Advances inEnvironmental Biology, 9(24) November 2015, Pages: 389-396
AENSI Journals
Advances inEnvironmental Biology
ISSN-1995-0756
EISSN-1998-1066
Journal home page: http://www.aensiweb.com/AEB/
Phytochemical Screening and Bio Assay of the Anti-Microbial Activity of Three
Species of Bamboo in Nueva Ecija, Philippines
1Mary
Jhane G. Valentino, 2Leslie S. Ganado, 2Michelle R. Ganado, 1Jerwin R. Undan
1
Faculty, Department of Biological Sciences, College of Arts and Sciences, Central Luzon State University, Science City of Munoz, Nueva
Ecija, Philippines, 3120.
2
BS Biology students, Department of Biological Sciences, College of Arts and Sciences, Central Luzon State University, Science City of
Munoz, Nueva Ecija, Philippines, 3120.
ARTICLE INFO
Article history:
Received 28 September 2015
Accepted 15 November 2015
Available online 24 November 2015
Keywords: bactericidal, bamboo,
colonization , eradicant, fungicidal,
protectant, inhibition
ABSTRACT
Bamboo has been utilized as one of the traditional Asian and Chinese medicine since
time immemorial. This study was undertaken to determine the phytochemicals present
in ethanol, acetone and hot water leaf- extracts of three species bamboo, which include
Bambusa blumeana var. Luzonensis, Bambusa blumeana and Bambusa vulgaris. Their
antimicrobial potentials were also assessed against bacterial pathogens (Escherichia
coli ATCC 25992 and Staphylococcus aureus ATCC 29213) and fungal pathogens
(Aspergillus niger and Fusarium semitectum). Results of the phytochemical screening
showed that tannins, steroids, saponins, terpenoids and cardiac glysosides are present
in all acetone and ethanol leaf extracts of bamboo while alkaloids are absent in all
bamboo leaf extracts. On the other hand, hot water extracts of B. blumeana lack
tannins, steroids, cardiac glycosides and terpenoids. Also, no traces of tannins and
steroids in B. blumeana var Luzonensis hot water extracts were observed. Bioassay of
the antimicrobial activity of the bamboo leaf extracts showed their potentials as
protectant against the bacterial and fungal test pathogens. For the bactericidal activity of
the bamboo leaf extracts, small zones of bacterial colonization were recorded ranging
from 7.0mm to 32.1mm for E.coli and 6.0mm to 16.9mm against S. aureus at 12 hrs
and 24 hrs of incubation. Whereas, eradicant test showed their inability to inhibit the
growth of the bacterial pathogens. For the fungicidal activity, results also revealed their
ability as protectant against A. niger and F. semitectum preventing fungal growth at 24
hrs of incubation. However, an observable decreased in their efficacy as protectant was
observed after 48 hrs of incubation which resulted to the colonization of the fungal
pathogens in all plates treated with different leaf extracts. Based on the results of the
study, it can be concluded that ethanol, acetone and hot water extracts of three species
of bamboo contain phytochemicals which can be attributed to their pharmacological
activities. Also, their antimicrobial potentials as protectant for both bacteria and fungi
were revealed suppressing the growth of the test pathogens.
© 2015 AENSI Publisher All rights reserved.
ToCite ThisArticle: Mary Jhane G. Valentino, Leslie S. Ganado, Michelle R. Ganado, Jerwin R. Undan, Phytochemical Screening and Bio
Assay of the Anti-Microbial Activity of Three Species of Bamboo in Nueva Ecija, Philippines. Adv. Environ. Biol., 9(24), 389-396, 2015
INTRODUCTION
Bamboo was once a symbol of the Orient that grows throughout the world and has originated from
Southeast Asia, where it is considered as a natural component of the forest ecosystem [1]. They are classified
under the Poaceae family and subfamily Bambusoidae which is composed of woody and some herbaceous
bamboo of about 1575 species and includes two tribes [2].
In the Philippines 62 species have been identified and cultivated, 21 species of which are regarded as
native or endemic. Some of the species are introduced in the prehistoric era while the rest is introduced by other
countries. In addition, bamboo is commonly referred to as “poor man’s timber” since most of its parts is being
utilized in building houses, furniture making and weaving. Furthermore, bamboo shoots are being utilized as
source of food [3, 4].
Several studies revealed the presence of phytochemicals in different parts of bamboo plant. Phytochemicals
are considered as non-essential nutrients and non-nutritive plant chemicals that have protective or disease
Corresponding Author: Mary Jhane G. Valentino, Department of Biological Sciences, Science City of Munoz, Nueva
Ecija, Philippines, 3120.
