Nanodiamond-reinforced nanofiber scaffold system for skin regeneration

Abstract

A nanodiamond-reinforced nanofiber scaffold system for skin regeneration, consisting of a multilayered, electrospun nanofiber matrix formed from one or more biocompatible polymers selected from polycaprolactone (PCL), poly(lactic-co-glycolic acid) (PLGA), chitosan, gelatin or collagen, wherein nanodiamonds are present in an amount of 0.1 to 5 wt.-% are incorporated and surface-functionalized with oxygen, hydroxyl, or carboxyl groups to ensure homogeneous dispersion within the polymer matrix, the scaffold being characterized by a controlled fiber diameter in the range of 100 to 800 nm, an interconnected porosity between 60 and 90%, a tensile strength of 2 to 10 MPa, a degradation time of 2 to 8 weeks, and the ability to improve cell adhesion, proliferation, and migration of fibroblasts and keratinocytes, while simultaneously exhibiting antioxidant activity, reduced formation of reactive oxygen species, and the optional incorporation of bioactive agents selected from growth factors, antioxidants, or antimicrobial compounds to accelerate wound healing and skin tissue regeneration.

Description

[0001] The present invention relates to the field of biomedical engineering and regenerative medicine. In particular, it relates to the development of nanofiber scaffold systems reinforced with nanodiamonds, which are produced from multiple materials by electrospinning. The invention is aimed at applications in the field of skin tissue engineering and offers improved mechanical strength, biocompatibility, and increased potential for cell regeneration.

[0002] Skin injuries such as deep burns, chronic wounds, and traumatic lacerations often fail to heal effectively due to a lack of suitable biological substitutes capable of mimicking the complex structure and function of natural skin. Conventional dressings, grafts, and synthetic scaffolds have limitations such as low mechanical strength, insufficient cell adhesion, risk of infection, and limited bioactivity, which restrict their effectiveness in promoting complete tissue regeneration. This necessitates the development of advanced biomaterials that can support cell growth, angiogenesis, and functional restoration.

[0003] Electrospinning technology has proven to be a powerful technique for producing nanofiber scaffolds that closely resemble the extracellular matrix (ECM). These nanofibers offer a high surface-to-volume ratio, adjustable porosity, and structural similarity to natural tissues, enhancing cell proliferation, nutrient diffusion, and tissue integration. However, single-material electrospun scaffolds often lack the necessary multifunctionality, such as sufficient mechanical stability, controlled biodegradability, and bioactive reinforcement, limiting their applicability in long-term skin tissue engineering.

[0004] Nanodiamonds (NDs) have recently attracted considerable attention as promising nanofillers due to their superior mechanical strength, excellent biocompatibility, potential for surface functionalization, and ability to enhance antioxidant and antibacterial properties. The incorporation of nanodiamonds into multilayer electrospun scaffolds offers an innovative approach to overcoming the limitations of conventional systems. Such reinforced scaffolds can synergistically combine structural support, biological activity, and durability, making them ideally suited for next-generation skin tissue engineering applications.

[0005] One objective of the present disclosure is to provide a nanofiber scaffold system reinforced with nanodiamonds that mimics the natural extracellular matrix for effective skin regeneration.

[0006] Another objective of the present disclosure is to ensure improved mechanical strength and structural stability of scaffolds suitable for long-term support of skin tissue.

[0007] Another objective of the present disclosure is to offer a biocompatible and biodegradable scaffold material that avoids harmful side effects.

[0008] Another objective of the present disclosure is to improve cell adhesion, proliferation and migration through optimized fiber morphology and surface properties.

[0009] Another objective of the present disclosure is to incorporate nanodiamonds as functional nanofillers to improve the durability and bioactivity of the scaffold.

[0010] Another objective of the present disclosure is to provide a multi-layered scaffold architecture that is very similar to the skin and epidermal layers of natural skin.

[0011] Another objective of the present disclosure is to facilitate the controlled incorporation of bioactive molecules for sustained therapeutic effects.

[0012] Another objective of the present disclosure is to maintain optimal porosity and moisture balance for accelerated wound closure and wound healing.

[0013] Another objective of the present disclosure is to support vascularization and tissue integration during the regeneration process.

[0014] Another objective of the present disclosure is to enable scalable manufacturing by electrospinning with multiple materials for clinical and commercial implementation.

