Available via license: CC BY-NC 3.0
Content may be subject to copyright.
409
INTRODUCTION
Pediculosis capitis (head lice infestation) is the most com-
mon type of pediculosis [1], an ectoparasitic disease in hu-
mans [2]. It is caused by the infestation of head lice (Pediculus
humanus capitis) [3], a 6-legged, blood sucking, wingless insect
that thrives on human scalp [4] (Fig. 1). Head lice are com-
mon among children aged 3-11 years, which can be transmit-
ted through either direct (head-to-head) contact with the in-
fested person or indirectly through personal items, such as
combs, hats, scarfs, beddings and towels [5]. Pediculosis capi-
tis presents a public health problem [6] as it can cause social
stigma, embarrassment, low self-esteem, loss of productivity,
and frustration among afflicted individuals [7]. It is not gener-
ally associated with morbidity [7] and is not currently consid-
ered to be vectors for human pathogens [8], but secondary
bacterial infections may occur as a result of subsequent skin
excoriation due to sensitization to louse saliva [6].
A number of both over-the-counter and prescription formula-
tions are available for the treatment of pediculosis capitis. These
includes permethrin, pyrethrin, malathion, and lindane. How-
ever, their use to control pediculosis globally has been ham-
pered due to the growing issues concerning the aforementioned
treatments [7]. There have been records of resistance in many
parts of the world due to a large treatment selection pressure in-
duced by conventional pediculicides. Resistance contributes to
treatment failures, which may result in chronic infestations [9].
In addition, these chemicals are neurotoxic [10] and cause pru-
ritus, erythema, and edema [11]. With the associated treatment
failures, drug resistance, and side effects of the currently avail-
able pediculicides, the search for safe and effective alternative
pediculicidal agents are comprehensively conducted.
ISSN (Print) 0023-4001
ISSN (Online) 1738-0006
Korean J Parasitol Vol. 55, No. 4: 409-416, August 2017
https://doi.org/10.3347/kjp.2017.55.4.409
▣ ORIGINAL ARTICLE
•Received 11 May 2017, revised 16 July 2017, accepted 19 July 2017.
*Corresponding author (ecarollado@up.edu.ph)
© 2017, Korean Society for Parasitology and Tropical Medicine
This is an Open Access article distributed under the terms of the Creative Commons
Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0)
which permits unrestricted non-commercial use, distribution, and reproduction in any
medium, provided the original work is properly cited.
Safety, Efficacy, and Physicochemical Characterization of
Tinospora crispa Ointment: A Community-Based
Formulation against Pediculus humanus capitis
Gerwin Louis Tapan Dela Torre1, Kerstin Mariae Gonzales Ponsaran1, Angelica Louise Dela Peña de Guzman1,
Richelle Ann Mallapre Manalo1, Erna Custodio Arollado1,2,
*
1
Institute of Pharmaceutical Sciences, National Institutes of Health, University of the Philippines Manila, Ermita, Manila 1000, the Philippines;
2
Department of Pharmacy, College of Pharmacy, University of the Philippines Manila, Ermita, Manila 1000, the Philippines
Abstract:
The high prevalence of pediculosis capitis, commonly known as head lice (Pediculus humanus capitis) infesta-
tion, has led to the preparation of a community-based pediculicidal ointment, which is made of common household items
and the extract of Tinospora crispa stem. The present study aimed to evaluate the safety, efficacy, and physicochemical
characteristics of the T. crispa pediculicidal ointment. The physicochemical properties of the ointment were characterized,
and safety was determined using acute dermal irritation test (OECD 404), while the efficacy was assessed using an in vi-
tro pediculicidal assay. Furthermore, the chemical compounds present in T. crispa were identified using liquid-liquid ex-
traction followed by ultra-performance liquid chromatography quadruple time-of-flight mass spectrometric (UPLC-qTOF/
MS) analysis. The community-based ointment formulation was light yellow in color, homogeneous, smooth, with distinct
aromatic odor and pH of 6.92± 0.09. It has spreadability value of 15.04 ± 0.98 g·cm/sec and has thixotropic behavior. It
was also found to be non-irritant, with a primary irritation index value of 0.15. Moreover, it was comparable to the pedicu-
licidal activity of the positive control Kwell
®
, a commercially available 1% permethrin shampoo (P> 0.05), and was signifi-
cantly different to the activity of the negative control ointment, a mixture of palm oil and candle wax (P<0.05). These find-
ings suggested that the community-based T. crispa pediculicidal ointment is safe and effective, having acceptable physi-
cochemical characteristics. Its activity can be attributed to the presence of compounds moupinamide and physalin I.
