International Journal of Herbs, Spices and Medicinal Plants

Vol. 4(2), pp. 050-057, June, 2019. © www.premierpublishers.org, ISSN: 2123-7362

Research Article
Characterization of the Volatile Components and Antimicrobial
Properties of the Ethanol Leaf Extract of Uvaria chamae Grown
in Eastern Nigeria
*1Iwu Irenus Chinonye, 2Oze Rita Nwanneamaka, 3Onu Uchenna Lynda, 4Onwumere Fidelis,
5Ukaoma Adanma Augustina
1,2,3,4Department of Chemistry, Federal University of Technology Owerri, Imo State, Nigeria
5Department of Biological Sciences, Federal University of Technology Owerri, Imo state, Nigeria

The characterization of the ethanol leaf extract of Uvaria chamae was carried out with the aim of
identifying and determining the chemical compounds present in the extract. Initial phytochemical
results showed the presence of flavonoids, saponins, tannins. Phenols, steroids and alkaloids.
Interpreted spectrum obtained from the GC-MS revealed twelve absorption peaks. Peak 1 was
identified as Benzene carboxylic acid with molecular weight of 122g and molecular formula
C7H6O2. Similarly, peaks 2-12 were identified as, hexadecanoic acid methyl ester, hexadecanoic
acid, 11-octadecenoic acid methyl ester, phytol, 6-octadecanoic acid, octadecanoic acid,
tetradecanamide. hexadecanoic acid-2,3-dihydroxpropyl ester, 9-octadecenamide, 9,12-
octadecadienoyl chloride and 13-octadecanal with corresponding molecular formulas of C17H34O2,
C16H36O2, C19H36O2, C20H40O, C18H34O2, C18H36O2, C14H29NO, C19H38O4, C18H35NO, C18H31ClO and
C18H34O respectively. The extract inhibited the growth of some selected human pathogen;
Pseudomonas aureginosa by 6mm with minimum inhibitory concentration (MIC) of 50mg/cm3.
Similar results were also obtained for Candida albicans 8mm with MIC 50mg/cm3 and
Trichophyton spp 4mm with MIC OF 100mg/cm3
Keywords: (Characterization, Gas Chromatography, Phytochemicals, Mass Spectrometry, Pathogens)

INTRODUCTION

Uvaria chamae is a plant tropical to eastern Nigeria and is
commonly called finger root plant which belongs to the
family of Annonaceae. It is a small tree that grows to about
4.5m high Moses et al.,( 2013) It is commonly found in the
savannah and rain forest regions of Nigeria and other
African countries. It is called “Mmimi ohia”, “Kas kaifi” and
“Akisan” amongst the Ibos, Hausas and Yorubas
respectively Adetunji, (1999). The fruits are yellow when
ripe and have a sweet pulp which is widely eaten. The fruit
carpels are in finger-like clusters. The plant has been used
for treatment of fevers, tumour growth,stroke and cases of
venereal disease, Ayenusi (1978).The leaf extracts of U.
chamae has been reported to possess antibacterial
Oluremi et al., (2009), antifungal Okwuosa et al.,( 2012),
antispasmodic, anti-trypanosomal and anti-inflammatory
properties. The roots of the plant has been reported to
possess antibacterial, antioxidant Kone et al., (2015), anti-
inflammatory, oxytocic. Okwu and Iroabuchi, (2009) and
anti-sickling Thierry et al., (2012) activities. Two benzyl
dihydrochalcones; chamuvaritin and chamuvarin have
been isolated from the roots of U. chamae. Uwaifo and
Bababunmi, (1984). The leaves of U. Chamae have been
used to treat wounds and sores, injuries, swellings, and to
treat yellow fever. The roots, barks and leaves of U.
chamae are used traditionally in the treatment of
diarrhoea, cough and urinary tract infections Chika et al.,
(2007).
The plant is rich in sesquiterpene hydrocarbons dominated
by germacrene D and γ-cadinene. (Okwwuosa et al.,
2012), Kone et al., 2015.) the root extract of U. Chamae
has been reported to contain flavonoids, alkaloids, cardiac
glycosides, terpenoid and terpenes, saponin, tannin,
*Corresponding Author: Iwu Irenus Chinonye, Department of
Chemistry, Federal University of Technology Owerri, Nigeria.
Tel.: +2348032444212. E-mail: iwu.chinonye@yahoo.com. Co-
Authors Email: 2ozerita@yahoo.com, Tel.: +2348038916979;
3ucheonu2018@gmail.com,
Tel.: +2348037676079;
4fideconwumere2@gmail.com,
Tel.: +2347038824111;
5ukaomaadanma@yahoo.com; Tel: +2348069380796

