IRJCCS

International Research Journal of Chemistry and Chemical Sciences

Vol. 3(2), pp. 037-042, November, 2016. www.premierpublishers.org. ISSN: 0278-5125.

Research Article

Free radical scavenging activity, phytochemistry

and antimicrobial properties of Tetrapleura
tetraptera Seeds
O. U. Igwe*1 and H. C. Akabuike2
1*,2

Department of Chemistry, Michael Okpara University of Agriculture, Umudike, P.M.B. 7267 Umuahia, Abia
State, Nigeria
This study is aimed at investigating the phytochemistry, free radical scavenging
activity and antimicrobial properties of Tetrapleura tetraptera seeds. The results of the
analysis of Tetrapleura tetraptera seeds showed the presence of phenols (0.34 %),
flavonoids (0.91 %), alkaloids (0.52 %), tannins (0.23 %) and saponins (0.51 %).
Vitamins include ascorbic acid (5.23 %), riboflavin (0.11 %), niacin (1.25 %), carotenoid (3.40 %) and thiamine (0.06 %). Proximate compositions were moisture
(14.73 %), ash (7.30 %), crude protein (17.36 %), crude fibre (6.68 %) and lipids (35.19
%). Mineral elements were calcium (1.41 %), magnesium (0.48 %), potassium (0.56 %),
sodium (0.21 %), phosphorus (0.32 %), iron (1.20 %), copper (0.13 %) and zinc (0.77 %).
The seed extract showed potent free radical scavenging activity (6.72 22.8 %) using
ascorbic acid as a standard (12.10 - 42.00 %). The seed extract exhibited significant
antimicrobial activity (7.00 20.00 mm) and the zones of inhibition were compared with
that of ciprofloxacin as standard (15.00 28.00 mm). From these investigations, seed
extract of Tetrapleura tetraptera has proven to be considered an excellent source of
nutriceuticals, a potential antioxidant and antimicrobial agents owing to its high level
of phytochemical and vitamin constituents.
Keywords: Tetrapleura tetraptera, Free radical, Antimicrobial, Phytochemicals, Seed extract

INTRODUCTION
Primary and secondary metabolites of plants have
together played an invaluable role to ensure that man
remains healthy. Natural products obtained from
plants are very important in disease control and
treatment which dates back several many years ago.
Till today people still patronize herbal preparations for
certain ailments especially when modern medicine is
considered to have failed to provide a solution.
However, substances obtained from plants which are
secreted by these plants in response to stress or
other factors have been harnessed and mostly used
as drugs and have proven to be efficacious. Most of
these plants are still understudied and underutilized.
Tetrapleura tetraptera is considered one of them.
T. tetraptera belongs to the family Fabaceae
(Gbadamosi and Obogo, 2013). It is commonly found
naturally in the rainforest belt of tropical Africa

especially in the lowlands and is single-stemmed.

The tree is robust and perennial possessing dark
green leaves, thick, woody base and spreading
branches (Adetunji, 2007). The fruits are green when
tender and dark red-brown when fully ripe and are
about 22-27 cm long and 4-5 cm wide (Adetunji,
2007; Enwereuzoh et al. 2015). The fruit consists of a
fleshy pulp with some small, brownish-black seeds
and possesses a fragrance, which has been
attributed to its insect repellent property (Okwu, 2003;
Adetunji, 2007; Enwereuzoh et al. 2015; Akintola et
al. 2015).
*Corresponding author: Dr. Okenwa U. Igwe,
Department of Chemistry, Michael Okpara University
of Agriculture, Umudike, P.M.B. 7267 Umuahia, Abia
State, Nigeria Email: okenwauigwe@gmail.com

Free radical scavenging activity, phytochemistry and antimicrobial properties of Tetrapleura tetraptera Seeds

Igwe and Akabuike

038

The mature dry pods contain sweet smelling aromatic

pulp and the brownish black seeds are found in them
(Okwu, 2003; Adetunji, 2007; Akintola et al. 2015).

seeds was obtained by sieving with 1 mm aperture

and the sample was stored in an airtight container for oil
extraction and laboratory analysis.