E-mail: [email protected]
390
Mary Jhane G. Valentino et al, 2015
Advances inEnvironmental Biology, 9(24) November 2015, Pages: 389-396
preventive properties [5]. Plants with phytochemicals have been proven as anti aging agent and it can also
reduce the risk factors of many diseases including cancer, heart disease, stroke, high blood pressure, cataracts,
osteoporosis, diabetes and urinary tract infections [6, 7, 8].
Phytochemicals in bamboo includes flavonoids and their glucosides, active polysaccharides, special amino
acids and their peptides, aromatic elements as well as microelements such as manganese, zinc and selenium
which was already been proven to have biological functions in the production of novel drugs [9]. Leaf extract of
different species of bamboo showed an antimicrobial against some bacteria which causes diseases in humans
[10]. Moreover, studies in recent years revealed several biological activity and pharmacological effects of
bamboo extracts, such as anti-free-radicals, anti oxidant, anti-aging, antifungal, insecticide, lipid-regulating and
cardio-cerebro-vascular protection. [11].
In the present study, different species of bamboo were surveyed in seven Municipalities of Nueva Ecija,
Philippines. Three species of which, namely Bambusa blumeana variety Luzonensis (Kawayang Bayog),
Bambusa blumeana J.A. & J.H. Schultes (Kawayang Tinik) and Bambusa vulgaris (Kawayang Kiling) were
selected as test plants to determine their phytochemical composition and to evaluate their anti microbial
functions.
MATERIALS AND METHODS
Prior to phytochemical screening and bioassay of the anti microbial activity of bamboo, a preliminary
survey was conducted in seven Municipalities of Nueva Ecija, Philippines which include San Jose, Lupao,
Llanera, Rizal, Science City of Munoz, Talavera and Sto. Domingo to identify and to determine the occurrence
of the different species of bamboo. Among the surveyed and identified species of bamboo, the three most
commonly utilized and frequently occurring bamboo (Bambusa blumeana variety Luzonensis (Kawayang
Bayog), Bambusa blumeana J.A. & J.H. Schultes (Kawayang Tinik) and Bambusa vulgaris (Kawayang Kiling))
were used as test plants.
Collection and preparation of the plant materials:
Matured leaves of B. blumeana var Luzonensis, B. blumeana and B. vulgaris were collected, shaded dried
and then crushed into powdered form.
Extraction procedures:
Bamboo leaves were extracted using three solvents which include 80% acetone, 80% ethanol and hot water.
Bamboo leaf extracts was prepared by mixing 25g of powdered bamboo leaves to 100 mL of solvent. The
solvents was then separately evaporated in rotary evaporator under reduced pressure at 50°C to yield ethanol
and acetone extracts. The filtrate was poured in a sterile amber bottle and placed in the refrigerator until used.
Screening of phytochemical composition of bamboo leaves:
Screening of phytochemical constituents was carried out following the standard methods described in
Laboratory Manual for the UNESCO (1986) [12]. The various phytochemical constituents tested were the
following: tannins, steroids, saponins, flavonoids, terpenoids, cardiac glycosides, and alkaloids.
Test for Tannins:
An aliquot of 0.5mL extract of the selected toxic plants was added to 10mL of distilled water on a test tube
and filtered. Two mL of 5% FeCl was added to the filtered sample. Brownish green or blue black coloration was
observed for the presence of tannins.
Test for Steroids:
Two milliliters of acetic anhydride was added to a 5 mL extract of plant sample with 2 mL of H2SO4.Violet
to blue or green precipitate was observed for the presence of steroids.
Test for Saponins:
A volume of 0.5mL of extract was added to 10mL distilled water and was shaken vigorously to obtain a
stable persistent froth. Persistent frothing indicated the presence of saponin.
Test for Flavonoids:
Few drops of 1% NH3 solution were added to 5 mL extract of plant sample in a test tube. Yellow coloration
indicates the presence of flavonoids.
391
Mary Jhane G. Valentino et al, 2015
Advances inEnvironmental Biology, 9(24) November 2015, Pages: 389-396
Test for Terpenoids:
Five milliliters extract of plant sample was added with 2 mL CHCl3 in a test tube. Then, 3 mL of H2SO4
was added carefully to the mixture to form a layer. The formation of the reddish brown of the interface was
observed for the presence of terpenoids.
Test for Cardiac Glycosides:
One milliliter of concentrated H2SO4 was prepared in a test tube. Five milliliters of extract from the plant
sample was mixed with 2 mL of glacial HCH3CO2 containing one drop of FeCl3. The mixture was added
carefully to 1 mL of concentrated H2SO4 so that the concentrated H2SO4was underneath the mixture. The
appearance of brown ring indicates the presence of cardiac glycosides.
Test for Alkaloids:
Five milliliters of the extract was prepared in a beaker and 200 mL of 10% HCH3CO2 in C2H5OH was
added. The mixture was filtered and the extract was allowed to become concentrated in a water bath until it
reached one fourth of the original volume. Concentrated NH4OH was added. Formation of the white precipitate
or turbidity indicates the presence of alkaloids (Trease and Evans,1983).
