RESUMO
A mild and efficient reduction of negatively charged glucuronate units of hyaluronic acid (HA) into less polar glucose units has not been reported yet. However, this modification could significantly affect physical and chemical properties. Here we show a one-pot procedure where HA is converted into its derivate with carboxyl groups reduced to primary alcohols (HA-Red) without severe polymer degradation. Optimized synthesis aimed at aqueous solutions allowed the preparation of polysaccharides with molecular weights up to 1000 kDa. The chemical structure of HA-Red was proved by 2-dimensional NMR methodologies, FT-IR, LC-MS and SECMALLS. The final materials were exposed to a higher temperature or digested with bovine testicular hyaluronidase (BTH). Obtained data proved higher stability of HA-Red compared to HA, and significant dependence of stability on the degree of modification was observed in most cases. Preliminary in vitro studies showed no negative effects of HA-Red on the growth of 3T3 fibroblasts, which may be promising for applications requiring biodegradable and biocompatible HA derivatives with increased resistance to degradation.
Assuntos
Fibroblastos , Ácido Hialurônico , Animais , Bovinos , Espectroscopia de Infravermelho com Transformada de Fourier , Cromatografia Líquida , Glucose , HialuronoglucosaminidaseRESUMO
The effect of hydrazide linkers on the formation and mechanical properties of hyaluronan hydrogels was intensively evaluated. The reaction kinetics of hydrazone formation was monitored by NMR spectroscopy under physiological conditions where polyaldehyde hyaluronan (unsaturated: ΔHA-CHO, saturated: HA-CHO) was reacted with various hydrazides to form hydrogels. Linear (adipic, oxalic dihydrazide) and branched (N,N´,N´´-tris(hexanoylhydrazide-6-yl)phosphoric triamide and 4-arm-PEG hydrazide) hydrazides were compared as crosslinking agents. The mechanical properties of hydrogels were also modified by attaching a hydrophobic chain to HA-CHO; however, it was found that this modification did not lead to an increase in hydrogel stiffness. Cytotoxicity tests showed that all tested hydrazide crosslinkers reduced the viability of cells only slightly, and that the final hyaluronan hydrogels were non-toxic materials.
Assuntos
Reagentes de Ligações Cruzadas/química , Ácido Hialurônico/análogos & derivados , Hidrazinas/química , Hidrazonas/química , Hidrogéis/química , Acilação , Animais , Materiais Biocompatíveis/síntese química , Materiais Biocompatíveis/química , Materiais Biocompatíveis/toxicidade , Reagentes de Ligações Cruzadas/síntese química , Reagentes de Ligações Cruzadas/toxicidade , Módulo de Elasticidade , Ácido Hialurônico/síntese química , Ácido Hialurônico/toxicidade , Hidrazinas/síntese química , Hidrazinas/toxicidade , Hidrazonas/síntese química , Hidrazonas/toxicidade , Hidrogéis/síntese química , Hidrogéis/toxicidade , Concentração de Íons de Hidrogênio , Cinética , Camundongos , Células Swiss 3T3RESUMO
Hyaluronic acid (HA) containing CC double bond in positions 4 and 5 of N-acetyl-glucosamine ring (ΔHA) is an unique material, which could be used for biomedical applications and cosmetics. The main advantage of the CC double bond is its ability to react with a wide range of oxidation agents. Location of the CC double bond directly on the glucopyranose ring allows to change the chemical capabilities and simultaneously to mimic the intrinsic physical properties of HA without introduction of linkers or other substances. The synthesis, structural analysis and basic chemical and biological characteristics of this novel biopolymer are described in details. In vitro cytotoxicity assays showed selective activity of ΔHA against cancer cell lines in comparison with standard human fibroblasts so this material has great potential in the field of anticancer drugs.
