Bioequivalence and Bioavailability Forum

Dear Vishal!

❝ As per my knowldge area threshold require to neglet those area of the

❝ chromatogram which are no need to quantify ...

So far so good.

❝ ... so you can put whatever value as area thresold but it should be

❝ less then the area which you found at LLOQ level.

As said in my previous post, it's not that easy - at least for internal standards methods...

❝ e.g. if the area of LLOQ is 12345 so you can put area thresold value less

❝ then the LLOQ area so whatever peak having the area less then area

❝ threshold will not reflect in your chromatogram.

Simple example (sample at LLOQ):
Analyte area 12345
IS area 246900
Ratio analyte/IS 0.05

Now you are loosing 5% somewhere in the sample transfer process, getting
Analyte area 11723
IS area 234555
Ratio analyte/IS 0.05 (=unaffected!)

If you would have set the threshold to 12345, the analyte would not have been integrated (in other words, you would report the sample as BLQ, which is not correct).
Just remember that it's OK at LLOQ to have an inaccuracy of ±20% together with precision of 20%.
If you set the area threshold to the analyte's response calculated at LLOQ for the average internal standard area from the CC, you will miss roughly 1/3 of samples by pure chance!
I agree with Rajendra, that the threshold should be set somewhere between LOD and 50% of LLOQ.