P H I L I P P I N E J O U R N A L O F

NATURAL AND SOCIAL SCIENCE

Vol. 4, No. 1, s. 2020

Atsuete (Bixa orellana L.) Seeds and Alugbati (Basella alba L.)
Stem Ethanolic Extract as Potential Hair Dye Tested via
Spectrophotometry
1John V. Bernardo, 2Hazel E. Laureta
1
Pangasinan State University, Binmaley Campus, Pangasinan, Philippines
2
PHINMA – University of Pangasinan, Dagupan City, Pangasinan, Philippines
1
bernardonhojv@gmail.com, 2hazellaureta29@yahoo.com

Abstract
Coloring agents were obtained from natural sources such as plants and animals.
The biosphere is gifted as more than 500 plant species which yield natural dyes. With the
worldwide concern over the use of eco-friendly and biodegradable materials, the use of
natural dyes has undoubtedly once again gained interest and momentum. This study aimed
to analyse the level of absorbance and (%) transmission of Atsuete seeds and Alugbati stem
dye extract through spectrophotometer as potential hair dye substitute synthetic hair dye.
Parallel group design was used in this study with three treatments groups and one control
group and was subjected to spectrophometric analysis at 200nm, 250m, and 400-550nm.
Results showed that Treatment3 is the most potential hair dye coloring agent because it
absorbs less light from ultraviolet to visible light and this may protect the hair strands from
being brittle due to light absorption of color applied in the hair stands. Further,
Treatment3 has the highest transmission among the treatment and control groups with
transmission of 6.028%-6.300% at 200-250nm respectively; and 99.46% at 400-550nm.
High Transmission property of Treatment3 can protect the hair from phototendering due
to light when applied by this dye. Generally, this study found that combination of dyes from
plant can enhance light transmission to help protect hair structure and color from sunlight
exposure and that atsuete and alugbati have a potential as hair dye. Recommendation for
home-made use for the said extract is recommended.

