Paraquat and diquat are two of most widely used herbicides in the world. Both compounds are toxic for humans and animals. Presence of paraquat and diquat in water is regulated by EPA. Paraquat and diquat have two quaternary amines, making them very polar molecules. Paraquat and diquat also have similar formula. Due to low hydrophobicity and highly basic properties, they show limited retention on reversed-phase columns and produce very poor peak shape due to the residual silanol interactions. Primesep AB and Obelisc R column retain and separate these two compounds with perfect peak shape. Method can be used for analysis of paraquat and diquat in soil, ground and drinking water, and other samples. Method allows to separate both compounds in one run without ion-pairing reagent.