Mass Spectrometry II

Ion trap

Magnetic Sector

FB

Magnetic Sector

Magnetic Sector

Time of Flight MS

In a quadrupole MS masses are separated based on their flight path

In a time-of-flight MS masses are separated based on their inertia

detectorIon-source+

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Field-free flight zone

Mass is determined by the time it takes the ion to move from source to detector

t =L (length)

(velocity)

=2V

(m/z)√+ -

(acceleration voltage)

(mass-to-charge)

Acceleration zone

235.16 to 235.19 amu

281.13 to 281.15 amu

193.12 to 193.14 amu

179.10 to 179.12 amu

169.09 to 169.08 amu

301.14 to 301.16 amu

MS Application

Environmental Issue:• Presence of bioloigcally-active compounds (pharmaceuticals…) that are not degraded in a wastewater treatment plant• Compound of interest is trimethoprim, an anti-microbial used to treat infectious diseases in humans

Analytical - want to analyze for trimethoprim• LC/GC?• Ionization source?

LC – why?ESI – why?

Hypothesis• Primary treatment of wastewater with nitrifying activated sludge will degrade trimethoprim

MS Application

Hypothesis• Primary treatment of wastewater with nitrifying activated sludge will degrade trimethoprim

Experiment• Small scale laboratory batch reactor was used to degrade trimethoprim

• LC-ESI coupled to a single quadrupole MS to determine the unit mass of degradation products

• LC-ESI coupled to an ion trap MS to determine structural features of the degradation products

• LC-ESI coupled to a quadrupole time of flight MS to determine the accurate mass of the degradation products to get a chemical formula

MS Application

Hypothesis• Primary treatment of wastewater with nitrifying activated sludge will degrade trimethoprim

Experiment• Small scale laboratory batch reactor was used to degrade trimethoprim

• LC-ESI coupled to a single quadrupole MS to determine the unit mass of degradation products

• LC-ESI coupled to an ion trap MS to determine structural features of the degradation products

• LC-ESI coupled to a quadrupole time of flight MS to determine the accurate mass of the degradation products to get a chemical formula

MS Application

Data fromLC-ESI-single quad MS

Parent [M+1]

MW: 290.32 g mol-1

Degradation products

Anything strange…

Total ion chromatogram

Single ion chromatogram

MS Application

Experiment• LC-ESI coupled to a single quadrupole MS to determine the unit mass of degradation products

• 2 degradation products• [M+H] of 307 m/z and 325 m/z• Increased mass but earlier LC elution• Suggests oxidation

• Mass difference between 291 m/z and 307 m/z is 16 m/z (oxygen)• Mass difference between 291 m/z and 325 m/z is 34 m/z (2O and 2 H)

MS Application

Data fromLC-ESI-Ion trap MS (MS2)LC mobile phase is H2O and CH3OH

291 (trimethoprim) trapped in ion trapAccelerated/dissociated and the products detected

[M+H] is 291

291 m/z → products

MS Application

MS Applications

• LC-ESI coupled to an ion trap MS to determine structural features of the degradation products

Parent •Demonstrates that the phenyl rings are recalcitrant to dissociation• Useful information for the ion trap analysis of the degradation products

Experiment• LC-ESI coupled to a single quadrupole MS to determine the unit mass of degradation products

• 2 degradation products• [M+H] of 307 m/z and 325 m/z• Increased mass but earlier LC elution• Suggests oxidation

• Mass difference between 291 m/z and 307 m/z is 16 m/z (oxygen)• Mass difference between 291 m/z and 325 m/z is 34 m/z

Data fromLC-ESI-Ion trap MS (MS2)

Degradation product 307 m/z trapped in ion trap Accelerated/dissociated and the products detected

D2O and CD3OD

307-18Loss of water[M + H – H2O]+

307 m/z → products

MS Application

Data fromLC-ESI-Ion trap MS (MS3)

Degradation product 307 m/z trapped in ion trap Accelerated/dissociated and 289 m/z trapped in ion trap Accelerated/dissociated and the products detected

D2O and CD3OD

Can be formed from the dissociation of 307m/z or 289 m/z

307 m/z → 289 m/z → products

MS Application

Data fromLC-ESI-Ion trap MS (MS3)

Degradation product 307 m/z trapped in ion trap Accelerated/dissociated and 274 m/z trapped in ion trap Accelerated/dissociated and the products detected

D2O and CD3OD

307 m/z → 274 m/z → products

Can be formed from the dissociation of

289 m/z and 274 m/z

MS Application

Products• Use MS3 data to propose a fragmentation pathway for 307 m/z degradation product• Structure confirmation

• LC-ESI coupled to an ion trap MS to determine structural features of the degradation products

Parent•Demonstrates that the phenyl rings are recalcitrant to dissociation• Useful information for the ion trap analysis of the degradation products

Experiment• LC-ESI coupled to a single quadrupole MS to determine the unit mass of degradation products

• 2 degradation products• [M+H] of 307 m/z and 325 m/z• Increased mass but earlier LC elution• Suggests oxidation

• Mass difference between 291 m/z and 307 m/z is 16 m/z (oxygen)• Mass difference between 291 m/z and 325 m/z is 34 m/z

MS Application

- H, - OH, - CH3

Data fromLC-ESI-Ion trap MS (MS2 and MS3)

Propose fragment structures

MS Application

Hypothesis• Primary treatment of wastewater with nitrifying activated sludge will degrade trimethoprim

Experiment• Small scale laboratory batch reactor was used to degrade trimethoprim

• LC-ESI coupled to a single quadrupole MS to determine the unit mass of degradation products

• LC-ESI coupled to an ion trap MS to determine structural features of the degradation products

• LC-ESI coupled to a quadrupole time of flight MS to determine the accurate mass of the degradation products to get a chemical formula

Answer• Degradation of trimethoprim is possible using nitrifying activated sludge• Degradation products identified using several MS techniques

MS Application

Data fromLC-ESI-Ion trap MSLC mobile phase is D2O and CD3OD

296 (d-trimethoprim) trapped in ion trapAccelerated/dissociated and the products detected

[M+D] is 296

MS Application

Data fromLC-ESI-Ion trap MS

D2O and CD3OD

H2O and CH3OH

MS Application