Identification of menaquinones (vitamin K2 homologues) as novel constituents of honey

Our recent research indicated that honey active macromolecules form colloidal particles that scatter the light and produce elaborate UV spectral profile dominated by double absorption peaks at 240-250 nm. The absorption at 240-250 nm signified the stable honey conformation that supported antibacterial activity and hydrogen peroxide production. Our aim was to identify the bioactive constituent relevant to this absorption. The methodology included activity-guided fractionation of honey through size-exclusion chromatography, solid-phase extraction and UPLC-UV-MS. UV spectral analysis of UPLC peaks revealed compounds with UV λ (max) typical of naphtoquinones. The MS chromatograms showed mass ions differing by [M-68n] indicating a polyisoprene structure and the fragmentation patterns typical for menaquinones. The exact mass measurements of menaquinones using a quadrupole-Orbitrap mass spectrometry confirmed their identification as a series of MK-3 to MK-7 aptimers. Detection of menaquinones, previously unknown constituents of honey, suggests that they might play role in honey redox and antibacterial activities.