A green approach to obtain stable and hydrophilic cellulose-based electrospun nanofibrous substrates for sustained release of therapeutic molecules

Abstract

Stable and (bio)-compatible nanofibrous matrices showing effective incorporation and release of nonsteroidal anti-inflammatory drugs (NSAIDs) hold a huge potential in tissue regeneration and wound healing. Herein, a two-step, water-based and needleless electrospinning method is used to fabricate thermally cross-linked multifunctional nanofibrous substrates from a hydrophilic cellulose derivative, i.e. carboxymethyl cellulose (CMC), and polyethylene glycol (PEG) with an in situ incorporated NSAID, diclofenac (DCF). Electrospun bi-component blend nanofibers, strongly linked together by ester bonds, with different degrees of cross-linking density are achieved by varying the concentrations of butanetetracarboxylic acid (BTCA, a green polycarboxylic cross-linker) and the sodium hypophosphite (SHP) catalyst, and the temperature. The results demonstrated that not only the dimensional stability and swelling properties could be better controlled but also the morphology, fiber diameter, surface area, pore volume, pore size, and functionality of the cross-linked nanofibers. Release kinetics of DCF from the nanofibrous substrates are controlled and prolonged up to 48 h, and the overall released mass of DCF decreased linearly with increasing cross-linking degree of BTCA and SHP. Fitting of release data using various kinetic models revealed that the release of DCF follows a non-Fickian (diffusion and erosion controlled) to Fickian mechanism (only diffusion-controlled process). Cell viability testing based on crystal violet dyeing showed that the DCF-incorporating nanofibers have excellent biocompatibility and no toxic effect on human skin fibroblast cells. Overall, the reported DCF-incorporating nanofibrous substrate demonstrates high potential to be used as a smart drug delivery system in wound healing, especially due to its noninvasive characteristics.

Fig. 1. Conductivity (left) and viscosity (right) of electrospinning CMC/PEG solution added with different concentrations of BTCA.

Fig. 2. SEM morphology of non cross-linked and cross-linked electrospun nanofibers with different BTCA concentrations at 160 °C.

Fig. 3. Average fiber diameter (A) and diameter distributions (B–D) of electrospun nanofibers cross-linked with different BTCA concentrations at different temperatures.

Fig. 4. (A) ATR-FTIR absorption spectra of neat PEG, Na-CMC and BTCA, and non-heated CMC/PEG nanofibers with added BTCA_10%. (B) Spectrum of electrospun nanofibers cross-linked with BTCA_10% at different temperatures, (C) the absorption peak intensity ratio (1720 cm⁻¹/1594 cm⁻¹) of electrospun samples cross-linked with different BTCA concentrations and temperatures, (D) spectrum of pure DCF and DCF incorporated electrospun nanofibers cross-linked with different BTCA concentrations at 160 °C.

Fig. 5. Water absorption capacity (WAC) and photo images of electrospun samples cross-linked with different BTCA concentrations at 160 °C, after 24 h soaking in water.

Fig. 6. (A and B) QCM-D change in frequency and dissipation shifts of electrospun samples cross-linked with different BTCA concentrations at 160 °C upon exposure to water. Electrospun nanofibers of BTCA_0% on QCM-D crystals before (C) and after (D) rinsing with water, (E) BTCA_3%, (F) BTCA_5%, (G) BTCA_7% and (H) BTCA_10% cross-linked electrospun nanofibers at 160 °C, after interaction with water.

Fig. 7. (A) DCF mass as a function of time, (B) the % release of DCF as a function from electrospun samples cross-linked with different BTCA concentrations at 160 °C, (C) total amount of drug release from the non cross-linked and cross-linked (at 160 °C) electrospun nanofibers with different BTCA concentrations, (D) linear fitting of the overall drug release from the cross-linked (at 160 °C) nanofibrous substrates with BTCA as in Fig. 8B. (E) First derivatives of the release data from (A).

Fig. 8. Kinetics models fitted to the release data of DCF, incorporated electrospun samples cross-linked with different BTCA concentrations at 160 °C. (A) First-order, (B) Higuchi-model, (C) Korsmeyer-peppas model.

Fig. 9. Viability of human skin derived fibroblasts after exposure to the DCF incorporated electrospun nanofibrous substrates cross-linked with different concentrations of BTCA at 160 °C. The shown results were calculated relative to control (pure cell growth media). The dashed red lines correspond to the calculated confidence intervals for the control sample. Values are expressed as percentage of the means ± SD. Statistical significance was defined as *P < 0.05 compared to control sample (ANOVA test).