Analytical Chemistry
Section E.14
Mass Spectrometer
<Instant Notes, D. Kealey & P.J. Haines>
 Contents
1. Basic of Mass Spectrometer
2. Advanced Mass Spectrometer
3. Conclusion
1. Basic of Mass Spectrometer

Terms of Mass Spectrometer

Principles

Mass Spectrometry

Ionization Techniques

Fragmentation

Mass Analyzer

Resolution

Isotope Peaks
 Terms of Mass Spectrometer (1)
Principles
Mass Spectrometry (MS) is a technique whereby materials are ionized and
dissociated into fragments characteristic of the molecule(s) or element(s) present in
the sample. The numbers of ions of each mass provide information for qualitative
and quantitative analysis.
Mass Spectrometric
A Mass Spectrometer, which is operated under high vacuum, incorporates a sample
inlet and ion source, a mass analyzer, an ion detector and a data processing system.
Ionization Techniques
Alternative ionization techniques are available differing in energy and applicability.
Some produce a high degree of dissociation of molecules, while others are used
primarily to establish an accurate relative molecular mass of a compound or to
facilitate elemental analysis.
 Terms of Mass Spectrometer (2)
Fragmentation
After ionization, molecules may dissociate into fragments of smaller mass, some
carrying a charge. The presence and relative abundances of the various charged
fragments provide structural information and enable unknown compounds to be
identified.
Isotope peaks
These are peaks in a mas spectrum arising from fragments containing naturally
occurring heavier isotopes of one or more elements.
Mass Spectra
Spectral data is either tabulated or shown graphically as a plot of the numbers of
ions of each mass detected. For ease of interpretation, these are presented as line
diagrams.
Related Topics : Inductively coupled plasma spectrometry (E5)
Combined techniques (Section F)
 Principles
- MS is an analytical technique in which gaseous ions formed
from the molecules or atoms of a sample are separated in
space or time and detected according to their mass-to-charge
ratio, m/z.
- Example of Mass Spectrum (m/z 31 for Methanol)
Base Peak
: the most
abundant ion
Fig. 1. Mass Spectrum of Methanol
 Mass Spectrometry
Fragment(in MSn)
- CID
- ETD
- EI, CI
- ESI
- MALDI
Fig. 2. Block diagram of a Mass Spectrometer
-
Single Focusing
Double Focusing
Quadrupole & Quadrupole Ion Trap
TOF
FT-ICR
Tandem (MS/MS)
 Ionization Techniques (1)
(70eV)
1. EI (Electron Ionization)
- EI Process (in gas phase)
M + e- → M+• + 2e(M is the analyte molecule being
ionized, e- is the electron and M+•
is the resulting ion.)
- Only possible in gas phase.
Fig. 3. Diagram of EI
 Ionization Techniques (2)
2. CI (Chemical Ionization)
- Collision of the analyte with
ions of a reagent gas.
- Reagent gas : Methane,
Ammonia, Isobutane
- Advantage to analyze mixture
compounds
- Variations : NCI(Negative),
APCI(Atmospheric Pressure)
Fig. 4. Mechanism of CI
 Ionization Techniques (3)
Fig. 4-1. Diagram of ESI
Fig. 4-2. Two processes of the conversion
of ions from droplets into the gas phase
(a) Charge Residue Model (b) Ion
Desorption Model
3. ESI (Electrospray Ionization)
-
Using the solvents. (water + Organic solvents + Acid)
Possible to analyze biological molecules and Polymers.
Using the nebulizer gas (inert). → rapid evaporation of solvents.
Being produced multiply charged ions.
 Ionization Techniques (4)
4. MALDI (Matrix-Assisted
Laser Desorption Ionization)
- Sample is mixed with a
