Antibacterial activity of green gold and silver nanoparticles using ginger root extract

Antibacterial activity of green gold and silver nanoparticles using ginger root extract | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Antibacterial activity of green gold and silver nanoparticles using ginger root extract Morteza Yadi, Abolfazl Akbarzadeh, Hassan Dianat-Moghadam, Mehdi Azizi, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-1352913/v2 This work is licensed under a CC BY 4.0 License Status: Posted Version 2 posted You are reading this latest preprint version Show more versions Abstract Background Recent studies demonstrated that the speed of synthesis, biocompatibility, and antimicrobial activity of gold (Au) and silver (Ag) metals is enhanced when biosynthesized in nano-sized particles. In the present study, Au NPs and Ag NPs were synthesized via a biological process using aqueous Ginger root extract and characterized by various spectroscopic methods. Methods & Results The NPs were found to be in hexagonal and spherical shapes. The average particle size for Au and Ag NPs was found to be 20 nm and 15 nm, respectively. The dynamic light scattering (DLS) method has shown that the zeta potential values of synthesized NPs were found to be 5.7 mv and 7.11mv, respectively. Gas chromatography-mass spectrometry (GC-MS) analysis of Ginger root extract revealed 25 compounds. The synthesized NPs showed significant activity against Staphylococcus aureus and Escherichia coli in vitro with IC50 and IC90 values for Au and Ag NPs, respectively, noted to be 7.5 and 7.3 µg/ml and 15 and 15.2 µg/ml for both bacterial strains. The protein leakage level was high and morphological changes occurred in bacteria treated with biosynthesized NPs. Conclusion These results suggest that the biosynthesized metallic NPs show potential for application as antibacterial agents with enhanced activities. green synthesis antibacterial agent ginger extract metallic nanoparticle Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 1. Background Biofilms are microbial populations enclosed in a matrix. Those grow in three steps, (i) initial adhesion, (ii) proliferation, and (iii) detachment. Bacterial cells bind together by extracellular polymeric substances and are connected to a substrate surface, involved in each cell-cell interaction and cell surface as a part of the developmental process. Biofilms show high resistance towards toxicants if compared to planktonic cells [1, 2]. They can cause of lead to infections in humans and animals and serious problems in the environment. Infectious diseases are one of the health threats on human society. The use of antibacterial drug for the control and treatment of infectious disease are common worldwide. However, continued use from the antibacterial drug leads to drug resistance [3]. In this condition use of the drug for control and treat of infectious diseases don't be effective. Eliminating this problem requires a new therapeutic agent for control and treatment of the infectious disease [4]. Nowadays, silver (Ag) and gold (Au) nanoparticles (NPs) are used in a wide range of medicine. There are several methods including laser ablation, gamma irradiation and use of the chemical agent as a reducing and capping agent, for the synthesis of the Au and Ag NPs [3]. The important problems with these methods are expensive, and use of toxic chemical agents that are not safe for human health and the environment [5]. The green synthesis of the NPs, also known as photosynthesis, is one of the emerging fields in nanotechnology. Green synthesis NPs have shown high activity against the primary biofilm. Plants are used for the synthesis of NPs and have advantages over physical and chemical processes [6]. In recent years, NP synthesis using plant extracts has been increasing because, these are available, environmentally friendly, and easy to use, and have a wide range of secondary metabolites that act as a reducing agent [7]. Ginger (Zingiber officinale), Roscoe belonging to the family Zingiber aceae, is a perennial herb with thick tuberous rhizomes. Ginger extracts have antibacterial activity. Malu and co-workers show that material such n-hexane, ethyl acetate and soxhlet which are in the ginger extract solution, have antibacterial effects. In the fact this material in addition to having bactericidal activity, inhibition of bacterial growth [8]. E. coli (gram-negative bacteria) and S. aureus ( gram-positive bacteria) have an important role in human infectious diseases. E. coli through penetration to lymphocytes and an inflammatory reaction with the host, by causing bloody diarrhea [9]. S. aureus is the main cause of food poisoning and surgical wounds infection which together with epidermidis syndrome, causes infections associated with medical equipment [10]. In this research, gold and silver nanoparticles were synthesized with green and chemical synthesis methods. For this purpose, Ginger root extract and citrate were used as reducing agents, respectively. The synthesized metal NPs were characterized by dynamic light scattering (DLS), transmission electron microscope (TEM), ultraviolet-visible spectroscopy UV-Vis, Atomic absorption spectroscopy (AAS), and Fourier transforms infrared spectroscopy (FTIR). The antibacterial activity of green and chemical synthesized Au and Ag NPs were investigated with E. coli and S. aureus strains in vitro. 2. Materials And Methods 2.1. Microorganisms Standard strains of S. aureus (ATTC 25923) and E. coli (ATTC 25922) were purchased from the Iranian Research Organization for Science and Technology (IROST). These strains were cultivated in the nutrient broth medium and were incubated at 37°C for 24h. For further experiments, small amount of bacterial colonies was stored in Tripticas soy broth containing glycerol at -70°C. 2.2. Preparation of ginger extracts Roots of ginger were purchased from the local market, the Islamic Republic of Iran, and washed frequently with ultra-pure deionized water. After shredding the ginger root, dried and crashed into powder by the steel hammer. After that, 2 gr of powder was mixed with 80 ml of ethanol and incubated at 40°C for 24 hours. Then for obtaining ginger extract, the solution was filtered with Whatman No.1 filter paper. 2.3. Green and chemical synthesis of gold and silver nanoparticles For the green synthesis of Au NPs, 1 mL ginger root extract was added to a 50 mL boiling solution of HAuCl 4 ·3H 2 O (1mM) and the boiling continued for 5 min. Then, the solution was kept undisturbed at room temperature until the colorless solution converted to a wine red color which indicated the formation of Au NPs. For the chemical synthesis of Au NPs, HAuCl 4 solution is boiled and then trisodium citrate dehydrate was added slowly into the boiling solution under stirring. A few minutes later, the color of the solution from light yellow converted to wine red. For green synthesis, like above, 1 ml of ginger extract was added to with 100 ml of silver nitrate (2mM) boiling solution. The reaction mixture was kept undisturbed at room temperature until the colorless solution converted to reddish brown color which indicated the formation of Ag NPs. For the chemical synthesis of silver nanoparticles, silver nitrate solution and citrate of sodium were used as a metal ion source and reduction agent, respectively. Also, citrate of sodium was used as a reducing agent. When the citrate sodium was added to the silver nitrate the color of the solution converted from pale yellow to pale brown color. And the final silver nanoparticle was purified by centrifugation. 2.4. Gas chromatography-mass spectrometry analysis (GC-MS) Ginger extract was analyzed by a mass scientific trace 2200 (gas chromatography) system with a thermalSaturnmass selective detector (Varian Company). The machine was equipped with a TG-5MS (mass spectroscopy) column (30 * 0.25 mm (5% phenyl) -methylpolysiloxane capillary column, film thickness * 0.25 micrometer), 220 centigrade temperature injector and 250 centigrade temperature transfer line. The temperature of the oven was programmed as follows: initial temperature; 50 centigrade for 5 min and then increase 4°C/min up to 250 centigrade. The gas carrier used was He at a flow rate of 1.0 ml/min. 1microliter sample was injected and the ionization energy was 70 eV. The base of the identification of individual components was based on their retention time and by comparison of their mass spectral pattern with standard library data. (National Institute of standards and technology). 2.5. Characterization of gold and silver nanoparticles The reduction of gold and silver ions was monitored by a UV-Vis spectrophotometer (SPECTOR 250, Analytic Jena) in the 350–800 nm wavelength range. Due to the evaluation of concentration of green and chemical synthesized NPs, a solution of synthesized NP was diluted, and then the amount of Au and Ag NPs were measured by atomic absorption spectroscopy (AAS) (NovaAA400, Analytic Jena Co). Also for determination of shape and size of the nanoparticle were used from Transmission electron microscopy (TEM) (Zeiss Leo q06) operating at 200 kV accelerating voltage. For the preparation of the sample; 10 microliter of aliquots of NPs solution was drop-casting onto a carbon-coated copper grid and then was placed on a piece of paper to get rid of excess solvent. For determination of the average particle size, distribution, and stability of the gold and silver NPs were used from ELSZ-1000 zeta-potential and particle sizer (Mastersizer 2000, Malvern, USA). For FTIR analysis, the powdered gold and silver NPs were recorded by FTIR spectrometer (Tensor 27, Bruker Co) over the 4000 − 400 cm − 1 frequency with 4 cm − 1 resolutions by using a KBr pellet method. 