Objective To develop an ultra performance convergence chromatography (UPC2) method for determining l-OTS level and optical purity. Methods Reference substance method was used for quantitative determination. The separation of enantiomers was achieved on a chiral chromatographic column ACQUITY UPC2TM TrefoilTMAMY1(3.0 mm×50 mm, 2.5 mm) at the column temperature of 30 ℃ with the mobile phase of supercritical carbon dioxide-0.2% DEA in 2-propanol (95∶5) at a flow rate of 1.0 mL/min. Detection wavelength was set at 227 nm (compensation reference was 310-360 nm). Results l-OTS and its enantiomer (d-OTS) were effectively separated with the resolution of 2.30. Peak area was linear to mass concentration within the concentration rang of 40-1 000 mg/L (r=0.999 9, n=7). Average recovery was 99.8% (n=9) and the sample was stable within 24 h (RSD=0.81%). The LOD was 5 ng (S/N=4). Conclusion This method is simple and quick and can be used for determining l-OTS level and analyzing optical purity, which has been validated by methodology.