Analysis of antibacterial activity of jalapa, Solanum
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Pure Appl. Biol., 12(1):683-693, March, 2023
http://dx.doi.org/10.19045/bspab.2023.120070
Research Article
Analysis of antibacterial activity of
indigenous Mirabilis jalapa, Solanum
nigrum and Aloe vera
Rameez Nisar1, Raja Tahir Mahmood2*, Ansar Ahmed Abbasi1, Khan
Muhammad3, Khalid Hameed1, Muhammad Asghar Khan4, Tauqeer
Ahmed Mughal1, Saiqa Khushal1, Rabia Shaukat1 and Maleeha
Masood2
1. Department of Zoology, Mirpur University of Science and Technology (MUST), Mirpur (AJK) Pakistan
2. Department of Biotechnology, Mirpur University of Science and Technology (MUST), Mirpur (AJK) Pakistan
3. Azad Jammu and Kashmir Medical College, Muzaffarabad, Pakistan
4. National University of Medical Sciences, Rawalpindi, Pakistan
*Corresponding author’s email: raja.tahir@must.edu.pk
Citation
Rameez Nisar, Raja Tahir Mahmood, Ansar Ahmed Abbasi, Khan Muhammad, Khalid Hameed, Muhammad
Asghar Khan, Tauqeer Ahmed Mughal, Saiqa Khushal, Rabia Shaukat and Maleeha Masood. Analysis of
antibacterial activity of indigenous Mirabilis jalapa, Solanum nigrum and Aloe vera. Pure and Applied Biology.
Vol. 12, Issue 1, pp683-693. http://dx.doi.org/10.19045/bspab.2023.120070
Received: 15/11/2022 Revised: 11/01/2023 Accepted: 16/01/203 Online First: 30/01/2023
Abstract
In this study, the antimicrobial potential of medicinal plants was determined against three
gram-negative: Klebsiella pneumonia, Pseudomonas aeruginosa and Escherichia coli, and
four gram-positive bacteria: Streptococcus pyogenes, Bacillus subtilis, Enterococcus and
Staphylococcus aureus. Aqueous extracts of three medicinal plants namely Mirabilis jalapa,
Solanum nigrum and Aloe vera were used to check the antimicrobial activity. The antimicrobial
potential of these plants was determined by agar well diffusion method. The antimicrobial
activity of aqueous extract of Aloe vera was better than Solanum nigrum, followed by Mirabilis
jalapa which exhibited lesser activity. Maximum zone of inhibition was shown by K.
pneumonia 22 mm at 100 µL concentration for A. vera extract. The was no antibacterial activity
against B. subtilis and Enterococcus by A. vera. Maximum zone of inhibition was exhibited by
K. pneumoniae and B. subtilis 20 mm at 100 µL concentrations of M. jalapa. The
Enterococcus, S. pyogenes and P. aeruginosa did not show any result against M. jalapa extract.
The highest zone of inhibition was exhibited by S. pyogenes 17 mm at 100 µL concentration.
E. coli and K. pneumoniae did show any activity against plant.
Keywords: Antibacterial activity; A. vera; B. subtilis; Medicinal Plants; S. nigrum
Introduction resistant microbes that are intensifying
It has been reported that medicinal herbs globally, which is a massive challenge to
would be the exceptional source to obtain healthcare. Therefore, there is an urgent
numerous antibiotics. The majority of need to take action so that antibiotics might
people in urbanized nations nearly 80% use lose their power to cure maladies [2]. It is
traditional medicines that contain difficult to cure serious community and
ingredients derived from medicinal herbs hospital-acquired infections caused by
[1]. World Health Organization giving multidrug-resistant bacteria by taking the
awareness to people about multidrug available antibiotics [3].
Published by Bolan Society for Pure and Applied Biology 683Nasir et al.
Multiple factors such as specific nature of medicinal herbs as sources for the
the connection between microorganisms extraction of potent pharmacological
and antibiotics, the use of antimicrobial substances [11].
agents, host features, and environmental Therapeutic and scented herbs and their
conditions, all contribute to the spread of soul are enriched in antimicrobial
drug resistance. Due to this problem, components which could be a substitute to
researchers have been forced to hunt for fight microbial infections even against few
novel antibacterial chemotherapeutic microscopic organisms that are becoming
agents from a variety of sources; resistant to various artificial drugs [12-14].
