Chem 551 :Instrumental Methods of Analysis

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Chem 551 Instrumental Methods of Analysis
Ralph Allen
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Instrumental Analysis

• There is much more than the Instrument
• Consider forensic analysis

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Analytical chemistry and the fight against
crime and terrorism



FBI National Academy
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First, there was one.
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CSI is coming to life as a video game USA
TODAY
Now, it's everywhere!

• Forensic Files (Court TV)
• I, Detective (Court TV)
• Investigative Reports (AE)
• American Justice (AE)
• Cold Case Files (AE)
• Justice Files (Discovery)
• The F.B.I. Files (Discovery)
• The New Detectives (Discovery)
• Historys Crimes and Trials (History)
• FBI The Untold Stories (History)
• Secrets of Forensic Science (TLC)
• Medical Detectives (TLC)
• Secrets of the Dead (PBS)
• NClS

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What do you know about the criminal justice
system?A NothingB What I learned
from televisionC First hand experience
(arrested )
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Federal Rules of Evidence

• Rule 104 Questions of admissibility generally -
  Qualification of the person to be a witness, or
  admissibility of evidence. Relevance conditioned
  on fact.
• Rule 403 Although relevant evidence may be
  excluded if probative value is substantially
  outweighed by danger of unfair prejudice,
  confusion, misleading the jury, or needless
  presentation of cumulative evidence.

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Federal Rules of Evidence

• Rule 702 If scientific, technical, or other
  specialized knowledge will assist the trier of
  fact, a witness qualified as an expert by
  knowledge, skill, experience, training, or
  education may testify.
• Rule 703 The facts or data in a particular case
  upon which an expert bases an opinion may be
  those perceived by or made known to the expert
  at, or before, the hearing. If reasonably relied
  upon, the facts or data need not be admissible in
  evidence.

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The Frye Rule

• , the thing from which the deduction is made
  must be sufficiently established to have gained
  general acceptance in the particular field in
  which it belongs.

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Daubert Rule

• Does the theory or technique involve testable
  hypotheses?
• Has the theory or technique been subject to peer
  review and publication?
• Are there known or potential error rates and are
  there standards controlling the techniques
  operation?, and
• Is the method or technique generally accepted in
  the scientific community?

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ENVS 470

• One of my other jobs is as Director of
  Environmental Health and Safety
• Measuring pollutants emitted by UVa in the air,
  water, or shipped out as waste

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Instrumental Analysis

• There is much more than the Instrument
• You are the analyst

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What do you do with

• This white powder (is it an illegal drug or
  anthrax or what)
• This sludge from the powerplant in Tennessee

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Could you take a look at this white powder?You
are the analyst. What questions would you ask???
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Why are you taking this class?A an easy AB
interestingC to learn
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What do you want to learn?
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What do I expect of you?

• Attend class on time (you will be allowed only 3
  late or missed classes)
• Pay attention (no games on your computers or
  reading Cav Daily)
• Attendance will be taken early with iClickers
• No text so your only expense is the Clicker

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What do I expect of you?

• You will form a group with people that you can
  meet with on a regular basis
• Your group will make at least one presentation to
  help guide the class through some subject
• You will have a special project to present orally
  and with a written report

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Analytical Chemistry

• art of recognizing different substances
  determining their constituents, takes a prominent
  position among the applications of science, since
  the questions it enables us to answer arise
  wherever chemical processes are present.
  
• 1894 Wilhelm Ostwald

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For this course

• Why bother doing any kind of analysis unless you
  are trying to learn something

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Off flavor cake mix (10)

• Send it off for analysis
• Do simple extractions
• Separation and identification by GC/MS
• Over 100 peaks but problem was in a valley
  between peaks (compare)
• Iodocresol at ppt
• Eliminate iodized salt that reacted with food
  coloring (creosolmethyl phenol)

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Techniques

• mass spectrometry
• NMR
• spectroscopy (UV, IR, AA)
• chromatography (GC, HPLC)
• measure radioactivity, crystallography, PCR, gas
  phase analysis

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You dont need a course to tell you how to run an
instrument

• They are all different and change
• Most of you wont be analysts
• We will talk about experimental design
• Learn about the choices available and the basics
  of techniques

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What I think you will learn

• analytical process and skills
• tools for research
• solve practical problems
• medical uses (including DNA)
• how instruments work and general concepts
• environmental and forensic applications
• new advances

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Organized as teams --but you are responsible for
your learning!

• Your project will be to take a crime scene and
  determine what physical evidence you would have,
  and how you would do the analysis.

