Thiolated chitosans are relatively new thiolated biopolymers exhibiting mucoadhesive, enzyme inhibitory, and permeation enhancing properties. A drawback, however, is their pH dependent reactivity. The aim of this study was therefore to develop a novel thiolated chitosan showing a non pH dependent reactivity of its thiol groups. For this purpose, 6-mercaptonicotinic acid (6-MNA) was covalently attached to chitosan by a carbodiimide mediated reaction. The obtained conjugate was characterized in vitro by quantification of immobilized thiol groups, cytotoxicity, in situ gelling properties, and disulfide bond formation at different pH. The synthesized conjugate displayed up to 973.80 micromol thiol groups per gram of polymer in reduced form. The polymer was nontoxic and showed in situ gelling properties. Furthermore, disulfide bond formation and therefore gelling properties occurred at various pH ranges. The reactivity of thiol groups was in the same range at pH 3 and pH 6.8. According to these results, chitosan-6 mercaptonicotinic acid seems to have some promising features to be used particularly for mucoadhesive formulations.