retention time and above the threshold. A Top N approach was opti-mized such that in a given time, when N abundant masses in theinclusion list within 10 ppm range are detected in the survey scan,N times dd-MS2are triggered to select the ion in the inclusion list,fragmented in HCD cell, collected in the C-trap and analyzed inthe Orbitrap, however, due to the N multiple ions fragmentationall entered Orbitrap at once, the mixed fragmentation spectrummay not be able to compare to the known fragment pattern. AGC(Automatic Gain Control) target and maximum ion time is used tocontrol the number of ions and maximum time taken to fill theC-trap, providing a duty cycle around 1.5 s.Chromatographic conditions were studied in order to achievethe best separation and retention for the compounds. First, sev-eral experiments were performed on different mobile phases