Trends Sci. 2025; 22(6): 9762

Fabrication of Antibiofilm-Based-Polymer Nanocomposite for Biophysical Applications

Rawaa A. Abdul-Nabi^1,2 and Ehssan Al-Bermany^2,*

¹Department of Electrical Engineering Techniques, Al-Mussaib Technical College, Al-Furat AL-Awssat Technical University, Najaf Governorate, Iraq

²Department of Physics, College of Education for Pure Sciences, University of Babylon, Babylon Governorate, Iraq

(^*Corresponding author’s e-mail: [email protected])

Received: 12 January 2025, Revised: 19 February 2025, Accepted: 26 February 2025, Published: 1 May 2025

Abstract

Polymer nanocomposites attracted significant interest because of their effective characterization and low cost. This investigation aims to fabricate newly cost-effective blended polymer polyethylene oxide (PEO), carboxymethyl cellulose (CMC), and conductive nano-polyaniline (PANI) were improved by different loading ratios of hybrid graphene oxide (Fixed ratio = 0.05 %) and different loading ratio of silicon nitride (0.05, 0.25, and 0.45 %), (GO-Si₃N₄) nanomaterials to fabricated nanocomposites. PEO-CMC-PANI/GO-Si₃N₄ composites were fabricated using the acoustic-ultrasonic method. The semicrystalline performance of samples was proved using X-ray diffraction, and Fourier-transform infrared spectroscopy exposed strong interfacial interaction. Field emission scanning electron microscopies showed homogenous samples with fine nanomaterial dispersion in the matrix, revealing significant changes. The transparency of the samples was increased with the rise in the nanomaterial’s ratio, with the main electron transitions at about 280 nm wavelengths due to reducing the optical band gap from 3.5 to 2.6 and 2.8 eV of allowed and forbidden transitions. The inhibition of biofilm formation by the nanocomposite at a concentration of 5 % for bacterial isolates of Escherichia coli and Streptococcus mutans was increased compared to untreated bacteria. The optical density of S. mutants and E. coli was improved from 0.95, 0.2, and 0.4 mm, respectively. These nanocomposites presented good and cheap materials with a high aptitude for killing bacteria. They could be used in various biological and related applications, such as coating operating rooms, tools, cleaning, and even replacing traditional disinfectants.

Keywords: Polymer, Silicon, Graphene, Biofilm, Escherichia coli, Streptococcus mutans

Introduction

An important field of research is the investigation of the optical properties of antibacterial materials for several probable reasons, including the following [1]: The presence and activity of antibacterial agents can be detected and monitored with the help of optical characteristics, which can be utilized for both detection and monitoring purposes [2]. Researchers can build optical sensing techniques to identify the presence of bacteria or evaluate the efficacy of antibacterial treatments by analyzing the interaction of light with antibacterial materials [3]. This allows the researchers to develop strategies using light. This is especially helpful in hospital settings, where it is necessary to guarantee

that sufficient cleanliness and sterilization are maintained [4,5].

Understanding the mechanism of action of antibacterial materials is possible by 1^st understanding their optical features [6]. For instance, if a substance demonstrates particular optical qualities when it comes into contact with bacteria, it can indicate that specific chemical or physical processes are taking place that contribute to the material’s antibacterial activity [7]. By examining these characteristics, researchers can acquire a more comprehensive comprehension of the functioning of antibacterial materials and potentially enhance their efficiency [8]. The surface characteristics of antibacterial materials can impact the material’s optical properties. Researchers can explore the optical response of modified surfaces to bacteria or bacterial byproducts to produce surfaces that are either naturally antibacterial or activated when bacteria come into contact [9,10].

Antibiotics are a significant breakthrough in the field of treatment, playing a vital role in saving many lives by rendering formerly fatal illnesses treatable [11]. The dependability of antibiotics is the cornerstone of modern medicine and has enabled the advancement of various medical operations that were before unattainable. Every facet of modern medicine encompasses surgical interventions and therapies for burns and injuries. The discovery of antimicrobial medications made all of this possible [12]. However, scholars and healthcare professionals need help identifying a resolution to the escalating issue of antibiotic resistance, which is especially prevalent in healthcare environments. Immediate action must be taken to resolve this issue as it threatens the fundamental principles on which modern medicine was founded [13]. Bacteria acquire resistance to antibacterial treatments in several ways, necessitating a novel approach to produce new bactericidal agents. The quest for novel antimicrobial drugs or enhancements in the efficacy of existing ones is imperative [14].

