Department of Pharmacognosy, Faculty of Pharmacy, Babu Banarasi Das National
Institute of Technology and Management Dr.
Akhilesh Das Nagar, Faizabad
Road, Lucknow 227 105 U.P (India)
ABSTRACT:
Trichosanthes dioica R.
(Cucurbitaceae), an important perennial herb found
throughout the Asia and is figured high in Indian medicine system for its
versatile therapeutic and prophylactic properties. To justify its traditional
claim, hair growth activity was performed with ethanol and aqueous extracts on wistar albino rats using minoxidil
as standard. The extracts were formulated as 5% ointments and formulations were
applied topically on the denuded back of rats. Both formulations exhibited
prominent hair growth activity as compared to control group (p<0.05) as well as minoxidil treated group. The number of hairs and hair
length were also measured which were more significant for formulations.
Formulation having 5% aqueous extract found more potent than other groups. The
results of this study revealed the presence
of strong hair growth potential in the leaves of Trichosanthes dioica R.
KEYWORDS: Alopecia,
minoxidil, ointment, hair growth.
INTRODUCTION:
Hair is one of
the vital parts of the body derived from ectoderm of skin, is protective
appendages on the body and considered accessory structure of the integument
along with sebaceous glands and sweat glands 1.
Hair loss is one
of the dermatological disorders to human race which is common throughout the
world and is of great concern for decades. Many factor such as metabolism,
hormones, heredity and side effects of anti-neoplastic
and immunosuppressant drugs, have been negatively affecting the healthy growth
of hair 2. Various disease state such as typhoid, malaria, jaundice
etc., also cause hair fall, although temporarily. Hair loss causes a very
stressful state of mind for hair fall sufferers 3.
Topical
application of biological response modifiers and anti-androgens are currently
available therapies for the management of alopecia. The patents claim that the
effect are due to stimulation of the hair follicle or scalp metabolism,
possibility due to an acceleration of blood circulation, activation of dermal
papilla, anti-testosterone action or increased nutrition to the hair follicles
through accelerated blood flow but the mechanism are not yet clear4,5.
The usage of
synthetic drugs, minoxidil and finasteride
have abbreviated due to their side effects. Though the use of drugs for its
side effect is not advisable, the drug of plant origin is necessary to replace
the synthetic one. Natural products in the form of herbal formulations are
available on the market and are used as hair tonic, hair growth promoter, hair
conditioner, hair-cleansing agent, antidandruff agents, as well as for the
treatment of alopecia and lice infection 6.
A number of herbal products have been acclaimed with
hair growth- promoting activity. The traditional system of medicine in India
acclaims a number of herbal drugs for hair growth promotion
The leaves of T. dioica are used in alopecia or hair loss ethnopharmacologically (Shaarangadhara
Samhita)7. Trichosanthes, a genus of family Cucurbitaceae is an
annual or perennial herb distributed in tropical Asia and Australia. Trichosanthes dioica is cultivated throughout the
plain of Northern India, extending to Assam and East Bengal8. The various chemical constituents
present in T. dioica
are vitamin A, vitamin C, tannins, and saponins9. Phytochemical
evaluations of aqueous and ethanol extracts have showed the presence of saponins and tannins10. The present study was aimed to evaluate the hair growth activity
of two herbal formulations, in which incorporated ethanol and aqueous extracts
of Trichosanthes dioica leaves
separately at same level of
concentration.
MATERIALS AND
METHODS:
Collection and authentication of plant material
Plant material (leaves) were collected from local area of Lucknow
(U.P.) India, during the month of October and authenticated by Division of
Taxonomy, National Botanical Research Institute (NBRI), Lucknow,
India and a voucher specimen was deposited for future references (Ref. No.
NBRI/CIF/128/2010).
Chemicals:
A commercially
available hair remover (Veet, having Minoxidil 2%), as reference standard for promoting hair
growth.
Animals:
Healthy
male Wistar albino rats each weighing 100-120g were
used for study. The rats were housed in polypropylene cages and maintained
under standard conditions (12 h light and dark cycles, at 25 ±3 C and
35-60% humidity). Standard pelletized feed and tap water was provided ad libitum.
