Raknam et al., IJPSR, 2020; Vol. 11(1): 146-155.

E-ISSN: 0975-8232; P-ISSN: 2320-5148

IJPSR (2020), Volume 11, Issue 1 (Research Article)

Received on 05 April 2019; received in revised form, 05 July 2019; accepted, 19 July 2019; published 01 January 2020

RUBBER SEED CLEANSING OIL FORMULATION AND ITS EFFICACY OF MAKEUP

REMOVER
*
Panithi Raknam, Sirirat Pinsuwan and Thanaporn Amnuaikit
Department of Pharmaceutical Technology and Drug Delivery System Excellence Center, Faculty of
Pharmaceutical Sciences, Prince of Songkla University, Hatyai, Songkhla 90112, Thailand.
Keywords: ABSTRACT: The objective of this study was to determine the safety
Rubber seed oil, of rubber seed oil in terms of toxicity, residual solvent and to develop
Cleansing oil, Cosmetic rubber seed cleansing oil as a makeup remover. The cytotoxicity of
application, Make-up remover rubber seed was determined on human fibroblast cell. It was not
Correspondence to Author: cytotoxic to human fibroblast cell at >1000 µg/ml. Rubber seed oil
Thanaporn Amnuaikit
was further examined for the presence of linamarin toxin and
Department of Pharmaceutical determining the solvent residual. FTIR spectrum revealed that there
Technology and Drug Delivery
was no presence of cyanide peak and, n-hexane residual was 60.35 ±
System Excellence Center, Faculty
of Pharmaceutical Sciences, Prince of 1.12 ppm. Based on these results, it was suggested that rubber seed oil
Songkla University, Hatyai, Songkhla was safe to be developed as a product for cosmetic applications. The
90112, Thailand. rubber seed cleansing oil was formulated in various compositions of
rubber seed oil (5, 10 and 15%) combined with sorbeth-30 tetraoleate
E-mail: [email protected]
(HLB 11.5) and sorbitan sesquioleate (HLB 3.7) and the ratio of
surfactant was 10, 15 and 20% respectively. The most suitable
formulation with good physical properties was obtained from 5% of
rubber seed oil, and 15% of surfactant. The makeup removal efficacy
of rubber seed cleansing oil was 89.89 ± 2.75% against the liquid
foundation.
INTRODUCTION: Cleansing oil is a cosmetic An example of synthetic oil that is often found in
formulation that can be used as a makeup remover cleansing oil is mineral oil. However, mineral oil
for daily routine. The main composition of which is obtained from distillation of petroleum
cleansing oil contains oil from two different may lack biological compounds which are essential
sources; synthetic and natural. Cleansing oil also for cosmetic application. Natural oil is a more
plays an important role as an emollient and can preferable choice to formulate as cleansing oil
easily dissolve cosmetic preparations 1, 2. Synthetic because of its antioxidant property, non-toxicity,
oil is often the choice for commercial use in and it contains biological compounds such as
cleansing oil because of its lower cost of essential fatty acids and active compounds which
production. are compatible with the skin.
QUICK RESPONSE CODE Another component of cleansing oil is a surfactant,
DOI:
10.13040/IJPSR.0975-8232.11(1).146-55 which is an ingredient that interacts with oil, facial
makeup and surface debris on the skin in a manner
that allows them to be rinsed off when they come in
The article can be accessed online on
www.ijpsr.com contact with water. The availability of different or
various sources of natural oil from which a
DOI link: http://dx.doi.org/10.13040/IJPSR.0975-8232.11(1).146-55 cleansing oil is formulated will always be

