Modification on antibacterial activity of PVC/PVDF blend filled with CuO NPs using laser ablation technique

Nanocomposite of polyvinyl chloride (PVC)/polyvinylidene fluoride (PVDF) have been in situ synthesized. Copper oxide nanoparticles (CuONPs) have been prepared via using the laser ablation technique. Nanoparticles were added to the blend. The properties of the blend were studied before and after adding CuONPs. These properties were characterized by different techniques. Antimicrobial activity of the prepared nanocomposite film was investigated. FTIR data show vibrational spectral bands and the shift of the bands is related to the interaction and the complexation that occurs between blend and nanoparticles. Structural properties and crystallinity of the samples were investigated using XRD diffraction. XRD results illustrated the effect of CuONPs at two new peaks 2θ = 26.25º and 38.41º. These results confirmed the interaction CuO NPs and PVDF/PVC matrix. UV–Visible analyses confirmed the existing of copper oxide nanoparticles and were also used for determining the optical absorption edge. The absorption edges have been obtained at 430–520 nm for all of the doping films. The obtained values for indirect and direct bandgaps were reduced by raising the nanoparticles because of the presence of charge transfer between PVC/PVDF and CuONPs. SEM images illustrateed the presence of CuONPs on the surface of the blend and the morphology changes which occurred to the blend. The antibacterial activity for the nanocomposite proved the antimicrobial effect of copper oxide nanoparticles. The prepared PVC/PVDF/CuONPs are potentially suggesting to be applied for biomedical applications.


Introduction
Nowadays, nanotechnology has a great attention in different fields because the materials with nanoscale size have new and improved physical, chemical and biological properties [1]. Nanoparticles can be prepared by chemical, electrochemical or photochemical methods. Scientists are attracted to metallic nanoparticles for years which are now deeply used in biomedical sciences and engineering [2,3]. Attention was focused on copper among the other metallic nanoparticles due to its catalytic and electrolytic characteristics and also it is more available and cheaper than other nanofiller material. CuO NPs were used as an anti-infective agent in various applications, for example, water remediation, food processing and medical instruments protection. Thin films of copper oxide are the most proper electrode material for electrochemical capacitors. CuONPs can be prepared by laser ablation technique which is a thermal or non-thermal process uses intense continuous wave or pulsed laser beam to break down apart from a matter forming nanoparticles [4][5][6][7]. Laser ablation technique has a great attention as it provides simple and low coast method in addition to providing not contaminated nanoparticles with ion residuals. There are different applications for CuONPs in super hydrophobic coatings and corrosion resistance. They also have variety uses in biomedical applications as they show anticancer, antimicrobial and antioxidant efficiency. They can combine with the microbial cells due to their nanoscale size and large surface area giving them attraction to be used in biomedical, industrial and agricultural applications [8].
Polymer is a chemical compound which composed of small repeating units called "monomers" linked together in long-chain losing some chemical groups. Atoms are linked together in the polymer backbone by covalent bond force. The mechanical and physical characterizations of the polymer are determined by the length of the monomer chain, while the chemical and electrical characteristics are determined by the polymer's structure [9]. Due to its properties, it has a wide range of uses. The combination between the polymer and the nanoparticle changes its optical, physical, thermal and electrical properties. This paved the way for producing materials with improved properties in different scientific applications [10][11][12].
Polyvinyl chloride (PVC) comes after polypropylene and in the world's fabrication demand [13]. It is a thermoplastic polymer which has a widespread uses in different fields [14]. It was synthesized by accident by German chemist Eugen Baumann. It is sorted into two forms rigid (RPVC) and flexible. RPVC is stronger than flexible PVC, while the properties of flexible PVC are more valuable than RPVC. PVC has a life time more over 100 years because it can be recycled seven times [15]. It could be prepared by polymerization of vinyl chloride monomer [16]. It is a good insulation material, resistant to all inorganic chemicals and resistant to weathering, chemical rotting, corrosion, shock and abrasion [17]. It has good physical as well as mechanical properties. It is tough and has lightweight. It has various applications including constructions, domestics, toys, wires and cables, packaging, clothing, automotive and medical materials [18][19][20][21].
Polyvinylidene fluoride (PVDF) is a semicrystalline polymer, which could be produced by free-radical polymerization of gaseous vinylidene fluoride (VDF) Polymer Bulletin (2023) 80:6247-6261 monomer. PVDF has at least five kinds of crystal phase, namely α, β, γ, ε and δ phases, while β phase is the most attractive type. It has great attention because of its pyroelectric and characterization. The common uses of PVDF are electricity, batteries, healthcare and pharmaceutics [22][23][24].
Our study aimed to prepare blend with composition (30 PVC/70 PVDF) wt/ wt incorporated via CuONPs, which have been synthesized by casting technique. Characterizations of PVC/PVDF blends were examined using various techniques. In addition, the antibacterial behavior of the papered sample was tested and could be used in wound healing applications.
Preparation of PVC/PVDF composite PVC and PVDF were dissolved in THF in a 1:3 weight ratio with continuous stirring at 50 °C for 7 h until a homogeneous solution was obtained.

