IJBPAS, June, 2016, 5(6): 1490-1500

ISSN: 22774998

COMPARATIVE STUDY ON ANTILEISHMANIAL AND CYTOTOXIC ACTIVITY OF

LAWSONIA INERMIS BARK AND ALOE VERA LEAVES
KASHIF IQBAL1*, JAVEID IQBAL1, MARIA SADAF AFREEN2
1

Department of Pharmacology, Faculty of Pharmacy and Health Sciences, University of

Balochistan, Quetta
2

Gomal University, Dera Ismail khan, KP, Pakistan

Corresponding Author: KASHIF IQBAL: Address: Department of Pharmacology,

Faculty of Pharmacy and Health Sciences, University of Balochistan, Quetta, Pakistan;
Tel.: +92 3356951284; E Mail: kashifiqbal321@gmail.com
ABSTRACT
This work describes an invitro, invivo antileishmanial and cytotoxic activity of Lawsonia
Inermis bark and Aloe Vera leaves extract. Methanolic extract of the plant material at
concentrations from 100 - 500 g/ml was tested invitro to get % inhibition activity on L. tropica
KWH23 promastigotes in comparison with negative control and Amphotericin-Bat 12-24 hours,

whereasinvivo antileishmanial activity was checked agianst L. tropica infected Albino miceand
cytotoxicity was analyzed via mammalian cell line. For Lawsonia Inermis bark, mean %
inhibition in extracellular promastigotes at four different concentrations (100 g/ml, 125 g/ml,
250 g/ml, and 500 g/ml) at 24th hour were 98.02 0.06, 98.70 1.09, 99.41 0.00 and 100.00
0.00 respectively, whereas after 8 weeks, mean lesion size decreased from 0.81 0.20mm to
0.10 0.11mm (p < 0.01)and % cure rate against intracellular amastigotes at dose 75mg/kg was
98.022 (95% C.I = 96.13-98.09) in Albino mice. IC50 value calculated for Lawsonia Inermis bark
to estimate the cytotoxic activity was 25.105 g/ml (95% C.I = 15.55-33.83) against
lymphocytes. The study proved to be Lawsonia Inermis bark as safe and potent inhibitor against
Leishmania tropica parasites.
Keywords: Lawsonia inermis, Aloe Vera, Leishmania tropica
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INTRODUCTION
Leishmaniasis is a major public health

non-volatile

problem in tropical and sub-tropical regions,

hydrocarbons, and alkaloids which possess a

and the disease is caused by parasites

variety

belonging to the genus Leishmania (Family

including

Trypanosomatidae). WHO have reported that

antiparasitic [3, 4].Aloe Vera belong to

people from 98 countries - covering 5

family Liliaceae; found in dry regions of

continents - are at high risk of Leishmaniasis,

Africa, Asia and Southern Europe used as

and it is estimated that approximately 12

therapeutic agent and have diverse action

million people are currently infected. In

against skin diseases - phytochemically

Pakistan, L. tropica and L. major are the

includes

main cause of cutaneous leishmaniasis. First

constituents which possessed antioxidant,

line therapy for cutaneous leishmaniasis in

antimicrobial and anticancer properties [5 -

Europe, Asia and Africa is pentavalent

7].

antimonials, i.e., sodium stibogluconate.

In the current work, comparison between

However, antimonials have severe side

antileishmanial

effects like myalgia, pancreatitis, cardiac

Inermis and Aloe Verawere examined on

arrhythmia, hepatitis, and accumulation of

Leishmania

the drug in liver and spleen. Thus, there is an

amastigotes whereas cytotoxic effect of plant

urgent need for new chemical entities for

extract was checked on lymphocyte cell line.

non-toxic

MATERIALS & METHODS

and

effective

treatment

of

terpenes),

of

aliphatic

pharmacological
antioxidant,

antiviral

Anthraquinones

activities

tropica

activities,

and

of

and

Phenolic

Lawsonia

promastigotes

and

leishmaniasis [1, 2].

