Journal of Cluster Science (2019) 30:1103–1113

https://doi.org/10.1007/s10876-019-01571-2(0123456789().,-volV)(0123456789().
,- volV)

ORIGINAL PAPER

Green Synthesis of Sliver Nanoparticles Using Avicennia officinalis

and Xylocarpus granatum Extracts and In vitro Evaluation
of Antioxidant, Antidiabetic and Anti-inflammatory Activities
Swagat Kumar Das1 • Supriti Behera1 • Jayanta Kumar Patra2 • Hrudayanath Thatoi3

Received: 8 March 2019 / Published online: 27 April 2019

 Springer Science+Business Media, LLC, part of Springer Nature 2019

Abstract
The present study involves biosynthesis of AgNPs using Avicennia officinalis and Xylocarpus granatum mangrove plants
along with evaluation of their potential biomedical applications. The synthesized AgNPs were characterized by UV–Vis
spectroscopy, FTIR analysis, scanning electron microscope, particle size analyzer, X-ray diffraction (XRD). The syn-
thesized AgNPs showed absorption maxima at 470 nm for X. granatum (XG-AgNPs) and 420 nm for A. offcinalis (AO-
AgNPs) which corresponds to their respective surface Plasmon resonance. The FTIR analysis reveals capping of phenolic
groups providing stability of synthesized AgNPs. The morphology of silver nanoparticles was confirmed by SEM tech-
nique. The dynamic light scattering study (DLS) also confirmed the size distribution of synthesized AgNPs. XRD peaks at
2h range of 20–70o corresponds (111), (200) and (220) reflection planes indicating the structure of metallic silver. AO-
AgNPs exhibited better DPPH scavenging, superoxide and protein denatuaration activity with IC50 values of 0.14, 0.32 and
0.21 mg/ml. However, XG-AgNPs exhibited better a-amylse and a-glucosidase inhibition potential as compared to AO-
AgNPs. It could be concluded that X. granatum bark extracts and A. officinalis leaf extract can be used efficiently for the
synthesis of biologically active silver nanoparticles which could be exploited pharmaceutical applications.

Keywords Silver nanoparticle  Mangrove  Antioxidant  Antidiabetic  Anti-inflammatory

Introduction biomedical properties such as antimicrobial, wound healing

Nanobiotechnology is presently one of the most dynamic
disciplines of research in contemporary science whereby
plants and different plant products are finding an impera-
tive use in the synthesis of nanoparticles (NPs) [1].
Amongst the different NPs, the silver nanoparticle
(AgNPs) have demonstrated improved electrical conduc-
tivity, chemical stability, catalytic activity, and distinct
& Swagat Kumar Das
das.swagat@gmail.com
1
Department of Biotechnology, College of Engineering and
Technology, Ghatikia, Kalinga Nagar, Bhubaneswar,
Odisha 751003, India
2 of greener syntheses of silver nanoparticles, plant mediated
Research Institute of Biotechnology and Medical Converged
Science, Dongguk University, Seoul, Republic of Korea
3 able cost-effective, environmentally friendly alternate
Department of Biotechnology, North Odisha University,
Baripada, Odisha 757003, India
ointments, also for food packaging, medical instruments,
textile coatings, cosmetics etc. [2]. The conventional
techniques used for synthesis of AgNPs such as laser
ablation, gamma irradiation, electron irradiation, photo-
chemical methods, and microwave processing are either
expensive, or labour intensive. Further, chemical methods
usually involves toxic or hazardous solvents and reductants
making these methods unsuitable [3]. Therefore, develop-
ment of simple and environmentally friendly alternative
approaches for preparation of nanoparticles using non-toxic
reagents and cost effectively is highly desirable. In this
context, green synthesis of nanoparticles using microbes,
marine organisms, and plant extracts have become popular
as these methods are biocompatibile, environmental
friendly and economic. Amongst the different approaches
synthesis of AgNPs is considered as a widely accept-
technology for rapid production of AgNPs. Although, the

