Gum karaya (GK), a natural hydrocolloid, was mixed with polyvinyl alcohol (PVA) at different weight ratios and electrospun to produce PVA/GK nanofibers. An 80 : 20 PVA/GK ratio produced the most suitable nanofiber for further testing. Silver nanoparticles (Ag-NPs) were synthesised through chemical reduction of AgNO3 (at different concentrations) in the PVA/GK solution, the GK hydroxyl groups being oxidised to carbonyl groups, and Ag+ cations reduced to metallic Ag-NPs. These PVA/GK/Ag solutions were then electrospun to produce nanofiber membranes containing Ag-NPs (Ag-MEMs). Membrane morphology and other characteristics were analysed using scanning electron microscopy coupled with energy dispersive X-ray analysis, transmission electron microscopy, and UV-Vis and ATR-FTIR spectroscopy. The antibacterial activity of the Ag-NP solution and Ag-MEM was then investigated against Gram-negative
Natural gums derived from plants have many potentially valuable uses as food additives and pharmaceutical ingredients as well as stabilising, suspending, gelling, emulsifying, thickening, binding, and coating agents [
Electrospinning, an environmentally friendly process capable of producing polymer nanofibers with high porosity and large surface area, allows for the use of a variety of polymers and polymer mixtures together with additives and fillers such as gums [
The properties of silver nanoparticles (Ag-NPs) make them particularly useful as antimicrobial materials, biosensors, composite fibres, cryogenic superconducting materials, cosmetic products, antibacterial medical textiles, wound dressing materials, and electronic components [
In this study, we describe a method for producing a new nanofiber membrane and film composed of PVA/GK coated with Ag-NPs. We assess the material’s morphology using various microscopy and spectroscopy techniques and assess its antibacterial activity using Gram-positive and Gram-negative bacteria. The results are discussed in the light of their potential usefulness in the medical, food packaging, and water treatment industries.
Commercial gum karaya (partially deacetylated) with molecular weight (Mw: 1.827 × 106 g/mole), PVA (Mw 88,000, 88% deacetylated), silver nitrate (AgNO3), and glutaraldehyde solution (Grade 1, 50% in water) were purchased from Sigma-Aldrich, USA. All other reagents used in the experiment were of analytical grade. Deionised water was used throughout.
A 10 wt% aqueous PVA solution and 1 wt% GA were prepared in deionised water. A range of PVA/GK electrospinning solutions were produced by mixing PVA (10 wt%) solution with GK (1.0 wt%) at different weight ratios (i.e., 100 : 0, 90 : 10, 80 : 20, 60 : 40, and 50 : 50) in order to identify that, giving the best spinnability and most uniform nanofiber size distribution. The mixtures were kept on a magnetic stirrer at 70°C for 5 h to ensure complete dissolution. The solutions were centrifuged to remove any suspended particles prior to electrospinning.
Based on the results of electrospinning different weight ratios of PVA/GK, the most suitable combination was found to be an 80 : 20 weight ratio mix. This was mixed with aqueous AgNO3 solutions of 1, 2, 4, 5, and 10 mmol L−1 and the resultant solutions stirred at room temperature for 12 h to obtain homogeneous solutions. Sufficient Ag-NP formation was indicated by a dark yellowish colour, whereupon the PVA/GK/Ag solution was deemed ready for electrospinning and testing for antibacterial activity.
The PVA/GK and PVA/GK/Ag solutionswere electrospun in order to produce nanofiber membranes. All electrospinning was carried out with a Nanospider electrospinning machine (NS IWS500U, Elmarco, Czech Republic) under the following parameters: spinning electrode width = 500 mm, effective nanofiber layer width = 200–500 mm; spinning distance = 130–280 mm, substrate speed = 0.015–1.95 m/min, process air flow = 20–150 m3/h, and voltage 0–50 kV. The PVA/GK and PVA/GK/Ag (Ag-MEM) membranes were then cross-linked through exposure to glutaraldehyde vapour in desiccators for 12 h. Both the membranes were then heated in an oven for 12 h at 110°C to complete the cross-linking process. Any excess of glutaraldehyde was removed by keeping membranes under vacuum for 24 h.
