Aloe vera - mediated silver-selenium doped fucoidan nanocomposites synthesis and their multi-faceted biological evaluation of antimicrobial, antioxidant and cytotoxicity activity

ABSTRACT

The synthesized Ag-SeO2-doped (Fu) nanocomposites are novel therapeutic approaches against multidrug-resistant bacteria. The present study explores the synthesis, characterization, and evaluation of the antibacterial, antioxidant, and in vivo cytotoxic activities of novel nanocomposites. The distinguishing characteristics of Ag-Se/Fu nanocomposites were revealed through the use of UV-visible spectroscopy, EDX, and FTIR analysis. The result of the SEM micrograph shows that Ag-Se NPs and Fu nanocomposites, with average nanoparticle diameters that vary from 97.28 ± 7.14 nm, display spherical shapes. It was found that the potency of antioxidant and antimicrobial agents against Enterococcus faecalis, Klebsiella pneumonia, Escherichia coli, and Methicillin-resistant Staphylococcus aureus (MRSA). Moreover, the cytotoxicity assay indicates the nanocomposite has shown a survival rate of 98% in zebrafish embryos at 150 mg/mL. Compared to the control group, the embryos from the nanocomposite group had normal morphology, which is evidence of its safety and effectiveness. The results of this research reveal a promising strategy for multidrug-resistant bacteria with less cytotoxicity.

KEYWORDS:

• Green synthesis
• Ag-Se doped fucoidan
• antimicrobial
• antioxidant
• cytotoxicity

Introduction

The field of nanotechnology, which is expanding, can be used to develop nanoscale structures. The creation of particles with diameters between one and one hundred nanometres (nm) is the focus of nanoproducts. Nanotechnology in living things is a fusion of wet, dry, and digital nanotechnologies. Biological components like membranes, organs, and enzymes are included in wet nanotechnology. Dry nanotechnology focuses on physical, chemical, and surface science as well as the creation of inorganic materials like silicon and carbon. Computational nanotechnology includes the modelling and simulation of intricate nanometre-scale structures [1]. Herbal remedies for wound care or therapy include debridement, disinfection, and the creation of a moist environment that promotes the emergence of the proper natural healing climate. Numerous plants are used in folklore cultures to treat burns, cuts, and other injuries [2]. The green synthesis technique makes it safer and more affordable to produce nanoparticles in large quantities. Aloe vera has been used as a therapeutic agent since ancient times. Compared to other medicinal plants, the biologically significant substances found in Aloe vera plant extract can be used to treat a wide range of disorders. Researchers have found that high-performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS) are good ways to separate the bioactive compounds from the Aloe vera plant extract [3]. Aloe vera has many natural bioactive substances that are both simple and complex. These include foaming agents, anthraquinones, aqueous polysaccharides, glycosides, pyrocatechol, plant-derived alcohol, and oleic acid. Aloe vera leaves, in both alcoholic and water-soluble forms, exhibit stronger antibacterial activity [4]. For the purpose to synthesising various metallic nanomaterials, the Aloe vera extract was chosen as a reducing and stabilising agent because of its low-cost, ecologically friendly, and chemically reduced properties. The ability of silver to exert excellent biological potential, including antifungal, antibacterial, antiviral, anti-infectious, wound healing, and anti-inflammatory characteristics at low concentrations, has led to the term ‘dynamic’ being applied to it [5]. Due to selenium’s crucial role in human health, interest in it has increased recently [6]. Selenium is essential for the metabolism of thyroid hormones and other vital metabolic processes, as well as for healthy immune system functioning. By being incorporated into antioxidant enzymes, it also stops cellular damage brought on by free radicals. Se NPs deficiency has been scientifically linked to a number of significant ailments, including cancer, heart disease, and contagious infections [7]. Fucoidan is a type of sulphated polysaccharide that has several biological uses, like antiviral, anticancer, anti-proliferative, anti-inflammatory, immunomodulator, anti-metastatic, anticoagulant, and other beneficial qualities [8]. Chronic abscess, including pressure sores, venous insufficient ulcers, and diabetic foot ulcers, doesn’t conform to the typical time sequence of cellular and molecular events that result in the healing of a healthy acute lesion [9]. To increase the incidence of multi-drug resistant pathogens in chronic wound infections, there is an immediate requirement to create noble therapeutic substances to address chronic wound infections [10]. For this reason, we use novel technology to synthesise nanoparticles to treat this infection. The present study deals with the synthesis of Ag-Se nanoparticles through a one-pot synthesis method. Through the green synthesis process, the cell-free extract of the plant contains phytochemicals. Then the fucoidan was doped with synthesised Ag-Se nanoparticles using the burette method, and the purified Ag-Se doped with fucoidan was further characterised with UV-Vis spectrometry, FT-IR, SEM, EDX, and XRD. The Ag-Se doped fucoidan nanocomposites furthermore use biomedical applications, including antimicrobial, antioxidant efficacy, and cytotoxicity studies.

