Abstract: Taking unknown impurities as the focal point,the optimum crystallization pH value was investigated at 25℃.The effect of temperature on crystallization process was explored at a certain pH value.Gradient elution was utilized to analyze components and impurities in product.Selectivity coefficients of effective components and total unknown impurities to spiramycin were compared with each other.In view of qualified effective compositions and lower impurities contents,the most suitable terminal pH value of crystallization operation was 7.5-8.0,temperature was 25-35℃.The variation of unknown impurities contents was first adopted to explore bitespiramycin crystallization in the existing studies,which would have a profound guiding significance for the practical production.