Sustainable drug release from polycaprolactone coated chitin-lignin gel fibrous scaffolds

Chronic wounds such as diabetic ulcers impose major health and economic burdens and often present hostile microenvironments that hinder regeneration and promote infection. Effective therapies require localized, controlled delivery of therapeutics to modulate concurrent healing processes and reduce systemic side effects. Electrospinning can produce ECM-mimicking fibrous scaffolds with tunable morphology and high surface area for drug loading and controlled release. Natural polymer hydrogels, particularly from chitin and lignin derived from abundant agricultural by-products, offer biocompatibility, biodegradability, antimicrobial and hemostatic properties and can form composite gels with pH/thermal responsiveness suitable for drug delivery. The authors previously electrospun ECM-like CL gel fibers and improved mechanical and antimicrobial performance by incorporating poly(glycerol sebacate) (PGS), but these scaffolds rapidly dissolved in aqueous media and exhibited burst release. To overcome this, the study investigates coaxial electrospinning to encapsulate CL/PGS hybrid fibers within a hydrophobic polycaprolactone (PCL) shell to prolong stability and achieve sustained drug release. The work evaluates morphology, chemistry, thermal and mechanical properties, drug release using methylene blue (MB), antibacterial performance with penicillin/streptomycin (PS), and cytocompatibility with BM-MSCs and NIH 3T3 cells.

Electrospinning enables production of nano/microfibers with controllable diameters by tuning polymer concentration, viscosity, voltage, feed rate, and humidity, supporting uniform dispersion of drugs and high loading capacity. Hydrogels are attractive for wound dressings due to moisture retention, barrier properties, and compatibility with aqueous processing that avoids drug denaturation. Natural polymers chitin (N-acetylglucosamine polysaccharide) and lignin (cross-linked phenolic biopolymer) are abundantly available yet underutilized, possessing moisturizing, anti-inflammatory, antimicrobial, and UV-absorbing characteristics. Chitin-lignin complexes can adsorb toxins and enable stimuli-responsive drug delivery. Coaxial electrospinning, which produces core-shell fibers, has been used to slow degradation and drug release (e.g., lawsone-loaded gelatin with PCL; glucosamine sulfate in PCL for cartilage). However, multicomponent gels with combinations of natural/synthetic polymers and drugs have been less explored in coaxial formats. PCL, a biocompatible hydrophobic polyester with moderate hydrophobicity and slow biodegradation, can act as a barrier to slow dissolution of CL gels, potentially mitigating burst release.

Materials and solution preparation: Chitin nanofibrils (2% water suspension), bio-lignin (CIMV, France), and PEOX were obtained from Nanoscience Centre (MAVI, Italy). Other chemicals were from Sigma-Aldrich. PGS was synthesized by polycondensation of glycerol and sebacic acid (1:1). The CL sol-gel solution was prepared by dispersing 30 wt.% chitin nanofibrils suspension, 0.1 wt.% bio-lignin, and 7 wt.% PEOX into 62.9 wt.% deionized water, adjusting pH to 10.5 with 0.1 M NaOH, and stirring for 48 h. PGS solution: 25% (w/v) PGS in ethanol (30 min stirring). The hybrid core solution was a 9:1 volume mixture of CL sol-gel and PGS solutions (30 min gentle stirring). PCL shell solution: 8% (w/v) PCL dissolved in chloroform:ethanol (9:1) for 3 h at room temperature.

Coaxial electrospinning: Conducted on a NANON-01A system under ~63% humidity. The outer (shell) PCL solution was delivered via system syringe pump through Teflon tubing to an ultra-thin coaxial spinneret at 0.5 mL/h. The inner (core) hybrid solution was delivered by an auxiliary syringe pump (KDS 100) through 20 cm Teflon tubing to a 27-gauge blunt needle at 0.2 mL/h. Voltage: 18 kV; tip-to-collector distance: 150 mm. Needle axially swayed over 8 cm at 10 cm/s; fibers collected on a stationary aluminum sheet. Electrospinning duration: 4 h; tip cleaned every 5 min. For single-component controls: hybrid fibers at 0.3 mL/h; PCL fibers at 0.9 mL/h. Process parameters (feed rates, solution concentrations, tube lengths) were optimized to ensure stable Taylor cone formation, appropriate solvent evaporation (chloroform vs water), and effective encapsulation without fiber fusion.