Materials and methods

Silver nitrate (AgNO3) and selenium dioxide (SeO2) were purchased from Loba Chemie Pvt. Ltd. All other chemicals are purchased in analytical grade from Himedia, India. Four different bacterial strains were obtained from the Department of Microbiology at Saveetha Medical College.

Sample collection

Aloe vera was collected in and around Poonamallee, Chennai, Tamil Nadu, and the taxonomic assessment of the samples was authenticated by Dr. N. Siva, Assistant Professor, Department of Botany, Raja Doraisingam Government Arts College Sivagangai, TamilNadu.

Preparation of aqueous leaf extract

The Aloe vera slices were rinsed three times in double distilled water and air-dried. The dried sample was ground into powder form using a mechanical grinder. 10 grams of powdered leaves were added to 200 ml of double distilled water and autoclaved. Subsequently, the solution was purified through the Whatman No. 1 paper, and the resulting extract was preserved at a temperature of 4°C [11].

Synthesis of Ag-Se nanoparticles

Aqueous extract was taken in the conical flask, and 25 mm of silver nitrate (AgNO3) was taken in the burette along with 25 mm of selenium dioxide (SeO2) precursors that were titrated using the dropwise titration method and adjusted to a pH level of 8.5 with sodium hydroxide NaOH. In addition, it was kept in a shaker for overnight, and the colour changes were noted before and after 24 hours of shaking. The hue of the plant extract shifted from a deep black to a shade of brown. Subsequently, the reaction mixture underwent centrifugation at a speed of 4,500 rpm for a duration of 30 minutes. After this step, the resulting solid residue was gathered and subjected to a wash using freshly distilled water, followed by centrifugation at the same speed for an additional 30 minutes. After the centrifugation process, the pellet was transferred to a plate and kept in the hot air oven for a period of 24 hours. The pellets were ground into a powder form using a mortar and pestle, and this powder was then preserved for subsequent characterization and application investigations [12].

Formulation of Ag-Se nanoparticles doped fucoidan nanocomposites

The 300 mg of Ag-Se nanoparticles were weighed and mixed with 50 ml of distilled water and 1 mm of EDTA solution. Furthermore, 50 ml of distilled water mixed with 150 mg of fucoidan was added to the solution of Ag-Se using the burette method through a magnetic stirrer. The pH of the samples was modified through the addition of sodium hydroxide (NaOH). Then, it was kept in a shaker overnight. The pellets were subjected to centrifugation at 4,500 revolutions per minute (rpm) for a duration of 15 minutes. Following centrifugation, the resulting pellets were carefully gathered and left to air-dry naturally at room temperature for a period of 24 hours. The dried Ag-Se/Fu nanocomposites were scrapped out and used for further study [13].

