Bioequivalence and Bioavailability Forum

Dear Deepak,

❝ The % recovery was determined by comparing the mean peak area of 6 replicates of extracted quality control samples at High, Middle and Low concentrations against respective mean peak area of 6 replicates of un-extracted quality control samples at High, Middle and Low concentrations.

Meaning that what you observe is a mix of extraction recovery and matrix effect. If you want to see the true extraction recovery you have to compare to samples spiked post-extraction. You can combine with matrix effects experiments.

❝ Internal standard used was deuterated.

Makes no difference. You are looking at peak areas, not peak area ratios.

❝ Kindly confirm recoveries markedly higher than 100% for LQC, MQC2 and MQC1 are acceptable as per USFDA guidelines??

Did you see any limit for absolute recovery in either direction in the FDA guidance ?

❝ Also specify reason for higher recoveries.

Multiple. The most probable: analytical variability (your results are well within the 15 % acceptance limits for precision and accuracy). Other possibilities include ion enhancement, sample preparation, calculation errors...