Abstract: To simultaneously, accurately and rapidly determine free lipoid components in tobacco, an optimized supercritical fluid chromatography-tandem mass spectrometry (SFC-MS-MS) method was established to quantify free lipoid levels. Through systematic optimization of sample pretreatment conditions and instrumental parameters including chromatographic column type, mobile phase, modifier and ion source, accurate determinations of 15 lipoid components including α-cembra-trienediol, β-cembratrienediol, α-tocopherol, α-tocotrienol, β-sitosterol, campesterol, brassicasterol, cholesterol, stigmasterol, ergosterol, lanosterol, squalene, labd-13-ene-8, 15-diol, cis-abienol and solanesol in tobacco were achieved. The results showed that: 1) All analytes exhibited good linearity within the tested ranges, with all of the correlation coefficients (r) more than 0.99. 2) Recoveries at different spiked levels ranged from 81.6% to 114.9%. Intra- and inter-day RSDs were 1.79%-6.29% and 1.74%-7.29%, respectively. Limits of detection and limits of quantification were 0.000 87-0.26 μg/mL and 0.002 9-0.85 μg/mL, respectively. 3) Application of this method to various tobacco samples showed that this method could successfully detect the target lipoid components in different types of tobacco samples. Compared with traditional methods, this method exhibited strong matrix compatibility, high separation efficiency and sensitivity. It reduced organic solvent consumption and was eco-friendly. It could achieve simultaneous, accurate and rapid quantitative determination of main free lipid components in tobacco.