionization and Appearance Energies

7.4 Fragmentation Reactions of Positive Ions

7.5 Fragmentation Reactions of Negative Ions

7.6 Charge Remote Fragmentation

7.7 Spectrum Interpretation

8 Analysis of Biomolecules

8.1 Biomolecules and Mass Spectrometry

8.2 Proteins and Peptides

8.3 Oligonucleotides

8.4 Oligosaccharides

8.5 Lipids

8.6 Metabolomics

9 Exercises

Questions

Answers

Appendices

1 Nomenclature

2 Acronyms and abbreviations

3 Fundamental Physical Constants

4A Table of Isotopes in Ascending Mass Order

4B Table of Isotopes in Alphabetical Order

5 Isotopic Abundances (in %) for Various Elemental Compositions CHON

6 Gas-Phase Ion Thermochemical Data of Molecules

7 Gas-Phase Ion Thermochemical Data of Radicals

8 Literature on Mass Spectrometry

9 Mass Spectrometry on Internet

Index

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Library of Congress Cataloging-in-Publication Data

Hoffmann, Edmond de.

[Spectrométrie de masse. English]

Mass spectrometry: principles and applications. – 3rd ed. / Edmond de Hoffmann, Vincent Stroobant.

p. cm.

Includes bibliographical references and index.

ISBN 978-0-470-03310-4

1. Mass spectrometry. I. Stroobant, Vincent. II. Title.

QD96.M3 H6413 2007

573′.65 – dc22

2007021691

British Library Cataloguing in Publication Data

A catalogue record for this book is available from the British Library

ISBN 978-0-470-03310-4 (HB)

ISBN 978-0-470-03311-1 (PB)

Preface to Third Edition

Following the first studies of J.J. Thomson (1912), mass spectrometry has undergone countless improvements. Since 1958, gas chromatography coupled with mass spectrometry has revolutionized the analysis of volatile compounds. Another revolution occurred in the 1980s when the technique became available for the study of non-volatile compounds such as peptides, oligosaccharides, phospholipids, bile salts, etc. From the discoveries of electrospray and matrix-assisted laser desorption in the late 1980s, compounds with molecular masses exceeding several hundred thousands of daltons, such as synthetic polymers, proteins, glycans and polynucleotides, have been analysed by mass spectrometry.

From the time of the second edition published in 2001 until now, much progress has been achieved. Several techniques have been improved, others have almost disappeared. New atmospheric pressure desorption ionization sources have been discovered and made available commercially. One completely new instrument, the orbitrap, based on a new mass analyser, has been developed and is now also available commercially. Improved accuracy in low-mass determination, even at low resolution, improvements in sensitivity, better detection limits and more efficient tandem mass spectrometry even on high-molecular-mass compounds are some of the main achievements. We have done our best to include them is this new edition.

As the techniques continue to advance, the use of mass spectrometry continues to grow. Many new applications have been developed. The most impressive ones arise in system biology analysis.

Starting from the very foundations of mass spectrometry, this book presents all the important techniques developed up to today. It describes many analytical methods based on these techniques and emphasizes their usefulness by numerous examples. The reader will also find the necessary information for the interpretation of data. A series of graduated exercises allows the reader to check his or her understanding of the subject. Numerous references are given for those who wish to go deeper into some subjects. Important Internet addresses are also provided. We hope that this new edition will prove useful to students, teachers and researchers.

We would like to thank Professor Jean-Louis Habib Jiwan and Alexander Spote for their friendly hospitality and competent help.

We would also like to acknowledge the financial support of the FNRS (Fonds National de la Recherche Scientifique, Brussels).

Many colleagues and friends have read the manuscript and their comments have been very helpful. Some of them carried out a thorough reading. They deserve special mention: namely, Magda Claeys, Bruno Domon, Jean-Claude Tabet, and the late François Van Hoof. We also wish to acknowledge the remarkable work done by the scientific editors at John Wiley & Sons.

Many useful comments have been published on the first two editions, or sent to the editor or the authors. Those from Steen Ingemann were particularly detailed and constructive.

Finally, we would like to thank the Université Catholique de Louvain, the Ludwig Institute for Cancer Research and all our colleagues and friends whose help was invaluable to us.