123
1104
potential of higher plants as source for this purpose is still
largely unexplored. Recently, synthesis of nanoparticles
using mangrove plants has recently got attention. The
mangrove plants are woody, specialized types of trees
growing in brackish wetlands in the tropical and sub-
tropical inter-tidal coastal zones and river deltas. These
mangrove plants possess different class of bioactive sec-
ondary metabolites such as alkaloids, glycosides, ter-
penoids, flavonoids, and other poyphenols that can be
exploited for various therapeutic needs such as antimicro-
bials, antioxidants, anti-inflammatory, antidiabetic, cyto-
toxic etc. [4]. The rich content of polyphenols in the
mangrove plants can be exploited as these polyphenols are
reported to be strong reducing agents and also have a
tendency to adsorb on the surface of NPs and also func-
tionalize the resultant NPs [5].
Considering the vast potentiality of mangrove plants as
source of reducing agents for nanoparticle synthesis, the
present work aims to apply a biological green technique for
the synthesis of silver nanoparticles as an alternative to
conventional methods. In this regard, leaf extract of Avi-
cennia officinalis L. and bark extract of Xylocarpus
granatum J. Koenig (www.theplantlist.org) two medici-
nally important mangrove plants were used for synthesis of
silver nanoparticles (AgNPs). The green synthesized
AgNPs were characterized by several techniques such as
UV–Vis, FT-IR, DLS and XRD, Moreover,the antioxidant,
antidiabetic and anti-inflammatory properties of the syn-
thesized AgNPs were evaluated.
Materials and Methods
Collection of Mangrove Plants and Extraction
The leaves of Avicennia officinalis L. and barks of Xylo-
carpus granatum J. Koenig mangrove plants were collected
from Mahanadi Delta mangrove forest of Odisha. The plant
specimens were identified by Prasanna Kumar Nayak,
Herbarium keeper, Integrated Coastal Zone Management
Project (ICZMP), Forest Department, Govt. of Odisha.
After collection, the plant samples were cleaned in tap
water, air dried under shade at room temperature cut into
small pieces and homogenized into a fine powder using
mechanical grinder. About 10 g of powdered leaf sample
of A. officinalis and bark sample X. granatum were taken
separately in two 500 ml conical flask containing 100 ml
of sterile distilled water and was boiled for 20 min. After
cooling the extracts were filtered through Whatman No.1
filter paper and the filtrate was evaporated to dryness. The
dried leaf and bark extracts of A. officinalis and bark
sample X. granatum respectively were used for the syn-
thesis of silver nanoparticle.
123
S. K. Das et al.
Green Synthesis of Silver Nanoparticle (AgNPs)
The silver nanoparticles (AgNPs) were synthesized fol-
lowing the method of Rao and Savithramma [6] with some
modification. Briefly, the leaf extracts of A. officinalis and
bark extracts of X. granatum and AgNO3 solution (10 mM)
were taken in separated conical flask in 1:19 ratio and
allowed to stand for 6 h at room temperature. The reduc-
tion of silver nitrate to AgNPs was confirmed by change in
colour of the solution. The extract contents were then
centrifuged at 10,000 rpm for 20 min. The Ag nanoparti-
cles were washed with distilled water for 2–3 times to
remove any unbound phytoconstituents. Then the prepared
AgNPs were dried at 50 C in a dry oven and stored at 4 C
till further use.
Characterization of Nanoparticles
The mangrove plants mediated synthesized AgNPs were
characterized by different analytical techniques such as,
UV–visible spectroscopy, Fourier transformation infrared
spectroscopy (FT-IR), dynamic light scattering (DLS) and
X-ray Powder diffraction (XRD).
UV–visible Spectral Analysis
The UV–Vis spectrophotometer UV–117 (SystronicsTM)
was used to measure UV–Vis absorbance spectra of AgNPs
synthesized by A. officinalis leaf and X. granatum bark
extracts. The absorbance was measured in 400–800 nm
range with a 1 nm step and the characteristic peaks were
detected. The spectra were recorded at the intervals of
1 min to 10 min and the distilled water was used as a
baseline.
FT-IR Analysis
The interactions of A. officinalis leaf and X. granatum bark
extracts and AgNPs were analyzed with FTIR spec-
troscopy. For FTIR measurements, sample ground with
KBr and pellet was analyzed using an FTIR spectropho-
tometer in the range of 400–4000 cm-1.
SEM Analysis
The field-emission scanning electron microscopy (FESEM)
was used to study the surface morphology of dried samples
of synthesized silver nanoparticles (JEOL, JSM7600F).
Green Synthesis of Sliver Nanoparticles Using Avicennia officinalis and Xylocarpus granatum… 1105

