Chromatography Forum

You said:
"Gabapentin is not totally unbound from protein but a lot of it is unbound."

and:
"since we are having problems with the recovery of the internal standard glibenclamide we decided to use it as external standard. i'm asking if this would be appropriate for bioanalytical methods."

Considering these two conditions, the answer is: No

ghie malig,
on the convention for IS and ES see any good books on HPLC, for instance Dolan and Snyder, Troubleshooting LC Systems., page 456. Some of us old guys get pretty much upset at some attempts to re-invent the world. (Nothing against improving things, but external to the unknown analysis, etc., is selfsustaining...)

I have question related to internal standard.
We want to determine 2 active substances which have extreme different in concentration from cream dosage form (each g of cream contains 1 mg of substance A and 50 mg of substance B). We would use centrifuge tube instead of volumetric flask for assay preparation. So, internal standard is the preferred option. The problem is we could not use one internal standard for both peaks due to their huge differences in area and properties. Is it all right to use two internal standards (one for each substance)?

It is not uncommon to use more than one internal standard in an assay, if there is a need to do so. In your case, it appears to be reasonable to use two internal standards.