Renewable Energy 130 (2019) 489e494

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Renewable Energy
journal homepage: www.elsevier.com/locate/renene

Immobilized lipase-catalyzed esterification for synthesis of

trimethylolpropane triester as a biolubricant
Heejin Kim a, Nakyung Choi b, Yangha Kim c, Hak-Ryul Kim d, Junsoo Lee e,
In-Hwan Kim a, b, *
a
Department of Public Health Sciences, Graduate School, Korea University, 145 Anam-Ro, Sungbuk-Gu, Seoul, 02841, Republic of Korea
b
Department of Integrated Biomedical and Life Sciences, Graduate School, Korea University, 145 Anam-Ro, Sungbuk-Gu, Seoul, 02841, Republic of Korea
c
Department of Nutritional Science and Food Management, Ewha Womans University, Seoul, 03760, Republic of Korea
d
School of Food Science and Biotechnology, Kyungpook National University, Daegu, 702-701, Republic of Korea
e
Division of Food and Animal Sciences, Chungbuk National University, Cheongju, Chungbuk, 28644, Republic of Korea

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

Article history: Synthetic oleochemical esters of polyols and fatty acids are biodegradable and possess desirable technical
Received 13 December 2017 and ecological properties. Trimethylolpropane (TMP) triester has been widely applied as a hydraulic fluid.
Received in revised form TMP triester was effectively synthesized by lipase-catalyzed esterification from TMP and high oleic fatty
25 April 2018
acid from palm oil using an immobilized lipase. The immobilized lipase was prepared with liquid Lip-
Accepted 21 June 2018
ozyme TL 100 L from Thermomyces lanuginosus with Duolite A568 as a carrier. The effects of temperature,
Available online 22 June 2018
enzyme loading, vacuum level, and water activity of the enzyme on the synthesis of TMP triester were
investigated. The optimum temperature, enzyme loading, and vacuum level were 60
C, 15% (based on
Keywords:
Biolubricant
total substrate), and 6.7 kPa, respectively. The optimum water activity range of the enzyme was 0.5e0.9.
Duolite A568 Under the optimum conditions, the maximum conversion reached up to 95% after 9 h. No significant
Immobilized lipase differences in physical properties were observed between TMP triester from this study and a commercial
Thermomyces lanuginosus TMP triester prepared by chemical catalyst.
Trimethylolpropane triester © 2018 Elsevier Ltd. All rights reserved.

1. Introduction have high thermo-oxidative stability, high viscosity indices, good

As pollution and environmental health have become increas-
ingly important public issues, interest in biolubricants has grown
because they are biodegradable and environmentally friendly [1].
Because of this trend, biodegradable base stocks have replaced
mineral oil base stocks. Vegetable oil based lubricants are biode-
gradable and have low eco-toxicity compared with mineral oil
based lubricants. However, they have several drawbacks, namely
low thermal, oxidative, and hydrolytic stabilities and poor low
temperature fluidity because of high pour points [2]. Synthetic
biolubricants have been developed to overcome these limitations.
Among the synthetic lubricants, synthetic esters of polyols and
fatty acids (FA) are considered as environmentally friendly sub-
stitutes to mineral oil based lubricants because they have suitable
properties for lubricant application. Synthetic esters of polyols and
FA generally show good performance at low temperatures, and
* Corresponding author. Department of Public Health Sciences, Graduate School,
Korea University, 145 Anam-Ro, Sungbuk-Gu, Seoul, 02841, Republic of Korea.
E-mail address: k610in@korea.ac.kr (I.-H. Kim).
antiwear performance, and low evaporation properties [1]. There-
fore, these esters are suitable for use as high performance lubri-
cants in industry. Among the polyols, trimethylolpropane (TMP) is
commonly used to synthesize TMP triester, because TMP has high
performance and moderate price level [3]. TMP triester is an
important lubricant and has been widely applied as a hydraulic
fluid, crank case lubricant, high temperature grease, and
compressor oil [4].
A number of studies have investigated the synthesis of TMP
esters via esterification of TMP with free FA or FA methyl esters
using a homogeneous or heterogeneous chemical catalyst [3,5e7].
Meanwhile, there have also been several reports on the synthesis of
TMP esters by esterification using lipases as the biocatalyst. The
lipases most frequently used for synthesis of TMP esters are
Novozym 435 from Candida antarctica, Lipozyme RM IM from
Rhizomucor miehei, and Candida rugosa lipase. For example,
Åkermanet al. [8] achieved 96% conversion using Novozym 435 for
esterification of TMP ester from oleic acid in 24 h. In studies of the
synthesis of TMP triester by transesterification using C. rugosa
lipase or Lipozyme RM IM [9,10], conversion of 64% was achieved

