https://orcid.org/0000-0001-5266-7796
Abstract
The rise of antibiotic resistance among pathogenic bacteria poses a critical challenge to global healthcare, necessitating innovative therapeutic strategies. This study addresses this gap by developing ciprofloxacin-loaded silver nanoparticles (CIP@Ag NPs), combining robust antibacterial efficacy with additional therapeutic benefits. CIP@Ag NPs were synthesised and characterised through UV–visible spectroscopy, revealing a distinct redshift to 401 nm and a reduced band gap of 2.24 eV, enhancing their photocatalytic and bioactive properties. Structural and morphological integrity was confirmed using X-ray diffraction and scanning electron microscopy analyses. The nanoparticles exhibited remarkable multifunctionality, with 84% 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity and a fourfold increase in total antioxidant capacity compared to bare Ag NPs. Anti-inflammatory activity was evidenced by 86.43% inhibition of egg albumin denaturation at 800 μg/ml. Additionally, CIP@Ag NPs demonstrated potent antidiabetic effects, achieving 73% α-amylase and 68% α-glucosidase inhibition at 60 μg/ml. Antibacterial assays of CIP@Ag NPs showed significant improvement, with inhibition zones of up to 34 mm against multidrug-resistant strains like Staphylococcus aureus and Escherichia coli, surpassing the efficacy of both Ag NPs and ciprofloxacin individually. These findings underscore the potential of CIP@Ag NPs as a multifunctional nanoplatform, offering a promising solution to combat antibiotic resistance while addressing inflammation, oxidative stress, and diabetes-related complications.
Introduction
Antibiotic resistance has emerged as one of the most pressing global health threats, with the World Health Organization (WHO) identifying it as a major challenge for contemporary medicine (Chinemerem Nwobodo et al., 2022). The rapid emergence of multidrug-resistant (MDR) bacteria, driven by the overuse and misuse of antibiotics, is rendering many conventional treatments ineffective, leading to prolonged illness, increased mortality, and significant economic burdens on healthcare systems (Catalano et al., 2022). Pathogens such as Pseudomonas aeruginosa, and Escherichia coli have evolved resistance mechanisms that render traditional antibiotics less effective, leading to severe complications in clinical settings (Qin et al., 2022). With the growing challenge of bacterial resistance outpacing the effectiveness of conventional antibiotics, there is an increasing imperative to develop novel approaches to effectively address these persistent infections (Arif et al., 2022; Hussein-Al-Ali et al., 2022; Ibraheem et al., 2022). In this context, nanotechnology has emerged as a powerful tool, offering new avenues for enhancing drug efficacy and overcoming resistance mechanisms. Ciprofloxacin, a widely used fluoroquinolone antibiotic, stands as a critical agent in the fight against bacterial infections; however, its effectiveness has been increasingly compromised due to the rise of bacterial resistance (Hussein-Al-Ali et al., 2022; Mohsen et al., 2020).
Silver nanoparticles (Ag NPs) have emerged as exceptional agents in biomedical research, celebrated for their remarkable antimicrobial properties (Husain et al., 2023; Laib et al., 2024b). These nanoscale agents compromise bacterial cell membranes, disrupt essential cellular functions, and trigger oxidative stress, ultimately causing bacterial cell death (Daoudi et al., 2024). Ag NPs demonstrate broad-spectrum antibacterial activity, effectively targeting both Gram-positive and Gram-negative bacteria, including strains that exhibit resistance (Haji et al., 2022). Beyond their potent antimicrobial capabilities, they possess antioxidant and anti-inflammatory effects, which further expand their therapeutic potential (Alzubaidi et al., 2023; Laib et al., 2024a). However, caution is warranted; higher concentrations of Ag NPs can produce reactive oxygen species (ROS) that may harm healthy cells (Doudi et al., 2024). Their band gap energy is crucial—lower band gaps boost antimicrobial action but increase oxidative damage risk, underscoring the need for optimization (Kumar et al., 2024). In the ongoing battle against bacterial infections, ciprofloxacin, a second-generation fluoroquinolone antibiotic, inhibits key enzymes essential for bacterial DNA replication (Ajaykumar et al., 2023; Laib et al., 2023). Yet, rising bacterial resistance mechanisms threaten its effectiveness (Daoudi et al., 2022). By combining ciprofloxacin with silver nanoparticles, researchers are unlocking a promising strategy that enhances the antibiotic's potency while mitigating resistance, paving the way for a more effective future in therapeutics (Jaber & Saadh, 2024).
Ciprofloxacin loading onto Ag NPs can be achieved through mechanisms such as electrostatic interactions, hydrogen bonding, and physical adsorption. Ag NPs provide a large surface area for drug binding, enabling efficient drug loading and controlled release (Jaber & Saadh, 2024). This nanoparticle–drug conjugate not only improves the stability of ciprofloxacin but also supports its gradual release at the infection site, maintaining extended antimicrobial effectiveness (He et al., 2021). The cooperative interaction between ciprofloxacin and Ag NPs enhances the bactericidal effect, making this combination particularly potent against resistant bacterial strains (Jaber & Saadh, 2024). In addition to their antimicrobial effects, silver nanoparticles hold promise across diverse biomedical applications, such as wound healing, cancer treatment, and biosensing. Due to their nanoscale size and capacity to cross biological barriers, Ag NPs can selectively target specific tissues and organs, making them well-suited for drug delivery systems. Their anti-inflammatory effects, which block pro-inflammatory cytokines and inhibit protein denaturation, also make them valuable for treating inflammatory diseases (Shen et al., 2023). Moreover, the antioxidant properties of Ag NPs neutralise ROSs, helping to reduce oxidative stress and protect cells from damage in various pathological states (Predoi et al., 2022).
This study synthesises ciprofloxacin-loaded silver nanoparticles (CIP@Ag NPs) and assesses their antimicrobial, antioxidant, anti-inflammatory, and antidiabetic properties. CIP@Ag NPs are tested against MDR strains, including S. aureus, B. subtilis, P. aeruginosa, Klebsiella pneumoniae, and E. coli. Antioxidant and anti-inflammatory activities are evaluated through radical scavenging and albumin denaturation assays, while antidiabetic potential is examined via α-glucosidase and α-amylase inhibition. This research aims to develop nano-antibiotics that combat resistance and expand Ag NPs biomedical applications.
Materials and methods
Materials
The research used analytical grade chemicals and reagents, such as silver nitrate (AgNO3, 99.9%), ascorbic acid (C6H8O6, ≥99%), ciprofloxacin (C17H18FN3O3, 99.9%), which were utilised in the experiment, along with other chemicals like potassium dihydrogen phosphate (KH2PO4, 99.5%), dibasic potassium phosphate (K2HPO4, 99.95%), 2,2-diphenyl-1-picrylhydrazyl (C18H₁2N5O6,≥ 95%), ammonium molybdate ((NH4)6Mo7O24•4H2O,99.98%), sodium phosphate (NaH2PO4, ≥ 99%), potassium ferricyanide (K3[Fe(CN)6], ≥ 99%), trichloroacetic acid (CCl3COOH, ≥ 99%), and ferric chloride (FeCl3, ≥ 98%). All these materials were sourced from Sigma-Aldrich (St. Louis, MO, USA).
The bacterial strains used in this study, including P. aeruginosa (ATCC 27853), S. aureus (ATCC 25923), B. subtilis (ATCC 25973), K. pneumoniae (ATCC 13885), and E. coli (ATCC 25922), were acquired from the Pasteur Institute laboratory in Algiers, Algeria.