Bioassay of the antibacterial activity of bamboo leaf extracts:
Preparation of Test Organisms:
Pure culture of Escherichia coli ATCC 25992 and Staphylococcus aureus ATCC 29213 were obtained from
the Department of Biological Sciences in CLSU, Science City of Muñoz, Nueva Ecija. Bacterial pathogens were
grown in Nutrient broth for 24 hours and were standardized to 1.5x108 cells /ml using 0.5McFarland standards.
Antibacterial activity of the bamboo leaf extracts were evaluated using the disc diffusion method at 12 and 24
hours of incubation.
Eradicant Test:
The paper discs were soaked in the leaf extracts. Then sterile plates were poured with 0.1ml of the bacterial
suspension. The discs were then soaked in different treatments were seeded equidistantly in the plates. And the
plates were incubated within 24 hours at room temperature. Zone of inhibition were measured at 12 and 24
hours of incubation.
Protectant test:
The paper discs were soaked in the bacterial suspension. The plates were then poured with 0.1 ml of the leaf
extracts. The paper discs were seeded equidistantly in the plates together with the control. The plates were
stored at room temperature and zones of bacterial colonization were measured after 12 and 24 hours of
incubation.
Antifungal activity of bamboo leaf extracts:
Pure culture of Aspegillus niger and Fusarium semitectum were grown in Potato Dextrose Agar for three
days and were standardized to 1.5x108 spores ml. The paper discs were soaked with fungal suspension. Then the
plates were poured with 0.1 ml of the bamboo leaf extracts. Then the paper discs were seeded equidistantly in
the plates together with the control. The plates were stored at room temperature and thee zones of mycelial
colonization were measured after 24 and 48 hours of incubation.
RESULTS AND DISCUSSION
Survey of Different Species of Bamboo in Seven Municipalities of Nueva Ecija:
As presented in Table 1, 10 species of bamboo were found and identified in seven Municipalities of
Nueva Ecija, Philippines. These include six species of bamboo under genus Bambusa (B. bluemeana, B.
blumeana var Luzonensis, B. vulgaris, B. multiplex var Riviereorum, B. vulgaris var striata and B. tuldoides), 3
species of genus Schizostachyum (S. brachycladum, S. lumampao) and Dendrocalamus asper. Among the
species of bamboo grown in Nueva Ecija, only B. vulgaris var Luzonenesis and S. lumampao are native or
endemic in the Philippines. Meanwhile, S. lumampao, B. bluemeana, B. blumeana var Luzonensis and B.
vulgaris were included in the 12 priority species based on their potentials for development in the Philippines [4].
In Nueva Ecija, bamboo are being utilized by the locals in building houses, weaving baskets, furniture
making, as fences, food and as ornamentals. But none utilized bamboo as remedies on different diseases.
392
Mary Jhane G. Valentino et al, 2015
Advances inEnvironmental Biology, 9(24) November 2015, Pages: 389-396
Table 1: Different species of bamboo in seven Municipalities of Nueva Ecija, Philippines
Local name
Scientific Name
Kawayang Tinik
Bambusa blumeana J.A. & J.H. Schultes
Kawayang Bayog
Bambusa blumeana var Luzonensis
Green Buho
Buho
Schizostachyum brachycladum Kurz
Schizostachyum lumampao
Kawayang Kiling
Bambusa vulgaris Schrad ex wendl.
Chinese Bamboo
Giant Bamboo
Golden Bamboo
Yellow Buho
Budhha Belly
Bambusa multiplex var. Riviereorum (R. Mire) Chia & Fung
Dendrocalamus asper
Bambusa vulgaris var striata Lodd ex Lindl
Schizostachyum brachycladum Kurz
Bambusa tuldoides Munro
Uses
Build houses, weaving basket, furniture
making
Build houses, weaving basket, furniture
making, food
Ornamental, fences
Ornamental, fences
Build houses, weaving basket, furniture
making
Ornamental
Build houses
Build houses
Ornamental, fences
Ornamental
Phytochemical Composition of Bamboo leaf Extracts:
The acetone, ethanol and hot water extracts of B. blumeana, B. blumeana var Luzonensis and B. vulgaris
were individually subjected to phytochemical analyses to establish the presence or absence of flavonoids,
saponins, tannins, steroids, terpenoids, alkaloids and cardiac glycosides using the prescribed techniques [12].
Results of phytochemical analyses are presented in the Table 2. All acetone and ethanol extracts of B. blumeana,
B. blumeana var Luzonensis and B. vulgaris indicated the presence of tannins, steroids, saponins, terpenoids
and cardiac glycosides. Meanwhile, tannins is absent in B. blumeana and B. blumeana var Luzonensis hot water
extracts. Steroids were also absent in all hot water extracts of the three species of bamboo. Terpenoids is absent
in B. blumeana hot water extracts while alkaloids was not detected in any of the tested bamboo leaf extracts.
