Green synthesis and Characterization of Acacia nilotica derived silver nanoparticles and their insecticidal potential against three important mosquito vectors

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Bruntha, S Lokeshwaran, SP Subala This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5353202/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract In the present study, the bark extract of Acacia nilotica plant was used to green synthesis and Characterization of Silver nanoparticles and check their insecticidal potential against three important mosquito vectors such as Aedes aegyptii, Anopheles stephensi and Culex quinquefasciatus . Our research aims to investigate the therapeutic potential of plant extract combined with nanoparticles. The synthesized silver nanoparticles were characterized by UV-Visible spectroscopy (UV-Vis), Fourier transform infrared Spectroscopy (FT-IR), and scanning electron microscopy (SEM) with EDax techniques (EDAX) and X-ray diffraction (XRD). UV-Vis confirms with the peak at 420 nm showed the synthesis of silver nanoparticles. SEM confirms the spherical shape of nanoparticles and EDAX revealed chemical components of Ag, N, and Oxygen with a strong elemental signal in association with distinct peak for Ag was confirmed at 3.15keV. XRD was determined the crystalline properties of the silver nanoparticles (AgNP). The antioxidant potential of Acacia nilotica bark extract using AgNP was determined by DPPH, hydroxyl and ABTS free radical scavenging assays. DPPH and hydroxyl activities of AgNP showed the highest inhibition of 63% and 84%, while ABTS analysis showed 58%. Antibacterial activity shows highest level of activity against S. pyrogens, P. aeruginosa and P.vulgaris . Larvicidal potential of AgNPs recorded good mortality rate against Aedes aegypti ., Anopheles stephensi., Culex quinquefasciatus . This study confirms the maximum activity of AgNPs and acts as larvicidal, adulticidal, antimicrobial, and antioxidant agents. Acacia nilotica Antimicrobial Antioxidant Smoke toxicity Larvicidal Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 1. Introduction Mosquitoes (Dipterans: Culicidae) are a major menace to millions of people worldwide because they can transmit diseases such Japanese encephalitis, yellow fever, dengue, zika, filariasis, and dengue (Jang et al., 2016). In addition, the Culicidae family disseminates significant parasites and illnesses, like dog heart worm, West Nile virus, and Eastern equine encephalitis, to which horses and dogs are especially susceptible (Mehlhorn, 2015). Unfortunately, there is no recognized treatment for most arboviruses that are carried by mosquitoes. Historically, Aedes aegypti has been the main vector in nearly all significant human epidemics caused by these four viruses. Geographically diverse disease prevalence is caused by differences in Aedes agypti competency within local populations. Anopheles stephensi was potentially linked to malaria epidemics, according to earlier reports (Faulde et al., 2014; de Santi, 2019). The main vector of transmission is the mosquito Culex quinquefasciatus . Elephantasis and irritation of lymphatic channels are the initial effects of the adult nematodes' obstruction of the lymphatic system's flow. Filariasis is found in China, Japan, Sri Lanka, and several Pacific islands, yet it is more prevalent in Africa and India. Chemical synthetic pesticides including methoprene, diflubenzuron, organophosphates, and bacterial larvicides are effective ways to control mosquitoes (Jesser et al., 2017). However, the regular use of chemical pesticides has had an influence on environmental problems (Ga'al, et al., 2018). An alternative approach to mosquito control is needed to address the issue. As a result, green synthesis-produced metal oxide nanoparticles are useful in lowering mosquito populations. There is a connection between cancer and mosquito-transmitted diseases because these infections alter human metabolic pathways, which can result in cancer. In India, An. stephensi is susceptible to DDT in Karnataka but resistant to it in Delhi, Gujarat, Rajasthan, Kerala, and Madhya Pradesh. Together with preventive chemotherapy and morbidity management, vector control in certain situations was a contributing factor. In contrast, it has been demonstrated that biologically produced nanoparticles have potent antimalarial activity against a variety of mosquito vectors (Chitra et al., 2015; Das et al., 2017). The development of nanomedicines holds promise for novel therapeutic approaches to various diseases spread by mosquitoes, as well as enhanced product and drug bioavailability and efficacy. Additionally, controlled release formulations requiring the right dosages and less side effects are anticipated. When compared to biological creatures like fungi, algae, and bacteria, plant extract-derived nanoparticles are far more readily available, safe, and generally nontoxic, requiring fewer steps during the downstream processing stage (Ali et al., 2016 a, c, d; Ali et al., 2017b; Pavela et al., 2017; Salari et al., 2017; Ali et al., 2018a). The tropical and subtropical plant Acacia nilcota (L.), generally known as "babul," is a significant decorative and medicinal plant. It produces a variety of active secondary metabolites that could be used as candidates for drugs in the near future, with the highest chance of success. Acacia nilotica contains volatile essential oils, terpenes, alkaloids, phenols, and phenolic glycosides as complex phytoconstituents. These specific phytoconstituents are responsible for Acacia nilotica 's therapeutic qualities. It is high in nutrients and has a great therapeutic value. Previous traditional descriptions have demonstrated its efficacy in treating and preventing a range of infectious disorders and degenerative conditions (Sadiq et al., 2015). An animal model study also highlighted the significant role that Acacia nilotica and its primary phytoconstituents play in the treatment of infections, inflammations, diabetes, cancer, and hypertension. It is thought to be a risk-free medicinal herb that alters a range of therapeutic benefits. The bark, gum, leaves, and pods of the tree are also widely used medicinally in Africa to treat gastrointestinal disorders, diarrhea, typhoid, cancer, colds, coughs, fever, convalescence, bleeding, and nerve stimulants (Duke, 1983). Acacia nilotica leaves and seeds have been found to have a range of multifunctional active ingredients (Pande et al. , 1981; Chaubal et al. , 2005). Consequently, fruit and leaf extracts from Acacia nilotica have demonstrated potential as molluscicides, bactericides, insecticides, and fungicides (Umalkar et al., 1976; Fagg and Greaves, 1990; Chaubal et al., 2005). According to traditional Unani medicine (Khan A, Muhit-i-A'zam, 2012), Acacia nilotica is a helpful medication for Zyabetus (diabetes), and this has been scientifically demonstrated in a number of in vivo, in vitro, and clinical investigations. There is a wide range of metabolites found in plants that can help in metal ion production, binding, and reduction (Kovendan et al. , 2012). While several nanomaterials such as AgNPs have been developed with varying applications, including but not limited to copper, zinc, titanium, magnesium, gold, and silver (Ali et al. , 2015; Ali et al. , 2016 b, e, f; Ali et al. , 2017a) AgNPs have demonstrated superior antibacterial activity against a wide range of pathogenic eukaryotic microorganisms, including viruses and bacteria. The synthesis of AgNPs using a variety of medicinal plants, including Azadirachta indica, Glycine max, Cinnamon zeylanicum, Pongamia pinnata, Annona squamosa , and Murraya koenigii , has been the subject of very few published reports (Sathishkumar et al., 2009; Tripathi et al., 2009; Vivekanandhan et al., 2009; Rajesh et al., 2010; Kumar et al., 2011; Suganya et al., 2013). Nevertheless, there is no information available regarding the larvicidal effects of silver nanoparticles made using bark extract from Acacia nilotica on mosquito species. 2. Materials and Methods 2.1. Preparation of bark extract The Acacia nilcotica was procured from Kolli Hills in the Salem District of Tamil Nadu, India. It is situated in the Eastern Ghats at an elevation of 1300 meters, between 10° 12′–11° 7′ N and 76°–77° 56′ E. Dr. D. Natarajan, an assistant professor in the department of biotechnology at Periyar University in Tamil Nadu, India, made the taxonomic identification. The plant's bark is cleaned with tap water to get rid of any undesired solid dust particles. It is then shade-dried at room temperature for about 15 days to make a powder that is then stored in the refrigerator for later use. A solution of 2 grams of bark powder and 20 milliliters of distilled water was placed in a heating mantle and heated to 90℃ for 30 min at temperature. After that, filter paper was used to filter the aqueous extract. The desired aqueous solution was obtained, and the filtrate was utilized to create the silver nanoparticles. 2.1. Biosynthesis of Silver Nanoparticles Silver nanoparticle synthesis was carried out using Kumar et al., 2017a. In order to create silver nanoparticles, 10 ml of plant extract were added to 90 ml of 1 millimeter AgNO 3 solution in conical flasks. For 10 minutes, the mixture was heated to 50 ± 2°C while being constantly stirred. The addition of aqueous bark extract converted the silver ions to AgNP in few minutes, as seen by the solution's gradual shift in color. The brown color transition that has been identified denotes the formation of silver nanoparticles. After centrifuging the mixture for one hour at 15,000 rpm, the AgNP that was produced was air-dried, cleaned again with deionized water, and then kept in the incubator. AgNP, the collected powder, was kept in the refrigerator for further characterizations. 2.2. Ultraviolet-Visible (UV-Vis) spectroscopy In order to quantify the AgNP synthesis and determine its optical characteristics, UV-Vis spectroscopy was used to confirm AgNP formation and calculate the high absorption peak caused by surface plasmon excitation. The UV-Vis 1800 spectrophotometer (Shimadzu, Japan) was used to measure the UV-Vis spectrum of the biosynthesized AgNP. At normal temperature, the device was run with a 1 nm resolution in the 200 and 800 nm ranges. 2.3. FTIR analysis The functional groups of brown synthesised AgNP were investigated by FTIR analysis (JASCO FT-IR 4100 spectrometer, Hachioji, Tokoyo, Japan). A disk was filled with roughly 0.2g of AgNP under high pressure. At a resolution of 4.0, the FT-IR spectra were scanned between 500 and 4000 cm − 1 . 2.4. Scanning electron microscopy with energy dispersive X-ray analysis The elemental structures of brown synthesised AgNP were determined using a SEM (JEOL JSM-6360LA, Tokyo, Japan), which was coupled to an energy dispersive spectroscope (EDX, Tokyo, Japan) to identify the surface characteristics and elemental compositions of various NPs (Shaheen and Fouda, 2018). The quantitative elemental structure of brown-synthesised AgNP was examined using EDX analysis. Data investigation revealed the weight percentages of chemical constituents in the artificial AgNP. The effective synthesis of AgNP employing filtrate metabolites is confirmed by the EDX result. 2.5. XRD analysis The X-ray diffraction (XRD) spectrum was used to assess the purity and formation of the compounds. The mixture was centrifuged for 10 minutes at 10,000 rpm in a cooled centrifuge, and the particle was then re-dispersed in acetone. The pellets were spread and dried for a week at 37°C in an incubator. The size of pure Ag nanoparticles was determined in a scan range of 10 to 80 degrees, 2 theta, in flat plate geometry with Ag radiation, and at a temperature of 25 oC using Ag-Kβ radiation (λ = 1.39222nm). The high intensity peak measurement was calculated using Debye-Schreure's equation (D = Kλ/β Cosθ). 2.6. In-vitro Antioxidant activity 2.6.1. DPPH assay Kirubakaran et al. (2024) described the DPPH study, which was detailed by AgNP. Ag-NP was held in sterile test containers at different concentrations (120, 160, 200, 240, and 300 µg/ml). 1.0 mL of 0.2 M DPPH was added to each test tube, and the mixture was then incubated for 30 min until the purple color turned yellow. The absorbance at 517 nm was then measured using UV-Vis spectrophotometry, with vitamin C serving as a standard for comparison. 2.6.2. Hydroxyl scavenging assay According to Rajeshwar et al. (2005), it was performed. Regarding this, 3 mL of hydrogen peroxide solution (1.0 mL of 1.5 mM FeSO 4 , 0.7 mL of 6 mM hydrogen peroxide, and 0.3 mL of 20 mM sodium salicylate) were combined with 1 mL of various AgNP concentrations (120, 160, 200, 240, and 300 µg/ml). The absorbance at 562 nm was measured after the reaction mixture was incubated for 37 ◦C. A 0 – Control; A 1 – absorbance of AgNP and A 2 - absorbance without sodium salicylate. 