ionization methods in mass · pdf fileionization methods in mass spectrometry at the scs mass...

Ionization Methodsin Mass Spectrometryat the SCS Mass Spectrometry Laboratory

Steven L. Mullen, Ph.D.Associate Director

School of Chemical Sciences, University of Illinois

SCS Mass Spectrometry Laboratory

Contact Information31 Noyes Laboratory (8:00-5:00 M-F)333-2545www.scs.uiuc.edu/[email protected]



StaffVera Mainz, Ph.D. (Acting Director)Steven L. Mullen, Ph.D. (Associate Director)Furong SunRachel TaylorRichard Milberg, Ph.D.



InstrumentationMicromass 70-VSE (3)Micromass 70-SE-4FMicromass QuattroMicromass Q-Tof UltimaApplied Biosystems Voyager DE-STRFinnigan LCQ Deca XP



70-VSE (A) 70-VSE (B)

70-VSE (C) 70-SE-4F



Quattro Q-Tof

Voyager LCQ Deca


Information from Mass Spectrometry

1. Molecular WeightFrom molecular ion

M+ or (M+H)+ [also (M+Na)+ or (M+K)+]

2. StructureFrom fragments

Hard ionizationMS/MS

3. Elemental CompositionFrom exact mass


Parts of a Mass Spectrometer

Inlet Source Analyzer Detector

EICIFI

FDFAB

MALDIESI

ProbeLeak Valve

SyringePlateGCLC

SectorQuadrupole

FTTOF

Ion Trap

Electron MultiplierPhotomultiplierArray Detector


Parts of a Mass Spectrometer

Inlet

Source

Analyzer

Detector


How a Mass Spectrometer Works

Source Analyzer Detector


Interpreting a Mass Spectrum

15

43

58

Acetone

CH3CH3

O 1543

MW=58


Isotopic Peaks

C6H6

C6H5Cl

C6H5Br

35Cl

37Cl

79Br 81Br

12C

13C


Resolution

R=1000

R=500

R=100

555.3

555.3

555.5

Leu-enkephalin Monoisotopic Mass = 555.3 Average Mass = 555.6


ChargeLeu-enkephalin Monoisotopic Mass = 555.3

555.3

556.3

557.3

277.6

278.6

278.1

+1 ionm/z = 555.3

+2 ionm/z = 277.6


HR Exact Mass Measurement

OH

HO

OCH3

O

OH

HO

OH

C21H22O4Nominal Mass: 338

Exact Mass: 338.1518Found Mass: 338.1518 (Error = 0.1 ppm)

C22H26O3Nominal Mass: 338

Exact Mass: 338.1882Found Mass: 338.1879 (Error = 1.0 ppm)


Ionization Techniques

Short name Full name How it works

EI Electron Ionization Impact of 70 eV electrons

CI Chemical Ionization Transfer of H+ from CH5+

FI Field Ionization High Field (12kV)

FAB Fast Atom Bombardment Beam of Xenon atoms

FD Field Desorption High Field (12kV)

MALDI Matrix-Assisted Laser Desorption Ionization Laser hits sample in UV-absorbing matrix

ESI Electrospray Ionization Spray and desolvation of preformed ions


Ionization Technique Summary

Ionization Volatile Thermal Size Amount Examples

EI Yes Stable Small 1-2 mg Organics

CI Yes Stable Small 1-2 mg Organics

FI Yes Stable Small 1-2 mg Organics

FAB No Labile Medium 50μg-1 mg Polar/ionic organics, peptides, biomolecules, organometallic

FD No Labile Medium 1-2 mgNon-polar organics, organometallics, porphyrins, PAHs

MALDI No Labile Large 250fmol-500pmol

Peptides, proteins, DNA/RNA, polymers, dendrimers, fullerenes

ESI No Labile Large 1-300pmol/μLPolar/ionic organics, peptides, proteins, biomolecules, organometallics, polymers


Electron Ionization (EI)


EI Yes Stable Small 1-2 mg Organics


EI Electron Ionization Impact of 70 eV electrons

M - e- M+

Small stable organicsExtensive fragmentation, 70eV libraries availableHR available


EI Sample

OH

O

Chemical Formula: C11H12O2Exact Mass: 176.0837

Molecular Weight: 176.2118


EI Sample

O

O


Molecular Weight: 188.2225(Typical EI sample)


EI Sample

N N

O O

Chemical Formula: C35H30N2O2Exact Mass: 510.2307



EI Sample

N

NH

Chemical Formula: C24H28N2Exact Mass: 344.2252



EI Sample

Br

Chemical Formula: C13H17BrExact Mass: 252.0514


(Typical EI sample)


Chemical Ionization (CI)


CI Yes Stable Small 1-2 mg Organics


CI Chemical Ionization Transfer of H+ from CH5+

M + CH5+ (M+H)+

Small organics that fall apart in EIExtensive fragmentationHR available


CI Sample

O O

O

OH




CI Sample

N

OH O

O

Chemical Formula: C18H25NO3Exact Mass: 303.1834



CI Sample

OHO

O

O

Si(C2H5)3

Si(C2H5)3

Chemical Formula: C18H38O4Si2Exact Mass: 374.2309



CI Sample

H2N

NH2

Chemical Formula: C19H34N2Exact Mass: 290.2722



CI Sample

O

O

OH



(Typical CI sample)


