Description:1. Background of the Invention
The rise of antimicrobial resistance (AMR) has significantly weakened the efficacy of conventional antibiotics, especially against multidrug-resistant (MDR) bacteria like Enterococcus spp., Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp. These pathogens contribute to prolonged hospital stays, increased healthcare costs, and higher mortality rates, necessitating innovative antimicrobial solutions. Silver nanoparticles (AgNPs) have emerged as promising alternatives due to their strong antibacterial properties, biofilm disruption ability, and low resistance potential. However, traditional AgNP synthesis methods often involve toxic chemicals, high costs, and environmental concerns, limiting their large-scale use.
To address these challenges, green synthesis approaches utilizing biological materials have gained traction. Quercetin, a naturally occurring flavonoid with antioxidant and antimicrobial properties, serves as an eco-friendly agent for AgNP synthesis. It enables the production of quercetin-functionalized silver nanoparticles (Q-AgNPs), which offer enhanced biocompatibility and sustainability. These nanoparticles are rigorously characterized using techniques such as UV-Vis, TEM, XRD, and FTIR, with studies confirming their antibacterial efficacy through bacterial membrane disruption, protein leakage analysis, and computational modeling.
Beyond medicine, Q-AgNPs show great potential in food preservation by preventing microbial contamination in packaging materials, thereby extending shelf life while maintaining safety standards. Additionally, they offer an eco-friendly alternative for mosquito control, demonstrating effectiveness against Aedes aegypti, the vector for dengue and Zika viruses.
By integrating green nanotechnology with antimicrobial strategies, Q-AgNPs provide a scalable and sustainable solution to global challenges in healthcare, food security, and pest management, all while minimizing environmental impact.

2. Materials and Method
2.1. Chemical required:
Silver nitrate (AgNO3), Quercetin (C15H10O7), DMSO (C2H6SO) and Methanol (CH4O) used in this current work were procured from sigma-aldrich, India.
2.2. Synthesis of Quercetin-Silver Nanoparticles (Q-AgNPs)
A novel method was adopted for synthesizing silver nanoparticles using quercetin. Quercetin is employed as both capping as well as reducing agent. A 1 mM quercetin (1 mM) solution is dissolved in DMSO and mixed with 100 mL AgNO3 dissolved in methanol solution in an Erlenmeyer flask at 30 ºC. The above solution is magnetically stirred at 300 rpm for 6 h, is kept in a dark condition and centrifuged at 10,000 rpm for 30 minutes. The supernatant liquid was removed and redisposed in deionized water. The process is repeated 3 to 5 times in order to remove the impurities, which can bind with Q-AgNPs. The absorbance value of the Q-AgNPs was evaluated using UV-VIS spectrometer and the stability was also monitored.
2. 3. Antibacterial Activity of Q-AgNPs
The well diffusion method was employed to evaluate the antibacterial activity of the synthesized Q-AgNPs against a panel of pathogenic bacteria, including S.a, E.c, K.p, and P.a. to assess antibacterial activity, 2 mL of nutrient broth was inoculated with equal volumes (100 μL) of each bacterial culture. Subsequently, varying concentrations of Q-AgNPs (25, 50, 75, and 100 μL) were introduced into the respective inoculated broths. Following incubation for 48 hours, each culture was plated onto nutrient agar plates to enumerate viable bacteria. Following incubation at 37°C for 48 hours, the Petri dishes were examined to enumerate colony-forming units (CFUs) of viable bacteria. Zones of inhibition (ZOIs) were then measured to quantify the antibacterial activity as described elsewhere. Each bacterial strain was assessed in six independent cultures with five replicates per culture.

2.4. Larvicidal bioassay:
Aedes aegypti larvae in their early fourth instar stage were collected from stagnant water sources in Chennai, India, and maintained in plastic enamel trays containing dechlorinated tap water at the Kings Institute of Preventive Medicines in Guindy, Chennai. The larvicidal activity of quercetin-silver nanoparticles (Q-AgNPs) was evaluated following a modified version of the WHO protocol from 1996, as previously described by Rahuman et al. A stock solution of Q-AgNPs was prepared by dissolving 1 mL of Q-AgNPs in 100 mL of pure water. For the initial bioassay, a 100 ppm solution was prepared by mixing the stock solution with dechlorinated tap water. Twenty larvae were placed in a beaker containing 249 mL of dechlorinated water and 1 mL of the test solution, with dechlorinated water serving as the control. After 24 hours of exposure, the number of dead larvae was counted, and the mortality percentage was calculated based on the average of five replicates. The initial test showed 100% mortality, prompting further dose-response bioassays.