Edmond de Hoffmann and Vincent Stroobant

Louvain-la-Neuve, March 2007

Introduction

Mass spectrometry’s characteristics have raised it to an outstanding position among analytical methods: unequalled sensitivity, detection limits, speed and diversity of its applications. In analytical chemistry, the most recent applications are mostly oriented towards biochemical problems, such as proteome, metabolome, high throughput in drug discovery and metabolism, and so on. Other analytical applications are routinely applied in pollution control, food control, forensic science, natural products or process monitoring. Other applications include atomic physics, reaction physics, reaction kinetics, geochronology, inorganic chemical analysis, ion–molecule reactions, determination of thermodynamic parameters (ΔGof, Ka, etc.), and many others.

Mass spectrometry has progressed extremely rapidly during the last decade, between 1995 and 2005. This progress has led to the advent of entirely new instruments. New atmospheric pressure sources were developed [1]–[4], existing analysers were perfected and new hybrid instruments were realized by new combinations of analysers. An analyser based on a new concept was described: namely, the orbitrap [5] presented in Chapter 2. This has led to the development of new applications. To give some examples, the first spectra of an intact virus [6] and of very large non-covalent complexes were obtained. New high-throughput mass spectrometry was developed to meet the needs of the proteomic [7, 8], metabolomic [9] and other ‘omics’.

Principles

The first step in the mass spectrometric analysis of compounds is the production of gas-phase ions of the compound, for example by electron ionization:

This molecular ion normally undergoes fragmentations. Because it is a radical cation with an odd number of electrons, it can fragment to give either a radical and an ion with an even number of electrons, or a molecule and a new radical cation. We stress the important difference between these two types of ions and the need to write them correctly:

These two types of ions have different chemical properties. Each primary product ion derived from the molecular ion can, in turn, undergo fragmentation, and so on. All these ions are separated in the mass spectrometer according to their mass-to-charge ratio, and are detected in proportion to their abundance. A mass spectrum of the molecule is thus produced. It provides this result as a plot of ion abundance versus mass-to-charge ratio. As illustrated in Figure 1, mass spectra can be presented as a bar graph or as a table. In either presentation, the most intense peak is called the base peak and is arbitrarily assigned the relative abundance of 100%. The abundances of all the other peaks are given their proportionate values, as percentages of the base peak. Many existing publications label the y axis of the mass spectrum as number of ions, ion counts or relative intensity. But the term relative abundance is better used to refer to the number of ions in the mass spectra.

Figure 1 Mass spectrum of methanol by electron ionization, presented as a graph and as a table.

Most of the positive ions have a charge corresponding to the loss of only one electron. For large molecules, multiply charged ions also can be obtained. Ions are separated and detected according to the mass-to-charge ratio. The total charge of the ions will be represented by q, the electron charge by e and the number of charges of the ions by z:

The x axis of the mass spectrum that represents the mass-to-charge ratio is commonly labelled m/z. When m is given as the relative mass and z as the charge number, both of which are unitless, m/z is used to denote a dimensionless quantity.

Ions provide information concerning the nature and the structure of their precursor molecule. In the spectrum of a pure compound, the molecular ion, if present, appears at the highest value of m/z (followed by ions containing heavier isotopes) and gives the molecular mass of the compound. The term molecular ion refers in chemistry to an ion corresponding to a complete molecule regarding occupied valences. This molecular ion appears at m/z 32 in the spectrum of methanol, where the peak at m/z 33 is due to the presence of the13C isotope, with an intensity that is 1.1% of that of the m/z 32 peak. In the same spectrum, the peak at m/z 15 indicates the presence of a methyl group. The difference between 32 and 15, that is 17, is characteristic of the loss of a neutral mass of 17 Da by the molecular ion and is typical of a hydroxyl group. In the same spectrum, the peak at m/z 16 could formally correspond to ions CH4•+, O+ or even CH3OH2+, because they all have m/z values equal to 16 at low resolution. However, O+ is unlikely to occur, and a doubly charged ion for such a small molecule is not stable enough to be observed.

The atomic mass units u or Da have the same fundamental definition:

However, they are traditionally used in different contexts: when dealing with mean isotopic masses, as generally used in stoichiometric calculations, Da will be preferred; in mass spectrometry, masses referring to the main isotope of each element are used and expressed in u.

There are different ways to define and thus to calculate the mass of an atom, molecule or ion. For stoichiometric calculations chemists use the average mass calculated using the atomic weight, which is the weighted average of the atomic masses of the different isotopes of each element in the molecule. In mass spectrometry, the nominal mass or the monoisotopic mass is generally used. The nominal mass is calculated using the mass of the predominant isotope of each element rounded to the nearest integer value that corresponds to the mass number, also called nucleon number. But the exact masses of isotopes are not exact whole numbers. They differ weakly from the summed mass values of their constituent particles that are protons, neutrons and electrons. These differences, which are called the mass defects, are equivalent to the binding energy that holds these particles together. Consequently, every isotope has a unique and characteristic mass defect. The monoisotopic mass, which takes into account these mass defects, is calculated by using the exact mass of the most abundant isotope for each constituent element.

