Chromatography Forum

Hello

I was hoping to get some recommendations on internal standards that can be used in HPLC.

I know the 1st inclination will be to ask details about the method and the analyte. But, really, the excrutiating details are not of any value. Bottom line: it is a relatively typical reversed phase run.

I'm looking for recomendations for internal standards that:

- are fairly stable

- and elute pretty late in reversed phase LC (so they are out of the way of all the analyte peaks)

Thanks very much in advance.

The choice of internal standard depends mainly on the nature of the analyte being analyzed and the detection method.

It is advisable to use chemical analogues of the analyte as internal standard. That assumes equal or near equal response factors from the detector to both analyte and internal standard. For example, analyte and the chosen internal standard have same or similar chromophore (when detection method is spectrophotometric).
In MS it is common practice to use isotope labeled substances analogues as internal standards.

Regards

relatively typical reversed phase run

There ain't no such thing!

If you're running isocratic with 10% organic, then an IS that requires 80% organic will be useless. If the detector response to your IS is wildly different than its response to your analytes, then you will probably see worse results rather than better.

I second zokitano's advice. That said, if you are using UV detection, then alkylphenones have been suggested as "last-resort" internal standards (you have to pick a chain length that is retained in the same range as your analytes).

Most analysts use external standard quantitation for most assays these days as the autosamplers are so reliable. At least in US.

There is no magic internal standard that will work 'typically' on most reverse-phase runs. You have to select one based on what your analyte is as many others have stated.

Most analysts use external standard quantitation for most assays these days as the autosamplers are so reliable. At least in US.

Disagree! Where do you get your facts?

If using MS detection, the labeled internal standards are the only way to go (if one is available). Suppression issues.