Abstract:

Objective To establish the method of high performance liquid chromatography with electrochemical detection (HPLC-ED) for determination of content of N-stearoyltyrosine (NsTyr) to monitor the purity of product during reactions, optimize the preparation process and evaluate the stability of NsTyr solution. Methods The separation was performed on an Agilent Eclipse XDB-C8 column (5 μm, 4.6×150 mm). The mobile phase was methanol-water (80∶20). Detection potential: 600 mV; flow rate: 0.7 mL/min; column oven temperature: 25 ℃; sample volume: 10 μL. The stability trials of NsTyr in methanol were tested under certain concentrations and varied storage circumstances, based on current methodology verified from content assays on NsTyr after numerous recrystallization. Results Amid 0.03-10 μg/mL of NsTyr, the area of peak was in linear correlation with concentration (r²＞0.999). The methodology was fully validated of rational precision, sensitivity and selectivity. The purity of NsTyr after recrystallization for three times surpassed 99%, and the concentration alternation in solution under short-term ambient temperature, long-term refrigeration storage and freezing-thaw circle test was within 5%. Conclusion Endowed with fairly high convenience and reliability, the established method is feasible for both quality control and quantity determination of NsTyr. The high purity and solubility of NsTyr may facilitate the cell culture and animal experiment.

Key words: high performance liquid chromatography, electrochemical detection, N-stearoyltyrosine, purity, stability