Preparation of Ag NPs
Ag NPs were synthesised using ascorbic acid as a reducing agent under controlled thermal conditions. A 0.01 M AgNO3 solution in 100 ml deionised water (DW) was prepared with continuous stirring, while a separate 0.02 M ascorbic acid solution in 100 ml DW was prepared. The AgNO3 solution was heated to 60 °C, and 10 ml of ascorbic acid solution was added dropwise over 5 min to control reduction rate and nanoparticle size. The mixture continued stirring at 60 °C for 2 hr, turning from colourless to deep yellow-brown, confirming Ag+ reduction and successful Ag NP formation. The nanoparticles were then centrifuged at 10,000 rpm, washed with DW, and dried at 100 °C for 2 hr. Finally, the dried powder was calcined at 500 °C for 4 hr to form Ag NPs powder (Suriati et al., 2014).
Preparation of drug-coated Ag nanoparticles (CIP@Ag NPs)
To prepare ciprofloxacin-loaded silver nanoparticles (CIP@Ag NPs), a validated protocol was followed to ensure synthesis efficiency and stability (Mohammed et al., 2023). CIP was dissolved in deionised water (0.5 mg/ml) and mixed with a 5 mg/ml dispersion of Ag NPs in distilled water. Prior to drug loading, the Ag NPs dispersion was subjected to ultrasonication using an ultrasonic bath sonicator (40 kHz) for 30 min at 42 °C. This step aimed to enhance the dispersion of Ag NPs by breaking down any agglomerates and ensuring uniform particle distribution in the solution. The Ag NPs solution was then added to the ciprofloxacin solution, with continuous stirring for 30 min at room temperature to facilitate drug adsorption. The final CIP@Ag NPs solution was stored at 4 °C to maintain stability until further experimentation.
Characterisation Ag NPs and CIP@Ag NPs surfaces
Advanced characterisation was performed to comprehensively investigate the properties of synthesised Ag NPs and ciprofloxacin-loaded Ag NPs (CIP@Ag NPs). The samples used for all analyses included silver nanoparticle powder roasted at 500 °C for 4 hr and CIP@Ag NPs. Fourier transform infrared spectroscopy (FTIR) analysis (Agilent Cary 630) was employed to identify functional groups and explore the interaction between ciprofloxacin and the nanoparticles, revealing chemical and structural properties (Allag et al., 2024). XRD, operated with CuKα radiation at 30 kV and 20 mA (λ = 0.154281 Å) with a scan speed of 0.05°, determined the crystallinity, phase purity, lattice parameters, and crystalline phases of the nanoparticles. Scanning electron microscopy (SEM: Thermo Scientific, Quatro, Thermo Fisher Scientific, Germany) provided detailed images of surface morphology and size distribution, further elucidating the structural characteristics of the nanoparticles. Additionally, UV–Vis spectroscopy (Shimadzu UV-2450, USA) was used to study the optical properties of Ag NPs and CIP@Ag NPs within the wavelength range of 200–800 nm (Meneceur et al., 2023; Zohra et al., 2023).
Antioxidant activity
DPPH free radical scavenging activity assay
To evaluate and compare the antioxidant potential of Ag NPs and CIP@Ag NPs, a DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging assay was conducted (Fouhma et al., 2024). Ag NPs and CIP@Ag NPs were prepared at six distinct concentrations (50, 100, 200, 250, 500, and 700 μg/ml). Each nanoparticle solution was then combined with a methanolic DPPH solution at a concentration of 4% (vol/vol). Following preparation, the mixtures were incubated at room temperature in darkness for 30 min, allowing sufficient interaction between the nanoparticles and DPPH radicals (Amor et al., 2023 ; Laib & Djahra, 2022). The reduction in DPPH radical concentration, indicative of antioxidant activity, was measured by recording absorbance at 517 nm using a UV–Vis spectrophotometer. The DPPH radical scavenging activity percentage was calculated using an established formula:
where (A control) represents the absorbance of the DPPH solution without nanoparticles and (A sample) denotes the absorbance of the solution containing Ag NPs or CIP@Ag NPs.
Reducing power assay
The antioxidant properties of Ag NPs and CIP@Ag NPs were evaluated through the ferric-reducing power assay. Nanoparticle solutions of varying concentrations (100, 200, 500, 700, and 1,000 μg/ml) were prepared, and 1 ml of each solution was mixed with 1 ml of 1% potassium ferricyanide and 1 ml of phosphate buffer (0.2 M, pH 6.6). This mixture was incubated at 50 °C for 20 min to facilitate the reduction process. After incubation, 0.5 ml of 10% trichloroacetic acid (wt/vol) was added, and the mixture was centrifuged at 3,000 rpm for 10 min to eliminate insoluble components. A 1 ml portion of the resulting supernatant was then mixed with 1 ml of deionised water and 0.125 ml of 0.1% ferric chloride solution. Finally, the absorbance of this solution was measured at 700 nm to quantify antioxidant activity (Laib & Djahra, 2023).
Assessment of total antioxidant capacity
To assess the total antioxidant capacity (TAC), a range of concentrations of Ag NPs and CIP-Ag NPs was prepared. Each nanoparticle solution was mixed with 1 ml of a reaction solution containing 0.6 M sulphuric acid, 0.1 M sodium phosphate buffer, and 0.28 M ammonium molybdate. This reaction mixture was then incubated at 95 °C for 90 min to promote the formation of a phosphomolybdenum complex, an indicator of antioxidant activity. The samples, after incubation, were cooled to room temperature. The absorbance at a wavelength of 695 nm was recorded using a UV–visible spectrophotometer. Then, TAC was determined and expressed as the equivalent ascorbic acid amount. This was done by making a comparison of the absorbance values obtained with that of a blank (nanoparticles free) and an ascorbic acid solution standard (Lu et al., 2011).
Evaluation of anti-inflammatory activity
The anti-inflammatory properties of Ag NPs and CIP@Ag NPs were assessed through an egg albumin denaturation inhibition assay. This assay involved the preparation of a mixture comprising 200 μl of fresh egg albumin, 2.8 ml of phosphate buffer (pH 6.4), and 2 ml of each nanoparticle solution at various concentrations (100, 300, 500, 700, and 800 μg/ml). For comparative purposes, 2 ml of diclofenac sodium, a well-established anti-inflammatory agent, was utilised as a positive control, while distilled water served as a negative control. The samples were incubated at 37 °C for 15 min and subsequently heated to 70 °C for 5 min to facilitate protein denaturation. Following the cooling period, the absorbance of each sample was measured at 660 nm using a UV–Vis spectrophotometer (Tlili et al., 2024). The percentage inhibition of albumin denaturation was calculated using the appropriate formula:
where A control refers to the absorbance measured for the control solution, which consists of distilled water, while A sample denotes the absorbance observed in the presence of Ag NPs or CIP@Ag NPs.
Antidiabetic activities
α-Amylase inhibitory activity for Ag NPs and CIP@Ag NPs
Inhibitory activity of α-amylase by AgNPs and CIP@Ag NPs was carried out through DNS method. Solutions of AgNPs and CIP@Ag NPs were prepared in phosphate buffer at various concentrations (5, 10, 20, 30, 40, 50, and 60 μg/ml) from a 1 mg/ml stock solution. Each sample (500 μl) was mixed with 500 μl of α-amylase enzyme solution (2 units/ml) and incubated at room temperature for 10 min. Subsequently, 500 μl of a 1% starch solution was added, followed by another 10-min incubation. The reaction was terminated by adding DNS reagent, bringing the mixture to a final volume of 500 μl. Following thorough mixing, the samples were subjected to a heating process in a boiling water bath for 15 min. Subsequently, they were allowed to cool and were diluted with 5 ml of distilled water. The absorbance of each sample was then measured at a wavelength of 540 nm using a spectrophotometer. Acarbose, an established antidiabetic agent, served as the positive control. Blank samples, prepared by substituting enzyme with buffer, were included for each concentration, and the control (without nanoparticles) was considered as the baseline (100% enzyme activity). The percentage of inhibition was calculated to quantify the inhibitory potential of AgNPs and CIP@Ag NPs on α-amylase activity.