Chemicals

Lawsonia inermis (Family Lythraceae) is

Fetal

native to Northern Africa and South-western

sulfoxide (DMSO), RPMI-1640 medium,

Asia - is cultivated in many tropical and sub-

Amphotericin B, penicillin, streptomycin,

tropical regions. Different parts of plant

formic

specially leaves including

flowers, stem,

(Ethanol, Methanol and Acetone) were

bark and root are used as traditional medicine

purchased from Sigma-Aldrich (St. Louis,

for years -havelarge number of identified

MO, USA). Water used for analysis was

compounds,

purified by deionization and 0.22 m

flavonoids,

coumarins,

bovine

acid,

serum

analytical

(FBS),

grade

dimethyl

solvents

alkylphenones, terpenes (volatile terpenes,

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Kashif Iqbal et al

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membrane filtration (Millipore, Billerica,

pressure at 46 C using a rotary evaporator,

MA, USA).

which afforded 500 g of crude extract.

Collection of plant material and preparation

In vitro antileishmanial activity

of crude extract

In vitro antileishmanial activity of Lawsonia

Plant material of Lawsonia Inermis bark and

inermisbark and Aloe Vera Leaveswere

Aloe VeraLeaves were collected from the

performed

territory of Dera Ismail Khan, Khyber

Leishmania tropica promastigotes (KWH23).

Pakhtunkhwa (KPK), Pakistan in August and

The in vitro antileishmanial growth inhibition

September

was

assay was adopted from Ozbilgin et al, 2014.

performed by Dr. Siraj ud-Din, Department

Promastigotes of Leishmania tropica were

of

specimen

cultured in RPMI-1640 medium containing

(accession number: Bot, 200110 (pup) and

10% fetal bovine serum, 200 U/mL of

200111 (pup) was deposited at Department

penicillin, and 0.2 mg/mL of streptomycin.

of Botany, University of Peshawar (UOP),

The parasites were cultured at 26 C for 4

KPK. The plant material was washed with

days in BOD incubator (Gallenkamp, Size 1,

distilled water before drying them in shade at

UK),

temperatures below 35 C. The dried material

harvested in sterile tubes. The number of

was stored in a cool dark place until use.

promastigotes were measured by transferring

Crude

by

5-10 l to a haemocytometer (REICHERT,

macerating 1 kg of powdered leaves in 2 liter

N.Y, U.S.A), and counting the number of

of methanol for 1 week with regular stirring

promastigotes under upright microscope

in the early morning. The extract was filtered

(CX31, OLYMPUS, Tokyo, Japan). The

through muslin cloth, and subsequently

number of promastigotes were calculated

through Whatman filter paper No. 41. The

using the formula:

Botany,

2015.

and

Identification

plant extract

voucher

was

obtained

where

with

after

clinically

isolated

promastigotes

were

filtrate was concentrated under reduced

Viable cell Count (live cells/ml) =

were

concentration of 2.1 10 6 promastigotes/mL

subsequently centrifuge at 4C at 2000 rpm

in the required volume (10ml). Distribute 10

for 10 min, the supernatant removed, and the

ml having 2.1 x 10 6promastigotes/mL in 96

pellet reconstituted in fresh RPMI-1640

wells of culture plate through multipipette

medium with 10% FBS to obtain a

and fraction mix well. Left the culture plate

The

harvested

promastigotes

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Research Article

in BOD incubator at 26C for two days.

RPMI-1640 medium with 10% FBS was

Haemocytometer was used to count the

added to get the required volume (10ml).

promastigotes under upright microscope

Add required volume (10l) of promastigotes

(lens 40x) at 12 and 24 hours. Performed the

(2.1 x 106 promastigotes/ml) into cardiac

in vitro promastigotes test in triplicate and in

cavity (Intraperitoneally) of BALB/c mice.

last the mean percentage inhibition of

The development of lesion was measured

parasite were taken in record.

weekly with dial micrometer during infection

Invivo test

period. The infection was well established

Animal

Model and

Administration of

and cleared visible lesions were seen with

Extract

naked eye.

To study the pathogenesis of leishmania

4 groups (n=6) of BALB/c were made in

strain, the BALB/c mouse was used by

which 2 groups comprised of drug control, 1

applying very low amount of drug agent.The

group have positive control and 1 group was

BALB/c mouse having weight (20-32 gm)

negative control. After 36 days, the lesion

sex male, age 6-8 week, cardiac route for

was established, so treatment process was

drug input and 4 groups were made for in

started. Dose of plant extract materail to

vivo screening.

