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Home > Books > Silver Micro-Nanoparticles - Properties, Synthesis, Characterization, and Applications

Silver Nanoparticles: Properties, Synthesis, Characterization, Applications and Future Trends

Submitted: 09 October 2020 Reviewed: 30 June 2021 Published: 06 August 2021

DOI: 10.5772/intechopen.99173

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Silver Micro-Nanoparticles - Properties, Synthesis, Characterization, and Applications

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Nanotechnology is an expanding area of research where we use to deal with the materials in Nano-dimension. The conventional procedures for synthesizing metal nanoparticles need to sophisticated and costly instruments or high-priced chemicals. Moreover, the techniques may not be environmentally safe. Therefore “green” technologies for synthesis of nanoparticles are always preferred which is simple, convenient, eco-friendly and cost effective. Green synthesis of nanoparticle is a novel way to synthesis nanoparticles by using biological sources. It is gaining attention due to its cost effective, ecofriendly and large scale production possibilities. Silver nanoparticles (AgNPs) are one of the most vital and fascinating nanomaterials among several metallic nanoparticles that are involved in biomedical applications. It has vital importance in nanoscience and naomedicines to treat and prevent vital disease in human beings especially in cancer treatment. In current work we discussed different methods for synthesis of AgNPs like biological, chemical and physical along with its characterization. We have also discussed vital importance of AgNPs to cure life threatnign diseases like cancer along with antidiabetic, antifungal, antiviral and antimicrobial alog with its molecular mode of action etc. Finally we conclude by discussing future prospects and possible applications of silver nano particles.

• green synthesis
• silver nanoparticles
• nonmaterials
• anticancer and antidiabetic

Author Information

Sunil t. galatage *.

• Sant Gajanan Maharaj College of Pharmacy, Mahagaon, Maharashtra, India

Aditya S. Hebalkar

Shradhey v. dhobale, omkar r. mali, pranav s. kumbhar, supriya v. nikade, suresh g. killedar.

*Address all correspondence to: [email protected]

1. Introduction

Currently, improving and protecting our environment using green chemistry have become important issues in many fields of research. The most promising approach for generating new fields in biomedical sciences is the pharmaceutical application of nanoparticles (NPs) [ 1 ]. Due to ascension of industrial era and explosion of world population large amount of hazardous chemicals and gases released in environment in which adversely affecting our nature. Due avoid this and to protect our nature currently we world is focusing on development of natural products nanoparaticles. Biomolecules are highly compatible with nanotechnology which makes unique assembly for development of metal nanoparticles of biological molecules which are authentic and coast effective [ 2 ]. From the ancient era noval medicinal potential of silver has been known and proven for its antimicrobial potential [ 3 ]. Silver nano particles (AgNPs) and its related products were tremendously venomous and showed broad spectrum antibacterial potential against sixteen bacterial species [ 4 , 5 ]. Nanotechnology is future era in material science which develops and upgrades qulaites of particles such as size, and morphology which provide entry of nonmaterial in future quality material building in almost every field [ 6 ]. Nanotechnologies have been used to develop nanoparticles-based targeted drug carriers [ 7 ]. Metal nanoparticles have a high specific surface area and a high fraction of surface atoms because of the unique physicochemical characteristics of nanoparticles [ 8 , 9 ]. In that they include catalytic activity, optical properties and electronic properties, antibacterial properties, and magnetic properties [ 10 , 11 ]. The nanoscale materials have emerged as novel “antimicrobial agents” due to their high surface area to volume ratio and their unique chemical and physical properties [ 12 , 13 ]. In recent years development of metallic nanoparicles is an emerging field of research in material science. Crystalline nanosilver gained prime importance and has superior applicability in detection of biomolecules, antibacterial, electronics, diagnostic applications in health care system etc. Apart from novel applicability of AgNPs researchers still in search of advance methods to synthesize eco-friendly and coast effective tools to develop AgNPs [ 14 , 15 ]. As silver posse’s broad spectrum potential against bacterial and microbial species which specially utilized in industries it has key role in healthcare systems [ 16 ]. Nitrate group of silver potentially responsible for its broad spectrum antibacterial potential and as it convert in to AgNPs surface area is drastically increased which improve microbial exposure time and area [ 17 , 18 , 19 ]. Different techniques are available to synthesize AgNPs such as physical, chemical and biological. Though chemical method is rapid it utilizes capping agents for synthesis which is costly and produces adverse and toxic effects. This demands development of safe, ecofriendly, coast effective tool for synthesis of AgNPs and focused on biological methods such as green synthesis which is non toxic and developed using plant origin materials and overcomes disadvantages of earlier approaches. Moreover, use of plant extracts also reduces the cost of microorganism’s isolation and culture media enhancing the cost competitive feasibility over nanoparticles synthesis by microorganisms [ 20 ]. Applicability of AgNPs is primly due to its nanoscale size and shape as compared to bulk. Due to these unique properties researchers are hunting of novel methods to synthesize AgNPs with prissily controllable size and shape [ 21 , 22 , 23 , 24 ]. Apart from excellent inhibitory potential of AgNPs in recent years most of the pathogenic bacteria developed resistant against it which is major concern of health care system. Chemical and physical approaches consumes ample of time, energy, money and generate toxic side effects. Nowadays green synthesis utilize microbes, fungi and medicinal plants which are easily available, convenient to handle and wide source of metabolites to synthesize AgNPs gained prime importance due to its nontoxic and ecofriendly properties [ 25 ]. Currently AgNPs are synthesized from natural herbs having medicinal potential such as synthesis of various metal nanoparticles using fungi like Aspergillus terreus , Paecilomyces lilacinus and Fusarium [ 26 ]. Penicillium sp. [ 27 ] Fusarium oxysporum [ 28 ] and Euphorbia hirata , green tea, neem, starch aloevera, lemon etc. [ 29 , 30 , 31 , 32 ]. AgNPs mainly binds to cell wall and penetrate deep inside the cell wall which produces cellular damage by interacting with DNA, proteins inside the cell which leads to cell death [ 33 , 34 , 35 , 36 , 37 ].

2. Need for green synthesis and silver nanoparticles

Silver is a basic element which is non-toxic belonging thermal and electrical potential [ 38 ]. Silver demand will likely to rise as silver find new uses, particularly in textiles, plastics and medical industries, surgical, dental resigns, coated water filters, sanitizers, detergents, soap and wound dressings. Applicability in healthcare for treatment of mental illness, convulsions, de addiction of narcotic products along with sexually transmitted diseases like syphilis and gonorrhea leads to changing the pattern of silver emission as these technologies and products diffuse through the global economy [ 39 , 40 , 41 ]. Green synthesis is an emerging approach which overcomes demerits of physiochemical approaches by utilization of natural herbs which are nontoxic [ 42 , 43 ]. Green synthesized nanosilver offer many advantages like utilization of phytochemicals, antioxidants acts as naturally occurring reducing agents, coast efficient, large scale manufacturing highly beneficial and usage of toxic chemicals, high pressure, energy are avoided. Nanosilver can be engineered by different techniques such as irradiation, reduction, electrochemical and chryochemical synthesis. Nanosilver can be molded in to desired shapes and bear unique properties like permeability by pH and dissolved ions as compare to routine metals [ 44 , 45 ]. As AgNPs generate larger surface area per unit mass which improves contact time nanosilver customer market an demand drastically raised in wide verity of industries along with healthcare, food packing, textiles, cosmetics etc. [ 46 ].

3. Silver nano particles

Generally AgNPs are nanoparticles of silver having size range between 1 and 100 nm in size having unique properties such as electrical, optical and magnetic having wide range of applicability [ 47 ]. Green chemistry is and encouraging approach mainly utilize nanosilver along with natural biomolecules such as polysaccharides, tollens which overcomes drawbacks of conventional methods and produce AgNPs which are ecofriedly, nontoxic and coast effective [ 48 , 49 ]. Metallic silver ions are inactive but once it come contact with reducing agent ionization occurs and it get converted in its active form. Ionic silver is active form of silver which binds to cell wall of bacteria leading to major structural changes in cell morphology. AgNPs causes de-naturation of RNA and DNA replication which further leads to cell death [ 50 ]. Silver is also called as oligodynamic due to its bactericidal potential at minimum concentration. That’s why it has been largely used in medical products [ 51 , 52 ].

4. Methods for synthesis of silver nanoparticle

4.1 physical approaches.

