IHJPAS. 36 (3) 2023 177 This work is licensed under a Creative Commons Attribution 4.0 International License *Corresponding Author: khloosa123aa@gmail.com Abstract In the current century, nanotechnology has gained great interest due to its ability to modify the size of metals to the nanoscale, which dramatically changes the physical, chemical, and biological characteristics of metals relative to their bulk counterparts. The approaches used to create nanoparticles (NPs) are physical, و chemical and وbiological. The shortcomings in physical and chemical synthesis approaches, such as the generation of toxic by-products, and energy consume as they require high temperature, pressure, power and lethal chemicals, contributed to an increased interest in biological synthesis by plants. Scientists have created a new filed called as "green nanotechnology" by fusing the idea of sustainability with nanotechnology. By substituting plant-based materials, it aims to reduce the amount of chemicals used in the manufacture of nanoparticles. Silver nanoparticles (AgNPs) attract the most attention due to their great stability and low chemical reactivity in comparison to other metals. The present review describes the fabrication of nanoparticles (NPs) via chemical and physical methods, as well as the use of plants, bacteria, and fungi. The current review also discusses certain analytical methods used to examine AgNPs, including UV-Vis spectroscopy, FT-IR, SEM, TEM, AFM, XRD, DLS, and zeta potential analysis. Keywords:Nanoparticles, silver nanoparticles, approach methods, green synthesis, characterization. doi.org/10.30526/36.3.3050 Article history: Received 30 September 2022, Accepted 15 December 2022, Published in July 2023. Ibn Al-Haitham Journal for Pure and Applied Sciences Journal homepage: jih.uobaghdad.edu.iq Synthesis and Characterization of Silver Nanoparticles: A Review *Ekhlas A. Abdul Kareem Department of Chemistry, Collage of Science, University of Diyala, Iraq. khloosa123aa@gmail.com Alaa E. Sultan Department of Chemistry, Collage of Science, University of Diyala, Iraq. alaesa1988@gmail.com Hadeel M. Oraibi Department of Biology, Collage of Science, University of Diyala, Iraq. hadeelmothher@gmail.com https://creativecommons.org/licenses/by/4.0/ mailto:khloosa123aa@gmail.com mailto:khloosa123aa@gmail.com mailto:khloosa123aa@gmail.com mailto:alaesa1988@gmail.com mailto:alaesa1988@gmail.com mailto:hadeelmothher@gmail.com mailto:hadeelmothher@gmail.com IHJPAS. 36 (3) 2023 178 This work is licensed under a Creative Commons Attribution 4.0 International License 1. Introduction Nanotechnology is an advanced technique that deals with nanometer-sized samples, which are referred to as nanoparticles (NPs). Nanomaterials are tiny, solid particles with sizes ranging between 1 and 100 nanometers[1]. Nanoparticles may be organic or inorganic, depending on the ingredients that were used in their synthesis. The first type (organic) is based on carbon, while the second type (inorganic) is a noble metallic or magnetic type [2]. Due to the many advantages of nanoparticles compared with bigger particles having the same chemical composition, they have gained great interest in overcoming the limitations of conventional therapies because of their improved and distinct physical, chemical, and optical properties, especially silver nanoparticles. There are many nanoparticles of great importance in the field of scientific research, including gold nanoparticles, copper nanoparticles, iron nanoparticles, zinc nanoparticles, and titanium dioxide nanoparticles [3]. Among these materials, AgNPs have drawn a lot of attention due to their exceptional biological activity. It has been demonstrated to have a potential candidate for biological activities that include anti-bacterial, anti-cancer, larvicidal, treatment of wounds, water purification, food preservation, wound healing, and cosmetics [4]. Moreover, AgNPs are playing an important role in a variety of applications, such as anti-oxidants, anti-microbial agents, nanomedicine, ointments, chemical sensing, the food industry, and information storage [5]. Despite the inconsistencies about the toxicity of silver nanoparticles (AgNPs), their efficacy as an antiseptic and antibacterial agent has received much praise. We decided to work on plant-mediated green synthesis because of the available documented data and community interest in this field [6]. The present review article is a comprehensive investigation of the environmentally friendly synthesis and characterization methods used for the synthesis of AgNPs by different biological sources using bacteria, fungi, and plants, and it provides an important database for researchers that may be useful in their future work. This review examines the published papers from 2012 to 2022. 