2.6. Antibacterial assay The antibacterial activity of chemical and green synthesizedAu and Ag NPs were performed by well diffusion agar method. Standard strains of bacteria were subculture on plates containing Muller Hinton agar using the pour plate method. Then the wells which have 6mm diameter were punctured onto the agar plates and 25µg/ ml of Au and Ag NPs solution and aqueous plant extract were loaded into the wells. After 24 hours of incubation, the inhibition of zone diameter around of wells was measured. For comparison of the effectiveness of Au and Ag NPs and ginger extract against tested bacteria, we used Streptomycin (30 µg/ml). For evaluation of the minimum inhibitory concentration (MIC) was studied using a two-fold dilution method with the first test concentration of 30 µg/ml [11]. The minimum inhibitory concentration was calculated as the minimum dose of the NPs inhibiting the visual growth of the test cultures on the agar plates. The culture tests were conducted in triplicates. 2.7. Intracellular protein leakage 30 microgram/ml of NPs for 8 h at 37 centigrade, were used for a treat of the bacteria cultures. After incubation, the bacteria were centrifuged at 5000 rpm for 10 min and then supernatants were collected. For evaluation of the intracellular protein leakage, the supernatants were assayed according to the method of Bradford (1976). The assay consisted of 1 ml of supernatant, 0.5 M NaOH (2 ml), and 0.1 N folin (0.1 ml) phenol reagent; absorbance of the solutions was read at 550 nm after 10 min. 3. Statistical Analysis Standard deviation (SD) was measured for antibacterial and protein leakage assays. For differences between treat and control group data in protein leakage assay were carried out by Student's t-test. P < 0.05 was significant. 4. Results 4.1. Phytochemical analysis GC-MS analysis for ginger extract was showed that 101 compounds (Table 1 ). The result showed that major compounds were Coronene (13.5%), trans-Caryophyllene (12.2%), Chavicol (11.9%), ACETONITRILE (11.8%), and EPOXYSPIRO (5.36%). Table 1 Compounds of ginger extract analyzed by GC-MS. NO: Type of Component Area (%) Time (min) 1 Chavicol 11.9% 27.13 2 Isopropyl 2% 26.52 3 ACETONITRILE 11.8% 26.41 4 Coronene 13.5% 26.33 5 Pentenamide 2.22% 26.23 6 Phosphine 1.27% 26.07 7 trans-Caryophyllene 12.2% 25.55 8 Benzoic acid 1.10% 25.42 9 Homobrend 4.49% 25.24 10 Benzaldehyde 1.10% 25.11 11 Tricyclo 3.26% 25.02 12 EPOXYSPIRO 5.36% 24.73 13 Longiverbenone 3.34% 24.57 14 Phenanthrenecarboxylic acid 3.18% 24.38 15 Seneciphylline 1.91% 23.21 16 Cyclohexene 4.06% 22.36 17 TETRAHYDROQUINOLINE 5.03% 22.1 18 Benzopyran 0.55% 9.35 19 1H-Benzimidazole 0.79% 8.14 20 2,3-Dimethylbenzofuran 0.63% 8.08 21 7-Methyl-1-indanone 0.48% 7.89 22 Benzimidazole 0.56% 7.8 23 Triazolo 0.09% 3.81 24 Quinoxaline 0.37% 3.55 25 5-ethyl-5-fluorobarbituric acid 0.61% 2.92 4.2. Characterization of biosynthesized nanoparticles The visual examination or color change test was used to indicate the green and chemic synthesis of Au NPs (Fig. 1 ) and Ag NPs (Fig. 2 ), which confirmed that the reduction of metal ions to metal NPs results in a color change of the solution. Thereafter, the UV-Vis spectrum was used to find out the stability and bioreduction of metal NPs in the solution. In the present study, UV-Vis analysis revealed the maximum absorption peaks of green synthesized Au NPs and Ag NPs were at 523 and 432.5 nm, respectively (Fig. 2 ). Also, the Au and Ag NPs amount were measured by atomic absorption spectroscopy (AAS), results show that the amounts of those are 1.531 and 2.025 mg/L, respectively. DLS method indicated that the average particle size for green synthesized Au and Ag NPs are 314 and 225 nm respectively (Fig. 3 ), also average particle size for chemical synthesized Au and Ag NPs are 42 and 27 nm respectively. Zeta potential values give information about the stability of the NPs that for green synthesized Au and Ag NPs value were − 7.11 and 4.83 mv, respectively, which confirm the high stability of biosynthesized NPs. TEM techniques were used for studying the morphology and sizes of Au NPs and Ag NPs (Fig. 4 ). TEM images of NPs showed the particles distributed individually in different shapes, such as hexagon and spheres, with sizes ranging from 15–25 nm for Au green synthesized nanoparticles and less than 15 nm for Ag green synthesized NPs. Also, TEM study was shown that sizes ranging for chemical synthesized Au nanoparticles from 15–25 and 20–70 nm for Ag NPs. FTIR spectrum (Fig. 5 ) of ginger root extract shows the band at 3441 cm − 1 which is assigned to O–H stretching of phenolic compounds, water, and fatty acids. The band 2933 cm − 1 is assigned to C–H stretching of methylene group in esters, fatty acids, and aliphatic hydrocarbons, 1738 cm − 1 is assigned to C = O stretching of aldehyde, esters, fatty acid, and ketones, 1620 cm − 1 is assigned to (H–O–H) bending of water, 1517 cm − 1 is due to C = C stretching of aromatic elements, 1462 cm − 1 is assigned to C–O–H in-plane bending of fatty acids and other compound’s, 1269 cm − 1 is assigned to C–O stretching of ester and fatty acid and 1044 cm − 1 is due to C–O stretching of alcohols, phenols. By comparing the infrared spectra of plant extract and green synthesized NPs (Fig. 5 ) it is observed that the intensity of the peaks at 3450 cm − 1 for Ag, and 3435 in Au, 2928 cm − 1 for Ag and 2928 cm − 1 in Au NPs, 1735 cm − 1 in Ag and 1738 cm − 1 in Au, 1460 cm − 1 in Ag and 1384 cm − 1 in Au, and1108 cm 1 in Au and 1107 cm − 1 in Ag in compare with plant extract spectrum has decreased/increased and then shifted to higher/lower wavenumbers. The band at 3450 cm − 1 is assigned to O–H stretching of water, 2928 cm − 1 is assigned to methyl C–H stretching of esters, 1620cm − 1 is assigned to H–O–H bending of water, 1383 cm − 1 is assigned to methyl symmetrical C–H bending of esters and band at 1107 cm 1 is assigned to C–O stretching of carbohydrates, ester. The peak of 2928 cm − 1 which is related to C-H stretch of aliphatic fatty acids, esters, and hydrocarbons in the plant root extract became less intense and shifted to 2923 cm − 1 . The band at 1620 cm − 1 assigned to H–O–H bending of water became less intense and shifted to 1628 cm − 1 . The band at1517 cm − 1 which is assigned to C = O stretching of esters, aldehyde, ketones, and fatty acid disappeared. Further, the peak at 1383 cm − 1 which is assigned to methyl symmetrical C–Hbending of esters became sharp. 4.3. Antibacterial activity Due to prove the antibacterial activity of the roots extract, in this study at the first we evaluated the antibacterial activity of ginger extract root (Table 2 ). Better than chemically synthesized once, the green synthesized Au and Ag NPs have shown acceptable bacterial growth inhibitory and also shown a mean zone of inhabitation of S. aureus and E. coli (Table 3 ). The IC 50 and IC 90 values for Au NPs and Ag NPs were noted to be 7.5 and 7.3 µg/ml and 15 and 15.2 µg/ml for both bacterial strains, respectively (Figs. 6 , and Fig. 7 ). Table 2 Antibacterial activity of ginger root extract. The pure ginger root extract (µg/ml) Concentration 500 250 125 62.5 31.25 15.62 7.81 3.90 S. aureus - - - - + + + + E. coli - - + + + + + + Table 3 Antibacterial activity of green and chemical synthesized Ag/Au NPs. Ag (µg/ml) Concentration 27.72 13.86 6.93 3.46 1.73 0.86 0.43 0.21 S. aureus - - - + + + + + E. coli - + + + + + + + Green-Ag S. aureus - - - - - + + + E. coli - - - - - - - + Au (µg/ml) Concentration 17.95 8.97 4.48 2.24 1.12 0.56 0.28 0.14 S. aureus - + + + + + + + E. coli + + + + + + + + Green-Au S. aureus - + + + + + + + E. coli + + + + + + + + 4.4. Protein leakage The total amount of protein leakages upon treatment with green synthesized AuNPs and Ag NPs were quantified. The result showed that protein leakage for treated bacterial cells with green and chemical synthesized NPs was higher when compared to the untreated groups, but the amount of that for Ag NPs was higher than from Au NPs (Table 4 ). This indicates that NPs disrupted the bacteria cells membrane and enhanced the protein leakage. Table 4 Quantification of protein leakage level in NPs treated bacterial species. Bacteria Control (%) G-AgNPs treated cell (%) G-AuNPs treated cell (%) S. aureus 7.18 ± 2.5 10.23 ± 1.77 a 10.07 ± 1.71 a E. Coli 11.01 ± 0.69 15.41 ± 0.37 a 13.19 ± 0.23 a a P < 0.05, Experiment performed in triplicates and statistical analysis using a student-t-test. 5. Discussion Earlier studies were showed that the ginger extract contains n. hexane, ethyl acetate, and soxhlet which those compounds have an antibacterial effect and also inhibit the growth of the bacterial biofilm [12]. In the present study chemical composition of ginger root extract is made up of gingerol, shogaols, zingerone, paradol, and starch. The rhizome, consisting of 6-gingerol and 6-shogaol, is the principal source of gingerol and shogaol, as previously reported were found in high levels in the ginger extract [13, 14]. The key compounds responsible for the reduction of Au and Ag ions to NPs are water-soluble ingredients present in the ginger root extract. Ginger holds chemical compounds like oxalic acid, ascorbic acid, phenylpropanoids, and zingerone. The Au NPs and Ag NPs can be reduced by the ascorbic acid and/or oxalic acid present in the ginger root extract. The possible stages of the formation of NPs from ginger extract during the chemical reaction include nucleation, condensation, surface reduction, and stabilization as previously described [15, 16]. Results indicated that during NPs