System for the microbial synthesis of bioactive silver nanoparticles for applications in cancer treatment

Abstract

System for the microbial synthesis of bioactive silver nanoparticles for anticancer applications, including: • a microbial culture vessel for cultivating bacterial strains that produce extracellular metabolites; • a separation unit for obtaining a cell-free culture filtrate from the microbial culture; • a reaction chamber for mixing the cell-free filtrate with a silver precursor solution; • a nanoparticle formation module for the production of silver nanoparticles by reduction of silver ions using microbial metabolites in the filtrate; wherein the produced silver nanoparticles exhibit cytotoxic activity against cancer cells.

Description

SCOPE OF THE INVENTION

[0001] The present disclosure relates to a system for the microbial synthesis of bioactive silver nanoparticles using microbial metabolites and their application to exert an antitumor or anticancer effect.

[0002] In particular, the invention relates to a biological synthesis system configured for the production of silver nanoparticles by microbial reduction of metal ions using extracellular metabolites of bacterial strains. The produced nanoparticles exhibit improved biological properties and are suitable for biomedical applications, including cancer therapy. BACKGROUND OF THE INVENTION

[0003] Nanotechnology has developed into an important interdisciplinary field of research with applications in medicine, biotechnology, environmental science, and materials engineering. Due to their nanoscale dimensions, nanoparticles exhibit unique physicochemical properties, including an increased specific surface area, improved catalytic properties, and enhanced interaction with biological molecules.

[0004] Among various metallic nanoparticles, silver nanoparticles have garnered particular attention due to their antimicrobial, antioxidant, and anticancer properties. These nanoparticles have been investigated for various applications, including antimicrobial coatings, diagnostic biosensors, targeted drug delivery systems, and cancer therapy.

[0005] Conventional methods for synthesizing nanoparticles generally rely on chemical reduction processes using hazardous reducing agents such as sodium borohydride or hydrazine. These chemicals can generate toxic byproducts and limit the biomedical applications of the resulting nanoparticles.

[0006] In contrast, biological synthesis methods utilize microorganisms or plant biomolecules to convert metal ions into nanoparticles under mild and environmentally friendly conditions. Microorganisms such as bacteria, fungi, and algae produce enzymes, proteins, and metabolites that can reduce metal ions and stabilize the resulting nanoparticles.

[0007] Bacterial strains of the genera Pseudomonas and Bacillus are known to secrete extracellular biomolecules that can reduce silver ions to metallic silver nanoparticles. These biomolecules also act as stabilizing agents, preventing nanoparticle aggregation and improving their stability.

[0008] Despite these advantages, there remains a need for improved synthesis systems for the production of stable and biofunctional silver nanoparticles with increased biological activity.

[0009] Therefore, the present invention provides a system for the microbial synthesis of silver nanoparticles using extracellular metabolites and for evaluating their cytotoxic effect on cancer cells. SUMMARY OF THE INVENTION

[0010] The following summary is intended to provide a simplified overview of some aspects of the disclosed invention and is not intended to limit the scope of the invention.

[0011] Accordingly, the present invention provides a system for the microbial synthesis of bioactive silver nanoparticles using microbial metabolites and for the evaluation of their anticancer activity.

[0012] The proposed system utilizes extracellular metabolites from microbial cultures to reduce silver ions to nanoscale silver particles. Nanoparticle formation is confirmed by UV-visible spectroscopy, while structural characterization is performed using transmission electron microscopy (TEM) and X-ray diffraction (XRD).

[0013] In one embodiment of the invention, a bacterial strain of the species Pseudomonas fluorescens is cultivated in a nutrient medium to produce extracellular biomolecules that can reduce silver ions.

[0014] A cell-free culture filtrate is obtained by centrifugation and filtration of the microbial culture. Subsequently, a silver nitrate solution is added to the filtrate under controlled reaction conditions to enable the formation of nanoparticles.

[0015] The formation of silver nanoparticles is visually confirmed by a color change of the reaction mixture from light yellow to reddish-brown, which is due to the surface plasmon resonance of the silver nanoparticles.

[0016] The synthesized nanoparticles are characterized by UV-visible spectroscopy, revealing a characteristic absorption peak at approximately 435 nm. Morphological analyses using transmission electron microscopy show quasi-spherical nanoparticles with particle sizes between 22 nm and 75 nm.

