INSTRUMENTAL ANALYSIS CHEM 4811

INSTRUMENTAL ANALYSIS CHEM 4811 CHAPTER 9 DR. AUGUSTINE OFORI AGYEMAN Assistant professor of chemistry Department of natural sciences Clayton state university

CHAPTER 9 MASS SPECTROMETRY I PRINCIPLES AND APPLICATIONS

PRINCIPLES Technique involves - Creating gas phase ions from the analyte atoms or molecules - Separating the ions according to their mass-to-charge ratio (m/z) - Measuring the abundance of the ions

PRINCIPLES Technique can be used for - Qualitative and quantitative analysis - Providing information about the mass of atoms and molecules - Molecular structure determination (organic & inorganic) - Identification and characterization of materials

PRINCIPLES - Instrument is mass spectrometer - Separates gas phase ionized atoms, molecules, and fragments of molecules - Separation is based on the difference in mass-to-charge ratio (m/z) m = unified atomic mass units (u) 1 dalton (Da) = 1 u = 1.665402 x 10-27 kg z = charge on the ion (may be positive or negative)

PRINCIPLES - Analyte molecule can undergo electron ionization M + e- → M●+ + 2e- - M●+ is the ionized analyte molecule called molecular ion - Radical cation is formed by the loss of one electron - Computer algorithms are used to deconvolute m/z values of multiply charged ions into the equivalent mass of singly charged ion - Permits easy determination of molecular weight of analyte

THE MASS SPECTRUM - A plot of relative abundance vs m/z - The most abundant peak is known as the base peak - The base peak is scaled to 100 - Spectrum shows fragmentation patterns - The m/z values and the fragmentation pattern are used to determine the molecular weight and structure of organic compounds - Provides the accurate mass of a given isotope not the weighted average

RESOLVING POWER - The ability of a mass spectrometer to separate ions of two different m/z values - Resolving power = M/∆M - M = mass of one singly charged ion - ∆M = difference in mass between M and the next m/z value - The resolving power of ions in the 600 range = 600 - The resolving power of ions in the 1200 range = 1200

RESOLVING POWER - Two methods used to calculate ∆M - Full width at half maximum (FWHM) = ∆M - 10% valley (overlap should not be > 10%) RESOLUTION - The value of ∆M at a given M - Expressed in ppm

INSTRUMENTATION Main components of the mass spectrometer - Sample input system - Ionization source - Mass analyzer - Detector - Vacuum pumps - Computer based data acquisition and processing system

SAMPLE INPUT METHODS Gas Expansion - Useful for gas samples and liquids with sufficiently high vapor pressures - The gas or vapor expands into an evacuated and heated vessel - Sample leaks through holes in a gold foil seal into the ionization source (termed molecular leak inlet) - Pressure in ionization is maintained at 10-6 – 10-8torr

SAMPLE INPUT METHODS Direct Insertion Probe - For liquids with high boiling points and solids with sufficiently high vapor pressure - The probe (with the sample in a glass capillary at the tip) is inserted into the ionization source - The probe is electrically heated to vaporize the sample - This method has a problem with contamination

SAMPLE INPUT METHODS Direct Exposure Probe - Sample is first dissolved in a solvent - A drop of solution is placed at the rounded glass tip of the probe - Solvent evaporates leaving a thin film of sample - The tip is inserted into the ionization source and heated to vaporize sample - Less likely to be contaminated

SAMPLE INPUT METHODS Chromatography and Electrophoresis Systems - Chromatographic instruments are used to separate mixtures of gases and liquids - Separated components are introduced into a mass spectrometer for detection - The GC-MS system - LC-MS system is used for nonvolatile organic compounds - Capillary electrophoresis (CE) can also be coupled to MS

IONIZATION SOURCES Electron Ionization (EI) - Commonly used for analysis of organic samples - Electrons are emitted from a heated tungsten filament cathode - Electrons are accelerated towards the anode with a potential of about 50 – 100 V - Electrons meet at right angles with the sample molecules - Interaction with the high energy electrons causes ionization of sample molecules and fragmentation into smaller ions

IONIZATION SOURCES Electron Ionization (EI) - Referred to as hard ionization source due to the high energy EI source - Ions are accelerated into the mass analyzer by an accelerating voltage of ~ 104 V - Both negative and positive ions are formed by EI - Negative ions form from molecules containing acid groups or electronegative atoms

IONIZATION SOURCES Electron Ionization (EI) - Collision between ions and molecules may also result in ions with higher m/z values than the molecular ion An example is the (M+1) peak - Reaction between analyte molecule and H+ to form MH+ or (M+H)+ in which charge equals a+1 - Low pressure in the ionization source minimizes reaction between ions and molecules