Group I and II for mice treatment were

Leishmania Tropica KWH23 promastigotes

15mg/kg for 5 days (Total dose 75mg/kg) in

were cultured in RPMI-1640 medium along

DMSO solvent up to final volume of 3 ml

with 10% Fetal bovine Serum, penicillin 200

and 10 ml respectively. Amphotericin was

U/ml and Streptomycin was 0.2 mg/ml. The

used as Standard drug at a dose of 15mg/kg.

parasite was cultured and growth multiplied

No drug agent was used in IV group

at 26 C for 4 days in BOD incubator and

(Negative Control). Injection dose 10 l, five

harvested parasite was used. The harvested

times with 3 day intervals were administered

promastigotes were taken in sterile tube and

to each mice and its result was recorded

was

by

regularly. Dial micrometer was used to note

haemocytometer under upright microscope.

the difference between size of the lesion in

Calculate the number of promastigotes and

infected and uninfected mice weekly. Before

centrifuge at 4 C at 2000 rpm for 10 min,

and

Supernatant

samples were taken from infected lesion [8].

checked

and

liquid

was

counted

removed

and

after

treatment,

needle

aspirations

discarded while pellet was left in tube. Fresh

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To

detect

Research Article
amastigotes

under

light

2000 rpm at 4C for 30 minutes. Puncture the

microscope, Giemsa stained was used in

lower portion of transparent layer in tube

th

with syringe, take liquid carefully, add 5 ml

and 60 day of infection, 60 mg of tissue

of RPMI-1640 medium and count the

sample was taken from the lesion for biopsy.

number

To search amastigote in leishmania strain,

haemocytometer under light microscope.

sample was smeared on the slides stained

Normally 10,000 cells (Lymphocytes) are

with Giemsa and light microscope was used

present in each milliliter. Put 100 l of

for examination.

lymphocyte media into 96 wells of culture

samples, under oil immersions. On the 48

th

Ethics Statement: BALB/c mice

were

of

lymphocytes

through

plate [10].

supplied by Department of Pharmacology

Take

(Animal center), University of Peshawar,

mammalian cell test were 500, 250, 125, and

KPK, Pakistan and this study was approved

100 g/mlin DMSO solvent up to final

by Animal and Ethics Committee [9], Faculty

volume of 3 ml and 10 ml respectively.

of Pharmacy and Health Sciences, University

Amphotericin having concentration 25 g/ml

of Balochistan, Quetta. Standard diet along

(positive control) was taken as reference drug

with water ad Lawsonia inermis Barkitum

while negative control was Leishmania

was

tropica KWH23 promastigotes. Add required

given

to

BALB/c

mice

during

the

of

dose of plant material

promastigotes

(2.1

for

106

experiments.

volume

Cytotoxicity test

promastigotes/ml) in 12 wells of culture plate

Fresh blood (10ml) was taken in BD

through multipipette. Left the culture plate in

Vacutainer K2E (EDTA) obtained from

incubator at 26 C for two days (at least

healthy volunteer to get mammalian cell

otherwise three days). Haemocytometer was

(lymphocyte) for cytotoxic analysis of plant

used to count the viable lymphocytes and

material. PBS (Phosphate Buffered Saline)

promastigotes under light microscope (lens

through 0.2 m filter under laminar flow

40x) at 12 and 24 hours. Performed the

hood (kept sterile conditions) and then add

cytotoxic test of mammalian cell in triplicate

equal quantity of PBS and blood in sterile

and in last the IC50 were taken in record.

tube. Put Ficol solution (ratio 1:2) carefully

Statistical analysis

at 165 angle in already mixed PBS and

In this study, invitro assay of plant extract

Blood tube. Centrifuge the 30ml mixture at

was expressed as the % Inhibition of parasite

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Research Article

growth, and calculated as the mean of three

replicate

measurements

with

deviation using the equation:

standard

Percentage inhibition =

Result record of cytotoxic activity was

ranging between 70 and 93%. At 24th hour,

expressed as % inhibitory concentration

parasite %age inhibition ranging between 65

(IC50) and was determined by non-linear

and 98 at 100g/ml and ranging between 81

regression

for invivo

and 100% when extract was checked at 500

(mm)

g/ml concentration.Lawsonia inermis bark

assays,

analysis

mean

whereas

lesion

size

and

percentage cure rate with 95% confidence

showed

level was taken in record, using GraphPad

reached 98% at 100 g/ml at 24 th hours (p <

Prism 5 software (GraphPad software, San

0.01). In comparison with Amphotericin-B,

Diego, CA). P <0.05 was the level of

having parasitic inhibition ranging between

significance.