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beneficial to the cosmetic market, as it provides the applications. Subsequently, the rubber seed
consumers more options to choose from, depending cleansing oil formulation was developed, and
on their preference. The oil derived from plants, makeup removal efficacy of the developed product
which are non-toxic and contain essential fatty was compared with commercial product available
acids is more preferable for use in cosmetic in the market.
applications 3. It was reported that the rubber seed
oil extracted from Hevea brasiliensis contains a MATERIALS AND METHODS: The rubber
significantly high value of essential fatty acid. seed oil was provided on special request by Thai-
China Flavors and Fragrances Co., Ltd. Almond
Many researchers have been conducted regarding oil, jojoba oil, macadamia oil, sunflower seed oil,
the cosmetic applications of rubber seed oil. caprylic/capric triglyceride, ascorbyl palmitate,
Kittigowittana et al., 4 studied the potential source phenoxyethanol and sorbitan sesquioleate (HLB
of antioxidant oil for cosmetic application, non- 3.7) was purchased from PC drug, Thailand.
toxic, and antimicrobial property. They reported Sorbeth-30 tetraoleate (HLB 11.5), cetyl ethyl
that the major components of fatty acids found in hexanoate, vitamin E acetate, and light mineral oil
rubber seed oil were linoleic and oleic acids, which were purchased from Namsiang, Thailand.
could inhibit the scavenging radical. Lourith et al.,
5
showed that rubber seed oil is unconventional oil Physicochemical Properties Study of Rubber
for cosmetic products due to the high level of fatty Seed Oil: Physicochemical properties of rubber
acid compositions. The fatty acids in rubber seed seed oil were determined by standard methods. The
oil comprise linoleic acid, oleic acid, linolenic acid, viscosity was measured in triplicate using a cup and
palmitic acid, and stearic acid. These fatty acids are bob viscometer (Brookfield® model DV-III ultra-
widely used in cosmetic in many products such as programmable rheometer, USA) at room
hair products, face preparations. temperature (25 ± 2 °C) at a speed of 200 rpm. The
refractive index and specific gravity were
Moreover, the utilization of rubber seed oil is determined by AOAC method number 921.08 7 and
facing various challenges; for example, the toxin USP30-NF25 Chapter 401, 8, respectively. The
and solvent residual in the oil can be harmful to the chemical properties of rubber seed oil like acid,
users. The toxin in the rubber seed is linamarin and iodine, peroxide and saponification value were
cyanogenic glucoside (C10H17O6) which can be determined by USP30-NF25. 8
catalytically hydrolyzed and release hydrogen
cyanide (CHN). However, different methods such The antioxidant activity was determined by ABTS
as soaking, boiling, and drying could reduce assay, which is more related to lipid peroxidation
cyanide content in rubber seed. Organic solvents and widely applied in the antioxidant evaluation of
extraction plays a vital role in oil production the vegetable oils and expressed as trolox
because it gives good physicochemical properties equivalent antioxidant capacity (TEAC). The fatty
and high percentage yield of oil, but the solvents acid composition of rubber seed oil was determined
cannot be eliminated by manufacturing techniques. by converting all fatty acids to the corresponding
As there is no benefit from residual solvents, all fatty acid methyl esters (FAME) followed by GC
residual solvents should be removed to the analysis 5, 9. Color parameters (CIELAB), which are
maximum extent possible. L*, a* and b*, were measured by CM-700d
Spectrophotometer (KONICA MINOLTA, INC.,
Therefore, residual solvents testing become one of Japan). The L* corresponds to the light ranging
the important parts of quality control in the from white (100) to black (0) color, the a* is for the
product. Particularly, in this study, the solvent green (-) and red (+) color, whereas b* is the hue of
which was extracted from rubber seed oil was n- blue (-) and yellow (+).
hexane. According to USP30-NF25 Chapter 4676,
n-hexane is classified as a class 2 solvent, which Cytotoxicity Test of Rubber Seed Oil: The
should be limited at 290 ppm. The rubber seed oil cytotoxicity assay of rubber seed oil against skin
was analyzed to determine the toxicity and n- fibroblast cells was measured using a
hexane residual, before use in cosmetic sulforhodamine B (SRB) assay. This assay relies on

