# Montgomery College Chemistry a Molecular Approach Worksheet

Spectrophotometric Determination of Iron in a Vitamin PillIntroduction
Iron is one of the essential elements in the human diet. In the body, iron is present in the form of
iron(II) or iron(III) ions. Natural sources of dietary iron are meat and leafy, green vegetables. Insufficient
dietary iron results in the condition of anemia. Many people take vitamin pills containing iron to
supplement their dietary intake of iron.
The technique of spectrophotometry is a method of instrumental analysis. Many different types
of spectrophotometers are available that transmit light in different regions of the electromagnetic radiation
spectrum. In this experiment, a visible spectrophotometer will be used. A visible spectrophotometer
measures the amount of light that a colored solution absorbs. When a beam of light of a particular
wavelength passes through a colored solution, the amount of light absorbed by the sample depends on
what the substance is that is absorbing the light. The absorbance also depends on the concentration of
that substance in the solution and the length of the path through which the light passes. The relationship
between absorbance, concentration, and pathlength is expressed by the equation known as Beer’s law:
A=εℓ c
where A is the absorbance of the solution, ε is a constant called absorptivity, which is a characteristic of
the substance and the wavelength of the absorption, c is the concentration of the absorbing substance in
the solution, and ℓ is the length of the path through which the light passes. Absorbance is a unitless
quantity, concentration can be expressed in any convenient concentration unit, and the pathlength is
measured in centimeters. The units of absorptivity depend on the concentration units used. In this
experiment concentration is measured in mg/mL. Therefore, the units of ε are mL/mg · cm.
To determine the absorptivity of a particular substance, a calibration curve is constructed. A
series of standard solutions in which the concentration of the absorbing species is known are prepared and
their absorbances are read. A plot of absorbance as a function of concentration is made. If the absorbing
species obeys Beer’s law, the plot will be a straight line, y = mx + b. When the pathlength is 1.0 cm, the
slope of the line is the absorptivity. The absorbance of a solution of unknown concentration can be read
and its concentration determined from the calibration curve.
In this experiment, a commercial vitamin pill will be analyzed for its iron content. The form of
iron present in the vitamin pill, most likely iron(II) sulfate, has a negligible visible absorbance. The
iron(II) will be converted to a species that has a large absorbance, allowing the absorbances of solutions
of low iron(II) concentration to be measured. The reagent 1,10-phenanthroline, Figure 1, reacts with
iron(II) to form a complex with three phenanthroline molecules bonded to one iron(II) ion. This complex
is red-orange and absorbs visible light most strongly at 508 nm. Iron(II) is easily oxidized to iron(III) in
aqueous solution. A reducing agent, hydroquinone, is added to the solution to prevent this oxidation from
occurring. In addition, the acidity of the solution must be controlled to ensure the formation of the
complex. In this experiment, sodium citrate is added to maintain the proper pH of the solution.
A series of standard solutions with known iron concentrations is prepared, the absorbances of the
solutions are read, and a calibration curve constructed. From the calibration curve, the concentration of
iron in a solution made from the dissolved vitamin pill will be determined and the total amount of iron in
the vitamin pill calculated.
Rev. 2
3/30/22 AC
Figure 1. The Structure of 1,10-Phenanthroline
H
H
H
C
C
H
C
C
C
H
C
C
C
C
H
C
N
N
C
H
H
PROCEDURE
coming to lab.
Do procedure 2a before starting procedure 1.
1. Preparation of the standard iron solutions for the calibration curve
a. Using the markings on the side, pour some (about 30 mL) of the stock iron solution into a clean,
dry beaker. Record the concentration of the stock solution.
b. Prepare a series of standard iron solutions as shown in Table 1 in clean 50-mL volumetric flasks.
Pipet 5.00 mL of the stock iron solution into one of the volumetric flasks. Add the sodium citrate,
hydroquinone, and phenanthroline solutions to the flask, then dilute to the mark with distilled
water. Mix well. Prepare the other solutions in a similar manner. Be sure to measure the stock
iron solution with a pipet. Let the solutions stand for about 10 minutes.
c. Pour some of each solution into separate cuvettes, filling each cuvette about three-fourths full.
Set the solutions aside until the vitamin pill solution is prepared.
Table 1. Preparation of Iron Standard Solutions
Solution
Drops citrate
mL hydroquinone
mL phenanthroline
4
mL stock iron
solution
5.00
30
1.00
1.50
3
3.00
18
1.00
1.50
2
2.00
12
1.00
1.50
1
1.00
6
1.00
1.50
Blank
0.00
0
1.00
1.50
Rev. 2
3/30/22 AC
2. Preparation of the vitamin pill solution
a. Obtain a vitamin pill and record its brand and the iron content listed on the label. Put it in a clean
125-mL Erlenmeyer flask. In the hood, measure 25 mL of 6 M HCl in a graduated cylinder and
add it to the flask and boil it gently on a hot plate for about 10 minutes to dissolve all of the
soluble matter in the pill. Remove the flask from the hot plate.
b. In the hood, add 25 mL of distilled water measured with a graduated cylinder to the solution.
