This research article investigates the bioactivity of the ethanolic leaf extract of Boehmeria macrophylla (BMET). Phytochemical screening revealed the presence of various compounds including alkaloids, tannins, and flavonoids in BMET. In vitro assays found the extract exhibited antioxidant, analgesic, cytotoxic, and antibacterial activities. Specifically, BMET showed free radical scavenging activity in the DPPH assay with an IC50 value of 39.76 μg/mL, comparable to ascorbic acid. In mice, BMET demonstrated 32.91% and 54.2% reductions in acetic acid-induced writhing at doses of 250 and 500 mg/kg, respectively. In br

1. RESEARCH ARTICLE
Bioactivity of Boehmeria Macrophylla (Urticaceae) leaf extract
Amir Hossain1
& Fahrima Islam1
& Md. Saifuzzaman1
& Md. Afjalus Siraj Saeed1
&
Md. Khirul Islam1
& Gazi Md. Monjur Murshid1
& Md. Mustafizur Rahman1
Received: 27 March 2016 /Accepted: 1 June 2016 /Published online: 30 June 2016
# Institute of Korean Medicine, Kyung Hee University and Springer Science+Business Media Dordrecht 2016
Abstract Boehmeria macrophylla is a perennial herb used as
folk medicine in Bangladesh for the healing of pathologic
conditions like fever, trauma, dermatitis, boils etc. After phy-
tochemical screening of ethanolic extract of B. macrophylla
(Urticaceae) leaf (BMET), in vitro antioxidant activity was
investigated by free radical (DPPH) scavenging assay. Total
tannin, phenolic and flavonoid contents were also estimated.
Analgesic, cytotoxic and antibacterial activities were evaluat-
ed by ‘acetic acid induced writhing inhibition in mice’, ‘brine
shrimp lethality bioassay’ and ‘disk diffusion assay’, respec-
tively. Phytochemical screening indicated the presence of al-
kaloids, tannins, flavonoids, saponins, steroids and terpenoids.
The extract showed scavenging activity with IC50 value of
39.76 μg/mL whereas IC50 was 6.86 μg/mL for ascorbic acid.
The extract also showed the total phenolic, flavonoid and tan-
nin content as 439.48, 356.84 and 395.42 mg QE/100 g of
dried plant extract. BMETshowed 32.91 % and 54.2 % writh-
ing inhibition in mice at doses 250 and 500 mg/kg, respective-
ly. BMET showed LC50 against brine shrimp nauplii at
33.16 μg/mL. The extract also showed antibacterial activity
against Vibrio cholerae, Escherichia coli, Staphylococcus
epidermidis and Streptococcus ferus. The results suggest that
BMET has antioxidant, analgesic, cytotoxic and antibacterial
activities.
Keywords Boehmeria macrophylla . Antioxidant .
Analgesic . Cytotoxic . Antibacterial . Mice
Introduction
Numerous disorders are shown to be healed by medicinal plants
(Das et al. 2011). For the management of infectious diseases,
majority of the people in both rural and urban areas rely on these
medicinal plants (Rahman and Junaid 2013). Natural products
have more ‘drug like and biological characteristics’ than fully
synthetic molecules which make them potent therapeutic candi-
dates (Minhajur et al. 2011). B. macrophylla (Urticaceae) is also
known as Boehmeria macrostachya, Boehmeria platyphylla. It is
a perennial shrub that is distributed throughout Bangladesh as
well as other sub-continent countries. Locally it is called Jangli
chotta. It is grown at river beds in the sub-Himalayan tract, edges
of forests, valleys and roadsides. It may grow up to 2.5 m. It is in
flower from June to September and the seeds ripen from Sep-
tember to January. The flowers are dioecious (Fei et al. 2003).
The plant is not self-fertile. B. macrophylla is suitable for sandy,
loamy and well-drained soil. It can grow in semi-shade or no
shade area but preferably in dry or moist soil. A fiber is obtained
from the stem. It is possibly useful for textiles. Wood is moder-
ately hard but too small for much use, it makes a good fuel.
