The study used for determine the spray-dried optimal conditions for making the powder bacteriocin from Lactobacillus acidophilus. The experiments were investigated at the center of these factors for investigating the influence: temperature of drying air, flow rate input and additional maltodextrin before drying. Optimal experiment was investigated based on the response surface method, center of structure, the number of laboratory experiments are 13 (center repeat 5 times) with two main factors are the drying temperature and the ratio additional maltodextrin. The results were as follows: the best drying temperature is 153.6°C, the best dry concentration use for drying is 15.1 Bx, flow rate input is 5 rpm (4.67 ml/min). The spray drying conditions as above the authors obtain bacteriocin active preparations 202.208 UI/ml. Bacteriocin the authors obtain was heat stable. Bacteriocin active was maintained during spray drying.