nevertheless, the manufacturing of artificial The most significant constituents of herbs
antibiotics is extremely expensive and has are alkaloids, flavonoids, tannins as well as
undesirable side effects when compared to phenolic components [15]. Due to the
antibiotics derived from plants [4]. presence of significant levels of antioxidant
Multi-drug resistance poses severe threat to and antibacterial phyto-constituents, many
the medicinal world and diseases brought plant species have been ingested. As a
about by multi-resistant microorganisms result, a significant source of antioxidant
particularly in the serious consideration and anti-aging characteristics can be found
units create an enormous issue [5]. As a in the extracts of medicinal plants and
result, herbs are a priceless source of natural goods [16]. Many human infections
healing substances. Additionally, the main are caused by oxidative pressure that results
components of many antibiotics are still from unevenness between the arrangement
derived from natural sources, unless and neutralization of pro-oxidants [17].
otherwise directed [6]. Man cannot live on Numerous secondary metabolites that are
our planet for a long, healthy, and generous produced by plants are used in the
existence without the help of the plant pharmaceutical industry as lead molecules
kingdom since herbal products and their or as direct precursors. However, only a
dynamic components are essential for small portion of the 400,000 plant species
sustaining excellent health. The world is on earth have had their antibacterial
blessed with a wealth of restorative herbs properties thoroughly researched [18].
[7]. In the last few decades, increased germ Materials and Methods
resistance to commonly used antibiotics has Plant sample collection
generated discussion on a global scale. As a The plant materials used in this study were
result, demand from consumers for natural obtained from different areas of Azad
anti-microbial compounds is progressively Jammu and Kashmir. Solanum nigrum was
rising. As an alternative therapeutic agent collected from Mirpur AJ&K. Aloe vera
in the pharmaceutical sector, natural was collected from Forest office Mirpur
antibacterial agents have attracted a lot of AJ&K, while, Mirabilis jalapa was
attention [8]. collected from Tatta pani District Kotli
These plants produce drugs that are easily AJ&K.
accessible, affordable, safe, effective, and Bacterial isolates
rarely come with adverse effects. The most The bacterial isolates selected for this study
obvious starting point for novel were collected from Microbiology section
therapeutically effective medications, such of the Pathology Labs of following
as anticancer drugs and antibacterial hospitals of Rawalpindi/Islamabad,
pharmaceuticals, is plants that have been Pakistan. Staphalococus aureus and
favoured for medical usage over thousands Enterococcus were collected from PIMS
of years [9, 10]. Despite recent Islamabad. The strains of Bacillus subtilis
breakthroughs in the field of chemotherapy, and Streptococcus pyogenes were obtained
the use of medicinal plants has grown. The from Combined Military Hospital (CMH)
explanations advanced include the use of Rawalpindi, while, Psedomanas
684Pure Appl. Biol., 12(1):683-693, March, 2023
http://dx.doi.org/10.19045/bspab.2023.120070
aeroginosa, Escherichia coli and Klebisella mL of 0.18 M H2SO4 with constant stirring.
pneumoniae were collected from Al-shifa After mixing, solution was autoclaved and
International hospital Islamabad. Bacterial stored in refrigerator for 24 hours. After 24
isolates were stored in the refrigerator at 4 hours, the turbidity of saline solution was
°
C. adjusted for comparing the test and
Culture media and chemicals standard were compared against a white
In this research, cultivation of cells was background with a contrasting black line.
done by using nutrient agar (oxoid # Antibacterial activity testing by well
CM003). The saline solution (0.9 % diffusion method
McFarland unit) was also used in this lab For well diffusion method, nutrient agar
work. was prepared and autoclaved. Then it was
Inoculum preparation poured into sterilized plates and then let it
The stock of each of the microbes were solidify. And plates were incubated at 37 oC
revived and kept up on supplement agar for 24 hours to check the sterility. Agar
plates. The cultures were prepared by plates without any contamination were
streaking on sterile nutrient agar plates and selected for inoculation. Pre-sterilized
kept in incubator at 37 °C for 24 hours and saline solution was inoculated with fresh
utilized as sub-cultures. Microbial cultures cultures and the turbidity of the cultures
were revived after every 3 to 5 days to was adjusted. Sanitized cotton swab was
evade contamination. dipped into diluted cultures and microbial
Preparation of plant extract lawn was arranged over the nutrient agar
Fresh extract plates. Wells having 6 mm diameter were
Before extraction, leaves of Solanum prepared by the help of sterilized cork borer
nigrum were washed with tap water and (dipped in spirit, followed by incineration)
then, with distilled water thrice times to in the seeded agar using the previously
remove contamination or dust. Then leaves marked places and the borer was used to
were crushed and ground with mortar pestle remove the agar from the marked points.
