Recovery issues for API in presence of excipients in UPLC
I have a situation where technical input from Waters or UPLC forum will be much appreciated. Essentially for a drug substance solution in buffer and methanol, we get 100% recovery in HPLC. We also get ~ 100% recovery with same solution in UPLC. Next, the drug substance is spiked with the excipients for linearity check and provided 100% recovery in HPLC. The same spiked solution in UPLC under identical condition as DS analysis now gives 95-96% recovery.The mobile phase is isocratic : Water: Methanol: TFA (47:53:0.1), wash solvents are water, methanol and TFA. The sample solvent is ammonium acetate buffer pH 5.0: methanol. PLNO mode 5 uL loop and 2 uL injection.Any clues why the UPLC recovery drops for the DS+excipient solution?.