For molecules of very high molecular weights, the differences between the different masses can become notable. Let us consider two examples.

In conclusion, the monoisotopic mass is used when it is possible experimentally to distinguish the isotopes, whereas the average mass is used when the isotopes are not distinguishable. The use of nominal mass is not recommended and should only be used for low-mass compounds containing only the elements C, H, N, O and S to avoid to making mistakes.

Diagram of a Mass Spectrometer

A mass spectrometer always contains the following elements, as illustrated in Figure 3: a sample inlet to introduce the compound that is analysed, for example a gas chromatograph or a direct insertion probe; an ionization source to produce ions from the sample; one or several mass analysers to separate the various ions; a detector to ‘count’ the ions emerging from the last analyser; and finally a data processing system that produces the mass spectrum in a suitable form. However, some mass spectrometers combine the sample inlet and the ionization source and others combine the mass analyser and the detector.

Figure 2 Mass spectra of isotopic patterns of two alkanes having the molecular formulae C20H42 and C100H202, respectively. The monoisotopic mass is the lighter mass of the isotopic pattern whereas the average mass, used by chemists in stoichiometric calculations, is the balanced mean value of all the observed masses.

A mass spectrometer should always perform the following processes:

1. Produce ions from the sample in the ionization source.

2. Separate these ions according to their mass-to-charge ratio in the mass analyser.

3. Eventually, fragment the selected ions and analyze the fragments in a second analyser.

4. Detect the ions emerging from the last analyser and measure their abundance with the detector that converts the ions into electrical signals.

5. Process the signals from the detector that are transmitted to the computer and control the instrument through feedback.

History

A large number of mass spectrometers have been developed according to this fundamental scheme since Thomson’s experiments in 1897. Listed here are some highlights of this development [11, 12]:

1886:

E. GOLDSTEIN discovers anode rays (positive gas-phase ions) in gas discharge [13].

1897:

J.J. THOMSON discovers the electron and determines its mass-to-charge ratio.

Nobel Prize in 1906.

Figure 3 Basic diagram for a mass spectrometer with two analysers and feedback control carried out by a data system.

1898:

W. WIEN analyses anode rays by magnetic deflection and then establishes that these rays carried a positive charge [14].

Nobel Prize in 1911.

1901:

W. KAUFMANN analyses cathodic rays using parallel electric and magnetic fields [15].

1909:

R.A. MILLIKAN and H. FLETCHER determine the elementary unit of charge.

1912:

J.J. THOMSON constructs the first mass spectrometer (then called a parabola spectrograph) [16]. He obtains mass spectra of O

2

, N

2

, CO, CO

2

and COCl

2

. He observes negative and multiply charged ions. He discovers metastable ions. In 1913, he discovers isotopes 20 and 22 of neon.

1918:

A.J. DEMPSTER develops the electron ionization source and the first spectrometer with a sector-shaped magnet (180°) with direction focusing [17].

1919:

F.W. ASTON develops the first mass spectrometer with velocity focusing [18].

Nobel Prize in 1922.

He measures mass defects in 1923 [19].

1932:

K.T. BAINBRIDGE proves the mass–energy equivalence postulated by Einstein [20].

1934:

R. CONRAD applies mass spectrometry to organic chemistry [21].

1934:

W.R. SMYTHE, L.H. RUMBAUGH and S.S. WEST succeed in the first preparative isotope separation [22].

1940:

A.O. NIER isolates uranium-235 [23].

1942:

The Consolidated Engineering Corporation builds the first commercial instrument dedicated to organic analysis for the Atlantic Refinery Company.

1945:

First recognition of the metastable peaks by J.A. HIPPLE and E.U. CONDON [24].

1948:

A.E. CAMERON and D.F. EGGERS publish design and mass spectra for a linear time-of-flight (LTOF) mass spectrometer [25]. W. STEPHENS proposed the concept of this analyser in 1946 [26].

1949:

H. SOMMER, H.A. THOMAS and J.A. HIPPLE describe the first application in mass spectrometry of ion cyclotron resonance (ICR) [27].

1952:

Theories of quasi-equilibrium (QET) [28] and RRKM [29] explain the monomolecular fragmentation of ions. R.A. MARCUS receives the

Nobel Prize in 1992.

