2 Afzelia Quanzensis Bark Extract for Green Synthesis of Silver 3 Nanoparticles and Study of Their Antibacterial Activity

' Int J Ind Chem CrossMark DOI 10.1007/s40090-015-0055-7

1 RESEARCH

2 Afzelia quanzensis bark extract for green synthesis of silver 3 nanoparticles and study of their antibacterial activity

4 Mambo Moyo1 *3®* Makore Gomba1 • Tichaona Nharingo1

5 Received: 14 September 2014/Accepted: 10 September 2015 6 © The Author(s) 2015. This article is published with open access at Springerlink.com

7 Abstract In the present study, Afzelia quanzensis bark such as their use in bio-labelling, sensors, drug delivery 32 8 extract was tested for the biosynthesis of silver nanoparti- system, antimicrobial agents and filters [1, 2], The syn- 33 JHJcles (AgNPs). Based on UV-Vis spectrum analysis, the thesized silver nanoparticles exhibit new or improved 34 10 characteristic absorption band was observed at 427 nm. properties depending upon their size, morphology and 35 11 Furthermore, the size and shape of the nanoparticles ranged distribution [3], The production of pure and well-defined 36 12 from 10 to 80 nm and were spherical in shape as observed metal-based nanoparticles by chemical reduction [4], 37 13 through SEM analysis. In addition, the X-ray diffraction thermal treatment, irradiation [5] and laser ablation [6] 38 URBlanalvsis showed that the silver nanoparticles are crystalline have been reported. On one hand, organic solvents, toxic 39 15 in nature and have a face-centred cubic structure. Based on reducing agents, high-pressure and high-temperature con- 40 16 FTIR analysis, the presence of phytochemical functional version which are potentially dangerous to the environment 41 17 groups such as carboxyl (-C=0) and amine (N-H) in are used [7], Hence, a pressing need to shift from physical 42 18 Afzelia quanzensis bark extract further confirmed the and chemical synthesis to ‘green’ chemistry and biopro- 43 19 responsible reducing agents for nanoparticles formation. cesses is of major interest. 44 20 Interestingly, the synthesized silver nanoparticles at 50 mg/ In the last decade, the biosynthesis of nanoparticles has 45 21 L concentration showed significant antibacterial activity received increasing attention due to the growing need to 46 22 against Escherichia coli and Staphylococcus aureus. develop environmentally benign technologies in material 47 synthesis [8, 9], Biological routes for the synthesis of metal 48 24 Keywords Biological synthesis • Silver nanoparti cl es ■ nanoparticles by exploiting bacteria [10, 11], marine fun- 49 25 Bark extract ■ Antibacterial activity gus Penicillium fellutanum [I], fungus Aspergillus foefidus 50 [12], yeast [13, 14], enzymes [15] have been reported. 51 However, one of the major drawbacks in using microbes 52 fth Introduction for nanoparticle synthesis is the elaborate process of 53 maintaining microbial cultures. The use of plant extracts 54 27 At present, nanotechnology is a rapidly developing field of for the synthesis of nanoparticles have gained momentum 55 28 importance since it deals with the synthesis and stabiliza- in recent years and could be advantageous over other 56 29 tion of different metal nanoparticles. Among the particles, environmentally benign biological processes by eliminat- 57 30 silver nanoparticles (AgNPs) have become the focus of ing the elaborate process of maintaining cell cultures, being 58 31 intensive research due to several important applications simply, eco-friendly and this could be an exciting possi- 59 bility that is relatively unexplored and under exploited 60 [16]. Green silver nanoparticles synthesis using various 61 1 E3 Mambo Moyo natural products like Magnolia kohus [17], Acacia leu- 62 A2 [email protected] cophloea extract [18], Zizyphus xylopyrus bark extract [19], 63 aloevera plant extract [20, 21], Cinnamon zeylanicum bark 64 3 1 Department of Chemical Technology, Faculty of Science, A4 Midlands State University, P. Bag 9055, Senga, Gweru, extract [22], Curcuma longa tuber powder [23] and Jat- 65 Zimbabwe ropha curcas [9] have been reported. However, 'the 66

