Summary Mass Spectrometry
By Jeanine Koelewijn – 2022
,Inhoudsopgave
Les 1: Mass spectrometry principles – Gary ...................................................................................................... 3
1.1 Intro mass spectrometry ............................................................................................................................... 3
1.2 What’s in a mass spec .................................................................................................................................. 5
Les 2: Electron and Chemical ionization ............................................................................................................ 8
2.1 Electron Ionization (EI).................................................................................................................................. 9
2.2 The molecular ion ....................................................................................................................................... 12
2.3 Isotope peaks .............................................................................................................................................. 12
2.4 Chemical Ionization (CI) .............................................................................................................................. 14
Les 3: MALDI and TOF..................................................................................................................................... 17
3.1 MALDI ......................................................................................................................................................... 18
3.2ToF – Time Of Flight..................................................................................................................................... 19
3.3 MALDI – MS imaging .................................................................................................................................. 21
Les 4: ESI, ion traps, quads ............................................................................................................................. 22
4.1 ESI – electrospray ionization ....................................................................................................................... 22
4.2 Iontraps ...................................................................................................................................................... 25
4.3 Quadrupoles ............................................................................................................................................... 27
Les 5: Scanning modes, protein identification, hybrid analysers..................................................................... 29
5.1 Scanning modes.......................................................................................................................................... 29
5.2 Protein Identification by MS ....................................................................................................................... 32
5.3 Hybrid Analysers ......................................................................................................................................... 34
Les 6: Fragmentation ...................................................................................................................................... 38
Les 7: LCMS – hyphenation ............................................................................................................................. 44
Les 8: Orbitrap & Ambient MS ........................................................................................................................ 49
8.1 Orbitrap ...................................................................................................................................................... 49
8.2 Ambient MS – Ambient ionization .............................................................................................................. 51
Les 9: Protein identification & Quantitation ................................................................................................... 56
9.1 Quantitative proteomics ............................................................................................................................ 56
9.2 Protein identification .................................................................................................................................. 59
Les 10: MS in life science ................................................................................................................................ 61
2
,Samenvatting -- Mass Spectrometry 2022/2023
Jeanine Koelewijn
Les 1: Mass spectrometry principles – Gary
1.1 Intro mass spectrometry
What is mass spectrometry
- Mass spectrometry is an analytical tool useful for measuring the mass-to-charge ratio
(m/z) of one or more molecules present in a sample
- These measurements can often be used to calculate the exact molecular weight of
the sample components as well
MOST often used for:
• Identify via molecular weight determination (match to known mass)
• To quantify known compounds
• Determine structure and chemical properties of molecules
HOW:
we get multiple levels of information
1. Molecular weight → molecular mass
2. Elemental composition → isotope pattern of atoms in molecules
3. Structural information → tandem mass spectrometry
IONISATION
Gas phase (for these the compounds need to be naturally volatile and thermally stable)
- Electron (EI)
- Chemical (CI)
Desorption phase (when the compounds are non-volatile and thermally non-stable)
- Electrospray (ESI)
- Matrix assisted laser desorption/ionization (MALDI)
MASS ANALYSERS
- Time of flight (TOF)
- Double focusing
- Quadrupoles (Q)
- Iontrap (IT)
- Fourier-transform ion cyclotron resonance (FTICR)
3
, - Orbitrap (Orbi)
- Qtof
- QIT/ORbi
All work on the same principle of ion movement and manipulation
F=m*a
F: magnetic or electric field to exert a force on an ion
a: this force accelerates ions differently
m: depending on the mass of the ion
Grams are not a good unit of mass for something this small
1 Da = 1.660538921 x 10-27 kg
1 Da = one twelfth (1/12) of the mass of an unbound neutral atom of carbon 12
The analyte must be charged (an ion)!
- We always measure m/q or m/z
- Mass of an ion differs from that of a neutral
q = -+ ze
q = the absolute/electric charge on an ion (SI unit:coulombs)
z = the unit charge (where q = z x e , the number of electrons/protons gained or lost, 1 2 3 4)
acceleration of ions VS deflection of ions
Acceleration:
Consider an ion of mass m and charge, q, in a potential difference of V.
The ion will accelerate through the potential gaining velocity.
