Mass Spectrometry (MS) Alternative names: Mass spectrometric (selective) detector (MSD) Spectrometry - methods based on interaction of matter and radiation Mass spectrometry - method based on formation of ions followed by interaction of ions with electric/magnetic fields
MS principle and outputs of measurements 1. Formation of ions (ionization) 2. Filtration of ions (mass analysis) 3. Measuring of ions abundance in dependence on m/z value m - relative molecular weight of ion z - charge of ion (1, 2, 3, 4, 5, 6...) 4. Mass spectrum * axis x corresponds to m/z value * axis y corresponds to abundance of ions (signal intensity) J. Poustka, VŠCHT Praha, ÚAPV 2016, http://web.vscht.cz/poustkaj
Mass spectrum - format Graphic (profiles X lines) Tabulated m / z Intenzita Relativní intenzita (%) 57 560 3,72 58 800 5,31 59 550 3,65 125 2532 16,81 126 2935 19,48 127 2580 17,13 128 2100 13,94 134 5785 38,40 135 15065 100,00 136 6256 41,53 137 10258 68,09 138 4618 30,65
Application possibilities Identification of compounds Empirical formula almost absolutely, if sufficiently high resolution is applied Identification of structure in dependence on ionization type (fragmentation) Comparison of spectra - identification based on characteristic formation of ions with specific m/z values and abundances (spectral libraries) Quantification of compounds Universal - LOD/LOQ differ according to ability to be ionized
Historical development of MS - summary 110 years From cathode tube to modern mass spectrometer 2010 GC-MS *1956 LC-MS(ESI) *1984 TOF-MS 1900 1899-1920 *1946 Development of spectra recording
IONS - formation and types * Various mechanisms - fundamental effect on spectrum * Ions charge - positive or negative (only one type at the time) positive or negative ionization - switching of modes * Regarding to original structure are distinguished: a) molecular ions: [M] + or [M] - b) pseudomolecular and adduct ions: [M+H] +, [M-H] -, [M+CH 4 ] + c) fragment ions: e.g. [M-CH 3 ] + J. Poustka, VŠCHT Praha, ÚAPV 2016, http://web.vscht.cz/poustkaj
Ionization in gas phase (a) (at reduced pressure - vacuum 10-4 to -8 torr) Electron Ionization - EI * loss of an electron after the Electron Impact or otherwise after the release of valence electrons - formation of ions: M + e - (70eV) M +. + 2e - * standard EI spectra (library) - ionization energy 70 ev (1 ev = 96,487 kj/mol = 23,06 kcal/mol) * "hard" ionization a) spectrum rich of fragment ions b) molecular ions - sometimes are not stable (not present in spectrum) * applicable for: finding of empirical formula, structure analysis, quantification * application: fundamental for GC-MS, probe, moving belt, particle beam J. Poustka, VŠCHT Praha, ÚAPV 2016, http://web.vscht.cz/poustkaj
J. Poustka, VŠCHT Praha, ÚAPV 2016, http://web.vscht.cz/poustkaj Ionization in gas phase (a) (at reduced pressure - vacuum 10-4 to -8 torr)
J. Poustka, VŠCHT Praha, ÚAPV 2016, http://web.vscht.cz/poustkaj Ionization in gas phase (b) (at reduced pressure - vacuum 10-4 to -8 torr) Field Ionization - FI * loss of an electron after the passing through the high voltage electrical field ( 5 kv) - formation of ions * "soft" ionization molecular and adduct ions are dominating ([M+Na] + etc.) * applicable for: finding of empirical formula; quantification * application: additional for GC-MS, probe
J. Poustka, VŠCHT Praha, ÚAPV 2016, http://web.vscht.cz/poustkaj Ionization in gas and liquid phase (at reduced or atmospheric pressure) Chemical ionization - CI * adduction of reagent or capture of low energy (thermal electron) - formation of (PCI) or (NCI) ions * various conditions - no standard spectra (no library availability) * "soft" ionization only most stable ions (usually molecular), fragment ions are mostly absent in source fragmentation (loss of molecular ions) * applicable for: finding of empirical formula; quantification * application: GC-MS, LC-MS (APCI), probe, moving belt, particle beam
