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Investigation of Antioxidant for Polyvinylpyrrolidone-Silver
Nanocomposite Using Pandanus atrocarpus Extract
Abdullah Hasan Jabbar 1*, Hayder Shkhair Obayes AL-Janabi 2, Maytham Qabel
Hamzah3, Salim Oudah Mezan 4, Alaa Nahad Tumah 5, Amira Saryati Binti Ameruddin 6,
Mohd Arif Agam 7*
1-7 Department of Physics and Chemistry, Faculty of Applied Sciences and Technology,
University Tun Hussain Onn Malaysia, Pagoh Campus, Jalan Pancor, 84600 Pancor, Johor,
Malaysia
1 Al-Hussein Teaching Hospital, Directorate of Al-Muthanna Health, Ministry of Health,
Republic of Iraq
2 Department of Biotechnology, Al-Qasim Green University, Iraq
3,4 Directorate of Education Al-Muthanna, Ministry of Education, Republic of Iraq.
Abstract
Nanocomposites are characterized as a multiphase material where one of the phases has a
dimension in the nanoscale. There has been huge enthusiasm for the commercialization of
nanocomposites for an assortment of uses including medicinal, electronic, and basic. The general
motivation behind this study was on the development of silver nanoparticles, due to the present
enthusiasm encompassing these metals due to their exceptional properties which are not quite the
same as the relating bulk material. A novel, simple, cost-effective, nontoxic, and environmentally
friendly technique was developed for synthesizing silver nanoparticle- (AgNP-)
Polyvinylpyrrolidone (PVP) nanocomposite using Pandanus atrocarpus aqueous extract. UV-
visible spectroscopic analysis was carried out to assess the formulation of AgNPs. The average
size of green AgNPs was about 40 nm. Images of spherical green nanoparticles were characterized
using field emission scanning electron microscopy (FESEM). The resultant green AgNPs were
added slowly to polymer (PVP) solution. The AgNPs encapsulated within the polymer chains were
characterized by X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR).
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Issn No : 1006-7930
Page No: 235
Modification of thermal stabilities of AgNP/PVP nanocomposites was confirmed using UV- vis
(450nm). The green AgNP/PVP nanocomposites showed tested for antioxidant activity by DPPH
and ABTS. Both tests showed good results ABTS 76% and DPPH 73.10%. Thus, the key findings
of the work include the use of a safe and simple nanocomposite, which had marked antioxidant
activity.
Keywords: ABTS; DPPH; Nanocomposites; Polyvinylpyrrolidone; Polymers
Introduction
Preparation of noble metal nanoparticles by reducing metal salts has been extensively studied
(Jamkhande et al., 2019). To stabilize the dispersion of nanoparticles, it is necessary to use
protective agents, such as polymers, surfactants, and chelating agents (Ramanathan & Aqra, 2019).
The metal/polymer nanocomposite is being used as additives in polymer matrix and they are
excellent candidates for various applications (Boomi et al., 2014). In the past, Poly-N-vinyl-2-
pyrrolidone (PVP) a polymeric substance was used in many pharmaceutical applications as it is
biocompatibile with control rate of drug release so as to improve the in vivo pharmacokinetics
(Fahmi et al., 2009). PVP is one of the major additives to increase the solubility and dissolution
rate (i.e. restrain the drug crystallization in solution) of poorly water soluble drugs in
gastrointestinal pharmaceutical preparations and enhance the bioavailability of drugs Chadha et
al., 2006). Nowadays, the PVP polymer has been used as thin films, detoxicants and bio-active
compounds (Li et al., 2010).These nanoparticles have widely been studied with a view to
improving the quality of catalysts (Marson et al., 2018). All living organisms are suffered from the
damage caused by the free-radical oxygen species. Free-radical oxygen species damage cells by
attacking unsaturated fatty acids in the cell membrane. Fortunately, a protective enzyme,
superoxide dismutase, completely converts these free-radical oxygen species into two water
molecules and oxygen (Carneiro et al, 2018).
The enhanced biological properties of PVP doped PVP/AgNPs are to be explored. Therefore in
the present study, PVP- AgNPs have been synthesized using Pandanus atrocarpus extract (PAE).