Key words:
Pediculus humanus capitis, Tinospora crispa, dermal irritation, ointment, pediculicidal assay
410
Korean J Parasitol
Vol. 55, No. 4: 409-416, August 2017
In the Philippines, the burden of pediculosis capitis among
schoolchildren was considered high at 54.7% during 1986
[12]. The disease burden increased significantly to 84% in the
year 2000, and in 2012, it was named as the second highest
medical problem among schoolchildren after tooth decay [13].
At the present time, the growing problem on pediculosis capi-
tis still persists. In this aspect, communities have resorted to
formulate their own pediculicidal treatment to resolve the
high prevalence of the disease. Community-based pediculici-
dal ointment formulation was documented by the Philippine
Institute of Traditional and Alternative Health Care (PITAHC)
utilizing common household items and the extract of Tinospo-
ra crispa stem as the active constituent [14]. The selection of T.
crispa may be attributed to its wide distribution throughout
the Philippines [15] and its traditional use as an anti-parasitic
agent on humans and domestic animals [16].
The aim of this work was to evaluate the safety and efficacy
of the community-based ointment preparation containing T.
crispa extract against head lice, and characterize its physico-
chemical properties.
MATERIALS AND METHODS
Plant sample
Fresh T. crispa stems were collected from the town of Alabat,
in the province of Quezon, Philippines. The plant was identi-
fied to be T. crispa by the National Museum of the Philippines
- Botany Division and was given a voucher specimen no. 16-
04-352.
Preparation of T. crispa ointment
The fresh stems of T. crispa were garbled and washed with
water to remove any clinging dirt and insect debris. The prepa-
ration of the ointment followed a community-based formula
[14]. The stems were chopped into small pieces. In a pot, a
cup of the chopped stems, together with a cup of palm oil,
were boiled for 15 min for the extraction of active constituents
from the stem. The oil extract was then filtered, and 1 white,
unscented, pulverized candle (diameter–1 cm; length-14.5
cm) was incorporated to the oil extract through continuous
stirring under the temperature range of 50-60˚C. It was then
transferred on opaque ointment jars, after the candle has com-
pletely melted.
Characterization of T. crispa ointment
The T. crispa ointment was subjected to physicochemical
tests of homogeneity, color, odor and pH. Spreadability mea-
surement was performed using the “slip” and “drag” character-
istics of the ointment [17]. Rheological behavior was deter-
mined using Brookfield Viscometer (Spindle – LV3) at 25˚C.
Experimental animals
Adult healthy albino rabbits of both sexes (8-10 weeks old;
1.5-2.0 kg) were procured from the St. Luke’s Medical Center,
Research & Biotechnology Division (Quezon City, Philip-
pines). The rabbits were housed individually at the animal fa-
cility of the National Institutes of Health, maintained in a con-
trolled room temperature (23-25˚C) under 12 hr/12 hr light/
dark cycle, with commercial rabbit pellet and water supplied
ad libitum. Acclimatization was done 1 week before the der-
mal irritation test. The experimental procedure was approved
by the Institutional Animal Care and Use Committee (IACUC)
of the University of the Philippines Manila (IACUC protocol
no. 2016-022).
Dermal irritation test
Acute dermal irritation study was carried out using the
OECD guideline 404 [18] with some modifications. Four rab-
bits (2 male and 2 female) were used in the study. Six areas of
approximately 6 cm2, 3 on each side of the dorsal area of each
rabbit, were shaved 24 hr before the test. The shaved areas on
the left side were applied with 0.5 g of T. crispa ointment using
a sterile cotton swab and held in place with a gauze patch and
non-irritating tape. The shaved areas on the right side served as
the control and received 0.5 g of the negative control ointment
Fig. 1. Microscopic picture of an adult Pediculus humanus capitis
(head louse).
Dela Torre et al.: Community-based Tinospora crispa pediculicidal ointment 411
(palm oil and pulverized candle only). After 4 hr, the gauze
patches were removed and the shaved areas were rinsed with
distilled water. The test areas were examined at approximately
1, 24, 48, and 72 hr after the removal of gauze for evidences of
primary irritation and scored according to the scale used in the
OECD guideline 404 (Table 1).
Primary irritation index (PII) was calculated by dividing the
sum of erythema and edema scores, with the product of the
number of test sites and the number of grading intervals [19].
