Overview of NETCHEM MSc & PhD courses: mass spectrometry in EFSC

Transcription

1 AQualEER Chemistry faculty (Belgrade, Serbia, December 2017) Overview of NETCHEM MSc & PhD courses: mass spectrometry in EFSC Emeritus Prof. Dr Jean-Claude Tabet 1,2,3 1 Institut Parisien de Chimie Moléculaire CNRS UMR 8232 (UPMC-Paris), France 2 Laboratoire d Etude du Métabolisme des Médicaments (CEA-Saclay), France 3 AQuaLEER, France Mass Spectrometry jean-claude.tabet@courriel.upmc.fr in EFSC - (Prof. Dr J.-C. Tabet )

2 General objectives Complex mixture analysis w.i. or w.o. extraction Qualitative: Identification & structural elucidation Molecular mass Elemental composition Structure, isomer, and stereochemistry Quantitative relative/absolute determination Concentration with LOD and LOQ

3 Dissociation spectrum of [Arg+H] +

4 L/I differentiation using complexes with Cu II (from 10 µl of 10 µm solution)

5 What areas can mass spectrometry help with molecular analysis? Chemistry 1. Organic chemical: small molecu les e.g. dyes, oils+ by-products, polymers 2. Doping and Forensic 3. Cosmetic and drugs 4. Food safety: e.g. toxico- 5. Environmental analysis: water and soil (e.g. POPs) 6. Homeland security. Biology 1. Biological fluids 2. Natural products 3. Surface imaging (e.g. brains, liver, cell) 4.Omics studies (metabolites, lipids, peptides, proteins, DNAs, carbohydrates, non- covalent complexes (SB vs. HB)

6 First of all, what is mass spectrometry as tool? It's based on what processes? Physic: ion motion and stable trajectory Analytical Chemistry: separation (GC, LC) and ion analysis Chemistry: formation of ions & dissociation

7 The most used general notions: - Pressure (from few mbar to mbar) - Temperature (condensed phase vs. macromolecular) - Voltage (DC, V RF ) and magnet field - mass unit (u or Da, defined by IUPAC, sept. 2013) - Natural isotopes, resolution - molecular mass (mono isotopic, average, nominal) - m/z ratio (z charge number), relative abundance - Ionization and ionization/desorption modes - Energy (E cin. & E com, E (vibr.) in fundamental & excited levels), bond energy - ΔH f of neutral and ions, activation energy E o, transi tion state, energy pathway (ZPE) - Collision (relaxation in E cin. & E vibr., ion-molecule reactions and ion dissociations) Overview -. of NETCHEM MSc and PhD courses:

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9 Principle of mass spectrometer

10 What kind of instrument choice??? Instruments with sectors, TOF/ MS & tandem with FT/MS Instrument Benchtop Q m/z filter QqQ, ITMS Orbitrap Miniaturized, portable and embedded instruments for in field analysis

11 Large variety of MS intruments

12 Minimum theoretical basis required to understand the necessary concepts used in mass spectrometry spectrometry in addition to chemical and possibly biological knowledge of the user

13 Competitive losses due to simple cleavage reactions Orientation depending on the thermochemistry: IE = 9.04 ev AE (M-Br) + = 12.7 ev AE (M-Cl) + = 13.2 ev

14 A same molecule and different electron ionization mass spectra (EI-MS) Molecular ion M + very weak in NIST databank mass spectra and diagnostic fragment ions at m/z 79, m/z 97 and m/z 128 recorded using quadrupole filter MS. Molecular ion M + accompanied by protonated molecule MH + & fragment ions (m/z 79, m/z 97, m/z 128) in EI mass spectrum recorded with iontrap MS.

15 EI mass spectra using ion-beam (a) and ion storage (b) MS modes display some surprising (instrument dependant) 1. Organic chemistry: ion formation & dissociation, 2. kinetic effect: M + becomes MH + & M 2 H + due to ion-molecule reaction depending on residen ce -time 2. Ion physic: space charge effect resulting in m/z shifting of M 2 H + M 2 + related to the change of the ion motion frequency.

