Atomic Absorption Spectrophotometer Hossein Piri Department of Biochemistry and Genetics, School of Medicine, Qazvin University of Medical Sciences, Qazvin, Iran September 2012

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

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

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

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.

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.

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.

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 and that emitted by excited atoms in the flame. To do this, the hollow-cathode light beam is modulated by inserting a mechanical rotating chopper between the light and the flame or by pulsing the electric supply to the lamp.

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.

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

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.

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.

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.