To ensure sterile penicillin sodium powder, strengthen airtight operation for the sterile process, the sampling method at the crystallization end-point was improved. Through the production test, the crystallization mother liquor moisture and distillate moisture were detected at the same time, the test data were analyzed by the least square method, it is found that there is obvious linear relationship between both. Therefore, the distillation liquid moisture detection instead of crystallization mother liquor moisture detection, is used as the judgment standard of crystallization end-point, and according to the relationship between the two methods, the determined distillate liquid moisture qualified moisture content < 7.50%. Through the improvement of the sampling method, the closure of the aseptic operation process is ensured.