1952:

E.G. JOHNSON and A.O. NIER develop double-focusing instruments [30].

1953:

W. PAUL and H.S. STEINWEDEL describe the quadrupole analyser and the ion trap or quistor in a patent [31]. W. PAUL, H.P. REINHARD and U. Von ZAHN, of Bonn University, describe the quadrupole spectrometer in

Zeitschrift für Physik

in 1958. PAUL and DEHMELT receive the

Nobel Prize in 1989

[32].

1955:

W.L. WILEY and I.H. McLAREN of Bendix Corporation make key advances in LTOF design [33].

1956:

J. BEYNON shows the analytical usefulness of high-resolution and exact mass determinations of the elementary composition of ions [34].

1956:

First spectrometers coupled with a gas chromatograph by F.W. McLAFFERTY [35] and R.S. GOHLKE [36].

1957:

Kratos introduces the first commercial mass spectrometer with double focusing.

1958:

Bendix introduces the first commercial LTOF instrument.

1966:

M.S.B. MUNSON and F.H. FIELD discover chemical ionization (CI) [37].

1967:

F.W. McLAFFERTY [38] and K.R. JENNINGS [39] introduce the collision-induced dissociation (CID) procedure.

1968:

Finnigan introduces the first commercial quadrupole mass spectrometer.

1968:

First mass spectrometers coupled with data processing units.

1969:

H.D. BECKEY demonstrates field desorption (FD) mass spectrometry of organic molecules [40].

1972:

V.I. KARATEV, B.A. MAMYRIM and D.V. SMIKK introduce the reflectron that corrects the kinetic energy distribution of the ions in a TOF mass spectrometer [41].

1973:

R.G. COOKS, J.H. BEYNON, R.M. CAPRIOLI and G.R. LESTER publish the book

Metastable Ions,

a landmark in tandem mass spectrometry [42].

1974:

E.C. HORNING, D.I. CARROLL, I. DZIDIC, K.D. HAEGELE, M.D. HORNING and R.N. STILLWELL discover atmospheric pressure chemical ionization (APCI) [43].

1974:

First spectrometers coupled with a high-performance liquid chromatograph by P.J. ARPINO, M.A. BALDWIN and F.W. McLAFFERTY [44].

1974:

M.D. COMISAROV and A.G. MARSHALL develop Fourier transformed ICR (FTICR) mass spectrometry [45].

1975:

First commercial gas chromatography/mass spectrometry (GC/MS) instruments with capillary columns.

1976:

R.D. MACFARLANE and D.F. TORGESSON introduce the plasma desorption (PD) source [46].

1977:

R.G. COOKS and T.L. KRUGER propose the kinetic method for thermochemical determination based on measurement of the rates of competitive fragmentations of cluster ions [47].

1978:

R.A. YOST and C.G. ENKE build the first triple quadrupole mass spectrometer, one of the most popular types of tandem instrument [48].

1978:

Introduction of lamellar and high-field magnets.

1980:

R.S. HOUK, V.A. FASSEL, G.D. FLESCH, A.L. GRAY and E. TAYLOR demonstrate the potential of inductively coupled plasma (ICP) mass spectrometry [49].

1981:

M. BARBER, R.S. BORDOLI, R.D. SEDGWICK and A.H. TYLER describe the fast atom bombardment (FAB) source [50].

1982:

First complete spectrum of insulin (5750 Da) by FAB [51] and PD [52].

1982:

Finnigan and Sciex introduce the first commercial triple quadrupole mass spectrometers.

1983:

C.R. BLAKNEY and M.L. VESTAL describe the thermospray (TSP) [53].

1983:

G.C. STAFFORD, P.E. KELLY, J.E. SYKA, W.E. REYNOLDS and J.F.J. TODD describe the development of a gas chromatography detector based on an ion trap and commercialized by Finnigan under the name Ion Trap [54].

1987:

M. GUILHAUS [55] and A.F. DODONOV [56] describe the orthogonal acceleration time-of-flight (oa-TOF) mass spectrometer. The concept of this technique was initially proposed in 1964 by G.J. O’Halloran of Bendix Corporation [57].

1987:

T. TANAKA [58] and M. KARAS, D. BACHMANN, U. BAHR and F. HILLENKAMP [59] discover matrix-assisted laser desorption/ionization (MALDI). TANAKA receives the

Nobel Prize in 2002.

1987:

R.D. SMITH describes the coupling of capillary electrophoresis (CE) with mass spectrometry [60].

