Clostridium novyi type B is an anaerobic bacterium that causes black disease (infectious necrotic hepatitis). The alpha toxin is considered as a major virulence factor. The disease has been controlled using toxoid vaccines. Production of a high yield of bacterial toxin is very important in the preparation of effective toxoids. The present study aimed to follow the most satisfactory growth requirements to produce a high yield of alpha toxin. This study tested the effect of different concentrations of meat pieces (0. 0.5, 1.5, 2 and 2.5%), adding yeast extract in concentration of (0.5%), variable incubation times (24, 48, 72, 96 and 120 hours) and different pH values (7, 7.5 and 8).The results showed that the best media used for producing alpha toxin of Clostridium novyi type B was the one that contains yeast extract in the concentration of 0.5% with absence of meat pieces and adjusting pH between 7.5- 8 and incubation for 24- 48 hrs. that leads to high bacterial growth and toxin yield production optimum physical condition as incubated period 24- 48 hrs. and favorable pH for growth production and toxin yield of Clostridium novyi type B which lead consequently to produce a high potent Clostridium novyi vaccine.