INTRODUCTION
Edible Bird’s Nest (EBN) is a secretion produced by swiftlets that has been known and used as a functional food and natural medicine 1,2. EBN has gained popularity for making various dietary supplements such as capsules, drinks and snacks 3,4. EBN produced from hardened swiftlet saliva is rich in nutrients such as protein, essential amino acids, and various minerals, making it a potential material for the development of health and wellness products 5,6,7. EBN has interesting potential to be used as a matrix in the form of nanofibers, especially because of its rich nutritional content, biocompatibility, bioactive properties, and multifunctional applications 5,7,8,9.
Electrospun nanofibers from pure EBN may not be possible due to the glycoprotein content, which provides bioactive and nutritional properties. These glycoproteins often have low solubility in common solvents, making it difficult for the electrospinning process to produce nanofibers from pure EBN 10. To overcome the solubility problem, EBN often needs to be mixed with other polymers. One of the polymers often used in composites with bioactive materials is polyvinylpyrrolidone (PVP) 11. PVP is a biocompatible and water soluble polymer commonly used in electrospinning 12,13. The use of PVP as a carrier material for EBN can increase the solubility of EBN in solvents and promote the formation of uniform and stable nanofibers during electrospinning, thus producing nanofibers with higher EBN content.
Nanofibers can be synthesized by several methods, such as self-assembly 14, phase separation 15, lyophilization 16 and electrospinning 17. However, the electrospinning technique is the most popular method for fabricating nanofibers due to its ability to produce fibers with very small diameters and good control over morphology 18.
Electrospinning is a versatile technique to produce nanofibers with high surface-to-volume ratios, small diameters, and excellent morphology. These unique properties make electrospun nanofibers useful in various fields such as biomedical engineering 17,19, energy storage 20, and food packaging 21,22.
Therefore, this study aims to investigate the optimization of electrospinning parameters for producing EBN/PVP nanofibers. Some parameters are studied, including the EBN to PVP weight ratio, electrospinning voltage, and collector distance on the diameter and morphology of the obtained fibers. Some physical characterizations were also conducted for the nanofibers, including scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy, and thermal analysis.
METHODS
Materials for Precursor Solution
EBN was obtained from a local market in Indonesia. Meanwhile, polyvinylpyrrolidone (PVP) with an average molecular weight of ~360,000 g/mol and absolute ethanol were obtained from Sigma-Aldrich Chemicals. The precursor solution was prepared by dissolving the PVP in ethanol to form a homogeneous solution with polymer concentration of 10 wt%. Then, EBN powder was added to the PVP solution at various EBN:PVP weight ratios of 10:90, 20:80, 30:70, 40:60, 50:50, 60:40, 70:30, 80:20, 90:10, and 100:0.
Electrospinning Process
Nanofiber were produced using an electrospinning machine (ILMI-N101 Electrospinning, Indonesia), as shown in Figure 1. The precursor solution was dispensed using a syringe pump at a constant flow rate of 0.8 mL h-1. A high voltage power supply was connected to the syringe needle which will draw the solution jet toward a grounded plate collector, forming fine nanofibers. The fabrication process was conducted at a constant temperature of 26 °C and relative humidity of 55%. Some parameters were varied for the electrospinning process, including the EBN ratio in the precursor solution, working voltage, and collector distance.
Nanofiber's Characterization
The morphology and diameter of the nanofibers were analyzed using Scanning Electron Microscopy (SEM, SU3500). The diameter distribution was determined from SEM images using ImageJ software (National Institutes of Health, Bethesda, MD, USA) 23. The functional groups of EBN/PVP nanofiber samples was investigated using Fourier Transform Infrared Spectroscopy (FTIR, Bruker, Alpha Platinum ATR A220/D-01) in wave number range of 500 to 4500 cm-1. Thermal characterization was also performed to determine the thermal stability of the nanofiber samples using thermogravimetric and differential thermal analysis (TG/DTA, STA7300, Hitachi). The TG/DTA measurements were performed using the temperature range of 30 °C to 600 °C, and heating rate of 10 °C/min. The statistical analysis was also performed to the samples using Tukey’s one-way ANOVA with p < 0.05 24. Minitab software version 17 (Minitab, LLC, State College, PA, USA) was used for this statistical analysis.
RESULTS AND DISCUSSION
Spinnability of Precursor Solution with Various EBN/PVP Ratio
To evaluate the spinnability of the precursor solutions, various weight ratios of EBN to PVP were prepared. The precursor solutions were prepared with various EBN:PVP weight ratios of 10:90, 20:80, 30:70, 40:60, 50:50, 60:40, 70:30, 80:20, 90:10, 100:0. Each solution was processed at a constant voltage of 15 kV, a fixed collector distance of 20 cm, and a constant flow rate of 0.8 mL h-1. The results indicate that smooth, beads free nanofibers were successfully produced from solutions with EBN:PVP ratio of 10:90, to 60:40. The smooth morphology of nanofibers can be obtained by careful selection of the EBN/PVP ratio. It is likely that the ratio between 10:90 and 60:40 provide a favorable balance of the solution conductivity and viscosity for successful electrospinning, resulting in the formation of uniform, defect-free nanofibers.