https://doi.org/10.1016/j.renene.2018.06.092
0960-1481/© 2018 Elsevier Ltd. All rights reserved.
490 H. Kim et al. / Renewable Energy 130 (2019) 489e494
with C. rugosa lipase after 24 h and conversion of 90% was achieved
with Lipozyme RM IM after 66 h. Overall, Novozym 435 is the most
effective lipase considering reaction rate and conversion in the
synthesis of TMP triester. However, when the immobilized lipase
prepared in this study was used for synthesis of TMP triester, the
reaction rate was much faster and the conversion was much higher
than with Novozym 435.
The goal of this study was to synthesize TMP triester from TMP
and FA using an immobilized lipase. The immobilized lipase was
prepared using liquid Lipozyme TL 100 L from Thermomyces lanu-
ginosus and Duolite A568 as a carrier. The effects of reaction tem-
perature, enzyme loading, vacuum, and water activity of the
enzyme were investigated. The physical properties of TMP triester
synthesized in this study were determined and compared with that
of a commercial TMP triester prepared by chemical catalyst.
2. Materials and methods
2.1. Materials
High oleic fatty acid (HOFA) from palm oil was used as the
substrate for synthesis of TMP triester. HOFA was donated by
ILSHINWELLS (Cheongju, Republic of Korea). The FA compositions
of the HOFA were oleic acid (C18:1n-9, 80%), linoleic acid (C18:2n-6,
11%), palmitic acid (C16:0, 6%), stearic acid (C18:0, 2%) and myristic
acid (C14:0, 1%). TMP and commercial TMP triester prepared by
chemical catalyst were donated by Ohsung Chemical Ind. Co., Ltd.
(Incheon, Republic of Korea). Liquid Lipozyme TL 100 L and Lip-
ozyme TL IM from T. lanuginosus, Novozym 435 from C. antarctica
and Lipozyme RM IM from R. miehei were purchased from Novo-
zymes (Seoul, Republic of Korea). Lipase OF from C. rugosa was
purchased from Meito Sangyo Co., Ltd. (Tokyo, Japan). Lipase PS
from Pseudomonas fluorescens and Lipase AYS from C. rugosa were
purchased from Amano Enzymes (Troy, VA, USA). Duolite A568 was
purchased from Rohm and Haas (Chauny, France). All of the other
chemicals used in this study were of analytical grade, unless
otherwise stated.
TMP triester
Conversionð%Þ ¼ 100
FA þ TMP monoester þ TMP diester þ TMP triester
2.2. Enzyme immobilization
Immobilization of enzyme was performed as described in our
previous study [11]. The enzyme solution was prepared by mixing
liquid Lipozyme TL 100 L (120 mL) with sodium phosphate buffer
(30 mL, 50 mM, pH 7.0). The enzyme solution (150 mL) was added
to a flask containing Duolite A568 (15 g), which acted as a carrier.
This mixture was shaken at 250 rpm and incubated at 30
C for 17 h
in an orbital shaker. The carrier was then separated from the
enzyme solution by filtration and immediately washed with buffer
solution (150 mL) to remove unbound enzyme. The carrier with the
immobilized enzyme was dried overnight at room temperature and
then dried in a vacuum oven for 12 h at 40
C. The immobilized
enzymes were stored at 4
C before use.
2.3. Equilibration of water activity
The immobilized enzyme was pre-equilibrated in individual
sealed containers with saturated salt solutions of known water
activity. The salts used were LiCl (aw ¼ 0.11), MgCl2 (aw ¼ 0.33),
Mg(NO3)2(aw ¼ 0.53), NaCl (aw ¼ 0.75), K2CO4(aw ¼ 0.97). The
equilibration process was carried out at 25
C for over 24 h.
2.4. Lipase-catalyzed esterification
Lipase-catalyzed esterification of TMP with FA were carried out
in a 50-mL water-jacketed glass vessel. The scheme of the reaction
was shown in Scheme 1. TMP (0.4 g, 3.1 mmol) and FA (2.6 g,
9.2 mmol) were placed in a reactor preheated to the desired tem-
perature using a water circulator. The reaction was initiated by
adding enzyme to the substrate mixture with stirring at 250 rpm
under vacuum. The vacuum level was controlled with a micro-
metering valve (Swagelok, Solon, OH, USA) and monitored using a
digital vacuum gauge (Teledyne, Thousand Oaks, CA, USA). Samples
(100 mL) were withdrawn from the reaction mixture at appropriate
intervals and dissolved in chloroform. Individual sample was then
filtered through a 0.45 mm nylon microfilter (Pall Corporation, Port
Washington, NY, USA). The samples were analyzed by gas chro-
matography. All experiments were conducted in triplicate.
2.5. Product analysis
The conversion in the reaction mixture was determined by
dissolving samples (10 mL) obtained under various reaction condi-
tions in chloroform (1 mL). A gas chromatograph (model 3800;
Varian Inc., 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 was used for the analysis. The column tem-
perature was held at 150
C for 2 min, increased to 370
C at a rate
of 25
C/min and then held at 370
C for 5 min. Helium at a flow rate
of 1.5 mL/min was used as the carrier gas and the split ratio was
1:50. The injector and detector temperatures were set at 340 and
350
C, respectively.
The conversion to TMP triester was calculated using the
following equation:
(1)
2.6. Determination of the physical properties
To determine the physical properties of the synthesized
TMP triester, a large-scale version of the lipase-catalyzed esterifi-
cation was conducted under the optimum conditions. After the
reaction, the final product was separated from the enzyme by
filtration.
The viscosity and viscosity index were determined according to
the ASTM D445 and ASTM D2270 methods, respectively. The vis-
cosity was calculated based on the time taken for the fluid to flow
through a glass capillary tube (Cannon-Fenske Routine viscometer,
Cannon Instrument Co., State College, PA). The kinetic viscosity was
obtained as the product of this time and the tube constant. The
viscosity index was calculated taking into account the product
viscosities at 40 and 100
C. The pour point and cloud point were
measured using a Tanaka Mini-Pour/Cloud Point Tester (Model
MPC-102 S, Tanaka Scientific Ltd., Tokyo, Japan) according to the
ASTM D2500 and ASTM D6749 methods, respectively. The color
was determined using a colorimeter (PFX195, The Tintometer Ltd,
 H. Kim et al. / Renewable Energy 130 (2019) 489e494 491

