PalladiumwrappedZnOnanocomposites synthesis through carrageenan were madeas bimetallic nanocomposites (C-Zn/Pd-Np) and their antibacterial, and anticancer efficacy were assessedunderin vitro condition.Further,they were physico-chemically characterized. The metal complex werecharacterized using a variety of physico-chemical techniques, including scanning electron microscopy (SEM), X-ray diffraction (XRD), and infrared spectroscopy (FT-IR). The bimetallic nanocomposites are more crystalline according to XRD analysis. IR spectra indicate faint multiple bands in the far infrared range because ofmetal-metal interactions and the absence of usual bands of acetate and nitrate groups. P. aeruginosa, S. aureus, B. subtilis andEscherichia coliwere used to test te antibacterial activity of the nanocomposites. The outcomes demonstrated significant antibacterial efficacy against all of the investigated pathogens. The antiproliferative effectiveness of the nanocomposites against A549 cancer cell types aresignificantly enhanced with anincreased concentration.It also resulted in cell cycle arrest, apoptosis, and necrosis found in bimetallic nanocomposites treated cells. In conclusion, (C-Zn/Pd-Np) has notable antibacterial action and is effective against at inhibiting the growth of lung cancer cells. According to this present study, C-Zn/Pd-Np may be useful for the effective clinical management of Human pathogensand lung cancer cells.
Research Article
Synthesis, characterization of C-Zn/Pd-Np as potential nanocomposites against Human Lung cancer cells (A549) and pathogenic microorganisms
https://doi.org/10.21203/rs.3.rs-3552644/v1
This work is licensed under a CC BY 4.0 License
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published 15 Jun, 2024
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Palladium
ZnO nanocomposites
Lung Cancer
Antimicrobial
Flowcytometry
A high molecular weight, water-soluble biopolymer and sulfated form of D-galactan, carrageenan is derived from many marine red algae species. Carrageenan has biological effects that have been demonstrated, including anticancer, antiviral, and anticoagulant effects. To determine the potential use carrageenan assessment is crucial now for various biological applications. To better understand the benefitconsequences of the nanoparticles, research on the toxicological impacts of nanoparticles on aquatic sources are required (Pacheco-Quito., et al., 2020).
A cost-effective and environmentally friendly method of creating nanoparticles is to use natural polymers. As a green stabilizer, numerous natural polymers are employed in the manufacturing of nanoparticles. An eco-friendly polymer called carrageenan is generated from a type of red seaweed. Due to its high viscosity and exceptional gelling properties, carrageenan serves valuable biological purposes. Carrageenan biological features, such as its anti-inflammation, antiviral, anticancer, immunomodulatory, antioxidant, and anticoagulant effects, may significantly alter how it is used in medicine (Bhardwaj et al., 2020).
From a scientific and technological perspective, bimetallic nanoparticles which are made of two different metals have attracted more interest than monometallic nanoparticles. The characteristics of the bimetallic nanoparticles are controlled by the constituent metals and their nanometric size. These are created by combining metallic nanoparticles with various architectural configurations. They truly give us the potential to enhance the metallic mixture's Plasmon absorption band's energy, giving us a flexible instrument for biosensing. These characteristics, which include special size-dependent optical, electrical, thermal, and catalytic effects, may be different from those of pure elemental particles. Just a decade ago, extensive research into bimetallic nanoparticles began (Dlamini et al., 2023).
For biomedical applications, zinc oxide (ZnO) has a low level of toxicity and strong biocompatibility. The US Food and Drug Administration has classified ZnO as Generally Recognized as Safe (GRAS). ZnO nanoparticles (NPs) have drawn a lot of attention in the last 10 years due to their potential use in the treatment of cancer and viruses, respectively. Additionally, recent studies have demonstrated the effectiveness of ZnO NPs against various types of bacteria. It is possible to create ZnO nanoparticles using a variety of chemical and physical processes. The use of hazardous substances that harm the environment makes the chemical and physical procedures less desirable. Recently, the easy, risk-free, environmentally friendly, and simple synthesis procedure of NPs employing biopolymers has been tremendously advantageous (Anjum et al., 2021).
