INSTRUMENTAL ANALYSIS CHEM 4811

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INSTRUMENTAL ANALYSIS CHEM 4811

• CHAPTER 5

DR. AUGUSTINE OFORI AGYEMAN Assistant professor
of chemistry Department of natural
sciences Clayton state university
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CHAPTER 4 ULTRAVIOLET AND VISIBLE MOLECULAR
SPECTROSCOPY (UV-VIS)
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UV-VIS SPECTROSCOPY
- Solutions allow a component of white light to
pass through and absorb the complementary color
of the component - The component that passes
through appears to the eye as the color of the
solution - This chapter deals with molecular
spectroscopy (absorption or emission of UV-VIS
radiation by molecules or polyatomic ions) -
The spectrum is absorbance or transmittance or
molar absorptivity versus wavelength
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UV-VIS SPECTROSCOPY
Complementary Colors
?max 380-420 420-440 440-470 470-500 500-520 520-
550 550-580 580-620 620-680 680-780
Color Observed Green-yellow Yellow Orange Red Pu
rple-red Violet Violet-blue Blue Blue-green Green
Color Absorbed Violet Violet-blue Blue Blue-green
Green Yellow-green Yellow Orange Red Red
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UV-VIS SPECTROSCOPY
Complementary Colors
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UV-VIS SPECTROSCOPY
Complementary Colors Ru(bpy)32 ?max 450
nm Color observed with the eye orange Color
absorbed blue Cr3-EDTA complex ?max 540
nm Color observed with the eye violet Color
absorbed yellow-green
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UV RADIATION
- Wavelength range is 190 nm 400 nm - Involved
with electronic excitations - Radiation has
sufficient energy to excite valence electrons in
atoms and molecules - Vacuum UV spectrometers
are available that uses radiation between 100 Å
200 nm (also electronic excitations)
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VISIBLE RADIATION
- Wavelength range is 400 nm 800 nm - Involved
with electronic excitations - Similar to UV -
Spectrometers therefore operate between 190 nm
and 800 nm and are called UV-VIS
spectrometers - Can be used for qualitative
identification of molecules - Useful tool for
quantitative determination
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ELECTRONIC EXCITATION
- Electrons in molecules move in molecular
orbitals at discrete energy levels - Energy
levels are quantized - Molecules are in the
ground state when energy of electrons is at a
minimum - The molecules can absorb radiation and
move to a higher energy state (excited state) -
An outer shell (valence) electron moves to a
higher energy orbital
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ELECTRONIC EXCITATION
- Is the process of moving electrons to higher
energy states - Radiation must be within the
visible or UV region in order to cause
electronic excitation - The frequency absorbed
or emitted by a molecule is given as ?E h? ?E
E1 Eo E1 excited state energy Eo ground
state energy
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ELECTRONIC EXCITATION
Three Distinct Types of Electrons Involved in
Transition Electrons in a Single Bond
(Alkanes) - Single bonds are called sigma (s)
bonds - Amount of energy required to excite
electrons in d bonds are higher than photons
with wavelength greater than 200 nm - Implies
alkanes and compounds with only single bonds do
not absorb UV radiation - Used as transparent
solvents for analytes
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ELECTRONIC EXCITATION
Three Distinct Types of Electrons Involved in
Transition Electrons in Double or Triple Bonds
(Unsaturated) - Alkenes, alkynes, aromatic
compounds - These bonds are called pi (p)
bonds - p bond electrons are excited relatively
easily - These compounds absorb in the UV-VIS
region
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ELECTRONIC EXCITATION
Three Distinct Types of Electrons Involved in
Transition Electrons Not Involved in Bonding
Between Atoms - Called the n electrons (n
nonbonding) - Organic compounds containing N, O,
S, X usually contain nonbonding electrons - n
electrons are usually excited by UV-VIS
radiation - Such compounds absorb UV-VIS
radiation
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ELECTRONIC EXCITATION
s
?E
Energy
s orbital
s orbital
s
- Two s orbitals on adjacent atoms overlap to
form a s bond - Two molecular orbitals is the
result - Sigma bonding orbital (s) is of lower
energy than the atomic orbitals (filled with
the two 1s electrons) - Sigma antibonding orbital
(s) is of higher energy than the atomic orbitals
(empty) ?E energy difference between s and s
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ELECTRONIC EXCITATION
- p orbitals of atoms can also overlap along axis
to form sigma bonds - There are three p
orbitals in a given subshell - One of these p
orbitals from adjacent atoms form sigma