In physical approach of synthesis of AgNPs evaporation and condensation has major importance. Temperature gradient play important role in cooling of vapors at desired rate. A chance of contamination by solvent has been removed by physical approach as no solvent has been used in physical method and uniform distribution of particle size precisely obtained [ 53 , 54 ]. Minimum inhibitory concentration in toxicity studies can be easily achieved by production of nano scale nanoparticles in high concentration [ 55 ]. AgNPs also synthesized by laser ablation of metallic particles [ 56 ]. One important advantage of laser ablation technique compared to other methods for production of metal colloids is the absence of chemical reagents in solutions. Therefore, pure and uncontaminated metal colloids for further applications can be prepared by this technique [ 57 ]. Wide range of material can be synthesized in nanoparticels by physical method such as Au, Au and PbS etc. Synthesis of AgNPs by tube furnace has ample of disadvantages such as require larger space, high power, rapid rise of environmental temperature etc. AgNPs synthesized by laser ablation strongly depend on laser wavelength, time of laser pulse, laser fluence, the ablation time duration and the effective liquid medium. Ejection of AgNPs synthesized by laser ablation requires little power and particle size is precisely depends on laser fluence. However morphology, size and shape of AgNPs mainly depend on contact of laser light passing. Also, the formation of nanoparticles by laser ablation is terminated by the surfactant coating. The nanoparticles formed in a solution of high surfactant concentration are smaller than those formed in a solution of low surfactant concentration. One advantage of laser ablation compared to other conventional method for preparing metal colloids is the absence of chemical reagents in solutions. Therefore, pure colloids, which will be useful for further applications, can be produced by this method [ 58 , 59 ].

4.2 Chemical approaches

Chemical reduction is the most frequently applied method for the preparation of AgNPs as stable, colloidal dispersions in water or organic solvents. Most commonly used reductant is citrate. In aqueous solution reduction of silver occurs and nanosize colloidal silver ions are generated. Stability of any colloidal dispersion has prime importance and which could be achieved by stabilizing agent (dodecanethiol) which adsorbed on surface and produce protective sheath. It can avoid agglomeration and crystal growth of the system. During the synthesis of AgNPs minute changes in parameters (Polymers) makes drastic changes in size, shape, morphology, polydispersibility index, self assembling and zeta potential (Stability). Frequently used ingredients in synthesis of AgNPs and AuNPs are glycol derivatives Polyvinyl pyrrolidone (PVP) and Polyethylene glycol (PEG). Polyacrylamide play dual function such as reducing and stabilizing agent in synthesis of AuNPs [ 59 , 60 ]. Surfactants containing functional groups such as amines, thoils and acids play important role in stability of colloidal dispersion which protects the system from crystal growth, coaleseces and agglomeration. Currently AuNPs developed by modified tollens method utilize saccharides and silver hydrosols and reducing agent which yield AgNPs in the range of 50–200 nm and 20–50 nm respectively [ 61 ].

4.3 Biological approaches

Biotechnology is an emerging tool to develop biological synthesis of AgNPs. Besides this magnetic nanoparticles has great antibacterial potential due to improved surface area to treat raised microbial resistant against many antibiotics and medicines [ 62 ]. Currently green chemistry is rapidly growing technique utilized for synthesis of AgNPs with naturally occurring stabilizing, reducing and capping agents to synthesize AgNPs without toxic adverse effects [ 63 ]. Reduction of metal ions by combined efforts of herbs and certain enzymes, proteins, microorganisms, bacteria and fungi etc. in biological synthesis has been successfully reported [ 64 ].

4.4 Synthesis of silver nanoparticles by fungi

High production yield AgNPs synthesized by fungi obtained when compared to bacteria due to fungi secret higher amount of proteins that directly responsible for increased production [ 65 ]. Higher production rate is mainly due to silver ions entered in to fungal cell wall which leads to reduction of silver ions by fungal enzymes such as naphthoquinones and anthraquinones [ 66 ]. Slower rate and process is only disadvantage associated with fungal synthesis of AgNPs hence green synthesis approach is more preferred over the other techniques [ 67 ].

4.5 Synthesis of silver nanoparticles by bacteria

Pseudomonas stutzeri which is the first strain of bacteria form which AgNPs were synthesized and isolated form Ag amine [ 68 ]. Many of the bacterial strains and microorganism developing resistance to metal at lower concentration. Resistance mainly produced due to efflux, change in solubility, toxicity via oxidation/reduction and precipitation of metals [ 69 ]. There are evidences that at lower conc. Microorganisms are alive but once exposed to high conc. Metal ions leads to microbial death. In biosynthesis of silver enzyme nitrate reductase convert nitrate to nitrite [ 70 ].

4.6 Synthesis of silver nanoparticles by plants

Green synthesis is an excellent tool that can be utilized for synthesis of AgNPs as it uses natural origin medicinal herbs and its extracts which contain wide range of metabolites specifically water soluble flavones, quiones causes rapid rapid and quick reduction of silver when compared to fungi and microbes. Green chemistry approach is safe, cosat efficient, easily scalable to mass productions, easily availability of raw materials at cheaper coast. Phytochemicals directly take part in reduction process of the silver ions a during synthesis of AgNPs ( Figure 1 ) [ 71 ].

Biological methods of silver nanoparticles.

5. Mechanism of action of silver nanoparticles

5.1 agnp’s antimicrobial moa.

When AgNP reaches toward cell they release Ag+ ions. These released ion then interact with sulfur and phosphorus containing compound present in cell wall. This lead to disarranged cell wall formation and small pits forms in the cell wall. Formed pit gives access to entry of ions and other foreign material to entry into cell. This increase intracellular osmotic pressure. As pressure built up in the cell, it begins to swell. Finally all these event lead to bursting of cell wall and cell lysis take place. This type of antimicrobial activity is more in gram −ve cell than gram +ve cell. As gram +ve cell have more cross linked peptidoglycan layer and teichic acid in their cell wall. The gram −ve cell have less or no peptidoglycane layer and have more lipopolysaccharide in their cell wall. So the AgNP’s easily interact with gram −ve cell due less barrier [ 72 ].

5.2 AgNP’s anticancer MOA

As described in above when pit formation takes place in the cell wall, the Ag+ ions released by AgNP’s get entered into cell. Then they reaches to mitochondria where they interact with thiolgoups and bind to NADPH dehydrogenase enzyme and liberates ROS. These formed ROS in mitochondria interacted with respiratory enzymes damage ATP formation and respiratory cycle of cell. Formed ROS also interact with protein, sulfur and phosphorus containing cell constituent. Also these formed ROS also bind to phosphorus elements of DNA and RNA which lead to inhibit cell replication and protein synthesis. Due to binding with DNA aggregation of damage protein sysnthesis which lead to cell death. Another possible action is by autophagy. AgNP’s have ability to induce autophagy by accumulation of autophagolysosmes in human ovarian cancer cell. This autophagy work by mainly 2 ways; at lower level they increases cell life i.e. surviving rate, but when its level increase it lead to cell death ( Figure 2 ) [ 73 ].

Anticancer mechanism of action of silver nano particles.

6. Factors affecting bactericidal potential of AgNP’s

Primarily morphology i.e. size and shape along with reactivity of AgNP’s were responsible for bactericidal potential of AgNP’s. Size and surface are inversely proportional to each other as size decreases area increases leads of rapid rise in surface-area to volume ratio. Bactericidal potential inhibit cell wall and free radicals Ag-thiol groups of enzymes Preventing biofilm formation Intercalates between bases Attaching to the surface of the cell membrane Bacterial peptides that can affect cell signaling Attaches to 30 s subunit ( Figure 3 ). Silver nanoparticles showing multiple bactericidal actions [ 74 ].

Factors affecting to the bactericidal effect of silver nanoparticles.

7. Charactrisation of AgNP’s

7.1 visual and uv: visible study.

To ascertain either AgNPs are developed or not visual and calorimetric appearance of samples checked by UV–Visible spectrophotometer before and after formulation of AgNPs at different time intervals. Before synthesis of AgNPs silver nitrate is colorless and herbal extract has definite color. Once AgNPs synthesized silver nitrate solution develop yellowish brown color after interacting with herbal extract which is confirmed by surface Plasmon resonance SPR and UV visible absorption in the specific range of 400–475 nm [ 75 ].

7.2 FTIR analysis

FTIR spectroscopy is an investigational tool to determine/conform functional groups priesnt in the moiety which is characteristic of that compound. Major functional moieties present in AgNPs and herbal extract were identified by scanning the samples in the range of 4000 to 400 cm−1 [ 76 ].

7.3 SEM/TEM analysis

Scanning electron microscopy/Transmission electron microscopy mainly used to study surface morphology of synthesized AgNPs. SEM/TEM plates were prepared by addition of silver nitrate to develop smear of solution on slides. Conductivity was incorporated in system by making thin film of platinum which was coated on slides. Once the slides were ready they were scanned at 20 KV accelerating voltage and high quality images were captured [ 77 ].