Table 2: Phytochemical compostion of the different extracts of three species of bamboo
PHYTOCHEMICAL COMPOSITION
BAMBOO LEAF EXTRACTS
Tannins
Steroids
Saponins
Terpenoids
B. blumeana acetone extract
+
B.blumeana ethanol extract
+
B. blumeana hot water extract
B. blumeana var Luzonensis acetone extract
+
B. blumeana varLuzonensis ethanol extract
+
B. blumeana var Luzonensis hot water extract
B. vulgaris acetone extract
+
B. vulgaris ethanol extract
+
B. vulgaris hot water extract
+
(+) = presence of phytochemical; (-) = absence of phytochemical
+
+
+
+
+
+
-
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
Alkaloids
-
Cardiac
glycosides
+
+
+
+
+
+
+
+
+
Flavonoids, lactones and certain phenolic acids are the main chemical constituents of the leaves of most
bamboo leaves [13]. Similarly, Nazreen et al. [14], Zhang et al. [15] and Zhang et al. [16] revealed that extract
of bamboo leaf mainly contains flavone glycosides, phenolic acids, coumarin lactones, flavanoids,
polysaccharides and peptides. In addition, phytochemical analysis of the aqueous leaf extract of Bambusa
vulgaris revealed the presence of alkaloids, tannins, phenolics, glycosides, saponins, flavonoids and
anthraquinones [17].
Antimicrobial Activity of Bamboo Leaf Extracts Against Bacterial and Fungal Pathogens:
1. Antibacterial activity of Bamboo leaf extracts:
In-vitro antibacterial activity of acetone, ethanol and hot water extracts of B. blumeana, B. vulgaris and B.
blumeana var Luzonensis were tested using the protectant and eradicant test.
1.1. Protectant Test:
Protectants or prophylactics are substances which are being supplemented before the prognosis of
symptoms of disease brought about by the pathogenic microorganisms. Table 3 presents the zones of
colonization of E.coli and S. aureus in different bamboo leaf extracts. At 12 hrs of incubation, the smallest zone
of colonization of E.coli was observed in plates treated with B. blumeana var. Luzonensis ethanol and hot
water extracts both with 7.0 mm, while the widest zone of colonization of 14.5mm was observed in B. vulgaris
ethanol extract. At 24 hrs of incubation, increased in bacterial colonization of E.coli was observed in all plates
treated with bamboo leaf extracts. Meanwhile, smallest zone of colonization was obtained in B. blumeana var.
Luzonensis hot water extract with 8.28mm, followed by B. blumeana var Luzonensis (8.33mm) and B. vulgaris
(8.72mm). While B. vulgaris acetone extract had the biggest zone of colonization with 32.09 mm. Statistical
analysis at 12 hrs of incubation showed that the zone of colonization of E.coli in all acetone, ethanol and hot
water extracts of B. blumeana, B.blumeana var Luzonensis and B. vulgaris were comparable to strepmycin
393
Mary Jhane G. Valentino et al, 2015
Advances inEnvironmental Biology, 9(24) November 2015, Pages: 389-396
sulphate (wherein no zone of bacterial colonization was measured). They are also significantly smaller than the
zones of bacterial colonization of E.coli in negative control with 46.9mm. This greatly suggests the antibacterial
activity of all bamboo extracts as protectant against E.coli at 12 hrs and 24 hrs of incubation.
For the protectant test against S. aureus, results showed no zones of bacterial colonization in all plates
treated with acetone, ethanol and hot water extracts of B. blumeana, B. blumeana var Luzonensis and B.
vulgaris at 12 hrs of incubation. While at 24 hrs of incubation, zone of bacterial colonization of S. aureus were
observed in all plates treated with bamboo extracts except for plates with B. blumeana var Luzonensis hot
water extract. Plates treated with B. blumeana var Luzonensis acetone extract had the least zone of colonization
with 6.6mm while the highest zone of colonization was observed in B. vulgaris hot water extract with 18.3mm.
Statistical analysis revealed that zones of colonization of S. aureus produced in all bamboo extracts were
comparable to streptomycin sulphate and significantly smaller than the negative control (29.8mm). Thus, this
depicts the potential protectant activity of all the tested bamboo extracts against S. aureus at 12 and 24hrs of
incubation.
