IN VITRO ANTIBACTERIAL AND ANTIOXIDANT PROPERTIES OF METHANOL LEAVES

EXTRACTS OF EMILIA ABYSSINICA (SCH. BIP. EX A. RICH) C. JEFFREY (COMPOSITAE) AND

CISSAMPELOS OWARIENSIS P. BEAUV. EX DC. (MENISPERMACEAE)

AROWONA, I.T.1*, SHITTU, I.A.1, FAMOJURO, O.B.2 AND FRED-JAIYESIMI, A.A.1

1
Department of Pharmacognosy, Olabisi Onabanjo University, Sagamu, Ogun State, Nigeria
2
Department of Pharmaceutical Microbiology, Olabisi Onabanjo University, Sagamu, Ogun State, Nigeria

ABSTRACT
Background: Emilia abyssinica and Cissampelos owariensis are medicinal plants used traditionally for the treatment
of wounds. It is important to identify potential natural remedies for diseases caused by pathogenic microorganisms
and oxidative stress. This study aimed to evaluate the antibacterial and antioxidant activities of Emilia abyssinica and
Cissampelos owariensis leaves.
Methodologies: The phytochemical analysis was carried out using standard protocols. Methanol crude extracts from
leaves of these plants were tested against various bacterial strains using the agar diffusion and agar dilution methods.
The antioxidant potential of the extracts was evaluated using DPPH free radical scavenging assay. Total Flavonoid
and Phenolic Compounds Content were quantified using Aluminium chloride test and Folin-Ciocalteu method
respectively. The assays were done in triplicate and data were expressed as mean ± SEM.
Results: The phytochemical analysis of these plants revealed the presence of saponins, flavonoids, tannins, alkaloids,
and sterols. The results indicated that both plants exhibited remarkable antibacterial and antioxidant activities. Emilia
abyssinica extract had lower MIC values for all the isolates as compared to Cissampelos owariensis extract. The DPPH
antioxidant activity of both plants showed comparable percentage inhibition with ascorbic acid the standard used (EA
– 78.2%; CO – 78.5%; AA – 80.3%). The Total Flavonoid Content for Emilia abyssinica and Cissampelos owariensis
methanol crude extracts were 204.3 µg/mL (Rutin equivalent) and 291.3 µg/mL (Rutin equivalent) while Total
Phenolic Compounds Content gave 37.6 µg/mL (GAE) and 38.8 µg/mL (GAE) respectively.
Conclusion: These findings suggest that Emilia abyssinica and Cissampelos owariensis leaves extracts have
promising antibacterial and antioxidant properties, and further studies are needed to identify and isolate the bioactive
compounds responsible for these activities.

Keywords: Antimicrobial, Antioxidants, Cissampelos owariensis, DPPH, Emilia abyssinica

*Correspondence: ismotarowona@yahoo.com, 08077573403

INTRODUCTION demonstrated potent activities and numerous researchers

have utilized natural products to combat bacterial
Bacteria are single-celled organisms that are found in resistance [4 - 6]. These compounds have exhibited
soil, water, and in the human body. They can either be promising results in overcoming the emergence of
beneficial or harmful, depending on the species. Some antibiotic resistance in bacterial pathogens [7]. They can
bacteria help to break down organic matter, while others restore the clinical application of older antibiotics by
cause infections and diseases in humans and animals [1]. increasing their potency and consequently, avoid
These pathogenic bacteria can enter the human body development of resistance [8].
through various routes, including ingestion, inhalation, Emilia abyssinica also called Senecio
or physical contact [2]. Once inside the body, they cause abyssinicus is a useful plant in tropical Africa,
a wide range of infections, from mild skin infections to commonly called “Amunimuye” by the Yoruba
life-threatening systemic infections [3]. Plant-derived speaking tribe in Nigeria [9 – 11]. The parts of the plant
chemicals are a wide group of chemical compounds that mostly used are leaves, stems, and flowers. The plant is
have been found naturally in plants. The extensive believed to possess potent medicinal properties in ethno-
existence of these compounds in antimicrobial agents, medicine [12]. It is an annual herb that grows in open
natural or synthetic substances that kill or inhibit the places to heights of about 50 cm and found in lowlands
growth of microorganisms like bacteria and fungi, have and mountain elevations in North and Southern Nigeria,

Nigerian Journal of Scientific Research, 23(1): 2024; journal.abu.edu.ng; https://njsrabuzaria.com.ng 1

Arowona et al. (2024); In vitro antibacterial and antioxidant properties of methanol leaves extracts

