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Journal of the Taiwan Institute of Chemical Engineers

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A novel strategy of biodiesel production from wet microalgae by

direct saponification–esterification conversion (DSEC)
Ying-Ru Fang, Yun Yeh, Hwai-Shen Liu∗
Department of Chemical Engineering, National Taiwan University, Taipei, Taiwan

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

Article history: Microalgae has become a potential resource of biodiesel in recent years for its fast growth, non-food
Received 5 October 2017 source, and CO2 capture ability. However, efficient and economic production of biodiesel from microalgae
Revised 30 November 2017
is still a challenge. In this study, we presented a novel strategy of direct saponification–esterification
Accepted 1 December 2017
conversion (DSEC) from wet microalgae (Chlorella vulgaris) to biodiesel and its associated kinetic model.
Available online xxx
The process was accomplished with two consecutive additions into wet algae. Firstly, addition of
Keywords: NaOH/methanol solution, lipids including triglycerides (TG) and free fatty acids (FFA) of wet algae were
Biodiesel released from disrupted algae and primarily converted into soaps by means of saponification. Then with
Wet microalgae further HCl/methanol addition, soaps were acidified into FFA and esterified into fatty acid methyl ester
Direct saponification–esterification (biodiesel). The experimental results supported the proposed mechanism and observed several advan-
conversion tages of DSEC process. Primarily, DSEC eliminates lipid extraction/purification step in the conventional
process, which usually accounts for more than half of production cost. Secondly, compared with the al-
kali catalyzed transesterification process, DSEC completely alleviates the restriction of incapable of dealing
with high content of water and FFA. On the other hand, compared with acid catalyzed transesterification
process, the production time of DSEC was only a fraction, less than 15 mins.
© 2017 Taiwan Institute of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

1. Introduction
Biodiesel is an eco-friendly alternative fuel [1–3] that has
gained significant attention these days. As a next-generation feed-
stock for biodiesel, microalgae can be cultivated in non-arable land
and compete very little with agricultural crops [4,5] plus many
of them could grow fast and accumulate high lipid content more
than 20 wt% of their dried biomass [6,7]. Meanwhile, their ability
to capture carbon dioxide and tolerate NOx and SOx from fuel gas
makes microalgae very favorable and feasible for biodiesel produc-
tion [8–10].
Biodiesel can be generated from microalgal lipid, mainly,
triglyceride (TG) and free fatty acid (FFA). Generally speaking, there
are two approaches to produce biodiesel from microalgae. One is
the conventional process that includes steps of cell disruption and
lipid extraction/purification, then followed by lipid conversion to
biodiesel. The other one is to combine lipid extraction and lipid
conversion into one operation, sometimes called direct conversion
process [11]. The major disadvantage of the former is the inten-
sive energy requirement for the extraction/purification of cellu-
lar lipid, while the latter could simplify the production process,
∗
Corresponding author.
E-mail address: hsliu@ntu.edu.tw (H.-S. Liu).
consequently the cost [12]. Either the conventional process or di-
rect conversion process, biodiesel is produced by lipid conversion
through transesterification of TG and esterification of FFA. Transes-
terification of TG [13] perhaps is the most common route of con-
verting feedstock into biodiesel. And the process combining the
lipid extraction and transesterification into one unit operation to
produce biodiesel is often called direct transesterification (DT). DT
was firstly described using macerated sunflower seeds incubated
in acid methanol. [11,14]. Furthermore, several studies concluded
that the direct process might achieve higher biodiesel yield com-
pared with the conventional process because of continuous opera-
tion without lipid loss in the extraction step [14–16].
Biodiesel production from microalgae by transesterification in-
volving the mixing of lipid with alcohol in the presence of the cat-
alysts to accelerate the reaction rate. Compared with acid catalysts,
transesterification with alkaline catalysts (such as NaOH, KOH, or
CH3 ONa) is superior in both reaction rate and conversion [17–19].
However, alkaline-catalyzed transesterification in the presence of
water or FFA [20] could significantly lower the biodiesel yield due
to saponification reaction [21–23]. That is, the alkaline hydrolysis
of both TG and FFA forms stable salt known as soap instead of
biodiesel. For example, Naik et al. [24] found that, in a KOH cat-
alyzed transesterification, the yield of biodiesel dropped from 97%
to 6% when the FFA content in the feedstock changed from 0.3 wt%

https://doi.org/10.1016/j.jtice.2017.12.001
1876-1070/© 2017 Taiwan Institute of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