synthesis, the biodegradable components of root extract can act both as reducing and capping agents, thus promoting the formation of NPs while inhibiting their aggregation via increasing their stability [17]. This finding also presents the potentials of plants root extract as biological “nano-factories” providing non-toxic reducing-capping agents and offering a clean, highly tunable, and environmentally benign method for producing desired NPs [18]. Although the idea of utilizing living plants is revolutionary; nevertheless, the difficulty of purification of the intracellularly formed NPs directed studies to utilize the extracts of plants for extracellular syntheses of NPs [19]. The UV-Vis analysis-peaks indicate that green NPs were synthesized and consistent with the results of previous studies that have shown the range of 400–450 nm for Ag NPs and in the range of 500–550 nm in case of Au NPs [20]. According to Zeta potential data, the surface charge of Ag NPs is more positive than Au NPs, which might potent them for better binding to the outer membrane of Gram-negative bacteria with a negative charge and thereby modulate their activity [21]. Also, Zeta potential for chemical synthesized Au and Ag nanoparticles values are 0 and − 10.1mv, respectively. Elia et al. synthesized the Au nanoparticle from P. granatum and characterized them with using DLS spectroscopy which particle size range was 34–312 nm [22]. Inconsistency with our study, Sujitha et al. [23] reported that the lower concentration of the plant extract leads to Au NPs with a lower ZP value. These findings reveal that biological extracts from plants' roots provide the method for producing NPS with a broad range of sizes [3], and since they are also originally naturals, so covering the NPs surface improves their biocompatibility for in vivo applications [18]. Addressing the TEM results, as previously studied, the ratio of plant extract, type of components, and the initial metal salt in the reaction medium affected the Au NPs' size and the shape [24]. Similarly, a study showed that the synthesis of NPs using a marigold flower, where TEM analysis showed spherical and hexagonal shape particles in the range between 10 to 90 nm [25]. Thus, the TEM and DLS studies gave similar results for the size range of the NPs. The FTIR analysis indicated the presence of phenolic groups which are suggested responsible for the reduction of silver ions [26]. The presence of other FTIR-assocated peaks confirmed that the NPs were covered by ginger root extract with functional groups such as carboxylic acid, ketone, aldehyde, and other functional groups. The presence of these functional groups is due to the biostability of the NPs. It confirms that NPs synthesized from the ginger root extracts are stabilized by phytoconstituents through functional groups [27, 28]. Prakash Patil groups synthesized Ag NPs using flower extract of MadhucaIongifolia as a reduction agent and synergic effect. Green synthesized NPs show potential antibacterial activity against Gram-negative and Gram-positive bacteria. MadhucaIongifolia flower is a good source for NPs synthesis. According to obtained data, synthesized Ag NPs are applicable as an antibacterial agent in therapeutics. This was explained by the fact that the antibacterial activity was due to the change in membrane permeability [29]. After entering the cytoplasm, Ag NPs induce reactive oxygen species (ROS) production and by binding the phosphate group of effector molecules disturbing the protein synthesis and thus, causes bacterial growth inhibition or killing [30]. Due to the high prevalence, antibiotic resistance and pathogenicity of S. aureus and E. coli we studied them. These two pathogens are the causative agents for several infections, such as endocarditis, urinary tract infection, osteomyelitis, and septicemia [31, 32]. Metal NPs especially once having a relatively large size/surface ratio or smaller than 20 nm act as destroyers of the cell membrane through binding to cells, causing structural alterations and eventually the loss of the semi-permeability of the membrane [6]. Studies have shown that bacterial cells membrane disruption by Psidiumguajava leaf extracts is in agreement with the result of this study [33]. Overall, the synthesized NPs exhibited pronounced antibacterial activities on S. aureus and on E. coli. Similar to these findings, Janaki et al showed that zinc oxide nanoparticle (ZnO NPs) which was synthesized using ginger extracted root has an efficient antimicrobial activity [34]. In the other study, Kumor and co-workers utilized ginger extract green synthesized gold NPs and evaluated of blood compatibility of them. The result showed that biosynthesized NPs are suitable vectors for medical applications [13]. These findings explain the possible antibacterial actions of green synthesized NPs: (i) may be due to DNA damage, (ii) protein synthesis inhibition and denaturation, and (iii) formation of free radicals causing cell wall damage [35] (Fig. 8 ). 6. Conclusion The present study reports the chemical and green synthesis of Au and AgNPs using citric acid and aqueous root extract of ginger, respectively. Biosynthetic NPs were characterized that are spherical and hexagonal shapes with an average size ≤ 100 nm. The biosynthesized Au and AgNPs showed an excellent antibacterial effect against S. aureus and E. coli in comparing chemical synthesized NPs. Further, the possible anti-bacterial mechanisms were studied by protein leakage. The Result of this study proves that biosynthesized Au and Ag NPs could be used as an alternative therapy to control and eliminate the infection caused by E. coli and S. aureus. Abbreviations FTIR, Fourier transforms infrared spectroscopy; MIC, minimum inhibitory concentration; NPs nanoparticles; ROS, reactive oxygen species; TEM, Transmission electron microscopy Declarations Acknowledgements The authors appreciate the personnel of Drug Applied Research Center for help and guide Availability of data and materials The data that support the findings of this study are available from the corresponding authors upon reasonable request. Contributions MY was a significant contributor to doing and writing the manuscript. MA, HDM and MM collaborated in doing the thesis that results in the paper. AA and MMD designed and supervised the manuscript. All authors read and approved the final manuscript. Ethics declarations Ethics approval and consent to participate Not applicable. Consent for publication Not applicable. Competing interests The authors declare that they have no competing interests. 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Proceedings of the National Academy of Sciences. 2014;111(15):5694-9. Henie E, Zaiton H, Suhaila M. Bacterial membrane disruption in food pathogens by Psidium guajava leaf extracts. Int Food Res J. 2009;16(3):297–311. Janaki AC, Sailatha E, Gunasekaran S. Synthesis, characteristics and antimicrobial activity of ZnO nanoparticles. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy. 2015;144:17–22. Kim S-H, Lee H-S, Ryu D-S, Choi S-J, Lee D-S. Antibacterial activity of silver-nanoparticles against Staphylococcus aureus and Escherichia coli. Korean J Microbiol Biotechnol. 2011;39(1):77–85. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 2 posted You are reading this latest preprint version Show more versions Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-1352913","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":89265607,"identity":"5f3f4ca0-2860-4fb8-91a7-218078706996","order_by":0,"name":"Morteza Yadi","email":"","orcid":"","institution":"Tabriz University of Medical Sciences","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Morteza","middleName":"","lastName":"Yadi","suffix":""},{"id":89265608,"identity":"beba1e0a-cc81-4021-8220-6744a9473291","order_by":1,"name":"Abolfazl Akbarzadeh","email":"","orcid":"","institution":"Tabriz University of Medical Sciences","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Abolfazl","middleName":"","lastName":"Akbarzadeh","suffix":""},{"id":89265609,"identity":"4020ecb1-a531-415f-99dd-b80287be7a89","order_by":2,"name":"Hassan Dianat-Moghadam","email":"","orcid":"","institution":"Isfahan University of Medical Sciences","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Hassan","middleName":"","lastName":"Dianat-Moghadam","suffix":""},{"id":89265610,"identity":"630fa26a-1bf1-4133-bdf1-2e877435e579","order_by":3,"name":"Mehdi Azizi","email":"","orcid":"","institution":"Hamadan University of Medical Sciences","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Mehdi","middleName":"","lastName":"Azizi","suffix":""},{"id":89265611,"identity":"e0f84b7b-cc84-4b72-996c-de96e062d3a8","order_by":4,"name":"Morteza Milani","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA2klEQVRIie3RoQrCUBTG8W8MTBdXFWE+gXBlYNnLKIaVTRYNhiuDLdl9DEEwKweuZWIdLPgMuqiIOkEE4TpMhvtv58CPEw6g0/1jpikADlhg5Vgu12piPEkzrkzwIPe4fCPKOokxLcJwZTuHGZ2ysQsrWRs0VpAeGVFrznOnJ+t9x089NNI+NqmaiBbj+WAlGR8GMQEZsBFfrpwfZBkzTsGV0K5A4vLKosa6USAIvApx78RpyNHQ9KXHuulAqMmeKGeX3LaiHRX+xLXtLdFRRT5ir0/pdDqd7vduKvBNrqAnukAAAAAASUVORK5CYII=","orcid":"","institution":"Tabriz University of Medical Sciences","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Morteza","middleName":"","lastName":"Milani","suffix":""}],"badges":[],"createdAt":"2022-02-12 11:29:08","currentVersionCode":2,"declarations":"","doi":"10.21203/rs.3.rs-1352913/v2","doiUrl":"https://doi.org/10.21203/rs.3.rs-1352913/v2","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":19038065,"identity":"d25dba21-813a-47ce-84c1-402674aa5ca1","added_by":"auto","created_at":"2022-03-09 19:10:08","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":460449,"visible":true,"origin":"","legend":"\u003cp\u003eSchematic representation of the synthesis of Au and Ag NPs using ginger root extract and Trisodium Citrate. \u003cstrong\u003ea\u003c/strong\u003e) Green synthesis of Au NPs using Ginger root extract. \u003cstrong\u003eb\u003c/strong\u003e) Chemical synthesis of Au NPs with use of Trisodium Citrate. \u003cstrong\u003ec\u003c/strong\u003e) Green synthesis of Ag NPs using Ginger root extract. \u003cstrong\u003ed\u003c/strong\u003e) Chemical synthesis of Ag NPs using trisodium citrate.