Ethical approval:
The experiments
were performed as per approved protocols by Institutional Animal Ethical
Committee under CPCSEA (Approval No. BBDNITM/IAEC/23/2010).
Preparation of
herbal extracts:
Dried, coarsely
powdered leaves of T. dioica (200 g) were soxhlet
extracted with ethanol. The solvent from ethanol extract was evaporated in rota evaporator (Buchi
type). The marc was air dried, and water extract was obtained by
boiling it with distilled water for 2 h. It was filtered, concentrated,
and dried in an oven at 40-50oC. These prepared ethanol extract and
aqueous extract of T. dioica
leaves were used in preparation of formulations.
Preparation of
formulations:
Ointments were
prepared by using the ointment base (I.P) containing white bees wax 2%, hard
paraffin 3%, cetostearyl alcohol 5%, white soft
paraffin 90%. Method reported in Indian Pharmacopoeia was followed for its
preparation11. These ointments named T1 and T2 containing 5% ethanolic extract and 5% aqueous extract of T. dioica
leaves respectively.
Primary
skin irritation test:
Measured quantity
(1g) of ointment was applied over the respective preselected test sites of all the
animals. The test sites were observed for erythema
and edema for 48 hours after application of test ointment. The ointment was not
showed erythema and edema then it was considered as a
safe preparation12.
Hair growth activity test:
Four groups of adult albino rats weighing (100-120 g),
five animals per group were used for the study. Four parameters were studied
for hair growth activity namely hair initiation time, hair completion time,
hair length and hair count. Commercially available Hair remover (Veet) was used to remove hair from dorsal portion of all
test animals nearly 2 sq. cm.
Denuded rats were divided into four groups (each having
five rats). Group-1 control, Group-2 treated with standard 2% solution of minoxidil (Mintop), Groups-3 and
4 treated with the two different solvents, ethanolic
and aqueous extracts formulations T1 and T2 respectively. Test ointments were applied topically
to the depilated area daily for 21 days. The numbers of hairs and hair length
was measured on 7th, 14th and 21st day during
experiment hair growth initiation and completion time were recorded for each
group of animals and compared with control13.
Qualitative hair growth analysis
was undertaken by visual observation of two parameters: hair growth initiation
time (i.e. minimum time to initiate hair growth on denuded skin region) and
hair growth completion time (i.e. minimum time taken to complete cover the
denuded skin region with new hair). Therefore, the hair growth initiation and
hair growth completion time were recorded for each group of animals with
respect to standard and control, during the treatment. Hair length and count
were considered to be important factors of growth. Hair was plucked randomly
from the denuded area of selected rats at 7th, 14th and
21st days. Manual measurement of plucked hair was carried out with
capillary holding the fine hair, with the aid of magnification lens. Random
samples of 25 strands of (from each group) hair were measured and the average
length (mm) was computed. At the same time intervals the numbers of hairs per square mm. was also
counted with the help of magnification lens
and computed average hair count.14, 15.
Statistical
analysis:
Statistical
analysis was carried out employing one way analysis of variance (ANOVA) by
application of Dunnett’s
multiple comparison test , p values <0.05 were considered as
significant.
RESULTS AND DISCUSSION:
The pH of the prepared
formulations, T1 (5% ethanol extract ointment) and T2 (5%aqueous extract
ointment) were found to be 6.8 and 6.7 respectively. The ointments showed
characteristic odour and different textures (Table 1). Primary
skin irritation test was carried out to evaluate the skin irritation by the
prepared formulations on intact skin of rats. Both the prepared ointments did
not show any erythema and edema in the duration of 48
h, this indicates that the prepared formulations were nonirritant and nontoxic
on skin of rats. Thus, the prepared ointments were considered safe for topical
administration.
