Sciencemadness Discussion Board

HBTU and Other Peptide Reagents - Preparation

Sauron - 6-7-2007 at 22:48

Someone was asking about where to buy HBTU, PyBOP etc that would be amateur friendly.

As there does not seem to be any simple cost effective way of doing that (importing from China being rather costly due to shipping and formalities) I hereby suggest an end run: make your own.

HBTU is dead easy. Attached is the one step preparation of 1-hydroxybenzotriazole from 2-nitrophenylhydrazine. The reagents are conc ammonium hydroxide and conc HCl. The reaction is rapid and followed easily by color change (red to dark brown).

The starting material is itself easily made from o-nitroaniline in the usual manner of preparing phenylhydrazines from anilines, q.v. This is diazotization followed by reduction of the diazonium salt, preferably with sodium or ammonium sulfite. Alternatively, SNCl2 or Sn/HCl conc, or Zn dust/HCl conc can be used but only the calculated amount to reduce the diazonium salt, as excess would reduce the nitro group as well. Org.Syn. recommends preparing the sodium sulfite fresh from NaOH and SO2 from a cylinder. Where I am this is impossible as SO2 is restricted.

ortho-Nitroaniline is cheap and readily available. I have a Kg of it sitting around here from Merck. You can of course make your own by nitrating sulfanilic acid. See Org.Syn. or Vogel for these facile and classical transformations, which start from acetanilide.

I have the Berichte reference if anyone wants to mud-wrestle with the German.
Have fun.

All in all even if you go all the way back to acetanilide it isn't too much work, and probably will cost less than buying the HBTU from abroad, or domestically.


[Edited on 7-7-2007 by Sauron]

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Sandmeyer - 7-7-2007 at 10:18

Sometimes I wonder if not certain brands pay you to promote them ;) For such synthesis you need chemicals that aren't OTC. Hence, it is much smarter alternative to simply buy 1 unregulated chemical of good purity than a couple of them needed for the time-consuming synthesis of that same chemical. For example, in kitchen situation, it is easier to obtain the coupling reagent directly than to obtain sodium nitrite.

[Edited on 7-7-2007 by Sandmeyer]

Sauron - 7-7-2007 at 16:08

Sorry, but your premises are erroneous.

No one is paying me to "promote" anything on this forum. I am getting tired of this petty accusation, why don't you come up with something new and original?

And it is absurd to suggest that buying a coupling reagent that is classified as EXPLOSIVE, for an amateur, is easier than obtaining sodium nitrite.

You have such a narrow eurocentric viewpoint that you apparently think everyone everywhere has trouble buying analytical grade sodium nitrite. That is not true. I can and do buy sodium nitrite of any desired grade ex stock locally with a delivery time of a few days and low prices.

And frankly, I regard your assumption of a "kitchen situation" for peptide synthesis, or for advanced organic chemistry employing peptide reagents, as an absurdity.

HBTU is horribly expensive (about $1.50 a gram in 250 g packing, and $10 a gram in 5 g packing) so as far as I am concerned it is a perfect candidate for making your own, particularly because of all the hassles associated with its having been STUPIDLY reclassified by the EU as explosive.

This is a simple unlaborious prep that one can enter at the o-nitroaniline stage and be three simple steps from HBTU. I have elaborated how to make the o-nitroaniline from acetanilide (or somewhat more conveniently from sulanilic acid) for the sake of those who have a hard time buying the nitroaniline - but I don't.

Can't do a diazotization for lack of NaNO2? Try moving to a free country. After all you DO call yourself Sandmeyer so I assume you don't dislike diazotizations.

[Edited on 8-7-2007 by Sauron]

Sandmeyer - 7-7-2007 at 16:49

I was unable to get sodium nitrite at one point, this even if it is used to preserve consumer food -- it is classified as poison where I am.

HBTU is certainly not the only coupling agent, have you considered cheaper alternatives.
Quote:
And frankly, I regard your assumption of a "kitchen situation" for peptide synthesis, or for advanced organic chemistry employing peptide reagents, as an absurdity.


I'm not sure I understand what you meen, but there is at least one published example of kitchen chemistry involving the use of a coupling reagent to make a lysergic acid amide in very high yield. Can't get operationally more simple

[Edited on 8-7-2007 by Sandmeyer]

Sauron - 7-7-2007 at 17:25

The choice of coupling reagent was not mine, it was redbaron's.

He mentioned only HBTU and PyBOP and PyBOP is simply a derivative of HBTU.

I certainly don't regard lysergic acid chemistry as kitchen chemistry, and if I thought that's what redbaron was up to I wouldn't be trying to help him. Silly me, I thought that peptide reagents might be being used to make peptides. How droll.