and West Cameroon. It is widely distributed in central reagent (Loba Chemie Pvt. Ltd), Sodium carbonate,
and east tropical Africa. Emilia abyssinica is used for Distilled water, Gallic acid (Molychem), Aluminium
various medicinal purposes by traditional healers in the chloride (Loba Chemie Pvt. Ltd), Potassium acetate
treatment of stomach-ache, dysentery, fever, and (Loba Chemie Pvt. Ltd.), Rutin (Loba Chemie Pvt. Ltd),
wounds [11]. The leaves of the plant are boiled in water Ascorbic acid, Gentamicin, and Mueller Hinton agar.
and the resulting infusion is taken orally as a remedy for
stomach-ache and dysentery. It is also used as a Collection of materials
purgative and for the treatment of rheumatism, scabies, The leaves of Emilia abyssinica and Cissampelos
and skin infections [13]. Senecio abyssinicus is a plant owariensis were collected in Forestry Research Institute
species that has been studied for its phytochemical of Nigeria (FRIN), Ibadan, Oyo State during the month
constituents. The plant contains a variety of secondary of April 2022. Contaminants were removed by hand
metabolites, including alkaloids, flavonoids, saponins picking from the bulk, pest-infested specimen were
and terpenoids [14]. removed, and the leaves were washed with water to
Cissampelos owariensis is commonly called remove sand and other debris. The plants were identified
velvet leaves, “ewe jenjoko or jokoje” among the and authenticated at Forest Herbarium Ibadan, Ibadan,
Yoruba people and “Damal gwaraajii” in Hausa, Nigeria, with FHI number 113719 and 113715
Nigeria. The plant is a perennial climbing plant with respectively.
annual stems that scramble over the ground or twine into
the surrounding vegetation for support. It is found Preparation and extraction of plant samples
mostly in African countries like Nigeria, Sierra Leone, Both plants were collected, cleaned, and air-dried for a
Congo, Tanzania and Angola [15]. Cissampelos period of one month. The dried leaves were further dried
owariensis is commonly used to treat gastrointestinal using an oven for 30 minutes at a temperature of 40°C
disorders such as diarrhea, dysentery, and stomach pain to remove all moisture contents. It was then pulverized.
[16]. The plant's leaves are boiled and the decoction is About 50 g of the pulverized sample of each plant was
consumed. It is used traditionally for various healing macerated in methanol in a ratio of 1:3 (pulverized
properties such as purgative, fertility, wound healing. In leaves: methanol) in a container. The mixture was
addition, the aerial parts of the plant have been used in macerated for 72 hours at room temperature and stirred
the management of mental disorder; juice from the leaf intermittently for optimal extraction of bioactive
is squeezed and given to patients experiencing psychosis compounds. After 72 hours, the macerated samples were
in Nigeria [17]. Phytochemicals found in Cissampelos filtered using filter paper to remove solid particles. The
owariensis leaves are flavonoids, saponins, tannins and filtrates were then concentrated under vacuo using a
moderate amounts of alkaloids [18], two sesquiterpenes, rotary evaporator set at 50°C. Further drying was done
which are bis (2- methoxy ethyl) phthalate and using a regulated water bath at 50oC to obtain dried
hexahydro 1,3-dimethyl (-4-phenyl-1H-azepine-4- crude extracts [21].
carboxylic acid have been isolated from this plant [19].
The phytochemical investigation of the root and leaf Phytochemical screening
extracts of C. owariensis yielded bisbenzylisoquinoline, Phytochemical screening of the pulverized leaves of
isochrodrondendrin, berberine and cycleanine alkaloids Emilia abyssinica and Cissampelos owariensis were
[20]. carried out using standard procedures [22, 23].
The fascinating reports about the trado-medical
properties of these plants, along with its wide range of Evaluation of antibacterial activity
reported biological activities, inspired us to delve into Test microorganisms
the leaves of both plants. Hence, the aim is to investigate Ten clinical isolates including three Staphylococcus
the antibacterial and antioxidant properties of the aureus, two Pseudomonas aeruginosa, two Klebsiella
methanol leaves extract of Emilia abyssinicus and pneumoniae and one each of Proteus mirabilis,
Cissampelos owariensis. Escherichia coli and Salmonella typhi were obtained
from Pharmaceutical Microbiology Laboratory, Olabisi
MATERIALS AND METHODS Onabanjo University Sagamu, Ogun State while the
typed strains of Pseudomonas aeruginosa ATCC 27853,
Chemicals Staphylococcus aureus ATCC 29213, Staphylococcus
The chemicals and reagents used include; Ammonia aureus NCTC 6571, Klebsiella pneumoniae ATCC
solution 10%, Chloroform, Concentrated sulphuric acid, 700303, Escherichia coli ATCC 25922, Escherichia
Dragendorff’s reagent, Ferric chloride, Glacial acetic coli ATCC 700728 and Salmonella typhi ATCC 14028
acid, Hydrochloric acid, Lead acetate, Mayer’s reagent, were obtained from the Pharmaceutical Microbiology
Methanol, Methylene chloride, Olive oil, Sodium Laboratory, Faculty of Pharmacy, University of Ibadan.
hydroxide, Wagner’s reagent, Ethanol, Folin-Ciocalteu

Nigerian Journal of Scientific Research, 23(1): 2024; journal.abu.edu.ng; https://njsrabuzaria.com.ng 2

Arowona et al. (2024); In vitro antibacterial and antioxidant properties of methanol leaves extracts

