B-182 Advantage of Core-Shell Particle Morphology Over Fully Porous HPLC Column for the Analysis of Vitamin K Compounds by LC-MS/MS

Abstract

Abstract Background Vitamin K’s are a group of structurally similar, fat-soluble compounds. There are two natural forms K1 (Phylloquinone) and K2 (Menaquinone), where the later has short and longer chain homologues called Vitamin K2-MK-4 and Vitamin K2-MK-7 respectively. Due to the extremely hydrophobic nature and poor ionization characteristics of these compounds, LC-MS/MS analysis at low concentrations becomes challenging. In this communication we make parallel comparisons of three HPLC columns packed with fully porous and core-shell particles to showcase the superiority of core-shell columns for sensitive detection of vitamin K analytes by LC-MS/MS. Method We compared fully porous (Gemini and Luna, Phenomenex, Torrance, CA) and core-shell (Kinetex EVO, Phenomenex) C18 columns for this study. All columns evaluated were 50 × 2.1 mm with a 5 μm particle size to maintain low back pressure for the viscous organic modifier utilized. Deionized water and a combination of isopropanol and acetonitrile (1:1; v/v) were used as mobile phase A and B respectively for chromatographic separation. An Agilent 1290 Infinity series LC system was employed with a SCIEX 6500 QTRAP mass spectrometer under APCI for MS detection. Analyst software 1.7.2 used for data analysis. Results The LC-MS/MS method had a 4-minute runtime for chromatographic separation of the analytes including 1.5 min of equilibration time. The comparison data revealed the Gemini C18 column shows some higher selectivity and resolution values, whereas both EVO and Luna C18 columns were somewhat comparable. However, the core-shell EVO C18 column displayed much higher efficiency with narrower and compressed peak widths, which dramatically increased the peak height for all 3 analytes exhibiting the highest signal/noise ratio. Conclusion The core-shell Kinetex 5 μm, EVO C18 column with its unique organo-silica grafting shows much higher efficiency and sensitive detection capability for vitamin K analysis by LC-MS/MS compared to that of conventional fully porous silica particles.