Characterization study

Size and shape are two important factors investigated in the characterization of nanoparticles. The UV-visible spectrum of Ag-Se/Fu nanocomposites was collected on a spectrophotometer (JascoV-730) at 200–600 nm to estimate the absorbance maximum [14]. An investigation employing FTIR was conducted within the spectral range of 4000–500 cm^{− 1}, utilising a Bruker Fourier transform infrared spectrophotometer. This study aimed to explore the functions of various phytonutrients and biomolecules in their capacity as agents for capping, and stabilising during the synthesis process of Ag-Se/Fu nanoparticles [15]. Ag-Se/Fu nanoparticles were characterised using a diffractometer with X-rays (Malvern Panalytical) operating at a voltage level of 45 kV and an output power of 35 mA [16]. The scanning electron microscope detects secondary electrons generated from the sample when it interacts with the electron beam. JEOL JSM-IT800 at 1.00 kV examined the surface morphology and structural analyses of Ag-Se/Fu nanocomposite at Saveetha Dental College. Researchers have employed EDX spectroscopy to detect the related elemental composition of metal nanoparticles to confirm appropriate synthesis and purity [17].

Agar well diffusion test

The well diffusion method was used to test the antibacterial activity of the Ag-Se nanoparticles and Ag-Se/Fu nanocomposite materials made from Aloe vera. The antibacterial assessment of the biologically synthesised Ag-Se as well as Ag-Se/Fu nanocomposites towards Klebsiella pneumonia, Escherichia coli (gram -ve) and Enterococcus faecalis, and Methicillin-resistant Staphylococcus aureus (gram +ve) bacterial species involved measuring the zone of inhibition (ZOI) and determining the minimum inhibition concentration (MIC). The experiment was performed by using sterile Muller Hinton Agar (MHA) plates. 300 ml of Muller Hinton agar was prepared and kept in an autoclave at 121°C for 20 minutes. Sterilised Muller Hinton media was poured into sterile Petri plates and left to solidify. After solidification, an aseptic procedure was followed to create a hole in the media using a sterilised tip. The bacterial strains that were prepared were applied to the agar plate using a sterilised cotton swab. The various volumes of Ag-Se/Fu nanocomposites (20–100 µL) are desired concentrations introduced into the well. In the study, positive control (Streptomycin) and negative control (DMSO) were employed, with 30 μl of each added to the respective wells. Subsequently, the plates were placed in an incubator at 37°C for 24 hours. Following incubation, the Ag-Se/Fu nanocomposites diffused within the agar medium and exhibited inhibitory effects on the tested microbial strain. The observation of inhibition zones was made after a 24-hour incubation period, providing valuable insights into the antimicrobial properties of the nanocomposites [18].

Minimum inhibitory concentration (MIC)

Microorganisms that were used such as Klebsiella pneumonia, Escherichia coli, as Gram -ve bacterial species and Enterococcus faecalis, Methicillin-resistant Staphylococcus aureus as Gram +ve bacterial species. Additionally, Streptomycin served as the positive control in the study. The Minimum Inhibitory Concentration (MIC) of the fabricated Ag-Se/Fu nanocomposites was assessed using the micro-dilution method. The MIC experiment was conducted using a 96-microplate, with each test performed in duplicate. The nanoparticles, obtained at a concentration of 10 mg/mL, were prepared in wells with a volume of 100 mL [19].

Establishment for experimentation on 96 well-plates

In a 96-well round-bottom microtiter plate, 200 µl of MH broth was added to each well in Column 1. Columns 2 through 10 were filled with 100 µl of MHB broth. In Column 11, 200 µl of diluted standardized inoculums was introduced, and Column 12 received 200 µl of the medium broth, serving as a sterility control for monitoring purposes. The various sample concentrations were obtained by performing successive double serial dilutions ranging from columns 10 to 1. Subsequently, a multichannel pipette was employed to transfer and thoroughly blend the samples from column 1 to 9, ensuring that each well received 100 µl of the sample. Following this, a standardized suspension of microorganisms was diluted by a factor of 1:100 in MHB broth. Next, approximately 50 µl of the bacterial suspension with an adjusted optical density (OD 600) was introduced into all wells containing the samples as well as the control wells, leading to an approximate bacterial concentration of 5 × 10⁵ colony-forming units per millilitre (CFU ml⁻¹). The duration required for the preparation and addition of the OD-adjusted bacteria did not exceed 15 minutes. Following a 24-hour incubation period at 37°C, Resazurin solution (15 mg/ml) was introduced into all wells, with each well receiving 30 µl of the solution. Additionally, the samples were incubated for duration of 30 minutes to assess any alterations in colour. After the incubation period concluded, columns where no colour change occurred (indicating that the blue resazurin colour remained unaltered) were categorized as having values above the MIC (Minimum Inhibitory Concentration). Metabolically active cells trigger a transformation of resazurin from its initial purple-blue form into resorufin, which assumes a pinkish or colourless appearance. The Minimum Bacterial Concentration (MBC) was established by directly plating the contents of wells with concentrations exceeding the Minimum Inhibitory Concentration (MIC) value. The MBC value was ascertained when no colonies were observed to grow from the contents plated directly from the wells. Furthermore, in cases where the well contents exhibited signs of inhibiting bacterial growth, serial dilutions were carried out to determine the endpoint at which bacteria were effectively killed, as elaborated in the results section.