Please cite this article as: Y.-R. Fang et al., A novel strategy of biodiesel production from wet microalgae by direct saponification–
esterification conversion (DSEC), Journal of the Taiwan Institute of Chemical Engineers (2017), https://doi.org/10.1016/j.jtice.2017.12.001
JID: JTICE
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2 Y.-R. Fang et al. / Journal of the Taiwan Institute of Chemical Engineers 000 (2017) 1–9
to 5.3 wt%. In order to prevent saponification, Freedman et al. sug-
gested that feedstock be restricted below 0.3 wt% of water and
0.5 wt% of FFA [25] when an alkaline catalyst is used.
Because of high FFA content in microalgae cell [26] with wa-
ter, acid catalysts are often used to avoid soap formation. Acid-
catalyzed transesterification has the advantage that, in the pres-
ence of water, biodiesel can be produced via esterification with
FFA and via transesterification with TG simultaneously. However,
water could inhibit the TG transesterification rate significantly.
Thus, reaction in acid-catalyzed transesterification usually requires
higher temperature and longer time as compared with alkali-
catalyzed transesterification [27–29]. That is, in an acidic environ-
ment, in comparison with esterification of FFA, biodiesel forma-
tion via transesterification of TG is much slower [28,29]. Therefore,
transesterification with acid catalysts has the advantage that both
FFA and TG can convert into biodiesel in the presence of water, but
the efficiency is often unsatisfactory because of TG transesterifica-
tion inhibition by water.
The development of two-step direct conversion that produced
biodiesel directly from biomass was conducted by several studies
[30–34] to deal with the presence of water and FFA. In the di-
rect acid/alkali conversion process, i.e. acid-catalyzed followed by
alkali-catalyzed reaction, it firstly undergoes esterification of FFA
with acid catalysts for a short time to reduce the FFA content from
microalgae cell and therefore avoid soap formation. Next, the reac-
tion undergoes transesterification of TG with alkaline catalysts to
obtain higher biodiesel yield at an efficient rate. Shiu et al. [30] re-
ported that, with two-step direct conversion, the biodiesel yields
from rice bran (of 3% and 33% FFA content) were better than those
by one-step alkali-catalyzed and by one-step acid-catalyzed. How-
ever, energy and efforts were still needed to remove moisture con-
tent of wet microalgae so that undesired saponification in alkali-
catalyzed step would not occur.
Instead of the direct acid/alkali conversion, in this work, we
proposed a direct alkali/acid conversion for biodiesel produc-
tion from wet microalgae. Although the proposed strategy is
only to reverse the sequence of adding acid/alkali, the ratio-
nales/mechanisms are very different. First, wet algae are added
with alkali/methanol solution to release lipid (TG and FFA) and
carry out saponification for both TG and FFA because of water
existence. Then acid/methanol solution is added to esterify the
soap into biodiesel. Therefore, a name of “direct saponification–
esterification conversion” (DSEC) is given because of the mecha-
nisms behind this. The primary advantage of a DSEC is the elimina-
tion of concerns of both water and FFA existence in the feedstock.
This is critical in the case of algae cells as well as microbial sys-
tem where water is abundant. Complete drying of these feedstock
for biodiesel production is often costly. Secondly, both saponifi-
cation and esterification are fast reactions. Other merits of direct
conversion characteristics still remain, such as no cell disruption
step/lipid purification, and it is a fast “one-vessel” reaction.
However, similar reaction schemes as DSEC were reported be-
fore. For example, Griffiths et al. [35] proposed a direct process for
wet microalgae with a first step of alkalized-catalyzed transesteri-
fication while 0–50% water content in feedstock, and followed by a
second step of acid-catalyzed transesterification. With the sequen-
tial combination of direct alkali/acid catalyst, a higher biodiesel
yield could be obtained compared with direct one-step conversion,
particularly in the presence of water. Another related study con-
