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Current Research in Microbiology and Biotechnology

Vol. 1, No. 3 (2013): 98-103


Research Article
Open Access

ISSN: 2320-2246

Study of Citronella leaf based herbal mosquito


repellents using natural binders
Nandini Rani, Aakanksha Wany, Ambarish Saran Vidyarthi and Dev Mani Pandey*
Department of Biotechnology, Birla Institute of Technology, Mesra, Jharkhand- 835215, India

* Corresponding author: Dev Mani Pandey, email: dmpandey@bitmesra.ac.in

Received: 15 April 2013

Accepted: 27 April 2013

Online: 04 May 2013

ABSTRACT
Citronella grass has been serving from years as a mosquito repellent in the field of ancient and modern medicine.
Commercially available mosquito repellents are chemical based and disastrous to human health. An attempt has
been made to prepare a 100% herbal product based on Citronella leaf remains which is left out and of no use
after steam distillation. It is cheap, effective and environment friendly. It is a first and preliminary work based on
formulating and evaluating herbal mosquito repellent cakes using natural binders such as neem powder, potato
starch, corn starch, coconut shell powder, wood powder and cow dung. The efficacy of prepared citronella leaf
cakes were evaluated on three different parameters such as flammability, burning time and mosquito repellency
test. Also, the cakes were sprayed with different concentrations of Citronella oil. Based on the results obtained
from these parameters, the residual percentage of each combination of cakes was calculated and it suggested
that Neem powder cake has the most effective repellency activity when impregnated with 10% Citronella oil.

Keywords: Citronella leaf cakes; flammability; mosquito repellent; natural binders; residual percentage
INTRODUCTION
Controlling mosquitoes is of utmost importance in the
present day scenario with rising number of mosquito
borne diseases. An alarming increase in the range of
mosquitoes is mainly due to deforestation,
industrialized farming and stagnant water. Thus,
special products like mosquito repellents for combating
mosquitoes are required. The products used for
mosquito control have varying degrees of effectiveness.
Carbon dioxide and lactic acid present in sweat in
warm-blooded animals act as an attractive substance
for mosquitoes. The perception of the odor is through
chemoreceptors present in the antennae of mosquitoes.
Insect repellents work by masking human scent. A
number of natural and chemical mosquito repellents
were studied in many research papers and review
papers [3, 4, 7, 9, 10, 13, 12] that work to repel
mosquitoes. Mosquito repellents based on chemicals
has a remarkable safety profile, but they are toxic
against the skin and nervous system like rashes,
swelling, eye irritation, and worse problems, though
unusual including brain swelling in children,
anaphylactic shock, and low blood pressure [10, 11].
Hence, natural mosquito repellents were preferred
over chemical mosquito repellents.
http://crmb.aizeonpublishers.net/content/2013/3/crmb98-103.pdf

All of the mosquitoes coils registered and sold contain


solely synthetic pyrethroid, for example, d-allethrin, dtransallethrin and transfluthrin as active ingredients.
These coils provide a high degree of reduction in
numbers of host-seeking mosquitoes [16, 17, 5].
However, many people still dislike smell of the
mosquito coils containing synthetic pyrethroid when
they are burned; and these people also feel that the
coils may be harmful for their health. Attempts have
been made to find out new active ingredients,
especially those derived from natural plants to replace
the synthetic pyrethroid [4].
The nine potential plants namely Greater galangale
(Alpiniaga
langa),
Fingerroot
(Boesenbergia
pandurata), Turmeric (Curcuma longa), Cardamom
(Elettaria cardamomum), Neem (Azadirachta indica),
Siamese cassia (Cassia siamea), Citronella grass
(Cymbopogon
nardus),
Eucalyptus
(Eucalyptus
citriodora) and Siam weed (Eupatorium odoratum) that
expressed high degree of repellency against mosquitoes
are recommended as new active ingredients for
inclusion in mosquito coil formulations. These were
studied for their efficacy in reducing human-mosquito
contact when used in mosquito coils in urban areas of
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Nandini Rani et. al. / Curr Res Microbiol Biotechnol. 2013, 1(3): 98-103

