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2Massspec - Ch 2 Objectives Mass Spectrometry To learn the...

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2-1 Ch. 2 Mass Spectrometry Objectives To learn the principle of operation of a mass spectrometer To be able to interpret the mass spectra of atoms and small molecules containing isotopes To learn how mass spectrometry can be applied to biomolecules such as proteins To use mass spectrometry to observe the difference between normal adult hemoglobin and sickle cell hemoglobin To learn the principle of peptide mass mapping to determine the identity of a protein from the mass spectrum Introduction A mass spectrometer is an instrument that is used to measure atomic and molecular mass directly. This instrument was developed in the 1920’s and has been one of the most widely used analytical techniques because of its versatility. A mass spectrometer is capable of providing information about isotopic ratios of atoms in samples, qualitative and quantitative composition of inorganic and organic analytes in complex mixtures, and structures in a wide variety of molecules. Recent discoveries in the field of mass spectrometry have also enabled the analysis of large biomolecules, such as proteins. The major components of any mass spectrometer are shown in Figure 1. The purpose of the inlet system is to introduce a very small amount of sample (typically < 1 micromole) into the instrument and convert the sample to the gaseous phase. The ion source of the mass spectrometer converts the components of the sample into ions by bombardment with electrons, ions, molecules, or photons. The mass analyzer functions to disperse the ions based on mass to charge ratio and the detector quantifies the amount of ion with a given mass to charge ratio by measuring a current. Inlet System Ion Source Mass Analyzer Detector Vacuum (10 -5 10 -8 torr)
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Chapter Three 2-2 Figure 1: Simplified components of a mass spectrometer. There are several different types of mass spectrometers, but in this experiment we will describe only two different designs. The first is a simple design based on the deflection of ions in a magnetic field. Figure 2 shows a schematic of this instrument. The sample is introduced into the instrument and vaporized (by heating) if it is not already in the gaseous state. It is then ionized by collisions with high-energy electrons produced from a heated filament. Collisions with the electrons result in positively charged ions of the analyte (A). A (g) + e - A + (g) + 2e - Electron impact ionization is carried out with electron energies that are high enough to break the covalent bonds within a molecule, resulting in molecular fragments. Fragmentation is often useful in deducing the structure of the parent molecule. All the ions are then moved through the mass spectrometer by an electrostatic potential. They are focused into a narrow beam before passing through a magnetic field, which deflects the ions by varying amounts depending on their mass to charge ratio. Since most of the analyte atoms or molecules acquire a +1 charge in the ionization process, the end result is that the ions are separated spatially by mass.
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