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Author Topic: How do I remove the excess diethylamine from an Fmoc deprotection?  (Read 464 times)

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carbonylcookie

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I've used diethylamine to deprotect an Fmoc from a solution phase dipeptide that also carries several boc protecting groups. I tried to purify my compound by recrystalization, but the solubilities didn't work out. On the advice of another chemist in the lab I used a silica column (which was not a good idea). From my NMR, it looks like a large amount of diethylamine and at least some of my product came off of the column. I've tried coevaporating the DEA with both chloroform and toluene, but it looks like it's only reduced the amount of DEA a small amount. Is there anything else I can try to remove the DEA and recover my product?
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wildfyr

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Re: How do I remove the excess diethylamine from an Fmoc deprotection?
« Reply #1 on: November 14, 2017, 09:29:09 AM »

Distill it off under high vacuum. It boils at 55C at 1 atm. Maybe wash it with dilute weak base before hand to make sure none of it is in the nonvolatile protonated form.

You could also try to separate it out by vitrifying the product with cold ether or a linear alkane.
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carbonylcookie

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Re: How do I remove the excess diethylamine from an Fmoc deprotection?
« Reply #2 on: November 14, 2017, 10:28:20 AM »

Distill it off under high vacuum. It boils at 55C at 1 atm. Maybe wash it with dilute weak base before hand to make sure none of it is in the nonvolatile protonated form.

You could also try to separate it out by vitrifying the product with cold ether or a linear alkane.

wildfyr,
I have tried applying the high vacuum several times. I think your suggestion that it is protonated is likely. I tested a small amount of my solid and it appears that both my product and the diethylamine are water soluble, so I don't think I could wash with an aqueous base. Is there a way to deprotonate it without an aqueous base? Are there basic resins I could run it over?
I'm not familiar with vitrification, can you explain what you're suggesting a bit more? I'm a biochemist by training so I'm already batting out of my league with this synthesis.
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Babcock_Hall

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Re: How do I remove the excess diethylamine from an Fmoc deprotection?
« Reply #3 on: November 14, 2017, 11:39:40 AM »

I don't understand how protonated diethyl amine would elute from a silica column.  Can you provide a structure of your presumed product at this stage?
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wildfyr

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Re: How do I remove the excess diethylamine from an Fmoc deprotection?
« Reply #4 on: November 14, 2017, 02:07:24 PM »

Vitrification is basically when you reduce your compound down to an oil then crash it out using a poor solvent. However if its soluble in water my advice is unlikely to work. Do you feel that you can share your molecule? For one thing, it's synthesis could be available on scifinder, which would cut us out as middlemen in helping you with this.

And yes, there are basic resins.

Something I learned very recently is that you should be able to see if you have the protonated version of an amine by IR. A bunch of odd peaks will show up from 2400-2800 cm-1 that are highly characteristic. The SDBS database has the spectrum of diethylamine hydrochloride.
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carbonylcookie

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Re: How do I remove the excess diethylamine from an Fmoc deprotection?
« Reply #5 on: November 15, 2017, 07:16:56 AM »

Vitrification is basically when you reduce your compound down to an oil then crash it out using a poor solvent. However if its soluble in water my advice is unlikely to work. Do you feel that you can share your molecule? For one thing, it's synthesis could be available on scifinder, which would cut us out as middlemen in helping you with this.

And yes, there are basic resins.

Something I learned very recently is that you should be able to see if you have the protonated version of an amine by IR. A bunch of odd peaks will show up from 2400-2800 cm-1 that are highly characteristic. The SDBS database has the spectrum of diethylamine hydrochloride.

Unfortunately my molecule falls on the proprietary side, it's enroute to a potential drug candidate. I can try to verbally describe it. It has a tri-boc protected arginine on one end connected via a short diamine linker to a propargyl glycine on the other. The Fmoc was removed from the amine of the propargyl glycine. I was not expecting it to be water soluble at all, so my best guess is either that the small sample of powder I dissolved didn't have enough for me to visually confirm insolubility or it's forming micelles. Thank you for explaining vitrification. The molecule isn't really something that already exists, so I'm only guessing at solubilities, and with the large amount of diethylamine I'm not sure when visible insolubility is product or diethylamine or both.

Because my lab sits in an odd limbo between departments it's always a toss up on whether I can get access to equipment, but I will look into trying to run a small sample on IR. It's fairly obvious from my NMR that I have diethylamine. I'll take a closer look to see if there's a hint about its protonation state.
Thanks for your advice so far.
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carbonylcookie

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Re: How do I remove the excess diethylamine from an Fmoc deprotection?
« Reply #6 on: November 15, 2017, 07:39:25 AM »

I don't understand how protonated diethyl amine would elute from a silica column.  Can you provide a structure of your presumed product at this stage?

Well, in order to get anything after the dibenzofulvene off the column I ended up running 60% MeOH in chloroform through it. I'm not sure why it's protonated, I included 0.05% TEA in the solvents run through the column. It seems that if it were unprotonated it would have evaporated after this many rounds of vacuum and different solvents, but maybe something else is keeping it from coming off. I do know I have diethylamine, it's very clear from the NMR.
I described the compound in my reply to wildfyr above.
The solid I have right now seems to be white with yellow streaks through it, I'm not sure what it implies but it's... something.
Right now I've exhausted the advice of the chemists in the lab and my ability to find anything via searching, so here I am.
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wildfyr

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Re: How do I remove the excess diethylamine from an Fmoc deprotection?
« Reply #7 on: November 15, 2017, 07:56:39 AM »

Your molecule sounds pretty greasy... its odd that it is water soluble.

Do you think you could take a little and try to partition between chloroform and water (or even brine?) and see which layer it prefers? This would give some really critical data about what kind of workups are available. If it prefers chloroform You could then do an aqueous base wash followed by distillation.

Alternatively, you could try a dilute acetic acid wash, which should protonate the DEA, pull it in to the water layer, but leave your protecting groups intact.
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carbonylcookie

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Re: How do I remove the excess diethylamine from an Fmoc deprotection?
« Reply #8 on: November 16, 2017, 07:04:37 AM »

Your molecule sounds pretty greasy... its odd that it is water soluble.

Do you think you could take a little and try to partition between chloroform and water (or even brine?) and see which layer it prefers? This would give some really critical data about what kind of workups are available. If it prefers chloroform You could then do an aqueous base wash followed by distillation.

Alternatively, you could try a dilute acetic acid wash, which should protonate the DEA, pull it in to the water layer, but leave your protecting groups intact.

wildfyr,
Thank you for all your suggestions. I eventually bit the bullet and put a hefty scoop into water to make sure it wasn't soluble. It looks like it probably isn't soluble, it just makes such a fine dust I had to use a lot to see it. I tried simply filtering it out of the water after that and evaporated off as much solvent and water as I could. The NMR shows a lot less DEA, but it's still showing up and even if what I got back from the water was completely pure it's unfortunately not enough to move on to the next step of synthesis. It looks like most of what I got off that column was diethylamine salt nonsense. I'm going to need to start my synthesis again, and figure out a better purification ahead of time.
Thanks again for your responses.
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wildfyr

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Re: How do I remove the excess diethylamine from an Fmoc deprotection?
« Reply #9 on: November 16, 2017, 09:28:39 AM »

Ok, sorry you have to do it again. If its not soluble in water, then acid/base chemistry liquid/liquid extraction is going to be your best friend!
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