Table 3: Mean zone of colonization of E. coli in different extracts of bamboo
ZONE OF BACTERIAL COLONIZATION
TREATMENTS
E.coli
S. aureus
12 Hrs
24 Hrs
12 Hrs
24 Hrs
B. blumeana acetone extract
10.5b
12.2cd
6.0b
16.9bc
B.blumeana ethanol extract
7.4b
10.8cd
6.0b
7.8cd
b
cd
b
B. blumeana hot water extract
7.2
9.7
6.0
7.6cd
B. blumeana var Luzonensis acetone extract
12.6b
16.6c
6.0b
6.6d
B. blumeana varLuzonensis ethanol extract
7.0b
8.3cd
6.0b
11.5bcd
B. blumeana var Luzonensis hot water extract
7.0b
8.3cd
6.0b
6.0d
B. vulgaris acetone extract
14.5b
32.1b
6.0b
9.8bcd
b
cd
b
B. vulgaris ethanol extract
8.3
14.2
6.0
13.0bcd
B. vulgaris hot water extract
7.3b
8.7cd
6.0b
18.3b
Steptomycin sulphate (positive control)
6.0b
6.0d
6.0b
6.0d
Distilled water ( negative control)
33.0a
46.9a
10.1a
29.8a
*Means with the same letter superscript are not significantly different at 5% level of significance by DMRT; 6mm is the size of paper discs
Results of the protectant activity of bamboo extracts against the two bacterial pathogens can be attributed
to their phytochemical constituents. Several researches have indicated that a flavonoid-rich bamboo leaf extract
has multiple biological effects, such as anti-free radical, anti-oxidant, anti-aging, anti-fatigue, fungicidal,
bacteriocidal, anti-viral, and prevention of cardiovascular diseases, and can be used as a pharmaceutical interme
diate, dietary supplement, cosmetic ingredient, and food additive[18]. Also, Meng et al. [19], reported the anti
microbial activity of various medicinal plants containing phenol and flavonoids. Results also coincides with the
findings of Zhang et al. [20] which demonstrated that the phenolic compounds, antraquinones and coumarins of
the bamboo leaves are all of relatively strong antibacterial and bactericidal functionalities. It is also in agreement
with the findings of Lu et al. [21] and Zhang et al. [22], wherein various extracts made from different parts of
Bambusa vulgaris was found to have multiple biological activities which protect mankind from different
diseases.
In addition, as shown in the results of the study, bamboo extracts are more effective as protectant against S.
aureus than E. coli as it shows minimal to no zone of bacterial colonization of S. aureus at 12 and 24 hrs of
incubation. This is due to the absence of outer membrane in Gram-positive bacteria which is the S. aureus and
that its cell can be easily penetrated by bioactive compounds present in the extracts. In most cases, plant extracts
are much active against gram-positive bacteria than gram-negative bacteria [23]. In addition, Demetzos et al.
[24] investigated that antimicrobial activity of myricetin and its derivatives and the result revealed that they
could inhibit the Gram-positive bacteria compared to Gram-negative bacteria. Antibacterial activity against
Staphylococcus aureus is of great importance since it is an important attribute of skin cosmetics and the
proliferation of bacteria causes skin problems such as acne, comedo, papules, cellulitis and allergies [19, 25].
1.2. Eradicant Test:
Eradicant test was done to determine the ability of the different extracts of selected bamboo species in
inhibiting the growth of E. coli ATCC 25992 and S. aureus ATCC 29213 after 12 and 24 hours of incubation
using the disc diffusion method.
As presented in Table 4, streptomycin sulphate registered the highest zone of inhibition of 29.3mm and
30.2mm against E. coli at 12 hrs and 24 hrs of incubation. Among the bamboo leaf extract tested, B. blumeana
var Luzonensis ethanol extracts had the highest zone of bacterial inhibition in all incubation period of 10.8 mm
and 9.6 mm. Meanwhile, at 12hrs of incubation, the least zone of inhibition was recorded in B. vulgaris acetone
extract (7.4 mm) while B. blumeana var Luzonensis acetone extract of 6.5mm at 24 hrs. Meanwhile, for the
eradicant test against S. aureus, streptomycin sulphate produced 32.1mm and 27.6 mm at 12 and 24 hrs of
incubation, followed by B. blumeana var. Luzonensis ethanol extract of 12.8mm while B. blumeana var.
394
Mary Jhane G. Valentino et al, 2015
Advances inEnvironmental Biology, 9(24) November 2015, Pages: 389-396
Luzonensis hot water extract had the least with 7.1mm at 12 hrs. At 24 hrs of incubation, among the tested
bamboo extracts, B. vulgaris hot water extract (10.7 mm) registered the highest zone of inhibition while B.
vulgaris acetone extract attained the least zone of 6.9 mm. Though zones of inhibition were measured in all
bamboo leaf extracts tested, statistical analysis revealed no significant differences among all the bamboo leaf
extracts and the negative control (6mm). This indicates the lack of eradicant activity of the aforementioned
bamboo extracts against E.coli and S. aureus at 12 and 24 hrs of incubation.