Characterization of the Volatile Components and Antimicrobial Properties of the Ethanol Leaf Extract of Uvaria chamae Grown in Eastern Nigeria
Iwu et al. 051
proteins and sugars. The extracts and aspirin have been
reported to inhibit carrageenan-induced paw oedema on
albino rats and mice. Okwu and Iroabuch. (2009). The drug
benzyl benzoate used in antifungal preparations has a
mutagenic compound, chamuvaritin, a benzyl
dihydrochalcone that was isolated from U. Chamae.
Therry et al., (2012). Recently, uvarinol, a novel cytotoxic
tribenzylated flavanone compound has been isolated from
U. Chamae. Uwaifo and Bababunmi, (1984). The root is
used in Nigeria as a purgative. The root bark is used for
respiratory catarrh and the root extract is used in
phytomedicine for the treatment of piles, menrrhegia,
epiostaxis, haematuria and haemalysis. Oliver-Bever,
1986. Its root infusion is used to cure abdominal pains.
The juice from the roots, stems or leaves is commonly
applied to wounds and sore. Shukda and Shital. (1995.)
wrote that the extracts of the roots, barks and leaves are
used to treat gastroenteritis, vomiting, diarrhea, dysentery,
wounds, sore throats, inflamed gums and a number of
other ailments. Irvin, (1961). Uvaria chamae methanol
extract has been shown to neutralized some biological
effects of Naja nigricollis venom. The leaf extract had
shown anti venom activity in animal models and could
potentially be used for therapeutic purpose in case of
snake bite (Omale et al.,(2012). Methanolic extracts of the
root, stem and leaf of Uvaria chamae have been evaluated
for their antibacterial activity against Methicillin-resistant
Staphylococcus aureus. The stem bark extract inhibited
the growth of all the tested organisms but the leaf extract
showed the least antibacterial activity as reported by
Oluremi et al., (2009.). Uvaria chamae root contain
abundant cardiac glycosides. Similar reports in other plant
species were obtained by Okon et al., (2013). They
reported that cardiac glycosides can be used in the
treatment of diseases associated with the heart and are
currently used by herbalist, to treat tumour Piett, (2000).
The cardiac glycosides found in Uvaria chamae root can
similarly be used for the treatment of heart diseases.
Monosodium glutamate has detrimental effect on
haematological parameters but Uvaria chamae ethanolic
extract is a potent remedy against MSG-induced toxicity.
Ibukun et al., (2015) Despite its many uses for the
treatment of various diseases, its full content has not been
fully characterised.
MATERIALS AND METHOD
SAMPLE PREPARATION
The leaves of finger root (Uvaria chamae were obtained
from F.U.T.O farmland, Owerri North L.G.A. The plant was
identified by Dr Ibeawuchi of Crop Science Department
Federal University of Technology Owerri. The sample was
cut into bits, room dried, ground into powdery form and
stored in an air-tight container afterwards before the
commencement of the analysis.
FROTHING TEST FOR SAPONINS
This test is based on the ability of the saponins to produce
froth in aqueous solution. 5g of the plant extract was
Characterization of the Volatile Components and Antimicrobial Properties of the Ethanol Leaf Extract of Uvaria chamae Grown in Eastern Nigeria
weighed into a test tube and 50cm 3 of water was added
and extracted after two hours. The water extract was
shaken vigorously in a conical flask. The production of a
stable froth indicates the presence of saponins in the
sample. (Iwu et al., 2018a)
TEST FOR FLAVONOIDS
5g of the sample was soaked with 20cm 3 of water and left
to stand for 2 hours, it was then filtered and to the filtrate
drops of ammonia and 3cm 3 of concentrated H2SO4 was
added. A yellow precipitate which disappears on storage
indicates the presence of flavonoids. Iwu et al., (2018b)
TEST FOR ALKALOIDS
5g of the sample was extracted using 20% acetic acid in
ethanol.5cm3 of the extract was treated with Wagner’s
reagent (iodine crystals and KI). A yellowish brown
precipitate indicates the presence of alkaloids.
TEST FOR TANNINS
5g of the root sample was weighed into a beaker and
50cm3 of water was added and allowed to soak properly
for two hours and extracted. The extract was treated with
drops of ferric chloride. A blue-black precipitate indicates
the presence of tannins. Iwu et al., (2018b)
TEST FOR STEROIDS
5cm3 of the water extract was treated with concentrated
H2SO4 in acetic anhydride. The formation of a blue-green
colour indicates the presence of steroids.
TEST FOR PHENOLS
20cm3 of the water extract was treated with 5cm 3 of
concentrated sulphuric acid and drops of sodium nitrate
(NaNO3). 2cm3 of sodium hydroxide was added to the
mixture. A blue precipitate indicated the presence of
phenols. Iwu et al., (2016a)
TEST FOR GLYCOSIDES
20cm3 of the water extract was treated with Fehling
solutions of A and B in equal amount and boiled. A
brownish red precipitate indicates the presence of
glycoside.
PREPARATION OF SAMPLES FOR GC-MS ANALYSIS
Two hundred grams of sample was soaked in 400cm3
ethanol for 48 hours and then extracted. The extract was
re-extracted using chloroform to obtain chloroform soluble
extract. This was centrifuged at 10,000 rpm for 20 minutes
and the clear supernatant oil was subjected to GC-MS
analysis.