Nanomaterials can be considered based on their dimensions: 0, 1, 2, and 3-dimensional thin films and layers [15]. Studies have demonstrated significant interest in inorganic nanoparticles due to their potential for various uses, such as their antibacterial properties [16]. Graphene oxide (GO) has shown promising potential as both an antibacterial agent and an antibiotic carrier due to several mechanisms [17]. It uses a physical damage procedure using the sharp edges of their nanosheets that can penetrate and disrupt bacterial cell membranes, leading to cell death. The addition strategy of GO is oxidative stress, where it can generate reactive oxygen species (ROS) that are the bases of oxidative stress in bacterial cells, lipids, DNA, and damaging proteins. Moreover, photothermal effects as light irradiation by GO, can convert light energy into heat, enhancing its antibacterial activity by causing thermal damage to bacterial cells [18]. Studies have demonstrated that GO can effectively inhibit the growth of various bacteria, including E. coli and S. aureus [19]. GO can coat medical devices, reducing the risk of bacterial infections. Incorporating GO into wound dressings can prevent bacterial infections and promote healing [18]. The large surface area of GO is associated with controlling the release of antibiotics and carrying enormous quantities of antibiotics essential for effective concentrations to be maintained for an extended duration [20]. GO can be combined with different antibiotics, doxycycline and ciprofloxacin, which improve the effectiveness against bacteria due to the GO stability compared to antibiotics alone [18], and ensure its safety and efficacy in medical applications [21]. In addition, GO-based sensors can detect cancer biomarkers at deficient concentrations, aiding in early diagnosis [22].

Another interesting nanomaterial is silicon nitride (Si₃N₄), a ceramic material known for its exceptional mechanical properties, chemical stability, and biocompatibility [23]. Although its main applications are in bearings, cutting tools, and implants, research has also investigated its potential in medicinal fields, such as antibacterial, antibiotic administration, and anticancer therapy [24]. The rough surface shape of Si₃N₄ at the micro- and nanoscale prevents bacterial adherence and growth by disrupting bacterial cell walls. Si₃N₄ exhibits chemical inertness, which enhances its ability to resist bacterial colonization and the production of biofilms. By introducing additional elements, Si₃N₄ can be doped to improve its antibacterial capabilities by releasing antimicrobial ions [25]. Si₃N₄ is employed in orthopedic and dental implants because of its antibacterial qualities. Also, it can reduce the probability of infection during surgical procedures. That makes Si₃N₄ a nondonated material for coating medical devices that can efficiently inhibit bacterial infections [24]. Si₃N₄ exhibits an outstanding biocompatibility that is suitable for drug delivery systems. Si₃N₄ can be controlled to release antibiotics at a regulated rate, and therapeutic doses are maintained for extended periods. The antibiotic is guaranteed not to degrade because of the chemical stability of Si₃N₄ [26]. It is a practical material for delivering the drag of anticancer through the porosity and high surface area that gives it features for loading and control, realizing the therapeutic chemicals. It is also presented as an efficient composite for treating bone infections and other localized illnesses by deliberately discharging antibiotics [27]. PEO can dissolve in many substances and has a large molecular weight [28]. PEO is compatible with various applications. For instance, PEO is used in drug delivery, the environment, medicine, industry, etc. [29]. CMC can dissolve in water, is semicrystalline, is low-cost, and has low conductivity and strength. Also, it is compatible and non-toxic and is used in applications such as cosmetics, living organisms, industrial, pharmaceutical, food, etc. [30]. PANI has high electrical conductivity and thermal stability, lower density than metals, a significant visible light absorption ability, good energy storage capabilities, and low cost. Amazing PANI characterizations are used it in various applications, such as drug delivery, display devices, photovoltaic cells, plastic batteries, etc. [31]. This study focused on the impact of nanomaterials on improving nanocomposites as effective and low-cost antibacterial materials could replace traditional use.

Materials and methods

The details, molecular weight, chemical form, and supplier company of materials used in this study are shown in Table 1. PEO with MW, 100,000 g mol^–1 provided by Sigma-Aldrich Company, UK. CMC with MW, 700,000 g mol^–1 supplied by Cheng Du Micxy Chemical Co., Ltd., China. Polyaniline is a conductive polymer with nanoparticles (20 nm)-dark/light green-black, with MW 150,000 g mol^–1, manufactured by Panichem. Co., LTD, Korea. Silicon Nitride nanopowders with size (15 - 30 nm) and grey white and MW, 140.28 g mol^–1. Full characterization of the synthesized GO was provided in the publication [21].

