SURVEY OF CHEMISTRY LABORATORY I

CHEM 1151L
SPECTROPHOTOMETRIC DETERMINATION
OF CHROMIUM

CAUTION!!!
BE SURE TO WEAR SAFETY GLASSES AT
ALL
TIMES IN THE LABORATORY
NO EXCEPTIONS TO THIS RULE!

SPECTROPHOTOMETRY
- To measure the absorbance of different concentrations of
chromium
Students will
- Prepare chromium-EDTA complex solutions (colored complex)
- Determine the wavelength of maximum absorbance of the
colored complex (max)
- Determine the concentration of an unknown chromium solution

CHROMIUM SOLUTION
- The chromium ion (Cr3+) react with EDTA
(ethylenediaminetetraacetic acid) to form a colored complex
- Heat is required for reaction to go to completion
- The complex absorbs light at particular wavelength and reflects
others
- Intensity of color depends on the concentration of chromium
- The color intensity can be used to measure the concentration of
solutions

BEERS LAW
- The absorbance of radiation (light) is related to the
Wavelength of radiation
and
Concentration of the absorbing species
- The equation for the relationship between the optical
absorbance and concentration is known as
Beers Law

BEERS LAW
A = kbc
- A is the optical absorbance
- k is a constant known as the molar absorptivity
- b is the pathlength (distance through which light passes sample)
- c is the concentration (molarity) of the solution

BEERS LAW
A = kbc
- Optical absorbance (A) is measured using the spectrophotometer
(read directly from the instrument)
- The spectronic 20 will be used for this experiment
- The small test tube that holds the sample is known as the cuvet
- The diameter of the cuvet = 1 cm
- Implies the pathlength (b) for this experiment = 1 cm
- Beers law reduces to A = kc

PROCEDURE I
DETERMINATION OF THE WAVELENGTH
OF MAXIMUM ABSORBANCE
(max)

PREPARATION OF Cr3+-EDTA COMPLEX

- Set up a hotplate at your work
station
- Fill a beaker with water (about
half-filled)
- Place the beaker of water on the
hotplate to heat up

PREPARATION OF Cr3+-EDTA COMPLEX

- Use the buret for volume
measurements
- Obtain a regular large test tube
(not the small test tubes near
the instrument)
- Measure 5.00 mL of chromium
nitrate solution and 5.00 mL
of EDTA solution into the test
tube

PREPARATION OF Cr3+-EDTA COMPLEX

- Place the test tube in the

boiling water bath for at
least 10 minutes

PREPARATION OF Cr3+-EDTA COMPLEX

- Remove and cool the solution

by placing the test tube in
cold water

SPECTRONIC 20

- Refer to the General Procedure for

Spectrometer Use in Appendix II
- Turn on the Spectronic 20 instrument to
warm up for at least 5 minutes before
use

max DETERMINATION
- Obtain two clean cuvets (small test
tubes near the instruments)
- Fill one cuvet with EDTA solution
- Fill the other cuvet with the
prepared colored complex
- Fill each to at least three-quarters
full

max DETERMINATION

- Avoid holding the sides of the

cuvets (do not leave finger
prints on them)
- Carefully clean the sides with
kim wipes
Click on picture to play movie

max DETERMINATION
- Set the instrument to 350 nm
- Insert the EDTA cuvet into the
sample holder
- Mark on the cuvet should be in line
with mark on the sample holder
- Make sure instrument is in the
absorbance (A) mode
- Zero the instrument
Click on picture to play movie

max DETERMINATION
- Remove the EDTA cuvet
- Insert the COMPLEX cuvet into
the sample holder
- Mark on the cuvet should be in
line with mark on the sample
holder
- Read and record the abosrbance
displayed
Click on picture to play movie

max DETERMINATION
- Change the wavelength to 360 nm
- Insert the EDTA cuvet into the
sample holder
- Mark on the cuvet should be in
line with mark on the sample
holder
- Zero the instrument

max DETERMINATION
- Remove the EDTA cuvet
- Insert the COMPLEX cuvet into the sample holder
- Mark on the cuvet should be in line with mark on the sample
holder
- Read and record the absorbance displayed

max DETERMINATION
- Repeat steps at wavelength increments of 10 up to 600 nm
- Zero instrument with EDTA cuvet each time wavelength is
changed
- Clean cuvets with kim wipes each time before inserting into
the instrument
- Always aline marks on the cuvets and the sample holder

ABSORBANCE VS WAVELENGTH GRAPH

- Use the Chart Wizard in Excel
- Plot a graph of absorbance verses wavelength
- This is not a straight line so do not join the points with a line
- Only plot the points on scatter and print it out
- Use pencil to join the points with your free hand
- Determine the wavelength corresponding to the maximum absorbance
This is the max of the Cr3+-EDTA complex

