Bioequivalence and Bioavailability Forum

Dear Ohlbe,

Thank you for going through my query, please find below answers to your questions you have raised for my query

❝ 1. What internal standard are you using: stable-labelled, structural analogue, other ?

No, internal standard is not stable-labelled, structural analogue, but the internal standard response throughout analysis is reproducilbe with excellent % CV.

❝ 2. What type of sample preparation are you using: protein precipitation, liquid-liquid extraction, solid phase extraction ?

sample preparation is by liquid-liquid extraction method.

❝ 3. Any "strange result" in the validation ?

No, method validation was extremely smooth and satisfactory with hardly 3 to 4 QCs out of acceptance criteria throughout validation.

❝ 4. What do you mean by "variation in response" of the analyte: do you mean that for your QC samples, for a same concentration you get a very different response for the analyte, while the response for the internal standard is stable ? And how much "variation" do you see ?

variation in response means in system suitability test (prepared in matrix) first five injection shows reproducible response but the sixth injection shows different response for only that analyte while another analyte and IS shows good reproducibility. Similar results are observed during subject analysis after 4 to 5 injections there is change in response for that particular analyte and due to which we have to reject the batch (e.g. in cc and QC sample shows accuracy of 70 or 140 % for any of the in-between standard/QC sample).

❝ What LC/MS/MS parameter do you plan to change ?

I planning to change slightly Source parameters of LC/MS/MS and not the compound parameters because I think gases need to be optimized.

Once again thank you,
Regards
Ketan

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Edit: Standard quotes restored. [Helmut]