Evaluation of α-glucosidase inhibition by Ag NPs and CIP@Ag NPs
The inhibitory activity of α-glucosidase by Ag NPs and CIP@Ag NPs was evaluated through the measurement of 4-nitrophenol release from p-nitrophenyl α-D-glucopyranoside. Various concentrations of Ag NPs and CIP@Ag NPs were prepared, specifically at 10, 20, 40, 60, 80, 100, and 120 μg/ml. In the assay, 200 μl of each sample was mixed with a reaction mixture consisting of 300 μl of 10 mM p-nitrophenyl α-D-glucopyranoside, 1,000 μl of 0.1 M potassium phosphate buffer (pH 6.8), and 200 μl of α-glucosidase enzyme solution. The reaction mixtures were incubated at 37 °C for 30 min. Following this incubation period, the reaction was terminated by the addition of 2,000 μl of 100 mM sodium carbonate. The concentration of liberated 4-nitrophenol was determined by measuring the absorbance at 400 nm using a spectrophotometer. Acarbose was utilised as a positive control for comparison, and the percentage of inhibition was calculated to quantify the α-glucosidase inhibitory activity of both Ag NPs and CIP@Ag NPs (Yousefzadeh-Valendeh et al., 2023).
Antibacterial activity of Ag NPs and CIP@Ag NPs
The antibacterial efficacy of Ag NPs and CIP@Ag NPs was assessed using the agar diffusion method. This evaluation included several pathogenic bacterial strains, such as S. aureus, P. aeruginosa, B. subtilis, K. pneumoniae, and E. coli. Solutions of the nanoparticles at varying concentrations were prepared, and 10 μl of each solution was applied to sterile filter paper discs (Aouadi et al., 2023; Chihi et al., 2023). These discs were subsequently placed on nutrient agar plates inoculated with the aforementioned bacterial strains, while ciprofloxacin (10 μg/disc) served as a positive control. The plates were incubated at 37 °C for 24 hr, after which the zones of inhibition surrounding the discs were measured to assess antibacterial activity. To ensure accuracy and minimise error, three replicates of each test were conducted for each bacterial strain analysed (Geyesa et al., 2024).
Statistical analysis
All experiments were performed in triplicate to ensure the reliability and reproducibility of the results. Differences among the experimental groups were assessed using analysis of variance, with statistical si gnificance determined at a threshold of p < .05.
Results and discussion
UV–visible analysis
The UV–visible analysis results reveal distinct absorption peaks for the synthesised Ag NPs at 395 nm, CIP at 329 nm, and the CIP@Ag NPs at 401 nm (Figure 1A). The redshift observed in CIP@Ag NPs, from 395 to 401 nm, indicates successful functionalization of Ag NPs with CIP, likely due to electronic interactions between the nanoparticles and the drug. This shift suggests a change in the local electronic structure, which can enhance the properties of the nanoparticles for drug delivery or photocatalytic applications. The interaction may have increased the conjugation length of the system, allowing for improved electron transfer and stability.
(A) UV–visible absorption spectra of Ag NPs, CIP, and CIP@Ag NPs. Tauc plots illustrating the relationship between (αhv)2 and photon energy (hv) for (B) Ag nanoparticles and (C) CIP@Ag NPs.
The Tauc plots derived from the UV–Vis spectra further highlight the changes in optical properties between Ag NPs and CIP@Ag NPs. The band gap energy of Ag NPs is 2.51 eV (Aktafa et al., 2024), while CIP@Ag NPs exhibit a reduced band gap of 2.24 eV (Figure 1B, C). This reduction in band gap energy upon the incorporation of CIP suggests enhanced photocatalytic potential (Zeghdi et al., 2024), as a lower energy gap facilitates more efficient light absorption and excitation of electrons. This change indicates that the CIP@Ag NPs could perform better under visible light conditions, making them suitable for applications such as environmental remediation and antibacterial treatments.
XRD analysis
The XRD analysis of the synthesised Ag NPs reveals distinct diffraction peaks at 2θ values of 38.1°, 44.6°, 64.7°, and 77.5°, corresponding to the (111), (200), (220), and (331) crystallographic planes, respectively (Figure 2). These values are characteristic of the face-centred cubic (FCC) crystal structure of silver, as confirmed by JCPDS Card No. 00-003-0921. The consistent peak positions indicate that the silver nanoparticles were successfully synthesised with high crystallinity. When ciprofloxacin was incorporated to form CIP@Ag NPs, the same diffraction peaks were observed, suggesting that the crystalline structure of the Ag NPs remained unaffected by the drug coating. However, a slight reduction in the intensity of these peaks was noted, which can be attributed to the surface adsorption of ciprofloxacin on the Ag NPs, potentially leading to slight changes in crystallinity or particle size. The crystallite sizes, calculated using the Scherrer equation (Mohammed Mohammed et al., 2023), showed that the Ag NPs had an average size of 21 nm, while the CIP@Ag NPs exhibited a slightly larger size of 24 nm. This increase in crystallite size may result from the added organic layer of ciprofloxacin, which could influence nanoparticle aggregation or growth during the coating process (Figure 2).
XRD patterns of synthesised Ag NPs and CIP@Ag NPs.
FTIR analysis
In the FTIR analysis of silver nanoparticles (Ag NPs) and ciprofloxacin-loaded silver nanoparticles (CIP@Ag NPs), several characteristic absorption peaks were observed, each corresponding to distinct functional groups and interactions (Figure 3).
FTIR of Ag NPs and CIP@Ag NPs.
For Ag NPs, the peak at 3,861 cm−1 corresponds to O–H stretching vibrations (Yang et al., 2021), indicative of hydroxyl groups likely associated with adsorbed water molecules. The peak at 3,723 cm−1 is attributed to N–H stretching, possibly due to amine groups or nitrogen-containing compounds (Dindarloo Inaloo et al., 2020). At 2,650 cm−1, weak C–H stretching vibrations suggest the presence of aliphatic hydrocarbon residues (Dun et al., 2013). The peak at 2,092 cm−1 corresponds to C☰C or C☰N stretching, indicating the presence of triple bonds or nitrile groups. The absorption band at 1,716 cm−1 is due to C=O stretching, characteristic of carbonyl groups from organic compounds stabilising the nanoparticles (Herrero et al., 2011). At 1,520 cm−1, the peak is associated with N–O symmetric stretching, likely from nitrate groups (Mihaylov et al., 2021). The peak at 484 cm−1 is attributed to Ag–O vibrations, confirming the presence of silver oxide bonds (Ande & Prasad, 2023).
For CIP@Ag NPs, the spectrum exhibits additional and shifted peaks, indicating the successful conjugation of ciprofloxacin onto the silver nanoparticles. The peak at 3,766 cm−1 corresponds to O–H stretching vibrations, suggesting the presence of hydroxyl groups from water or ciprofloxacin (Mohammed et al., 2024). At 3,347 cm−1, N–H stretching vibrations characteristic of amine groups in ciprofloxacin are observed (He et al., 2022). The peak at 2,913 cm−1 represents C–H stretching, indicating aliphatic hydrocarbon chains (Igisu et al., 2018). Weak C–H stretching vibrations are also observed at 2,646 cm−1 (Pitsevich et al., 2015). The peak at 2,093 cm−1 is attributed to C☰C or C☰N stretching. A significant peak at 1,685 cm−1 is assigned to C=O stretching vibrations (Biniak et al., 2016), characteristic of carbonyl groups in ciprofloxacin, with a slight shift from the Ag NPs spectrum, suggesting interactions with the nanoparticle surface. The peak at 1,431 cm−1 corresponds to C–N stretching vibrations, while the peak at 1,232 cm−1 represents C–O stretching, indicating bonding interactions between ciprofloxacin and the nanoparticles (Alia et al., 2024; Romeu et al., 2022). Additional peaks at 1,006 and 721 cm−1 correspond to C–H bending and out-of-plane bending vibrations, respectively, indicating the presence of aromatic or aliphatic groups (Sun et al., 2010).
