In this present study we focused on the one pot green synthesis of AgNPs using a leaf bud extract of R. mucronata
which is simple, convenient and eco-friendly. The green synthesis of AgNPs has been investigated as an alternative to chemical and physical ones. A conical flask containing the extract of mangrove leaf bud R. mucronata
and aqueous silver nitrate (1
mM) was kept at 121°C for 5 minutes and the resulting solution was turned to yellowish brown, this color change was due to excitation of surface plasmon vibrations in the metal nanoparticles. This visible observation indicates the reduction of the Ag+
ions and the biosynthesis of AgNPs. This observation was further reconfirmed by UV-visible spectrum and XRD analysis. The (Figure ) shows the UV visible spectra recorded from the reaction medium which were characterized by UV–vis spectrophotometer; it is well observed that the silver surface plasmon resonance band occurs at 426
UV-Visible spectra of Silver nanoparticles synthesized from aqueous leaf bud extract of R.mucronata.
In metal nanoparticles such as in silver, the conduction band and valence band lie very close to each other and through these electrons move freely. These free electrons give rise to a surface plasmon resonance (SPR) absorption band, occurring due to the collective oscillation of electrons of AgNPs in resonance with the light wave. Classically, the electric field of an incoming wave induces polarization of the electrons with respect to much heavier ionic core of AgNPs. As a result a net charge difference occurs, which in turn acts as a restoring force. This creates a dipolar oscillation of all the electrons with the same phase. When the frequency of the electromagnetic field becomes resonant with the coherent electron motion, a strong absorption takes place, which is the origin of the observed color, which was yellowish brown in our observation. This absorption strongly depends on the particle size, dielectric medium and chemical surroundings. The UV/Vis absorption spectra of the silver nano particles dispersed in R. mucronata
extracts is shown in Figure . The absorption peak (SPR) was obtained in the visible range at 426
nm, the broad spectra is due to the size (4
nm) and shape (spherical) of the biosynthesized AgNPs which were documented by High Resolution TEM micrograph.
The result of XRD patterns (Figure ) showed AgNPs are reflected in the 2θ on 38.1, 44.3 and 64.4 that respect the Bragg model of diffraction. The peak corresponding to the 2θ
38.1°(111), 44.3°(200) and 64.4°(220) of the sample respects the JCPDS 652871 and it was confirmed the crystalline nature of the AgNPs, obtained biogenic AgNPs which indexed the planes 111, 200 and 220 of the face center cubic nature of silver nanopartilces. The angle measured by this method corresponds to the lattice planes were observed the face center cubic structures of silver matched with the database of Joint Committee on Powder Diffraction Standard [32
]. Thus XRD patterns clearly showed that the AgNPs formed by the reduction of Ag+
ions by the extract of R. mucronata
with aqueous silver nitrate are crystalline in nature.
XRD patterns of the silver nanoparticles synthesized from aqueous leaf extract of R. mucronata. (Inset showing the standard XRD pattern for silver: JCPDS 652871)
FTIR measurements were carried out to identify the possible biomolecule responsible for the reduction of the Ag+
ions and capping of the bioreduced AgNPs synthesized by mangrove leaf bud extract R. mucronata.
The FTIR spectrum shows peaks at 2368, 1618, 1540, 1384, 1325, 1265, 1053 and 788
. Curve of the mangrove leaf bud extract of R. mucronata
(Figure a) resulted a multiple broad peaks at 2368
corresponding to N-H stretching of any ammonium ions; the medium band at 1618
corresponding to stretching of C = N; the stronger band at 1540
corresponding to N-O stretching of nitro compounds. The weaker band at 1384
corresponding to N-O stretching of nitro compounds; the broad band at 1092
corresponding to C-X stretching of fluoroalkanes; the strong band at 788
corresponding to C-H stretching of aromatic benzene.1325
relating the stretching of amide.
(a) FT-IR spectra of plain R. mucronata. (b) Biologically synthesized silver nanoparticles using R. mucronata.