2.6.3. ABTS assay This assay was followed by (Giao et al. , 2007). Various concentrations (120, 160, 200, 240, and 300 µg/ml) of Ag-NP preparations were collected in clean test containers. In test tubes 1 mL of a 7 mM ABTS solution is added, followed by potassium persulfate 2.45 mM. Mixture was thoroughly mixed and all kept to incubate for 10 min followed by the absorbance were measured by using UV-Vis spectrophotometry at 734 nm with Vitamin C as standard for comparison. 2.7. Antimicrobial activity by disc diffusion method Disc diffusion method was carried out to check the antimicrobial activity of Acacia nilcotica derived AgNP. Different strains of bacteria. [ Escherichia coli ( ATCC 25922), Staphylococcus aureus ( ATCC 25923 ), Streptococcus aureus ( ATCC 700294 ) , Pseudomonas aeruginosa (ATCC 15442), Proteus vulgaris (ATCC 6380)] obtained from Department of Microbiology, ANJA college, Sivakasi), Active bacterial cultures were revived by inoculating a loop full of bacterial culture in nutrient broth from the stock maintained at 4°C and incubated overnight at 37°C in a shaker incubator at 800 rpm. Nutrient agar plates were prepared and spreading of 60 µl of each bacterial culture was carried out. Different concentrations (120, 160, 200, 240, and 300 µg/ml) of AgNP infused discs were prepared and placed over the bacterial spread plates followed by incubation overnight at 37°C. The observed zone of inhibition was measured in mm. 2.8. Antimicrobial activity by Minimal Inhibitory Concentration (MIC) To evaluate the minimal inhibitory concentration, different concentrations (120, 160, 200, 240, and 300 µg/ml) of AgNP extract were tested against [ Escherichia coli ( ATCC 25922), Staphylococcus aureus ( ATCC 25923 ), Streptococcus aureus ( ATCC 700294 ) , Pseudomonas aeruginosa (ATCC 15442), Proteus vulgaris (ATCC 6380)] For this experiment, 25 ml of nutrient broth was added to four different conical flasks containing different concentrations of the AgNP extract mentioned above followed by the addition of 100 µl of bacterial culture. After the addition of bacterial culture, OD was measured at 600 nm in every 2 h interval of time from 0 to 12 h followed by incubation at 37°C at 800 rpm. 2.9. Maintenance mosquito culture Aedes aegypti, Anopheles stephensi and Culex quinquefasciatus mosquito larvae were purchased from the Indian Council for Medical Research (ICMR), Madurai, India. The larvae were kept in plastic trays containing tap water and were maintained in the laboratory, and all the experiments were carried out at 27 ± 2°C and 75–85% relative humidity under 14:10 light and dark photoperiod cycles. Larvae were fed with dog biscuit and yeast powder in the ratio of 3:1. They were maintained and reared in the laboratory. 2.10. Larvicidal activity World Health Organization (2006) larval bioassay method was followed with some slight modifications. Culex quinquefasciatus, Anopheles stephensi , and Aedes aegypti larvae were released in a compartment tray with deionized water. After that, different concentrations of various AgNP were tested with mosquito populations. Bioassays were performed separately at five different concentrations of AgNP (120, 160, 200, 240, and 300 µg/ml). Controls were not received any test concentration (water only). Three replicates were kept for each test and 25 larvae were released in each concentration. The larval mortality was observed after 24h and the percentage of mortality were reported from the average of three replicates. The lethal concentration (LC50 and LC90) were calculated by probit analysis. 2.11. Adulticidal activity Adulticidal bioassay was performed by WHO method (1981) (Brogdon and McAllister 1998; CDC, 2004; Ramkumar et al. , 2014) using a clean glass test tube, and five different concentrations of AgNP (120, 160, 200, 240, and 300 µg/ml) were used. There were three replicates per concentration, and 20 adult mosquitoes were released in each replicates. The tubes were tightly covered with a net cloth and coated with a water alone that served as a control. The mortality of mosquitoes was determined at the end of a 24-h recovery period. LC50 and LC90 with their 95% confidence limits were determined using Log Probit analysis test (Finney, 1971). 2.12. Mosquito coil preparation Mosquito coils were prepared followed by the methods of Ramkumar et al. (2014) and Saini et al . (1986) with suitable modification. The mosquito coils were prepared by using shade dried 4g of plant crude extract and 2 g of sawdust and 2 g of coconut shell as a burning materials. All the materials were mixed thoroughly in distilled water to form a semi-solid paste, and the paste was prepared as mosquito coils with a thickness 0.5 cm. Coils were shade dried and used for further experiment. 2.13. Smoke toxicity test Smoke toxicity experiment was conducted in a glass chamber measuring 60×40×35 cm with mid bottom of the chamber. Fifty blood-fed adult mosquitoes were released to the chamber, and the mosquitoes were exposed to the smoke of burning coils for 40 min, and the mortality data were recorded after every 10 min. The smoke toxicity was compared with the commercially available mosquito coil as tested above (Singha et al . 2011). The commercial mosquito coil active ingredients contain extract allethrin as an active ingredient. 2.14. Statistical analysis Antioxidant assays were calculated by one way ANOVA by using Prism 8.0 software. Graph Pad Software was used to analyze Mean ± SD and one-way ANOVA Asterisk (***) indicates significant difference among treatments with respect compared to control (P < 0.001***). Average larval mortality data were subjected to probit analysis for calculating LC50, LC90 and other statistics at 95% confidence limits of upper confidence limit (UCL) and lower confidence limit (LCL) values, and chi-square test was calculated using the SPSS13.0 (Statistical Package of Social Sciences) software version 13.0. 3. Results 3.1. Silver nanoparticles of Acacia nilotica The primary basis of visual validation of silver nanoparticles was confirmed by seeing the complete reaction through colour changing pattern of silver nitrate (AgNO 3 ) and extraction of Acacia nilotica. After 24hrs of incubation in a dark room and heated in a hot plate at 80 ° C, colour change was observed from Black to reddish brown. This occurred due to reduction of silver nitrate present in this solution (Fig. 1 ). 3.2. UV-Visible spectrophotometer The inclusion of a reducing, capping, and stabilizing agent of silver nitrate in the solution combined with extracts of Acacia nilotica led to the synergy of silver nanoparticles. UV-Visible spectrophotometer has extended wavelength from 200nm to 800nm which was observed in bark extract of Acacia nilotica was mentioned in the Fig. 2 . UV-Visible examined broad peak at (427.00 nm) was occupied in the Acacia nilotica derived silver nanoparticles (Fig. 3 ). 3.3. FTIR analysis The numerous functional groups and their existence in the AgNP of Acacia nilotica was revealed by the FTIR spectrum examination. The FTIR light source absorption conveys the regular frequency of the vibration of various functional chemical groups or chemical bonds. Atomic mass shows an important part in confirming energy which is absorbed the peak of several noticeable chemical compounds or molecules in FTIR spectrum. The FTIR spectrum of the AgNP showed broad and strong absorbance peaks at 3901.73, 2932.56, 1747.39, and 427.20 cm − 1 which might be corresponding to functional groups (Fig. 4 ). A peak in absorption at 3444.63 cm − 1 correlates to O-H stretch, H-bonded of alcohols, phenols while peak value, 2932.56 cm − 1 examined C-H stretching, O-H stretching alkanes, carboxylic acids. A peak values 2873.74 cm − 1 can be credited with C-H stretching of alkanes. Absorbance peak values of 1747.39 cm − 1 can be credited with C = O stretching of esters, saturated aliphatic, carboxylic acids, carbonyls(general) Absorbance peak at 1619.13 cm − 1 examined N-H bend of Primary amines A peak in absorbance at 1513.05 cm − 1 can be N-O asymmetric stretching at nitro compounds Absorbance peak at 1451.33 cm − 1 C-H bend C-C Stretching (in-ring) of alkanes aromatics. A peak values 1317.29 cm − 1 C-O Stretching C-N Stretching N-O Symmetric stretching of alcohols, carboxylic acids, esters, ethers aromatic amines nitro compounds. Absorbance Peak at 894.91 cm − 1 , 817.7cm − 1 , 779.19cm − 1 , 623.93cm − 1 , 515.92 cm − 1 examined C-H “oop” N-H Wag = C-H, C-Cl Stretching, -C(triple bond)C-H C-H bend, C-Br Stretching bend represents aromatics primary, secondary amines alkenes, Alkyl halides, primary, secondary amines, alkenes, alkylhalides, alkynes, alkyl halides (Table 1 ). Table 1 FTIR spectrum analysis of functional groups present in AgNPusing A.nilotica bark extract. S.No Frequency absorption (cm − 1) Appearance Functional Group Compound Class 1. 427.2 ------- ------- ------- 2. 452.28 ------- ------- ------- 3. 515.92 Medium C-Br Stretch alkyl halides 4. 623.93 broad, sharp narrow -C(triple bond) C-H C-H bend alkynes alkynes 5. 779.19 Medium C-Cl Stretch alkyl halides 6. 817.76 Medium strong, broad strong C-Cl Stretch N-H wag C = H bend alkyl halides primary, secondary amines, alkenes 7. 894.91 Strong strong, broad strong C-H “oop” N-H Wag =C-H bend aromatics primary, secondary amines, alkenes 8. 1028.95 Medium Strong C-N Stretch C-O Stretch aliphatic amines alcohols, carboxylic acids, esters, ethers 9. 1111.89 Strong C-O Stretch alcohols,carboxylic acids, esters, ethers 10. 1161.07 Medium medium strong medium C-H wag (-CH2X) C-H wag (-CH2X) C-O Stretch C-N Stretch alkyl halides alkyl halides Alcohols, carboxylic acids, esters, aliphatic amines. 11. 1238.21 Medium Medium medium strong C-N Stretch C-H wag(-CH2X) C-H wag(-CH2X) C-O stretch aliphatic amines alkyl halides alkyl halides alcohols, carboxylic acids, esters, ethers 12. 1317.29 Strong strong medium C-O Stretch C-N Stretch N-O Symmetric stretch alcohols, carboxylic acids, esters, ethers aromatic amines nitro compounds 13. 1374.19 ------- ------- ------- 14. 1451.33 Medium Medium C-H bend C-C Stretch (in-ring) alkanes aromatics 15. 1513.05 Strong N-O asymmetric stretch nitro compounds 16. 1619.13 Medium N-H bend Primary amines 17. 1747.39 Strong Strong Strong C = O stretch C = O stretch C = O stretch esters, saturated aliphatic carboxylic acids, carbonyls (general) 18. 2321.17 ------- ------- ------- 19. 2380.96 ------- ------- ------- 20. 2873.74 Strong C-H stretch alkanes 21. 2932.56 Medium Medium C-H stretch O-H stretch alkanes carboxylic acids 22. 3444.63 Medium O-H stretch, H-bonded alcohols, phenols 23. 3722.36 ------- ------- ------- 24. 3769.61 ------- ------- ------- 3.4. SEM with EDAX The morphological pattern of AgNP was authorized by applying of SEM. The shapes of particles are predominantly spherical and aggregate into larger particles with no well-defined morphology. This aggregation may be due to the presence of secondary metabolites in the bark extracts. SEM yielding only inadequate evidence on the actual nature of the shape and size distribution of AgNP of Acacia nilotica extracts. The authentic diameters of the AgNP were measured between Maximum 300 nm and minimum 200 nm in diameter executed SEM characterization. The uniform size of silver suggests that the particles are observed. SEM analysis of AgNP was clearly distinguishable owing to their size difference of synthesized AgNP. Scanning electron microscope images of AgNP using Acacia nilotica bark extract at different magnification like 100x, 200x, 500x (Fig. 5 a-f). EDX bands were clarified to provide qualitative and quantitative signal of the chemical elements present in newly synthesized AgNP. Displaying of particular intensity peak region in electron beam of EDX identified distinct chemical elemental composition of AgNP. To identify the level of NKα1 weight percentage is 4.5%, Okα1 weight percentage is 42.7%, AgLα., AgLβ., AgLβ2 were weight is 52.8%, Atomic percentage is NKα1 9.3%, Atomic percentage is Okα1 76.6.