Atomic Absorption

Spectroscopy
(AAS)

Topics to be covered
Importance of elemental analysis .
Introduction to spectroscopy.
Atomic Absorption Spectroscopy (AAS
Atomic Emission Spectroscopy (AES).
Inductively Coupled Plasma Spectroscopy (ICP).

Importance of elemental
analysis
Monitoring levels of certain elements in samples ( eg.
pharmaceutical products and standards) to detect
the concentrations of these elements.
homodyalysis
protonialdyalysis
ringer lactate
Monitoring the levels of the toxic elements in
samples ( eg. cosmetics, food supplements, entire
plant or part of it) to ensure its safety.
With these information, we can take steps to approve
or withdraw products from the markets.
Widely used in clinical chemistry and environmental
laboratories.

Elements (Heavy Metals)

The term heavy metal refers to any metallic
chemical element that has a relatively high density
and is toxic or poisonous at low concentrations.
They have a specific gravity that is at least 5 times
the specific gravity of water.
Example: Arsenic 5.7,Cadmium 8.65 , Iron 7.9,
Lead 11.34 Mercury 13.546

Trace elements :
Heavy metals that are nutritionally essential for
a healthy life.
Examples are ( Iron, Manganese, Copper and
Zinc).

Commonly encountered
toxic heavy metals
Arsenic

Lead

Mercury

Cadmium

Iron

Aluminum

Types of samples for

analysis
Pharmaceutical

Food

supplements

product

Standards

Cosmetics

Entire

plant
or part of it

Mixture of
known & unknown
herb

Spectroscopy

Spectrometric methods are a large group

of analytical methods
Spectroscopy is the science that deals with
the interactions of radiation with matter
(atomic and molecular).
The most widely used spectrometric
methods are based on electromagnetic
radiation (light, gamma rays, X-rays, UV,
microwave, and radio-frequency).

Electromagnetic Radiation:

consists of discrete packets of energy, which

we call photons
A photon consists of an oscillating electric
field component, E, and an oscillating
magnetic field component, M.

 The characteristics of these fields are:

Orthogonal ( perpendicular ) to each other
Orthogonal to the direction of propagation
of the photon

They flip direction as the photon travels

All photons (in a given, non-absorbing
medium) travel at the same
velocity, v.

What is Frequency ( )?
The number of flips, or oscillations, that occur in one
second.
What is A Wavelength ( )?

The physical distance in the direction of propagation

over which the electric and magnetic fields of a photon
make one complete oscillation.
Unit: Angstrom, nm, m
Velocity Of Light = 2.99792 x 10 8 m/s

The electromagnetic nature of all photons is the same, but

photons can have different frequencies

The relationship between the light velocity, wavelength,

and frequency is :

v=
E = h = hv /
The energy, E, of one photon depends on its frequency of
oscillation :

where h is Planck's constant (6.62618x10-34 Js)

The relationship between the speed of light c ,

wavelength, and frequency is :

C=
When light passes through other media, the velocity
of light
Since the energy of a photon is fixed, the frequency of
a photon does not change.
Thus for a given frequency of light, the wavelength
must as the velocity decreases.

n=c/v
The decrease in velocity is quantitated by the
refractive index, n, which is the ratio of c to the
velocity of light in another medium, v:

Electromagnetic Spectrum:

1- Absorption of Radiation

When radiation passes through a layer of

solid, liquid, or gas, certain frequencies may
be absorbed, a process in which
electromagnetic energy is transferred to the
sample.

Absorption promotes these

particles from their ground state to
more higher-energy excited state.

Tow types of absorption spectra:

Atomic absorption spectrum.
Molecular absorption spectrum.

Beers Law

Many compounds absorb radiation. The

diagram below shows a beam of
monochromatic radiation of radiant
power P0 directed at a sample solution.

Absorption takes place and the beam of

radiation leaving the sample has radiant
power P.

The amount of radiation absorbed may

be measured in a number of ways:
Transmittance, T = P / P0
% Transmittance, %T = 100 T
Absorbance,
A=log10 P0 /P
A=log10 1/T
A=log10 100/%T
A=2-log10 %T

The last equation, A = 2 - log10

%T , is worth remembering
because it allows you to easily
calculate absorbance from
percentage transmittance data.
The relationship between
absorbance and transmittance is
illustrated in the following
diagram:

The equation representing the Beers law:

A=bc

Where
A is absorbance (no units, A = log10 P0 / P ).
is the molar absorbtivity (is a measure of
the amount of light absorbed per unit
concentration) with units of L mol-1 cm-1.
is the path length of the sample that is, the path
blength
of the cuvette in which the sample is
contained. We will express this measurement in
centimeters.

of the compound in solution,

c is the concentration
-1
expressed in mol L

Beers law tells us that absorbance

depends on the total quantity of the
absorbing compound in the light path
through the cuvette. If we plot
absorbance against concentration, we
get a straight line passing through the
origin (0,0).