\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Fig1.png","url":"https://assets-eu.researchsquare.com/files/rs-1352913/v2/6b3a95046e9bf40b4ee58e5f.png"},{"id":19037307,"identity":"b7809915-4abd-4d46-9036-c53fde7818d5","added_by":"auto","created_at":"2022-03-09 19:07:08","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":215766,"visible":true,"origin":"","legend":"\u003cp\u003eUV-vis spectra of green synthesized Au (right) and Ag (left) NPs.\u003c/p\u003e","description":"","filename":"Fig2.png","url":"https://assets-eu.researchsquare.com/files/rs-1352913/v2/a01c75bf9007a7e7538ffee5.png"},{"id":19038064,"identity":"d70c0856-c2b0-4826-bb56-b430bca9c0cf","added_by":"auto","created_at":"2022-03-09 19:10:08","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":440594,"visible":true,"origin":"","legend":"\u003cp\u003eDLS profile and zeta potential analysis of green and chemical synthesized NPs. \u003cstrong\u003ea\u003c/strong\u003e) size of biosynthesized Ag, \u003cstrong\u003eb\u003c/strong\u003e) size of biosynthesized Au, \u003cstrong\u003ec\u003c/strong\u003e) size of chemical synthesized Ag and D) size of chemical synthesized Au NPs. E) zeta potential of biosynthesized Au, F) zeta potential of biosynthesized Ag, \u003cstrong\u003eg\u003c/strong\u003e) zeta potential of chemical synthesized Ag, and \u003cstrong\u003eh\u003c/strong\u003e) zeta potential of chemical synthesized Au NPs.\u003c/p\u003e","description":"","filename":"Fig3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-1352913/v2/bdec2ef72bafe5c506bdfa83.jpg"},{"id":19036452,"identity":"12e1100a-1462-469f-a176-22228a956750","added_by":"auto","created_at":"2022-03-09 19:04:08","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":135892,"visible":true,"origin":"","legend":"\u003cp\u003eTEM image of NPs biosynthesized and chemical using ginger root extract and citrate. A) TEM imaging of biosynthesized Au NPs, B) TEM imaging of chemically synthesized Ag NPs, C) TEM imaging of biosynthesized Ag NPs, and D) TEM imaging of chemically synthesized Ag NPs.\u003c/p\u003e","description":"","filename":"Fig4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-1352913/v2/adfb23f60e7d999d8db66c9f.jpg"},{"id":19036459,"identity":"8b4323eb-0926-451a-8898-b6b14aa751b8","added_by":"auto","created_at":"2022-03-09 19:04:08","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":138850,"visible":true,"origin":"","legend":"\u003cp\u003eFTIR spectra of (a) ginger root extract and (b) capped reducing phytoconstituents responsible for the synthesis of Au (left) and Ag (right) NPs.\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Fig5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-1352913/v2/6b839628fb85d69093438409.jpg"},{"id":19037308,"identity":"4e318d7d-62b5-4bb5-8987-ce043c6f61ee","added_by":"auto","created_at":"2022-03-09 19:07:08","extension":"jpg","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":234654,"visible":true,"origin":"","legend":"\u003cp\u003eAntibacterial activity of chemical and biosynthesized silver NPs.\u0026nbsp;\u003c/p\u003e","description":"","filename":"Fig6.jpg","url":"https://assets-eu.researchsquare.com/files/rs-1352913/v2/4627a97452f54754bc0ab92e.jpg"},{"id":19036457,"identity":"5ca6b8d3-44d0-4ef1-9c59-44c23022ce18","added_by":"auto","created_at":"2022-03-09 19:04:08","extension":"jpg","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":232914,"visible":true,"origin":"","legend":"\u003cp\u003eAntibacterial activity of chemical and biosynthesized gold NPs.\u003c/p\u003e","description":"","filename":"Fig7.jpg","url":"https://assets-eu.researchsquare.com/files/rs-1352913/v2/b4c4bd901c7770b4fa92ccbc.jpg"},{"id":19037311,"identity":"ae74d3a4-9398-41ca-8780-f32de8b74106","added_by":"auto","created_at":"2022-03-09 19:07:08","extension":"jpg","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":111217,"visible":true,"origin":"","legend":"\u003cp\u003eA possible mode of antibacterial action of green synthesized NPs.\u003c/p\u003e","description":"","filename":"Fig8.jpg","url":"https://assets-eu.researchsquare.com/files/rs-1352913/v2/825106b611be0f7e279c51a6.jpg"},{"id":19699559,"identity":"bd5f2c8a-6a70-413f-9d0a-e1011bf6e682","added_by":"auto","created_at":"2022-03-28 18:29:29","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1881933,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1352913/v2/d08e04da-8a3f-4dbf-87f0-4e28683ddfb1.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Antibacterial activity of green gold and silver nanoparticles using ginger root extract","fulltext":[{"header":"1. Background","content":"\u003cp\u003eBiofilms are microbial populations enclosed in a matrix. Those grow in three steps, (i) initial adhesion, (ii) proliferation, and (iii) detachment. Bacterial cells bind together by extracellular polymeric substances and are connected to a substrate surface, involved in each cell-cell interaction and cell surface as a part of the developmental process. Biofilms show high resistance towards toxicants if compared to planktonic cells [1, 2]. They can cause of lead to infections in humans and animals and serious problems in the environment. Infectious diseases are one of the health threats on human society. The use of antibacterial drug for the control and treatment of infectious disease are common worldwide. However, continued use from the antibacterial drug leads to drug resistance [3]. In this condition use of the drug for control and treat of infectious diseases don't be effective. Eliminating this problem requires a new therapeutic agent for control and treatment of the infectious disease [4].\u003c/p\u003e \u003cp\u003eNowadays, silver (Ag) and gold (Au) nanoparticles (NPs) are used in a wide range of medicine. There are several methods including laser ablation, gamma irradiation and use of the chemical agent as a reducing and capping agent, for the synthesis of the Au and Ag NPs [3]. The important problems with these methods are expensive, and use of toxic chemical agents that are not safe for human health and the environment [5]. The green synthesis of the NPs, also known as photosynthesis, is one of the emerging fields in nanotechnology. Green synthesis NPs have shown high activity against the primary biofilm. Plants are used for the synthesis of NPs and have advantages over physical and chemical processes [6]. In recent years, NP synthesis using plant extracts has been increasing because, these are available, environmentally friendly, and easy to use, and have a wide range of secondary metabolites that act as a reducing agent [7].\u003c/p\u003e \u003cp\u003eGinger (Zingiber officinale), Roscoe belonging to the family Zingiber aceae, is a perennial herb with thick tuberous rhizomes. Ginger extracts have antibacterial activity. Malu and co-workers show that material such n-hexane, ethyl acetate and soxhlet which are in the ginger extract solution, have antibacterial effects. In the fact this material in addition to having bactericidal activity, inhibition of bacterial growth [8]. \u003cem\u003eE. coli\u003c/em\u003e (gram-negative bacteria) and \u003cem\u003eS. aureus (\u003c/em\u003egram-positive bacteria) have an important role in human infectious diseases. \u003cem\u003eE. coli\u003c/em\u003e through penetration to lymphocytes and an inflammatory reaction with the host, by causing bloody diarrhea [9]. \u003cem\u003eS. aureus\u003c/em\u003e is the main cause of food poisoning and surgical wounds infection which together with epidermidis syndrome, causes infections associated with medical equipment [10].\u003c/p\u003e \u003cp\u003eIn this research, gold and silver nanoparticles were synthesized with green and chemical synthesis methods. For this purpose, Ginger root extract and citrate were used as reducing agents, respectively. The synthesized metal NPs were characterized by dynamic light scattering (DLS), transmission electron microscope (TEM), ultraviolet-visible spectroscopy UV-Vis, Atomic absorption spectroscopy (AAS), and Fourier transforms infrared spectroscopy (FTIR). The antibacterial activity of green and chemical synthesized Au and Ag NPs were investigated with \u003cem\u003eE. coli\u003c/em\u003e and \u003cem\u003eS. aureus\u003c/em\u003e strains in vitro.\u003c/p\u003e"},{"header":"2. Materials And Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1. Microorganisms\u003c/h2\u003e \u003cp\u003eStandard strains of \u003cem\u003eS. aureus\u003c/em\u003e (ATTC 25923) and \u003cem\u003eE. coli\u003c/em\u003e (ATTC 25922) were purchased from the Iranian Research Organization for Science and Technology (IROST). These strains were cultivated in the nutrient broth medium and were incubated at 37\u0026deg;C for 24h. For further experiments, small amount of bacterial colonies was stored in Tripticas soy broth containing glycerol at -70\u0026deg;C.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2. Preparation of ginger extracts\u003c/h2\u003e \u003cp\u003eRoots of ginger were purchased from the local market, the Islamic Republic of Iran, and washed frequently with ultra-pure deionized water. After shredding the ginger root, dried and crashed into powder by the steel hammer. After that, 2 gr of powder was mixed with 80 ml of ethanol and incubated at 40\u0026deg;C for 24 hours. Then for obtaining ginger extract, the solution was filtered with Whatman No.1 filter paper.