One of the most transformative paths for the future is nanomedicine, which entails using nanotechnology's knowledge and technologies for illness detection, treatment, and prevention. Numerous cutting-edge research areas are very interested in nanometals(Patil et al., 2008). Metal nanoparticles, such as palladium (Pd), ruthenium (Ru), silver (Ag), platinum (Pt), and gold (Au) nanoparticles (NPs), vary in activity, chemical, biological, physical, optical, and electrical properties. Compared to many other metals, Pd has less negative effects and more potential for use in biological applications. Pd-NPs have been employed in Drug transporters, prodrug activators, antibacterial, anticancer, antifungal, and antioxidant applications(Ventola et al., 2012).
Strong antibacterial/antibiofilm activity and therapeutic benefits against cancer cells and drug-resistant bacteria were demonstrated by Pd-NPs. Additionally, Pd(0) shown excellent therapeutic efficacy for human ovarian cancer. Due to the substantial surface area of Pd-NPs show efficacy in a number of biological applications. Pd-NPs exhibited strong antibacterial, cytotoxic, and antiproliferative activity (Jayakumar et al., 2023). The objective of this study was to synthesize, characterize, and investigate the antibacterial and anti-proliferative properties of novel bimetallic (C-Zn/Pd-Np) composites.
Materials
Palladium chloride, Zinc nitrate, 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and annexin VFITC apoptosis detection kit were supplied by Merck (USA). Dulbecco's modified Eagle's medium (DMEM) and propidium iodide (PI) were purchased fromCambrex (New Jersey, USA) and (Tianjin Dingshengxin Chemical Industry Co., China), respectively. Human lung cancer cells (A549), were procured from NCCS, Pune.
Synthesis of C-Zn-Pd-Nps
The carrageenan was made using a previously documented method. For the production of C-Zn-Pd-Nps, 400 mL of precursor stock solutions of 1 mM PdCl2, 0.25 mM were produced. 100 mL of aq. carrageenan were added to these stock solutions one at a time to produce Zn-Pd (C-0.5) Zn-Pd (1) Zn-Pd (1:5) Zn-Pd (2), respectively. The reaction took place at pH 7, 100°C, and for two hours in a water bath that was stirring. C-Zn-Pd-Nps form when dark brown colour is present. To get rid of the superfluous precursor and carrageenan, the NPs solution was centrifuged (15,000 rpm, 5 min), and the pellet that was produced was washed three times. The resultant powder was employed for tests on colour toxicity and degradation, and the pellets obtained were dried at a temperature of 70°C (Yew et al., 2020).
Cell culture
The National Centre for Cell Sciences (NCCS), located in Pune, India, was where the A-549 lung cancer cells were purchased. Then the cells were sustained in Dulbecco's modified eagles medium (DMEM) enhanced by means of 2mM l-glutamine and adjusted basal salt solution(BSS) acclimated to encompass1.5 g/L Na2CO3,1.5 g/L glucose,2 mM l-glutamine, 1 mM sodium pyruvate, 10 mM (4-(2-hydroxyethyl)-piperazineethane sulfonic acid) (HEPES), 0.1 mM non-essential amino acids and 10% foetal bovine serum (GIBCO, USA). In agreement with 1 mL/L, the antibiotics penicillin and streptomycin (100 IU/100g) were utilized. The cells are then maintained at 37°C with 5% CO2(Kavithaa et al., 2016).
MTT
The inhibitory concentration (IC50) of the synthesized nanocomplex was determined using MTT [3-(4, 5-dimethylthiazol-2-yl)- 2, 5-diphenyltetrazolium bromide] analysis. To achieve 80% confluence of the cultivated cells, A-549 and IMR-90 cells were cultured (1x104 cells/well) in a 96-well plate for 48 hours. After changing the media to one containing various concentrations of nanocomplex (0-100 g/ml), the cells were incubated for 24 h. The media was then removed, and each well received 100L of the MTT solution before being incubated at 37°C for 4h. After removing the supernatant, each well received 50 L of DMSO, which was then given a 10-minute incubation period to help dissolve the formazan crystals that had developed during the process. Using an ELISA multi-well plate reader (ThermoMultiskan EX, USA), the optical density of the samples was then calculated to be 570nm. The following formula was applied with the acquired OD values to estimate the percent cytotoxicity (Kavithaa et al., 2016).