50.12 and 78.78% at 100 and 500 g/ml

RESULTS

concentration respectively, whereas at 24 th

Invitro Antileishmanial activity

hour showed parasite inhibition between

The current work describes the extract of

80.09 and 90.55% at 100 and 500 g/ml

Lawsonia inermis bark and Aloe Vera

concentrationagainst L. tropica, Lawsonia

Leaves-

inermis showed the significant results,

in

combination

with

invitro

promising

ranging

extract of all plant material was tested for its

antileishmanial activity at 24th hour at 100

ability to inhibit extracellular promastigotes

and 500 g/ml concentration respectively.

(2.1 x 106 cells) ofL. tropicaKWH23 at

Invivo Antileishmanial activity

concentrations of 500, 250, 125, and 100

Invivo activity of Lawsonia inermis bark and

g/ml, after 12-24 hours gave promising

Aloe Vera Leaves were analyzed on Albino

result

shown

whereasAmphotericin-B

in
was

98.02

and

activity,

antileishmanial profiling. Crude methanol

as

between

inhibitory

100%

Table

1,

mice infected with 0.02 ml clinically isolated

used

as

L.

Tropica

KWH23

having

2.1

Standard drug.

10 promastigotes via intraperitoneal route,

At 12th hour analysis, the plant extract

after 36-120 days gave promising result as

percentage inhibition ranging between 40.10

shown in Table 2.Four groups consisting of

and 59.01 % at 100g/mlconcentration while

Six Albino mice each were made, in which

at 500 g/ml concentration, %age inhibition

Group I and II were used for sample analysis.

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Research Article

Group III was used as positive control and

from the resultsthat Lawsonia Inermisbark

Group IV as negative control. Leishmania

exhibited significant invivo antileishmanial

lesion of mice was checked by measuring its

activity against L. tropica (p < 0.01) infected

size before and after treatment of sample and

mice whereas rest of the plant material have

difference in size of lesion was recorded and

promising activity as compared to negative

its mean value was taken in record. All the

control group.

experimental procedure were go through in

Cytotoxic effect

triplicate. In two (2) sample groups, the mean

Cytotoxic activity of Lawsonia inermis bark

lesion size of mice decreased significantly

and Aloe Vera Leaves were analyzed invitro

from0.51 0.20 mmto 0.10 0.10 mm after

against Lymphocytes having count 1.8x 10 4,

treatment with plant extract but negative

after

group reached 1.5 0.50 mm (p > 0.05),

summarized in Table 3.

whereas Amphotericin-B decreased from

The methanolic extract of Aloe Vera Leaves

0.85 0.60 mm to 0.15 0.60 mm, at the

showed highest cytotoxic activity at a

end of 8th week. The mice receiving

concentration of more than 100 g/ml.

methanolic extract of Lawsonia inermis bark

Lowest cytotoxic activity was observed when

and Aloe Vera Leaves have percentage cure

Lawsonia Inermis bark extract was exposed

rate was 98.02% and 75.27% respectively

to lymphocyte cells having IC50 values of

after 8th week treatment. It is evident that

25.105 g/ml.

12-24

hours

showed

result

as

Table 1: In vitro Antileishmanial activity of Plant extract; Data represent mean percent inhibition S.D of three replicates
Sample
Lawsonia inermis bark

Aloe vera Leaves

Amphotericin-B

Negative Control

Sample concentrations
(g/ml)
100
125
250
500
100
125
250
500
100
125
250
500
100
125
250
500

Percent inhibition at Time (h)

12
24
59.010.02
98.020.06
70.680.03
98.701.09
85.350.10
99.410.00
93.000.34
100.000.00
40.100.70
65.110.00
53.000.20
70.030.07
61.070.40
78.011.00
70.000.20
81.960.00
50.120.16
80.090.06
63.340.09
84.671.09
71.450.10
87.620.00
78.780.07
90.550.57
0.000.00
0.000.00
0.000.00
0.000.00
0.000.00
0.000.00
0.000.00
0.000.00

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Table 2: In vivo Antileishmanial activity of Plant extract; Data represent mean lesion size (mm) with percentage cure rate
with 95% confidence intervals
Sample

Dosing
Regimena,b
(For 5 Days)

Mean Lesion
(mm)
Pretreatment

Mean Lesion (mm)

After Treatment
(After 8 Weeks)

%age Cure Rate

(with 95% Confidence
intervals)

No: of mice
cured/No: of mice
Infected

Mean survival time

(Days)