IONIZATION SOURCES Chemical Ionization (CI) - A large excess of reagent gas (1000 – 10000 times) is introduced into the ionization region - Pressures in source are typically higher than EI - Electrons are allowed to bombard the gas-sample mixture Examples of reagent gas - Methane, ammonia, isobutane

IONIZATION SOURCES Chemical Ionization (CI) - Reagent gases are much more likely ionized by the electrons than sample due to large excess - Sample molecules are subsequently ionized by collision with ionized reagent gas molecules - Considered soft ionization source - Less fragmentation and molecular ion is much more abundant - Combination of CI and EI spectra provide good interpretation

IONIZATION SOURCES Chemical Ionization (CI) - For methane reagent gas Proton transfer occurs when sample molecules collide with

IONIZATION SOURCES Chemical Ionization (CI) The following may occur if analyte is a saturated HC

IONIZATION SOURCES Atmospheric Pressure Ionization (API) Sources - Two major types Electrospray Ionization (ESI) and Atmospheric Pressure Chemical Ionization (APCI) - Operate at atmospheric pressure - Modified version of ESI is the Ion Spray Source - Used for mixtures of nonvolatile high molecular weight compounds

IONIZATION SOURCES Atmospheric Pressure Ionization (API) Sources Applications - Pharmaceutical chemistry - Biochemistry - Clinical biomonitoring Electrospray - Fine spray of positively or negatively charged droplets

IONIZATION SOURCES Desorption Ionization - For direct ionization of solids - Excellent tool for analysis of large molecules - Solid samples are placed on a support and then bombarded with ions or photons - Different types are available

IONIZATION SOURCES Desorption Ionization Desorption Chemical Ionization - Used for nonvolatile compounds - Sample is directly introduced into the chemical ionization source on a tungsten or rhenium wire Secondary Ion Mass Spectrometry (SIMS) - For surface analysis - For large molecules

IONIZATION SOURCES Desorption Ionization Laser Desorption Ionization - Uses pulsed laser - Provides selective ionization by choosing appropriate λ - Laser is focused on a solid surface to ionize material Examples of Lasers - IR laser: CO2 laser - UV laser: Nd:YAG (yttrium aluminum garnet)

IONIZATION SOURCES Desorption Ionization Matrix-Assisted Laser Desorption Ionization (MALDI) - Matrix disperses large amounts of energy absorbed by the laser - Minimizes fragmentation of the molecule - Permits analysis of molecular weight over 10,000 Da - Used for study of polymers, proteins, peptides

IONIZATION SOURCES Desorption Ionization Matrix-Assisted Laser Desorption Ionization (MALDI) Matrix - must be stable in vacuum and not react chemically - must absorb strongly at laser λ (where analyte absorbs weakly) Examples - IR region : urea, alcohols, carboxylic acids - UV region: 3-hydroxypicolinic acid, 5-chlorosalicylic acid

IONIZATION SOURCES Desorption Ionization Fast Atom Bombardment (FAB) - Employs fast moving neutral inert gas atoms (Ar) to ionize large molecules - Sample is dissolved in glycerol and spread in a thin layer on a metal probe - Probe is then inserted into the mass spectrometer and a beam of fast moving atoms probe the surface

IONIZATION SOURCES Desorption Ionization Fast Atom Bombardment (FAB) - Used for analysis of surfactants and proteins (MW > 10,000) - For large and thermally unstable molecules - Technique works well at room temperature - Simple and high sensitivity - Sample can be recovered

IONIZATION SOURCES Desorption Ionization Fast Atom Bombardment (FAB) - Modified technique is the continuous flow FAB (CFFAB) - Sample introduction is through a fused silica capillary tube - Solvent flows continuously and sample is introduced by continuous flow injection - For analysis of blood, urine, other body fluids, waste water

IONIZATION SOURCES Inorganic MS Ionization Sources Solid Samples - Glow Discharge (GD) and Spark sources - For sputtering and ionizing species from solid surfaces - Primarily for atomic mass determination of elements - GD has better S/N and able to sputter more material from sample

IONIZATION SOURCES Inorganic MS Ionization Sources Liquid Samples - Inductively coupled plasma (ICP) - Has high ionization efficiency - Provides very simple mass spectra

MASS ANALYZERS - Differentiates ions according to their m/z - Different designs are available Scanning Instruments - Only ions of a given m/z pass through the analyzer at a given time - Magnetic Sector Mass Analyzer - Quadrupole Mass Analyzer

MASS ANALYZERS • Simultaneous Transmission Instruments • - Allow transmission of all ions at the same time • - Time-of-flight (TOF) • - Ion Trap • - Ion Cyclotron Resonance Mass Analyzer • Dispersive Magnetic Mass Analyzer • Tandem Mass Spectrometer (MSn) • - Two or more mass analyzers in sequence