% of viability = OD value of experimental sample/OD value of experimental control ×100
Morphological analysis
The chosen Human MDA-MB-231 cancer cells were cultured on cover slips (1x105 cells/cover slip), then exposed to -Santalol, chitosan NPs, and Sn-CNP (IC50), before being fixed in a 3:1 ethanol:acetic acid solution. Each experimental group had three monolayers that were micrographed. Additionally, the cover slips were carefully mounted on glass slides for the morphometric analysis, which was then focused using Nikon (Japan) bright field inverted light microscopy at a 40x magnification.
Analysis of apoptotic cell death using fluorescent microscopy
On clean microscope cover slips, 1 L of a dye solution (100 mg/mL acridine orange (AO) and 100 mg/mL ethidium bromide (EtBr) in distilled water) was combined with 0.9 mL of cell suspension (1 105 cells/mL). Cancer cells that had previously undergone treatment were gathered, cleaned with PBS (pH 7.2), and stained with 10 mL of AO/EtBr. The cells were given a 2-minute incubation period, then were washed twice in PBS for five minutes each. The cells were then examined using a fluorescence microscope (Nikon Eclipse, Inc., Japan) with a 400x magnification and an excitation filter at 580nm. On glass coverslips in a 6-well plate, the cells were similarly treated for 24 hours with -Santalol, chitosan NPs, and Sn-CNP (IC50). The fixed cells were permeabilised with 0.2% triton X-100 (50 l) for 10 min at room temperature, and with the aid of positioning the coverslip over the cells, which then allows equal dispersion of the stain, they were incubated for 3 min with 10 l of DAPI. Under a fluorescent microscope (Nikon Eclipse, Inc., Japan), the cells were examined.
Cell cycle evaluation
The A-549 lung cancer cells (1x105 cells/ml) were cultured for 48 hours with and without different amounts of synthesized nanocomplex to perform the cell cycle study. The cells were then gathered and suspended for 30 minutes at room temperature in the dark in 500 l of phosphate buffer saline containing propidium iodide (PI) with 0.025 mg and RNase with 50g. After that, BD Facsverse USA performed flow cytometry on the samples.
Apoptosis detection using an Annexin-V/FITC-Flow Cytometer
One 105 A-549 lung cancer cells were grown in a six-well plate. Chitosan-oleanolic acid nanocomplex, synthesized at its IC50concentration, was used in place of the medium. After about 24 hours of incubation, the cells were collected using trypsin, rinsed in PBS before being fixed in 70% ethanol and kept at -20°C for an hour. The cell's nuclear DNA was stained using Annexin V-FITC, which was then let to sit at 37°C for 30 minutes. The BD FACS Verse flow cytometer was then used to perform flow cytometry in duplicate. 10,000 times of each sample were collected, and CellQuest and Modifit recorded and evaluated the fluorescence intensity.
Evaluation of Antimicrobial Activity of metallic-NPs
The produced materials were evaluated for antibacterial activity against P. aeruginosa, S. aureus, B. subtilis, and Escherichia coli. In wells created in the solid media (bacteria "Nutrient Agar," fungi Potato dextrose Agar) that had previously been inoculated by distributing a freshly prepared inoculum throughout the entire agar, various concentrations of metallic Zn-Pd-NPs composite were added. The yeast and bacteria were cultivated for 48 hours at 30°C while the fungi were incubated for 6 days at 26°C. After looking over the plates, the percentage of inhibition was determined as follows:
Inhibition growth percentage = MC– MT /MC x 100 (1), Where MCis the diameter of inhibition zone around control: MT is the diameter of inhibition zone around treated material (El-Sheekh et al., 2014).Minimum Inhibitory concentrations (MICs) were determined. All the experiments were repeated three times.
Statistical analysis
Data were analyzed using the SPSS software. Results are presented as mean ± SD. Differences among various groups were evaluated by conducting one-way analysis of variance (ANOVA); P < 0.05 were considered statistically significant groups.