For the sake of those who are NaNO2 challenged, or simply lazy, here is a very likely route to o-nitrophenylhydrazine that does NOT require any diazotization step:

o-nitrochlorobenzene + hydrazine sulfate (or 60-64% soln of hydrazine in water) -> o-nitrophenylhydrazine

I don't have any direct references for this but see Vogel, who uses this reaction to prepare 2,4-dinitrophenylhydrazine from 2,4-dinitrochlorobenzene. The corresponding dinitroaniline cannot be diazotized.

A similar reaction of o-nitrochlorobenzene is the reaction illustrated in Org.Syn. with 2 mols of the nitrochlorobenzene and one of sodium sulfide; this is used to prepare orthanilic acid. This shows that the Cl in o-nitrochlorobenzene is sufficiently activated by the adjacent nitro group to be readily substituted.

Assuming this works, then one is only two steps from o-nitrochlorobenzene to HBTU. o-nitrochlorobenzene is rather nasty stuff so use a hood and gloves etc.

Now I await the Greek chorus of wails and moans about hydrazine not being OTC, but I do seem to recall several popular threads about making hydrazine, prominently featuring Roscoe Bodine, so maybe chemistry isn't quite dead on this forum yet, just wheezing a little.

Meanwhile here is the Org.Syn. prep of 2,4-dinitrophenylhydrazine. Tweaking the procedure for o-nitrochlorobenzene should be easy.



[Edited on 8-7-2007 by Sauron]

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Sauron - 9-7-2007 at 01:05

According to the literature, 2,4-dinitrochlorobenzene is about 2000 times more reactive at the halogen toward SnAr than o-nitrochlorobenzene, and p-nitrochlorobenzene is less reactive in many instances than the ortho isomer. However there is much literature on nucleophilic substitution reactions of o-nitrochlorobenzene and 0-nitrobromobenzene, including many instances of reactions with primary and secondary amines, so I think there is every reason to assume that in a suitable solvent, hydrazine hydrate or hydrazine sulfate will attack the halogen and result in the desired o-nitrophenylhydrazine. DMF or DMSO suggest themselves. MeCN also.

I found the Beilstein references for o-nitrophenylhydrazine in Federoff Vol. 7 H-207.

Beil. 15 454 (127) & [177]

Anyone have access to the books or the online service?



[Edited on 10-7-2007 by Sauron]

Sauron - 10-7-2007 at 04:01

Further literature searching has not been particularly encouraging about the SnAr approach in the case of the mononitrochlorobenzenes..

Ullmann's in their Hydrazine Derivatives section says that all phenylhydrazines are made by diazotization and reduction (with sodium sulphite) of the phenyldiazonium chloride. The exceptions are the two dinitrophenylhydrazines and picry hydrazine (2,3,6-trinitrophenylhydrazine) which are made as described upthread.

I did find some specific descriptions of the preparation of o-nitrophenylhydrazine - interestingly enough they were lurking all along in the forum library.

Shirley's "Preparation of Organic Intermediates" p 232 and referencing Muller and coworkers, Helv.Chim.Acta, 20, 1432 (1937). This procedure employs freshly prepared potassium sulfite in the reduction.

Fierz-David, "Fundamental Processes of Sye Chemistry" pp 95-97. This one is particularly interesting because the author employs sodium bisulfite and NaOH rather than sodium sulfite in the reduction. (The preparation is of the p-isomer but states that same instructions are used for o-sisomer except for a deviation in workup which is detailed.) The prep references Bamberger and Kraus, Ber., 29, 1829-1835 (1896). This at least is a reference I can easily retrieve.

The Org.Syn. prep (of phenylhydrazine itself) insisted that the sodium sulfite needed to be made fresh from SO2 and NaOH because commercial sodium sulfite was unsatisfactory.

Also, the o-nitroaniline required is made directly from o-nitrochlorobenzene affording a somewhat shorter route than the acetanilide or sulfanilic acid ones. This is however at the expense of having to sort out the two isomers from the nitration of chlorobenzene.

Also, in Saul Patai's "The Chermistry of the Hydrazo, Azo, and Azoxy Groups, Vol. 1" I found a section (Chapter 16, p.677) on the intramolecular reaction between the terminal nitrogen and the nitrogroup of o-nitrophenylhydrazine to give HBTU, which details the mechanism, and specifies the reaction conditions required to give this product rather than the N-oxide. This volume is not in the forum library, I got it a while back from the (MadHatter) ftp site.


[Edited on 10-7-2007 by Sauron]

Sauron - 11-7-2007 at 01:54

o-Nitrochlorobenzene is converted in 95% yield to o-nitroaniline by autoclaving for 10 hours at 170 C with a tenfold excess of conc NH4OH.

Therefore, I believe that hydrazine hydrate (60-65% aq soln) will work similarly to give the o-nitrophenylhydrazine.