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the property of SRB, a bright-pink aminoxanthene at 1.0 ml/min; the split mode was set at 1: 10 and
dye, which binds to basic amino acid residues in the temperature were set at 250 C, for the
protein components of cells under mildly acidic detection. The determination was carried out at an
conditions, and then can be extracted under basic ion source temperature of 200 C and ionization
conditions. The amount of dye extracted from energy of 70 eV.
stained cells is directly proportional to the cells
mass. Briefly, the cells were seeded onto a 96-well Oxidative Stability Study of Rubber Seed Oil:
plate at the density of 3 × 103 cells/well. The cells Accelerated oxidative stability study of samples
were incubated at 37 C, 5% CO2 for 24 h. The was determined using a Metrohm Rancimat model
cells were then treated with or without various 743 (Herisau, Switzerland) according to the AOCS
concentrations (0-1000 µM) of rubber seed oil for Official method Cd 12b-92. 14 Approximately, 3 gm
48 h. After incubation, 40% (w/v) trichloroacetic samples of rubber seed oil and cleansing oil were
acid (TCA) was gently added to each well and was placed separately in reaction glass test tubes. These
then incubated at 4C for 1 h. samples were analyzed under a constant airflow of
rate 20 l/h within the thermostat controlled-block
Then the medium was removed and the cells were heater at a temperature of 110 C. The volatile
washed with tap water. After the last wash, the oxidation products were carried through the
plate was allowed to air dry at room temperature detector chamber and transported to measuring
(25 ± 2 °C). 0.4% (w/v) SRB solution (100 µl) was vessel containing 60 ml of Millipore water, where
dropped into each well, incubated at room the conductivity was measured as a function of
temperature for 1 h and then the plates were time. The volatile compounds can be detected by
quickly rinsed 3 times with 1% (v/v) acetic acid to an increase in conductivity. The time that was
remove unbound dye. The plates were then allowed taken to detect the volatile compounds was
to dry at room temperature, and after drying, 10 calculated as the oxidative stability index (OSI) 15.
mM tris base solution was added to each well to
dissolve the protein-bound dye and absorbance was Preparation and Characterization of Rubber
calculated at 492 nm using a microplate reader 10, Seed Cleansing Oil: Rubber seed cleansing oil was
11
. prepared by varying the composition of rubber seed
oil at the ratio of 5, 10 and 15 wt. %. Different
Determination of Linamarin: Fourier transform concentrations of non-ionic surfactants including
infrared spectroscopy (FTIR) was carried out sorbeth-30 tetraoleate (HLB 11.5) and sorbitan
according to Salimon et al. 12 Rubber seed oil was sesquioleate (HLB 3.7) were used at 10, 15, and 20
applied as films on a NaCl disc (25 mm i.d. × 4 mm wt. % as emulsifying agent based on the HLB value
thickness) for this analysis. FTIR spectra of rubber of each formulation Table 1. All ingredients were
seed oil were determined and recorded on an FTIR mixed at room temperature (25±2 C) until a
spectrophotometer (Perkin Elmer Spectrum GX, completely homogenous and clear liquid was
USA). The recording was done between 400-4000 produced. The most suitable rubber seed cleansing
cm-1. FTIR was used to determine functional oil was chosen according to physicochemical
groups of rubber seed oil 13, i.e., nitrile (C≡N) properties, cleansing property, and satisfying
groups, at wavelength number 2260-2220 cm-1. evaluation of product efficacy (emulsification,
greasiness, removal property).
Residual Solvent in Rubber Seed Oil: This
method was used for analyzing n-hexane levels that Study of Physicochemical Properties and
were involved in the production of rubber seed oil Stability Test of Rubber Seed Cleansing Oil: The
according to the USP30-NF25 Chapter 467. 6 physical properties of rubber seed cleansing oil like
Instrument used in this study was Gas pH and viscosity were measured by pH meter
Chromatography-Flame Ionization Detector GC/ (Seven Compact, Mettler Toledo, Switzerland) and
FID (GC6850 Hewlett Packard, USA). The Brookfield rheometer (DV-III Brookfield
determining column used was CP-SIL 5CB (50 m x Engineering Laboratories Inc., Middleboro, MA,
0.32 mm i.d., 5 µm, Chrompack, Middleburg EA, United States) at 25 C, respectively. The color
The Netherlands). The gas flow rate was constant measurement was examined by CM-700d Spectro-