%, AgLα., AgLβ., AgLβ2 Atomic percentage is 14.1%, Error percentage is NKα1 13.0%, Error percentage is Okα1 10.5%, Error percentage is AgLα., AgLβ., AgLβ2 2.4%, K ratio is NKα1 0.0172, K ratio is Okα1 0.0756, K ratio is AgLα., AgLβ., AgLβ2 0.4649. It indicated the higher content of AgNP. But, a strong elemental signal with distinct peak for Ag was confirmed at 3.15keV in EDX analysis (Figure: 6). Moreover, weight percentage, atomic percentage and K ratio was mentioned in Table 2 . Table 2 Smart Quant Results Element Weight % Atomic % Error % K ratio N K 4.5 9.3 13.0 0.0172 O K 42.7 76.6 10.5 0.0756 Ag L 52.8 14.1 2.4 0.4649 Table 3 Larvicidal activity of Acacia nilotica bark extracts against fourth instar larvae of Aedesagypti., Anopheles stephensi., Culexquiquefasciatus. Mosquito species Sample n a LC 50 (mg/ml) LC 90 (mg/ml) X 2 df (95%CL) (95%CL) Aedes Agypti AgNP Plant extract 60 60 240.714 (235.18–241.61) 313.116 (309.28–316.82) 367.006 (361.1–370.01) 417.820 (414.16–421.8) 0.579 0.703 3 3 Anopheles stephensi AgNP Plant extract 60 60 104.13 (99.811–106.161) 296.73 (291.11–298.12) 158.13 (151.03–160.82) 450.08 (441.18–453.16) 0.426 0.499 3 3 Culex Quinquefasilitus AgNP Plant extract 60 60 99.602 (95.116–103.82) 293.126 (289.72–298.11) 153.59 (148.26–159.12) 451.754 (443.16–455.16) 0.453 0.597 3 3 n a -total number of mosquitoes larvae used; n = 20/replicate; LC50-Lethal Concentration 50% mortality, LC90-Lethal concentration 90% mortality, LCL-Lower confidence limits, UCL-Upper confidence limits, X 2 -chi-square, df- degrees of freedom (Note: Chi square values with a static are significant P < 0.05). Table 4 Adulticidal activity of Acacia nilotica bark extract species against Aedesagypti.,Anophlesstephensi., Culexquinquefasciatus. Adult Mosquito Species Sample n a LC 50 (mg/ml) LC 90 (mg/ml) X 2 df Aedes agypti AgNP Plant extract 60 60 13.560 (9.4–17.3) 14.314 (11.31–16.41) 24.35 (20.47–34.43) 257.76 (218.61–329.62) 2.113 1.421 3 3 Anopheles Stephensi AgNP Plant extract 60 60 16.854 (13.06–19.011) 17.269 (15.09–23.259) 28.411 (24.24–39.2) 31.130 (27.620–33.03) 1.039 1.642 3 3 Qulex Quinquefasciatus AgNP Plant extract 60 60 17.012 (14.458–20.05) 18.312 (15.458–21.05) 31.230 (27.18–34.011) 33.204 (29.18–36.016) 1.267 1.675 3 3 * indicates significantly difference (p < 0.01) between different solvent extracts. LC50—lethal concentration that kills 50% of the exposed larvae, LC90—lethal concentration that kills 90% of the exposed larvae, UCL—upper confidence limit (95%), LCL—lower confidence limit (95%), df—degrees of freedom. Table: 5 Smoke toxicity test of A. nilotica plant powder compared with commercial mosquito coils and mosquito coil without any plant powder on major three vectors Table: 5 Smoke toxicity test of A. nilotica plant powder compared with commercial mosquito coils and mosquito coil without any plant powder on major three vectors Mosquito Species Observation (in Minutes) % Mortality from A. nilotica Powder Mosquito Coil % Mortality from CommercialMosquito Coil % Control Mortality Aedes agypti 10 20 30 40 00 02 11 92 00 10 53 100 0 1 1 0 Anopheles stephensi 10 20 30 40 00 08 24 85 00 14 50 100 0 1 1 0 Culex quinquefasciatus 10 20 30 40 00 15 24 83 00 10 46 100 0 1 1 0 3.5. XRD (X-ray Diffraction) The formation and quality of compounds were checked by X-ray diffraction (XRD) spectrum. The size of the purified Silver nanoparticles was analysed by X-ray powder diffraction crystallography (SEIFERT JSO-DEBYEREX-2002, Germany) diffractometer with Cu-Kα1 radiation. A scan rate of 0.020 per second and a scan range between 10–80 2 theta in flat plate geometry with Ag radiation. The crystalline domain size was calculated using D. Scherrer ’s equation (D = kλ/βcosθ) (Hendricks, 1941; Suryanarayana, 1998) where, D = average crystalline domain size, β denotes Full Width Half Maximum (FWHM); K = 0.94, λ = wavelength of X-ray (1.54 A 0 ) and θ is the Brag’s angle). XRD pattern of synthesized to AgNP was mentioned in the Fig. 7 . 3.6. In-vitro antioxidant activity 3.6.1. DPPH (2, 2-diphenyl-1-picrylhydrazyl) Utilizing DPPH, ABTS, and H 2 O 2 tests, the antioxidant capacity of the AgNP-containing Acacia nilotica bark extract was assessed. Percentage of DPPH radical scavenging activity inhibition measured at various AgNP doses (20 µg/mL, 40 µg/mL, 60 µg/mL, 80 µg/mL, and 100 µg/mL). The present investigation examined DPPH activity in a dose-dependent fashion. Similar, dose-dependent action was discovered in AgNPs by the use of bark extracts from Acacia nilotica (Fig. 8 a). 3.6.2. Hydroxyl (H 2 O 2 ) scavenging activity According to the findings of the antioxidant assays, all extracts may have some degree of radical scavenger activity (Fig. 8 b). The variable chemical makeup of different phytochemicals, which may react differently with different kinds of free radicals, may be the cause of the varying scavenging actions of different extracts. 3.6.3. ABTS assay The ABTS assay was used to evaluate the free radical scavenging activity. A widely used colorimetic technique for determining the ability of bark extracts to scavenge radicals is the 1,1-diphenyl-2-picrylhydrazl (DPPH) assay. This technique, which screens the overall activity of antioxidants and is based on the stable synthetic radical DPPH, is precise, simple to use, and reasonably priced. ABTS is a chemically stable and highly water soluble substance. It generates a metastable cation and functions as a substrate for peroxide. Spectrophotometric measurement shows that the synthesis of ABTS is inhibited when an antioxidant molecule is present. The potential of the antioxidant chemical increased with decreasing test solution absorption at 734 nm. In order to reduce interference from sample turbidity and other absorbing materials, the hydrogen donating potential in the antioxidant samples was measured at 734 nm (Fig. 8 c). 3.7. Antibacterial activity using Disc diffusion method Figure 9 showed the results of an antimicrobial activity test using the Disc Diffusion Method against several bacterial strains. Using a ruler or caliper, measure the zones during incubation to the closest millimeter; make careful to account for the disk's diameter in your measurement. Measure from the disk's center to a point on the zone's perimeter where a clear edge is present (the radius), then multiply the result by two to find the diameter if the zone's size or placement prevent you from reading the diameter. Swarming bacterial strains require a different method of measurement than non swarming organisms. Measure the outer boundary in an otherwise obvious zone of inhibition, ignoring the narrow vein of swarming. The small layer of swarming development within an otherwise clear zone of inhibition should be ignored when measuring the zone of inhibition for swarming organisms. Only the categories of susceptible, intermediate, and resistant are reported for the Well Diffusion technique susceptibility test results. AgNP's higher antimicrobial activity was facilitated by the phytonutrients that helped maintain it, and its increased antibacterial effectiveness can be attributed to its surface area and macromolecules. The antibacterial survivability of AgNP may be due to its tiny size, dispersion, and spherical morphology, which provide a broad surface for maximum interaction with bacteria and cause much more damage than bigger particle sizes. AgNP's antibacterial activity was evaluated using the four different bacterial species. Analysis was done on the effects of AgNP as a time feature in fixed amounts of bacterial species' cell morphology (Fig. 10 a–f). 3.7. Antimicrobial activity by using minimum inhibitory conentration To determine the antimicrobial dosage of our AgNP, Minimum Inhibitory Concentration (MIC) was done using different concentrations (120 µg/ml, 160 µg/ml, 200 µg/ml 240 µg/ml and 300 µg/ml) AgNP (Fig. 11 ). Antimicrobial activity test was done with Acacia nilotica derived AgNP Minimum inhibitory concentration using S. aureus (Fig. 12 ). Minimum inhibitory concentration of Acacia nilotica bark extract was and increasing concentration of synthesized ANL-AgNPs in terms of the zone of inhibition against five types of bacterial strains [ Escherichia coli ( ATCC number of 25922), Staphylococcus aureus ( ATCC number of 25923 ), Streptococcus aureus ( ATCC number of 700294 ) , Pseudomonas aeruginosa (ATCC number of 15442), Proteus vulgaris (ATCC number of 6380)]. 3.8. Larvicidal potential of AgNP The larvicidal efficacy and synthesized to AgNP were tested as per the guidelines of world Health organization procedure with some modifications. Different concentration (120µg/ml; 160µg/ml; 200µg/ml; 240µg/ml) AgNP were used, respectively, to test larvicidal activity on Aedes aegypti, Anopheles stephensi, and Culex quinquefasciatus mosquito larvae and a control with test samples. The number of dead larvae was counted after 24 hr. of exposure and their percentage mortality was calculated using Abbott’s formula Mortality (%) where n = number of larvae, T = treated, C = control. This has made it necessary to look for and create mosquito-repelling insecticides that are more affordable, target-specific, and safe for the environment. Botanical derivatives, or natural compounds derived from plants, are a newer and alternate method of controlling mosquitoes. Numerous natural botanical products are affordable, efficient, safe for the environment, and readily biodegradable. They have a variety of unique ways of action and don't harm non-target creatures. Results of bioassays using bark extracts from Acacia nilotica against the larvae of three significant vector mosquito species— Aedes Agypti, Anopheles Stephensi , and Culex Quinquefasciatus —showed the larvicidal efficacy of the extracts (table:3). The plant extracts with LC50 and LC90 values of -0.02, -2.560, -3.961 and.015, 2.332, 1.075 mg/ml, respectively, showed the best larvicidal activity. The degree of freedom (df), chi-square, and 95% confidence limits, LC50 and LC90, were also computed. We did not see any noteworthy mortality in the control assays. 3.9. Adulticidal potential of AgNP Since adulticides may only temporarily lower the adult population, mosquito control programs are primarily focused on eradicating adult mosquitoes at their nesting places with larvicides. Therefore, focusing on the larvae would be a more effective way to lower the mosquito population. Our findings unequivocally show that Acacia nilotica was more successful in suppressing immature phases. Aedes agypti , Anopheles stephensi , and Culex quinquefasciatus are the three major vector mosquito species against which the adulticidal action of Acacia nilotica bark extract has been recorded (table:4). The mortality rate of the bark extract rose in direct proportion to dosage. The bark extract that was examined showed the strongest adulticidal activity against Culex quinquefasciatus , Anopheles stephensi , and Aedes aegypti . When the control group was tested without a solvent, there was no death. The adulticidal activity of Aedes agypti , Anopheles stephensi , and Culex quinquefasciatus was measured by Acacia nilotica bark extracts, and the LC50 and LC90 values were 2.039 and 2.671;603 and 494; 2.930; 6.379 mg/ml, respectively. After 48 hours of PT, the adulticidal mortality reached its peak, and in extracts from the bark of Acacia nilotica , the mortality reached 100% at 1500 PPm. The following are the lethal concentrations: 120, 160, 200, and 240 µg/ml. In both aqueous and bark extracts, the larval death rate for the conventional positive control was 100% at 24 hours and after treatment, respectively. Declarations Funding Declaration There is no financial support for this study. Ethical approval Not applicable. Consent for publication All the authors agreed to publish the data in this journal. Consent to participate Not applicable. CRediT authorship contribution statement R.Bruntha : Writing – review & editing, Writing –original draft, Visualization, Validation, Software, Resources, Methodology, Investigation, Funding acquisition, Formal analysis, Data curation, Conceptualization. S.Lokeshwaran : Visualization, Data curation. Kirubakaaran Dharmaligam: Software, Formal analysis. S.P.Subala: Supervision, Methodology, Investigation, Formal analysis, Data curation, Conceptualization. Declaration of Competing Interest All of the authors are declared there is no conflict of interest. Data Availability The authors do not have permission to share data. Acknowledgment The authors recognize and thank the Department of Biotechnology, Ayya Nadar Janaki Ammal College (autonomous), Sivakasi-626125, Tamil Nadu, India, for providing infrastructural facilities. 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We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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-5353202","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":376892544,"identity":"a6a47a86-ec8c-49ba-bb93-1a5c9fa38523","order_by":0,"name":"R. 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legend\u003c/p\u003e","description":"","filename":"10.png","url":"https://assets-eu.researchsquare.com/files/rs-5353202/v1/910f12a895067e0165f383e3.png"},{"id":70108448,"identity":"16b3c6b6-c687-473b-9e61-2ab9c8749a1c","added_by":"auto","created_at":"2024-11-28 11:58:22","extension":"png","order_by":11,"title":"Figure 11","display":"","copyAsset":false,"role":"figure","size":684589,"visible":true,"origin":"","legend":"\u003cp\u003eSee image above for figure legend\u003c/p\u003e","description":"","filename":"11.png","url":"https://assets-eu.researchsquare.com/files/rs-5353202/v1/a522f1e9ba22cbfac559f9a5.png"},{"id":70108630,"identity":"7d0e5faf-896c-474d-a3da-29b4e1f71271","added_by":"auto","created_at":"2024-11-28 12:06:22","extension":"png","order_by":12,"title":"Figure 12","display":"","copyAsset":false,"role":"figure","size":84143,"visible":true,"origin":"","legend":"\u003cp\u003eSee image above for figure legend\u003c/p\u003e","description":"","filename":"12.png","url":"https://assets-eu.researchsquare.com/files/rs-5353202/v1/9801fd6aa54a101fb3b6be85.png"},{"id":70316179,"identity":"f9d8fcbd-138f-4e9c-b82f-3d03ed16d467","added_by":"auto","created_at":"2024-12-02 05:40:11","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":10024980,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5353202/v1/96797afe-4b40-4dc3-b114-04f0db4ab52d.