ducted by Kumar et al. [36], the authors investigated that, for ei-
ther wet microalgae (water content ∼80 wt%) or freeze-dried mi-
croalgae (water content ∼0 wt%), reaction with alkaline catalyst
followed by acid catalyst had an optimum yield. Although these
authors investigated the process of direct alkali/acid conversion
similar to our proposed DSEC, “transesterification” (from TG) was
improperly referred as a main route for biodiesel production. In
Please cite this article as: Y.-R. Fang et al., A novel strategy of biodiesel production from wet microalgae by direct saponification–
esterification conversion (DSEC), Journal of the Taiwan Institute of Chemical Engineers (2017), https://doi.org/10.1016/j.jtice.2017.12.001
[m5G;December 15, 2017;20:5]
this work, we will elucidate that, in the DSEC, biodiesel is pro-
duced, not directly from TG by transesterification, but by esteri-
fication from “acidified soaps” . Meanwhile, the kinetic model is
proposed to further support the mechanisms of DSEC.
2. Experimental section
2.1. Microalgae preparation and culture condition
The freshwater green microalgae of Chlorella sp. ESP-6 was
a kind gift from professor Duu-Jong Lee at National Tai-
wan University and used as a model algal system. This mi-
croalgae was cultivated in a 2 L bottles (SCHOTT, Germany)
at 26 °C for 14 days. The medium using BBM [37] (pH
6.4∼7.0) consisted of 0.43 m M K2 PO4 , 1.29 mM KH2 PO4 , 0.3 mM
MgSO4 ·7H2 O, 2.94 mM NaNO3 , 0.17 mM CaCl2 ·2H2 O, 2.94 mM NaCl,
0.12 mM ETDA-Na4 /KOH, 0.55 mM FeSO4 ·7H2 O/H2 SO4, 0.1 mM
H3 BO3 , 4.9 × 10−3 mM ZnSO4 ·7H2 O, 1.2 × 10−3 mM MnCl2 ·2H2 O,
1 × 10−3 mM CuSO4·5H2 O, 2.75 × 10−4 mM CO(NO3 )2 ·6H2 O and
8.77 × 10−4 mM NaMoO4 ·2H2 O. The microalgae were cultivated at
a constant light intensity of 1325 LM and aerated the first 3 days of
air and 10% CO2 (with air) for later 11 days at a rate of 20 mL/min.
The wet microalgae used in the experiments were harvested and
washed by centrifugation at 60 0 0 rpm for 10 min, rinsed in dis-
tilled water and re-centrifuged at 60 0 0 rpm for 10 min.
2.2. Biodiesel production by DSEC
In a typical case, the wet biomass sample (25–100 g-dried-
cell/L) was mixed with 1 mL methanol containing 2 M NaOH in a
test tube. Each tube was sealed with a PTFE screw cap and vor-
texed for 10 s, then placed in the water bath for reaction. Initially,
in the first step, lipid in the microalgae cell was released and pri-
marily converted into soap (fatty acid sodium salts) by saponifi-
cation with NaOH as alkaline catalyst at 100 °C for 20 min. After
that, 2 mL methanol containing 1.8 M HCl was added to the mix-
ture for esterification for another 15 min. The soap was first neu-
tralized into FFA and esterified into fatty acid methyl esters (FAME,
i.e. biodiesel).
Biodiesel production from wet microalgae by DSEC process was
accomplished with two consecutive additions and biodiesel yield
was calculated based on a percentage of dried microalgae biomass
as Eq. (1) :
mass of biodiesel obtained (g )
Biodiesel yield(g/g ) = (1)
dried microalgae weight (g )
2.3. Microalgae concentration in DSEC
To investigate the extraction model and kinetic model of the
DSEC process, different concentrations of wet microalgae biomass
were carried out. Because volume change, concentrations of wet
microalgae were carefully calculated in both saponification and es-
terification steps as defined in Eqs. (2) and (3), respectively for
clarity.
dried weight of microalgae biomass(g )
[C]I = (2)
reaction volume in the saponification step (L )
dried weight of microalgae biomass(g )
[C]II = (3)
reaction volume in the esterification step (L )
where subscripts Ⅰ and Ⅱ stand for the reaction in the first step as
the saponification step and the second step referred to the esteri-
fication step, respectively. The ranges of [C ]Ⅰ = 8.67 g/L ∼24.67 g/L,
[C ]Ⅱ = 5.14 g/L∼15.43 g/L were evaluated.
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Fig. 1. Reactions in the saponification step (a) transesterification of TG; (b) saponification of TG; (c) saponification of FAME; (d) saponification of FFA.
2.4. Analysis
Experimental samples were extracted in hexane with an inter-
nal standard hexadecane and analyzed by a GC-FID (Perkin Elmer)