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photometer (KONICA MINOLTA, INC., Japan) at after the stability test period for product screening.
25 C. Further, the selected formulations were The selected formulations of rubber seed cleansing
subjected to stability test by freeze-thaw cycle test oil were analyzed at accelerated conditions of
(4 C for 24 h and 45 C for 24 h) for 6 cycles. All temperature and humidity to assess their stability
parameter measurements in terms of pH, viscosity, again, according to the ICH guidelines Q1A R216.
and color were done at the initial time and also
TABLE 1: COMPOSITION OF RUBBER SEED CLEANSING OIL
Ingredients Formulation no.
(w/w) 1 2 3 4 5 6 7 8 9
Rubber seed oil 5 10 15 5 10 15 5 10 15
Mixed oil 30 30 30 30 30 30 30 30 30
Sorbeth-30 tetraoleate 6.24 5.3 4.36 8.75 7.19 5.65 10.74 8.44 6.17
Sorbitan sesquioleate 3.76 4.7 5.64 6.25 7.81 9.35 9.26 11.56 13.82
Caprylic/capric triglyceride √ √ √ √ √ √ √ √ √
Cetyl ethylhexanoate √ √ √ √ √ √ √ √ √
Vitamin E acetate √ √ √ √ √ √ √ √ √
Ascorbyl palmitate √ √ √ √ √ √ √ √ √
Phenoxyethanol √ √ √ √ √ √ √ √ √
Light mineral oil q.s. to 100 100 100 100 100 100 100 100 100

Physical and chemical stability of rubber seed content of liquid foundation (True Match, L’Oréal,
cleansing oil was examined for 6 months at 3 China) in 95% ethanol using a UV-visible
different conditions of room temperature which spectrophotometer (UV-1800, Shimadzu, Japan).
were 25 ± 2 C, 30 ± 2 C/ 75 ± 5% RH, and 40 ± 2 Validation method to evaluate the specificity,
C/ 75 ± 5% RH respectively. After the storage linearity, accuracy, and precision of rubber seed
period, the rubber seed cleansing oil was subjected cleansing oil on liquid foundation removal were
to a test by the Rancimat method. performed according to the method followed by
Chaloennit and Lourith18 with slight modifications.
Evaluation of Cleansing Oil Product Efficacy:
The cleansing efficacy of all the 9 formulations of Specificity: Each of the selected rubber seed
rubber seed cleansing oil prepared were cleansing oil or commercial product was weighted
determined. To evaluate cleansing efficacy, a 0.080 g in a conical flask, 95% of ethanol was
slightly modified method of NIKKOL group 17 was added and then sonicated for 2 min. The mixture
followed; 2 × 2 cm2 areas with lipstick and 4 × 3 was filtered through a Whatman no. 1 paper and
cm2 areas with liquid foundation were made on the adjusted to the final volume of 50 ml. Also, the
Petri dishes, left on for 5 min and then, pictures liquid foundation was weighted 0.035 gm and
were captured. Next, 0.5 ml and 1 ml of rubber extracted by 95% ethanol in the same method as
seed cleansing oil were dropped on the lipstick and described above. Absorbance patterns were
liquid foundation respectively, and massaged by a recorded for comparing each of these cleansing oil
gloved hand; 10 times on lipstick, and 25 times on products in ethanol.
liquid foundation. Then, 5 ml of water was poured
in both the areas and was gently massaged again Linearity: Different amounts of a liquid
for 25 times. Next, the Petri-dishes were dipped in foundation in the range of 0.04-1.4 g were
water and immediately taken out. Pictures were weighted into a series of 50 ml volumetric flasks,
taken before and after, for comparison. The best and adjusted to the final volume with ethanol to
formulations were selected after comparison, for obtain the concentration 0.8-2.8 mg/ml,
testing the cleansing ability by UV- respectively. The extraction process was performed
spectrophotometer method. as the previous conditions mentioned, and then the
absorbance was reported for the specified
UV-Spectrophotometer Validation of the wavelength. The correlation coefficient (r 2) was
Rubber Seed Cleansing Oil Efficacy: The calculated, which had to be equal to or greater than
cleansing efficacy of the selected rubber seed 0.995 to demonstrate acceptable linearity of the
cleansing oils were determined by measuring the analytical method.