Figure 1 shows the absorption spectrum of Zn-Pd-NPs nanocomposites. The absorption spectrum of Zn-Pd-NPs showed the absorption peak at 318nm. Usually for zinc metal nanoconjugates the absorption spectra ranges between 200 to 380nm. Here the UV visible spectrum of different concentration of carrageenannot showing any significant difference between each other, corresponding to composite formation. It further confirms that Pd is firmly attached to the surface of Zinc particles formed as bimetallic composites.
IR spectra
The IR spectrum of the synthesized composite (Fig. 2) showed absence of the characteristic bands assigned to M-O or M-C bonds in the range 500–650 cm − 1 which indicates the presence of oxides or carbides materials, respectively in the chemical structure of the synthesized composite. Also, the spectrum showed no bands in the ranges 1400–2300 cm − 1 and 2382–2991 cm − 1 assigned to bending and stretching vibrations of H2O molecules, respectively, indicating that no chemical bonded or physical adsorbed water molecules which confirmed the anhydrous structure of the metallic composite. The multiple weak bands observed at 540–650 cm − 1 are due to metal-metal interaction.
The peak at 2991cm− 1 indicates the N-H stretching and 841cm− 1 corresponds to the pyranose ring vibration of zinc nanoparticles. The peak by 2382 cm− 1 indicates the presence of C-H stretching, the peak at 1603 cm− 1 represents in-plane bending vibration of N-H molecule for Pd molecules. Further the peak at 3490 cm− 1 represents N-H2 stretching, 3511 cm− 1 corresponds to O-H stretching, represents the presence of C-N stretching and the peak at 825 cm− 1 indicates out-of plane bending vibration of N-H molecule for -Zn/Pd-Nps as shown in the figure.
XRD
XRD was used to confirm the structure of the bimetallic nanocomposites (Fig. 3). The positions of high intensity peaks refer to the formation of pure C-Zn/Pd-Nps. Peaks appeared at 2θ = 40.11, 46.15, and 68.31 which correspond to (111), (200), and (220), respectively, and are consistent with a conventional Pd-NPs phase pattern. Ag NPs showed typical peaks corresponding to the (111), (200), (220), and (311) planes at 2θ values of 38.14, 44.35, 65.00, and 77.5, respectively. For the hexagonal structure of Zn-NPs, five different diffraction peaks were found and indexed with the planes (100), (002), (101), (102), and (110) at 38.42°, 42.12°, 43.98°, 58.32°, and 69.42°, respectively. The size of the crystals was determined using the Scherrer equation.
Morphological SEMstudy
The morphology of the synthesized composite was examined using SEM at different magnifications. As depicted in Fig. 4, the synthesized NPs exhibited highly homogenous structure. The size and shape of the synthesized composite were assessed by SEM. It appears spherical in shape and the electron micrographs exhibit the presence of spherical nanomaterials of the synthesized materials which was depicted in the figure. This confirmed that the particles synthesized were zinc crafted with Pd molecules.
MTT assay
A549 cells were exposed to varied doses of C-Zn/Pd-Nps (10, 50, 100, and 200µg/mL) to determine how they affected the viability and cytotoxicity of selected cells (Fig. 5). The fact that variations in C-Zn/Pd-Npsconcentration had no impact on cell viability suggests that the C-Zn/Pd-Npsdid not have any cytotoxic effects on human lung cancer cells.
Cell morphology analysis
The A549 morphological changes in the absence and presence of C-Zn/Pd-Npat various concentrations are shown in Fig. 6. It could be observed from Fig. that control cells did not show any remarkable changes on their morphology. However, in the presence of C-Zn/Pd-Npthe cells show the improved cell shrinkage, membrane blebbing and forms floating cells in a dose-dependent manner. It is well accepted that cytological investigations elucidate the antiproliferative effect routed through membrane blebbing, membrane instability and distressing the cytoskeleton of the cells by the C-Zn/Pd-Np.
Acridine Orange Ethidium Bromide (AO/EtBr) staining
The percentage of dead cells by apoptosis was checked by simple staining techniques such as AO/EtBr under fluorescent microscope. Based on the concentration of the C-Zn/Pd-Npsthe percentage of apoptotic cells started increasing. The concentration of C-Zn/Pd-Npswas taken in 3 different concentrations such as 5µg/ml, 10µg/ml, 25µg/ml and an untreated group as control. The apoptotic cells alone were stained using AO/EtBr stain and were represented in percentage as shown in the Fig. 7.
