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Vitamin K in Serum/Plasma LC-MS/MS Analysis Kit

Vitamin K is a cofactor for enzymatic modification of glutamic acid residues (Glu) to gamma-carboxyglutamic acid residues (Gla) in vitamin K-dependent Gla proteins. These Gla proteins are necessary for hemostasis, bone metabolism, vascular calcification and cell proliferation. Vitamin K is found in nature as phylloquinone (vitamin K1) in green leafy vegetables, algae and some plant oils, and menaquinones (vitamin K2 also termed MK-n, where n is the number of isoprenoid units) can be found in meat, eggs and fermented food. A synthetic form of vitamin K menadione (vitamin K3) can be found as an additive in some animal feeds.

  • Highlights

    Simultaneous measurement the concentrations of 5 forms of vitamin K.

    Total run time is 12 min.

    One sample preparation for all forms.

    Easy liquid-liquid extraction step following by protein precipitation for the sample treatment- no need for SPE.

    Long life span of HPLC column.
  • Parameters

    Vitamin K1-Phylloquinone, Vitamin K1-2,3-Epoxide, Vitamin K2-7-Menaquinone, Vitamin K2-2,3-Epoxide and Vitamin K2-4-Menaquinone
  • Matrix

    Serum, plasma

Sample Preparation

  • Step 1
    Pipette 400 µL of serum sample/working calibration standard into a glass centrifuge tube.
  • Step 2
    Add 800 µL of Reagent 1, additionally vortex for 5 sec.
  • Step 3
    Add 25 µL of internal standard working stock solution, then 2 ml Reagent-2 and vortex for 3 sec.
  • Step 4
    Centrifuge at 4000 rpm for 5 min.
  • Step 5
    Transfer 2.4 ml of the upper phase to a new glass tube (tube-3) and evaporate under nitrogen stream. (Upper phase can be evaporated at 7 psi during ~15 min using water bath at the temperature of 30 °C).
  • Step 6
    Reconstitute tube-3 with 100 µl of Reagent-3 and transfer to a HPLC vial prior to injection.
  • Chromatogram

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