In contrast, nanofibers were not formed for the EBN:PVP ratio of 70:30, 80:20, 90:10, 100:0. This suggests that EBN alone lacks sufficient electrospinnability and must be blended with polymeric additives such as PVP. The inability to form fibers at higher EBN content may be attributed to excessive solution viscosity, which impedes the electrospinning process 10. Since maximizing the functional benefits of EBN is essential, nanofiber samples with the highest EBN content were selected for further optimization of electrospinning process parameters. Accordingly, the EBN:PVP ratio of 60:40 was chosen as the precursor formulation for subsequent optimization studies.
Effect of Process Parameters on the Nanofibers Diameter
The diameter of nanofibers produced by electrospinning is highly dependent on the electrospinning voltage. Figure 2 shows the morphology and diameter distribution of EBN/PVP nanofibers (EBN:PVP ratio of 60:40) obtained from various electrospinning voltages at constant solution flow rate of 0.8 mL h-1, and a needle-to-collector distance of 20 cm. Generally, as the voltage increases, the average diameter of the nanofiber decreases as indicated by Figure 3. Statistical analysis using Tukey’s test, represented by different alphabetical groupings, further confirms the significant differences in fiber diameters obtained at varying electrospinning voltages. These results are consistent with previous studies on electrospinning of biopolymers at different voltages 25,26. The decrease of nanofiber diameter with increasing voltage can be attributed to the increase of electric force acting on the polymer solution during electrospinning 27,28,29. This force causes the solution jet to stretch and form whipping mode before reaching collector 30. Stronger electrostatic forces cause greater stretching on the polymer jet, resulting in a more reduction in nanofiber diameter 31,32.
These results indicate that proper control of the electrospinning voltage is essential to produce EBN/PVP nanofibers with the desired diameter. While higher voltage may result in smaller diameter nanofibers, it may also increase the possibility of beads formation and fiber discontinuity. Therefore, the optimal electrospinning voltage should be carefully selected based on the desired nanofiber morphology and diameter.
In addition to the voltage, the distance between the needle and the collector affects the size of the EBN/PVP nanofiber produced during the electrospinning process. Figure 4 shows the effect of varying distance of the collector on the diameter distribution of the EBN/PVP nanofiber. The experiment used fixed voltage of 15 kV and solution flow rate of 0.8 mL h-1 for EBN:PVP ratio of 60:40. Figure 5 shows that the larger the distance between the needle and collector during electrospinning, the smaller the average diameter of the nanofiber. This is in agreement with research conducted by Z AL-Hazeem, (2020), which shows that by increasing the distance between the needle and the collector, a smaller fiber diameter and a more uniform size distribution can be obtained 33.
A shorter needle-to-collector distance can lead to larger fiber diameters, as the jet has less time to undergo full elongation during the stretching process 33. The jet may reach the collector before sufficient stretching occurs, resulting in thicker fibers. Additionally, the reduced distance may prevent complete solvent evaporation, causing the fibers to remain wet upon deposition and potentially leading to fused fibers. Fused fibers often appear flattened and may resemble very large fibers in SEM images, potentially leading to misinterpretation of fiber diameter. In this case, the SEM image on Figure 4(a) indicates the fused fiber morphology with large fiber diameter.
FTIR Analysis
The FTIR spectra of PVP, EBN, and EBN/PVP nanofibers are presented in Figure 6. The FTIR spectrum of EBN displays several characteristic peaks that reflect its complex chemical composition 34. The presence of proteins, particularly glycoproteins and sialoproteins, is indicated by the appearance of amide I and II bands, as well as broad O-H and N-H stretching vibrations. Carbohydrate components, such as sialic acid, are associated with pronounced C-O stretching peaks 35. A broad peak at 3448.72 cm-1 corresponds to hydroxyl groups (O-H), which may be attributed to moisture content and the hydrophilic nature of protein constituents like sialoproteins and glycoproteins 36,37,38. The absorption band at 2926.01 cm-1 represents aliphatic C–H stretching, commonly associated with the carbon chains of amino acids and lipids 35,38,34. The peak at 2372.44 cm-1 indicates the presence of a triple bond between carbon and nitrogen (C≡N). A peak at 1242.16 cm-1 is attributed to the stretching of carbon–nitrogen triple bonds (C≡N). Peaks at 1242.16 cm-1 and 1660.71 cm-1 are indicative of C–N and C=O functional groups, which are fundamental to protein and amino acid structures. The band at 1242.16 cm-1 also reflects C–O stretching associated with carbohydrate residues such as sialic acid and glucosamine found in EBN 39,40,41,42,43,44.