[0015] The present invention relates generally to a nanofiber scaffold system reinforced with nanodiamonds, designed for skin regeneration, wherein the scaffold mimics the extracellular matrix and provides structural support for cell growth. It offers improved mechanical, biological, and architectural properties essential for tissue repair.

[0016] One embodiment of the present invention relates to the incorporation of nanodiamonds as nanofillers into polymeric nanofibers to improve the durability, bioactivity, and stability of the scaffold while maintaining biocompatibility. This ensures suitability for biomedical applications.

[0017] Another embodiment of the invention provides for a multilayer scaffold architecture fabricated using multimaterial electrospinning. This allows for differentiation between dermal and epidermal layers for more physiologically relevant tissue engineering.

[0018] Another embodiment of the invention aims to optimize the porosity and surface morphology of the scaffold. Such features support effective cell adhesion, proliferation, nutrient diffusion, and vascularization during skin tissue regeneration.

[0019] Another embodiment of the invention is the integration of bioactive molecules or therapeutic agents into the nanofiber matrix. This enables a controlled release of molecules to support wound healing, antimicrobial activity, and tissue regeneration.

[0020] Another embodiment of the invention is the maintenance of an ideal balance between biodegradability and mechanical strength. This ensures the integrity of the scaffold during the healing process while simultaneously allowing for gradual resorption.

[0021] Another embodiment of the invention focuses on improving cytocompatibility and reducing cytotoxic effects through careful selection of polymer mixtures and reinforcing agents. This enables safe use in clinical wound healing settings.

[0022] Another embodiment of the invention aims at the scalable and reproducible fabrication of scaffolds using advanced electrospinning techniques. This enables large-scale production and clinical implementation of the system for skin tissue engineering.

[0023] The present invention relates to a nanodiamond-reinforced nanofiber scaffold system for skin regeneration, developed using a multi-material electrospinning approach to accurately mimic the extracellular matrix. The system integrates nanodiamonds into polymer nanofibers to enhance mechanical strength and bioactivity, while providing a multilayered architecture with controlled porosity to support cell adhesion and growth. Designed with biocompatibility in mind, the invention enables sustained therapeutic support and effective tissue regeneration, making it ideally suited for wound healing and skin tissue engineering applications. EXAMPLE 1: Production of polymer solutions

[0024] Polyvinyl alcohol (PVA) and polyacrylonitrile (PAN) were selected as the base polymers for the scaffold due to their proven biocompatibility, stability, and suitability for electrospinning. A 10% w / v PVA solution was prepared by dissolving 1.5 g of PVA in distilled water at 80 °C with continuous stirring until a clear, homogeneous solution was obtained. Similarly, a 10% PAN solution was prepared by dissolving 1.5 g of PAN in N,N-dimethylformamide (DMF) at 50 °C with continuous stirring for 12 hours. Nanodiamonds (NDs) were incorporated into each polymer solution at predetermined concentrations (1% w / v) and uniformly dispersed using probe ultrasound for 30 minutes to prevent agglomeration.

[0025] Therapeutic agents were also incorporated into the respective polymer matrices. Ceftriaxone sodium (CS), a broad-spectrum antibiotic, was dissolved in deionized water, while sinapic acid (SA), a natural antioxidant with anti-inflammatory properties, was dissolved in DMF. Each drug solution was then combined with the respective polymer-ND solution to achieve the desired concentration, maintaining consistent polymer ratios across all formulations. This ensured reproducibility and uniformity in scaffold fabrication. EXAMPLE 2: Production of multilayer nanofiber scaffolds

[0026] The multilayer scaffolds were fabricated using a high-precision electrospinning system. The polymer-drug solutions were filled into disposable syringes and attached to syringe pumps to control the flow rates (350–500 µL / h). Electrospinning was performed under ambient conditions (25 ± 2 °C, 45% relative humidity) with an applied voltage of 13.5–18 kV and a fixed tip-to-collector distance of 18 cm. At approximately 15 kV, a stable Taylor cone was achieved, enabling the formation of uniform nanofibers that deposited onto an aluminum foil-covered collector plate.