Water was used as the environment benign reaction medium and plant metabolites as reducing and
capping agent, making the process greener one. Interaction between metal nanoparticles and PVP
is not much documented and properties that nanoparticles may acquire as a result of these
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Issn No : 1006-7930
Page No: 236
interactions are even less reported. Antioxidants may be defined as any substance that when
present at low concentrations compared with those of an oxidizable substrate, significantly delays
or prevents oxidation of that substrate in a chain reaction (Moharram & Youssef, 2014).
Antioxidants have become a popular research topic because they cannot be generated by the human
body and hence have to be consumed in the diet. Many fruit and vegetables have been found to be
rich sources of antioxidants. Since a large portion of the human diet is based on fruit and
vegetables, it is important to understand the biological and biochemical interactions between these
dietary antioxidants and living systems.
Materials and Methods
Preparation of Pandanus atrocarpus aqueous extract
Pandanus atrocarpus (PA) plant was supplied by Ethno Resources Sdn. Bhd. (Selangor,
Malaysia). The plant sample was stored at 4 C until further use. The protocol described by Awad
et al. (2019) was used for the preparation of aqueous extract. In brief, 2 g of PE whole plant powder
was boiled in 100 mL distilled water. The filtered extract was concentrated to dryness in a hot air
oven at 40 ℃ for 48 hours to obtain a dark brown semisolid mass which was weighed and labelled
as PAE. The MCE was stored at 4 ℃ and used for further experiments.
Synthesis of PA-AgNPs
For the synthesis of the silver nanoparticles, the PAE (0.8) ml was added to the AgNO3 solution
and the volume was adjusted to 10 ml with de-ionized water. The final concentration of Ag+ was
1 × 10−3 M. The solution was stirred for 2 min. The reduction process Ag+ to Ag0 nanoparticles
was followed by the color change of the solution from yellow to brownish-yellow to deep brown
depending on parameters studied such as the extract concentration, temperature and pH. The
nanoparticles were prepared at different pH values, the pH of the solutions was adjusted using 0.1
N H3PO4 or 0.1 N NaOH solutions (Khalil et al., 2014).
Synthesis of PVP/Ag Nanocomposite
PVP-Ag NCs synthesized by the method of (Awad et al. 2015) with slightly modification. The
PVP dissolved in distilled water to prepare a 20 % stock solution. The synthesized PAE-AgNPs
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(Previous section) mixed with of PVP (40%) in different ratios (1:9, 2:8, 3:7, 4:6, 5:5, 6:4, 7:3, 8:2
and 9:1) The resulting reaction mixture monitored for change in color visually as well as by
recording UV-vis spectra at regular intervals within the wavelength range of 300 to 700 nm. The
PVP-Ag nanocomposite, further characterized.
Characterization of PVP-AgNCs
The synthesis of PVP-AgNCs was confirmed by periodically scanning the absorption maxima of
aliquots (1.0 mL) of the reaction solution in1-cm path-length disposable cuvettes. The UV–
visspectrawere recorded every 30 minutes within the wavelength of 300 to 700 nm (Ahmed et al.,
2016). The possible biomolecules of PAE responsible for bioreduction of silver ions were
identified by recording the Fourier transform infrared (FTIR) spectrum of AgNPs using Perkin
Elmer Spectrum FTIR in the range of 4000–450 cm-1 at a resolution of 4 cm-1 (Ovais et al., 2016).
The powder AgNP sample was used for X-ray diffraction measurements performed on Shimadzu
XRD 6000 diffractometer operating at 40 kV in the region of 2θ from 30ᵒ to 80ᵒ at a speed of
0.02ᵒ/min (Ali et al., 2016). For size and shape analysis of PVP-AgNPs, the field emission
scanning electron microscope (FESEM) images were captured on JEOL JSM-7600F at an
accelerating voltage of 10 keV. The size analysis from FESEM images was performed on Image J
1.52g software (Mohseni et al., 2019).
Antioxidant Activity (ABTS free radical scavenging activity)
The ABTS free radical scavenging assay was performed by following the procedure as described
by Garcia-Leis et al. (2016). ABTS reagent was dissolved in water to a 7 mM concentration, the
ABTS stock solution was reacted with 2.45 mM potassium per sulfate (final concentration). The
ABTS•1 solution was diluted with ethanol in such a way that the absorbance was between 0.7-0.8.