GC-MS EXPERIMENTAL PROCEDURES
GC-MS analysis was carried out with SHIMAZU Japan
Gas Chromatography 5890-11 with a fused GC column
OV 101 coated with polymethyl silicon (0.25 mm x 50 m)
and the conditions are as follows: Temperature
programming from 80 – 200oC held at 80oC for 1 minute,
the rate is 5oC/min and at 200oC for 20 minutes. FID
Temperature of 300oC, injection temperature of250oC,
carrier gas is Nitrogen at a flow rate of 1 cm 3/min and split
ratio of 1:75. GC-MS Gas chromatography, Mass
spectrum analysis were conducted using GC-MS QP 2010
Plus Shimazu Japan with injector Temperature at 230oC
and carrier gas pressure of 100kpa. The column length
was 30 m with a diameter of 0.25 mm and the flow rate of
50m/min. The eluents were automatically passed into the
Mass Spectrometer with a detector voltage set at 1.5kv
and sampling rate of 0.2 seconds. The Mass Spectrometer
was also equipped with a computer fed Mass Spectra data
bank, HERMCE Z 233 M-Z centrifuge Germany was used.
Reagents and solvents such as Ethanol, Chloroform,
Diethyl ether, hexane all of analytics grade was obtained
from Merck Germany. Iwu et al., (2016b, 2018c)
ANTIMICROBIAL ANALYSIS
The microorganisms; Staphylococcus aureus,
Streptococcus spp Pseudomonas aeruginosa, Aspergillus
niger, Candida albicans and Trycophton spp were used for
the analysis. They are clinical isolates of human
pathogens obtained from the Federal Medical Centre
Umuahia and were brought to the laboratory and
resuscitated in buffered peptone broth (Secharian chemie)
and thereafter into nutrient agar medium and incubated at
37oC for 24 hrs. Iwu et al., (2018b)
ANTIBACTERIAL ASSAY
The test solution of each extract was prepared by
dissolving 0.1 g of the plant extract separately in 1.0cm3 of
dimethyl sulphoxide (DMSO) to get a concentration of
100mg/cm3. The antibacterial activity was performed by
filter paper disc diffusion technique. Filter paper disc
(Whatman No 1.6 mm diameter) were placed in glass petri
dishes and sterilized in hot air over. Iwu and Onu,
(2018).The media (10g nutrient Agar in 200cm 3 distilled
water, autoclaved at115oC for 30 minutes) was cooled to
50oC. The sterile nutrient Agar media were poured into the
sterile petri dish and allowed to solidify. The bacteria were
swabbed with a sterile wire loop. Each disc was
impregnated with 0.2cm 3 of plant extract. .The discs were
used after drying them in an incubator at 40oC to remove
any trace of solvent. Discs were introduced into the surface
of the medium. The plates were microbated at 37oC for 24
hours to obtain zones of inhibition. The experiments were
repeated three times for each extract and twice for
reference antibiotics to minimize error and the average of
these values were recorded. Kanayo and Ezeugo,(2006)
Characterization of the Volatile Components and Antimicrobial Properties of the Ethanol Leaf Extract of Uvaria chamae Grown in Eastern Nigeria
Int. J. Herbs, Spices Med. Plants 052
MINIMUM INHIBITORY CONCENTRATION (MIC).
The minimum inhibitory concentration of the extract was
determined by incorporating constant volume 0.2cm3 of
each diluents of the extract into the perforated disc on a
seeded nutrient agar plate as described in the anti-
microbial susceptibility test section. 0.1g of each extract
was dissolved in 1cm 3 of DMSO to obtain 100mg/cm 3. This
concentration of DMSO was then doubled to obtain
50mg/cm3 then doubled again to obtain 12.5mg/cm 3 and
again to obtain6.25mg/cm 3. Each concentration was then
used in the method earlier described to obtain zone of
inhibition. The least concentration that showed inhibitory
zones was taken as the MIC.
RESULTS AND DISCUSSION
The results obtained from the phytochemical screening of
the leaf extract of sample are presented in table 1 below.
Initial results showed the presence of flavonoids,
saponins, tannins. phenols , steroids and alkaloids
Table 1. Phytochemical content of the leaf extract of uvaria
chamae
Phytochemical Inference
Alkaloid ++
Flavonoid ++
Saponons ++
Steroids ++
Glycosides ++
Tannins ++
Phenols ++
Key: ++ present
Alkaloids are vast and vary a lot in their activity when
ingested by man and livestock. Some alkaloids are useful