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3. Green and chemical synthesis of gold and silver nanoparticles\u003c/h2\u003e \u003cp\u003eFor the green synthesis of Au NPs, 1 mL ginger root extract was added to a 50 mL boiling solution of HAuCl\u003csub\u003e4\u003c/sub\u003e\u0026middot;3H\u003csub\u003e2\u003c/sub\u003eO (1mM) and the boiling continued for 5 min. Then, the solution was kept undisturbed at room temperature until the colorless solution converted to a wine red color which indicated the formation of Au NPs. For the chemical synthesis of Au NPs, HAuCl\u003csub\u003e4\u003c/sub\u003e solution is boiled and then trisodium citrate dehydrate was added slowly into the boiling solution under stirring. A few minutes later, the color of the solution from light yellow converted to wine red.\u003c/p\u003e \u003cp\u003eFor green synthesis, like above, 1 ml of ginger extract was added to with 100 ml of silver nitrate (2mM) boiling solution. The reaction mixture was kept undisturbed at room temperature until the colorless solution converted to reddish brown color which indicated the formation of Ag NPs. For the chemical synthesis of silver nanoparticles, silver nitrate solution and citrate of sodium were used as a metal ion source and reduction agent, respectively. Also, citrate of sodium was used as a reducing agent. When the citrate sodium was added to the silver nitrate the color of the solution converted from pale yellow to pale brown color. And the final silver nanoparticle was purified by centrifugation.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.4. Gas chromatography-mass spectrometry analysis (GC-MS)\u003c/h2\u003e \u003cp\u003eGinger extract was analyzed by a mass scientific trace 2200 (gas chromatography) system with a thermalSaturnmass selective detector (Varian Company). The machine was equipped with a TG-5MS (mass spectroscopy) column (30 * 0.25 mm (5% phenyl) -methylpolysiloxane capillary column, film thickness * 0.25 micrometer), 220 centigrade temperature injector and 250 centigrade temperature transfer line. The temperature of the oven was programmed as follows: initial temperature; 50 centigrade for 5 min and then increase 4\u0026deg;C/min up to 250 centigrade. The gas carrier used was He at a flow rate of 1.0 ml/min. 1microliter sample was injected and the ionization energy was 70 eV. The base of the identification of individual components was based on their retention time and by comparison of their mass spectral pattern with standard library data. (National Institute of standards and technology).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e2.5. Characterization of gold and silver nanoparticles\u003c/h2\u003e \u003cp\u003eThe reduction of gold and silver ions was monitored by a UV-Vis spectrophotometer (SPECTOR 250, Analytic Jena) in the 350\u0026ndash;800 nm wavelength range. Due to the evaluation of concentration of green and chemical synthesized NPs, a solution of synthesized NP was diluted, and then the amount of Au and Ag NPs were measured by atomic absorption spectroscopy (AAS) (NovaAA400, Analytic Jena Co). Also for determination of shape and size of the nanoparticle were used from Transmission electron microscopy (TEM) (Zeiss Leo q06) operating at 200 kV accelerating voltage. For the preparation of the sample; 10 microliter of aliquots of NPs solution was drop-casting onto a carbon-coated copper grid and then was placed on a piece of paper to get rid of excess solvent. For determination of the average particle size, distribution, and stability of the gold and silver NPs were used from ELSZ-1000 zeta-potential and particle sizer (Mastersizer 2000, Malvern, USA). For FTIR analysis, the powdered gold and silver NPs were recorded by FTIR spectrometer (Tensor 27, Bruker Co) over the 4000\u0026thinsp;\u0026minus;\u0026thinsp;400 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e frequency with 4 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e resolutions by using a KBr pellet method.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e2.6. Antibacterial assay\u003c/h2\u003e \u003cp\u003eThe antibacterial activity of chemical and green synthesizedAu and Ag NPs were performed by well diffusion agar method. Standard strains of bacteria were subculture on plates containing Muller Hinton agar using the pour plate method. Then the wells which have 6mm diameter were punctured onto the agar plates and 25\u0026micro;g/ ml of Au and Ag NPs solution and aqueous plant extract were loaded into the wells. After 24 hours of incubation, the inhibition of zone diameter around of wells was measured. For comparison of the effectiveness of Au and Ag NPs and ginger extract against tested bacteria, we used Streptomycin (30 \u0026micro;g/ml). For evaluation of the minimum inhibitory concentration (MIC) was studied using a two-fold dilution method with the first test concentration of 30 \u0026micro;g/ml [11]. The minimum inhibitory concentration was calculated as the minimum dose of the NPs inhibiting the visual growth of the test cultures on the agar plates. The culture tests were conducted in triplicates.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e2.7. Intracellular protein leakage\u003c/h2\u003e \u003cp\u003e30 microgram/ml of NPs for 8 h at 37 centigrade, were used for a treat of the bacteria cultures. After incubation, the bacteria were centrifuged at 5000 rpm for 10 min and then supernatants were collected. For evaluation of the intracellular protein leakage, the supernatants were assayed according to the method of Bradford (1976). The assay consisted of 1 ml of supernatant, 0.5 M NaOH (2 ml), and 0.1 N folin (0.1 ml) phenol reagent; absorbance of the solutions was read at 550 nm after 10 min.\u003c/p\u003e \u003c/div\u003e"},{"header":"3. Statistical Analysis","content":"\u003cp\u003eStandard deviation (SD) was measured for antibacterial and protein leakage assays. For differences between treat and control group data in protein leakage assay were carried out by Student's t-test. P\u0026thinsp;\u0026lt;\u0026thinsp;0.05 was significant.\u003c/p\u003e"},{"header":"4. Results","content":"\u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003e4.1. Phytochemical analysis\u003c/h2\u003e \u003cp\u003eGC-MS analysis for ginger extract was showed that 101 compounds (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). The result showed that major compounds were Coronene (13.5%), trans-Caryophyllene (12.2%), Chavicol (11.9%), ACETONITRILE (11.8%), and EPOXYSPIRO (5.36%).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eCompounds of ginger extract analyzed by GC-MS.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNO:\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eType of Component\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eArea (%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eTime (min)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eChavicol\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11.9%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e27.13\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eIsopropyl\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e26.52\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eACETONITRILE\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11.8%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e26.41\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCoronene\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e13.5%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e26.33\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePentenamide\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.22%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e26.23\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePhosphine\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.27%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e26.07\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003etrans-Caryophyllene\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e12.2%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e25.55\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eBenzoic acid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.10%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e25.42\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eHomobrend\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e4.49%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e25.24\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eBenzaldehyde\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.10%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e25.11\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e11\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTricyclo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.26%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e25.02\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e12\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eEPOXYSPIRO\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e5.36%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e24.73\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e13\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eLongiverbenone\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.34%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e24.57\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePhenanthrenecarboxylic acid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.18%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e24.38\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSeneciphylline\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.91%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e23.21\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCyclohexene\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e4.06%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e22.36\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e17\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTETRAHYDROQUINOLINE\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e5.03%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e22.