2. Approaches for silver nanoparticles (AgNPs) synthesis Silver nanoparticles can be prepared by three methods: physical, chemical, and biological, which result in various forms and sizes for usage in a variety of applications. These synthesis methods are classified into two categories: top-down and bottom-up approaches. Various top- down and bottom-up approaches for the manufacture of AgNPs are shown schematically in Figure 1 [10]. https://creativecommons.org/licenses/by/4.0/ IHJPAS. 36 (3) 2023 179 Figure 1. Systematically synthetic approaches of nanoparticles[2]. 2.1 Top- Down Approach (physical Approach) In this method of synthesis, silver nanoparticles are created using a top-down approach. This approach starts by breaking down large silver particles into smaller units, and these units are then transformed into useful nanoparticles. The most popular physical approaches for the manufacture of nanoparticles are chemical itching, sputtering, mechanical milling, and laser ablation[3, 4]. The evaporation-condensation method is carried out using a tube furnace at atmospheric pressure. The sample is evaporated into a carrier gas within the boat positioned at the furnace. However, using a physical approach (tube furnace) to produce AgNPs has many drawbacks, including large area requirements, slow synthesis, raising the temperature of the environment around the source material, and low yield[5]. This method needs a long time to accomplish thermal stability. Additionally, a typical tube furnace needs a lot of energy and requires high concentration [5-7]. In the laser ablation method, AgNPs have been created using metallic bulk materials in solution. Their creation depends on a number of factors, including the wavelength of the laser, the duration of the laser pulses, the amount of the ablation time, and the liquid medium's efficiency. The nanoparticles that form in a solution with a high concentration of surfactants, with or without the presence of surfactants, are smaller than those that form in a solution with a low concentration of surfactants [8]. However, a benefit of using a laser to create colloids over other methods is that there are no chemical reagents present in the solution. In light of this, pure colloids will be beneficial for more applications.[9]. IHJPAS. 36 (3) 2023 180 2.2 Bottom-up Approach This approach is a constructive that involves arranging small units-to -form nanoparticles (NPs). In this approach nanoparticles (NPs) can be manufactured using green or chemical process[10]. 2.2.1. Chemical Approach This approach include various chemical methods such as chemical reduction, chemical 1vapor deposition (CVD), sol–gel process, hydrothermal, solvothermal, photo-chemical, chemical, laser pyrolysis, aerosol pyrolysis, plasma or flame spraying, templates, spinning[11]. The synthetic approach involves fabrication of metallic nanoparticles in solution by use three main components of the synthetic method: metal precursors, reducing agents and stabilizing agents or capping agents. In this approach sodium citrate, sodium borohydride (NaBH4), ascorbate, - N,N,dimethylformamide (DMF), ascorbic- acid , hydrazine (NH2-NH2), ammonium format (NH4COOH), ethylene glycol and glucose, are the most frequently used reducing agents [12]. These reducing -agents are accountable for the reduce metal ions (like, silver ions (Ag+)) to the metal (Ag0) followed by the nucleation stage and finally lead to the formation of metal nanoparticles (MNPs). Stabilizing agents like borohydride are added during preparation, as they help the growth of nanoparticles in addition to protecting nanoparticles from forming agglomeration[5]. The seed-mediated growth method, includes the adding of seeds into the growth medium to produce size-controlled metal nanoparticles, in this approach yields nanoparticles of various morphologies. Depending on the capping agent utilized[13]. The essential advantage of chemical method is producing particles that are easily distributed in organic media, which is acknowledged by scientists across a wide range of disciplines, but one of the disadvantages of this method is that it requires a high cost and high energy in addition to hazardous chemicals, so it was necessary to search for an environmentally friendly and economical technique for the production of nanoparticles. The green approach became inevitable and vital rather than other approaches, and it deserved extensive research [14]. 