and important in medicine and constitute most of the
valuable drugs currently used by humans. They are
reported to have marked physiological effect on animals.
Edeoga and Eriata, (2001). Iwu et al., (2018c)
Flavonoids have been shown to be highly effective
scavengers of most oxidizing molecules. Tukappa and
Londonkar, (2013). Flavonoids are the major nutraceutical
ingredients that are in plants. The best described property
of almost every group of flavonoids is their capacity to act
as anti-oxidants. The flavones seem to be the most
powerful flavonoid for protecting the body against reactive
oxygen species (ROS). Antibacterial activity has been
displayed by a number of flavonoids,. Quercetin has been
reported to completely inhibit the growth of
Staphylococcus aureus. Havesteen (1983). Flavonoids
also possess anti-inflammatory and analgesic effect as
well as anti-ulcerogenic activity. Shahid et al., (1998)
The infusions of Momordica charantia and Uvaria chamae
are taken as a remedy for gonorrhea and jaundice
Draughton, (2004), Mohammed et al.,(2013) This is
probably due to antibacterial action of saponins. Saponins
are foam forming in nature and have been implicated as a
Iwu et al. 053
Fig 1: GC/MS GC-MS spectra of the the ethanolic leaf sample of Uvaria chamae
bioactive antibacterial agent of plant. Mandal et al., (2005.}
Saponins are also useful for utilization in foods that need
sustained foam volume such as ice-creams. Plant
saponins may serve as anti-feedants to protect the plant
against microbes and fungi. Some plant saponins may
enhance nutrient absorption and aid in food digestion.
Saponins have been used as a pharmacological and/or
immunological agent that modifies the effect of other
agents in vaccines. Saponins from plants have been
shown to significantly augment the cytotoxicity of
immunotoxins and other target toxins directed against
human cancer cells. Tannins are astringent, bitter plant
polyphenol compounds that bind to and precipitate
proteins and various other organic compounds including
amino acids and alkaloids. The tannin compounds are
widely distributed in many species of plants where they
play a role in protection from predation and perhaps also
as pesticides and in plant growth regulation. The
astringency from tannin is what causes the dry puckery
feeling in the mouth following the consumption of unripe
fruits or red wine. Tannins are important ingredients used
in process of making tannin leather. Medicinally, tannins
are used as anti-diarrhea, haemostatic and anti-
hemorrhoid compounds Plant leaves with high tannin
content has been used successfully as hops alternative in
beer. Hutchinson and Dalziel,.(1963).
Characterization of the Volatile Components and Antimicrobial Properties of the Ethanol Leaf Extract of Uvaria chamae Grown in Eastern Nigeria
The presence of phenolic compounds in the leaf of Uvaria
chamae indicates that this plant might be an anti-microbial
agent. This is because phenols and phenolic compounds
have been extensively used in disinfections and remains
the standard with which other bactericides are compare.
Okwu and Okwu,( 2004}. Phenolic compounds acts as
electron donors and are readily oxidized to form phenolate
ions. This gives rise to protonated phenol which is used as
a cleaning agent. Extracts from leaves of Uvaria chamae
therefore have potent antiseptic or bactericidal properties.
The presence of phenol further indicates that Uvaria
chamae could act as anti-inflammatory, anti-clotting,
immune enhancers and hormone modulators.
Glycosides are molecules in which a sugar is bound to
another functional group via a glycosidic bond. Glycosides
play numerous important roles in living organisms. Many
plant store chemicals in form of inactive glycosides. Many
such plant glycosides are used as medications. Some
glycosides have shown some evidence of pharmacological
effects in patients with hypertension or with type-2
diabetes but concluded that further study was required to
determine the proper dosages
The result of the GC/MS analysis of Uvariae chamae is
contained in fig 1 above;
 Int. J. Herbs, Spices Med. Plants 054

