Mass Spectrometry PDF

Biotechniques (BMS 34010A) Fall semester 2023 -2024 Dr. Tania Tahtouh [email protected] Mass spectrometric techniques Learning outcomes:  Define mass spectrometry and describe its principle.  List the components of a mass spectrometer.  Describe what happens in a mass spectromet...

Biotechniques (BMS 34010A) Fall semester 2023 -2024 Dr. Tania Tahtouh [email protected] Mass spectrometric techniques Learning outcomes:  Define mass spectrometry and describe its principle.  List the components of a mass spectrometer.  Describe what happens in a mass spectrometer.  Explain some applications. ▪ Mass spectrometry | Atomic structure and properties | AP Chemistry | Khan Academy: https://youtu.be/myolF-h1kKI ▪ Mass Spectrometry: https://youtu.be/mBT73Pesiog ▪ Spectrometry | Chemical Tests | Chemistry | FuseSchool: https://youtu.be/sTi--ixdAME ▪ Flashcards: https://www.cram.com/flashcards/five-stages-in-mass-spectrometry-724750  Mass spectrometry  Mass spectrometry (mass spectroscopy) is an analytical tool useful for measuring the mass-to- charge ratio (m/z or sometimes m/e) of one or more molecules present in a sample. ▪ These measurements can often be used to calculate the exact molecular weight of the sample components.  Mass spectrometry can be used to: ▪ Identify unknown compounds via determination of the molecular weight and elemental composition of a sample or molecule. ▪ Quantify known compounds. ▪ Determine structure and chemical properties of molecules.  The instruments used in such studies: ▪ Mass spectrometers: moving ions are detected electrically. ▪ Mass spectrographs: moving ions detected by photographic or other nonelectrical means.  Principle  In this technique, molecules are bombarded with a beam of energetic electrons.  The molecules are ionized and broken up into many fragments, some of which are ions.  If something (e.g. ions) is moving and you subject it to a sideways force, instead of moving in a straight line, it will move in a curve - deflected out of its original path by the sideways force.  The amount of deflection you will get for a given sideways force depends on the mass of the ion.  If you know the speed of the ion, the size of the force, and the deflection, you can calculate the mass of the ion.  LTQ XL Linear Ion Trap Mass Spectrometer (LITMS)  Every mass spectrometer consists of at least these three components: ▪ Ionization source. ▪ Mass filter/analyzer. ▪ Ion detection system.  What happens in a mass spectrometer? 1- Ionization 2- Acceleration 3- Deflection 4- Detection  Ionization source/chamber  Samples are converted to gas-phase ions so that they can be easily moved. The atom or molecule is bombarded with a stream Detector of electrons and ionised by knocking one or more electrons off to Mass filter/analyzer give a positive ion. Most mass spectrometers work with positive ions. This is true even for things which you would normally expect to form negative ions (chlorine, for example) or never form ions at all (argon, for example). Ionization source  Mass filter/analyzer  The mass analyzer is the heart of the mass spectrometer. It takes ionized masses and uses magnetic field to separate them based Mass filter/analyzer Detector on mass-to-charge ratios. ▪ Once the ions enter the mass analyzer, they are exposed to the magnetic field. ▪ Different ions are deflected by the magnetic field by different amounts. Ionization source  The amount of deflection depends on: ▪ The mass of the ion: lighter ions are deflected more than heavier ones. ▪ The charge on the ion: ions with 2 (or more) positive charges are deflected more than ions with only 1 positive charge.  Ion detection system  When an ion hits the metal box, its charge is neutralized by an electron jumping from the metal on to the ion. Mass filter/analyzer Detector  A flow of electrons is detected as an electric current which can be amplified and recorded.  The current will be proportional to the number of ions arriving. Ionization source  Calibration Mass filter/analyzer Detector Mass filter/analyzer Detector Mass filter/analyzer Detector Ionization source Ionization source Ionization source These ions collide with the walls where they will pick up electrons and be neutralised. They get removed from the mass spectrometer by the vacuum pump.  Mass spectrometer output  The output from the chart recorder is usually simplified into a "stick diagram".  It shows the relative current produced by ions of varying mass/charge ratio. Interpreting spectra  One molecule can show expression at multiple m/z values due to isotopical peaks. Isotopes are atom species of the same chemical element that have different masses, caused by a different number of neutrons.  Molybdenum consists of 7 different isotopes: the commonest ion has a mass/charge ratio of 98; other ions have mass/charge ratios of 92, 94, 95, 96, 97 and 100. Interpreting spectra If there were also 2+ ions present, there would be lines at exactly half its m/z value. Those lines would be much less tall than the 1+ ion lines because the chances of forming 2+ ions are much less than forming 1+ ions.  Interpreting spectra  One peak could indicate several biomolecular ions: ▪ Isomers are organic compounds with the same molecular formula (= same mass) but different structures, and may have completely different chemical properties. ▪ Distinct molecules may be closer to each other in mass-to-charge ratio than the mass spectrometer can distinguish (mass resolution).  Applications  Environmental monitoring and analysis (soil, water and air pollutants, water quality, etc.).  Determination of pesticides residues in food.  Drug testing and discovery. Identification of drugs abuse and metabolites of drugs of abuse in blood.  Quality control in chemical and petrochemical industry.  Geochemistry: age determination, soil and rock composition, oil and gas surveying.  Determination of molecular mass of peptides, proteins, and oligonucleotides.  Identification of biomolecule sequences & structures. References  THE MASS SPECTROMETER. https://www.chemguide.co.uk/analysis/masspec/howitworks.html  Introduction to mass spectrometry data analysis. https://www.aspect- analytics.com/media/blog/2020-05-30-introduction-to-mass-spectrometry- data-analysis/  Mass Spectrometry Applications Areas. Thermo Fisher Scientific. https://www.thermofisher.com/ae/en/home/industrial/mass- spectrometry/mass-spectrometry-learning-center/mass-spectrometry- applications-area.html