compound capable of absorbing
energy from the laser.
⇒Analyte/Matrix Mixture
- Possible to analyze solid phase
samples.
- Soft ionization technique.
- Being produced proton ions.
Fig. 5. Diagram of MALDI
- Possible to use Genomics,
Proteomics.
 Fragmentation (1)
Fig. 6. Diagram of Fragmentation of peptides
- The backbone of a peptide can fragment at three bonds CH-CO, CONH and NH-CH with each dissociation producing two fragments named
according to the location of the charge and the amino acid position
(n-terminus = a-, b-, c-; c-terminus = x-, y-, z-)
- Using in a Tandem MS (MSn)
 Fragmentation (2)
Fig. 7. Diagram of comparison between CID & ETD
CID (Collision-Induced Dissociation)
ETD (Electron-Transfer Dissociation)
- Breaking the weakest bonds and
producing a characteristic series of
fragments.
- ETD cleaves selectively on the
peptide backbone, leaving PTMs
intact.
- ETD produces a different set of
fragments that are complementary to
CID, so sequence coverage is more
complete.
- Many PTMs are fragile and are lost
in the CID process.
 Mass Analyzer (1)
Fig. 8-1. Diagram of a Principle of Single
Focusing Magnetic Mass Analyzer
Fig. 8-2. Diagram of a principle Double
Focusing Magnetic Mass Analyzer
- Related Equations
𝒎
𝐁𝟐 𝐫 𝟐
=
𝒛
𝟐𝐕
(r : radii of curvature, B : magnetic field strength, V : accelerating
voltage)
 Mass Analyzer (2)
Fig. 9-1. Diagram of a Principle
Quadrupole Mass Analyzer
Fig. 9-2. Diagram of a Principle
Quadrupole Ion Trap Mass Analyzer
- The same principle both Quadrupole and Quadrupole Ion Trap
- Advantages of Quadrupole Ion Trap
→ High Sensitivity, Trapping the specific ions, Specialized for
Qualitative Analysis.
 Mass Analyzer (3)
- Linear-TOF
- Reflector-TOF
• High Sensitivity • Low Sensitivity
• Low Resolution • High Resolution
- The analytical technique has been
extremely useful for proteomics
using MALDI-TOF/MS systems.
Fig. 10. Diagram of comparison of Linear
and Reflector
 Mass Analyzer (4)
Fig. 11-1. Diagram of FT-ICR
Fig. 11-2. Diagram of a Principle
of Ion Cyclotron Resonance
- The excited ions pass a set of metal detector plates with each orbit.
- Very Strong Magnetic Field : 5~12 Tesla
- The image current is recorded and Fourier Transformed to produce
the mass spectrum.
- Extremely High Price, Vacuum needs, Resolution and Mass Accuracy
 Mass Analyzer (5)
Fig. 12. Diagram of a Principle of Tandem MS
- Hyphenated techniques (MS-MS)
- Tandem MS modes
→ Precursor Ion Scan, Product Ion Scan, Neutral Loss Scan,
Selected Reaction Monitoring
- High Resolution, Selectivity
 Resolution
- MS are designed to give a specified Resolving Power.
→ The minimum acceptable Resolution = One mass unit.
- Two masses are considered to be resolved when the valley
between their peaks is less than 10% of the smaller peak height.
m1
m2
m2
Resolving Power =
m2 − m1
- Requirement for unit mass resolution increase with the
magnitudes of m1 and m2.
 Isotope Peaks (1)
- Most elements occur naturally as a mixture of isotopes, all of
which contribute to peaks in a mass spectrum.
- Isotope Peaks are of importance in the interpretation of mass
spectra.
Table. 1. Empirical formulae and isotope peak ratios for a nominal RMM
value of 70 (M=100%)
 Isotope Peaks (2)
- The intensities in mass spectra of Isotope Peaks of C24H22O7
(Using the Table. 2.)
(M)+
(M+1)+, (M+2)+
Table. 2. Natural isotopic abundances of some common elements as a
percentage of the most abundant isotope
2. Advanced Mass Spectrometer