Preparation of PVC/PVDF/CuONPs by laser ablation process
CuONPs have been fabricated by laser ablation in liquids technique "PLAL." Polyvinyl chloride (PVC)/polyvinylidene fluoride (PVDF) blend was doped with different concentration of CuONPs (0, 1, 2, 3, 4 and 5). Nd:YAG laser emits its fundamental wavelength (λ) at 1064 nm and was used as irradiation source. The plate was sterilized before the experiment. Quartz convex lens of focal length 100 mm has been utilized to focus the laser beam perpendicularly on the target surface. The power of laser was 3.6 W, repetition rate was 10 Hz, and pulse duration was 7 nm [21,22]. The pure blend of PVC/PVDF was dried in the oven at 50 °C for 1 h before using it to get rid of any moisture. The PVC/PVDF blend with weight ratio (30/70) was dissolved in tetrahydrofuran THF solvent with constant stirring at room temperature for 1 h until obtaining a homogenous solution. The different concentrations of CuONPs were added to the blend solution. The obtained blend solution has been poured in clear Petri dishes left in oven at 50º for 48 h to dry and remove any remaining traces of solvent.

Characterization techniques
XRD scans have been investigated using (Shimadzu 7000, Japan) occupied by Cu-Kα radiation (λ = 0.154060 nm) at 30 kV and 30 mA; the patterns were collected in the Bragg's angle (2θ) ranging between 5° and 80°. FT-IR spectra of the fabricated nanocomposite were performed within the wave number range 400-4000 cm −1 using a spectrometer type (Jasco, 6100, Japan). UV-Visible spectroscopy was investigated by JASCO (V-570) spectrophotometer in the wavelength region of 200-1000 nm. Morphological investigations of the prepared films have been obtained via FE-SEM type (Quanta FEG 250, USA).

The antibacterial activity
The prepared PVC/PVDF/CuONPs films have a dynamic effect versus a broadspectrum bacteria including both gram-negative (Pseudomonas aeruginosa and Escherichia coli) and gram-positive bacteria (Staphylococcus aureus and Bacillus subtilis). The MIC was used to estimate the minimal nanoparticle concentration required to prevent pathogenic bacteria growth, whereas the MBC was done on Muller-Hinton agar plates [23]. The test took place by pouring 3 ml of blend/ copper oxide nanoparticles in Petri dishes containing agar; the dishes were incubated at 36 ο C for approximately 5 h. The antibacterial activity was announced after 24 h from incubation by measuring the inhibition zone and calculating the activity index at room temperature. THF was also performed against bacteria, so we measured the activity index for THF; then, we subtract the value of THF from the samples to obtain an activity index belonging to copper oxide nanoparticles. The test was done three times to obtain the standard error (SD) ( ±).

Results and discussion
XRD XRD measurements were utilized to illustrate the nature of the crystallinity of the polymer [25]. The XRD of PVC/PVDF blend showed the semicrystalline nature of the film. The X-ray patterns of PVC polymer illustrated the amorphous nature and characterized at two peaks located at 2θ = 18.69º and 23.72º as shown in Fig. 1.
XRD pattern of PVDF illustrated a semicrystalline nature and characterized at multiple peaks located at 2θ = 18.27º, 19.95º, 26.52º, 32.94º, 35.88º, 38.68º and 56.28º [26]. The 2θ = 26.52º, 32.94º, 35.88º, 38.68º and 56.28º peaks observed for pure PVDF disappeared in the blend because of the addition of pure PVC which reducing the long-range order in PVDF. The spectrum of the PVC/PVDF blend showed the amorphous nature characterized by two peaks referred to the complexation and miscibility of the two polymers.
XRD illustrated the nature of the crystallinity of the polymer film doped with CuONPs and characterized with two main peaks located at 2θ = 18.04º and 19.86º. Some changes occurred by adding CuONPs to the blend [27,28]. The effect of CuONPs appeared at two new peaks 2θ = 26.25º and 38.41º. The intensity of the peaks increased by adding CuONPs at the first concentration (Cu1) and then reduced gradually by adding the rest of the concentration. XRD spectra illustrated the crystalline nature of the nanocomposite blend at the lowest concentration which gradually disappeared by increasing CuONPs concentration including increasing the volume of the amorphous region. The amorphous phase of the nanocomposite blend appeared clearly at the highest concentration.
For many decades, the strategy of development polymers with high conductivity had been achieved by reaching the maximum amorphicity of the polymer. By comparing between the pure blend and the nanocomposite polymers, the most effective conductivity obtained from the polymer with the highest concentration of nanoparticle (PVC/PVDF/CuONPs5).
The average crystal size (D) of CuO NPs was calculated from XRD by Scherrer's formula: The average crystal size of CuO NPs was 42 nm.