However, there are very few reports on the medicinal use of
B. macrophylla.
It is reported that Chinese people are using B. macrophylla as
herbal medicine for the treatment of several disorders like- trau-
ma, hemorrhoids, fever and other diseases. In addition, 8 com-
pounds were isolated and identified from B. macrophylla which
include β-sitosterol, β-sitosterolglucoside, ursolic acid, hydrox-
yl-6-methoxycoumarin, emodin, cinnamic acid, sweroside, 5, 7-
dihydroxy4’ methoxy dihydroflavonol (Min et al. 2011).
To make public the potentials of medicinal plants accessi-
ble in Bangladesh, we have selected B. macrophylla
(Urticaceae) leaf as the material of medicinal values. As no
reports were found on the ethanol extract of B. macrophylla
(Urticaceae) leaf (BMET), we aimed to investigate the
* Md. Mustafizur Rahman
dipti0103@yahoo.com
1
Pharmacy Discipline, Life Science School, Khulna University,
Khulna -9208, Bangladesh
Orient Pharm Exp Med (2016) 16:233–241
DOI 10.1007/s13596-016-0230-1
Online ISSN 2211-1069
Print ISSN 1598-2386

2. possible antioxidant, analgesic, cytotoxic and antibacterial ac-
tivities of BMET.
Materials and methods
Plant collection and extraction
B. macrophylla leaf was collected from Jamalpur area of
Rajbari district, Bangladesh in November, 2012 and was identi-
fied by experts at Bangladesh National Herbarium, Mirpur, Dha-
ka (voucher specimen no.: DACB 37855). Leaf of the plant was
then properly washed with fresh water to eliminate dirty mate-
rials and was shade dried for few days. Then the dried leaves
were crushed into a coarse powder with the help of a grinding
machine. The plant materials were stored for future at room
temperature. About 450 g of the dried crushed materials were
put in an amber colored glass container. The materials were then
soaked with about 1600 mL ethanol. The contents in the con-
tainer were sealed keeping for a period of about 8 days undergo-
ing occasional shaking and stirring. Then the entire combination
endured filtration with a piece of white and clean cotton. Then it
was filtered through Whatman No. 1 filter paper. The filtrate
obtained was evaporated under the air of ceiling fan. The yield
of BMET was found 2.05 %.
Chemicals
Sodium carbonate and Folin–Ciocalteus’s reagent were ob-
tained from Merck (Damstadt, Germany). 1, 1-diphenyl-2-
picrylhydrazyl (DPPH), potassium dichromate, gallic acid
(GA), ascorbic acid (AA), AlCl3, H2SO4, Nitric acid, NaOH,
quercetin (QU), DMSO, EDTA, Diclofenac-Na, sodium ni-
trous, tween-80 and FeCl3 were obtained from the Sigma
Chemical Co.
Phytochemical screening
To identify the chemical constituents of the plant extract some
positive control processes were followed. The BMET was
examined qualitatively to categorize chemical components
with the help of the following reagents and some chemicals,
reducing sugar with Fehling’s Solution and Benedict’s re-
agent, alkaloid with Mayer’s and Dragendroff’s reagent, sa-
ponins with distilled water, Glycosides with sodium hydrox-
ide solution, Steroids with H2SO4, Steroids with Sulphuric
acid, Tannin with Ferric Chloride and Potassium dichromate
solution, gum with Molish’s reagent (Ghani 1995).
DPPH free radical scavenging assay
DPPH (2, 2-diphenyl-1-picrylhydrazyl) free radical scaveng-
ing assay described by Choi et al. was used to determine the
antioxidant activity of BMET (Khan et al. 2013). Firstly,
0.004 % (w/v) solution of DPPH in ethanol was prepared.
From this 3 mL solution was mixed with 1 mL crude extract
in ethanol at a different concentration. Subsequently, the mix-
ture was shaken properly and kept it in a dark place at room
temperature for about 30 min. Spectrophotometrically absor-
bance was measured at 517 nm. Percentage scavenging activ-
ity was measured by following equation:
%DPPH free radical scavenging activity ¼ Ao–A1ð Þ=Ao½ Š Â 100
Here, A0 is absorbance of control, and A1 is absorbance of
BMET/positive control. The % scavenging activity was then
plotted against log concentration and a graph for IC50 was
calculated.