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How do you know if you have the correct answer

• A Hope for the best
• B Run standards to see if you can get them
  correct
• C Compare the results with the values that
  result from other techniques
• D Use a technique that others have used and
  gotten good results

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Pan Am 10385 of the Maid of the
Searecovered16,000 pieces of
propertyrecovered
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Instrumental Analysis

• There is much more than the Instrument
• You are the analyst

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Questions to ask???

• Why? Is sample representative
• What is host matrix?
• Impurities to be measured and approximate
  concentrations
• Range of quantities expected
• Precision accuracy required

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More things to ask.

• Where is analysis to be conducted
• How many samples (per day total)
• How soon are results needed
• Are there standards (analytical QC)
• Long term reliability
• Form of answer required
• Special facilities available

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The Analytical Approach

• Identify the problem.what do you want to know
• What instrumental methods can provide necessary
  results
• Which method is best
• What do the results mean

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Characterization of Properties

• chemical state
• structure
• orientation
• interactions
• general properties

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Reason to understand how an instrument works

• What results can be obtained
• What kind of materials can be characterized
• Where can errors arise

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Where to see Power Points

• http//ehs.virginia.edu/ralphs_chemistry_courses/5
  51/
• Or go to Collab site

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Design of instrumentation to probe a material

• Signal Generation-sample excitation
• Input transducer-detection of analytical signal
• Signal modifier-separation of signals or
  amplification
• Output transducer-translation interpretation

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Molecular Methods

• macro Vs micro
• pure samples Vs mixtures
• qualitative Vs quantitative
• surface Vs bulk
• large molecules (polymers, biomolecules)

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Molecular SpectroscopyIR, UV-Vis, MS, NMR

• What are interactions with radiation
• Means of excitation (light sources)
• Separation of signals (dispersion)
• Detection (heat, excitation, ionization,
  molecule)
• Interpretation (qualitative easier than
  quantitative)

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Elemental Analysis

• bulk, micro, contamination (matrix)
• matrix effects
• qualitative Vs quantitative
• complete or specific element
• chemical state

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Extreme trace elemental analysis

• Direct instrumental determination - multi-element
  - direct excitation---should be least expensive
• These are relative physical methods requiring
  appropriate standards systematic errors like
  spectral interferences occur
• NAA, XRF, sputtered neutral MS

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Extreme trace elemental analysis

• Multi-stage procedures --- sample separation
  and preparation before quantitation
• Standards are less of a problem
• Time consuming subject to losses or
  contamination
• Chromatography coupled with analysis

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Comparing MethodsWhat is important?
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What are important factors in Comparing
Analytical Methods

• A Detection limits
• B Dynamic range
• C Possible Interferences
• D Simplicity
• E All of the above

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Comparing Methods

• cost
• sensitivity
• accuracy/precision
• time
• compatibility
• conditions
• availability

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An analytical checklist

• Have the analytical tasks and goals been defined?
• Have issues of sampling been defined?(eg. size,
  homogeneity, composites)
• Are there facilities for sample storage (custody)
  available and is there a means of identification
  and retreival)

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Checklist 2

• Is pretreatment (eg. extraction, dissolution)
  necessary? (facilities, equipment, reagents)
• Is the sample analyzed representative? (mixing,
  weighing, size)
• Are the instruments appropriate for the required
  measurements? (sensitivity, sample state)

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Checklist 3

• What is the time required for each analysis?
• What expertise is needed to prepare, analyze, and
  interpret?
• How is data captured, calculated, presented, and
  stored for future comparisons?
• Are there appropriate quality controls?
• Define time line for tasks and analysis and then
  calculate overall costs

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Statisticsare no substitute for judgment

• Common sense put into a mathematical form
• Analysis of results - accuracy precision
• Elimination of errors
• Detection limits - signal to noise
• Chemometrics - what do the results mean

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• Accuracy and precision are both needed when you
  have a very limited amount of sample to analyze
• A is true
• B is false

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There is a difference - you
need both
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Errors in Analytical Measurements

• Determinant - unidirectional errors ascribable to
  a definite cause
  
• Indeterminate - uncertainties from unknown or
  uncontrollable factors - generally random -
  noise

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Systematic errors - sources

• Inhomogeneity - handling storage
• Contamination - sampling to reagents
• Adsorption on surface or volatilization
• Unwanted or incomplete chemical reactions
• Matrix effects on generation of analytical signal
• Incorrect standards or calibration

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Recognition of systematic errors

• Reproducibility gives NO information on accuracy
  (high std. dev. hints at problems)
• Make comparisons with other methods
• Check standard reference materials (available
  from NIST)
• Run blanks (be sure background is small and
  reproducible)