Antioxidant activity

The synthesized Ag-Se/Fu nanocomposites studied the antioxidant efficacy and scavenging activity using 2,2diphenyl -1-picrylhydrazyl DPPH assay [20]. In this procedure, distinct volumes of Ag-Se/Fu at varying concentrations ranging from 0 to 500 mg/mL were individually prepared, each containing 2 mL and the 1 mL of 1.9 mg DPPH (2,2diphenyl -1-picrylhydrazyl) solution was added to the sample and also 800 µl of tris-HCL buffer added to the mixer. Subsequently, the falcons were positioned upright and maintained in darkness for duration of 60 minutes. After this period, the absorbance reading was taken at a wavelength of 517 nm. The percentage of DPPH radical inhibition was determined utilizing the following formula.

$\%\mathrm{of}\mathrm{radicals}=\frac{(\mathrm{Abs}\mathrm{control}-\mathrm{Abs}\mathrm{sample})}{(\mathrm{Abs}\mathrm{control})}\times 100$

Cytotoxicity study of zebra fish

Adult zebrafish, aged approximately five months, were procured from the NSK fish aquarium located in Chennai, Tamil Nadu, India. This experiment conduct with IAEC Guidelines and approval number: (BRULAC/SDCH/SIMATS/IAEC/06–2023/15O). Upon acquisition, they underwent a one-month acclimatization period in our laboratory facilities. During this period, the zebrafish were housed in controlled conditions with a 12-hour light and 12-hour dark (LD) cycle. To ensure their well-being, routine water changes were performed, and their diet primarily consisted of Artemia salina. The zebra fish were kept for breeding to obtain the embryos. The experiments were performed. Fertilized embryos were placed into individual wells within 6-well plates, each containing 3 mL of solution. Various exposure groups were then introduced to these embryos. In each experiment involving zebrafish embryos, a sample size of n = 50 per group was employed, and the experiment was replicated three times. The investigations were conducted within four different treatment groups, untreated groups serve as a control group, and Ag-Se/Fu with various concentration ranges from the 75, 150, as well as 300 mg/mL. The untreated embryos were used as control. The different groups were exposed to the zebra fish embryos for 1 day. After the exposure different toxicity parameters such as malformation and survival rate were investigated.

Toxicity analysis in zebra fish embryos

The zebra fish embryos exposed to the toxins develop a malformation such yolk sac oedema and pericardial oedema. Using this condition, the toxic level of drug can be calculated. The 48 hours post fertilized (hpf) zebra fish embryos were used for measuring the survival rate of the embryos. Based on the survival of embryos after the exposure the results are expressed in percentage.

Statistical analysis

The data was reported as mean ± SD. All in vitro studies consisted of three independent experiments, each with three replicate measurements. The statistical differences between untreated and treated cells were analysed using IBM SPSS Statistics 27. The multiple comparison test was performed using one-way analysis of variance (ANOVA) and the P-values < 0.05 were considered statistically significant.

Results and discussion

The synthesis of Ag-SeO2 nanoparticles derived from an aqueous extract constitutes a significant contribution to the field of nanomaterials. The utilisation of natural extracts in nanoparticle synthesis is gaining momentum due to their eco-friendly and sustainable nature.