16 Resonant vs. non-resonant excitation in the MS/MS experiments (ion-trap vs quad. cell) Resonant mode: only an ion with the selected frequen cy is excited Non-resonant: All ions are excited independen tly of their frequencies

17 It would be desirable to be able to explain some accidental artifacts. To cite some examples, let s takes the case where the used ion analyzer is an ion trap (Paul s trap or 2D ion trap). Particular situations such as the apparition of ghost peaks or the m/z ratio shift can occur due to the charge space effect, if : - AGC (or ICC) is accidently turned off (or defective) or - a too large number of ions is injected in the ion trap

18 Space charge effect on resolution and accurate mass measurements The use of the reverse scan mode i.e., called as zoom scan

19 Non-linear résonances (hexa-, octa-, dodecapolars) and «ghost peaks» Being like a hen in front of a toothbrush X4

20 Non-linear resonances and «ghost peaks» X4

21 Space charge and «ghost peaks» X4 Injection duration: 1 ms Injection duration: 10 ms Injection duration: 100 ms Ion ejection at b z = 1/3 yields the ghost peak observation by quadrular field à b z =1 due to the space charge in the ion trap. From the external ESI source, the ion injection duration plays a main role.

22 Prompt fragile ion cleavage prior to reach the limit of ion stability How is it possible to prevent the omelet and the eggs loss?

23 Finally, competencies required for understanding, & to practice mass spectrometry as tool in analytical chemistry & biology science are: 1. Basic physical part 2. Physical-chemistry 3. Thermochemistry and kinetic 4. Organic chemistry and basic biology This allows going towards an enhanced efficiency of its using especially in the EFSC

24 Mass spectrometry courses: a program A. formation of ions 1. Gas phase ionization modes (volatils from compounds in : doping, environment, foods, forensic) - In vacuum : EI, CI, FI - Atmospheric pressure: APCI, APPI, DART 2. Ionization/desorption modes - In vacuum: LD, MALDI - Atmospheric pressure: ESI, DESI, LDESI, AP-MALDI (and other ambient ionizations)

25 Mass spectrometry courses: a program B. Dissociation of monocharged species 1.Kinetic theory & thermochemistry (gas phase basicity & acidity): competitive dissociation (rearrangement vs. simple cleavage). 2.Prompt dissociation in the ion source - In vacuum from odd- and even-electron ions. Distonic, ion-neutral species. Cleavage rules. - In reduced pressure (after desolvation steps for charged molecular species ) 3.Dissociation of selected ions by activation - collision by gas (CID: reso nant or not) or electron (EID) - by photon (IRMPD), and BIRD - ETD (or ECD and EDD) for multiply charged species,

26 Mass spectrometry courses: a program 1. Analyzers based on: C. Instrumentation - ion beam: time-of-flight, ion mobility, quadrupole - ion storage: Paul s trap (3D and 2D), Penning trap and orbitrap 2. Analyzers in tandems - Homogene ous tandems: triple quad, TOF/TOF - Tandem hybrids: Q/qTrap, QqTOF, Q/Trap/TOF, IMMS derivated tandems and with very HR (Qh/FTICR, LTQ/Orbitrap) 3. GC/MS and LC/MS coupling

27 How to distribute the MS courses between the master and PhD levels Master (15/8h cour se /lab) 1. Gas phase ionization, desorption meaning 2. Mechanismes of fragmentation in EI* and CI* (and exercices) 3. MS/MS with TQ/ITMS (low energy collision on even-electron ions) 4. Instrumentation MS (quad., ion trap, TOF) 5. GC/MS in EI &CI (lab) PhD (20/12h cour se /lab) 1. Ionization/desorption (ESI, MALDI..) 2. Tandem hybrides: IMMS, and FT/MS 3. MS/MS with ergodic and non-ergodic modes). Databank 4. MS 2 analysis in HR on biological molecules 5. LC/MS and MS/MS in ESI +/- (lab)

28 Thank you for your attention