Abstract: The tobacco sample was treated with high-temperature resistant amylase at 95℃ for thirty minutes, during which the starch in tobacco was liquefied into dextrin, then dealt with glucoamylase at 55℃ for thirty minutes, during which the dextrin was hydrolyzed into glucose.The glucose in the resultant sample solution was determined with an autoanalyzer.The results showed that the relative standard deviation of the method was 2.0%, and the recoveries were from 96% to 100%.The starch in some tobacco leavers were also determined.