Extract standardization Cissampelos owariensis leaf extracts obtained from

The extracts were reconstituted with 50% methanol. stock extract solution (1 mg/mL) were added to 3.9 mL
Serial dilutions were carried out to obtain four different of DPPH solution and the reactants incubated at 25°C
concentrations of 100 mg/mL, 50 mg/mL, 25 mg/mL, room temperature for 30 minutes in a dark environment.
and 12.5 mg/mL. In place of an extract, the ascorbic acid solution (a
positive control) was used as standard. The mixture was
Inocula standardization shaken, and the solution was allowed to stand in the dark
Mueller-Hinton Agar (MHA) served as a medium for at room temperature for 30 minutes. Free radical
sub-culturing the isolates on sterile plates. The plates scavenging activity was calculated from the absorbance
were incubated at 37ºC for 24 hours. The broth cultures values at 517 nm. Blank (methanol) was also measured
of the isolates were adjusted to the exact turbidity and recorded [27].
equivalent to 0.5 McFarland Standard as recommended
by Clinical and Laboratory Standard Institute (CLSI). % Inhibition = Abs control – Abs sample x 100
Abs control
Antibacterial susceptibility testing
The antibacterial activity of the extracts was determined Total phenol content estimation
using agar well diffusion assay method [24, 25]. Mueller The Total Phenol Content (TPC) of Emilia abyssinica
Hinton Agar (MHA) was prepared as recommended by and Cissampelos owariensi methanol leaves extracts
the manufacturer and allowed to cool to 50ºC before were determined by spectrophotometric method using
pouring into pre-labeled sterile petri plates. Overnight the Folin-Ciocalteu method, gallic acid was set as
culture of the test organism adjusted to 0.5 McFarland standard curve. 0.25 mL of each extract was mixed with
turbidity standard was used to inoculate the surface of 0.5 mL Folin-Ciocalteu phenol reagent. After 5 min, 5
solidified Mueller Hinton agar plate using a sterile swab mL of 7% Na2CO3 solution was added to the mixture,
stick. A sterile cork borer (9 mm) was used to punch six followed by the addition of 65 mL distilled water. The
wells at equidistant wells on the Mueller Hinton Agar mixture was thoroughly mixed, allowed 90 min
plates. Wells 1 – 4 contained four different incubation at 25°C and the absorbance was read at 765
concentrations of each extract (12.5 mg/mL – 100 nm. The TPC value of the leaves crude extract (1
mg/mL). Wells nine and ten were filled with 50% mg/mL) was calculated from the gallic acid standard
methanol and 20 µg/mL gentamicin, respectively. The curve expressed as GAE mL/g of extract. Blank was
plates were left on the bench for 45 minutes allowing for distilled water and the values were in triplicates [28].
proper pre-diffusion of the extracts into the agar and The various concentrations of gallic acid used for the
incubated at 37ºC for 24 hours. Subsequently, standard curve were 500 µg/mL, 250 µg/mL, 125
antibacterial activity was determined by measuring the µg/mL, 62.5 µg/mL, 31.25 µg/mL, 15.6 µg/mL, 7.8
diameter of zones of inhibition in millimeters. µg/mL and 3.91 µg/mL.
Determination of minimum inhibitory concentration
(MIC) Total flavonoid content estimation
Using the agar dilution method, different concentrations Total Flavonoid Content (TFC) was determined using
of plant extracts were prepared by adding 2 mL of stock Aluminium chloride method [29]. One mL of sample (1
concentration of each plant extract to 18 mL of molten mg/mL) was mixed with 3 mL of methanol, 0.2 mL of
Mueller Hinton agar, cooled down to 45°C to obtain the 10% aluminium chloride, 0.2 mL of 1 M potassium
desired final concentrations of 50 mg/mL, 25 mg/mL, acetate and 5.6 mL of distilled water and kept at room
12.5 mg/mL, 6.25 mg/mL, 3.125 mg/mL, 1.563 mg/mL temperature for 30 min. The absorbance of the reaction
and 0.781 mg/mL. Bacterial suspensions adjusted to 105 mixture was measured at 420 nm using UV-Visible
CFU/mL were used to inoculate the MHA plates spectrophotometer. The total flavonoid content was
containing different concentrations of the plant extracts. determined from standard rutin calibration curve (0-100
The plates were incubated at 37°C for 24 hours. The µg/mL in methanol). The concentration of total
MIC was determined as the lowest concentration of flavonoid was expressed as mg of rutin equivalent/g of
plant extract in solid media where no growth was plant extract. The concentrations of rutin used for the
observed after 24 or 48 h [26]. standard curve were; 500 µg/mL, 250 µg/mL, 125
µg/mL, 62.5 µg/mL, 31.25 µg/mL, 15.6 µg/mL and 7.8
Antioxidant activity evaluation µg/mL.
DPPH radical scavenging assay
The DPPH radical solution was prepared by dissolving
2.5 mg of DPPH in 100 mL of methanol (0.025 g/L). RESULTS AND DISCUSSION
Different concentrations of Emilia abyssinica and

Nigerian Journal of Scientific Research, 23(1): 2024; journal.abu.edu.ng; https://njsrabuzaria.com.ng 3

Arowona et al. (2024); In vitro antibacterial and antioxidant properties of methanol leaves extracts

Phytochemical screening Antibacterial activity of Emilia abyssinica and

The qualitative phytochemical screening for the Cissampelos owariensis leaves extracts
presence of different classes of metabolites in Emilia Antimicrobial Sensitivity Testing involves testing the
abyssinica and Cissampelos owariensis leaves indicated sensitivity of microorganisms to various concentrations
presence of saponins, tannins, flavonoids, alkaloids, of plant extracts. The microorganisms were cultured in
steroids, and cardiac glycosides, but absence of the presence of different concentrations of the extract,
cyanogenic glycosides, anthraquinones and amino acids and the results were evaluated based on the degree of
in pulverized leaves of both plants (Table 1). Egharevba, growth inhibition [32]. Furthermore, Minimum
et al. [30], reported that the leaves extract of Senecio Inhibitory Concentration (MIC) testing is a quantitative
abyssinicus showed presence of polyphenols, steroids, test that determines the minimum concentration of plant
terpenoids, alkaloids, and cardiac glycosides. In extract required to inhibit completely the growth of
addition, C. owariensis root methanol extract was found microorganisms using agar dilution method [33]. It is a
to contain flavonoids, alkaloids, resins, tannins, valuable tool for identifying the most effective
balsams, steroids, saponins, terpenes, cardiac concentration of plant extract against a specific
glycosides, carbohydrate and phenols [31]. These microorganism.
previous studies supported the result of our study. Emilia abyssinica methanol extract exerts
antibacterial activity against some clinical bacterial
Table 1: Preliminary phytochemical screening of isolates at used concentrations (Table 2) with zones of
Emilia abyssinica and Cissampelos owariensis inhibition ranging from 10 to 20 mm while the extract
pulverized leaves did not show any activity on the following clinical
TEST E. C. isolates: K. pneumoniae 112, P. mirabilis, S. aureus 02,
abyssinica owariensis E. coli 71, P. aeruginosa 31 and P. aeruginosa 81. All
Anthraquinones - - the clinical isolates were not susceptible to the extract at
Flavonoids + + 12.5 and 25 mg/mL. Higher antibacterial activity was
Cyanogenetic - - observed against the typed strains than the clinical
Saponins + + isolates. Only K. pneumoniae ATCC 700303 and E. coli
Sterols + + ATCC 700728 had zones of inhibition at lower
Amino acids - - concentration of 12.5 mg/mL (Table 2). MIC values
Alkaloids + + ranged between 0.78 – 1.56 mg/mL and 1.56 – 3.13
Tannins + + mg/mL among the clinical isolates and typed strains,
Key: + Present, - Absent respectively (Table 3).