The pod is therefore used as a seasoning spice in

Southern and Eastern Nigeria. The plant possesses
a hard wood that is reddish in colour used for building
houses and other construction work (Okwu, 2003;
Adetunji, 2007; Enwereuzoh et al. 2015; Akintola et
al. 2015).

Extraction of Oil

T. tetraptera has numerous traditional herbal

applications in the treatment of diseases and
infections. A decoction of T. tetraptera pod is used in
the treatment of jaundice (Gbadamosi and Obogo,
2013). In West Africa, the fruit is widely used in
traditional remedies for the treatment of several
conditions including convulsion, gastric ulcer,
rheumatism, fevers, whitlow, skin rashes, smallpox,
malaria and dysentery (Dalziel, 1955; Fleischer et al.,
2006). An infusion of the whole fruit is taken as a
recuperative tonic (Ojewole and Adesina, 1983). The
plant is used in the management of leprosy and
inflammation (Adetunji, 2007). It has been reported
that an infusion of the whole fruit is usually taken by
convalescents to bathe in order to get relief from
feverish conditions, for constipation and as an emetic
(Adetunji, 2007). The methanolic extract of the leaf
has been reported to possess antioxidant effect on
carbon tetrachloride-induced hepatoxicity (Atawodi et
al., 2014) while the aqueous extract of the fruit
possesses hypoglycaemic properties (Lekana-Douki
et al., 2011). The plant is used in the treatment of
hypertension, asthma, epilepsy, schistosomiasis, and
for prevention of post-partum contraction (Ojewole
and Adewunmi, 2004).
More emphasis and concern had been given to T.
tetraptera fruits/pods, pulp and leaf by researchers
and it appears there is a paucity of documented
information on the seeds. Therefore, this research is
aimed at investigating the free radical scavenging
activity, phytochemistry and antimicrobial properties
of T. tetraptera seeds.

The method reported by Igwe and Onuoha (2016)

was employed. In a typical extraction of oil from the
seeds of T. tetraptera, 200 g of the powdered sample
was introduced into a conical flask and 500 ml of 100
% methanol was added followed by continuous
stirring which lasted for about 5 h. The flask was
covered with filter paper and cotton fastened with a
paper tape to make it airtight. Stirring continued the
next day for another 5 h. The mixture was separated
using a Whatman No. 1 filter paper. The process was
repeated on the residue to ensure exhaustive
extraction. The filtrate was pooled together and the
solvent was eventually evaporated in a rotary
evaporator to get the oil.
Determination of Plant Chemicals
Alkaloids and phenols were determined according to
the method of Harborne (1973) while tannin was
determined using the method of Van-Burden and
Robinson (1981). Saponin was determined using the
method of Obadoni and Ochuko (2002). Flavonoids
were determined according to the method of Boham
and Kocipia (1994). Ascorbic acid was determined
using the method of the Association of Vitamin
Chemists described by Kirk and Sawyer (1991). The
B-complex vitamins (thiamine, riboflavin, and niacin)
were determined according to SKALAR Analyzers
method of Barakat et al. (1993) while carotenoid was
determined according to the method described by
James (1995). The macro and micro elements
comprising potassium, sodium, magnesium, calcium,
phosphorus, nitrogen, iron, copper and zinc were
determined according to the method of Shahidi et al.,
(1999). Protein, crude fibre, lipids, ash and moisture
were determined by the method described by James
(1995).
Antioxidant Activity Determination

MATERIALS AND METHODS

Sample Collection and Preparation
Mature T. tetraptera fruits were bought from Ariaria
International Market, Aba, Abia State, Nigeria. The fruits
were identified and authenticated at the Taxonomy
Unit of Forestry Department, Michael Okpara
University of Agriculture, Umudike, Abia State,
Nigeria. The fruit brown pulp was separated from the
seeds by scraping it off with the aid of a sharp knife.
Removal of the seed coats was done with the aid
of mortar and pestle to separate the endosperms
from the seed coat. The endosperms were ground
into powder using a blender. Fine powder of the