Fabrication of Ag-SeO₂ nanoparticles from aqueous extract

In this study, we employed an aqueous extract as a reducing and stabilizing agent for the fabrication of silver selenite (Ag-SeO₂) nanoparticles, exploring the potential of this approach for green synthesis. The natural reducing agents present in the extract played a crucial role in the reduction of silver ions and the formation of selenium dioxide, leading to the production of well-defined nanoparticles as depicted in Figure 1.

Figure 1. Synthesis of Ag-SeO₂ nanoparticles.

Synthesized of fucoidan doped Ag-SeO₂ NPs

The synthesis of fucoidan doped Ag-SeO2 nanoparticles was carried out successfully through a controlled process. During the doping process, the addition of fucoidan to the Ag-SeO2 nanoparticle solution, we observe the colour changes from the initial pale yellow to a deep brown, indicating the reduction of silver ions and the formation of fucoidan doped Ag-SeO2 nanoparticles, as shown in Figure 2.

Figure 2. Synthesis process of Ag-SeO₂ doped fucoidan.

Characterization of Ag-Se/Fu nanocomposites

UV-Spectrophotometer

Ag-Se/Fu nanoparticles that were synthesized displayed peaks at 320, 360 and 410 nm. These findings allowed us to confirm that silver (Ag) peak at 410 nm, selenium (Se) peak at 320 nm, and fucoidan (Fu) peak at 360 nm are all present in the synthesized nanocomposites show in Figure 3. The reason for this spectral absorption is that when certain wavelengths of light excite electrons on the metal surface, they oscillate [21,22].

Figure 3. Ultraviolet – visible spectrum of the Ag-Se/Fu suspension.

Fourier transform infrared spectroscopy (FT-IR analysis)

FTIR analysis successfully identified sulphate functional groups of fucoidan in Ag-Se/Fu nanocomposites, as illustrated in Figure 4. The crucial information regarding the sulphate groups localization falls within the wavelength range of 4000-500 cm⁻¹. Previous research has established that the broad signal observed at 12,259 cm⁻¹ serves as an indicative measure of the overall sulphate esters present in polysaccharides [23]. In our doped fucoidan doped Ag-SeO₂ nanocomposite results, spectral peaks at 671.01 cm⁻¹ and 1020.87 cm⁻¹. The functional groups like C=O bending and C-N Stretching respectively. This finding further supports the successful incorporation of fucoidan into the Ag-SeO₂ nanocomposite, emphasizing its potential contribution to the nanocomposite properties.

Figure 4. FT-IR spectra of biologically synthesized Ag-Se/Fu nanocomposites.

X-Ray diffraction analyses (XRD)

The X-Ray Diffraction analyses are important for identification of sample purity, crystalline size [24]. Our synthetic Ag-Se/Fu nanocomposites were more crystalline (68.2%) and less amorphous (31.8%) components as a result of this investigation. so that the crystalline character of the Ag-Se/Fu nanocomposites we generated is exceedingly stable in Figure 5. Similar findings were made by previous study, which showed that the XRD analysis of the silver nanoparticles corroborated their crystalline character [25,26]. By using X-ray diffraction (XRD) analysis, the crystalline structure of as-formed Se-NPs was examined. The pattern showed eight absorption peaks with the following values: (100), (101), (110), (102), (111), (201), (112), and (202), which matched Bragg diffraction at 2θ values of 23.5°, 29.3°, 41.3°, 45.51°, 52.53°, 55.71°, and 62.74°, respectively.

Figure 5. Powder X-ray crystallography spectra of Aloe vera mediated Ag-se doped fucoidan heterostructures.

Scanning electron microscopy (SEM)

SEM images and particle size distribution were confirmed the surface morphology of the synthesis of Ag-Se/Fu nanocomposites at different magnifications at 1 & 0.5 μm (Figure 6(a,b)). The Ag-Se/Fu nanocomposites showed irregular shapes were agglomerated size ranges from 20–80 nm and the mean particle size of produced Ag-Se/Fu nanocomposites was found 48 ± 2 nm. Previous study evidence conducted by Nassar et. al., 2023 revealed that the biosynthesized SeNPs prevented a monodisperse spherical morphology with an approximate size of 100 nm [26]. Another study showed that the Silver nanoparticles using Ceratonia siliqua extract were Spherical in shape with particle size range from 5 to 40 nm [27].

