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BATON
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CINEMA
Chapter
Molecular and Atomic
17
Spectroscopic Methods
Molecular Absorption Spectrometry

•Molecular spectroscopy is used for the

identification and determination of a huge
number of inorganic, organic and
biochemical species.

•Infrared absorption spectroscopy is one of

the most powerful tools for determining the
structure of both inorganic and organic
compounds.
UV/VISIBLE MOLECULAR ABSORTION SPECTROSCOPY

In the UV/visible region, many types

of inorganic compounds absorb radiation
directly. Others can be converted to
absorbing species by means of a
chemical reaction.
 Absorption by Organic Compound
Two types of electrons responsible
for the absorption of ultraviolet and visible
radiation by organic molecules

1. shared electrons that participate

directly in bond formation
2. unshared outer electrons that are
largely localized on atoms such as oxygen,
the halogens, sulfur, and nitrogen.
 Absorption by Organic Compound
Chromophores
Unsaturated organic
functional groups that absorb in the
UV/visible region.

Thus, species with unsaturated

bonds generally absorb in the UV.
 Absorption by Inorganic Compound
In general, the ions and complexes
of elements in the first two transition series
absorb broad bands of visible radiation in
at least one of their oxidation states and
are, as a consequence, colored.

Absorption involves transitions

between filled and unfilled d orbitals of the
metal ion with energies that depend on the
bonded ligands.
 Charge-Transfer
Charge-Transfer Absorption
Absorption
For quantitative purposes, charge-
transfer absorption is particularly important
because molar absorptivities are unusually
large, a circumstance that leads to high
sensitivity.

Charge-Transfer Complex
consists of an electron-donor group
bonded to an electron acceptor. The
excited state is thus the product of a kind
of internal oxidation/reduction process.
Charge-Transfer Absorption
The Standard Addition Method
The difficulties associated with production
of standards with an overall composition closely
resembling that of the sample can be formidable.
Under such circumstances, the method of
standard additions may prove useful.

In the single-point standard addition

method, a known amount of analyte is introduced
into a second aliquot of the sample and the
difference in absorbance is used to calculate the
analyte concentration of the sample.
Alternatively, multiple additions can be made to
several aliquots of the sample and multiple
standard addition calibration curve obtained.
The Standard Addition Method
 SpectrophotometricTitrations
Spectrophotometric Titrations
The method requires that one or
more of the reactants or products absorb
radiation or that an absorbing indicator
be present. In spectrophotometric
titrations, the spectrophotometer serves
as the detect or that monitors the
transmittance or absorbance or the
solution at a suitable wavelength during
the addition of increments or the titrant.
Spectrophotometric Titrations
TitrationCurves: The plot of absorbance versus
titrant volume is called a spectrophotometric
titration curve; the shapes depend on the
species that absorbs radiation. Normally, the
absorbances are corrected for dilution by the
titrant by multiplying the measured values by
(VT+VA)/VA, where VA and VT are the
volumes of the analyte solution and titrant,
respectively. Ideally, the end point is located
by a sharp change in absorbance; often,
conditions are arranged so that two straight-
line regions of differing slopes intersect at the
end point.
Spectrophotometric Titrations
Spectrophotometric Titrations
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THINK
INFRARED ABSORPTION SPECTROSCOPY

Infrared absorption spectroscopy is

also widely employed in analytical
chemistry for identification. Its scope is
nearly as broad as that of UV/visible
methods

IR absorption is one of the premier

techniques for qualitative analysis and
functional group identification.
Molecules That Absorb Infrared Radiation

With the exception of homonuclear

diatomic molecules, such as O2, Cl2, and N2, all
molecules, organic and inorganic, absorb infrared
radiation. Absorption of IR radiation involves
transitions among the vibrational energy levels of
the lowest excited electronic energy levels of
molecules. The number of ways a molecule can
vibrate is related to the number of bond sit
contains and thus the number of atoms making up
the molecule. The number of vibrations is large
even for a simple molecule.
Qualitative and Structural Applications

Infrared absorption spectroscopy

is one of the most powerful and
important tools available to the
chemist for identifying and
determining the structure of organic,
inorganic and biochemical species.
Qualitative and Structural Applications
Making Infrared Absorption Spectroscopy Quantitative

Quantitative applications of IR
spectroscopy are much more limited than
are such applications in the UV/visible
region.
Making Infrared Absorption Spectroscopy Quantitative

Limitations:
1. low molar absorptivities in the
IR region
2. the presence of significant
solvent or background absorption
3. the narowness of IR absorption
bands
 Measuring Absorbance

The use of matched cuvettes for

solvents and analyte, is common in
UV/visible methods, is seldom practical
for IR measurements because of the
difficulty in obtaining cells with identical
transmission characteristics.
 Measuring Absorbance

With Fourier transform systems, it is

common to store the spectrum of the
solvent in memory and to obtain the ratio
of the sample spectrum to that of the
solvent.
 Applying IR Spectroscopy

Infrared spectrophotometry has the

potential for determining an unusually
large number of substances because
nearly all species absorb in this region.

Infrared absorption is used widely to

determine atmospheric pollutants.
 Applying IR Spectroscopy

The primary use of IR methods is

for identification, determination of
confirmation of molecular structure.
 Applying IR Spectroscopy
Infrared Photometers for Routine
Determination of Atmospheric Pollutants
 Applying IR Spectroscopy
Infrared Photometers for Routine
Determination of Atmospheric Pollutants
Applying IR Spectroscopy (VIDEO)
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HANDICAP
ERRORS IN SPECTROPHOTOMETRY

The relative error in the result

can be directly related to the
relative instrumental error
associated with the
measurement.
ERRORS IN SPECTROPHOTOMETRY
MOLECULAR FLUORESCENCE SPECTROSCOPY

Fluorescence
a photoluminescence
process in which atoms or
molecules are excited by
absorption of electromagnetic
radiation.
INFRARED ABSORPTION SPECTROSCOPY

Fluorescence spectroscopy
a very sensitive technique in
determining some molecules.