The working curves are used to

1.

Determine the concentration of an

unknown sample.

2.

To calibrate the linearity of an

analytical instrument.

What are the Processes by which a molecule

can absorb radiation?
1- Rotational transition:
The molecule rotate about various axes, the
energy of rotation being at definite energy
levels, so the molecule may absorb radiation
and be raised to a higher rotational energy level.

2- Vibrational transition:
The atoms or group of atoms within a
molecule vibrate relative to
each other. The molecule may then absorb a
discrete amount of energy
and be raised to a higher vibrational energy
level..

3- Electronic transition:
The electrons of molecule may be raised to a
higher electron Energy.

The three types of internal energy are

quantized

Rotational transitions: low energy E [long (microwave or

far-infrared)]
Vibrational transitions: takes place at high energy E [ near,
far infrared region]
Electronic transitions: takes place at higher energy E
[visible and U.V region]
Type of Radiation

Frequency Range (Hz)

Wavelength Range

Type of Transition

gamma-rays

1020-1024

<1 pm

nuclear

X-rays

1017-1020

1 nm-1 pm

inner electron

Ultraviolet

1015-1017

400 nm-1 nm

outer electron

Visible

4-7.5x1014

750 nm-400 nm

outer electron

near-infrared

1x1014-4x1014

2.5 m-750 nm

outer electron molecular

vibrations

Infrared

1013-1014

25 m-2.5 m

molecular vibrations

Microwaves

3x1011-1013

1 mm-25 m

molecular
rotations,
electron spin flips*

radio waves

<3x1011

>1 mm

nuclear spin flips*

Cont. Introduction to
Spectroscopy
TRANSITION

SPECTRUM

TECHNIQUE

MAIN USE

Electronic
Transitions

UV-vis

UV-vis spectroscopy

Quantitative Analysis

Atomic Absorption
Spectroscopy

Quantitative Analysis

Vibrational
Transitions

IR

IR Spectroscopy

Functional groups
Structural
Identification

Radio

NMR

Spin Orientation

Structure
Identification

 Which molecules or atoms can absorb radiation?

Molecules:

For absorption to occur there must be change in the dipole moment

(polarity) of the molecule.
i.e polar covalent bond in which a pair of electrons is shared
unequally.
eg: of a molecule that can not exhibit a dipole moment.

NN

Can not exhibit a dipole and will not absorb in the I.R region.

eg. of a molecule that can exhibit a dipole moment.

O C O

and so

Unsymmetrical diatomic molecule, does have a permanent dipole

will absorb light.

O C O Vibration mode symmetry and no dipole moment

O C O By induced dipole dipole moment and the molecule can absorb I.R radiation.

 Atoms:

Incase of atoms only electronic transition

occurs.
P
O

Atomic absorption spectra

Molecular absorption
spectra

The outer most electrons

-1
occupy one of the atomic
orbitals and have its energy
, ......levels [K, L, M,
N
[ s, s,p s,p,d s,p,d,f

The outer most electrons

occupy , or n electronic
.energy in the ground state

Upon excitation electrons are- 2 Upon excitation electrons

promoted to any permissible
raised to * or * energy.
higher atomic energy levels
levels
Since there are no bonds-3
there are no vibrational or
rotational energy levels in
either the ground or excited
. state

Since there are bonds, there are

vibrational and rotational
energy levels in both the
ground and excited states

The analytical wavelength is- 4

the resonance wavelength of
the analyte

The analytical wavelength is

.the max

. The spectra are line form- 5

The spectra are in the form

of bands due to the presence
of very close, superimposed
and unresolved vibrational and

Atomic Absorption Spectroscopy (AAS)

AAS was employed in the 1950s
Used for qualitative and quantitative detection.
Its used for the determination of the presence and
concentrations of metals in liquid samples.
Metals that can be detected include Fe, Cu, Al, Pb,
Ca, Zn, Cd and many more.
Concentrations range is in the low mg/L (ppm)
range.