EI/MS
Agilent 7000 Series Triple Quadrupole GC/MS

ESI/ETD/MS
Thermo Orbitrap Elite

MALDI/MS
Waters MALDI SYNAPT G2-S HDMS
 EI/MS
- Manufacturer
Agilent
- Model
7000 Series Triple Quadrupole
GC/MS
- Ionization Type
EI, PCI, NCI
- Resolution
0.7 to 2.5 Da
- Scanning Speed
Up to 6250 u/s
- Mass Range
Fig. 13. Agilent 7000 Series Triple Quadrupole
GC/MS
1.2 to 1050 m/z
 EI/MS - Characteristics
Fig. 14. Diagram of a Principle of Agilent 7000 Series Triple Quadrupole GC/MS
• Characteristics Video
- GC/MS/MS
- Gold Quadrupole
- Hexapole Collision Cell
- Triple-Axis HED-EM Detector
 EI/MS – Agilent Techniques (1)
Fig. 15-2. Gold Plated Hyperbolic
Quartz Quadrupole
Fig. 15-1. Diagram of Agilent 7000
Series Triple Quadrupole GC/MS
- Heated gold plated hyperbolic
quartz quadrupoles
- Reliability, Stability
 EI/MS – Agilent Techniques (2)
Fig. 16-2. Diagram of a Principle
of Helium Quenching
Fig. 16-1. Diagram of Agilent 7000
Series Triple Quadrupole GC/MS
- Using a Helium buffer gas
- Reduction of Chemical Noise
- High Sensitivity, Resolution
 EI/MS – Agilent Techniques (3)
Fig. 17-2. Diagram of a Principle
of Triple-Axis Detector
Fig. 17-1. Diagram of Agilent 7000
Series Triple Quadrupole GC/MS
– Ultra low neutrals noise
– Long life and high linearity
– Superior sensitivity
 ESI/ETD/MS
- Manufacturer
Thermo Scientific
- Model
Orbitrap Elite
- Ionization Type
ESI
- Fragmentation Type
CID, ETD, HCD
- Resolution
>240,000 at m/z 400
- Mass Accuracy
< 3 ppm with external calibration
Fig. 18. Thermo Orbitrap Elite
< 1 ppm with internal calibration
 ESI/ETD/MS - Characteristics
Fig. 19. Diagram of a Principle of Thermo Orbitrap Elite
• Characteristics Video
- Ion Optics
- Ion Trap with Neutral Blocker
- Trap-HCD
- Orbitrap
 ESI/ETD/MS – Thermo Techniques (1)
Fig. 20-1. Diagram of Thermo Orbitrap
Elite
- Variable Spaced Stacked Lenses.
→ Increasing spacing = increasing
field penetration to focus ion beam
Fig. 20-2. Ion Optics
- Robustness, High Sensitivity
 ESI/ETD/MS – Thermo Techniques (2)
Fig. 21-1. Diagram of Thermo Orbitrap
Elite
- Rotated 45o Quadrupole
- Blocking the Neutral Beams.
- Separation of Neutrals and Ions
Fig. 21-2. Ion Trap (Square
Quadrupole with Neutral Blocker)
- More Robustness
 ESI/ETD/MS – Thermo Techniques (3)
Fig. 22-1. Diagram of Thermo Orbitrap
Elite
Fig. 22-2. Diagram of Trap-HCD
- Trap-HCD fragmentation (HCD,CID,
PQD,ETD)
- Dual Pressure Trap
- No low mass cut off
- High Resolution
 ESI/ETD/MS – Thermo Techniques (4)
Fig. 23-1. Diagram of Thermo Orbitrap
Elite
- New type of Ion Trap
- Faster Scanning
- High Resolution
Fig. 23-2. Diagram of Orbitrap
 MALDI/MS
- Manufacturer
Waters
- Model
MALDI SYNAPT G2-S HDMS
- Ionization Type
MALDI, ESI, APPI, APCI, ESCi®
- Fragmentation Type
CID, ETD
- Resolution
> 40,000 FWHM
- Mass Range
Max. 100,000 m/z
Fig. 24. Waters MALDI SYNAPT G2-S
HDMS
 MALDI/MS - Characteristics
Fig. 25. Diagram of a Principle of Waters
MALDI SYNAPT G2-S HDMS
• Characteristics Video
- Variable Ionization Techniques
- T-Wave Ion Guide
- TRIWAVE
- QUANTOF
- HDMS™ instruments
 MALDI/MS – Waters Techniques (1)
Fig. 26. Diagram of Ion Source of Waters
MALDI SYNAPT G2-S HDMS
-Very simple to exchange Ion Source
-Extend Compound Coverage
-High Flexibility
 MALDI/MS – Waters Techniques (2)
Fig. 27-1. Diagram of Waters MALDI
SYNAPT G2-S HDMS
- Positive and Negative RF fields are
applied to each ring electrode pair.
- New type of Ion Optics
Fig. 27-2. Diagram of T-Wave
Ion Guide
- Outstanding linearity, Sensitivity
 MALDI/MS – Waters Techniques (3)
Fig. 28-1. Diagram of Waters MALDI
SYNAPT G2-S HDMS
- Ion Mobility Separation
→ Being separated by Size,
Shape and Charge
- Fragmentation(CID, ETD)
Fig. 28-2. Diagram of TRIWAVE
- Increasing Peak Capacity and
Detection limit
 MALDI/MS – Waters Techniques (3)
Fig. 29. Diagram of Waters MALDI SYNAPT
G2-S HDMS
-Dual Stage Reflectron
-Hybrid Ion Detection System
-Compatible with HDMS™ analysis
-High Resolution : Over 40,000 FWHM
3. Conclusion


Summary
Reference
 Summary
-
Basic principle of Mass Spectrometer
Ionization, Fragmentation
Several Types of Mass Analyzer
Identification of Mass Spectra
Application
⇒ Future Works : Advanced Hybrid Mass Spectrometer
Contributing to analyze and interpret
Biological Molecules in Proteomics quickly and
accurately.
 Reference
- Agilent Technologies
http://www.chem.agilent.com/en-US/Products/Instruments/ms/gcms/systems/7000triplequadrupolegcms/pages/default.aspx
- Thermo Scientific
http://www.thermoscientific.com/ecomm/servlet/productsdetail_1115
2_L10710_87170_13901130_-1
- Waters
http://www.waters.com/waters/nav.htm?cid=134614100
- Instant Notes, Analytical Chemistry, Kealey & Haines