The GC-MS spectrum of finger root (fig 1) has 12 and % oil composition of 45.04. Peak 7 was identified as
absorption peaks. Peak 1 was identified as benzene Octadecanoic acid with molecular formula of C18H36O2 and
carboxylic acid with percentage oil composition of 0.39 and molecular weight 284g with % oil composition of 9.49.
a molecular weight of 122g, its molecular formula is Peak 8 was identified as Tetradecanamide. Its molecular
C7H6O2.. Similarly, peak 2 was identified as hexadecanoic weight and formula are 227g and C14H29NO respectively.
acid methyl ester having a % oil composition of 0.93 with Peak 9 was identified as Hexadecanoic acid, 2,3-
a molecular weight of 270g and a molecular formula of dihydroxypropyl ester with a molecular weight and formula
C17H34O2. Peak 3 was identified as hexadecanoic acid, its of 330g and C19H38O4 and % oil composition of 2.99. Peak
% oil composition is 10.61 and its molecular weight is 256g 10 was identified as 9-octadecenamide with molecular
with a molecular formula of C16H36O2. Peak 4 was weight and formula of 281g and C18H35NO. Its % oil
identified as 11-octadecenoic acidFigmethyl ester with a composition is 2.32. Peak 11 was identified as 9,12-
molecular weight of 296g and 1molecular formula of octadecadienoyl chloride with molecular weight and
C19H36O2 and its % composition was 4.75. Peak 5 was formula of 298g and C18H31ClO. Its % oil composition is
identified as Phytol with molecularGC- weight of 296g and 12.55. Peak 12 was identified as 13-octadecanal with
molecular formula C20H4OO. Its % oil MScomposition is 1.92. molecular weight and formula of 266g and C18H34O
Peak 6 was identified as 6-octadecanoic
spe acid with respectively. Its % oil composition is 4.46.
molecular formula C18H34O2 and molecular weight 282g
ctr
Table 2. Interpreted values from thea GC/MS
of spectrum of Uvaria chamae leaf extract
Chromatographic % oil Chemical name Molecular Molecular
peak composition the formula weight
1 0.39 pla
Benzene carboxylic acid C7H6O2 122
2 0.93 Hexadecanoic acid methyl ester
nt C17H34O2 270
3 10.61 Haxadecanoic acid C16H32O2 256
ext
4 4.75 11-octadecenoic acid methyl ester C19H36O2 296
5 1.92 rac
Phytol C20H40O 296
6 45.04 t6-octadecenoic acid C18H34O2 282
7 9.49 Octadecanoic acid C18H36O2 284
8 4.55 Tetradecanamide C14H29NO 227
9 2.99 Hexadecanoic acid,2,3 dihydroxypropyl ester C19H38O4 330
10 2.32 9-Octadecenamide C18H35NO 281
11 12.55 9,12-octadecadienoyl chloride C18H31ClO 298
12 4.46 13-octadecanal C18H34O 266

