动物性食物不同形式维生素K检测方法的建立

Determination of vitamin K in animal foods

目的建立不同形式维生素K在线分析技术,探讨动物性食物中维生素K的含量分布。方法动物性食物样品在pH为8、脂肪酶(0.2 g)、蛋白酶(0.1 g)条件下水解4 h,再用异辛烷提取,经旋转蒸发、复溶后测定。使用Atlantis T3(4.6 mm×250 mm,5μm)色谱柱分离,以900 mL甲醇∶100 mL四氢呋喃(包含5 mmol冰乙酸、11 mmol氯化锌、6 mmol无水乙酸钠)为流动相,激发波长为320 nm,发射波长为410.3 nm,高效液相色谱-荧光检测法分离维生素K_1、四烯甲萘醌(menaquinone-4, MK-4)和七烯甲萘醌(menaquinone-7,MK-7)。结果方法线性范围为0.01～0.40μg/mL(决定系数>0.999),加标回收率为84.4%～124.2%,相对标准偏差<6%(n=6)。MK-4是猪、鸡肉中维生素K的主要形式,牛肉中以维生素K_1含量最高,白贝和鱿鱼中可检测到MK-7。结论建立的高效液相色谱柱后衍生法适用于动物性食物维生素K的测定,不同形式维生素K在不同动物体内的分布特征不同。

OBJECTIVE To establish a high performance liquid chromatography method with fluorescence detection for a variety of vitamin K and to assess the content of vitamin K in animal foods.METHODS Animal foods were hydrolyzed by 0.2 g lipase and 0.1 g protease in pH 8 for 4 hours,extracted with isooctane followed by rotary evaporation and reconstitution.The mobile phase was 900 mL methanol and 100 mL tetrahydrofuran which contained 5 mmol glacial acetic acid,11 mmol zinc chloride and 6 mmol anhydrous sodium acetate.The content of vitamin K1,menaquinone-4(MK-4),and menaquinone-7(MK-7)were separated on Atlantis T3 column(4.6 mm×250 mm,5μm)by high-performance liquid chromatography.The fluorescence detector was set at a wavelength of 320 nm for excitation and 410.3 nm for emission.RESULTS The linear range of the method was 0.01-0.40μg/mL,and coefficient of determination was>0.999.The spiked recoveries were 84.4%-124.2%with relative standard deviation was<6%(n=6).MK-4 was the main form of vitamin K in pork and chicken.The highest content of vitamin K1 was found in beef,and MK-7 could be detected in aquatic products.CONCLUSION The proposed method is successfully applied for the determination of vitamin K in animal foods.A variety of vitamin K are distributed differently in distinct animals.