and was filtered through filter paper and Wells were filled with 100 µL of each plant
kept in the refrigerator for the future use. extract by the help of micropipette using
The A. vera juice and M. jalapa extracts sterilized tips and then incubated for 24
were taken through filtration with the help hours at 37 oC. The zones of inhibition were
of mesh cloth. The 40 mL of A. vera juice observed after 18 to 24 hours incubation.
was soaked in 40 mL of distilled water and The same well diffusion method was used
placed in water bath for 24 hours and placed to check the minimum inhibitory
in refrigerator. concentration of plant extracts. Four wells
Preparation of saline solution and of 100 µL, 40 µL, 20 µL and 10 µL were
McFarland solution made in each agar plate. Each well of
By mixing 0.9 g of NaCl with 100 mL of different concentration was filled with
distilled water and sterilising it in an respective concentration by sterilized
autoclave, saline solution was created and micro-pipettes and then incubated for 24
placed in an incubator at 37 oC for 24 hours. hours at 37 °C. The zones of inhibition were
On next day, saline solution was inoculated measured in millimeters after 18 to 24
with bacterial cultures and kept on shaking hours.
until the cultures were mixed in saline Results
solution. Then, McFarland (0.5 McFarland Antimicrobial activity of A. vera, M. jalapa
turbidity standard) solution was used to and S. nigrum were determined against
adjust the turbidity of the strains by adding seven bacterial species namely: S.
sterile distilled water. The McFarland pyogenes, P. aeruginosa, K. pneumonia,
standard solutions was prepared by adding Enterococcus, E. coli, S. aureus and B.
0.05 mL of barium chloride (BaCl2) in 9.95 subtilis. All these herbs showed effective
685Nasir et al.
inhibitory activity against some of the The antimicrobial activity of aqueous
tested microbes and ineffective against extract of Aloe vera was better than S.
others. The result of antimicrobial activity nigrum, followed by M. jalapa, which
of aqueous extracts of three plants against exhibited lesser activity.
seven bacterial isolates are presented in the
(Fig. 1).
25
Inhibitory zones
20 E.coli
15 P.aeruginosa
(mm)
K.pneumoniae
10
Enterococcus
5
S.aureus
0
A.vera M.jalapa S.nigrum S.pyogenes
Plant extracts B.subtilis
(µL)
Figure 1. Zones of inhibition of three plant extracts against pathogenic bacteria in mm
A. vera plant showed antimicrobial activity level activity of activity was revealed by S.
against five pathogens and two pathogens pyogens, P. aeruginosa, E. coli and
showed resistance as shown in (Fig 2). Four Enterococcus with a zone of inhibition 20
different concentrations 10, 20, 40 and 100 mm, 15 mm, 14 mm, 14 mm, respectively
µL concentrations were used against each at 100 µL concentration. The B. subtilis and
tested pathogen. Maximum zone of Enterococcus did show any antimicrobial
inhibition was shown by K. pneumonia 22 activity against A. vera.
mm at 100 µL concentration. Moderate
25
20
Inhibitory zones
E.coli
15 P.aeruginosa
(mm)
K.pneumoniae
10
Enterococcus
5 S.aurues
0 S.pyogenes
10 20 40 100 B.subtilis
Concentrations
(µL)
Figure 2. Zones of inhibition against seven bacterial strains by extract of A. vera
686Pure Appl. Biol., 12(1):683-693, March, 2023
http://dx.doi.org/10.19045/bspab.2023.120070
M. jalapa shown activity against four and B. subtilis 20 mm at 100 µL
pathogens and three pathogens showed concentrations. Moderate level of activity
resistance as shown in (Fig. 3). Four was shown by E. coli and S. aureus 14 and
different concentrations 10, 20, 40 and 100 12 mm respectively at 100 µL
µL concentrations were used against each concentrations. The Enterococcus, S.
tested pathogen Maximum zone of pyogenes and P. aeruginosa did not show
inhibition was exhibited by K. pneumoniae any result against M. jalapa.