Prepare a funnel with filter paper. Filter the solution into a clean 100-mL volumetric flask. Rinse
the Erlenmeyer flask three times with a few mL of distilled water and add the rinsings to the
funnel over the volumetric flask. Then wash the residue on the filter paper with a few mL of
distilled water, letting the washings filter into the volumetric flask. Be sure that the volume of the
solution does not go over the mark on the neck of the flask. When the flask has cooled to room
temperature, fill it to the mark with distilled water and mix thoroughly. This is solution A.
c. Using the markings on the side, pour some (about 20 mL) of solution A into a clean, dry beaker.
Pipet 5.00 mL of solution A into a clean 50-mL volumetric flask. Fill to the mark with distilled
water and mix well. This is solution B.
d. Using the markings on the side pour some (about 20 mL) of solution B into a clean, dry beaker.
Pipet 5.00 mL of Solution B into a clean 50-mL volumetric flask. Add the following solutions to
the flask: 50 drops of citrate, 1.00 mL of hydroquinone, and 1.50 mL of phenanthroline. Fill the
flask to the mark with distilled water and mix well. Let the solution stand for about 10 minutes.
This is solution C. Pour some solution C into a cuvette.
e. Calibrate a spectrophotometer at 508 nm with the blank solution. Read and record the
absorbances of the solutions for the standard curve and solution C.
WASTE DISPOSAL
Pour all waste materials in the waste bottle in the hood.
Rev. 2
3/30/22 AC
Name _______________________
Partner _______________________
DATA
1. Preparation of the standard iron solutions for the calibration curve
Concentration of stock iron solution (include units)
____________________
Absorbances of solutions:
Solution
Absorbance
Blank
_____________
1
_____________
2
_____________
3
_____________
4
_____________
2. Preparation of the vitamin pill solution
Brand of vitamin _________________
Absorbance of solution _____________
Stated iron content _______________
CALCULATIONS
1. Preparation of the standard iron solutions
a. Calculate the iron concentration in the solutions prepared for the standard curve.
Solution
Concentration of Iron (mg/mL)
Blank
____________
1
____________
2
____________
3
____________
4
____________
Rev. 2
3/30/22 AC
b.
Using Excel, create a worksheet and graph, including the best-fit line and the equation for the
line.
2. Amount of iron in the vitamin pill
a. Using the Excel directions, determine the concentration of iron in Solution C. Copy that value in
the space below. Include units.
__________________
b. Calculate the concentration of iron in Solution B. Use the dilution formula, i.e., ccvC = cBvB. The
concentration of Solution C is recorded in part a. Recall the volume of Solution C that was made and
the volume of Solution B that was used to make Solution C. Show your work.
___________________
c. Calculate the concentration of iron in solution A. Use the dilution formula, i.e., cBvB = cAvA. The
concentration of Solution B was determined in part b. Recall the volume of Solution B that was made
and the volume of Solution A that was used to make Solution B. Show your work.
___________________
d. Calculate the number of milligrams of iron in the vitamin pill. (Hint: The dissolved vitamin pill
was used to make Solution A. What was the total volume of Solution A?)
______________________
e. Calculate the percent error in your determination. Use the value stated on the label as the actual
value of iron in the vitamin pill.
______________________
Attach a copy of your Excel work to this lab report. Hand in your Advance Study Assignment and
hand-drawn graph as well.
Rev. 2
3/30/22 AC
Determination of Iron in a Vitamin Pill
Name________________________
1. The concentration of iron in a solution is 0.0400 mg/mL. A new solution is prepared from this
solution by diluting 5.00 mL of the original solution to a final volume of 50.00 mL. What is the
concentration of iron in the diluted solution?
2. A solution is determined to have a concentration of 0.0912 mg/mL of iron. How many milligrams of
iron are contained in 100.00 mL of this solution?
3. Graph the following data using correct graphing procedure. Have the graph fill as much of the paper
as possible, give the graph a title, label the axes with a label and units, draw the best straight line through
the points using a straight edge, and calculate the slope of the line. Plot absorbance on the y axis and
concentration on the x axis. Attach the graph to this paper. (Excel graphs will not be accepted)
Solution
Absorbance
Blank
1
2
3
4
0.000
0.205
0.423
0.618
0.996
Concentration,
mg/mL
0.00
1.00
2.00
3.00
5.00
Using your graph, determine the concentration of a solution with an absorbance of 0.560.
____________________
Slope calculation: Remember to use points on the line, not data points to determine the slope!
Slope = _______________
Rev. 2
3/30/22 AC

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