Background: Bioanalysis of conventional methods for compounds with permanent positive charge leads to peak tailing in separation and matrix effects in MS. This study describes a novel, rapid and sensitive method for quinolinium-containing compounds quantification. Results & methodology: A charged surface hybrid chromatography-tandem MS/MS using one-step protein precipitation dilution technique has been developed for determining analytes in plasma. We found symmetric peak and high recoveries for the analytes without matrix effect. All calibration curves had good linearity (r 0.991). The intra- and inter-assay precision was within 15% and the accuracy ranged from 88 to 103%. The method has been successfully applied to the PK study. Conclusion: The proposed method was sensitive, reproducible and applicable to other permanent positively charged compounds.