Lawsonia inermis
bark
Aloe vera Leaves

15mg/kg

0.810.20

0.100.10

6/6

60

15mg/kg

0.720.30

0.510.20

98.02
(96.13-98.09)
75.27
(74.20-76.80)

5/6

60

Amphotericin-B

15mg/kg

0.850.60

0.150.60

96.00
6/6
(94.28-96.01)
15mg/kg
0.760.50
1.50.50
0.000
0/6
a: Total dose 75 mg/kg; b: Route of administration: Intraperitoneally (i.p) Injection

Negative Control

60
30

Table 3: Cytotoxic Effect of Plant extract; Data represent IC 50(g/ml) with 95% confidence intervals
Sample

IC50 (g/ml)
95% confidence intervals
25.105
(15.55-33.83)
>100

Lawsonia inermis bark

Aloe vera Leaves

DISCUSSION

showed significant antileishmanial activity

In Algeria, L. majorpromastigotes were

agianst L. tropica [13]. Crude methanolic

successfully treated with 10l hydroalcoholic

extracts of four Sudanese plants (Azadirachta

extracts of Juglan regia, Lawsonia ienrmis

indica, Allium Sativum, Acacia nilotica and

and Salvia officinalis [11]. Helietta apiculata

Balanites aegyotiaca) were analyzed against

was reported to have significant invitro

L. major promastigotes inwhich Balanites

antileishmanial activity on L. amazonensis,

aegyotiaca showed moderate antileishmanial

Leishmania

Leishmania

activity [14]. The leishmanicidal activity of

Braziliensis whereas invivo antileishmanial

Aloe vera leaf exudate was reported to have

activity against L. amazonensis [12].

promising invivo antileishmanial effect when

In literature, there are few studies having

checked against L. donovani strains [15].

antileishmanial activity of plant extracts and

In this work, methanol was selected as a

there

solvent for extraction due to its polar nature

Infantum

isolated

and

natural

compounds

and

secondary metabolites agianst L. tropica.

in preparation of plant extract. Due to

In a study conducted in Pakistan on

methanolic extract, secondary metabolites

antileishmanial activities of 5 plant species

play crucial role in antileishmanial and

Stellaria media, Sida cordata, Asparagus

cytotoxic activity. Bark of Lawsonia Inermis

gracilis, Jurinea dolomiaea and Opuntia

showed invitro % inhibition100 0.00 at 500

ficus-indica

g/ml concentration (p < 0.01), after 24

inwhich

Jurinea

dolomiaea

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Kashif Iqbal et al
hours

Research Article

which

confirmed

antileishmanial

CONCLUSION

activity from previous findings[11]. Aloe

Antileishmanial and cytotoxic evaluation

Vera leaves possessed 81.96 % at 500 g/ml

stated that bark of Lawsonia Inermisis safe

(p < 0.04),

after 24 hours which is an

and promising antileishmanial source against

agrrement with the results reported for the

L. tropicaparasites as compared to Aloe Vera

antileishmanial activity as described earlier

leaves.

in literature [15].Thus, it was confirmed that

REFERENCES

plant

materail

secondary

[1] Singh S, Sivakumar R. Challenges

metabolites such as Flavonoids, Tannins,

and new discoveries in the treatment

Coumarins,

of

Alkaloids

possessed

Terpenes,
[4,

12]

Saponins
which

and

exhibited

antileishmanial and cytotoxic activity against

leishmaniasis. J

Infect

Chemother2004; 10: 307-315.

[2] Brooker S, Mohammed N, Adil K,

L. Tropica.

Agha S, Reithinger R, Rowland M,

Extract of Lawsonia inermis possessed very

Ali I, Kolaczinski J. Leishmaniasis in

interesting and significant (p < 0.01) invitro

refugee

and invivo activity against L. tropica

populations.

promastigotes

2004; 10(9):1681-1684

than

Aloe

Vera

and

and

local

Emerg.

Pakistani
Infect.

Dis.

Amphotericin-B. All Albino mice were cured

[3] Liou JR, ElShazly M, Du YC, Tseng

at or after 60 day of infection, when extract

CN, Hwang TL, Chuang YL, Hsu

of Lawsonia iermis was given whereas after

YM, Hsieh PW, Wu CC, Chen SL, et

8th week, mean lesion size was decreased

al., 1,5-Diphenylpent-3-en-1-ynes and

upto 0.10 0.10 mm in mice. That effect

methyl

may

from Lawsonia inermis and their anti-

be

due

to

killing

of

naphthalene

carboxylates

promastigotes/amstigotes or by inhibiting

inflammatory

extra/intracellular metaboic pathways of L.