Scheme 1. Lipase-catalyzed esteirification of TMP with fatty acid using immobilized lipase.

Amesbury, UK) according to ASTM D1209. All experiments were
conducted in triplicate.
3. Results and discussion
3.1. Synthesis of TMP triester via lipase-catalyzed esterification
3.1.1. Enzyme screening
Six commercial lipases and the immobilized lipase prepared in
this study were screened for their activity in the synthesis of TMP
triester. The immobilized lipase was prepared by physical binding
of liquid Lipozyme TL 100 L to Duolite A568 as a carrier. The enzyme
activity was defined based on the conversion to TMP triester. In
these experiments, temperature, enzyme loading, vacuum level,
molar ratio of FA to TMP, and water activity of the enzyme were
kept constant at 60
C, 10% (based on total substrate), 0.67 kPa, 3:1,
and 0.3, respectively.
Only two lipases, namely, Novozym 435 and our immobilized
lipase, were effective for the synthesis of TMP triester (Fig. 1.).
Meanwhile, the other five commercial lipases did not synthesize
100
80
TMP triester (wt%)
TMP triester. For the experiments with Novozym 435, the
maximum conversion (ca. 84%) was achieved at 12 h. On the other
hand, with our immobilized lipase, a similar conversion (ca. 83%)
was achieved within only 6 h and the maximum conversion (ca.
95%) was obtained after 12 h. Therefore, our immobilized lipase is
more effective for the synthesis of TMP triester than Novozym 435,
which is the most effective lipase identified to date. In our previous
study [11], the same immobilized lipase was also effective for the
triacylglycerol (TAG) synthesis from a-linolenic acid-rich FA and
glycerol by esterification. Because the structure of TMP is similar to
that of glycerol, the immobilized lipase was applied for synthesis of
TMP triester. Consequently, TMP triester was also synthesized
efficiently via esterification of TMP with FA using the immobilized
lipase as a biocatalyst. Therefore, the immobilized lipase prepared
in this study was chosen as the lipase to investigate the effects of
the process parameters.
3.1.2. Effect of temperature
The reaction temperature is a crucial factor affecting lipase-
catalyzed esterification. Generally, increasing the temperature re-
duces the viscosity of solution and improves the solubility of a
compound. This facilitates interactions between the enzyme and
substrate and thus increases the reaction rate [12]. Even though
high temperatures cause an increase of the reaction rate, temper-
atures that are too high can deactivate enzymes and reduce the
enzyme stability and half-life [13]. The optimum temperature for

