Purple wisteria

Abstract

In the current study, we reported the green synthetic route of TiO2 nanoparticles using WS
acting as both reducing and capping agent. Various characteristic parameters like
morphology, size of particle, crystalline structure, elemental composition and stability were
studied using UV-VIS spectroscopy, FTIR, Scanning Electron Microscopy (SEM), X-ray
diffraction (XRD), Energy dispersive spectroscopy (EDS). In case of UV-VIS spectroscopy,
characteristic peak of TiO2 was observed at 350 nm which confirm the formation of TiO2
nanoparticles. Then through morphological studies, shape was found to be spherical with
elemental composition of 5.22 % and 32.29 % for titanium and oxygen respectively. From
FTIR, different stretching and bending peak were observed for TiO2 particles. Afterwards,
antimicrobial activity was performed against various gram positive and gram negative
bacteria.
1. Introduction

The application of nanotechnology in many aspects of daily life, including medical,

pharmaceuticals, electronics, energy, the environment, chemistry, food, and, more recently,
agriculture, is increasing relevance in society. In the latter field, nanotechnology is utilised in
the formulation, manufacturing, and application of nano fertilisers, nano insecticides, and
nano herbicides. Nanotechnology is also utilised in the development of goods like zeolites,
clays, and chitosan that are specifically designed to improve plant nutrient absorption. These
substances avoid their interaction with soil, water, air, and microbes, which exponentially
decreases the loss of micro- and macro-elements in plants. Nanobiotechnology, which
focuses on the creation and use of nanoparticles (NPs), has had a substantial impact on the
growth of numerous economic sectors, including those related to food, cosmetics, agriculture,
medicine delivery, cancer theranostics, and many more. Nanoscale materials differ from
microscale materials in many ways, including their chemical, mechanical, magneto-optical,
and surface-to-volume ratio, making them candidates for use in a variety of biological
applications.

For the synthesis of NPs, a variety of physical and chemical techniques have been used,
including sol-gel chemistry, electro deposition, combustion, the solvothermal approach, and
co-precipitation. However, these synthesis techniques have significant negative side effects
that restrict the large range of biological applications they can be used for. When compared to
physical processes, chemical synthesis most typically produces toxic substances that are
harmful to the environment and result in non-biodegradable products. Scientists have created
a more dependable, environmentally friendly way to solve these issues. Different biological
sources, including plants, algae, microbes, and others, can be employed to synthesise required
NPs that are risk-free, affordable, green, and environmentally friendly. The necessary MNPs
can be synthesised from a variety of biological sources, including plants, algae, microbes, and
others, and are safer, more affordable, environmentally friendly, and green. As plant extracts
have a rich phytochemical profile (alkaloids, flavonoids, and phenolics), which are thought to
be excellent stabilising and chelating agents for biogenic nanoparticles, green synthesis
utilising plant extracts is growing in popularity. Plant extract may also be used to alter the
size and form of NPs as well as act as a potent capping and lowering agent. Scientists have
therefore concentrated on the plant-mediated production of metallic nanoparticles.
In recent years, scientists have gained interested in the reduction of metal salts to NPs, which
may lead to the development of an affordable and ecologically friendly way to make NPs.
Wisteria sinensis is a member of the Fabaceae family and is a species of the genus Wisteria.
With their numerous tall, blue-violet, fragrant flowers, its dangling racemes make a stunning
visual. It is a climbing plant that can grow up to 20 metres tall. The oil of wisteria flowers has
previously been used to extract phenylpropanoids and -chromenes, according to a review of
the literature. This plant contains lectins, triterpene Saponins, and isoflavone phytochemicals.
It has antibacterial, antioxidant, and cancer-fighting qualities.

