LCMS of primary amides

[pigfarmer]

Hello all, my boss wants me to look into a method to increase detection sensitivity by LCMS (or possibly HPLC) of amide containing compounds. Any ideas? It would be best if the reaction would be specific to the amide and not label other groups such as carboxylic acids. Thanks!

[Don_Hilton]

The reactions that I can come up with for amides either atack the carbonyl - and would result in loss of the nitrogen - making it tough to differentiate between acids and amides - or would abstract a proton from the amide nitrogen - which would react with many other active hydrogen compunds long before the reaction would touch the amide... Perhaps react the amide with an acid halide - off the top of my head the acid anhydride that would form from the organic acids would be more easily decomposed than the amide. The trick is to do this in a way that you add some useful functionality to the amide so you can light it up in the LC/MS or drag in a chromaphore for the UV.

Any other functionality on these molecules that might make a better handle to grab onto?

[pigfarmer]

Sadly, no there is no other functional groups present. What we are looking at are fatty acid amides, so its basically a fatty acid which is an amide instead of the acid.

I pretty much feel I am at the same place as far as reacting with the carbonyl, and am thinking I can use a protecting group to block the carboxylic acid and prevent it from having the carbonyl, then hit the carbonyl. One glaring problem is that I am not an organic chemist by any means, and I don't have access to much in the way of equipment here. I am hoping to be able to find some fairly easy chemistry and then throw some Girard's on it!

Thank you for the response too! Almost forgot to say that!

[Don_Hilton]

Transforming an analyte to somethign else becomes tricky because you need to be able to demonstrate that you did not already have the reaction product present and that you did not transform anything else to the product. The Grignard reaction (I assume this is what you were talking about throwing in the mix?) is a bit drastic for derivitization - and you have to work up the resulting product. Many steps for an analytical derivitization. You can add aromatic character (a chomophore for UV) by making the Grignard reagent from something like benzyl bromide, but you are going to have a less polar compound in the end and not adding anything that will ionize easily for LCMS.

Fatty acid amides do nicely with GC...

[pigfarmer]

I wasn't talking about a Grignard reaction, did one way back in undergrad, definitely not something we are set up to do!

Have you ever seen the Girard's reagents? I know we have Girard's P in our lab, and Girard's T is the other main one used (that I have seen anyways). I have actually used the Girard's P for looking at steroids by MALDI (long time ago) and it worked pretty good.

I don't know how to put a picture in here...but I can give you a link:

http://www.sigmaaldrich.com/catalog/pro ... &region=US
and P
http://www.sigmaaldrich.com/catalog/pro ... &region=US

They contain a quaternary amine and also have a hydrazine group which reacts with carbonyls.

Right now the LC/MS our lab owns is a bruker FTICR, but I should be able to get access to an absciex 4000, and doing SRM may eliminate the need for all this rigamarole!

[Don_Hilton]

I have my dobts about that one. You can form a hydrazone easily with a ketone or aldehyde, but an amide -- from my looking around, amides do not form hydrazones as easily. But, if you can push that one to work, it would be cool.

[pigfarmer]

I have my dobts about that one. You can form a hydrazone easily with a ketone or aldehyde, but an amide -- from my looking around, amides do not form hydrazones as easily. But, if you can push that one to work, it would be cool.

I know, its not ideal. If I did it this way I would have to probably do some chemistry on the amide first...again, getting a little complex! I'll let you know if I ever figure something out!