Table 1: Physical parameters of formulations
|
Formulations |
pH |
Texture |
Odour |
|
T1(
5% ethanol extract ointment) |
6.8 |
Blackish
green |
Characteristic |
|
T2
(5% aqueous extract ointment) |
6.7 |
Greenish
yellow |
Characteristic |
These ointments were applied to
the denuded area of albino rats once a day for 21 days. The results
of preliminary hair growth initiation tests were encouraging. They were
recorded on the basis of time taken for initiation of growth of hair follicles.
This parameter was recorded for finding effective extract formulation of drug. Minoxidil and T1 showed significant initiation growth after
4th day but T2 showed initiation growth after 3rd day of
the experiment. The formulation T2 showed fastest rate of initiation of hair
growth when compared with other. It was also observed that the time taken for
complete hair growth was 18 days in Minoxidil and T1
treated groups, but completion time 17 days in T2 treated group indicating
early initiation and completion of hair growth (Table 2, Fig-1). The results show that formulation T2 has
contributed most significant hair growth activity when compared to the control.
Table 2: Effect of different formulations on qualitative hair growth
|
S.
No. |
Formulation |
Hair
growth (days) |
|
|
Initiation
time |
Completion
time |
||
|
1. |
Control |
6 ± 0.44 |
21 ± 0.44 |
|
2. |
Minoxidil (2%) |
4 ± 0.54* |
18 ± 0.31** |
|
3. |
T1 |
4 ± 0.63* |
18 ± 0.54** |
|
4. |
T2 |
3 ± 0.44** |
17 ± 0.31** |
Values are expressed as mean ±
SEM, n = 5, *p < 0.05 and **p < 0.01 considered
significant (compared to control group) by using one way Analysis of Variance followed
by Dunnett’s test
Figure 1: Graph showing effect of formulations on hair initiation and completion
time
Hair length was measured at the
end of 7th, 14th, 21st days in all the groups
(Table 3, Fig-2). The hair lengthening properties was found to be in the
order of T2 > T1 > Minoxidil > Control.
Therefore it can be suggested that formulation T2 (5% aqueous
extract ointment) showed the best hair lengthening properties as compared to
others.
Table 3: Hair length of groups treated
with different formulations
|
S. NO. |
Treat ment |
Length of hair (mm) |
||
|
Day 7th |
Day 14th |
Day 21st |
||
|
1. |
Control |
1.0±0.071 |
6.5±0.17 |
12.1±0.13 |
|
2. |
Minoxidil (2%) |
2.8±0.22** |
9.8±0.07** |
16.9±0.09** |
|
3. |
T1 |
3.1±0.04** |
11.2±0.07** |
17.6±0.04** |
|
4. |
T2 |
3.4±0.08** |
12.4±0.14** |
18.5±0.07** |
Values are expressed as mean ±
SEM, n = 5, **p < 0.01 considered significant (compared to control
group) by using one way Analysis of Variance followed by Dunnett’s test
Figure 2: Graph
showing hair length of groups treated with different
formulations
Hair count was done at the
end of 7th, 14th, 21st days in all the groups
(Table 4, Fig-3). The numbers of hair follicle were found to be in the
order of T2 > T1 > Minoxidil > Control.
Therefore it can be suggested that formulation T2 (5% aqueous
extract ointment) showed the best hairs growing property as compared to others.
Table 4: Hair count of groups treated with different formulations
|
S. No. |
Treatment |
Hair count (Day) |
||
|
7th |
14th |
21st |
||
|
1 |
Control |
24±0.70 |
81±0.70 |
152±2.30 |
|
2 |
Minoxidil (2%) |
36±1.0** |
120±1.6** |
186±0.94** |
|
3 |
T1 |
40±1.7** |
136±2.0** |
210±3.53** |
|
4 |
T2 |
42±0.70** |
150±1.7** |
234±1.49** |
Values are expressed as mean±
SEM, n = 5, **p<0.01 considered significant (compared to control group) by using one way
Analysis of Variance followed by Dunnett’s
test
From the results of hair growth
activity experiment in rat model, it can be strongly suggested that both the
leaves extracts of T. dioica
have potential components to stimulate the hair growth. It was also
resulted that T2 (5% aqueous extract ointment) showed the best hair growth
activity with no skin irritation. It showed minimum time to initiate as well as
to complete the hair growth at denuded surfaces. As compared to others, it was
also found to improve the hair length and hair count (Fig.-4).
