The invention discloses a production method of ligno-cellulosic bio-ethanol. Specifically, the method includes the steps of: 1) pretreating cellulose raw material to obtain a water-containing material; 2) using ammonia water to adjust the pH value of the water-containing material obtained in step 1) to 4-6, and then conducting enzyme hydrolysis treatment to obtain mash; and 3) using an alkaline agent to adjust the pH value of the mash to 4-6, then conducting fermentation treatment to obtain an ethanol-containing material, and separating ethanol from the ethanol-containing material, with the alkaline agent being at least one of sodium hydroxide, potassium hydroxide, sodium carbonate, calcium oxide and calcium hydroxide. The production method of ligno-cellulosic bio-ethanol provided by the invention guarantees the production of ligno-cellulosic bio-ethanol, at the same time significantly reduces the production cost and environmental pollution.