20
Inhibitory zones
E.coli
15
P.aeruginosa
(mm)
10 K.pneumoniae
5 Enterococcus
S.aurues
0
10 20 40 100 S.pyogenes
Concentrations B.subtilis
(µL)
Figure 3. Zones of inhibition of M. jalapa against seven bacterial strains in µL
Antimicrobial activity of seven pathogens inhibition was exhibited by S. pyogenes 17
were checked against S. nigrum as shown in mm at 100 µL concentration. Moderate
(Fig. 4). E. coli and K. pneumoniae did level of activity was shown by P.
show any activity against plant. Four aeroginosa, S. aureus and B. subtilis 13, 11
different concentrations 10, 20, 40 and 100 and 12 mm respectively at 100 µL
µL concentrations were used against each concentration.
tested pathogen. The highest zone of
20
Inhibitory zones
E.coli
15
P.aeruginosa
(mm)
10 K.pneumoniae
5 Enterococcus
S.aurues
0
S.pyogenes
10 20 40 100
Concentrations B.subtilis
(µL)
Figure 4. Zones of inhibition of S. nigrum against seven bacterial strains
687Nasir et al.
Discussion vera gel showed inhibition on S. pyogenes,
As herbal combinations have made P. aeruginosa, E. coli and S. aureus. Ferro
significant pledges to human prosperity, the et al. [30], found A. vera gel can inhibit
herbs have typically been a source of hope gram positive bacteria S. pyogenes, which
for novel antibiotic components. Utilizing was not similar to our work.
plant extracts with well-known A. vera exhibited minimum inhibitory
antibacterial properties can be extremely activity against gram negative microbes
important for therapeutic purposes [19]. In due to the thick murein layer present in their
the last decade, Aloe vera has been utilized structure, which inhibits the entry of growth
broadly in health-care products like; inhibitors [31]. The MIC of A. vera gel for
ointments, cosmetics and health drinks, Gram-positive microbes, which has
with its antimicrobial properties [20]. additionally been accounted for already,
In this study, it was observed that aqueous might be associated with the variation in
extract of A. vera exhibited excellent cell wall structures of the microbes [30, 32].
antibacterial action against the selected In this study, the results depicted that
bacterial species: P. aeruginosa, K. aqueous abstract of S. nigrum showed
pneumoniae, E. coli, S. aureus and S. antimicrobial action on P. aeruginosa,
pyogenes. Results of the conducted Enterococcus, S. aureus, S. pyogenes and
research work were similar to that of B. subtilis, respectively. Results indicated
Kaithwas et al. [21] and also to the studies that gram positive microorganisms were
conducted by Mangena, 1999 in which it more sensitive to the aqueous extract of S.
was found out that A. vera gel was rich in nigrum than gram negative
various types of secondary metabolites like, microorganisms, which showed more
tannins, polysaccharides, anthraquinone, resistance. The reason would be that gram
glycosides, enzymes, sterols, vitamins, positive bacteria have peptidoglycan in
minerals and organic acids [22, 23]. their cell wall and no outer membrane,
The antimicrobial activity of A. vera which couldn’t block the passage of
extracts containing anthraquinone aloe hydrophobic molecules across the cell wall
emodin [24]. Selvamohan et al. [25], found [33]. Whereas, gram negative microbes
the aqueous extract of A.vera exhibited possess lippolysaccharides (LPS) layer in
moderate antibacterial activity against E. outer membrane presents as a strong barrier
coli, Klebisella sp., Pseudomonas sp. and due to high hydrophobicity [34].
Staphylococcus sp., which was similar to Chauhan, et al. [35], tested aqueous extract
our present work. of S.nigrum against five bacterial species
In the previous research, it was shown that named as: B. subtilis, P. aeruginosa,
A. vera exhibited inhibitory activity against Enterobacter aerogenes, S. aureus, and E.
S. aureus K. pneumoniae and E. coli, which coli which is partially similar to our present
was in accordance to our work [21, 26, 27]. research. In the present study, aqueous
The present study was in accordance with extract of S. nigrum prevented the
the results stated by Johnson et al., 2011, development of B. subtilis, P. aeruginosa
who found aqueous extract of A. vera had and S. aureus while exhibited no inhibitory
remarkable inhibitory activity against two activity against E. coli, respectively.
bacterial species i.e. E. coli and S. aureus Aqueous extract of S.nigrum possess
[28]. excellent hypoglycemic effect [36]. Abbas
Arun kumar, et al. [29], reported the et al. [37] determined water extract of S.
maximum inhibitory activity of A. vera nigrum to be the most active candidate,
extract against four bacterial species: E. possessing antimicrobial activity, which
coli, S. pyogenes, S. aureus and P. was in consistency to our present work. The
aeruginosa, which was similar to our reason would be that water is a polar
present study. The aqueous extract of A. compound and polar solvents possess
688Pure Appl. Biol., 12(1):683-693, March, 2023
http://dx.doi.org/10.19045/bspab.2023.120070
higher potential for antibacterial action than inhibitory concentration (MIC). The
those extracts with lesser polarity. Also, the observed variation in sensitivity testing
antibacterial activity always depends upon pattern of pathogenic microorganisms
the concentration of extracted antibacterial utilized in this work might be related to the
secondary metabolites, which can be genetic diversity among the organisms,
increased with the increase in polarity of which played a major role in resistance to
solvents [38]. the effects of abstracts [44].