Thailand. The potential of volatile oils extracted from


turmeric, citronella grass and hairy basil as topical
repellents against both day- and night-biting
mosquitoes has been demonstrated [13].
In the case of Citronella species, for example, the
components present in the oil are responsible for the
desirable repellent characteristics of the plant against
mosquitoes [2]. The repellent efficiency of 38 essential
oils against mosquito bites was compared, including
the species Aedes aegypti and found Citronella oil as the
most effective and provided 2 hours of repellency [14,
15]. The chemical composition of citronella oil was
studied and it was found that the crude essential oil
consists of active ingredients that markedly suppressed
the growth of several species of Aspergillus, Penicillium
and Eurotium [6]. The most active compounds among
the 16 volatiles examined in Citronella oil, consisting of
6 major constituents of the essential oil and 10 other
related monoterpenes, were citronellal and linalool.
Most of the essential oil-based repellents tend to give
short-lasting protection for less than two hours.
Citronella oil has demonstrated good efficacy against
44 mosquitoes in concentrations ranging from 0.05 %
to 15 % (w/v) alone or in combination with other
natural or commercial insect repellent products [1].
The characteristic of the oil is due to the presence of
four main components, citronellal, eugenol, geraniol
and limonene [8, 11]. A coil product preparation using
sawdust, rice husk and corncob based fillers along with
herbal oils and herbal powders indicating towards the
improvisation of mosquito repellents by supplementing
it with natural fillers and binders have been devised
[10].
The present study was carried out to evaluate
repellency of mosquito cakes derived from plants. An
attempt has been made to develop a Citronella based
herbal mosquito repellent cake which is more effective,
cheap and keeps environment pleasant and health
friendly using different binders such as sawdust,
coconut shell powder, Neem powder etc.

MATERIALS AND METHODS


Collection of Citronella leaf remains left after steam
distillation
Citronella (Cymbopogon winterianus) leaf remains were
collected from Pharmaceutical Medicinal Plant garden,
BIT Mesra, Ranchi, India after steam distillation from
steam distillation plant. Bundles of leaves were cut into
small pieces with the help of sterile sharp scissors.
Preparation of Citronella leaf cakes
Cut leaf pieces were grounded into paste using
electrical grinder by adding distilled water. 50-100g of
Citronella leaf paste was taken and was plated
accordingly (Fig 1). Wet weights of cakes were taken.
For the determination of dry weight cakes were
allowed to dry in sun for 24 hrs and dry weight was
taken.

http://crmb.aizeonpublishers.net/content/2013/3/crmb98-103.pdf

Figure 1. Preparation of Citronella leaf cake and drying

Formulation of cakes using different natural


binders
The different natural binders (500g each) were
purchased commercially from local vendors. Different
natural binders used were wood powder, potato starch,
corn starch, coconut shell powder, neem powder and
cow dung. The same procedure was followed for
different combinations using different natural binders
for the preparation of cakes.
Supplementation of Citronella leaf paste with
binders and impregnation with oil
Each cake was prepared as 20% (w/w) binder (i.e.
wood powder, coconut shell powder, Neem powder,
cow dung, cornstarch, potato starch) in each cake
formulation, whereas the citronella leaf paste (80%) in
all cake formulations was same. As a result, a total of six
cake formulations comprising different binders with
Citronella leaf as active ingredients were prepared for
testing. Citronella leaf cakes with no supplementation
were used as reference cake. Wet weight and dry
weight of each cake were taken after 24 hours of
drying. Simultaneously, different concentrations of
citronella oil such as 5%, 10%, 15% using methanol as
a carrier alcohol was evenly sprayed on different
combinations of cakes. Ten cakes (replicates) were
prepared from each combination.
Evaluation of mosquito repellent activity
For investigating mosquito repellent activity the
prepared cakes were checked for its flammability,
burning efficiency with respect to burning time and
eventually its effective repellent activity. Flammability
test of these cakes were conducted to check its
consistent combustibility.
Further the time taken to burn the cake, smoke
produced and its causal effect such as irritation,
coughing, tears were observed and recorded. Ash
produced by cakes were weighed and recorded. The
cakes were burnt in selected mosquito prone areas in
the evening and night period such as bushes, shrubs,
laboratory corners, department premises and cafeteria.

RESULTS AND DISCUSSION


In the present study we aimed to formulate a natural,
herbal mosquito repellent based on citronella leaves
which are a natural source of essential oils. Among the
plant families with promising essential oils used as
mosquito repellents, Cymbopogon spp., Ocimum spp.
and Eucalyptus spp. are the most cited [12]. Individual
compounds present in these mixtures with high
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Nandini Rani et. al. / Curr Res Microbiol Biotechnol. 2013, 1(3): 98-103