Keywords: Absorbance level, Alugbati stem, Atusete seeds, Percent Transmssion,

Spectrophotometric analysis

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INTRODUCTION
Coloring agents were obtained from
natural sources such as plants and animals. The
biosphere is gifted as more than 500 plant species
which yield natural dyes. Coloring agents of
these plants are derived from leaves, roots,
trunks, barks, or fruits. Natural dyes are
environmental friendly, nontoxic, non-
carcinogenic, non-allergic and biodegradable
substance, which have high regenerating,
compatibility and renewable source [1].
However, the replacement of natural dyes could
happen until the introduction of synthetic dyes.
[2]. With the advent of widely available and
cheaper synthetic dyes in 1856, the use of natural
dyes with poor to moderate wash and light
fastness declined drastically and was replaced by
more moderate to excellent color fastness
properties of synthetic materials [3].
In the research conducted in 1970, it
reported the use of eighty indophenol – which are
inorganic coloring matters [4], and indamine
synthetic dyes evaluated for their substantivity to
human hair, their fastness to light, and the
stability to storage of the dye solution. Research
of Tucker published in 1971 is one of the
research evidences that synthetic dyes are being
promoted in color/dye industry and over-
throwing natural dyes.
Despite the market competition between
natural dyes and synthetic hair dyes, natural dyes
made a way to be in demand again due to
environmental and health issues pose about by
the use of synthetic dyes. A published article in
2012 reports that the demand for natural dyes in
the modern world is increasing day by day and
back-to-nature trend is being practiced by the
developed world [5]. In fact, in 1995 Food and
Agriculture Organization (FAO) reported that
the food sector is now experiencing a trend back
towards natural colorants. This change has not
been driven by the food industry but by
consumers in developed countries who are
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concerned over possible health risks associated
with synthetic food additives [6].
Natural dyes are now a days in demand
not only in textile industry but in cosmetics,
leather, pharmaceuticals [7] food [7] [3],
handicrafts articles, drawings [3], and cotton
fabrics [1] [2]. Natural dyes from plants have
been given much interest in recent years due to
the threat and harmful effects raised by synthetic
dyes and environmental awareness created by
researchers [2]. With the worldwide concern
over the use of eco-friendly and biodegradable
materials, the use of natural dyes has
undoubtedly once again gained interest and
momentum. [7]. Today, an undeniable fact is that
there are increasing demands for natural dyes in
textile industries all over the world [3]. The
aspect of producing products without negatively
altering ecological balance, affecting both human
and environmental health, is an important focal
point to be pursued [9].
Extracted dyes from plants undergo
several testing for its effectiveness to color
materials and retain its color even for a long
period of exposure to sunlight. This property of
dyes is called as color fastness to light.
Colorfastness testing is used to determine the
loss and change of color [10]. Some dyes can
exhibit a reversible colour loss (change) on
exposure to light [11]. Colorfastness to light is an
essential quality for any dye/fabric combination
intended for sunlight exposure for a significant
length of time. One need only look at garments
or fabric that has been in a shop display window
to see that fading due to sunlight is a problem for
most fabric dyes [12].
With the interest of producing natural
dyes that has low fading, machine has been used
to study dye from plant extracts. Nearly all
natural dyes will fade badly during an exposure
to articial light, or to a much smaller dose of
daylight [13]. It is of great importance to
manufacturers of all types of materials and
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colorants that the effects of radiation, particularly
visible and ultraviolet, can be measured in some
quantitative way [Pugh].
Nowadays, dyes are being subjected to
spectrophotometric analysis. Spectrophotometric
analysis was introduced in 1941 [14] and
spectrophotometry is mainly used for the
measurement of specific substances and
aggregate parameters in water and wastewater.
However, other applications for physical
parameters like measurement or characterisation
of color had also been reported [15]. Some other
uses of spectrophotometric analysis in research
field are in biostain [16] [17], anti-oxidant [18],
spray dye [19], dye for solar cell [20], and bixa
dye for food that is tested for absorbance level;
and optical property test of biosynthesized silver
nanoparticles [21]. This study aimed to conduct
spectrophotometric analysis on the different ratio
mixture of Atsuete (Bixa orellana L.) seeds and
Alugbati (Basella alba L.) stem ethanolic extract
as potential source of hair dye.
Atsuete (Bixa orellana L.). Bixa
orellana L., a representative of Bixaceae is rich
in bixin and nor-bixin pigments. These pigments
were proved to be non-toxic, non-carcinogenic
and non-mutagenic. The pigment was found to be
a potent antioxidant as well as bactericidal
against opportunistic bacteria [Nathan]. Further,
Atsuete has been documented that it contains
important amounts of tocotrienols, tocopherols,
terpenes and flavonoids both in the seeds and in
the leaves. Vitamin E is the generic name given
to all compounds that exerts the biological
functions of a-tocoferol. Atsuete is widely
cultivated in all tropical regions around the world
[22], thus, availability is not a problem. Two
thirds of the production is commercialized as
dried seeds and the rest as colorant. Latin
America produces 60% of the total world
production, followed by Africa (27%) and Asia
(12%) [22].
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On the other hand, Alugbati (Basella
alba L.) a plant that thrives in tropical Asia,
Africa, Malaya and is found cultivated
throughout the Philippines. Basella alba fruit
with dark blue skin and deep red violet flesh is a
potential source of natural colorant. [sk reshmi].
Different studies have proved that the plant is
rich in vitamin A and vitamin C along with
flavonoids, saponins, carotenoids, many amino
acids and organic acids. Various in vivo and in
vitro studies revealed that the plants is enriched
with active substances/principles having
medicinal potential. Physical and biological
properties of Basella alba stem mucilage has
studied and reported that mucilage is mainly a
polysaccharide with pH ranging between 5.3 to
5.4, contains Dgalactose as a major
monosaccharide and exhibits slow swelling
capacity, mild antioxidant activity and no
tyrosinase inhibiting activity [24]. These two
plants will be utilized for the production and
formulation of potential hair dye that can replace
synthetic hair dye in the market.
OBJECTIVES OF THE STUDY
This study aimed to analyse the level of
absorbance and (%) transmission of Atsuete
seeds and Alugbati stem extract through
spectrophotometer.
Specifically, this study aimed to:
1. determine the absorbance and
transmission of treatment groups
and control group at 200-250nm;
2. determine the absorbance and
transmission of treatment groups
and control group at 400-550nm;
and
3. determine if there is a significant
difference exist between treatment
groups and control groups in terms
of absorbance and transmission.
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MATERIALS AND METHODS Research Design