The synthesized PVP-Ag NCs solution of different concentrations (100 µL, 200 µL, 300 µL, 400
µL and 500 µL, diluted to a total volume of 1 mL with distilled water) was mixed with 4 mL of
ABTS•1 solution in test tubes. The absorbance was measured at 734 nm exactly after 6 minutes
using UV-vis spectrophotometer (GENESYSTM 10S, Thermo Scientific, USA). ABTS•1 free
radical scavenging activity was expressed as percent inhibition (% I) calculated by the following
formula:
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% I = (C - S
C)×100
Where; C is the absorbance of control and S is the absorbance of sample.
All experiments were repeated thrice and the data were reported as means (±SD for triplicates).
Antioxidant Activity (DPPH free radical scavenging activity)
The DPPH free radical scavenging activity of CA-CrA was estimated according to the
procedure described by Rasheed et al. (2017), with slight modification. Briefly, 0.5 mL of 0.1 mM
DPPH (dissolved in ethanol) was mixed with 0.5 mL each of synthesized PVP-Ag NCs solution
of different concentrations (100 µL, 200 µL, 300 µL, 400 µL and 500 µL, diluted to a total volume
of 0.5 mL with distilled water, where needed) in test tubes. The absorbance was recorded at 517
nm by using UV-vis spectrophotometer using UV-vis spectrophotometer (GENESYSTM 10S,
Thermo Scientific, USA). DPPH free radical scavenging activity was expressed as percent
inhibition (% I) calculated by the following formula:
% I = (C - S
C)×100
Where; C is the absorbance of control and S is the absorbance of sample.
All experiments were repeated thrice and the data were reported as means (±SD for triplicates).
Results and Discussion
Uv-Vis (Ultraviolet–visible Spectroscopy)
The formation of PVP-AgNC was confirmed by UV–vis spectral study. The AgNPs was first
checked for change in color from colorless to brown and UV-Vis spectroscopy. The change in
color showed the formation of AgNPs because of the reduction of silver metal particles Ag+ to
nanoparticles Ag0. This color refers to the excitation of SPR. As shown in Figure 1, characteristic
SPR band for AgNPs and PVP-AgNCs was observed at around 432 nm and 420 nm respectively
(Edison & Sethuraman, 2017).
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Figure 1: UV-VIS absorbance spectra for various weight-of (wt %) AgNO3 and different concentrations
Fourier Transformed Infrared Spectrometry (FTIR)
The prominent peaks present in PAE are located at 3338, 2131, 1638, 1295, and 428 as shown in
Fig. 2. It may be inferred from that peak shifts after PVP-Ag NCs synthesis by varying the
concentration of PAE do not considerably differ from each other as all the five spectra very closely
overlap each other. In all of the PVP-Ag NCs samples, the same two peak shifts after Ag NCs
synthesis were observed as: 3338 to 3325/3326 cm-1 and 1638 to 1634/1635 cm-1, as recorded for
Ag NPs samples 1 to 5. The peak at 3338 belongs to the –OH group which, in this case, can be
attributed to the phenolic and flavonoid compounds of PAE. Second important stretching vibration
is recorded from 1638 to 1634/1635 cm-1 which, although may be considered as amide groupf of
proteins as well as benzene ring containing aromatic compounds, it may also be taken into account
for phenolic and flavonoid compounds as they exhibit strong vibration on this wavenumber
(Shivakumar et al., 2014; Prathna et al., 2011). In the context of all these stretching vibrations
found in the FTIR spectra of PVP-Ag NCs, it may be assumed that they have played a role in
fastening the rate of Ag NCs synthesis reaction leading to the nucleation along with stalling the
secondary growth of the nuclei.
0.00.20.40.60.81.01.21.41.61.82.0
300 400 500 600 700
Ab
sorb
ance
Wavelength (nm)
30 min 60 min 90 min 2 h 3 h
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Figure 2: Fourier Transformed Infrared Spectrometry (FTIR) spectrum of conductive film.
Field Emission Scanning Electron Microscopy (FESEM)
The particle size and morphology analysis was performed by scanning electron microscope (SEM).
The SEM micrographs of the AgNPs and PVP-AgNCs are shown at the magnification of 200,000×
in figure 3. The shape of the PVP-AgNCs is almost spherical and the smallest particle size is 27.34
nm. The function of PVP in the PVP-AgNCs nanocomposites is not only as a binder, but it also
prevents the process of agglomeration of Ag nanoparticles and limits the diameter of the
nanoparticles formed (Zhang et al., 2011; Flahaut et al., 2000). As for the morphology of particles
in nanocomposite, it is evident that the particles obtained in this study are mostly spherical in
shape.