Table 2: Antibacterial activity of Emilia abyssinica against clinical isolates and typed cultures (Zones of Inhibition
in millimeters)
ORGANISMS 100 50 25 12.5 50% methanol Gentamycin
mg/mL mg/mL mg/mL mg/mL + sterile water
Klebsiella pneumoniae (112) 0 0 0 0 0 12mm
Staphylococcus aureus (8571) 11mm 0 0 0 0 24mm
Salmonella typhi (111) 12mm 11mm 0 0 0 16mm
Proteus mirabilis 0 0 0 0 0 20mm
Staphylococcus aureus (02) 0 0 0 0 0 15mm
Escherichia coli ATCC 25922 0 0 0 0 0 26mm
Staphylococcus aureus ATCC 11mm 0 0 0 0 16mm
29213
Klebsiella pneumoniae ATCC 11mm 11mm 10mm 10mm 0 17mm
700303
Pseudomonas aeruginosa ATCC 12mm 12mm 13mm 0 0 22mm
27853
Escherichia coli ATCC 700728 16mm 14mm 13mm 11mm 0 20mm
Salmonella typhi ATCC 14028 17mm 0 0 0 0 25mm
Escherichia coli (71) 0 0 0 0 0 0
Klebsiella pneumoniae (287) 10mm 10mm 0 0 0 22mm
Staphylococcus aureus (13) 20mm 15mm 0 0 0 23mm

Nigerian Journal of Scientific Research, 23(1): 2024; journal.abu.edu.ng; https://njsrabuzaria.com.ng 4

Arowona et al. (2024); In vitro antibacterial and antioxidant properties of methanol leaves extracts

Pseudomonas aeruginosa (31) 0 0 0 0 0 22mm

Pseudomonas aeruginosa (81) 0 0 0 0 0 0

Table 3: Total Minimal Inhibitory Concentration (MIC) of Emilia abyssinica leaves extract against different isolates
Test Organism MIC (mg/mL)
Staphylococcus aureus (8571) 1.56
Salmonella typhi (111) 1.56
Klebsiella pneumoniae (287) 0.78
Staphylococcus aureus (13) 3.12
Klebsiella pneumoniae ATCC 700303 1.56
Pseudomonas aeruginosa ATCC 27853 1.56
Escherichia coli ATCC 700728 3.12
Salmonella typhi ATCC 14028 3.12

The crude methanol extract of Cissampelos owariensis diffuse into the agar medium, as opposed to the low
were tested against different strains of microorganisms minimum inhibitory concentrations seen using the agar
using agar dilution method 34]. The degree of growth dilution method. According to Eloff [35], Agar diffusion
inhibition was determined by measuring the diameter of experiments use an aqueous agar substrate, and many
the inhibition zone around the microbial growth. From antimicrobial chemicals found in plant extracts are
the AST result in Table 4, the highest degree of activity relatively non-polar, meaning that they do not diffuse
was observed against E. coli ATCC 700728 (ZOI range well in this matrix.
of 12 – 14 mm) and S. typhi ATCC 14028 (ZOI of 13 Several researchers have documented the
mm). Moderate degree of activity was observed against antimicrobial activity of Emilia species. Antibacterial
S. aureus (13) and (02) (ZOI of 11 mm and 10 – 12 mm activity of aqueous, ethanol and methanol extracts of
respectively), S. aureus (8571) (ZOI of 10 – 11 mm), K. Emilia sonchifolia was reported [36, 37]. Teke et al. [38]
pneumonia (112), (287) and ATCC 700303 (ZOI of 10 also reported the antidiarrheal effects of both methanol
mm, 10 mm and 10 – 12 mm, respectively), and P. and aqueous extracts of the leaves of Emilia coccinea.
aeruginosa (81) (ZOI of 11 mm). Strains of bacteria that To the best of our knowledge, this is the first report on
showed no degree of activity include S. aureus ATCC the antibacterial activity of Emilia abyssinica leaves
29213, E. coli (71), ATCC 25922, P. aeruginosa (31), extract.
ATCC 27833, and S. typhi (111). The test bacterial that The antimicrobial activity of Cissampelos
were susceptible were further tested for the Minimum species has also been reported [39, 40]. Ngoci et al. [39],
Inhibitory Concentration (MIC) to determine the lowest found that Cissampelos pareira exhibits antibacterial
concentration of the extract. Staphylococcus aureus efficacy against both Gram-positive and Gram-negative
(8571), Salmonella typhi (111), Staphylococcus aureus bacteria. Crude methanol root extract of Cissampelos
(02), Klebsiella pneumoniae (287) gave an MIC value owariensis was reported to be active against
of 6.25 mg/mL while Staphylococcus aureus (13), Mycobacterium tuberculosis, Staphylococcus aureus,
Pseudomonas aeruginosa (31) and Pseudomonas Salmonella typhi, Escherichia coli and Candida
aeruginosa (81) gave an MIC value of 12.5 mg/mL albicans [41]. This study also reported lower MIC
(Table 5). values against the typed bacterial strain and clinical
The absence of growth inhibition zones using bacterial isolates including Staphylococcus aureus,
the agar well diffusion method may be caused by the Salmonella typhi, Klebsiella pneumoniae and
active component in the plant extract not being able to Pseudomonas aeruginosa

Nigerian Journal of Scientific Research, 23(1): 2024; journal.abu.edu.ng; https://njsrabuzaria.com.ng 5

Arowona et al. (2024); In vitro antibacterial and antioxidant properties of methanol leaves extracts

