SURVEY OF CHEMISTRY LABORATORY I SPECTROPHOTOMETRIC DETERMINATION 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 (λmax) - 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

BEER’S 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 Beer’s Law

BEER’S 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

BEER’S 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 - Beer’s law reduces to A = kc

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

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

λmax 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

Click on picture to play movie λmax 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

Click on picture to play movie λmax 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

Click on picture to play movie λmax 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

λmax 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

λmax 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

λmax 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 λmax of the Cr3+-EDTA complex

DETERMINATION OF THE RELATIOSHIP BETWEEN ABSORBANCE AND CONCENTRATION PROCEDURE II DETERMINATION OF THE RELATIOSHIP BETWEEN ABSORBANCE AND CONCENTRATION OF THE Cr3+ AT THE λmax 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 Cr3+ (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 λmax - 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 λmax - 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 Beer’s 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