Biochemical instrumental analysis-2
CLS 332
Dr. Maha Al-Sedik
2015
Atomic absorption spectrophotometer
Atomic absorption spectrophotometer
• Atomic absorption spectroscopy is a quantitative method of
analysis that is applicable to many metals such as Ca , Fe , Al and
copper.
• Only a drop of sample is needed.
• Atomic absorption is so sensitive that it can measure down to
parts per billion of a gram in a sample.
in AAS, the sample is
atomised – ie converted
into free atoms in the
vapour state.
The
Atomisation
Process of converting an analyte in solid, liquid or solution form to a
free gaseous atom.
Principle:
 Atoms of different elements absorb characteristic wavelengths
of light.
 The metal vapor absorbs energy from an external light source,
and electrons jump from the ground to the excited states.
 The greater the number of atoms in the vapor, the more
radiation is absorbed.
How it works?
 For example with lead, a lamp containing lead emits light from
excited lead atoms that produce the right mix of wavelengths to
be absorbed by any lead atoms in the sample.
 Beam of electromagnetic radiation emitted from the lamp is
passed through the vaporized sample. Some of the radiation is
absorbed by the lead atoms in the sample.
Principle:
 The greater the number of atoms
in the vapor, the more radiation is
absorbed.
OR
 The amount of light absorbed is
directly proportional to the
number of lead atoms.
Components
Components :
 The light source ( hollow cathode lamp ).
 flame
 Monochromater
 photomultiplier tube
 Detector
The light source:
hollow cathode lamp
hollow cathode lamp for Aluminum (Al) is shown below
hollow cathode lamp:
 This contains a tungsten anode and a cylindrical hollow cathode
made of the element to be determined. These are sealed in a glass
tube filled with an inert gas. eg neon or argon.
 The shape of the cathode concentrates the radiation into a beam
which passes through window.
•The cathode lamps are stored in
a
compartment inside the AA
spectrometer. The specific lamp
needed for a given metal analysis
is rotated into position for a
specific experiment.
The flame:
 A flame is created, usually using ethyne & oxygen (fuel).
 The flame gases flowing into the burner create a suction that pulls
the liquid into the small tube from the sample container.
 This liquid is transferred to the flame where the sample is
atomized. The metal atoms then absorb light from the source
(cathode lamp).
 The flame is used here as cuvet and for atomization.
The Manganese Flame
The Potassium Flame
The Copper Flame
The Calcium Flame
Monochromater:
 The monochromator is used to select a particular wavelength of
light for observation.
Photomultiplier tube:
 The intensity of the light is fairly low, so a photomultiplier tube
(PMT) is used to strengthen the signal intensity.
 It can multiply the current produced by incident light by as much
as 100 million times.
photomultiplier tube
Detector:
 Transform light to electrical impulses.
 The electricity is directly proportional to the intensity of light.
http://www.youtube.com/watch?v=_KZjb9G3hB8irst.pptx
http://www.youtube.com/watch?v=HBegTB_WDxQ
Flameless atomic absorption
spectrophotometry
 It differs from atomic absorption spectrophotometer in how the
compounds are atomized.
 In AAS a flame is used to produce individual atoms.
 In the other it is carried out by putting the sample in a small
graphite tube and passing an electrical current through to heat it.
Summary of atomic absorption spectrophotometry :
 Atomic absorption spectrophotometer is used in the measurement
of many metal such as Ca , Fe , Al and copper.
 Only a drop of sample needed.
 Atomic absorption is so sensitive that it can measure down to parts
per billion of a gram in a sample.
Principle:
Atoms of different elements absorb characteristic wavelengths of light
to be transformed to the exited state.
The greater the number of atoms in the vapor, the more radiation is
absorbed.
Components:
 The light source
 Flame
 Monochromater
 Photomultiplier tube
 Detector
Function of each component:
 Light source: produce the right mix of wavelengths to be absorbed
by specific atoms in the sample.
 Flame: Atomization of the sample.
 Monochromator: selection a particular wavelength of light for
observation).
 photomultiplier tube: used to strengthen the signal intensity.
 Detector: transform light to electrical impulses.
Differences between atomic absorption spectrophotometer and the
flameless atomic absorption spectrophotometer:
 They differ in how the compounds are atomized.
 In one case a flame is used to produce individual atoms.
 In the other it is carried out by putting the sample in a small
graphite tube and passing an electrical current through to heat it.