Journal of the Japan Institute of Energy, 94, 763-768（2015） 763

Production of Biodiesel from Rubber Seeds (Hevea Brasiliensis)

by In situ Transesterification Method

Bashir Abubakar ABDULKADIR ※ 1, Yoshimitsu UEMURA ※ 2, Anita RAMLI※ 1, Noridah Bt OSMAN※ 2,

Katsuki KUSAKABE ※ 3, and Takami K AI ※ 4
(Received April 4, 2014)

Biodiesel is defined as the mono-alkyl esters of vegetable oils (both edible and non edible) or animal fats, obtained
by transesterifying oil or fat with an alcohol. In this research, biodiesel is produced by in situ transesterification
(direct transesterification) method from the seed of rubber tree using KOH as a catalyst. The influence of seed to
methanol mass ratio, reaction time, temperature, seeds sizes, and catalyst loading was investigated. Important properties of
biodiesel such as free fatty acid (FFA), viscosity, pour point, density etc were also investigated. The result showed that
the best ratio of seeds to methanol was 1:6 (10 g seeds with 60 g methanol), 120 min reaction time at 60 °C reaction
temperature, and catalyst loading of 3.0 g with 150 µm seeds size. The maximum yield obtained was 96 %, and also most of
the properties obtained were in agreement with the standards and with other FAMEs. This study supports the production of
biodiesel from the rubber seeds using in situ (direct) transesterifcation method from the seeds as an alternative to diesel
fuel.