The free radical scavenging activity of the oil fraction

of the sample extract was determined using the 1,1diphenyl-2-picrylhydrazyl
(, -diphenyl-picrylhydrazyl; DPPH) method described by
Manzocco et al., (1998). 1.0 g of DPPH, a stable
radical was dissolved in 100 mL of methanol. 3.0 mL
of different concentrations of the test sample were
added to 3.0 mL of a 0.004 % methanol solution of
DPPH and incubated for 30 min. at room
temperature. The decrease in absorbance of the
solution brought about by the test samples was
measured at 517 nm using a spectrophotometer.
Ascorbic acid, which is a known antioxidant, was
used as a reference standard. The radical
scavenging activity was calculated as the percentage

Free radical scavenging activity, phytochemistry and antimicrobial properties of Tetrapleura tetraptera Seeds

Int. Res. J. Chem. Chem. Sci.

inhibition of DPPH discoloration using the equation

below:

% inhibition of DPPH radical

Where,

Ablank Asample

100
Ablank

Ablank is the absorbance of the control

reaction solution (containing all reagents except the

test sample) and Asample is the absorbance of the test
sample.
Antimicrobial Screening
The in vitro antimicrobial activity of the sample extract
was carried out for 24 h culture of three selected
each of bacteria and fungi. The test organisms were
obtained from stock cultures at the Central
Laboratory services Unit of National Root Crops
Research Institute, Umudike, Abia State, Nigeria.
With the aid of a single hole punch office paper
perforator, circular discs of 5 mm diameter were cut
from Whatman No 1 filter paper. The paper discs
were boiled in distilled water for an hour to remove
any residual preservatives. The boiled paper discs
were allowed to drain dry and they were wrapped in
aluminium foil and sterilized in an autoclave at 121
for 15 min. They were however used within 48 h of
production.
The
sensitivity
of
each
test
microorganism to the extract was determined using
the Disc Diffusion Technique (Pelczar and Chan,
1977; Cheesbrough, 2000). A loopful of each test
sample organism was aseptically transferred into the
surface of a sterile solid medium, appropriate for the
test organism. Using a flamed glass hockey, the
inoculum was spread evenly over the surface of the
medium, and then with the aid of a flamed pair of
forceps, the extract bearing paper discs were
carefully placed on the surface of the inoculated
medium at some distance from one another. The
inoculated plates were incubated for 24 h in an
incubator at 37 . They were examined daily for
growth and for the presence of inhibition zones
around the paper discs. The level of sensitivity was
determined by the diameter of the inhibition zone as
measured with a transparent millimeter rule.
Statistical Analysis
Data were replicated three times and expressed as
mean standard deviation (SD) in tables.

RESULTS AND DISCUSSION

The phytochemical constituents of T. tetraptera
seeds are shown in Table 1. The seeds contain
phenols, flavonoids, alkaloids, tannins and saponins.
Flavonoids were the highest constituents followed by