[0027] After electrospinning, the scaffolds were carefully removed and crosslinked for 96 hours at room temperature using a 0.5% w / v ethanol glutaraldehyde (GA) solution. This step improved the mechanical stability, water resistance, and durability of the fibrous architecture, which is crucial for tissue engineering applications. Layered scaffold design

[0028] A layer-by-layer (LbL) electrospinning strategy was used to construct multifunctional scaffolds, with each layer tailored to a specific therapeutic function: • Inner contact layer (PAN + SA + NDs): This layer was developed for the interface with the wound bed and contains sinapic acid and nanodiamonds to achieve a strong antioxidant and anti-inflammatory effect while improving scaffold bioactivity and cell adhesion. • Middle adhesive layer (PVA + CS + NDs): This layer contains ceftriaxone sodium and nanodiamonds to provide long-lasting antibacterial protection and controlled release, thereby preventing infection and promoting wound healing. • Outer protective layer (PAN + NDs): This layer serves as a barrier against external contaminants and mechanical stresses while ensuring structural integrity. EXAMPLE 3: Scaffolding composition

[0029] Various scaffold compositions were developed to investigate the influence of nanodiamonds and therapeutic agents (Table 1). Formulation Code Description Ceftriaxone sodium (CS) Sinapic acid (SA) Nanodiamonds (NDs) S1 Pure dummy frame (PVA + PAN only) 0.00 g 0.00 g 0% S2 ND-reinforced dummy frame 0.00 g 0.00 g 1% S3 CS-loaded scaffold (PVA matrix) 0.40 g 0.00 g 1% S4 SA-loaded scaffold (PAN matrix) 0.00 g 0.40 g 1% S5 Scaffold loaded with two active ingredients (PVA + PAN, CS + SA + NDs) 0.20 g 0.20 g 1%

[0030] This innovative multi-layered scaffold system, reinforced with nanodiamonds and containing two therapeutic agents, addresses the main challenges in skin tissue engineering by combining mechanical robustness, antimicrobial protection, antioxidant activity, and biomimetic nanofiber architecture. Example 4: Swelling study

[0031] In another example, the swelling behavior of the fabricated multilayer nanofiber scaffolds was investigated. Square patches (1 × 1 cm) were cut from each formulation and immersed in phosphate-buffered saline (PBS, pH 7.4, 37 °C). The swelling weight (Ws) was measured at specific intervals after excess surface fluid had been carefully removed with filter paper. The swelling ratio was calculated using the following formula: Swelling ratio=Ws−Wd Wd×100 where Wd represents the dry weight of the scaffold. All experiments were performed three times and the mean values ​​are reported to provide insight into the hydration capacity of the scaffold.

[0032] The swelling study revealed significant differences in the hydration behavior of the nanofiber scaffolds. Formulation S4 exhibited the highest swelling rate, peaking after 60 minutes, which is related to its high porosity (50.15%) and the partial hydrophilic interactions of sinapic acid within PAN. S5 showed moderate swelling, as its sandwich structure allowed water uptake through the inner PVA-CS layer but limited excessive hydration through the outer PAN-SA layers. S3 showed the least swelling due to its denser PVA-CS matrix, while S1, the drug-free control, exhibited minimal water uptake. These results suggest that drug incorporation and porosity have a strong influence on fluid uptake.Importantly, S5 maintained an optimal balance between hydration and stability, making it a suitable candidate for wound dressing applications. Example 5: Porosity measurement

[0033] In another example, the porosity of the nanofiber scaffolds was determined using the ethanol displacement method. The samples were immersed in absolute ethanol, and the porosity (P%) was calculated using the following relationship: P=W2−W1ρ×V×100P=ρ×VW2−W1×100 where W1 is the dry weight of the scaffold, W2 the weight saturated with ethanol, ρ the ethanol density, and V the volume of the scaffold. This method enabled a quantitative assessment of the scaffold's porosity, which is crucial for nutrient diffusion and tissue integration.

[0034] S4 exhibited the highest porosity (50.15%), indicating a loosely packed fiber network that promotes solvent penetration and extended drug diffusion. S2 and S5 also showed relatively high porosity values ​​of 43.46% and 42.98%, respectively, suggesting open fiber structures with the potential for controlled release. In contrast, S3 exhibited the lowest porosity (32.09%), which is related to its densely packed PVA-CS matrix that restricts solvent uptake and leads to faster drug availability at the surface. The correlation between porosity and drug release shows that higher porosity favors sustained release, while denser matrices support rapid release. This structural diversity allows for the tailoring of drug release kinetics to specific therapeutic requirements. Example 6: In vitro study on drug release

[0035] In another example, the drug release profile of the fabricated scaffolds was investigated using the dialysis bag method in PBS (pH 7.4, 37 °C, 50 rpm). Aliquots were withdrawn at predetermined time intervals and replaced with fresh buffer solution. Drug concentrations were measured by UV-Vis spectrophotometry at 236 nm for sinapic acid (SA) and at 241 nm for ceftriaxone sodium (CS). The cumulative release was calculated using the following formula: Drug release (%) = MtMα × 100 where Mt is the amount of drug released at time t and Mα is the total drug content. The release data were further fitted to zero-order, first-order, Higuchi, and Korsmeyer-Peppas models to determine the release mechanism.