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Green synthesis and Characterization of Acacia nilotica derived silver nanoparticles and their insecticidal potential against three important mosquito vectors","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eMosquitoes (Dipterans: Culicidae) are a major menace to millions of people worldwide because they can transmit diseases such Japanese encephalitis, yellow fever, dengue, zika, filariasis, and dengue (Jang et al., 2016). In addition, the Culicidae family disseminates significant parasites and illnesses, like dog heart worm, West Nile virus, and Eastern equine encephalitis, to which horses and dogs are especially susceptible (Mehlhorn, 2015). Unfortunately, there is no recognized treatment for most arboviruses that are carried by mosquitoes. Historically, \u003cem\u003eAedes aegypti\u003c/em\u003e has been the main vector in nearly all significant human epidemics caused by these four viruses. Geographically diverse disease prevalence is caused by differences in \u003cem\u003eAedes agypti\u003c/em\u003e competency within local populations. Anopheles stephensi was potentially linked to malaria epidemics, according to earlier reports (Faulde et al., 2014; de Santi, 2019). The main vector of transmission is the mosquito \u003cem\u003eCulex quinquefasciatus\u003c/em\u003e. Elephantasis and irritation of lymphatic channels are the initial effects of the adult nematodes' obstruction of the lymphatic system's flow. Filariasis is found in China, Japan, Sri Lanka, and several Pacific islands, yet it is more prevalent in Africa and India.\u003c/p\u003e \u003cp\u003eChemical synthetic pesticides including methoprene, diflubenzuron, organophosphates, and bacterial larvicides are effective ways to control mosquitoes (Jesser et al., 2017). However, the regular use of chemical pesticides has had an influence on environmental problems (Ga'al, et al., 2018). An alternative approach to mosquito control is needed to address the issue. As a result, green synthesis-produced metal oxide nanoparticles are useful in lowering mosquito populations. There is a connection between cancer and mosquito-transmitted diseases because these infections alter human metabolic pathways, which can result in cancer. In India, \u003cem\u003eAn. stephensi\u003c/em\u003e is susceptible to DDT in Karnataka but resistant to it in Delhi, Gujarat, Rajasthan, Kerala, and Madhya Pradesh.\u003c/p\u003e \u003cp\u003eTogether with preventive chemotherapy and morbidity management, vector control in certain situations was a contributing factor. In contrast, it has been demonstrated that biologically produced nanoparticles have potent antimalarial activity against a variety of mosquito vectors (Chitra et al., 2015; Das et al., 2017). The development of nanomedicines holds promise for novel therapeutic approaches to various diseases spread by mosquitoes, as well as enhanced product and drug bioavailability and efficacy. Additionally, controlled release formulations requiring the right dosages and less side effects are anticipated. When compared to biological creatures like fungi, algae, and bacteria, plant extract-derived nanoparticles are far more readily available, safe, and generally nontoxic, requiring fewer steps during the downstream processing stage (Ali et al., 2016 a, c, d; Ali et al., 2017b; Pavela et al., 2017; Salari et al., 2017; Ali et al., 2018a).\u003c/p\u003e \u003cp\u003eThe tropical and subtropical plant \u003cem\u003eAcacia nilcota\u003c/em\u003e (L.), generally known as \"babul,\" is a significant decorative and medicinal plant. It produces a variety of active secondary metabolites that could be used as candidates for drugs in the near future, with the highest chance of success. \u003cem\u003eAcacia nilotica\u003c/em\u003e contains volatile essential oils, terpenes, alkaloids, phenols, and phenolic glycosides as complex phytoconstituents. These specific phytoconstituents are responsible for \u003cem\u003eAcacia nilotica\u003c/em\u003e's therapeutic qualities. It is high in nutrients and has a great therapeutic value. Previous traditional descriptions have demonstrated its efficacy in treating and preventing a range of infectious disorders and degenerative conditions (Sadiq et al., 2015). An animal model study also highlighted the significant role that \u003cem\u003eAcacia nilotica\u003c/em\u003e and its primary phytoconstituents play in the treatment of infections, inflammations, diabetes, cancer, and hypertension. It is thought to be a risk-free medicinal herb that alters a range of therapeutic benefits. The bark, gum, leaves, and pods of the tree are also widely used medicinally in Africa to treat gastrointestinal disorders, diarrhea, typhoid, cancer, colds, coughs, fever, convalescence, bleeding, and nerve stimulants (Duke, 1983). \u003cem\u003eAcacia nilotica\u003c/em\u003e leaves and seeds have been found to have a range of multifunctional active ingredients (Pande \u003cem\u003eet al.\u003c/em\u003e, 1981; Chaubal \u003cem\u003eet al.\u003c/em\u003e, 2005). Consequently, fruit and leaf extracts from \u003cem\u003eAcacia nilotica\u003c/em\u003e have demonstrated potential as molluscicides, bactericides, insecticides, and fungicides (Umalkar et al., 1976; Fagg and Greaves, 1990; Chaubal et al., 2005). According to traditional Unani medicine (Khan A, Muhit-i-A'zam, 2012), \u003cem\u003eAcacia nilotica\u003c/em\u003e is a helpful medication for Zyabetus (diabetes), and this has been scientifically demonstrated in a number of in vivo, in vitro, and clinical investigations.\u003c/p\u003e \u003cp\u003eThere is a wide range of metabolites found in plants that can help in metal ion production, binding, and reduction (Kovendan \u003cem\u003eet al.\u003c/em\u003e, 2012). While several nanomaterials such as AgNPs have been developed with varying applications, including but not limited to copper, zinc, titanium, magnesium, gold, and silver (Ali \u003cem\u003eet al.\u003c/em\u003e, 2015; Ali \u003cem\u003eet al.\u003c/em\u003e, 2016 b, e, f; Ali \u003cem\u003eet al.\u003c/em\u003e, 2017a) AgNPs have demonstrated superior antibacterial activity against a wide range of pathogenic eukaryotic microorganisms, including viruses and bacteria. The synthesis of AgNPs using a variety of medicinal plants, including \u003cem\u003eAzadirachta indica, Glycine max, Cinnamon zeylanicum, Pongamia pinnata, Annona squamosa\u003c/em\u003e, and \u003cem\u003eMurraya koenigii\u003c/em\u003e, has been the subject of very few published reports (Sathishkumar et al., 2009; Tripathi et al., 2009; Vivekanandhan et al., 2009; Rajesh et al., 2010; Kumar et al., 2011; Suganya et al., 2013). Nevertheless, there is no information available regarding the larvicidal effects of silver nanoparticles made using bark extract from \u003cem\u003eAcacia nilotica\u003c/em\u003e on mosquito species.\u003c/p\u003e"},{"header":"2. Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1. Preparation of bark extract\u003c/h2\u003e \u003cp\u003eThe \u003cem\u003eAcacia nilcotica\u003c/em\u003e was procured from Kolli Hills in the Salem District of Tamil Nadu, India. It is situated in the Eastern Ghats at an elevation of 1300 meters, between 10\u0026deg; 12\u0026prime;\u0026ndash;11\u0026deg; 7\u0026prime; N and 76\u0026deg;\u0026ndash;77\u0026deg; 56\u0026prime; E. Dr. D. Natarajan, an assistant professor in the department of biotechnology at Periyar University in Tamil Nadu, India, made the taxonomic identification. The plant's bark is cleaned with tap water to get rid of any undesired solid dust particles. It is then shade-dried at room temperature for about 15 days to make a powder that is then stored in the refrigerator for later use. A solution of 2 grams of bark powder and 20 milliliters of distilled water was placed in a heating mantle and heated to 90℃ for 30 min at temperature. After that, filter paper was used to filter the aqueous extract. The desired aqueous solution was obtained, and the filtrate was utilized to create the silver nanoparticles.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.1. Biosynthesis of Silver Nanoparticles\u003c/h2\u003e \u003cp\u003eSilver nanoparticle synthesis was carried out using Kumar et al., 2017a. In order to create silver nanoparticles, 10 ml of plant extract were added to 90 ml of 1 millimeter AgNO\u003csub\u003e3\u003c/sub\u003e solution in conical flasks. For 10 minutes, the mixture was heated to 50\u0026thinsp;\u0026plusmn;\u0026thinsp;2\u0026deg;C while being constantly stirred. The addition of aqueous bark extract converted the silver ions to AgNP in few minutes, as seen by the solution's gradual shift in color. The brown color transition that has been identified denotes the formation of silver nanoparticles. After centrifuging the mixture for one hour at 15,000 rpm, the AgNP that was produced was air-dried, cleaned again with deionized water, and then kept in the incubator. AgNP, the collected powder, was kept in the refrigerator for further characterizations.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.2. Ultraviolet-Visible (UV-Vis) spectroscopy\u003c/h2\u003e \u003cp\u003eIn order to quantify the AgNP synthesis and determine its optical characteristics, UV-Vis spectroscopy was used to confirm AgNP formation and calculate the high absorption peak caused by surface plasmon excitation. The UV-Vis 1800 spectrophotometer (Shimadzu, Japan) was used to measure the UV-Vis spectrum of the biosynthesized AgNP. At normal temperature, the device was run with a 1 nm resolution in the 200 and 800 nm ranges.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.3. FTIR analysis\u003c/h2\u003e \u003cp\u003eThe functional groups of brown synthesised AgNP were investigated by FTIR analysis (JASCO FT-IR 4100 spectrometer, Hachioji, Tokoyo, Japan). A disk was filled with roughly 0.2g of AgNP under high pressure. At a resolution of 4.0, the FT-IR spectra were scanned between 500 and 4000 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e2.4. Scanning electron microscopy with energy dispersive X-ray analysis\u003c/h2\u003e \u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eThe elemental structures of brown synthesised AgNP were determined using a SEM (JEOL JSM-6360LA, Tokyo, Japan), which was coupled to an energy dispersive spectroscope (EDX, Tokyo, Japan) to identify the surface characteristics and elemental compositions of various NPs (Shaheen and Fouda, 2018). The quantitative elemental structure of brown-synthesised AgNP was examined using EDX analysis. Data investigation revealed the weight percentages of chemical constituents in the artificial AgNP. The effective synthesis of AgNP employing filtrate metabolites is confirmed by the EDX result.\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e2.5. XRD analysis\u003c/h2\u003e \u003cp\u003eThe X-ray diffraction (XRD) spectrum was used to assess the purity and formation of the compounds. The mixture was centrifuged for 10 minutes at 10,000 rpm in a cooled centrifuge, and the particle was then re-dispersed in acetone. The pellets were spread and dried for a week at 37\u0026deg;C in an incubator. The size of pure Ag nanoparticles was determined in a scan range of 10 to 80 degrees, 2 theta, in flat plate geometry with Ag radiation, and at a temperature of 25 oC using Ag-Kβ radiation (λ\u0026thinsp;=\u0026thinsp;1.39222nm). The high intensity peak measurement was calculated using Debye-Schreure's equation (D\u0026thinsp;=\u0026thinsp;Kλ/β Cosθ).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e2.6. In-vitro Antioxidant activity\u003c/h2\u003e \u003cdiv id=\"Sec10\" class=\"Section3\"\u003e \u003ch2\u003e2.6.1. DPPH assay\u003c/h2\u003e \u003cp\u003eKirubakaran et al. (2024) described the DPPH study, which was detailed by AgNP. Ag-NP was held in sterile test containers at different concentrations (120, 160, 200, 240, and 300 \u0026micro;g/ml). 1.0 mL of 0.2 M DPPH was added to each test tube, and the mixture was then incubated for 30 min until the purple color turned yellow. The absorbance at 517 nm was then measured using UV-Vis spectrophotometry, with vitamin C serving as a standard for comparison.