Journal of Molecular Catalysis B: Enzymatic 107 (2014) 17–22

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Journal of Molecular Catalysis B: Enzymatic

journal homepage: www.elsevier.com/locate/molcatb

Novel strategy for lipase-catalyzed synthesis of biodiesel using

blended alcohol as an acyl acceptor
TingTing Zhao a,b , Da Som No a,b , Yangha Kim c , Young Soon Kim a , In-Hwan Kim a,b,∗
a
Department of Food and Nutrition, Korea University, Seoul 136-703, Republic of Korea
b
BK21PLUS Program in Embodiment: Health-Society Interaction, Department of Public Health Sciences, Graduate School, Korea University, Seoul 136-703,
Republic of Korea
c
Department of Nutritional Science and Food Management, Ewha Womans University, Seoul 120-749, Republic of Korea

a r t i c l e i n f o a b s t r a c t

Article history: As a novel strategy, blended alcohols consisting of methanol and ethanol were used as acyl acceptors for
Received 5 February 2014 biodiesel synthesis from soybean oil by lipase-catalyzed transesterification. Based on enzyme screening,
Received in revised form 16 April 2014 Novozym 435 from Candida antarctica was selected for the reaction. The effects of the molar proportion
Accepted 11 May 2014
of methanol in the blended alcohol, temperature, and enzyme loading were investigated for optimization
Available online 20 May 2014
of the reaction. In addition, the relative consumption rates of methanol and ethanol during the trans-
esterification were studied. Among six proportions tested, 0 (100 mol% ethanol), 20, 40, and 60 mol%
Keywords:
methanol in the blended alcohols exhibited high yields of biodiesel. For the optimum temperature, 30 ◦ C
Biodiesel
Blended alcohol was selected. The highest yield of biodiesel, over 95 wt%, was obtained at an enzyme loading of 5–10 wt%
Ethanol loading. In the lipase-catalyzed transesterification, the reactivity of methanol was significantly higher
Lipase than that of ethanol.
Methanol © 2014 Elsevier B.V. All rights reserved.

1. Introduction makes high equilibrium conversion due to highly reactive

Biodiesels are defined as monoalkyl esters of long-chain fatty
acids derived from vegetable oils or animal fats, for use in diesel
engines [1,2]. Biodiesel can be used in any mixture with petroleum
diesel as it has very similar characteristics, but it has lower exhaust
emissions. Biodiesel has better properties than those of petroleum
diesel in that it is renewable, biodegradable, non-toxic, and essen-
tially free of sulfur and aromatics [3]. The use of biodiesel fuel
has the potential to reduce levels of pollutants and of potential or
probable carcinogens [4].
The fundamental reaction in biodiesel production is the trans-
esterification reaction, which can be catalyzed either chemically
or enzymatically. Enzymatic transesterification has certain advan-
tages over the chemical catalysis of transesterification, because it
allows less energy intensity and easy recovery of glycerol [5–8].
In the biodiesel production, methanol is most widely used
because of its economic feasibility and accessibility in most
countries [9,10]. In addition, with a chemical catalyst, methanol
∗ Corresponding author at: Department of Food and Nutrition, Korea University,
Jeongneung-dong, Seongbuk-Gu, Seoul 136-703, Republic of Korea.
Tel.: +82 2 940 2855; fax: +82 2 940 2850.
E-mail address: k610in@korea.ac.kr (I.-H. Kim).
intermediate methoxide [11,12]. However, for enzymatic trans-
esterification, methanol has a critical problem that methanol has
a stronger denaturing activity compared with longer aliphatic
alcohols [13,14]. Chen and Wu [15] reported that the degree of
deactivation is inversely proportional to the number of carbon
atoms in the alcohol. In general, lipases are known to have a
propensity to act on long-chain fatty alcohols more readily than on
short-chain ones [16,17]. To overcome drawbacks of methanol, tri-
als using stepwise addition of methanol to reaction mixtures have
been performed in previous studies [18–21].
For the biodiesel production, ethanol is also used as an acyl
acceptor because ethanol less likely triggers enzyme deactivation.
From the literatures, the efficiency of the enzymatic transester-
ifiaction of triglycerides with methanol (methanolysis) was low
compared to that of ethanol in systems without a solvent. Mittel-
bach and Tritthart [22] reported transesterifications of the primary
alcohols, i.e., methanol, ethanol, and 1-butanol, with and without
solvents; the ester yields from ethanol and 1-butanol were signifi-
cantly higher than that from methanol. Abigor et al. [23] also found
that in the conversion of palm kernel oil to alkyl esters using Pseu-
domonas cepacia lipase, ethanol gave the highest conversion, i.e.,
72%, whereas the methyl ester yield from methanol was only 15%.
However, ethanol is still used not as often as methanol because of
its higher cost.

