正己烷提取-高效液相色谱法及其串联质谱法分别测定婴幼儿配方奶粉中维生素A、D_(3)含量

Determination of vitamin A and D_(3) in infant formula milk powder by n-hexane extraction-high performance liquid chromatography and tandem mass spectrometry

目的建立正己烷提取,高效液相色谱法(high performance liquid chromatography,HPLC)和高效液相色谱-串联质谱法(high performance liquid chromatography-tandem mass spectrometry,HPLC-MS/MS)分别测定婴幼儿配方奶粉中维生素A(vitamin A,VA)和维生素D_(3)(vitamin D_(3),VD_(3))含量的方法。方法样品中加入VD_(3)-d_(3)内标后皂化,正己烷提取,采用去离子水清洗正己烷提取液以去除提取液中的氢氧化钾。加入氯化钠促进水和正己烷两相分层,将洗至中性的正己烷提取液浓缩至干。加入甲醇定容至10 mL,取1 mL溶液用高效液相色谱法测定VA。剩余的9 mL定溶液浓缩至干后再用1 mL甲醇复容,采用高效液相色谱-串联质谱法测定VD_(3)。结果VA的线性范围为0.2~6.0μg/mL,方法检出限为30μg/100 g,方法定量限为100μg/100 g;VD_(3)的线性范围为0.01~0.20μg/mL,方法检出限为0.5μg/100 g,方法定量限为1.0μg/100 g。批内相对标准偏差和批间相对标准偏差均小于5%。使用该方法成功通过了2020年婴幼儿配方奶粉中VA、VD_(3)样品能力验证考核。结论该方法所用提取试剂少且操作简单快捷,适用于婴幼儿配方奶粉中VA和VD_(3)的测定。

Objective To establish a method for the determination of vitamin A(VA)and vitamin D_(3)(VD_(3))in infant formula milk powder by high performance liquid chromatography(HPLC)and high performance liquid chromatography-tandem mass spectrometry(HPLC-MS/MS).Methods The sample was saponified after the addition of VD_(3)-d_(3) as internal standard,and was further extracted with n-hexane,cleaned with deionized water to remove potassium hydroxide in the extract.Sodium chloride was added to promote the two-phase stratification of water and n-hexane,and the n-hexane extract washed to neutrality was concentrated to dryness.Ten millilitres of methanol was added to dissolve the dried sample,and 1 mL of the solution was sampled for the determination of VA by HPLC.The remaining 9 mL of the fixed solution was concentrated to dryness and reconstituted with 1 mL of methanol for the determination of VD_(3) using HPLC-MS/MS.Results The linear range of VA was 0.2-6.0μg/mL,the method limit of detection was 30μg/100 g,and the method limit of quantitation was 100μg/100 g;the linear range of VD_(3) was 0.01-0.20μg/mL,the method limit of detection was 0.5μg/100 g,and the method limit of quantitation was 1.0μg/100 g.The relative standard deviations within and between batches were less than 5%.The method has been successfully used to verify the capability of VA and VD_(3) samples in infant formula milk powder in 2020.Conclusion This method has the advantages of less extraction reagents and simple and quick operation,and is suitable for the determination of VA and VD_(3) in infant formula milk powder.