Solution-sonication-casting methods were developed and used to prepare the samples following the procedures: First, all the polymer was dissolved in distilled water (DW) at a ratio of 5 g/100 mL independently.

Secondly, after dissolving each polymer, it was mixed for 24 h to prepare the blended polymers at a ratio of (60:30:10 %) of (PEO: CMC: PANI) at room temperature (RT) (25 ± 3 °C) for 24 h to achieve optimal dissolution and homogeneity.

Thirdly, the nanomaterial was separately dispersed in deionized water using a stirrer with 100 mg/100 mL, followed by 10 min of bath sonication for every hour of mixing to achieve optimal dispersion.

Fourthly, to achieve the best possible results of the novel quinary nanocomposites. Nanomaterials were mixed with different concentrations (1, 3, and 5 %) of (GO: Si₃N₄) with a fixed concentration of GO and various concentrations of Si₃N_4, together with the assistance of sonication for 30 minutes.

Fifthly, combined nanomaterials were loaded in blend polymers for 7 days using a stirrer and 30 minutes of sonication in a bath.

Finally, samples were stored in class-celled pots for the biofilm test, and others were dried on a glass slide for XRD and FESEM and utilized as solutions for other tests. The thickness of the samples was 60 microns.

Table 1 The ratios of micing sample components.

Results and discussion

Figure 1 presents the XRD patterns for the pure (CMC/PANI/PEO) blend sample and its nanocomposites filled with the mixture of GO + Si₃N₄ nanoparticles at different ratios (1, 3, and 5 %). The XRD was set up to run 28 times for 30 min for each XRD test for reliable results. Because the PEO polymer has a semi-crystalline structure, the XRD pattern of ternary blended polymers (B) showed that the polymer matrix is semi-crystalline. Where PEO is associated with the primary peaks at 18.4 and 22.7 °. PANI showed additional tiny peaks, whereas the CMC semi-crystalline peak, seen at 22.3 °, overlapped with PEO peaks. The peaks of nanocomposites revealed a minor movement in the locations of most of the peaks from their initial positions. The contribution of (GO + Si₃N₄) nanomaterials at various ratios resulted in the shifting of most peaks with increased peak intensity and nanomaterial loading ratios in the matrix.

The XRD patterns of the blend filled with the contribution of (GO-Si₃N₄) nanomaterials at different ratios (1, 3, and 5 wt.%) revealed a reduction of the peak intensity with the addition and raising of the (Si₃N₄) nanoparticles ratio. Moreover, the peaks revealed a slight shifting in most of the peaks from their original positions. Specifically, NC1 revealed a sign of GO in 11.1 and 40.02 ° [32], and all other XRD diffraction peaks might be indexed to the Si₃N_4, a hexagonal structure matched (JCPDS Card no. 41-0360), and other reports [33]. In addition, the obtained XRD results proved the strong interaction and good distribution of the (GO-Si₃N₄) nanoparticles and the matrix blend, as demonstrated by FT-IR results and literature [34,35]. Most peaks were shifted, and the intensity at around 19 ° increased to become higher compared with other peaks and blended samples; moreover, the dissolution of (GO-Si₃N₄) nanoparticles within the structure of the PEO-CMC-PANI polymer blend.

X-ray diffraction determines any material’s structural factors, which are crucial in explaining many of the material’s physical properties. When X-ray light of wavelength (𝜆) is projected at a Bragg diffraction angle (𝜃) onto a crystal lattice, the incoming X-rays interact constructively with the sample if the circumstances meet Bragg’s law, as shown in Figure 1 [36].

(1)

where means distance and is diffraction rating.

The crystal size (D) in nm units was considered using the Scherrer Eq. (2) [37].

(2)

(D) means the size of the crystal (k = 0.9), and (β) means the complete breadth at half the highest point (FWHM).

In Table 1, calculations showed that the crystalline size of samples loaded with NPs increased significantly after increasing the loading of GO-Si₃N₄ by 16.71, 19.5, and 20.2, then reduced to 17.2 nm for B, NC1, NC2, and NC3, respectively. At the same time, the samples had an increase in lattice strain of up to 8.58, 6.77, and 8.80 % for NC1, NC2, and NC3, respectively.