[0036] Drug release studies revealed different profiles depending on the drug type and scaffold structure. S3 (PVA-CS) showed rapid release, with more than 80% of the ceftriaxone being released within 30 minutes and complete release within 120 minutes due to the hydrophilicity of PVA. S4 (PAN-SA) showed more sustained release, with 70% of the sinapic acid being released within 30 minutes and release gradually completing after 3 hours, reflecting the hydrophobicity of PAN and the stronger interactions between the polymer and the drug. The dual-drug scaffold S5 showed the longest release, with only about 40% of the ceftriaxone being released within 30 minutes and controlled diffusion continuing for up to 4 hours. This layered design formed a diffusion barrier through the outer PAN layers and regulated the release of both drugs.These results confirm that the scaffold design strongly influences the release kinetics, with S5 exhibiting superior sustained therapeutic drug delivery. Example 7: Antibacterial effect

[0037] In one example, the antibacterial efficacy of scaffolds was tested against Pseudomonas aeruginosa and Klebsiella pneumoniae using the agar disc diffusion method. Circular scaffold discs (6 mm diameter) were placed on agar plates inoculated with the bacterial strains and incubated for 24 hours at 37 °C. Antibacterial activity was quantified by measuring the diameter of the zone of inhibition (ZoI) in millimeters (mean ± SD, n = 3). Pure ceftriaxone sodium discs served as positive controls.

[0038] The antibacterial activity of nanofiber formulations (SI-S5), nanodiamonds (NDs), and pure ceftriaxone sodium was evaluated against Pseudomonas aeruginosa and Klebsiella pneumoniae. Of all the formulations tested, the dual-action S5 formulation showed the strongest activity, producing zones of inhibition of 24.0 ± 1.0 mm and 22.7 ± 0.6 mm, which were significantly larger than those achieved with pure ceftriaxone sodium (19.0 ± 1.0 mm and 16.0 ± 1.0 mm), representing improvements of 26% and 42%, respectively. This synergistic enhancement between ceftriaxone sodium and sinapic acid was evident compared to S3 (21.3 ± 0.9 mm and 19.0 ± 1.0 mm), which contained only ceftriaxone sodium, and showed an almost 13% and 20% higher efficacy of S5, respectively. Interestingly, S3 still outperformed the pure drug by 12% and 19%, respectively, suggesting that the PAN / PVA / ND scaffold improved the stability and release of the drug.In contrast, S1 and S2 showed only moderate inhibition, while S4, containing only sinapic acid, had the weakest effect, consistent with the limited antibacterial efficacy of phenolic acids. NDs alone showed no activity against P. aeruginosa but demonstrated activity (14.7 ± 1.5 mm) against K. pneumoniae, confirming previous findings on ND-induced membrane disruption and ROS generation. Overall, the results underscore that the PAN / PVA / ND scaffold is a versatile platform for drug delivery, and the co-loading of ceftriaxone sodium with sinapic acid offers superior antibacterial efficacy, making S5 a promising multifunctional nanofiber dressing for combating resistant infections. Example 8: DPPH radical scavenger assay

[0039] In another example, the antioxidant activity of scaffold formulations was determined using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. The scaffold extracts were diluted to 0.5 mM in dimethyl sulfoxide (DMSO). A freshly prepared 0.3 mM DPPH solution was mixed with the test samples in a 96-well plate and incubated for 30 minutes at 37 °C in the dark. The absorbance was measured at 517 nm using a microplate reader. The radical scavenging activity was calculated relative to the DMSO control, and the IC50 was determined. 50 Values ​​were determined. Standard antioxidants such as gallic acid, N-acetylcysteine, and ascorbic acid were used for comparison.