DLS Analysis
where DOD = Abs after 15 min - Abs at 0 min
The dynamic light scattering (DLS) is used to characterize
Antidiabetic Study
the surface charge and the size distribution of the AgNPs
suspended in a liquid [7]. The AgNP solutions after fil-
a-Amylase Inhibition Assay
tration were transferred to the cuvette and the hydrody-
namic radius was measured in the particle analyzer
Amylase activity was assayed using chromogenic 3,5-
(Malvern). All measurements were performed at room
dinitrosalicylic acid (DNS) method [11]. AgNPs of dif-
temperature.
ferent concentrations (0.1 mg/ml to 0.5 mg/ml) were
allowed to incubate with 200 ll of the a-amylase enzyme
XRD Analysis
and 100 ll of 2 mM phosphate buffer (pH 6.9) for 5 min
followed by addition of 0.5% starch solution. After incu-
XRD is used for the phase identification and characteri-
bation for 5 min at 37 C, 400 ll of reaction mixture was
zation of the crystal structure of the silver nanoparticles
removed and then 200 ll of dinitrosalicylic acid reagent
[8]. X-rays penetrate into the nanomaterial and the result-
was added and were boiled for 15 min at 90 C. The
ing diffraction pattern is compared with standards to obtain
reaction mixture was then diluted with 1.8 ml of distilled
structural information. The crystalline phase of A. offici-
water. The absorbance was recorded at 540 nm. The % of
nalis leaf and X. granatum bark extracts mediated syn-
enzyme inhibition was calculated.
thesized Ag nanoparticles were determined by X-ray
diffraction (XRD) using Cu–Ka radiations (k 1.5406 Å)
a-Glucosidase Inhibitory Activity
in 2h range from 20 to 70. The XRD method is suitable to
determine the crystal structures by analyzing the positions
The a-glucosidase was assayed following the method of
and intensities of diffraction peaks.
Apostolidis et al. [12]. a-Glucosidase was premixed with
AgNPs at various concentrations (0.1 mg/ml and 0.5 mg/
Bioactivity Studies ml) and incubated at 37 C for 10 min. 3 mM PNPG in
0.2 M sodium phosphate buffer (pH 7.4) as substrate was
Antioxidant Study
added to the reaction mixture to start the reaction. The
reaction was incubated at 37 C for 20 min and stopped by
DPPH Scavenging Assay The free radical scavenging
adding 2 mL of Na2CO3. The a-glucosidase activity was
activity of the synthesized AgNPs on the stable radical 1,1-
determined by measuring the p-nitrophenol release from
diphenyl-2-picrylhydrazyl was estimated (DPPH) [9].
pNPG at 405 nm. The percentage inhibition of enzyme
Synthesized AgNPs particles at different concentrations
activity was calculated.
(0.1 mg/ml to 0.5 mg/ml) were incubated with 3.0 ml of a
DPPH methanol solution (0.1 mM) in dark for 30 min at
Anti-inflammatory Assay
room temperature followed by measuring the absorbance at
517 nm.
The protein denaturation activity study was carried fol-
lowing the method of Dey et al. [13]. Briefly, the reaction
Superoxide Scavenging Activity The superoxide scav-
mixture consisting of 0.2 ml of egg albumin (from fresh
enging was determined by the nitro blue tetrazolium
hen’s egg), 2.8 ml of phosphate buffered saline (PBS, pH
reduction method [10]. The reaction mixture containing
6.4) and AgNPs of varying concentrations (0.1 mg/ml and
ethylene diaminetetraacetic acid (EDTA) (0.1 mM), nitro
0.5 mg/ml) were incubated at 37 C for 15 min followed
blue tetrazolium (1 mM), Na2CO3 (50 mM) and various
by heating at 70 C for 5 min. After cooling, their absor-
concentrations of synthesized AgNPs (0.1 mg/ml to
bance was measured at 660 nm. The percentage inhibition
0.5 mg/ml) were taken. About 1.5 ml of (3 mM) of
of protein denaturation was calculated by using the fol-
hydroxylamine hydrochloride was added to initiate the
lowing formula:
reaction. The absorbance was measured at the start of the
reaction and after an interval of 15 min at 560 nm. The Percentage of inhibition ¼ 100  ½Vt =Vc  1
control was simultaneous run without plant extract. where Vt = absorbance of test sample, Vc = absorbance of
% of superoxide inhibition control.
DOD of control  DOD of treated sample
¼  100
DOD of control