Figure 1 XRD patterns of (a) B, (b) N1 %, (c) N3 %, and (d) N5 %.

Table 1 provides an overview of the diffraction angle (2θ), FWHM (β), d-spacing, average crystallite size, and size of the crystallites for both blended polymers and samples loaded with different amounts of GO-Si₃N₄.

Figure 2 FTIR spectrum for blend polymers and their nanocomposites.

Figure 2 displays the doping of (GO-Si₃N₄) nanoparticles in the blended (PEO-CMC-PANI) polymers. Several major results are presented in the FTIR spectrum of the pristine blended polymer and (PEO-CMC-PANI/GO-Si₃N₄) nanocomposites. The FTIR spectrum was set up to run 16 times in each FTIR run, and the sample was run for reliable results. The most functional peaks of the spectrum of blended polymer and quinary (N1, N3, and N5 %) nanocomposites match peaks in the contained polymers peaks spectrum. In general, samples exposed strong O-H peaks at 3,346 and 1,636 cm⁻¹. The nanomaterial contribution in nanocomposites looked to be responsible for presenting the C-O-C bond and helped to form a complex network between the oxygen functional groups of GO-Si₃N₄ nanomaterial and blended polymers, which strongly agrees with other findings [38].

All samples generally showed strong absorption peaks at 3,346, 1,636, 1,088, and 696 cm⁻¹, as hydrogen interactions between the ternary blended polymer (B) and nanomaterials. The contribution of the binary nanomaterial in nanocomposites appeared, which is responsible for the decrease in the intensity of most peaks. This is connected with creating the network among GO and Si₃N₄ nonmaterial and polymers, which strongly agrees with other findings [38]. Structural features of the blended polymer were not affected after the fabrication of the nanocomposites in agreement with another report [39]. Table 3 displays the functional group and corresponding FTIR peaks.

Table 3 The functional group and corresponding FTIR peaks.

Figure 3 FESEM images of (A) B, (B) N1 %, (C) N3 %, and (D) N5 %.

When the polymers are mixed, the sample of blended polymer B shows a significant surface change, as shown in Figure 3(A). It presents a rough, uneven, granular appearance with some agglomerations and particles of different sizes and a cauliflower-like structure with deep cavities. When nanomaterials were loaded at 1 % to the blended polymer mixture, the bonding between the polymers improved and became uniformly distributed, the surface became smoother with the attached nanomaterials, the granular shape was lost, and the deep cavities and some tangled threads appeared for a similar network of PEO, CMC, PANI, and nanomaterials. When nanomaterials were loaded at 1 % into the polymer mixture Figure 3(B), the bonding between the polymers improved and became uniformly distributed. The surface became smoother with the attached nanomaterials. The granular shape was lost, and the deep cavities and some tangled threads appeared for a similar PEO, CMC, PANI, and nanomaterials network. Increasing the Si₃N₄ ratio in the sample to 3 %, as shown in Figure 3(C), helped provide a porous structure resulting from PANI nanoparticles and nanofibers with complex network shapes where polymers are linked together in an arm-like manner, interspersed with some pores as well as particles of various clusters. The sharp edges of the graphene oxide layers were also revealed. In Figure 3(D), increasing the nanomaterials to 5 % increased the width of the porous structure resulting from the nanoparticles and nanofibers of PANI; in addition, the surface became smoother in other parts, although its increased value was modest compared to the blended sample [54].

Figure 4 Transmittance of samples with wavelength.

The change in the transmittance spectrum as a function of the wavelength of the pure mixed polymer and the compound loaded with different percentages (1, 3, and 5 %) of the binary nanomaterial GO-Si₃N₄ appears in Figure 4. The UV-Vis spectrum was set up to run 5 times for each sample, and the average was taken for reliable results. The transmittance was the lowest possible at the basic absorption edge (short wavelengths). The transmittance increased with increasing wavelength, showing a sudden and strong increase until it stabilizes after the wavelength (340 nm). On the other hand, the greatest value of transmittance was in the near-infrared and visible rays )0.77 (and then gradually decreased with an increase in the percentage of GO-Si₃N₄, as loading the binary material GO-Si₃N₄ led to the accumulation of NPs, which increased the surface roughness, and thus the light surface fragmentation increases with increasing loading rates, which causes a decreased permeability. The optical energy gap values of the indirect transition were calculated according to Tauc’s relation [55].

(3)

The values of r = 2 and 3 for permitted and forbidden indirect transitions, while (B), (h), and (E_g) represent constants, photon energy, and the energy gap, respectively.