use of an enzyme depends on its source, the nature of immobili-

zation or chemical modification, the solvent, and the pH of the
60 reaction medium [14].
The effect of temperature on the synthesis of TMP triester via
lipase-catalyzed esterification was investigated (Fig. 2). The tem-
40 A
B perature range tested was 40e80
C. In these experiments, enzyme
C loading, vacuum level, molar ratio of FA to TMP, and water activity
D of the enzyme were kept constant at 10% (based on total substrate),
20
E 0.67 kPa, 3:1, and 0.3, respectively.
F
As the temperature increased from 40 to 60
C, both the reaction
G
0 rate and conversion increased significantly, but no significant dif-
0 2 4 6 8 10 12 ference was observed at temperatures between 60 and 70
C.
Meanwhile, the conversion decreased drastically as the tempera-
Reaction time (h) ture increased from 70 to 80
C. These results were consistent with
those from our previous study [11] on the synthesis of a-linolenic
Fig. 1. Enzyme screening for synthesis of TMP triester by lipase-catalyzed esterifica-
tion. The following enzymes were used: A, Lipase OF (from Meito Sangyo Co., Ltd.) acid enriched TAG using the same immobilized lipase as in this
from Candida rugosa; B, Lipase AYS (from Amano Enzymes) from Candida rugosa; C, study. These results indicated that the immobilized lipase
Lipase PS from Pseudomonas fluorescens; D, Lipozyme RM IM from Rhizomucor miehei; remarkably lost its activity in esterification of FA with TMP at
E, Lipozyme TL IM from Thermomyces lanuginosus; F, Novozym 435 Candida antarctica; temperatures higher than 70
C. Even though similar maximum
and G, the immobilized lipase from Thermomyces lanuginosus. In these experiments,
temperature, enzyme loading, vacuum level, molar ratio of FA to TMP, and water ac-
conversions were obtained after 12 h at temperatures between 60
tivity of the enzyme were kept at 60
C, 10% (based on total substrate), 0.7 kPa, 3:1, and and 70
C, 60
C was chosen as the optimum temperature in
0.3, respectively. All experiments were conducted in triplicate. consideration of energy requirements and stability of the lipase.
492 H. Kim et al. / Renewable Energy 130 (2019) 489e494
Fig. 2. Effect of temperature on the synthesis of TMP triester by immobilized lipase-
catalyzed esterification. In these experiments, enzyme loading, vacuum level, molar
ratio of FA to TMP, and water activity of the enzyme were kept at 10% (based on total
substrate), 0.7 kPa, 3:1, and 0.3, respectively. All experiments were conducted in
triplicate.
3.1.3. Effect of enzyme loading
It is necessary to use an adequate amount of enzyme for an
effective reaction. Even though the reaction rate is proportional to
the enzyme loading [15,16], to reduce the enzyme loading is also
important for economic feasibility. Therefore, it is important to
determine the optimum enzyme loading for an enzymatic reaction
to achieve the highest level of efficiency.
The effect of enzyme loading on the synthesis of TMP triester via
lipase-catalyzed esterification was investigated (Fig. 3). The
enzyme loading range tested was 5e20% (based on total substrate).
In these experiments, temperature, vacuum level, molar ratio of FA
to TMP, and water activity of the enzyme were kept constant at
60
C, 0.67 kPa, 3:1, and 0.3, respectively.
During the first 4 h, the reaction rate and conversion increased