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Accuracy: Three different concentrations of liquid avocado oil (<10 mg eq of peroxide/kg, 177-198
foundation (1, 1.4 and 2.2 mg/ml) were applied mg KOH/g) 22, 23. The fatty acid of rubber seed oil
onto cotton sheets, then extracted using 95% consisted of saturated fatty acid (17.07%) and
ethanol (10 ml × 4), and filtered through Whatman unsaturated fatty acid (78.73%). The saturated fatty
no. 1 filter paper into 50 ml volumetric flasks. The acids were palmitic acid (9.33 ± 0.01%) and stearic
recovery (%) of the extracted liquid foundation was acid (7.74 ± 0.01%), and unsaturated fatty acids
assessed. were mainly oleic acid (23.60 ± 0.01%), linoleic
acid (40.83 ± 0.02%) and linolenic acid (14.30 ±
Precision: This experiment was done to evaluate 0.00%). The color of rubber seed oil presents as a
and compare the precision of various formulations pale yellow based on b* value (9.30 ± 0.02).
of rubber seed cleansing oil and a commercial
product. In this experiment, liquid foundation (3 × TABLE 2: THE PHYSICOCHEMICAL PROPERTIES
3 cm) was applied on to a Petri dish, then 2 drops AND FATTY ACID CONTENT OF RUBBER SEED OIL
Parameter Rubber seed oil
of cleansing oil were dropped, and a cotton sheet Viscosity 25 °C, 200 rpm 75.31±0.54
folded into 4 layers was placed over it. A standard Refractive index 1.4632-1.4812
weight (100 g, Class F2) was placed on top of the Specific gravity 0.9244±8.11×10-5
cotton sheet. Next, the cotton sheet was moved Acid value (mg KOH/g) 6.91±0.00
Iodine value (mg I2/g) 128.49±0.59
horizontally in all directions without taking off the Peroxide value (mg eq of peroxide/kg) 2.45±0.01
standard weight atop. The residues in the cotton Saponification value (mg KOH/g) 183.46±2.25
sheet were extracted as previously mentioned. The TEAC (mg/100g oil) 42.23±3.53
experiment was done in triplicate for each Palmitic acid (%) 3.47±0.01
Stearic acid (%) 8.33±0.01
cleansing oil product. Oleic acid (%) 32.05±0.01
Linoleic acid (%) 37.09±0.02
Statistical Analysis: All experiment data were Linolenic acid (%) 16.40±0.00
presented as mean ± SD. Analysis of variance Color L* 30.65±0.00
(ANOVA) was used to test the statistical Color a* 1.99±0.01
Color b* 9.30±0.02
significance of difference among the group. The
significance of the difference between the mean Cytotoxicity Effect of Rubber Seed Oil on Skin
was tested using the Student's t-test. The Fibroblast Cells: Cytotoxicity test of rubber seed
differences were considered statistically significant oil on skin fibroblast cells was tested by SRB
when p<0.05. assay. Skin fibroblast cells were treated with or
without various concentrations of rubber seed oil
RESULTS AND DISCUSSION:
(0-1000 µg/ml).
Physicochemical Properties and Fatty Acids
Content of Rubber Seed Oil: The
physicochemical properties of rubber seed oil are
shown in Table 2. The viscosity, refractive index,
and the specific gravity was in the same range as
previously reported by Lourith et al., 5 and Raknam
et al. 19 The acid value (6.91 ± 0.00 mg/KOH/g)
and iodine value (128.49 ± 0.59 mg I2/g) was
similar to the general seed oils used in cosmetics
such as sunflower (<6 mg KOH/g) and wheat germ
(128 mg I2/g) 5, 20. Peroxide and saponification
value of rubber seed oil were in an acceptable
FIG. 1: CYTOTOXICITY OF RUBBER SEED OIL ON
range for natural oil in cosmetic products 21. All SKIN FIBROBLAST CELL
parameters of chemical properties of rubber seed
oil were in the normal range for a natural oil, and After 48 h of treatment, the result showed that
comparable to macadamia (<10 mg eq of IC50>1000 µM in these concentrations, which
peroxide/kg, 150-200 mg KOH/g), rice bran (<15 meant that rubber seed oil was not toxic to the skin
mg eq of peroxide/kg, 190-200 mg KOH/g) and fibroblast cell. Furthermore, the results also showed