repellent activity include -pinene, limonene,


citronellol, citronellal, camphor and thymol. Still,
synthetic chemicals are still more frequently used as
repellents than essential oils, these natural products
have the potential to provide efficient, and safer
repellents for humans and the environment.
Collection of plant material
In pharmaceutical medicinal plant garden, BIT Mesra,
steam distillation plant is operated for extraction of
citronella oil from Cymbopogon winterianus (Java
Citronella). After steam distillation the left out leaves
were collected for this study and it was selected as the
base for mosquito repellent citronella leaf cakes.
Approximately, three kg of Citronella leaf remains were

collected from steam distillation plant and were finely


grounded.
Preparation of Citronella leaf cakes in combination
with natural binders
For the preparation of mosquito repellent coils,
different fillers can be used, however, in this study
natural binders were utilized [10, 4]. Cow dung based
formulation along with optimized ingredients like
Neem, Tulsi, rice; sawdust etc. has also been reported
[4]. Citronella leaf cakes were prepared by plating 60g
of leaf paste in a sterile petri dish. Wet weight of cakes
and dry weights (after 24 hours) of prepared cakes
were recorded (Table 1 and Fig 2). A significant amount
of water loss is recorded i.e. 53.57g in all combinations.

Table 1. Average wet and dry weights of Citronella leaf cakes


S.
No
1
2
3
4
5
6
7

Name of the sample


Citronella Leaf Paste
Leaf Paste + 20% Wood powder
Leaf Paste + 20% Neem powder
Leaf Paste + 20% Potato starch
Leaf Paste + 20% Corn starch
Leaf Paste + 20% Cow dung
Leaf paste + 20% Coconut shell
powder

Average
wet
weight* (g)
54.48
65.03
74.68
70.63
71.35
76.69

Average
dry weight*
(g)
7.53
12.64
16.14
18.82
22.00
14.88

63.16

9.00

Amount
lost (g)
46.95
52.39
58.54
51.81
49.35
61.81
54.16
Average =
53.57

*The wet and dry weights are average of 10 replicates

Table 2. Parameters to check flammability of citronella leaf cakes


Name of the sample

Dry
weight
(g)

Ash
weight
(g)

Time
taken to
burn
(Minutes)

Residual
(%)

Irritation

Remarks

Citronella leaf

7.43

3.23

12

43.47

Less Irritation

Fully burnt

Citronella + 20% wood powder


Citronella + 20% coconut shell
powder

13.59

2.9

21

21.34

Less Irritation

Fully burnt

9.5

2.4

25.26

Less Irritation

Fully burnt

Citronella + 20% Neem powder

21.29

5.27

22

24.75

Less Irritation

Fully burnt

Citronella + 20% cowdung

13.48

5.22

19

38.72

High Irritation

Fully burnt

Citronella + 20% corn starch

21.17

9.85

26

46.53

No Irritation

Citronella + potato starch

21.21

10.25

36

48.33

Very less
Irritation

S.
No.

These natural binders are easily available and can be


purchased. It provides excellent binding to all the
ingredients and holds it strongly together. Suitable
binder is the one which gives slow and prolonged
burning along with uniform binding ability. Neem
powder cake and wood powder cake proved efficient
candidates for mosquito repellent activity based on
Citronella leaves when added in 20% concentration.
Cow dung based cake produced more smoke and thus is
a potential antioxidant which is in accordance with the
result [4].
Evaluation of Citronella leaf cakes
The efficacy of prepared citronella leaf cakes were
evaluated on three different parameters such as
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Not burnt
completely
Not burnt
completely

flammability, burning time and mosquito repellency


test.
a) Flammability test and burning time
To observe the flammability of the cakes, the cakes
were burnt using candles (Fig 3). The quantity of ash,
irritation produced by different combination of cakes
and the time taken to burn completely were recorded
(Table 2). The ash weights of each combination and
their burning time were recorded. Depending upon ash
weight and dry weight, residual percentage was
calculated. Residual percentage is calculated by:
Residual (%) = Ash weight / Dry weight X 100

100

Nandini Rani et. al. / Curr Res Microbiol Biotechnol. 2013, 1(3): 98-103

For a good and consistently burning mosquito repellent


cake, it is necessary that the cake should be burnt
completely, producing low smoke and irritation and
less residual percentage. A high residual percentage
suggests incomplete burning of cakes and hence two
combinations gave highest residual percentage i.e. corn
flour and potato starch (46.53% and 48.33%
respectively). Also, the burning time of cakes suggested
that the cakes with starchy combinations took much
time to burn (26 minutes corn flour and 36 minutespotato starch) and thus increased their residual
percentage. Subsequently, wood powder, coconut shell
powder and neem powder combinations took less time
to burn giving less residual percentage. Thus, it can be
suggested that the neem powder and wood powder
combinations were good for making mosquito repellent
cakes as the burning time and residual percentage and
irritation caused were very less as compared to other
combinations. On the other hand, coconut shell powder,
irrespective of giving a lesser residual percentage burnt
in a very short period of time i.e. 6 minutes which
cannot be selected for formulation of mosquito
repellent cakes. Hence, the flammability test and
burning time gave two best combinations for
formulating mosquito repellent cakes i.e. wood powder
and neem powder, wood powder does not have any
medicinal property but is easily available. However,
neem powder cake is the best option, being medicinally
active and its easy availability.