2.2.2. Green Approach The conventional methods (physical and chemical) used to produce nanoparticles (NPs) are costly, hazardous, and not environmentally friendly. To overcome these problems, researchers turned to biological methods in the production of nanoparticles because of their simplicity, environmental, and high productivity features, in addition to the fact that they do not require dangerous chemicals, high temperatures, or pressures [23–25]. Biological methods include biological microorganisms such as bacteria, algae, yeast, fungi, or plant extracts, which are precursors to the synthesis of metallic nanoparticles (MNPs) [26]. Green synthesis is a suitable approach when silver nanoparticles are synthesized using plant extracts and microbes. They contain reducing agents that reduce 1Ag+ to 1Ag0 and also contain capping or stabilizing agents that prevent the aggregation of the nanoparticles [27]. Especially with the green biological synthesis method that has attracted the interest of researchers recently due to its being environmentally friendly, in a short period of time a large amount of nanoparticles (NPs) can be manufactured, which is time-saving, cost-effective, less toxic, and produces different sizes of nanoparticles in comparison with other biological methods [23], which makes it preferred over IHJPAS. 36 (3) 2023 181 others. With the advancement of technology towards green chemistry, a lot of work has been done to synthesize a number of metallic nanoparticles like silver, gold, copper, zinc, platinum, and lead [28]. Interestingly, these nanoparticles have been exploited in a variety of environmental applications [29]. Figure 2 shows different green approaches to the synthesis of AgNPs. In general, the green synthesis of nanoparticles can be summarized as follows- :  Biosynthesis; use of Microorganisms like Yeast, bacteria, fungi, and algae.  Phyto synthesis; using plants (leaves, stems, latex, flowers, seeds, roots, fruits, and peel) and their extract  Utilization of templates such as membranes, DNA, diatoms, and viruses [15]. The green synthesis via plants, bacteria, and fungi is described in the further sections. Figure 2. Different green approaches to synthesis of AgNPs . 3. Biosynthesis of silver nanoparticles (AgNPs) 3.1 Using Plant Extract Plant extracts are a significant branch of biosynthesis processes, as plants have the ability to production metal nanoparticles in different ways inside and outside cells by reducing, metal ions. Silver ion is converted into silver nanoparticles by biologically active molecules found in plants. Proteins, terpenoids, polysaccharides, phenols, alkaloids, flavonoids, amino acids, and enzymes of alcoholic substances are some examples of these biological molecules. The major variables that can affect the creation of the nanoparticles (NPs) are temperature , the concentration of the extract, metal salt, contact duration and acidic function (pH) [11]. The importance of using-plants in the synthesis of nanoparticles (NPs) because all parts of plants, including seeds, stems, roots, latex , and leaves, contain a variety of active ingredients that can be used to reduce silver ions (Ag+)[16]. When comparing green synthesis for nanoparticle, synthesis using plant extracts was considered several times faster than synthesis Templates Diatoms, membranes, DNA and viruses, Green Approaches Microorganisms Yeast, fungi, bacteria, and algae Plants Leaves, stem, latex, flowers, seeds, roots, fruits, and peel IHJPAS. 36 (3) 2023 182 using microorganisms (such as bacteria, fungi, algae and yeast). The latter is not feasible and requires more sterile circumstances, a laborious procedure, and a longer incubation period [17]. Therefore the use of plant extracts in green synthesis has attracted attention due to its quick development and ability to produce nanoparticles (NPs) in a single step at a low cost while being non-pathogenic and environmentally friendly protocol [17]. Figure 3, shows a schematic-diagram for the synthesis of silver nanoparticles using plants. Figure 3. Schematic diagram for green synthesis of AgNPs by using plan/plant extracts[18]. The protocol used to prepare the plant extract and nanoparticles synthesize can be summarized as follows  If the plant extract is made from the leaves or peels of some plants, a portion of the leaves or peels of the plant of interest is collected from the available sites and thoroughly rinsed two or three times in tap water to remove dust and soil particles, followed by distilled water to remove any accompanying debris. Clean, fresh leaves that are dried in the shade for (5-7) days and then crushed using a domestic blender. Finally, to prepare the plant broth, about 0.5–10 g of the dried powder is boiled with a suitable volume of distilled water [19]. The resulting extracts were then filtered using filter paper, and each filtrate was collected in a separate volumetric flask (250 ml) and kept at (4°C) for later use[20]. This extract was used for generating silver nanoparticles (AgNPs). This bio-extract is always used fresh[21].  