Estimation of total phenolics
Folin–Ciocalteu technique described by Wolfe et al. was used
to estimate total phenolic content of BMET (Borah et al.
2011). An aliquot of BMET or positive control was mixed
with 2 mL Folin–Ciocalteu reagent that is previously diluted
10 times with water (1:10 v/v) and 2 mL sodium carbonate
(75 g/l). Tubes were shaken for about 15 s. Then the mixtures
were allowed to stand for 20 min at 25 °C for the purpose of a
color development. At 760 nm absorbance was measured with
the help of a UV-spectrophotometer. At concentration of
0.1 mg/mL BMET or positive control was evaluated. Total
phenolic content was expressed as gallic acid equivalent,
GAE (mg of GA/g of dried plant extract). Then a gallic acid
standard calibration curve was developed where the positive
control curve equation was y = 0.1854x + 0.0412, R2 = 0.9895
[def of x,y & R].
Determination of total flavonoids
Total flavonoid content was determined by the following
Method (Raju et al. 2013). Here, 1 mL of samples or positive
control was added to 4 mL of distilled water and 0.3 mL 5 %
sodium nitrous solution (NaNO2) and it kept for 5 min. At the
6th minute, 2 mL 1 M sodium hydroxide (NaOH) was added.
This preparation was then shaken well and the final volume
was adjusted to 10 mL. After that, the UV absorbance was
measured at 510 nm against blank. Total flavonoid content
was expressed as quercetin equivalent, QE (mg of Q/g of dried
plant extract). The positive control curve equation was
y = 0.158x + 0.2612, R2 = 0.9669.
Determination of total tannins
Determination of total tannin content of the BMET was per-
formed using Folin-Ciocalteu (FC) reagent described by
Marinova et al. (2005). Firstly, 0.1 mL positive control or
234 A. Hossain et al.

3. sample solution was added to 7.5 mL distill water and 0.5 mL
FC reagent with shaking and then kept for 5 min. Then 1 mL
of 35 % sodium carbonate solution was added to it and diluted
with 10 mL distilled water. Then they were kept for 30 min at
room temperature. After 30 min the UVabsorbance was mea-
sured at 725 nm against blank. Total tannin content was
expressed as gallic acid equivalent, GAE (mg of GA/g of dried
plant extract). The positive control curve equation was
y = 0.0177x + 0.0121, R2 = 0.9856.
Acetic acid induced writhing inhibition test
Acetic acid induced writhing method was performed on mice
to evaluate the analgesic activity of the BMET (Ahmed et al.
2004). Some young Swiss-albino mice of any sex (n = 5)
balancing 18–22 g were used. All animals were kept fasted
for 2 h before starting the experiment. Then they were divided
in four groups. Group I was given 1 % tween-80 solution in
water (10 mL/kg) as the control, Group II received pos-
itive control (Diclofenac-Na) at a dose of 25 mg/kg
body weight. The groups III and IV were set with 250
and 500 mg/kg BMET, respectively. After 30 min, the
animals were treated with 0.7 % acetic acid intraperito-
neally. After 5 min of acetic acid injection, the number
of abdominal constrictions (writhing) was calculated for
the period of 15 min.
Brine shrimp lethality bioassay
The cytotoxic activity of BMET was evaluated by Brine-
Shrimp Lethality bioassay as it is described by McLaughlin
et al. (1998). Artemia was used for the activity test. The brine
shrimp eggs were hatched through the artificial sea water that
is 3.8 % NaCl solution for about 24 h to mature the shrimp
(nauplii). 5 mL stock solution was added to a test tube and a
serial dilution was made to make concentrations 320, 160, 80,
40, 20, 10 and 5 μg/mL. 5 mL stock solution of positive
control (vincristine sulfate) was added to a test tube and a
serial dilution was made to provide 5, 2.5, 1.25, 0.625 and
0.325 μg/mL. Ten (10) nauplii were added to each test tubes
and volume was adjusted to 10 mL with sea water. The
concentration of DMSO didn’t exceed 10 μL/mL. After
24 h, the number of alive shrimps was counted with the help
of a 3 times magnifying glass against light. Then the LC50
values were measured from Probit Analysis Chart using
computer software BLdP line software^, (Bauer et al. 1996).