7.4 X-ray diffraction (XRD) analysis

X-ray diffraction is a modern technique mainly utilized to identify state of matter either it is crystalline or amorphous in nature at different radiation angles. X-ray diffraction determines phases either crystalline/amorphous and cell dimension [ 78 ].

8. Application of silver nanoparticles

8.1 antimicrobial activity.

Products prepared with silver nanoparticles have been permitted by no. of accredited bodies including USFDA, USEPA, Korea’s testing body and SIAA of japan institute of research. Antimicrobial and antimicrobial potential potential of AgNPs containing silver sulfadiazine is incorporated in to medicines and used in burns to avoid infections. Nowadays AgNPs involved in extending field of nanotechnology and appears in many consumer products that include acne vulgaris cream and for deodorizing sprays. The antimicrobial properties of silver nanoparticles depend on size, environmental conditions (size, Ph, tonic strength) and capping agent. Recently an improvement in antimicrobial activity synergistic effect has been reported when silver naqnoparticles combined with ampicillin, amoxicillin and chloramphenicol on the contrary reports showed antagonistic interaction between silver nanoparticles and amoxicillin or oxacillin antibiotic combined with silver nanoparticles have suggested improve therapeutic activity ( Figure 4 ) [ 79 , 80 ].

Applications of silver nanoparticles.

8.2 Antiviral activity

Antiviral activity of silver nanoparticles have proven to exert antiviral activity against HIV-1 at non cytotoxic concentration but the mechanism underlying their HIV inhibitory activity has not been fully elucidated. The study from intranasal silver nano particles administration in mice increased survical, lower lung viral titer levels, minor pathologic lesions in lung disease, and remarkable survival benefit after infection with the H3N2 influenza virus, suggesting that AgNPs had significant role in mice survival. Biologically prepared silver nano particles inhibited the viability in herpes simplex virus (HSV) types 1 and 2 and human para influenza virus type 3 based on size and zeta potential. The treatment of vero cells with non-cytotoxic concentrations of silver nanoparticles significantly inhibited by the replication of paste des petits ruminants virus (PPRV). The mechanisms of viral replication are due to the interaction of silver nanoparticles with the virion core. Tannic acid mediated synthesis of various various sizes of silver nanoparticles capable of reducing HSV-2 infectivity both in in-vitro and in-vivo through direct interaction, blocked virus attachment, penetration and further spread [ 81 , 82 ].

8.3 Antibacterial activity

Silver nano particles are one of the most attractive nonmaterial’s for commercialization applications. As antibacterial agents silver nanoparticles were used for wide range of applications from disinfecting medical devices and home appliances to water treatment. AgNPs promisingly used in drastic fields such as healthcare products, food storage, textile and medicinal devices. In antibacterial potential AgNPs free silver ions are released at slower rate along with higher surface area which produces noxious environment and this is the main reason for broad spectrum antibacterial potential of AgNPs [ 83 ].

8.4 AgNP’s in cancer control

AgNPs has prominent anticancer potential as it discourage mitochondrial respiratory chain, increase reactive oxygen species (ROS) rate of synthesis which finally leads to DNA damage and cancerous cell death. Yu-Guo Yuan in 2018 revealed that the combination of camptothecin and silver nanoparticles treatment significantly increases the levels of cancer cells. It increases oxidative stress markers and decrease ant oxidative stress markers compared to single treatment. Overall these results suggested that camptothecin and silver nanoparticles cause cell death by inducing the mitochondrial membrane permeability change and activation of caspase. The synergistic cytotoxicity and apoptosis effect seems to be associated with enhanced ROS formation and depletion of antioxidant. Certainly a combination of CPT and silver nano particles provide advantageous effect in treatment of cervical cancer compared to immunotherapy [ 84 ].

8.5 Antidiabetic activity of AgNP’s

Tephrosiatinctoria stem extracts mediated silver nano particle synthesis was evaluated for control of blood suger levels. AgNP’s scavenged free radicals, reduced the levels of enzymes that bring about hydrolysis of complex carbohydrates ( α − glucosidaseα − amylase ) and as a result of which there is an increase in consumption ratr of glucose. The promising antidiabetic activity of shown by Ananascomosus (L.) silver nanoparticles. In dose dependent manner. AC-AgNP’s inhibit α-glucosidase enzyme in stomach. Which is helpful in non-insulin diabetic patient. Also the silvernanoparticles synthesized with Argyreia nervosa leaf extract shown great antidiabetic activity. They inhibit mainly enzymes that digest the carbohydrates into monosaccharide and reduce blood glucose level [ 85 , 86 ].

8.6 Different field application of AgNp’s

Studies can contracting on the therapeutic applications of AgNP’s in the gastrointestinal tract have displayed that gastric cells can be sensitized to radiation by the use of AgNP’s and they may bypass the stomach and instead release the drug in small intestine. Apart from the health related applications; Silver Nanoparticles are act as a brilliant heterogeneous catalyst used for reduction of halogenated organic pollutants. Also it increases the bleaching power of organic dyes. The tubular shaped silver Nanoparticles have a very potent catalytic activity so they can used as a catalyst. In case of water treatment when the biosynthesized Silver Nanoparticles which are biologically synthesized on nitrocellulose membrane filters, can used for the promising inhibition and inactivation of microbes like E. coli and Enterococcus faecalis , etc. Rather as the silver Nanoparticles are the very good antimicrobial agents so they are used as the preservatives in various food and agricultural products [ 87 ].

8.7 Antifungal activity of AgNPs

AgNP’s play important role as antifungal agents against various diseases caused by fungi. Biologically synthesized AgNP’s shows enhanced antifungal activity with fluconazole against phomaglomerata , Candida albicans species. AgNP’s stabilized by sodium dodecyl sulphate showed greater antifungal activity against Candida albicans compared to conventional antifungal agents. The AgNP’s synthesized by bacillus species exhibit strong antifungal activity against the plant pathogenic fungus fusariumoxysporum at concentration of 8 μg/ml. AgNP’s shown promising antifungal activity on T. asahii with MIC of 0.5ug/ml by damaging cell wall and components of cell. Due to size of nanoparticles they easily penetrate into cell. Where it binds to different cell components and inhibits cell functions. In combination with antimicrobial agents like ketoconazole shown great antifungal activity with MIC less than 0.5 mg/ml against the Malassezia where they give synergistic effect with ketoconazole it form pores in cell to show antifungal activity [ 88 , 89 ].

8.8 Anti angiogenic activity of AgNP’s

Antiangiogenic potential of green synthesized AgNP’s in retinal endothelial cells model mainly produced by inhibition, proliferation and migration of BRECs at 500 nM concentration. In CAM model (chicken embryo chorioallantoic membrane) the silver nanoparticles inhibit angiogenesis approximately up to 73%. In comparison to other antiangiogenic molecules. They give dose dependent cytototoxic action on endothelial cell present in blood vessels to inhibit formation of new blood vessel in tumor region. Also the by using the same model i.e. by CAM assay the silver nanoparticle synthesized by Rubinatinctorum shown antigiogenic activity. Ru-AgNP’s shown inhibitory action on blood vessels. In CAM model, there is decrease in length of embryo resulted out due to the antiangiogenic action of Ru-AgNP’s [ 90 , 91 ].

8.9 Diagnostic, biosensor and gene therapy applications of AgNP’s

Nanoparticles have advantage over today’s therapies because they can be engineered to have certain properties or to in certain way. They are helpful in cellular imaging. Silver plays an important role in imaging systems due to its stronger and sharper Plasmon resonance. Currently biosensor made with silver used as powerful tool to detect cytochrome P53 of squamous cell cancer of head and neck. Due to the colorimetric sensing property the silver Nanoparticles are applicable to detect the heavy metal ions of nickel, cobalt and mercury along with the sulfide traces. Among all the types of silver Nanoparticles, especially the triangular shaped silver Nanoparticles have higher anisotropy and lightening rod effect which leads to its wide use in manufacturing of Plasmon sensors or Plasmon detectors which are used to detect the mercurial ions in the solution. Also the silver Nanoparticles are used to develop the electrochemical sensor which is used to detect common herbicide atrazine. On the other hand the in situ growth and development of silver Nanoparticles on polydopamine traced filter paper is responsible for the quick collection and detection of green colored residue of malachite [ 92 ].

