J Nanostruct 13(1): 151-158, Winter 2023

RESEARCH PAPER

Green Synthesis of MnO2 Nanoparticles Using Cumin Extract

Composited with Hypericum Plant: Investigation of Antibacterial
and Anticancer Properties
Sareh Ghorbani 1, Younes Mirzaei 1,2, Mohammad Mahdi Bordbar 3 Ali Gholami 2*
1
Ghiaseddin Jamshid Kashani Student Research Institute, kashan, 38576704, Iran
2
Department of Analytical Chemistry, Faculty of Chemistry, University of Kashan, Kashan, 87317-51167, Iran
3
Personal Laboratory, Fasa, 74614, Iran

ARTICLE INFO ABSTRACT

Cancer is a major cause of morbidity and mortality. Although there are
Article History: many treatment techniques, these methods still have many limitations.
Received 11 October 2022
Nanotechnology is rapidly growing and flourishing in the field of diagnosis
Accepted 26 December 2022
and treatment of various diseases. The antibacterial and anticancer
Published 01 January 2023
properties of nanoparticles (NPs) have made them promising for the
Keywords: treatment of diseases. In this study, manganese dioxide (MnO2) NPs are
green-synthesized by the cumin extract as a regenerator and stabilizer in
Antibacterial properties
a co-precipitation method. The synthesized NPs are then composited with
Anticancer propertie
the Hypercom perforatum plant. In order to characterize the structure
Cumin extract
and morphology of MnO2 NPs, XRD, FTIR, SEM, and TEM analyses are
Hypercom perforatum performed. MnO2 NPs composited with the Hypericum plant were tested
Manganese dioxide nanoparticles for their antibacterial efficacy against strains of Staphylococcus aureus and
Pseudomonas aeruginosa. The MTT cytotoxicity assay was used to assess
the anticancer effects of MnO2 NPs composited with the Hypericum plant.
In this instance, it was looked into how the NPs affected the human Hela
cervical cancer cell line and the cancerous fibroblast cells. Based on these
results, MnO2 NPs can damage Hela cells without damaging healthy cells.
This investigation can provide fundamental insights for understanding
the antibacterial activity and cytotoxicity of these NPs and point out their
applicability for use in the biomedical field.

How to cite this article

Ghorbani S., Mirzaei Y., Bordbar M., Gholami A. Green Synthesis of MnO2 Nanoparticles Using Cumin Extract Composited
with Hypericum Plant: Investigation of Antibacterial and Anticancer Properties. J Nanostruct, 2023; 13(1):151-158. DOI:
10.22052/JNS.2023.01.017

INTRODUCTION
Nanoscience and nanotechnology are and low degradability [1]. Due to their small size
significant scientific fields that deal with the and high surface-to-volume ratio, NPs have high
use and development of materials made at the effectiveness and durability, being employed
nanoscale. Nanoparticles (NPs) with sizes in the in various fields, including biological, physical,
range of 1‒100 nm can be produced through chemical, pharmaceutical, and engineering
numerous methods. However, these methods sciences [2].
use chemicals that are with a high risk of toxicity Although not all bacterial species are pathogenic,
* Corresponding Author Email: agholami@kashanu.ac.ir dangerous diseases such as cholera, tuberculosis,

This work is licensed under the Creative Commons Attribution 4.0 International License.
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 S. Ghorbani et al. / Antibacterial and Anticancer Properties of Green Synthesis of MnO2 NPs

