lecture 14 lc-ms ionization. gc computer ms gc-ms
TRANSCRIPT
Lecture 14
LC-MS
Ionization
GC
Computer
MS
GC-MS
LC
Computer
MS
LC-MS
MS Analysis
MS
Gas phaseIons LC?
OH
H3CH2CH2C CH2CH2CH3
NO2O2N
CF3
S
F F
F F
F F
FO
O
O
Condensed Phase Ionization
Electrospray Ionization (ESI)
Atmospheric Pressure Chemical Ionization (APCI)
Atmospheric Pressure Photoionization (APPI)
Matrix Assisted Laser Desorption/Ionization (MALDI)
Desorption Electrospray Ionization (DESI)
Direct Analysis in Real Time (DART)
Liquid phase(LC)
Solid phase
PCBs
Production started1920s
Env Detection1968
PFOS
Production started1930s
Env Detection2001
Difference?
GC-MS and LC-MS ionization are very different
GC – Producing ions from gas phase analytes
Producing gas phase compounds from charged analytesLC –
MS Analysis
Necessary for certain important analytes:– Biological molecules:
• Proteins• Amino acids/nucleic acids• sugars
– Strong acids without the ability to absorption UV-vis
Problem solved…
John Fenn
Electrospray Ionization
(ESI)
Electrospray Ionization(ESI)
Gas
Nebulized
Spray
LC
Heater
ESI Key Features:
- Nebulized flow through capillary
- Heat applied to evaporate solvent
- Capillary (LC) 90o to Cone (MS)
Sample
Cone
MS
CapillaryIonization
Chamber
760 torr
10-6 torr
- Atmospheric pressure ion source
- Voltage difference applied between capillary and cone
-
+ - Detect (+) ions Cap (+), Cone (-)
- Detect (-) ions Cap (-), Cone (+)
What happens in the droplet?
Electrospray Ionization(ESI)
Interested in Positive Ions
- Apply (+) Voltage to capillary
- (+) ions repelled from capillary
- (-) ions attracted to capillary
- Results in droplets enriched in (+) ions
P. Kebarle, J. Mass Spectrom. 2000, 35, 804–817.
+++
+++
Electrospray Ionization(ESI)
Summary– Ions are formed in condensed phase
• In mobile phase droplets
– Ions move into the gas phase by two mechanisms:• Partition into gas phase due to columbic repulsion in droplet• Solvent in droplet evaporates leaving ion behind
OH
H3CH2CH2C CH2CH2CH3
NO2O2N
CF3S
F F
F F
F F
FO
O
O
Works well for compounds that can be charged in the condensed phase
Strong or weak Acids and Bases
Atmospheric Pressure Chemical Ionization(APCI)
Gas
Nebulized
Spray
LC
Heater
APCI Key Features:
- Nebulized flow through capillary
- Heat applied to evaporate solvent
- Capillary (LC) 90o to Cone (MS)
MS
CapillaryIonization
Chamber
760 torr
10-6 torr
- Atmospheric pressure ion source
- No voltage applied to capillary
- droplets are not charged
-
- Voltage still applied to cone
- Detect (-) ions Cone (+)
- Detect (+) ions Cone (-)
Sample
Cone
- Heat evaporates solvent, analytes partition into the gas-phase uncharged
- Need to make ions!
Corona discharge
needle
Atmospheric Pressure Chemical Ionization(APCI)
Summary– First step is solvent evaporation
• Compounds in gas-phase as neutrals
– Ions produced in the gas-phase• Corona discharge ionizes carrier gas and mobile phase• Acid/base reactions in the gas phase with mobile phase
OH
H3CH2CH2C CH2CH2CH3
NO2O2N
CF3
S
F F
F F
F F
FO
O
O
Works well for compounds that are protic but neutral at a pH between 2 and 9
Weak Acids / Bases
Atmospheric Pressure Photoionization(APPI)
Gas
Nebulized
Spray
LC
Heater
APPI Key Features (same as APCI):
- Nebulized flow through capillary
- Heat applied to evaporate solvent
- Capillary (LC) 90o to Cone (MS)
MS
CapillaryIonization
Chamber
760 torr
10-6 torr
- Atmospheric pressure ion source
- No voltage applied to capillary
- droplets are not charged
-
- Voltage still applied to cone
- Detect (-) ions Cone (+)
- Detect (+) ions Cone (-)
Sample
Cone
- Heat evaporates solvent, analytes partition into the gas-phase uncharged
- Need to make ions!
UV lamp
Summary– First step is solvent evaporation
• Compounds in gas-phase as neutrals
– Ions produced in the gas-phase• UV lamp ionizes compound by photon absorption or
• Charge exchange with dopant or
• Dopant ionizes mobile phase, followed by;
• Acid/base reactions in the gas phase with mobile phase
OH
H3CH2CH2C CH2CH2CH3
NO2O2N
CF3
S
F F
F F
F F
FO
O
O
Works well for compounds that are absorb UV radiation or are protic but neutral at a pH between 2 and 9
Weak Acids / BasesNonpolar compounds that are UV active
Atmospheric Pressure Photoionization(APPI)
Summary LC-MS Ionization
www.agilent.com
m/z(MW)
Matrix-Assisted Laser Desorption/Ionization (MALDI)
Matrix molecule
Analyte molecule
Adapted from Dass, C. Fundamentals of Contemporary Mass Spectrometry, Wiley-Interscience, 2007.
Laser beam
+
+ -
MALDI Key Features: - Analyte deposited on surface with excess matrix compound
- Shine laser on the surface
- h aborbed by matrix molecules
- Matrix molecules transfer energy to analyte molecules
- Results in desorption/ionization of both matrix and analyte
MS
Vacuum
Matrix-Assisted Laser Desorption/Ionization (MALDI)
Common dopants:
Analytes of interest:– Proteins– Polymers
HO
HC
C
C
C
O
OH
N
HO COOH
OH
– High MW, ionization not possible using ESI, APCI, APPI
– High MW separates these compounds from the background signal of the matrix
Desorption Electrospray Ionization(DESI)
Z. Takáts et al. Science 2004, 306, 471-473.
Direct Analysis in Real Time(DART)