[0040] The antioxidant activity of the nanofiber formulations was evaluated using the DPPH radical scavenger assay, and the results showed significant differences depending on the active ingredients. Pure sinapic acid (SA) exhibited strong radical scavenging potential with an inhibition of 93.2% at 0.5 mg / ml and an IC50 of 1000. 50 -value of 22.2 ± 0.6 µg / ml, which is comparable to the standard gallic acid (95.3% inhibition; IC50). 50 = 3.69 µg / ml). When incorporated into nanofibers, SA in formulation S4 retained high activity (89.4% inhibition, IC10). 50 = 103 ± 1.14 µg / ml), however, with reduced efficacy due to encapsulation in the polymer matrix. In contrast, the dual-action formulation S5 showed moderate activity (70.2% inhibition, IC50). 50= 293.9 ± 3.17 µg / ml), which was lower than that of S4, probably because the coexistence of ceftriaxone sodium impaired the radical scavenging activity of SA. The pure ceftriaxone formulation S3 and nanodiamonds (NDs) showed only negligible antioxidant activity with negative inhibition values ​​(-10.6% and -16.1%, respectively), confirming their lack of intrinsic radical scavenging property. After adjusting for dilution factors, the effective IC50 values ​​increased. 50 -values, with S4 shifted to 447 µg / ml and S5 to 351 µg / ml, while S3 showed a weak but detectable effect with an IC 50 showed a level of 53 µg / ml. Example 9: Cytotoxicity activity (MTT method)

[0041] In one example, the cytotoxicity of the fabricated scaffolds was evaluated using the MTT assay on mouse fibroblast 3T3 cells. The cells were cultured in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 5% fetal bovine serum, penicillin, and streptomycin under standard conditions (37 °C, 5% CO2). Cell suspensions (5 × 10 4 Cells (µL / ml) were seeded into 96-well plates and exposed to different concentrations of the scaffold extract. After 48 hours of incubation, MTT solution was added, followed by a further 4-hour incubation. The resulting formazan crystals were dissolved in DMSO, and the absorbance was measured at 540 nm. Cell viability and the percentage of growth inhibition were calculated in comparison to untreated controls.

[0042] The cytocompatibility of the fabricated nanofiber scaffolds was evaluated using the MTT assay in 3T3 fibroblasts at 30 µg / ml, and the results showed varying responses depending on the formulation. Doxorubicin, used as a positive cytotoxic control, exhibited the highest inhibition of cell viability (93.68%), thus validating the assay. Among the scaffolds, the ceftriaxone-loaded formulation (S3) showed the strongest cytotoxic effect with an inhibition of 81.8%, indicating that high concentrations of ceftriaxone significantly impair fibroblast survival. The dual-agent scaffold (S5) showed moderate inhibition (62.95%), suggesting that the presence of sinapic acid partially attenuated the cytotoxic effect of ceftriaxone while maintaining its antibacterial potential.In contrast, the sinapic acid-loaded scaffold (S4) and pure sinapic acid (SA) showed only moderate cytotoxicity with inhibition of 22.05% and 28.26%, respectively, supporting the protective role of the phenol compound. Nanodiamonds (NDs) alone showed minimal toxicity with only 5.45% inhibition, confirming their biocompatibility as a structural nanofiller.

[0043] Taken together, these results suggest that while S3 is highly effective, it is cytotoxic, whereas the dual-drug scaffold S5 has a more balanced profile in terms of antimicrobial efficacy and fibroblast tolerance, making it a promising candidate for wound healing applications where both infection control and tissue regeneration are critical. Example 10: In-vivo wound healing experiment

[0044] In another example, the wound-healing efficacy of the scaffolds was evaluated in vivo using an excision wound model in rabbits. Healthy rabbits were divided into four groups (n = 4 each): an untreated control group, Quench® cream (commercially available ointment), an empty nanofiber scaffold, and a nanodiamond-reinforced, two-agent-loaded multilayer scaffold (S5). Under anesthesia, a full-thickness skin wound (1.5 cm) was created. 2 ) generated and the respective treatments applied. Wound contraction was measured over 21 days using a metric scale. The percentage of wound closure was calculated as follows: Wound closure (%) = A∘−At A∘×100 where A° is the initial wound area and At is the wound area at time t.

