Thiol addition to hyaluronic acid oligomer non-reducing terminus - synthesis, analysis, stability, and unexpected...

The properties of hyaluronic acid (HA) can be enhanced by targeted modification. HA oligosaccharides can be further modified at the ends of their chains. However, methods to derivatize the non-reducing terminus are underdeveloped. In this work, various thiols were attached to the non-reducing terminus of unsaturated HA oligosaccharides (ΔHA), maintaining the linear character of HA-conjugates and their reducing terminus in the native form. Esters of ΔHA were successfully used as Michael thiol acceptors, while amides of ΔHA were unreactive. Based on the thiol structure, their addition provided mixtures of l-altro, d-galacto, and an unexpected d-gluco isomer. On the other hand, photo-induced radical (thiol-ene) addition to native ΔHA, its esters or amides, provided exclusively d-galacto products. This approach allowed the synthesis of sulphur-bridged oligosaccharides with defined sulphur position and a length range of 3 to 28 saccharides. No cytotoxic effect on 3T3, NHDF, HaCaT, and HT-29 cells was found with eight selected compounds. The sulphur bridge resisted cleavage with both hyaluronan hydrolases and lyases. These findings allow a synthesis of covalent conjugates of HA-oligosaccharides and active substances with native reducing end. Higher resilience against enzymatic cleavage and tunable affinity to HA cell-receptors can be another benefit for biomedical applications.