It was then classified according to Draize method of classifica-
tion using PII scoring as non-irritant (if PII<0.5), slightly irri-
tant (if PII<2), moderately irritant (if PII <2-5), and severely
irritant (if PII >5) [20].
Ethical Consideration
The study was reviewed and approved by the University of
the Philippines Manila Review and Ethics Board (UPMREB)
and was given the study code no. UPMREB 2016-277-01. Prior
to screening and collection of head lice, verbal assent was ob-
tained from the children and informed written consent was
secured from their parents or guardians. All children and their
families were given an over-the-counter 1% permethrin sham-
poo (Kwell®) after the study.
Screening of schoolchildren with head lice
Schoolchildren aged 5-11, of either sex around the 5th Dis-
trict of Manila, were screened for their possible inclusion in
the study. Screening was made by the parent or guardian of
the child. The hair was divided into 4 quadrants by a plastic
head lice comb. Next, the hair on each quadrant was combed
6 times starting from the scalp and ending at the hair tips [21].
The number of lice collected was counted. Children with at
least 5 living lice and did not use any form of pediculicidal
agents in the previous month were included in the study.
Head lice collection
Adult head lice were collected from the screened schoolchil-
dren. The parents or guardians extracted the head lice at the
day of the researcher’s visit to ensure the viability of the in-
sects. Head lice were placed over a plastic container covered
with filter paper, moistened with distilled water to prevent
death by desiccation. The collected head lice were transported
to the Institute of Pharmaceutical Sciences’ laboratory for the
in vitro assay. Only fully active and intact adult head lice with
the size range of 2-3 mm long were used, irrespective of sex.
Within an hour after the collection, the assay was performed.
In vitro pediculicidal assay
The T. crispa ointment was tested for its pediculicidal activity
along with the negative control ointment (palm oil and pul-
verized candle only) and positive control (1% permethrin
shampoo, Kwell®). A total of 252 head lice were used in the
study (n =84 per group). The analyses were tested in batches
of 84 (n =28 per group) at room temperature (25 ±3˚C). Petri
dishes lined with filter paper were prepared. Head lice were
then placed on the filter paper together with some hair
strands. To prevent the desiccation of head lice, the filter pa-
pers were moistened with distilled water. Samples were intro-
duced through direct application. Each louse received either
15 mg of T. crispa ointment, 15 mg of negative control or a
single drop of the positive control. After 20 min of exposure to
the samples, the head lice were washed with tap water to sim-
ulate treatment on an infested host. Afterwards, head lice were
placed into new petri dishes with unused moistened filter pa-
pers and were examined under a dissecting microscope. Three
pre-defined criteria for the evaluation of the pediculicidal ac-
tivity of samples were used. Lice were defined as ‘active’ if no
changes in the activity or behavior were observed post-treat-
ment. In the ‘some vital signs’ category, the reduced activity
was shown by the inability of the lice to walk in a progressive
fashion and the absence of righting reflex when rolled onto
the back, but still with peristalsis. Lastly, the complete absence
of any vital signs such as peristalsis and movement of anten-
nae or legs, even after stimulating with a single hair strand,
Table 1. Grading of skin reactions
Erythema and eschar formation Edema formation
No erythema 0No edema 0
Very slight erythema (barely perceptible) 1Very slight edema 1
Well defined erythema 2Slight edema (edges of area well defined by definite rating) 2
Moderate to severe erythema 3Moderate edema (raised approximately 1 mm) 3
Severe erythema (beef redness) to eschar formation
preventing grading of erythema
4Severe edema (raised more than 1 mm and extending beyond
area of exposure)
4
412
Korean J Parasitol
Vol. 55, No. 4: 409-416, August 2017
were defined as ‘no vital signs’. Observations were conducted
at 30, 60, 90, 120, 150, and 180 min post-treatment [22,23].
Compound identification
The solution (50 ml) obtained from the T. crispa extraction
with palm oil was extracted 3 times with equal amount of di-
methylsulfoxide (DMSO). The DMSO portion was pooled
and was diluted to 5% using distilled water. The solution was
filtered using 0.2 μm syringe filter. Palm oil underwent the
same treatment and was used as blank. The resulting filtrates
were subjected to ultra-performance liquid chromatography
quadruple time-of-flight mass spectrometric (UPLC-qTOF/
MS) analysis for the identification of compounds present [24].