Table 4: Antibacterial activity of Cissampelos owariensis against clinical isolates and typed cultures (zones of
inhibition in millimeters)
Test organism 100 50 25 12.5 50% methanol Gentamycin
mg/mL mg/mL mg/mL mg/mL + sterile water
Klebsiella pneumoniae (112) 10mm 10mm 0 0 0 12mm
Staphylococcus aureus (8571) 11mm 10mm 10mm 0 0 23mm
Salmonella typhi (111) 0 0 0 0 0 25mm
Proteus mirabilis 0 0 0 0 0 0
Staphylococcus aureus (02) 12mm 11mm 10mm 11mm 0 13mm
Escherichia coli ATCC 25922 0 0 0 0 0 22mm
Staphylococcus aureus ATCC 0 0 0 0 0 23mm
29213
Klebsiella pneumoniae ATCC 12mm 11mm 0 0 0 16mm
700303
Pseudomonas aeruginosa ATCC 0 0 0 0 0 22mm
27853
Escherichia coli ATCC 700728 14mm 12mm 0 0 0 21mm
Salmonella typhi ATCC 14028 13mm 13mm 0 0 0 18mm
Escherichia coli (71) 0 0 0 0 0 17mm
Klebsiella pneumoniae (287) 10mm 0 0 0 0 25mm
Staphylococcus aureus (13) 11mm 0 0 0 0 20mm
Pseudomonas aeruginosa (31) 0 0 0 0 0 22mm
Pseudomonas aeruginosa (81) 11mm 0 0 0 0 0

Table 5: Minimum Inhibitory Concentration (MIC) of Cissampelos owariensis methanol leaves extract against
different isolates
Test organisms MIC (mg/mL)
Staphylococcus aureus (8571) 6.25
Salmonella typhi (111) 6.25
Staphylococcus aureus (02) 6.25
Klebsiella pneumoniae (287) 6.25
Staphylococcus aureus (13) 12.5
Pseudomonas aeruginosa (31) 12.5
Pseudomonas aeruginosa (81) 12.5
Klebsiella pneumoniae ATCC 700303 6.25
Pseudomonas aeruginosa ATCC 27853 6.25
Escherichia coli ATCC 700728 6.25
Salmonella typhi (14028) 6.25

abyssinica and Cissampelos owariensis possess highest

Antioxidant assay percentage inhibition of 78.17% and 78.51% at
DPPH radical is scavenged by antioxidants through the concentration of 1.0 mg/mL when compared to ascorbic
donation of proton forming the reduced DPPH. The acid standard with percentage inhibition of 80.28% at
colour changes from purple to yellow after reduction, the same concentrations (Figure 1). This implies that
which can be quantified by its decrease of absorbance at both plants contain high amount of antioxidant
wavelength 517 nm. Radical scavenging activity principles that mop up free radicals in the body when
increased with increasing percentage of the free radical used. Preceding studies confirmed the antioxidant
inhibition. The degree of discoloration indicates the free property of the flower extract and fractions of Senecio
radical scavenging potentials of the sample/antioxidant abyssinicus leaves [14, 30]. Several species of Senecio
by their hydrogen donating ability. The electrons have been reported to possess strong antioxidant
become paired off and solution loses color property. Senecio salsuginea was reported to possess an
stoichiometrically depending on the number of electrons inhibitory concentration (IC50) of 26.23 μg/mL [43], and
taken up [42]. The result of DPPH showed that Emilia ethyl acetate fraction of Senecio stabianus showed an

Nigerian Journal of Scientific Research, 23(1): 2024; journal.abu.edu.ng; https://njsrabuzaria.com.ng 6

Arowona et al. (2024); In vitro antibacterial and antioxidant properties of methanol leaves extracts

IC50 value of 35.5 mg/mL [44]. Furthermore, Omotoso of E. abyssinica and Cissampelos owariensis (1 mg/mL)
et al. [45] reported that methanol extract of C. were 204.27 µg/mL and 291.28 µg/mL, respectively
owariensis possesses abundant phenolic compounds, with reference to rutin standard curve. Comparatively,
which explains its antioxidant property. we discovered that C. owariensis have higher flavonoid
content than E. abyssinica. The total flavonoid content
of the methanolic extract of Biophytum sensitivum
82% whole plant methanol extract was found to be 9.49 Âμg
% Inhibition

80% with respect to quercetin standard curve [55]. The

78%
antioxidant activity, total phenolic and flavonoid
contents of whole plant extracts of Torilis leptophylla
76% have also been reported by Saeed et al. [49]. The
0.2mg/ml
0.6mg/ml
methanol leaves extract of Costus afar was reported to
1.0mg/ml contain 3.13% of flavonoid and 0.77% of phenol
respectively [56]. These previous reports support our
Standard Abscorbic Acid results and confirm the presence of different quantity of
S. abyssinicus flavonoids and phenolic compounds in medicinal plants.
C. owariensis

Table 6: Concentration of phenolic compounds in

Figure 1: DPPH scavenging activity of methanol leaf methanol leaf extracts of Emilia abyssinica and
extracts of E. abyssinicus and C. owariensis with respect Cissampelos owariensis
to standard Ascorbic acid Plant samples Concentration
(µg/mL)
Phenolic compounds in plants are known to act as free
radical scavengers and it has been opined that the Emilia abyssinicus 37.61
antioxidant activity of most of the plant produce is
mainly due to the presence of phenolic compounds [46, Cissampelos owariensis 38.78
47]. Basically, antioxidant mechanism of polyphenolic
compounds is based on their hydrogen donating and
metal ion chelating abilities [48]. The total phenol Table 7: Concentration of flavonoid contents in
content is used to analyze tannins, anthocyanins, methanol leaf extracts of Emilia abyssinica and
monomeric phenolic compounds and polymeric Cissampelos owariensis
pigments present in a sample. A correlation between the Plant sample Concentration
phenolic content and antioxidant activity has been (µg/mL)
shown earlier by number of researchers [49, 50]. The
amount of total phenol was investigated by Folin- Emilia abyssinica 204.27
Ciocalteu method [50]. The total phenol content of
Emilia abyssinica and Cissampelos owariensis Cissampelos owariensis 291.28
methanol leaf extracts (1 mg/mL) were 37.61 µg/mL and
38.78 µg/mL of gallic acid equivalent per gram of plant CONCLUSION
extract with reference to gallic acid standard curve.
Flavonoids are a group of naturally occurring These findings suggest that Emilia abyssinica and
compounds that have been shown to possess antioxidant Cissampelos owariensis have promising antibacterial
activity, which makes them useful in various and antioxidant properties. The activities possessed by
applications, including pharmaceuticals, food additives, these plants are due to the phytoconstituents present.
and cosmetics [51]. The TFC assay involves the Further studies are warranted to identify and isolate
quantification of total flavonoids present in a plant bioactive compounds responsible for these activities.
sample. One of the most widely used methods for
determining TFC is an aluminum chloride colorimetric ACKNOWLEGDEMENTS
assay. The potential antioxidant activity of flavonoid is
associated with the chemical structures o-diphenolic The authors are grateful to Mr Adedokun and Mr Adeoti
group, a 2–3 double bond conjugated with the 4-oxo of the Pharmacognosy Department, Olabisi Onabanjo
function and hydroxyl groups in positions 3 and 5 [52, University, for their technical assistance during the
53]. Various concentrations of standard rutin solution bench work.
were used to prepare a standard calibration curve [54].
The total flavonoid content of the methanol leaf extract

