Page 1 of 1
Guanidine HCl compatability of C18
Posted: Sat Jul 08, 2006 6:06 pm
by Amar
Helo every body,
Is C18 RP-HPLC column compatible with Gunidine chloride? My peptide is insoluble without Gunidine. If not, which column can be used with denaturant?
Thanks
Amar
Posted: Mon Jul 10, 2006 2:51 am
by MartinG
I have been able to inject a protein in 6M GnHCl on a R1 POROS reversed phase column, which has a selectivity similar to a C4 column. The retention time was the same than when the same protein was injected without GnHCl. So it worth trying.
Good luck
Martin
Posted: Mon Jul 10, 2006 8:08 am
by HW Mueller
If one does a lit search on protein RPHPLC one sees a lot of applications with all kinds of columns using guanidine HCl. Having used only urea I would still venture a guess that the only cause for worry is an acidic pH, with all that Cl- around. On the latter: Check into all those chains about Cl-, HCl, and stainless steel.
is urea ok?
Posted: Mon Jul 10, 2006 4:49 pm
by Amar
Helo HW,
do you mean you have used urea on C18 RPHPLC columns? is it okay to use? coz I use either GuCl or urea.
Thanks for previous reply.
Amar
Posted: Mon Jul 10, 2006 9:39 pm
by Mark Tracy
Urea won't hurt your column at all. Just be sure that the sample does not precipitate when it mixes with the mobile phase.
Posted: Mon Jul 10, 2006 11:16 pm
by Amar
Hi Mark,
Thanks. That information was of great help. How about DTT? is it going to damage C18 column?
Amar
Posted: Mon Jul 10, 2006 11:29 pm
by Mark Tracy
DTT is also harmless, so is iodoacetamide (at least harmless to your C18 column).
Posted: Mon Jul 10, 2006 11:46 pm
by Amar
Hi mark, I really appreciate your answers. I will post if I have any question in future.
Amar
Posted: Tue Jul 11, 2006 6:49 am
by HW Mueller
Yes I did use C-18 columns with Urea, I didn´t mention it specifically as there are also sundry examples in the literature, and also because I can´t see any possibility for reaction. Again, the Cl- of guanidine should be watched if you use low pH, because of the stainless steel.