LAB #2: INTRODUCTION TO THIN LAYER AND COLUMN CHROMATOGRAPHY: THE SEPARATION OF FERROCENE AND ACETY

Prelab
*Before lab begins, read the following:
Zubrick’s Chapters on Chromatography, Thin-layer Chromatography and Wet-column Chromatography (Ch 26-28). Note that you won’t have to prepare TLC plates; we buy them already made, so you just have to spot and develop them.
Zubrick’s chapter on Infrared Spectroscopy (Ch 33). We will do our sample preparation differently (directions are at the end of this experiment).
*Make a table of physical constants and safety considerations.
*Draw the structures of ferrocene and acetylferrocene.
*Design a flow chart outlining your activities in lab.

Grading.
Completion 5 pts
(includes lab and a report turned in on time)
Prelab 3 pts
Experimental 2 pts
Results 5 pts
Conclusion 5 pts

In lab

In this experiment you will use chromatography to separate a mixture of ferrocene and acetylferrocene according to their differing polarities. Normally, you would first perform thin layer chromatography to determine what would be the best solvent to use as the eluent in column chromatography. To save time, so you will only need to perform TLC at the end to assess how well you separated the mixture.
Assemble the apparatus as shown at the end of this experiment. Note that your column has a porous frit in the bottom, so you do not need to add cotton or glass wool as described in Zubrick. Prepare the column as directed (see section in Zubrick called Preparing the Column). As the eluent you will use petroleum ether, a mixture of C6H14 isomers. The alumina should be added in a fine, steady stream for best results.

Preparing the Sample for Chromatography:
Usually the sample is added exactly as described in Zubrick (Compounds on the Column); however this sample is too insoluble in pet. ether to add this way. Instead, you will get the sample to stick to some alumina and add it to the column as a powder.
Dissolve 90 mg of the ferrocene/acetylferrocene mixture in the minimum amount of dichloromethane (methylene chloride). see note*** Add to this solution, 300 mg of alumina. Stir and evaporate the dichloromethane under a gentle stream of nitrogen.
Drain the solvent level just to the top of the sand and add your sample/alumina powder to the top of the column. To wash this sample onto the column, add 1 mL of pet ether 2 or 3 times, letting the solvent level drain just to the top of the sand before each new addition. At that point you can fill the column with as much eluent as you need and start collecting the compounds in 10 mL Erlenmeyer flasks.
When the first compound has been collected, begin to add a 1:1 mixture of pet ether:t-butyl methyl ether as the eluent.

***note. While the compound is dissolved in CH2Cl2, spot an alumina TLC plate with the mixture and set the TLC plate aside. You will be doing Multiple Spotting along with the two pure products after column chromatography. We have pre-prepared TLC plates and pre-made capillaries (ignore Preparation of Plates and The Plate Spotter and just follow Spotting the Plates). Because this solution is dilute, you may need to touch the capillary to the same position 5-10 times (drying after each time) in order to get enough material on the plate so you can see it well.

When finished, spot the previous alumina TLC plate with the purified fractions of ferrocene and acetylferrocene. Elute this TLC plate with pet ether in the TLC chamber provided in lab. The spots visible on the developed plate fade pretty quickly, so circle them with a soft pencil, and mark the solvent front, too.
Transfer the separated ferrocene and acetylferrocene to two round bottom flasks and evaporate the solvents under vacuum with the rotary evaporator. Transfer the products using a small amount of the appropriate solvent and store them in an open preweighed vial in your lab drawer so that the solvents can evaporate. Tape the vials to the sides of your drawer so that they will not spill. When the solvent has fully evaporated, weigh the vials again in order to determine the percent yield and the percent composition.

Postlab.
Calculate the percent composition of the ferrocene/acetyl-ferrocene mixture and obtain the melting point of acetylferrocene (ferrocene sublimes). Calculate the percent recovery of the entire mixture. Include the TLC plates for the mixture, ferrocene, and acetylferrocene. Calculate the Rf values for all three. These data should go in your results tables. Comment briefly on this procedure and the quality of the separation.
Obtain infrared spectrum of ferrocene and acetylferrocene. Include these spectra with your lab report. These spectra should always have your filename (automatically printed) and a title (can be handwritten). Make data tables for each IR spectrum including all signals of major importance. The IR correlation chart in Zubrick will tell you what types of bonds are present in your sample. You may also compare the IR spectra to the spectra in the Aldrich Library of Infrared Spectroscopy (available in the organic lab). Comment on the purity of both fractions based on TLC and IR spectra. Discuss possible sites of error in your procedure.