The FTIR spectrum of PVP reveals several well-defined peaks corresponding to its polymeric structure 45,46,47. A strong absorption at 1658.78 cm-1 represents the C=O stretching of the amide group, characteristic of the pyrrolidone ring in PVP 44,47,48. The band at 3429.43 cm-1 indicates the presence of hydroxyl (O–H) groups, a reflection of PVP’s hydrophilic nature 47,49. The peak at 2924.09 cm-1 corresponds to aliphatic C–H stretching, while the peak at 2372.44 cm-1 indicates the presence of (C≡N) triple bonds. The absorption at 1436.97 cm-1 is associated with C–N stretching, confirming the presence of amide linkages within the polymer backbone 47,50. Additionally, a peak at 1033.85 cm-1 suggests the presence of C–O bonds, consistent with ether groups in the polymer structure 12,47.
The FTIR spectrum of the EBN/PVP nanofibers displays a combination of characteristic features from both EBN and PVP, indicating successful blending of the two components. A broad peak corresponding to O–H stretching is observed, which may originate from moisture, hydroxyl groups in EBN, or hydrogen bonding between EBN and PVP. The C=O stretching band confirms the presence of carbonyl groups derived from both the amide bonds in PVP and the protein content of EBN. A peak at 2954.95 cm-1 is attributed to aliphatic C–H bonds, indicative of PVP’s carbon backbone. Furthermore, the absorption bands at 1442.75 cm-1 and 1242.16 cm-1 correspond to C–N and C–O stretching, respectively, reflecting the amide and ether structures contributed by PVP and the carbohydrate content of EBN 46,51.
Thermal Analysis
Thermal analysis was performed on EBN powder, PVP powder, and electrospun EBN/PVP composite nanofibers to evaluate their thermal stability and decomposition behavior. The results are presented as thermogravimetric (TG), derivative thermogravimetric (DTG), and differential thermal analysis (DTA) curves, which collectively provide insights into mass loss, decomposition kinetics, and associated enthalpy changes during thermal degradation 52,53. Figure 7 illustrates the thermal profiles of pure EBN, pure PVP, and the EBN/PVP nanofibers, enabling direct comparison of their thermal performance.
Figure 7(a) presents the TG curve of EBN powder. A substantial weight loss is observed in the temperature range of approximately 250°C to 500°C, indicating the thermal decomposition of organic constituents 54,55,56,57. The corresponding DTG curve exhibits a prominent peak at around 295°C, representing the temperature at which the maximum rate of decomposition occurs. Additionally, the DTA curve reveals an endothermic peak, suggesting a decomposition process or a possible phase transition. These findings indicate that EBN powder is relatively susceptible to thermal degradation at moderate temperatures.
Figure 7(b) illustrates the thermal behavior of pure PVP powder. The TG curve demonstrates that PVP exhibits superior thermal stability, with significant weight loss initiating only above 400°C. A prominent DTG peak is observed at approximately 435°C, indicating the temperature at which the maximum decomposition rate occurs 58. The DTA curve also displays an endothermic peak corresponding to the decomposition process. These results confirm that PVP possesses high thermal stability and is capable of withstanding elevated temperatures without substantial degradation 59,60.
Figure 7(c) shows the TG curve of the electrospun EBN/PVP nanofiber composite, revealing two distinct phases of weight loss: the first occurring between 200°C and 350°C, and the second above 450°C 61,62. The corresponding DTG peaks within these ranges indicate a two-step decomposition process, which can be attributed to the sequential degradation of EBN and PVP components, respectively. The DTA curve also displays two endothermic peaks, supporting the occurrence of gradual thermal transitions resulting from the composite nature of the material. These findings suggest that, although each component retains its inherent thermal behavior, the EBN/PVP nanofibers exhibit enhanced thermal stability compared to pure EBN, likely due to the stabilizing effect of the PVP matrix 63,64,65.
CONCLUSION
The electrospun EBN/PVP nanofibers were successfully synthesized. Smooth, homogeneous, and defect-free fibers were obtained from precursor solutions with EBN/PVP ratios ranging from 10:90 to 60:40. The fiber diameter decreased with increasing electrospinning voltage and collector distance. FTIR analysis confirmed the successful synthesis of EBN/PVP nanofibers by displaying characteristic peaks from both pure PVP and EBN. Thermal analysis indicated enhanced thermal stability of EBN/PVP nanofibers compared to pure EBN. Owing to their favorable morphology, stability, and biocompatibility, these nanofibers hold promising potential for future applications in functional food delivery systems, nutraceutical encapsulation, or biomedical fields where controlled release and protection of bioactive compounds are essential.