UPLC-qTOF/MS analysis was performed using Waters Ac-
quityTM UPLCTM I-Class/Xevo equipped with Xevo G2-XS qTOF
mass spectrometer. Chromatographic separation was per-
formed using Waters HSS T3 C18 column (2.1×1
00 mm i.d.,
1.8 μm) maintained at 40˚C. The mobile phase was composed
of acetonitrile+0.1% formic acid (A) and water+0.1% formic
acid (B). The following gradient program was employed: 5% A
for 0.5 min, linearly increased to 95% A for 10 min, followed
by an isocratic phase for 4.5 min, then 99% A for 2.5 min and
linearly decreased to 5% A for 25 min. The flow rate 0.4 ml/
min and the injection volume was 5 μl.
The Waters Xevo G2-XS qTOF MSE was run in positive ion
mode. The capillary and cone voltages were set to 1.0 kV and
40 V, respectively, with the cone gas flow at 40 L/hr. The desol-
vation temperature was 550˚C, gas flow at 950 L/hr, and
source temperature at 120˚C. Data were collected between the
mass range of 100 and 1,200 Da, with a scan time of 0.150
sec. MS analysis was carried out to identify the compounds
present in the T. crispa extract and the data were processed us-
ing UNIFI Scientific Information System with the Traditional
Chinese Medicine Library (TCML) for identification of puta-
tive compounds present in the sample.
Statistical analysis
The data were expressed as mean±standard error of the
mean (SE). The differences between groups per observation
time were compared by 1-way analysis of variance (ANOVA)
followed by Tukey’s post-hoc test. To investigate the efficacy
between T. crispa ointment and positive control over time,
2-way mixed ANOVA was used. All statistical analyses were
performed using the Statistical Package for the Social Sciences
(SPSS, Chicago, Illinois, USA) 23.0. Results with P<0.05 were
considered statistically significant.
RESULTS
Characterization of T. crispa ointment
The community-based formulation of T. crispa ointment
was light yellow, smooth and homogenous, having a distinc-
tive aromatic odor and pH of 6.92±0.09. It has a spreadability
value of 15.04±0.98 g.cm/sec. Rheological study revealed that
t
he ointment has a thixotropic behavior (Fig. 2).
Pediculicidal activity study
The results of the in vitro pediculicidal assay are shown in
Fig. 3. It was observed that all groups displayed fluctuations in
their activities over time except for active head lice group treat-
ed with T. crispa ointment. For the active head lice group, the T.
crispa ointment treatment showed comparable activity with
the positive control at 30 (P=0.728), 60 (P=0.165), 90 (P=
0.214), 120 (P=0.240), 150 (P=0.096), and 180 (P=0.056)
min (Fig. 3A). On the other hand, positive control and T. crispa
ointment treatment groups demonstrated significantly differ-
ent activities with negative control at all observation times
(P<0.05). Comparable activities were also observed in the
head lice group with some vital signs treated with T. crispa
ointment relative to positive (P>0.05) and negative (P>0.05)
controls (Fig. 3B) at all time points. The mortality or the num-
ber of head lice with no vital signs treated with T. crispa oint-
ment decreased over time but displayed comparable activity
with the positive control at 30 (P=0.961), 60 (P=0.229), 90
(P=0.416), 120 (P=0.334), 150 (P=0.278), and 180 (P=
0.131) min (Fig. 3C). Significantly different activities in the
mortality of head lice were observed in positive and T. crispa
ointment treatment groups in comparison to the negative con-
trol (P<0.05). It should be noted that at 30 min observation
Fig. 2. Rheogram of Tinospora crispa ointment at 25
˚
C.
140
120
100
80
60
40
20
Shear stress (
τ
)
0 200 400 600 800 1,000 1,200 1,400
Shear rate (Y)
Dela Torre et al.: Community-based Tinospora crispa pediculicidal ointment 413
time, the percentage mortality of head lice in the positive con-
trol group is lower than that of T. crispa ointment group, which
is the highest mortality of the group at 76.80%. Two-way
mixed ANOVA was utilized to further assess the comparability
of the positive control and T. crispa ointment groups based on
time and treatment. Time (F(5,20)=0.371, P=0.863,
η
p2
=0.085) and treatment (F(1,4)=2.034, P=0.227,
η
p2= 0.337)
have no significant effect on the overall mortality of head lice,
Fig. 3. Percent activity of T. crispa ointment, positive control (1%
permethrin, Kwell
®
shampoo) and negative control ointment (palm
oil+pulverized candle) on head lice. (A) Percentage of head lice
that are active, post-treatment. (B) Percentage of head lice con-
sidered having ‘some vital signs’, post-treatment. (C) Percentage
mortality of head lice considered having ‘no vital signs’, post-
treatment. Different lower case letters in mean significantly differ-
ent, P< 0.05, whereas similar letters mean not significantly differ-
ent from each other.
