Hello All,

I am trying to develop a CEC-HPLC method with column heating at 45 celsius. My analytes are 80 and 84 aa peptides. My question is, is there a possiblity that peptides (or even any small molecules) can degrade/decompose at high temperatures when we try to use temperture as a variable in method development?.
I think it is possible but it depends on the stability of the anlyte at the temperture you are using??Or since there is only a few seconds exposure of the anlyte inside the column to the high temperature it is not possiible for any degrdation?
I appreciate your comments and sharing your experience on this.

Thanks in advance!

Ananda S.

It's unlikely that you will experience degradation problems unless you're working at extreme pH values while working at this temperature. The peptide bond is somewhat unstable but the major issue with temperature is the problem of denaturing native proteins, not degradation, in general. Since you are working with peptides, this isn't likely to be an issue. In fact, it's not always true that elevated temperature is bad even for native proteins. We've recently developed separations with monoclonal antibodies which perform well even at 60 Celsius.

Chris,

Thank you so much for your comments.

Best Regards!

Ananda S.

Ananda,

Why have you decided to use 45 degrees for your study in the first place? is this because you are not confident of efficiently thermostatting your column at a temperature closer to room temperature or for some other reason?