The invention discloses a method for preparing a tobacco ointment from distiller's yeast through fermentation. The method is characterized in that the tobacco ointment is prepared by the following steps: A, performing steam sterilization on tobacco powder; B, inoculating the distiller's yeast for fermentation; C, performing ethanol extraction, filtration and concentration, thereby obtaining the tobacco ointment. Due to adoption of a wine making fermentation process, the method is gentle in production condition, high in efficiency and small in pollution, and as common distiller's yeast used in the wine brewing industry, the distiller's yeast is low in price, good in culture activity and good in infectious microbe resistance, and fermentation conditions are easy to handle and control. The tobacco ointment prepared by using the method is excellent in stability and repeatability and has a relatively remarkable fragrance enhancing effect and unique style characteristics, and in addition, selective regulation and control can be achieved by changing the distiller's yeast. The method is simple in production process, low in production cost and applicable to industrial production.