8.10 Anti-inflammatory activity of AgNP’s

AgNPs have been known for its antimicrobial but the anti-inflammatory response is still limited. Rats treated intra colonic ally with 4 mg/kg or orally with 40 mg/kg of nanocrystalline silver (NP32101) showed significantly reduced colonic inflammation. AgNPs showed rapid healing and improved cosmetic appreance occurring in dose dependent manner. Silver Nanoparticles made by using the extraction method with petroleum ether and some small amount of ethyl acetate are having potent cyclooxigenase-2 inhibition property. So, as one can add the natural extract of anti-inflammatory activity to this silver Nanoparticles extracted with petroleum ether, the anti-inflammatory activity of the resulted silver Nanoparticles get increased. Recently some scientists were done the extraction of soft coral named nephthea sp. Which already possessing the anti-inflammatory activity and extracted the silver Nanoparticles with petroleum ethers then the produced silver Nanoparticles of nephthea sp. having very potent anti-inflammatory activity which were estimated by analysis and molecular docking methods [ 93 ].

9. Future prospects

AgNPs has potential applications in healthcare system and treating infectious diseases and it is emerging as remedies for large no of resistant bacteria infections along with it is known for its anti-inflammatory potential. Apart from it has numerous application in biological and research fields such as electrochemistry, biochemistry, nanoprism synthesis, garments, detergents and soap industry, involved in devising water purification system, and surgical instrument. Nowadays Ag-NPs opened new era as it has used in artificial implants which decreeing dependency on antibiotics. Studies have been revealed that Ag-NPs have novel potential in development of new pharmaceutical dosage forms and AgNPs cures inflammation of bladder which tremendous application in healthcare systems. AgNPs useful in animal models for detection of biosensors [ 94 ]. A reliable mechanism responsible for the impressive biological activity of AgNPs is considered to be a key factor in future research. Wide scope to aware control the release of silver and improving the stability of AgNPs used in medical and mechanical devices.

10. Concslusion

Over the past few decades, nanoparticles of noble metals such as silver exhibited significantly distinct physical, chemical and biological properties from their bulk counterparts. Current chapter specifically encounters synthesis, characterization, and bio-applications of silver nanoparticles, with special emphasis on anticancer, antimicrobial activity and its mechanisms. Green chemistry is being exploited for developing silver nanoparticles. Several methods utilized to create silver nanoparticles utilizing plant extracts as reducing or capping agents. Current chapter represents different methods of preparation silver nanoparticles and application of these nanoparticles in different fields.

Acknowledgments

The authors are thankful to Department of Pharmaceutics Sant Gajanan Maharaj College of Pharmacy Mahagaon and Trustees of Sant Gajanan Maharaj College of Pharmacy Mahagaon for providing required guidance and support for completion of this work.

Conflict of interest

The authors declare no conflict of interest.

Notes/thanks/other declarations

Special thanks to Shivtej for continuous support throughout the work.

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Materials Advances

Green synthesis of silver nanoparticles: methods, biological applications, delivery and toxicity.

* Corresponding authors

a Department of Medicinal Chemistry, Faculty of Ayurveda, Institute of Medical Sciences, Banaras Hindu University, Varanasi, India E-mail: [email protected]

b Department of Microbiology, Institute of Medical Sciences, Banaras Hindu University, Varanasi, India

The advent of nanotechnology profoundly transformed the pharmaceutical sciences and greatly enhanced the diagnostics and treatment of various diseases that threaten human life. Several metallic nanoparticles are extensively used as nanomedicines due to their potential therapeutic applications. Among them, silver nanoparticles are remarkable due to their unique chemical and physical properties. This review discusses types of nanoparticles, and green synthesis methods along with their reduction mechanisms, involving economically viable reducing materials like algae, seaweeds and flowers. Apart from environment-friendly methods, several biological activities such as wound healing, antibacterial, antifungal, anti-tumour, anti-viral, etc. , are described in detail. Consequently, we have focused on how silver nanoparticles enhance targeted drug delivery and the mechanism of drug release along with their toxic effects.

• This article is part of the themed collection: Recent Review Articles

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Vidyasagar, R. R. Patel, S. K. Singh and M. Singh, Mater. Adv. , 2023,  4 , 1831 DOI: 10.1039/D2MA01105K

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Foeniculum Vulgare leaf extract loaded synthesis of silver nanoparticles in different volume ratios for antimicrobial and antioxidant activities: Comparative study of composition

• Original Paper
• Published: 16 August 2024

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• Eneyew Tilahun Bekele   ORCID: orcid.org/0000-0003-1308-0292 1 ,
• Fasika Dereje Ambecha 1 ,
• C. R. Ravikumar 2 ,
• Taymour A. Hamdalla 3 ,
• H. C. Ananda Murthy 4 , 5 &
• Defaru Negera Duke 1  

The current world is exposed to immense classes of challenges, of which antimicrobial-caused infectious diseases have been ranked the third killer disease due to their high resistance capability. Oxidative stress is also the other problem faced by the current world. In the current study, a leaf of Foeniculum Vulgare was employed for the synthesis of silver nanoparticles (Ag NPs) within the 1:3, 1:1, and 3:1 compositions using 0.1 M of AgNO 3 . The calculated average crystalline size from X-ray diffraction (XRD) was found to be 12.6, 13.7, and 21.6 nm for the 1:3, 1:1, and 3:1 volume ratios, respectively. Scanning electron microscope coupled with energy dispersive spectroscopy (SEM-EDS) analysis depicts the quasi-spherical shape with an intense Ag peak at around 3.00 eV. Transmission electron microscope coupled with high-resolution transmission microscope with surface area electron diffraction pattern (TEM-HRTEM with SAED) showed spherical shaped Ag NPs. The electronic bandgap energy was calculated as 3.1, 3.2, and 3.3 eV for the 1:1, 3:1, and 1:3 volume compositions, respectively. Ag NPs show 13.5, 12.5, and 11.0 mm zones of inhibition for the 1:3, 1:1, and 3:1 ratios, respectively. The antifungal activity was found to be 16.9, 11.2, and 10.5 mm for the 3:1, 1:3 and 1:1 ratios, respectively. Lastly, the antioxidant activity was estimated to be 42.4, 28.94, and 27.39% RSA for the 1:1, 3:1, and 1:3 volume ratios respectively. All the ratios of Ag NPs showed promised antimicrobial and antioxidant activity in the presence of 2, 2-Diphenyl-1-picrylhydrazyl (DPPH) due to the enhanced generation of reactive oxygen species (ROS).

Graphical Abstract

Green Synthesis of Ag NPs within three volume ratios in the presence of Foeniculum vulgare leaf extract.

Investigating the potential antibacterial, antifungal, and antioxidant activities.

Comparative study of the volume ratios on the antimicrobial and antioxidant activities.

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Authors greatly acknowledge Adama Science and Technology University, Ethiopia, and Department of Research Centre, Department of Science, East-West Institute of Technology, Bangalore 560091, India.

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Eneyew Tilahun Bekele, Fasika Dereje Ambecha & Defaru Negera Duke

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Bekele, E.T., Ambecha, F.D., Ravikumar, C.R. et al. Foeniculum Vulgare leaf extract loaded synthesis of silver nanoparticles in different volume ratios for antimicrobial and antioxidant activities: Comparative study of composition. J Sol-Gel Sci Technol (2024). https://doi.org/10.1007/s10971-024-06476-9

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Antibacterial activity and characteristics of silver nanoparticles biosynthesized from Carduus crispus

• Enerelt Urnukhsaikhan 1 ,
• Bum-Erdene Bold 1 ,
• Aminaa Gunbileg 1 ,
• Nominchimeg Sukhbaatar 1 &
• Tsogbadrakh Mishig-Ochir 1  

Scientific Reports volume  11 , Article number:  21047 ( 2021 ) Cite this article

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• Nanoscale materials

In recent years’ synthesis of metal nanoparticle using plants has been extensively studied and recognized as a non-toxic and efficient method applicable in biomedical field. The aim of this study is to investigate the role of different parts of medical plant Carduus crispus on synthesizing silver nanoparticles and characterize the produced nanoparticle. Our study showed that silver nanoparticles (AgNP) synthesized via whole plant extract exhibited a blue shift in absorption spectra with increased optical density, which correlates to a high yield and small size. Also, the results of zeta potential, X-ray diffraction, photon cross-correlation spectroscopy analysis showed the surface charge of − 54.29 ± 4.96 mV (AgNP-S), − 42.64 ± 3.762 mV (AgNP-F), − 46.02 ± 4.17 mV (AgNP-W), the crystallite size of 36 nm (AgNP-S), 13 nm (AgNP-F), 14 nm (AgNP-W) with face-centered cubic structure and average grain sizes of 145.1 nm, 22.5 nm and 99.6 nm. Another important characteristic, such as elemental composition and constituent capping agent has been determined by energy-dispersive X-ray spectroscopy and Fourier transform infrared. The silver nanoparticles were composed of ~ 80% Ag, ~ 15% K, and ~ 7.5% Ca (or ~ 2.8% P) elements. Moreover, the results of the FTIR measurement suggested that the distinct functional groups present in both AgNP-S and AgNP-F were found in AgNP-W. The atomic force microscopy analysis revealed that AgNP-S, AgNP-F and AgNP-W had sizes of 131 nm, 33 nm and 70 nm respectively. In addition, the biosynthesized silver nanoparticles were evaluated for their cytotoxicity and antibacterial activity. At 17 µg/ml concentration, AgNP-S, AgNP-F and AgNP-W showed very low toxicity on HepG2 cell line but also high antibacterial activity. The silver nanoparticles showed antibacterial activity on both gram-negative bacterium Escherichia coli (5.5 ± 0.2 mm to 6.5 ± 0.3 mm) and gram-positive bacterium Micrococcus luteus (7 ± 0.4 mm to 7.7 ± 0.5 mm). Our study is meaningful as a first observation indicating the possibility of using special plant organs to control the characteristics of nanoparticles.