The shifts and additional peaks in the FTIR spectrum of CIP@Ag NPs confirm the successful functionalization of silver nanoparticles with ciprofloxacin. The observed interactions, such as hydrogen bonding and coordination between functional groups like C=O, C–N, and C–O with the nanoparticle surface, provide critical insights into the chemical environment and validate the effective loading of ciprofloxacin onto the nanoparticles.
SEM analysis
The SEM images, shown in Figure 4, offer significant insights into the morphological characteristics and surface structure of the synthesised nanoparticles. In the SEM image of the Ag NPs (Figure 4A), a predominantly spherical morphology with uniform size distribution is observed, suggesting controlled nucleation and growth during synthesis. The spherical shapes indicate efficient reduction by ascorbic acid, producing well-dispersed Ag nanoparticles. In contrast, the SEM image of CIP@Ag NPs (Figure 4B) reveals changes in surface texture, implying successful ciprofloxacin coating on the nanoparticle surfaces. This coating effect is evidenced by a rougher and slightly aggregated appearance compared to bare Ag NPs, indicating ciprofloxacin's interaction with the nanoparticles. The observed morphological changes validate the ciprofloxacin coating process, which could enhance the nanoparticles' functionality for potential antibacterial applications by providing a stable drug layer on the particle surface.
(A) SEM image of Ag NPs; (B) CIP@Ag NPs.
Evaluation of antioxidant activity
Evaluation of DPPH radical scavenging activity
The DPPH scavenging assay results highlight the remarkable antioxidant activity of both Ag NPs and CIP@Ag NPs (Figure 5). Notably, CIP@Ag NPs exhibits superior scavenging efficiency across all concentrations, with a significant enhancement observed at higher doses. At 50 μg/ml, CIP@Ag NPs show 25.11% inhibition compared to 22% for Ag NPs, and this difference becomes more pronounced at 500 μg/ml, where CIP@Ag NPs reaches 74.22%, nearly double that of Ag NPs (42.44%). At 700 μg/ml, the inhibition percentage of Ag NPs-CIP soars to 84%, far surpassing Ag NPs' 45.77%. This significant improvement is attributed to the synergistic action between ciprofloxacin and the silver nanoparticles, where the antibiotic enhances the radical scavenging capacity of Ag NPs. Silver nanoparticles are known to generate ROSs, but in controlled environments, they also demonstrate antioxidant behaviour (Khorrami et al., 2019). The presence of ciprofloxacin further boosts this by facilitating electron transfer to neutralise DPPH radicals. The substantial antioxidant activity of CIP@Ag NPs at higher concentrations points to their potential to mitigate oxidative stress, inflammation, and tissue damage (Ibraheem et al., 2022). Compared to bare Ag NPs, CIP@Ag NPs exhibit superior free radical neutralisation, making them effective in reducing oxidative damage and inflammation often associated with bacterial infections (Hussan et al., 2024). This dual antioxidant and antimicrobial functionality highlights their potential as a powerful therapeutic option in oxidative stress-related conditions (Figure 5). Moreover, the findings suggest that CIP@Ag NPs could be explored as adjunctive therapies in antibiotic treatments. The ability of CIP@Ag NPs to neutralise free radicals while maintaining robust antimicrobial activity can be especially valuable in treating infections associated with biofilm formation (Mohsen et al., 2020). Biofilms notoriously resistant to conventional antibiotics often create oxidative microenvironments that contribute to chronic diseases. The dual action of CIP@Ag NPs in combating biofilm-associated bacteria and neutralising ROS could address these challenges effectively, as supported by recent studies (Hussein-Al-Ali et al., 2022).
Free DPPH radical scavenging activity of Ag NPs, CIP@Ag NPs, and ascorbic acid.
Assessment of reducing power
The reducing power assay results reveal the outstanding antioxidant potential of CIP@Ag NPs compared to Ag NPs alone, showcasing remarkable electron-donating capabilities (Figure 6). At 100 μg/ml, CIP@Ag NPs exhibits an absorbance of 0.13, significantly surpassing Ag NPs (0.09). This difference becomes even more pronounced at higher concentrations. At 500 μg/ml, CIP@Ag NPs displays an absorbance of 0.30, nearly double that of Ag NPs (0.15), and at 1,000 μg/ml, CIP@Ag NPs reaches a peak absorbance of 0.38, vastly outperforming AgNPs (0.22) (Figure 6). The exceptional reducing power of CIP@Ag NPs can be attributed to the synergistic interaction between ciprofloxacin and the silver nanoparticles, enhancing their electron-donating and radical-scavenging capabilities (Nikparast & Saliani, 2018). This synergism amplifies their antioxidant activity, positioning CIP@Ag NPs as a superior candidate for combating oxidative stress and inflammation, offering a dual function as a potent antimicrobial and antioxidant agent (Hussein-Al-Ali et al., 2022). This remarkable enhancement makes CIP@Ag NPs highly promising for therapeutic applications. Furthermore, the enhanced performance of CIP@Ag NPs suggests their applicability in advanced biomedical materials, such as antioxidant coatings, drug delivery systems, and bioactive scaffolds. The ability to scavenge radicals and donate electrons can be leveraged to improve the efficacy of existing treatments for inflammatory and oxidative stress-related disorders (Liu et al., 2022). For instance, their integration into polymer-based drug delivery matrices could enhance the localised therapeutic effects while minimising systemic toxicity. Future studies should explore the mechanistic pathways through which CIP@Ag NPs exert their reducing power in complex biological systems. In vivo models could provide deeper insights into their pharmacodynamics and pharmacokinetics, ensuring their safety and efficacy in clinical applications (Mohsen et al., 2020). These findings underscore the tremendous promise of CIP@Ag NPs as a transformative tool in nanomedicine, offering solutions to some of the most pressing challenges in healthcare today (Lin et al., 2016).
Total reducing power of Ag NPs, CIP@Ag NPs, and ascorbic acid at different concentrations.
Total antioxidant capacity
The TAC data demonstrate a striking enhancement in antioxidant potential with CIP@Ag NPs compared to Ag NPs (Figure 7). CIP@Ag NPs exhibits a TAC of 130.17 ± 2.02 μgAA/mg, representing a dramatic increase over AgNPs’ 42.92 ± 1.18 μgAA/mg (Figure 7). This impressive fourfold rise underscores the exceptional synergistic effect of ciprofloxacin and silver nanoparticles, significantly amplifying their collective antioxidant power (Figure 7). The superior TAC of CIP@Ag NPs highlights their enhanced ability to neutralise free radicals and mitigate oxidative stress, positioning them as a transformative advancement in nanomedicine (Zor et al., 2024). This extraordinary improvement not only showcases CIP@Ag NPs's potential in therapeutic applications but also sets a new standard for efficacy in combating oxidative damage, promising revolutionary impacts in clinical treatments and beyond. The synergistic effect observed in CIP@Ag NPs can be attributed to the unique properties of both ciprofloxacin and silver nanoparticles. Ciprofloxacin, a broad-spectrum antibiotic, is known for its ability to inhibit bacterial DNA replication, making it effective against a wide range of bacterial infections (Mohsen et al., 2020). When combined with silver nanoparticles, which possess inherent antimicrobial and antioxidant properties, the resulting CIP@Ag NPs exhibit enhanced therapeutic potential. The silver nanoparticles act as a delivery vehicle, ensuring sustained release and targeted delivery of ciprofloxacin, while also contributing to the overall antioxidant activity (Nawaz et al., 2021). Moreover, the enhanced antioxidant potential of CIP@Ag NPs suggests that they could be particularly beneficial in treating conditions characterised by high levels of oxidative stress, such as chronic inflammation, neurodegenerative diseases, and certain types of cancer. By effectively neutralising free radicals, CIP@Ag NPs can help protect cells from oxidative damage, thereby promoting tissue repair and regeneration (Din et al., 2024). This dual-action mechanism—combining antimicrobial and antioxidant properties—makes CIP@Ag NPs a versatile tool in the arsenal of nanomedicine. However, while the potential of CIP@Ag NPs is immense, further research is needed to fully understand their mechanisms of action, optimise their formulation, and ensure their safety and efficacy in clinical settings. Comprehensive in vivo studies and clinical trials will be crucial in translating these promising findings into practical applications. Additionally, exploring the long-term stability and potential side effects of CIP@Ag NPs will be essential for their successful integration into mainstream medicine (Begum et al., 2022).
