Towards the semi-synthesis of phosphorylated mimics of glycosaminoglycans: Screening of methods for the regioselective phosphorylation of chondroitin

Glycosaminoglycan (GAG) mimics carrying phosphate rather than sulfate anionic groups have been poorly investigated, in spite of their interesting perspectives. While some GAG-mimicking phosphorylated polymers have been reported, to the best of our knowledge no phosphorylated polysaccharides having the same backbone of natural sulfated GAGs have been accessed yet. To fill this gap, in this work two standard phosphorylation protocols and two recently reported procedures have been screened on a set of polysaccharide species composed by microbial sourced chondroitin and three partially protected, semi-synthetic derivatives thereof. A detailed structural characterization by 1H, 13C and 31P NMR spectroscopy revealed the higher versatility of the innovative, biomimetic reaction employing monopotassium salt of phosphoenolpyruvate (PEP-K) with respect to standard phosphorylating agents (phosphoric acid or phosphorus oxychloride). Indeed, PEP-K and H3PO4 gave similar results in the regioselective phosphorylation of the primary hydroxyls of unprotected chondroitin, while only the former reacted on partially protected chondroitin derivatives in a controlled, regioselective fashion, affording chondroitin phosphate (CP) polysaccharides with different derivatization patterns. The reported results represent the first, key steps towards the systematic semi-synthesis of phosphorylated GAGs as a new class of GAG mimics and to the evaluation of their biological activities in comparison with native sulfated GAGs.