DAPI (4′,6-diamidino-2-phenylindole)staining
DAPI was used to stain the nucleic acid which was visible under fluorescence microscope. It is mainly used to determine the chromosome condensation and DNA damage. The concentration of C-Zn/Pd-Npswas taken in 3 different concentrations such as 5µg/ml, 10µg/ml, 25µg/ml and an untreated group as control. When the lung cancer cell line A-549 was treated with various concentrations of the synthesized C-Zn/Pd-Nps, the DNA damaged apoptotic cells was stained by the dye DAPI and was represented in percentage as shown in the Fig. 8.
Cell cycle analysis
The cell cycle analysis was done by using flow cytometry to check at which stage the cells got arrested. In control group (untreated group) the cells were evenly distributed in G0-G1, S and G2M phase. The lung cancer cells were treated with different concentration of 5, 10 and 25µg/ml of C-Zn/Pd-Nps. The cells got arrested atG2M phase for the concentrations such as 5µg/ml and 25µg/ml whereas for the concentration 50µg/ml the cells got arrested at S phase. This indicated that the when the concentration got increased it arrested the synthesis of lung cancer cells when treated with C-Zn/Pd-Nps as shown in Fig. 9.
Apoptosis through flowcytometry
In order to measure the percentage of dead cells using Annexin V/FITC, flow cytometry was performed. In control group the percentage of live cells remains the same whereas the percentage of dead cells increased markedly on the late apoptotic stage based on the concentration of C-Zn/Pd-Nps. The percentage of live cells decreased significantly for the same concentration of 5, 10 and 25µg/ml which was represented in the Fig. 10.
Antimicrobial activity
P. aeruginosa, S. aureus, B. subtilis andEscherichia coli were chosen to evaluate the antimicrobial activity of the C-Zn/Pd-Nps. The minimum inhibitory concentration (MIC) was determined using four different dosages of nanomaterials (0.0125, 0.025, 0.05, and 0.1 mg/mL) (Fig. and Table) and was found to be 0.0125 mg/mL. For comparison, antimicrobial experiments were carried out for each single metal as depicted in Table 2 and Fig. 11. The findings indicated that C-Zn/Pd-Nps demonstrated high antimicrobial activity in comparison to different isolate species of bacteria.
Non-small cell lung cancer (NSCLC), which accounts for 85% of occurrences of lung cancer and is the second biggest cause of cancer-related death worldwide, is the most common type. The main risk factors for developing NSCLC are genetic predisposition, way of life, environmental variables, and smoking (Molina et al., 2008). There is a need for alternative therapy because the current treatment options for NSCLC include surgical excision and chemotherapy, both of which suffer from serious drawbacks like side effects caused by poor target specificity, recurrence, and drug degradation before it reaches the target site (Boffetta et al., 2004). Noble metal nanoparticle-based therapy, in particular, has recently become a focus of research due to its potential as an imaging and diagnostic tool for cancer therapy and its compact structure with large surface area, targeted drug administration, improved stability, sustainable drug release, and enhanced EPR impact.
We investigated the anticancer potential of bimetallic nanoparticles as a result of the success of nanoformulation-based therapies for advanced tumours, such as albumin-bound paclitaxel and noble metal nanoparticles (Hwang et al., 2003; Li et al., 2004). Bimetallic nanoparticles created by conventional methods had a negative impact on the environment, which drew researchers' attention to natural resources. Unique physiochemical characteristics and biocompatibility make biogenic nanoparticles ideal for biological applications [Bailey-Wilson et al., 2004]. In the current study, bimetallic nanoparticles C-Zn/Pd-Np were successfully synthesized from a biogenic source, bio-polymer carrageenan, and their antibacterial and anticancer properties were evaluated in relation to lung cancer cell lines.
The creation of C-Zn/Pd-Np is visually indicated by a change in colour. This was validated by the UV-Visible absorption spectrum, the most used technique for identifying individual noble metal nanoparticles based on their surface Plasmon resonance (Thorgeirsson et al., 2008).PdNPs encapsulation dampens the SPR peak of Zn/Pd, as reported previously (Amos et al., 2008), the production of Zn/Pd bimetallic nanoparticles is confirmed by the absence of a distinct surface Plasmon resonance peak (SPR) for Zn/Pd NPs.