Research Materials Table 1. The Parallel Group Design
The research materials were Atsuete Table 1 shows the research design used
(Bixa orellana L.) seeds and Alugbati in this study. This research design is known
(Basella alba L.) stem collected at the locale as Parallel Group Design which composed of
of San Jacinto and brought at National three treatment groups and one positive
Museum Botany Division for authentication. control group. Each treatment groups and
Plate 1 shows the certification of the plant control group have their three replicates.
used in this study. One kilogram (1kg) of
each plant was set aside for the extraction
process. Ratio
Other research materials such as the Treatments Atsuete Alugbati
Bremod synthetic hair dye was bought in the dye dye
cosmetics market. Spectrophotometer was Treatment 1 (1:1) 10mL 10mL
used to determine the level of absorbance Treatment 2 (1:2) 10mL 20mL
and transmission at 200-250nm and at 400- Treatment 3 (2:1) 20mL 10mL
550nm. General Procedures

a. Ethanolic extraction of Dye stuff

One kilogram of Atsuete (seeds were

macerated with 1200mL of ethanol for
three days, afterwards it was extracted
and evaporated until the pure extract is
acquired. The same process was done to
Alugbati (Basella alba L.) stem. This
was performed at Virgen Milagrosa
University Foundation San Carlos City,
Pangasinan.

Plate 1. Certificate of
Plant Authentication
Plate 2. One kilogram of Atsuete and
Alugbati and laboratory materials

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b. Collection and Stocking of dye for

treatment groups and control group

Table 2. Mixture Ratio of Atsuete and

Alugbati dye extract

Obtained dye from Atsuete and

Alugbati was prepared by dividing into
three formulation ratio shown in Table 2.
o T1 = 10mL of Atsuete dye extract +
10mL of Alugbati dye extract (1:1
ratio);
o T2 = 10mL of Atsuete dye extract +
20mL of Alugbati dye extract (1:2
ratio);
o T3 = 20mL Atsuetedye extract +
10mL Alugbati dye (2:1 ratio).
Stocking of the dyes was carried out
at San Jacinto National High School, San
Jacinto, Pangasinan (See Plate).
Plate 4. Researcher with their consultant
conducting Spectrophotometric Analysis
d. Statistical Analysis
Average mean was employed to raw
data from spectrophotometer. Average
mean data was then subjected to One
way Analysis of Variance (ANOVA) to
determine if there is a significant
difference exist between treatment
groups and control group in their
absorbance and transmission at 200 and
250nm and spectrum at 400-550nm.

