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Topic: Phenylfluorenyl PG removal + isolation of small aminoacid  (Read 948 times)

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Offline kriggy

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Phenylfluorenyl PG removal + isolation of small aminoacid
« on: October 27, 2021, 08:51:09 AM »
Hey guys,
its somehow related to my previous question but its probably better to start a new topic.
Anyway, Im strugling to remove my protecting group: phenylfluorenyl. I can get the protected intermediate without much trouble but the removal is pain. Using TFA with various scavengers (EtSiH, TIPS, MeOH...) I clearly see the starting material gone (by TLC) but afterwards, I dont see the mass of the product in my LCMS analyssis or only traces to be more specific and the spectrum further shows various unknown impurities. Furthermore, only very weak oftentimes none ninhydrin reaction is observed with TLC analysis.
I tried to deprotect the PhSe intermediate hoping the double bond is the cause of the problems with similar results observing some kind of PhSe transfer to get double selenylated product. Hydrogenation of the PhSe at 2.5 atm did not show any traces of reaction.

Furthermore, I recently started the same sequence with trityl protected aminoacid (trt is more labile to acid) but during first step (KHMDS enolate then PhSe quench) im observing poor reactivity and cleavage of the trityl group. Anyone observed this happening? I did quench the reaction with acetic acid but I dont think trt is that labile.

Im glad for any tips you can give me. Esp. some tricks for LCMS detection of aminoacids are wellcome (i detect by SIR but from that its hard to extimate the amounf of the aminoacid in my sample

thank you

Offline rolnor

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #1 on: October 27, 2021, 11:53:56 AM »
Is it necessary to use TFA? The trityl goes with 80%HOAc, maybe you can try that, maybe some heat? Maybe scavenger is bad in this case, I would try without.

Offline kriggy

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #2 on: November 04, 2021, 03:22:13 AM »
Pf is about 6000x times more stable so I think TFA is needed. I might try HCl in ether and maybe the hydrochloride crashes out. I think I did try without the scavenger and the results are fairly the same. I get rid of the Pf group (TLC) but then im unable to isolate the product.

I think I might be actually getting the pyroglutamic derivative or some kind of michael addition.
« Last Edit: November 04, 2021, 05:23:17 AM by kriggy »

Offline Babcock_Hall

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #3 on: November 04, 2021, 09:36:08 AM »
I work a good deal with unsaturated amino acids, but I doubt that I can be of much help in this instance.  KMnO4 dipping has worked well for visualizing our TLC plates.  The amino group should not be performing a Michael addition, unless one is under basic conditions; therefore, I am puzzled.  Regarding MS, if I could  spare a small portion of a product with a free amino group, I would at least consider using heptafluorobutyric anhydride to make a derivative.

Offline rolnor

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #4 on: November 04, 2021, 10:37:25 AM »
OK, maybe you can run a deprotection in a NMR-tube to see what goes wrong, if the double-bond is affected this should be easy to spot.

Offline phth

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #5 on: November 04, 2021, 01:43:41 PM »
Protect the seleno compound from oxygen. It is stable to acid. Seleno compounds can oxidize during column chromatography... Why not just deprotect then do the elimination step???

Offline rolnor

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #6 on: November 04, 2021, 04:36:09 PM »
The amino group can be oxidized if its unprotected I think, I guess you use hydrogen peroxide to get the selen oxide before elimination? Also the unsaturated product is a Michael addition substrate so a free amino group can add to the double bond. Its also a di ester so it can ringclose or polymerize via aminolysis.

Offline phth

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #7 on: November 04, 2021, 07:59:43 PM »
The amino group can be oxidized if its unprotected I think, I guess you use hydrogen peroxide to get the selen oxide before elimination? Also the unsaturated product is a Michael addition substrate so a free amino group can add to the double bond. Its also a di ester so it can ringclose or polymerize via aminolysis.

Yes MCPBA may be the wrong reagent. I would try a chlorine source under aqueous and buffered conditions.

Offline kriggy

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #8 on: November 05, 2021, 02:33:39 AM »
Protect the seleno compound from oxygen. It is stable to acid. Seleno compounds can oxidize during column chromatography... Why not just deprotect then do the elimination step???

Im able to purify on column without any issues - except the starting material spot is very cloes to the product as much as they are almost coeluting but its doable.

The amino group can be oxidized if its unprotected I think, I guess you use hydrogen peroxide to get the selen oxide before elimination? Also the unsaturated product is a Michael addition substrate so a free amino group can add to the double bond. Its also a di ester so it can ringclose or polymerize via aminolysis.

Yes MCPBA may be the wrong reagent. I would try a chlorine source under aqueous and buffered conditions.

The original procedure for the elimination on this cpd used mcpba, I find NaIO4 bit better and easier to do (just mix and stir) but the deprotection is then not described because they do ester reduction, dihydroxylation and deprotection later.

Protect the seleno compound from oxygen. It is stable to acid. Seleno compounds can oxidize during column chromatography... Why not just deprotect then do the elimination step???

I did attempt that as well (one experiment) and while I observed the product in LCMS only traces were there and mostly it was some uknown which was assigned from m/z to compound having 2 SePh groups so some kind of rearrangement or transfer probably happened.

I work a good deal with unsaturated amino acids, but I doubt that I can be of much help in this instance.  KMnO4 dipping has worked well for visualizing our TLC plates.  The amino group should not be performing a Michael addition, unless one is under basic conditions; therefore, I am puzzled.  Regarding MS, if I could  spare a small portion of a product with a free amino group, I would at least consider using heptafluorobutyric anhydride to make a derivative.

Thank you. Im used to using CAM which I thought it should work but maybe KMnO4 is better in this case.


OK, maybe you can run a deprotection in a NMR-tube to see what goes wrong, if the double-bond is affected this should be easy to spot.

That is actually great idea. Dont know why I did not think of that already.

Offline rolnor

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #9 on: November 05, 2021, 03:22:12 AM »
The seleno compound will also stain with KMnO4, it will oxidize.

Offline rolnor

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #10 on: November 05, 2021, 05:44:12 AM »
I repeat also, this compound with the amino group as free base is not stable.

Offline kriggy

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #11 on: November 05, 2021, 08:02:29 AM »
I repeat also, this compound with the amino group as free base is not stable.

Why do you think so? Im fine with it being whatever kind of salt that allows subsequent amide bond formation or even deprotecting it in-situ before the coupling

Offline Babcock_Hall

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #12 on: November 05, 2021, 10:00:58 AM »
I like to think of the proton as a poor man's protecting group.

Offline rolnor

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #13 on: November 05, 2021, 10:43:36 AM »
The amino group can be oxidized if its unprotected I think, I guess you use hydrogen peroxide to get the selen oxide before elimination? Also the unsaturated product is a Michael addition substrate so a free amino group can add to the double bond. Its also a di ester so it can ringclose or polymerize via aminolysis.

As I wrote here, the free base is problematic but it depends on the timescale, it could be stable for short time.

Offline kriggy

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Re: Phenylfluorenyl PG removal + isolation of small aminoacid
« Reply #14 on: November 10, 2021, 09:08:45 AM »
So I did run the NMR experiment and after addition of TFA the double bond signal shifted towards lower ppm. Im not sure if its because of the N protonation or because of the deprotection is happening but after about hour or so, the double bond signals dissapeared completely. I dont have much idea whats going on but seems I need to do the deprotection quick.

Im running the same sequence with trityl and for some reason, the Phenylselenyl derivative is decomposing to something which is neither alkene nor selenoxide and for some reason, it does not get oxidized using same condition as the Pf derivative..

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