Bioequivalence and Bioavailability Forum

Dear chari,

Is the response systematically significantly higher in your unknown samples compared to the standards and QCs ? If you are using a LC/MS/MS method, it could be a matrix effect issue. Could also be a different behaviour during sample preparation, leading to a different recovery. A few questions and ideas:

- what method are you using for sample preparation ?
- are you using the same anticoagulant in your standards/QCs and in your unknown samples ?
- how did you investigate matrix effect during method development and validation ? Did you try any post-column infusion as part of development ?
- how variable is your IS response in your unknown samples ? I understand the response is higher, but is the CV of the IS response similar in unknown samples and in standards/QCs ?
- if you have more variability in your unknown samples, does it follow any specific pattern, for instance depending on the sampling time (looking like a PK curve) ? If this is the case you may have some interference from a metabolite. Enhancement is not frequent with electrospray ionisation (suppression is much more frequent), but it does happen with APCI.
- Oh, and what kind of IS are you using ? Stable isotope labelled, or structural analogue (what structure compared to your metabolites), or totally different structure ?

Have a look at this thread for previous discussions on this topic.

Regards
Ohlbe