Key Words
Biodiesel, FAME, Free fatty acid, in situ

in the diesel engine and in the existing fuel distribution and

1. Introduction
According to the Brazilian biodiesel programme,
biodiesel is defined as a fuel obtained from mixtures, in
different proportions of fossil diesel and alkyl esters of
vegetable oils or animal fats 1). However, under the chemical point
of view, biodiesel is composed mainly of fatty acids mono-alkyl
esters. Transesterification is a process used for obtaining
biodiesel and involves the catalyzed reaction of triglycerides
(major compounds of oils and fats) with short-chain alcohols
such as methanol and ethanol. Most transesterification in
industrial processes employ alkaline catalyst (KOH, NaOH, and
NaOCH3) and methanol 1). Its use (pure or blended) does not
demand any modification
※ 1 Department of Fundamental and Applied Sciences,
Universiti Teknologi PETRONAS
3150 Tronoh,Malaysia
※ 2 Centre for Biofuel and Biochemical Research (CBBR),
Universiti Teknologi PETRONAS
Tronoh, Malaysia
※ 3 Department of Nanoscience, Faculty of Engineering, Sojo
University
4-22-1 Ikeda, Kumamoto 8600082, Japan
※ 4 Department of Chemical Engineering, Faculty of
Engineering, Kagoshima University 1-21-40
Korimoto, Kagoshima 890-0065, Japan
storage infrastructure. Moreover, biodiesel has a high energetic
yield, fixes the solar energy and contains insignificant
amounts of sulphur. Therefore, biodiesel is currently the best
substitute for fossil diesel 1) 2).
In recent discussions of biodiesel production by
transesterification process, a controversial issue has been
whether biodiesel can be produced by in-situ (direct)
transesterification process. Widayat et al. 3), reported that,
biodiesel production from either vegetable oil or animal fat has a
major drawback. The essence of Widaya’s argument is that to
obtain the vegetable oil by conventional method, the seed is
pressed by mechanical process or extraction with chemical
solvent. The oil is further pretreated by degumming process. Of
course this process requires a lot of manufacturing cost. The
biodiesel production from rubber seed oil used two stage-
method of esterification reaction followed by transesterification
reaction 4) 5). This process still has the same drawbacks that are
the pretreatment process of the oil. Most studies in the field
of biodiesel production have only focused on vegetables oil and
animal fat. Therefore, to solve these drawbacks, the
(trans)
 764 J. Jpn. Inst. Energy, Vol. 94, No. 7, 2015
esterification process by in situ method will be one of the best
possible alternatives.
In-situ (trans)esterification is a method used to
produce biodiesel where the raw material (seed) is contacted
directly with an alcohol assisted with an acid or alkaline
catalyst. Thus extraction and transesterification take place in
one step and the alcohol is used for extracting oil from the
seeds as well as transesterifying it 6). In this process, mass ratio
of seeds to methanol must be higher than the value calculated
based on the stoichiometry. Excess methanol will play dual
role as the solvent extraction as well as the reactant 7). The cost
of biodiesel production process could be minimized if in situ
(trans) esterification method is used. In this process, costs for
solvent extraction and oil purification can be reduced. So,
biodiesel production becomes easier and simpler 8). In-situ
transesterification of sunflower oil with acidified methanol
produces fatty acid methyl esters in yield significantly greater
than those obtained from conventional process 3).
In-situ transesterification was first introduced by
Harrington and D’Arcy-Evans 7). They used sunflower seed as
a raw material. Marinkovic et al. 9), has done experiments using
sunflower seed with the same process. Harrington et al. 7), have
done in-situ process from soybeans. Ozgul-Yucel and Turkay 10)
11)
performed in situ esterification process with rice bran as raw
material. Also Ginting et al. 12), performed in situ
transesterification from castor seed. Ozgul and Turkay 13)
investigated in situ esterification of high-acidity rice bran oil
with methanol and ethanol, and with sulfuric acid as catalyst.
The transesterification with in situ method gave similar ester
yields to those obtained from conventional transesterification,
being an alternative, efficient, and economical process 13).
The objectives of this research are to produce
biodiesel from rubber seed by in-situ transesterification method
and to investigate the influence of reaction parameters on the
FAME yield. The merit of this method is that this new
process just requires a single process, where the normal oil
extraction process can be avoided. In addition, in this process,
the purification of products after transesterification reaction is
much simple compared to the common method. Therefore, this
new process can offer an alternative way to convert the seeds
directly to methyl esters by a simpler-shorter production
process.
2. Experimental Procedure
2.1 Sample collection and preparation
Rubber trees yield 3-seeded ellipsoidal capsules, each
capsule with three seeds. Rubber seeds are ellipsoidal, variable
in size, 2.5-3 cm long, mottled brown, lustrous,
weighing 2-4 g each. The capsules of the seeds are spread over
the ground in plantation. The seeds were collected from a
plantation nearby Universiti Teknologi PETRONAS, Perak
Malaysia and dried under the sun for about 3 h to remove
the remaining moisture, then, the shell and kernels were
separated by breaking the capsules. The kernels were dried in
the oven for about 24 h to remove the remaining moisture. The
kernels were immediately ground using food processing
machine ( blender) in order to weaken and rupture the cell.
Subsequently, the ground rubber seed was taken into the oven
and kept overnight at 104 °C in order to deactivate the remaining
enzyme as well as to remove the moisture and then sieved using
300, 350, and 400 µm opening screens. Finally, three different
size fractions were obtained; 150 µm (0-300 µm), 325 µm
(300-350 µm), and 375 µm (350-400 µm). After drying, the
moisture content of the kernels was determined using halogen
moisture content analyzer (Mettler Toledo, HR83) and also after
the water is removed from the seeds by evaporation using
furnace, and in both cases it was found to be 1.5 %. The oil
content of the kernels was determined through soxhlet
extraction using hexane as a solvent for 4-5 h. It was found that,
the seed has oil content of 40 %.
2.2 Rubber seeds characterization
The recommended methods of the Association of
Official Analytical Chemists (AOAC, 1984) were adopted to
determine the levels of ash and water content from the seeds.
2.3 Transterification process
Methanol and the KOH catalyst were mixed in a
three neck round bottom flask connected to a reflux condenser
and heated until catalyst completely dissolved. Then, 10 g of the
ground rubber seeds sample was added into the mixture and the
blend was heated at 60 °C and at atmospheric pressure for 120 min
at 500 rpm stirring speed. The product was filtered using
Whatman filter paper and poured into the separating funnel to
separate the excess of alcohol. The reaction was repeated to
study the effect of rubber seeds to methanol mass ratio, reaction
temperature, reaction period, catalyst loading, and sizes of the
seeds on the FAME yield. The FAMEs analysis was quantified
by Agilent Technologies 7890A Gas Chromatography equipped
with Flame Ionization Detector (GC-FID) with HP-5 5%
Phenyl Methyl Siloxane capillary column (30 m by 320 µm by
0.25 mm). Automatic injection sampler by Agilent was used.
Methyl heptadecanoate (0.630 mg internal standard) was
dissolved in 1 mL hexane to prepare the standard
 J. Jpn. Inst. Energy, Vol. 94, No. 7,
765
2015