1988:

J. FENN develops the electrospray (ESI) [61]. First spectra of proteins above 20 000 Da. He demonstrated the electrospray’s potential as a mass spectrometric technique for small molecules in 1984 [62]. The concept of this source was proposed in 1968 by M. DOLE [63]. FENN receives the

Nobel Prize in 2002.

1991:

V. KATTA and B.T. CHAIT [64] and B. GAMEN, Y.T. LI and J.D. HENION [65] demonstrate that specific non-covalent complexes could be detected by mass spectrometry.

1991:

B. SPENGLER, D. KIRSCH and R. KAUFMANN obtain structural information with reflectron TOF mass spectrometry (MALDI post-source decay) [66].

1993:

R.K. JULIAN and R.G. COOKS develop broadband excitation of ions using the stored-waveform inverse Fourier transform (SWIFT) [67].

1994:

M. WILM and M. MANN describe the nanoelectrospray source (then called microelectrospray source) [68].

1999:

A.A. MAKAROV describes a new type of mass analyser: the orbitrap. The orbitrap is a high-performance ion trap using an electrostatic quadro-logarithmic field [5, 69].

The progress of experimental methods and the refinements in instruments led to spectacular improvements in resolution, sensitivity, mass range and accuracy. Resolution (m/δm) developed as follows:

m/δm

1913

13

Thomson [16]

1918

100

Dempster [17]

1919

130

Aston [18]

1937

2000

Aston [70]

1998

8 000 000

Marshall and co-workers [71]

A continuous improvement has allowed analysis to reach detection limits at the pico-, femto- and attomole levels [72, 73]. Furthermore, the direct coupling of chromatographic techniques with mass spectrometry has improved these limits to the atto- and zeptomole levels [74, 75]. A sensitivity record obtained by mass spectrometry has been demonstrated by using modified desorption/ionization on silicon DIOS method to measure concentration of a peptide in solution. This technique has achieved a lower detection limit of 800 yoctomoles, which corresponds to about 480 molecules [76].

Regarding the mass range, DNA ions of 108 Da were weighed by mass spectrometry [77]. In the same way, non-covalent complexes with molecular weights up to 2.2 MDa were measured by mass spectrometry [78]. Intact viral particles of tobacco mosaic virus with a theoretical molecular weight of 40.5 MDa were analysed with an electrospray ionization charge detection time-of-flight mass spectrometer [6].

The mass accuracy indicates the deviation of the instrument’s response between the theoretical mass and the measured mass. It is usually expressed in parts per million (ppm) or in 10−3 u for a given mass. The limit of accuracy in mass spectrometry is about 1 ppm. The measurement of the atomic masses has reached an accuracy of better than 10−9 u [79].

In another field, Litherland et al. [80] succeeded in determining a14C/12C ratio of 1:1015 and hence in dating a 40 000-year-old sample with a 1% error. A quantity of 14C corresponding to only 106 atoms was able to be detected in less than 1 mg of carbon [81].

Ion Free Path

All mass spectrometers must function under high vacuum (low pressure). This is necessary to allow ions to reach the detector without undergoing collisions with other gaseous molecules. Indeed, collisions would produce a deviation of the trajectory and the ion would lose its charge against the walls of the instrument. On the other hand, ion–molecule collisions could produce unwanted reactions and hence increase the complexity of the spectrum. Nevertheless, we will see later that useful techniques use controlled collisions in specific regions of a spectrometer.

According to the kinetic theory of gases, the mean free path L (in m) is given by

(1)

(2)

(3)

Table 1 is a conversion table for pressure units. In a mass spectrometer, the mean free path should be at least 1 m and hence the maximum pressure should be 66 nbar. In instruments using a high-voltage source, the pressure must be further reduced to prevent the occurrence of discharges. In contrast, some trap-based instruments operate at higher pressure.

However, introducing a sample into a mass spectrometer requires the transfer of the sample at atmospheric pressure into a region of high vacuum without compromising the latter. In the same way, producing efficient ion–molecule collisions requires the mean free path to be reduced to around 0.1 mm, implying at least a 60 Pa pressure in a region of the spectrometer. These large differences in pressure are controlled with the help of an efficient pumping system using mechanical pumps in conjunction with turbomolecular, diffusion or cryogenic pumps. The mechanical pumps allow a vacuum of about 10−3 Torr to be obtained. Once this vacuum is achieved, the operation of the other pumping systems allows a vacuum as high as 10−10 Torr to be reached.

Table 1 Pressure units. The official SI unit is the pascal.