Determination of antibacterial activity
The disc diffusion method was used for the determination of
antibacterial activity of the extract (Fakruddin et al. 2012).
The extract was evaluated against 7 g-positive (Staphylococ-
cus aureus, Staphylococcus epidermidis, Streptococcus ferus,
Streptococcus pyogens, Shigella boydii, Enterococcus
faecalis, Shigella sonnei) and 5 g-negative (Escherichia coli,
Shigella dysenteriae, Proteus species, Shigella flexneri, and
Vibrio cholerae) bacteria. Mueller-Hinton agar media was
used for the preparation of test plates. Filter paper discs
(Whatman No. 1) of 5 mm diameter were loaded with two
concentrations of crude leaf extracts (25 μg/μL and 50 μg/
μL). Discs were then completely dried. A 100 μL of culture
was spread on sterilized agar media and the impregnated discs
were placed on it and incubated for 24 h at 37 °C.
Ciprofloxacin discs (5 μg/disc) were used as positive
control. The diameter of zone of inhibition in mm was
recorded after the incubation period. A sterilized blank
disc was used as control.
Statistical analysis
All analyses were passed in two replications. Data were pre-
sented as mean ± SEM. All experimental parameters were
evaluated for their significance level by correlation and regres-
sion analysis, the t-tests (P < 0.05) was used. A free R-
software version: 2.15.1 (http://www.r-project.org/) and
Microsoft Excel 2010 were used for both statistical and
graphical evaluations.
Results
Phytochemical screening
Phytochemical screening of the BMET leaf extract showed
the presence of alkaloids, tannins, flavonoids, saponins, ste-
roids and terpenoids which are summarized in Table 1.
Table 1 Result of
phytochemical screening
of ethanol leaves extract
of BMET
Chemical groups BMET
Reducing sugar -
Combined reducing sugar -
Tannins +
Flavonoids +
Saponin +
Gums -
Steroids +
Alkaloids +
Glycoside -
Proteins -
Terpenoid +
Acidic compounds -
BMET denotes for ethanolic extracts of
BMET; (+) = present; (−) = absent
Bioactivity of Boehmeria Macrophylla (Urticaceae) Leaf Extract 235

4. DPPH free radical scavenging activity
Antioxidant activity of BMET was determined by DPPH free
radical scavenging method. The IC50 value for free radical
scavenging activity was measured and compared with stan-
dard ascorbic acid (Fig. 1). IC50 value for BMET leaf extract
was found to be 39.76 μg/mL whereas for standard ascorbic
acid it was 6.86 μg/mL.
Total phenol, tannin and flavonoid content
The total phenolic, total tannin and total flavonoid contents of
BMETwere determined and expressed as gallic acid and quer-
cetin equivalents (Fig. 2). Total phenolic and total tannin con-
tent were 439.48 mg GAE/100 g and 395.42 mg GAE/100 g
of BMET leaf extract where the total flavonoid content was
356.84 mg QE/100 g of dried plant extract as shown in
Table 2.
Analgesic activity
The result of analgesic activity for BMET exhibits a
significant inhibition of writhing impulse by 32.91 %
and 54.2 % at 250 mg/kg and 500 mg/kg doses, respec-
tively. The positive control (Diclofenac Na) showed
76.13 % writhing inhibition at a dose of 25 mg/kg body
weight (Table 3).
Brine shrimp lethality bioassay
In the brine shrimp lethality bioassay, the percent mortality of
the nauplii caused by the leaf extract as well as the positive
control Vincristine sulfate is represented in Fig. 3. The LD50
of both sample and positive control was calculated by Probit
analysis software (LdP Line software, USA) and was found to
be 33.16 μg/mL for BMET whereas the positive control Vin-
cristine sulfate showed 0.72 μg/mL (Table 4).
