OBJECTIVE： To optimize the solution preparation in related substance test of cefotaxime sodium. METHODS： HPLC method was adopted to determine the total amount of impurities in cefotaxime sodium. Using phosphoric acid buffer solution pH， placing temperature， standing time and illumination as factor， the total amount of impurities as indexes， the preparation condition of solution was optimized by L9（34） orthogonal test. The validation test was carried out. RESULTS： The optimal preparation method was as follows as the sample was dissolved with a solvent containing pH 6.50 phosphate buffer protected from light. The test temperature and the sample temperature were set at 5 ℃， and the sample was injected within 5 min after the preparation of the solution. CONCLUSIONS： The optimized method is reproducible and reliable.