Journal of Research in Medical and Dental Science

2024, Volume 12, Issue 2, Page No: 19-23

Copyright CC BY-NC 4.0
Available Online at: www.jrmds.in
eISSN No. 2347-2367: pISSN No. 2347-2545

Antimicrobial Activity of Piperine Based Zinc Oxide Nanoparticle against

Dental Pathogens

Kritheka CK*, Sarita Bandari

Department of Orthodontics, Saveetha Institute of Medical and Technical Sciences, Tamil Nadu, India

ABSTRACT
Background: The Piperaceae family, including black pepper, is known for its antibacterial properties. The oral cavity is
a major source of microorganisms, including plaque biofilms, which cause infections. Advancements in nanotechnology
have led to a growing interest in antibacterial therapies based on nanoparticles. Black pepper, a member of the
Piperaceae family, contains polyphenolic substances used in traditional medicine for treating illnesses like melano
derma and leprosy. In this study, Piperine incorporated with zinc oxide nanoparticles was tested for efficacy against
dental pathogens.
Aim: To determine the antimicrobial activity of Piperine based zinc oxide nanoparticle against dental pathogens
Materials and methods: Zinc acetate dehydrate and NaOH were used as precursors in the direct precipitation technique
to create zinc oxide nanoparticles, which were then examined under a scanning electron microscope. Using the 2,
2-diphenylpicrylhydrazyl (DPPH) experiment, antioxidants' capacity to scavenge radicals is assessed. 2.2’-azino-bis
(3-ethylbenzothiazoline-6-sulfonate) are the ABTS test used to quantify the interaction between an antioxidant and
the cation of the pre-generated ABTS+ radical. The Zone of Inhibition test is used to assess a pathogen's sensitivity to
antibacterial agents or resistance to them. The outcomes were assessed.
Results and discussion: In this study zinc oxide nanoparticles were doped with piperine and SEM was demonstrated.
The SEM images revealed that the uniform distribution and almost spherically shaped piperine mediated ZnO NPs is
observed. The DPPH assay & ABTS assay shows antioxidant activity of ascorbic acid, piperine & piperine doped ZnO
NP against oral pathogens. In DPPH assay & ABTS assay, there was an increased inhibition rate in piperine mediated
ZnO NP when compared ascorbic acid and piperine. Therefore results concluded that piperine mediated zinc oxide
nanoparticles showed very good antimicrobial activity against all oral pathogens such as S. mutans, C. albicans and
E. faecalis.
Conclusion: Nano-dentistry is a developing field with the potential to address new and improved applications in dentistry,
and it has provided a new avenue for revolution in oral care. Our study conclusions suggested that PP-mediated ZnO
NPs may provide a useful nanomaterial for the creation of drugs intended to treat oral infections. ZnO nanoparticles
mediated by PP showed remarkably potent antibacterial activity against S. mutans, Candida albicans, and E. faecalis,
among other oral pathogens. When using 28-nm PP ZnO NPs against E. faecalis, a greatest inhibition zone was seen.
Key words: Piperine mediated Zinc oxide nanoparticles, S.mutans, Candida albicans, E. faecalis, and Nano dentistry.

HOW TO CITE THIS ARTICLE: Kritheka CK, Sarita Bandari. Antimicrobial Activity of Piperine Based Zinc Oxide Nanoparticle against Dental
Pathogens. J Res Med Dent Sci, 2024, 12(2):19-23.

Corresponding author: Kritheka CK harmful germs from external pharmacological agents

e-mail: [email protected] and evading the host defensive mechanisms. It is a
Received: 26-January-2024, Manuscript No. jrmds-23-120434; primary cause of dental illnesses, including caries
Editor assigned: 29-January-2024, PreQC No. jrmds-23-120434(PQ); and periodontitis [1]. Despite the fact that many
Reviewed: 12-February-2024, QC No. jrmds-23-120434(Q); research have been conducted in an attempt to
Revised: 17-February-2024, Manuscript No. jrmds-23-120434(R); develop antimicrobial agents to address this problem,
Published: 23-February-2024
most of these efforts have not been able to provide
the desired outcomes since antibacterial medications
INTRODUCTION quickly break down and release, which causes low
efficacy and safety concerns. With the growth in
An abundance of bacteria is always present in the bacterial resistance, it is imperative to conduct long-
mouth cavity. Plaque biofilm is a complex community term research on suitable alternatives to standard
of bacteria or fungi that produces infection by shielding antibacterial treatments. The developments in