http://dx.doi.org/10.1016/j.molcatb.2014.05.002
1381-1177/© 2014 Elsevier B.V. All rights reserved.
18 T. Zhao et al. / Journal of Molecular Catalysis B: Enzymatic 107 (2014) 17–22

Methanol and ethanol, which are acyl acceptors used for synthe-
sis of biodiesel, have their own advantages and disadvantages. Thus,
using a blended alcohol of methanol and ethanol as an acyl accep-
tor for lipase-catalyzed transesterification could be an innovative
strategy for overcoming the drawbacks of each alcohol.
In this study, lipase-catalyzed transesterification was carried out
to explore the effect of a blended alcohol of methanol and ethanol
on biodiesel synthesis in a solvent-free system. Enzyme screening
of six lipases from different sources was performed. The optimum
reaction parameters with the selected lipase were also determined.
The effect of the proportion of methanol in the blended alcohol was
thoroughly studied as the major parameter, and other parameters
such as temperature and enzyme loading were also investigated.
In addition, changes in the molar proportion of methanol in the
residual blended alcohol during the reaction were also explored.
2. Materials and methods
2.1. Materials
The refined soybean oil (Sajo Haepyo® , Seoul, Korea) used in this
study was purchased from a local market (Seoul, Korea). Novozym
435 from Candida antarctica, Lipozyme TL IM from Thermomyces
lanuginosa, and Lipozyme RM IM from Rhizomucor miehei were
purchased from Novo Nordisk Bioindustry Ltd. (Seoul, Korea). Free-
type lipase OF from Candida rugosa was purchased from Meito
Sangyo Co., Ltd. (Nagoya, Japan). Free-type lipase PS from Pseu-
domonas fluorescence and lipase AYS from C. rugosa were purchased
from Amano Enzymes (Troy, VA, USA). Tricaprin was purchased
from Sigma Aldrich Co. (Seoul, Korea). The other chemicals used in
this study were of analytical grade unless otherwise noted.
of biodiesel was calculated as follows:
Yield of biodisel (wt%)
fatty acid methyl and ethyl ester (mg)
= × 100
reaction mixture (mg)
2.4. Methanol and ethanol residues after transesterification
To determine the molar proportion of methanol in the residual
blended alcohol in the reaction mixture by gas chromatography,
a distillation was firstly carried out using a distillation apparatus
(Catalog No. 751351-0005, Kimble Chase, Vineland, NJ, USA). In
brief, the reaction mixture (4 g) and toluene (8 mL) were placed in a
50 mL round-bottomed flask, which was connected to a Dean–Stark
distillation receiver. The temperature of the condenser in the distil-
lation system was kept at −20 ◦ C using a cooling circulator (model
CW-05G; Jeio Tech, Seoul, Korea). The alcohols and toluene in the
reaction mixture were distilled by heating with a heating man-
tle. The distilled fraction was analyzed by gas chromatography,
using a gas chromatograph (model 3800; Varian, Palo Alto, CA,
USA) equipped with a Supelcowax 10 (30 m × 0.25 mm i.d.; Sigma
Aldrich Co., Seoul, Korea) column and a FID. The column was ini-
tially held at 40 ◦ C for 2 min, and programmed to rise to 115 ◦ C at a
rate of 5 ◦ C/min. The carrier gas was helium, and the total gas flow
rate was 50 mL/min. The injector and detector temperatures were
both 220 ◦ C.
3. Results and discussion
3.1. Enzyme screening

There have been numerous reports on biodiesel synthesis using

2.2. Lipase-catalyzed transesterification various lipases. In particular, three immobilized lipases, namely
Novozym 435 [24–26], Lipozyme TL IM [27–29], and Lipozyme RM
For the lipase-catalyzed transesterifications, blended alcohols IM [18] have been studied widely. However, in these previous stud-
of methanol and ethanol were prepared for use as acyl accep- ies, transesterifications were carried out with only a single acceptor
tors. The molar proportions of methanol in the blended alcohols such as methanol or ethanol.
were 0, 20, 40, 60, 80, and 100 mol%; 0 mol% methanol indicates
100 mol% ethanol. The reaction was performed in a 25 mL screw-
capped Erlenmeyer flask. A substrate (3 g) consisting of a mixture
of soybean oil and the blended alcohol in a molar ratio of 1:3
were combined in the flask and a lipase (2.5 to 15% of total sub-
strate weight) was added. The reaction mixture was agitated in
an orbital shaker water-bath (model G76; New Brunswick Scien-
tific Co., Inc., New Brunswick, NJ, USA) operating at 300 rpm and
temperatures from 20 to 50 ◦ C. Individual samples were removed
at selected times and analyzed. All trials were conducted in
duplicate.