123
1106
Statistical Analysis
All data were presented as means ± standard deviations
(SD) where appropriate. The means of all the parameters
were examined for significance by analysis of variance
(ANOVA) and in case of significance mean separation
were accomplished by SPSS (Edition 20) statistical soft-
ware package. Differences were considered significant at a
level of p \ 0.05 with Bonferroni post hoc test.
Results and Discussion
The plants are an excellent source of important secondary
metabolites of medicinal importance compared to bacteria
and fungi, their by providing greater scope for the pro-
duction of biocompatible nanoscale particles. The present
study deals with the green synthesis of silver nanoparticle
using extracts of two medicinally important mangrove
plants viz. X. granatum and A. officinalis, their character-
ization and evaluation of their antioxidant, carbohydrate
metabolizing enzyme inhibition and anti-inflammatory
potential.
Green Synthesis and Characterization of AgNPs
Addition of leaf and bark extracts of A. officinalis and X.
granatum to AgNO3 solution resulted in distinct change in
colour of the reaction mixture. The initial yellow AgNO3
solution turned into dark brownish which indicates the
gradual reduction of AgNO3 by mangrove plant extracts
(Fig. 1a, b). The Ag? ion reduction was confirmed by the
apparent transformation in the colour. Under a similar set
a
Before After
b accountable for the reduction of silver ions. The FT- IR
Before After
Fig. 1 Colour change during synthesis of silver nanoparticles using
the extracts of a X. granatum and b A. officinalis
123
S. K. Das et al.
of circumstances, no colour transformation was noticed in
the absence of plant extract. The colour change arises due
to excitation of surface plasmon resonance (SPR) in syn-
thesized AgNPs [1] and corroborated with the earlier
studies [14, 15]. The visual change in colour of the solu-
tions was different for both A. officinalis and X. granatum
extracts may be due to the presence of different phyto-
components present in them which causes a varied range of
bioreduction. Variation in the availability of phyto-
molecules in two different mangrove plant extracts is
responsible for this difference in the absorbance values.
UV–Vis Spectral Analysis
The formation of AgNPs was further confirmed by UV–Vis
spectral study, which is an authentic technique to monitor
the progress of the reaction during the reduction of Ag ?
ions. Typically, UV–Vis spectroscopy serves as a prelim-
inary tool to confirm the formation of NPs. The UV–Vis
absorption in the visible light region of 420–450 nm is an
evidence of the presence of surface plasmon resonance
(SPR) of AgNPs [16]. From the results of UV–Vis
absorption spectra of the synthesized NPs in solution it is
clear that the AgNPs showed their characteristic absorption
maxima between 420 and 490 nm. The present analysis
showed characteristic UV–Vis absorption maxima at
420 nm for AO-AgNPs and 470 nm for XG-AgNPS
(Fig. 2a, b). The difference in the position of absorption
peak of AgNPs may be due to the difference in nanopar-
ticle particle size and shape. This observation is in accor-
dance with the earlier reports on absorption spectra for
AgNPs [15, 17].
FT-IR Analysis
FT-IR spectroscopic analysis of XG-AgNPs and AO-
AGNPs were carried out to investigate the functional
groups present in X. granatum bark and A. officinalis leaf
extracts which may be responsible for synthsis silver
nanoparticles and surface capping. The FT-IR spectra of
XG-AgNPs shown in Fig. 3a reveals the possible biomo-
lecules present in the bark extract of X. granatum which is
spectrum of XG-AgNPs shows absorption peaks at
3441.37, 1621.11 and 1384.2 cm-1. The broad band at
3434.59 cm-1 corresponds to the strong stretching vibra-
tions of hydroxyl group (–OH) of phenolic compounds that
suggests the binding of silver ions with hydroxyl group
(OH). The intense bands at 1621.11 can be attributed to
C=C and 1384.20 cm-1 from C–H stretching which may
be due to ring stretching vibration of heterocyclic com-
pound. Similarly, the FT-IR spectra of the AO-AgNPs was
recorded in order to identify the functional groups present
Green Synthesis of Sliver Nanoparticles Using Avicennia officinalis and Xylocarpus granatum…
Fig. 2 UV–Vis spectroscopy
analysis of silver nanoparticles
using the extracts of a X.
granatum and b A. officinalis
in the aqueous extract of A. officinalis leaf involved in the
synthesis and stabilization of silver nanoparticles (Fig. 3b).
The FT-IR spectra showed absorption peaks at 3436.88,
1631.3, 1384.35, 1114.21 and 825.14 cm-1. A broad peak
at 3436.88 cm-1 strongly suggests the binding of silver
ions with hydroxyl group (OH). The other four bands at
1631.3, 1384.35 and 1114.21 825.14 cm-1 were due to
C=C, C–H, C–O functional groups. The bioactive com-
pounds present in the X. granatum and A. officinalis
extracts might have been responsible for the reduction and
stability of the silver nanoparticles.
SEM Analysis
The FESEM study provided further information on surface
morphology of XG-AgNPs and AO-AGNPs. The FESEM
revealed that both XG-AgNPs and AO-AGNPs are poly-
dispersed (Fig. 4a, b). The clusters of silver nanoparticles
were also observed which may be due to aggregation of
1107
nanoparticles. It has been reported elsewhere that aggre-
gation of nanoparticles can be induced during sample
preparation and evaporation of solvent which may be the
possible reason behind aggregation of XG-AgNPs and AO-
AGNPs [18].
DLS Analysis
DLS is a very efficient and effective tool in analyzing
quantitative size distributions and quantity of monodis-
persity in colloidal solutions. The differential intensity,
number related to particle size distributions of the A.
officinalis and X. granatum extracts mediated AgNPs were
obtained from DLS study (Fig. 5a, b). The DLS study
demonstrated that X. granatum bark mediated silver
nanoparticle (XG-AgNP) particle ranged between 20 and
1000 nm with the average particle intensity at 98.77 nm.
Similarly, the population of particle for A. officinalis leaf
mediated silver nanoparticle (AO-AgNP) varied between
123
1108 S. K. Das et al.