remarkably as the enzyme loading increased from 5 to 20%. For the
experiments using enzyme loadings of 15 and 20%, the conversion
Fig. 3. Effect of enzyme loading on the synthesis of TMP triester by immobilized
lipase-catalyzed esterification. In these experiments, temperature, vacuum level, molar
ratio of FA to TMP, and water activity of the enzyme were kept at 60
C, 0.7 kPa, 3:1, and
0.3, respectively. All experiments were conducted in triplicate.
approached equilibrium after 6 h. Although the reaction rate with
an enzyme loading of 20% was slightly faster than with a loading of
15%, the conversions obtained with these loadings were not
significantly different after the reaction reached equilibrium.
Meanwhile, with an enzyme loading of 5%, the conversion
increased very slowly and did not reach equilibrium throughout the
entire period studied. Therefore, taking into consideration the cost
of the operation, an enzyme loading of 15% was chosen as the op-
timum loading for synthesis of TMP triester.
3.1.4. Effect of vacuum
The amount of water in lipase-catalyzed esterification is also a
crucial factor. Water affected the conversion and reaction rate for
lipase-catalyzed esterification in nonaqueous media [17]. For
esterification/hydrolysis reactions, if the water produced by the
reaction of TMP with FA is removed, the reaction equilibrium can be
moved toward esterification. There are some methods for control-
ling the water content in a reaction mixture [15,18]. A vacuum can
be an effective method for controlling the water content in the
reaction mixture [19].
The effect of the vacuum level on the synthesis of TMP triester
via lipase-catalyzed esterification was investigated (Fig. 4). In these
experiments, temperature, enzyme loading, molar ratio of FA to
TMP, and water activity of the enzyme were kept constant at 60
C,
15% (based on total substrate), 3:1, and 0.3, respectively. The range
of pressures tested was 0.7e40.0 kPa. At 40.0 kPa, the conversion
was only 88% at 12 h. Meanwhile, the reaction rate and conversion
increased significantly as the vacuum level increased from 40.0 to
13.3 kPa. The reaction rate and conversion increased slightly when
the vacuum level was increased further to 6.7 kPa. However, the
reaction rate decreased when the vacuum level was increased
beyond 6.7 kPa. After 6 h, no significant differences among the
conversions were observed for vacuum levels between 0.7 and
6.7 kPa. Although a high vacuum level can cause a reduction of
enzyme activity because of the deficiency of essential water for the
lipase activity [20], application of a suitable vacuum level is
essential to inhibit hydrolysis and improve the yield. Hong et al.
[21] reported that Novozym 435 required a low water content for
enzymatic esterification of conjugated linoleic acid with glycerol,
and the equilibrium could be shifted toward the synthesis reaction
by the removal of water. However, as the vacuum level was too
Fig. 4. Effect of vacuum level on the synthesis of TMP triester by immobilized lipase-
catalyzed esterification. In these experiments, temperature, enzyme loading, molar
ratio of FA to TMP, and water activity of the enzyme were kept at 60
C, 15% (based on
total substrate), 3:1, and 0.3, respectively. All experiments were conducted in triplicate.
 H. Kim et al. / Renewable Energy 130 (2019) 489e494 493

