Formation of water-resistant hyaluronic acid nanofibers by blowing-assisted electro-spinning and non-toxic post treatments

A unique blowing-assisted electro-spinning process has been demonstrated recently to fabricate hyaluronic acid (HA) nanofibers. In this article, effects of various experimental parameters, such as air-blowing rate, HA concentration, feeding rate of HA solution, applied electric field, and type of collector on the performance of blowing-assisted electro-spinning of HA solution were investigated. With the assistance of air-blowing, the solution-feeding rate could be increased to 40 μl/min/spinneret and the applied electric field could be decreased to 2.5 kV/cm. The optimum conditions for consistent fabrication of HA (with a molecular weight of ∼3.5 million) nanofibers involved the use of an air-blowing rate of around 70 ft³/h and a concentration range between 2.5 and 2.7% (w/v) in aqueous solution. Two benign methods to fabricate water-resistant HA nanofibrous membranes without the use of reactive chemical agents were demonstrated: (a) the exposure of HA membranes in hydrochloric acid (HCl) vapor, followed by a freezing treatment at -20 °C for 20-40 days; and (b) the immersion of HA membranes in an acidic mixture of ethanol/HCl/H₂O at 4 °C for 1-2 days. Although both methods could produce hydrophilic, substantially water-resistant HA nanofibrous membranes (the treated membranes could keep their shape intact in neutral water at 25 °C for about 1 week), the immersion method (6) was shown to be more versatile and effective. IR spectroscopy was used to investigate this 'cross-linking' mechanism in the solid HA membrane. Viscosity studies of acidic HA solutions under varying freezing conditions were also carried out. It was found that when the freezing time exceeded 8 h, the HA solution became gel-like and exhibited a large increase in the hydrogen-bond concentration. Thus, the resistance to water solubility could be due to the high density of hydrogen bonds in the solid HA membranes that were treated by the 'freezing' approach.