Measurement of vitamin K homologues in biological fluids and tissues by APCI LC/MS

TL;DRAbstract

Vitamin K (K) is a family of molecules that contains a 2‐methyl‐1,4‐napthoquinone nucleus, including phylloquinone (K 1 ) with a isoprenyl side chain, menaquinones with unsaturated isoprenyl side chains (K 2 ; MK‐ n ) and menadione (K 3 ; MK‐0). Here we describe a sensitive and specific liquid chromatography mass spectrometry (LC/MS) method to measure K 1 , K 2 (MK‐4 & MK‐7), and K 3 in biological fluids and tissues. For all assays a Micromass ZQ 2000 MS detector coupled to a Waters 2695 HPLC was used. For K 1 & K 2 analysis an isocratic method consisting of 100% methanol delivered at 1.0 mL/min for 30 min was used. For K 3 , a gradient of 50–90% methanol delivered over 30 min at 0.2 mL/min was used. Internal standards of ubiquinone‐4 and ‐2 were used for K 1 /K 2 and K 3 , respectively. Calibration curves for the K forms generated a linear response from 1 to 50 nM, with a lower limit of detection of 0.2 nM (S/N of 3/1). K 1 was detectable in human plasma, but MK‐4 was not. In

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