The structural elucidation of the compounds obtained from 5. Phytol

the GC/MS analysis of the sample are contained in fig 2
below.
1. Benzene carboxylic acid

6. 6-octadecenoic acid

2. Hexadecanoic methyl ester acid

7. Octadecanoic acid

3. Hexadecanoic acid

8. Tetradecanamide
4. 11-octadecenoic acid methyl ester

Characterization of the Volatile Components and Antimicrobial Properties of the Ethanol Leaf Extract of Uvaria chamae Grown in Eastern Nigeria
Iwu et al. 055

9. Hexadecanoic acid, 2, 3-dihydroxypropyl ester inflammation with associated pains, pus, swelling redness
and impaired vision Pseudominas spp causes bacterial
blight in guinea corn. Similarly the extract showed activity
against Candida albican with 8mm diameter of inhibition.
Candida albicans is a fungus which causes yeast infection.
Candida yeast normally live in the skin and mucous
membranes without causing infection; however,
overgrowth of these organisms can cause symptoms to
10. 9-octadenamide develop. They cause diseases based on the area of the
body they affect; candidiasis that develop in the mouth or
throat is called thrush while in the vagina is called yeast
infection. Invasive candidiasis occurs when candida
species enter the bloodstream and spread throughout the
body. The extract also showed activity against
Trichophyton spp 4mm. This pathogen is a fungus that
11. 9, 12-octadecadienyl chloride cause tinea, athletes’ foot, ring worm, jock itch, nail, beard
and skin infections. The extracts exhibited some level of
inhibitory effects against some of the studied pathogens
which have been implicated in one bacterial infection to the
other in human and plant. Okwuosa et al, (2012), Kone et
12. 13-octadecanal al., (2015)

CONCLUSION

The work so far carried out has shown that the leaf extract
of Uvaria chamae is rich in phytochemicals and other
volatile compounds. The compounds have far reaching
medicinal applications. The extract may be applied in
Fig 2: Structures of the compounds obtained from the GC- cases involving bacteremia, pneumonia, folliculitis,
MS spectrum OF Uvaria chamae leaf extract swimmer ear which is an ear infection accompanied with
swelling, ear pus. Itching, discharge and difficulty in
Antimicrobial analysis of Uvaria chamae hearing, eye inflammation with associated pains, pus,
swelling redness and impaired vision, candidiasis, tinea,
The antimicrobial analysis result obtained from the leaf athletes foot, ring worm, jock itch , nail , beard and skin
extract of Uvaria chamae are shown in table 3 below. infections.
Table 3: Result of the antimicrobial analysis of Uvariae
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Accepted 16 May 2019

Citation: Iwu IC, Oze RN, Onu UL, Onwumere F, Ukaoma

AA (2019). Characterization of the Volatile Components
and Antimicrobial Properties of the Ethanol Leaf Extract of
Uvaria chamae Grown in Eastern Nigeria. International
Journal of Herbs, Spices and Medicinal Plants. 4(2): 050-
057.

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Characterization of the Volatile Components and Antimicrobial Properties of the Ethanol Leaf Extract of Uvaria chamae Grown in Eastern Nigeria