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that the high dose of rubber seed oil (125-1000 Oxidative Stability of Rubber Seed Oil:
µg/ml) Fig. 1 significantly enhanced fibroblast cell Accelerated oxidative stability test of rubber seed
growth. These results indicated that rubber seed oil oil was carried out by measuring the oxidative
was safe to apply to human skin. stability index (OSI). Higher OSI indicates that it
takes a longer time to create the volatile oxidation
Analysis of Linamarin: Linamarin in rubber seed products, which mean the stability would also be
oil was determined by FTIR technique. The result higher. The OSI of rubber seed oil in this study was
of the characteristic absorbance band and their 5.21 ± 0.1 h at 110 C. This showed that the OSI of
assignment to functional groups are shown in Fig. rubber seed oil was even better than other oils
2. The FTIR spectrum indicates the various which are used commercially, such as sunflower oil
contents of rubber seed oil. Wavenumber (1743 cm- (2.40 h) and grape seed oil (3.20 h) 24. Moreover,
1
) indicates the presence of the ester carbonyl the presence of adequate content of saturated fatty
functional group (C=O). Spectrum peaks at 3009, acids and moderately low monounsaturated fatty
2922 and 2853 cm-1 indicates the presence of CH 2 acids in rubber seed oil might support a high
and CH3 in rubber seed oil. The spectra ranging thermal stability 25.
between 1461-1419 cm-1 belongs to double bond
(C=C), and 1376-1236 cm-1 indicates the presence Physicochemical Properties and Stability of
of carboxylic acid. The obtained FTIR spectra of Rubber Seed Cleansing Oil: Nine formulations of
rubber seed oil did not show regions which rubber seed cleansing oils were prepared and tested
indicated the presence of cyanide. It meant that for their stability by freeze-thaw cycle, as shown in
linamarin or hydrogen cyanide was eliminated Table 3. The physical appearance, color, viscosity,
during the process of extraction with an adequately pH, and cleansing power were used as the criteria
high temperature. to select the product of rubber seed cleansing oil
with the highest quality. Rubber seed cleansing oil
was stable in terms of appearance, pH, and color,
with ΔE less than 1.5 26, 27, which means the color
change was not visually identifiable. All the
formulations showed good results in terms of
physical appearance and stability after the freeze-
thaw cycle. However, the best product was selected
based not only on good physical appearance and
stability but also on good cleansing efficacy and
overall property.

Evaluation of Rubber Seed Cleansing Oil

FIG. 2: THE FTIR SPECTRUM OF RUBBER SEED OIL
Product Efficacy: The various properties of rubber
Residue Solvent in Rubber Seed Oil: Hexane is seed cleansing oil related to its cleaning efficacy is
one of the most commonly used solvents, not only reported in Table 4. The results show the efficacy
in the edible oil industry but also in the process of in terms of the ability to emulsify when in contact
production of herbal medicine and bioactive with water, non-greasiness after use, and efficacy
components. So, it is possible that hexane may be as a makeup remover. Formulation no. 2, 4, and 7
present in trace amounts in these kinds of products. showed a high score in the ability to emulsify,
A study showed that 2, 5-hexanedione, which is removal property and efficacy power with low
derived from the metabolism of hexane, is greasiness after evaluation Fig. 3. This result shows
neurotoxic. According to USP30-NF25 that formulation no. 2, 4, and 7 were superior over
concentration limit, n-hexane concentration should the other formulations. Therefore, the formulation
be lower than 290 ppm. In this study, n-hexane no. 2, 4, and 7 were selected to evaluate their
residual in rubber seed oil was determined. It was cleansing efficacy by UV method. A blind test was
found that n-hexane in rubber seed oil was 60.35 ± conducted between rubber seed cleansing oil
1.12 ppm, which is acceptable and not harmful. formulation no. 2, 4, 7, and the commercial

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product. Their efficacy was compared, as shown in quality assessment. Cleansing oil formulation no. 4
Fig. 4 and 5. was selected for oxidative stability study by
Rancimat method at 3 conditions for 6 months to
Oxidative Stability of Rubber Seed Cleansing determine the shelf life of cleansing oil, as shown
Oil Formulation: Oxidative stability of cleansing in Table 5.
oil is one of the most important parameters for its
TABLE 3: PHYSICAL APPEARANCES OF RUBBER SEED CLEANSING OIL BEFORE AND AFTER FREEZE
AND THAW CYCLE (6 CYCLE)
Formulation Before After
no. pH Viscosity Color ΔE pH Viscosity
1 7.90±0.23 34.27±0.000 0.15±0.00 7.40±0.12 34.43±0.000
2 8.00±0.45 35.55±0.069 1.21±0.01 7.11±0.32 35.87±0.000
3 7.58±0.64 36.71±0.000 1.11±0.01 7.00±0.53 36.19±0.183
4 8.43±0.67 37.87±0.069 0.95±0.10 7.70±0.45 38.07±0.069
5 7.38±0.33 39.75±0.183 0.86±0.02 7.15±0.64 38.79±0.069
6 8.32±0.21 38.79±0.069 0.27±0.01 6.72±0.11 39.55±0.346
7 8.73±0.15 47.23±0.069 0.08±0.00 7.58±0.46 42.75±0.183
8 8.29±0.53 46.91±0.120 1.24±0.01 6.93±0.33 45.91±0.183
9 7.82±0.32 51.47±0.120 0.31±0.01 6.66±0.23 51.51±0.069