Figure 3: Graph showing hair count of groups treated with different formulations
Figure 4:
Photographs of
albino rats (a) control, (b) standard, (c) T1, (d) T2 at 14th
day of experiment
CONCLUSION:
From the pharmacological study for hair
growth activity of Trichosanthes dioica leaves
it can be concluded that both the formulations 5%
aqueous extract ointment and 5% ethanol extract ointment of T. dioica leaves have potential components to stimulate
the hair growth. It can also be concluded that 5% aqueous extract ointment
showed the best hair growth activity with no skin irritation. It showed minimum
time to initiate as well as to complete the hair growth at denuded surfaces. As
compared to others, it was also found to improve the hair length and hair
count.
The present study therefore provides a basis
for further detailed investigations in the direction of isolation of active
compounds and elucidating the mechanism of hair growth activity, which is also
recommended by the authors.
REFERENCES:
1.
Rathi V, Rathi JC, Tamizharasia S and Pathak AK. Plants used for hair growth promotion: A Review.
Pharmacognosy Reviews 2008; 2(3): 185-187.
2.
Olsen EA. Androgenetic
alopecia: Disorders of hair growth, Diagnosis and treatment. New York: McGraw
Hill Inc; 1995. pp. 651-655.
3.
Ranganathan S. and Shobhana S. Evaluation of herbal hair oil in reducing hair
fall in human volunteers. Indian Drugs
2008; 45(6): 451-455.
4.
Kameyama S. Application and
Consideration of medicinal plants for hair care product. Frag
J. 1995; 23: 28-34.
5.
Lee OS, Kang, HH and Han SH. Oriental herbs in
Cosmetics: Cosmet Toiletries, 1997; vol.112, pp.
57-64.
6.
Olsen EA. Androgenetic
alopecia: Disorders of hair growth, Diagnosis and Treatment, New York; MC Grow
Hill Inc; 1993. p. 257-287.
7.
Khare CP. Encyclopedia of Indian medicinal plants. Springer science business
media; 2007. pp. 458.
8.
Kirtikar KR, and Basu B.D. Indian medicinal plant. 2nd
edn. Oriental Enterprises, Dehradun; 2001. vol.5, pp.1543.
9.
Chopra RN, Nayar SL and Chopra IC. Glossary of Indian Medicinal
plants. New Delhi: CSIR; 2002. pp. 93-94.
10.
Ghaisas MM, Tanwar MB. Ninave
PB, Navghare VV and Deshpande
T. Hepatoprotective activity of aqueous and ethanolic extract of Trichosanthes dioica in ferrous sulphate
induced liver injury. Pharmacologyonline 2008; 3:
127-135.
11.
Roy RK, Thakur M and Dixit
VK. Effect of Cuscuta reflexa
Roxb. on hair growth activity of albino rats. Indian Drugs 2006; 43(12): 951-956.
12.
Roy RK, Thakur M, and Dixit
VK. Effect of Citrullus colocynthis
on Hair Growth in albino Rats. Pharmaceutical
Biology 2007; 45(10): 739-744.
13.
Suraja R, Rejitha G, Anandarajagopala
K and Promwichita P. In vivo hair growth activity of Prunus dulcis seeds
in rats. Biology and Medicine 2009; 1(4): 34-38.
14.
Patni P, Varghese D, Balekar N and Jain DK. Formulation and evaluation of herbal
hair oil for alopecia management. Planta Indica 2006; 2(3): 27-30.
15. Sabarwal N,
Dhanila V, Barik R, Khandelwal A, Jain A, and Jain S.Development and evaluation of polyherbal formulations for hair growth activity. Phcog J 2009; 1(2): 165-170.
Received on 30.10.2010
Accepted on 03.12.2010
© A&V Publication all right reserved
Research Journal of Pharmacognosy and Phytochemistry. 3(1): Jan. - Feb. 2011, 30-33