Nigerian Journal of Scientific Research, 23(1): 2024; journal.abu.edu.ng; https://njsrabuzaria.com.ng 7

Arowona et al. (2024); In vitro antibacterial and antioxidant properties of methanol leaves extracts

12. ODUBANJO, V.O., OBOH, G., OYELEYE, S.I. &

ADEFEGHA, S.A. (2018). Anticholinesterase
REFERENCES activity and phenolic profile of two medicinal
plants (Quassia undulata and Senecio
1. GIELECIŃSKA, A., KCIUK, M., MUJWAR, S., abyssinicus) used in managing cognitive
CELIK, I., KOŁAT, D., KAŁUZIŃSKA- dysfunction in Nigeria. Journal of Food
KOŁAT, Ż. & KONTEK, R. (2023). Biochemistry, 42(4): e12497.
Substances of Natural Origin in Medicine: 13. SOFOWORA, A. Plants in African traditional
Plants vs. Cancer. Cells, 12(7): 986. medicine-an overview. In: EVANS, W.C. (ed)
2. RYAN, K.J. & RAY, C.G. (Eds.). (2004). Sherris Trease and Evans pharmacognosy, Edn 15.
Medical Microbiology, 4th edition. McGraw W.B. Saunders, Edinburgh, 2000, 488-496 pp.
Hill. 14. EGHAREVBA, G.O., DOSUMU, O.O.,
3. TODAR, K. (2012). Pathogenic Bacteria. In Todar's OGUNTOYE, S.O., NJINGA, N.S.,
Online Textbook of Bacteriology. ABDULMUMEEN. H.A. & ADEBAYO,
4. KHAMENEH, B., DIAB, R., GHAZVINI, K. & M.A.B. (2018). Phytochemical screening,
FAZLY BAZZAZ, B.S. (2016). Breakthroughs Antimicrobial and Antioxidant Activities of
in bacterial resistance mechanisms and the Crude Extracts of Senecio abyssinicus Flower.
potential ways to combat them. Microbial Journal of Pharmaceutical Research and
Pathogenesis, 95: 32–42. Development Practice, 2(1): 16-24.
5. FAZLY BAZZAZ, B.S., SARABANDI, S., 15. TROPICAL PLANTS DATABASE, Ken Fern.
KHAMENEH, B. & HOSSEINZADEH, H. tropical.theferns.info. (2021, August 06).
(2016). Effect of catechins, green tea extract 16. OKOLI, R.I., AIGBE, O., OHAJU-OBODO, J.O. &
and methylxanthines in combination with MENSAH, J.K. (2007). Medicinal plants used
gentamicin against Staphylococcus aureus and for managing some common ailments among
Pseudomonas aeruginosa: Combination Esan people of Edo State, Nigeria. Pakistan
therapy against resistant bacteria. Journal of Journal of Nutrition,
Pharmacopuncture, 19(4): 312-318. 17. SONIBARE, M.A., SOLADOYE, M.O. &
6. SHAKERI, A., SHARIFI, M.J., FAZLY BAZZAZ, SUBULOYE, T.O. (2008). Ethnobotanical
B.S., EMAMI, A., SOHEILI, V. & survey of anti-psychotic plants in Lagos and
SAHEBKAR, A. (2018). Bioautography Ogun States of Nigeria. European Journal of
detection of antimicrobial compounds from the Science Research. 19: 634–43.
essential oil of Salvia pachystachys. Current https://www.researchgate.net/publication/2885
Bioactive Compounds, 14(1): 80–5. 68972
7. ROSSITER, S.E., FLETCHER, M.H. & WUEST, 18. EKEANYANWU, R.C., UDEME, A.A.,
W.M. (2017) Natural products as platforms to ONUIGBO, A.O. & ETIENAJIRHEVWE,
overcome antibiotic resistance. Chemical O.F. (2012). Anti-diabetic Effect of Ethanol
Review, 117(19): 12415–74. Leaf Extract of Cissampelos owariensis
8. BARBIERI, R., COPPO, E., MARCHESE, A., (lungwort) on Alloxan induced Diabetic Rats.
DAGLIA, M., SOBARZO-SANCHEZ, E., African Journal of Biotechnology, 11(25):
NABAVI, S.F. ET AL. (2017) Phytochemicals 6758-6762. DOI: 10.5897/AJB12.512
for human disease: An update on plant-derived 19. EFIOM, O.O. (2010). Isolation and Characterization
compounds antibacterial activity. of Bis (2 – Methoxyethyl) Phthalate and
Microbiology Research, 196: 44–68. Hexashydro-1 3–Dimethyl–4–Phenyl–1h–
9. ADEKUNLE, M.F., OLUWALANA, S.A., Azepine4–Carboxylic Acid from the Root of
AKLILU, N. & MENGISTEAB, H. (2002). Cissampelos owariensis (P. Beauv). Nigerian
Exploratory Survey of Forest Plants in Journal of Basic and Applied Science, 18(2):
Traditional Treatment of Guinea Worm 189 - 192.
Infections (GWI) (Dracunculus medinensis 20. SOUTHERN, I.W. & BUCKINGHAM, J. (1989).
Linn): Experiences from Nigeria and Ethiopia. Dictionary of alkaloid. Chapman Hall, New
African Research Review, 1(3): 108-124. York. 628 pp.
10. ODUGBEMI, T. (2008). A textbook of medicinal 21. AROWONA, I.T., SONIBARE, M.A., ALIYU,
plants from Nigeria. University of Lagos Press, A.B. & IQBAL, J. (2023). Inhibition of D –
Lagos, Nigeria, 20-323 pp. Amino Acid Oxidase by Chromatographic
11. GBILE, Z.O. (1984). Vernacular names of Nigerian Fractions and Kaempferol-3-O-rutinoside
plants (Yoruba). 1st edn. Forestry Research Isolated from Philenoptera cyanescens Leaves.
Institute of Nigeria, Ibadan, Nigeria, 101 pp. Tropical Journal of Natural Product Research.

