Determination of Vitamin K1 and Vitamin K2 in Foods by HPLC-FLD
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摘要: 建立高效液相色谱-荧光检测法（HPLC-FLD）测定食品中维生素K1和维生素K2含量的分析方法。方法：配方奶粉、酸奶、纳豆冻干粉和鸡胗冻干粉经酶解、异丙醇分散、正己烷萃取、旋转蒸发复溶后，使用Thermo AcclaimTM 120 C18色谱柱（150 mm×4.6 mm，5 μm）分离，锌粉还原柱（50 mm×4.6 mm）衍生，以900 mL甲醇：100 mL四氢呋喃（含冰醋酸0.3 mL，氯化锌1.5 g，无水乙酸钠0.5 g）为流动相，激发波长为243 nm，发射波长为430 nm，分离维生素K1（Vitamin K1，VK1）、四烯甲萘醌（menaquinone-4，MK-4）、七烯甲萘醌（menaquinone-7，MK-7）和九烯甲萘醌（menaquinone-9，MK-9）。结果：VK1、MK-4、MK-7和MK-9在20~1000 μg/L范围内与峰面积有良好线性关系。配方奶粉、酸奶、纳豆冻干粉和鸡胗冻干粉中VK1、MK-4、MK-7、MK-9含量的相对标准偏差（RSD）均小于5%，表明精密度好。配方奶粉、酸奶和纳豆冻干粉不同水平加标回收率分别为85.0%~107.8%、85.8%~109.6%、85.1%~102.7%，回收率的RSD均小于5%，表明回收率高，相对标准偏差小。结论：实验建立的方法重复性好、准确度高，可以精准定量食品中维生素K1和维生素K2的含量。
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关键词:
- 高效液相色谱-荧光检测法(HPLC-FLD) /
- 维生素K1(VK1) /
- 维生素K2(VK2) /
- 四烯甲萘醌(MK-4) /
- 七烯甲萘醌(MK-7) /
- 九烯甲萘醌(MK-9)
Abstract: An analytical method was developed for determining vitamin K1 and vitamin K2 in foods by high performance liquid chromatography with Fluorescence Detector(HPLC-FLD). Milk powder,yoghurt,natto freeze-dried powder and chicken gizzard freeze-dried powder were hydrolyzed by enzyme,dispersed by isopropanol,extracted by n-hexane,evaporated by rotation and redissolved. The separation of vitamin K1(VK1),menaquinone-4(MK-4),menaquinone-7(MK-7)and menaquinone-9(MK-9)were achieved on Thermo AcclaimTM120 C18 column(150 mm×4.6 mm,5 μm)and derivatived by zinc powder reduction column(50 mm×4.6 mm)using 900 mL methanol:100 mL tetrahydrofuran(containing 0.3 mL glacial acetic acid,1.5 g zinc chloride,0.5 g anhydrous sodium acetate)as the mobile phase at excitation wavelength was 243 nm,emission wavelength was 430 nm. The results showed that the linear range of VK1、MK-4、MK-7、MK-9 were 20~1000 μg/L. The relative standard deviation(RSD)of VK1,MK-4,MK-7,MK-9 in dried milk,yoghurt,natto freeze-dried and chicken gizzard were less than 5%,indicating high precision. The recovery rate of VK1,MK-4,MK-7 and MK-9 in dried milk,yoghurt,natto freeze-dried were 85.0%~107.8%,85.8%~109.6%,85.1%~102.7% and RSD were less than 5%,indicating high recovery rate and small relative standard deviation. Therefore,this developed method is high reproducibility and accuracy. It can be applied to accurate determine of vitamin K1 and vitamin K2 in foods. -
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