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Materials Today: Proceedings 18 (2019) 920–927 www.materialstoday.com/proceedings

ICN3I-2017

Study of Synergistic Effects of Antibiotics And Triangular Shaped

Silver Nanoparticles, Synthesized Using UV-Light Irradiation, on S.
Aureus and P. Aeruginosa

Sonali Sahaa, M.M.Malikb, M.S. Qureshic

a
IES College, Bhopal
b, c
Department of Physics, M.A.N.I.T., Bhopal, India

Abstract

Now days, resistance of bacteria to bactericides and antibiotics has increased. Some antimicrobial agents are
extremely irritant and toxic. Hence there is a need to find ways to formulate green and less toxic materials. In the
present work, a study of synergistic effects of as-synthesised triangular shaped silver nanoparticles and two standard
antibiotics, ampicillin and gentamycin, was studied on Staphylococcus Aureus and Pseudomonas Aeruginosa. A
green route is taken to synthesise silver nanoparticles using silver oxalate as precursor; tea extract and chitin as
capping and stabilizing agents respectively. A grey coloured colloidal solution of silver nanoparticles is obtained.
XRD, TEM and UV-visible spectroscopy, are used tocharacterise the as-synthesised silver colloidal solution.

© 2019 Elsevier Ltd. All rights reserved.

Selection and/or Peer-review under responsibility of International Conference on Nanotechnology: Ideas, Innovations & Initiatives-2017 (ICN:3i-
2017).

Keywords: Silver colloidal solution, Tea extract, UV-Irradiation, Ampicillin and Gentamycin

2214-7853© 2019 Elsevier Ltd. All rights reserved.

Selection and/or Peer-review under responsibility of International Conference on Nanotechnology: Ideas, Innovations & Initiatives-2017 (ICN:3i-
2017).
 S. Saha et al. / Materials Today: Proceedings 18 (2019) 920–927 921

1. Introduction

Among metal nanoparticles, silver is gaining more attention as it exhibits completely new or improved properties
based on specific characteristics such as size distribution and morphology. It is well known from ancient time that
silver nanoparticles have very useful biomedical application due to their antibacterial activity. Antibacterial activity
of the silver-containing materials can be used, for example, in medicine to reduce infections in burn treatment[1,2]
and arthroplasty,[3] as well as to prevent bacteria colonization on prostheses,[4] catheters,[5,6] vascular grafts,[7]
dental materials,[8]stainless steel materials,[9] and human skin.[10,11]. At nano scale range this particle size leads
to ultra large surface area per mass where a large number of atoms are in immediate contact and available for
reaction. The antibacterial activity can be tuned by controlling the size and morphology of nanoparticles. This size,
morphology and stability of the metal nanoparticles are strongly influenced by the experimental conditions [12, 13].
Hence, the design of a synthesis method in which the size, morphology, stability and properties are controlled has
become a thrust area[14] of research.

Previous studies done by researchers on the synthesis and the antibacterial activity of silver nanoparticles, we find that
most of the studies are limited upon interaction of spherical silver nanoparticles with bacterial cell wall.[15, 16] The
work done by Pal.et.al [17]was the first comparative study of the effect of shape of nanoparticles on bacterial cell.
The use of silver had reduced as an anti-infection agent due to the advent of antibiotics and other
disinfectants and the poorly understood mechanisms of their toxic effects. However, resistance of bacteria to
bactericides and antibiotics has increased in recent years. Some antimicrobial agents are extremely irritant and toxic.
Hence there is a need to find ways to formulate green and less toxic materials? Therefore study of the synergistic
effects of silver nanoparticles with antibiotics on bacteria has become the requirement of the present time.
In the present synthesis route the author has developed a green route by using tea leaf extract as capping
agent, UV-irradiation source as a reducing agent and chitin flakes as stabilizing agent to synthesised triangular
shaped nanoparticles. Tea, especially black tea, contains antioxidants. The antioxidants contained in tea play a major
role in protecting the body against certain illnesses such as cancer and heart ailments. In view of utilizing this
property of tea extract the green synthesis process discussed in this paper, uses tea extract as capping agent. Silver
oxalate (Ag2C2O4) is photosensitive and yield metallic silver upon the exposure to UV light, those compounds can
be photo chemically decomposed to obtain Ag nanoparticles in the presence of capping agents. Previous studies
show that antimicrobial formulations of silver in the form of nano particles could be used as effective bactericidal
materials. In the present work synergetic effects of silver nanoparticles with antibiotics is being studied against gram
positive and gram negative bacteria.