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Introduction.

Nanotechnology is a science that deals with the manipulation and fabrication of nanoparticles 1 . At least one or two dimensions of nanoparticles are within the range of 100 nm or less 2 . The nanometer-scaled particles have a unique property that differs them from their counterpart bulk material 3 , their small size offers a large surface-to-volume ratio which causes a substantial biochemical and catalytic activity compared to the particles with the same composition 1 , 4 . Nanoparticles are employed in the areas of drug delivery, biomedical sciences, gene delivery, chemical industries, optics, mechanics, catalysis and etc. 5 . Among metal nanoparticles, silver nanoparticles (AgNP) garner much attention due to their strong antibacterial and anti-inflammation effect. AgNPs are utilized in various physical, biological and pharmaceutical fields, for instance, cream or ointment containing AgNPs are applied for burn and wounds to inhibit bacterial infection 6 . Although AgNP is integrated into many areas, the exact mechanism explaining the particle formation is not fully uncovered yet. The traditional method for the synthesis of AgNP is to use physical and chemical approach to produce nanoparticles with controlled and well-defined size and shapes 7 . However, the use of toxic substances, high pressure and energy such as laser ablation, hydrothermal synthesis, solvothermal synthesis, pyrolysis and inert gas condensation brought a demand for more biologically compatiblenanoparticles 8 , 9 . Recently, the synthesis of AgNP through biological method has been studied intensely. The biological method offers nanoparticles with high yield and stability compared to the conventional physical and chemical approach 10 . AgNP can be biosynthesized by bacteria, fungi, yeast, actinomycetes and plant, thus avoiding the use of toxic substances and enabling for further application in medical and pharmaceutical field 11 . The application of plants for the synthesis of AgNP has gained significant attention. Plant-mediated synthesis of AgNP has many advantages, it can be obtained under ambient temperature with low cost and the process is relatively fast compared to bacteria, where a long process of maintaining cell culture is required 12 . Plants contain a wide range of metabolites that can aid in reducing silver ion, stabilizing and capping AgNP 13 , therefore the concentration and composition of AgNP will vary depending on the plant type 3 . This is especially the case for the medicinal plant as it is a rich source of complex phytochemicals and antioxidants. The main antioxidants of medicinal plants are polyphenols, carotenoids, and vitamins. The medicinal plants display a wide range of anti-inflammatory, antibacterial, antiviral, anti-aging, and anti-cancer activities 14 . In addition to polyphenols found in plants, there are other biomolecules responsible for reducing and capping AgNP 15 , these include polysaccharides, aldehydes, ketones, proteins, enzymes, amino acids, and caffeine 16 . The complex biomolecules found in medicinal plant assist in the reduction of metal ions and stabilization of nanoparticles into desired shape and size 17 . The plant-mediated synthesis of AgNP is relatively simple as it requires only plant extract and silver salt, thereafter it undergoes a reduction process 18 . There are many reports published regarding a medicinal plant-mediated synthesis of AgNP, these include Gmelina aroberea 19 , Tecomella undulata 20 , Artemisia absinthium 21 , Datura stramonium 22 , Calliandra haematocephala 23 , Carica papaya 24 etc. Carduus crispus is a plant species of the family Asteraceae that can be found in Mongolia. The medicinal effect ranges from a stomachache, rheumatism, atherosclerosis to cancer. And due to its medical properties, it is broadly applied in Mongolian traditional medicine 25 . The main activity of Carduus crispus is coagulation, antioxidant and anticonvulsive activity 26 . According to the study done by Baumberger the major compounds detected in Carduus crispus are flavonoids and coumarins, also alkaloids saccharides, essential oil, rubber, lipids contained in small quantities 27 . There are no available reports on the synthesis of AgNP using Carduus crispus as the plant extract. The mechanism of action of AgNP is not yet completely understood, however there are several hypotheses available explaining the antibacterial, anti-inflammatory and anti-cancer activity. It is known that nanoparticles have a large surface area that either penetrates the cell or attaches itself to the cell wall 11 , causing a disturbance in the membrane permeability making it porous 28 , and this action leads to a further leakage of cell content. Moreover, the appearance of pores on membrane result to diffusion of nanoparticles into the cell where it binds with sulfur and phosphorus-containing proteins, thus leading to the inactivation of proteins and DNA 17 . Another hypothesis suggests that the antibacterial activity of AgNPs results from the release of Ag + ions through the oxidation dissolution process. Silver ions oxidized from AgNP mainly interact with thiol groups of various enzymes and protein, thereby interfering with the respiratory chain and disrupting the bacterial cell wall. Silver ions also facilitate the generation of reactive oxygen species (ROS), which is considered as the main cause for most cell death through the inactivation of DNA replication and ATP production 29 . The present study aimed to synthesize silver nanoparticles with medicinal plant Carduus crispus extracts and characterize the final product, and evaluate their antibacterial activities.

Results and discussion

Uv–vis spectra analysis and color change.

The visual color change from pale yellow to dark brown in response to time can be seen as evidence of silver ion reduction to AgNP. The change in color of biosynthesized AgNP is due to the excitation of surface plasmon resonance (SPR). Several studies done on the synthesis of AgNP via medicinal plant suggest the absorption peak around 412–470 nm with the duration of synthesis from 4 h till 24 h, these include medicinal plants, such as Abutilon indicum, Aegle marmelos, Azadirachta indica, Calliandra haematocephala, Calotropis procera, Carica papaya, Helicteres isora, Lawsonia inermi, Leptadenia reticulate, Rheum palmatum, Tecomella undulata, Tagetes erecta, Urtica dioica . The rate of color change from light yellow to dark brown varied in these studies, the earliest color change began within 1 h till 4 h 4 , 20 , 23 , 24 , 30 , 31 , 32 , 33 , 34 , 35 , 36 , 37 , 38 . Alternatively, different studies utilizing non-medicinal plants for the AgNP synthesis, such as Allium cepa, Chenopodiastrum murale, Cyperus rotundus, Eleusin indica, Euphorbia hirta, Melastoma malabathricum, Musa acuminate, Pachyrhizus erosus, Rubus glaucus exhibited absorption peak from 401–780 nm and was synthesized for 72 h till 14 days. The color change of AgNP synthesized via C. murale turned to brown color after incubating overnight 39 , 40 , 41 , 42 , 43 . The difference in color change rate might be due to the different properties of the plant, specifically, the medicinal plant contains a wide range of phytochemicals, such as flavonoids, polyphenols, terpenoids, etc. 44 that assist in the formation of silver nanoparticles. Iravani et al. 5 reported in their studies that flavonoids, polyphenols, terpenoids, alkaloids and proteins are the main constituents responsible for the reduction and stabilization of silver nanoparticles. Figure  1 shows the result of color change of the synthesized silver nanoparticle with different organs of Carduus crispus , such as stem, flower and the whole plant. It can be seen that different plant organs affected differently on silver nanoparticle synthesis, and particularly whole plant extract facilitated better silver nanoparticle formation compared to the stem and flower extract. The synthesis of silver nanoparticles with whole plant extract exhibited a darker color change. The variation in color change might be due to the different phytochemical content in the plant organs. Following the visual color change study, the formation and stability of silver nanoparticles synthesized with flower, stem, and whole plant of Carduus crispus were characterized using a UV–Vis spectrophotometer (Fig.  2 ). The results revealed that silver nanoparticles synthesized with whole plant (AgNP-W) exhibited higher absorption compared to silver nanoparticles synthesized using plant organs such as flower (AgNP-F) and stem (AgNP-S). The higher absorption is directly proportional to the higher yield of silver nanoparticles in colloidal solution 45 . Additionally, the size of the synthesized silver nanoparticle was studied by observing the shift of the absorption peak towards a longer or shorter wavelength 8 , 46 . In Fig.  2 b-d, silver nanoparticles were measured at various times, and according to our results, the AgNP-W exhibited blueshift in contrast to AgNP-F and AgNP-S, which can be interpreted as the formation of smaller-sized silver nanoparticles.