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67 potential of other plants as sources of biomaterial for the In the present study, we used plant bark extracts for 101 68 synthesis of new nanoparticles are yet to be entirely synthesis of silver nanoparticles by monitoring their con 102 69 explored. version using UV-visible spectroscopy. We also investi 103 70 It has been reported that plants contain different phy gated the effects of reaction conditions such as 104 71 tochemical products [24] which are able to breakdown the temperature, pH, quantities of bark extract and AgN03 105 72 silver nitrate, a complex hazardous chemical into Ag+ and concentration on the synthesis rate. The silver nanoparti 106 73 N 03_ ions. In the process, the toxic Ag+ ions are further cles were further characterized by X-ray diffraction (XRD), 107 74 reduced to the nontoxic (Ag°) metallic nanoparticles scanning electron microscopy (SEM), energy dispersive 108 75 through the use of different functional groups on the sur X-ray (EDX) spectrometer, and Fourier transform infrared 109 76 face of the extract [25], In the present study, we selected spectroscopy (FTIR). Furthermore, the antibacterial activ 110 77 Afzelia quanzensis, Pod mahogany (English), Mujar- ity of the silver nanoparticles on a strain of Escherichia coli 111 78 akamba (Shona name in Zimbabwe) bark as a biomaterial and Staphylococcus aureus was qualitatively evaluated by 112 79 for supplying the different phytochemicals required for the the zone inhibition method. 113 80 synthesis of silver nanoparticles and the mechanism 81 involved in the synthesis is illustrated in Fig. 1. The plant 82 is economic and abundantly available in Zimbabwe. It is a Materials and methods 114 83 medium-sized to large deciduous tree and has a bark which 84 is greyish-brown, flaking and leaving pale patches [26]. It Preparation of Afzelia quanzensis bark powder 115 85 produces fruits which consist of a large flattened pod, and extract 116 |86 thickly woody, 10-17 cm, and splitting to reveal large, *87 shiny black seeds with a bright red aril. In medicine, roots The bark was obtained from Afzelia quanzensis tree in 117 88 are used to treat gonorrhoea, chest pains, kidney problems, Chivi rural district area, Zimbabwe. The bark was washed 118 89 bilharzia, eye problems and snakebites, and a small piece to remove any impurities and dried under sunlight for a 119 90 of bark is applied to an aching tooth. The plant can be week to completely remove the moisture. The bark was cut 120 91 categorized as: into small pieces, powdered in a mixer and then sieved 121 92 Taxonomy using a 20-mesh sieve to get uniform size range. The 122 93 Kingdom: Plantae sieved powder was used for all further studies. For the 123 94 Division: Tracheophyta production of an extract, 50 g of powdered bark was added 124 95 Subdivision: Spermatophytina to a 500-mL Erlenmeyer flask containing 200 mL deion 125 96 Class: Magnoliopsida ized water and then boiled for 15 min. After cooling, the 126 97 Order: Fabales mixture was filtered through Whatman filter paper no. 1 127 98 Family: Fabaceae and the extract was kept at 4 °C prior to silver nanoparti 128 99 Genus: Afzelia Sm.—mahogany cles synthesis. 129 Species: Afzelia quanzensis Welw.—pod mahogany

Fig. 1 Mechanism involved in the synthesis of silver nanoparticles using Afzelia Silver Nitrate quanzensis extract Chemistry (AgNOj)

Afzeiia quanzensis Afzelia quanzensis baric extract Reduction process baric Ag+ NO3-

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130 Synthesis of silver nanoparticles 4000 cm-1. FTIR measurements were made to identify the 173 possible biomolecules responsible for capping and efficient 174 131 Silver nitrate (AgN03) analytical grade used as a precursor stabilization of the synthesized silver nanoparticles. 175 132 was purchased from Sigma-Aldrich (Pretoria, South 133 Africa). For the AgNPs synthesis, 5 mL bark extract were Study for the influence of different parameters 176 134 added to 50 mL of 1 mM aqueous AgN03 solution in a 135 250-mL Erlenmeyer flask. The flask was then incubated in Influence of different temperatures (30, 40, 50, 60, 70, 80, 177 136 a rotary shaker at 160 rpm in the dark. The reduction of 90 °C), pH values (3, 5, 7, 8, 11), bark extract amounts (2, 178 137 silver ions was routinely monitored visually for colour 4, 6, 8, 10, 12, 14 mL), substrate concentrations (1, 3, 5, 7, 179 138 change at regular intervals. Thereafter, the silver 9, 11, 13 mM AgN03) and incubation periods (20, 40, 60, 180 139 nanoparticle solution thus obtained was purified by repe 80, 100, 120, 140 min) were investigated by varying the 181 140 ated centrifugation at 500 rpm for 20 min followed by re parameters one at a time. A sample of 1 mL was with- 182 141 dispersion of the pellet of silver nanoparticles into a 10 mL drawn at different time intervals and the absorbance was 183 142 of deionized water. After freeze drying, lyophilization measured at 427 nm. 184 143 process was performed on the purified silver nanoparticles 144 to obtain the powdered form which was stored in brown Measuring concentration of AgNPs using 185 145 bottles prior to physical characterization and antibacterial inductively coupled plasma optical emission 186 146 activity. spectrophotometer (ICP-OES) 187