\u003c/p\u003e \u003cp\u003e\u003cimg src=\"https://myfiles.space/user_files/127393_c7e80a1c9bb65875/127393_custom_files/img1732794349.png\"\u003e\u003cbr\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec11\" class=\"Section3\"\u003e \u003ch2\u003e2.6.2. Hydroxyl scavenging assay\u003c/h2\u003e \u003cp\u003eAccording to Rajeshwar et al. (2005), it was performed. Regarding this, 3 mL of hydrogen peroxide solution (1.0 mL of 1.5 mM FeSO\u003csub\u003e4\u003c/sub\u003e, 0.7 mL of 6 mM hydrogen peroxide, and 0.3 mL of 20 mM sodium salicylate) were combined with 1 mL of various AgNP concentrations (120, 160, 200, 240, and 300 \u0026micro;g/ml). The absorbance at 562 nm was measured after the reaction mixture was incubated for 37 ◦C.\u003c/p\u003e \u003cp\u003e\u003cimg src=\"https://myfiles.space/user_files/127393_c7e80a1c9bb65875/127393_custom_files/img1732794426.png\"\u003e\u003cbr\u003e\u003c/p\u003e \u003cp\u003eA\u003csub\u003e0\u003c/sub\u003e \u0026ndash; Control; A\u003csub\u003e1\u003c/sub\u003e \u0026ndash; absorbance of AgNP and A\u003csub\u003e2\u003c/sub\u003e - absorbance without sodium salicylate.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section3\"\u003e \u003ch2\u003e2.6.3. ABTS assay\u003c/h2\u003e \u003cp\u003e\u003cdiv class=\"BlockQuote\"\u003e\u003cp\u003eThis assay was followed by (Giao \u003cem\u003eet al.\u003c/em\u003e, 2007). Various concentrations (120, 160, 200, 240, and 300 \u0026micro;g/ml) of Ag-NP preparations were collected in clean test containers. In test tubes 1 mL of a 7 mM ABTS solution is added, followed by potassium persulfate 2.45 mM. Mixture was thoroughly mixed and all kept to incubate for 10 min followed by the absorbance were measured by using UV-Vis spectrophotometry at 734 nm with Vitamin C as standard for comparison.\u003c/p\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003e\u003cimg src=\"https://myfiles.space/user_files/127393_c7e80a1c9bb65875/127393_custom_files/img1732794506.png\"\u003e\u003cbr\u003e\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003e2.7. Antimicrobial activity by disc diffusion method\u003c/h2\u003e \u003cp\u003eDisc diffusion method was carried out to check the antimicrobial activity of \u003cem\u003eAcacia nilcotica\u003c/em\u003e derived AgNP. Different strains of bacteria. [\u003cem\u003eEscherichia coli (\u003c/em\u003eATCC 25922), \u003cem\u003eStaphylococcus aureus (\u003c/em\u003eATCC 25923\u003cem\u003e), Streptococcus aureus (\u003c/em\u003eATCC 700294\u003cem\u003e)\u003c/em\u003e, \u003cem\u003ePseudomonas aeruginosa\u003c/em\u003e (ATCC 15442), \u003cem\u003eProteus vulgaris\u003c/em\u003e (ATCC 6380)] obtained from Department of Microbiology, ANJA college, Sivakasi), Active bacterial cultures were revived by inoculating a loop full of bacterial culture in nutrient broth from the stock maintained at 4\u0026deg;C and incubated overnight at 37\u0026deg;C in a shaker incubator at 800 rpm. Nutrient agar plates were prepared and spreading of 60 \u0026micro;l of each bacterial culture was carried out. Different concentrations (120, 160, 200, 240, and 300 \u0026micro;g/ml) of AgNP infused discs were prepared and placed over the bacterial spread plates followed by incubation overnight at 37\u0026deg;C. The observed zone of inhibition was measured in mm.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003e2.8. Antimicrobial activity by Minimal Inhibitory Concentration (MIC)\u003c/h2\u003e \u003cp\u003eTo evaluate the minimal inhibitory concentration, different concentrations (120, 160, 200, 240, and 300 \u0026micro;g/ml) of AgNP extract were tested against [\u003cem\u003eEscherichia coli (\u003c/em\u003eATCC 25922), \u003cem\u003eStaphylococcus aureus (\u003c/em\u003eATCC 25923\u003cem\u003e), Streptococcus aureus (\u003c/em\u003eATCC 700294\u003cem\u003e)\u003c/em\u003e, \u003cem\u003ePseudomonas aeruginosa\u003c/em\u003e (ATCC 15442), \u003cem\u003eProteus vulgaris\u003c/em\u003e (ATCC 6380)] For this experiment, 25 ml of nutrient broth was added to four different conical flasks containing different concentrations of the AgNP extract mentioned above followed by the addition of 100 \u0026micro;l of bacterial culture. After the addition of bacterial culture, OD was measured at 600 nm in every 2 h interval of time from 0 to 12 h followed by incubation at 37\u0026deg;C at 800 rpm.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003e2.9. Maintenance mosquito culture\u003c/h2\u003e \u003cp\u003e \u003cem\u003eAedes aegypti, Anopheles stephensi and Culex quinquefasciatus\u003c/em\u003e mosquito larvae were purchased from the Indian Council for Medical Research (ICMR), Madurai, India. The larvae were kept in plastic trays containing tap water and were maintained in the laboratory, and all the experiments were carried out at 27\u0026thinsp;\u0026plusmn;\u0026thinsp;2\u0026deg;C and 75\u0026ndash;85% relative humidity under 14:10 light and dark photoperiod cycles. Larvae were fed with dog biscuit and yeast powder in the ratio of 3:1. They were maintained and reared in the laboratory.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003e2.10. Larvicidal activity\u003c/h2\u003e \u003cp\u003eWorld Health Organization (2006) larval bioassay method was followed with some slight modifications. \u003cem\u003eCulex quinquefasciatus, Anopheles stephensi\u003c/em\u003e, and \u003cem\u003eAedes aegypti\u003c/em\u003e larvae were released in a compartment tray with deionized water. After that, different concentrations of various AgNP were tested with mosquito populations. Bioassays were performed separately at five different concentrations of AgNP (120, 160, 200, 240, and 300 \u0026micro;g/ml). Controls were not received any test concentration (water only). Three replicates were kept for each test and 25 larvae were released in each concentration. The larval mortality was observed after 24h and the percentage of mortality were reported from the average of three replicates. The lethal concentration (LC50 and LC90) were calculated by probit analysis.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003e2.11. Adulticidal activity\u003c/h2\u003e \u003cp\u003eAdulticidal bioassay was performed by WHO method (1981) (Brogdon and McAllister 1998; CDC, 2004; Ramkumar \u003cem\u003eet al.\u003c/em\u003e, 2014) using a clean glass test tube, and five different concentrations of AgNP (120, 160, 200, 240, and 300 \u0026micro;g/ml) were used. There were three replicates per concentration, and 20 adult mosquitoes were released in each replicates. The tubes were tightly covered with a net cloth and coated with a water alone that served as a control. The mortality of mosquitoes was determined at the end of a 24-h recovery period. LC50 and LC90 with their 95% confidence limits were determined using Log Probit analysis test (Finney, 1971).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec18\" class=\"Section2\"\u003e \u003ch2\u003e2.12. Mosquito coil preparation\u003c/h2\u003e \u003cp\u003eMosquito coils were prepared followed by the methods of Ramkumar \u003cem\u003eet al.\u003c/em\u003e (2014) and Saini \u003cem\u003eet al\u003c/em\u003e. (1986) with suitable modification. The mosquito coils were prepared by using shade dried 4g of plant crude extract and 2 g of sawdust and 2 g of coconut shell as a burning materials. All the materials were mixed thoroughly in distilled water to form a semi-solid paste, and the paste was prepared as mosquito coils with a thickness 0.5 cm. Coils were shade dried and used for further experiment.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec19\" class=\"Section2\"\u003e \u003ch2\u003e2.13. Smoke toxicity test\u003c/h2\u003e \u003cp\u003eSmoke toxicity experiment was conducted in a glass chamber measuring 60\u0026times;40\u0026times;35 cm with mid bottom of the chamber. Fifty blood-fed adult mosquitoes were released to the chamber, and the mosquitoes were exposed to the smoke of burning coils for 40 min, and the mortality data were recorded after every 10 min. The smoke toxicity was compared with the commercially available mosquito coil as tested above (Singha \u003cem\u003eet al\u003c/em\u003e. 2011). The commercial mosquito coil active ingredients contain extract allethrin as an active ingredient.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec20\" class=\"Section2\"\u003e \u003ch2\u003e2.14. Statistical analysis\u003c/h2\u003e \u003cp\u003eAntioxidant assays were calculated by one way ANOVA by using Prism 8.0 software. Graph Pad Software was used to analyze Mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD and one-way ANOVA Asterisk (***) indicates significant difference among treatments with respect compared to control (P\u0026thinsp;\u0026lt;\u0026thinsp;0.001***). Average larval mortality data were subjected to probit analysis for calculating LC50, LC90 and other statistics at 95% confidence limits of upper confidence limit (UCL) and lower confidence limit (LCL) values, and chi-square test was calculated using the SPSS13.0 (Statistical Package of Social Sciences) software version 13.0.\u003c/p\u003e \u003c/div\u003e"},{"header":"3. Results","content":"\u003cdiv id=\"Sec22\" class=\"Section2\"\u003e \u003ch2\u003e3.1. \u003cem\u003eSilver nanoparticles of Acacia nilotica\u003c/em\u003e\u003c/h2\u003e \u003cp\u003eThe primary basis of visual validation of silver nanoparticles was confirmed by seeing the complete reaction through colour changing pattern of silver nitrate (AgNO\u003csub\u003e3\u003c/sub\u003e) and extraction of \u003cem\u003eAcacia nilotica.\u003c/em\u003e After 24hrs of incubation in a dark room and heated in a hot plate at 80\u003cb\u003e°\u003c/b\u003eC, colour change was observed from Black to reddish brown. This occurred due to reduction of silver nitrate present in this solution (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec23\" class=\"Section2\"\u003e \u003ch2\u003e3.2. UV-Visible spectrophotometer\u003c/h2\u003e \u003cp\u003eThe inclusion of a reducing, capping, and stabilizing agent of silver nitrate in the solution combined with extracts of \u003cem\u003eAcacia nilotica\u003c/em\u003e led to the synergy of silver nanoparticles. UV-Visible spectrophotometer has extended wavelength from 200nm to 800nm which was observed in bark extract of \u003cem\u003eAcacia nilotica\u003c/em\u003e was mentioned in the Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e2\u003c/span\u003e. UV-Visible examined broad peak at (427.00 nm) was occupied in the \u003cem\u003eAcacia nilotica\u003c/em\u003e derived silver nanoparticles (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec24\" class=\"Section2\"\u003e \u003ch2\u003e\u003cem\u003e3.3. FTIR analysis\u003c/em\u003e\u003c/h2\u003e \u003cp\u003eThe numerous functional groups and their existence in the AgNP of \u003cem\u003eAcacia nilotica\u003c/em\u003e was revealed by the FTIR spectrum examination. The FTIR light source absorption conveys the regular frequency of the vibration of various functional chemical groups or chemical bonds. Atomic mass shows an important part in confirming energy which is absorbed the peak of several noticeable chemical compounds or molecules in FTIR spectrum. The FTIR spectrum of the AgNP showed broad and strong absorbance peaks at 3901.73, 2932.56, 1747.39, and 427.20 cm\u003csup\u003e− 1\u003c/sup\u003e which might be corresponding to functional groups (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e4\u003c/span\u003e). A peak in absorption at 3444.63 cm\u003csup\u003e− 1\u003c/sup\u003e correlates to O-H stretch, H-bonded of alcohols, phenols while peak value, 2932.56 cm\u003csup\u003e− 1\u003c/sup\u003e examined C-H stretching, O-H stretching alkanes, carboxylic acids. A peak values 2873.74 cm\u003csup\u003e− 1\u003c/sup\u003e can be credited with C-H stretching of alkanes. Absorbance peak values of 1747.39 cm\u003csup\u003e− 1\u003c/sup\u003ecan be credited with C = O stretching of esters, saturated aliphatic, carboxylic acids, carbonyls(general) Absorbance peak at 1619.13 cm\u003csup\u003e− 1\u003c/sup\u003e examined N-H bend of Primary amines A peak in absorbance at 1513.05 cm\u003csup\u003e− 1\u003c/sup\u003e can be N-O asymmetric stretching at nitro compounds Absorbance peak at 1451.33 cm\u003csup\u003e− 1\u003c/sup\u003e C-H bend C-C Stretching (in-ring) of alkanes aromatics. A peak values 1317.29 cm\u003csup\u003e− 1\u003c/sup\u003e C-O Stretching C-N Stretching N-O Symmetric stretching of alcohols, carboxylic acids, esters, ethers aromatic amines nitro compounds. Absorbance Peak at 894.91 cm\u003csup\u003e− 1\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e 817.7cm\u003csup\u003e− 1\u003c/sup\u003e, 779.19cm\u003csup\u003e− 1\u003c/sup\u003e, 623.93cm\u003csup\u003e− 1\u003c/sup\u003e, 515.92 cm\u003csup\u003e− 1\u003c/sup\u003e examined C-H “oop” N-H Wag = C-H, C-Cl Stretching, -C(triple bond)C-H C-H bend, C-Br Stretching bend represents aromatics primary, secondary amines alkenes, Alkyl halides, primary, secondary amines, alkenes, alkylhalides, alkynes, alkyl halides (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\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\u003eFTIR spectrum analysis of functional groups present in AgNPusing \u003cem\u003eA.nilotica\u003c/em\u003e bark extract.