Experiment 3b Column Chromatography

How do you find a colorless sample?

Collect fractions of a specific volume and run TLC on them to determine where the compound is and then combine the fractions that contain the compound

Why is it important to limit the sand bath temperature to 60°C when evaporating solvent from your collected fractions?

The reason that the sand bath must be kept at 60 degrees Celsius when evaporating the solvent is because the desired compound may melt or vaporize if the temperature of the sand bath is too high. For example, the melting point of acetlyferrocene is only around 80 degrees Celsius, so if the sand bath were set to 140 degrees Celsius to try to speed up the evaporation process, the acetlyferrocene compound will melt and negatively affect the yield the students calculate. In addition, it is possible that the organic compounds that the students are trying to isolate can decompose into their substituents or ignite if they are heated too rapidly in the sand bath. Obviously, both of these outcomes would introduce large sources of error into the students' results.

Description

a column is filed with a larger amount of adsorbent and the mixture is loaded on top of it. An eluent is allowed to percolate through the column by gravity. As the eluent is moving down the column it carries the soluble compounds with it.

Column Chromatography

a preparative method for separating and isolating compounds from mixtures. Method is used for obtaining compounds from natural sources or for purifying products from reaction mixtures. It is an upside down version of TLC

Ferrocene

is less polar than acetylferrocene

Suppose you are given 150 mg of an unknown mixture that contains three compounds, and you are told to separate the compounds using column chromatography, using alumina as a stationary phase. How would you decide what solvent or solvent system to use?

Given an unknown mixture, the first step would be to run multiple TLC plates spotted with the mixture and run with various mobile phases to determine which solvent results in the best separation of the mixture. Once this has been determined and one or two good solvents have been found, column chromatography can be run. The first solvent in the column chromatography should be the least polar of the chosen solvents, and the more polar of the chosen solvents should be introduced midway into the elution process. Using this method, the components of the mixture should be easily isolated for subsequent analysis.

In running column chromatography under the following conditions, you get results that are less than ideal. Consider the situation in each case and suggest a correction. a.You run a column on a mixture of two unknown halogenated alkenes, but hey both elute from the column at the same time. The solvent used was methylene choride (dichloromethane). b.You run an alumina column on a mixture of a thiol and an amine, but you never get any compound off the column (i.e. neither compound elutes form the column). The solvent was a mixture of petroleum ether and dichloromethane.

A. If the two compounds that you desire to separate elute from the column at the same time, you can conclude that the two halogenated alkenes have too similar of retardation factors for a solvent such as dichloromethane to separate. In order to fix this problem, the student should use two different solvents, starting with a less polar solvent such as petroleum ether and then switch to a more polar solvent such as MtBE. The petroleum ether would elute out the less polar of the two halogenated alkenes while the MtBE would elute out the more polar of the two halogenated alkenes. The use of two different solvents would result in a separation of the two mixed compounds. B. If neither of the two compounds elute off the column, then we can conclude that the solvent is not polar enough to strip the compounds from their affinity to the stationary phase. To fix this, a more polar solvent should be selected for use, but it cannot be too polar otherwise both of the compounds will elute off at the same time. A list of possible solvents to use is located on the back of the carbonless "Organic Chemistry Student Laboratory Notebook".

Procedure

Dry column packing: -Take a 6 inch micro column and clamp the column onto a stand -Using a small plastic filter funnel add sand -Gently tap the column -Add alumina until the column is a little less than half filled Dry column loading: -Weigh 100mg of ferrocene/acetylferrocene mixture and transfer the sample into a small conical vial -Add 100mg of alumnia to the vial and 2-3 drops of MtBE -Stir with a spatula -Transfer the solid mixture into the packed micro column -Cover with an additional layer of sand Eluting the Column: -Place first tube under column -Measure 10mL of petroleum ether in a graduated cylinder -Add the mobile phase (using a plastic pipet) to the top of the column -Continue adding more mobile phase until the column is almost full -Use air pressure to facilitate the solvent through the column -Collect the first solvent to elute from the column until the yellow orange band just begins to elute from the bottom of the column -Switch to new test tube until yellow band has been collected -Switch to third test tube -Stop adding more petroleum ether, start adding more polar solvent, MtBE -Switch to fourth test tube and continue eluting with MtBE until band has been collected TLC Analysis: -Obtain silica gel TLC plates and analyze the individual fractions to determine the extent of separation using a TLC solvent system 15% MtBE and 85% petroleum ether -Transfer fractions to a pre-weighed Erlenmeyer flask with boiling chips

The compounds used in Experiment 3b were both highly colored, so it was easy to see their progress as they moved down the column during elution. Most organic compounds however are colorless in solution. How might you monitor the progress of a column chromatography given a colorless sample?

In order to monitor the progress of a column chromatography if the bands produced are colorless, the student would have to set up a very slow drip rate and run TLC plates as the elution is occurring in order to determine which compounds are being eluted. Based off of the information the student obtains from the TLC plates, they will know when to switch to the more polar solvent in order to elute the more polar compound, and they will know when the elution has been completed.

Ferrocene eluted from the column first but it ended up as the higher of the two spots during TLC analysis. explain.

It is important to note that TLC and column chromatography are almost exact opposites in that the flow of solvent is down the column in column chromatography but up in TLC. Thus, due to the direction of solvent flow in the two methods, column chromatography will have its least polar compounds toward the bottom and its most polar compounds toward the top. TLC, on the other hand, will have its least polar compounds toward the top and more polar compounds toward the bottom. This explains why ferrocene (relatively non-polar) was at the bottom of the columnchromatography and the top of the TLC.

If benzyl alcohol, benzyl acetate, p-nitrophenol, and napthalene were separated by column chromatography with ether on silica gel, which compound would elute first?

Naphthalene would elute first because it is the least polar of the given compounds.

Why does ferrocene elute from the column first? Why was the solvent changed in the middle of the column procedure?

The ferrocene elutes from the column first because it is less polar than the acetlyferrocene and is thus more attracted to the relatively non-polar mobile phase compared to the polar stationary phase. This high affinity for the petroleum ether resulted in the ferrocene eluting out of the column first. The reason that the mobile phase was switched to MtBE in the middle of the column procedure was because acetlyferrocene needed a more polar mobile phase in order to be eluted out of the column. With the more polar mobile phase, the acetlyferrocene had a high affinity for the MtBE and thus moved with the MtBE down the column instead of adsorbing strictly to the stationary phase.

You run a column to separate a mixture of three compounds: naphthalene, o-toluic acid, and fluorenol. The column is run with a solvent system based on hexanes, but which becomes more polar with the addition of more and more dichloromethane over time. Predict the elution order.

The predicted elution order will go from least to most polar compounds.This order corresponds with naphthalene fluorenol o-toluic acid. I came to this conclusion by knowing that acids are generally more polar than alcohols, which are generally more polar than alkenes.

Why do you add a sand layer into the column before adding the alumina?

To ensure that the alumina layer will not interact with the glass filter at the bottom of the column (you probably didn't even notice the glass filter... it was white)

Why do you change solvents in the middle of the separation?

You change to a more polar solvent in order to elute the more polar acetylferrocene

In this experiment

You will be separating a mixture of ferrocene and acetylferrocene using a dry-packed column for chromatography