orbitals - The other two p orbitals can overlap
sideways to form p orbitals - The result is pi
bonding (p) and pi antibonding (p) orbitals - p
orbital filled with 2 electrons has no tendency
of forming bonds
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ELECTRONIC EXCITATION
Relative Energy Diagram of s,p, and n electrons
s
p
n
p
s
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ELECTRONIC EXCITATION
- Energy required to excite electrons from s to
s is very high (higher than those available in
the UV region) - UV radiation is however
sufficient to excite electrons in p to p and n
to p or s antibonding - Molecular groups
that absorb UV or VIS light are called
chromophores
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ABSORPTION BY MOLECULES
- Review quantum mechanics (beyond the scope of
this text) - Quantum mechanical selection rules
indicate that some transitions are allowed and
some are forbidden - Electrons move from highest
occupied molecular orbital (HOMO) to the lowest
unoccupied molecular orbital (LUMO) during
excitation - LUMO is usually an antibonding
orbital - p electrons are excited to antibonding
p orbitals
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ABSORPTION BY MOLECULES
- n electrons are excited to either s or p
orbitals p ? p Transition - A molecule must
possess a chromophore with an unsaturated
bond (CO, CC, CN, etc) n ? p or n ? s
Transition - A molecule must contain atoms with
nonbonding electrons (N, O, S, X) - Lists or
organic compounds and their ?max are available
(table 5.3)
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TRANSITION METAL COMPOUNDS
- Solutions are colored - Absorb light in the
visible portion of the spectrum - Absorption is
due to the presence of unfilled d orbitals ?max
is due to - The number of d electrons - Geometry
of compound - Atoms coordinated to the transition
metal
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ABSORPTIVITY (a)
- Defines how much radiation will be absorbed by
a molecule at a given concentration and
wavelength - Is termed molar absorptivity (e) if
concentration is expressed in molarity (M,
mol/L) - Can be calculated using Beers Law (A
abc ebc) - If units of b is cm and c is M then
e is M-1cm-1 or Lmol-1cm-1 - Magnitude of e is
an indication of the probability of the
electronic transition
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ABSORPTIVITY (a)
- High e results in strong absorption of light -
Low e results in weak absorption of light - e is
constant for a given wavelength but different at
different wavelengths - emax implies e at ?max
(see table 5.4 for some emax values) - e is 104
105 for allowed transitions and 10 100 for
forbidden transitions
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UV ABSORPTION CURVES
- Broad absorption band is seen over a wide range
of wavelengths - Broad because each electronic
energy level has multiple vibrational and
rotational energy levels associated with it -
Each separate transition is quantized -
Vibrational energy levels are very close in
energy - Rotational energy levels are even
closer - These cause electronic transitions to
appear as a broad band
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SOLVENTS
- Many absorbing molecules are usually dissolved
in a solvent - Solvent must be transparent over
the wavelength range of interest - Solute must
completely dissolve in solvent - Undissolved
particles may scatter light which will affect
quantitative analysis - Solvent must be
colorless - Examples acetone, water, toluene,
hexane, chloroform
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INSTRUMENTATION
- Radiation source - Monochromator - Sample
holder - Detector - Computer
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RADIATION SOURCE
- Constant intensity over all wavelengths -
Produce light over a continuum of wavelengths -
Tungsten lamp and deuterium discharge lamp are
the most common
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RADIATION SOURCE
Tungsten Filament Lamp - Just like an ordinary
electric light bulb - Contains tungsten filament
that is heated electrically - Glows at a
temperature near 3000 K - Produces radiation at
wavelengths from 320 to 2500 nm - Stable, robust,
and easy to use - Modern lamps are
tungsten-halogen lamps (has quartz
bulb) Disadvantage - Low radiation intensity at
shorter wavelengths (lt 350 nm)
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RADIATION SOURCE
Dueterium (D2) Arc Lamp - Made of deuterium gas
(D2) in a quartz bulb - D2 molecules are
electrically excited and dissociated - Produces
continuum radiation at ?s from 160 to 400 nm -
Stable, robust, and widely used - Emission
intensity is 3x that of hydrogen at short ?s
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RADIATION SOURCE
Xenon Arc Lamps - Electric discharge lamps -
Xenon gas produces intense radiation over 200
1000 nm upon passage of current - Produce very