Formation of Ag-NPs was confirmed through UV-Vis spectroscopy (UV-1601, Shimadzu, Japan) and transmission electron microscopy (TEM; Tecnai F30, Japan; acceleration voltage 15 kV) was used to analyse Ag-NP size distribution. The morphology of the PVA/GK nanofibers (different weight ratios) and the Ag-MEM was assessed using scanning electron microscopy with energy dispersive X-ray spectroscopy (SEM-EDXA; Zeiss, Ultra/Plus, Germany). Attenuated total reflectance-fourier transform infrared spectroscopy (ATR-FTIR; NICOLET IZ10, Thermo Scientific, USA) was used to characterise the functional groups of PVA, GK, PVA/GK, and Ag-MEM. Conductivity and viscosity of the electrospinning solutions were recorded using a Toledo FG3 electric conductivity meter (Mettler, USA) and a rotational viscometer (Brookfield Engineering Laboratories, USA). The concentration of Ag NPs in PVA/GK/Ag solution and Ag-MEM was established by ICP-AES.
The bacterial strains of Gram-negative
We determined the antibacterial activity of four PVA/GK/Ag solutions (1, 2, 4, 5, and 10 mM) and samples of Ag-MEM (each containing the equivalent of 1 mM of AgNO3). The PVA/GK/Ag solutions were pipetted onto a sterilised membrane filter and placed onto an inoculated agar plate, while 6 mm diameter circles of Ag-MEM were placed directly onto inoculated agar plates. Similarly sized samples of PVA/GK solution (10 mg/mL) and samples of nanofiber membrane without Ag-NP were used as controls. The samples and inoculated agar plates were then incubated for 24 h at 37°C. The zone of inhibition (ZOI) was determined as the total diameter (mm) of PVA/GK/Ag-filter paper or Ag-MEM sample plus the halo zone where bacterial growth was inhibited. All measurements were performed in triplicate for the PVA/GK/Ag solutions and repeated three times (once for each bacterial strain, i.e., nine runs) for the Ag-MEM.
One-way ANOVA and the Mann-Whitney test (GraphPad Prism Software, CA, USA) were used to compare differences among the mean ZOIs for the PVA/GK/Ag solutions and Ag-MEM on
The colour change of the PVA/GK solution with a ratio of 80 : 20 to dark yellow following formation of Ag-NPs is shown in Figures
PVA/GK solution (a) before formation of Ag NPs and (b) after Ag NPS formation; and (c) UV-vis spectra of PVA/GK aqueous solution containing Ag-NP prepared at various concentrations of AgNO3 (0, 1, 2, 4, 5, and 10 mM).
GK comprises around 60% neutral sugars and 40% acidic sugars and a range of hydroxyl, carbonyl, carboxyl, and acetyl functional groups [
Presence of Ag-NPs in the PVA/GK/Ag solution was confirmed by SEM imaging of freshly formed Ag-NP (Figure
SEM image of (a) Ag-NP prepared using PVA/GK; (b) EDS of Ag-NP, showing the presence of Ag; and (c) TEM image of Ag-NP prepared using PVA/GK and 10 mM AgNO3; and (d) particles diameter distribution of Ag NPs (7–10 nm).
We prepared a range of PVA/GK weight ratio mixtures (100 : 0, 90 : 10, 80 : 20, 60 : 40, and 50 : 50) in order to optimise the electrospinning solution, that is, to obtain optimal spinnability and uniform nanofiber size. SEM images of the resultant nanofibers (Figures
SEM images of electrospun PVA/GK mixed with different weight ratios: (a) PVA/GK (50/50); (b) PVA/GK (60/40); (c) PVA/GK (70/30); (d) PVA/GK (80/20); (e) PVA/GK (90/10); and (f) neat PVA (100/0) 10
Further, the pure GK solution proved too viscous for electrospinning as GK is an acidic polymer with high viscosity and molecular weight [
Not only were the nature and morphology of the nanofibers affected by polymer solution viscosity and conductivity (both affected by the PVA/GK weight ratio used), but we also found that the viscosity of the PVA/GK/Ag electrospinning solution increased from 300 to 500 mPa·s and its conductivity from 2500 to 3200 mS·cm−1, with increasing AgNO3 concentration (1, 2, 4, 5, and 10 mmol L−1). The levels at 1 mM, however, were within acceptable limits for electrospinning using the 80 : 20 PVA/GK weight ratios and provided reasonable Ag-NP coverage in the final Ag-MEM (Figure
Digital image of (a) PVA/GK membrane and (b) Ag-MEM prepared by electrospinning of PVA/GK and PVA/GK/Ag NP solution, respectively.