high, the reaction rate decreased. A similar tendency was also different optimum water activities. For the synthesis of butyl
observed in this study. butyrate by transesterification, Chowdary et al. [23] found the op-
Consequently, vacuum levels higher than 6.7 kPa were not timum water activities for lipases from C. rugosa and Penicillium
suitable because the excessively high vacuum level removed the roqueforti were both 0.33, those for lipases from Mucor javanicus
essential water in the lipase and reduced the reaction rate. Mean- and Rhizopus oryzae were both 0.54, and that of lipase from
while, a vacuum level of 40.0 kPa was also not suitable because Aspergillus niger was 0.75. For the synthesis of phospholipid con-
water formed in the reaction was not removed effectively, which taining n-3 PUFA via acidolysis, Kim et al. [24] found that the op-
could lead to hydrolysis. Hence, a vacuum of 6.7 kPa was selected as timum water activity of immobilized phospholipase was 0.65. It is
the optimum level for synthesis of TMP triester. clear that different lipases in organic media have quite different
water activity requirements for optimum efficacy.
For the synthesis of structured lipid by acidolysis with olive oil
3.1.5. Effect of the initial water activity
and capric acid using Lipozyme TL IM from T. lanuginosus, Oh et al.
There is a critical lower limit for the water content, and below
[25], indicated that capric acid incorporation into olive oil increased
this, enzymes will not be able to maintain their catalytic activities
when the water activity was continuously increased up to 0.80.
[14,22]. This critical water content is needed to maintain the three-
Meanwhile, Svensson and Adlercreutz [26] studied the effect of acyl
dimensional configuration of the enzyme necessary for its catalytic
migration in Lipozyme TL IM-catalyzed esterification using TAG,
activity. However, excessive water can induce negative effects.
and found that the reaction rate with a water activity of 0.35 was
Therefore, to maximize the efficiency, to identify the optimum
the fastest and that with a water activity of 0.84 was the slowest.
water content in the enzyme is important for an enzymatic
These results show that the optimum water activity of lipase from
reaction.
T. lanuginosus varies depending on the type of substrate used or
The effect of the water activity of the enzyme on the synthesis of
reaction system.
TMP triester via lipase-catalyzed esterification was investigated
(Fig. 5). The water activity range tested was 0.11e0.97. In these
3.2. Physical properties of the final product
experiments, temperature, enzyme loading, molar ratio of FA to
TMP, and vacuum level were kept constant at 60
C, 15% (based on
After the reaction parameters were optimized, the physical
total substrate), 3:1, and 6.7 kPa, respectively.
properties of the TMP triester were tested. As a polyolester, the
As the water activity increased from 0.11 to 0.33, the reaction
physical properties of TMP triester meet the requirements for use
rate and the conversion increased significantly throughout the
as a lubricant. The physical properties of the TMP triester synthe-
entire reaction (Fig. 5). With a water activity of 0.11, the reaction
sized in this study were compared with those of a commercial TMP
rate was significantly slower than at the other water activities
triester prepared by chemical catalyst as the reference, and to the
tested in this study. This was probably caused by a lack of essential
properties of the HOFA substrate. The physical properties investi-
water for the catalytic activity of the enzyme. Meanwhile, during
gated in this study were the viscosity, viscosity index, pour point,
the first 4 h of reaction, the reaction rate increased slightly as the
cloud point, and color (Table 1).
water activity increased from 0.33 to 0.75. However, increasing the
Viscosity is one of the most important properties of a lubricant,
water activity from 0.75 to 0.97 did not have a remarkable effect on
because it is related to the ability of the lubricant to efficiently
the conversion. Although slight differences in the reaction rates
lubricate the contact surface. The viscosities of the TMP triester
were observed with water activities between 0.53 and 0.97, similar
from this study and the commercial TMP triester were not signifi-
maximum conversions (ca. 95%) were achieved after 9 h. These
cantly different. The viscosity index indicates the change in vis-
results indicate that the immobilized lipase is very effective for
cosity depending on the temperature change. The high viscosity
synthesis of TMP triester when the water activity is higher than 0.5.
index implies that the change in viscosity is small. In general,
It has been reported that lipases from different sources have
mineral oils show a viscosity index of approximately 100, but
vegetable oil-based lubricants have higher viscosity index than
100 mineral oils [27]. Both the TMP triester from this study and the
commercial TMP triester had high viscosity indices of approxi-
mately 200 which implies that these TMP triesters are desirable for
80 a wide temperature range. The cloud point and pour point are also
TMP triester (wt%)