The kinetic energy gained (1/2 mv2) = potential energy (qV)
4
By Jeanine Koelewijn – 2022
,Inhoudsopgave
Les 1: Mass spectrometry principles – Gary ...................................................................................................... 3
1.1 Intro mass spectrometry ............................................................................................................................... 3
1.2 What’s in a mass spec .................................................................................................................................. 5
Les 2: Electron and Chemical ionization ............................................................................................................ 8
2.1 Electron Ionization (EI).................................................................................................................................. 9
2.2 The molecular ion ....................................................................................................................................... 12
2.3 Isotope peaks .............................................................................................................................................. 12
2.4 Chemical Ionization (CI) .............................................................................................................................. 14
Les 3: MALDI and TOF..................................................................................................................................... 17
3.1 MALDI ......................................................................................................................................................... 18
3.2ToF – Time Of Flight..................................................................................................................................... 19
3.3 MALDI – MS imaging .................................................................................................................................. 21
Les 4: ESI, ion traps, quads ............................................................................................................................. 22
4.1 ESI – electrospray ionization ....................................................................................................................... 22
4.2 Iontraps ...................................................................................................................................................... 25
4.3 Quadrupoles ............................................................................................................................................... 27
Les 5: Scanning modes, protein identification, hybrid analysers..................................................................... 29
5.1 Scanning modes.......................................................................................................................................... 29
5.2 Protein Identification by MS ....................................................................................................................... 32
5.3 Hybrid Analysers ......................................................................................................................................... 34
Les 6: Fragmentation ...................................................................................................................................... 38
Les 7: LCMS – hyphenation ............................................................................................................................. 44
Les 8: Orbitrap & Ambient MS ........................................................................................................................ 49
8.1 Orbitrap ...................................................................................................................................................... 49
8.2 Ambient MS – Ambient ionization .............................................................................................................. 51
Les 9: Protein identification & Quantitation ................................................................................................... 56
9.1 Quantitative proteomics ............................................................................................................................ 56
9.2 Protein identification .................................................................................................................................. 59
Les 10: MS in life science ................................................................................................................................ 61
2
,Samenvatting -- Mass Spectrometry 2022/2023
Jeanine Koelewijn
Les 1: Mass spectrometry principles – Gary
1.1 Intro mass spectrometry
What is mass spectrometry
- Mass spectrometry is an analytical tool useful for measuring the mass-to-charge ratio
(m/z) of one or more molecules present in a sample
- These measurements can often be used to calculate the exact molecular weight of
the sample components as well
MOST often used for:
• Identify via molecular weight determination (match to known mass)
• To quantify known compounds
• Determine structure and chemical properties of molecules
HOW:
we get multiple levels of information
1. Molecular weight → molecular mass
2. Elemental composition → isotope pattern of atoms in molecules
3. Structural information → tandem mass spectrometry
IONISATION
Gas phase (for these the compounds need to be naturally volatile and thermally stable)
- Electron (EI)
- Chemical (CI)
Desorption phase (when the compounds are non-volatile and thermally non-stable)
- Electrospray (ESI)
- Matrix assisted laser desorption/ionization (MALDI)
MASS ANALYSERS
- Time of flight (TOF)
- Double focusing
- Quadrupoles (Q)
- Iontrap (IT)
- Fourier-transform ion cyclotron resonance (FTICR)
3
, - Orbitrap (Orbi)
- Qtof
- QIT/ORbi
All work on the same principle of ion movement and manipulation
F=m*a
F: magnetic or electric field to exert a force on an ion
a: this force accelerates ions differently
m: depending on the mass of the ion
Grams are not a good unit of mass for something this small
1 Da = 1.660538921 x 10-27 kg
1 Da = one twelfth (1/12) of the mass of an unbound neutral atom of carbon 12
The analyte must be charged (an ion)!
- We always measure m/q or m/z
- Mass of an ion differs from that of a neutral
q = -+ ze
q = the absolute/electric charge on an ion (SI unit:coulombs)
z = the unit charge (where q = z x e , the number of electrons/protons gained or lost, 1 2 3 4)
acceleration of ions VS deflection of ions
Acceleration:
Consider an ion of mass m and charge, q, in a potential difference of V.
The ion will accelerate through the potential gaining velocity.
The kinetic energy gained (1/2 mv2) = potential energy (qV)
4