100
90
80
70
60
50
40
30
20
10
0
Active head lice (%)
30 60 90 120 150 180
Time (min)
A
30
25
20
15
10
5
0
Head lice with some Vital Signs (%)
30 60 90 120 150 180
Time (min)
B
100
90
80
70
60
50
40
30
20
10
0
Head lice with No Vital Signs (%)
30 60 90 120 150 180
Time (min)
C
Fig. 4. Base peak ion chromatogram of T. crispa extract, showing 3 unique peaks as compared to palm oil (blank). (A) Moupinamide. (B)
Physalin I. (C) An unknown compound.
Tinospora crispa ointment
Positive control
Negative control
Tinospora crispa ointment
Positive control
Negative control
Tinospora crispa ointment
Positive control
Negative control
414
Korean J Parasitol
Vol. 55, No. 4: 409-416, August 2017
with the positive control (x
̅=81.35%) and T. crispa ointment
(x
̅=67.06%) performing slightly similar overall activities.
Dermal irritation test
All the rabbit’s skin areas treated with negative control oint-
ment were devoid of both erythema and edema, thus, having a
PII value of 0.00. No signs of edema but with a very slight ery-
thema on a single area of all rabbits were observed an hour after
the application of T. crispa ointment, which disappeared there-
after. It was classified as a non-irritant formulation because of
the PII value of 0.15. Additionally, the skin of the rabbits on all
areas tested were intact throughout the 72 hr study duration.
Compound identification
T. crispa oil extract was analyzed by UPLC-qTOF/MS for
compound identification, using palm oil as blank. The chro-
matogram generated is shown in Fig. 4. It was observed that
there were 3 distinct peaks in the T. crispa extract as compared
to palm oil, with retention times at 5.28, 6.66 and 8.26 min.
Using the TCML, the compound with retention time of 5.28
min and an observed mass of 313.1311 Da was identified to be
moupinamide, while the compound with retention time of
6.66 and an observed mass of 558.2083 Da was recognized to
be physalin I. Although the last compound with retention
time of 8.26 min was not identified from the TCML, it was
known to have an observed m/z of 230.2471.
DISCUSSION
Pediculosis capitis has been well-known since antiquity [25]
and hitherto, remains a public health problem worldwide
[26]. With the presence of resistance to conventional pediculi-
cidal agents [9], the combat against head lice became more
difficult. To address the problem, the search for alternative
treatments were initiated, focusing on natural products as they
can provide good pediculicidal activity and can offer a valid al-
ternative to conventional treatments [27]. In the Philippines, a
community-based ointment formulation against head lice is
currently being used. The ointment is composed of edible oil
and candle wax as bases, and T. crispa extract as active ingredi-
ent [14]. The formulation has an almost neutral pH value
range of 6.92±0.09, which is close to pH value range of the
skin (pH 4.0-7.0), and similar to the pH of a viable epidermis
(around pH 7.0) [28]. The ointment’s pH is acceptable be-
cause the use of substances with neutral pH can maintain the
physiological pH of the skin, preventing the overgrowth of mi-
croorganisms [29]. The T. crispa ointment exhibits thixotropy
at 25˚C, which means that the viscosity decreases with increas-
ing rate of shear. This rheological behavior is preferred due to
its low flow resistance confirming the ointment’s high spread-
ability, satisfying the ideal quality in topical application [30].
Assessment of irritation for a topical product, especially for
a community-based ointment formulation, is a significant step
in the determination of their acceptability and safety. The 0.00
PII score of the negative control ointment can be attributed to
the previous non-irritancy claims of palm oil [31] and the in-
ertness of candle wax [32]. On the other hand, the 0.15 PII
score of T. crispa ointment, depicted as very slight erythema,
may be due to various compounds extracted from the plant.
The data gathered revealed that T. crispa extract has a negligible
skin irritancy effect, inferring that the community-based oint-
ment formulation of T. crispa is safe to be used topically.
The general activity of pediculicides occurs by penetrating
the cuticle barrier on the skin of the lice and then disrupting
the sodium channel in the nerve membrane, thereby causing
delayed repolarization leading to impairment of the nerve re-
sponsible for breathing [33]. As a result, lice experience suffo-
cation and expire. Permethrin, is one of the most extensively
used pediculicide available in the market due to its great effi-
cacy and wide margin of safety [34]. In the experiment, a small
percentage of the lice treated with permethrin were still active
after 180 min observation period. This may be attributed to
lice that developed resistance against pyrethroids (i.e. perme-
thrin). It is caused by a substitution of an amino acid in a pro-
tein responsible for “knockdown resistance”, that results to in-
sensitivity of the cholinergic nerves to pediculicides [35,36].
