[35] Assay of phylloquinone in plasma by high-performance liquid chromatography with electrochemical detection

The extraction of vitamin K from plasma is usually achieved by first denaturing the vitamin K-lipoprotein complex with a polar solvent followed by extraction into a nonpolar solvent. Solid-phase extraction (SPE), a more contemporary technique for sample preparation that has been widely adopted in many other fields of clinical analysis, has only occasionally been adopted for the direct extraction of phylloquinone from plasma although it has been used extensively for the purification of vitamin K-rich fractions from solvent extracts. Three physicochemical techniques have been used to detect vitamin K in high-performance liquid chromatography (HPLC) analyses, namely, UV spectrophotometry, electrochemistry, and spectrofluorimetry. HPLC with fluorescence detection of the K vitamins exploits the highly fluorescent nature of vitamin K hydroquinone produced either by postcolumn electrochemical or chemical reduction. A number of chemical methods for the reduction of vitamin K have been employed including use of sodium borohydrate (NaBH4), tetramethylammonium octahydridotriborate [(CH3)4NB3H8] contained either in a postcolumn reaction tube or incorporated into the mobile phase, zinc metal in the presence of zinc ions, and platinum catalysts.