79
a 78

77

76

75

74
%T

73

72

71 1621.11cm-1, 71.53%T

70
1384.20cm-1, 70.45%T

69

68

67 3441.37cm-1, 66.36%T

66
4000 3500 3000 2500 2000 1500 1000 500 400

cm-1

b 78
75

70
825.14cm-1, 72.50%T
1114.21cm-1, 68.98%T 613.68cm-1, 72.56%T
65

60 1631.30cm-1, 63.66%T

55

50
%T

45 3436.88cm-1, 47.30%T

40

35

30

25
1384.35cm-1, 18.78%T
20
17
4000 3500 3000 2500 2000 1500 1000 500 400

cm-1

Fig. 3 FT-IR analysis of silver nanoparticles using the extracts of a X. granatum and b A. officinalis

50 and 1000 nm with the average particle intensity at
181.4 nm. This observation is in accordance with the ear-
lier DLS study for AgNPs [14].
XRD Analysis
The X- ray diffraction technique is used to analyze the
metallic nature of particles. The X-ray diffraction (XRD)
pattern of the X. granatum and A. officinalis extract
mediated AgNPs are shown in Fig. 6a, b. The XRD
pattern shows peak in the whole spectrum of 2h values
ranging from 20o to 70o. The pattern of XG-AgNPs
showed three distinct characteristic peaks at 38.09, 46.25
and 64.55 in 2h range which can be indexed to the (111),
(200) and (220) reflection planes. Similarly, three dis-
tinct peaks were observed in AO-AgNPs at 38.21, 44.38
and 64.58 which were indexed as (111), (200) and (220).
The reflection planes obtained from XG-AgNPs and AO-
AgNPs data obtained were matched with the database of
Joint Committee on Powder Diffraction Standards
(JCPDS file No.04-0783) and were interpreted for the
structure of metallic silver with face-centered cubic
structure of AgNPs Comparison of the sharp peaks of the
obtained data in the present study with the standard,
clearly indicate the crystalline nature of the synthesized
nanoparticles which is in nano regime and agreement
with the earlier reports [16, 17]. In the above XRD
profile, apart from normal peaks, some additional and
unknown peaks were also noticed at the vicinity of the
characteristic peaks of Ag, which might have resulted
due to the presence of bioorganic compounds of the
extract that act as capping agent in stabilizing the
nanoparticle [19].
Bioactivity Studies
X. granatum and A. officinalis extract mediated Silver
nanoparticles were tested for their possible therapeutical
potential for antioxidant, anti-diabetic anti-inflammatory
activities.