Although the source of the lice did not use permethrin or any
pediculicides for the past month prior to collection, it may be
caused by the transfer of permethrin-resistant head lice from
another host. The prevalent occurrence of resistant head lice is
due to misdiagnosis, overuse and improper use of pediculi-
cides [27,37]. On the other hand, presence of inactive head
lice in the group treated with palm oil was noted. Although it
is anticipated for palm oil not to have any effect on the mor-
tality of the lice, the presence of inactive head lice may be
caused by the hydrophobic nature of the oil, which was re-
ported to work by occluding the respiratory spiracle of the
head louse [35]. Several papers have also reported that mayon-
naise, petroleum jelly and olive oil are capable of causing mor-
tality in head lice but only supplies transient period of stasis
Dela Torre et al.: Community-based Tinospora crispa pediculicidal ointment 415
[22,36]. However, over 180 min period, it was observed that
the percentage of head lice with no vital signs decreased and
so it may also be due to a period of temporary stasis called
“sham death” as insects (i.e., head louse) exhibit this ability
when exposed to chemicals for less than 6 hr [22,38]. Head
lice utilize their anatomical structure and physiologic charac-
teristics to exhibit this ability as a defense mechanism when
exposed to either too much water or to various chemicals that
cause stupor, in a period insufficient to kill them [39,40]. This
phenomenon is characterized by a momentary period where
in head lice are thought to be inactive due to the absence of
their gut and limb movements, but after some time they re-
commence and resume, respectively [39]. Fluctuations in the
activities seen in all treatment groups may also be attributed to
sham death.
It was confirmed that the community-based ointment for-
mulation of T. crispa is an effective pediculicide, with the high-
est percent mortality (78.60%) at 30 min post-treatment, and
is comparable to the commercially available permethrin on
the basis of pediculicidal efficacy on adult head lice. Although
there was an observed decrease in the number of head lice
considered to have no vital signs over time, T. crispa ointment
still performed better than palm oil and relatively the same
with permethrin. The efficacy of T. crispa may be attributed to
the 3 compounds detected in the UPLC-qTOF/MS analysis.
The 2 compounds were identified to be moupinamide and
physalin I, and a third unknown compound with a m/z of
230.2471. The structural aspect of these compound may con-
tribute to the pediculicidal efficacy [41-43]. Moupinamide be-
longs to the class benzenoids, and its direct parent methoxy-
phenol [44]. Studies have shown that benzenoids and me-
thoxyphenol, have ability to elicit insecticidal activity, however
their mode of action were not reported [41,45]. Physalin, on
the other hand, belongs to the class of lipids [44] because of
its steroidal nature. Lipids act on the spiracle of head louse and
causes suffocation [35]. These studies provide supportive evi-
dence for moupinamide and physalin I, as the compounds re-
sponsible for the pediculicidal efficacy of T. crispa ointment.
ACKNOWLEDGMENTS
We would like to express our sincerest gratitude to the Phil-
ippine Institute of Traditional and Alternative Health Care (PI-
TAHC) for funding this project. We would also like to thank
Ms. Camille Vien D. Sayson and Mr. Patrick Gabriel G. More-
no for their technical assistance in the project.
CONFLICT OF INTEREST
The authors of this article have no issues that might lead to
a conflict of interest.
REFERENCES
1. Diaz JH. The epidemiology, diagnosis, management, and pre-
vention of ectoparasitic diseases in travelers. J Travel Med 2006;
13: 100-111.
2. McNair CM. Ectoparasites of medical and veterinary impor-
tance: drug resistance and the need for alternative control meth-
ods. J Pharm Pharmacol 2015; 67: 351-363.
3. Austin NI. Pediculosis among school children, in Owerri north
local government area of Imo State, South Eastern Nigeria. Int J
Infect Dis 2016; 45; 352-353.
4. Roberts RJ. Clinical practice. Head lice. N Engl J Med 2002; 346:
1645-1650.
5. Moshki M, Zamani-Alavijeh F, Mojadam M. Efficacy of peer edu-
cation for adopting preventive behaviors against head lice infes-
tation in female elementary school students: a randomised con-
trolled trial. PLoS One 2017; 12: e0169361.