Das et al. [39] tested the aqueous extract of Devi, et al. [45], tested methanolic extract
S. nigrum against different pathogenic and of M. jalapa, which showed potent
food-borne microorganisms. From which antimicrobial action against two gram
the extract showed excellent inhibitory positive microbes i.e. B. subtilis and S.
activity against B. subtilis, which was aureus and two gram negative
similar to the present study. S. nigrum microorganisms i.e. P. aeruginosa and E.
contains phytochemical constituents coli, respectively [45]. Though, in this
including, alkaloids, saponins, tannins, research, aqueous abstract of M. jalapa
flavinoids, proteins etc, which are exhibited excellent antimicrobial action
responsible for the antimicrobial activity against three gram positive bacteria i.e.
[40]. Enterococcus, B. subtilis and S. aureus) and
Singh et al. [41] determined the gram negative bacteria i.e. E. coli,
antimicrobial potential of aqueous and respectively. A variety of leaf extracts of M.
alcoholic extract of S. nigrum by Agar jalapa have been shown to be extensively
diffusion method against S. aureus, B. efficient against gram positive as well as
subtilis, E. coli, P. mirabilis, K. pneumonia negative microbes [46].
and P. aeruginosa and found out that E. Oladunmoye [47] and Kaladhar, et al. [48],
coli, P. mirabilis, K. pneumonia, B. subtilis determined that methanolic extract of M.
and S. aureus exhibited resistance to both jalapa showed MIC against S. aureus and
the extracts of S. nigrum, while, P. Aspergillus flavus, which was in
aeruginosa showed sensitivity to both the contradiction to the present work. The
extracts [41]. However, in this research, aqueous extract of M. jalapa was used to
results revealed the antibacterial potential check the MIC against different pathogenic
of aqueous extract of S. nigrum against S. and food-borne microorganisms,
aureus, P. aeruginosa, Enterococcus, B. respectively.
subtilis and S. pyogenes whereas, K. The antimicrobial activity exhibited by M.
pneumoniae and E. coli exhibited jalapa tends to concur with earlier
resistance, respectively. observations because of the presence of
Many studies were conducted which have terpenoids, essential oils, tannins,
been reported on antibacterial potential of flavonoids, and alkaloids [49]. Poovendran
M. jalapa. Akintobi et al. [42] studied the et al. [50] reported the aqueous extract of
antibacterial activity of M. jalapa using M. jalapa lack antimicrobial potential
four types of extracts against human against uro-pathogenic E. coli, which was
pathogenic microorganisms and reported dissimilar to the present research work. The
from the results that ethanol extract showed aqueous extract of M. jalapa exhibited
more inhibitory activity, followed by antibacterial action against E. coli with
methanolic extract than aqueous and zone of inhibition in the range of 14mm in
petroleum ether extracts, which was in diameter, respectively. The reason would
accordance to the findings of Obi and be that the antibacterial potential of herbs
Onuoha [43], whereas, in our work aqueous varies greatly, depending on the type and
extract exhibited inhibition on both gram kind of herbs, test medium and microbes.
positive as well as negative Oladunmoye [47] found that M. jalapa
microorganisms with the lowest minimum exhibited inhibitory potential against some
689Nasir et al.
human food-borne and pathogenic [55]. In the current study, the antibacterial
microorganisms. These antibacterial potential of three indigenous plants was
activities explain many advantages of herbs analyzed and this will help to develop drugs
in ethno-medicine from the indigenous sources.
Aiyelaagbe et al. [51] found out in their Authors’ contributions
study that gram positive bacteiria (GPB) Conceived and designed the experiments: R
used in the study were more sensitive to the Nisar & AA Abbasi, Performed the
herbal extracts as compared to the gram experiments: R Nisar, TA Mughal, S
negative bacteria (GNB), which was similar Khushal & R Shaukat, Analyzed the data:
to the prior studies that plant extracts RT Mahmood, K Hameed, MA Khan & M
exhibited excellent antimicrobial potential Masood, Contributed materials/ analysis/
against gram positive and negative tools: RT Mahmood & AA Abbasi, Wrote
microorganisms [52]. The outer membrane the paper: R Nisar, K Muhammad & M
of gram-negative microorganisms serves as Masood.
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