Chromatography Forum

Newbie question:

I experienced the following: running a 10%-90% Water/MeOH gradient (10min) there is a constant peak at ~60% MeOH - even without injection of a sample/blank.

If this peak is some residual compund dwelling inside my column: why does it appear in peak-form instead of washing out continuously?

If it does not live in my column: where else could it be located?

Thanks!

This is a common occurance. The peak is most likely from an impurity in your water which binds to the surface of your reverse-phase column and is then released when the methanol reaches a certain percent. You can verify this by running at 10% water for 10 minutes before running your gradient and then doing another run where you run at 10% water for 30 minutes before running the gradient. If the peak is from the water, you will see a much larger peak at 60% methanol on the second run. There are several posts about water impurities in the HPLC section so you will have to search them out for more information. Good luck!

That question comes up frequently enough that we've actually put together a "mini-seminar":

http://www.lcresources.com/more_resourc ... hp?f=3&t=5

You will have to register (free) on the LC Resources site to view it.

Thanks, problem solved - and: great collection of mini-seminars!