Nigerian Journal of Scientific Research, 23(1): 2024; journal.abu.edu.ng; https://njsrabuzaria.com.ng 8

Arowona et al. (2024); In vitro antibacterial and antioxidant properties of methanol leaves extracts

7(1): 2251 - 2257. Chemistry, 7(6): 225-230.

22. EVANS, W.C. (2009). Trease and Evans 32. GOLUS, J., SAWICKI, R., WIDELSKI, J. &
Pharmacognosy. Elsevier Health Sciences. GINALSKA, G. (2016). The agar
23. JUNAID, R.S. & PATIL, M.K. (2020) Qualitative microdilution method–a new method for
tests for preliminary phytochemical screening: antimicrobial susceptibility testing for essential
An overview. International Journal of oils and plant extracts. Journal of Applied
Chemical Studies, 8(2): 603-608 Microbiology, 121(5): 1291-1299.
24. THITILERTDECHA, N., TEERAWUTGULRAG, 33. SILVA, A.C.O., SANTANA, E.F., SARAIVA,
A. & RAKARIYATHAM, N. (2008). A.M., COUTINHO, F.N., CASTRO, R.H.A.,
Antioxidant and antibacterial activities of PISCIOTTANO, M.N.C. &
Nephelium lappaceum L. extracts. LWT- Food ALBUQUERQUE, U.P. (2013). Which
Science and Technology, 41(10): 2029-2035. approach is more effective in the selection of
25. AMEYA, G., GURE, A., & DESSALEGN, E. plants with antimicrobial activity? Evidence-
(2016). Antimicrobial activity of Echinops Based Complementary and Alternative
kebericho against human pathogenic bacteria Medicine, 2013
and fungi. African Journal of Traditional, 34. BARON, E.J. & FINEGOLD, S.M. (1994). Methods
Complementary and Alternative Medicines, of testing antimicrobial effectiveness (In:
13(6): 199-203. Bailey and Scott’s Diagnostic
26. WECKESSER, S., ENGEL, K., SIMON- Microbiology). New York: Mosby.
HAARHAUS, B., WITTMER, A., PELZ, K., 35. ELOFF, J.N. (2019). Avoiding pitfalls in
& SCHEMPP, C.M. (2007). Screening of plant determining antimicrobial activity of plant
extracts for antimicrobial activity against extracts and publishing the results. BMC
bacteria and yeasts with dermatological Complementary and Alternative Medicines,
relevance. Phytomedicine, 14(6): 508–516 19(1): 106. doi: 10.1186/s12906-019-2519-3.
27. EBRAHIMZADEH, M.A., NABAVI, S.M., PMID: 31113428; PMCID: PMC6530048.
NABAVI, S.F., BAHRAMIAN, F. & 36. YOGA LATHA, L. JR., DARAH, I.,
BEKHRADNIA, A.R. (2010). Antioxidant and SASIDHARAN, S. & JAIN, K. (2009).
free radical scavenging activity of H. officinalis Antimicrobial Activity of Emilia sonchifolia
L. var. angustifolius, V. odorata, B. hyrcana DC., Tridax procumbens L. and Vernonia
and C. speciosum, Pakistan Journal of cinerea L. of Asteracea Family: Potential as
Pharmaceutical Sciences 23(1): 29-34. Food Preservatives. Malaysian Journal of
28. KIM, J.H., KIM, S.J., PARK, H.R., CHOI, J.I., JU Nutrition, 15(2): 223 - 31. PMID: 22691820.
Y.C., NAM, K.C., KIM, S.T. & LEE, S.C. 37. UGWOKE, G.I., IKORO, N.C. & ESENWAH, E.C.
(2009). The different antioxidant and (2021). Comparative analysis of the
anticancer activities depending on the colour of antibacterial effects of Emilia sonchifolia
oyster mushroom. Journal of Medicinal Plants (tassel flower) and selected antibiotics on
Research, 3(11): 1016-1020. ocular bacteria, in vitro. International Journal
29. SHEN, Y., JIN, L., XIAO, P., LU, Y. & BAO, J. of Health Science Research, 11(3): 61-68.
(2009). Total phenolics, flavonoids, 38. TEKE, G.N., KUIATE, J.R., NGOUATEU, O.B. &
antioxidant capacity in rice grain and their GATSING, D. (2007). Antidiarrhoeal and
relations to grain color, size and weight. antimicrobial activities of Emilia coccinea
Journal of Cereal Science, 49: 106-111. (Sims) G. Don extracts. Journal of
30. EGHAREVBA, G.O., DOSUMU, O.O, Ethnopharmacology, 12(2): 278-83. doi:
EVBUOMWAN, I.O., NJINGA, N.S., 10.1016/j.jep.2007.03.007. Epub 2007 Mar 12.
OLUYORI, A.P., HAMID, A.A. & AFOLABI, PMID: 17433589.
Y.T. (2023). In vitro Antimicrobial, 39. NGOCI, S.N., RAMADHAN, M., NGARI, S.M., &
Antioxidant, and Antidiabetic Activities of LEONARD, P.O. (2013). Screening for
Extracts of Senecio abyssinicus Leaves. Antimicrobial Activity of Cissampelos pareira
Journal of Applied Science Enviironmental L. Methanol Root Extract. European Journal
Management, 27(5): 939-948 of Medicinal Plants, 4(1): 45–51.
31. OLUTAYO, O., ABAYOMI, O., OLAKUNLE, F., 40. AMIT, M.D. & SHARMA, K.R. (2020). In vitro
TEMITOPE, A. & IBRAHIM, I. (2013). The biological study of seven Nepalese medicinal
chemical composition of essential oil from the plants and isolation of Chemical constituents
root of Cissampelos owariensis (P. Beauv.) and from Cissampelos pareira. Asian of Journal
free radical scavenging activities of its extracts. Pharmacy and Clinical Research, 13(9): 1-7
African Journal of Pure and Applied 41. AKANDE, R., OKWUTE, S.K., ILIYA, I. &