Total antioxidant capacity (TAC, μg AA/mg NPs) of Ag NPs and CIP@Ag NPs.
Anti-inflammatory activity of Ag NPs and CIP@Ag NPs
The anti-inflammatory effectiveness of AgNPs and CIP@Ag NPs was evaluated via egg albumin denaturation inhibition, demonstrating a remarkable enhancement in CIP@Ag NPs (Figure 8). At 100 μg/ml, Ag NPs exhibited 54.13% inhibition, while CIP@Ag NPs showed 44.16%, suggesting a slight initial advantage of Ag NPs. However, the disparity becomes increasingly significant at higher concentrations. CIP@Ag NPs outperformed Ag NPs at 300 μg/ml with 57.87% inhibition compared to 51.60% and further demonstrated superior activity at 500 and 700 μg/ml with 62.63% and 62.52% inhibition versus 60% and 64.34% for Ag NPs, respectively (Figure 8). The most striking result was observed at 800 μg/ml, where CIP@Ag NPs achieved 86.43% inhibition, dramatically surpassing Ag NPs’ 73.21%. This substantial increase in inhibition underscores the exceptional anti-inflammatory potential of CIP@Ag NPs (Algarni et al., 2023; Zafar et al., 2022). The enhanced activity can be attributed to the synergistic effects of ciprofloxacin, which not only augments the anti-inflammatory properties of silver nanoparticles but also effectively prevents protein denaturation (Liu et al., 2022). The extraordinary dual action of CIP@Ag NPs lies in their ability to integrate silver nanoparticles' potent antibacterial properties with ciprofloxacin's targeted therapeutic benefits. Ciprofloxacin, a widely studied fluoroquinolone, disrupts bacterial DNA replication by inhibiting DNA gyrase and topoisomerase IV, while also enhancing the permeability of bacterial membranes, enabling deeper nanoparticle penetration (Liu et al., 2022). In parallel, silver nanoparticles attack bacterial cells through multi-pronged mechanisms, including the generation of ROSs, disruption of the cell membrane integrity, and inhibition of key metabolic enzymes. This powerful synergy not only amplifies antimicrobial efficacy but also equips CIP@Ag NPs to overcome challenges posed by MDR pathogens, representing a significant leap forward in nanomedicine (Hashemikia et al., 2021).
Comparative anti-inflammatory activity of Ag NPs and CIP@Ag NPs at various concentrations (100–800 μg/ml).
In addition to their unparalleled antimicrobial capabilities, CIP@Ag NPs demonstrate robust anti-inflammatory properties, further solidifying their transformative therapeutic potential. They effectively downregulate pro-inflammatory cytokines such as TNF-α, IL-6, and IL-1β, while neutralising excessive oxidative stress by scavenging free radicals, thus mitigating the underlying factors driving chronic inflammation (Zafar et al., 2022). Unlike traditional Ag NPs, which primarily offer antimicrobial activity, the ciprofloxacin coating of CIP@Ag NPs extends their therapeutic profile, enhancing their bioavailability, stability, and precision in targeting inflamed tissues. This makes them an exceptional therapeutic candidate for conditions where infection and inflammation are closely intertwined, such as chronic inflammatory disorders and infections associated with drug-resistant bacteria. This study not only highlights their immense potential but also aligns with the broader movement toward multifunctional nanocarriers, cementing CIP@Ag NPs as a paradigm shift in managing complex, inflammation-related disorders (Helmy et al., 2020). The remarkable inhibition achieved by CIP@Ag NPs makes them a compelling choice for advanced therapeutic applications, offering significant advantages in managing chronic inflammatory conditions and showcasing their superior efficacy over traditional Ag NPs.
In vitro antidiabetic activities
α-Amylase inhibition activity of of Ag NPs and CIP@Ag NPs
The graph illustrates the comparative α-amylase inhibition of Ag NPs and CIP@Ag NPs across varying concentrations, revealing a clear distinction in their antidiabetic potential (Figure 9). While both Ag NPs and CIP@Ag NPs exhibit a dose-dependent increase in enzyme inhibition, CIP@Ag NPs consistently outperform bare Ag NPs at every concentration, indicating enhanced bioactivity. Compared with acarbose, a standard antidiabetic drug, CIP@Ag NPs show competitive inhibition, suggesting that the conjugation of ciprofloxacin with Ag NPs significantly improves the nanoparticles' ability to inhibit α-amylase. The improved inhibition by CIP@Ag NPs may stem from the synergistic interaction between the antibiotic and the nanoparticles, which likely enhances their binding affinity to the enzyme or alters their surface chemistry, making them more effective (Debnath et al., 2020; Patel & Ishnava, 2024). On the other hand, bare Ag NPs, although exhibiting notable inhibitory effects, demonstrate comparatively weaker activity, likely due to their limited interaction with the enzyme without the additional pharmacological influence of ciprofloxacin (Seekonda & Rani, 2022). The superior performance of CIP@Ag NPs highlights the importance of functionalization in nanoparticle design, suggesting that drug-loaded nanoparticles hold greater potential for therapeutic applications in antidiabetic treatments, particularly in α-amylase inhibition, positioning CIP@Ag NPs as a more promising candidate over Ag NPs for mitigating postprandial hyperglycemia (Figure 9). Furthermore, the inclusion of ciprofloxacin in the nanoparticle formulation not only enhances the enzyme inhibitory activity but also provides an added antimicrobial effect, which may be beneficial in preventing or treating infections that are common in diabetic patients, who are prone to bacterial infections (Zafar et al., 2022). This dual therapeutic role of CIP@Ag NPs makes them a promising candidate for integrated management of diabetes and related infections. Additional studies on the pharmacokinetics, long-term safety, and potential side effects of CIP@Ag NPs are crucial to fully understand their therapeutic potential and optimise their clinical applications. Moreover, exploring the ability of CIP@Ag NPs to target specific tissues or cells could further improve their effectiveness, ensuring that they exert their effects in the appropriate biological compartments, minimising side effects, and maximising therapeutic benefits (Patel et al., 2020).
α-Amylase inhibition activity of Ag NPs and CIP@Ag NPs.
α-Glucosidase inhibitory activity Ag NPs and CIP@Ag NPs
The graph illustrates a remarkable comparison of α-glucosidase inhibitory activity between AgNPs, CIP@Ag NPs, and the benchmark antidiabetic drug, acarbose, across increasing concentrations (10–120 μg/ml). Both Ag NPs and CIP@Ag NPs exhibit a concentration-dependent inhibitory effect; however, CIP@Ag NPs consistently demonstrates superior activity across all concentrations, notably approaching the potent inhibitory profile of acarbose at higher doses (80–120 μg/ml) (Figure 10). This heightened activity of CIP@Ag NPs suggests a profound enhancement in bioactivity due to the conjugation with ciprofloxacin, which may modify the surface chemistry of the nanoparticles, leading to an increased affinity for the α-glucosidase enzyme (Yousefzadeh-Valendeh et al., 2023). The synergistic interplay between the antibiotic and the nanoparticles likely boosts their efficacy, offering a dual advantage: not only do they inhibit the enzyme more effectively than bare Ag NPs, but they also rival acarbose (Zeng et al., 2023). In contrast, bare Ag NPs, while displaying notable inhibitory effects, lag behind CIP@Ag NPs, highlighting the transformative impact of functionalization on nanoparticle efficacy.