A crucial role in the production of bimetallic Zn/Pd NPs as a reductant and capping agent is played by glycosides, alkaloids, flavonoids, phenols, carbohydrates, steroids, saponins, reducing sugars, proteins, and terpenoids, according to phytochemical analyses of the biopolymer carrageenan. FTIR data showed the presence of functional groups related to sugars, polyphenols, and tannins, which may be responsible for the synthesis of C-Zn/Pd-Np as a reducing and stabilizing agent. As mentioned in earlier papers, the XRD pattern closely resembles the standard patterns of silver (JCPDS No. 04–0783) and palladium (JCPDS No. 87–0641), showing the synthesis of C-Zn/Pd-Np with face-centered cubic crystalline structure.
FeSEM and HRTEM studies further supported the XRD findings by confirming the presence of uniformly shaped C-Zn/Pd-Np with an average size of 20 nm (size range 2–40 nm). For cellular uptake and the penetration of biological barriers, DLS and ZP measurements provide an idea of the size and charge of the nanoparticles.
The most used method to measure cellular toxicity and cytolysis, the MTT and LDH assay, was utilized to examine the anticancer potential of Zn/Pd bimetallic nanoparticles against A549 cells. The results of MTT and LDH experiments, along with phase contrast microscopy examination, support the hypothesis that C-Zn/Pd-Np cause apoptosis by impairing cell membrane integrity and reducing cell proliferation. Normal cellular biological processes like cell division and death depend heavily on reactive oxygen species. However, the level of ROS in malignant cells is higher than in normal cells, where it might have a double-edged effect depending on its concentration. A moderate quantity of ROS accelerates the development of cancer, while an excessively high level of ROS activates the apoptotic pathway, which leading to cell death.
The growth inhibitory property of C-Zn/Pd-Np was seen at a concentration of 23µg/mL, resulting in a 50% reduction in growth. Priya et al., 2020 also documented comparable results, noting the dose-dependent anticancer effects of biogenic CS-Ag NPs on hepatocellular carcinoma cells. Metal nanoparticles were generated using biogenic method and encapsulated with polymer shown significant anti-cancer effect against HeLa cells [Sukumar et., 2022]. According to the paper, the anticancer characteristics of Ag NPs were found to be contingent upon the morphology, size, and reducing agents of the nanoparticles.
Numerous apoptosis-based studies utilizing fluorescence dyes have been conducted to assess the cytotoxic effect of C-Zn/Pd-Np on A549 cells because the apoptotic route is the main mode of action in most chemotherapy agents to kill cancer cells (Yamato et al., 2001). A549 cells treated with C-Zn/Pd-Np demonstrated apoptosis-mediated cell death as a result of AO/EtBr dual labeling. A549 cells treated with Zn/Pd NPs showed DNA strand breakage, which was further confirmed by the findings of the DAPI and comet analysis. Overall, the findings show that AgPd bimetallic nanoparticles have the ability to fight cancer by boosting ROS production through respiratory chain malfunction, which results in the loss of MMP and ROS-mediated DNA damage, activating the apoptotic cascade pathway, and ultimately killing cells.
The utilization of a fluorescence microscope allowed for the observation of apoptosis in A549 cells that were treated with the generated C-Zn/Pd-Np. The control cells had a green hue, while the treated cells exhibited a red hue, so providing evidence for the inductive nature of C-Zn/Pd-Np. The manifestation of a crimson tint signified the existence of apoptotic entities and exhibited cellular contraction along with membrane blebbing. In the majority of instances, the occurrence of cell death is accompanied by many morphological and biochemical changes. These changes include cell shrinkage, membrane blebbing, disruption of membrane integrity, fragmentation of the nucleus, and condensation of the cytoplasm. These alterations collectively play a role in initiating the apoptotic cascade, leading to cell demise. This biological mechanism was accountable for the demise of cellular entities [Aswini et al., 2021; Kannan et al., 2022]. The ability of nanoparticles (NPs) to induce apoptosis is contingent upon the permeability of metal (Zn) ions into cancer cells. This permeability facilitates cell damage, DNA breakage, and subsequently triggers apoptosis [Bharathi et al., 2019].