Nigerian Journal of Scientific Research, 23(1): 2024; journal.abu.edu.ng; https://njsrabuzaria.com.ng 9

Arowona et al. (2024); In vitro antibacterial and antioxidant properties of methanol leaves extracts

EFIOM, O.O. (2013). Chemical constituents 49. SAEED, N., KHAN, M.R. & SHABBIR, M. (2012).
and anti-tuberculosis activity of the root Antioxidant activity, total phenolic and total
extracts of Cissampelos owariensis (P. Beauv.) flavonoid contents of whole plant extracts
Menispermaceae. Africa Journal of Pure Torilis leptophylla L. BMC complementary
Applied Chemistry, 7(1): 21-30 and Alternative Medicine, 12: 1-12.
42. SANNIGRAHI, S., MAZUMDER, U.K. & PAL, 50. SINGLETON, V.L. & ROSSI, J.A. (1965).
D.M. (2009). Hepatoprotective potential of Colorimetry of total phenolics with
methanol extract of Clerodendrum phosphomolybdic-phospho-tungstic acid
infortunatum Linn. against CCl4 induced reagents. American Journal of Enology and
hepatotoxicity in rats. Indian Journal of Viticulture, 16(3): 144-158.
Experimental Biology, 5(20): 394-399. 51. ORDOÑEZ, A.A., GOMEZ, J.D., VATTUONE,
43. ALBAYRAK, S., AKSOY, A., YURTSEVEN, L. & M.A. & ISLA, M.I. (2006). Antioxidant
YAŞAR, A. (2015). A comparative study on activities of Sechium edule (Jacq.) Swartz
antioxidant and antibacterial activities of four extracts. Food Chemistry, 97(3): 452-458.
Senecio Species from Turkey. International 52. LEE, L.S., CHOI, E.J., KIM, C.H., SUNG, J.M.,
Journal of Secondary Metabolite, 2(2): 26-36. KIM, Y.B., SEO, D.H. & PARK, J.D. (2016).
44. TUNDIS, R., MENICHINI, F., LOIZZO, M.R., Contribution of flavonoids to the antioxidant
BONESI, M., SOLIMENE, U. & properties of common and tartary
MENICHINI, F. (2012). Studies on the buckwheat. Journal of Cereal Science, 68:
potential antioxidant properties of Senecio 181-186.
stabianus Lacaita (Asteraceae) and its 53. BRAVO, L. & MATEOS, R. (2008). Analysis of
inhibitory activity against carbohydrate- flavonoids in functional foods and
hydrolysing enzymes. Natural Product nutraceuticals. Methods of Analysis for
Research, 26(5): 393–404. Functional Foods and Nutraceuticals, 145-
45. OMOTOSO, D.R., OKWUONU, U.C., LAWAL, 204.
O.S., & OLATOMIDE, O.D. (2021). 54. PAL, D., SANNIGRAHI, S. & MAZUMDER, U.
Assessment of the antiproliferative potential of (2009). Analgesic and anticonvulsant effects of
Cissampelos owariensis (P. Beauv) methanolic saponin isolated from the leaves of
extract in Wistar rats. Journal of Physiology, Clerodendrum infortunatum Linn. in mice.
13: 178-183. doi: 10.25081/jp. 2021.v13.7247 Indian Journal of Experimental Biology, 47:
46. MILIAUSKAS, G., VENSKUTONIS, P.R. & VAN 743-747.
BEEK, T.A. (2004). Screening of radical 55. KALITA, P., TAPAN, B.K., PAL, T.K., &
scavenging activity of some medicinal and KALITA, R. (2013). Estimation of total
aromatic plant extracts. Food Chemistry, 85(2): flavonoids content (TFC) and antioxidant
231-237. doi: activities of methanolic whole plant extract of
10.1016/j.foodchem.2003.05.007. Biophytum sensitivum Linn. Journal of Drug
47. SKERGET, M., KOTNIK, P., HADOLIN, M., delivery and Therapeutics, 3(4): 33-37.
HRAS, A.R., SIMONIC, M. & KNEZ, Z. 56. SONIBARE, M.A., ISOLA, A.O. &
(2005). Phenols, proanthocyanidins, flavones AKINMURELE, O.J. (2023).
and flavonols in some plant materials and their Pharmacognostic standardization of the leaves
antioxidant activity. Food Chemistry, 89: 191– of Costus afer Ker Gawl. (Zingiberaceae) and
198. Palisota hirsuta (Thunb.) K. Schum.
48. JACOBO-VELAZQUEZ, D.A. & CISNEROS- (Commelinaceae). Future Journal of
ZEVALLOS, L. (2009). Correlations of Pharmaceutical Sciences, 9: 19.
antioxidant activity against phenolic content
revisited: a new approach in data analysis for
food and medicinal plants. Journal of Food
Science, 74: 107–113.

Nigerian Journal of Scientific Research, 23(1): 2024; journal.abu.edu.ng; https://njsrabuzaria.com.ng 10