Table 4: Mean zone of bacterial inhibition of different bamboo extracts against E. coli and S. aureus
ZONE OF INHIBITION
TREATMENTS
E.coli
S. aureus
12 Hours
24 Hours
12 Hours
24 Hours
B. blumeana acetone extract
6.9c
7.2bc
10.9b
9.3b
B.blumeana ethanol extract
8.6bc
7.6bc
8.9b
7.2b
B. blumeana hot water extract
7.7bc
7.4bc
8.6b
7.4b
B. blumeana var Luzonensis acetone extract
6.5c
6.6c
10.3b
7.0b
B. blumeana varLuzonensis ethanol extract
10.8b
9.8b
12.8b
9.3b
B. blumeana var Luzonensis hot water extract
7.4c
7.3bc
7.1b
7.3b
c
bc
b
B. vulgaris acetone extract
7.3
7.5
7.7
6.9b
B. vulgaris ethanol extract
7.5c
7.5bc
9.1b
7.8b
bc
bc
b
B. vulgaris hot water extract
7.7
7.2
10.2
10.7b
Streptomycin Sulphate
29.3a
30.2a
32.1a
27.6a
Distilled Water
6.0c
6.0c
6.0b
6.0b
*Means with the same letter superscript are not significantly different at 5% level of significance by DMRT; 6mm is the size of paper discs
Minimal zones of bacterial inhibition produced by the bamboo leaf extracts imply their lack of bactericidal
activity as eradicant against the two bacterial pathogens. This can be due to the absence of alkaloids in all the
tested extracts. Alkaloids are toxic to microbes that effectively kill bacteria and viruses, as antiprotozoa and
antidiarrheal, are detoxification which can neutralize toxins. They are known to have antimicrobial antifungal
and anti-inflammatory effects and also act as anti-hypertensive agents. Alkaloids have been liked to specific
actions against infections and diseases [5, 26, 27].
Anti-Fungal Activity of Different Bamboo Leaf Extracts:
Aspergillus niger and Fusarium semitectum were used as test pathogens for the fungicidal activity as
protectant of bamboo. Apparently, Aspergillus niger produce mycotoxins that can cause crown rot of can
induce a crown rot of fruits and vegetables, and deformities and curling of several plants [28]. Similarly, F.
semitectum causes several diseases such as canker of walnut, pod rot, seed rot and root rot of beans and storage
rot of banana [29,30]. Moreover, A. niger and F. semitectum are also regarded as human pathogens causing
respiratory and skin diseases.
As shown in Table 4, no zones of mycelial colonization of F. semitectum was observed in plates treated
with different bamboo extracts while fungal growth was recorded in plates treated with distilled water of
33.0mm zone of colonization at 12 hrs of incubation. After 48 hrs of incubation, mycelial growth was observed
in all plates treated with bamboo extracts. In addition, B. blumeana acetone extracts had the least zone of
colonization of 11.15mm while B. blumeana var Luzonensis of 18.80mm registered the highest zone of fungal
colonization. Statistical analysis revealed that all treatments were significantly different from the negative
control at 12 hrs of incubation. While at 48 hrs of incubation, treatments were significant lower than the
negative control however it is also not comparable with the positive control. This suggests the protectant
potential of bamboo extracts at 12 hrs of incubation while reduced efficacy was observed at 48hrs of incubation.
Results of the fungicidal activity against A. niger showed no zone of fungal colonization in all plates treated
with bamboo leaf extracts except for B. vulgaris acetone extracts of 8.67 mm. At 48 hrs of incubation, rapid
mycelia growth was observed colonizing the entire plates. Statistical analysis revealed that at 12 hrs of
incubation no significant difference between the positive control and the tested bamboo extracts. This implies
the protectant potentials of bamboo leaf extracts only at 12hrs of incubation.
Table 5: Mean zones of fungal colonization of F. semitectum and A. niger
ZONE OF FUNGAL COLONIZATION
TREATMENTS
F. semitectum
24 Hours
48 Hours
B. blumeana acetone extract
6.0b
11.15e
B.blumeana ethanol extract
6.0b
13.22de
B. blumeana hot water extract
6.0b
14.79cd
B. blumeana var Luzonensis acetone extract
6.0b
17.46abc
b
B. blumeana varLuzonensis ethanol extract
6.0
18. 80ab
B. blumeana var Luzonensis hot water extract
6.0b
16.18bc
b
B. vulgaris acetone extract
6.0
12.63de
B. vulgaris ethanol extract
6.0b
13.22de
A. niger
24 Hours
6.0c
6.0c
6.0c
6.0c
6.0c
6.0c
8.67b
6.0c
48 Hours
90.0b
90.0b
90.0b
90.0b
90.0b
90.0b
90.0b
90.0b
395
Mary Jhane G. Valentino et al, 2015
Advances inEnvironmental Biology, 9(24) November 2015, Pages: 389-396
B. vulgaris hot water extract
6.0b
12.57de
6.0c
90.0b
Benlate solution
6.0b
6.00f
6.0c
6.0a
Distilled Water)
33.0a
19.98a
12.48a
90.0b
*Means with the same letter superscript are not significantly different at 5% level of significance by DMRT; 6mm is the size of paper discs
Fungicidal activity of certain plants are can attributed to the action of flavonoids, terpenoids and tannins in
bamboo extracts tested. In addition, bamboo is a rich source of phenols and flavonoids which have an
antimicrobial activity. Flavonoids found in bamboo extracts are flavonides, such as orientin, homoorientin,
vitexin and Isovitexin, 3- bonded oxygen flavonol glycosides such as 3- bonded pilocarpine glycosylation and 6bonded glucose [31, 32].
