Experiment 11:

Spectrophotometry of Co

2+

Introduction:
A very wide variety of analytical techniques exist to identify and quantify chemical substances.
Modern instrumentation can separate hundreds or thousands of components into individual species, identify
each one, and calculate the exact amounts present. Paper chromatography, a technique we learned earlier
this year, is a simple variation on a separation method which could be used.
Qualitative analysis is a branch of chemistry that determines what chemicals are present.
Quantitative analysis tells how much. Spectrophotometry is a quantitative method that is used
to determine very low concentrations, usually in the 10-3 to 10-7 M ranges. As its name implies, spectrophoto-metry measures the light absorbed in a particular part of the electromagnetic spectrum (usually the
visible spectrum from about 350 nm to 700 nm). The method used is based on Beer's Law, which states
that for a specific wavelength
A C and A b
where A = absorbance, b = path length, and C = concentration (moles/L)
Therefore, A = b C
where is the = coefficient of extinction
With a little reflection, experience will tell you that this is obvious. Consider two test tubes, X and Y,
with different concentrations of a colored chemical species. Hold them both up to the light. Which one is
darker? Logic tells you that the darker one is more concentrated; and if it's twice as dark, it must be twice
as concentrated. But why does test tube X appear darker? When you hold it up to the light, the light
passes through the test tube and absorbed by a small amount every time a light particle encounters a lightabsorbing (colored) chemical species. The more light absorbing particles, the greater the concentration,
and the more that the light intensity is diminished as it emerges on the other side of the test tube.
Refer to Figure 10.1. The transmittance is defined for a sample as T =

I
. The "lost" intensity has
Io

been absorbed and the spectrophotometer measures this absorbance. Notice that the effect is also
dependent oh how far the light must travel. The path length, b, is usually measured in centimeters, and the
standard cuvette is exactly 1 cm wide.

Figure 11.1

Spectrophotometry is probably more widely used than any other analytical technique and is the
method of choice in the field of medicine. Chemical substances that are not colored can often be "tagged"
by reacting them with a dyeing agent. For example, diabetics can monitor their blood sugar levels at home
with a simple portable spectrophotometer. They take a blood sample by finger pin-pick and touch the drop
of blood to a strip of paper. Chemicals in the paper react with glucose. The end of the paper is placed in
the device and the absorbance of the glucose-colored species is recorded. The device then displays a
calculated blood sugar level. Thousands of laboratory tests are conducted in hospitals and clinics by this
technique. Some blood tests can measure 24 or more different substances on one sample relatively
quickly.
In this experiment you will determine the wavelength of maximum absorbance for Co2+ ions,
construct a Beer's Law calibration curve, and determine the concentration of an unknown Co2+ solution.

Procedure
Materials
Apparatus
100-mL volumetric flasks
cuvettes
10-mL graduated pipet
pipet bulb
spectrophotometer
Kimwipes

Chemicals
CoCl2.6H2O

Part I: Wavelength of Maximum Absorbance

1. Prepare 250.00 mL of 0.150 M CoCl2 in a 250-mL volumetric flask.
2. Transfer about 5.0 mL of the 0.150 M CoCl2 to a cuvette making certain that it reaches to a level at
the distance up the white dot on the side of the cuvette.
3. Prepare a "blank" by placing 5.0 mL of distilled water in another cuvette.
4. Obtain the absorption spectrum of aqueous Co2+ by measuring the absorbance of the CoCl2 solution
at 25 nm intervals between 400 nm and 700 nm. Go back and read at 5 nm intervals in the 25 nm
interval that indicates the wavelength of maximal absorbance.
5. Remember to "zero" the spectrophotometer at each wavelength. Also, remember to adjust the
spectrophotometer to 100% transmittance, using the blank, at each wavelength.
6. Record the data in Table 11.1.

Part II: Beer's Law Calibration Curve

1. From the 0.150 M CoCl2 stock solution prepare the following dilutions in 100-mL volumetric flasks:
0.120 M, 0.090 M, 0.060 M, and 0.030M.
2. As to save on the number of volumetric flasks. Pour the prepared solutions into a 150-mL beaker
each time. Rinse the volumetric and prepare the solution higher in concentration.

4. Construct a Beer's Law calibration curve using a regression line to obtain the slope.

Part III: Determination of Concentration of Unknown Co2+

1. Obtain a CoCl2 solution of unknown concentration.
2. Measure the absorbance and use interpolation to determine the unknown concentration.

Experiment 11

Name _______________________

___________________________
Date

Report Sheet

_______________________

Period _____________

Spectrophotometry of Co

2+

Prelab Questions
1. In your own words, define the following terms: spectrophotometry, absorbance, transmittance, and
pathlength.
___________________________________________________________________
___________________________________________________________________
___________________________________________________________________
___________________________________________________________________
___________________________________________________________________
___________________________________________________________________
___________________________________________________________________
___________________________________________________________________
2. The absorbances of a series of solutions containing Cr3+ were measured at a fixed wavelength and a
path length of 1.1 cm. The following data were recorded:
Tube #
mL 0.100 M Cr3+
solution
mL water
Absorbance

4.0

3.0

2.0

1.0

1.0

2.0

3.0

4.0

0.394

0.193

0.807

0.608

[Cr3+] M

a.) Complete the above table by calculating the concentration of each solution.
b.) Construct a Beer's Law calibration curve.
c.) Determine the slope and the value for .

d.) What would be the concentration of a Cr3+ solution determined to have an absorbance of 0.654.
3. When 10.0 mL of a Cr3+ solution was diluted to 500 mL, an absorbance of 0.126 was measured. Both
the wavelength and pathlength were as described in question 1. Determine, mathematically, the
concentration of Cr3+ in both the diluted solution and the undiluted solution.
4. The absorbance of a solution of Cr3+ is 0.224 at the wavelength and path length described in question
What absorbance would occur if the pathlength were changed to 4.0 cm?
5. The percent transmittance obtained from a Cr3+ solution is 20.0% at the wavelength and pathlength
described in question 1. Calculate the absorbance and [Cr3+]. (Hint: Absorbance is defined as the
negative logarithm of transmittance; A = -log T or 2-log(%T)).

Data and Observations

Table 11.1: Data for absorption spectrum of aqueous Co2+
Wavelength (nm)
400
425
450
475
500
525
550
575
600
625
650
675
700

Absorbance

Table 11.2: Data for Beer's Law calibration curve of Co2+

Concentration Co2+

Absorbance

0.150
0.120
0.090
0.060
0.060

Interpolated Concentration value for Unknown # _________

Absorbance _______________
Interpolated Concentration _________________

Data Analysis
1. Plot a graph of absorbance vs. wavelength using the data from Part I. Draw a smooth curve to fit the
experimental points. Identify the maximum in the absorption curve to the nearest multiple of 25 nm and record.
2. Construct a Beer's Law calibration curve for the data in Part II. Apply an manual curve fit to determine the slope
and the equation of the "best-fit" line. Measure the pathlength of the cuvette and calculate the value of .

Questions:
1. Calculate the concentration of Cl- in the undiluted sample of your unknown solution.
2. Why must the regression line pass through the origin in a Beer's Law plot?
3. What experimental problems would confront you if you attempted to determine the concentration of the
metal ion in solutions of each of the following reagents?

a.) Cr3+ which has a maximum absorbance at 407 nm with

with

= 13 M(cm)

= 15 M(cm) and another at 574 nm

1
b.) Mn2+ which has a maximum absorbance at 530 nm with = 0.050 M(cm)