[0017] Furthermore, the cytotoxic activity of the synthesized nanoparticles is evaluated by cell viability tests on cancer cells, revealing a dose-dependent inhibition of cancer cell growth. DETAILED DESCRIPTION

[0018] The embodiments of the present invention and their advantageous features are explained in more detail below.

[0019] To simplify the presentation, commonly known laboratory techniques and analytical instruments are not described in detail.

[0020] In one embodiment, a microbial strain of the species Pseudomonas fluorescens is used for the synthesis of the silver nanoparticles.

[0021] The bacterial culture is cultivated in a nutrient medium that includes the following components: Pepton - 6 g / L Yeast extract - 2 g / L Glucose - 9 g / L

[0022] The culture is incubated at approximately 32°C under shaking conditions of 170 rpm for approximately 36 hours to allow microbial growth and metabolite production.

[0023] After incubation, the culture is centrifuged at approximately 6500 rpm for 12 minutes to separate microbial cells from the extracellular medium.

[0024] The resulting supernatant, which contains extracellular metabolites, is collected and filtered through 0.22 µm membrane filters to obtain a sterile cell-free culture filtrate. Biosynthesis of silver nanoparticles

[0025] A freshly prepared 0.8 mM silver nitrate solution (AgNO3) is added to the microbial filtrate in various volume ratios, including: 1:3 1:6 1:8

[0026] The reaction mixture is incubated at approximately 29°C for 20 hours under dark conditions.

[0027] The formation of silver nanoparticles is indicated by a color change from light yellow to reddish-brown, which confirms the reduction of silver ions and the formation of nanoparticles. Optimization of reaction conditions

[0028] The synthesis conditions can be optimized by varying various parameters, including: pH range: 5,5 - 8,5 Temperature: 26°C - 35°C Response time: 8 - 36 hours

[0029] The highest nanoparticle yield is typically observed at approximately pH 7.0, 30°C and a reaction time of 18 hours. Spectroscopic analysis

[0030] UV-visible spectroscopy is used to confirm the formation of nanoparticles.

[0031] The absorption spectrum shows a characteristic surface plasmon resonance peak at about 433-435 nm, confirming the formation of silver nanoparticles. Morphological analysis

[0032] Transmission electron microscopy (TEM) is used to investigate particle morphology and size.

[0033] The results show quasi-spherical nanoparticles with sizes between 22 nm and 75 nm. Cytotoxic activity

[0034] The anticancer activity of the synthesized nanoparticles is investigated through cell viability tests.

[0035] The results show a dose-dependent reduction in cancer cell viability with increasing nanoparticle concentration.

[0036] The estimated IC 50The value is approximately 63 µg / mL, which indicates a significant cytotoxic effect against cancer cells.

Claims

System for the microbial synthesis of bioactive silver nanoparticles for anticancer applications, comprising: • a microbial culture vessel for cultivating bacterial strains that produce extracellular metabolites; • a separation unit for obtaining a cell-free culture filtrate from the microbial culture; • a reaction chamber for mixing the cell-free filtrate with a silver precursor solution; • a nanoparticle formation module for generating silver nanoparticles by reducing silver ions using microbial metabolites in the filtrate; wherein the generated silver nanoparticles exhibit cytotoxic activity against cancer cells. System according to claim 1, wherein the microbial strain comprises Pseudomonas fluorescens. System according to claim 1, wherein the formation of the nanoparticles is confirmed by a color change from light yellow to reddish-brown. System according to claim 1, wherein the synthesized nanoparticles have a surface plasmon resonance peak between 433 nm and 435 nm. System according to claim 1, wherein the nanoparticles have a quasi-spherical morphology with particle sizes between 22 nm and 75 nm. System according to claim 1, wherein the nanoparticles exhibit cytotoxic activity against cancer cells with an IC50 value of about 63 µg / mL. Bioactive nanoparticle composition comprising silver nanoparticles synthesized using microbial metabolites of a bacterial strain of Pseudomonas fluorescens, wherein the nanoparticles have a quasi-spherical morphology with particle sizes between 22 nm and 75 nm and exhibit cytotoxic activity against cancer cells.