with the injection volume of 1 μL. The quantification and qualifica-
tion of products in each step is given below.
2.4.1. Analysis of the saponification step
At the end of saponification step, reaction broth might include
triglycerides, diglyceride, monoglyceride and fatty acid sodium salt
(soap). Thus, 0.33 mL HCl/CH3 OH of 9.22 M was added to convert
soap into free fatty acids before hexane extraction. Then acidified
samples after hexane extraction were analyzed by a GC with a ZB-
5HT capillary column (15 m × 0.32 mm i.d. × 0.32 μm film thickness,
Zebron Corporation, USA). Conditions of the GC were 5 mL/min car-
rier gas (nitrogen), 320 °C injection temperature, 320 °C detector
temperature, with the column temperature 100 °C initially, heated
up to 155 °C (2 min hold) at 10 °C /min followed by increasing
to 175 °C at 5 °C/min and then increasing to 310 °C (2 min hold)
at 30 °C /min. Trioleate (Triglyceride, C18:1), Dioleate (Diglyceride,
C18:1), Palmitic acid (Free fatty acid, C16:0), Oleic acid (Free fatty
acid, C18:1) and FAME mix C16-C20 of GLC-20 from Sigma-Aldrich
Corporation, USA were used as standard.
2.4.2. Analysis of the esterification step
FAME produced from wet Chlorella sp. microalgae was obtained
in the esterification step. As a result, samples after hexane extrac-
tion in this step were mainly FAME and possible some unreacted
FFA. Samples in the mixture were quantified in a GC-FID with a
BPX-70 capillary column (30 m × 0.53 mm i.d. × 0.5 μm film thick-
ness, SGE analytical science, Australia) with 5 mL/min N2 as the
carrier gas, 250 °Cinjector temperature, 250 °Cdetector temperature
and a column temperature increased from 85 °C to 150 °C (4 min
hold) at 30 °C/min followed by increasing to 170 °C (4 min hold)
at 8 °C/min. Palmitic acid (Free fatty acid, C16:0), oleic acid (Free
Please cite this article as: Y.-R. Fang et al., A novel strategy of biodiesel production from wet microalgae by direct saponification–
esterification conversion (DSEC), Journal of the Taiwan Institute of Chemical Engineers (2017), https://doi.org/10.1016/j.jtice.2017.12.001
[m5G;December 15, 2017;20:5]
3
fatty acid, C18:1) and FAME mix C16-C20 of GLC-20 and GLC-50
from Sigma-Aldrich were used as standard.
3. Reaction kinetics
3.1. Theoretical background of DSEC
Since TG and significant FFA (ca 5.11 wt% [38]) exist in mi-
croalgae cell, lipid is expressed to include both TG and FFA in this
paper. The following schemes are proposed for biodiesel produc-
tion from wet microalgae by DSEC. First, cells are disrupted and
lipid is released from wet microalgae cell by NaOH together with
methanol [39]. Secondly, as soon as lipid is released to the solu-
tion, lipid undergoes the reactions shown in Fig. 1 in the presence
of water. That is, TG reacts with methanol through transesterifi-
cation and converts into FAME [40–42] (Fig. 1a). Meanwhile both
TG (Fig. 1b) and FFA (Fig. 1d) may also undergo alkaline hydrolysis
of esters, known as saponification [21,43]; and therefore, soap is
produced. Likewise, because of high water content, FAME from TG
undergoes saponification and converts into soap (Fig. 1c), too. Ac-
tually, soap formation in the presence of water is primarily what
happen for both TG and FFA in this saponification step. Finally,
with HCl/methanol addition in the 2nd step, soap is acidified into
FFA and then FFA undergoes acid hydrolysis, referred to esterifica-
tion [44], and then converts into FAME as shown in Fig. 2. That is,
biodiesel production primarily comes from the esterification step
by DSEC from lipid of microalgae.
In Figs. 1 and 2, Ri represents different numbers of carbon
atoms of fatty acids, either saturated or unsaturated. For simplic-
ity in describing reaction schemes, we name fatty acid base (–FA)
as shown in Fig. 3a and the structure of glycerol base (G–) as
shown in Fig. 3b so that TG can be expressed as one glycerol base
and three fatty acids base, G(FA)3 and other structures involved
in our reaction schemes could be found in Table 1. Since DSEC is
accomplished with two consecutive additions into the microalgae
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4 Y.-R. Fang et al. / Journal of the Taiwan Institute of Chemical Engineers 000 (2017) 1–9