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eBenzopyran\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.55%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e9.35\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e19\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1H-Benzimidazole\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.79%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e8.14\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2,3-Dimethylbenzofuran\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.63%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e8.08\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e7-Methyl-1-indanone\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.48%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e7.89\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e22\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eBenzimidazole\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.56%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e7.8\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e23\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTriazolo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.09%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e3.81\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e24\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eQuinoxaline\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.37%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e3.55\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e25\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5-ethyl-5-fluorobarbituric acid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.61%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e2.92\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003e4.2. Characterization of biosynthesized nanoparticles\u003c/h2\u003e \u003cp\u003eThe visual examination or color change test was used to indicate the green and chemic synthesis of Au NPs (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e) and Ag NPs (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e), which confirmed that the reduction of metal ions to metal NPs results in a color change of the solution.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThereafter, the UV-Vis spectrum was used to find out the stability and bioreduction of metal NPs in the solution. In the present study, UV-Vis analysis revealed the maximum absorption peaks of green synthesized Au NPs and Ag NPs were at 523 and 432.5 nm, respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Also, the Au and Ag NPs amount were measured by atomic absorption spectroscopy (AAS), results show that the amounts of those are 1.531 and 2.025 mg/L, respectively.\u003c/p\u003e \u003cp\u003eDLS method indicated that the average particle size for green synthesized Au and Ag NPs are 314 and 225 nm respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e), also average particle size for chemical synthesized Au and Ag NPs are 42 and 27 nm respectively. Zeta potential values give information about the stability of the NPs that for green synthesized Au and Ag NPs value were \u0026minus;\u0026thinsp;7.11 and 4.83 mv, respectively, which confirm the high stability of biosynthesized NPs.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eTEM techniques were used for studying the morphology and sizes of Au NPs and Ag NPs (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). TEM images of NPs showed the particles distributed individually in different shapes, such as hexagon and spheres, with sizes ranging from 15\u0026ndash;25 nm for Au green synthesized nanoparticles and less than 15 nm for Ag green synthesized NPs. Also, TEM study was shown that sizes ranging for chemical synthesized Au nanoparticles from 15\u0026ndash;25 and 20\u0026ndash;70 nm for Ag NPs.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eFTIR spectrum (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e) of ginger root extract shows the band at 3441 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e which is assigned to O\u0026ndash;H stretching of phenolic compounds, water, and fatty acids. The band 2933 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is assigned to C\u0026ndash;H stretching of methylene group in esters, fatty acids, and aliphatic hydrocarbons, 1738 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is assigned to C\u0026thinsp;=\u0026thinsp;O stretching of aldehyde, esters, fatty acid, and ketones, 1620 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is assigned to (H\u0026ndash;O\u0026ndash;H) bending of water, 1517 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is due to C\u0026thinsp;=\u0026thinsp;C stretching of aromatic elements, 1462 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is assigned to C\u0026ndash;O\u0026ndash;H in-plane bending of fatty acids and other compound\u0026rsquo;s, 1269 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is assigned to C\u0026ndash;O stretching of ester and fatty acid and 1044 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is due to C\u0026ndash;O stretching of alcohols, phenols.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eBy comparing the infrared spectra of plant extract and green synthesized NPs (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e) it is observed that the intensity of the peaks at 3450 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e for Ag, and 3435 in Au, 2928 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e for Ag and 2928 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e in Au NPs, 1735 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e in Ag and 1738 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e in Au, 1460 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e in Ag and 1384 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e in Au, and1108 cm\u003csup\u003e1\u003c/sup\u003e in Au and 1107 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e in Ag in compare with plant extract spectrum has decreased/increased and then shifted to higher/lower wavenumbers. The band at 3450 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is assigned to O\u0026ndash;H stretching of water, 2928 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is assigned to methyl C\u0026ndash;H stretching of esters, 1620cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is assigned to H\u0026ndash;O\u0026ndash;H bending of water, 1383 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e is assigned to methyl symmetrical C\u0026ndash;H bending of esters and band at 1107 cm\u003csup\u003e1\u003c/sup\u003e is assigned to C\u0026ndash;O stretching of carbohydrates, ester. The peak of 2928 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e which is related to C-H stretch of aliphatic fatty acids, esters, and hydrocarbons in the plant root extract became less intense and shifted to 2923 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e. The band at 1620 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e assigned to H\u0026ndash;O\u0026ndash;H bending of water became less intense and shifted to 1628 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e. The band at1517 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e which is assigned to C\u0026thinsp;=\u0026thinsp;O stretching of esters, aldehyde, ketones, and fatty acid disappeared. Further, the peak at 1383 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e which is assigned to methyl symmetrical C\u0026ndash;Hbending of esters became sharp.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003e4.3. Antibacterial activity\u003c/h2\u003e \u003cp\u003eDue to prove the antibacterial activity of the roots extract, in this study at the first we evaluated the antibacterial activity of ginger extract root (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Better than chemically synthesized once, the green synthesized Au and Ag NPs have shown acceptable bacterial growth inhibitory and also shown a mean zone of inhabitation of \u003cem\u003eS. aureus\u003c/em\u003e and \u003cem\u003eE. coli\u003c/em\u003e (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). The IC\u003csub\u003e50\u003c/sub\u003e and IC\u003csub\u003e90\u003c/sub\u003e values for Au NPs and Ag NPs were noted to be 7.5 and 7.3 \u0026micro;g/ml and 15 and 15.2 \u0026micro;g/ml for both bacterial strains, respectively (Figs.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e, and Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eAntibacterial activity of ginger root extract.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"9\"\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colspan=\"8\" nameend=\"c9\" namest=\"c2\"\u003e \u003cp\u003eThe pure ginger root extract (\u0026micro;g/ml)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eConcentration\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e500\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e250\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e125\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e62.