TABLE 4: EVALUATION OF RUBBER SEED CLEANSING OIL PRODUCT

Formulation Evaluation protocol
no. Ability to emulsify Greasiness Removal property
1 + ++ ++++
2 ++ + ++++
3 ++ +++ +++
4 +++ + +++++
5 + + ++++
6 + ++++ ++
7 +++ ++ ++++
8 ++ + ++
9 + +++++ +
+++++ the highest value, ++++ very high, +++ high value, ++ fair value and + less value

FIG. 3: THE APPEARANCE OF RUBBER SEED CLEANSING FORMULATION NO. 2, 4 AND 7

FIG. 4: THE PETRI DISHES BEFORE EVALUATION (FORMULATION NO. 2, 4, 7 AND COMMERCIAL
PRODUCT WHEN APPLIED ON A, B, C, AND D RESPECTIVELY)

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FIG. 5: THE PETRI DISHES AFTER EVALUATION (FORMULATION NO. 2, 4, 7 AND COMMERCIAL PRODUCT
WHEN APPLIED ON A, B, C AND D RESPECTIVELY)

The result revealed that when the cleansing oil was UV-Spectrophotometer Validation of the
stored at room temperature (25 ± 2 C) for 6 Rubber Seed Cleansing Oil Efficacy:
months, OSI was the highest (66.64 ± 1.45 h). This Specificity: The wavelength 250 nm was found to
may be due to the antioxidant in the formulation be the most suitable for measuring the content of
such as vitamin E and ascorbyl palmitate 21, which liquid foundation. This wavelength only indicated
delayed the oxidation in the product. Also, the the content of liquid foundation and was not
process of drying rubber seed before oil extraction interfered by any of cleansing oil product in
helped the oil have good stability for long time extracted solution.
storage 28.
Linearity: The solutions were scanned on a
After storing for 6 months at 30 ± 2 C / 75 ± 5% spectrophotometer in the UV range of 200-400 nm.
RH, OSI was 55.75 ± 0.11 h, and after storing at 45 The spectrum was recorded at 250 nm as maximum
± 2 C / 75 ± 5% RH, OSI was 48.35 ± 0.62 h, (λmax). The calibration plot was plotted as
which indicated that rubber seed cleansing oil was concentration vs. absorbance. The coefficient of
stable in terms of oxidative stability. determination (r2) was found to be 0.9995 (y=
0.2634x + 0.0050). This method indicated good
Moreover, we can minimize the variations between linearity and also represented the precision of the
shelf life predictions and long-term storage study extracted amount of liquid foundation.
by selecting the correct levels in the Rancimat
method according to the product type. Thus, using Accuracy: A known amount of three different
the Rancimat technique, we can rapidly evaluate solutions that represent low, medium and high (1,
the quality of cleansing oil, and determine if it can 1.4 and 2.2 mg/ml) concentrations of a liquid
be stored at room temperature for 6 months. foundation in cotton sheets were extracted and
tested to validate the accuracy of this method.
TABLE 5: OXIDATIVE STABILITY OF RUBBER Recovery (%) was found in the range of 88-95%, as
SEED CLEANSING OIL FORMULATION NO. 4 FOR 6
shown in Table 6. The validity and reliability of
MONTHS
Storage condition Oxidative stability index (h) this method were obtained by calculating the %
Room temperature (25±2˚C) 66.64±1.45 RSD, which was < 2% (0.25-0.76%) 29, indicating
30±2˚C/75±5% RH 55.75±0.11 that this analytical method was accurate.
45±2˚C/75±5% RH 48.35±0.62

TABLE 6: RECOVERY (%) OF THIS UV-SPECTROPHOTOMETRIC DETERMINATION (N=3)

Added amount (mg/mL) Mean of measured conc. (mg/mL) Average recovery (%) % RSD
1 0.89±0.02 88.88±1.78 0.31
1.4 1.32±0.03 94.40±2.18 0.76
2.2 2.11±0.06 95.53±2.51 0.25