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"5\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS.No\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eFrequency absorption (cm\u003csup\u003e− 1)\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAppearance\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eFunctional Group\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eCompound Class\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e427.2\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\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e2.\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e452.28\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\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e3.\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e515.92\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedium\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-Br Stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealkyl halides\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e623.93\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ebroad, sharp\u003c/p\u003e \u003cp\u003enarrow\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-C(triple bond) C-H\u003c/p\u003e \u003cp\u003eC-H bend\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealkynes\u003c/p\u003e \u003cp\u003ealkynes\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e779.19\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedium\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-Cl Stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealkyl halides\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e817.76\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedium\u003c/p\u003e \u003cp\u003estrong, broad\u003c/p\u003e \u003cp\u003estrong\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-Cl Stretch\u003c/p\u003e \u003cp\u003eN-H wag\u003c/p\u003e \u003cp\u003eC = H bend\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealkyl halides\u003c/p\u003e \u003cp\u003eprimary, secondary amines, alkenes\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e894.91\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eStrong\u003c/p\u003e \u003cp\u003estrong, broad\u003c/p\u003e \u003cp\u003estrong\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-H “oop”\u003c/p\u003e \u003cp\u003eN-H Wag\u003c/p\u003e \u003cp\u003e=C-H bend\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003earomatics\u003c/p\u003e \u003cp\u003eprimary, secondary amines, alkenes\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1028.95\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedium\u003c/p\u003e \u003cp\u003eStrong\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-N Stretch\u003c/p\u003e \u003cp\u003eC-O Stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealiphatic amines\u003c/p\u003e \u003cp\u003ealcohols, carboxylic acids, esters, ethers\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1111.89\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eStrong\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-O Stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealcohols,carboxylic acids, esters, ethers\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1161.07\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedium\u003c/p\u003e \u003cp\u003emedium\u003c/p\u003e \u003cp\u003estrong\u003c/p\u003e \u003cp\u003emedium\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-H wag (-CH2X)\u003c/p\u003e \u003cp\u003eC-H wag (-CH2X)\u003c/p\u003e \u003cp\u003eC-O Stretch\u003c/p\u003e \u003cp\u003eC-N Stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealkyl halides\u003c/p\u003e \u003cp\u003ealkyl halides\u003c/p\u003e \u003cp\u003eAlcohols, carboxylic acids, esters, aliphatic amines.\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1238.21\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedium\u003c/p\u003e \u003cp\u003eMedium\u003c/p\u003e \u003cp\u003emedium\u003c/p\u003e \u003cp\u003estrong\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-N Stretch\u003c/p\u003e \u003cp\u003eC-H wag(-CH2X)\u003c/p\u003e \u003cp\u003eC-H wag(-CH2X)\u003c/p\u003e \u003cp\u003eC-O stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealiphatic amines\u003c/p\u003e \u003cp\u003ealkyl halides\u003c/p\u003e \u003cp\u003ealkyl halides\u003c/p\u003e \u003cp\u003ealcohols, carboxylic acids, esters, ethers\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1317.29\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eStrong\u003c/p\u003e \u003cp\u003estrong\u003c/p\u003e \u003cp\u003emedium\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-O Stretch\u003c/p\u003e \u003cp\u003eC-N Stretch\u003c/p\u003e \u003cp\u003eN-O Symmetric stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealcohols, carboxylic acids, esters, ethers\u003c/p\u003e \u003cp\u003earomatic amines\u003c/p\u003e \u003cp\u003enitro compounds\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1374.19\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\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e14.\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1451.33\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedium\u003c/p\u003e \u003cp\u003eMedium\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-H bend\u003c/p\u003e \u003cp\u003eC-C Stretch (in-ring)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealkanes\u003c/p\u003e \u003cp\u003earomatics\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1513.05\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eStrong\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eN-O asymmetric stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003enitro compounds\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1619.13\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedium\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eN-H bend\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ePrimary amines\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e1747.39\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eStrong\u003c/p\u003e \u003cp\u003eStrong\u003c/p\u003e \u003cp\u003eStrong\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC = O stretch\u003c/p\u003e \u003cp\u003eC = O stretch\u003c/p\u003e \u003cp\u003eC = O stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eesters, saturated aliphatic\u003c/p\u003e \u003cp\u003ecarboxylic acids, carbonyls (general)\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e2321.17\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\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e19.\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e2380.96\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\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e20.\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e2873.74\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eStrong\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-H stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealkanes\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e2932.56\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedium\u003c/p\u003e \u003cp\u003eMedium\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eC-H stretch\u003c/p\u003e \u003cp\u003eO-H stretch\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealkanes\u003c/p\u003e \u003cp\u003ecarboxylic acids\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e3444.63\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedium\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eO-H stretch, H-bonded\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ealcohols, phenols\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=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e3722.36\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\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e24.\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e3769.61\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\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec25\" class=\"Section2\"\u003e \u003ch2\u003e3.4. SEM with EDAX\u003c/h2\u003e \u003cp\u003eThe morphological pattern of AgNP was authorized by applying of SEM. The shapes of particles are predominantly spherical and aggregate into larger particles with no well-defined morphology. This aggregation may be due to the presence of secondary metabolites in the bark extracts. SEM yielding only inadequate evidence on the actual nature of the shape and size distribution of AgNP of \u003cem\u003eAcacia nilotica extracts.\u003c/em\u003e The authentic diameters of the AgNP were measured between Maximum 300 nm and minimum 200 nm in diameter executed SEM characterization. The uniform size of silver suggests that the particles are observed. SEM analysis of AgNP was clearly distinguishable owing to their size difference of synthesized AgNP. Scanning electron microscope images of AgNP using \u003cem\u003eAcacia nilotica\u003c/em\u003e bark extract at different magnification like 100x, 200x, 500x (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e5\u003c/span\u003ea-f).\u003c/p\u003e \u003cp\u003eEDX bands were clarified to provide qualitative and quantitative signal of the chemical elements present in newly synthesized AgNP. Displaying of particular intensity peak region in electron beam of EDX identified distinct chemical elemental composition of AgNP. To identify the level of NKα1 weight percentage is 4.5%, Okα1 weight percentage is 42.7%, AgLα., AgLβ., AgLβ2 were weight is 52.8%, Atomic percentage is NKα1 9.3%, Atomic percentage is Okα1 76.6.%, AgLα., AgLβ., AgLβ2 Atomic percentage is 14.1%, Error percentage is NKα1 13.0%, Error percentage is Okα1 10.5%, Error percentage is AgLα., AgLβ., AgLβ2 2.4%, K ratio is NKα1 0.0172, K ratio is Okα1 0.0756, K ratio is AgLα., AgLβ., AgLβ2 0.4649. It indicated the higher content of AgNP. But, a strong elemental signal with distinct peak for Ag was confirmed at 3.15keV in EDX analysis (Figure: 6). Moreover, weight percentage, atomic percentage and K ratio was mentioned in Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\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\u003eSmart Quant Results\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"5\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eElement\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eWeight %\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAtomic %\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eError %\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eK ratio\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eN K\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e4.5\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e9.3\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e13.0\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.0172\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eO K\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e42.7\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e76.6\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e10.5\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.0756\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAg L\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e52.8\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e14.1\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e2.4\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.4649\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e \u003cp\u003e\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\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\u003eLarvicidal activity of \u003cem\u003eAcacia nilotica\u003c/em\u003e bark extracts against fourth instar larvae of \u003cem\u003eAedesagypti., Anopheles stephensi., Culexquiquefasciatus.