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e31.25\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e15.62\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e7.81\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e3.90\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cspan type=\"BoldItalic\" class=\"BoldItalic\" name=\"Emphasis\"\u003eS. aureus\u003c/span\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cb\u003e-\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e-\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e-\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cspan type=\"BoldItalic\" class=\"BoldItalic\" name=\"Emphasis\"\u003eE. coli\u003c/span\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eAntibacterial activity of green and chemical synthesized Ag/Au NPs.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"9\"\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colspan=\"9\" nameend=\"c9\" namest=\"c1\"\u003e \u003cp\u003eAg (\u0026micro;g/ml)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eConcentration\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cb\u003e27.72\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e13.86\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e6.93\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e3.46\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e1.73\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e0.86\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u003cb\u003e0.43\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cb\u003e0.21\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cspan type=\"BoldItalic\" class=\"BoldItalic\" name=\"Emphasis\"\u003eS. aureus\u003c/span\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cspan type=\"BoldItalic\" class=\"BoldItalic\" name=\"Emphasis\"\u003eE. coli\u003c/span\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"9\" nameend=\"c9\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eGreen-Ag\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cspan type=\"BoldItalic\" class=\"BoldItalic\" name=\"Emphasis\"\u003eS. aureus\u003c/span\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cspan type=\"BoldItalic\" class=\"BoldItalic\" name=\"Emphasis\"\u003eE. coli\u003c/span\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"9\" nameend=\"c9\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eAu (\u0026micro;g/ml)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eConcentration\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cb\u003e17.95\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e8.97\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e4.48\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e2.24\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e1.12\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e0.56\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u003cb\u003e0.28\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cb\u003e0.14\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cspan type=\"BoldItalic\" class=\"BoldItalic\" name=\"Emphasis\"\u003eS. aureus\u003c/span\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cspan type=\"BoldItalic\" class=\"BoldItalic\" name=\"Emphasis\"\u003eE. coli\u003c/span\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"9\" nameend=\"c9\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eGreen-Au\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cspan type=\"BoldItalic\" class=\"BoldItalic\" name=\"Emphasis\"\u003eS. aureus\u003c/span\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cspan type=\"BoldItalic\" class=\"BoldItalic\" name=\"Emphasis\"\u003eE. coli\u003c/span\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003e4.4. Protein leakage\u003c/h2\u003e \u003cp\u003eThe total amount of protein leakages upon treatment with green synthesized AuNPs and Ag NPs were quantified. The result showed that protein leakage for treated bacterial cells with green and chemical synthesized NPs was higher when compared to the untreated groups, but the amount of that for Ag NPs was higher than from Au NPs (Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). This indicates that NPs disrupted the bacteria cells membrane and enhanced the protein leakage.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eQuantification of protein leakage level in NPs treated bacterial species.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eBacteria\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eControl (%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eG-AgNPs treated cell (%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eG-AuNPs treated cell (%)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eS. aureus\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e7.18\u0026thinsp;\u0026plusmn;\u0026thinsp;2.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e10.23\u0026thinsp;\u0026plusmn;\u0026thinsp;1.77\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e10.07\u0026thinsp;\u0026plusmn;\u0026thinsp;1.71\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eE. Coli\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e11.01\u0026thinsp;\u0026plusmn;\u0026thinsp;0.69\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e15.41\u0026thinsp;\u0026plusmn;\u0026thinsp;0.37 \u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e13.19\u0026thinsp;\u0026plusmn;\u0026thinsp;0.23 \u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"4\" nameend=\"c4\" namest=\"c1\"\u003e \u003cp\u003e\u003csup\u003ea\u003c/sup\u003e P\u0026thinsp;\u0026lt;\u0026thinsp;0.05, Experiment performed in triplicates and statistical analysis using a student-t-test.\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"5. Discussion","content":"\u003cp\u003eEarlier studies were showed that the ginger extract contains n. hexane, ethyl acetate, and soxhlet which those compounds have an antibacterial effect and also inhibit the growth of the bacterial biofilm [12]. In the present study chemical composition of ginger root extract is made up of gingerol, shogaols, zingerone, paradol, and starch. The rhizome, consisting of 6-gingerol and 6-shogaol, is the principal source of gingerol and shogaol, as previously reported were found in high levels in the ginger extract [13, 14].\u003c/p\u003e \u003cp\u003eThe key compounds responsible for the reduction of Au and Ag ions to NPs are water-soluble ingredients present in the ginger root extract. Ginger holds chemical compounds like oxalic acid, ascorbic acid, phenylpropanoids, and zingerone. The Au NPs and Ag NPs can be reduced by the ascorbic acid and/or oxalic acid present in the ginger root extract. The possible stages of the formation of NPs from ginger extract during the chemical reaction include nucleation, condensation, surface reduction, and stabilization as previously described [15, 16].\u003c/p\u003e \u003cp\u003eResults indicated that during NPs synthesis, the biodegradable components of root extract can act both as reducing and capping agents, thus promoting the formation of NPs while inhibiting their aggregation via increasing their stability [17]. This finding also presents the potentials of plants root extract as biological \u0026ldquo;nano-factories\u0026rdquo; providing non-toxic reducing-capping agents and offering a clean, highly tunable, and environmentally benign method for producing desired NPs [18]. Although the idea of utilizing living plants is revolutionary; nevertheless, the difficulty of purification of the intracellularly formed NPs directed studies to utilize the extracts of plants for extracellular syntheses of NPs [19].\u003c/p\u003e \u003cp\u003eThe UV-Vis analysis-peaks indicate that green NPs were synthesized and consistent with the results of previous studies that have shown the range of 400\u0026ndash;450 nm for Ag NPs and in the range of 500\u0026ndash;550 nm in case of Au NPs [20].\u003c/p\u003e \u003cp\u003eAccording to Zeta potential data, the surface charge of Ag NPs is more positive than Au NPs, which might potent them for better binding to the outer membrane of Gram-negative bacteria with a negative charge and thereby modulate their activity [21]. Also, Zeta potential for chemical synthesized Au and Ag nanoparticles values are 0 and \u0026minus;\u0026thinsp;10.1mv, respectively. Elia et al. synthesized the Au nanoparticle from P. granatum and characterized them with using DLS spectroscopy which particle size range was 34\u0026ndash;312 nm [22]. Inconsistency with our study, Sujitha et al. [23] reported that the lower concentration of the plant extract leads to Au NPs with a lower ZP value. These findings reveal that biological extracts from plants' roots provide the method for producing NPS with a broad range of sizes [3], and since they are also originally naturals, so covering the NPs surface improves their biocompatibility for in vivo applications [18].\u003c/p\u003e \u003cp\u003eAddressing the TEM results, as previously studied, the ratio of plant extract, type of components, and the initial metal salt in the reaction medium affected the Au NPs' size and the shape [24]. Similarly, a study showed that the synthesis of NPs using a marigold flower, where TEM analysis showed spherical and hexagonal shape particles in the range between 10 to 90 nm [25]. Thus, the TEM and DLS studies gave similar results for the size range of the NPs.