Wisteria sinensis leaf extract was utilised in this study to create an innovative, affordable, and
ecologically friendly method for the synthesis of NPs. In order to create nanoparticles in an
environmentally friendly manner and to evaluate the photo-catalytic and biological properties
of these nanoparticles, wisteria sinensis extract has been employed for the first time. We
think this is the first time Wisteria sinensis has been used to create nanoparticles. The
polyphenolic compounds, or phytochemicals, contained in this plant are particularly
significant due to the multitude of functions for which they are used. This plant was chosen
for this study because it is readily available, financially viable, and non-toxic. It was
discovered that plant extract acts as a reducing agent and that each plant or biomass has the
same reduction effect. Although there may be some variation in the phytochemicals of the
plants, their main function is to serve as reducing and stabilising agents. Therefore, the major
goals of this study are to (i) manufacture NPs using Wisteria sinensis leaf extract at a low
cost and without harming the environment. (ii) Research possible uses such dye degradation
(iii) Examine the antioxidant and antibacterial qualities of NPs.
2. Material and methodology

All of the chemicals used to carry out this study were of the good quality and the solvent was
of analytical grade. Titanium isopropoxide, ammonia solutions were purchased from LOBA
Chemie and Sicsco research laboratories respectively. Double distilled water was used in all
the experiments. Fresh leaves of the “Wisteria Sinensis” (Chinese wine flower) were obtained
from the locality of Palampur, Himachal Pradesh, India.

2.1 Methodology

2.1.1 Preparation of plant extract

The leaves of the plant were thoroughly cleaned with tap water and then with double-distilled
water to get rid of any contaminants. To establish a constant mixture that was used
throughout the investigation, the mixture was air dried and carefully milled. Afterwards,
20gm of leaf powder was mixed in 200ml of distilled water. Then the mixture was allowed to
heat at 80 oC for nearly 4 hours (until solution get reduced to half of its initial volume).
Before storing the solution in the refrigerator, the concentrated extract was filtered through
Whatman filter paper.

2.1.2 Synthesis of TiO2 nanoparticles using Wisteria Sinensis

To prepare plant mediated TiO2 nanoparticles, a solution was prepared by mixing TiO2
(10gm) in ethanol (100 ml). Solution was put on magnetic stirrer and then 30 ml of plant
extract (mixed in 100 ml of water) was added into the solution to get homogenized solution.
After two hour of stirring, ammonia was added slowly and was allowed to stir for another one
hour. After the completion of reaction, the solution was kept for overnight at room
temperature. Next day the precepitates were filtered and then kept in oven for 2 hrs (200 0C).
For conduct further studies, the nanoparticles were calcinated at 550 oC, 750 oC, 1050 oC.

2.1.3 Instrumentation

UV-Vis absorption spectra were recorded on Shimadzu UV-2450 model spectrophotometer.

All the measurements were recorded between the range 190-900 nm wavelengths. For FTIR
analysis, Agilent technologies carry 600 spectrometer was used. To identify possible
functional groups, the range was selected to be 4000-400 cm-1. To collect the information on
crystal phase and crystal structure, X-ray diffraction analysis was carried out using Rigaku
corporation Smart lab model having 9kw rotating anode diffractometer.
The average crystalline size was calculated by Debye-Scherrer equation:

D = Kλ/βCosθ

Where ‘D’ represents the average grain size of the material, ‘K’ is Scherrer’s constant, λ is
the wavelength, and β is the full width half maxima of the peak. ‘θ’ is the Bragg’s angle.

For determining the surface morphology of the sample, scanning electron microscopy (SEM-
EDS) was depicted using JSM-6610-JEOL/EO SEM model.

2.1.4 Antimicrobial activity

For the antimicrobial activity of the synthesized nanoparticles, four bacterial strains, namely,
Staphylococcus aureus, Pseudomonas aeruginosa, Escheria Coli, and Klebsiella pneumonia
were taken. For this experiment, glassware and samples were sterilized before use. Agar well
diffusion method was used to determine the potential antimicrobial activity against the as-
synthesized nanoparticles.
3. Results and discussion
The synthesis, characterisation, and antimicrobial activity of the synthesized plant mediated
particles are covered in this section. An environmentally friendly method involving
coprecipitation technique was used to synthesise WS-TiO2. For characterising synthesised
catalysts, various physicochemical techniques including XRD, SEM-EDX, UV-VIS, and
FTIR were used. Antimicrobial activity was carried out on four different bacteria using agar
well diffusion method.
3.1 Synthesis of WS-TiO2
WS-TiO2 was synthesized by mixing TiO2 and plant extract using coprecepitation method.
Afterwards it was calcinated at different temperatures such as 550, 750 and 1050 oC (Fig. 1).
With the change in temperature, structure, morphology and particle size vary and leads to
change in their applications. The obtained catalyst was characterised using different
techniques.
Fig.1 Images of nanoparticles calcinated at different temperatures (a) WST-550 (b) WST-750
(c) WST-1050