Antibacterial assay
BMETshowed antibacterial activity against four bacterial species
(Vibrio cholerae, Staphylococcus epidermidis, Escherichia coli
and Streptococcus ferus) out of twelve bacterial strains used in
disk diffusion assay (Table 5). The extract showed 27, 14, 11.5
and 10 mm zone of inhibition against Vibrio cholerae, Staphylo-
coccus epidermidis, Escherichia coli and Streptococcus ferus,
respectively at a dose of 500 μg. However, at 250 μg dose the
zone of inhibition against the corresponding microorganisms
were found to be 20, 7.5, 7.5 and 7 mm respectively. On the
other hand, diclofenac sodium showed 36.5, 25, 36 and 26 mm
of zone of inhibition at 5 μg dose.
Discussion
Plants produce secondary compounds as their natural defense
(Sahu and Padhy 2013). Among these secondary compounds,
few are toxic for animals, but the others are not. Certainly,
majority of those compounds have been utilized as whole
plants or crude extracts for the purpose of medical ap-
plications in human being, (Lucy and DaSilva 1999).
Plants are also natural sources of different therapeutic
activities, (Raja et al. 2013). These crude medicines
are gaining much more popularities for healthcare pur-
pose, (Joshi et al. 2011). Compounds like glycosides
and flavonoids have been reported to possess antidiabet-
ic and antioxidant activity, (Katsube et al. 2010). Phy-
tochemical study revealed that BMET contains tannins,
flavonoids, alkaloids, saponins, terpenoids and steroids
(but lacks reducing sugar, combined reducing sugar and
gums) (Table 1). Different studies proposed that various
sorts of polyphenolic compounds like phenolic acids,
flavonoids and tannins have copious biological effects
including antioxidant activity (Prakash et al. 2009).
The BMET showed a good amount of total phenolic,
total tannin and total flavonoid contents (table 2) that
is comparable to the standards (Fig. 2). Many diseases
occur as the result of oxidative stress and antioxidants
are the mean of benefits for treating those diseases
(Nascimento et al. 2013). In the present study the ex-
tract of BMET showed antioxidant activity by scaveng-
ing DPPH (Fig. 1). However, there are reports that an-
tioxidants like tea, phenolics can directly inhibit cancer
cells (Ewan 2005). Plant’s antioxidant activity is due to
the presence of phenolic components such as flavonoids,
tannins (Morshed et al. 2011). There is an established
positive relationship between phenolic content and anti-
oxidant activity because of the scavenging ability of
0
50
100
0 0.5 1 1.5 2 2.5 3
%ScavengingacƟvity
Log concentraƟon (μg/ml)
B macrophylla
Ascorbic acid
Fig. 1 DPPH scavenging activity of BMET and positive control
(Ascorbic acid)
236 A. Hossain et al.

5. hydroxyl groups (Wang et al. 2007). It is also reported
that antioxidants are effective hydrogen donor or elec-
tron acceptor (Vinson et al. 2011). Moreover, tannins
are generally defined as naturally occurring polypheno-
lic compounds of high molecular weight to form com-
plexes with the proteins. Tannins are important source
of protein in animals but unfortunately the amounts of
tannins that they contain vary widely and largely unpre-
dictable and their effects on animals range from benefi-
cial to toxicity and death (Yang and Russell 1992).
Consumption of foods containing antioxidant and anal-
gesic activity prevents health risks (Hossain et al. 2012).
The extract was evaluated for analgesic activity by acetic
acid induced writhing in Swiss albino mice. Intraperitoneal
administration of acetic acid induces pain sensation (Barua
et al. 2011) and produces writhing reflex in animals by trig-
gering the chemo-sensitive nociceptors (Onasanwo and
Elegbe 2006). Acetic acid, used to induce writhing, causes
algesia by liberation of endogenous substances, which in turn
excite the pain nerve endings (Taesotikul et al. 2003). Acetic
acid-induced writhing model represents pain sensation
by triggering localized inflammatory response. Also, it
is found that in case of acetic acid-induced models the
analgesia level is indicated by the percent reduction in
the number of abdominal constrictions (Machioro et al.