Results are revealed in Table 4 and Figure 5. The obtained result matches the same behavior of the literature that used NiCl₂ꞏ6H₂O in (PVDF-PHFP). The nanofiber had an optical bandgap reduced from 3.24 to 3.10 eV [56]. These highly transparent samples in this wide wavenumber range could open a wide way for various optical applications such as optical sensors, solar cells, etc.

Table 4 The optical energy gap values of the indirect transition in eV for samples.

Figure 5 Depict the allowed and forbidden optical energy gap for the B, N1, N3, and N5 %.

Antibiofilm activity

Biofilm is one of the characteristics of virulence that can be formed by gram-negative and gram-positive bacteria, regardless of whether they are isolated from the body or from outside the body. This is especially true for bacteria that are found on medical devices such as catheters and dialysis devices, as well as in burns, industrial water, and other surfaces of organic and inorganic materials [57]. There are 4 stages to the life cycle of a microbial biofilm as shown in Figure 6(A) [58].

Figure 6 Schematic diagrams of (A) the different steps of Biofilm formation and (B) evaluation of the antibiofilm activity of the Nanoantigen.

The procedure was performed using a sterile 96-hole microtiter plate assay method (Crystal violet staining), as shown in Figure 6(B). According to the literature, individuals with urinary tract infections brought on by E. coli bacteria that form biofilms are more likely to contract the illness again after recovering. The study examined how the crystal violet color compound attaches to adherent cells. The studied bacterial isolates were able to produce biofilms. The results of the CV test revealed that 0.1 % that the tested bacterial isolates could form biofilms, and 100 % for E. coli and Streptococcus. Studies showed that people with urinary tract infections caused by biofilm-producing E. coli bacteria are more susceptible to re-infection with the disease after recovery from it [59,60]. In addition, treatment with (100 microliters) of Si₃N₄-GO NPs results in rapid biofilm removal. The samples were run 3 times to confirm the obtained results, and the standard deviation was ± 0.02. Figures 7(A) - 7(C) depict the results of E. coli and S. mutans isolates treated with GO-Si₃N₄. The 5 % nanocomposite inhibits biofilm formation (-) with a value of 0.2 and 0.4 for S. mutans and E. coli bacteria, respectively. In addition, there is an inverse correlation between the optical density and the addition of the nanomaterial, as a decrease in the optical density is noticed with the treatment of bacteria with the nanomaterial.

This demonstrates the ability of GO to penetrate the EPS of biofilms and destroy their 3-dimensional structure, as shown in Figure 7(C). It prevents the formation of biofilms either by preventing the synthesis of exogenous sugars, which is considered an important step in the formation of biofilms by entering the aquaporin channels that carry water and nutrients through the layers of sugars present on the bacterial cell wall, or the formation of biofilms is reduced by another behavior, which is intervention. All produced biofilms were detached from the wells, in the proteins and enzymes necessary for adhesion and entry into the bacterial cell. As shown in Table 5. It was found that the rate of optical density of S. mutans was better than that of E. coli. This indicates more bacterial inhibition, consistent with the literature [61-64]. The samples were run 3 times to confirm the obtained results, and the standard deviation was ± 0.02.

Figure 7 (A) Antibiofilm (optical density) of isolates of S. mutans compared to E. coli of control (without bacteria) and N5 % sample, (B) image of E. coli and S. mutans bacteria adhered with crystal violet in a microtiter plate, of 5 % GO-Si₃N₄ concentration compared to control (without bacteria) and (C) image of E. coli and S. mutans after adhered bacteria stained with crystal violet by microscopy.

Table 5 The optical density of nano-fabricated antibiofilm.

Conclusions

Effective and low-cost new polymer nanocomposites were successfully fabricated. The samples showed significant interaction between their components, and the addition of nanomaterials increased this interaction without affecting the main structure. Samples showed reduced transmittance, optical band gap, and significantly increased electron transitions that helped enhance the inhibition in biofilm formation, increasing with increasing concentrations for the 2 bacterial isolates. The promising nanocomposite exhibited excellent optical and antibacterial characteristics that can be employed in optical devices, filters, and antimicrobials.

Acknowledgments

RA Abdul-Nabi did the investigation, methodology, 1^st analysis, and wrote the 1^st draft. E Al-Bermany is responsible for project administration, suggestions, investigation, review, and editing of the final manuscript.

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