3.2 X-ray diffraction (XRD) pattern of WS-TiO2

When calcinated at a lower temperature (550 0C), the primary peak was detected at 25o
according to XRD spectra (Fig.2 (a). The major phase is precisely ascribed to the anatase
phase, which is represented by the diffraction peaks 25.540(101), 38.040(112), 48.230(200),
54.190(105), 55.240(211), and 62.980(213).When calcinated at 750⁰C , the diffraction peaks
were seen at 25.54⁰(101), 27.64⁰(110), 36.36⁰(004), 37.14⁰(004), 38.02⁰(112), 38.81⁰(112),
48.29⁰(200), 54.11⁰(105), 55.32⁰(211), 62.94⁰(002), 68.98⁰(116), 70.57⁰(220), 75.33⁰(107)
which is also showed that the anatase phase is the primary phase (Fig.2 (b)). Rutile is the
predominant phase that is seen at 27 0C after synthesised nanoparticles are calcined at 1050
0
C. At 27.700(110), 29.370 (110), 36.320 (101), 39.460(200), 41.490(111), 44.310(210),
54.560(105), 56.870(220), 62.980(002), 64.330(310), 69.240(301), 69.410(301), and
70.030(220), the diffraction peaks were noted (Fig.2 (c)).
Fig. 2 XRD pattern of synthesized nanoparticles (a) WST-550 (b) WST-750 (c) WST-1050

Due to the assimilation of organic molecules found in the plant extract, the anatase to rutile
phase transition is taking place. The created TiO2 nanoparticles are crystalline in form and
exhibit anatase and rutile phases. The rutile phase is seen at a higher temperature of 1050 0C,
while the anatase phase is shown at a lower temperature of 550 0C.
Table 1 provides the average crystalline size and lattice strain as determined by Scherer's
equation and the lattice strain equation. Lattice strain and crystalline size were determined to
be 17.51 nm, 35.11 nm, and 64.93 nm, respectively, and to be 0.0089, 0.0044, and 0.0022.
Table 1. Phase composition, crystalline size (nm) and lattice strain of WST-550, WST-750
and WST-1050
NPs 2θ of D- Axial Crystalline Lattice Phase Structure
crystalline spacing angle size (nm) strain
plane

WST- 25.5 3.48 α=β= 17.51 0.0089 Crystalline Tetragonal

550 γ = 900

WST- 25.5 3.48 α=β= 35.11 0.0044 Crystalline Tetragonal

750 γ = 900
WST- 27.7 3.21 α=β= 64.93 0.0022 Crystalline Tetragonal
1050 γ = 900

3.3 FTIR spectra of WS-TiO2

WS-TiO2 nanoparticle FTIR spectra exhibit many peaks between 4000 and 400 cm-1. The
peaks between 3400 and 3200 cm-1 show the existence of alcohol and phenol stretching. The
peak between 3000 and 2800 cm-1 corresponds to CH2 stretching. The peak at position 2342
cm-1 represents the presence of CO2 (C=O stretching) but as the temperature increases, the
carbon dioxide peak decreases. The C-C and C=C bonding are represented by the peak
between 1700 and 1600 cm-1. The peaks in the range 1600–1500 cm-1 indicates the existence
of the nitro group (Fig. 3). The C-O stretching is shown by the peaks between 1300 and 1000
cm-1 (which specify the presence of alkyl aryl, ether, ester, amine, 10, 20, and 30 alcohol
groups). The metal-oxygen bonding, or the titanium and oxygen bonding, is represented by a
range between 1000 and 400 cm-1. These bonds are related to the chemical composition of
plant i.e “Wisteria sinesis”. From FTIR, it is observed that these functional groups are present
the plant extract and also act as reducing agents for the metal ion.

