Analytical Techniques

2015 4 17

12:04

Reasons for using analytical chemistry

Structure determination
Composition analysis of substances
Determining purity
Structural determination
Infrared spectroscopy (bonds)
Mass spectroscopy (Ar/Mr, arrangement)
NMR spectroscopy (chemical environment of isotopes)

Modern Analytical Chemistry Page 1

Spectroscopy Principles
2015 4 17

12:08

Electromagnetic Spectrum

Range of all possible frequencies of light

Uses:
NMR
Bond Length Bond Type
Vis-UV
Process:
Molecular Rotation
Electronic Transition
Spectra
Absorption Spectrum
Spectrum of electromagnetic radiation transmitted through a substance
Lines correlate to frequencies that are absorbed
Absorbed frequencies excite e- from ground state to excited state
Production
EM radiation passed through sample
Incident EM radiation compared to recorded EM radiation
Emission Spectrum
Spectrum of electromagnetic radiation emitted by a substance
Lines correlate to frequencies that are emitted
Emitted frequencies are from e- moving from excited state to ground
state
Production
EM radiation measured from excited sample
Absorption
Vibrations
Rotations
Electronic rotation

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Infrared Spectroscopy
2015 4 17

12:37

Diagram

Monochromator
Allows only radiation of particular wavelength to pass through
Beam Splitter
Splits beam in half to sample and half to reference
Detector
Compares the two samples (photomultiplier)
Bond Stretching/Bending
Types of Vibrational Motions
Stretching of bonds
Bending of bonds (changes in bond angle)
Dipole moment is necessary for vibrational motion
Energy Required for Vibration
Increases with stronger bond
Increases with greater atomic mass
Each bond type has characteristic wavenumber
Wavenumber = 1/
Low wavenumber - greater energy required for vibration

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Absorption of Radiation
2015 4 17

12:48

Vibrations
Not all vibrations absorb IR
For IR absorption, dipole moment must occur
H2O/SO2

CO2

-CH2-

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Nuclear Magnetic Resonance

2015 4 17

1:01

NMR in MRI

Human body consists of 70% water

Protons in water molecules/proteins/lipids/carbohydrates detected by MRI
Water is in different environments
Organs have different water-lipid ratio
Protons have different environments have varied absorption
Tetramethylsilane
Internal standard is (CH3)4Si
Chemically inert, and does not react with sample
All protons in are in the same environment, giving strong single peak
As CH3 is bonded to Si, TMS absorbs radiowaves much more upfield than protons
attached to carbon
Si has low electronegativity
Spin-Spin Coupling
Proton can align with or against the magnetic field
Number of Adjacent Protons Splitting Pattern Type of Splitting

0
1

1
1

Singlet
1

Doublet

1 2 1

Triplet

1 3 3 1

Quartet

1 4 6 4 1

Quintet

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Atomic Absorption Spectroscopy

2015 4 17

1:03

Uses of AA Spectroscopy

Detection of metal ions in samples

Aluminum in blood
Calcium in blood, plants, soil samples, water (hardness)
Iron in plants
Copper in alloys
Chromium in seawater
Measures low concentration of metals/metal ions
Principles of AA spectroscopy
When metal atoms are excited by heat, atoms absorb light of certain frequency
e- goes from ground state to excited state
Ratio of transmitted/incident light is proportional to concentration of atoms
present
Sample is compared to known calibration curve
Diagram

Monochromatic Light Source (Hollow Cathode Lamp)

Produces light of same frequency as is absorbed by specie being detected
Metal being detected is used as cathode of lamp
Atomizer
Liquid sample converted into small droplets
Evaporates and atomizes solvent
Flame
e- excited at flame
Monochromator
Only detects radiation of same/certain frequency
Detector
Converts photons into electric current/signal

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Chromatography
2015 4 17

1:23

Chromatography Uses
Separation of mixtures
Identification of compounds in mixtures
Determine purity of a substance
Stationary and Mobile Phases
Stationary phase does not move
Components adsorb onto stationary phase
Mobile phase moves through the stationary phase
Components dissolve in mobile phase
Adsorption
Adhesion of atoms/ions/molecules of gas/liquid/dissolved solid to a surface
Separation
Depends on different abilities of the components to adsorb and dissolve

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Paper, Thin-Paper, and Column Chromatography

