Development of a non-derivatization high-performance liquid chromatography method with resonance Rayleigh scattering detection for the detection of sisomicin in rat serum

A resonance Rayleigh scattering (RRS) detection approach was developed to detect sisomicin (Siso) in rat serum following chromatographic separation. The detection principle is based on the enhancement of RRS intensity of ion-association complex formed from aminoglycosides and pontamine sky blue (PSB) used as molecular recognition probe. The high-performance liquid chromatography (HPLC) coupled with RRS detection scheme was implemented post-column by mixing a PSB solution with the column eluent prior to detection. The RRS signal was detected by fluorescence detector at λex = λem = 365 nm. Separation and detection conditions were optimized. Siso and etimicin (Eti) chosen as the internal standard (IS) were separated on a C18 reversed phase column with the mobile phase consisting of a ternary mixture of 20 mM sodium acetate aqueous solution–methanol (92:8, v/v) containing 0.22% TFA (v/v). The limit of detection (S/N = 3) for Siso was 18 ng. A calibration curve ranged from 25 ng to 700 ng shown to be linear. The presented method was applied for the determination of Siso in rat serum and used for the pharmacokinetics study of Siso in rat.