Biochemical analysis

To quantify DHS in the plasma of mice, the method for resveratrol detection by Muzzio et al.³⁴ was adapted and performed for its analogue. HPLC-UV-ESI/MS and analyses have been carried out on a ThermoFisher Scientific HPLC/UV/MS system (Thermo Scientific LCQ FLEET). Separation of DHS from plasma components was achieved using a Luna C18 3μm 2×100 mm column maintained at r.t., with a flow rate of 0.2 mL/min, and injection volume 20 μl. The mobile phase consisted of 5 mM ammonium acetate in water containing 2% propan-2-ol and methanol with 2% propan-2-ol. A mobile phase gradient was performed. An Electro Spray Ionization (ESI) interface was used as ion source, operating both in negative and in positive ion mode. Acquisition was performed in full scan mode (mass range 50–1000 Da). Ion spray voltage, capillary voltage were −5000V and −0.5 V in negative ion mode and + 5000V and +35V in positive ion mode. The capillary temperature was 220 °C. The DHS quantification in plasma was performed using a calibration standard curve (5–5000 ng/mL) starting from stock solution (100 mM) in DMSO.