2005). It has been reported that flavonoids show anal-
gesic action by enhancing the endogenous serotonin lev-
el or interact with 5-HT2A and 5-HT3 receptors
(Annegowda et al. 2010). The extract produced signifi-
cant writhing inhibition comparable to the standard drug
Diclofenac-Na (Table 3). The polar compounds present
in the plant extract may be responsible for the obtained
analgesic activity. The results showed a higher inhibi-
tion at 500 mg/kg dose and the inhibitory effect of
positive control was greater and the inhibitory effects
were statistically significant (control vs. BMET 250 mg/
kg, P < 0.01 and control vs. BMET 500 mg/kg, P < 0.05).
The results demonstrated that BMET at the given doses
significantly reduced the acetic acid induced writhing re-
flex in mice.
The cytotoxic activity of the BMET was tested by brine
shrimp lethality bioassay. The plant is reported to contain Sa-
ponins. There is growing interest in natural saponins as by the
fact of their wide spectrum of pharmacological activities, for
instance, bactericidal, antiviral, cytotoxic, and anti-cancer
(Ksouri et al. 2009; Balasundaram et al. 2006). Brine shrimp
lethality bioassay indicates cytotoxicity (Ahmed et al. 2013).
In toxicity evaluation of plant extracts by Brine shrimp
y = 0.1854x + 0.0412
R² = 0.9895
0.0
0.2
0.4
0.6
0.8
1.0
1.2
1.4
15.7 31.25 62.5 125 250 500
Absorbance
ConcentraƟon (mg/l)
Total phenolic content
Gallic acid
Linear (Gallic acid)
y = 0.1885x + 0.0339
R² = 0.983
0.0
0.2
0.4
0.6
0.8
1.0
1.2
15.7 31.25 62.5 125 250
Absorbance
ConcentraƟon (mg/l)
Total tannin content
Gallic acid
Linear (Gallic acid)
y = 0.158x + 0.2612
R² = 0.9669
0.0
0.2
0.4
0.6
0.8
1.0
1.2
20 40 60 80 100
Absorbance
ConcentraƟon (mg/l)
Total flavonoid content
querceƟn
Linear (querceƟn)
a
c
b
Fig. 2 Determination of total
phenolic (a), total tannin (b) and
total flavonoid (c) contents
Table 2 Polyphenolic content of ethanol leaves extract of BMET
Polyphenolic compounds Content
Total phenol 439.48 ± 0.009a
Total tannin 395.42 ± 0.001a
Total flavonoid 356.84 ± 0.001b
Here, each value represents the average of two analysis ± standard error of
mean. a, expressed in terms of mg GAE /100 g dried plant extract; and b,
expressed in terms of mg QE/100 g of dried plant extract
Bioactivity of Boehmeria Macrophylla (Urticaceae) Leaf Extract 237

6. Table 3 Percentage analgesic activity of BMET in acetic acid induced pain model
Animal group Mean of writhing % Writhing SD SE % Inhibition of writhing T-test (value of p)
Negative control 31 100 3.16 1.41 0
Standard (25 mg/kg) 7.4 23.87 1.95 0.87 76.13 14.24*
Extract (250 mg/kg) 20.8 67.09 1.30 0.58 32.91 6.69**
Extract (500 mg/kg) 14.2 45.8 1.09 0.49 54.2 11.25*
Here BMET denotes B. macrophylla ethanol leaves extract. *P < 0.05, **P < 0.01, SD = Standard Deviation, SE = Standard Error. Standard used
Diclofenac-Na
Fig. 3 Graphical representation
of brine shrimp lethality bioassay
and LC50 for BMET A and
Positive control B (Vincristine
sulphate) by Ldp line software
238 A. Hossain et al.

7. lethality bioassay LC50 values lower than 1000 μg/mL are
considered bioactive, (Martin-Cordero et al. 1995). A dose
dependent cytotoxic activity was shown in this study and the
LC50 value suggests the growing cell killing capacity of
BMET (Table 4). The extract showed a very good lethality
against brine shrimp which requires further cell line assay to
evaluate cytotoxic activity in more sensitive way (Fig. 3). It is
also reported that there is correlation between cytotoxic activ-
ity and brine shrimp lethality, (Abu Bakar et al. 2010;
Gurkan et al. 1995; Mongelli et al. 1995). Antibacterial
activity was tested by using the disc diffusion method.
