Abstract: In order to determine the content of free solanesol in tobacco leaves quickly and efficiently, a rapid Ultra-high Pressure Liquid Chromatography (UPLC) method was developed. By ultrasonic extraction with methanol, the extracting solution was membrane filtered to analyze by UPLC. The extract was eluted with methanol onto a short 1.7 μm BEH C18 column, and solanesol was separated rapidly in 5 min. According to the ion peaks of m/z 613.57 and 631.58, qualitative analysis was carried out by Electrospray Ionization-Quadropole Time-of-Flight Mass Spectrometry (Q-TOF-MS). The results showed that the linearity range of the proposed method was 0.765-76.5 μg/mL, the RSD% was 2.62%, and the average recovery rate was 91.1%. The method was appropriate for quick and sensitive determination of free solanesol in tobacco.