Abstract: Two Bacillus licheniformis strains Pb-WC09001 and Pb-WC09002 were isolated from the tropical aquacultural systems in Hainan. The effects of salinity, temperatures, and pH on their relative protease activities were determined by the lipid-excluded milk plate assay. With the increase of salinity, the relative enzyme activities of these two strains were on a downward trend, and when salinity increased to 50, Pb-WC09001 still had the ability producing protease and the relative enzyme activity was 1.56 U•mL^-1, which suggested that Pb-WC09001 might be more adaptable to high salt environment. In the range of 18—38 ℃(4 ℃as gradient difference), both strains had the highest relative enzyme activities at 34 ℃. When the culture temperature were at 22 ℃, the relative enzyme activities of Pb-WC09001 were highest; when the culture temperature were at at 30 ℃, the relative enzyme activities of Pb-WC09002 were highest, which suggested that the two strains have produced different type protease under the same temperature, and at low temperatures, Pb-WC09001 has a higher relative enzyme activity than Pb-WC09002. When they were cultured into 2×10³/mL at 30 ℃ and the salinity of 30, the most suitable pH values of the highest relative enzyme activity were 8.0 for Pb-WC09001, 7.5 for Pb-WC09002, their relative enzyme activities were 2.74 U•mL^-1. The results suggested that Pb-WC09001 had higher relative enzyme activities in marine aquaculture environment, especially in lower-temperature seasons.