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目的建立高效液相色谱法检测葡萄酒中溶菌酶的含量。方法葡萄酒样品经5%盐酸溶液酸化后,采用流动相A乙腈∶三氟乙酸∶水(50:0.2:49.8,V:V:V)和流动相B乙腈∶三氟乙酸∶水(1:0.2:98.8,V:V:V)进行梯度洗脱,经反相苯基柱分离后,用高效液相色谱仪荧光检测器或紫外检测器进行测定,通过保留时间定性,外标法定量,并优化流动相的配比、梯度洗脱程序和样品的酸化条件。结果本方法在50、200、500和750 mg/L添加水平下的回收率为93.5%~99.2%,相对标准偏差为1.62~5.74%(n=6),方法定量限为50 mg/L。结论本方法具有分离效能高、重现性好、准确可靠,灵敏度高等优点,测定结果与酶联免疫法结果一致。该方法可以满足葡萄酒中溶菌酶的检测需求。
Objective To establish a method for the determination of lysozyme in wine by high performance liquid chromatography. Methods The samples of wine were acidified with 5% hydrochloric acid solution and the mobile phase A consisted of acetonitrile: trifluoroacetic acid: water (50: 0.2: 49.8, V: V: V) and mobile phase B acetonitrile: trifluoroacetic acid : 98.8, V: V: V) gradient elution after reversed-phase phenyl column separation, high performance liquid chromatography with a fluorescence detector or UV detector was determined by the retention time of qualitative, external standard and Optimize the mobile phase ratio, gradient elution program and sample acidification conditions. Results The recoveries of this method were 93.5% -99.2% at 50, 200, 500 and 750 mg / L with the relative standard deviations of 1.62-5.74% (n = 6). The limit of quantification was 50 mg / L. Conclusion The method has the advantages of high separation efficiency, good reproducibility, accuracy and reliability, and high sensitivity. The determination results are consistent with the results of enzyme-linked immunosorbent assay. The method can meet the detection requirements of lysozyme in wine.