Figure 2. Different combination of Citronella leaf cake

Figure 3. Burning of Citronella leaf cake and its ash


Table 3. Mosquito repellency test in different areas of
Department of Biotechnology
S. No.
1
2
3
4

Areas
Laboratory
corners
Department
premises
Supervisor's
room
Cafeteria

Reports given by
people
Smoke caused
irritation
Mosquitoes escaped
Mosquitoes moved
outside the room
Mosquitoes escaped

Remarks
Mosquito repelled
Mosquito repelled
Mosquito repelled
Mosquito repelled

Table 4. Parameters to check mosquito repellency of citronella leaf cakes with different concentrations of citronella oil
S. No.

Name of the
sample

Citronella leaf

Citronella leaf
+ 20% Wood
powder

Citronella leaf
+ 20% Potato
starch

Citronella leaf
+ 20% Corn
starch

Citronella leaf
+ 20% Cow
dung

Citronella leaf
+ 20% Coconut
shell powder

Citronella leaf
+ 20% Neem
powder

Concentration
of citronella
oil (%)

Dry
weight
(g)

Ash
weight
(g)

0
5
10
15
0
5
10
15
0
5
10
15
0
5
10
15
0
5
10
15
0
5
10
15
0
5
10
15

7.1
7.87
7.83
6.98
15.22
13.73
13.96
14.41
18.07
19.39
21.21
20.26
25.7
26.09
24.06
19.79
12.16
16.85
14.1
14.48
9.26
9.39
8.86
8.69
16.2
16.35
15.72
16.54

3.27
3.15
3.05
3.1
3.69
4.28
3.99
5.11
9.9
10.25
10.54
10.78
9.5
9.95
10.01
9.5
5.69
7.02
6.58
6.57
3.85
3.29
3.2
3.19
3.91
5.49
5.16
4.13

http://crmb.aizeonpublishers.net/content/2013/3/crmb98-103.pdf

Time
taken
to burn
(mins)
20
15
18
15
20
21
18
19
45
50
40
45
50
45
44
40
25
20
22
21
18
15
20
20
30
25
20
25

Residual
(%)

Irritation

Remarks

46.05
40.02
38.95
44.4
24.24
31.17
28.58
35.46
54.7
52.8
49.6
53.2
36.9
38.13
41.6
48.0
46.7
41.6
46.6
45.37
41.5
35.03
36.1
36.7
24.1
33.5
32.8
24.96

No
No
No
No
No
No
No
No
No
No
No
No
Less
Less
Less
Less
High
High
High
High
Less
Less
Less
Less
Less
Less
Less
Less

No mosquitoes while burning


No mosquitoes while burning
No mosquitoes while burning
No mosquitoes while burning
No mosquitoes while burning
No mosquitoes while burning
No mosquitoes while burning
No mosquitoes while burning
Not burnt completely
Not burnt completely
Not burnt completely
Not burnt completely
Not burnt completely
Not burnt completely
Not burnt completely
Not burnt completely
No mosquitoes but irritating
No mosquitoes but irritating
No mosquitoes but irritating
No mosquitoes but irritating
2-3 mosquitoes were flying
2-3 mosquitoes were flying
2-3 mosquitoes were flying
2-3 mosquitoes were flying
No mosquitoes after 1 hour
No mosquitoes after 1 hour
No mosquitoes after 1 hour
No mosquitoes after 1 hour