high radiation intensity - Widely used in
visible region and long ? end of UV
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MONOCHROMATORS
- Disperse radiation according to selected
wavelengths - Allow selected wavelengths to
interact with the sample - Diffraction gratings
are used to disperse light in modern
instruments Refer chapter 2
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DETECTOR
- Earlier detectors were human eye observation of
color and intensity - Modern instruments make
use of photoelectric transducers (detection
devices that convert photons into electric
signal) Examples - Barrier layer cells -
Photomultiplier tubes - Semiconductor detectors
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DETECTOR
Barrier Layer Cells - Also called photovoltaic
cells - A semiconductor (selenium) is joined to
a strong metal base (iron) - Silver is coated on
the semiconductor - Current is generated at the
metal-semiconductor interface (requires no
external electrical power) - Response range is
350 nm 750 nm
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DETECTOR
Photomultiplier Tubes - The most common -
Photoemissive cathode is sealed in an evacuated
transparent envelope - Also contains anode and
other electrodes called dynodes - Electrons from
cathode hit dynodes which causes more electrons
to be emitted - Process repeats until electrons
fall on anode (collector)
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DETECTOR
Semiconductor Detectors - Silicon and germanium
are the most widely used elements - Others
include InP, GaAs, CdTe - Covalently bonded
solids with ? range 190 nm 1100
nm Photodiode Array - Consists of a number of
semiconductors embedded in a single crystal in a
linear array - Used as detector for HPLC and CE
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SAMPLE HOLDER
- Called sample cells or cuvettes or cuvets -
Different types of sample holders are designed
for solids, liquids, and gases - Cells must be
transparent to UV radiation - Quartz and fused
silica are commonly used as materials - Glass or
plastic cells can be used for only VIS region -
Material must be chemically inert to solvents
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SAMPLE HOLDER
- HF and very strong bases should not be put in
cells - Standard cell is the 1 cm pathlength
rectangular cell - Holds about 3.5 mL sample -
Flow through cells are available (for
chromatographic systems) - Larger pathlength or
volume cells are used for gases - Thin solid
films can be analyzed using a sliding film holder
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SAMPLE HOLDER
Fiber Optic Probes - Enables spectrometer to be
brought to sample for analysis - Enables
collection of spectrum from microliter samples -
Can collect spectrum from inside almost every
container - Useful for hazardous samples
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ABSORPTION DEFINITIONS
Chromophore - A group of atoms that gives rise to
electronic absorption Auxochrome - A
substituent that contains unshared electron pairs
(OH, NH, X) - An auxochrome attached to a
chromophore with p electrons shifts the ?max to
longer wavelengths
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ABSORPTION DEFINITIONS
Bathochromic - A shift to longer wavelengths or
red shift Hypsochromic - A shift to shorter
wavelengths or blue shift Hyperchromism - An
increase in intensity of an absorption band
(increase in emax) Hypochromism - A decrease in
intensity of an absorption band (decrease in emax)
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SOLVENT EFFECTS
- Molecules with absorption due to p ? p
transition exhibit red shift when dissolved in
polar solvents as compared to nonpolar
solvents - Used to confirm the presence of p ?
p transitions in molecules - Molecules with
absorption due to n ? p transition exhibit blue
shift when dissolved in solvents that are able
to form hydrogen bonds (same with n ? s
transition) - Used to confirm the presence of n
electrons in a molecule - Blue shift of n ? s
puts molecules into the vacuum UV region
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SOLVENT EFFECTS
- A compound that contains both p and n electrons
may exhibit two absorption maxima with change in
solvent polarity - p ? p transitions absorb
10x more strongly than n ? p transition - n ?
p transition occur at longer wavelengths than p
? p - Such a compound will exhibit two
characteristic peaks in a nonpolar solvent such
as hexane - The two peaks will be shifted closer
to each other in a polar and hydrogen bonding
solvent such as ethanol
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ANALYSIS OF A MIXTURE
- Occurs when there is more than one absorbing
species - All absorbing species will contribute
to absorbance at most ?s - Absorbance at a given
? sum of absorbances from all species AT
e1b1c1 e2b2c2 e3b3c3 . For the same
sample cell b1 b2 b3 b AT b(e1c1 e2c2
e3c3 .)
