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通过对高效液相色谱条件中色谱柱、流动相、流速、检测波长、柱温优化的试验,建立了反相高效液相色谱对鲜牛乳中α-乳白蛋白、β-乳球蛋白、乳铁蛋白同时微量检测的分析方法,最终确定采用Agela Venusil ASB C8(5μm,4.6 mm×250 mm,150μm)色谱柱;三氟乙酸/乙腈/水流动相,流速0.8 m L/min,检测波长215 nm,柱温30℃的色谱条件,并利用外标法定量。结果表明,α-乳白蛋白、β-乳球蛋白、乳铁蛋白3种组分的线性关系良好,相关系数分别为0.9999、0.9991、0.9998,检出限为0.001~0.012 mg/m L,样品平均回收率在86.5%~109.08%之间,精密度和重现性试验RSD值均小于5%。该方法的前处理简单、耗时少,样品损失少,分离效果好,能够实现牛乳中3种蛋白的同时检测。
Through high performance liquid chromatography column conditions, mobile phase, flow rate, detection wavelength, column temperature optimization test established reverse phase high performance liquid chromatography in fresh milk alpha-lactalbumin, beta lactoglobulin, lactoferrin The analytical method was used to detect the protein at the same time. Finally, the mobile phase consisted of Acetone, acetonitrile and water with a flow rate of 0.8 m L / min with Agela Venusil ASB C8 (5 μm, 4.6 mm × 250 mm, 150 μm) , Column temperature 30 ℃ chromatographic conditions, and the use of external standard method. The results showed that there was a good linear relationship between the three components of α-lactalbumin, β-lactoglobulin and lactoferrin with the correlation coefficients of 0.9999, 0.9991 and 0.9998 respectively. The detection limits were 0.001 ~ 0.012 mg / mL. The average The recoveries ranged from 86.5% to 109.08%, and the RSD values for precision and reproducibility were less than 5%. The method has the advantages of simple pretreatment, less time consumption, less sample loss, good separation effect, and simultaneous detection of three proteins in milk.