If the plant extract is made from fresh fruit, the fruit is thoroughly washed with tap water and then distilled water before being cut and squeezed through a fine nylon mesh to obtain the extract. Then, the obtained extract was centrifuged at 10,000 rpm for 10 minutes to remove any unwanted impurities. This extract was collected in a dark volumetric flask (100 ml) and stored at 4 °C for further experiments[22].  To prepare the plant extract from the other plant parts, the required plant part must thoroughly washed, then mixed in distilled water, boiled for a short time, and filtered. Finally, the filtrate can be used immediately or stored at a low temperature for later use [23]. The solvent used also has an effect on the extraction rate. The phytochemical content of the IHJPAS. 36 (3) 2023 183 phenolic and alcoholic extracts increases [24]. Several studies summarized in Table 1 have been carried out on the synthesis of AgNPs using different plant extracts. Figure 4. Greens synthesis of metal nanoparticles by plant parts [25]. Table 1. Use of various plant extract in the synthesis of AgNPs Name of plants Part of plant used Shape Size (nm) References Angelicae- pubescenis Extract of leaf Spherical 12.48 [26] Andrographis-echiodes Extract of leaf Cubic 68-91 [27] Amomumvilosum Extract of fruit Spherical 5-15 [28] Artemsia vulgaris Extract of leaf Round 25 [29] Alium sativum Extract of fruit Spherical 3-6 [30] Acacia- seyal Gum Round 81.45 [31] Acalypha- hispida Extract of leaf Spherical 20–50 [32] A. millefolium Aqueous extract Spherical /rectangular, and cubical 14-20.77 [33] Barleria-buxifolia Extract of leaf Spherical 80 [34] Butea -monosperm Extract of leaf Spherical, and triangular 20-80 [35] Camellia- sinensis (green tea) ------------- Spherical 11 [36] Coriandrum -sativum Extract of leaf Spherical 37 [37] Cornus officinali Extract of fruit Quasi Spherical 11.7 [38] Curcuma- aromatica Irregular 10–30 [39] Erythrina- indica Extract of leaf Spherical 20-118 [40] Eucalyptus - chapmaniana Extract of leaf Spherical Different sizes [41] Datura- stramonium Extract of leaf Spherical, and triangular 18 [42] IHJPAS. 36 (3) 2023 184 3.2 Using Bacteria Bacteria is a type of microorganism, which are regarded as one of the best choices for the manufacture of AgNPs, the use of bacteria as environmentally sustainable precursors for the manufacture of nanoparticles such as gold and silver has been very successful. Due to their amazing capacity to reduce heavy metal ions and their relative simplicity in handling them.[18]. Nanoparticles (NPs) are manufactured, by bacteria either intracellular or extracellular, green synthesis is made flexible, reasonable, and appropriate method by the choice of bacteria[43]. Figure 5 shows the schematic diagram for the synthesis of AgNPs by using different bacteria. The following are a few instances of bacterial strains that have been widely used to synthesize bio-reduced AgNPs with distinctive shape/size morphologies include: Bacillus indicus, Antarctica, Pseudomonas, Escherichia-coli, Bacillus Amylolique- faciens, Arthrobacter 5gangotriensis, Bacillus-cereus, Aeromonas sp. SH10 Phaeocystis proteolytica, Bacillus cecembensis, Lactobacillus casei, Enterobacter cloacae, Geobacter spp., and Corynebacterium sp. SH09 [44]. These species of bacterial are commonly utilized for commercial biotechnological applications including bioremediation,, bioleaching and genetic engineering [45]. According to some recent scientific studies , some types of bacteria including Pseudomonas-aeruginosa and Pseudomonas -stutzeri possess a high skill in using defense mechanisms to combat stresses such as heavy metal ion toxicity and can even thrive and survive in the presence of high concentrations of heavy metal ions[46]. Also, several recent scientific reports confirmed that the ability of microbes to synthesize metallic nanoparticles mainly depends on several factors, including culture conditions, and the improvement of reaction parameters such as 33acidic function, temperature, and 2nutrient concentrations affects the production of microbial enzyme-activity[47]. The numerous studies have reported the production of AgNPs using bacteria. The first of these studies was the study conducted by Klaus et al. in which AgNPs were synthesized with