In this paper,some high fatty acids in tobacco were analyzed by Ultra-high performance liquid chromatography with evaporative light-scatter detector(UPLC/ELSD).The saponification influence factors of the high fatty acids,including temperature,reaction time,hydrogen ion concentration(pH),volume of extracting agents,etc,were systematically studied through the orthogonal experiments.The best saponification conditions were as follows:saponification temperature:60 ℃,KOH:3 mol/L,CH2Cl2:15 mL,reaction time:60 min.In addition,a new UPLC/ELSD method was established for the determination of six high fatty acids,including linolenic acid,linoleic acid,palm acid,oleic acid,myristic acid and stearic acid in tobacco on the C18 chromatography column by gradient elution with acetonitrile-acetic acid aqueous solution(80∶20,by volume) as mobile phase at a flow rate of 0.3 mL/min,a drift tube temperature of 50 ℃ and a gas flow rate of 2.89 L/min.The linear range of the method was in the range of 42.4 -520.0 mg/L，with detection limits of 3.64-11.98 mg/L.The RSDs(n=7) were in the range of 4.9%-13.5%,and the recoveries ranged from 83% to 104%.This method is rapid,convenient and sensitive,and is suitable for the determination of high fatty acids in tobacco.