Amine purification,

[grace]

Can any body explain the real chemistry behind Chromatography??

 

I have very polar amine, with primary, secondry and tertiary N's. I want to purify it.

I tried using alumina TLC with even 10% MeOH/DCM with trietyhyl ammonia, but my compound is not even moving from base line.

What should I do????

[hypervalent_iodine]

Amines are typically quite sticky and hard to purify via silica chromatography. MeOH/DCM with TEA is what I would have recommended, but you seem to have tried that already! Can I ask what you're using to develop your plates and are you pre-treating your plates with your solvent? Depending on what else is in the crude, you might be able to purify it using a simple acid base work up. Do you have any idea of what else is in there?

Edited August 29, 2011 by hypervalent_iodine

[spin-1/2-nuclei]

Can any body explain the real chemistry behind Chromatography??

 

I have very polar amine, with primary, secondry and tertiary N's. I want to purify it.

I tried using alumina TLC with even 10% MeOH/DCM with trietyhyl ammonia, but my compound is not even moving from base line.

What should I do????

 

Hello,

 

Have you tried reverse phase HPLC?

If not, you can spray your TLCs with ET3N prior to use.. Also, when working with amines you must first wash your silica with ET3N prior to use. I typically dry pack my columns, push them down with air (before adding compound and sand at the top) and then run the ET3N through the column a few times, since I dry pack this has to be done until the column is fully saturated. I then push the excess ET3N out with air - but am careful not to dry out the silica - then I add my solvent system as normal.. This always works great for me whenever I have to deal with alkaloids.

 

Get on Sci-Finder and grab some SIs for papers that have a synthetic route containing an alkaloid or other polar amines. Some of them will have good procedures for purifying all manner of polar amines and alkaloids. There are many procedures - some that involve the use of ET3N washed silica etc as I described before, and some use buffers in mobile phase with pHs typically ranging from 8 to 12. There are too many methods to try to describe them all here (especially from memory) and I'd hate leave something out and mess you over - so, if my above method doesn't seem like it will work for you - or if you have already tried something like it - time to dig in and read a bunch of SIs..

 

Unfortunately, running columns on amines and alkaloids is a lot like black magic. You bubble a little of this, add a little of that, spin around on your head a few times, sacrifice an undergrad or first year [if you are either my apologies (and you will need to find a premed major ;-)]clap twice to the east - and hopefully you get good results..

 

If you really need more info on how chromatography works, I'd be happy to walk through that with you, but I am assuming (please tell me if I'm wrong) that you were just being facetious..

 

Either way,

Best of luck, you have my sincerest sympathies..

Cheers.

Edited August 29, 2011 by spin-1/2-nuclei

[hypervalent_iodine]

Hello,

 

Have you tried reverse phase HPLC?

If not, you can spray your TLCs with ET3N prior to use.. Also, when working with amines you must first wash your silica with ET3N prior to use. I typically dry pack my columns, push them down with air (before adding compound and sand at the top) and then run the ET3N through the column a few times, since I dry pack this has to be done until the column is fully saturated. I then push the excess ET3N out with air - but am careful not to dry out the silica - then I add my solvent system as normal.. This always works great for me whenever I have to deal with alkaloids.

 

 

Alternatively, you can just make up your silica 'slurry' with your solvent + 5% TEA and run the column with 1%, which would seem less wasteful than using straight TEA. It works for the benzyl amines I use, at any rate.

 

As I mentioned before, gka, you will need to pretreat your plates with your solvent + TEA. If you haven't done this, it may explain why you aren't seeing any movement on your plates. Run your plate in the solvent before you spot your stuff onto them, let it dry and then run them with your compound spotted onto it. Additionally, if you are using KMnO₄ or Goofy dip to develop your plates, you might be finding your plate difficult to visualise. Vanillin stain has always worked well for me, so you might want to give that a try as well (if you haven't already).

 

 

 

[spin-1/2-nuclei]

Alternatively, you can just make up your silica 'slurry' with your solvent + 5% TEA and run the column with 1%, which would seem less wasteful than using straight TEA.