Ionization in liquid and solid phase (a) (at reduced pressure) Ionization by accelerated ions or atoms a) fast ion bombardment FIB, e.g. Cs + b) fast atom bombardment FAB, e.g. Xe, Ar * suitable for high molecular compounds * applicable for: finding of empirical formula; quantification * application: probe, continuous flow (CF FIB / FAB), moving belt / wire, LC-MS
Ionization in liquid and solid phase (a) (at reduced pressure) CF FIB / FAB Moving belt / wire
J. Poustka, VŠCHT Praha, ÚAPV 2016, http://web.vscht.cz/poustkaj Particle beam
Ionization in liquid and solid phase (b) (at reduced pressure) Desorption ionization a) chemical (DCI): Pt wire wetted by sample - fast heating b) plasma ( 252 Cf PD): californium decays into radioactive isotopes - formation of adducts c) laser (LDI): pulse evaporation by laser d) laser at matrix assistance (MALDI): LDI with matrix e) field (FD): ionization direct from emitter (see field ionization - FI) * structure analysis * application: probe
Ionization in liquid and solid phase (b) (at reduced pressure) MALDI
Ionization in liquid phase (c) (at reduced pressure) Thermospray - TSP * Ionization by fast evaporation at elevated temperature ( 300 C) at simultaneous nebulization to evacuated space * "soft" ionization predominant molecular and pseudomolecular ions * applicable for finding of empirical formula * application: LC-MS (first interface similar to API)
Ionization in liquid phase (c) (at reduced pressure) [M+H] + [M+NH 4 ] + [M-H] - (not [M + zh] z + ) t 150 300 C flow rate < 1 ml/min
Ionization in liquid phase (a) (atmospheric pressure ionization - API) Electrospray - ESP, Electrospray ionization - ESI * Ionization by high voltage ( 5 kv) at simultaneous nebulization * "soft" ionization predominant molecular and pseudomolecular ions single and/or multicharged ions * applicable for: finding of empirical formula; quantification * application: LC-MS
Ionization in liquid phase (a) (atmospheric pressure ionization - API)
Ionization in liquid phase (a) (atmospheric pressure ionization - API) ESI [M+H] + [M+NH 4 ] + [M-H] - [M ± zh] z ± t N2 50 400 C 2-8 kv Flow rate 0.001-1 ml/min Volatile modifiers: ammonium acetate, formic acid Nonvolatile modifiers: phosphate buffers
Ionization in liquid phase (a) (atmospheric pressure ionization - API) Types of electrospray Ionspray (ISP) - pneumatic nebulization Ultraspray (USP) - ultrasound nebulization Turbospray (TBSP) - thermal nebulization
Ionization in liquid phase (b) (atmospheric pressure ionization - API) Atmospheric pressure chemical ionization - (APCI) * ionization based on combination of evaporation at high temperature ( 400 C) and electric discharge ( 5 kv) at simultaneous nebulization * "soft" ionization predominant molecular and pseudomolecular ions * applicable for: finding of empirical formula; quantification * application: LC-MS
Ionization in liquid phase (b) (atmospheric pressure ionization - API) APCI [M+H] + [M+NH 4 ] + [M-H] - ([M ± zh] z ± ) t 200 650 C Discharge: 2-8 kv Flow rate 0.2-2 ml/min Volatile modifiers: ammonium acetate, formic acid Nonvolatile modifiers: phosphate buffers
Ionization in liquid phase (c) (atmospheric pressure ionization - API) Atmospheric pressure photo-ionization - (APPI) * ionization based on combination of evaporation at high temperature ( 400 C) and UV radiation ( 10 ev) at simultaneous nebulization * "soft" ionization predominant molecular and pseudomolecular ions * applicable for: finding of empirical formula; quantification * application: LC-MS
Ionization in liquid phase (c) (atmospheric pressure ionization - API) APPI [M+H] + [M+NH 4 ] + [M-H] - ([M ± zh] z ± ) t 200 650 C UV radiation 10 ev Flow rate: 0.2-2 ml/min Dopant - ionization support: toluene, benzene
Ionization in liquid phase (d) (atmospheric pressure ionization - API) DESI Desorption ESI
Ionization in liquid phase (d) (atmospheric pressure ionization - API) DART Direct Analysis in Real Time
Spray techniques Z-spray
Molecular weight Applicability of ionization techniques J. Poustka, VŠCHT Praha, ÚAPV 2016, http://web.vscht.cz/poustkaj 100 000 ESI 10 000 APPI 1 000 APCI Particle Beam Thermospray FIB FAB GC/MS (EI/CI/FI) Nonpolar Analyte polarity Polar