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b) Mosquito repellency test


Mosquito repellency test was done by simply selecting
the mosquito prone areas in the evening and night
period such as bushes, shrubs, laboratory corners, and
cafeteria. The public remarks were noted down after
allowing them to burn the cakes and checking if the
mosquitoes are present or escaping away from the
burning cakes (Table 3). Also, the mosquito repellency
was checked by addition of citronella oil mixed with
carrier alcohol (methanol) in different concentration
i.e. 5%, 10%, 15% (Table 4).
Kim et.al (2005) evaluated repellent efficacies of two
medicinally active components and two natural aroma
compounds citronella and citronellal, against
mosquitoes, Culex pipiens pallens, was evaluated both in
field and in vitro. The experiment conducted in vitro
was carried out with hand bands impregnated with
30% citronella extract, 15% citronella extract and 30%
citronellal extract, and with bands impregnated 30%
citronella in field. They analyzed the data by the means
of counting numbers bitten by mosquitoes per unit
time, namely human bait method and percentage
repellencies were calculated and statistically confirmed
by t-test compared between control group and each
experimental group. Based on the above study,
different concentrations of Citronella oil (5%, 10% and
15%) mixed with methanol in all the different
combinations were tried. 10% addition of Citronella oil
in Neem powder mosquito cakes was found to be the
best alternative for repellency activity.
Citronella grass for their efficacy in reducing humanmosquito contact when used in mosquito coils has been
studied [12]. They reported that mosquito coil
containing leaves of citronella grass showed highest
efficacy whereas that containing rhizomes of turmeric
was least effective. Mosquitoes caught in their study
included 12 species belonging to five genera (Aedes,
Anopheles, Armigeres, Culex and Mansonia), but Cx.
vishnui, Culex gelidus and Cx. quinquefasciatus were
most predominant species.
The best combination of citronella leaf cake was with
neem powder showing less residual percentage of
24.1%, less irritating and average burning time of 21
minutes. Similarly, combination of citronella leaf cake
with wood powder also showed low residual
percentage of 24.2%, no irritation and an average
burning time of 20 minutes. The effectiveness of both
the combination was further increased by adding 10%
citronella oil. Neem powder with citronella leaf as a
base is the best option as it has antibacterial and other
medicinal properties. Instead nascent citronella leaf
cakes can also be used as a mosquito repellent. Thus, a
good attempt was made in formulating citronella leaf
based herbal mosquito repellent cake comprising Neem
powder impregnated with 10% citronella oil.
On the basis of public remarks and different areas
tested for mosquito repellency, neem powder cake with
0%, and 15% citronella oil addition gave less residual
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percentage of 4.1% and 24.96 % and took 30 minutes


and 25 minutes for complete burning, respectively. It
showed less irritation and was able to repel mosquitoes
for more than one hour. Similarly, wood powder cake
gave less residual percentage of 24.24% and 28.58%
with 0% and 10% citronella oil addition and showed
burning time of 20 and 18 minutes, respectively. On the
other hand, cakes with starch as a components showed
a high residual percentage (potato starch with 0%
Citronella oil has a residual percentage of 54.7%, while
that of corn starch having 15% oil concentration
showed residual percentage of 48 % with an average
burning time of 40 minutes). On the other hand,
nascent Citronella leaf cake having high residual
percentage of 46.05%, showing no irritation has a
tendency to repel mosquitoes away from the place.
Thus, it can be concluded from the mosquito repellency
test that neem powder and wood powder cakes with
10% citronella oil addition showed lesser residual
percentage with average burning time of 20 minutes
and can be considered as good mosquito repellent
cakes because of its consistent burning ability with less
irritating smoke and low residual percentage and high
mosquito repellent ability.

CONCLUSION
Currently, the use of synthetic chemicals to control
insects and arthropods raises several concerns related
to environment and human health. An alternative is to
use natural products that possess good efficacy and are
environmentally friendly. Among those chemicals,
essential oils from plants belonging to several species
have been extensively tested to assess their repellent
properties as a valuable natural resource. Citronella
(Cymbopogon winterianus) oil is the essential oil whose
repellent activities have been demonstrated, as well as
the importance of its leaf remains which are generally
thrown as waste are the main focus of this study.
Citronella leaf remains in the form of cakes is the first
and preliminary report. In addition, the use of other
natural products in the mixture, such as binders, could
increase the protection time, potentiating the repellent
effect of some essential oils. On the basis of results
obtained it can be concluded that the citronella leaf
cakes with neem powder as a binder and impregnated
with 10% citronella oil having less residual percentage,
less burning time and irritation is able to repel
mosquitoes effectively is the most suitable mosquito
repellent. This report is the preliminary work done
using Citronella leaf cakes as herbal mosquito
repellents using natural binders. However, further
extensive study by collecting specific number of
mosquitoes in a glass chamber covered with cloth sieve
and exposing them to the smoke generated by the
herbal product with the varying concentrations and
recording of mortality time and comparing with
chemical based formula in the artificial mosquito coils
need to be performed.

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