Atomic Absorption Spectrometry

Background: Atomic absorption spectrometry involves the absorption of radiant energy by neutral atoms in the gaseous state. The element being determined must be reduced to the elemental state, vaporized, and imposed in the beam of radiation from the source. This process is most frequently accomplished by drawing a solution of the sample, as a fine mist, into a flame. The flame thus serves a function analogous to that of the cell and solution in conventional absorption spectroscopy. The light beam from the source is passed directly through the flame.

Quantitative analysis is easily performed by measuring the radiation absorbed by the sample in the flame. Typically a calibration curve is first prepared by measuring the absorbance of a series of solutions of known concentration. The sample(s) is then measured under same conditions. Excellent results are generally obtained. About 65 elements can be determined by atomic absorption with sensitivities down to 0.1 µg/mL in same cases.

Reference: Skoog, Holler and Nieman Principles of Instrumental Analysis

Equipment: Buck model 200A atomic absorption spectrophotometer, Dynac centrifuge, centrifuge tubes.

Reagents: 1.4 N HCl, 100 ppm Stock Solution of Copper (made by diluting the appropriate weight of copper sulfate pentahydrate with 1.4 N HCl), 20% (v/v) trichloroacetic acid.

Unknown: Student's will choose there own unknowns. Obtain samples that will have relatively low, yet measurable copper concentrations. The procedure given below will work for almost any sample. Biological or environmental samples are usually the most interesting. Consult with the laboratory Teaching Assistant or Instructor for details and/or procedures.

Procedure

Begin the digestion of your real sample. There are two 30 minute periods of incubation during which time you will continue with the instrument startup procedure below.

Sample and Standards

1. Make 10, 20, and 40 ppm Cu standards from the stock solution by diluting with 1.4 N HCl.
   
   
2. To five labeled centrifuge tubs, add 2.0 mL of the following solutions: 0 (Blank), 20, 40, 50, ppm Cu Standards. To the fifth tube add 2.0 mL of unknown sample. Add 2.0 mL of 1.4 N HCl to all tubes and mix. Allow tubes to sit for 30 minutes at room temperature.
   
   
3. Add 2.0 mL of 20% trichloracetic Acid (TCA) to ALL tubes and allow to sit for an additional 20 minutes.
   
   
4. Centrifuge all tubes for 10 minutes at 2000 RPM. (Use setting of 80 on Dynac Centrifuge to provide 2000 RPM)
   
   
5. Pour off supernatent into labeled clean centrifuge tubes.

Measurements

See your instructor, he or she will demonstrate the operation of the Buck 200A Atomic Absorption Spectrophotometer.

• Adjust the gas flow and light the flame with the assistance of the instructor according to the instructions listed under Flame Types and Lighting Instructions. DO NOT LIGHT THE FLAME UNTIL YOU ARE READY TO ASSAY YOUR TREATED SAMPLES AND STANDARDS.

• Be sure the instrument is set to the primary emission line of Copper (~324 nm). Use only the minimum volume of sample necessary to achieve a stable absorbance value (usually only 10 seconds aspiration time is required).

• Adjust the meter to read zero absorbance while aspirating the Blank solution. Next, read each solution while aspirating it into the flame. It may be necessary to repeat the measurements several times to obtain reproducible values. It is also advised that the zero absorbance setting be rechecked every 3-4 samples for drift.

• Take readings of absorbance of each solution, beginning with the tube 'Blank'.

• Return the instrument to the secondary emission line as indicated by the COPPER INFORMATION SHEET. Once again, set the zero absorbance level and read the absorbance of each of the copper samples.

• Adjust the meter to read zero absorbance while aspirating the Blank solution. Next, read each solution while aspirating it into the flame. It may be necessary to repeat the measurements several times to obtain reproducible values. It is also advised that the zero absorbance setting be rechecked every 3-4 samples for drift.

• After your samples have been measured to give sufficient reliability, allow the flame to aspirate distilled water for 5-10 minutes to wash out the burner head of any salts or particulate material.

• Finally, shut the burner off with assistance from the instructor. Then, turn off the electronics with the main power switch.

Questions

1. What was the Cu concentration in the unknown sample?
   
   
2. Was the standard curve you used to calculate the Cu concentration in the unknown sample adequate? If not, how would you have changed it?
   
   
3. Why did you add trichloroacetic acid to the sample?
   
   
4. Were there any sources of interference? If so, what were they?