Crosslinking with GTA was carried out by placing samples of the scaffolds electrospun from S1, hereafter identified as M1, in a sealed glass container. An open recipient containing 10 ml of a GTA solution (5 wt.%) had been inserted inside this container in order to create an atmosphere with GTA vapor. The whole system was put in an oven at 40 °C from 0.5 to 24 h. After this, samples were placed in a desiccator for 3 days to remove any residual GTA.
The DHT crosslinking methodwas also applied to samples of M1 andwas accomplished in an oven under a vacuumof 2 mbar produced by a diaphragm pump. The temperature of the oven was chosen based on a preliminary test performed with different temperatures ranging from
100 °C to 160 °C with increments of 20 °C, and exposure times of 24 h, 48 h and 72 h. Well preserved fibers were observed for scaffoldstreated at a minimum of 140 °C. At 160 °C degradation of gelatin was noticed. Therefore, we chose 140 °C to proceed to the weight loss study for temperatures from 8 to 96 h.
Six days after being prepared, the gelatin in solution S2 had been partially crosslinked by the genipin present in the solution, but the viscosity was sufficiently low for electrospinning. After 6 days, the viscosity increases steeply and it becomes impossible to electrospin this solution. However, the scaffolds obtained from S2 aged for
6 days, which will be referred to as M2, were not stable when immersed in water and dissolved. Thus, it was necessary to further crosslink these scaffolds by immersing them into a bath of 2% genipin dissolved in a mixture of ethanol and water (90EtOH:10H2O in wt.%) for 1 to 5 days. Then they were placed in distilled water for 1 day and finally dried in a desiccator for 3 days.
Genipin crosslinking of gelatin scaffolds obtained from S1 was also attempted, by immersing the scaffolds in the same genipin solution.However, these scaffolds were not stable and dissolved, so this approach was abandoned.