A detailed MS(n) study on an ion trap instrument of the quaternary ammonium pesticides paraquat, diquat, difenzoquat, mepiquat and chlormequat reveals a number of ions not reported previously, and has allowed examination of the fragmentation pathways. A number of transitions that are highly specific to each quat have been identified. Optimal ion trap operating conditions determined using Simplex optimisation can promote either detection of a particular fragmentation transition or a range of MS/MS product ions with a high overall signal response. Thus, fragmentation conditions were optimised to enhance the specificity or sensitivity of MS/MS methods.