[Kate]

Hello.

So in my biology class, the Prof put cellulose TLC plates overnight with a mixture of olive oil and benzine. I thought she did this only because it creates a separation line (the oil won't go up after a while and the benzine will) and then you don't have to mark the application zone with a pencil. But is the stationary phase non-polar now?

[MOTOBALL]

1. My understanding is that cellulose chromatography is partition chromatography, in which the analyte is partitioned between the developing, mobile phase and the water (naturally present from the atmosphere) in the cellulose matrix.  The cellulose (paper sheet or TLC plate) is there only as a physical support for the water.  Therefore, the polarity of the cellulose plate (Water) has not changed; I am unclear as to what "the oil won't go up after a while" means.

2. The benzene will evaporate very readily from the cellulose.

3. I'm VERY surprised that your Prof. is using benzene for this; suspected carcinogen; should only be used in a fume hood, if at all.  Should be replaced by toluene !!

[Kate]

1. My understanding is that cellulose chromatography is partition chromatography, in which the analyte is partitioned between the developing, mobile phase and the water (naturally present from the atmosphere) in the cellulose matrix.  The cellulose (paper sheet or TLC plate) is there only as a physical support for the water.  Therefore, the polarity of the cellulose plate (Water) has not changed; I am unclear as to what "the oil won't go up after a while" means.

Well, I thought because the plate is polar (OH groups from the cellulose), the compounds that will move more rapidly will be apolar. But she used petroleum ether and olive oil overnight and then eluted a mixture of compounds with a polar mobile phase. And the less non-polar compounds moved more rapidly in the plate. So, my question is, did she reverse the polarity of the plate? You said it didn't but then the less polar compound should have moved more rapidly and it didn't.

I'm so confused by this.

I am unclear as to what "the oil won't go up after a while" means.

Well, if you saw the TLC plate before application of the sample you'd see a separation line in the plate due only to the olive oil/petroleum ether mixture in which it stayed overnight. So I think it's because the olive oil just went up a little bit along the plate while the petroleum ether went all the way. Does that make sense?

3. I'm VERY surprised that your Prof. is using benzene for this; suspected carcinogen; should only be used in a fume hood, if at all.  Should be replaced by toluene !!

Benzine/petroleum ether, not benzene. I don't think there's any benzene in petroleum ether.

[MOTOBALL]

 But she used petroleum ether and olive oil overnight and then eluted a mixture of compounds with a polar mobile phase. And the less non-polar compounds moved more rapidly in the plate.

I think that I understand now. Your confusion is due to my initial poor understanding of the situation.

The Prof. used pet. ether/olive oil overnight to impregnate the cellulose with olive oil; analytes are then separated by partition between the (non-polar) olive oil phase and the (polar) developing phase. "the less non-polar compounds" moved more rapidly in the plate, i.e. the less-non-hydrophilic, i.e. the less-hydrophobic compounds moved more rapidly.

Yes, the less-hydrophobic (i.e. more hydrophilic) spend more time in the polar developing phase and so run faster.  The more-hydrophobic compounds will spend more time in the olive oil phase and will be retarded (will run more slowly).

You are correct, I was wrong---the "polarity" of the plate has been reversed, from polar to non-polar.

As a matter of interest, did the Cellulose plate become semi-transparent after the petroleum ether/olive oil treatment ?

How did you set up to put my earlier response in a blue box ?

[Kate]

You are correct, I was wrong---the "polarity" of the plate has been reversed, from polar to non-polar.

But the petroleum ether isn't adsorbed to the cellulose. So when we impregnate the plate in the polar mobile phase, can't the ether be displaced as the mobile phase moves up?

As a matter of interest, did the Cellulose plate become semi-transparent after the petroleum ether/olive oil treatment ?

Sorry, didn't notice it.

How did you set up to put my earlier response in a blue box ?

I used the quote function, at the top right corner. Then selected which text I want to quote.

[MOTOBALL]

The Prof. used pet. ether/olive oil overnight to impregnate the cellulose with olive oil;

The pet. ether here is used to carry the olive oil up the cellulose plate overnight; it then evaporates very quickly once the plate is removed from the developing chamber.  The cellulose is now the support matrix for olive oil, instead of the water that it was the support for initially. The pet. ether plays no subsequent role in the chromatography, because it is not present.

An atomizer could theoretically have been used to spray a solution of olive oil in pet. ether onto the plate.  This would have been much quicker than the overnight procedure.  However, it is difficult to obtain even coverage using a spray and would not have given the uniform coating of oil on the plate that is required for good, reproducible chromatography.

[Kate]

Got it now. Many thanks.