typhus, and anthrax, as well as infections such as medicine. In recent decades, a number of
blood and urinary infections, wound infections, pharmaceutical and phytochemical studies
and many other diseases can be caused by some has shown that the antitumor, antidepressant,
species of bacteria [3]. Staphylococcus aureus is antibacterial and anti-inflammatory activities of
a gram-positive bacterium and one of the most Hypericum species are due to the presence of
prominent pathogenic species of staphylococci. bioactive compounds, including phloroglucinol,
Nevertheless, it has become one of the public xanthone, and flavonoid derivatives [12].
health concerns due to its intrinsic resistance to NPs can cause three types of damage to
antimicrobial agents and drugs [4]. In the last 50 bacteria: i) they can be located near the bacterial
years, Staphylococcus aureus has undergone many cell and destroy the cell without effectively
genetic changes. This bacterium has a flexible connecting to its external structure (e.g., the
genome, leading to an increase to its pathogenic and cell wall or cytoplasmic membrane), ii) they can
drug-resistant strains [5]. Staphylococcus aureus destroy the cell through electrostatic attraction,
settles in the nose and skin (especially damaged hydrogen bonding, or van der Waals interaction,
skin), vagina, armpit, and navel of newborn babies and iii) they can penetrate into the cell and
[6]. Pseudomonas aeruginosa is a gram-negative directly damage the components inside it (e.g.,
bacterium belonging to the Pseudomonadaceae cytoplasmic proteins or DNA), thus destroying the
family, having the possibility to survive in a wide cell [13]. On the other hand, NPs and drugs can be
range of environments. Pseudomonas aeruginosa used with precise targeting to treat cancer cells.
is known as an opportunistic pathogen and the This approach is called targeted therapy, being
most common bacterium associated with hospital a growing part of treatment for many types of
infections [7]. This bacterium leads to fatal cancer.
infections in immunocompromised people such Due to its excellent physical and chemical
as patients with cancer post-surgery and severe properties, cost-effectiveness, structural diversity
burns or infected with human immunodeficiency and biocompatibility, manganese dioxide (MnO2)
virus [8]. with semiconducting properties has been widely
Cancer is one of the health problems of today’s used in various fields of science and technology,
societies, and many efforts are being made to deal including sensors [14], energy storage [15],
with it. In fact, cancer is complex, inconsistent catalysts [16], bioimaging, and drug release [17,
and heterogeneous, arising from genetic changes. 18]. In general, MnO2 NPs can act as a multipurpose
Many factors are responsible for the proper therapeutic agent to improve tumor therapy [19].
growth and proliferation of normal cells, being With regard to structural properties, MnO2 NPs
appropriate only when the cell proliferation is tend to have a linear geometry in the form of
carefully controlled. In other words, cancer occurs O-Mn-O chains, comprising flat zigzag chains in
if the regulation of cell proliferation processes one plane [20].
is not made correctly, thereby disrupting the The use of plant extracts as reducers and
normal function and behavior of cells [9, 10]. surfactants is much more efficient than the
As the fourth most common cancer in women, chemical method, owing to the presence of various
cervical cancer accounts for approximately 12% of biological molecules in plants [21]. Many effective
women’s cancers, and is considered the seventh approaches and methods, including hydrothermal,
most common cancer in the world. The majority co-precipitation, sol-gel, and microwave have so
of global cancer cases occur in less developed far been proposed for the synthesis of MnO2 NPs
countries [11]. [22]. The above-mentioned synthesis methods
Hypericum perforatum is a valuable medicinal involve the utilization of chemical surfactants
plant from the Hypericaceae family. Hypercom that can have effects on the environment. This
species are used in different parts of the world can be reduced to some extent by employing
such as Europe, America, Africa, and Asia, having green chemistry and plants as surfactants. Green
astringent, diuretic, analgesic, antidepressant, chemistry is a new innovative method that aims
and disinfection properties. In addition to the to minimize possible harm to humans and the
aforementioned properties, Hypericum species environment. The production of NPs by the
are used to treat wounds, burns, and bites of green chemistry method together with the use
poisonous animals in traditional and Chinese of plants has attracted considerable attention as

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 S. Ghorbani et al. / Antibacterial and Anticancer Properties of Green Synthesis of MnO2 NPs