123
Green Synthesis of Sliver Nanoparticles Using Avicennia officinalis and Xylocarpus granatum…
Fig. 4 FESEM images of silver nanoparticles using the extracts of
a X. granatum and b A. officinalis
Antioxidant Activity
DPPH is a stable and well-known free radical based on the
reduction of accepting hydrogen or electron from donors.
The DPPH reducing ability of the AgNPs was assessed by
observing colour change. When AgNPs added to DPPH
solution, change in colour occurred due to the scavenging
of DPPH by donation of hydrogen atom to stable the DPPH
molecule [20]. The DPPH free radical scavenging potential
of the synthesized XG-AgNPs and AO-AgNPs were eval-
uated at 0.1, 0.2 and 0.5 mg/ml and the result is presented
in Fig. 7a. The study revealed that both XG-AgNPs and
AO-AgNPs possess free radical scavenging activity in a
dose dependent manner. The DPPH scavenging activity
ranged between 25.6 and 74.85% where as AO-AgNPs
activity ranged between 31.6 and 100%. The study further
1109
demonstrates that AO-AgNPs was effective as compared
with XG-AgNPs in scavenging DPPH radicals as the for-
mer was having an IC50 value of 0.14 mg/ml in comparison
to the later one having 0.25 mg/ml (Table 1). The standard
antioxidant compound BHT under similar condition could
scavenge DPPH radical with IC50 value of 0.04 mg/ml.
Earlier study have shown that another mangrove plant
Sonneratia apetala mediated AgNPs could scavenge DPPH
radical at 65.31% at 100 lg/ml [21]. Similarly, it has also
been reported that AgNPs synthesized using extracts of two
mangrove plants Heritiera fomes and S. apetala could
scavenge DPPH radicals with IC50 values \ 0.2 mg/ml
[14] which is in accordance with the present study.
The antioxidant potential of the synthesized AgNPs was
evaluated in terms of the superoxide radical scavenging
activity also. Superoxide radicals were generated in vitro
chemically by hydroxylamine hydrochloride, which redu-
ces nitro blue tetrazolium (NBT) and forms a chromophore,
diformazan [10]. The superoxide radical scavenging
activity of XG-AgNPs and AO-AgNPs were found to be
dose-dependent (Fig. 7b). The superoxide scavenging
activity XG-AgNPs and AO-AgNPs at 0.1 mg/ml was
18.55 and 17.02% where as at 0.5 mg/ml the activities
were 48.19 and 71.52%. The result of the present study
demonstrated that the IC50 values of XG-AgNPs, AO-
AgNPs and standard antioxidant compound Ascorbic acid
were 0.49, 0.32 and 0.08 mg/ml (Table 1). Hence, AO-
AgNPs showed better superoxide scavenging potential as
compared to XG-AgNPs. The antioxidant potential of
AgNPs could be attributed to functional groups viz. phe-
nolics adhered to them which were originated from the X.
granatum and A. officinalis extracts.
Antidiabetic Activity
The inhibition of carbohydrate metabolizing enzymes like
a-amylase and a-glucosidase is considered as a useful tool
to assess the in vitro antidiabetic potential of different
compounds [22]. In the presents study, both XG-AgNPs
and AO-AgNPs were evaluated for their inhibition poten-
tial against carbohydrate metabolizing enzymes like a-
amylase and a-glucosidase at three different concentration
viz. 0.1, 0.2 and 0.5 mg/ml concentrations. The study
revealed that both XG-AgNPs and AO-AgNPs were able to
inhibit a-amylase and a-glucosidase enzymes in a dose
dependent manner (Fig. 7c, d). The a-amylase inhibition
potential of XG-AgNPs varied between 35.51 and 89.5%
whereas for AO-AgNPs, it ranged between 18.21 and
76.13%. From the results of the a-amylase inhibition study,
it can be concluded that XG-AgNPs possessed better
123
1110
Fig. 5 Dynamic Light
Scattering analysis of silver
nanoparticles using the extracts
of a X. granatum and b A.
officinalis
inhibition activity as compared to AO-AgNPs as the former
is having an IC50 value of 0.19 mg/ml and the latter with
0.28 mg/ml. The a-glucosidase inhibition study also
demonstrated that XG-AgNPs possessed better inhibition
activity as compared to AO-AgNPs as indicated by their
IC50 value. The IC50 values of XG-AgNPs and AO-AgNPs
were recorded as of 0.13 and 0.15 mg/ml respectively
(Table 1). Under the similar condition standard antidiabetic
drug Acarbose exhibited a-amylase and a-glucosidase
inhibition activity with IC50 values of 0.15 and 0.11 mg/ml
respectively. The findings of the present study is in
agreement with earlier study where it has been shown that
mangrove plant extracts mediated silver nanoparticle could
inhibit a-amylase enzyme [14].
Anti-inflammatory Activity
Protein (Albumin) denaturation is a process in which pro-
teins lose their tertiary structure and secondary structure by
123
S. K. Das et al.
application of external stress or compound, such as strong
acid or base, a concentrated inorganic salt, an organic
solvent or heat. Protein denaturation is a marker for
inflammatory and arthritic diseases [13]. Most biological
proteins lose their biological function when denatured.
Denaturation of proteins (Albumin) is a well-documented
cause of inflammation. Agents that can prevent protein
denaturation, therefore, would be possible candidate for
anti-inflammatory drug development. As part of the
investigation on anti-inflammation activity, the ability of
nanoparticles to inhibit protein (egg Albumin) denaturation
was evaluated in the present study. Both XG-AgNPs and
AO-AgNPs were evaluated at 0.1, 0.2 and 0.5 mg/ml for
their capacity to inhibit the denaturation of heat induced
egg albumin (Fig. 7e). The study demonstrated that both
XG-AgNPs and AO-AgNPs could inhibit the protein
denaturation of albumin in a dose dependent manner and it
ranged between 25.6 and 100%. Amongst the two, AO-
AgNPs showed better protein denaturation capacity with
Green Synthesis of Sliver Nanoparticles Using Avicennia officinalis and Xylocarpus granatum… 1111