Color changes in biosynthesized silver nanoparticle with different parts of Carduus crispus . S-stem, F-flower and W-whole plant.

UV–Vis spectra for the reaction mixture containing of silver nanoparticles synthesized from Carduus crispus flower (AgNP-F), stem (AgNP-S) and whole plant (AgNP-W). Shown are the UV–Vis absorption spectra from 370 to 700 nm of all plant organs and synthesized ( A ) AgNPs, ( B ) AgNP-W, ( C ) AgNP-S, and ( D ) AgNP-F.

Zeta potential analysis

Zeta potential explains the stability, dispersion and surface charge of the nanoparticles. The zeta potential greater than + 30 mV or less than − 30 mV indicates high stability of nanoparticles in dry powder form 31 . The high negative value produces repulsion between similarly charged particles in suspension, therefore resisting aggregation 47 . Several studies were done on silver nanoparticle synthesis with a medicinal plant such as Pot entilla fulgens, Alpinia calcarata, Pestalotiopsis micospora, Urtica dioica, Jatropha curcas which resulted inzeta potential of − 18 mV, − 19.4 mV, − 35.7 mV, − 24.1 mV, and − 23.4 mV respectively 4 , 6 , 12 , 47 , 48 . Our results showed that zeta potential of the synthesized AgNP-W, AgNP-S, AgNP-F had an average zeta potential of − 46.0 2 ± 4.17 (AgNP-W), − 54.29 ± 4.96 (AgNP-S) and − 42.64 ± 3.762 (AgNP-F) (Table 1 ). The zeta potential of AgNP-S exhibited a higher average value compared to the AgNP-W and AgNP-F, this may be due to the presence of different phytochemicals in each sample that reduces and cap silver nanoparticles. The results of the zeta potential analysis suggest that silver nanoparticles synthesized with Carduus crispus exhibit high stability and resist agglomeration. Figure  3 showed that zeta potential values of AgNP-W, AgNP-S, and AgNP-F fall within the normal distribution curve, which indicates that synthesized silver nanoparticles are fairly monodisperse.

Zeta potential analysis of ( A ) AgNP-W, ( B ) AgNP-F and ( C ) AgNP-S.

FTIR spectral analysis of synthesized AgNP by Carduus crispus

The presence of the functional groups capping AgNP synthesized using Carduus crispus is analyzed by FTIR and shown in Fig.  4 . The presence of various organic compounds in the plant reveals multiple peaks compared to the chemical method where only a few and strong peaks are displayed 49 , 50 . The results of our FTIR analysis showed the presence of several functional groups in AgNP-W, AgNP-S, AgNP-F. Additionally, the functional groups in AgNP-F and AgNP-S were present in AgNP-W samples as well, this may be attributed to the various phytochemicals capping the silver nanoparticles that are found both in flower and stem of Carduus crispus . The strong characteristic bands at ~ 3418 cm −1 to 3429 cm −1 and 2361 cm −1 in all samples AgNP-S, AgNP-F, AgNP-W are assigned to the O–H stretching/N–H stretching of amides and 2361 cm −1 to the C≡C stretching. Additionally, the weak band at ~ 1017 cm −1  to 1022 cm −1 and ~ 828 cm −1 assigned to carbohydrates and –C = O bending were found in all samples AgNP-S, AgNP-F, and AgNP-W. C–O stretching is present in AgNP-F which was observed from the very strong band at 1353 cm −1 . The weak bands at 2922 cm −1 and 2857 cm −1 of CH 3 stretch of alkane/carboxylic acids present in AgNP-F and were absent in AgNP-S. The band detected at ~ 3418 cm −1 to 3429 cm −1 and 1618.35 cm −1 correspond to the presence of phenolic compounds and flavonoids, and the band found on 1021.35 cm −1 indicates carboxylic acid, ester, and ether groups of proteins and metabolites that may be involved in the synthesis of nanoparticles 33 . Our result show that the strong band detected at 1611 cm −1 and 1017 cm −1 from AgNP-F correspond to the presence of flavonoids and proteins. On the other hand, weak bands detected at ~ 1696 cm −1 to 1371 cm −1 correspond to alcohol, carboxylic acids, alkyl halides/carboxylic acids/ester, alkenes/alkyl halides/aromatics, alkynes/alkyl halides stretch that peaks found from AgNP-S. According to Baumberger 27 the major compounds detected in Carduus crispus are flavonoids and coumarins, in addition, alkaloids, saccharides, essential oil, rubber and lipids contained in small quantities which is in line with the presence of flavonoids and phenolic compounds in our synthesized AgNP. The AgNP-F and AgNP-S contained different functional groups that correspond to various compounds, and AgNP-F revealed that it has a strong correlation with flavonoids from Carduus crispus . The results of FTIR and UV–Vis spectra analysis confirm that these functional groups are the capping and reducing agents responsible for the synthesis of AgNPs.

Fourier transform infrared spectra of ( a ) AgNP-W, ( b ) AgNP-S, and (c) AgNP-F.

XRD, PCCS, SEM/EDX and AFM analysis

The crystalline nature of the synthesized AgNP was confirmed by X-ray crystallography. The XRD pattern of the nanoparticles was analyzed with an XRD instrument and shown in Fig.  5 . Bragg reflection of the 2θ peaks was observed at 32.25˚ to 81.62˚ and corresponded to (111), (200), (220), (311), (222) plane lattice which can be indexed to the face-centered cubic crystal nature of the silver. The average crystallite size was calculated using the Scherrer equation. The average crystallite sizes were 13 nm (AgNP-F), 14 nm (AgNP-W) and 36 nm (AgNP-S). The results of our study are in line with other published literature, the crystal nature of silver nanoparticles synthesized with Tagetes erecta 31 , Urtica dioica 4 , Aegle marmelos was face-centered cubic with diffraction peaks of (111), (200), (220), (311) respectively 34 . PCCS is a technique based on the Brownian motion that measures the average nanoparticle size (grain size). In Fig.  6 , the average particle size of AgNP-W, AgNP-F and AgNP-S was 99.6 nm, 22.5 nm and 145.1 nm respectively. The difference between PCCS and XRD analysis lies in the measurement method of the particle. Application of the Scherrer equation on XRD data gives the average crystallite size, specifically the size of a single crystal inside the particle or grain. The morphological and elemental analysis was done on Scanning Electron Microscope (SEM) and Energy Dispersive X-Ray Spectroscopy (EDX). The elemental composition of the synthesized silver nanoparticle was assessed using EDX spectroscopy (Table 2 ). The results in Fig.  7 showed that AgNP-W, AgNP-S, and AgNP-F contained silver and potassium elements together with several other elements that differed in AgNP-F and AgNP-S samples, i.e. AgNP-F included phosphorus 2.8%, potassium 15.2%, and AgNP-S had calcium 7.5%, pottassium 15.5% elements. In contrast, AgNP-W contained all the elements including the elements that differed in AgNP-F and AgNP-S. Interestingly, the silver element in AgNP-F had the highest content of 82% compared to AgNP-W and AgNP-S which had a silver content of 79% and 77% respectively. Another observation on EDX analysis revealed that AgNP-W, AgNP-F, AgNP-S did not show the presence of nitrogen peak, this indicates that trace ions from AgNO 3 are absent in the samples. The size of biosynthesized AgNP-W, AgNP-F and AgNP-S was determined with Atomic Force Microscopy (AFM). Figure  8 show that the size of nanoparticles differed, for instance, AgNP-W had a size of 70 nm, AgNP-F with size 33 nm and AgNP-S with size 131 nm. Figure  8 (A-C, E–G and I-K) represents the two dimensional images of AgNP-W, AgNP-F and AgNP-S. Figure  8 (D, H and L) shows the three dimensional image of AgNP-W, AgNP-F and AgNP-S respectively. The different composition of plant organs, such as stem, flower and whole plant could be the reason for the observed variability in, color change, UV–Vis absorption, EDX, FTIR. In addition, the results of AFM data and XRD show that the synthesis of AgNP can be manipulated with different plant organs.

XRD spectra of ( a ) AgNP-W, ( b ) AgNP-F, ( c ) AgNP-S. Peaks are appeared at 111, 200, 220, 311 and 222.

PCCS analysis: particle number distribution of synthesized AgNP-W ( A ), AgNP-F ( B ) and AgNP-S ( C ).