important factors to indicate the lubricants properties. The cloud

point and pour point of the TMP triester from this study were as
60 low as those of the commercial TMP triester. Generally, a lubricant
with low pour point has efficiently worked in low temperature
environments. Meanwhile, the TMP triester from this study was
40 much lighter in color than the commercial TMP triester, which was
aw 0.11
aw 0.33
similar to the result reported by Åkerman et al. [8]. Consequently, it
was demonstrated that the TMP triester from this study has iden-
20 aw 0.53
tical physical properties when compared to the commercial TMP
aw 0.75
triester.
aw 0.97
0
4. Conclusions
0 2 4 6 8 10 12
Reaction time (h) The immobilized lipase from T. lanuginosus with Duolite A568 as
a carrier is a promising biocatalyst for synthesis of TMP triester. The
Fig. 5. Effect of the water activity of the enzyme on the synthesis of TMP triester by conversion to TMP triester decreased markedly as temperature
immobilized lipase-catalyzed esterification. In these experiments, temperature,
enzyme loading, molar ratio of FA to TMP, and vacuum level were kept at 60
C, 15%
increased from 70 to 80
C. TMP triester was synthesized effectively
(based on total substrate), 3:1, and 6.7 kPa, respectively. All experiments were con- when the water activity of the immobilized lipase was higher than
ducted in triplicate. 0.53. The final product, composed of 95 wt% TMP triester, 3.5 wt%
494 H. Kim et al. / Renewable Energy 130 (2019) 489e494
Table 1
Physical propertiesa of TMP triester synthesized in this study, a commercial TMP triester prepared by chemical catalyst, and high oleic fatty acid (HOFA).
Product Viscosity (mm2/s) Viscosity index, (VI)
40
C 100
C
TMP triester Ib 46.2 9.5 195
TMP triester IIc 46.6 9.5 194
HOFA 18.7 4.8 189
a
Values represent the average of triplicate determinations from different experiments.
b
TMP triester was synthesized by immobilized lipase-catalyzed esterification under the optimum conditions.
c
Commercial TMP triester prepared by chemical catalyst.
TMP diester, and 1.5 wt% FA, was obtained after 9 h under the op-
timum conditions. These results are even better than those from
earlier studies on the synthesis of TMP triester using other com-
mercial lipases. The TMP triester synthesized in this study has
physical properties that are similar to a commercial TMP triester
prepared by a chemical method.
Acknowledgement
This research was supported by the Yang Young Foundation.
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