 

I think you've misunderstood. It's not wasteful because you don't need more ET3N than you normally would, you just run it through without the rest of the solvent - i.e. - you push it through alone. So the volumes between the two methods are very similar. I typically add 1ml of ET3N for every 5 to 6 grams of silica gel. This is why you must use the air to push this through. I simply keep collecting that ET3N and redistributing it through the column. When I get to the 3rd or 4th run and it seems like the silica has been mostly wetted with it, I add the solution made up for washing/packing the column. Typically in my case petroleum ether. I then run that through the column a few times and clean out the ET3N - since the silica is already deactivated I don't need excess ET3N hanging around and messing up my separation.

 

This procedure always works well for me when I am working with alkaloids and polar amines, but everyone has their own preferences. I prefer to dry pack because I run columns on both micro and large scales. I've found that for me, dry packing works best with both, but is an absolute must for microscale columns. They're easier to set up, there is less mess (no residue hanging out on the side walls of the column), it is easier to control the exact height of the silica gel in the column (which is paramount for me), and if you know what you're doing you can get these columns evenly packed and cracked free easily.

 

but, no matter which method you decide to use, you will find many many experimental procedures for purifying polar amines if you check Sci-Finder and grab some SIs.

 

Cheers

[hypervalent_iodine]

I think you've misunderstood. It's not wasteful because you don't need more ET3N than you normally would, you just run it through without the rest of the solvent - i.e. - you push it through alone. So the volumes between the two methods are very similar. I typically add 1ml of ET3N for every 5 to 6 grams of silica gel. This is why you must use the air to push this through. I simply keep collecting that ET3N and redistributing it through the column. When I get to the 3rd or 4th run and it seems like the silica has been mostly wetted with it, I add the solution made up for washing/packing the column. Typically in my case petroleum ether. I then run that through the column a few times and clean out the ET3N - since the silica is already deactivated I don't need excess ET3N hanging around and messing up my separation.

 

This procedure always works well for me when I am working with alkaloids and polar amines, but everyone has their own preferences. I prefer to dry pack because I run columns on both micro and large scales. I've found that for me, dry packing works best with both, but is an absolute must for microscale columns. They're easier to set up, there is less mess (no residue hanging out on the side walls of the column), it is easier to control the exact height of the silica gel in the column (which is paramount for me), and if you know what you're doing you can get these columns evenly packed and cracked free easily.

 

but, no matter which method you decide to use, you will find many many experimental procedures for purifying polar amines if you check Sci-Finder and grab some SIs.

 

Cheers

 

Ah right, gotcha. I thought you were loading it with neat TEA and then running it with a 1% TEA/solvent, which would be incredibly wasteful. Yes, each to their own. Since I've started in research labs I think I've been taught about a thousand different ways to run columns, but in the end it comes down to what you're most comfortable with in your hands.

 

Another thought I just had is maybe using an alumina column instead of silica. I'm not too familiar with using such columns at present, but I do know you can buy them activated and they can work in purifications where silica does not. I myself am about to embark on series of reactions using basic alumina with some alkaloids that are annoyingly very sensitive to silica; I'm just waiting for CSIRO to send me some goodies . Perhaps spin-1/2-nuclei might know more about it (I would also be keen to hear your opinion on alumina actually as no one in my group who isn't busy writing up their thesis seems to have worked with it before).

Edited August 30, 2011 by hypervalent_iodine

[spin-1/2-nuclei]

Ah right, gotcha. I thought you were loading it with neat TEA and then running it with a 1% TEA/solvent, which would be incredibly wasteful. Yes, each to their own. Since I've started in research labs I think I've been taught about a thousand different ways to run columns, but in the end it comes down to what you're most comfortable with in your hands.