Disc diffusion method is widely acceptable for the pre-
liminary screening of antibacterial activity. It is essen-
tially a qualitative or semi qualitative test indicating the
sensitivity or resistance of microorganisms to the test
materials. It is well known that plant containing various
phytochemical constituents such as flavonoids, saponins
and steroids have antimicrobial activity (Tim Cushnie
and Andrew 2005). The plant rich in flavonoids from
the species of Hypericum, Capsella, (Dall’Agnol et al.
2003) and Chromolaena (Aladesanmi et al. 1986) have
revealed antibacterial activity. Phytochemical preparations
rich in flavonoid contents have also shown antibacterial activ-
ity (Toyin et al. 2012; Daswani et al. 2010; Lamoral-Theys
et al. 2010; Brijesh et al. 2009; Naowaratwattana et al. 2010;
Ramalingam et al. 2009). It has been reported that saponins
have potent antimicrobial activity. In antibacterial screening,
twelve bacteria were selected for the test by disk diffu-
sion assay, where zone of inhibitions were observed
among the four bacteria (Table 5). These are Vibrio
cholerae, Escherichia coli, Staphylococcus epidermidis,
and Streptococcus ferus. A significant zone of inhibition
was observed against V. cholera which was comparable
to positive control (ciprofloxacin). BMET leaf extract
showed good zone of inhibitions against Escherichia
coli, Staphylococcus epidermidis, and Streptococcus
ferus. The maximum activity was demonstrated against
Vibrio cholerae and lowest against Streptococcus ferus.
Better antibacterial activity was demonstrated with
500 μg concentration than 250 μg.
Conclusions
The present study justifies the claimed traditional uses of
BMETand provides a livelihood for traditional medicine. This
study summarizes that BMET has potential in free-radical
scavenging activity, analgesic and cytotoxic as well as anti-
bacterial activities. Since, BMET showed both antibacterial
and cytotoxic activity it can be presumed that numerous active
secondary metabolites are present in the extract. However,
further studies are compulsory to explain the probable mech-
anisms related with these effects. However, we suggest for
further studies to explain the probable mechanisms related
with these effects.
Table 4 LC50 for extract and standard with lower and upper limit
Conc. (μg/ml) Lower limit (μg/ml) Upper limit (μg/ml)
Extract 33.16 27.32 40.04
Standard 0.72 0.41 1.03
Table 5 In vitro antibacterial activity of Ethanol leaves extract of BMET
Bacterial strains Type of bacterial
strains
Diameter of zone of inhibition in mm
Standard (Ciprofloxacin)
5 μg
Extract
250 μg
Extract 500 μg Negative control
Staphylococcus epidermidis Gram (+) 25 7.5 14 na
Streptococcus ferus Gram (+) 26 7 10 na
Streptococcus pyogens Gram (+) 33.5 na na na
Shigella boydii Gram (+) 29 na na na
Enterococcus faecalis Gram (+) 25.5 na na na
Staphylococcus aureus Gram (+) 27.5 na na na
Shigella sonnei Gram (+) 32 na na na
Shigella flexneri Gram (−) 38 na na na
Proteus species Gram (−) 27.5 na na na
Shigella dysenteriae Gram (−) 25 na na na
Escherichia coli Gram (−) 36 7.5 11.5 na
Vibrio cholerae Gram (−) 36.5 20 27 na
Gram (−) = Gram Negative Bacteria; Gram (+) = Gram Positive Bacteria; na = not active
Bioactivity of Boehmeria Macrophylla (Urticaceae) Leaf Extract 239

8. Acknowledgments The authors are thankful to the Pharmacy
Discipline of Khulna University, Bangladesh for providing laboratory
facilities and instrumental support as well as chemicals & reagents.
Also we acknowledge our heartfelt thanks to the authority of
BInternational Centre for Diarrheal Disease and Research, Bangladesh^
(ICDDR, B) for providing experimental mice as well as bacterial strains.
Our special thanks to the authority of Bangladesh National Herbarium
(BNH) for identification of the plant. Except these, no other institution/
foundation has an involvement in providing us any sort of research grant/
support.
Compliance with ethical standards
Ethical Statement N/A.
Conflict of Interest The authors declare that they have no competing
interests.
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