Fig. 6 XRD analysis of silver a 1800

nanoparticles using the extracts
of a X. granatum and b A. 1600

[4]
officinalis 1400

1200

Intensity (cps)

[1]
1000
800

[2]

[6]
600
400

[5]

[7]
[8]
200

[3]

[10]
[9]
0
10 20 30 40 50 60 70
2-theta (deg)

b 2400
2200
2000
1800

[3]
1600
Intensity (cps)

1400
1200
1000

[6]
[1]
800
600

[2]
400

[7]
[4]

[8]

[10]
200

[9]
[5]
0
10 20 30 40 50 60 70
2-theta (deg)

IC50 value of 0.17 mg/ml. Under the similar condition, the
standard drug Asprin could inhibit the denaturation of
albumin with IC50 value of 0.41 mg/ml (Table 1). The
present study corroborated with the earlier findings that
showed mangrove plant extract mediated AgNPs possesses
anti-inflammatory potential [14].
Earlier study have shown that different mangrove plants
such as Rhizophora mucronata, Ceriops tagal, Heritiera
fomes and Sonneratia apetala, S. alba, S. caseolaris were
being used for synthesis of silver nanoparticles and their
pharmaceutical applications in have also been studied
[14, 15, 23, 24]. However the present study on silver
nanoparticle synthesis using X. granatum and A. officinalis
extracts is a new attempt. The therapeutic potential of the
synthesized AgNPs using X. granatum and A. officinalis
plant extracts justified its previously claimed potential
applications [25, 26].
Conclusion
Nanotechnology research has grabbed considerable atten-
tion because of their unique ability to control the matter in
atomic and molecular scale. AgNPs can release of Ag ions
rapidly and exhibit various biological properties and hence
attracted tremendous attentions of researchers globally.
Different methods are being developed to find new,
greener, safer, economical, and rapid synthesis methods for
the preparation of silver nanoparticles. In the present study
a simple, safer, greener and non-toxic method of synthe-
sizing silver nanoparticles has been developed successfully
using X. granatum and A. officinalis plant extracts. The
UV–Vis, FT-IR, DLS and XRD analysis revealed that the
synthesized silver nanoparticles are stable in nature. The
results of the present study also concluded that these syn-
thesized silver nanoparticle showed excellent pharmaco-
logical properties such as antioxidant, antidiabetic, and
anti-inflammatory activities. The synthesized silver
nanoparticles from the mangrove plant could find potential
applications in biomedical and pharmaceutical industries
and hence, indicate the value of further studies. Thus,
synthesis of silver nanoparticles with medicinal phyto-
chemicals derived from barks of X. granatum and leaves of
A. officinalis may result in unprecedented opportunities
directed at large-scale production of silver nanoparticles
and can be used in many medicinal applications.

123
1112 S. K. Das et al.

Fig. 7 Bioactivities of silver nanoparticles using the extracts of X. e Protein denaturaion assays. All the experiments were done in
granatum and A. officinalis. a DPPH scavenging; b Superoxide triplicate and data are represented as mean ± SD. Different letters in
scavenging; c a-Amylase inhibition; d a-Glucosidase inhibition; the bar are significantly different (p \ 0.05)

Table 1 Bioactivities (expressed in IC50, mg/ml) of synthesized silver nanoparticles using extracts of X. granatum and A. officinalis
Sample DPPH scavenging Superoxide scavenging a-Amylase inhibition a-Glucosidase inhibition Protein denaturation

XG-AgNP 0.25 ± 0.004 0.49 ± 0.035 0.19 ± 0.011 0.13 ± 0.005 0.27 ± 0.01
AO-AgNP 0.14 ± 0.001 0.32 ± 0.017 0.28 ± 0.002 0.15 ± 0.012 0.21 ± 0.001
Standard 0.04 ± 0.001 0.08 ± 0.002 0.15 ± 0.003 0.11 ± 0.001 0.41 ± 0.01