SEM micrographs of the final electrospun Ag-MEM (Figure
SEM micrograph of (a) Ag-MEM showing the presence of Ag NPs on the surface membrane; (b) EDXA analysis of Ag-NP on Ag-MEM; (c) EDXA layered image indicating the presence of Ag-NP on the surface of Ag-MEM.
In examining the bonding between Ag-NPs and the Ag-MEM (also GK, PVA, and PVA/GK) using ATR-FTIR, we noted a broad absorption peak centred around 3318–3350 cm−1 for all samples, attributable to O–H stretching vibration in the hydrogen bonded hydroxyl groups (Figure
ATR-FTIR spectra of PVA, GK, PVA/GK blend, and Ag-MEM.
The peaks at 1430 cm−1 and 1326 cm−1 are characteristic of O–H groups and C–H deformation vibration in PVA, respectively, while the peak at 1000–1100 cm−1 can be assigned to C–O stretching and O–H bending vibrations arising from the PVA chain. The appearance of a new peak at 1561 cm−1 in the PVA/GK blend represents O–H group deformation vibration with the H bond, suggesting the formation of an H bond between PVA and GK when forming the PVA/GK blend. Structurally, GK has abundant hydroxyl groups; hence, H bonding interactions between GK and PVA occur readily on blending with PVA. The O–H bond absorption band at 3300–3500 cm−1 indicates that the O–H bond was involved in bonding with the Ag-NPs. Carboxylate group stretching vibration at 1419 cm−1 was considerably reduced in the PVA/GK/Ag-MEM spectrum, demonstrating binding of Ag+ ions with the PVA/GK nanofibres. These results are in agreement with earlier reported studies on the binding of Ag-NPs with other natural gums [
We tested the antibacterial activity of the PVA/GK/Ag and Ag-MEM composites synthesised in this study against Gram-negative
For the PVA/GK/Ag solution, zone of inhibitions (ZOIs) for Gram-negative
Diameter (mm) of zone of inhibition (ZOI) for PVA/GK/Ag solutions produced with different concentrations of AgNO3 (1, 2, 4, 5, and 10 mM) and Ag-MEM (1 mM AgNO3). Means were calculated from in triplicate tests on the PVA/GK/Ag solution and nine replicates for the Ag-MEM (±SD).
PVA/GK/Ag solution | Ag-MEM | |||||
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AgNO3 (mM) | 1 | 2 | 4 | 5 | 10 | 1 |
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The ZOI of Gram-positive
The bacterial growth inhibition zones of
While the mechanism for Ag-NP action is still not fully understood, it has been documented that Ag-NPs cause structural changes when they interact with the outer membrane of bacteria [
In this study, we produced an electrospun nanofiber membrane from GK, a natural hydrocolloid, blended with PVA. Uniform PVA/GK nanofibers were obtained at a PVA/GK weight ratio of 80 : 20. The 80 : 20 PVA/GK was blended with various concentrations of AgNO3 solution to produce a PVA/GK/Ag NP solution. PVA/GK/Ag NP solution was then used to produce nanofibers containing AG-NPs, from which an antibacterial nanofiber membrane (Ag-MEM) was fabricated. The PVA/GK/Ag solution and Ag-MEM showed clear antibacterial activity toward Gram-negative
The authors declare no conflict of interests.
The research reported in this paper was supported in part by Project LO1201, the financial support of the Ministry of Education, Youth and Sports in the framework of the targeted support of the “National Programme for Sustainability I,” the OPR & DI Project and OP VaVpI of the Centre for Nanomaterials, Advanced Technologies and Innovation, CZ.1.05/2.1.00/01.0005, the “Project Development of Research Teams for R & D Projects’’ at the Technical university of Liberec, CZ.1.07/2.3.00/30.0024, and a Grant from the Competence Centre, TE01020218.