Precision: As shown in Table 7, the % RSD of the commercial product, and the result found that
three formulations of rubber seed cleansing oil and formulation no.4 exhibited the highest efficacy
the commercial product was less than 2, which (89.89 ± 2.75%) in makeup removal against the
indicates that the method used was precise. The liquid foundation.
rubber seed cleansing oil was compared to

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Raknam et al., IJPSR, 2020; Vol. 11(1): 146-155. E-ISSN: 0975-8232; P-ISSN: 2320-5148

After blind test evaluation, the result indicated that 6. The United States Pharmacopeia-30: The National
Formulary-25, General Chapter 467 Residual Solvents.
the efficacy of formulation no 4 was almost Baltimore, Maryland: United State Pharmacopeial
identical to the commercial product, and was better Convention, United Book Press 2017: 1494
than formulation 2 and 7. Therefore, formulation 7. Collison MW: Official methods and recommended
practices of the American Oil Chemists' Society (AOCS).
no 4 was the best cleansing oil product. Official method 921.08 Index of refraction of oils and fats.
Champaign, Edition 7th, 2017.
TABLE 7: PRECISION OF EFFICACY OF RUBBER 8. The United State of Pharmacopeia-30: The National
SEED CLEANSING OIL AND COMMERCIAL Formulary-25, General Chapter 401 Fats and Oils.
PRODUCT AND SATISFIED EVALUATION Baltimore, Maryland: United State Pharmacopeial
Formulation no. Recovery (%) % RSD Convention, United Book Press 2017: 1486.
2 87.28±3.23 1.00 9. Bialek A, Bialek M, Jelinska M and Tokarz A: Fatty acid
4 89.89±2.75 1.01 profile of new promising unconventional plant oils for
cosmetic use. International Journal of Cosmetic Science
7 85.17±3.33 0.56
2016; 38(4): 382-88.
Commercial product 88.08±6.09 0.55 10. Vichai V and Kirtikara K: Sulforhodamine B colorimetric
assay for cytotoxicity screening. Nature Protocol 2006; 1:
CONCLUSION: This study proved that the rubber 1112-1116.
seed oil obtained from solvent reflux extraction 11. Orellana EA and Kasinski AL: Sulforhodamine B (SRB)
assay in cell culture to investigate cell proliferation. Bio-
was safe for use in topical applications. Hence, we protocol 2016; 6(21): e1984.
were able to develop rubber seed oil into a 12. Salimon J, Abdullah BM and Salih N: Rubber (Hevea
cleansing oil product. It showed good brasiliensis) seed oil toxicity effect and Linamarin
compound analysis. Lipid in Health and Disease 2012; 11:
physicochemical properties, good stability, high 74-82.
ability in emulsification, removal property, and 13. Abdullah BM: Analysis of linamarin compound toxicity
cleansing efficacy with low greasiness. The best on rubber seed oil for health disease. Journal of
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rubber seed cleansing oil product composed of 5% 131.
of rubber seed oil and 15% of surfactants, which 14. Collison MW: Official methods and recommended
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Edition 7th, 2017.
ACKNOWLEDGEMENT: This research was 15. O'Brien RD: Fats and oils analysis. Fat and oils
financially supported by the Natural Rubber formulating and processing for application. CRC Press,
Innovation Research Institute (Grant No. Boca Raton, 2009: 766.
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PHA581138S) and Faculty of Pharmaceutical Stability Testing of New Drugs Substances and Products,
Sciences, Prince of Songkla University, Thailand. in Proceedings of the International Conference on
Harmonisation, Geneva, 2003. [cited 2018 January 25].
CONFLICTS OF INTEREST: The authors Available from: http://www.ich.org/fileadmin/Public_
Web_Site/ICH_Products/Guidelines/Quality/Q1A_R2/Ste
declared that they have no conflict of interest. p4/Q1A_R2_Guideline.pdf.
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How to cite this article:

Raknam P, Pinsuwan S and Amnuaikit T: Rubber seed cleansing oil formulation and its efficacy of makeup remover. Int J Pharm Sci &
Res 2020; 11(1): 146-55. doi: 10.13040/IJPSR.0975-8232.11(1).146-55.
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