[0045] The in vivo wound healing study demonstrated accelerated closure in the nanofiber scaffold groups compared to the Quench cream-treated or untreated controls. Over a 21-day period, the drug-loaded scaffolds (S5) achieved the fastest wound closure, closely followed by the empty nanofibers, highlighting the intrinsic wound-healing capacity of PAN. By day 14, the nanofiber-treated wounds were almost completely closed, while the untreated wounds showed slower healing. Drug-loaded scaffolds exhibited enhanced antibacterial and anti-inflammatory effects, further improving the healing outcomes. Notably, despite a significantly lower sinapic acid content compared to the pure SA control, S5 achieved comparable or even better wound closure, underscoring its dose-saving and synergistic effect.These results suggest that multilayer nanofibers represent a powerful platform for wound healing. Example 11: Histological analysis

[0046] In one example, a histological examination of wound tissue taken on day 21 was performed. The tissue was fixed in formalin, embedded in paraffin, sectioned, and stained with hematoxylin and eosin (H&E). Microscopic analysis was conducted to assess re-epithelialization, collagen deposition, neovascularization, and inflammatory cell infiltration. These results confirmed the regenerative potential of the nanodiamond-reinforced, dual-agent-loaded scaffolds compared to the controls.

[0047] Histopathological evaluation confirmed the biocompatibility and therapeutic efficacy of the S5 scaffold in skin tissue. Microscopic analysis revealed mild epidermal hyperplasia and a slight accumulation of mononuclear cells around the hair follicles, indicating an active but controlled healing response. No signs of necrosis, fibrosis, or severe inflammatory reactions were observed, confirming the scaffold's safety for tissue integration. The observed cell invasion suggested enhanced regenerative activity in the wound area. Overall, S5 demonstrated a favorable interaction with biological tissues, supporting its use as a safe and effective wound healing material.

Claims

[1] A nanodiamond-reinforced nanofiber scaffold system for skin regeneration, comprising a multilayered, electrospun nanofiber matrix formed from one or more biocompatible polymers selected from polycaprolactone (PCL), poly(lactic-co-glycolic acid) (PLGA), chitosan, gelatin or collagen, wherein nanodiamonds are present in an amount of 0.1 to 5 wt.-% are incorporated and surface-functionalized with oxygen, hydroxyl, or carboxyl groups to ensure homogeneous dispersion within the polymer matrix, the scaffold being characterized by a controlled fiber diameter in the range of 100 to 800 nm, an interconnected porosity between 60 and 90%, a tensile strength of 2 to 10 MPa, a degradation time of 2 to 8 weeks, and the ability to improve cell adhesion, proliferation, and migration of fibroblasts and keratinocytes, while simultaneously exhibiting antioxidant activity, reduced formation of reactive oxygen species, and the optional incorporation of bioactive agents selected from growth factors, antioxidants, or antimicrobial compounds to accelerate wound healing and skin tissue regeneration. [2] The nanodiamond-reinforced nanofiber scaffold system according to claim 1, wherein the scaffold comprises at least two or more different electrospun layers arranged to mimic the dermal and epidermal structure of natural skin. [3] The nanodiamond-reinforced nanofiber scaffold system according to claim 1, wherein the nanodiamonds are surface functionalized by acid treatment to introduce carboxyl groups and thereby improve the polymer-nanodiamond bond. [4] The nanodiamond-reinforced nanofiber scaffold system according to claim 1, wherein the porosity of the scaffold allows permeability for oxygen and nutrients to promote the growth of skin cells. [5] The nanodiamond-reinforced nanofiber scaffold system according to claim 1, wherein the scaffold has a swelling capacity between 100% and 400% to maintain a moist wound environment. [6] The nanodiamond-reinforced nanofiber scaffold system according to claim 1, wherein the scaffold supports the migration of vascular endothelial cells to promote angiogenesis during skin regeneration. [7] The nanodiamond-reinforced nanofiber scaffold system according to claim 1, wherein the scaffold is produced using a multi-material electrospinning technique to form gradient structures with different fiber orientations and polymer compositions. [8] The nanodiamond-reinforced nanofiber scaffold system according to claim 1, wherein the bioactive agents included comprise epidermal growth factor (EGF), vascular endothelial growth factor (VEGF) or curcumin to accelerate wound healing. [9] The nanodiamond-reinforced nanofiber scaffold system according to claim 1, wherein the scaffold exhibits antimicrobial activity against gram-positive and gram-negative bacteria. [10] The nanodiamond-reinforced nanofiber scaffold system according to claim 1, wherein the scaffold exhibits an antioxidant potential of at least 60% radical scavenging effect in vitro compared to non-reinforced scaffolds.