\u003c/em\u003e\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"7\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eMosquito species\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eSample\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003en\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eLC\u003csub\u003e50\u003c/sub\u003e (mg/ml)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eLC\u003csub\u003e90\u003c/sub\u003e (mg/ml)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eX\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c7\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003edf\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003e(95%CL)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003e(95%CL)\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\u003eAedes Agypti\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAgNP\u003c/p\u003e \u003cp\u003ePlant extract\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e60\u003c/p\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e240.714\u003c/p\u003e \u003cp\u003e(235.18–241.61)\u003c/p\u003e \u003cp\u003e313.116\u003c/p\u003e \u003cp\u003e(309.28–316.82)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e367.006\u003c/p\u003e \u003cp\u003e(361.1–370.01)\u003c/p\u003e \u003cp\u003e417.820\u003c/p\u003e \u003cp\u003e(414.16–421.8)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.579\u003c/p\u003e \u003cp\u003e0.703\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e3\u003c/p\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eAnopheles stephensi\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAgNP\u003c/p\u003e \u003cp\u003ePlant extract\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e60\u003c/p\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e104.13\u003c/p\u003e \u003cp\u003e(99.811–106.161)\u003c/p\u003e \u003cp\u003e296.73\u003c/p\u003e \u003cp\u003e(291.11–298.12)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e158.13\u003c/p\u003e \u003cp\u003e(151.03–160.82)\u003c/p\u003e \u003cp\u003e450.08\u003c/p\u003e \u003cp\u003e(441.18–453.16)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.426\u003c/p\u003e \u003cp\u003e0.499\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e3\u003c/p\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eCulex Quinquefasilitus\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAgNP\u003c/p\u003e \u003cp\u003ePlant extract\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e60\u003c/p\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e99.602\u003c/p\u003e \u003cp\u003e(95.116–103.82)\u003c/p\u003e \u003cp\u003e293.126\u003c/p\u003e \u003cp\u003e(289.72–298.11)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e153.59\u003c/p\u003e \u003cp\u003e(148.26–159.12)\u003c/p\u003e \u003cp\u003e451.754 (443.16–455.16)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.453\u003c/p\u003e \u003cp\u003e0.597\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e3\u003c/p\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003ctfoot\u003e\u003ctr\u003e\u003ctd colspan=\"7\"\u003en\u003csup\u003ea\u003c/sup\u003e-total number of mosquitoes larvae used; n = 20/replicate; LC50-Lethal Concentration 50% mortality, LC90-Lethal concentration 90% mortality, LCL-Lower confidence limits, UCL-Upper confidence limits, X\u003csup\u003e2\u003c/sup\u003e-chi-square, df- degrees of freedom (Note: Chi square values with a static are significant P \u0026lt; 0.05).\u003c/td\u003e\u003c/tr\u003e\u003c/tfoot\u003e\u003c/table\u003e\u003c/div\u003e \u003cp\u003e\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\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\u003eAdulticidal activity of \u003cem\u003eAcacia nilotica\u003c/em\u003e bark extract species against \u003cem\u003eAedesagypti.,Anophlesstephensi., Culexquinquefasciatus.\u003c/em\u003e\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"7\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAdult Mosquito Species\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSample\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003en\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eLC\u003csub\u003e50\u003c/sub\u003e(mg/ml)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eLC\u003csub\u003e90\u003c/sub\u003e(mg/ml)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eX\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003edf\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\u003eAedes agypti\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAgNP\u003c/p\u003e \u003cp\u003ePlant extract\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e60\u003c/p\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e13.560\u003c/p\u003e \u003cp\u003e(9.4–17.3)\u003c/p\u003e \u003cp\u003e14.314\u003c/p\u003e \u003cp\u003e(11.31–16.41)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e24.35\u003c/p\u003e \u003cp\u003e(20.47–34.43)\u003c/p\u003e \u003cp\u003e257.76\u003c/p\u003e \u003cp\u003e(218.61–329.62)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e2.113\u003c/p\u003e \u003cp\u003e1.421\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e3\u003c/p\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eAnopheles Stephensi\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAgNP\u003c/p\u003e \u003cp\u003ePlant extract\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e60\u003c/p\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e16.854\u003c/p\u003e \u003cp\u003e(13.06–19.011)\u003c/p\u003e \u003cp\u003e17.269\u003c/p\u003e \u003cp\u003e(15.09–23.259)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e28.411\u003c/p\u003e \u003cp\u003e(24.24–39.2)\u003c/p\u003e \u003cp\u003e31.130 (27.620–33.03)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1.039\u003c/p\u003e \u003cp\u003e1.642\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e3\u003c/p\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eQulex Quinquefasciatus\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAgNP\u003c/p\u003e \u003cp\u003ePlant extract\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e60\u003c/p\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e17.012 (14.458–20.05)\u003c/p\u003e \u003cp\u003e18.312 (15.458–21.05)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e31.230\u003c/p\u003e \u003cp\u003e(27.18–34.011)\u003c/p\u003e \u003cp\u003e33.204\u003c/p\u003e \u003cp\u003e(29.18–36.016)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1.267\u003c/p\u003e \u003cp\u003e1.675\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e3\u003c/p\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003ctfoot\u003e\u003ctr\u003e\u003ctd colspan=\"7\"\u003e* indicates significantly difference (p \u0026lt; 0.01) between different solvent extracts. LC50—lethal concentration that kills 50% of the exposed larvae, LC90—lethal concentration that kills 90% of the exposed larvae, UCL—upper confidence limit (95%), LCL—lower confidence limit (95%), df—degrees of freedom.\u003cb\u003eTable: 5 Smoke toxicity test of\u003c/b\u003e \u003cb\u003eA. nilotica\u003c/b\u003e \u003cb\u003eplant powder compared with commercial mosquito coils and mosquito coil without any plant powder on major three vectors\u003c/b\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tfoot\u003e\u003c/table\u003e\u003c/div\u003e \u003cp\u003e\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eTable: 5 Smoke toxicity test of \u003cem\u003eA. nilotica\u003c/em\u003e plant powder compared with commercial mosquito coils and mosquito coil without any plant powder on major three vectors\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 126px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eMosquito Species\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eObservation\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(in Minutes)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 131px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e% Mortality from \u003cem\u003eA. nilotica\u003c/em\u003e Powder Mosquito Coil\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 113px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e% Mortality from CommercialMosquito Coil\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 133px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e% Control Mortality\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 126px;\"\u003e\n \u003cp\u003e\u003cem\u003eAedes agypti\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 131px;\"\u003e\n \u003cp\u003e00\u003c/p\u003e\n \u003cp\u003e02\u003c/p\u003e\n \u003cp\u003e11\u003c/p\u003e\n \u003cp\u003e92\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 113px;\"\u003e\n \u003cp\u003e00\u003c/p\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003cp\u003e53\u003c/p\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 133px;\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 126px;\"\u003e\n \u003cp\u003e\u003cem\u003eAnopheles stephensi\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003cp\u003e\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 131px;\"\u003e\n \u003cp\u003e00\u003c/p\u003e\n \u003cp\u003e08\u003c/p\u003e\n \u003cp\u003e24\u003c/p\u003e\n \u003cp\u003e85\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 113px;\"\u003e\n \u003cp\u003e00\u003c/p\u003e\n \u003cp\u003e14\u003c/p\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 133px;\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 126px;\"\u003e\n \u003cp\u003e\u003cem\u003eCulex quinquefasciatus\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 99px;\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003cp\u003e\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 131px;\"\u003e\n \u003cp\u003e00\u003c/p\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003cp\u003e24\u003c/p\u003e\n \u003cp\u003e83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 113px;\"\u003e\n \u003cp\u003e00\u003c/p\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003cp\u003e46\u003c/p\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 133px;\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec26\" class=\"Section2\"\u003e \u003ch2\u003e3.5. XRD (X-ray Diffraction)\u003c/h2\u003e \u003cp\u003eThe formation and quality of compounds were checked by X-ray diffraction (XRD) spectrum. The size of the purified Silver nanoparticles was analysed by X-ray powder diffraction crystallography (SEIFERT JSO-DEBYEREX-2002, Germany) diffractometer with Cu-Kα1 radiation. A scan rate of 0.020 per second and a scan range between 10–80 2 theta in flat plate geometry with Ag radiation. The crystalline domain size was calculated using \u003cem\u003eD. Scherrer\u003c/em\u003e’s equation (D = kλ/βcosθ) (Hendricks, 1941; Suryanarayana, 1998) where, D = average crystalline domain size, β denotes Full Width Half Maximum (FWHM); K = 0.94, λ = wavelength of X-ray (1.54 A\u003csup\u003e0\u003c/sup\u003e) and θ is the Brag’s angle). XRD pattern of synthesized to AgNP was mentioned in the Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e7\u003c/span\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec27\" class=\"Section2\"\u003e \u003ch2\u003e3.6. In-vitro antioxidant activity\u003c/h2\u003e \u003cdiv id=\"Sec28\" class=\"Section3\"\u003e \u003ch2\u003e3.6.1. DPPH (2, 2-diphenyl-1-picrylhydrazyl)\u003c/h2\u003e \u003cp\u003eUtilizing DPPH, ABTS, and H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e tests, the antioxidant capacity of the AgNP-containing \u003cem\u003eAcacia nilotica\u003c/em\u003e bark extract was assessed. Percentage of DPPH radical scavenging activity inhibition measured at various AgNP doses (20 µg/mL, 40 µg/mL, 60 µg/mL, 80 µg/mL, and 100 µg/mL). The present investigation examined DPPH activity in a dose-dependent fashion. Similar, dose-dependent action was discovered in AgNPs by the use of bark extracts from \u003cem\u003eAcacia nilotica\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e8\u003c/span\u003ea).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec29\" class=\"Section3\"\u003e \u003ch2\u003e3.6.2. Hydroxyl (H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e) scavenging activity\u003c/h2\u003e \u003cp\u003eAccording to the findings of the antioxidant assays, all extracts may have some degree of radical scavenger activity (Fig.\u0026nbsp;\u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e8\u003c/span\u003eb). The variable chemical makeup of different phytochemicals, which may react differently with different kinds of free radicals, may be the cause of the varying scavenging actions of different extracts.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec30\" class=\"Section3\"\u003e \u003ch2\u003e3.6.3. ABTS assay\u003c/h2\u003e \u003cp\u003eThe ABTS assay was used to evaluate the free radical scavenging activity. A widely used colorimetic technique for determining the ability of bark extracts to scavenge radicals is the 1,1-diphenyl-2-picrylhydrazl (DPPH) assay. This technique, which screens the overall activity of antioxidants and is based on the stable synthetic radical DPPH, is precise, simple to use, and reasonably priced. ABTS is a chemically stable and highly water soluble substance. It generates a metastable cation and functions as a substrate for peroxide. Spectrophotometric measurement shows that the synthesis of ABTS is inhibited when an antioxidant molecule is present. The potential of the antioxidant chemical increased with decreasing test solution absorption at 734 nm. In order to reduce interference from sample turbidity and other absorbing materials, the hydrogen donating potential in the antioxidant samples was measured at 734 nm (Fig.