Forensic files with GC use: Deadly formula

Organic and Inorganic Chemical Analysis Chapter 5 and 6 Organic and Inorganic Chemical Analysis

Ch 5/6: Chemical Analysis (Analytical Methods) Elements and compounds Solids, liquids, and gases (phase changes) Organic vs. inorganic compounds Qualitative vs. quantitative analysis Chromatography Retention time and Rf value Electrophoresis Spectrophotometry and spectrometry

Phase Changes: (physical state changes) Melting: change from the solid directly into the liquid state Freezing: change from the liquid directly into the solid state Vaporization: change from the liquid directly into the gaseous state Condensation: change from the gas directly into the liquid state Sublimation: change from the solid directly into the gaseous state Deposition: change from the gas directly into the solid state

Phase Diagrams:

Matter Pure Substance Mixture Compound Element Homogeneous Uniform Composition? Heterogeneous Can be separated by physical methods Pure Substance Can it be broken down further ? Compound Element Yes No Homogeneous (solution) Mixture

Selecting an Analytical Technique  Organic: a substance composed of carbon and hydrogen (often contain smaller amounts of oxygen, nitrogen, chlorine, phosphorus, or other elements)  Inorganic: a chemical compound not based on carbon

Quantitative or qualitative required Questions to consider in choosing an analytical (chemical) method: Quantitative or qualitative required Sample size and sample preparation requirements What level of analysis is required (e.g., ± 1.0% or ± 0.001%) Detection levels Destructive or non-destructive Availability of instrumentation Admissibility

What Is Chromatography? Laboratory technique 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

Chromatography Chromatographic systems have a stationary phase (can be solid or liquid) and a mobile phase (usually liquid or gas). The mixture is placed at the beginning of the chromatographic system ( on the stationary phase). The mobile phase then “pushes” the components of the mixture through the system. Each component adsorbs on the stationary phase with a different strength (stronger means moves more slowly through the system). Each component comes out the end of the system at a different time (retention time).

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

Street Drugs in Real Time Amphetamine Methamphetamine and MDMA Hydrocodone Cocaine Oxycodone

Thin Layer Chromatography (TLC) Gas Chromatography (GC) Types of Chromatographic Separation Paper Chromatography Thin Layer Chromatography (TLC) Gas Chromatography (GC) High Performance Liquid Chromatography (HPLC)

Paper Chromatography Stationary phase: a sheet or strip of paper Mobile phase: a liquid solvent (substances must be soluble in the solvent) Mixture is spotted onto the paper Capillary action moves mobile phase across the stationary phase Components appear as separate spots spread out on the paper after drying

Thin Layer Chromatography (TLC) Stationary Phase: a thin layer of adsorbent coating on a sheet of plastic or glass Mobile Phase: a liquid solvent Sample mixture spotted onto the adsorbent material - Solids must first be dissolved - Liquids can be directly applied Some components bind to the TLC plate strongly, others weakly Components appear as separate spots after development

TLC

Measuring Retention Factor (Rf) Quantitative indication of how far a compound travels in a particular solvent Good indicator of whether an unknown and a known compound are similar, if not identical Rf = distance the solute (D1) moves divided by the distance traveled by the solvent front (D2) Rf = D1 / D2

Gas Chromatography (GC) Stationary phase: a solid or very syrupy liquid lines a tube (column) Mobile phase: an inert gas ( also called a carrier gas) Usually nitrogen or helium GC Columns

Schematic of a GC A mixture is injected into the GC and is vaporized The carrier gas “pushes” the mixture through a GC column, where the compounds become separated and enters a detector. Detectors measure separation as a function of time. Each peak corresponds to a component

GC Analysis Retention time can be used as a characteristic of a substance BUT may not be unique An extremely sensitive technique allows quantitation of sample area under a peak is proportional to the quantity of substance present Retention time: time between when the sample is injected and when it exits the column reaching the detector

Retention Time tm is the time it takes for the mobile phase to pass through the column

Pyrolysis Gas Chromatography Used when a sample does not readily dissolve in a solvent (paint chips, fibers)

High Performance Liquid Chromatography (HPLC) Stationary phase: fine solid particles that are chemically treated packed into a column Mobile phase: liquid solvent pumped through the column Advantage: takes place at room temperature Used for organic explosives that are heat sensitive as well as heat sensitive drugs

Underlying Ideas - Atomic and Molecular Weights Mass Scale Underlying Ideas - Atomic and Molecular Weights Uses Atomic Mass Scale Most elements in nature exist as mixtures of isotopes (atoms of an element that have different numbers of neutrons but same number of protons).

Mass Spectrometry (MS or mass spec) Basic Principle: Creates charged particles (ions) Separation of ions –according to mass-to-charge ratio Detection of ions The differences in mass spectrometer types are the different ways they carry out these three functions The MS analyzes ions to provide information about the molecular weight of the compound and its chemical structure.