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Journal of Research in Medical and Dental Science | Vol. 12 | Issue 2 | February 2024
Kritheka, et al. J Res Med Dent Sci, 2024, 12 (2):19-23

nanotechnology have led to a surge in interest MATERIALS AND METHODS

in the dental field for antibacterial treatments
based on nanoparticles because of their broad Characterization of Nanoparticle
antibacterial spectrum, stability, and outstanding Analyzing nanoparticles using a Scanning Electron
antibacterial properties at an inexpensive price [2]. Microscope (SEM) involves specific sample
And by overcoming the limitations of single therapy preparation methods to ensure proper imaging and
and boosting osteogenesis and remineralization accurate characterization. If the nanoparticles are
based on antibacterial qualities, the application of part of a larger structure or matrix, consider using
multifunctional nanomaterials has considerably fixation methods to preserve the overall structure. If
increased the long-term prevention and treatment of nanoparticles tend to agglomerate, employ methods to
oral disorders [3].
de-agglomerate them. This may involve sonication or
Nanomaterials are being used more and more in other dispersion techniques. Attach the nanoparticles
items like semiconductors and antimicrobial surface onto a clean SEM stub using a conductive adhesive,
coatings. Nanoparticles can differ significantly double-sided carbon tape, or another suitable
from micrometer-sized particles in terms of their mounting method. Ensure good electrical contact.
hardness, active surface area, chemical reactivity, and If the nanoparticles are non-conductive, consider
biological activity. In fact, it has been proposed that coating them with a thin layer of conductive material
the biocidal efficacy of metallic nanoparticles stems to enhance conductivity and reduce charging effects
from their large size as well as their high surface-to- during imaging. Carefully load the sample stub
volume ratio [4]. They should be able to intimately into the SEM chamber using the designated sample
interact with microbial membranes owing to these holder. Ensure the sample is securely mounted.
qualities, and the effect won't just come from the Calibrate the SEM instrument, including adjusting
release of metal ions [5].Despite this, because of their the electron beam energy, focus, and stigmation. Use
large specific surface area and high charge density, calibration standards to ensure accurate imaging
nano-antibacterial agent’s exhibit broad antibacterial and measurement. Choose SE imaging mode for
range and persistent antibacterial efficacy. Therefore, surface characterization of the nanoparticles. This
Reactive Oxygen Species (ROS) are produced by mode provides detailed surface information. Set the
catalytically active nanomaterials and can cause
working distance and magnification based on the size
fatal oxidative stress. It also has a high drug loading
and features of the nanoparticles. Start with lower
capacity, can enter eukaryotic cells, and release ions
magnifications for an overview before moving to
[6].
higher magnifications. Use image analysis software to
In this study piperine incorporated with zinc oxide measure particle size, shape, and distribution.
nanoparticles was used to assess the efficacy against DPPH free radical scavenging assay
dental pathogens. The Piperaceae family, which
includes black pepper (Piper nigrum L.), is widely In the DPPH free radical scavenging assay, test
samples are prepared at various concentrations, and
harvested for its fruit. The majority of the time,
equal volumes are mixed with a freshly prepared
pepper is used in curry preparations and is used in
DPPH solution in ethanol. The reaction mixture
Ayurveda and other traditional medicine cures [7].
is incubated in the dark at room temperature,
Because of the polyphenolic chemicals including
allowing for the scavenging of DPPH radicals by
tannins, volatile compounds, and phenols, it is used in
antioxidants present in the test samples. After the
folk medicine to cure a variety of conditions including incubation period, the absorbance of the reaction
leprosy, rheumatoid arthritis, cough, melano-derma, mixture is measured at around 517 nm using a
and peripheral neuropathy. It is also used as an spectrophotometer. The decrease in absorbance
antiseptic & antimicrobial agent [8].One of the main indicates the free radical scavenging activity of
reasons ZnO NPs are being studied as partners of the test samples. A blank control without the test
antimicrobial drugs is because zinc is an essential sample is included for background correction. The
trace element that is present in muscle, bone, skin, and percentage of DPPH radical scavenging activity is
the hard tissues of teeth [9].The "Trojan Horse Effect," calculated using the formula, and a dose-response
a recent idea, explains this by claiming that an acidic curve is plotted to visualize the scavenging activity at
lysosomal environment stimulates the breakdown of different concentrations. The IC50, representing the
nanoparticles, which in turn causes the conversion concentration required to scavenge 50% of the DPPH
of core metals into ions and the release of hazardous radicals, can be determined, providing a quantitative
chemicals that subsequently stop cell division [10]. measure of antioxidant efficacy. The assay is
Therefore the aim of this study is to determine the performed in triplicate or more, with appropriate
antimicrobial activity of piperine based zinc oxide controls and standards, to ensure accuracy and
nanoparticles against dental pathogens. reliability of the results.