α-Glucosidase inhibitory activity of Ag NPs and CIF@Ag NPs.
The superior performance of CIP@Ag NPs highlights their potential as an innovative and effective treatment for diabetes, offering an exciting alternative to conventional drugs like acarbose. The combination of ciprofloxacin with silver nanoparticles not only enhances the nanoparticles' ability to inhibit α-glucosidase but also improves the pharmacokinetics, ensuring more controlled and prolonged therapeutic effects, which could reduce the need for frequent dosing (Ibne Shoukani et al., 2024). Future studies should explore the effectiveness of CIP@Ag NPs in in vivo models to evaluate their impact on blood glucose regulation and their long-term safety. Additionally, surface modifications to target specific tissues or organs could enhance their precision, maximizing their benefits while minimising side effects.
Antibacterial activity of Ag NPs and CIP@Ag NPs
The findings indicate a notable difference in antibacterial effectiveness between Ag NPs and CIP@Ag NPs (Figures 11 and 12). Across different concentrations, Ag NPs alone showed moderate inhibition zones against S. aureus, P. aeruginosa, B. subtilis, K. pneumoniae, and E. coli, with values ranging from 8.5 to 14 mm (Table 1). This outcome suggests that Ag NPs disrupt bacterial cell membranes, produce ROSs, and lead to bacterial cell death (Albert et al., 2024; Mukherjee et al., 2023). In contrast, the CIP@Ag NPs demonstrated outstanding antibacterial efficacy, with inhibition zones reaching up to 34 mm for P. aeruginosa, B. subtilis, K. pneumoniae, and S. aureus at the highest concentration. This significant enhancement stems from the synergistic interaction between ciprofloxacin and Ag NPs (Ibraheem et al., 2022). Compared to ciprofloxacin alone (15.5–23 mm), CIP@Ag NPs outperform both the antibiotic and Ag NPs individually (Figures 13 and 14). The synergy of combining traditional antibiotics with nanotechnology represents a promising strategy for overcoming bacterial resistance, as CIP@Ag NPs exploit both ciprofloxacin’s ability to interfere with DNA replication and the ROS generation mechanism of Ag NPs (Tawre et al., 2022).
Comparison of the antibacterial effects of AgNPs and ciprofloxacin (CIP) alone on the growth of pathogenic bacteria at different concentrations.
The synergistic effects of CIP@Ag NPs on inhibiting the growth of pathogenic bacteria, compared to ciprofloxacin alone at 1,000 μg/ml.
Antibacterial activity of Ag NPs and CIF@Ag NPs.
| Sample | Zone of inhibition (nm) | |||||
|---|---|---|---|---|---|---|
| Concentartion (μg/ml) | S. aureus | P. aeruginosa | B. subtilis | Klebsiella pneumoniae | E. coli | |
| Ag NPs | 250 500 750 1,000 | 8.5 ± 0.70 9.5 ± 0.70 11.5 ± 0.70 14 ± 0.00 | 8.5 ± 0.70 9.5 ± 0.70 10.5 ± 0.70 11.5 ± 0.7 | 8.5 ± 0.7 10.5 ± 0.7 11.00 ± 0.00 11.5 ± 0.7 | 8.5 ± 0.7 9.5 ± 0.7 11 ± 1.41 11.5 ± 0.7 | 9.0 ± 0.7 10.0 ± 0.7 11.0 ± 0.7 12.0 ± 0.7 |
| Ag NPs-CIP | 5 (S1) | 31 ± 1.41 | 33.5 ± 0.7 | 34 ± 1.41 | 34 ± 1.41 | 30.0 ± 0.9 |
| Ciprofloxacin | 15.5 ± 0.7 | 19 ± 0.7 | 17.5 ± 0.7 | 23 ± 1.41 | 18 ± 0.7 | |
| DMSO | — | Nul | Nul | Nul | Nul | Nul |
| Sample | Zone of inhibition (nm) | |||||
|---|---|---|---|---|---|---|
| Concentartion (μg/ml) | S. aureus | P. aeruginosa | B. subtilis | Klebsiella pneumoniae | E. coli | |
| Ag NPs | 250 500 750 1,000 | 8.5 ± 0.70 9.5 ± 0.70 11.5 ± 0.70 14 ± 0.00 | 8.5 ± 0.70 9.5 ± 0.70 10.5 ± 0.70 11.5 ± 0.7 | 8.5 ± 0.7 10.5 ± 0.7 11.00 ± 0.00 11.5 ± 0.7 | 8.5 ± 0.7 9.5 ± 0.7 11 ± 1.41 11.5 ± 0.7 | 9.0 ± 0.7 10.0 ± 0.7 11.0 ± 0.7 12.0 ± 0.7 |
| Ag NPs-CIP | 5 (S1) | 31 ± 1.41 | 33.5 ± 0.7 | 34 ± 1.41 | 34 ± 1.41 | 30.0 ± 0.9 |
| Ciprofloxacin | 15.5 ± 0.7 | 19 ± 0.7 | 17.5 ± 0.7 | 23 ± 1.41 | 18 ± 0.7 | |
| DMSO | — | Nul | Nul | Nul | Nul | Nul |
Note. Sa = S. aureus ATCC 25923; Pa = P. aeruginosa ATCC 27853; Bs = Bacillus subtilis ATCC 25973, Kp = K. pneumoniae ATCC 13885, Ec = E. coli ATCC 25922, DMSO = dimethyl sulfoxide.
Antibacterial activity of Ag NPs and CIF@Ag NPs.
| Sample | Zone of inhibition (nm) | |||||
|---|---|---|---|---|---|---|
| Concentartion (μg/ml) | S. aureus | P. aeruginosa | B. subtilis | Klebsiella pneumoniae | E. coli | |
| Ag NPs | 250 500 750 1,000 | 8.5 ± 0.70 9.5 ± 0.70 11.5 ± 0.70 14 ± 0.00 | 8.5 ± 0.70 9.5 ± 0.70 10.5 ± 0.70 11.5 ± 0.7 | 8.5 ± 0.7 10.5 ± 0.7 11.00 ± 0.00 11.5 ± 0.7 | 8.5 ± 0.7 9.5 ± 0.7 11 ± 1.41 11.5 ± 0.7 | 9.0 ± 0.7 10.0 ± 0.7 11.0 ± 0.7 12.0 ± 0.7 |
| Ag NPs-CIP | 5 (S1) | 31 ± 1.41 | 33.5 ± 0.7 | 34 ± 1.41 | 34 ± 1.41 | 30.0 ± 0.9 |
| Ciprofloxacin | 15.5 ± 0.7 | 19 ± 0.7 | 17.5 ± 0.7 | 23 ± 1.41 | 18 ± 0.7 | |
| DMSO | — | Nul | Nul | Nul | Nul | Nul |
| Sample | Zone of inhibition (nm) | |||||
|---|---|---|---|---|---|---|
| Concentartion (μg/ml) | S. aureus | P. aeruginosa | B. subtilis | Klebsiella pneumoniae | E. coli | |
| Ag NPs | 250 500 750 1,000 | 8.5 ± 0.70 9.5 ± 0.70 11.5 ± 0.70 14 ± 0.00 | 8.5 ± 0.70 9.5 ± 0.70 10.5 ± 0.70 11.5 ± 0.7 | 8.5 ± 0.7 10.5 ± 0.7 11.00 ± 0.00 11.5 ± 0.7 | 8.5 ± 0.7 9.5 ± 0.7 11 ± 1.41 11.5 ± 0.7 | 9.0 ± 0.7 10.0 ± 0.7 11.0 ± 0.7 12.0 ± 0.7 |
| Ag NPs-CIP | 5 (S1) | 31 ± 1.41 | 33.5 ± 0.7 | 34 ± 1.41 | 34 ± 1.41 | 30.0 ± 0.9 |
| Ciprofloxacin | 15.5 ± 0.7 | 19 ± 0.7 | 17.5 ± 0.7 | 23 ± 1.41 | 18 ± 0.7 | |
| DMSO | — | Nul | Nul | Nul | Nul | Nul |
Note. Sa = S. aureus ATCC 25923; Pa = P. aeruginosa ATCC 27853; Bs = Bacillus subtilis ATCC 25973, Kp = K. pneumoniae ATCC 13885, Ec = E. coli ATCC 25922, DMSO = dimethyl sulfoxide.
