Ch 5 - Organic Analysis Elements and compounds. Solids, liquids, and gases. Phase Organic vs inorganic compounds. Qualitative vs quantitative analysis. Henry’s law and equilibrium state. Chromatography. Retention time.

Thin-layer vs gas chromatography. Rf value. Electrophoresis. Theories of light: wave vs particle. Electromagnetic spectrum. Relationship between color and absorption of light by molecules. Beer’s law. Absorption spectrophotometer. Ultraviolet (UV) and Infrared (IR) spectrum for identification of organic compounds.

Mass Spectrometry (MS) Significance of mass spectrum in forensic science.

Matter: all things of substance. Matter is composed of atoms or molecules. Element: a fundamental particle of matter. An element cannot be broken down into simpler substances by chemical means. Periodic table: chart of elements arranged in a systematic fashion. Vertical rows are called groups or families; horizontal rows are called series. Elements in a given row have similar properties. Compound: a pure substance composed of two or more elements

Physical state: a condition or stage in the physical being of matter; a solid, liquid, or gas Solid: a state of matter in which the molecules are held closely together in a rigid state Liquid: a state of matter in which molecules are in contact with one another but are not rigidly held in place Gas (Vapor): a state of matter in which the attractive forces between molecules are small enough to permit them to move with complete freedom Sublimation: a physical change from the solid directly into the gaseous state

Phase: a uniform piece of matter; different phases are separated by definite visible boundaries Organic: a substance composed of carbon and hydrogen, and, often, smaller amounts of oxygen, nitrogen, chlorine, phosphorus, or other elements Inorganic: a chemical compound not based on carbon Spectrophotometry: an analytical method for identifying a substance by its selective absorption of different wavelengths of light

Chromatography: any of several analytical techniques whereby organic mixtures are separated into their components by their attraction to a stationary phase while being propelled by a moving phase Pyrolysis: the decomposition of organic matter by heat Fluoresce: to emit visible light when exposed to light of a shorter wavelength-i.e., ultraviolet light

Electrophoresis: a technique for the separation of molecules through their migration on a support medium while under the influence of an electrical potential Proteins: polymers of amino acids that play basic roles in the structures and functions of living things Enzyme: a type of protein that acts as a catalyst for certain specific reactions Visible light: colored light ranging from red to violet in the electromagnetic spectrum

Wavelength: the distance between crests of adjacent waves Frequency: the number of waves that pass a given point per second Electromagnetic spectrum: the entire range of radiation energy from the most energetic cosmic rays to the least energetic radio waves X-ray: a high energy, short wavelength form of electromagnetic radiation

Laser: light amplification by the simulated emission of radiation. Light that has all its waves pulsating in unison Photon: a small pocket of electromagnetic radiation energy. Each photon contains a unit of energy equal to the product of Planck’s constant and the frequency of radiation: E= hf Monochromator: a device for isolating individual wavelengths or frequencies of light Monochromatic light: light having a single wavelength or frequency

Ultraviolet: Invisible long frequencies of light beyond violet in the visible spectrum Infrared: invisible short frequencies of light before red in the visible spectrum Ion: an atom or molecule bearing a positive or negative charge

Gas Chromatograph (GC)

Thin-Layer Chromatography (TLC)

Electrophoresis

Electromagnetic Spectrum

UV-VIS Spectrophotometer

UV-VIS Spectrum

IR Spectrophotometer

IR Spectrum

Mass Spectrometer (MS)

Mass Spectrometer

Mass Spectrum

Ch.5 Organic Analysis Chromatography

Separation of Mixtures

What Is Chromatography? A family of laboratory techniques for separating mixtures into their component compounds Uses some version of a technique in which two phases, one mobile, one stationary, flow past one another The mixture separates as it interacts with the two phases

Basic Principle Different compounds will stick to a solid surface with different degrees of strength or vary in the efficiency with which they dissolve in a liquid

Basic Principles A mobile phase sweeps the sample over a stationary phase –like the wind sweeps the swarm over the flower bed

Basic Principles When a mixture of compounds flows over a surface, the molecules will stick to the surface If a molecule does not stick to the surface too strongly, the molecule stick & unstick many times as it is swept along the surface Over time, the molecules will become physically separated from each other

Basic Principles When the molecules reach the far end of the surface, they are detected or measured one at a time as they emerge Chromatography is non-destructive –does not alter the molecular structure of the compounds

Types of Chromatographic Attraction Adsorption Chromatography –depends on physical forces such as dipole attraction to cause the molecules to “stick” to the stationary phase column, TLC, HPLC

Types of Chromatographic Attraction Partition Chromatography –depends on the relative solubility of the mixture’s molecules in the stationary phase coating –polarity may also have some effect gas chromatography

Types of Chromatographic Attraction Size-exclusion –the relative sizes of the molecules determine how fast the molecules move through the stationary phase –large molecules flow right through –small molecules spend time trapped in the pores of the stationary phase gel filtration chromatography