123
Green Synthesis of Sliver Nanoparticles Using Avicennia officinalis and Xylocarpus granatum…
Acknowledgements The authors are thankful to PCCF (Wildlife),
Govt. of Odisha, for giving the necessary permission for the research
work. The authors are also thankful to the DFO, Rajnagar, Odisha and
their field staff for their kind help and cooperation during the field
study. The authors would like to acknowledge SAIF, Punjab, India to
carry out the 1H, 13C NMR and MS analysis of plant extracts.
Compliance with Ethical Standards
Conflict of interest The authors declare that they have no conflict of
interest.
References
1. U. Mani, S. Dhanasingh, R. Arunachalam, E. Paul, P. Shan-
mugam, C. Rose, and A. B. Mandal (2013). Prog. Nanotechnol.
Nanomater. 2, (1), 21–25.
2. M. Rai, K. Kon, N. Duran, S. Galdiero, and M. Galdiero (2014).
Appl. Microbiol. Biotechnol. 98, 1951–1961.
3. A. Henglein (1998). Chem. Mater. 10, (1), 444–450.
4. W. M. Bandaranayake (2002). Wetl. Ecol. Manag. 10, 421–452.
5. M. Khan, M. Khan, S. F. Adil, M. N. Tahir, W. Tremel, H.
Z. Alkhathlan, A. Al-Warthan, and M. R. H. Siddiqui (2013). Int.
J. Nanomed. 8, 1507–1516.
6. L. M. Rao and N. Savithramma (2012). Res. Biotechnol. 3, (3),
41–47.
7. J. Jiang, G. Oberdorster, and P. Biswas (2009). J. Nanopart. Res.
11, 77–89.
8. S. Sun, C. Murray, D. Weller, L. Folks, and A. Moser (2000).
Science. 287, 1989–1992.
9. W. Brand-Williams, M. E. Cuvelier, and C. Berset (1995). LWT-.
Food Sci. Technol. 28, (1), 25–30.
10. Y. Kono (1978). Arch. Biochem. Biophys. 186, (1), 189–195.
11. F. J. Gella, G. Gubern, R. Vidal, and F. Canalias (1997). Clinica.
Chimica. Acta. 259, 147–160.
1113
12. E. Apostolidis, Y. I. Kwon, and K. Shetty (2007). Innov. Food
Sci. Emerg. Technol. 8, 46–54.
13. P. Dey, P. Chatterjee, S. Chandra, and S. Bhattacharya (2011). J.
Adv. Pharma. Edu. Res. 1, 271–277.
14. P. Thatoi, G. K. Rout, S. Gouda, G. Das, K. Pramanik, H.
N. Thatoi, and J. K. Patra (2016). Photochem. Photobiol. 163,
311–318.
15. M. Bakshi, S. Ghosh, and P. Chaudhuri (2015). BioNanoSci. 5,
162–170.
16. R. Bhanumathi, K. Vimala, K. Shanthi, R. Thangaraj, and S.
Kannan (2017). New J. Chem. 41, 14466–14477.
17. S. Bhakya, S. Muthukrishnan, M. Sukumaran, and M.
Muthukumar (2016). Appl. Nanosci. 6, 755–766.
18. G. Suresh, P. H. Gunasekar, D. Kokila, D. Prabhu, D. Dinesh, N.
Ravichandran, et al. (2014). Spectrochim. Acta Part A Mol.
Biomol. Spectrosc. 127, 61–66.
19. M. M. Poojary, P. Passamonti, and A. V. Adhikari (2016). Bio-
NanoSci. 6, 110–120.
20. N. Kanipandian, S. Kannan, R. Ramesh, P. Subramanian, and R.
Thirumurugan (2014). Mater. Res. Bull. 49, 494–502.
21. P. Nagababu and V. U. Rao (2017). J. Appl. Pharm. Sci. 7,
175–182.
22. H. Ali, P. J. Houghton, and A. Soumyanath (2006). J.
Ethnopharmacol. 107, 449–455.
23. J. Umashankari, D. Inbakandan, T. T. Ajithkumar, and T. Bala-
subramanian (2012). Saline Syst. 8, 1–7.
24. S. P. Dhas, A. Mukerjhee, and N. Chandrasekaran (2013). Int.
J. Pharma. Pharma. Sci. 5, 349–352.
25. H. N. Thatoi, J. K. Patra, and S. K. Das (2014). Acta Physiol.
Plant 36, 561–579.
26. H. Thatoi, S. K. Das, and D. Samantaray (2016). Front. Life Sci.
9, 267–291.
Publisher’s Note Springer Nature remains neutral with regard to
jurisdictional claims in published maps and institutional affiliations.
123