Knutsen's Chem 140 Lab: Visible Spectroscopy

Experiment: Measuring Concentrations Using Visible Spectroscopy

Purpose
In this experiment you will learn to use visible spectroscopy to determine the concentration of a solution, and will use MS Excel to analyze your experimental data. You will also gain additional experience in preparing solutions and using molarity.

Introduction
There are many different chemical methods that can be used to determine the concentration of a solution. You have already used one method, titration, to find the concentration of Ca⁺² ions in a sample of water. In this lab you will use a method called visible spectroscopy to determine the concentration of a solution of CoCl₂. This method relies on the fact that colored solutions will absorb light, and that the amount of light the solution absorbs is directly proportional to its concentration.

This relationship is expressed by Beer’s Law,

A = kc

where A is the Absorbance (amount of light absorbed), k is a constant, and c is the concentration of the solution. In other words, the higher the concentration, the higher the absorbance. From this relationship you should be able to see that a graph of Absorbance (y–axis) vs concentration (x–axis) should be a straight line.

You will use an instrument called a spectrophotometer to measure the amount of light the solution absorbs. The spectrophotometer can emit light of several different wavelengths. Usually there is one wavelength at which the absorption of light is most efficient. Your first task will be to determine this optimum wavelength, l_max (pronounced “lambda max”), using a stock solution of CoCl₂ you have prepared.

Once you have determined l_max, you will prepare several standard solutions by diluting the stock solution. The absorbance of these standard solutions will be measured and used to prepare a calibration curve. The calibration curve can then be used to determine the concentration of an unknown solution.

Note: In this laboratory you will be working with a lab partner.

Procedure

Using the solid CoCl₂•6H₂O provided, prepare a stock solution of 250.0 mL of 0.1500 M CoCl₂•6H₂O. This solution must be prepared with care, for the success of the experiment is dependent upon the precision with which you mix solutions. Be sure to record your calculations in your lab book.

Using 50.0 mL volumetric flasks, prepare five standard solutions from the stock solution, each with the concentrations listed below. You may want to check with the instructor to see that you have done your calculations correctly before preparing the solutions. Use tape to label the solutions with your name. Again, be sure to record your calculations in your lab book.

Solution	Concentration (M)
Solution #1	0.1200
Solution #2	0.0900
Solution #3	0.0600
Solution #4	0.0450
Solution #5	0.0300

Instructions on the use of the spectrophotometer are provided in the lab. Be sure to follow them carefully. Use your stock solution to determine l_max as follows:
1. Fill the sample tube with your stock 0.1500 M CoCl₂•6H₂O solution, and place it in the spectrophotometer.
2. Set the mode to absorbance.
3. Set the wavelength at 400 nm, and note the absorbance of the sample in your notebook. Record additional readings at 450, 500, 550 and 600 nm.
4. Return to the wavelength setting with the maximum reading, and then explore on either side of that wavelength to see if there is one that gives a higher reading. The setting with the highest reading is l_max. Once it has been determined, leave the machine set at this wavelength for the duration of the experiment.
Calibrate the spectrophotometer according to the instructions in the lab.
Measure and record the absorbance of each of the five standard solutions.
Obtain an unknown and record the number in your lab notebook. The unknown will be quite concentrated and will likely have an absorbance greater than any of the standard solutions. You must dilute the unknown such that its absorbance falls within the range of the five standard solutions. It will be up to you to determine the proper dilution factor; be sure to do it in the same manner you made Solutions 1–5. Do be sure to record, in your laboratory notebook, how you dilute your sample since you will need this information later.
Measure and record the absorbance of the diluted unknown.

Note: Steps 5–7 must be completed on the same day! Do not start step 5 unless you are sure you have time to complete step 7 as well.

Analysis and Calculations

Use MS Excel to prepare a graph of absorbance vs concentration for the five standard solutions. Make sure that the chart has a name, the axes are labeled (including appropriate units), that there are major and minor gridlines, and then place the chart on a new sheet. Determine the slope and intercept using linear regression, and include the equation for the line on your graph. Remember to include a copy of the graph (with the best regression line and equation of the line) in your laboratory report.

Use the equation for the line to calculate the concentration of your diluted unknown. Then determine the concentration before dilution and check your results with your instructor. For maximum credit your results must be within 5% of the true value.

In your discussion be sure to evaluate your graph, with attention to the precision of your data points. Is your graph consistent with Beer’s Law?

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