 

Another thought I just had is maybe using an alumina column instead of silica. I'm not too familiar with using such columns at present, but I do know you can buy them activated and they can work in purifications where silica does not. I myself am about to embark on series of reactions using basic alumina with some alkaloids that are annoyingly very sensitive to silica; I'm just waiting for CSIRO to send me some goodies . Perhaps spin-1/2-nuclei might know more about it (I would also be keen to hear your opinion on alumina actually as no one in my group who isn't busy writing up their thesis seems to have worked with it before).

 

Yes you are right, you can use the alumina columns but unfortunately there are some disadvantages to doing so and I personally prefer the silica adsorbent for my columns (but as you said yours are sensitive to silica so that is a bit of a bust)..

 

Still, some people in my lab have had success with these..

 

Some things to keep in mind though..

For starters, you must mind the water content, since the polar sites will go down with increasing water content - so solvents like MeOH start to become problematic with alumina adsorbent. When you purchase your alumina you'll have to chose the activity (from 1 - 3 and acidic, neutral, or basic), which is just another way of describing it's water content. In our lab we buy activity 1 and deactivate it with water as necessary and as with silica you have to also mind your mesh size in comparison to your chosen flow rate.

 

I have a good reference on using alumina columns with alkaloids somewhere, but I cannot find it on my desk at the moment - so I will have to try to search for it later. I will post here with it again if/when I find it..

 

Cheers

 

p.s. if I can't find this reference I will ask my lab mate to come here and post an overview of these columns for you.

Edited August 30, 2011 by spin-1/2-nuclei

[hypervalent_iodine]

Yes you are right, you can use the alumina columns but unfortunately there are some disadvantages to doing so and I personally prefer the silica adsorbent for my columns (but as you said yours are sensitive to silica so that is a bit of a bust)..

 

Still, some people in my lab have had success with these..

 

Some things to keep in mind though..

For starters, you must mind the water content, since the polar sites will go down with increasing water content - so solvents like MeOH start to become problematic with alumina adsorbent. When you purchase your alumina you'll have to chose the activity (from 1 - 3 and acidic, neutral, or basic), which is just another way of describing it's water content. In our lab we buy activity 1 and deactivate it with water as necessary and as with silica you have to also mind your mesh size in comparison to your chosen flow rate.

 

I have a good reference on using alumina columns with alkaloids somewhere, but I cannot find it on my desk at the moment - so I will have to try to search for it later. I will post here with it again if/when I find it..

 

Cheers

 

p.s. if I can't find this reference I will ask my lab mate to come here and post an overview of these columns for you.

 

 

We have a container of each of them in our lab. Unfortunately the basic one I needed looked a little bit dodgy, so I had some sent up from my collaborators with some alumina plates, which should be in tomorrow I honestly wouldn't even be bothering with alumina if it weren't for the fact that my stuff is tricky to purify on the small scales I do it on and that it decomposes if I so much as put a silica container within 2 meters of it. Once I build up to larger scales, I'm just going to do a Kugelrohr distillation, which is much easier and a personal favourite of mine (if only because I get to use the word Kugelrohr). That being said, I'll still need to use alumina for the next bit so I figure I may as well become an expert now. The reference you mention would be very much appreciated, thank you

 

 

[spin-1/2-nuclei]

We have a container of each of them in our lab. Unfortunately the basic one I needed looked a little bit dodgy, so I had some sent up from my collaborators with some alumina plates, which should be in tomorrow I honestly wouldn't even be bothering with alumina if it weren't for the fact that my stuff is tricky to purify on the small scales I do it on and that it decomposes if I so much as put a silica container within 2 meters of it. Once I build up to larger scales, I'm just going to do a Kugelrohr distillation, which is much easier and a personal favourite of mine (if only because I get to use the word Kugelrohr). That being said, I'll still need to use alumina for the next bit so I figure I may as well become an expert now. The reference you mention would be very much appreciated, thank you

 

Hi all

 

What spin said already covers the basics. The rest depends on the compound details. Since you're familiar with silica columns you will catch on quick. Look at this book: Preparative Chromatography Techniques: Applications in Natural Product Isolation by K. Hostettmann, Andrew Marston and Maryse Hostettmann (Dec 7, 2010) & you will be an expert in no time.