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目的:建立高效液相色谱法检测乳及乳制品中羟脯氨酸的方法。方法:乳制品中的胶原蛋白经过酸水解后生成包括羟脯氨酸在内的氨基酸混合物,用6-氨基喹啉基-N-羟基琥珀酰亚胺基甲酸酯(AQC)试剂衍生后测定羟脯氨酸,以缓冲液-乙腈水溶液为流动相,经过C18柱分离,紫外248 nm检测,以顺式-L-羟脯氨酸为内标定量分析。结果:在2μg/ml～30μg/ml范围内呈良好线性关系,40μg/g、100μg/g和300μg/g三个水平添加平均回收率99.7%～120.5%,精密度RSD<5%,方法定量限为4μg/ml(液态奶),方法检测限为2.0μg/ml(液态奶)和10μg/ml(奶粉,乳酪)。结论:方法灵敏度高,准确度好,实用性强,可用于乳及乳制品中羟脯氨酸的检测。
Objective: To establish a method for the determination of hydroxyproline in milk and dairy products by high performance liquid chromatography. Methods: Collagen in dairy products was acid-hydrolyzed to generate amino acid mixture including hydroxyproline, which was determined by derivatization with 6-aminoquinolyl-N-hydroxysuccinimide ester (AQC) reagent Hydroxyproline, buffer - acetonitrile aqueous solution as the mobile phase, after C18 column, UV detection at 248 nm, cis-L-hydroxyproline as an internal standard quantitative analysis. Results: There was a good linear relationship in the range of 2μg / ml ~ 30μg / ml. The average recoveries were 99.7% ~ 120.5% and the precision RSD was less than 5% at 40μg / g, 100μg / g and 300μg / Limit of 4μg / ml (liquid milk), the method detection limit of 2.0μg / ml (liquid milk) and 10μg / ml (milk powder, cheese). Conclusion: The method has high sensitivity, good accuracy and practicability. It can be used to detect hydroxyproline in milk and dairy products.