Starch and cellulose are pivotal determinants of tobacco quality. In this work, starch and cellulose in flue-cured tobacco leaf were extracted through a sequential extraction process. First, an aqueous solution of tobacco leaves was prepared, and starch was extracted by boiling the leaves in water. The precipitate was separated, and the cellulose within it was hydrolyzed using a sulfuric acid solution. Then, starch content was quantified by a colorimetric reaction with iodine solution, while cellulose content was determined by using anthrone as a chromogenic agent. The reaction conditions were optimized to ensure accuracy, and the recovery rate and relative standard deviation (RSD) were calculated under these optimum conditions. The recovery rates for starch and cellulose in the tobacco samples ranged from 90 to 105%, with the RSD of interday and intraday determination being less than 5%. These results demonstrate that the proposed method is both sensitive and reproducible, making it suitable for the rapid and precise quantification of starch and cellulose in tobacco samples.