Elements that are highlighted

in pink are detectable by AAS

the AAS instrument

:

The simple diagram for the

AAS
4. The element in the
sample will absorb
some of the light, thus
reducing its intensity
5. The
monochromator
isolates the line of
interest

3. A beam of UV light
will be focused on the
sample

1. We set the
instrument at
certain wavelength
suitable for a
certain element

2. The element
in the sample
will be atomized
by heat

6. The detector
measures the
change in intensity

7. A computer data
system converts the
change in intensity
into an absorbance

1 . L ig h t s o u r c e
H o llo w C a th o d e L a m p

Laser

The disadvantage of both the HCL and laser is that they

have narrow-band light sources and so only one element is
measurable at a time.

2. Laser:
They are intense enough to excite atoms to higher
energy levels. This allows AA and atomic fluorescence
measurements in a single instrument.

2 . A to m is e r
F la m e

G ra p h ite F u rn a c e

H y d rid e

3 .L ig h t S e p a r a tio n
M o n o c h ro m a to r:
u s e d t o i s o l a t e t h e a b s o r p t io n l i n e f r o m
b a c k g r o u n d lig h t d u e to in te r fe r e n c e s .
Note:
Simple dedicated AA instruments often replace the
monochromator with a bandpass interference filter.

4 .D e te c to rs
P h o t o m u lt ip lie r t u b e s

A. Flame Atomic Absorption Spectroscopy:

The technique requires a liquid sample to be

aspirated, aerosolized, and mixed with
combustible gases, such as acetylene and air or
acetylene and nitrous oxide.
The mixture is ignited in a flame whose
temperature ranges from
2100 to 2800 C.

1. Nebulizer:
4. The mixture flows
immediately into the
burner head.
5. It burns as a smooth,
laminar flame evenly
distributed along a
narrow slot.

1. mixes acetylene (the fuel)

and oxidant
(air or nitrous oxide).

3. The result is a heterogeneous

mixture of gases (fuel +
oxidant) and suspended aerosol
(finely dispersed sample).

2. A negative pressure is
formed at the end of the
small diameter, plastic
nebulizer tube (aspiration).

6. Liquid sample not

flowing into the flame
collects in the waste.

Note:
When do we use NO2 ?

2. Ignition:
The process of lighting the AAS flame involves:
turning on first the fuel then the oxidant and then
lighting the flame with the instrument's auto ignition
system.
The flame breaks down the analyte's matrix create the
elemental form of the analyte atom.

During combustion, atoms of the element of

Interest in the sample are reduced to
free,
unexcited ground state atoms, which absorb
light at characteristic wavelengths,
as shown in the figure.

Optimization is accomplished by :

1.

Aspirating a solution containing the element

2.

Adjusting the fuel/oxidant mix until the

maximum light absorbance is achieved.

3.

Careful control of the fuel/air mixture is

important because each element's response
depends on that mix in the burning flame.

3. Lamp (Hollow Cathode Lamb):

Consists of a cathode and an anode. The cathode is made of the
element of interest

1.
2.
3.

A large voltage across the anode and cathode will cause the
inert gas to ionize.
The inert gas ions will then be accelerated into the cathode,
sputtering off atoms from the cathode.
Both the inert gas and the sputtered cathode atoms will in turn
be excited by collisions with each other.

4- When these excited atoms decay to lower

energy levels they emit a few spectral
lines characteristic of the element of
interest.
5- The light is emitted directionally through
the lamp's window, a window made of a
glass transparent in the UV and visible
wavelengths.
6- The light can then be detected and a
spectrum can be determined.

4. Monochromator
The light passes from the HCL through the element in
the sample to the monochromator.
Its function is:
It isolates the specific light of the element of
interest from the other background lights and
transfers it to the photomultiplier tube (detector).

5. Photomultiplier Tube (PMT)

:
Before an analyte is aspirated, a measured signal is
generated by the PMT as light from the HCL passes
through the flame. When analyte atoms are present in
the flame--while the sample is aspirated--some of that
light is absorbed by those atoms. This causes a
decrease in PMT signal that is proportional to the
amount of analyte
.

PMT

The PMT detects the amount of reduction of

the light intensity due to absorption by the
analyte, and this can be directly related to
the amount of the element in the sample.
The PMT converts the light signal into an
electrical
signal and a computer system translates
it into
absorbance.

Photomultiplier Tube
Light
Dynode

Photocathode
eDynode

Dynode
Current

Convert light energy to electrical energy

Gases used in the FAAS:

Different flames can be achieved using different mixtures of gases, depending on the
desired temperature and burning velocity.
Some elements can only be converted to atoms at high temperatures.
Some metals form oxides that do not re-dissociate into atoms. To inhibit their
formation, conditions of the flame may be modified to achieve a reducing, nonoxidizing flame.