\u003c/p\u003e \u003cp\u003eThe FTIR analysis indicated the presence of phenolic groups which are suggested responsible for the reduction of silver ions [26]. The presence of other FTIR-assocated peaks confirmed that the NPs were covered by ginger root extract with functional groups such as carboxylic acid, ketone, aldehyde, and other functional groups. The presence of these functional groups is due to the biostability of the NPs. It confirms that NPs synthesized from the ginger root extracts are stabilized by phytoconstituents through functional groups [27, 28].\u003c/p\u003e \u003cp\u003ePrakash Patil groups synthesized Ag NPs using flower extract of MadhucaIongifolia as a reduction agent and synergic effect. Green synthesized NPs show potential antibacterial activity against Gram-negative and Gram-positive bacteria. MadhucaIongifolia flower is a good source for NPs synthesis. According to obtained data, synthesized Ag NPs are applicable as an antibacterial agent in therapeutics. This was explained by the fact that the antibacterial activity was due to the change in membrane permeability [29]. After entering the cytoplasm, Ag NPs induce reactive oxygen species (ROS) production and by binding the phosphate group of effector molecules disturbing the protein synthesis and thus, causes bacterial growth inhibition or killing [30]. Due to the high prevalence, antibiotic resistance and pathogenicity of \u003cem\u003eS. aureus\u003c/em\u003e and \u003cem\u003eE. coli\u003c/em\u003e we studied them. These two pathogens are the causative agents for several infections, such as endocarditis, urinary tract infection, osteomyelitis, and septicemia [31, 32].\u003c/p\u003e \u003cp\u003eMetal NPs especially once having a relatively large size/surface ratio or smaller than 20 nm act as destroyers of the cell membrane through binding to cells, causing structural alterations and eventually the loss of the semi-permeability of the membrane [6]. Studies have shown that bacterial cells membrane disruption by Psidiumguajava leaf extracts is in agreement with the result of this study [33].\u003c/p\u003e \u003cp\u003eOverall, the synthesized NPs exhibited pronounced antibacterial activities on S. aureus and on E. coli. Similar to these findings, Janaki et al showed that zinc oxide nanoparticle (ZnO NPs) which was synthesized using ginger extracted root has an efficient antimicrobial activity [34]. In the other study, Kumor and co-workers utilized ginger extract green synthesized gold NPs and evaluated of blood compatibility of them. The result showed that biosynthesized NPs are suitable vectors for medical applications [13]. These findings explain the possible antibacterial actions of green synthesized NPs: (i) may be due to DNA damage, (ii) protein synthesis inhibition and denaturation, and (iii) formation of free radicals causing cell wall damage [35] (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"6. Conclusion","content":"\u003cp\u003eThe present study reports the chemical and green synthesis of Au and AgNPs using citric acid and aqueous root extract of ginger, respectively. Biosynthetic NPs were characterized that are spherical and hexagonal shapes with an average size\u0026thinsp;\u0026le;\u0026thinsp;100 nm. The biosynthesized Au and AgNPs showed an excellent antibacterial effect against \u003cem\u003eS. aureus\u003c/em\u003e and \u003cem\u003eE. coli\u003c/em\u003e in comparing chemical synthesized NPs. Further, the possible anti-bacterial mechanisms were studied by protein leakage. The Result of this study proves that biosynthesized Au and Ag NPs could be used as an alternative therapy to control and eliminate the infection caused by E. coli and S. aureus.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eFTIR, Fourier transforms infrared spectroscopy; MIC, minimum inhibitory concentration; NPs nanoparticles; ROS, reactive oxygen species; TEM, Transmission electron microscopy\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cbr\u003e\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors appreciate the personnel of Drug Applied Research Center for help and guide\u003c/p\u003e\n\u003ch3\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/h3\u003e\n\u003cp\u003eThe data that support the findings of this study are available from the corresponding authors upon reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eContributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMY was a significant contributor to doing and writing the manuscript. MA, HDM and MM collaborated in doing the thesis that results in the paper. AA and MMD designed and supervised the manuscript. All authors read and approved the final manuscript.\u003c/p\u003e\n\u003ch3\u003e\u003cstrong\u003eEthics declarations\u003c/strong\u003e\u003c/h3\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis work was financially supported by Tabriz University of Medical Sciences (Morteza Yadi M.Sc. Thesis NO: 96/2-3/16).\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003eDavey ME, O'toole GA. Microbial biofilms: from ecology to molecular genetics. Microbiology and molecular biology reviews. 2000;64(4):847\u0026thinsp;\u0026minus;\u0026thinsp;67.\u003c/span\u003e\u003c/li\u003e \u003cli\u003eO'Toole GA, Gibbs KA, Hager PW, Phibbs PV, Kolter R. The global carbon metabolism regulator Crc is a component of a signal transduction pathway required for biofilm development by Pseudomonas aeruginosa. Journal of Bacteriology. 2000;182(2):425\u0026thinsp;\u0026minus;\u0026thinsp;31.\u003c/span\u003e\u003c/li\u003e \u003cli\u003eR\u0026oacute;nav\u0026aacute;ri A, Igaz N, Adamecz DI, Szerencs\u0026eacute;s B, Molnar C, K\u0026oacute;nya Z, et al. Green Silver and Gold Nanoparticles: Biological Synthesis Approaches and Potentials for Biomedical Applications. Molecules. 2021;26(4):844.\u003c/span\u003e\u003c/li\u003e \u003cli\u003eWu X, Lu Y, Zhou S, Chen L, Xu B. Impact of climate change on human infectious diseases: Empirical evidence and human adaptation. Environment international. 2016;86:14\u0026ndash;23.\u003c/span\u003e\u003c/li\u003e \u003cli\u003eAhmed S, Ahmad M, Swami BL, Ikram S. A review on plants extract mediated synthesis of silver nanoparticles for antimicrobial applications: a green expertise. Journal of advanced research. 2016;7(1):17\u0026ndash;28.\u003c/span\u003e\u003c/li\u003e \u003cli\u003eSalleh A, Naomi R, Utami ND, Mohammad AW, Mahmoudi E, Mustafa N, et al. The potential of silver nanoparticles for antiviral and antibacterial applications: a mechanism of action. Nanomaterials. 2020;10(8):1566.\u003c/span\u003e\u003c/li\u003e \u003cli\u003eChitsazi MR, Korbekandi H, Asghari G, Bahri Najafi R, Badii A, Iravani S. Synthesis of silver nanoparticles using methanol and dichloromethane extracts of Pulicaria gnaphalodes (Vent.) Boiss. aerial parts. 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Korean J Microbiol Biotechnol. 2011;39(1):77\u0026ndash;85.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"green synthesis, antibacterial agent, ginger extract, metallic nanoparticle","lastPublishedDoi":"10.21203/rs.3.rs-1352913/v2","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-1352913/v2","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground\u003c/strong\u003e\u003c/p\u003e\u003cp\u003eRecent studies demonstrated that the speed of synthesis, biocompatibility, and antimicrobial activity of gold (Au) and silver (Ag) metals is enhanced when biosynthesized in nano-sized particles.\u0026nbsp;In the present study, Au NPs and Ag NPs were synthesized \u003cem\u003evia\u003c/em\u003e a biological process using aqueous Ginger root extract and characterized by various spectroscopic methods. \u003c/p\u003e\u003cp\u003e\u003cstrong\u003eMethods \u0026amp; Results\u003c/strong\u003e\u003c/p\u003e\u003cp\u003eThe NPs were found to be in hexagonal and spherical shapes. The average particle size for Au and Ag NPs was found to be 20 nm and 15 nm, respectively. The dynamic light scattering (DLS) method has shown that the zeta potential values of synthesized NPs were found to be 5.7 mv and 7.11mv, respectively. Gas chromatography-mass spectrometry (GC-MS) analysis of Ginger root extract revealed 25 compounds. The synthesized NPs showed significant activity against \u003cem\u003eStaphylococcus aureus \u003c/em\u003eand \u003cem\u003eEscherichia coli\u003c/em\u003e in vitro with IC50 and IC90 values for Au and Ag NPs, respectively, noted to be 7.5 and 7.3 µg/ml and 15 and 15.2 µg/ml for both bacterial strains. The protein leakage level was high and morphological changes occurred in bacteria treated with biosynthesized NPs. \u003c/p\u003e\u003cp\u003e\u003cstrong\u003eConclusion\u003c/strong\u003e\u003c/p\u003e\u003cp\u003eThese results suggest that the biosynthesized metallic NPs show potential for application as antibacterial agents with enhanced activities. \u003c/p\u003e","manuscriptTitle":"Antibacterial activity of green gold and silver nanoparticles using ginger root extract","msid":"","msnumber":"","nonDraftVersions":[{"code":2,"date":"2022-03-09 19:04:06","doi":"10.21203/rs.3.rs-1352913/v2","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}},{"code":1,"date":"2022-02-16 17:46:19","doi":"10.21203/rs.3.rs-1352913/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"05f3152d-dd30-4211-b582-3dbd81fb5020","owner":[],"postedDate":"March 9th, 2022","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2022-03-28T18:29:19+00:00","versionOfRecord":[],"versionCreatedAt":"2022-03-09 19:04:06","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v2","identity":"rs-1352913","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-1352913","identity":"rs-1352913","version":["v2"]},"buildId":"rHA-KDH7Qsr4HCuvH75dn","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}