a fast, affordable, and biocompatible approach. Hypericum plant and stirred for 24 h to form the
However, there is no report on the use of cumin desired nanocomposite.
extract and Hypericum perforatum plant for the
synthesis of MnO2 NPs and the investigation of RESULTS AND DISCUSSION
their antibacterial and anticancer properties, X-ray diffraction (XRD) pattern
according to the best of our knowledge. The X-ray diffraction (XRD) pattern of MnO2
In this paper, MnO2 NPs are synthesized by NPs measured in 2θ range of 10°‒80° is shown in
using co-precipitation and green chemistry Fig. 1. Crystal structure of the NPs is found to be
methods with cumin extract. The resulting NPs α-tetragonal phase, corresponding to the standard
are then composited with Hypericum plant. card number (044-0141-0141) [23]. No impurity
They are also characterized in terms of crystal or secondary phase is observed in the pattern.
structure, morphology, and chemical state. The size of the crystallites was estimated using
Alternatively, antibacterial properties of the Scherrer equation (indicating the dependence
MnO2 NPs in inhibiting Staphylococcus aureus of particle size on the spread and broadening of
and Pseudomonas aeruginosa bacteria, and their diffraction lines) as given below:
anticancer properties against cervical cancer cells 𝐾𝐾𝐾𝐾
are investigated. These results give insights into 𝐷𝐷 = (1)
𝛽𝛽 cos 𝜃𝜃
the efficient and potential applications of the
green synthesized MnO2 NPs for targeted therapy. where D is the average size of the crystallites,
𝜆 is the wavelength of the X-ray radiation, K is
MATERIALS AND METHODS the shape factor (0.9), β is the full width at half
Co-precipitation and green chemistry methods maximum (FWHM) of the main peak, and θ is the
were used to synthesize MnO2 NPs. To this end, Bragg angle. Accordingly, the average crystallite
0.47 g KMnO4 precursor was dissolved in 20 size of the green synthesized MnO2 NPs is equal to
ml of deionized water. Cumin extract was then D= 20 nm.
added drop by drop to the previous solution and
stirred at 40 °C for 2 h using a magnetic stirrer. Scanning and transmission electron microscopy
The resultant solution was dried in an oven at 80 (SEM and TEM) images
°C. The powder obtained was calcined at 400 °C To study morphological properties of the MnO2
for 2 h. Finally, the NPs were combined with the NPs, scanning electron microscopy (SEM) and

Fig. 1. XRD pattern of MnO2 NPs synthesized by the co-precipitation and green chemistry methods.

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Fig. 2. (A) SEM image of MnO2 NPs synthesized by co-precipitation and green chemistry methods. The inset shows the
corresponding nanoparticle size distribution histogram. (B) SEM image of MnO2 NPs composited with the Hypericum plant.

transmission electron microscopy (TEM) analyses of MnO2 NPs composited with the Hypericum
were carried out, and the results obtained are perforatum plant. As can be seen, the resulting
shown in Figs. 2 and 3, respectively. From Fig. 2(A), composite is less agglomerated than the as-
SEM image shows that the as-calcined MnO2 NPs calcined product. In other words, the MnO2 NPs
have sizes in the range between 10 and 45 nm (see are spread homogeneously on the surface of the
the inset). The average size of the NPs is found to plant, confirming their efficient attachment.
be approximately 25 nm. It should be noted that On the other hand, TEM images of the as-
the MnO2 NPs are not homogenous and uniform calcined and composited MnO2 NPs are depicted in
in all areas due to adhesion and agglomeration. Fig. 3. The TEM image (Fig. 3(A)) of the as-calcined
This can be caused by the calcination and product reveals that the MnO2 NPs have spherical-
exposure to the high temperature, leading to a like morphology. The accumulation of some NPs
tendency of the NPs to clump together to reach and the enlargement of their dimensions can also
the minimum energy. Fig. 2(B) shows SEM image be observed. The TEM image (Fig. 3(B)) of the NPs

Fig. 3. (A) TEM image of MnO2 NPs synthesized by co-precipitation and green chemistry methods. (B) TEM image of MnO2 NPs
composited with Hypericum plant.

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 S. Ghorbani et al. / Antibacterial and Anticancer Properties of Green Synthesis of MnO2 NPs

composited with the Hypericum plant indicates 500‒600 cm-1 are related to Mn-O-Mn vibrations.
the uniform combination of these two materials, These peaks are also observed in Fig. 4(C), thereby
confirming the results obtained from the SEM confirming the binding of the NPs to the plant
analysis. [24]. The absorption peaks in the 3400‒1630 cm-1
region are related to the stretching and bending
Fourier transform infrared (FTIR) spectra vibrations of H-O bond, which is adsorbed on the
The formation of ligands and identification surface of the molecule. The 1385 cm-1 band is
of the molecules and functional groups were related to bending vibrations of the H-C bond. The
performed by obtaining Fourier transform infrared absorption peak at 1270 cm-1 in the Hypericum
(FTIR) spectra of the as-calcined MnO2 NPs, plant (Fig. 4(B)) and NPs composited with the
Hypericum plant, and MnO2 NPs composited with plant indicates the presence of C-O aromatic
Hypericum. These results are shown in Fig. 4. From carbon compounds. As well, the existence of CO-
Fig. 4(A), the absorption peaks in the range of O-CO stretching vibrations is observed due to the

Fig. 4. FTIR spectra of: (A) as-calcined MnO2 NPs, (B) Hypericum plant, and (C) MnO2 NPs
composited with Hypericum plant.