\u0026nbsp;\u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e8\u003c/span\u003ec).\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec31\" class=\"Section2\"\u003e \u003ch2\u003e3.7. Antibacterial activity using Disc diffusion method\u003c/h2\u003e \u003cp\u003eFigure \u003cspan refid=\"Fig13\" class=\"InternalRef\"\u003e9\u003c/span\u003e showed the results of an antimicrobial activity test using the Disc Diffusion Method against several bacterial strains. Using a ruler or caliper, measure the zones during incubation to the closest millimeter; make careful to account for the disk's diameter in your measurement. Measure from the disk's center to a point on the zone's perimeter where a clear edge is present (the radius), then multiply the result by two to find the diameter if the zone's size or placement prevent you from reading the diameter. Swarming bacterial strains require a different method of measurement than non swarming organisms. Measure the outer boundary in an otherwise obvious zone of inhibition, ignoring the narrow vein of swarming. The small layer of swarming development within an otherwise clear zone of inhibition should be ignored when measuring the zone of inhibition for swarming organisms. Only the categories of susceptible, intermediate, and resistant are reported for the Well Diffusion technique susceptibility test results.\u003c/p\u003e \u003cp\u003eAgNP's higher antimicrobial activity was facilitated by the phytonutrients that helped maintain it, and its increased antibacterial effectiveness can be attributed to its surface area and macromolecules. The antibacterial survivability of AgNP may be due to its tiny size, dispersion, and spherical morphology, which provide a broad surface for maximum interaction with bacteria and cause much more damage than bigger particle sizes. AgNP's antibacterial activity was evaluated using the four different bacterial species. Analysis was done on the effects of AgNP as a time feature in fixed amounts of bacterial species' cell morphology (Fig.\u0026nbsp;\u003cspan refid=\"Fig14\" class=\"InternalRef\"\u003e10\u003c/span\u003ea–f).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec32\" class=\"Section2\"\u003e \u003ch2\u003e3.7. Antimicrobial activity by using minimum inhibitory conentration\u003c/h2\u003e \u003cp\u003eTo determine the antimicrobial dosage of our AgNP, Minimum Inhibitory Concentration (MIC) was done using different concentrations (120 µg/ml, 160 µg/ml, 200 µg/ml 240 µg/ml and 300 µg/ml) AgNP (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e11\u003c/span\u003e). Antimicrobial activity test was done with \u003cem\u003eAcacia nilotica\u003c/em\u003e derived AgNP Minimum inhibitory concentration using \u003cem\u003eS. aureus\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig15\" class=\"InternalRef\"\u003e12\u003c/span\u003e). Minimum inhibitory concentration of \u003cem\u003eAcacia nilotica\u003c/em\u003e bark extract was and increasing concentration of synthesized ANL-AgNPs in terms of the zone of inhibition against five types of bacterial strains [\u003cem\u003eEscherichia coli (\u003c/em\u003eATCC number of 25922), \u003cem\u003eStaphylococcus aureus (\u003c/em\u003eATCC number of 25923\u003cem\u003e), Streptococcus aureus (\u003c/em\u003eATCC number of 700294\u003cem\u003e)\u003c/em\u003e, \u003cem\u003ePseudomonas aeruginosa\u003c/em\u003e (ATCC number of 15442), \u003cem\u003eProteus vulgaris\u003c/em\u003e (ATCC number of 6380)].\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec33\" class=\"Section2\"\u003e \u003ch2\u003e3.8. Larvicidal potential of AgNP\u003c/h2\u003e \u003cp\u003eThe larvicidal efficacy and synthesized to \u003cem\u003eAgNP\u003c/em\u003e were tested as per the guidelines of world Health organization procedure with some modifications. Different concentration (120µg/ml; 160µg/ml; 200µg/ml; 240µg/ml) AgNP were used, respectively, to test larvicidal activity on \u003cem\u003eAedes aegypti, Anopheles stephensi, and Culex quinquefasciatus\u003c/em\u003e mosquito larvae and a control with test samples. The number of dead larvae was counted after 24 hr. of exposure and their percentage mortality was calculated using Abbott’s formula Mortality (%)\u003c/p\u003e\u003cp\u003e\u003cimg src=\"https://myfiles.space/user_files/127393_c7e80a1c9bb65875/127393_custom_files/img1732794594.png\"\u003e\u003cbr\u003e\u003c/p\u003e \u003cp\u003ewhere n = number of larvae, T = treated, C = control.\u003c/p\u003e \u003cp\u003eThis has made it necessary to look for and create mosquito-repelling insecticides that are more affordable, target-specific, and safe for the environment. Botanical derivatives, or natural compounds derived from plants, are a newer and alternate method of controlling mosquitoes. Numerous natural botanical products are affordable, efficient, safe for the environment, and readily biodegradable. They have a variety of unique ways of action and don't harm non-target creatures. Results of bioassays using bark extracts from \u003cem\u003eAcacia nilotica\u003c/em\u003e against the larvae of three significant vector mosquito species—\u003cem\u003eAedes Agypti, Anopheles Stephensi\u003c/em\u003e, and \u003cem\u003eCulex Quinquefasciatus\u003c/em\u003e—showed the larvicidal efficacy of the extracts (table:3). The plant extracts with LC50 and LC90 values of -0.02, -2.560, -3.961 and.015, 2.332, 1.075 mg/ml, respectively, showed the best larvicidal activity. The degree of freedom (df), chi-square, and 95% confidence limits, LC50 and LC90, were also computed. We did not see any noteworthy mortality in the control assays.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec34\" class=\"Section2\"\u003e \u003ch2\u003e3.9. Adulticidal potential of AgNP\u003c/h2\u003e \u003cp\u003eSince adulticides may only temporarily lower the adult population, mosquito control programs are primarily focused on eradicating adult mosquitoes at their nesting places with larvicides. Therefore, focusing on the larvae would be a more effective way to lower the mosquito population. Our findings unequivocally show that \u003cem\u003eAcacia nilotica\u003c/em\u003e was more successful in suppressing immature phases. \u003cem\u003eAedes agypti\u003c/em\u003e, \u003cem\u003eAnopheles stephensi\u003c/em\u003e, and \u003cem\u003eCulex quinquefasciatus\u003c/em\u003e are the three major vector mosquito species against which the adulticidal action of \u003cem\u003eAcacia nilotica\u003c/em\u003e bark extract has been recorded (table:4). The mortality rate of the bark extract rose in direct proportion to dosage. The bark extract that was examined showed the strongest adulticidal activity against \u003cem\u003eCulex quinquefasciatus\u003c/em\u003e, \u003cem\u003eAnopheles stephensi\u003c/em\u003e, and \u003cem\u003eAedes aegypti\u003c/em\u003e. When the control group was tested without a solvent, there was no death.\u003c/p\u003e \u003cp\u003eThe adulticidal activity of \u003cem\u003eAedes agypti\u003c/em\u003e, \u003cem\u003eAnopheles stephensi\u003c/em\u003e, and \u003cem\u003eCulex quinquefasciatus\u003c/em\u003e was measured by \u003cem\u003eAcacia nilotica\u003c/em\u003e bark extracts, and the LC50 and LC90 values were 2.039 and 2.671;603 and 494; 2.930; 6.379 mg/ml, respectively. After 48 hours of PT, the adulticidal mortality reached its peak, and in extracts from the bark of \u003cem\u003eAcacia nilotica\u003c/em\u003e, the mortality reached 100% at 1500 PPm. The following are the lethal concentrations: 120, 160, 200, and 240 µg/ml. In both aqueous and bark extracts, the larval death rate for the conventional positive control was 100% at 24 hours and after treatment, respectively.\u003c/p\u003e "},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding Declaration\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThere is no financial support for this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthical approval\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\u003eAll the authors agreed to publish the data in this journal.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCRediT authorship contribution statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eR.Bruntha\u003c/strong\u003e: Writing \u0026ndash; review \u0026amp; editing, Writing \u0026ndash;original draft, Visualization, Validation, Software, Resources, Methodology, Investigation, Funding acquisition, Formal analysis, Data curation, Conceptualization. \u003cstrong\u003eS.Lokeshwaran\u003c/strong\u003e: Visualization, Data curation. Kirubakaaran Dharmaligam: Software, Formal analysis. \u003cstrong\u003eS.P.Subala:\u003c/strong\u003e Supervision, Methodology, Investigation, Formal analysis, Data curation, Conceptualization.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDeclaration of Competing Interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll of the authors are declared there is no conflict of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors do not have permission to share data.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgment\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors recognize and thank the Department of Biotechnology, Ayya Nadar Janaki Ammal College (autonomous), Sivakasi-626125, Tamil Nadu, India, for providing infrastructural facilities.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAdhavan, R., Selvam, K., Prakash, P., Kirubakaran, D., Shivakumar, M. 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WHO/VBC/81.806.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eXiao, M., Jia, X., Wang, N., Kang, J., Hu, X., Goff, H.D., Cui, S.W., Ding, H., Guo, Q., 2024. Therapeutic potential of non-starch polysaccharides on type 2 diabetes: from hypoglycemic mechanism to clinical trials. Crit. Rev. Food Sci. Nutr. 64 (4), 1177\u0026ndash;1210. https://doi.org/10.1080/10408398.2022.2113366.\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":"Acacia nilotica, Antimicrobial, Antioxidant, Smoke toxicity, Larvicidal","lastPublishedDoi":"10.21203/rs.3.rs-5353202/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5353202/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eIn the present study, the bark extract of \u003cem\u003eAcacia nilotica\u003c/em\u003e plant was used to green synthesis and Characterization of Silver nanoparticles and check their insecticidal potential against three important mosquito vectors such as \u003cem\u003eAedes aegyptii, Anopheles stephensi\u003c/em\u003e and \u003cem\u003eCulex quinquefasciatus\u003c/em\u003e. Our research aims to investigate the therapeutic potential of plant extract combined with nanoparticles. The synthesized silver nanoparticles were characterized by UV-Visible spectroscopy (UV-Vis), Fourier transform infrared Spectroscopy (FT-IR), and scanning electron microscopy (SEM) with EDax techniques (EDAX) and X-ray diffraction (XRD). UV-Vis confirms with the peak at 420 nm showed the synthesis of silver nanoparticles. SEM confirms the spherical shape of nanoparticles and EDAX revealed chemical components of Ag, N, and Oxygen with a strong elemental signal in association with distinct peak for Ag was confirmed at 3.15keV. XRD was determined the crystalline properties of the silver nanoparticles (AgNP). The antioxidant potential of \u003cem\u003eAcacia nilotica\u003c/em\u003e bark extract using AgNP was determined by DPPH, hydroxyl and ABTS free radical scavenging assays. DPPH and hydroxyl activities of AgNP showed the highest inhibition of 63% and 84%, while ABTS analysis showed 58%. Antibacterial activity shows highest level of activity against \u003cem\u003eS. pyrogens, P. aeruginosa and P.vulgaris\u003c/em\u003e. Larvicidal potential of AgNPs recorded good mortality rate against \u003cem\u003eAedes aegypti\u003c/em\u003e., \u003cem\u003eAnopheles stephensi., Culex quinquefasciatus\u003c/em\u003e. This study confirms the maximum activity of AgNPs and acts as larvicidal, adulticidal, antimicrobial, and antioxidant agents.\u003c/p\u003e","manuscriptTitle":"Green synthesis and Characterization of Acacia nilotica derived silver nanoparticles and their insecticidal potential against three important mosquito vectors","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-11-28 11:58:17","doi":"10.21203/rs.3.rs-5353202/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":"7a00e92c-dd1b-4724-ae9f-6915d26f3853","owner":[],"postedDate":"November 28th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2026-01-03T17:38:28+00:00","versionOfRecord":[],"versionCreatedAt":"2024-11-28 11:58:17","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-5353202","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5353202","identity":"rs-5353202","version":["v1"]},"buildId":"WrCJVZZCHTDjtuVLN7oU0","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}