Figure 6. Scanning electron microscope images and the particle size distribution of Ag-Se doped fucoidan nanocomposites.

Energy-dispersive X-ray spectroscopy (EDX)

EDX analysis was employed to ascertain the elemental composition of the Ag-Se/Fu nanocomposites. In Figure 7, the EDX spectra revealed the presence of signals from Ag, U, and Se in the Ag-Se nanocomposites, with approximate proportions of 88.2%, 10.9%, and 0.9%, respectively. This verified the bonding of silver (Ag) to selenium (Se) on the nanocomposite surface. This study compared with other study revealed that the presence of Ag and O elements in both phyto-synthesized Ag₂O via Mentha Pulegium and Ficus Carica extracts signals in the EDX spectrum [28].

Figure 7. EDX images of synthesized Ag-Se/Fu nanocomposites.

Antibacterial activity

Ag-Se/Fu nanocomposites from Aloe vera

The antimicrobial efficiency of Ag-Se/Fu nanocomposites and Ag-Se nanoparticles’ zones of inhibition were compiled in a Tables 1 and 2 and Figures 8 and 9. The ZOI values recorded by Ag-Se/Fu were 20 mm for Klebsiella pneumoniae, and 19 mm for MRSA and 20 mm for Escherichia coli and 18 mm for Enterococcus faecalis while Ag-Se recorded low ZOI values of 13 mm for E. coli as well as 18 mm for Methicillin resistant Staphylococcus aureus (MRSA) and 13 mm and 14 mm for Klebsiella pneumoniae and Enterococcus faecalis respectively. The successful synthesis of Fucoidan doped on the Ag-Se surface is indicated by a gradual rise in the zone of inhibition with increasing Ag-Se/Fu concentration. These findings show that a synergistic impact between Fucoidan and Ag-Se/Fu nanocomposites enhances their antibacterial activity. The largest zone of inhibition diameter was obtained against the Bacillus subtilis bacterial strain [29].

Figure 8. Antimicrobial activity of Ag-Se/Fu nanocomposites.

Figure 9. Antimicrobial activity of (Ag-Se NPs) silver- selenium nanoparticles.

Table 1. Zone of inhibition: Ag-Se/Fu nanocomposites.

S.no	Organism	Concentration/Mean Zone diameter in MM
		20 mg	40 mg	60 mg	80 mg	Positive control	Negative control
1	Enterococcus faecalis	14 mm	15 mm	17 mm	18 mm	27 mm	-
2	Escherichia coli	16 mm	18 mm	19 mm	20 mm	30 mm	-
3	Klebsiella pneumoniae	15 mm	17 mm	19 mm	20 mm	32 mm	-
4	MRSA	14 mm	15 mm	17 mm	19 mm	25 mm

Table 2. Zone of inhibition: Ag-Se nanocomposites.

S.no	Organism	Concentration/Mean Zone diameter in MM
		20 mg	40 mg	60 mg	80 mg	Positive Control	Negative Control
1	Enterococcus faecalis	11mm	13mm	14mm	14mm	15mm	-
2	Escherichia coli	10mm	12mm	12mm	13mm	25mm	-
3	Klebsiella pneumoniae	8mm	10mm	12mm	13mm	23mm	-
4	MRSA	14mm	14mm	16mm	18mm	21mm	-

Minimum inhibitory concentration (MIC)

The Ag-Se/Fu nanocomposites were shown to have a minimum bactericidal concentration of 31.75 µg/mL for E. coli, 15.625 µg/mL for Enterococcus faecalis, 124.9 µg/mL for K. pneumoniae, and 31.25 µg/mL for MRSA, as depicted in Table 3. The results showed that the Ag-Se/Fu nanocomposites had a great antibacterial property against the majority of bacteria. Furthermore, it’s worth noting that streptomycin, a broad-spectrum antibiotic, demonstrated its highest effectiveness against Enterococcus faecalis, Klebsiella pneumoniae, MRSA, and Escherichia coli, exhibiting inhibitory effects.