Plate 3. Collecting and Stock formulation of dye per

Treatment

c. Spectrophotometric Analysis

Using Spectrophotometer, the

researchers tested the level of absorbance
and transmission of Atsuete-alugbati dye and
Bremod synthetic dye at 200nm, 250nm, and
at 400-550nm [12][25][27][28]. Effect of
light to colorfastness of dye has been tested
through light exposure that can reach up to
10 hours before the dye is rated as
‘satisfactorily’ [4].
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RESULTS AND DISCUSSIONS
Spectrophotometric Analysis of Atsuete-
alugbati dye and Bremod synthetic dye
Table 3. Mean Level of Absorbance of
Atsuete and Alugbati dye and Bremod synthetic
dye
Treatment groups
+Co
Wavelength group
T1 T2 T3 Bremod
200nm 2.7336 1.9913 1.2253 2.1316
250nm 2.063 1.986 1.168 2.112
400-550nm 3.448 1.776 0.002 2.973
Table 3 shows that at 200nm
wavelength, Treatment3 has the lowest
absorbance level followed by Treatment2 and
Bremod dye respectively. Treatment1 has the
highest absorbance level at 200nm. At 250nm,
Treatment3 has the lowest absorbance level still
followed by Treatment2 and Treatment1
respectively. Bremod dye has the highest
absorbance level at 250nm.
Further, the table shows that in visible
light wavelength at 400-500nm, Treatment3 still
has the lowest absorbance level and again
followed by Treatment2 and Bremod dye
respectively. Treatment1 has the highest
absorbance level of visible light at 400-550nm
The higher the absorbance level, the
faster the color fading and higher risk of
photodegradation [25] and phototendering [11].
Thus, the researcher implied that Treamtent3 –
combination of 20mL of Atsuete and 10mL of
Alugbati –, is the most potential dye to be used
as hair coloring agent because it absorbs less
light and will not be too sensitive to light that can
cause fading [25]. Further, the table shows that
Treatment3 has decreasing absorbance level in
increasing wavelength. Therefore, the researcher
can say that exposure to sunlight will not fade the
color produced by Treatment 3 fast. Furthermore,
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Treatment 3 can be used as coloring agent
compared to Bremod synthetic dye.
Analysis of Variance showed there is a
significant difference exist between the treatment
groups and control group in terms of absorbance
level at 200nm (F(0.05; 3,8) Fc 22.67>Ft 4.07); 250nm
(F(0.05; 3,8) Fc 13.04>Ft 4.07); spectrum at 400-550nm
(F(0.05; 3,8) Fc 4.95>Ft 4.07). Generally, Scheffe’s test
revealed that Treatment3 gives the significance.
Table 4. Mean Percent Transmission of Atsuete
and Alugbati dye and Bremod synthetic dye
+Co
Treatment groups(%)
Wavelength group
T1 T2 T3 Bremod
200nm 0.2639 1.022 6.028 0.748
250nm 1.055 1.035 6.300 0.763
400-550nm 1.383 3.171 99.46 0.732
Table 4 shows that Treatment3 has the
highest light transmission at all wavelength used
in this study. In the ultraviolet wavelength,
approximately 6% of light was transmitted. This
is highest among the experimental treatments and
most importantly higher than the synthetic dye
with transmission percentage ranging only from
0 to 0.748% at 200-250nm. Further, in the visible
light wavelength, Treatment3 was able to
transmit 99.46% of the light compared to
syntethic dye which only transmitted 0.732% of
light at 400-550nm.
The results in the table 4 further justify
that Treatment3 is the most ideal dye as coloring
agent presented in this study that can be used
rather than bremod syntethic dye. High
Transmission property of Treatment3 can protect
the hair from phototendering [13] when applied
by this dye.
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CONCLUSIONS AND
RECOMMENDATIONS
The researchers conclude the following:
1. Treatment3 is the most potential hair dye
coloring agent because it absorbs less
light from ultraviolet to visible light.
2. Treatment3 has the highest transmission
among the treatment and control groups
with transmission of 6.028%-6.300% at
200-250nm respectively; and 99.46% at
400-550nm. High Transmission
property of Treatment3 can protect the
hair from phototendering [13] when
applied by this dye.
The researchers recommend the following:
1. Atsuete and Alugbati dye extract should
be used as alternative hair dye coloring
agent.
2. Higher amount of Atsuete dye than
Alugbati dye extract should be
formulated to decrease light absorbance
and increase light transmission than can
last for long period of time in sunlight
exposure.
ACKNOWLEDGMENTS
Special thanks to the Laboratory Staff of
Virgen Milagros University Foundation for the
technical assistance given to the researchers during
the laboratory part of the study.
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