Types of Chromatographic Attraction Ion-exchange –depends on the relative strength with which ions interact with an ionic resin –less strongly held ions are displaced by more strongly attaching ions –one kind of ion is exchanged for another ion exchange chromatography

Paper Chromatography Stationary phase –a sheet or strip of paper mobile phase –a liquid solvent Sample mixture spotted onto the paper Capillary action moves mobile phase through stationary phase

Paper Chromatography Components appear as separate spots spread out on the paper after drying Can be used for ink analysis

Paper Chromatography 2D Chromatography –accomplished by running another chromatography with the paper turned 90 o –Can complete separation of overlapping compounds

Thin Layer Chromatography Stationary Phase –a thin layer of adsorbent coating on a sheet of plastic or glass usually Al 2 O 3 (alumina) or SiO 2 (silica) Mobile Phase –a liquid solvent Sample mixture spotted onto the adsorbent

TLC Some components bind to the adsorbent strongly; some weakly Components appear as separate spots after development

Retention Factor (R f ) quantitative indication of how far a compound travels in a particular solvent good indicator of whether an unknown & a known compound are similar, if not identical –If the R f value for the unknown compound is close to or the same as that for the known compound, the two compounds are most likely similar or identical

Retention Factor (R f ) R f = distance the solute (D 1 ) moves divided by the distance traveled by the solvent front (D 2 ) R f = D 1 / D 2 R f <1.0

Gas Chromatography Stationary phase –a solid or very syrupy liquid lines a tube (column) silicone polymers (like Silly Putty) commonly used Mobile phase –an inert gas nitrogen helium

GC Columns A packed column A capillary column

Schematic of a GC

Retention Time

The time between when the sample is injected & when it exits the column reaching the detector Tm is the time taken for the mobile phase to pass through the column

Analysis Using the GC Retention time can be used as an identifying characteristic of a substance –retention times may not be unique –GC is not an absolute method of identification An extremely sensitive technique –area under a peak is proportional to the quantity of substance present –allows quantitation of sample

Identification of Accelerants unevaporated gasoline 90% evaporated gasoline unevaporated kerosene 90% evaporated kerosene

Pyrolysis Gas Chromatography Used when sample does not readily dissolve in a solvent If heating such sample at high temp ( C) decomposes it into gaseous products, the products can be analyzed by CGC A pyrogram is obtained

High Pressure Liquid Chromatography (HPLC) Stationary Phase –fine solid particles Mobile Phase –a liquid solvent The solvent is pumped through the column The sample components are retarded by different amounts by interaction with the column packing

Schematic of HPLC

Advantages of HPLC Separation occurs at room temperature Does not decompose heat sensitive materials –explosives –heat sensitive drugs like LSD

Analysis of Components of Sake Sake is composed of several chemical components –sugar –organic acids –amino acids Each component relates to the taste of the sake

Sugar & organic acids can be analyzed by HPLC

Spectroscopy Compound Identification

Wave Nature of Light Wavelength –distance between crests Frequency –number of crest that pass a given point in one second

Wave Nature of Light Energy –frequency & energy are proportional

The Electromagnetic Spectrum

The Hydrogen Atom The electron structure of an atom is quantized –electrons can only exist in discrete energy levels

Excitation When a “packet” of energy equal to the energy difference between two energy levels is absorbed –electron is promoted

Excitation Excitation causes wavelengths to be absorbed & removed

Deexcitation As the electrons falls back to the ground state –an energy “packet” is emitted

Deexcitation Deexcitation causes wavelengths to be emitted

Visible Spectroscopy

The amount of light absorbed depends on the number of ions or molecules present

Beer’s Law Absorption is proportional to concentration

Example Determination of the wavelength of light absorbed by a sample of grape soda Determination of the amount of dilution of a sample of grape soda

Absorption of Grape Soda

Dilution of Grape Soda

UV-VIS Spectrophotometry Can give information about the structure of the compound(qualitative info) –positions of the absorption maxima Can quantitate the sample –Beer’s Law analysis

Electromagnetic Radiations Each type of electromagnetic radiation is composed of wavelengths possessing a range of energies related to the frequency of the wave Each type of electromagnetic radiation will cause different types of excitation in a sample of matter –dependent on the amount of energy the wave possesses

IR Radiation Exposing molecules to the correct frequencies of infra-red light will result in some of that light being absorbed –gives that molecule more energy –results in more energetic motion of the atoms in the molecule

IR Probes different molecular vibrations –absorption occurs when the frequency of the IR wave matches a vibrational frequency of the molecule Most molecules have numerous vibrations –bond stretching vibrations detect different kinds of A-B bonds – bond bending involves several atoms at once

Molecular Motions

Cocaine

The Mass Spectrometer A detector Allows the identification of a chemical compound In the MS, a compound is bombarded with a stream of electrons –compound breaks into fragments –each compound gives a unique set of fragments “fingerprint”

GC/MS As individual compounds elute from the GC column, they enter the MS detector Fragmented by electron bombardment –fragments are charged ions with a certain mass –mass to charge ratio (M/Z) Z is usually +1 M/Z represents that molecular weight of the fragment