2015 4 17

1:25

Paper Chromatography

Stationary Phase
Water within fibres of paper
Mobile Phase
Water or ethanol
Thin-Layer Chromatography
Stationary Phase
Silica or Alumina supported on glass/plastic
Mobile Phase
Water or ethanol
Rr Values
Ratio of distances moved by solute and solvent
Rf = Distance Travelled by Component / Distance Travelled by Solute
Paper vs. TLC
Similarities
Both used for separation
Differences
TLC
Gives better separation, as particles are finer than pores in paper
Withstands stronger solvents
Separation is more easily recoverable
Paper
May need to be developed using UV/Iodine/Acid
Sugars and H2SO4 reagent
Column Chromatography
Stationary Phase
Silica or Alumina
Mobile Phase
Eluent
Procedure
Stationary phase packed on top of glass wool
Eluting solvent saturates powdered stationary phase
Sample, dissolved in eluent, is added at the top of the column
More eluting solvent is added at the top of the column
Components move at different rates and have different retention times
Collected separately as they leave the column
Eluent is removed by evaporation

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High-Pressure Liquid and Gas-Liquid Chromatography

2015 4 17

1:34

High-Pressure Liquid Chromatography

Stationary Phase
Silica or Alumina
Mobile Phase
Liquid (e.g. water or alcohol)
Procedure
Mobile phase forced through column under pressure at constant temperature
Sample placed in liquid stream/components separate as they pass through
tube
Components reach detector at different times
Relative attraction to stationary/mobile phases determines retention
time
Detected by absorption of UV/fluorescence/conductivity
Gas-Liquid Chromatography
Stationary Phase
Liquid (long-chain alkane/hydrocarbon) coated solid support
Mobile Phase
Inert gas (H2 or N2)
Procedure
Sample injected into mobile phase
Sample vaporized at high temperatures
Sample is carried by inert gas through column
Detector at end of column
Change of detector signal with corresponding time is recorded
Components are separated by partition between alkane and gas
Components have different retention times
Dependent on mass, polarity, and volatility
Speed of separation is temperature dependent
Area under peak proportional to quantity of component
Gas-Liquid Chromatography vs. High-Pressure Liquid Chromatography
Gas-Liquid Chromatography
High-Pressure Liquid Chromatography

Identifies volatile/stable compounds

Analysis of urine
Underground mine gas analysis
Blood-alcohol level determination

Analyzes temperature-sensitive compounds

Oil analysis
Alcoholic beverages
Antioxidants/sugars/vitamins in food
Pharmaceuticals
Polymers
Insecticide/Herbicide quality control
Biotechnology and Biochemistry research

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Visible Spectroscopy and UV Spectroscopy

2015 4 17

1:33

Ligands and d-orbital Splitting

Charged and polar ligands cause d-orbitals to split

Repulsion between d electrons and non-bonding electrons on ligand
Interactions of different orbitals of ligands vary
3 low energy : 2 high energy split
Different ligands and extent of split (spectrochemical series)
ClH2O

NH3

Transition Metal Color

Result of e- transition between d-orbital split

Increased d-orbital splitting leads to higher frequency radiation absorption
Complementary color of color absorbed is observed
Wavelength Absorbed Wavelength Emitted
400 nm (Violet)

560 nm (Green-Yellow)

490 nm (Blue-Green)

620 nm (Red)

580 nm (Yellow)

430 nm (Dark Blue)

650 nm (Red)
520 nm (Green)
Factors Affecting Transition Metal Color
Element being considered (especially nuclear charge)
[Mn(H2O)6]2+ is almost colorless, and [Fe(H2O)]3+ is yellow-brown
Charge of the ion, which affects d-orbital splitting due to numbers of e- present
[Fe(H2O)6]2+ is pale-green and [Fe(H2O)6]3+ is yellow-brown
[Ar] 3d6
[Ar] 3d5
Identity of ligand
[CuCl4]2- is yellow, [Cu(H2O)6]2+ is pale blue, and [Cu(NH3)]2+ is dark blue
Stereochemistry
Double Bonds and Absorption
Chromophore
Part of molecule responsible for absorbing radiation
Conjugated System
Alternated double and single bonds
More likely to absorb longer wavelengths
Delocalization of pi bonds over large areas absorb light in far UV/Visible light region
2
sp hybridization enables delocalization
Phenolphthalein in acid vs. alkali
Retinol (UV) vs. Retinal (Visible)
Beer-Lambert Law
Direct relationship between absorption and concentration
Absorbance = log (I0/I) = * c * l
= molar absorption coefficient
c = concentration
l = distanced travelled by light
Transmittance = (Io/I) * 100

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