Evaluation of antibacterial effects of Ag NPs compared to CIP alone against (A) gram-positive bacteria and (B) gram-negative bacteria. CIP = ciprofloxacin.
Synergistic effects of Ag NPs and CIP@Ag NPs compared to CIP alone on the growth inhibition of (A) gram-positive pathogenic bacteria and (B) gram-negative pathogenic bacteria. CIP = ciprofloxacin.
Although the precise antibacterial mechanisms of CIP@Ag NPs are still under active investigation, multiple hypotheses can be drawn based on their well-documented properties. Ciprofloxacin in the Ag NP matrix further enhances bacterial inhibition through its classic mechanism—interference with bacterial DNA replication by inhibiting DNA gyrase (Bhusal et al., 2024). This coupled with the ROS generation by Ag NPs, results in an amplified bactericidal effect, targeting bacterial structures and metabolic functions on multiple fronts. Ciprofloxacin’s presence also promotes greater nanoparticle penetration into bacterial cells, boosting the overall efficacy of the CIP@Ag NPs complex (Mohsen et al., 2020). In scenarios where bacteria exhibit resistance to either Ag NPs or ciprofloxacin, the CIP@Ag NPs complex overcomes this hurdle by utilising the dual mechanisms of ROS production and enhanced drug delivery. This synergistic action disrupts bacterial cell walls and inhibits cellular function, making CIP@Ag NPs more effective than either component alone (Abdolhosseini et al., 2019; Zeng et al., 2023). Thus, CIP@Ag nanoparticles demonstrate significant promise in tackling the escalating issue of antibiotic resistance, particularly in treating infections caused by S. aureus, E. coli, and P. aeruginosa. Table 2 introduces comparison of the results of antibacterial activity of present study with literature, indicating the efficiency of CIP@Ag NPs.
Comparison of the results of antibacterial activity of this study with literature.
| Sample | Method of synthesis | Bacteria tested | Zone of inhibition (nm) | References |
|---|---|---|---|---|
| Ag NPs | Chemical reduction | Staphylococcus aureus–Escherichia coli | 07 06 | Mohsen et al. (2020) |
| Ag NPs-CIP | Ultrasonication-assisted drug loading | S. aureus–E. coli | 09 09 | Mohsen et al. (2020) |
| CIF@Ag NPs | Ultrasonication-assisted drug loading | Acinetobacter baumannii Serratia marcescens S. aureus | 28 31 32 | Ibraheem et al. (2022) |
| Ag NPs-PEG-CIP | Ultrasonication-assisted drug loading | A. baumannii S. marcescens S. aureus | 35 36 37 | Ibraheem et al. (2022) |
| HAPNWs@AgNPs-CIP | Sunlight-assisted Agnp immobilisation | E. coli S. aureus | 16 14 | Xiong et al. (2017) |
| Ag NPs | Chemical reduction | S. aureus Pseudomonas aeruginosa Bacillus subtilis Klebsiella pneumoniae E. coli | 14 11.5 111.5 11.5 12 | Present study |
| CIF@Ag NPs | Ultrasonication-assisted drug loading | S. aureus P. aeruginosa B. subtilis K. pneumoniae E. coli | 31 33.5 34 34 30 | Present study |
| Sample | Method of synthesis | Bacteria tested | Zone of inhibition (nm) | References |
|---|---|---|---|---|
| Ag NPs | Chemical reduction | Staphylococcus aureus–Escherichia coli | 07 06 | Mohsen et al. (2020) |
| Ag NPs-CIP | Ultrasonication-assisted drug loading | S. aureus–E. coli | 09 09 | Mohsen et al. (2020) |
| CIF@Ag NPs | Ultrasonication-assisted drug loading | Acinetobacter baumannii Serratia marcescens S. aureus | 28 31 32 | Ibraheem et al. (2022) |
| Ag NPs-PEG-CIP | Ultrasonication-assisted drug loading | A. baumannii S. marcescens S. aureus | 35 36 37 | Ibraheem et al. (2022) |
| HAPNWs@AgNPs-CIP | Sunlight-assisted Agnp immobilisation | E. coli S. aureus | 16 14 | Xiong et al. (2017) |
| Ag NPs | Chemical reduction | S. aureus Pseudomonas aeruginosa Bacillus subtilis Klebsiella pneumoniae E. coli | 14 11.5 111.5 11.5 12 | Present study |
| CIF@Ag NPs | Ultrasonication-assisted drug loading | S. aureus P. aeruginosa B. subtilis K. pneumoniae E. coli | 31 33.5 34 34 30 | Present study |
Comparison of the results of antibacterial activity of this study with literature.
| Sample | Method of synthesis | Bacteria tested | Zone of inhibition (nm) | References |
|---|---|---|---|---|
| Ag NPs | Chemical reduction | Staphylococcus aureus–Escherichia coli | 07 06 | Mohsen et al. (2020) |
| Ag NPs-CIP | Ultrasonication-assisted drug loading | S. aureus–E. coli | 09 09 | Mohsen et al. (2020) |
| CIF@Ag NPs | Ultrasonication-assisted drug loading | Acinetobacter baumannii Serratia marcescens S. aureus | 28 31 32 | Ibraheem et al. (2022) |
| Ag NPs-PEG-CIP | Ultrasonication-assisted drug loading | A. baumannii S. marcescens S. aureus | 35 36 37 | Ibraheem et al. (2022) |
| HAPNWs@AgNPs-CIP | Sunlight-assisted Agnp immobilisation | E. coli S. aureus | 16 14 | Xiong et al. (2017) |
| Ag NPs | Chemical reduction | S. aureus Pseudomonas aeruginosa Bacillus subtilis Klebsiella pneumoniae E. coli | 14 11.5 111.5 11.5 12 | Present study |
| CIF@Ag NPs | Ultrasonication-assisted drug loading | S. aureus P. aeruginosa B. subtilis K. pneumoniae E. coli | 31 33.5 34 34 30 | Present study |
| Sample | Method of synthesis | Bacteria tested | Zone of inhibition (nm) | References |
|---|---|---|---|---|
| Ag NPs | Chemical reduction | Staphylococcus aureus–Escherichia coli | 07 06 | Mohsen et al. (2020) |
| Ag NPs-CIP | Ultrasonication-assisted drug loading | S. aureus–E. coli | 09 09 | Mohsen et al. (2020) |
| CIF@Ag NPs | Ultrasonication-assisted drug loading | Acinetobacter baumannii Serratia marcescens S. aureus | 28 31 32 | Ibraheem et al. (2022) |
| Ag NPs-PEG-CIP | Ultrasonication-assisted drug loading | A. baumannii S. marcescens S. aureus | 35 36 37 | Ibraheem et al. (2022) |
| HAPNWs@AgNPs-CIP | Sunlight-assisted Agnp immobilisation | E. coli S. aureus | 16 14 | Xiong et al. (2017) |
| Ag NPs | Chemical reduction | S. aureus Pseudomonas aeruginosa Bacillus subtilis Klebsiella pneumoniae E. coli | 14 11.5 111.5 11.5 12 | Present study |
| CIF@Ag NPs | Ultrasonication-assisted drug loading | S. aureus P. aeruginosa B. subtilis K. pneumoniae E. coli | 31 33.5 34 34 30 | Present study |
The enhanced antibacterial activity of CIP@Ag NPs can also be attributed to their ability to modulate multiple bacterial pathways. The combination of ciprofloxacin and silver nanoparticles not only inhibits bacterial DNA replication but also disrupts bacterial cell membranes and generates ROSs, leading to bacterial cell death. This multi-target approach makes CIP@Ag NPs particularly effective against a broad spectrum of bacteria, including those that have developed resistance to traditional antibiotics (Mohsen et al., 2020). Moreover, the targeted delivery of ciprofloxacin via silver nanoparticles ensures that the drug is released in a controlled manner, enhancing its bioavailability and therapeutic efficacy. This targeted approach not only improves the antibacterial response but also reduces the risk of systemic side effects associated with high doses of ciprofloxacin (Hussein-Al-Ali et al., 2022). The biocompatibility and low toxicity of silver nanoparticles further enhance the safety profile of CIP@Ag NPs, making them suitable for long-term use in treating chronic bacterial infections. In addition to their antibacterial properties, CIP@Ag NPs hold promise for treating infectious diseases where inflammation and infection often coexist (Arif et al., 2022). The antimicrobial properties of silver nanoparticles, combined with the antibiotic activity of ciprofloxacin, make CIP@Ag NPs a powerful tool for combating bacterial infections while simultaneously reducing inflammation. This dual-action mechanism is particularly beneficial in treating conditions such as sepsis, where uncontrolled inflammation and bacterial infection can lead to severe tissue damage and organ failure (Asadi et al., 2022). Additionally, Table 2 comparison of antibacterial activity with existing literature emphasises the efficiency of the CIP@Ag NPs complex, establishing it as a powerful tool in antimicrobial therapy, poised to revolutionise the treatment of resistant infections (Mohsen et al., 2020).