solution. Approximately 100 mg crude methyl ester was

dissolved in 1 mL standard solution for GC-FID analysis.
About 1 µL sample was injected into the GC at an oven
temperature of 210 °C with helium as the carrier gas. The GC oven
was programmed at 210 °C, isothermally for 15 min. Then, the
FAMEs content and the FAME yield were calculated. The
experimental design and response/FAME conversion conducted
in this research paper is similar to a conventional method of
FAME production from rubber seed oil by Junaid et al. 14).
The FAME yield was calculated using the following
expressions 15):
Moil phase×XFAME
FAME Yield = ×100 %
Mseed×Xoil×F
Moil phase = Mass of oil phase after transeterification (g). XFAME
= Mass fraction of FAME in oil phase.
Mseed = Mass of the seeds (g).
Xoil = Mass fraction of oil in seeds.
F = 3MWFAME/MWoil (MW= molecular weight);
approximately unity.
3. Result and Discussion
3.1 Rubber seeds characterization
Water content or moisture content is the quantity of
water contained in a material, such as in soil (called soil
moisture), oil, fruit, seeds or wood. Water content is used in a
wide range of scientific and technical areas, and is expressed as
a ratio, which can range from 0 (completely dry) to the value of
the materials' porosity at saturation 16). The moisture content in
this paper was determined by measuring the mass of rubber
seeds before and after the water is removed from the seeds by
evaporation using furnace while the water content for the oil
was determined using Karl Fischer titration process based on
AOCS standard.
For many samples, the water content may be an
extremely important index used for establishing the
relationship between the samples/rubber seed oil and its
properties. The uniformity of a fine-grained seeds largely
depends on its moisture content 17). In this research, a
Malaysian rubber seeds was found with 1.5 % moisture
content while rubber seed oil was found with less than 3.3 %
water content as shown in Table 1.
Ash content is the name given to all non-aqueous residue
that remains after a sample is burned, which consists mostly of
metal oxides. Ashing is the process of mineralization for pre-
concentration of trace substances prior to chemical analysis 18).
In this research, Malaysian rubber seeds was found to have 2.88
% ash content as shown in Table 1, and the result was found
to be very
Table 1 Characteristics of rubber seed/rubber seed oil
Component Content (wt %)
Seed
Moisture 1.50
Ash 2.88
Oil 40.00
Seed oil
Water 3.30
similar with that of Widayat and Suherman 19).
3.2 Effect of seed to alcohol mass ratio
The mass ratio of seeds to alcohol is one of the
important factors that affect the FAME yield as well as
production cost of biodiesel. Mass ratio is the ratio of weight
of seeds to the weight of alcohol. The FAME yield is defined
as the yield of the process represented in terms of percentage 20).
In this research, various mass ratios of the seeds to alcohol
(methanol) starting with 1:2 and followed by (1:3, 1:4, 1:5, 1:6,
1:7 and 1:8) were tested. It was found that, the maximum yield
was achieved at the mass ratio of 1:6 (96 %) as shown in Fig. 1
and further increase in mass ratio, there was no any improvement
in the conversion efficiency, instead a decrease was observed.
3.3 Effect of reaction temperature
The FAME yield was only 67 % at 40 °C, but increased
rapidly with increasing in temperature. The optimum
temperature for this reaction was found to be at 60 °C, with 96
% FAME yield with 1:6 mass ratio, 3 g KOH catalyst and 120
min reaction time as shown in Fig. 2.
The reason for the increase in yield as the temperature
increases is that, increase in temperature is believed to facilitate
the breaking of the seeds cell walls, thereby creating a void
which serves as migratory space