^4 7 Characterization The original concentration (80 mg/L) of the Afzelia 188 quanzensis bark extract mediated AgNPs was measured 189 148 UV-Vis spectrophotometer analysis using ICP-OES. Then, by diluting this solution, samples of 190 different concentrations (10, 25, 50 mg/L) were used to 191 149 The preliminary reduction of synthesized silver nanopar investigate the concentration dependence of the antibacte- 192 150 ticles was monitored using UV-visible spectrophotometer rial effect of Ag nanoparticles. 193 151 (Shimadzu UV-1601, Japan) by scanning the absorbance 152 spectra in the range of 300-600 nm. All sample solutions Evaluation of antibacterial activity of synthesized 194 153 were diluted using a ratio of 1:10. silver nanoparticles 195

154 Electron microscopic study The synthesized silver nanoparticles were tested for their 196 antibacterial activity by using the disk diffusion method. 197 155 SEM analysis of the synthesized silver nanoparticles was The cultures of Staphylococcus aureus (ATCC-25923) and 198 156 done using a Hitachi S-4500 SEM machine (Japan). A thin Escherichia coli (ATCC-39403) were obtained from 199 157 layer of gold was used to coat the samples through vacuum American Type Culture Collection Center, USA. S. aureus 200 158 evaporation so that the nanoparticles conducts evenly and and E. coli were grown on Mueller-Hinton agar medium. 201 159 provides a homogeneous surface for analysis and imaging The disk diffusion was performed by placing different 202 160 on an aluminium slab. The elemental analysis was per types of disks including bark extract (50 pL), synthesized 203 161 formed using EDX (JEOL-JSM-5800LV, Tokyo, Japan) silver nanoparticles (50 pL/10, 25, 50 mg/L), standard 204 which is an attachment to the SEM. The sample powder of antibiotic (erythromycin 50 pL) and synthesized' silver 205 m AgNPs was compressed to form tablets before analysis nanoparticles with standard antibiotic on the surface of the 206 164 with EDX spectrum. agar in plates and incubated for 24 h at 37 °C. The zones of 207 inhibition were measured by the Hi-Media scale. 208 165 XRD analysis

166 The dried synthesized silver nanoparticles were analysed Results and discussion 209 167 using an X-ray diffractometer (D8 Bruker, Germany) with 168 Cu Ky radiation in the range of 30° < 2 0 < 90° at 40 kV Visual analysis 210 169 and 40 mA. Afzelia quanzensis bark extract when incubated with 211 170 FTIR analysis AgN03 solution under dark conditions changed its colour 212 from colourless to light reddish and finally to reddish 213 171 FTIR (PerkinElmer, US) spectra were obtained at room brown. The colour of the filtrate changed to intense browning! 14 172 temperature in the spectral range between 480 and after 120 min of incubation (Fig. 2b, c). The centrein g ! 15

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Fig. 2 Optical photograph of 1 mM AgN03 solution (a) filtrate with silver ions at the beginning of reaction (b) and after 120 min of reaction (c)

Fig. 3 The UV-visible absorption spectra of synthesized AgNPs. The inset shows the change in SPR as a function of time

Wavelength (nm) — 'A fter 140 min

216 AgNOi solution (without Afzelia quanzensis bark extract) From Fig. 3, it can be observed that the plasmon band 229 217 showed no change in colour (Fig. 2a). The possible was symmetric, indicating that the solution does not have 230 218 chemical reactions involved in the preparation of the much aggregated particles [27]. The evolution of an 231 p 9 AgNPs can be represented as: absorption spectra for the AgNPs shows an increasingly 232