C-Zn/Pd-Np exhibit strong antibacterial activity against E.coli and P. aeruginosa, according to antimicrobial investigations. C-Zn/Pd-Np antimicrobial activity may result from the synergistic effects of C-Zn/Pd-Np, and the mechanism of action is linked to the interaction of nanoparticles with cell membranes, which alters their fluidity and releases intracellular content, causes cell death, or induces the formation of free radicals, which results in oxidative stress-mediated cell death, as reported for Zn and Pd NPs (Alen et al., 2006). Size, shape, and concentration of nanoparticles all affect their ability to have an antimicrobial effect. Particles smaller than 25 nm have been found to be extremely hazardous to bacterial cells.The average size of C-Zn/Pd-Np in the current investigation was found to be 20 nm, which may be the cause of its antibacterial action. Zn/Pd NPs with strong antibacterial properties can be used in medical devices like catheters, probes, dressings, and more to help prevent the colonization of pathogen microorganisms.
With regard to therapy and diagnostics, nanomaterials have gained significant attention over the past 20 years. This calls for the safety assessment of nanoparticles in order to achieve biocompatibility and desired activity. Human PBMC and RBC, the most commonly used model system to evaluate the immunotoxicity and blood cell compatibility under in vitro conditions, were used in the present study to assess the safety aspects of AgPd bimetallic nanoparticles as an alternative to animal studies (Kyriakides et al., 2021). In all doses tested, AgPd NPs showed no cytotoxic effects and hemolytic activity in PBMC and RBC, demonstrating their biocompatibility.
The current study used a biopolymer carrageenanused to synthesizeC-Zn/Pd-Npsin a biogenic and economically advantageous manner. The as-prepared C-Zn/Pd-Npswere discovered to be crystalline, stable, and extremely small (20–70 nm). Against the A549 cell line, C-Zn/Pd-Npsexhibited dose-dependent antiproliferative activity. The mechanism's clarification showed that C-Zn/Pd-Nps activate the apoptotic signals, which in turn activated the mitochondria's intrinsic apoptotic pathway. C-Zn/Pd-Npsalso demonstrated antibacterial action against bacterial strains. Preclinical investigations on the molecular mechanism of action and other applications of C-Zn/Pd-Npsin the treatment of infectious and cancerous diseases need to be conducted.
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Author Contributions
All authors contributed to the study conception and design. Initial ideology of the study was given by Niveditha dhoraiand Rajendran Venkatachalam. Material preparation, Biochemical and Staining assays were performed by Niveditha dhoraiand Suriya prabha. Nanoparticles characterization and analysis was done by Kosiha Arumugam, Giriraj Kalaiarasi. The initial draft of the manuscript was written by Niveditha dhoraiand Suriya prabha and overviewed by Rajendran Venkatachalam. The final manuscript was read and approved by all authors.
Funding
No fund supported for this work.
Competing interests
The authors declare no competing interests.
Availability of data and materials
All data generated and analysed are included in the submitted manuscript and supplementary materials.
Acknowledgement
The authors thank The Management of Dr.N.G.P College of Arts & Science, and PSGR Krishanammal College for Women, Coimbatore, for conducting this research.
Financial interests
The authors have no relevant financial or non-financial interests to disclose.
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Table .1. Cytotoxic activity of C-Zn/Pd-Np against Human lung cancer cells
IC50 – Values of respective sample (at 24 hrs)
Table; 2. Antibacterial activity of C-Zn/Pd-Np against human microbial pathogens
|
S.No |
Strain |
C-Zn/Pd-Np Zone of inhibition level (mm) |
||||
|
|
Concentration |
+Control |
5 |
10 |
15 |
20 |
|
1 |
E. coli |
20 |
11 |
13 |
14 |
17 |
|
2 |
P. aeruginosa |
23 |
13 |
14 |
16 |
18 |
|
3 |
S. aureus |
20 |
12 |
13 |
15 |
17 |
|
4 |
B. subtilis |
21 |
11 |
12 |
13 |
16 |