Fig. 1 Effects of mass ratio on the FAME yield (%) (temp= 60

°C, seeds size= 150 µm, time = 120 min)
766 J. Jpn. Inst. Energy, Vol. 94, No. 7, 2015

Fig. 2 Effects of reaction temperature on the FAME yield (%)

(seeds to MeOH mass ratio= 1:6, seed size= 150 μm, time
= 120 min)

for the contents of the oil bearing cells. Temperature also

lowers the viscosity of the oil, thus facilitating the release of the
oil out of the kernel. However, at higher temperature, there is
substantial loss of moisture leading to a hardening of samples
and oil degradation. This may account for the decrease in the
yield at higher temperature of 65 °C 21).
As said by Ramadhas et al. 4) reaction temperature greater
than 60 °C should be avoided in the case of rubber seed, because
they tend to accelerate saponification before transesterification
reaction completed. Fig. 3 Effects of catalyst concentration on the FAME yield (%)
(Seeds to MeOH mass ratio= 1:6, seed size= 150 µm, 60 °C,
3.4 Effect of catalyst concentration. 120 min)
The KOH concentration of 1.0-4.0 g was used for the
analysis. The maximum FAME yield of 96 % was
achieved at 3.0 g KOH loading with 1:6 mass ratio of seeds to
methanol, 120 min reaction time, and at 60 °C reaction
temperature, as shown in Fig. 3. It was reported 22) that, in- situ
transesterification is inactive without catalyst. Acid or alkali
catalyst in in-situ transesterification helps to break the cell
wall of the seeds, thereby facilitating methanol to access the
oil in cotyledon cells.
As mentioned by Ramadhs et al. 23) from physical
appearance, the higher the catalyst concentration, the darker
biodiesel will be produced. Therefore the addition of a suitable
catalyst concentration is important to the physical appearance of
biodiesel.

3.5 Effect of reaction duration

The maximum FAME yield obtained in this
experiment (effect of reaction duration) was 96 % at 120 min
reaction duration with 1:6 mass ratio of seeds to methanol, 3 g KOH Fig. 4 Effects of reaction time on the FAME yield (%) (seeds to
catalyst loading, and at 60 °C reaction temperature as shown in MeOH mass ratio= 1:6, temp = 60 °C, seed size= 150
Fig. 4.
µm, time = 120 min)

Fig. 5 Effect of mesh size (µm) on ester content (%) (seeds to

MeOH mass ratio= 1:6, temp = 60 °C, time = 120 min)

3.6 Effect of sizes of the seed.

Various sizes of the seeds ranging from 300 to 400
µm in size were used. It has been observed that, the maximum
yield of 96 % was achieved at the smallest mesh sizes of 150
µm as shown in Fig. 5.
This phenomenon could be attributed to the fact that
smaller particles have larger amount of surface area as well
 J. Jpn. Inst. Energy, Vol. 94, No. 7,
767
2015