PROCEDURE II
DETERMINATION OF THE RELATIOSHIP
BETWEEN ABSORBANCE AND CONCENTRATION
OF THE Cr3+ AT THE max
DETERMINATION OF THE CONCENTRATION
OF AN UNKNOWN Cr3+ SOLUTION

HOMEWORK EXERCISE

Do the following calculations before coming to the next class

- Calculate moles of Cr3+ for test tubes 1 through 5
- Calculate molarities (M) of Cr3+ for test tubes 1 through 5
- Complete the table with moles and molarities of Cr3+

HOMEWORK EXERCISE
Use the dilution method: M1V1 = M2V2
Molarity (concentration) of stock Cr3+ solution = M1 = 0.00750 mol/liter
Final volume after mixing for each test tube = V2 = 10.00 mL
For example
Mixing 2.50 mL (V1) of stock Cr3+ solution with 7.50 mL EDTA
2.50 mL Cr3+ + 7.50 mL EDTA = 10.00 mL total solution = 0.0100 L
Moles of Cr3+ = Stock concentration (Molarity) x Volume of Cr 3+ (liters)
= 0.00750 moles/liter x 0.00250 liter
= 0.00001875 mole of Cr3+
Molarity (M2) = Moles Cr3+ / Total volume of solution (liters)
= 0.00001875 mole/ 0.0100 L
= 0.001875 M (or moles/liter) Cr3+
M means Molarity or moles/liter

HOT WATER BATH

- Set up a hotplate at your work
station
- Fill a beaker with water (about
half filled)
- Place the beaker of water on the
hotplate to heat up

PREPARATION OF KNOWN SOLUTIONS

- Use the buret to measure volumes
of solutions
- Obtain five large test tubes and
label them 1 through 5
- Carefully measure out the Cr3+ and
EDTA solutions directly into the
test tubes using volumes in the
table that was designed as the
homework assignment

PREPARATION OF AN UNKNOWN SOLUTION

- Obtain an unknown sample of 5.00 mL Cr(N03)3 of an unknown

concentration
- Record the unknown number on your data sheet
- Add 5.00 mL of EDTA solution to give you a total of 10.00 mL

HEAT SOLUTIONS

- Place all the test tubes (knowns

and unknown) in the boiling
water bath for at least 10
minutes
- Remove any caps on the tubes
before heating

COOL SOLUTIONS

- Remove and cool the solutions

by placing the test tubes in cold
water

SPECTRONIC 20

- Refer to the General Procedure for

Spectrometer Use in Appendix II
- Turn on the Spectronic 20 instrument to
warm up for at least 5 minutes before
use

ABSORBANCE DETERMINATION

- Obtain one clean cuvet (small test

tubes near the instruments)
- Fill the cuvet with EDTA solution
- Fill to at least three-quarters full

ABSORBANCE DETERMINATION

- Avoid holding the sides of the cuvet

(do not leave finger prints on them)
- Carefully clean the sides with kim wipes

ABSORBANCE DETERMINATION
- Set the instrument to the determined
max
- Insert the cuvet into the sample holder
- Mark on the cuvet should be in line
with mark on the sample holder
- Make sure instrument is in the
absorbance (A) mode
- Zero the instrument

ABSORBANCE DETERMINATION
- Remove the cuvet
- Pour out EDTA into a waste beaker
- Rinse cuvet twice with bits of solution #1
(pour rinsing solution into the waste beaker)
- Fill cuvet with solution #1

ABSORBANCE DETERMINATION
- Wipe sides with kim wipes
- Insert cuvet into the sample holder
- Mark on the cuvet should be in line with mark on the sample
holder
- Read and record the absorbance displayed

ABSORBANCE DETERMINATION
- Repeat steps for the rest of the test tubes including the unknown
- Use the same cuvet for all the samples
- Always pour out used sample back into its tube and rinse twice with the next
solution before refilling (do not rinse with water)
- Clean cuvet with kim wipes each time before inserting into the instrument
- Always aline marks on the cuvet and the sample holder
- Wavelength remains at the max
- Clean up when done and pour all solutions into the appropriate waste bottle

ABSORBANCE VS WAVELENGTH GRAPH

- Use the Chart Wizard in Excel
- Plot a graph of absorbance verses wavelength
- This is the Beers Law line graph
- Line is expected to be a straight line so join the points with a
trendline (the best straight line)
- Include the equation of the trendline

ABSORBANCE VS WAVELENGTH GRAPH

- Use the equation of the trendline and the optical absorbance of

the unknown to determine the concentration of the unknown
- Also estimate the concentration of the unknown visually from
the graph
- Include the two graphs in your laboratory report