Mass Spectrometer (MS) As each gas particle leaves the GC, it enters the mass spec A beam of electrons is “shot” at the substance breaking it down into fragments and giving them a charge These fragments pass through an electric field which separates them by their masses The fragment masses are then recorded on a graph Each substance breaks down into its own characteristic pattern

Mass Spectrometer Cl C P Atomic Spectra 35Cl: 75% abundant Spectrum Mass Spectrum Mass Spectrum Int. Int. Int. Cl C P 35 31 12 37 13 mass number (amu) mass number (amu) mass number (amu) 35Cl: 75% abundant 37Cl: 24% abundant 12Cl: 98.9% abundant 13Cl: 1.11% abundant 31P: 100% abundant

Allows for identifying chemical substance Under carefully controlled conditions, no two substances produce the same fragmentation pattern! Allows for identifying chemical substance Unknown white powdery substance ingested by unconscious patient. What do you do? Is it Heroin, Cocaine, Caffeine? Mass Spectrum of Unknown Compound

Mass Spectrometer MS Library Heroin MS of Unknown

MS Library Cocaine MS of Unknown

MS Library Caffeine MS of Unknown

Mol. Mass = 194 Mass Spectrum Caffeine

GC-Mass Spectrometry

Electrophoresis Separates materials based on their migration rates on a stationary solid phase Passes an electrical current through and allows for classification of proteins

Most useful applications of Electrophoresis Characterization of proteins and DNA in dried blood Proteins migrate at speeds that vary according to their electrical charge and size resulting in characteristic band patterns

Spectroscopy and Spectrophotometry Based on the study of absorption of light by chemical substances Used for identification of various organic materials or for presence of trace elements Electromagnetic spectrum – entire range of “light waves” Visible Light (colors) Ultraviolet or infrared radiation (either side of visible region) X-ray – high energy, short wavelength

Photon - small packet of electromagnetic energy that carries a quantum Each photon contains a specific amount of energy E = hν E = energy of a photon (Joules) ν = frequency of radiation (the number of waves that pass per second) h = Plank’s constant (6.626x10-34 Js)

Atomic Emission Spectroscopy (AES) Used to detect the types of elements present in a sample Can use measurement of the emissions from excited atoms to determine concentration. The Hydrogen Discharge Tube H2 molecules are split into excited H atoms by an electric discharge As the atoms return to lower energy states, light is emitted

Flame Tests Atomic Emission

Atomic Absorption Spectroscopy (AAS) Atoms exposed to radiation emitted from a discharge tube Atoms absorb radiation and become excited The amount of radiation absorbed is recorded Usually used to determine the presence of specific elements

Beer’s Law: absorption is proportional to the concentration The amount absorbed is determined based on a calibration curve

Absorption of Grape Soda Example: Determination of the wavelength of light absorbed by a sample of grape soda Absorption of Grape Soda

Example: Determination of the amount of dilution of a sample of grape soda

Atomic Absorption (AAS) Example: A child becomes ill and is taken to the hospital It is found that the child is suffering from possible lead poisoning. A forensic laboratory is contacted and asked if it can determine the source of the lead which the child has ingested. Paint samples from a number of objects are collected Paint on the child's crib, paint from his toys, and paint from the child's swing are sent to the laboratory. Each sample undergoes AAS to detect if lead is present

The Spectrophotometer Instrument used to measure and record the absorption spectrum of a chemical substance (compounds)

UV Spectrophotometry Measures absorbance of ultraviolet and visible light of a compound as a function of wavelength or frequency Allows tentative identification Ex. White powder with UV spectrum comparable to that of heroin results in a tentative identification

UV-VIS Spectrum

Infrared (IR) Spectrophotometry Different materials always have distinctively different infrared spectra Each IR spectrum is therefore equivalent to a “fingerprint” of that substance and no other Extensive catalogue of IR spectra of organic compounds allows for identification of organic substances

IR Spectrum

Neutron Activation Analysis (NAA) Neutrons interact with a target nucleus to form a compound nucleus in an excited state. The compound nucleus will decay into a more stable configuration through emission of one or more gamma rays. This new configuration may yields a radioactive nucleus which also decays by emission of delayed gamma rays, but at a much slower rate according to the unique half-life of the radioactive nucleus. Quantitation in parts per billion but requires a nuclear reactor

Neutron Activation Analysis Rate depends on half-life of isotope Prompt gamma ray formation measurement taken during irradiation Delayed gamma ray formation measurements taken after irradiation more common About 70% of elements have properties suitable for measurement by NAA

Gamma-ray Spectra Continuation of medium- & long-lived elements

Comparison of Techniques Qual.* or Quant. Sample Size Detection levels Destructive Instr. Avail. Mass Spec. Qual. 0.1 mL to 10-8 mL * Yes Easy Infrared 0.001 g No UV-visible AES Quant. 10-4 g/L Moderate AAS NAA 1 x 10-9 g Possibly Difficult * Primary use is in qual. analysis, although it can be used quantitatively in some cases.