2.2.4. In Vitro Release Studies

The TA release from WPI hydrogels was measured using a spectrophotometer (Multi-Mode Reader Synergy H1, BioTek, Winooski, VT, USA) at 48 h after incubation. A dried hydrogel sample was weighed accurately and then incubated in PBS at room temperature for up to 48 h. At the indicated time, a few drops of 0.5 N iron(III) chloride were added

to the selected aliquot, and the optical density of the solutions was measured at 586 nm (Figures S1 and S2) [24]. The tests were conducted on six independent replicates.

#### 2.2.5. Cell Viability Test

Cells were seeded in 96-well plates at the density described in the individual experiments. The following day, the excised hydrogel discs (diameter 3 mm) were added to triplicate wells. Fresh medium was added to each of 96 wells. Subsequently, the cells were incubated (Innova CO-170, New Brunswick Scientific, Enfield, CT, USA) at 37 ◦C for 48 h, together with the added materials. In the last step, 10 μL of AlamarBlue dye was added to each well and the intensity was measured using a spectrophotometer (Multi-Mode Reader Synergy H1). The experiment showed the capability of metabolically active cells to convert the AlamarBlue reagent into a fluorescent and colorimetric indicator. [25].

A commercially available laryngeal cancer cell line, Hep-2 (ATCC, CCL-23) was kindly provided by the center "Symbiosis" IBPPM RAS (Saratov, Russia).