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Figure 3. FESEM image of PVP/AgNCs
X-Ray Diffraction Analysis (XRD)
The XRD analysis of PVP/AgNC was carried out to confirm the crystalline nature of the
synthesized nanomaterials. Figure 4 show the XRD spectrum of PVP/AgNCs, respectively. Figure
4 show the XRD spectrum of MCE-AgNPs and PVP/AgNC, respectively. The X-ray
diffractograms, at 2θ from 20°-80°, clearly indicate the crystallinity of the tested samples as
indicated by the broadening of Bragg’s peaks. The presence of sharp peaks indicates the bioactive
compounds present on the surface of the NPs (Shabestarian et al., 2017). The Bragg’s reflection
values of 38°, 44°, 64° and 77°, obtained for MCE-AgNPs and PVP/AgNCs, corresponding to the
set of lattice planes, that is, (111), (200), (220) and (311) have been obtained at 2θ indicating the
formation of face centered cubic (fcc) crystalline structure of silver nanostructure. The peak related
to the lattice plane (111) is the most intense suggesting its most predominant orientation in addition
to the crystalline nature of the synthesized silver colloids. The XRD spectrum did not show any
other crystallographic peak confirming the high purity of synthesized Ag colloids. Similar results
have been reported recently for the synthesis of AgNPs-mediated nanocomposite (Awad et al.,
2015).
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Table 1. XRD Peak list of PVP/AgNCs
Pos. [°2Th.] Height [cts] FWHM Left [°2Th.] d-spacing [Å] Rel. Int. [%]
38.0589 4981.03 0.4428 2.36445 100.00
44.2253 1324.48 0.4723 2.04803 26.59
64.5287 1127.84 0.2066 1.44417 22.64
77.4782 1191.13 0.4723 1.23197 23.91
20 30 40 50 60 70 80
0
1000
2000
3000
4000
5000
6000
(311
)
(220
)
(200
)
(111
)
Inte
nsi
ty
2 Theta
PVP- AgNCs
Figure 4. XRD pattern of PVP/AgNCs
Antioxidant activity of PAE-AgNPs and PVP/AgNCs
Many research reports indicate that there is an intimate relationship between oxidative stress and
age-related neurodegenerative disorders. In this context, many studies have been performed to
examine the beneficial effects of antioxidants aiming at reducing or blocking the neuronal death
which is usually a pathophysiological consequence in such diseases (Ramassamy, 2006). As the
radical scavenging capacity of a compound is usually attributed to its antioxidant potential, two
antioxidant activity assays of as-synthesized PAE-AgNPs and PVP/AgNCs were performed for
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the evaluation of their antioxidant potential, that is, their capacity to inhibit ABTS and DPPH free
radicals.
ABTS Free Radical Scavenging Activity
The dose-response bar chart of ABTS scavenging assay of as-synthesized PAE and PVP/AgNC
are shown in Fig. 5. It can be clearly observed that all samples and PAE have good antioxidant
potential with effective ABTS radical inhibitory activity in a dose-dependent manner. At 500 µL
sample, the ABTS radical scavenging activity (% I) of PAE was recorded at 58.20 %, and
PVP/AgNC at 76.00 % (Table 2). The antioxidant activity of PVP/AgNC was higher than PAE.
However, the maximum ABTS radical scavenging activity was shown by ascorbic acid recorded
at 81.40 %.
This result is in conformity to the previous reports in which MNPs were reported to have
higher antioxidant activity as compared to the extract (Gomaa, 2017; He et al., 2017; Khan et al.,
2016). The antioxidant activity of ascorbic acid was higher than that of the other samples as shown
in Table 2.
Figure 5. ABTS free radical scavenging assay of PVP/AgNCs and PAE. Error bars represent 95%
confidence interval (n=3).
