1 / 58

Spectrométrie de masse: tour d’horizon de la technique et applications

Spectrométrie de masse: tour d’horizon de la technique et applications. Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department habib@chim.ucl.ac.be. Layout. Mass spectrometry Elemental composition Radiosterilization of solid metoclopramide Phytosterols Glucosamine Carnitines

sheryl
Download Presentation

Spectrométrie de masse: tour d’horizon de la technique et applications

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. Spectrométrie de masse: tour d’horizon de la technique et applications Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department habib@chim.ucl.ac.be

  2. Layout • Mass spectrometry • Elemental composition • Radiosterilization of solid metoclopramide • Phytosterols • Glucosamine • Carnitines • Proteomics Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  3. Mass spectrometry Technical realizations sample introduction Ionization methods vacuum systems type of mass analyzers combination of mass analyzers modes of operation coupling of separation device Fundamentals ionization processes internal energy time scale of events isotopic distribution isotopic mass MS Applications identification quantification Mass spectral interpretation fragmentation pathways characteristic ions rules Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  4. Mass spectrometry 2000 Spectrum of noncovalently associated macromolecules > 106 Da 2005 Orbitrap Makarov 1988 ESI by Fenn First spectra of protein > 20000 Da 1987 MALDI Karas & Hillenkamp 1918 1982 First complete MS spectrum of Insulin and first commercial triple Quad 1974 First LC/MS and FTICR description 1968 First commercial Quad 1958 First commercial TOF 1956 First GC/MS 1953 Quad an Trap description Paul & Steinwedel 1942 First commercial Mass spectrometer 1912 First mass spectrometer Thomson 1897 Thomson discovers e- Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  5. Mass spectrometry Relative intensity m/z EIMS of 2-methyl-3-pentanone (MW = 100) Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  6. Mass spectrometry http://physics.nist.gov/PhysRefData/Compositions/index.html http://www.webelements.com/ Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  7. Mass spectrometry For high molecular weights, the difference between the various masses becomes important : Insulin : C257H383N65O77S6 Chemical mass 5807.6559 Nominal monoisotopic mass : 5801 Exact monoisotopic mass : 5803.6375 The instrument response will depend on its resolution Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  8. Mass spectrometry Sample Introduction HPLC GC Ion Source Analyser(s) Vacuum Detector Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  9. Mass spectrometry Analyzers Magnetic and electric sectors Quadrupoles Time of Flight (Linear, Reflectron) Ion Trap ( 3D, Linear, Orbitrap) Fourier Transform Ion Cyclotron Resonance Hybrid instruments Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  10. Mass spectrometry Ion sources Electron Ionisation Chemical ionisation Fast Atom Bombardment Laser Desorption Ionisation Matrix-Assisted Laser Desorption Ionisation AP-MALDI Electrospray Atmospheric Pressure Chemical Ionisation Atmospheric Pressure Photoionisation Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  11. Mass spectrometry resolved ions sample ions mass analyzer detector source Mass spectrum intensity m/z Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  12. Mass spectrometry : tandem mass spec or MS2 Full spectrum source Collision Induced Dissociation MS2 spectrum detector Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  13. Elemental composition C13H14NO3Cl Masse : 267.0662 M+1 : 268.0740 M+3 : 270.0640 Peak matching results : 268.0735 12C13H1535ClNO3 270.0705 12C13H1537ClNO3 Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  14. Measured : 150.0701 ± 0.010 C : 0 – 30 H : 0 – 60 N : 0 – 10 O : 0 - 15 Measured : 150.0701 ± 0.050 Elemental composition 14 compositions 60 compositions Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  15. Measured : 322.0151 ± 0.010 C : 0 – 30 H : 0 – 60 N : 0 – 10 O : 0 - 15 Measured : 322.0151 ± 0.010 Elemental composition 31 compositions 152 compositions C : 0 – 30 H : 0 – 60 N : 0 – 10 O : 0 – 15 S : 0 - 10 Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  16. Radiosterilization of solid-state metoclopramide Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  17. Radiosterilization of solid-state metoclopramide Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  18. Radiosterilization of solid-state metoclopramide Peak 2 Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  19. Radiosterilization of solid-state metoclopramide Peak 3 Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  20. Radiosterilization of solid-state metoclopramide Peak 4 Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  21. Phytosterols in Spelt Spelt is said to have better nutritive value than winter wheat. Functional properties have traditionally been attributed to spelt. Lack of scientific evidence for these