Table of the characteristics of

various flames.
Max. flame speed (cm/s)

Max. temp. (oC)

Air-Coal gas

55

1840

Air-propane

82

1925

Air-hydrogen

320

2050

Air-50% oxygen-acetylene

160

2300

Oxygen-nitrogen-acetylene

640

2815

Oxygen-acetylene

1130

3060

Oxygen-cyanogen

140

4640

Nitrous oxide-acetylene

180

2955

Nitric oxide-acetylene

90

3095

Nitrogen dioxyde-hydrogen

150

2660

Nitrous oxide-hydrogen

390

2650

B. Graphite Furnace Atom Absorption

Spectroscopy:
(GFAAS) Graphite Furnace Atomic Absorption
Spectroscopy
(ETAAS) Electro thermal Atomic Absorption
Spectrometry

5. The monochromator isolates the

light of the element of interest from
the background lights to the PMT.
The PMT tube measures the change
intensity.

2. A controlled voltage is
applied at the ends of the
graphite tube, which is heated
rapidly to high temperatures
(up to 2600C).

4. The graphite tube is

permanently flushed with
argon while it is in
operation

1. The source of light

(lamp) emits light with a
wavelength specific to
the element of interest

3. Samples are deposited in the graphite

tube heated to vaporize and atomize
the analyte atoms absorb ultraviolet
or visible light and make transitions to
higher electronic energy levels.

Small

sample
Formation of stable Thermal oxide (Al, Mo, Ti
Residual elements
Biological samples
Effective atomization
Sample stay for Long time in graphite
tube>>>high sensetivety
Suitable for solid samples
Ionic interferase rather than chemical or
physical due to high temp.

Mercury Cold Vapor

-Free mercury atoms can exist at room
temperature can be analysed using atomic
absorption without employing flame and graphite
furnace techniques at high temperatures.

-Mercury is reduced in solution using stannous

chloride or sodium borohydride in a closed
system.
-The reaction quantitatively releases mercury
(from the sample solution) and is carried by a
stream of air or argon through a quartz
sample sell placed in the light path of an AA
instrument for analysis.
The detection limit for mercury by this cold
vapor technique is approximately 0.02 mg/L.

Hydride Generation Atomic Absorption

Spectroscopy (HGAASC.):
Its used for elements that are forming volatile hydrides
(e.g. As, Se, pb, Sb, Te, Sn, Bi)
(HAAS) is very useful in case of interferences, poor
reproducibility, and poor detection limits .

4. The PMT detects the amount of

reduction of the light intensity due to
absorption by the analyte and can be
directly related to the amount of the
element in the sample

3. The monochromator isolates

analytical lines' photons
passing through the optical cell
and removes the scattered
light of other wavelengths from
the optical cell

1. The HCL emits the light

with a wavelength
characteristic to the
element of interest

2. The Light passes from the

HCL through the optical cell
to the monochromator then
to the detector

sample flow in the (HGAAS)

4. In the optical cell the flame is ignited
automatically by the air/C2 H 4 and the gaseous
metal hydride form decomposes into the
elemental form which can absorb the
HCL's beam. The light passes to the Mon. and
then to the PMT

3. The liquid mixture flows

into a gas/liquid separator where
the hydride and some gaseous hydrogen are
purged (via a high purity inert gas) into the
optical cell via a gas transfer line

Optical cell

Volatile Hydride
Gas liquid separator

peristaltic
pump
1. The metal oxyanions reacts
with Sodium Borohydride and HCl
and produces a volatile hydride:
H2Te, H2Se, H3As, H3Sb, etc.

Mixing
reaction
coil

2. The peristaltic pump and the flowing reagents

along with the tubing of specific lengths controls
the time from reagent mixing and separation of
the volatile hydride from the
liquid and sending the sample to the optical cell.

AAS Interferences
Any factor that affects the ground state population
of the analyte element.
Factors that may affect the ability of the
instrument to read this parameter.
A) Spectral interferences: due to radiation
overlapping than of the light source.
B) Formation of compounds that do not dissociate in
the flame.
The most common example is the formation of
calcium and strontium phosphates.

C) Ionization of the analyte reduces the

signal.
This is commonly happens to barium, calcium,
strontium, sodium and potassium.
D) Matrix interferences: due to differences
between surface tension and viscosity of test
solutions and standards
.
E) Broadening of a spectral line
1. Doppler effect
2. Lorentz effect
3. Quenching effect
4. Self absorption or self-reversal effect