Conclusion:
Based on the results of the study, it can be concluded that ethanol, acetone and hot water extracts of B.
blumeana, B. blumeana var Luzonensis and B. vulgaris contains possess phytochemicals such as tannins,
steroids, saponins, flavonoids, terpenoids and cardiac glycosides. In addition, bamboo leaf extracts showed
protectant activity against E. coli and S.aureus at 12 to 24 hrs of incubation while their effectivity against A.
niger and F. semitectum was only observed at 12 hrs of incubation.
ACKNOWLEDGMENT
To God be the Glory. Thy will be done!
REFERENCES
[1] Dannenmann, B.M.E., C. Choocharoen, W. Spreer, M. Nagle, H. Leis, A. Neef and J. Mueller, 2007. The
potential of bamboo as a source of renewable energy in Northern Laos. Conference on International
Agricultural Research for Development, University of Kassel-Witzenhausen and University of Gottingen.
[2] Goyal, A.K. and B.K. Brahma, 2014. Antioxidant and nutraceutical potential of bamboo: An overview.
International Journal of Fundamanetal and Applied Sciences, (3): 2-10.
[3] Lantican, C., A. Palijon and C. Saludo, 1985. Bamboo research in Philippines. College of Forestry,
University of the Philippines at Los Banos,College, Laguna, Philippines.
[4] Virtucio, F.D. and C.A. Roxas, 2003. Bamboo production in the Philippines. Department of Environment
and Natural Resources, Ecosystem Research and Development Bureau.
[5] Okwu, D.E. and M.E. Okwu, 2004. Chemical composition of spondia mombin plants. Journal of
Sustainable Agriculture and Environement, 6: 140-147.
[6] Liu, R.A., 2004. Potential synergy of phytochemicals in cancer prevention: mechanism of action. Journal of
Nutrition, 134(12): 3479s-3485s.
[7] Roa, A.V. and L.G. Rao, 2007. Carotenoids and human health. Pharmacological Research, 55(3): 207-16.
[8] Barjesteh van Waalwijk van Doorn-khosrovani, S., J. Janssen, I.M. Maas, R.W. Godschalk, J.G. Nijhuis,
F.J. Van Schooten, 2007. Dietary flavonoids induce MLL translocations in primary human CD34+ cells.
Carcinogenesis, 28(8): 1703-1709.
[9] Wang, J., Y.D. Yue, F. Tang and J. Sun, 2012. TLC Screening for Antioxidant Activity of Extracts from
Fifteen Bamboo Species and Identification of Antioxidant Flavone Glycosides from Leaves of Bambusa
textilis McClure. Molecules, 17: 12297-12311.
[10] Singh, V.K., R. Shukla, V. Satish, S. Kumar, S. Guptas and A. Mishra, 2010.Antibacterial activity of leaves
of bamboo. International Journal of Pharma and Bio Sciences, 4(2).
[11] Lu, B., J. Chen, W. Huang, D. Wu, W. Xu, Q. Xie, X. Yu and L. Li, 2011. Determination of flavonoids and
phenolic acids in the extract of Bamboo leaves using near-infrared spectroscopy and multivariate
calibration. African Journal of Biotechnology, 10(42): 8448-8455.
[12] AOAC. Official Methods of Analysis, 1990. 15th edition. Association of Official Analytical Chemists,
Washington DC, USA.
[13] Zhang, Y., Y. Zhang and T. Ying, 2008. Reduction of acrylamide and its kinetics by addition of
antioxidant of Bamboo leaves (AOB) and extract of green tea (EGT) in asparagines - glucose microwave
heating system. Journal of Food Science, 73 (2).
[14] Nazreen, S., M.S. Alam, H. Hamid, G. Kaur, M.M. Alam, S. Haider and S. Shafi, 2011. Phytochemical
investigation of Bambusa arundinacea Retz.Internationl Journal of Natural Product Science, 3: 1-7.