claims. What about the phytosterol contents ? b-sitosterol Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  22. Phytosterols in Spelt Fine bran ground to 1 mm sieve Saponification in KOHaq (500g/l) / EtOH (1/3), overnight, RT, dark Extraction with diethyl ether Washing with water Filtration (MN 616 wa filter) Evaporation to dryness and redissolution in EtOH SPE (C18, CHCL3/MeOH; 95/5) Evaporation to dryness and redissolution in absolute EtOH Filtration on DynaGardTM filters Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  23. Phytosterols in Spelt SG FS ASG SE LC/APCI-MS TIC trace of 19 standard compounds Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  24. Phytosterols in Spelt ASG SG [M+H-H2O]+ FS ES Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  25. Phytosterols in Spelt FS SG LC/APCI-MS TIC trace Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  26. Phytosterols in Spelt Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  27. Phytosterols in Spelt Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  28. Phytosterols in Spelt Combined Steryl Glycoside and Acylated Steryl Glygoside (µg/g) Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  29. Phytosterols in Spelt Combined FS and ES (µg/g) Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  30. Glucosamine • Stimulates the synthesis of proteoglycans, which, in combination with collagen, form the connective tissue destroyed in osteoarthritis. • Anti-inflammatory action ~ 21 millions adults in the USA live with osteoarthritis 25% of visits to primary care physicians and half of all non-steroidal anti-inflammatory prescriptions* * Y. Shao et al J. Pharma. Biomed. Anal. 35 (2004) 625 Dietary supplements as alternative treatment huge business Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  31. Glucosamine Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  32. Glucosamine Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  33. Glucosamine D-Glucosamine chlorhydrate (C6H11O5NH2) . HCl MW 215.6 83.1% of free base D-Glucosamine sulfate (C6H11O5NH2)2 . H2SO4 . 2KCl MW 605.5 59.2% of free base HPLC Methods Difficulties : ► no chromophore ► highly polar ► positively charged at pH 7 Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  34. Glucosamine H2O : CH3CN / 50:50 M+1+CH3CN - H2O M+1 - H2O M+Na+CH3CN Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  35. Glucosamine H2O : MeOH / 50:50 M+1 Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  36. Glucosamine NEUTRAL LOSS SCANNING METHOD TRIPLE QUAD INSTRUMENTS MS1 CID MS2 Precursor m scan Fragmentation Fragment detection m-a scan Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  37. Glucosamine 180 84 Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  38. Glucosamine C18, H2O:MeOH / 50:50 200 µl/min 10 µl Y = 56026 + 1571*x R2 = 1.0000 25 – 25000 ng/ml LOQ ~0.5 ng on column Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  39. 10mg of powder dissolution in 10 ml MeOH:H2O / 50:50 dilution 100 x in PBS Results and Reproducibility : 1 capsule , 5 injections : 224 ± 5% mg/g capsule 1 : 233 mg/g capsule 2 : 231 mg/g capsule 3 : 233 mg/g Glucosamine Commercial capsule Commercial solution 10mg of liquide dilution in 10 ml MeOH:H2O / 50:50 dilution 10 x in PBS 5 analysis : 0.52 mg/g ± 10% Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  40. Detection of carnitines in urine Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  41. Detection of carnitines in urine Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  42. Detection of carnitines in urine FABMS spectrum of crude sample Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  43. Detection of carnitines in urine FABMS2 spectrum of crude sample detection of the precursors of 85 Scanning Selective detection at 85 Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  44. Detection of carnitines in urine : discovery of a new carnitine FABMS2 spectrum of a crude sample detection of the precursors of 85 ? Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  45. ethylsuccinylcarnitine Detection of carnitines in urine : discovery of a new carnitine Daughter spectrum of 290 m/z MS RMN Synthesis Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  46. Detection of carnitines in urine : discovery of a new carnitine Ethylsuccinylc. 2,6-dimethyl- Heptanoylc. Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  47. Short introduction to proteomics Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  48. Short introduction to proteomics separation Protein mixture Proteins digestion digestion separation Peptide mixture Peptides MS analysis MS data database search Separation: always 2D 2D gel or LC - LC Identification Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  49. Short introduction to proteomics pump injector sample SCX column RP column MS system Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

  50. Short introduction to proteomics : mass fingerprinting Digestion Tryptic peptides Peptide mass list … 1529.7348 measured mass … Pr. Jean-Louis Habib Jiwan UCL - Chemistry Department May 23d 2006

More Related