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目的:建立高效液相色谱快速测定血管紧张素转换酶I抑制剂活性的方法。方法:以马尿酰-组胺酰-亮氨酸为底物,血管紧张素转换酶Ⅰ为催化剂,酶促反应产物马尿酸为检测指标,卡托普利为抑制剂的代表进行方法学研究。采用Phenomenex Luna C18色谱柱,流动相为甲醇和乙酸溶液,等度洗脱,检测波长为228 nm。根据保留时间定性,外标峰面积法定量。结果:马尿酸最低检测限为0.01μmol/L,加标回收率为93.8%~103%。应用该方法测得实验室制备的鳕鱼寡肽浓度与其ACE抑制率呈自然对数关系,计算得鳕鱼寡肽对ACE的IC50为4.56 mg/ml。结论:该方法可作为体外测定血管紧张素转换酶抑制剂活性的方法,具有操作简便快捷、精密度和准确性高的特点。
Objective: To establish a method for rapid determination of angiotensin converting enzyme inhibitor activity by high performance liquid chromatography. Methods: Using hippuryl-histidyl-leucine as substrate and angiotensin converting enzyme Ⅰ as catalyst, the enzymatic reaction product hippuric acid was used as a detection index, and captopril was used as a representative of inhibitors for the methodological study . The Phenomenex Luna C18 column was used. The mobile phase consisted of methanol and acetic acid and wasocratically eluted. The detection wavelength was 228 nm. According to the retention time qualitative, external standard peak area method. Results: The detection limit of hippuric acid was 0.01μmol / L and the recoveries were 93.8% -103%. Using this method, the natural logarithm of the concentration of cod oligopeptide prepared in the laboratory and its ACE inhibition rate was calculated. The IC50 of cod oligopeptide to ACE was calculated to be 4.56 mg / ml. Conclusion: This method can be used as a method to determine the activity of angiotensin converting enzyme inhibitor in vitro. It has the characteristics of simple and quick operation, high precision and high accuracy.