2.3. Analysis of products

When the reaction was complete, the product mixtures were

filtered through a 0.45 ␮m nylon microfilter to remove lipase. The
yield of fatty acid esters in the reaction mixture was determined
using samples (30 mg) of the products obtained using the differ-
ent reaction conditions dissolved in 1 mL of chloroform. Tricaprin
(1 mg) was used as an internal standard. A gas chromatograph
(model 3800; Varian, Palo Alto, CA, USA) equipped with a DB-1ht
column (15 m × 0.25 mm i.d.; J&W Scientific, Folsom, CA, USA) and
a flame ionization detector (FID) was used for analysis. The col-
umn temperature was initially held at 120 ◦ C for 3 min, increased
to 370 ◦ C at a rate of 20 ◦ C/min, and held for 3 min at 370 ◦ C. The
carrier gas was helium, and the total gas flow rate was 50 mL/min.
The injector and detector temperatures were both 370 ◦ C. The yield
Fig. 1. Enzyme screening for synthesis of biodiesel by transesterification of soybean
oil and blended alcohols. Enzymes: A, Novozym 435 (from Candida antarctica); B,
Lipozyme RM IM (from Rhizomucor miehei); C, Lipozyme TL IM (from Thermomyces
lanuginose); D, lipase PS (from Burkholderia cepacia); E, lipase AYS (from Candida
rugosa); F, lipase OF (from Candida rugosa). The reaction was performed at a molar
ratio of 1:3 (soybean oil to alcohol), temperature of 40 ◦ C, and enzyme loading of 5%
of total substrate weight for 12 h. The molar proportions of methanol in the blended
alcohols were 0 (100 mol% ethanol) (), 20( ), 40( ), 60( ), 80( ), and 100 mol%
( ).
 T. Zhao et al. / Journal of Molecular Catalysis B: Enzymatic 107 (2014) 17–22 19

100 In our study, six enzymes, including four immobilized lipases,

were screened for their activities in the synthesis of biodiesel from
soybean oil and blended alcohols of methanol and ethanol as sub-
80 strates. The enzymes tested were Novozym 435 (from C. antarctica),
Yield of biodiesel (wt%)

Lipozyme TL IM (from Thermomyces lanuginose), and Lipozyme RM

IM (from R. miehei), lipase PS (from P. fluorescence), lipase AYS
60 (from C. rugosa) and lipase OF (from C. rugosa). Before the trials,
the enzymes were stored at 4 ◦ C until use and all the enzymes were
used at a condition as it was offered by a supplier. For these tri-
40
als, the molar ratio of oil to alcohol, temperature, enzyme loading,
and reaction time were maintained constant at 1:3, 40 ◦ C, 5% (of
total substrate weight), and 12 h, respectively. The molar propor-
20
tions of methanol in the blended alcohols were 0, 20, 40, 60, 80, and
100 mol%. Among the six enzymes tested, Novozym 435 exhibited
the highest esterification capability (Fig. 1). For Novozym 435, as
0
0 5 10 15 20
the molar proportion of methanol in the blended alcohol increased
from 0 to 60 mol%, no significant differences were observed in the
Reaction time (h) yields of biodiesel, and the maximum yield was ca. 95 wt%. How-
ever, a significant decrease in the yield of biodiesel was observed
Fig. 2. Effect of molar proportion of methanol in blended alcohols on the yield of
biodiesel in lipase-catalyzed transesterification of soybean oil and blended alcohols:
when the molar proportion of methanol was increased from 60
0 (100 mol% ethanol) (䊉), 20(), 40(), 60(), 80(), and 100 mol% (). The reaction to 100 mol%, and the ester yield was only 5 wt% with 100 mol%
was performed with Novozym 435 at a molar ratio of 1:3 (soybean oil to blended methanol as the acyl acceptor. Lipozyme TL IM showed similar
alcohol), temperature of 40 ◦ C, and enzyme loading of 5% of total substrate weight. trends to those observed with Novozym 435, except in the trial
with 60 mol% methanol, but the yields were significantly lower than
those with Novozym 435, in all trials. In contrast, for Lipozyme RM
IM, a yield of 46 wt% was obtained at 0 mol% methanol (100 mol%