EDX spectra for ( A ) AgNP-F, ( B ) AgNP-S and ( C ) AgNP-W along with SEM image area (inset).

Atomic force microscopy images (2D and 3D) of silver nanoparticles on siliconized cover slide; AgNP-W ( A – D ), AgNP-F ( E – H ) and AgNP-S ( I – L ).

Antibacterial activity

The antibacterial activity of silver nanoparticles was studied against pathogenic bacterial strains of gram-negative E.coli and gram-positive M.luteus using the well diffusion method (Fig.  9 ). Standard antibiotics such as Penicillin G and Chloramphenicol, plant extracts, AgNO 3 and distilled water were chosen as the control group. The results of the antibacterial activity showed that all synthesized silver nanoparticles had efficient antibacterial activity against both gram-negative E.coli and gram-positive M.luteus bacterial strains. The inhibition zone of AgNP-F, AgNP-W and AgNP-S against E.coli and M.luteus were 6.5 ± 0.3, 6 ± 0.2, 5.5 ± 0.2 and 7.5 ± 0.3, 7 ± 0.2, 7.7 ± 0.4 mm respectively. The plant extract and AgNO 3 did not reveal any antibacterial activity against both E.coli and M.luteus , which can be interpreted that AgNP-W, AgNP-F, and AgNP-S are solely responsible for the antibacterial activity. The mode of action of AgNPs against bacteria is not completely understood yet. However, several hypotheses are explaining the antibacterial activity of silver nanoparticle: (1) generation of reactive oxygen species; (2) release of Ag + ions from AgNPs denaturize proteins by bonding with sulfhydryl groups; (3) attachment of AgNPs on bacteria and subsequent damage to bacteria 4 , 11 , 24 . The multiple published reports on the antibacterial activity of silver nanoparticles against gram-negative and gram-positive bacteria showed that silver nanoparticles had a slight antibacterial activity on gram-positive bacteria 6 , 22 , 31 , 36 . Interestingly, AgNP synthesized by Carduus crispus exhibited effective inhibition on both gram-positive and gram-negative bacteria which can be interpreted that the antibacterial activity of silver nanoparticles (AgNP-W, AgNP-F and AgNP-S) is not affected by the difference in the bacterial wall.

Petri dishes showing the zone of inhibition of synthesized AgNP-W on ( A ) M. luteus and ( B ) E. coli , and AgNP-F on ( C ) M. luteus and ( D ) E. coli , AgNP-S on ( E ) M. luteus and ( F ) E. coli (AgNP: silver nanoparticle, AgNO 3 : silver nitrate, DW: distilled water, PE: plant extract).

In-vitro cytotoxicity assay

Cytotoxicity is considered as an important indicator for cell viability, therefore in this study we employed crystal violet assay to investigate the effect of different concentration of AgNP-W, AgNP-F and AgNP-S on the adherent human hepatoma cell line HepG2 (Fig.  10 ). The liver is an important organ with detoxifying effect, additionally, it is considered as an accumulation site for AgNPs 51 . In this study, the untreated HepG2 cell lines revealed significant adherence to the well plate. On the other hand, the treated cells with nanoparticles exhibited small decrease in cell viability after 24 h incubation at 3 to 17 µg/ml. The cell viability of these treated groups with AgNP-W, AgNP-F and AgNP-S were 87.93 ± 4.87%, 92.24 ± 1.21% and 86.20 ± 2.43% at 17 µg/ml. The toxicity of AgNPs to bacteria and human cells is widely known, however, the result of our study suggests that AgNPs synthesized by medicinal plant Carduus crispus with concentration of 3 to 17 µg/ml have low toxicity on HepG2 cell line (Fig.  10 A,B). In addition, biosynthesized silver nanoparticles possessed efficient antibacterial activity against Gram-negative and Gram-positive bacteria (Fig.  9 ). The antibacterial activity of the synthesized AgNPs and their low toxicity to human cells may enable further application in biomedical field. The low toxicity of biosynthesized AgNPs to adherent human cells are similar to other published reports 52 .

A microscopic pictures of HepG2 cells treated with AgNPs for 24 h in cell culture: control ( A ), AgNP-W ( B ), AgNP-F ( C ) and AgNP-S ( D ). After 24 h, the cell toxicity effect was examined with Crystal Violet ( E ).

The synthesis of silver nanoparticles via biological method, specifically plant extracts provides a natural, eco-friendly, cost-effective, rapid synthesis of silver nanoparticles. The present study reports the synthesis of silver nanoparticles with medicinal plant Carduus crispus in reducing silver ions and stabilizing the silver nanoparticles. It has been reported that medicinal plants are a rich source of phenolic compounds such as flavonoids and phenolic acids, etc. Additionally, plant organs contain different contents of phenolic compounds, therefore flower, stem, and whole plant of Carduus crispus were chosen for this study. Afterwards, the synthesized silver nanoparticles were characterized using visual color change, UV–Vis spectroscopy, zeta potential, FTIR, XRD, PCCS, SEM–EDX and AFM. The characterization of AgNP-W, AgNP-F, and AgNP-S revealed that AgNP-W had a higher yield, synthesis rate, and smaller-sized silver nanoparticles. The zeta potential conveys the stability and the result of all the synthesized silver nanoparticles showed the zeta potential value of − 46.0 2 ± 4.17 (AgNP-W), − 54.29 ± 4.96 (AgNP-S), and − 42.64 ± 3.762 (AgNP-F) which indicates highly stable silver nanoparticles. The variation in zeta potential may be due to the different phytochemical properties of the plant. Then FTIR analysis was utilized to study the role of phytochemical properties in plants for the synthesis of silver nanoparticles, the results showed that different functional groups in AgNP-F and AgNP-S were also present in AgNP-W samples as well. And based on the UV–Vis spectra analysis, AgNP-W and AgNP-F had the highest absorbance compared to AgNP-S, therefore we can conclude that the functional groups present and coincided in both AgNP-F and AgNP-W may play a contributing role in capping and synthesis of silver nanoparticles, these include functional groups with bands at 2922.28 cm −1 , 2857.66 cm −1 , 1711.90 cm −1 , 1611.59 cm −1 , 1079.22 cm −1 and 1017.49 cm −1 which correspond to alkanes, carboxylic acids, ketones, alkenes, amides, esters/ethers/amides, alkyl halides. Furthermore, strong bands at 3418 cm −1 to 3429 cm −1 , 1618.35 cm −1 to 1611 cm −1 , and 1017 cm −1 correlates to flavonoids and phenolic compounds. The EDX analysis detected the following elements, such as silver, potassium, phosphorus in AgNP-F; silver, potassium, calcium, chloride, and phosphorus in AgNP-W; finally, silver, potassium, calcium in AgNP-S samples. The synthesized silver nanoparticles had an average crystallite sizes of 14 nm (AgNP-W), 13 nm (AgNP-F) and 36 nm (AgNP-S) with face-centered crystal structure and average grain sizes of 99.6 nm (AgNP-W), 22.5 nm (AgNP-F) and 145.1 nm (AgNP-S). The sizes detected in AFM was 70 nm (AgNP-W), 33 nm (AgNP-F) and 131 nm (AgNP-S). Although the method of synthesis varied in AgNP-F, AgNP-W, and AgNP-S, their antibacterial activity showed efficient inhibition on both gram-negative and gram-positive bacteria. Based on these results, we can conclude that silver nanoparticles synthesized by whole plant of Carduus crispus have a faster rate of synthesis, higher yield with a smaller size, and high antibacterial activity against both gram-negative and gram-positive bacteria. The overall results show that the effectiveness of the synthesis of the flower for AgNP appears similar to using whole plant. Additionally, we have shown that the process of synthesizing nanoparticles can be manipulated with specific organs of plant, for example, particle size and synthesis duration, biological effect, etc. Our study is meaningful as a first observation indicating the possibility of using special plant organs to control the characteristics of nanoparticles. Moreover, further studies are required in this area.

Chemicals and plant

The Carduus crispus was collected from Khuder soums, Selenge province of Mongolia (GPS coordinates: N 49.641772, E 107.80935) and the taxonomy was determined by a botanist Kh.Khaliunaa from National University of Mongolia. The Carduus crispus used for the study does not violate the local regulations of Mongolia, the permission for the plant collection was granted from the Ministry of Environment and Tourism of Mongolia. The collected plant specimen of Carduus crispus was deposited into the publicly available herbarium of National University of Mongolia with deposition number UBU0002509. The Silver Nitrate (AgNO 3 ) with ≥ 99.0% purity was purchased from Sigma Aldrich. All the other relevant reagents are up to the standard.

