Inhabitants of my column

[sscala]

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!

[skunked_once]

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!

[tom jupille]

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.

[sscala]

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