Fig. 3. Effect of temperature on biodiesel yield in lipase-catalyzed transesterification of soybean oil and blended alcohols: (a) 0 mol% methanol (100 mol% ethanol), (b)
20 mol% methanol, (c) 40 mol% methanol, and (d) 60 mol% methanol. Symbols: 20 ◦ C (䊉), 30 ◦ C (), 40 ◦ C (), 50 ◦ C (♦). The reaction was performed with Novozym 435 at a
molar ratio of 1:3 (soybean oil to blended alcohol) and enzyme loading of 5% of total substrate weight.
20 T. Zhao et al. / Journal of Molecular Catalysis B: Enzymatic 107 (2014) 17–22
Fig. 4. Effect of enzyme loading on yield of biodiesel in lipase-catalyzed transesterification of soybean oil and blended alcohols: (a) 0 mol% methanol (100 mol% ethanol), (b)
20 mol% methanol, (c) 40 mol% methanol, and (d) 60 mol% methanol. Symbols: 2.5%(䊉), 5%(), 10%(), 15%(♦). The reaction was performed with Novozym 435 at a molar
ratio of 1:3 (soybean oil to blended alcohol) and temperature of 30 ◦ C.
ethanol), but the yields of biodiesel were negligible in all blended
alcohols. For the other enzymes tested, the yields of biodiesel were
minimal in all tests. Novozym 435 was therefore selected as a suit-
able biocatalyst for further experimental parameters.
3.2. Effect of methanol and ethanol proportion in blended alcohol
The effect of the molar proportion of methanol in the blended
alcohol on the synthesis of biodiesel by lipase-catalyzed transes-
terification of alcohol and soybean oil is shown in Fig. 2. The molar
proportions of methanol in the blended alcohols tested were 0
(100% ethanol), 20, 40, 60, 80, and 100 mol%. In these trials, the
molar ratio of oil to alcohol, temperature, and enzyme loading
were maintained constant at 1:3, 40 ◦ C, and 5% (of total substrate
weight), respectively. The yield of biodiesel increased significantly
when the molar proportion of methanol in the blended alcohol
decreased from 100 to 60 mol% during the first 8 h of reaction. How-
ever, when the molar proportion of methanol in the blended alcohol
was further decreased from 60 to 40 mol%, the incremental increase
in the yield of biodiesel was small. Significant differences were
not observed among the yields of biodiesel at 0, 20, and 40 mol%
methanol throughout the reaction. In these trials, a maximum yield
of ca. 95 wt% was obtained after 8 h. A similar maximum yield was
obtained at 60 mol% methanol at a reaction time of 24 h. However,
the yields of biodiesel at 100 and 80 mol% were only 8 and 32 wt%,
respectively.
Nelson et al. [30] reported that methanol was less efficient than
ethanol in a solvent-free system and the low methyl ester yield
of 25.7% was attributed to inhibition by methanol of C. antarctica
lipase activity. This was confirmed by Abigor et al. [23] who
reported that the conversions of palm kernel oil using methanol
and ethanol were 15% and 72%, respectively.
For subsequent temperature trials, four blends of the two acyl
acceptors, i.e., 0, 20, 40, and 60 mol% methanol, were used, because
the yields of biodiesel using 100 and 80 mol% methanol were low.
3.3. Temperature
The reaction temperature is one of the most important param-
eters in lipase-catalyzed transesterification. An increase in the
temperature can reduce mass transfer limitations by reducing the
mixture viscosity and enhancing the mutual solubility or diffusion
processes of substrates, and can also increase interactions between
enzyme particles and substrates [31]. However, temperatures that
are too high can induce irreversible denaturation of the enzyme
protein and shorten the half-life of the enzyme activity, although
some enzymes from microorganisms are stable and active at high
temperatures [12,32].
The effect of temperature on lipase-catalyzed transesterification
for biodiesel synthesis was investigated as a function of reaction
time in the range from 20 to 50 ◦ C (Fig. 3). The acyl acceptors used
were blended alcohols of methanol and ethanol containing 0, 20, 40,
 T. Zhao et al. / Journal of Molecular Catalysis B: Enzymatic 107 (2014) 17–22 21

and 60 mol% methanol. For these trials, the molar ratio of oil to alco- 30

hol and enzyme loading were maintained constant at 1:3 and 5% (of (a)
total substrate weight), respectively. At 20 ◦ C, the yields of biodiesel 25