Table 3. Minimum inhibitory concentration of Ag-Se/Fu nanocomposites.

Organism	MIC
Enterococcus faecalis	15.625µg/ml
Methicillin- resistant Staphylococcus aureus	31.25 µg/ml
Klebsiella pneumoniae	124.9 µg/ml
Escherichia coli	31.75 µg/ml

Antioxidant assay

A DPPH radical scavenging assay was used to determine the Ag-Se/Fu nanocomposites substantial antioxidant capabilities. The findings demonstrated that several Ag-Se/Fu nanocomposite concentrations (100, 200, 300, 400, and 500 μg/ml) were used to calculate free radical scanning ability, as depicted in Figure 10. Similarly, a previous study conducted by Abdelghany et al. (2023) on the green fabrication of nanocomposite-doped selenium nanoparticles has shown antioxidant properties [30]. The findings strongly suggest using Ag-Se/Fu nanocomposites as natural antioxidants to protect against various forms of oxidative stress linked to developing illnesses.

Figure 10. Antioxidant activity of synthesized Ag-Se/Fu nanocomposites.

Malformation and survival rate

Zebrafish embryos are widely employed as a valuable model organism in toxicity research due to their transparency, rapid development, and physiological similarity to humans. To introduce the medication under investigation, it is administered by incorporating it into the environment surrounding the embryos [31]. This can be achieved by introducing the medication into the water in which the embryos are housed. The embryos are closely observed for any detectable changes or effects induced by the substance. These changes could encompass variations in survival rates, behaviour, or developmental sequences. The information gathered is then analysed to gauge the substance’s toxicity. This information can be utilized to assess potential risks associated with the medication and inform decisions regarding its use [32]. In comparison to the control group, zebrafish embryos exposed to varying concentrations of Aloe Ag-Se/Fu (75 mg/mL and 150 mg/mL) exhibited regular morphological features in Figure 11. At the concentration of 75 mg/mL, Aloe Ag-Se/Fu exhibited a survival rate of 98%, and at 150 mg/mL, it showed a survival rate of 93%, which were similar to one another. However, when exposed to a concentration of 300 mg/mL, there was a slightly notable decrease in the survival rate (85%) in comparison to the control group.

Figure 11. (a) abnormalities in morphology and (b) survival rates of zebrafish blastula following exposure to various groups, including clove Ag-Se/Fu, were assessed. The experiments were conducted three times, and asterisks (*) indicate statistically significant differences at a significance level of p < 0.05.

Conclusion

In Taken Together, aqueous leaf extract from Aloe vera proved effective in successfully synthesising Ag-Se/Fu nanocomposites, demonstrating potent antimicrobial properties against Klebseilla pneumoniae and Methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, and Enterococcus faecalis bacteria. Furthermore, the synthesised Ag-Se/Fu nanocomposites exhibit antioxidant capabilities and demonstrated cytotoxicity, with a notable 85% survival rate in zebrafish embryos at a concentration of 300 mg/mL. These findings underscore the multifaceted benefits of the Ag-Se/Fu nanocomposites, pointing towards their potential applications in various biomedical and antimicrobial avenues.

Author contribution

Conception and design: M.S & T.A; analysis and interpretation of the data: M.S, S.B.V, P.G.S & B.R; the drafting of the paper: M.S, S.S, & M.A.A, revising it critically for intellectual content; M.S, T.A, S.S & B.R. All author are approve the final version manuscript.

Acknowledgments

The authors express their sincere appreciation to the researchers supporting project number (RSP2024R228) King Saud University, Riyadh, Saudi Arabia. Dr. Saravanan Muthupandian acknowledges to ICMR, Govt. of India for the Financial support IIRP-2023-8109.

Disclosure statement

No potential conflict of interest was reported by the author(s).