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Errors in Analytical Measurements

• Determinant - unidirectional errors ascribable to
  a definite cause
• Indeterminate - uncertainties from unknown or
  uncontrollable factors - generally random -
  noise

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Gaussian Distribution

• Random fluctuations
• Bell shaped curve
• Mean and standard deviation
• 1sigma 68.3, 2sigma 95.5, 3sigma 99.7
• Absolute Vs Relative standard deviation
• Accuracy and its relationship to the measured mean

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Limit of detection

• signal - output measured as difference between
  sample and blank (averages)
• noise - std dev of the fluctuations of the
  instrument output with a blank

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Limit of detection

• signal - output measured as difference between
  sample and blank (averages)
• noise - std dev of the fluctuations of the
  instrument output with a blank
• S/N 3 for limit of detection
• S/N 10 for limit of quantitation

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Sources of Noise

• Environmental - 60 Hz electrical, vibrational
  (shield)
• Johnson (thermal) noise - random fluctuations in
  charge carriers (cool)
• Shot noise - pulses
• 1/f (flicker) noise - important at low frequencies

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Noise Reduction

• Avoid (cool, shield, etc.)
• Electronically filter
• Average
• Mathematical smoothing
• Fourier transform

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If noise is random you know that

• A you can use Gaussian Statistics
• B you can collect data longer to average the
  noise
• C you can determine the average and calculate
  the uncertainty in that value
• D all of the above

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Single channel scanning

• 3 objects each measured 3 times (averaged to
  reduce noise)
• Balance requires 9 measurements
• Monochromator - broad band source to dispersive
  device and then wavelengths are selected one at a
  time
• Increase intensity by scanning slower or
  increasing bandpass

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Multidetector Spectrometer

• Get 3 balances and measure all 3 samples
  simultaneously on separate balances
• Can make measurements in 1/3 time or measure 3
  times as much (noise is random and proportional
  to square root of number of measurements)
• Use of diode arrays instead of slits

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Signal Transformation

• Double pan balance - mesure multiple objects
  simultaneously measure linear combinations
• y(1)X(1) X(2)
• y(2)X(1) X(3)
• y(3)X(2) X(3)
• 3 equations 3 unknowns (each object measured
  twice in half the time)

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The problem with making a measurement using a
double pan balance is..

• A they are hard to use
• B you need to determine the relationship
  between the angle of the pointer and the weight
  difference

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Hadamard multiplexing (transform)

• Use one detector and replace the slit with a mask
  of slits at certain locations (n)- some are open
  others closed (2n-1 slits in mask with just
  more than half open)
• For n3 a mask of 11011 (1 is open) can be slid
  to give 110, 101, 011
• Linear equations improve S/N

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Fourier advantage

• Put all weights on 2 pan balance at the same time
• Change what is measured (not weights but angle of
  pointer showing difference in the 2 pans)
• Z(1)X(1) X(2) - X(3)
• Z(2)X(1) - X(2) X(3)
• Z(3) -X(1) X(2) X(3)

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h(t) a cos 2 pi freq. x time

• sum cos(2pi((f1f2)/2)t
• beat or difference cos(2pi((f1-f2)/2)t
• 5104-sine-wa

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Fourier transform - beat frequency (time domain)

• We can sample the time domain at N equally spaced
  time intervals
• Represent each measurement in terms of a series
  of frequencies
• Decoding procedure to decode N algebraic
  equations
• Fourier transform requires a computer

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Good news / Bad newsAnthrax
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INSTRUMENTATION

• Look at some part of a molecule that has
  characteristic absorption of radiation (color)
• IR / Raman
• Look at the size of a molecule (separation in a
  vacuum mass spectroscopy)
• Separation based on size or chemical affinity
  chromatography

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Correlation Between Bonds and Absorption Peaks
Propanoic Acid

• High wave number (4000 1500 cm-1) peaks are
  produced by X-H, XY, X Y (X, Y can be O,
  N, C) stretching --- Functional Group Region
• Low wave number (1500-600 cm-1) peaks are
  produced by single bonds. --- Fingerprint Region

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Portable Infrared (solids)
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Portable IR
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Raman
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Laser excitation of solids or liquids/ Raman
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Spectral interpretation
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Portable Raman with computer for spectral
interpretation
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Gas Chromatography/ Mass Spectrometry
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QUADRUPOLE MASS SPECTROMETER
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Quadrupole mass analyzer
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New rapid response columns
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Ion Chromatograph (sniffer)
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Sample collector for dogs
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