DISPOSAL PROCEDURES
Ferrocene and acetylferrocene will be collected in containers in the hood. Alumina from the chromatography columns will be collected in a waste container in the hood.
t-butyl methyl ether and petroleum ether from the chromatography will be collected in a labeled container.
IR waste can be collected in a waste container by the IR.



INSTRUCTIONS FOR TAKING THE IR SPECTRUM OF A SOLID SAMPLE

1) Transfer about 100 mg of dry KBr (from dessicator bottle) into the mortar and grind this until it has the consistency of fine powder. Be sure to put the KBr into the dessicator.

2) Pack the end sample holder with ground KBr: the surface should be evenly flat.

3) Add 1-2 mg of solid sample to the ground KBr and grind it until the total is homogeneous.

4) Pack the middle holder with KBr and sample. With brush, remove excess Kbr and sample from the holder, being careful not to disturb the flat surfaces of sample and background.

5) Carefully place the holder into the diffuse reflectance attachment and situate the KBr holder at the center of the device.

6) On the computer follow the normal instructions for getting into the program and take a background scan.

7) Move the holder such that the sample + KBr are at the center of the device.

8) Take spectrum and process it as usual. Do not remove the sample holder from the Diffuse Reflectance attachment until you have Quit.

 



Directions for IR Operation

To begin:

1. Check to see that the instrument is ready to collect a spectrum. If it is in standby, press stop/rest. If you need to log on to the computer, a common username and password is posted by the monitor.
2. Double clink on the Winfirst icon, which brings a Control Panel box (with a green SCAN button on it) onto the screen, with the Winfirst page in the background. If the control Panel is not there, pull down Tools from the menu bar and click on Control Panel.

To take a background spectrum:

3. To obtain a background spectrum with a solid sample, place the sample cup with the pure ground KBr in the beam. If you are working with a liquid cell or salt plates, you will take your background scans with nothing in the beam.
4. Choose Background in the Control Panel and click on Scan. While this is happening, the LCD on the spectrophotometer should read start background interferogram.

To take a sample spectrum:

5. When the Control Panel appears again on the screen, you are ready to take a spectrum of your sample. If you are taking the spectrum of a solid sample, position the sample cup containing a mixture of your sample ground with KBr in the diffuse reflectance attachment. If you are using salt plates, place a drop of your sample between the two plates and position the plates on the salt plate holder.
6. Click on Scan. When the spectrophotometer is finished collecting scans, there will be a prompt asking you for a file name. Type in your filename and hit enter. Your filename should be your three initials followed by a dash, the experiment number and the spectrum letter. (e.g., JBK-1a, JBK-1b, etc.)
7. An absorbance spectrum will appear on the computer screen. To convert from absorbance to transmittance, click on Math, then Convert, then Transmittance.

At this point, check the quality of your spectrum:
At 4000 wavenumbers, the transmittance should be greater than 80% and your strongest absorption between 4000 and 1500 wavenumbers should be between 20% and 60%.

If your strongest absorption is greater than 60%, you need more sample.

If your strongest absorption all the way to the bottom of the chart and has square corners, you have too much sample.



8. To plot, click on Plot under File. Then click on plot and enter your name, date, the sample name and any comments. When you click on ok, the plotting will begin. You may need to click ok again.


To quit:

You may quit while the printer is operating by selecting close under the spectrum icon. You can then begin collecting the background for the next spectrum (even while the printer is still working).

If you are using salt plates, rinse them with ANHYDROUS acetone over the waste container and return the salt plates to the dessicator.

If you are using the diffuse reflectance attachment, clean the solid from the sample cups with a spatula and/or brush.

If no one is immediately following you, touch standby on the instrument. This reduces wear and tear on the moving mirror.

Do NOT turn anything off. Do not log out or even close the Winfirst program.



Notes to faculty and TAs:

If there is an error message on the spectrophotometer, remove everything from the beam’s path (sample or diffuse reflectance attachment) and open Winfirst and the Control Panel. Click on Diagnostics, then Reset Bench. The instrument will go through a reboot cycle and when it is complete, the LCD on the instrument will read ready to collect spectrum. To ensure that the instrument is communicating with the computer, click on Init Bench. If you get an error message, ignore it and initialize again. Then Exit Diagnostics.