039

alkaloids and then saponins, phenols and lastly,

tannins. According to scientific studies these detected
phytochemicals possess medicinal properties. Also,
according to Mallikharjuna et al. (2007) plants have
the limitless ability to synthesize phenols or their
derivatives and their presence in all types of tissues
is a characteristic feature of plants. Phenolic
compounds are reported to have antioxidant, antiinflammatory and antimicrobial properties (Yadav and
Agarwala, 2011). Flavonoids are polyphenolic
compounds that have been reported to have
antiallergic, antioxidant, anticancer, vasodilatory, antiinflammatory, antimicrobial and immune-stimulating
activities (Igwe and Okwu, 2013a). Alkaloids are plant
bases with pharmacological properties such as
hypotensive,
anticonvulsant,
antiprotozoal,
antimicrobial and antimalarial activities (Aiyelaagbe
and Osamudiamen, 2009). Tannins are reported to
possess anti-tumor, antiviral and antibacterial
activities (Chung et al., 1998). The presence of
tannins in a plant has been associated with ulcer
management, wound healing, control of bleeding and
burns in herbal medicine (Igwe and Okwu, 2013a).
Saponins possess antioxidant, anticancer, antiinflammatory,
hyperglycaemic
and
antifungal
properties (Rausch et al., 2006). Phytochemicals
possess a lot of physiological properties which are
beneficial to human. Phytochemicals in plant-based
foods can improve glucose metabolism as well as
enhance the overall health of diabetic patients by
improving lipid metabolism, antioxidant status,
improving capillary function, and lowering blood
pressure and cholesterol (Kelble, 2006; Kasolo et al.,
2010).
The vitamins constituents of T. tetraptera seeds is
shown in Table 2. The seeds are sources of ascorbic
acid, riboflavin, niacin, -carotenoid and thiamine.
Ascorbic acid and -carotenoid were the most
abundant analytes in Table 2. Ascorbic acid is a good
antioxidant compound and has been involved in
wound healing and prevention of scurvy (Igwe and
Okwu, 2013b). -Carotenoid has also been touted as
having the ability to scavenge free radicals (Igwe and
Okwu, 2013b). The detection of ascorbic acid and carotenoid in the seeds of T. tetraptera suggests that
the seeds may have high nutritional and medicinal
values. Riboflavin, niacin, and thiamine are present in
lower amounts. The proximate composition of the
seed is displayed in Table 3. Lipid content was so
high meaning the seed is a good source of fat and oil.
Lipids play a significant role in the body metabolism.
Apart from their energy-yielding function, they also
constitute a component of the membrane structure.
They protect the body from mechanical injury
(Ononogbu, 2002). High protein content was also
observed in the seeds of T. teraptera. It means that
these seeds are indeed nutritionally rich. The fibre
content will provide bulky stools and this helps to
improve
gastrointestinal
functions,
preventing
constipation and may thus reduce the incidence of

Free radical scavenging activity, phytochemistry and antimicrobial properties of Tetrapleura tetraptera Seeds

Igwe and Akabuike

040
Table 1. Phytochemical constituents of T.
tetraptera seeds

Phytochemicals
Phenols
Flavonoids
Alkaloids
Tannins
Saponins

Composition (%)
0.34 0.10
0.91 0.12
0.52 0.08
0.23 0.02
0.51 0.01

Values are means standard deviation of

three determination

Table 2. Vitamin Composition of T.

tetraptera seeds

Vitamin
Ascorbic acid
Riboflavin
Niacin
-Carotenoid
Thiamine

Composition (%)
5.23 0.21
0.11 0.02
1.25 0.25
3.40 0.64
0.06 0.02

Values are means standard deviation

of three determinations

Table 3. Proximate composition of T.

tetraptera seeds

Constituents
Moisture content
Ash content
Crude protein
Crude fibre
Lipids

Composition (%)
14.73 0.34
7.30 0.75
17.36 1.10
6.68 0.71
35.19 0.18

Values are means standard deviation of

three determinations .

Table 4. Mineral elements composition of

T. tetraptera seeds

Mineral elements
Calcium
Magnesium
Potassium
Sodium
phosphorus
Iron
Copper
Zinc

Composition (%)
1.41 0.21
0.48 0.02
0.56 0.03
0.21 0.03
0.32 0.05
1.20 0.05
0.13 0.03
0.77 0.04