Characterization: Morphology by FESEM (JSM 7600F, JEOL); fiber size distributions quantified via custom MATLAB image processing with Gaussian fitting. Core-shell architecture by TEM (JEM-2100F, JEOL). Shell thickness estimated from 12,000× TEM images using ImageJ; 100 measurements across positions. Bulk composition by ATR-FTIR (Thermo Fisher, 400–4000 cm−1). Surface composition by XPS (SPECS XR-50, Mg Kα; binding energies referenced to C–C at 284.6 eV). Thermal behavior by DSC (MICRO DSC3 EVO, Setaram): heat to 180 °C and cool to 25 °C under N2 at 35 mL/min. Uniaxial tensile testing (Lloyd Instruments): strips 4 cm × 1 cm, extension speed 10 mm/min; thickness measured by caliper.

Biodegradation and drug release: 20 mg fibrous mats immersed in 5 mL PBS (pH 7.4) at 37.5 °C. Mass change recorded over time via microbalance (0.01 mg precision). UV–Vis spectra of supernatant by NanoDrop 2000. Visual color images recorded; MB release quantified using image processing of RGB channels in MATLAB with calibration: C_MB = 41.202(G − R)^0.95 + 319.05 (R^2 = 0.9766), reporting concentration in ng/mL. MB (2 mg/mL) was incorporated into the core solution pre-electrospinning for release studies.

Antibacterial testing: PS loading by adding penicillin (1000 U/mL) and streptomycin (1000 µg/mL) to the core solution prior to electrospinning. Agar diffusion assays against E. coli and S. aureus: overnight cultures in LB (ampicillin 100 µg/mL) spread on LB-agar with ampicillin (100 µg/mL). Triplicate disks of hybrid and core-shell mats with PS placed on inoculated plates and incubated overnight; non-loaded scaffolds as controls. Inhibition zone diameters recorded.

Biocompatibility: PS-loaded scaffold pieces (~2 mg) placed in 48-well plates and seeded with BM-MSCs (1×10^4 cells/well). MTT assay (0.5 mg/mL) used at 24, 48, 72 h; absorbance at 570 nm (ref 630 nm) measured (SpectraMax). NIH 3T3 viability assessed via PrestoBlue (10% v/v, 1 h incubation) with fluorescence read at 540/600 nm (Cytation 5). Statistical analysis by one-way ANOVA (OriginPro 8.0); p<0.05 and p<0.01 denoted by single and double asterisks.