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 S. Ghorbani et al. / Antibacterial and Anticancer Properties of Green Synthesis of MnO2 NPs

Fig. 5. The effect of different concentrations of MnO2 NPs composited with Hypericum plant on the
reduction of biofilm formation in the standard strains of Staphylococcus aureus ATCC 43300 and
Pseudomonas aeruginosa PAO1.

Table 1. Minimum inhibitory concentration (MIC) and minimum bacterial concentration (MBC) of MnO2 NPs composited with
Hypericum plant for Staphylococcus aureus ATCC 43300 and Pseudomonas aeruginosa PAO1.

MnO2 NPs composited with Hypericum plant

MIC MBC
Microorganism
Minimum−Maximum Minimum−Maximum
Mean ± SD Mean ± SD
(mg/ml) (mg/ml)

S.aureus ATCC 43300 9.50-9.75 9.6±0.13 19.5-39 32.5±11.2

P.aeruginosa PAO1 13.00-13.10 13.50±0.050 27 27

appearance of the peak at 1000 cm-1. the Pseudomonas aeruginosa PAO1 and
Staphylococcus aureus ATCC 43300, the resulting
Antimicrobial activity minimum−maximum MIC and MBC indicate
Antibacterial activity of MnO2 NPs composited that the NPs can be used as a medicine to kill
with the Hypericum plant was investigated bacteria. The effect of different concentrations
for Staphylococcus aureus and Pseudomonas of NPs on the reduction of biofilm formation
aeruginosa strains, and the results obtained are in the Staphylococcus aureus ATCC 43300 and
shown in Fig. 5 and Table 1 [25]. Pseudomonas Pseudomonas aeruginosa PAO1 standard strains
aeruginosa is a gram-negative bacterium that is was also studied. Each microplate was used for
widespread in all parts of the world and is one of two bacteria, and a stock of NPs with the same MIC
the essential hospital-causing bacteria [26]. concentration was prepared for each bacterium.
Based on the tests performed against The concentration of NPs in the wells was 1/8

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 S. Ghorbani et al. / Antibacterial and Anticancer Properties of Green Synthesis of MnO2 NPs

Fig. 6. The toxicity graph of MnO2 NPs composited with Hypericum plant in Hela and fibroblast
cells. The vertical and horizontal axes show the cell viability percentage and different
concentrations of the compound treated with the cells, respectively.

MIC, 1/4 MIC, and 1/2 MIC. Moreover, the control the co-precipitation method using cumin extract,
was considered with no NPs in one row. As can be and then composited with Hypericum plant. XRD
seen in Fig. 5, the biofilm formation percentage analysis confirmed the high purity of the NPs
for the two bacteria decreases significantly with with α-tetragonal phase, and the crystallite size
increasing the concentration. of the NPs was estimated to be around 20 nm.
The functional groups were investigated and
Anticancer properties shown by FTIR analysis. Also, the morphology of
In order to evaluate anticancer properties of NPs was investigated by SEM and TEM analyses.
MnO2 NPs composited with the Hypericum plant, The resulting nanocomposite was tested in
MTT cytotoxicity test was performed. In this the laboratory to check its antibacterial and
case, the effect of the NPs on the human cervical anticancer properties (MTT test), demonstrating
cancer cell line (Hela) and fibroblast cancer cells good results in inhibiting the formation of biofilms
was investigated, and the results obtained are of Staphylococcus aureus and Pseudomonas
shown in Fig. 6. As inferred, the inhibition of Hela aeruginosa bacteria. Thus, this nanocomposite
and fibroblast cancer cells increases by increasing can be used in media with the above-mentioned
the concentration of the NPs. Note that the rate bacteria. Furthermore, MnO2 NPs composited
of inhibition in Hela cells is higher than that in with the Hypericum plant provided favorable
fibroblast cancer cells. Accordingly, the MnO2 effects on cervical cancer cell inhibition, making
NPs composited with the Hypericum plant can be them suitable for delivering to the target cancer
selected as a good candidate for the treatment of cell by targeted drug delivery while also preventing
Hela cancer. In addition, owing to the presence the harmful effects of chemotherapy drugs and
of NPs in the composition, this cancer cell can be radiation in radiation therapy on healthy cells.
treated in a targeted manner.
CONFLICT OF INTEREST
CONCLUSION The authors declare that there is no conflict
MnO2 NPs have been initially synthesized by of interests regarding the publication of this

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 S. Ghorbani et al. / Antibacterial and Anticancer Properties of Green Synthesis of MnO2 NPs

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