Curve of AgNPs biosynthesized using the mangrove leaf bud extract of R.mucronata
(Figure b) resulted a broad band at 3396
corresponding to O-H stretching of high concentration of alcohols or phenols; the multiple broad band at 2375
corresponding to N-H stretching of ammonium ions; the medium band at 1636
corresponding to stretching of C = N; the weak to strong band at 1444
corresponding to C-C stretching of aromatic C = C. the medium band at 1264
corresponding to C-O stretching of any carboxylic acids. A band shift from 1094
corresponding to C-X stretching of ordinary fluroalkanes to strong band at 798
corresponding to C-H stretching of benzene of aromatic compounds. The disappearance of 1325
relating the stretching of amide which cause the capping of AgNPs.
FTIR spectroscopy from the absorption of IR radiation through resonance of non-centro symmetric (IR active) modes of vibration and is a useful tool for quantifying secondary structure in metal nanoparticle–biomolecules interaction. Figure a-b confirmed that the N-H stretching vibration of primary amines and C-N stretching and over lapping of aliphatic amines has the stronger ability to bind metal, so that the secondary metabolites from mangrove leaf bud extract of R.mucronata could most possibly form a coat covering the metal nanoparticles (i.e. capping of silver nanoparticles) to prevent agglomeration of the particles and stabilizing in the medium. This evidence suggests that the biological molecules could possibly perform the function for the formation and stabilization of the silver colloids in aqueous medium. The exact mechanism leading to the reduction of metal ions is yet to be elucidated for mangrove leaf bud extract of R.mucronata.
The size and shape of the biosynthesized AgNPs were documented by High Resolution TEM micrograph (Figure a, b, and c). The enlarged TEM graphs helped to plot a histogram (Figure d), which revealed the particle distribution according to the size. The TEM micrograph of the AgNPs confirmed that the particles were likely to be spherical in shapes with a size range from 4 to 26
nm. However a maximum number of particles were observed at 4
nm in size. The selected area electron diffraction (SAED) pattern of the nanoparticles explained the face centered cubic (fcc) crystalline structure of silver with different diffracting index (Figure c).
Figure 4 TEM images of silver nanoparticle in two magnifications.(a) 50nm (b) 20nm (c) SAED pattern and (d) Histogram showing the particle size distribution.
Antimicrobial effects of one pot green synthesized AgNPs against Pseudomonas florescence, Proteus spp.
and Flavobacterium spp.
of marine aquatic pathogen were confirmed by the circular inhibition zone formed around the well impregnated with different concentration of AgNPs. The antimicrobial effect varies according to the species and the effect was higher at 75
μg/μL concentration of AgNPs. A maximum zone was recorded as 16, 14 and 14
mm for Pseudomonas florescence, Proteus spp.
, and Flavobacterium spp.
respectively at 75
μg/μL concentrations. A maximum zone was also recorded as 17, 14 and 15
mm for Pseudomonas florescence, Proteus spp.
, and Flavobacterium spp.
respectively when treated with Chloramphenicol (1
ppm) as a control. No zone formations were observed for the plant extract alone. Thus the zone observed in control was significantly equal at 75
μg/μL concentration that confirms the antimicrobial effect of biosynthesized AgNPs to control the marine microbial pathogens and the Figure shows the rate of inhibition against marine ornamental fish pathogens. The antibacterial effect of AgNPs inside bacterial cells will form a strong association with bacterial cellular components. Once inside the cell, nanoparticles would interfere with the bacterial growth signaling pathway by modulating tyrosine phosphorylation of putative peptide substrates critical for cell viability and division. Inside a bacterium, nanoparticles can interact with DNA, thus losing its ability to replicate which may lead to the cell death. Interaction between such nanoparticles and the cell wall of bacteria would be facilitated by the relative abundance of negative charges on the gram-negative bacteria, which was affable to the fact that growth of gram-negative bacteria was more profoundly affected by the AgNPs than that of the gram-positive organisms [32
] Thus Figure represents the maximum zone formation was found in Pseudomonas fluorescence
highest zone formation in 16
mm at 75
mm at 50
μg/μL and 12
mm in 25
Graph showing the rate of inhibition of silver nanoparticles against marine ornamental fish pathogens. (Inset showing the antimicrobial activity against (A) Proteus spp., (B) Pseudomonas florescence and (C) Flavobacterium spp.