6. Sim S, Lee IY, Lee KJ, Seo JH, Im KI, Shin MH, Yong TS. A survey
on head lice infestation in Korea (2001) and the therapeutic effi-
cacy of oral trimethoprim/sulfamethoxazole adding to lindane
shampoo. Korean J Parasitol 2003; 41: 57-61.
7. Greive K, Barnes T. In vitro comparison of four treatments which
discourage infestation by head lice. Parasitol Res 2012; 110:
1695-1699.
8. Badiaga S, Brouqui P. Human louse-transmitted infectious dis-
eases. Clin Microbiol Infect 2012; 18: 332-337.
9. Durand R, Bouvresse S, Berdjane Z, Izri A, Chosidow O, Clark
JM. Insecticide resistance in head lice: clinical, parasitological
and genetic aspects. Clin Microbiol Infect 2012; 18: 338-344.
10. Gupta A, Kaul N, Palma K. Isopropyl myristate 50% as a novel
pediculicide in the treatment of head lice. J Am Acad Dermatol
2008; 58: AB8.
11. Verma P, Namdeo. Treatment of pediculosis capitis. Indian J
Dermatol 2015; 60: 238-247.
12. Cabrera BD, Rampal L. A study on the problem of Pediculus
humanus capitis infestation among primary school children in
selected areas in the Philippines. Acta Med Philipp 1986; 22:
1-4.
13. Heading Off Lice “Getting Rid of the Itchiness”. cited 2017 Apr 5.
Available from: http://m.thefilipinodoctor.com/health-care/
health-articles/499/.
14. Philippine Institute of Traditional and Alternative Health Care.
cited 2016 Jun 9. Available from: https://pitahc.wordpress.com/
2016/05/12/paggawa-ng-makabuhay-ointment-para-sa-galis-
416
Korean J Parasitol
Vol. 55, No. 4: 409-416, August 2017
aso/.
15. Ragasa CY, Cruz MC, Gula R, Rideout JA. Clerodane diterpenes
from Tinospora rumphii. J Nat Prod 2000; 63: 509-511.
16. Ahmad W, Jantan I, Bukhari SNA. Tinospora crispa (L.) Hook. f. &
Thomson: a review of its ethnobotanical, phytochemical, and
pharmacological aspects. Front Pharmacol 2016; 7: 59.
17. Shukr MH, Metwally GF. Evaluation of topical gel bases formu-
lated with various essential oils for antibacterial activity against
methicillin-resistant Staphylococcus aureus. Trop J Pharm Res
2013; 12: 877-884.
18. Organization for Economic Cooperation and Development
(OECD). Guideline for the testing of chemicals, 404: acute der-
mal irritation/corrosion. 2002.
19. Soiefer AI, Rauckman EJ. Toxicity associated with single chemi-
cal exposures. In Jabcoson-Kram D, Keller KA eds, Toxicology
Testing Handbook Principles, Applications, and Data Applica-
tions. New York, USA. Marcel Dekker. 2001, pp 28.
20. Lehman AJ. Appraisal of chemicals in foods, drugs and cosmet-
ics. The Association of Food and Drugs Officials of the United
States. 1959, pp 46.
21. Wolf L, Eertmans F, Wolf D, Rossel B, Adriaens E. Efficacy and
safety of a mineral oi-based head lice shampoo: a randomized,
controlled, investigator-blinded, comparative study. PLoS One
2016; 11: e0156853.
22. Heukelbach J, Canyon DV, Oliveira FA, Muller R, Speare R. In vi-
tro efficacy of over-the-counter botanical pediculicides against
head louse Pediculus humanus var capitis based on a stringent
standard for mortality assessment. Med Vet Entomol 2008; 22:
264-272.
23. Oladimeji FA, Orafidiya OO, Oqunniyi TAB, Adewunmi TA. Pe-
diculicidal and scabicidal properties Lippia multiflora essential
oil. J Ethnopharmacol 2000; 72: 305-311.
24. Dela Torre, GLT, Arollado, EC, Atienza AA, Manalo RAM. Evalu-
ation of antioxidant capacity and identification of bioactive
compounds of crude methanol extracts of Caesalpinia pulcherri-
ma (L.) Swartz. Indian J Pharm Sci 2017; 79: 112-123.
25. Davarpanah MA, Kazerouni A, Rahmati H, Neirami RN,
Bakhtiary H, Sadeghi M. The prevalence of pediculus capitis
among the middle schoolchildren in Fars province, southern
Iran. Caspian J Intern Med 2013; 4: 607-610.