MAGNETIC SECTOR MASS ANALYZER - Gas phase molecules are ionized by a beam of high energy electrons - Electrons may be ejected from molecules (ionization) or bonds in molecules may rapture (fragmentation) - Ions are then accelerated in a field (sector) at a voltage V - Sector can have any apex angle (60o and 90o are common) - Most modern instruments combine both electric sector and magnetic sector (double-focusing MS)

MAGNETIC SECTOR MASS ANALYZER - The electric sector acts as an energy filter - m/z range is 1 – 1400 for single-focusing and 5,000 – 10,000 for double-focusing instruments - Energy of each ion = zV - Kinetic energy depends on charge and voltage but not on mass of ion - Ions with small masses must travel at a higher velocity than ions with larger masses

MAGNETIC SECTOR MASS ANALYZER - For single positively charged ions m = mass of ion v = velocity of ion z = charge of ion V = accelerating voltage - V changes as m varies such that ½ mv2 is constant

MAGNETIC SECTOR MASS ANALYZER - Ions enter a curved section of a homogeneous magnetic field B after acceleration - Ions move in a circle with radius r - Attractive force on magnet = Bzv - Centrifugal force on the ion = mv2/r - The two forces are equal if the ion follows the radius of curvature of the magnet

MAGNETIC SECTOR MASS ANALYZER Substituting for v and rearranging gives

MAGNETIC SECTOR MASS ANALYZER - Radius of circular path depends on m/z if V and B are kept constant - Ions with different m/z travel in circles with different radii - Basis of separation by m/z - Ions with the right m/z reach the detector and others hit the sides of the instrument and be lost - Which m/z to reach the detector can be selected by varying V or B - B is varied and V is kept constant in modern instruments

TIME OF FLIGHT (TOF) ANALYZER - Makes use of a drift tube - Pulses of ions are accelerated into the an evacuated drift tube (free of field or external force) - Velocity of an ion depends on m/z (depends on mass if all ions have the same charge) - Lighter ions move faster along the tube than heavier ions - Ions are separated in the drift tube according to their velocities (v)

TIME OF FLIGHT (TOF) ANALYZER - V = accelerating voltage - If L is the length of tube (typically 1-2 m) and t is the flight time of ion, then v = L/t - Implies mass-to-charge ratio and flight time can be found from - An ion mirror called a reflectron is used to increase resolution

QUADRUPOLE MASS ANALYZER - Separates ions in an electric field (the quadrupole field) - Field is varied with time - Oscillating radio frequency (RF) voltage and a constant DC voltage are used to create the field - These are applied to four precisely machined parallel metal rods - The result is an AC potential superimposed on a DC potential - Ion beam is directed axially between the four rods

QUADRUPOLE MASS ANALYZER - Opposite pairs of rods are connected to opposite ends of a DC source - Ions follow an oscillating (corkscrew) path through the quadrupole to the detector - For a given ratio of DC to RF at a fixed frequency, only ions of a given m/z value will pass through the quadrupole - Other ions with different m/z values will collide with the rods and be lost

QUADRUPOLE MASS ANALYZER - The quadrupole acts as a filter so is often called the mass filter - Sample must be ionized and in the gas phase - m/z range is 1 – 1000 Da - Has smaller range and lower resolution than magnetic sector but faster - Is the most common analyzer - Rugged, inexpensive, and compact

MS – MS (TANDEM MS) INSTRUMENTS - Employs two or more stages of mass analyzers - Example is two quadrupoles coupled in series - First analyzer selects ion (precursor ion) and second analyzer selects the fragments of the precursor ion - Used to obtain more information about the structure of fragment ions - Fragment ions may be dissociated into lighter fragment ions or converted into heavier ions by reaction with neutral molecule

ION TRAP - A device in which gaseous ions are formed and/or stored for periods of time Two commercial types - Quadrupole Ion Trap (QIT) and - Ion-Cyclotron Resonance Trap (ICR)

ION TRAP Quadrupole Ion Trap (QIT) - Also called Paul Ion Trap - Uses a quadrupole field to separate ions - A 3D field is created using a ring-shaped electrode between two end cap electrodes - A fixed frequency RF voltage is applied to the ring electrode - The end cap electrodes are either grounded or under RF or DC voltage

ION TRAP Quadrupole Ion Trap (QIT) - Ions are stored in trap by moving in trajectories between electrodes - This is done by changing signs of electrodes to repel ions as they approach the electrodes - Ions of a given m/z pass through an opening to the detector when the RF of the ring electrode is changed - m/z range is 10 – 1000 Da

INSTRUMENTAL ANALYSIS CHEM 4811