Proportion of methanol (mol%)

were significantly lower than those at other temperatures through-

in residual blended alcohol

out the reaction for all acceptors tested. In the temperature trials
20
using 0 mol% methanol and 20 mol% methanol as acyl acceptors, a
similar trend was observed throughout the reaction, although there
was a slight difference between the yields of biodiesel at 20 ◦ C. For 15
both of these acyl acceptors, there was a significant increase in the
yield of biodiesel during the first 12 h of reaction when the reac-
10
tion temperature was increased from 20 to 40 ◦ C (Fig. 3(a) and (b)).
However, there was no significant difference between the yields of
biodiesel at 40 and 50 ◦ C throughout the reaction, and a maximum 5

yield of ca. 95 wt% was obtained at each temperature after 12 h. In

contrast, at 30 ◦ C, a maximum yield of ca. 95 wt% was obtained after 0
24 h. 0 5 10 15 20 25
In temperature trials involving 40 mol% methanol (Fig. 3(c)) as
an acyl acceptor, no significant differences among the yields of Reaction time (h)
biodiesel were observed throughout the reactions when the reac-
tion temperature was increased from 30 to 50 ◦ C; a maximum yield 50

of ca. 95 wt% was obtained after 24 h. In the temperature trials (b)

involving 60 mol% methanol as the acyl acceptor, a similar trend

Proportion of methanol (mol%)

40
was observed to those in the trials involving 40 mol% methanol.

in residual blended alcohol

However, the overall yields of biodiesel with 60 mol% methanol
were lower than those with 40 mol% methanol at all temperatures
30
tested.
In previous studies, enzymatic synthesis of biodiesel was con-
ducted at temperatures between 20 and 60 ◦ C [18,33]. In terms of
20
economy, the optimum reaction temperature should be as low as
possible but still lead to the highest conversion in the shortest reac-
tion time. Enzyme stability should also be considered when the
10
optimum temperature is selected. The relationships among tem-
perature, transesterification rate, and stability of Lipozyme TL IM in
repeated-batch processes were reported by Du et al. [34]. In their 0
study, although a rise in temperature from 30 to 40 ◦ C increased 0 5 10 15 20 25
the reaction rate, the enzyme stability in continuous-batch reac-
tions decreased by 50% at 40 ◦ C, whereas 90% enzymatic activity Reaction time (h)
was maintained at 30 ◦ C after 10 successive batches.
Overall, even though the reaction rate was faster at 40 ◦ C than 70
30 ◦ C, the similar maximum yield of biodiesel was obtained at both (c)
temperatures after 24 h. In general, a higher temperature enables 60
Pproportion of methanol (mol%)

to achieve the maximum yield of biodiesel faster, but it can also

in residual blended alcohol

induce lipase deactivation by nucleophiles. Thus we carried out the 50

subsequent experiments at 30 ◦ C and 40 ◦ C.
40
3.4. Enzyme loading
30
The number of active sites available for a reaction is proportional
to the enzyme loading, and is an important factor in enzymatic
20
reactions. For the synthesis of biodiesel using enzymatic methods,
a wide range of enzyme loadings, between 4 and 50% (of total sub-
10
strate weight) have been employed. The optimum enzyme loading
for transesterifications of sesame oil and mustard oil with ethanol
0
and methanol were found to be 5–10% in several previous stud-
0 5 10 15 20 25
ies [35,36]. Modi et al. [37] reported an optimum enzyme loading
of 10% for solvent-free transesterification of sunflower oil. In cases Reaction time (h)
where tert-butanol was used as the solvent, low enzyme loadings
Fig. 5. Changes in molar proportion of methanol in residual blended alcohol in reac-
of 4–6% gave good conversions in the transesterification of waste
tion mixtures obtained at different reaction times: (a) 20 mol% methanol, (b) 40 mol%
palm oil [36]. In contrast, Hernández-Martín and Otero [33] used methanol, and (c) 60 mol% methanol. The reaction was performed with Novozym
a high enzyme loading of 50% for the transesterification of fatty 435 at a molar ratio of 1:3 (soybean oil to blended alcohol), temperature of 30 ◦ C,
acids from various vegetable oils with excess ethanol. However, and enzyme loading of 5% (for 20 and 40 mol% methanol) and 10% (for 60 mol%
it is a crucially important to choose the optimum enzyme loading methanol) of total substrate weight.