Fig. 3 FTIR spectra of WST-750 nanoparticles

Table 2 include the detailed description about the functional group and their presence at
different positions when calcined at different temperatures such as 550, 750, and 1050 0C.

S. Functional Characteristic WST-550 WST-750 WST-1050

No. group absorption (cm-)

1. OH stretching 3400-3200 3277.6 3253.2 3230.6

2. CH2 stretching 3000-2800 2875.8 2871.8 2876.7

(alkane)

3. CO2 stretching 2400-2200 2342.1 2359.3 -

4. C=C stretching 1800-1600 1627.2 1627.3 1627.7

3.4 UV-VIS spectra of WS-TiO2

.
Fig. 4 displays the UV-VIS spectra of WS-TiO2 that was calcined at various temperatures.
Between 200 and 800 nm in wavelength, UV-Visible absorption spectra for synthesised
nanoparticles were noted. At 350 nm, the absorbance peaks are seen. The graph between E
(hυ) vs (αhυ)½ for synthesised WS-TiO2 was used to get the bandgap investigation, as
illustrated in Figure 6. As a result, the predicted bandgap energy values for WS-550 and WS-
750 bandgaps are 2.75 eV and 2.43 eV, respectively. The bandgap decreases with rising
temperature as a result of the presence of various crystal forms.
 Fig. 3 UV-VIS spectra of (a) WST-550 (b) WST-750

3.5 Morphological study and elemental composition of WS-TiO2

The form and morphological characteristics of synthetic TiO2 nanoparticles are described
by SEM microscopy. The form of the nanoparticles is visible in the first photograph (Fig. 5).
The spherical and aggregation-like shape of the nanoparticles can be seen in the photograph.
And in EDX spectra, titanium and oxygen peaks were seen with atomic compositions of
5.22% and 32.29%, respectively (Fig. 6). Additionally, two additional elemental peaks,
nitrogen and carbon, were seen in the EDX spectra. These two elements are found in the plant
extract of "Wisteria sinensis" and operate as an organic reducing agent to promote stability
with atomic compositions of 14.25% and 48.25%, respectively.

Fig. 4 SEM images of synthesized WST-550

Fig. 5 EDS micrograph of synthesized WST-550

3.4 Antimicrobial activity of WS-TiO2

The gram positive bacteria Staphylococcus aureus (ATCC 43300) and the gram negative
bacteria Pseudomonas aeruginosa (PA01), Escherichia coli, and Klebsiella pneumonia
(B5055) were used to test the antibacterial activity of the synthesised WST-550 nanoparticles
(Fig. 6).
 TiO2 nanoparticles, which have a positive charge, and microorganisms, which have a
negative charge, are attracted to one another by electromagnetic forces, which results in an
oxidation reaction. Due to their small size, TiO2 nanoparticles can easily enter bacterial cell
walls. By forming coordinating bonds with substances including thiols, amides,
carbohydrates, indoles, and hydroxyls, they aid in the inactivation of cellular enzymes and
nucleic acids of germs.
 Fig. 6 Antibacterial activity of WST-550 towards various gram positive and gram negative
bacteria.
The bacterial cell membrane develops pores during this process, breaking the cell wall and
resulting in cell death.
4. Conclusion
The present work describes the plant mediated synthesis of TiO2 nanoparticles using leave
extract of WS that act as reducing agent and capping agent both. The obtained nanoparticles
were characterized using different techniques like UV-VIS, FTIR, SEM-EDX, and XRD. All
these techniques confirm the formation of nanoparticles by giving characteristic peak in
different analysis. In addition, antimicrobial activity was performed against different gram
positive and gram negative bacteria using synthesised nanoparticles. Size of the plant
mediated nanoparticles has peculiar effect on biological characteristic of the bacteria. With
the help of morphological study, different aspects are discussed in order to establish
mechanism of antimicrobial activity of the nanoparticles..

Acknowledgment

The authors are thankful to the Director, Sant Longowal Institute of Engineering and
Technology, Longowal, Punjab for providing essential assistance in conducting experimental
work.
Conflict of interest
There is no conflict of interest to be declared.
References