Pressurized hot (subcritical) water and ethanol were used to extract betulin and antioxidants from birch bark. Betulin was found to be the major compound (around 26% (w/w)), which was able to be extracted with ethanol (120 °C, 50 bar, 15 minutes) but not with water at any of the temperatures tested (40–180 °C, 50 bar). The obtained extraction result for betulin is supported by theoretical solvation parameter calculations. Furthermore, high antioxidant activity of the extract was obtained using both ethanol and water as solvent. The antioxidant activity, as determined by a DPPH (2,2-diphenyl-1-picrylhydrazyl) assay, was found to be highest for the water extract of finely ground bark and it markedly increased with elevated extraction temperatures (90–180 °C). To elucidate if this was due to increased extraction efficiency or chemical reactions, a set of experiments was performed in which the samples were pre-treated with water at different temperatures before extraction. Results from liquid chromatography showed some differences in molecular composition between samples pre-treated at ambient and 180 °C, respectively. However, more detailed studies have to be performed to distinguish between hot-water extraction and reaction kinetics.
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