as an increased number of broken cells resulting in a high oil

concentration at the smaller particle sizes 21).
The FAME yield obtained in all the optimum
conditions was 96 % as discussed. This is one of the
observed drawbacks of in-situ transesterification process that it
can not go beyond 96 % based on this paper and other previous
work, and this might be due to the extraction and
transesterification taking place at the same time, and also single
solvent (methanol) used for extracting the oil as well as
transesterifying it unlike conventional method of
transesterification. But further studies may improve the yield
beyond the acquired result in this paper.
4. Properties of the Produced FAME
The fuel properties of rubber seed methyl ester in
relationship with those of other esters are shown in Table 2. The
majority of the properties of the obtained FAME through in
situ method are relatively equivalent to those of other esters.
The GC-FID analysis shows that, the biodiesel contains
large amount of C18. The present result shows that, in situ
transesterification process enhanced the fuel properties of the
oil with respect to specific gravity, viscosity, density, and acid
value. The viscosity of biodiesel is in the range of biodiesel
standard. The properties of the obtained biodiesel are in
comparable with ASTM standards. The observed properties of
methyl esters from rubber seed by in situ transesterification
methood are found to be in reasonable conformity with
ASTM 6751.
The present results obtained show that, the
transesterification process improved the fuel properties of the oil
with respect to specific gravity, density, viscosity, pour point,
and cloud point. The comparison of these properties with
diesel shows that, the methyl ester has a relatively closer fuel
property values to that of diesel. Viscosity is the most
important property of biofuel since it affects the operation of fuel
injection equipment, particularly at low temperatures where the
increase in viscosity affects the fluidity of the fuel 25). Biodiesel
has a viscosity close to that of diesel fuels. High viscosity leads
to poorer
atomization of the fuel spray and less accurate operation of the
fuel injectors 26). In this study the viscosity obtained was 4.68 mPa
s as shown in Table 2 which falls within the range of ASTM
standard. This shows that FAME produced through an in situ
process will serve as an excellent alternatives to the energy
crisis since it has low viscosity.
Density is an important property of biodiesel. Density is
the mass per unit volume of any liquid at a given temperature
while specific gravity is the ratio of the density of a liquid to the
density of water. Density and specific gravity has importance in
diesel-engine performance since fuel injection operates on a
volume metering system 25). Also, the density of the liquid
product is required for the estimation of the Cetane index 27).
Two important parameters for low-temperature
applications of a fuel are cloud point (CP) and pour point (PP). The
CP is the temperature at which a cloud of crystals first appears
in a liquid when cooled under conditions as described in
ASTM D2500-91. The PP is the temperature at which the
amount of wax from solution is sufficient to gel the fuel; thus
it is the lowest temperature at which the fuel can flow. The
PP is the lowest temperature at which an oil specimen can still
be moved. It is determined according to ASTM D97-96. These
two properties are used to specify the cold-temperature usability of
a fuel 28). In this experiment, both the pour point and cloud point
fall within the ASTM standard described in Table 2. Also, it has
been observed in this study that, the acid value was less than
1 mg-KOH/g which is within the acceptable limit of ASTM.
5. Conclusion
This research revealed that, biodiesel can be produced by
in situ (direct) transesterification method from the seeds of
rubber tree, and it was found that, the maximum biodiesel yield
obtained was 96 % at 60 °C with mass ratio of 1:6, 3.0 g of KOH
as catalyst, stirring speed of 500 rpm for 120 min. The results
were also in agreement with the findings of a previous study by
Widayat et al. 3) on Production Process of Biodiesel from Rubber
Seeds (Hevea

Table 2 Properties of methyl esters of rubber seed in comparison with other

esters
Biodiesel
Testing standard Rape seed Soybean oil RSO RSO RSO
S/N Property procedure This work oil FAME 4) FAME 4) FAME 14) FAME 4) FAME 24)
ASTM 6751-02

1 Specific gravity at 20 °C ASTM D4052 0.87–0.90 0.874 0.882 0.885 - 0.874 0.881
2 Density at 20 °C (g/cm3) ASTM D4052 - 0.897 NA NA - - -
3 Viscosity (mPa s) at 40 °C ASTM D445 1.9–6.0 4.68 4.5 4.08 3.89 5.81 4.059
4 Pour point (°C) ASTM D97 -15 to 10 -4.0 -12 -3 -2.0 -8.0 -
5 Cloud point (°C) ASTM D2500 -3 to 12 4.8 -4.0 -2.0 3.2 4.0 -
6 Acid value (mg KOH/g) ASTM D974 0.8 0.65 NA NA 0.42 0.11 0.22
 768 J. Jpn. Inst. Energy, Vol. 94, No. 7, 2015
Brasiliensis) by In Situ Transesterification Method with
Alkaline Catalyzed and also by that of Junaid et al. 14) by
conventional method.
Acknowledgement
The authors are grateful to the Mitsubishi
Corporation Education Trust Fund and University Teknologi
PETRONAS for research facilities.
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