The sample must be introduced into the ionization source so that vacuum inside the instrument remains unchanged. Samples are often introduced without compromising the vacuum using direct infusion or direct insertion methods. For direct infusion, a capillary is employed to introduce the sample as a gas or a solution. For direct insertion, the sample is placed on a probe, a plate or a target that is then inserted into the source through a vacuum interlock. For the sources that work at atmospheric pressure and are known as atmospheric pressure ionization (API) sources, introduction of the sample is easy because the complicated procedure for sample introduction into the high vacuum of the mass spectrometer is removed.

References

1. Robb, D.B., Covey, T.R. and Bruins, A.P. (2000) Anal. Chem.,72, 3653.

2. Laiko, V.V., Baldwin, M.A. and Burlingame, A.L. (2000) Anal. Chem.,72, 652.

3. Takats, Z., Wiseman, J.M., Gologan, B. and Cooks, R.G. (2004) Mass spectrometry sampling under ambient conditions with desorption electrospray ionization. Science,5695, 471–3.

4. Cody, R.B., Laramée, J.A. and Durst, H.D. (2005) Versatile new ion source for analysis of materials in open air under ambient conditions. Anal. Chem,77, 2297–302.

5. Makarov, A. (2000) Electrostatic axially harmonic orbital trapping: a high performance technique of mass analysis. Anal. Chem.,72, 1156–62.

6. Fuerstenauw, S., Benner, W., Thomas, J. et al. (2001) Whole virus mass spectrometry. Angew. Chem. Int. Ed.,40, 541–4.

7. Reinders, J., Lewandrowski, U., Moebius, J. et al. (2004) Challenges in mass spectrometry-based proteomics. Proteomics,4, 3686–703.

8. Naylor, S. and Kumar, R. (2003) Emerging role of mass spectrometry in structural and functional proteomics. Adv. Protein Chem.,65, 217–48.

9. Villas-Boas, S.G., Mas, S., Akesson, M. et al. (2005) Mass spectrometry in metabolome analysis. Mass Spectrom. Rev.,24, 613–46.

10. Cooks, R.G. and Rockwood, A.L. (1991) The ‘Thomson’. A suggested unit for mass spectroscopists. Rapid Commun. Mass Spectrom.,5, 93.

11. Grayson, M.A. (2002) Measuring Mass: from Positive Rays to Proteins, Chemical Heritage Press, Philadelphia.

12. http://masspec.scripps.edu/MSHistory/mshisto.php (21 March 2007).

13. Goldstein, E. (1886) Berl. Ber.,39, 691.

14. Wien, W. (1898) Verhanal. Phys. Ges.,17.

15. Kaufmann, R.L., Heinen, H.J., Shurmann, L.W. and Wechsung, R.M. (1979) Microbeam Analysis (ed. D.E. Newburg), San Francisco Press, San Francisco, pp. 63–72.

16. Thomson, J.J. (1913) Rays of Positive Electricity and Their Application to Chemical Analysis, Longmans Green, London.