carefully to ensure economic feasibility. The effect of enzyme load-

ing on biodiesel synthesis by lipase-catalyzed transesterification of
blended alcohols and soybean oil as a function of reaction time is
shown in Fig. 4. The enzyme loading range was between 2.5 and
22 T. Zhao et al. / Journal of Molecular Catalysis B: Enzymatic 107 (2014) 17–22
15% of the total substrate weight, and the reaction temperatures
were 30 ◦ C and 40 ◦ C.
When the reaction temperature is 30 ◦ C, for all trials involv-
ing enzyme loadings greater than 10%, yields of ca. 95 wt% were
obtained after12 h, but there were no significant increases in the
yield when the reaction time was further increased. In the enzyme
loading trials, for an enzyme loading of 2.5%, the yields were lower
than those obtained using enzyme loadings greater than 5%. Mean-
while the yields differed depending on the molar proportion of
methanol in the blended alcohol. In the trials of three blends of
the two alcohols, i.e., 0, 20, and 40 mol% methanol, at an enzyme
loading of 5%, yields of over 95 wt% were observed for a reaction
time of 24 h; However, for the trials with 60 mol% methanol, yields
of over 95 wt% were obtained at an enzyme loading of 10% and a
reaction time of 24 h, whereas only ca. 85 wt% yield was obtained
at an enzyme loading of 5%.
In the trials of 40 ◦ C, very similar reaction trends were obtained
to the trials of 30 ◦ C (data not shown). Even though the reaction
rate was slightly faster compared to the trials of 30 ◦ C, there was
no difference between the maximum yields of biodiesel at 30 ◦ C
and 40 ◦ C at all the enzyme loadings tested. Ognjanovic et al. [38]
reported higher deactivation effect of the nucleophile on the lipase
as temperature increased. Taking the lipase stability and economic
reasons into consideration, we selected 30 ◦ C for the optimum tem-
perature.
Consequently, at 30 ◦ C selected as an optimum temperature, to
achieve yields of over 95 wt% within 24 h, the optimum enzyme
loadings for 0, 20, and 40% methanol were 5%, whereas the optimum
enzyme loading for 60 mol% methanol was 10%.
3.5. Relative consumption rates of methanol and ethanol
In order to compare the consumption rates of methanol and
ethanol during the transesterification, the changes in the molar
proportion of methanol in the residual blended alcohol were stud-
ied (Fig. 5). Transesterification was carried out at the optimum
condition selected in the previous sections. Regardless of the ini-
tial proportion of methanol in the blended alcohol, the proportion
of methanol in the residual alcohol decreased significantly dur-
ing the initial 12 h. For trials with 20, 40, and 60 mol% methanol,
the proportions of methanol in the residual alcohols after 24 h
were 7, 19, and 40 mol%, respectively. These results indicate greater
consumption preference for methanol than for ethanol during
transesterification. In chemical transesterification with both alka-
line and acid catalysts, it has been reported that the reactivity of
methanol was higher than that of ethanol, because methanol has
a higher nucleophilicity and lower stearic hindrance than ethanol
[12,31,32].
In the present study, ethanol acts not only as a solvent but also as
a reactant. Thus, when a blended alcohol of methanol and ethanol is
used as an acyl acceptor, it is anticipated that the higher reactivity
of methanol results in more ethanol remaining in the reaction mix-
ture, which can possibly minimize lipase deactivation by methanol.
In addition, the increased mutual solubility of the alcohol and oil
as a result of the presence of ethanol can increase the reaction rate
for the production of biodiesel.
4. Conclusions
A blended alcohol of methanol and ethanol was used as an acyl
acceptor for lipase-catalyzed transesterification. Use of blended
alcohols engendered the successful results, although proportions
of methanol higher than 60 mol% in blended alcohol adversely
affected the yield of biodiesel. On the other hand, the reactivity of
methanol in the transesterification was higher than that of ethanol.
Our results importantly indicate that higher methanol consump-
tion results in more ethanol remaining in the reaction mixture and
show possible extension of lipase activity. It also increases the sol-
ubility of the oil in the alcohol, yielding faster reaction rate. Thus,
the employment of the blended alcohols of methanol and ethanol
as an acyl acceptor for the transesterification has positive effect on
the enzymatic biodiesel production.
Acknowledgment
This research was supported by the Basic Science Research Pro-
gram through the National Research Foundation of Korea (NRF),
funded by the Ministry of Education, Science, and Technology
(2013R1A1A2006050).
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