Determination of apramycin sulfate in premix by precolumn derivatization-HPLC

Objective: To establish a pre-column derivatization-HPLC for determination of apramycin sulfate in premix. Methods: The separation was performed on a Thermo C18column( 150 mm × 4. 6 mm,5 μm) with pre-column derivatization by DNFB and the mobile phase was acetonitrile-water( containing 0. 5% acetic acid) using gradient elution program at the flow rate of 1. 0 m L·min-1. The detection wavelength was 350 nm and the column temperature was 30 ℃. And the contents of apramycin sulfate in three batches of premix were determined. Results: A linearity between derivative peak area and apramycin sulfate concentration was achieved in the range of 0-100. 0 μg·m L-1( r = 0. 999 8). The average recovery( n = 6) was 98. 70%,RSD = 1. 3%. The contents of apramycin sulfate in three batches( n = 3) were 24. 42 mg·g-1,24. 39 mg·g-1and 24. 63 mg·g-1,respectively,the corresponding RSDs were 0. 27%,0. 49% and 1. 0%. Conclusion: It is proved by methodology validation that the method can accurately determine the content of apramycin sulfate in the premix.