0.0
10.0
20.0
30.0
40.0
50.0
60.0
70.0
80.0
90.0
100 200 300 400 500
AB
TS %
I
Sample Concentration (µL)
PVP-AgNC
Extract
Ascorbic acid
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Table 2. ABTS free radical scavenging assay of PVP/AgNCs and PAE
Sample Concentration (µL) PA-AgNPs±SD PAE±SD Ascorbic acid±SD
100 21.4±1.35 12.0±1.19 41.7±1.25
200 34.6±1.36 21.8±1.26 49.9±0.94
300 48.4±1.54 34.9±1.20 60.9±1.10
400 59.0±1.13 48.9±1.73 71.2±1.40
500 64.2±1.95 58.7±0.85 81.9±1.85
DPPH Free Radical Scavenging Potential
The DPPH free radical scavenging potential of as-synthesized PVP/AgNCs and PAE show that
they have the hydrogen donating ability and hence they may serve as free radical inhibitors or
scavengers, acting possibly as potent antioxidants. The dose-response bar chart of DPPH free
radical scavenging assay of as-synthesize, PVP-AgNCs and PAE have been shown in Fig. 4.9. It
can be clearly observed that both the silver colloids and PAE have good antioxidant potential with
DPPH radical inhibitory activity in a dose-dependent manner. At 500 µL sample, the DPPH radical
scavenging activity (% I) of PVP/AgNCs recorded at 73.10 % and PAE at 58.70 % as shown in
Fig. 6. However, it is noteworthy that the antioxidant activity of PVP/ AgNCs is higher than CAP
that may be due to comparatively higher concentration of bioreductants adsorbed onto the surface
(Shabestarian et al., 2017). The antioxidant activity of ascorbic acid was significantly higher than
that of the PVP-AgNCs and PAE as shown in table 3.
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Figure 6. DPPH free radical scavenging assay of PVP/AgNCs and PAE. Error bars represent 95%
confidence interval (n=3).
Table 3. ABTS free radical scavenging assay of PVP/AgNCs and PAE
Concentration (µL) PVP/AgNCs±SD PAE±SD Ascorbic acid±SD
100 28.0±0.84 12.0±1.19 41.7±1.25
200 42.1±0.94 21.8±1.26 49.9±0.94
300 55.5±1.10 34.9±1.20 60.9±1.10
400 66.4±1.16 48.9±1.73 71.2±1.40
500 73.1±0.99 58.7±0.85 81.9±1.85
The electron transferring potential of a compound determines its reducing power and as such it
may be considered in terms of its antioxidant activity (Gooding et al., 2014). The presence of rich
polyphenols and flavonoids content, responsible for the biosynthesis of PAE-AgNPs, has been
reported for its antioxidant capacity (Gray et al., 2018). These antioxidant compounds of flavonoid
content might got adsorbed onto the surfaces of metallic NPs (Wang et al., 2009). Surface reactions
of the synthesized NPs, by virtue of the adsorbed antioxidant moieties onto the NPs surfaces, along
with a high surface area to volume ratio of the NPs, produce a tendency in the NPs to interact and
0.0
10.0
20.0
30.0
40.0
50.0
60.0
70.0
80.0
90.0
100 200 300 400 500
DP
PH
% I
Sample Concentration (µL)
PVP/AgNC
PAE
Ascorbic acid
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scavenge the free radicals (Dauthal & Mukhopadhyay, 2013). Nobel MNPs like AgNPs and GNPs
have been used under in vitro and in vivo conditions for the scavenging of free radicals (Suganthy
et al., 2017). It implies that the findings of this study signify the potential of PAE-AgNPs as
promising antioxidant moieties.
Conclusion
The introduced work demonstrated the fast greener synthesis of AgNPs utilizing TPandanus
atrocarpus and their composite with the PVP polymer. The technique here is nontoxic, ecologically
cordial, and straight forward and involves minimal effort and has no lethal chemicals. The
formation of greener AgNPs was determined by FESEM and UV-Vis spectroscopy, where UV-
vis recorded at 450 nm in the UV-Vis range. FESEM demonstrated the average size of the resulting
nanoparticles to be 40 nm. The nanocomposite was characterized utilizing FTIR spectroscopy and
XRD techniques. The nanocomposite films showed significant antioxidant activity both on ABTS
and DPPH assay. This guarantees a promising potential utilization of the nanocomposite as
antioxidant.
Acknowledgment
This work was funded by the Fundamental Research Grant awarded by the Ministry of Higher
Education (FRGS/1/2019/STG07/UTHM/02/5(FRGS K171).
Conflict of interest: The authors have no conflicts of interest to declare
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