For the dose-response bioassay, concentrations of 25, 20, 15, 10, and 5 ppm were prepared by diluting the stock solution. Twenty larvae were exposed to each concentration, with distilled water as the control. Mortality rates were recorded after 24 hours, with five replicates for each concentration. The study confirmed the larvicidal efficacy of Q-AgNPs against Aedes aegypti larvae, demonstrating dose-dependent toxicity and highlighting their potential as an effective larvicidal agent.
Summary of the Invention:
This invention focuses on the synthesis, characterization, and multifunctional applications of Quercetin-mediated Silver Nanoparticles (Q-AgNPs), leveraging the combined benefits of quercetin—a natural flavonoid with antioxidant and antimicrobial properties—and silver nanoparticles (AgNPs), renowned for their broad-spectrum antimicrobial activity. Employing a green chemistry approach, the synthesis yields monodisperse spherical nanoparticles averaging 30 nm in size, ensuring enhanced antibacterial, larvicidal, and cytotoxic properties. These attributes make Q-AgNPs highly suitable for diverse biomedical and environmental applications, including drug delivery, food packaging, and vector control.
Q-AgNPs present a promising solution across multiple fields, addressing critical challenges in healthcare, food safety, and pest management. As nanotherapeutic agents, they offer a targeted approach to combating bacterial infections, while their integration into food packaging materials effectively prevents microbial contamination, prolonging shelf life and ensuring safety. Furthermore, their eco-friendly insecticidal properties make them a valuable tool for controlling Aedes aegypti, a primary vector of dengue and Zika viruses, thereby contributing to sustainable vector management and public health initiatives. This innovation highlights the potential of Q-AgNPs as a scalable and environmentally responsible alternative for addressing global concerns in medicine, food preservation, and insect control.
3.1 Key Features and Advantages:
3.1.1. Antibacterial Activity:
Q-AgNPs demonstrate significant antibacterial efficacy against both Gram-positive (Staphylococcus aureus) and Gram-negative (Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa) bacteria. The Minimum Inhibitory Concentration (MIC) values for Q-AgNPs were determined to be 12.5 µg/mL for S. aureus and 6.25 µg/mL for Gram-negative bacteria. The nanoparticles disrupt bacterial cell membranes, leading to protein leakage and cell death. The antibacterial activity is further validated by Zone of Inhibition (ZOI) studies, where Q-AgNPs outperformed standard antibiotics like streptomycin.
3.1.2. Larvicidal Activity:
Fourth-instar larvae of Aedes aegypti were collected from static aquatic habitats in Chennai, India, and acclimatized in enamel-coated plastic containers filled with dechlorinated tap water at the Kings Institute of Preventive Medicines, Guindy. The larvicidal efficacy of Quercetin-functionalized silver nanoparticles (Q-AgNPs) was assessed using a modified 1996 WHO protocol, as adapted by Rahuman et al. A stock solution was synthesized by homogenizing 1 mL of Q-AgNPs in 100 mL of ultrapure water. For preliminary bioassays, a 100 ppm test solution was formulated by diluting the stock in dechlorinated tap water. Cohorts of 20 larvae were introduced into 249 mL of dechlorinated water amended with 1 mL of the test solution, while controls received dechlorinated water alone. Post 24-hour exposure, larval mortality was quantified by averaging five experimental replicates, revealing complete lethality (100%) in treated groups, necessitating dose-response analyses.
Subsequent bioassays evaluated concentrations of 25, 20, 15, 10, and 5 ppm, generated via serial dilution of the stock. Each concentration was tested against 20 larvae, with distilled water as the negative control. Mortality rates, recorded after 24 hours across five replicates per concentration, demonstrated concentration-dependent toxicity. The study validated Q-AgNPs as a potent larvicidal agent against Aedes aegypti, underscoring their potential for vector control applications.
, Claims:Claims
Claim 1: Green Synthesis of Quercetin-Silver Nanoparticles (Q-AgNPs)
A green eco-friendly process synthesizes silver nanoparticles based on the use of quercetin as reducing agent and also capping agent. A 1 mM quercetin in DMSO solution is added to 100 mL methanolic AgNO₃ and stirred at 30°C (300 rpm, 6 hours, dark). Q-AgNPs resulting from centrifugation of the mixture at 10,000 rpm for 30 minutes are washed and redispersed in water. Nanoparticle synthesis and stability are ascertained by UV-VIS spectroscopy.
Claim 2: Broad-Spectrum Antibacterial Efficacy
Q-AgNPs display effective antibacterial efficacy against Gram-positive (S. aureus, MIC: 12.5 µg/mL) as well as Gram-negative bacteria (E. coli, K. pneumoniae, P. aeruginosa, MIC: 6.25 µg/mL). Their efficacy against Gram-negative bacteria is outstanding with low MIC values versus Gram-positive bacteria.
Claim 3: Antibacterial Mechanism and Superiority to Antibiotics
Q-AgNPs cause bacterial membrane disruption and protein leakage leading to rapid bacterial death. They are superior in Zone of Inhibition (ZOI) tests to streptomycin with superior efficacy against antibiotic-resistant strains. This establishes Q-AgNPs as a contender to fight antimicrobial resistance.
Claim 4: Concentration-Dependent Larvicidal Activity
Q-AgNPs provide 100% mortality in Aedes aegypti larvae at 100 ppm (24-hour exposure). Dose-response assays (5–25 ppm) establish concentration-dependent toxicity confirmed using a modified WHO protocol. Their efficacy indicates mosquito vector control implications in public health scenarios.
Claim 5: Multifunctional Food Packaging Applications and Challenges
Q-AgNPs upgrade food packaging with antimicrobial activity, UV stability, and barrier property improvement (oxygen/moisture permeability reduction). Spoilage detection smart packaging and biopolymer (e.g., chitosan) compatibility for sustainable application are facilitated. Challenges are nanoparticle migration prevention, safety certification, cost minimization, and environment/regulatory issues.