Atomic Absorption Spectrophotometer

1. Atomic Absorption Spectrophotometer Hossein PiriDepartment of Biochemistry and Genetics, School of Medicine, Qazvin University of Medical Sciences, Qazvin, Iran September 2012

2. Atomic absorption spectrophotometer • Is used to measure concentration by: • detecting absorption of electromagnetic radiation by atoms rather than by molecules.

5. Usual light source • Hollow-cathode lamp, consists of: • Anode • Cylindrical cathode • Inert gas (helium or argon) • When voltage is applied: • gas is ionized • Ions collide with metal, and cause atoms to be excited • When they return to ground state, light energy is emitted that is characteristic of metal in cathode • Separate lamp is required for each metal

6. Electrodeless discharge lamps • Are a relatively new light source • A bulb is filled with argon and element to be tested • A radiofrequency generator around the bulb supplies the energy to excite the element

7. Sample • Sample must contain reduced metal in atomic vaporized state • This is done by using the heat of a flame to break chemical bonds and form free, unexcited atoms • The flame is the sample cell in this instrument, rather than a cuvet. • There are various designs; however, the most common burner is the premix long-path burner.

8. Sample • The sample is aspirated as a spray into a chamber, where it is mixed with air and fuel. • This mixture passes through baffles, where large drops fall and are drained off. • Only fine droplets reach the flame. • The burner is a long, narrow slit, to permit a longer path length for absorption of incident radiation. • Light from the hollow-cathode lamp passes through the sample of ground-state atoms in the flame.

9. Sample • The amount of light absorbed is proportional to the concentration. • When a ground-state atom absorbs light energy, an excited atom is produced. • The excited atom then returns to the ground state, emitting light of the same energy as it absorbed. • The flame sample thus contains a dynamic population of ground-state and excited atoms, both absorbing and emitting radiant energy.

10. Mechanical rotating chopper • The emitted energy from the flame will go in all directions, and it will be a steady emission. • Because the purpose of the instrument is to measure the amount of light absorbed, the light detector must be able to distinguish between the light beam emitted by the hollow-cathode lamp andthat emitted by excited atoms in the flame. • To do this, the hollow-cathode light beam is modulated by inserting a mechanical rotating chopperbetween the light and the flame or by pulsing the electric supply to the lamp.

11. Detecting the pulsed signal • Because the light beam being absorbed enters the sample in pulses, the transmitted light also will be in pulses • There are, therefore, two light signals from the flame— • Alternating signal from hollow-cathode lamp • Direct signal from flame emission • Interference from the constant flame emission is electronically eliminated by accepting only the pulsed signal from the hollow cathode.

12. Monochromator • Is used to isolate the desired emission line from other lamp emission lines • It serves to protect the photodetector from excessive light emanating from flame emissions • A PM tube is the usual light detector

13. Flameless atomic absorption • Uses an electric furnace to break chemical bonds (electrothermal atomization) • An electric current, evaporates the solvent, ashes the sample and, finally, heats the unit to incandescence to atomize the sample. • This instrument, like the spectrophotometer, is used to determine the amount of light absorbed. • Again, Beer’s law is used for calculating concentration.

14. Problems • For example, phosphate may interfere with calcium analysis by formation of calcium phosphate. • This may be overcome by adding cations that compete with calcium for phosphate. • Routinely, lanthanum or strontium is added to samples to form stable complexes with phosphate.

15. Problems • Another possible problem, • Ionization of atoms, which can be decreased by reducing the flame temperature. • Matrix interference, can be another source of error. • This interference may be overcome by pretreatment of the sample by extraction.