Chitosan with enhanced deprotonation for accelerated thermosensitive gelation with β-glycerophosphate

Chitosan (CS)/β-sodium glycerophosphate (β-GP) gels have low temperature sensitivity, which limits their biomedical application. The protonation and deprotonation of the CS amino groups are regulated by pH. It is possible to accelerate thermosensitive gelation by increasing the pH of the CS solution to maximize the degree of CS amino deprotonation and enhance hydrophobic interactions. In this study, a CS solution with a higher pH (HpHCS; pH 6.19, close to pKa 6.2) was obtained by means of the consumption of hydrogen ions using excessive chitosan. Raman spectroscopy showed that strong hydrogen bonds might exist in HpHCS. Gelation was accelerated significantly at 37 °C using HpHCS due to the decreased sol–gel transition temperature. Support vector machine and t-distributed stochastic neighbor embedding analyses confirmed that the gel derived from HpHCS was significantly different from that of the other groups. In addition, the HpHCS gel exhibited a low degradation rate and low cytotoxicity. Furthermore, a mathematical equation was established to predict the gelation time of HpHCS/β-GP at 37 °C. A better concentration of HpHCS for in vivo injection was obtained due to the shorter gelation time and lower viscosity. We found that human mesenchymal stem cells survived and proliferated in vitro in the HpHCS/β-GP hydrogel. Therefore, this study provides an optimized HpHCS solution with enhanced deprotonation for accelerated thermosensitive gelation with β-GP.