FT-IR
IR data were determined by FTIR spectrophotometer. The samples signals appeared in from 4000 cm −1 to 500 cm −1 as Fig. 2. Each chemical structure has a distinctive fingerprint so FTIR analysis is a good choice for chemical structure identification. IR spectra assignments of pure PVC were reported as the following; CH 2 wagging vibration was obtained at 609 cm −1 [24]. C-Cl stretching  [20]. Trans CH wagging mode was cleared at 976 cm −1 . C-H rocking mode near Cl has been observed at 1200 cm −1 [26]. CH bending vibration mode was illustrated at 1427 cm −1 . CH 2 asymmetric stretching mode was obtained at 2908 cm −1 [29]. CH asymmetric stretching vibrational mode was obtained at 2982 cm −1 [30].
FT-IR spectrum for pure PVDF demonstrated the following bands; the bands appeared at 613, 763 and 976 cm −1 assigned to α form of PVDF. The β phase has been illustrated at 874 and 1180 cm −1 . The absorbance band at 874 cm −1 corresponded to the CH 2 rocking/CF 2 stretching of β phase of PVDF [31]. The bands at 1180 and 1068 cm −1 referred to the symmetrical stretching of single bond CF 2 groups. The band at 972 cm −1 referred to out of plane C-H bending [32]. Scissoring or in plane bending of CH 2 was obtained at 1401 cm −1 [33]. The band at 1454 cm −1 related to -CH 3 or -CH 2 stretching [34].
IR spectrum for PVC/PVDF blend without adding copper oxide nanoparticles showed; the bands overlapping of PVC with PVDF and not expected appearing new bands due to presence of the covalent bonds of PVC and PVDF. It is difficult to break these covalent bonds so no chemical interactions took place between PVC and PVDF.
FTIR spectra demonstrated the present of characteristic bands for the pure blend and the shift in some characteristic bands was due to the interaction between polymeric matrix and nanoparticles. When CuO nanoparticles were added, a change in intensity of the characteristic peaks in the fingerprint region was observed. Besides, a minor change was observed in the region 600-500 cm −1 due to the small concentration of CuO additives.

UV-Vis
Ultraviolet (UV) is an electromagnetic radiation form that comes from the sun. UV-Vis spectra are shorter than visible light and longer than X-rays. UV-Visible spectroscopy is an analytical technique used in the solid state mainly to confirm the formation of the complexes. The molecules could absorb ultraviolet or visible light. This absorption is compatible with the electrons excitation in the involved molecules. The occupied molecular orbitals with the lowest energy are called σ orbitals, and those at slightly higher energy are called π orbitals and non-bonding orbitals at still a higher energy. The highest energy orbitals are called σ * and π *. Figure 3 obtains the absorption spectrum of PVC/PVDF blend and PVC/PVDF/ CuONPs films.
The absorption edges attributed to the semicrystalline behavior of the prepared films have been obtained at approximately 250 nm in the spectra. As CuONPs content increased, absorption edge intensity decreased which is evidence for the reaction between all the components due to adding CuONPs. The absorption bands showed in the 220-238 nm range have been attributed to π → π* transition. The bands appeared in the range 430-520 nm belonging to copper nanoparticles [35].
The optical absorption coefficients α of the synthesized PVC/PVDF blend and PVC/PVDF/CuONPs films are illustrated in Fig. 4. It could be calculated by Beer-Lambert's formula [36]: By increasing the laser ablation time, it is noticed that with increasing hυ, optical absorption coefficients are increased.
The values of direct and indirect transitions were determined via plotting the relations between α 1/2 , α 2 and energy (E = hυ), respectively, and it is shown in Figs. 5, 6 and recorded in Table 1.   From Table 1, it was concluded that the values of indirect and direct optical bandgaps of PVC/PVDF composite were 4.18 eV and 3.56 eV, respectively. By raising the ratios of CuONPs, these values were reduced to 2.81 eV and 2.07 eV, respectively, as the laser ablation time was increased. This happened due to the chemical reaction between PVC/PVDF blend and CuONPs.
The refractive index (n) can be calculated by Dimitrov and Sakka equation [37] in terms of indirect energy bandgap and recorded in Table 1, Rising laser ablation time raised the values of (n) for PVC/PVDF/CuONPs samples owing to increasing the number of non-bridging oxygen bonds (NBO) bonds between copper oxide nanoparticles. Also, the refractive index increased by decreasing direct and indirect energy bandgap which confirmed the effect of the laser ablation time.