specific structures and shapes well using Ag resistant bacterial strains Pseudomonas stutzeri (AG259). These cells accumulate AgNPs in large quantities up to (200) nm [48]. Table 2 lists many other bacteria that can be employed in the synthesis of AgNPs. However, the main disadvantage of using bacteria as nanofactories is the slow rate of synthesis, the possibility of culture contamination, lengthy procedures, difficulty in controlling the nanoparticle size and reduction rate, and restricted morphologies compared to other biological sources such as fungi and algal species that contain many proteins that play an important role in the biosynthesis of metallic nanoparticles (NPs) of various shapes [9]. Table 2 shows h green synthesis of AgNPs using different bacteria. IHJPAS. 36 (3) 2023 185 Figure 5. Diagram depicting the use of bacteria in the synthesis of AgNPs [18]. Table 2. Green synthesis of AgNPs using different bacteria. Bacteria Species Intracellular/extracellular Shape Size (nm) References Esciheichia coli Extracellular Spherical 1.2–62 [49] Bacilus- cereus Extracellular Spherical 20–40 [50] Lactobcillus- casei Extracellular Spherical 20–50 [51] Rhodococcus spp. , Intracellular Spherical 5–50 [52] Endosymbiotic Bacterium Extracellular Spherical, cubic, hexagonal, crystalline, and oval 10 –60 [53] Aeromonas spTHG- FG1.2 Extracellular Spherical, and FaceCentered Cubic 8 –16 [54] Bacillus strainCS 11 Extracellular Spherical and FCC 45 ± 0.15 [55] Novosphingobium sp.HGC3 Extracellular Spherical) and crystalline, 8– 25 [56] Bacillus- methylotrophicus Extracellular Spherical 10–30 [57] Marine- Ochrobactrum sp , Intracellular Spherical 38–85 [58] Actinobacteria , Intracellular Spherical 5–50 [59] KinneretiaTHGSQ14 Extracellular Spherical, Mono-disperse, FCC. 15–20 [60] Nocardiopsis- spp. Extracellular Spherical 50 ± 0.15 [61] 3.3 Using Fungi Fungi are good biological agents, it acts as a “Nano factory” for the biosynthesis of metal oxide /metal nanoparticles especially AgNPs and due to their high capacity for metal IHJPAS. 36 (3) 2023 186 bioaccumulation, their tolerance, high binding capacity with the metal ions in the intracellular region, and intracellular uptake[62]. Fungi are better than bacteria as biological agents as the specialized fungi can produce well-defined structured nanoparticles with good monodispersed compared to bacteria due to the presence of a variety of enzymes / proteins / reducing components on their surfaces and intracellular[63]. Which directly affects the higher productivity of nanoparticles [64]. The main advantage of nanoparticles manufactured from fungi is that they contain a large amount of pure enzyme and are free of cellular protein. In Fig.6. A schematic representation showing the synthesis of AgNPs using fungi. The expected mechanism for the synthesis of nanoparticles in fungi is due to the occurrence of electrostatic interaction between the negatively charged carboxylate groups in the enzymes and the positively charged Ag ions. This extracellular extraction simplifies biomass recovery in downstream procedures. This method was more beneficial than bacterial synthesis. It was observed that rapid reduction and extracellular formation of metallic nanoparticles occur within 10 minutes. Many studies which can be used for the synthesis of AgNPs from fungi are shown in Table 3. Figure 6. Schematic diagram for synthesis of AgNPs by using fungi[18]. Table 3. Green synthesis of silver nanoparticles (AgNPs) using different fungi. Fungi Species Intracellular/extracellular Shape Size (nm) References Penicillium italicum Extracellular Face-centered cubic lattice 39.5 nm. [65] Botryodiplodia theobromae Extracellular Spherical 62.77 –103 [66] Schizophyllum commune Extracellular Spherical 51–93 [67] Beauveria bassiana Extracellular Spherical, triangular, hexagonal 10–50 nm, [68] IHJPAS. 36 (3) 2023 187 Beauveria bassiana Extracellular Spherical 40.14–289.13 nm [69] Aspergillus niger Extracellular Poly dispersed spherical 1–20 [70] Trichoderma harzianum Extracellular Spherical Different size [71] Guignardia mangiferae Extracellular Spherical 5–30 [72] Arthroderma fulvum Extracellular Spherical 21 [73] Candida glabrata Extracellular Spherical 2–15 [74] Tritirachium oryzae W5H Extracellular Monodispersed and sphericalto ovular 7-75 [75] 4. Characterization m of silver nanoparticle (AgNPs). The first qualitative indicator for the synthesis of AgNPs is the color-change of the solution from yellow to brown [76].There are many techniques employed for characterization of nanoparticles, based on their size, shape, morphology, surface area, optical activity, thermal stability and dispersity. These techniques include UV-Visible spectrophotometry, Fourier Transform Infrared Spectroscopy (FT-IR), Scanning Electron Microscopy (SEM), Transmission Electron Microscopy (TEM), Atomic Force Microscopy (AFM), X – ray diffraction (XRD), Dynamic Light Scattering (DLS), and Zeta-potential measurements [15]. Figure 7 shows techniques for the characterization of AgNPs. Figure 7. Characterization techniques of AgNPs[77]. Characterization FTIR AFM TEM UV-Visible Spectroscopy DLS Zeta- potential XRD SEM IHJPAS. 