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ABTS free radical scavenging assay as a valuable tool for screening and comparing the
In the ABTS (2, 2’-azino-bis (3-ethylbenzothiazoline- antimicrobial potential of different agents.
6-sulfonic acid)) free radical scavenging assay, a stock Statistics
solution of ABTS is reacted with potassium persulfate
The data presented in this study are the mean of
to generate the ABTS radical cation (ABTS). The ABTS
three replicates and their respective Standard
solution is then diluted to an absorbance of around
Deviation (SD). Also, the data were subjected to one-
0.70 at a specific wavelength usually 734 nm. Test
way ANOVA and post-ANOVA using Graph Pad Prism
samples, such as antioxidants or plant extracts, are
(version 5.0). Results were evaluated.
prepared at various concentrations and mixed with
the diluted ABTS solution. The reduction of the RESULTS
ABTS radical by antioxidants leads to a decrease
in absorbance, which is measured after a suitable In this study zinc oxide nanoparticles were doped
incubation period using a spectrophotometer. The with piperine and SEM was demonstrated. The SEM
scavenging activity is calculated as the percentage images revealed that the uniform distribution and
inhibition of the ABTS radical compared to a almost spherically shaped piperine mediated nO NPs
control without the test sample. A dose-response is observed in the SEM image in [Figure 1].
curve is generated, and the IC50, representing the
concentration at which 50% of the ABTS radicals Some piperine mediated ZnO NPs are present with
are scavenged, can be determined. The assay is a hexagonal shape and the average particle size is
performed in triplicate or more, with appropriate around 25-30 nm. Then DPPH Assay & ABTS assay
controls, to ensure the accuracy and reliability of were accessed, The DPPH assay & ABTS assay
the results. shows antioxidant activity of ascorbic acid, piperine
& piperine doped ZnO NP against oral pathogens
Antibacterial activity [figures 2 & 3].
The Zone of Inhibition assay is employed to
In DPPH assay & ABTS assay, there was an increased
evaluate the antimicrobial activity of substances.
inhibition rate in piperine mediated ZnO NP when
Initially, a microbial culture is evenly spread on a
compared ascorbic acid and piperine. And also
solid agar medium. Sterile paper discs or wells are
the zone of inhibition against Candida albicans,
then impregnated with the test substance, such as
streptococcus mutans, and Enterococcus faecalis were
antibiotics or plant extracts, and placed onto the
tabulated below [figure 4].
inoculated agar surface. Following an appropriate
incubation period at the microorganism's optimal The largest inhibition zone of PP ZnO NPs against
growth temperature, the plates are examined for E. faecalis were observed with 28-nm and while
the presence of clear zones surrounding the discs the largest inhibition zones against S. mutans and
or wells, indicating inhibition of microbial growth. C. albicans were observed with 26-nm & 23-nm.
The diameter of these zones is measured as the Zone Therefore results concluded that piperine mediated
of Inhibition, providing a quantitative measure of zinc oxide nanoparticles showed very good
the antimicrobial efficacy of the test substance. The antimicrobial activity against all oral pathogens such
assay is performed in triplicate or more for statistical as S. mutans, C. albicans and E. faecalis. In that the
reliability, and appropriate controls, including a blank piperine mediated zinc oxide nanoparticles show
disc or well, are included to account for any non- increased activity when compared with standard
specific effects. The Zone of Inhibition assay serves drugs.

Figure 1: Piperine mediated Zinc oxide SEM image.

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Kritheka, et al. J Res Med Dent Sci, 2024, 12 (2):19-23

Figure 2: DPPH Assay.

Figure 3: ABTS Assay.

Figure 4: zone of inhibition rate.

DISCUSSION zinc oxide nanoparticles against S. mutans, Candida

albicans, and Enterococcus faecalis, three oral
Antimicrobial nanomaterials were incorporated pathogens. For each of the three pathogens, the PP-
into dental materials as a promising substitute for mediated ZnO NPs displayed a zone of inhibition.
conventional antibacterial medications, leading to Compared to the other two, the zone of inhibition
numerous advancements in treatment approaches for E. faecalis was larger, indicating a stronger effect
[11]. Nanomaterial-based system advancements can of PP-mediated ZnO nanoparticles against the same.
give unique solutions in the prevention and treatment The highest inhibition zone of PP ZnO NPs against E.
of dental illnesses, particularly in the control of oral faecalis was seen in the current investigation at 28 nm,
pathogenic microorganisms [12]. which was consistent with prior studies revealed that
lower concentrations of nZnO eliminated E. faecalis
Zinc oxide nanoparticles can bind to bacterial cells in contrast to other microbes [14].The generation
and subsequently enter them, producing structural of Active Oxygen Species (AOS), which cling to the
alterations in the cell membrane such as increased cell surface or collect in the cytoplasm of the cell, is
permeability and cell death [13]. This study examined responsible for the significant antibacterial action of
the antimicrobial activity of Piperine mediated nZnO on E. faecalis [15].
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