Stirring at room temperature sharp absorbance at 427 nm with increase in time, which 233 Ag(aq) + Afzelia quanzensis [Stirring for J 20 min at 70° C] steadily increased in intensity as a function of reaction time 234 (i) without showing any shift of the maximum wavelength [2]. 235 [Ag {Afzelia quanzensis)]H This phenomenon may be linked to polarization of the free 236 221 [Ag (Afzelia quanzensis)}++ R-CHO conduction electrons with respect to the much heavier ionic 237 —> [Ag (Afzelia quanzensis)] + R-COOH (2) core of AgNPs, resulting in electron dipolar oscillation 238 after exposure of AgNPs to light [12], For quality assur 239 223 ance and comparison, AgNO:i solution in deionized water 240 224 UV-Vis spectra analysis showed no absorption peak at the same wavelength range 241 (data not shown). In the present bark extract/Ag investi 242 225 The development of metal silver nanoparticles in aqueous gation, the reaction mixtures showed a single SPR band 243 226 solution was described by UV-Vis spectroscopy. The revealing the spherical shape of AgNPs which is in good 244 227 absorption spectra of AgNPs formed at different durations agreement to the Mie’s theory [28]. Consequently, this 245 228 are shown in (Fig. 3).

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0 2 4 6 8 10 12 14 lAgNO,] / mM

Fig. 5 Effect of AgN03 concentration (error bar ±SD and n = 3) Fig. 4 Effect of different pH on nanoparticle production (error bar ±SD and n = 3)

246 validates the application of Afzelia quanzensis bark extract 247 as a precursor for AgNPs synthesis.

248 Study for the influence of different parameters

^4 9 The biosynthesis of AgNPs is affected by a variety of 450 factors (substrate concentration, electron donor, incubation 251 time, pH, temperature, buffer strength, etc.) which control 252 the shape and size as well as achieving the monodispersity 253 in solution phase. In this study, the effects of pH, AgN03 254 concentrations, different quantities of bark extract, and 255 different temperatures were investigated. Fig. 6 Effect of different quantities of Afzelia quanzensis bark 256 Effect of pH extract (error bar ±SD and n = 3)

257 The size and morphology of nanoparticles are mainly 258 affected by the pH of solution [29-31]. As shown by UV- There was an increase in synthesis of AgNPs with 278 259 Vis spectroscopy (Fig. 4), when the pH of the reaction respect to Ag+ ion concentration in the range 1-9 mM. 279 260 mixture was increased, an increase in absorbance was However, the absorbance was found to decrease at con 280 261 observed. This might be due to the increase in production centrations greater than 9 mM. Comparable results were 281 262 of colloidal silver nanoparticles and reduction rate. Visual obtained for the synthesis AgNPs using Pinus eldarica bark 282 263 observation showed the amount of nanoparticles to be pH extract [31]. Consequently, 9 mM was used for further 283 264 value dependent. The reaction mixture colouring acceler studies. 284 265 ated when the pH was increased. At acidic pH, large-sized 9 * silver nanoparticles were observed, whilst at higher pH Effect of different quantities o f Afzelia quanzensis bark 285 267 highly dispersed, small-sized nanoparticles were formed. extract 286 268 The results are in agreement with those reported in litera 269 ture [32-34], Control experiments (AgN03 solution incu The production of AgNPs was monitored as a function of 287 270 bated at different alkaline pH 8, 9, 10) showed no synthesis different amounts of Afzelia quanzensis bark extract 288 271 of nanoparticles. amount. Figure 6 shows the effect of extract amount on 289 AgNPs production. The increase in the extract amount 290 272 Effect of AgNO3 concentration from 2 to 10 mL caused a considerable increase in peak 291 absorbance in UV-Vis spectrum. The increase in the 292 273 The concentration of AgN03 which might be converted to amount of soluble phytochemical reducing agents in the 293 274 a final product is one of the important measures required to extract would mean more Ag+ ion reduction, and subse 294 275 make the reaction more economical and efficient [31]. The quently more nanoparticle production. Furthermore, a 295 276 effect of AgN03 concentration on synthesis of AgNPs is decrease in amount of Ag nanoparticles was observed due 296 277 shown in Fig. 5. to an increase in extract amount above 10 mL. 297

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T C’C)

Fig. 7 Effect of reaction temperature (error bar ±SD; and n = 3)

298 Effect of temperature

299 The role of temperature on the reaction rate from 30 to 300 70 °C was investigated. An increase in absorbance was 301 noted as the temperature increased (Fig. 7). The enhanced ^ 0 2 rate of synthesis of AgNPs might be due to the reaction R )3 temperature increasing the kinetic energy of the reacting 304 molecules thus more Ag+ ions were in collision with the 305 reducing molecules of the extract. The maximal synthesis 306 of AgNPs was achieved at 70 °C.