17. Dempster, A.J. (1918) Phys. Rev.,11, 316.

18. Aston, F.W. (1919) Philos. Mag.,38, 707.

19. Aston, F.W. (1942) Mass Spectra and Isotopes, 2nd edn, Edward Arnold, London.

20. Bainbridge, K.T. (1932) Phys. Rev.,42, 1; Bainbridge, K.T. and Jordan, E.B. (1936) Phys. Rev.,50, 282.

21. Conrad, R. (1934) Trans. Faraday Soc.,30, 215.

22. Smythe, W.R., Rumbaugh, L.H. and West, S.S. (1934) Phys. Rev.,45, 724.

23. Nier, A.O. (1940) Rev. Sci. Instrum.,11, 252.

24. Hipple, J.A. and Condon, E.U. (1946) Phys. Rev.,69, 347.

25. Cameron, A.E. and Eggers, D.F. (1948) Rev. Sci. Instrum.,19, 605.

26. Stephens, W. (1946) Phys. Rev.,69, 691.

27. Sommer, H., Thomas, H.A. and Hipple, J.A. (1949) Phys. Rev.,76, 1877.

28. Rosenstock, H.M., Wallenstein, M.B., Warhaftig, A.L. and Eyring, H. (1952) Proc. Natl. Acad. Sci. USA.,38, 667.

29. Marcus, R.A. (1952) J. Chem. Phys.,20, 359.

30. Johnson, E.G. and Nier, A.O. (1953) Phys. Rev.,91, 12.

31. Paul, W. and Steinwedel, H.S. (1953) Z. Naturforsch.,8a, 448.

32. Paul, W., Reinhard, H.P. and von Zahn, U. (1958) Z. Phys.,152, 143.

33. Wiley, W.L. and McLaren, I.H. (1955) Rev. Sci. Instrum.,16, 1150.

34. Beynon, J. (1956) Mikrochim. Acta, 437.

35. McLafferty, F.W. (1957) Appl. Spectrosc.,11, 148.

36. Gohlke, R.S. (1959) Anal. Chem.,31, 535.

37. Munson, M.S.B. and Field, F.H. (1966) J. Am. Chem. Soc.,88, 2681.

38. McLafferty, F.W. and Bryce, T.A. (1967) Chem. Commun., 1215.

39. Jennings, K.R. (1968) Int. J. Mass Spectrom. Ion Phys.,1, 227.

40. Beckey, H.D. (1969) Int. J. Mass Spectrom. Ion Phys.,2, 500.

41. Karataev, V.I., Mamyrin, B.A. and Smikk, D.V. (1972) Sov. Phys.–Tech. Phys.,16, 1177.

42. Cooks, R.G., Beynon, J.H., Caprioli, R.M. and Lester, G.R. (1973) Metastable Ions, Elsevier, New York, p. 296.

43. Horning, E.C., Carroll, D.I., Dzidic, I. et al. (1974) J. Chromatogr. Sci.,412, 725.

44. Arpino, P.J., Baldwin, M.A. and McLafferty, F.W. (1974) Biomed. Mass Spectrom., 1, 80.

45. Comisarov, M.B. and Marshall, A.G. (1974) Chem. Phys. Lett.,25, 282.

46. Macfarlane, R.D. and Torgesson, D.F. (1976) Science,191, 920.

47. Cooks, R.G. and Kruger, T.L. (1977) J. Am. Chem. Soc.,99, 1279.

48. Yost, R.A. and Enke, C.G. (1978) J. Am. Chem. Soc.,100, 2274.

49. Houk, R.S., Fassel, V.A., Flesch, G.D. et al. (1980) Anal. Chem.,52, 2283.

50. Barber, M., Bardoli, R.S., Sedgwick, R.D. and Tyler, A.H. (1981) J. Chem. Soc., Chem. Commun., 325.

51. McFarlane, R. (1982) J. Am. Chem. Soc.,104, 2948.

52. Barber, M. et al. (1982) J. Chem. Soc., Chem. Commun., 936.

53. Blakney, C.R. and Vestal, M.L. (1983) Anal. Chem.,55, 750.

54. Stafford, G.C., Kelley, P.E., Syka, J.E. et al. (1984) Int. J. Mass Spectrom. Ion Processes,60, 85.

55. Dawson, J.H.J. and Guilhaus, M. (1989) Rapid Commun. Mass Spectrom., 3, 155–9.

56. Dovonof, A.F., Chernushevich, I.V. and Laiko, V.V. (1991) Proceedings of the 12th International Mass Spectrometry Conference, 26–30 August, Amsterdam, the Netherlands, p. 153.

57. O’Halloran, G.J., Fluegge, R.A., Betts, J.F. and Everett, W.L. (1964) Technical Documentary Report No. ASD-TDR-62-644, The Bendix Corporation, Research Laboratory Division, Southfield, MI.

58. Tanaka, T., Waki, H., Ido, Y. et al. (1988) Rapid Commun. Mass Spectrom.,2, 151.

59. Karas, M., Bachmann, D., Bahr, U. and Hillenkamp, F. (1987) Int. J. Mass Spectrom. Ion Processes,78, 53.

60. Olivares, J.A., Nguyen, N.T., Yonker, C.R. and Smith, R.D. (1987) Anal. Chem.,59, 1230.

61. Fenn, J.B., Mann, M., Meng, C.K. et al. (1989) Science,246, 64.

62. Yamashita, M. and Fenn, J.B. (1984) J. Chem. Phys.,80, 4451.

63. Dole, M., Mach, L.L., Hines, R.L. et al. (1968) J. Chem. Phys.,49, 2240.

64. Katta, V. and Chait, B.T. (1991) J. Am. Chem. Soc.,113, 8534.

65. Gamen, B., Li, Y.T. and Henion, J.D. (1991) J. Am. Chem. Soc.,113, 6294.

66. Spengler, B., Kirsch, D. and Kaufmann, R. (1991) Rapid Commun. Mass Spectrom.,5, 198.

67. Julian, R.K. and Cooks, R.G. (1993) Anal. Chem.,65, 1827.

68. Wilm, M. and Mann, M. (1994) Proceedings of the 42nd ASMS Conference, Chicago, IL, p. 770.

69. Makarov, A. (2000) Electrostatic axially harmonic orbital trapping: a high performance technique of mass analysis. Anal. Chem.,72, 1156–62.