2 Materials and Method

2.1 Chemicals.
Materials used for the synthesis of silver nano particles are AR grade silver nitrate (AgNO3) and oxalic acid
purchased from Merck, India, Red label tea leaves, deionized water (Ultra pure) and chitin flakes.

2.2 Preparation of silver oxalate

Silver oxalate was prepared by mixing 50ml solution of 0.5M AgNO3(Merk,99%) with 30ml of 0.5M oxalic acid
(Merk,99%).The white formed precipitate was filtered washed with distilled water, dried in an air oven for one hour
and stored in a dark bottle.

2.3 Preparation of tea extract

In 50ml volumetric flask was filled up to the mark with boiling water,0.5gm of tea grains were weighed and
transferred to this flask and filtered immediately.

2.4 Synthesis of Colloidal Silver nano particles

For the synthesis of colloidal solution, 10ml of tea extract is mixed with 0.02 gm of Ag2C2O4 and 20ml doubly
distilled water in a three neck round bottom flask and stirred for 35mint in dark. Vacuum is created in the flask
followed by UV radiation. During the irradiation process no cut-off filter was used. A plane yellow colloidal
922 S. Saha et al. / Materials Today: Proceedings 18 (2019) 920–927

solution was formed. After this 0.01gm chitin flakes used as stabilizing agent was added to the solution and finally
the colour becomes grey. The resultant colloid was washed by centrifugation several times.Fig.1 shows the set up
for the above procedure.

Fig.1 Experimental Set-up

3. Results and discussions

As-synthesised silver oxalate was characterized using powder XRD using (Rigaku Miniflex II). The lines presented
in the powder XRD pattern match the standard sample, primitive monoclinic system. Moreover it is also clear from
the XRD graph (shown in fig.2) that no other phase of silver oxalate is present in the as-synthesised silver oxalate.

Fig.2 XRD graph of silver oxalate

 S. Saha et al. / Materials Today: Proceedings 18 (2019) 920–927 923

Further characterization was done by SEM (JSM 6390 A Models). The SEM images of as-synthesised silver oxalate
is shown in fig.3 (a, b) which indicates irregular shaped particles approximately of size in the range of 500-750 nm.

(a) (b)

Fig.3 (a, b) SEM images of silver oxalate

Fig.4 XRD pattern of sample (for 30mint of uv-irradiation)

The XRD pattern of the sample shown in fig.4 showed that when sample was exposed for different time interval (30
mints),silver oxalate was partially decomposed to generate silver, the main remaining were silver oxalate. Thus the
time of UV-irradiation was increased to one hour and further it was observed that silver oxalate decomposed
completely. Graph in fig. 5shows pure peaks of pure silver well in accordance with JCPDS file No. 04-0783. The
diffraction profile of as-synthesised colloidal silver is fcc and are obviously broadened as compared with bulk silver
confirming the formation of silver nanoparticles.
Silver oxalate decomposes under UV irradiation to give metallic silver and CO2gas. This is due to the high
photosensitivity of Ag2C2O4. [18]. Since this decomposition of Ag2C2O4 is thermodynamically favourable due to the
suitable reduction potentials of oxalate [19].This decomposition of oxalate occurs rapidly under UV radiation to
yield metallic Ag as shown in Equation given below:

Ag2C2O4(s) →Ag(s) +2CO2(g)

924 S. Saha et al. / Materials Today: Proceedings 18 (2019) 920–927

In the presence of UV-irradiation dianion of silver oxalate get excited and thus decomposes into CO2, formation of
CO2 can be proved by the appearance of the white precipitate in the baryata solution [20]. To understand the
mechanism of the reaction during the decomposition we see that electron is continuously transferred from silver ion
to form silver metal [18].This formed silver atoms starts to form particles after getting sufficient concentration.