• Process optimization: Stable coaxial electrospinning achieved at 18 kV with core feed 0.2 mL/h and shell feed 0.5 mL/h; reducing PCL to 8 wt.% facilitated stability.
• Fiber morphology and size: SEM showed smooth fibers. Mean diameters (mean ± SD): hybrid CL/PGS 176 ± 24 nm; PCL 936 ± 99 nm; core-shell 168 ± 22 nm, indicating the core governed overall fiber diameter due to higher solution conductivity (CL sol-gel ~7.8 mS vs PCL <0.04 µS).
• Core-shell structure: TEM revealed distinct core-shell architecture with thinner PCL shell than core. XPS carbon spectra indicated surface signals consistent with thin PCL shell; analysis depth (~10 nm) suggested shell thickness <10 nm, aligning with TEM measurements.
• Chemistry: FTIR of core-shell resembled hybrid fibers with observable PCL carbonyl peak ~1723 cm−1, indicating dominance of core composition in bulk.
• Mechanics and thermal behavior: PCL fibers were more flexible than hybrid. Core-shell scaffolds showed similar tensile behavior to hybrid, indicating minimal impact of thin PCL shell. DSC showed combined endothermic peaks corresponding to PEO (core) and PCL; two exothermic crystallization peaks on cooling, consistent with phase-separated crystallizations.
• Aqueous stability/degradation: In PBS (pH 7.4, 37.5 °C), CL-only fibers dissolved <15 min; adding PGS extended to ~2 h. Core-shell fibers exhibited a three-stage dissolution and lasted >24 h. UV absorbance (200–230 nm) increased with time, consistent with chitin/lignin dissolution aligning with core dissolution kinetics.
• Drug release (MB model): Hybrid and PCL controls showed initial burst releases. Core-shell release tracked the staged dissolution of the core with mitigated burst and prolonged release over 24 h, quantified by RGB-based imaging model.
• Antibacterial efficacy: PS-loaded hybrid and core-shell mats produced clear inhibition zones against E. coli and S. aureus, with core-shell exhibiting larger zones (stronger inhibition). For S. aureus, a double inhibition zone with a darker inner zone was observed, reflecting higher tolerance compared to E. coli.
• Cytocompatibility: BM-MSC adhesion and proliferation over 72 h were comparable between PS-loaded and unloaded scaffolds; no significant cytotoxicity detected. NIH 3T3 metabolic activity at 24 h was similar across scaffolds and controls, indicating good biocompatibility.

Encapsulation of CL/PGS hybrid fibers within a thin PCL shell via coaxial electrospinning effectively addressed rapid dissolution and burst release inherent to hydrophilic CL-based fibers. The high conductivity of the core solution dominated jet stretching, yielding core-shell fiber diameters similar to uncoated hybrids despite the thick PCL-only fibers, and resulting in a very thin shell (<10 nm) that preserved mechanical compliance and thermal transitions of the core. This shell acted as a diffusion barrier, producing a staged dissolution over >24 h and translating to sustained, non-burst drug release with MB. The enhanced and more sustained antibacterial zones for PS-loaded core-shell scaffolds demonstrate that controlled, localized antibiotic release can improve inhibitory performance against both Gram-negative (E. coli) and Gram-positive (S. aureus) strains, potentially maintaining therapeutic levels while reducing risks associated with systemic delivery. Biocompatibility with BM-MSCs and NIH 3T3 cells suggests suitability for wound contact applications. Overall, the findings support the hypothesis that a hydrophobic PCL shell can extend wet stability and modulate release kinetics of CL-based gel fibers without compromising desirable mechanical properties, making the platform promising for wound dressings requiring sustained therapeutic delivery.

Core-shell fibrous scaffolds comprising a CL/PGS hybrid gel core encapsulated by a thin PCL shell were fabricated by coaxial electrospinning. The PCL shell preserved core fiber morphology, did not significantly alter mechanical or thermal properties, and substantially delayed aqueous dissolution, extending scaffold stability beyond 24 h. Drug release was sustained with reduced burst, and antibiotics loaded into the core-shell fibers showed superior antibacterial activity against common skin pathogens without observable cytotoxicity. These results underscore the potential of PCL-coated CL gel fibers as controlled drug-releasing wound dressings and highlight the broader utility of natural polymer-based, coaxially engineered biomaterials derived from abundant agricultural by-products.

• Experiments were conducted in vitro; no in vivo wound healing or infection models were assessed, limiting direct clinical translatability.
• Drug release kinetics were evaluated over 24 h and with a model dye (methylene blue) and PS antibiotics; broader therapeutic classes and longer-term release profiles were not investigated.
• Antibacterial activity was measured by agar diffusion, which may not fully represent performance in complex wound environments.
• Shell thickness and structure were inferred from TEM and XPS with approximate depth limits; more precise 3D shell thickness mapping was not performed.