*2.3. Extraction*

The extraction process was carried out based on a DoE (Design of Experiments) developed using Modde software, version 11.0 (Sartorius Stedim Data Analytics AB, Umeå, Sweden). The D-optimal design type, which is based on the selection of the experiments so that they are spread over the largest area of the variability matrix, allowed the study of two quantitative factors which varied on different levels, namely the microwave power, with 4 levels of variation (300, 450, 600, 800 W) and the extraction time, varied over 3 levels (2, 3, 4 min). The amount of extracted total phenolic content (TPC) using different solvent types (water, ethanol–water 50:50 and 80:20) was introduced in the DoE as 3 separate responses, which allowed a good comparison of the solvents' TPC extraction capacities (Table 1).



Notes: mg GAE/g = gallic acid equivalents per gram plant material.

Bark was weighed (2 g) and mixed with 20 mL of solvent (water, 50:50 ethanol–water, 80:20 ethanol–water) in Falcon tubes. The Microwave extraction (MAE) was performed using a domestic microwave oven (Samsung MS23K3513). 7. After extraction, solvent was added to give a final volume of 20 mL. Each tube was centrifuged (Hettich, Micro 22R, Andreas Hettich GmbH & Co., Tuttlingen, Germany) 15 min at 3000 rpm, maintaining the extraction temperature. The supernatant was carefully separated, and the solvent was removed under vacuum at 40 ◦C using a rotary evaporator (Hei-VAP, Heidolph Instruments GmbH & Co., Schwabach, Germany).