Fig.5 XRD graph of as-synthesised Ag nano particles for 1hour UV-reduction

TEM characterization of the as-synthesized colloidal silver in fig. 6 (a,b) shows formation of triangular silver
nanoparticles. The reason for the triangular shaped silver nanoparticles can be attributed to the reason that less time
of uv-irradiation leads to slower reduction rate [21]. The average particle size was around 50nm. The size control of
metal nano particles is determined by such factors as precursor concentration, molar ratio between surfactant and
precursor, as well as the selective absorption of surfactant to different crystal facets [22-23].

(a) (b)
Fig.6 (a, b) Tem images of as-synthesised silver nanoparticles for Reduction time 1hr
 S. Saha et al. / Materials Today: Proceedings 18 (2019) 920–927 925

After the TEM measurement it becomes also very clear that in the present process tea extract is only acting as
capping agent nor as a reducing agent because tea extract is a weak reducing agent [24] and the particle size what
we are getting in the present method is not possible by the reduction by weak reducing agent.

The next characterisation which was done to characterise the as-synthesised nanoparticles was UV-visible
spectroscopy.

Fig.7 UV-visible spectroscopy of as-synthesised nanoparticles for Reduction 1hrs

Figure 7 it is very clear that the peak for the graph is nearly at 520nm which is the characteristic peak for triangular
shaped silver nanoparticles [25].
4. Antimicrobial study
The bactericidal properties of silver nanoparticles of different shaped were conducted [26]. In the present study
antibacterial activity of triangular shaped nanoparticles along with antibiotics ampicillin and gentamicin was tested
against Staphylococcus Aureus and Pseudomonas Aeruginosa because triangular nano prism with sharp vertices and
edges also display a good antibacterial activity in comparison to other shaped nanoparticles, these nanoparticles can
be useful for biomedical applications [27]. As in the present route the shape of the synthesised nanoparticles is
triangular which can give best antibacterial result for silver nanoparticles. Zone of inhibition were obtained for the
combination of silver nanoparticles and antibiotics. The comparison table of zone of inhibition is shown in table 1.
For the comparative study, 1ml of as-synthesized colloidal solution of silver nanoparticles was taken with
10mcg of ampicillin for 15 minutes and the total solution was placed on Soyabean casein digest agar and incubated
at 370 C for 48 hrs. The process was repeated with 10mcg of gentamycin.The test was conducted Micro bio
laboratory Thane (Maharashtra, India).
Table 1-Comparitive study of effect of silver nanoparticles in combination with two antibiotics

SR. NO. ORGANISM ZONE FOR SAMPLE ZONE FOR SAMPLE

+ AMPICILLIN + GENTAMICIN
P1 P2 MEAN P1 P2 MEAN
1. Staphylococcus Aureus 22 20 21 20 18 19

2. Pseudomonas Aeruginosa 19 23 21 20 16 18
926 S. Saha et al. / Materials Today: Proceedings 18 (2019) 920–927

Plate 1=P1 Plate 2=P2

The mechanism of the bactericidal effect of Ag NPs is that they may attach to the surface of the cell membrane
disturbing permeability and respiration functions of the cell [28]. Sharp vertexes and sharp edges of triangular nano
prism are more toxic in damaging the bacterial cell [27].
Thus we can summarise from the size of inhibition zone that the as-synthesised sample in
combination with ampicillin and gentamicinis equally effective for killing of both the bacteria Staphylococcus
Aureus and Pseudomonas Aeruginosa
5. Conclusions

In the present work the author to best of her knowledge has tried a new way to synthesise silver
nano particle of triangular shape using the basic principle of green technology. The average particle size obtained for
the nanoparticles was 50nm. The most important feature of this procedure is that it is free from any chemical
process, which satisfies the green biogenic approach of synthesis and gives an edge over various chemical
procedures usually applied for the synthesis of silver nanoparticles. A comparative antibacterial study of as-
synthesised silver nanoparticles in combination with two standard antibiotics was analysed for a gram positive and a
gram negative bacteria and it was found that sample in combination with ampicillin and gentamicin are equally
effective for both the bacteria.

Acknowledgement
Author is thankful to the Director, M.A.N.I.T., Bhopal, India for the facilities at the institute, AMPRI Bhopal for
XRD Bhopal, for providing XRD facility, HSADL, Bhopal for the TEM facility, Civil Engineering department
M.A.N.I.T., Bhopal for providing UV vis-spectrophotometer facility. I am thankful to all my colleagues for the
valuable help and suggestions.

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