26. Dalimi A, Mahdavi Poor B, Rashedi J, Asgharzadeh M, Abdolal-
izadeh J. Is hair lice still a public health problem? Iran J Public
Health 2016; 45: 1671-1672.
27.
Di Campli E, Di Bartolomeo S, Delli Pizzi P, Di Giulio M, Grande
R, Nostro A, Cellini L. Activity of tea tree oil and nerolidol alone
or in combination against Pediculus capitis (head lice) and its
eggs. Parasitol Res 2012; 111: 1985-1992.
28. Lambers H, Piessens S, Bloem A, Pronk H, Finkel P. Natural skin
surface pH is on average below 5, which is beneficial for its resi-
dent flora. Int J Cosmet Sci 2006; 28: 359-3370.
29. Mukhopadhyay P. Cleansers and their role in various dermato-
logical disorders. Indian J Dermatol 2011; 56: 2-6.
30.
Dantas MG, Reis SA, Damasceno CM, Rolim LA, Rolim-Neto PJ,
Carvalho FO, Quintans-Junior LJ, Almeida JR. Development and
evaluation of stability of a gel formulation containing the mono-
terpene borneol. ScientificWorldJournal 2016; 2016: 7394685.
31. Andersen FA. Final report on the safety assessment of Elaeis
guineensis (palm) oil, Elaeis guineensis (palm) kernel oil, hydro-
genated palm oil and hydrogenated palm kernel oil. Int J Toxi-
col 2000; 19: 7-28.
32. Caraccio TR, McGuigan MA. Over-the-counter products. In Dart
RC ed, Medical Toxicology. Philadelphia, USA. Lippincott Wil-
liams & Wilkins. 2004, pp 1059.
33. Connolly M. Recommended management of head lice and sca-
bies. Prescriber 2013; 24: 17-29.
34. Mac-Mary S, Messikh R, Jeudy A, Lihoreau T, Sainthillier JM,
Gabard B, Schneider C, Auderset P, Humbert P. Assessment of
the efficacy and safety of a new treatment for head lice. ISRN
Dermatol 2012; 2012: 460467.
35. Paller AS, Mancini AJ. Hurwitz Clinical Pediatric Dermatology.
5th ed. Canada. Elsevier Health Sciences. 2016, pp 439.
36. Burkhart CN, Burkhart CG. Recommendation to standardize
pediculicidal and ovicidal testing for head lice (Anoplura; pedic-
ulidae). J Med Entomol 2001; 38: 127-129.
37. Burkhart CG. Relationship of treatment-resistant head lice to the
safety and efficacy of pediculicides. Mayo Clin Proc 2004; 79:
661-666.
38. Oliveira FA, Speare R, Heukelbach J. High in vitro efficacy of
Nyda L, a pediculicide containing dimeticone. J Eur Acad Der-
matol Venereol 2007; 21: 1325-1329.
39. Nuttall GHF. The biology of Pediculus humanus. Parasitology
1917; 10: 80-185.
40. Burkhart CN, Burkhart CG, Gunning WT. Scanning electron mi-
croscopy of adult head lice (Pediculus humanus capitis) with focus
on clinical implications. J Cutan Med Surg 2000; 4: 181-185.
41. Bagavan A, Rahuman AA, Kamaraj C, Elango G, Zahir AA, Jayas-
eelan C, Santhoshkumar T, Marimuthu S. Contact and fumigant
toxicity of hexane flower bud extract of Syzygium aromaticum and
its compounds against Pediculus humanus capitis (phtiraptera: pe-
diculidae). Parasitol Res 2011; 109: 1329-1340.
42. Priestley CM, Burgess IF, Williamson EM. Lethality of essential
oil constituents towards the human louse, Pediculus humanus,
and its eggs. Fitoterapia 2006; 77: 303-309.
43. Rossini C, Castillo L, González A. Plant extracts and their com-
ponents as potential control agents against human head lice.
Phytochem Rev 2008; 7: 51-63.
44. Afendi FM, Okada T, Yamazaki M, Hirai-Morita A, Nakamura K,
Ikeda S, Takahashi H, Ataf-Ul-Amin M, Darusman LK, Saito K,
Kanaya S. KNApSAcK family databases: integrated metabolite-
plant species databases for multifaceted plant research. Plant
Cell Physiol 2012; 53: 1-12.
45. Silva CGV, Zago HB, Hugo JGS, da Camara CAG. Composition
and insecticidal activity of the essential oil of Croton grewioides
Baill. against Mexican bean weevil (Zabrotes subfasciatus Bohe-
man). J Essent Oil Res 2012; 20: 179-182.