Preparation of plant extract

The whole plant was washed with tap water in order to remove the adhering dust and soil particles, followed by washing with distilled water. 100 ml of distilled water was added to 5 g of Carduus crispus and boiled for 15 min, then cooled at ambient temperature. Afterward, it was filtered by Whatman filter paper and centrifuged twice at 10,000 rpm to obtain a plant extract. Finally, the extract was ready for the synthesis of AgNP.

Synthesis of silver nanoparticle

The aqueous plant extract of Carduus crispus and AgNO 3 (1 mM) were mixed with the ratio of 1:16, then the solution was exposed to the daylight and the reaction took place at the various time at room temperature. In order to obtain silver nanoparticles in powdered form, the solution was vaporized on a vacuum evaporator, and the final product of AgNP was kept inside the oven at a temperature of 300 °C for 4 h.

Characterization of AgNP synthesized by Carduus crispus

AgNP was successfully synthesized by using Carduus crispus . A color change from pale yellow to colloidal dark brown indicated the formation of silver nanoparticles. UV–Vis spectra analysis offers an insight into the synthesis and stability of the AgNP. Formation of the biosynthesized AgNP was determined by the UV–Vis spectrophotometer (Shimadzu UV-2500PC Series) at 30 min, 1 h, 2 h, 3 h, 4 h, 6 h, 12 h, 24 h and was carried out at 350–700 nm range. FTIR spectrum was recorded in the range of 500 to 4000 cm −1 through the potassium bromide powder method using FTIR spectrophotometer (Prestege-21, Shimadzu, Japan) for understanding the constituent capping and reducing agents of silver nanoparticles. Also, elemental composition of the synthesized silver nanoparticles was analyzed with an energy dispersive X-ray spectroscope instrument (TM-10000 with EDX). To identify the structural phase present in the AgNP, XRD was performed by XRD instrument (Shimadzu, Maxima-X-7000) operating at 40 kV with a current of 30 mA and Co-Ka radiation. And crystalline size was determined by Scherrer equation. In order to understand the size distribution and surface charge, the zeta potential (ZetaCompact, CAD Instruments, France) and Photon Cross-correlation Spectroscopy (PCCS) (NANOPHOX 1 nm to 10,000 nm, Sympatec GmbH, Germany) methods was used for dispersed nanoparticles of silver.

Atomic force microscope (AFM) measurement

The size of the synthesized AgNP was analyzed with Atomic Force Spectroscopy (SPA 300, Seiko Inc., Japan). First, the siliconized glass cover slides selected for the AFM measurement were immersed in ultra-pure water and sonicated for 10 min with ultra sonicator, afterwards the siliconized glass cover slides were rinsed with ethanol solution and air-dried in laminar box at RT, then the samples for AFM analysis were prepared by drying the AgNP suspension on prepared siliconized glass cover slide at RT. Finally, Atomic Force Microscope was used to analyze the morphology and size of the samples via golden silicon probe (GSG11) with tip curvature radius of 10 nm.

Determination of anti-bacterial activity using well diffusion method

The agar well diffusion method was used to study the antibacterial activity of the synthesized silver nanoparticle. Broth medium was used to subculture bacteria and was incubated at 37 °C for 24 h, afterwards, overnight cultures were taken and spread on the agar plates to cultivate a uniform microbial growth plate. The bacterial strains were gram-negative Escherichia coli and gram-positive Micrococcus luteus . And silver nitrate, plant extract, antibiotics (Penicillin G against Micrococcus luteus and Chloramphenicol against Escherichia coli ) were chosen as the control group for the study of antibacterial activity. Finally, the petri dishes were incubated for 24 h at 37 °C. In order to evaluate the antibacterial activity of the synthesized silver nanoparticle, the diameter of the inhibition zone was measured and compared with the control groups.

Cell culture

The cell line HepG2 was cultured in Dulbecco’s Modified Eagle Medium (DMEM) medium supplemented with 10% Fetal Bovine Serum (FBS), 1% penicillin, 1% streptomycin and maintained in standard condition with 5% CO 2 humidified incubator at 37 °C temperature. Prior treatment, the cells were seeded in 96-well tissue culture plates with a seeding density of 2 × 10 4 cells and incubated overnight. The cell line HepG2 were sub-cultured at 70–80% confluence and used for further study.

Crystal violet assay

The crystal violet assay was performed to determine the cell viability according to the method described by Feoktistova et al. Crystal violet is a dye which binds to DNA and protein of cells and used for studying viable cells that are adhered to the cell culture plates. In order to study the cytotoxicity of the synthesized AgNP-W, AgNP-F and AgNP-S, the nanoparticles were first filtered with 0.45 µm filter. Different concentration of AgNP-W, AgNP-F and AgNP-S suspension (3–17 μg/ml) were added to triplicate well and incubated for 24 h. After treatment the medium was removed and the cells were washed twice with PBS, followed by staining with 50 µl of 0.5% crystal violet dye for 20 min at room temperature. Thereafter, crystal violet dye was removed and the wells were washed in a stream of tap water and left to air-dry for 3 h at room temperature. The crystal violet dye was dissolved with the addition of methanol to each well and the absorbance was measured at 570 nm with ELISA reader.

Statistical analysis

All experiments were performed at least three times, independently and data were analyzed by a Student’s t-test, and a value of p < 0.05 was considered significant.

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Acknowledgements

We gratefully acknowledge the assistance from Associate Professor Lkhagvasuren.D Department of Biology, National University of Mongolia and Associate Professor Tegshjargal.Kh Department of Chemical and Biological Engineering. This work was supported by the Science Technology Foundation grant funded by the Mongolian Government.

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E.U., B.B. and T.M. designed the experiments. E.U., B.B., A.G. and N.S. conducted the experiments. E.U. and B.B. analyzed the data. E.U. and B.B. wrote the manuscript.

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Urnukhsaikhan, E., Bold, BE., Gunbileg, A. et al. Antibacterial activity and characteristics of silver nanoparticles biosynthesized from Carduus crispus . Sci Rep 11 , 21047 (2021). https://doi.org/10.1038/s41598-021-00520-2

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DOI : https://doi.org/10.1038/s41598-021-00520-2

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Biosynthesis of silver nanoparticles using nitrate reductase from aspergillus terreus n4 and their potential use as a non-alcoholic disinfectant., proksee: in-depth characterization and visualization of bacterial genomes, regulation of the formation and structure of biofilms by quorum sensing signal molecules packaged in outer membrane vesicles., bionanofactories for green synthesis of silver nanoparticles: toward antimicrobial applications, tygs and lpsn: a database tandem for fast and reliable genome-based classification and nomenclature of prokaryotes, evaluating green silver nanoparticles as prospective biopesticides: an environmental standpoint., silver nanoparticles biosynthesis, characterization, antimicrobial activities, applications, cytotoxicity and safety issues: an updated review, metal-pseudomonas veronii 2e interactions as strategies for innovative process developments in environmental biotechnology, review on green nano-biosynthesis of silver nanoparticles and their biological activities: with an emphasis on medicinal plants, online self-powered cr(vi) monitoring with autochthonous pseudomonas and a bio-inspired redox polymer, related papers.

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Article Versions Notes

Deonas, A.N.; Souza, L.M.d.S.; Andrade, G.J.S.; Germiniani-Cardozo, J.; Dahmer, D.; de Oliveira, A.G.; Nakazato, G.; Torezan, J.M.D.; Kobayashi, R.K.T. Green Synthesis of Silver Nanoparticle from Anadenanthera colubrina Extract and Its Antimicrobial Action against ESKAPEE Group Bacteria. Antibiotics 2024 , 13 , 777. https://doi.org/10.3390/antibiotics13080777

Deonas AN, Souza LMdS, Andrade GJS, Germiniani-Cardozo J, Dahmer D, de Oliveira AG, Nakazato G, Torezan JMD, Kobayashi RKT. Green Synthesis of Silver Nanoparticle from Anadenanthera colubrina Extract and Its Antimicrobial Action against ESKAPEE Group Bacteria. Antibiotics . 2024; 13(8):777. https://doi.org/10.3390/antibiotics13080777

Deonas, Anastácia Nikolaos, Lucas Marcelino dos Santos Souza, Gabriel Jonathan Sousa Andrade, Jennifer Germiniani-Cardozo, Débora Dahmer, Admilton Gonçalves de Oliveira, Gerson Nakazato, José Marcelo Domingues Torezan, and Renata Katsuko Takayama Kobayashi. 2024. "Green Synthesis of Silver Nanoparticle from Anadenanthera colubrina Extract and Its Antimicrobial Action against ESKAPEE Group Bacteria" Antibiotics 13, no. 8: 777. https://doi.org/10.3390/antibiotics13080777

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