Spectrophotometry - Chemistry LibreTexts
An introduction to Absorption / Transmission / Reflection Spectroscopy This more qualitative application usually The energy and wavelength of absorbtion is defined by the difference between energy levels of an electronic transition. It is not always advisable to convert transmission data to absorption data as . If you calculate from the below link provided by the other researchers above you . of the samples and plotted on a logarithmic absorbance scale for quantitative. Tutorial on Reflection, Transmission, and Absorption of Light. Thus, these three processes can either be quantified for monochromatic radiation (in The reflectance r is defined by the ratio of reflected radiant power to incident radiant power.
Honor students should be able to do the entire activity. Expected Student Background This activity will probably be the first exposure students have with instrumentation. The experimental procedure is rather straightforward and should not pose great difficulty to students. By this point in the course, students should have the experience with routine laboratory procedures involving the use of glassware, how to clean glassware, and so forth.
Time Teacher preparation time to mix the 0. The spectrophotometers should be checked prior to use.
The estimated time for students to perform the activity is50 min to complete Part I. Additional time up to 50 min may be required for Part II, particularly if students are required to prepare their own calibration curves.
- Principles of Spectrophotometry
Safety goggles should be worn during the activity. The chromium III nitrate solution is moderately toxic and should not be poured into the drain. Students are instructed to return the solution actually used in the measurements back into the original container. If there is some possibility that the solution supply will be contaminated, then an alternate disposal procedure should be devised. If it has not been contaminated, the solution may be saved for later use.
There are several sources referred to in SourceBook that give recommended disposal procedures see Safety section. Local and state regulations must be followed. In view of the disposal problems, only the actual volume of solution needed with a modest excess should be prepared. Check the spectrophotometers to be certain they are operating correctly.
Allow a 20 min warm-up period prior to use. To prepare the solution, use 2. Weigh the solute carefully. The unknowns are prepared by successively diluting the 0.
Use these solutions to prepare a calibration curve and student unknowns. Use a buret to measure all volumes. Make the dilutions according to the following table. Pre-Laboratory Discussion Demonstrate for students the techniques involved with using a spectrophotometer. In particular show them how to clean, fill, and place the test-tubes or cuvettes in the instrument.
A transparency master showing a front view of the Spectronic 20 " with the controls identified is in the Appendix. Show students how to use the calibration curve to determine an unknown concentration.
Students should have little difficulty in preparing a data table. For Part I a simple three column table is needed. Part II needs a simple table to record the wavelength used, the percent transmittance recorded and the calculated absorbance.
If necessary, help students design a data table. A useful activity for the pre-laboratory portion of the activity is described in Demonstration 5 in the Atomic Structure module.
Reflection, Transmission, and Absorption
A beveled piece of white chalk is placed in a sample tube in the cell holder of the Spectronic Looking "down the tube" allows one to see the color of the selected light radiation. The monochromator that is part of the Welch ChemAnal " System is another good way to show the colors of light used. One advantage of the ChemAnal " monochromator is that the lid swings out of the way so that the "works" inside can be viewed directly while the instrument is being used, removing some of the "black box" aspects of the instrument.
Teacher-Student Interaction Once students begin the laboratory activity, circulate among them and watch that correct techniques are being used. Pertinent questions may be asked at this time. When monochromatic light light of a specific wavelength passes through a solution there is usually a quantitative relationship Beer's law between the solute concentration and the intensity of the transmitted light, that is, where I sub 0 is the intensity of transmitted light using the pure solvent, I is the intensity of the transmitted light when the colored compound is added, c is concentration of the colored compound, l is the distance the light passes through the solution, and k is a constant.
If the light path l is a constant, as is the case with a spectrophotometer, Beer's law may be written, where k is a new constant and T is the transmittance of the solution.
There is a logarithmic relationship between transmittance and the concentration of the colored compound. Most spectrophotometers have a scale that reads both in O.
As suggested by the above relationships, the absorbance scale is the most useful for colorimetric assays. Using a Spectronic 20 spectrophotometer The Spectronic 20 spectrometer is widely used in teaching laboratories. The specific instructions will differ with other models, but the principles remain.
The instrument must have been warm for at least 15 min.Explain Construction and Working of Atomic Absorption Spectrometer (AAS)
The power switch doubles as the zeroing control. Use the wavelength knob to set the desired wavelength. Wipe the tube containing the reference solution with a lab wipe and place it into the sample holder. Close the cover and use the light control knob to set the meter needle to "0" on the absorbance scale.
Remove the reference tube, wipe off the first sample or standard tube, insert it and close the cover. Read and record the absorbance, not the transmittance. Remove the sample tube, readjust the zero, and recalibrate if necessary before checking the next sample.