FE-SEM
SEM data illustrated the composition and morphology of the polymer. SEM photographs of PVC/PVDF blend without and with doped in different concentrations of CuONPs presented in Fig. 7. SEM photographs showed the change occurred in the blend surface after adding different concentration of CuONPs. Figures 7 (a, b) are the SEM micrographs of the pure PVC/PVDF (30/70) blend without adding any nanoparticles "CuNPs 0." It appeared soft, homogenous and coherent. Figures 7  (c-h) obtain the distribution of CuONPs in PVC/PVDF blend for the samples (c, d) CuONPs 1, (e, f) CuONPs 3 and (g, h) CuONPs 5. Figures 7 (c, d) are the micrographs of the polymer blend doped with the first concentration of CuONPs. They demonstrated appearing pores with random shape and size after adding CuONPs 1. Figures 7 (e, f) are the micrographs of the polymer blend with CuONPs 3. They showed that the pores became bigger and round and the number of pores decreased which could be due to increasing CuONPs concentration. Figures 7 (g, h) are micrographs of the polymer blend with CuONPs 5. They illustrated that the pores became closer by increasing the concentration of CuONPs. It is concluded that the number of pores was decreased by increasing CuONPs concentration while the size of the pores was increased.

Antibacterial activity
Antibacterial activity of PVC/PVDF/CuONPs prepared by PLAL in organic solvent "THF" was studied against four types of bacteria strains by determining minimal inhibitory concentrations (MIC). E. coli and P. aeruginosa were used as a twomodel for gram-negative bacteria and S. aureus and B. cereus were also used as a two-model for gram-positive bacteria. The antibacterial activity percent (%) was measured via the following equation [38,39]: % Activity index =

Zone of inhibition by test compound (diameter)
Zone of inhibition by standard (diameter) × 100.
The data in Table 2 showed the activity index of the prepared samples. The data revealed that the nanoparticles were active against the microorganisms under study. The inhibition of antibacterial growth depended on the concentration of nanoparticles. The interaction between the nanoparticle and the cell wall of bacteria resulted in a hole of cell wall of bacteria due to penetrating the ions into the cell wall attributing to the antibacterial activity of copper oxide nanoparticles. The antibacterial activity of PVC/PVDF/CuONPs5 as shown in Fig. 8  The results showed that the antibacterial activity increased gradually by increasing the concentration of nanoparticles [40]. The figure illustrated that S. aureus and B. cereus (gram-positive bacteria) were more affected by the concentration of PVC/PVDF/CuONPs than E. coli and P. aeruginosa (gram-negative . That could be referred to the stronger cell wall of gram-negative bacteria than gram-positive bacteria which prevent the ions from penetrating it. The interaction between the nanoparticle and the cell wall of bacteria resulted in a hole of cell wall of bacteria attributing to the antibacterial activity of the nanoparticles. It was observed that the gram-positive bacteria were more deeply affected by CuONPs than gram-negative bacteria.

Conclusions
PVC/PVDF blend doped with various ratios concentrations of CuONPs had been prepared by laser ablation technique. The obtained films had been examined via different routes to study the structural and morphological properties before and after adding nanoparticles. X-ray data illustrated the intensity of CuO nano-metal which was characterized with two new peaks located at 2θ = 26.25º and 38.41º, confirming that the interaction between CuO nano-metal and PVDF/PVC blend occurred in the amorphous area. FT-IR data showed the composition and chemical structures of the PVC/PVDF blend and demonstrated the possible interactions took place after adding CuONPs. UV-Vis data showed the effect of the laser ablation time and revealed the interaction between the blend and CuONPs and the peak at around 430-520 nm approved the doping by CuONPs. SEM images revealed the morphological changes happened to the surface due to adding CuONPs to PVC/PVDF blend. The images become rough with pores compared to the pure blend. The antibacterial activity test proved the antimicrobial effect of copper oxide nanoparticles against the tested organisms as the activity index was increased by increasing the concentration of the doped nanoparticles. These researches provide an easy, low cost and clean route for fabricating nanocomposite systems that can be used in clinical and biomedical applications such as wound healing applications.