Field Ionization (FI)


FI Yes Stable Small 1-2 mg Organics


FI Field Ionization High Field (12kV)

M + kV M+

Small organics that fall apart in EI, Isotopic enrichmentLittle fragmentationHR not available


Field Desorption (FD)


FD No Labile Medium 1-2 mgNon-polar organics, organometallics, porphyrins, PAHs


FD Field Desorption High Field (12kV)

M + kV M+

Non-polar compoundsNo fragmentationHR not available


FD Sample

NN

OO

O O

HN



(Typical FD sample)


FD Sample

NN

O

O

NN




FD Sample

N

N

N

O

OO




FD Sample

(CO)4Os

(OC)3Os Os(CO)3H

H

Chemical Formula: C10H2O10Os3Exact Mass: 857.8492


(Typical FD sample)


FD Sample

Chemical Formula: C42H24Exact Mass: 528.1878



Fast-Atom Bombardment (FAB)


FAB No Labile Medium 50μg-1 mg Polar/ionic organics, peptides, biomolecules, organometallic


FAB Fast Atom Bombardment Beam of Xenon atoms

M + H+ (M+H)+ M+ M+

Polar/ionic compounds in liquid matrix (DTT/DTE or 3-nba)Some fragmentationHR availableRarely used, superseded by ESI


FAB Sample

HNN

NH N

O

OO




Electrospray Ionization (ESI)


ESI No Labile Large 1-300pmol/μLPolar/ionic organics, peptides, proteins, biomolecules, organometallics, polymers


ESI Electrospray Ionization Spray and desolvation of preformed ions

M + H+ (M+H)+ M+ M+ M - H+ (M-H)- M- M-

Polar/ionic compounds, very little sample neededLittle fragmentationHR available


ESI SampleNH2

CH

C

H2C

O

CHO

O

NH

CH

C

H2C

O

H2C

H2C

HNCH2N

NH

NH

CH

C

CH

OCH3

H3C

HN

CH

CH2C

O

HO

HN CH C

CH

O

CH3

CH2

CH3

HN CH C

CH2

O

N

NH

N

C

O

NH

CH

C

CH2

O NH

CH

C

H2C

O

N

NH

NH

CH

C

H2C

OH

O

CH

CH3

CH3



Asp-Arg-Val-Tyr-Ile-His-Pro-Phe-His-Leu

Angiotensin

(Typical ESI sample)


ESI SampleO

HO

O

O

O

O

O




ESI Sample

O O

OO

O

NO2

OO

O

Chemical Formula: C32H25NO10Exact Mass: 583.1478



ESI Sample

N

Ir

N

S

HH

OO

PF6

Chemical Formula: C31H37F6IrN2O2PSExact Mass: 839.1847


(Typical ESI sample)


ESI Sample

N

HN

N

O

O




Matrix-Assisted Laser Desorption Ionization (MALDI)


MALDI No Labile Large 250fmol-500pmol

Peptides, proteins, DNA/RNA, polymers, dendrimers, fullerenes


MALDI Matrix-Assisted Laser Desorption Ionization Laser hits sample in UV-absorbing matrix

M + laser (M+H)+

Large compounds, very little sample neededLittle fragmentationHR not available


MALDI SampleOTg

OTg

OTg

OTg

TgO

TgO

MW=1907


MALDI Sample

CO2Tg

TIPSTIPS

12

MW=3822


MALDI Sample

N

NN

N

NH2

O

HO

HH

HH

PO

O-

HO

O-

n

MW=34,000


MALDI Sample

H2N-UNPSTFPGHSALMICGTLTLK-OH

MW=1852


MALDI Sample

OBr

=

O

O

HO

OHO

OH

*

*

7

MW = 1270


MS Experiment Summary

Experiment Ionization Comments

LR All Mass Spectrum

HR EI, CI, FAB, ESI Exact Mass – Elemental Composition

MS/MS ESI, FAB, MALDI Structure, e.g. Peptide Sequence

Isotope Ratio FI Isotopic Enrichment

GC/MS EI, CI, FI

LC/MS ESI HPLC or Capillary HPLC

School of Chemical Sciences, University of IllinoisGraphic by Dorothy Loudermilk


Further InformationMass Spectrometry: A Textbook

Jürgen H. Gross2004, Springer-Verlag, ISBN 3-540-40739-1

Interpretation of Mass Spectra, 4th editionF. W. McLafferty1993, University Science Books, ISBN 0-93-570225-3

Mass Spectrometry for BiotechnologyGary Siuzdak1996, Academic Press, ISBN 0-12-647471-0


Use of this fileThis slide show was prepared by

Steven L. Mullen, Ph.D.Mass Spectrometry LaboratorySchool of Chemical SciencesUniversity of Illinois(217) 333-2545

It may be used for non-commercial educational purposes, withappropriate credit and no alterations.

ionization methods in mass · pdf fileionization methods in mass spectrometry at the scs mass...

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