36 (3) 2023 188 4.1 UV–Visible, Spectrophotometry UV-visible spectrophotometry is one of the simplest and most reliable techniques that is also efficient and selective for many nanoparticles. The basis of spectroscopy is that light in the UV and visible spectrums is absorbed or scattered by metal nanoparticles. which results in a strong absorption band known as surface plasmon resonance (SPR) in the 400–500 nanometer, due to the interaction between light and mobile surface electrons of AgNPs. The concentration, form, and size of the metal ions investigated by UV-visible spectroscopy influence the degree of excitation[78]. Table 4 showing green synthesis of AgNPs using plant extracts and a characterization method. Table 4 . Showing green synthesis of silver nanoparticles using plant extracts and a characterization method. 1Plants name Plant Part used Shape Size- (nm) Characterization techniques used. References Alliumsativum Extract of fruit Spherical 3–6 Uv- Visible spectroscopy, FT-IR, XRD/EDX, and TEM. [30] Acacia -seyal Gum Round 81.45 Uv- Visible spectroscopy, FT-IR, AEM, and XRD. [31] Acalypha- hispida Extract of leaf Spherical 20–50 Uv- Visible spectroscopy, FT-IR, TEM, XRD, and GC- mass. [32] A. millefolium Aqueous extract Spherical, rectangular and cubical 14-20.77 Uv – visible spectroscopy, FT-IR, SEM, and XRD. [33] Barleria -buxifolia Extract of leaf Spherical 80 Uv- Visible spectroscopy, FT-IR, TEM, EDS, SEM, and XRD. [34] (Buteamonosperma Extract of leaf Spherical and triangular 20–80 Uv- visible spectroscopy, FT-IR, TEM, DLS, TEXRD, and Fluorescence- Microscopy. [35] Alhagi -graecorum Extract of fruit Spherical 22- 36 Uv- visible spectroscopy, -FT-IR, and TEM. [79] Cornusofficinalis Extract of fruit Spherical 11.7 Uv- Visible spectroscopy, FT-IR, DLS, XRD/ EDAX, and FE-SEM. [38] Erythrinaindica - Extract of root Spherical 20-118 Uv- Visible spectroscopy, FTIR, TEM, XRD/ EDX, and DLS. [40] Eucalyptus - chapmaniana Extract of leaf Spherical Different sizes Uv-Visible spectroscopy, FT-IR, and XRD. [41] Datura- stramonium Extract of leaf -/ Spherical, and triangular 18 Uv- Visible spectroscopy, FT-IR, TEM, and XRD. [42] IHJPAS. 36 (3) 2023 189 Nauclea -latifolia Extract of fruit Shape of irregular 12 Uv- visible spectroscopy, FT-IR, EDX, andSEM [80] Oryza- sativa L. (rice) Extract of fruit Different shape 346.4±36.8 Uv- Visible spectroscopy, FT-IR, and DLS. [81] P. subpeltata Extract of leaf Spherical 22.6 Uv-visible spectroscopy, FT-IR, FE-SEM, and XRD [82] Punica granatum L. (pomegranate Extract of peels, leaves and seeds Shape of spherical, and regular 50 Uv- Visible spectroscopy, FT-IR, SEM, and XRD. [83] Indigofera tinctoria Extract of leaf Spherical 9–26 Uv-visible spectroscopy, FT-IR, TEM, XRD/ EDX and AFM. [84] Seaweed (S. swartzii) Extract of leaf Spherical 20–40 Uv-visible spectroscopy, FTIR spectroscopy, XRD, TEM, FE-SEM, and AFM. [85] Sambucus ebulus Extract of Aerial Spherical, and cubic 35 - 50 Uv- Visible spectroscopy, FT-IR, TEM, XRD, EDX, and HPLC. [86] Scindapsus officinalis Extract of Fresh fruits Spherical Different size Uv- Visible - spectroscopy, FT-IR, TEM, FE-SEM/EDX and XRD. [87] Hylocereus undatus Extract of peel Spherical 10- 50 Uv-visible- spectroscopy, FT-IR, SEM, XRD, and EDX. [88] Humulus lupulus Extract of crushed Spherical 17.40 Uv- Visible spectroscopy, FT-IR, TEM, XRD,DLS, BET, XPS, Raman Spectroscopy , SEM/EDAX, and AFM analysis. [89] Ixora brachypoda Extract of leaf Spherical 18 - 50 Uv- visible spectroscopy, FT-IR, TEM, XRD/FE-SEM, and EDS. [90] 4.2 Fourier transform infrared spectroscopy (FTIR) Analysis FT-IR technique is used to investigate and identify the functional groups (such as ketones, ammines, and aldehydes) of both plant extracts and AgNPs. The spectrum depends on the principle that particles absorb electromagnetic energy in the infrared region of the spectrum and reasons the subatomic particles to vibrate. The wave number ranges from (4000 - 400) cm- 1. The form of nanoparticles (NPs) based on the peak position, while the size of the nanoparticle (NPs) based on the intensity of the peaks measured in the (FT-IR) spectrum[91]. Table 5 summarizes the FTIR data of the reviewed articles. https://www.sciencedirect.com/topics/earth-and-planetary-sciences/raman-spectroscopy https://www.sciencedirect.com/topics/earth-and-planetary-sciences/raman-spectroscopy https://www.sciencedirect.com/topics/earth-and-planetary-sciences/electron-microscopes IHJPAS. 