UPLC Separation and Detection of Vitamin K

The UPLC system consisted of the ACQUITY UPLC sample manager and a binary solvent manager (Waters). Sample separation was performed using a VanGuard BEH C18 (1.7 µm) as the precolumn (Waters) and an ACQUITY UPLC BEH C18 (1.7 µm, 2.1 mm × 100 mm) column (Waters) as the main column. Vitamin K₁ and MK-7 were detected with the ACQUITY UPLC fluorescent detector (Waters) after post-column reduction using CQ-R 2.0 × 20 mm column (OSAKA SODA, Osaka, Japan). The column temperature was maintained at 50 °C during analysis. The mobile phase was a mixture of Milli-Q water (solvent A) and liquid chromatography-grade methanol (solvent B). The extraction and emission wavelengths of the fluorescence detector were set to 244 nm and 430 nm, respectively.