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APPLICATIONS
- Environmental monitoring - Industrial quality
control or process control - Pharmaceutical
quality control - For measuring kinetics of a
chemical reaction - For measuring the endpoint
of spectrophotometric titrations - For
spectroelectrochemistry in which redox reactions
are studied by measuring the electrochemistry
and spectroscopy simultaneously
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OTHER TECHNIQUES
- Methods for nontransparent particles suspended
in a liquid (colloidal suspensions,
precipitates) - Used for analyzing the clarity
of drinking water, liquid medications,
beverages Nephelometry - Measures the amount of
radiation scattered by the particles Turbidimetry
- Measures the amount of radiation not scattered
by the particles
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LUMINESCENCE
- Molecular emission - Includes any emission of
radiation Emission Intensity (I) I kPoc k is
a proportionality constant Po is the incident
radiant power c is the concentration of emitting
species - Only holds for low concentrations
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LUMINESCENCE
Photoluminescence (PL) - Excitation by absorption
and re-radiation (very short lifetime) - Examples
are fluorescence and phosphorescence Chemilumines
cence (CL) - Excitation and emission of light as
a result of a chemical reaction Electrochemilumin
escence (ECL) - Emission as a result of
electrochemically generated species Bioluminescen
ce - Production and emission of light by a living
organism
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LUMINESCENCE
Fluorescence - Instantaneous emission of light
following excitation - Excitation by photon
absorption to a vibrationally excited singlet
state followed by relaxation resulting in
emission of a photon - Emitted photon has lower
energy (longer ?) than absorbed energy (due to
the radiationless loss) - Called the stokes
fluorescence (excited state lifetime 1-20
ns) - A molecule that exhibits fluorescence is
called fluorophore
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LUMINESCENCE
Phosphorescence - Similar to fluorescence -
Excited state lifetime is up to 10 s -
Excitation by absorption of light to an excited
singlet state, then an intersystem crossing (ISC)
to the triplet state, followed by emission of a
photon - Photon associated with phosphorescence
has lower energy than fluorescence
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MOLECULAR EMISSION SPECTROSCOPY
- Two electrons occupying a given orbital have
opposite spins - There are two possible
electronic transitions - The excited state is
known as a singlet state if one of the electrons
goes to the excited state without changing its
spin - The excited state is known as a triplet
state if one of the electrons goes to the
excited state and changes its spin for both to
have same spin
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MOLECULAR EMISSION SPECTROSCOPY
- Singlet state energy levels (S) are higher than
triplet state energy levels (T) - Ground state
is a singlet state (So) - Excited state singlet
can undergo radiationless transition to excited
state triplet (ISC) Transition from ground state
singlet to excited state triplet is
forbidden Relative energy of transition Absorptio
n gt Fluorescence gt Phosphorescence
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MOLECULAR EMISSION SPECTROSCOPY
Intersystem crossing (radiationless)
Excited Singlet State (S1)
ISC
Excited Triplet State (T1)
Relative Energy
Absorption
Fluorescence
Phosphorescence
Ground Singlet State (So)
Radiationless transition to the lowest
vibrational level in the excited state
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MOLECULAR EMISSION SPECTROSCOPY
Instrumentation Components of the Fluorometer
Source ? selector sample
monochromator (? selector)
readout
detector
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MOLECULAR EMISSION SPECTROSCOPY
Applications - Analysis of clinical samples,
pharmaceuticals, environmental samples,
steroids Advantages - High sensitivity and
specificity - Large linear range Disadvantage -
Quenching by impurities and solvent -
Temperature, viscosity and pH must be controlled
to minimize quenching