Values are means standard deviation of

three determinations

metabolic diseases. The mineral elements analyzed

and found in the seeds of T. tetraptera were calcium,
magnesium, potassium, sodium, phosphorus, iron,
copper and zinc (Table 4). Of all these elements,
calcium, iron and zinc were found in high amounts.
Calcium is important for strong bone and teeth
formation. Iron is needed in building red blood cells
while zinc is important for wound healing and in
carbohydrate metabolism (Igwe and Okwu, 2013b).
The free radical scavenging activity of the seed
extract of T. tetraptera is shown in Table 5. The

extract showed comparable free radical scavenging

activity with the ascorbic acid used as a standard.
The presence of phenols, flavonoids, saponins,
ascorbic acid and carotenoid in the extract was
responsible for the free radical scavenging activity
exhibited by this extract (Rausch et al., 2006; Yadav
and Agarwala, 2011; Igwe and Okwu, 2013a; Igwe
and Okwu, 2013b). Table 6 shows the antimicrobial
activity of T. tetraptera seed extract. The extract
showed
significant
inhibitions
against
the
microorganisms and zones of inhibition were
comparable to that obtained using ciprofloxacin as a

Free radical scavenging activity, phytochemistry and antimicrobial properties of Tetrapleura tetraptera Seeds

Int. Res. J. Chem. Chem. Sci.

041

Table 5. Free radical scavenging activity of extract of T. tetraptera seeds

Concentration (mg/ml)

Radical Scavenging (%)

Ascorbic acid
Extract

4.0
8.0
12.0
16.0
20.0

12.10 0.92
22.00 0.45
25.00 1.75
38.00 0.01
42.00 0.06

6.72 0.54
10.0 0.3
15.5 1.55
18.9 0.08
22.8 0.18

Values are means standard deviation of triplicate results.

Table 6. Antimicrobial activity (zone of inhibition) of T. tetraptera seed extract

Microorganism
S. aureus
E. coli
P. aeruginosa
C. albicans
A. niger
P. notatum

T. tetraptera (mm)
7.00 1.41
9.00 0.14
15.00 0.80
14.00 0.70
20.00 0.71
16.00 0.90

Ciprofloxacin (mm)
18.00 0.70
24.00 0.28
15.00 0.80
21.00 0.90
23.00 0.70
28.00 0.20

Values are means standard deviation of triplicate results.

standard antimicrobial agent. The order of inhibition

against the tested organisms was A. niger > P.
notatum > P. aeruginosa > C. albican > E. coli > S.
aureus. An important characteristic of plant extracts
and their components is their hydrophobicity, which
enables them to partition the lipids of the bacterial
cell membrane and mitochondria, disturbing the cell
structures and rendering them more permeable
(Sikkema et al., 1994). The presence of
phytochemicals in the extract, such as phenols,
flavonoids, alkaloids, tannins and saponins could be
the reason behind the strong antimicrobial activity
exhibited by the extract (Chung et al., 1998; Rausch
et al., 2006; Aiyelaagbe and Osamudiamen, 2009;
Yadav and Agarwala, 2011; Igwe and Okwu, 2013a).

CONCLUSION
From this investigation, the seeds of T. tetraptera are
well endowed with phytochemicals such as alkaloids,
flavonoids, phenols, tannins, and saponins. Vitamins
and mineral elements compositions proved the seeds
to be a good source of materials needed by the body
for sound health. The extract of T. tetraptera seeds
exhibited marked antimicrobial and free radical
scavenging activities which are attributable to the
presence of bioactive phytochemicals in them. T.
tetraptera seeds could find application in food and
flavour industries where they could be used as spices
cum antimicrobial and antioxidant agents.

ACKNOWLEDGEMENTS
The authors are grateful to Mr. I. K. Ndukwe of Plant
Taxonomy Section, Forestry Department, Michael
Okpara University of Agriculture, Umudike, for
identifying and authenticating the plant sample.

COMPETING INTERESTS
Authors have declared that no competing interests
exist.

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Accepted 28 September 2016.
Citation: Igwe OU, Akabuike HC (2016). Free radical
scavenging activity, phytochemistry and antimicrobial
properties of Tetrapleura tetraptera Seeds. International
Research Journal of Chemistry and Chemical Sciences,
3(2): 037-042.

Copyright: 2016 Igwe and Akabuike. This is an openaccess article distributed under the terms of the
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unrestricted use, distribution, and reproduction in any
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Free radical scavenging activity, phytochemistry and antimicrobial properties of Tetrapleura tetraptera Seeds