Phytochemistry 2013; 88: 6773.

tropica strains.Cytotoxic effect of Lawsonia

[4] Semwal RB, Semwal DK, Combrinck

inermisleaves was moderate (IC50 value:

S, Cartwright-Jones C, Viljoen A.

25.105g/ml) were known to results of test

Lawsonia

extract [16].

Ethnobotanical, Phytochemical and

Inermis

pharmacological

activity.

L.

(henna):

aspects.

J.

Ethnopharmacol 2014; 155: 80-103.

1498
IJBPAS, June, 2016, 5(6)

Kashif Iqbal et al

Research Article

[5] Sydiskis RJ, Owen DG, Lohr JL,

Rosler

KH,

Blomster

RN,

CIOMS, Inc.; c2002 [cited 2016 May

21].

Available

from

http://eur-

Inactivation of Enveloped Viruses

lex.europa.eu/LexUriServ/LexUriSer

by Anthraquinones Extracted from

v.do?uri=OJ:L:2010:276:0033:0079:

Plants,

Agents

En:PDF.

Chemother. 1991; 35 (12): 2463-

[10] Byum,

Antimicrob.

2466.

A.

lymphocytes,

[6] Lopez A, Tangil MS, Vega-Orellana

O, Ramirez AS, Rico M. Phenolic
constituents,

antioxidant

Isolation

of

granulocytes

and

macrophages.

Scand

Immunol.1976, 5, 915.

and

[11] Serakta M, Djerrou Z, Mansour-

antimycoplasmic

Djaalab H, Kahlouche-Riachi F,

activities of leaf skin and flowers of

Hamimed S, Trifa W, Belkhiri A,

Aloe Vera (L). Burm. F. (syn. A.

Edikra

Barbadensis Mill) from the canary

Antileishmanial activity of some

Islands (Spain) Molecules 2013; 18:

plants growing in Algeria: Juglans

4942-4954.

regia, Lawsonia Inermis and salvia

preliminary

[7] Kim HS, Lee BM. Inhibition of

Benzopyren DNA Adduct Formation
by

Aloe

Barbadensis

N,

Pacha

YH.,

officinalis. Afr J Tradit Complement

Altern med. 2013; 10 (3): 427-430.

Miller,

[12] Ferreira ME, Arias AR, Yaluff G,

Carcinogenesis, 1997; 18 (4): 771-

Bilbao NV, Nakayama H, Torres S,

776.

Schinini A, Guy I, Heinzen H,

[8] Ozbilgin A, Durmuskahya C, Kayalar

Fournet A. Antileishmanial activity

H, Ertabaklar H, Gunduz C, Ural IO,

of furoquinolines and Coumarins

Zeyrek

from

F,

Kurt

O,

Cavus

I,

Baalcioglu C et al., Antileishmanial

activity of selected turkish medicinal

Helietta

Phytomedicine 2010; 17: 375-378.

[13] Shah NA, Khan MR, Nadhman A.

plants. Trop J Pharm Res 2014; 13

Antileishmanial,

(12): 2047-2055

phytochemical

[9] Directive
protection

2010/63/EU on
of

animals

used

apiculata.

toxicity
evaluation

and
of

the

medicinal plants collected from

for

Pakistan. BioMed Res. Int. 2014; 7.

scientific purposes Hinsdale. (IL):

1499
IJBPAS, June, 2016, 5(6)

Kashif Iqbal et al

Research Article

[14] Khalid FA, Abdalla NM. Mohomed

HE, Toum AM, Magzoub MM, Ali
S. In vitro assessment of anticutaneous leishmaniasis activity of
some

Sudanese

plants.

Turkiya

Parasitol Derg, 2005; 29 (1): 3-6.

[15] Dutta A, Sarkar D, Gurib-Fakim A,
Mandal C, Chatterjee M. Invitro and
invivo activity of Aloe vera leaf
exudate in experimental visceral
leishmaniasis. Parasitol res 2008;
102: 1235-1242.
[16] Endrini S, Rahmat A, Ismail P,
Taufiq-Yap YH. Comparing of the
cytotoxicity

properties

and

mechanism of Lawsonia Inermis

and Strobilanthes crispus extract
against several cancer cell lines. J
Med Sci. 2007; 7 (7): 1098-1102.

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