70. Aston, F.W. (1937) Proc. R. Soc., Ser. A, 163, 391.

71. Shi, S.D.H., Hendrickson, C.L. and Marshall, A.G. (1998) Counting individual sulfur atoms in a protein by ultrahigh-resolution Fourier transform ion cyclotron resonance mass spectrometry: experimental resolution of isotopic fine structure in proteins. Proc. Natl. Acad. Sci. USA,95, 11532–7.

72. Solouki, T., Marto, J.A., White, F.M. et al. (1995) Attomole biomolecule mass analysis by matrix-assisted-laser-desorption ionization Fourier-transform ion-cyclotron resonance. Anal. Chem.,67, 4139–44.

73. Morris, H.R., Paxton, T., Panico, M. et al. (1997) A novel geometry mass spectrometer, the Q-TOF, for low-femtomole/attomole-range biopolymer sequencing. J. Protein Chem.,16, 469–79.

74. Valaskovic, G.A., Kelleher, N.L. and McLafferty, F.W. (1996) Attomole protein characterization by capillary electrophoresis mass spectrometry. Science,273, 1199–1200.

75. Wang, P.G., Murugaiah, V., Yeung, B. Vouros, P. and Giese, R.W. (1996) 2-Phosphoglycolate and glycolate-electrophore detection, including detection of 87 zeptomoles of the latter by gas chromatography electron-capture mass spectrometry. J. Chromatogr. A, 721, 289–96.

76. Trauger, S.A., Go, E.P., Shen, Z. et al. (2004) High sensitivity and analyte capture with desorption/ionization mass spectrometry on silylated porous silicon. Anal. Chem.,76, 4484–9.

77. Chen, R., Cheng, X, Mitchell, D. et al. (1995) Anal. Chem.,67, 1159.

78. Sanglier, S., Leize, E., Van Dorsselaer, A. and Zal, F. (2003) J. Am. Soc. Mass Spectrom.,14, 419.

79. Di Fillip, F., Natarajan, V., Bradley, M. et al. (1995) Phys. Scr.,T59, 144.

80. Litherland, A.E., Benkens, R.P., Lilius, L.R. et al. (1981) Nucl. Instrum. Methods,186, 463.

81. Vogel, J.S., Trteltaub, K.W., Finkel, R. and Nelson, D.E. (1995) Anal. Chem.,67, 353A.

1

Ion Sources

In the ion sources, the analysed samples are ionized prior to analysis in the mass spectrometer. A variety of ionization techniques are used for mass spectrometry. The most important considerations are the internal energy transferred during the ionization process and the physico-chemical properties of the analyte that can be ionized. Some ionization techniques are very energetic and cause extensive fragmentation. Other techniques are softer and only produce ions of the molecular species. Electron ionization, chemical ionization and field ionization are only suitable for gas-phase ionization and thus their use is limited to compounds sufficiently volatile and thermally stable. However, a large number of compounds are thermally labile or do not have sufficient vapour pressure. Molecules of these compounds must be directly extracted from the condensed to the gas phase.

These direct ion sources exist under two types: liquid-phase ion sources and solid-state ion sources. In liquid-phase ion sources the analyte is in solution. This solution is introduced, by nebulization, as droplets into the source where ions are produced at atmospheric pressure and focused into the mass spectrometer through some vacuum pumping stages. Electrospray, atmospheric pressure chemical ionization and atmospheric pressure photoionization sources correspond to this type. In solid-state ion sources, the analyte is in an involatile deposit. It is obtained by various preparation methods which frequently involve the introduction of a matrix that can be either a solid or a viscous fluid. This deposit is then irradiated by energetic particles or photons that desorb ions near the surface of the deposit. These ions can be extracted by an electric field and focused towards the analyser. Matrix-assisted laser desorption, secondary ion mass spectrometry, plasma desorption and field desorption sources all use this strategy to produce ions. Fast atom bombardment uses an involatile liquid matrix.

The ion sources produce ions mainly by ionizing a neutral molecule in the gas phase through electron ejection, electron capture, protonation, deprotonation, adduct formation or by the transfer of a charged species from a condensed phase to the gas phase. Ion production often implies gas-phase ion–molecule reactions. A brief description of such reactions is given at the end of the chapter.