The olive biophenol hydroxytyrosol in neutral aqueous solutions – a UPLC-MS/MS investigation of its stability and oxidative color formation

Olive pomace is a by-product of the olive oil industry rich in biophenols such as hydroxytyrosol, with potential to be valorized as a functional ingredient. To investigate the stability of hydroxytyrosol in neutral aqueous solutions, the oxidative transformation of 50 ppm hydroxytyrosol in real and simulated tap waters over 5 days was studied with UV/Vis spectroscopy and UPLC-MS/MS. Simulated tap water systems consisted of 241 mg/L NaHCO_3, with or without 139 mg/L CaCl₂, and a pH range of 6.5-8.0. Hydroxytyrosol in real tap water was completely transformed to various oxidation products after 1 day, exhibiting a marked red color (λ_max = 490 nm). Similarly, the simulated tap water system with a chemical composition closest to that of real tap water was largely able to reproduce this behavior with 85% hydroxytyrosol oxidation. A number of hydroxytyrosol oxidation products were annotated, including the red chromophore 2-(2-hydroxyethyl)-5-hydroxy-benzoquinone, which was formed through hydroxytyrosol autoxidation in the presence of bicarbonate ions. The chromophore's formation without addition of transition metals, enzymes or H₂O₂ is presented for the first time. The results have implications for the stability and color behavior of olive pomace-derived biophenols applied to neutral or mildly alkaline environments.