Abstract: The nicotine in tobacco was extracted with mixture of 0.5% acetic acid glacial and 99.5% isopropyl alcohol and determined directly by gas chromatography. The relationship between quantity of nicotine and peak area ratio was linear in the range of 0-18.05mg/25mL and its correlation coefficient was 0.999 9. The mean recovery of nicotine was from 98.18% to 100.06%. It was proved that the method was accurate, reliable and suited to determine the content of nicotine in tobacco rapidly.