36 (3) 2023 190 Table 5. FTIR data from chosen plant extracts. Plant name FTIR absorption bands (cm−1) Possible functional group References Ixora brachypoda 3403 Stretching of COOH in group of carboxylic acids [90] 2923 Stretching - vibrations of C–H bond in alkane (CH4) 2853 Stretching-vibrations of C–H bond in aldehydes (CHO) 1603 Bending of N–H bond in primary amines 1403 Bending of C–H bond in alkanes 1384 bending of N=O bond in nitro compound (NO2) 1318 Bending of S=O bond in sulfates 1262 Bending of esters (COOR) 1117 Bending of C-F bond in compound. 859, 620, 788, and 467 Stretching of C-X bond in aromatic compounds like (C-F, , C-Cl, C-Br, C-I). Seaweed (S. swartzii) 3748 and 3523 Stretching of O–H bond. [85] 2357 and 1737 Stretching of – C≡C– bond 1540 Stretching of -C–N- 1356 and 668 The bond related to C–H 1645 The bond related to (C = O) A. millefolium 1524, 1035 and 558 C-C bond in aromatic compounds [33] 3357 Stretching of – O–H bond 2917 and 2846 Stretching of C-H bond 3348.19 Stretching of O-H bond [86] 1623.95 Stretching of –C = O 1401.65 –C-H bend of alkanes or –C-C- the stretch of aromatics 1273.12 Bending of C-O bond in 8anhydrides group Acer oblongifolium 2865.16 The bond related to (C-H) in alkanes [92] 2031.92 The bond related to – C–H 1478.96 the N-C and N=C groups 845.35 The bond related to -C-O Alhagi graecorum 3294 The bond related to –N-H [79] 2178 Stretching of C-H 1631 for amide I 1488 The bond related to C-H-N 590 and 540 hydroxyl (-OH) group Barleria buxifolia 3410.15   Stretching vibration of -OH bond [34] 2926.06 stretching of -CH(Aliphatic) 1710 Stretching vibration of Carbonyl in the acid 1593.2 Stretching of C=O bond in amide, 2926 Stretch of C–H bond in (alkanes); 1348 Stretch of C=C bond in (aromatic ring) 1382 -C–H (aromatics) IHJPAS. 36 (3) 2023 191 P. subpeltata 1050 Bending Strong of S=O bond [82] 1347 Stretch Strong C–F 1605 Bending of N–H Primary and secondary 2836 Stretch C–H amines and amides 3078 Stretch C–H Aldehyde 7Fagonia- cretica 3864 The bond related to O-H 3729 The bond related to O-H 3626 Stretching of O-H 3467 The bond related to N-H 2916 Stretch of C-H in alkanes 2358 Stretching of N-H 1636 Stretching of C=O 1472 Bending vibration of N-H 1401 C-O-H 1114 C-O-C 1061 Stretching of C-O 869 Bending of C-C, C-OH, C-H (ring) 627 and 643 The bond related to C-H / bending Citrus tangerina, Citrus sinensis, and Citrus limon 2950 and 3670 The bond related to O-H/ stretching-frequency [93] 1636 Stretching of C=O, and aromatic C=C Stretching. 2115 alkyne group present in phytochemicals 597 Silver nanoparticles bonding with oxygen from OH group Tagetes -erecta 3415 Stretching-vibration of O-H in phenol and alcohol compounds. [94] 2922 The -C-H stretching mode in alkanes 1060, 1399, and 1829 Stretching of C-O, and Stretching of C=O in alcohol, ethers,and esters 1643 and 1564 Bending vibration of N-H in amides 1490 Stretching of C=C 670 Bending of O-H 609 Bending of C-H 4.3 Scanning Electron Microscopy (SEM) Analysis SEM is a common technique; it is used to determine the morphology and surface topology of nanoparticles. The principle behind scanning electron microscopy (SEM) is that when an electron beam incident on the surface of the sample, an interaction occurs between electrons and atoms inside the sample that leads to the emission of secondary electrons. The emission of these electrons depends on the surface geometry and composition of the sample [95]. This set of reflected electrons is captured by the detectors in the SEM and translated into images. SEM is paired with EDX. The advantage of using SEM is that it can resolve particles smaller IHJPAS. 36 (3) 2023 192 than 10 nm, but the disadvantage is that it cannot determine the interior structure of the particle [95]. 