307 Characterization

308 Electron microscopic study

309 The SEM images of the AgNC>3 (Fig. 8a) and synthesized 310 silver nanoparticles (Fig. 8b) were clearly noticeable. The 311 size of the silver nitrate particles used as control in this 312 study was greater than 1 000 nm size (Fig. 8a); whereas, 313 the synthesized silver nanoparticles measured 10-80 nm in Fig. 8 Scanning electron microscopic observation of a silver nitrate 314 size (Fig. 8b). Similar to our study, the same pattern of and b synthesized 315 silver nanoparticles were also reported [24]. The EDAX 316 spectroscopy results confirmed the significant presence of clearly illustrates that the silver nanoparticles formed in 330 317 61.66 % silver, 30.47 % carbon and oxygen 7.87 % this present synthesis are crystalline in nature. There were 331 (Fig. 9). Metallic silver nanocrystals generally show a no other corresponding peaks observed from the XRD 332 § 1 typical optical absorption peak approximately at 3 keV due pattern which showed that the formed AgNPs have a high 333 320 to surface plasmon resonance [35]. The weak signals at purity. Previous studies have also reported the crystalline 334 321 0.25 and 0.50 keV were are for carbon and oxygen, nature of biosynthesized AgNPs using different plant 335 322 respectively, which might arise from the functional com extracts [3, 9, 22, 23, 36], The average crystalline size of 336 323 pounds present in the aqueous extract. silver nanoparticles synthesized using Afzelia quanzensis 337 bark extract can be calculated using the Scherrer equation 338 324 XRD: purity and crystalline nature of AgNPs [25]: 339 KX 325 The phase of the vacuum dried nanoparticles was investi D = -,(3) 326 gated by XRD and corresponding patterns are shown in [ICosO' 327 Fig. 10. However, silver nanoparticles have shown clear where D is the crystallite size of AgNPs, X is the wdve-EBS>41 328 peaks of cubic phases (JCPDS No. 03-0921) at 38.1 (111), length of the X-ray source (0.1541 nm), (i is the full Width 342 329 42.9 (200), 65.5 (220) and 77.9 (311). XRD pattern thus at half maximum of the diffraction peak, K is the Scherrer 343

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F u ll seal® counts: 1266 Bns«(2681)_(»t1

Fig. 9 The EDX spectrum for AgNPs a

4000 3500 3000 2500 2000 1500 1000 500 Wavenumber (cm'1) • t Fig. 11 FTIR spectra of Afzelia quanzensis extract (a) and synthe sized AgNPs (b)

and alkyl group decreased in intensity, and shifted to 363 2937 cm” 1. The disappearance of the peaks at 1445 and 364 1264 cm” 1 and formation of new intense peak at 365 1384 cm” 1, signify the involvement of the secondary 366 constant with a value from 0.9 to 1, and 0 is the Bragg amines in the reduction process. The shift of the band from 367 angle. The average crystalline size was 19.8 nm. The 1635 to 1637 cm” 1 is attributed to the binding of (NH) CO 368 obtained average crystalline size coupled with the presence group with nanoparticles. It can be concluded from the 369 of structural peaks in XRD patterns clearly illustrated that FTIR study that the carboxyl (-C=0) and amine (N-H) 370 the AgNPs synthesized were nanocrystalline in nature. groups in Afzelia quanzensis bark extract were mainly 371 involved in reduction of Ag+ ions to Ag° nanoparticles. 372 FTIR analysis Stability studies 373 The FTIR spectra of Afzelia quanzensis extract and syn thesized AgNPs were examined (Fig. 11). The strong band A vital aspect in colloid chemistry is how nano-scale par 374 at 3455 cm-1 on the Afzelia quanzensis extract is charac ticles are stabilized in their reaction media as smaller 375 teristic of N-H and O-H stretching vibrations [35], The particles are prone to agglomeration. The stability of-the 376 characteristic absorption band at 2933 cm” 1 is due to alkyl AgNPs in solution was checked using UV-Vis spec 377 chains. The FTIR spectra also show bands at 1637 and troscopy by observing changes in the SPR band peak'[37]. 378 1445 cm” 1 identified as amide I and amide II which arise Figure 12 shows UV-Vis spectra periodically obtained 379 due to carbonyl (C=0) and amine (-NH) stretching over 60 days. The AgNPs produced from bark extract were 380 vibrations in the amide linkages of the proteins, respec observed to be very stable in solution when monitored 381 tively. The peaks at 1264 and 1060 cm” 1 may be due to a periodically over a period of 40 days; with no evidence of 382 carboxylate group (COO”) and phosphate group, respec flocculation or change in SPR, measured at 427 nm. 383 tively. After reduction of AgN03, the band characteristic of However, after 50 days the surface plasmon resonance 384 N-H and O-H stretching vibrations shifted to 3424 cm” 1 absorption band decreased and can be attributed to 385