Ciprofloxacin, a fluoroquinolone antibiotic, is effective against a wide range of Gram-positive and Gram-negative bacteria. However, its clinical efficacy is often compromised due to the following limitations: Resistance development, where pathogens such as P. aeruginosa and Staphylococcus aureus have developed resistance mechanisms, including efflux pumps and mutations in DNA gyrase, reducing ciprofloxacin's effectiveness (Jaber & Saadh, 2024; Shariati et al., 2022); biofilm penetration, as ciprofloxacin struggles to penetrate biofilms that protect bacteria, diminishing its efficacy in treating biofilm-associated infections (Shariati et al., 2022); and toxicity and side effects, such as gastrointestinal disturbances, tendon damage, and potential neurotoxic effects, especially with prolonged use (Shariati et al., 2022).
Silver nanoparticles (AgNPs) possess broad-spectrum antimicrobial properties through mechanisms such as bacterial cell membrane disruption and the generation of ROSs. However, AgNPs face several challenges, including cytotoxicity, where high concentrations of AgNPs can exhibit toxic effects on human cells, limiting their standalone therapeutic use (Arif et al., 2022); environmental concerns, as the release of silver ions raises issues about environmental toxicity and bioaccumulation in aquatic ecosystems (Arif et al., 2022); and limited spectrum, where AgNPs may exhibit reduced efficacy against certain resistant bacterial strains without modification or combination with other agents (Arif et al., 2022).
The combination of ciprofloxacin and silver nanoparticles addresses these limitations, providing enhanced antibacterial efficacy through synergistic mechanisms. Studies have demonstrated that ciprofloxacin-loaded AgNPs show significantly improved efficacy against resistant pathogens compared to either agent used alone. This improvement is attributed to enhanced penetration, biofilm disruption, and sustained drug release (de Lacerda Coriolano et al., 2021; Mohsen et al., 2020). The synergistic effects arise from ciprofloxacin’s ability to inhibit bacterial DNA replication by targeting DNA gyrase, while AgNPs disrupt cell membranes and generate ROS. This dual action effectively overcomes resistance mechanisms (de Lacerda Coriolano et al., 2021; Mohsen et al., 2020). Moreover, by using AgNPs as carriers, the effective dose of ciprofloxacin can be reduced, minimising side effects associated with higher doses (de Lacerda Coriolano et al., 2021).
Mechanistically, the combination works by targeting multiple bacterial pathways. AgNPs facilitate ciprofloxacin delivery to infection sites, enhancing local drug concentrations and disrupting biofilms, thereby improving the efficacy of ciprofloxacin against biofilm-associated bacteria (de Lacerda Coriolano et al., 2021; Mohsen et al., 2020). This synergistic approach holds great promise for addressing the challenges posed by antibiotic resistance and warrants further exploration in clinical studies.
Conclusion
This study highlights the exceptional potential of ciprofloxacin-loaded silver nanoparticles (CIP@Ag NPs) as a powerful therapeutic platform with multifaceted biomedical applications. The CIP@Ag NPs demonstrated remarkable antimicrobial efficacy, exhibiting broad-spectrum antibacterial activity with inhibition zones reaching up to 34 mm against MDR strains such as S. aureus and E. coli. Notably, their effectiveness surpassed that of both silver nanoparticles and ciprofloxacin alone. Antioxidant assays confirmed the impressive DPPH radical scavenging activity (84%) and a fourfold increase in total antioxidant capacity, underscoring the role of CIP@Ag NPs in mitigating oxidative stress. Furthermore, anti-inflammatory studies revealed a striking 86.43% inhibition of albumin denaturation, positioning CIP@Ag NPs as potent anti-inflammatory agents. In addition, their significant inhibition of α-amylase and α-glucosidase enzymes demonstrates promising antidiabetic potential, offering a novel approach to managing postprandial hyperglycaemia. The multifunctional properties of CIP@Ag NPs suggest their potential as an integrative solution for treating infections, inflammation, oxidative stress, and diabetes. Future research, particularly in vivo studies, is essential to fully exploit their transformative therapeutic capabilities in diverse biomedical applications.
Data availability
All data generated or analysed during this study are included in this published article.
Author contributions
Conceptualization, I.L. and H.A.M.; Methodology, I.L.; Software, S.E.L.; Validation, H.A.M., S.E.L., and A.B.; Formal Analysis, M.M.S.A.; Investigation, A.B.; Resources, M.A.A.L.; Data Curation, Q.A.E.; Writing—Original Draft Preparation, I.L.; Writing—Review & Editing, H.A.M., S.E.L., and A.B.; Visualisation, T.T.; Supervision, S.E.L.; Project Administration, I.L.; Funding Acquisition, M.A.A.L. All authors have read and agreed to the published version of the manuscript.
Ibtissam Laib (Writing—original draft [equal], Writing—review & editing [equal]), Hamdi Mohammed (Investigation [equal], Writing—original draft [equal]), Salah Laouini (Supervision [equal], Validation [equal]), Abderrhmane Bouafia (Writing—original draft [equal], Writing—review & editing [equal]), Mahmood Abdullah (Investigation [equal], Methodology [equal]), Hamad Al-Lohedan (Investigation [equal], Methodology [equal]), Qudama Al-Essa (Conceptualization [equal], Formal analysis [equal]), and Tomasz Trzepieciński (Validation [equal], Visualisation [equal]).
Funding
The authors would like to thank the Algerian Directorate General for Scientific Research and Technological Development-DGRSDT for financial assistance, Laboratory of Biotechnology Biomaterial and Condensed Matter, Faculty of Technology, El-Oued University, El-Oued 39000, Algeria. The authors acknowledge the financial support through Researchers Supporting Project number (RSP2025R54), King Saud University, Riyadh, Saudi Arabia.
Conflicts of interest
The authors declare that they have no competing interests.
Acknowledgements
None declared.
References
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