4.4 Transmission Electron Microscopy (TEM) Analysis Transmission Electron Microscopy (TEM) is one of the most common analysis and described as high-resolution techniques used to study the, morphology of NPs. This analysis provides information about, particle shape, size, and distribution. The basic principle of this technique is that an electron beam is transmitted through the surface of the metal nanoparticles, and the interaction of the transmitted electrons produces in an image [47, 96]. Electron microscopy (EM) can be used to obtain a better resolution on the sample. The (SEM) and (TEM) techniques are used to study topography and surface morphology for nanoparticles. The key distinction between (TEM) and (SEM) is that the former offers the good resolution and data of the internal structure than the SEM. 4.5 Atomic Force Microscopy (AFM) Analysis Atomic Force Microscopy (AFM) is a microscopy technique used to describe topography, 0morphology, the surface texture, roughness, and particle -size distribution of nano- particles[97]. A disadvantage of the atomic microscopy technique is that the lateral dimensions of the sample are overestimated, so it is necessary remove the error which requires more attention [98]. 4.6 X- ray Diffraction (XRD) Analysis XRD is the essential technique used to identify the crystalline nature of nanoparticles (NPs), polymers, various biomolecules, and superconductors. The basic idea behind the method is that when a monochromatic beam of X-ray is focused on a crystal, it produces a variety of diffraction patterns that can be analyzed using the Bragg’s equation: 2d sinө = nλ, where, d: Is the spacing between the diffracting planes, ө: Is the, incident angle, n is any integer, and λ: Is the, wavelength of the beam used to know the, characteristics of crystalline or poly- crystalline material[47]. It is also used to determine the size of the crystal using the Scherrer equation: D = 0.94 𝜆 /𝛽 cos 𝜃. 𝐷 represent size of particles nm, (𝜆) is the wavelength X-ray KCu (K𝛼 = 15406 ˚), (𝛽 ) is the full width XRD peak, and (𝜃) is t angle . The disadvantage of this XRD technique is that sometimes there is difficulty growing the crystals. This is the only disadvantage of the XRD technique [76]. 4.7 Dynamic Light Scattering (DLS) The most common simplistic technique used to determine the average size of nanoparticles in aqueous solution [99]. This method's basic idea is that it gauges the particle's hydrodynamic radius while it is in Brownian motion. The foundation of this technology is the interaction of light and particles. When exposed to laser light, the particle in the solution scatters the light in various intensities. Accordingly, the Stokes-Einstein equation was used to determine the corresponding particle sizes. This technique can be used to determine the size of nanoparticles ranging from 20 to 200 nm. When comparing the sizes of the particles IHJPAS. 36 (3) 2023 193 obtained by using DLS, TEM, and SEM techniques, it was noted that the size obtained from DLS was larger than the size obtained from TEM and SEM [95]. . 4.8 Zeta potential Analysis Zeta potential analysis is one of an important, techniques in interpreting the properties of nanoparticles, especially silver nanoparticles. This technique is used to determine the stability and aggregation of nanoparticles in a state of dispersion [100]. Also, zeta potential analysis allows detecting the surface charges of nanoparticles by giving information about those charges. The principle of this technique is the electrostatic attraction between the charges on the surface of nanoparticles with the oppositely charged ions present in the solution. 5. Conclusions The present review describes methods for the synthesis of nanoparticles, especially silver nanoparticles, exemplified by the top-down and bottom-up approaches. This article also explained the shortcomings in chemical and physical methods, which increased the interest of researchers in biological synthesis by bacteria, fungi, and plants. The green synthesis of AgNPs mediated by many plant materials is more advantageous than alternative biological approaches because plant extracts are easy to handle, widely available, safe, and readily available. The present review, combining several recently published works, shows the significance of plant extracts facilitated bio-synthesis of AgNPs, and these studies are described as being cost-effective, environmentally friendly, and highly suitable for producing nanoparticles free of toxic contaminants needed in bio applications due to their unique properties. Silver nanoparticles will play a significant role in many nanotechnology-based processes, the present review also includes several analytical techniques that are utilized for the examination of these AgNPs, including UV-Vis spectroscopy, FT-IR, SEM, TEM, AFM, XRD, DLS, and Zeta potential analysis Authors' declaration The researchers declare that there are no conflicts of interest regarding the current manuscript. IHJPAS. 36 (3) 2023 194 References 1.Yazdanian, M.; Rostamzadeh, P.; Rahbar, M.; Alam, M.; Abbasi, K. Tahmasebi, E., Tebyaniyan, H., Ranjbar, R., Seifalian, A.; Yazdanian, A. 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