Lecture 14 LC-MS Ionization

GC-MS GC MS Computer

LC-MS LC MS Computer

MS Analysis MS Gas phase Ions LC?

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

Difference? PCBs PFOS Production started 1920s Env Detection 1968 2001

MS Analysis GC-MS and LC-MS ionization are very different GC – Producing ions from gas phase analytes Producing gas phase compounds from charged analytes LC – Necessary for certain important analytes: Biological molecules: Proteins Amino acids/nucleic acids sugars Strong acids without the ability to absorption UV-vis

Electrospray Ionization Problem solved… Electrospray Ionization (ESI) John Fenn

Electrospray Ionization (ESI) LC ESI Key Features: - Capillary (LC) 90o to Cone (MS) Capillary Ionization Chamber - Atmospheric pressure ion source - Nebulized flow through capillary - Heat applied to evaporate solvent Heater Gas - Voltage difference applied between capillary and cone Nebulized Spray + MS Sample Cone - Detect (+) ions  Cap (+), Cone (-) - Detect (-) ions  Cap (-), Cone (+) - 10-6 torr What happens in the droplet? 760 torr

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 Works well for compounds that can be charged in the condensed phase Strong or weak Acids and Bases

Atmospheric Pressure Chemical Ionization (APCI) APCI Key Features: LC - Capillary (LC) 90o to Cone (MS) - Atmospheric pressure ion source Capillary Ionization Chamber - Nebulized flow through capillary - Heat applied to evaporate solvent - No voltage applied to capillary - droplets are not charged Heater Gas Nebulized Spray - Voltage still applied to cone - Detect (-) ions  Cone (+) - Detect (+) ions Cone (-) Sample Cone MS - - Heat evaporates solvent, analytes partition into the gas-phase uncharged 10-6 torr Corona discharge needle 760 torr - Need to make ions!

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 Works well for compounds that are protic but neutral at a pH between 2 and 9 Weak Acids / Bases

Atmospheric Pressure Photoionization (APPI) APPI Key Features (same as APCI): LC - Capillary (LC) 90o to Cone (MS) - Atmospheric pressure ion source Capillary Ionization Chamber - Nebulized flow through capillary - Heat applied to evaporate solvent - No voltage applied to capillary - droplets are not charged Heater Gas Nebulized Spray - Voltage still applied to cone - Detect (-) ions  Cone (+) - Detect (+) ions Cone (-) Sample Cone MS - UV lamp - Heat evaporates solvent, analytes partition into the gas-phase uncharged 10-6 torr 760 torr - Need to make ions!

Atmospheric Pressure Photoionization (APPI) 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 Works well for compounds that are absorb UV radiation or are protic but neutral at a pH between 2 and 9 Weak Acids / Bases Nonpolar compounds that are UV active

Summary LC-MS Ionization m/z (MW) www.agilent.com

Matrix-Assisted Laser Desorption/Ionization (MALDI) MS MALDI Key Features: Vacuum - Analyte deposited on surface with excess matrix compound + - - Shine laser on the surface - hn aborbed by matrix molecules - Matrix molecules transfer energy to analyte molecules - Results in desorption/ionization of both matrix and analyte Laser beam Analyte molecule Matrix molecule Adapted from Dass, C. Fundamentals of Contemporary Mass Spectrometry, Wiley-Interscience, 2007.

Matrix-Assisted Laser Desorption/Ionization (MALDI) Common dopants: Analytes of interest: Proteins Polymers 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)