Luminescence phenomena
include fluorescence,
phosphorescence, and
chemiluminescence
Principles of Molecular Fluorescence

Molecular fluorescence is
measured by exciting the sample at
the absorption wavelength, also
called the excitation wavelength and
measuring the emission at a longer
wavelength called the emission or
fluorescence wavelength.
Principles of Molecular Fluorescence

The short-lived emission that

occurs is called fluorescence whereas
luminescence that is much longer
lasting is called phosphorescence.

Vibration relaxation takes place

during collisions between excited
molecules and molecules of the solvent.
Principles of Molecular Fluorescence
How does concentration influence fluorescence intensity?
How does concentration influence fluorescence intensity?
How does concentration influence fluorescence intensity?
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 Fluorescence Instrumentation

Fluorometer
- two wavelength selectors are
both fillers.

Spectrofluorometer
– both wavelength selectors are
monochromators.

Corrected Spectrofluorometers
– instruments that correct for the
source spectral distribution.
SPECTROFLUOMETER (VIDEO)
FLUOROMETER (VIDEO)
 Fluorescence Instrumentation

Fluorescence
– radiant power emitted is
directly proportional to the
source intensity.

Absorbance
– essentially independent of
source intensity; related to the
ratio of radiant powers.
Applying Molecular Fluorescence Spectroscopy
Fluorescence
– valuable tool used in oil spill
identification. broad at
room temperature.

Fluorescence method
– used to study chemical equilibria
Relatively and kinetics.

Quantitative fluorescence method

– developed for inorganic, organic,
and biochemical species.
Applying Molecular Fluorescence Spectroscopy
Two classes of inorganic
fluorescence method:
1. Direct- based on the reaction of
the analyte with a complexing agent
form a fluorescent complex.

2. Indirect- based on the result of

interaction between analyte and
fluorescent reagent.
Principles of Molecular Fluorescence

Quenching method
– used for the determination of
anions and dissolved oxygen.

Quenching
– depend on the decrease in
fluorescence.
 Chemiluminescence Methods
Chemiluminescence
– produced when a chemical
reactions yields an electronically
excited molecule, which emits
light as it returns to the ground
state.

Bioluminescence
– chemiluminescence reactions
encountered in a number of
biological systems.
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GUIDANCE
 Molecular Scattering Method
Scattering of electromagnetic radiation – An
additional process, aside from absorption and
emission of radiation by molecules, which occurs
when radiation interacts with matter.

Two classes of Scattering:

1. Elastic Scattering – Scattered radiation
has the same energy as the incident
radiation.
2. Inelastic Scattering – Scattered
radiation has higher or lower energy than
the incident radiation
 Molecular Scattering Method
Turbidimetry
– A method that measures the
decrease which occurs in the
transmitted radiation as a result
of scattering from particles.

Nephelometry
- a method that directly measure
the intensity of the scattered
radiation
 Molecular Scattering Method
Laser light scattering method
– a method used to determine the
molecular weights of the compound
and to determine particle sizes.

Raman spectroscopy
- involves inelastic scattering of radiation
caused by vibrational and rotational
transition
- it is complimentary to IR spectroscopy
- it can be used to obtain qualitative
structural and quantitative information
about molecular species
Raman Spectroscopy (VIDEO)
ATOMIC ATOMIC SPECTROSCOPY
SPECTROSCOPY
Atomic Spectroscopy
– it is used for the qualitative and
quantitative determination of 70 to
80 elements.

Detection limits lie in the sub-parts-

per-million range. This method is based on
absorption, emission, or fluorescence . The
widely used technique of atomic
spectroscopy is Atomic absorption (AA)
spectroscopy.
ATOMIC ABSORPTION SPECTROSCOPY

In AA spectroscopy, the sample

must be converted into an atomic vapor
also known as atomization. The sample is
volatilized and decomposed to produce
atom and some ions in the gas phase.

Flame Atomic Absorption Spectroscopy

Furnace Atomic Absorption Spectroscopy
Flame Atomic Absorption Spectroscopy (VIDEO)
Furnace Atomic Absorption Spectroscopy (VIDEO)
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Some other types of Atomic Spectroscopy

In addition to atomic absorption

spectroscopy, atomic emission and
atomic fluorescence are also
encountered. Of the last two, atomic
emission (AE) is much more widely
employed at present. Some AE methods
use flames to produce excited atoms,
which then emit characteristic radiation
as they return to the ground state
Some other types of Atomic Spectroscopy

In contrast to atomic absorption,

atomic emission methods can be used for
qualitative analysis. Complete spectra
can be recorded that are then used to
identify the elements present based on
the wavelength of the lights emitted.

Flame emission was at one time

widely used in the clinical laboratory for
the determination of sodium and
potassium.
Some other types of Atomic Spectroscopy

Atomic fluorescence (AF)

spectroscopy is the most sensitive and
selective of the atomic spectrometric
method. In AF, the fluorescence following
radiational excitation is measured, usually
at the same wavelength.
Chapter/
Molecular and Atomic
17
Spectroscopic Methods
MAYO, CHARLENE MENCH M.
MENDOZA, ANGIE LYKA JOY A.
MENDOZA,CHARLENE MAE B.
NISORTADO, HAROLD P.
PANALIGAN, KERBY Z.
PANOPIO, JOHN ROMAR C.