[15] Zhang, Y., X.Q. Wu and Z.Y. Yu, 2002. Comparisions of total content of flavonoids in leaves of bamboo
and ginkgo and their free-radical resistance. Chinese Journal of Chinese Medicine, 27(4): 254-257.
[16] Zhang, J., J. Gong, Y. Ding, B, Lu, X. Wu and Y. Zhang, 2010. Antibacterial activity of water-phase
extracts from bamboo shavings against food spoilage microorganisms. African Journal of Biotechnology,
9(45): 7710-7717.
396
Mary Jhane G. Valentino et al, 2015
Advances inEnvironmental Biology, 9(24) November 2015, Pages: 389-396
[17] Yakubu, M.T. and B.B. Bukoye, 2009. Abortifacient potentials of the aqueous extract of Bambusa vulgaris
leaves in pregnant Dutch rabbits. Contraception., 80(3): 308-313.
[18] Soni, V., A.K. Jha and J. Dewedi, 2012. A Review on Ethanomedical Uses, Phytochemicals and
Pharmacological Profile of Bambusa arudinacea Retz: Indian Journal of Novel Drug delivery, 4(4): 264271.
[19] Meng, T.X., S. Furuta, S. Fukamizu, R. Yamamoto, H. Ishikawa, E.T. Arung, K. Shimizu and R. Kondo,
2011. Evaluation of biological activities of extracts from the fruiting body of Pleurotus citrinopileatus for
skin cosmetics. Journal of Wood Science, 57: 452-458.
[20] Zhang, Y.X.B. Yao, B.l. Bao and Y. Zhang, 2006. Anti-fatigue activity of a triterpenoid-rich extract from
Chinese bamboo shavings (Caulis bamfusae in taeniam). Phytotherapy Research, 20: 872-876.
[21] Lu, B.Y., X.Q. Wu, X.W. Tie and Y. Zhang, 2005. Toxicology and safety of antioxidant of Bamboo leaves
part 1: Acute and subchronic toxicity studies on antioxidant of bamboo leaves. Food and Chemical
Toxicology, 43: 783-792.
[22] Zhang, Y., X.W. Tie, B.L. Bao, X.Q. Wu and Y. Zhang, 2007. Metabolism of flavones C-glycosides and
p-coumaric acid from antioxidant of bamboo leaves (AOB) in rats. British Journal of Nutrition, 97: 484494.
[23] Vlientinck, A.J., J. Totte, A. Lasure, B.D. Vanden, P.C. Rwangabo and J. Mvukiyuwami, 1995. Screening
of Hundred Rwandese medicinal plants for antimicrobial and antiviral properties. Journal of
Ethnopharmacology, 46: 31-47.
[24] Demetzos, C., D. Angelopoulou, A. Kolocouris, I. Daliani and T. Mavromoustakos, 2001. Structure
elucidation, conformational analysis and thermal effects on membrane bilayers of an antimicrobial
myricetin ether derivative. Journal of Heterocyclic Chemistry, 38: 703-710.
[25] Breuer, K., S. Haussler, A. Kapp and T. Werfel, 2002. Staphylococcus aureus: Colonizing features and
influence of an antibacterial treatment in adults with atopic dermatitis. British Journal of Dermatology, 147:
55- 61.
[26] Sofowora, A., 1993. Medicinal plants and traditional medicine in Africa. Spectrum books Ltd. Ibadan,
Nigerian: 289.
[27] Supriyatin, Rahayu S. and D. Sukmawati, 2014. Phytchemical analysis of Manggong bamboo leaf extracts.
Proceeding of International Conference On Research, Implementation And Education. B-165.
[28] Sharma, R., 2012. Pathogenecity of Aspergillus niger in plants. Cibtech Journal of Microbiology, 1(1): 4751.
[29] Zaccardelli, M., V. Balmas, C. Altomare, L. Corazza and C. Scotti, 2006. Characterization of Italian
isolates of Fusarium semitectum from alfalfa (Medicago sativa) by AFLP analysis, morphology,
pathogenicity and toxin production. Journal of Phytopathology, 154: 454-460.
[30] Latiffah, Z., M.S. Nurul Huda and T.M.A. Tengku Ahmad Akram, 2013. Characterization of Fusarium
semitectum from isolates vegetable fruits. Sains Malaysiana, 42(5): 629-633.
[31] Xiao, Y. Zhang, Z.T. and J.D. Hou, 2001. Analyses the flavonoids in the leaves of Pleioblastus amarus
using liquid chromatography-mass spectrography. Journal of Ningbo Technical College., 13(3): 123-126.
[32] Zhang, S.X., F.H. Liu and X.Z. Sun, 2002. Isolation of effective components from bamboo leaves. Journal
of Sichuan University (Enginerring Sciences). 34(6): 116-118.