Abstract:

A method for the determination of vitamin K₁ and vitamin K₂ in modulation milk powder was developed by high performance liquid chromatography (HPLC) coupled with post-column reduction. The samples were dissolved in water, lipase hydrolyzed, saponified with 2.5 mol/L sodium hydroxide solution and ethanol solution, extracted with n-hexane, and dissolved in methanol after concentration. The vitamin K were first separated on an Xbridge C₁₈ column and then on a zinc powder reduction column, and detected using a fluorescence detector. The excitation and emission wavelengths were 326 nm and 432 nm, respectively. An external standard method was used for quantification. The results showed that the linearities of vitamin K₁ and vitamin K₂ was in the ranges of 0.0025-2.0 μg/mL and 0.01-2.0 μg/mL, respectively, with correlation coefficients both greater than 0.999. The spiked recoveries were 80.39%-94.39% and the precisions were 0.85%-3.98%. The limits of detection of vitamin K₁ and vitamin K₂ were 0.07 μg/100 g and 0.2 μg/100 g, respectively. The limits of quantification of vitamin K₁ and vitamin K₂ were 0.2 μg/100 g and 0.8 μg/100 g, respectively. The method has high sensitivity and good repeatability, and gives accurate results. It is suitable for the analysis and determination of the vitamin K₁ and vitamin K₂ in formula milk powder.

Key words: high performance liquid chromatography (HPLC), fluorescence detector, post-column reduction, vitamin K₁, vitamin K₂, modulation milk powder