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Fig. 12 Stability studies

Fig. 13 Graphical 30 representation of inhibition zones by plant extract (a), 10 mg/L synthesized AgNPs (b), 25 mg/L synthesized AgNPs (c), 50 mg/L synthesized AgNPs (d), erythromycin (e), erythromycin plus 10 mg/L synthesized » AgNPs (/), erythromycin plus 25 mg/L synthesized AgNPs (g), erythromycin plus 50 mg/L synthesized AgNPs (h), for a Escherichia coli, b Staphylococcus aureus (error bar ±SD and n = 3) Escherichia coli Staphylococcus aureus Name of organism

386 destabilization of AgNPs in solution due to aggregation. the phytofabricated silver nanoparticles were found to be 408 387 Between 50 and 60 days secondary peaks appear at longer more effective against Escherichia coli as compared to that 409 388 wavelength which is a sign of destabilization. of Staphylococcus aureus. From Fig. 13a, b, it can be 410 confirmed that combining the antibiotics with AgNPs 411 389 Evaluation of antibacterial activity of synthesized resulted in a greater bactericidal effect on test pathogens 412 390 silver nanoparticles than either of the antibacterial agents used alone. Inhibition 413 of Escherichia coli might have been facilitated by silver’s 414 391 The antimicrobial activity of bark extract (a), synthesized high affinity for phosphorus and sulphur which are part of 415 392 AgNPs of different concentrations (b-d), the antibiotic the cell membranes of Gram-negative bacteria [38].'.The 416 393 (erythromycin) (e) and antibiotic plus different concentra sulphur and phosphorus reacts with AgNPs causing dys 417 tions of synthesized AgNPs (f-h) were evaluated by dif function of enzymes on bacteria cell wall and also’it dis 418 T95P 4 fusion method. The bacterial cultures used were Gram turbs DNA’s moieties process thereby denying replication 419 396 positive bacteria, i.e. Staphylococcus aureus and Gram [39]. On the other hand, the cell wall of Gram-positive 420 397 negative bacteria, i.e. Escherichia coli. The increase in bacteria is much more rigid due to the presence of a thick 421 398 zone of inhibition of silver nanoparticles at different con peptidoglycan layer, which is superficial to the cell mem 422 399 centrations (10, 25, 50 mg/L) compared with antibiotic for brane hence a difference in activity observed. 423 400 both bacterial cultures (Fig. 13a, b) demonstrated the lesser 401 antibacterial potential of silver nanoparticles with that of 402 antibiotics. The zone of inhibition also increased with Conclusion 424 403 AgNPs concentration in both cultures. The bark extract 404 exhibits the highest activity against E. coli than S. aureus. The silver nanoparticles were green synthesized using bark 425 405 The obtained results support, at least in part, the use of this extract of Afzelia quanzensis. The method represents an 426 406 plant as traditional medicine against Gram-negative bac example of clean, nontoxic and eco-friendly method for 427 407 teria due to the presence of phytochemicals. Furthermore, obtaining silver nanoparticles. Colour changes occur due to 428

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429 surface plasmon resonance during the reaction with Afzelia 11. Ghorbani HR (2013) Biosynthesis of silver nanoparticles using 485, 430 quanzensis bark extract resulting in the formation of silver Salmonella typhirium. J Nanostruct Chem 3(29): 1-4 486 12. Swarup R, Kumar DT (2014) Biosynthesis of silver nanoparticles 487 431 nanoparticles, which was confirmed by XRD, FTIR, UV- by Aspergillus foetidus: optimization of physicochemical 488 432 Vis spectroscopy, and EDAX. The silver nanoparticles parameters. Nanosci Nanotechnol Lett 6(3): 181 489 433 were found to be stable in water for 40 days. The zone of 13. Prasad K, Anal KJ, Kulkami AR (2007) Lactobacillus assisted 490 434 inhibition test showed that the synthesized nanoparticles synthesis of titanium nanoparticles. Nanoscale Res Lett 2: 491 248-25 0 4 92 435 have some antibacterial activity. Further studies will be 14. 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