Fig. 2. Reactions in the esterification step (a) neutralization of soap; (b) esterification of FFA.

in FFA (one –FA) and TG (three –FA’s) shown in Eq. (4):

[Lipid] = 3[G(FA )3 ] + [H − FA] (4)

As in Fig. 1, lipid is the reactant and soap (Na–FA) is the main

product through saponification with possible some un-reacted
CH3 –FA. Hence, lipid can also be calculated by Eq. (5) if lipid avail-
able is completely transformed (with excess NaOH):

[Lipid] = [Na − FA]I + [CH3 − FA]I (5)

Recent studies have proposed a first-order differential equation

Fig. 3. The expression of (a) fatty acid base (-FA); (b) glycerol base (G-). of extraction model to describe how lipid was released during the
extraction step in the conventional process [45–47]. In view of the
Table 1 previous studies and our experimental data, we also adopt this
The expression of components based on the form of -FA and G-.
common model to describe lipid release in DSEC. That is, if the
Component Structure Expression mass transfer of lipid harvest during DSEC follows the first-order
differential equation.

d
[Lipid] = kc ([Lipid]max − [Lipid] ) (6)
dt
where kc (min−1 ) is the overall mass transfer coefficient and
[Lipid]max (mM) is the maximum amount of lipid could be re-
leased.
Triglyceride (TG) G(FA)3

3.3. Kinetic model in DSEC

Fatty acid methyl ester (FAME)
Free fatty acid (FFA)
Fatty acid sodium salt (soap)
Glycerol
solution, as shown in Figs. 1 and 2 that are (I) saponification and
(II) esterification steps, respectively. Thus, subscripts Ⅰ and Ⅱ are
used to represent the individual step in DSEC in the following sec-
tions.
3.2. Extraction model
Different from the lipid extraction/recovery followed by the
biodiesel conversion in the conventional process, DSEC combines
lipid extraction and direct conversion into biodiesel without dry-
ing/purification. Lipid could be released from the microalgae cell
when NaOH/methnaol acts as a cell disruption agent. Further, lipid
is expressed as the total fatty acid base (-FA) which could be found
CH3 –FA
The DSEC process is fulfilled by two consecutive additions. The
first is the saponification step with NaOH catalyst and the sec-
H–FA
ond is the esterification step with HCl catalyst. For the first step
with alkali catalyst, there would be only transesterification of TG
involved [27,41,48] if very little water and free fatty acid in the so-
Na–FA lution. Otherwise, both transesterification and saponification occur
and the soap is formed [43,49–51]. Meanwhile, Eze et al. [43] have
recently shown the mechanism of the equilibrium between the hy-
droxide and methoxide (CH3 O− ) anions in the alkaline hydroly-
sis. In addition, according to Reyero et al. [50], they reported that
the rate of the hydroxide–methoxide anions reaction was relatively
high than those of transesterification and saponification in the case
G(OH)3 of NaOH− catalyzed sunflower oil.
Therefore, a kinetic model is proposed for this DSEC pro-
cess based on the following understanding. (1) The hydroxide–
methoxide equilibrium with methanol is faster than other reac-
tions when NaOH used as the catalyst. That is, methoxide would
be first formed and other reactions proceed. (2) The transesterifi-
cation reaction constituted by three consecutive reactions is sim-
plified into an overall reaction by neglecting intermediates of di-
glyceride and mono-glyceride. Similarly, the saponification reaction
is involved with TG is consolidated into an overall reaction. (3) In
the first step, saponification reactions are taken as irreversible re-
actions. (4) In the esterification (or the 2nd) step, some HCl is neu-
tralized with NaOH from the 1st step and then the rest converts
soap into free fatty acid.
In the first stage, according to the model assumptions and the
proposed mechanisms in Fig. 1, the scheme in the saponification

Please cite this article as: Y.-R. Fang et al., A novel strategy of biodiesel production from wet microalgae by direct saponification–
esterification conversion (DSEC), Journal of the Taiwan Institute of Chemical Engineers (2017), https://doi.org/10.1016/j.jtice.2017.12.001
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Fig. 4. The routes in the saponification step.

step of DSEC from microalgae lipid catalyzed by NaOH can be in-

dicated in Fig. 4 where k1 denotes the rate constant of transes-
terification of G(FA)3 , and k2 , k3 and k4 are the rate constants of
saponification of CH3 –FA, G(FA)3 and H–FA, respectively.
In addition, the hydroxide–methoxide anions equilibrium is
shown in Eq. (7) and the equilibrium constant (KM ) is expressed
in Eq. (8).
KM
CH3 OH + OH− ←→ CH3 O− + H2 O (7)
Fig. 5. The routes in terms of ion forms in the saponification step.

[CH3 O− ]x [H2 O]x

KM = (8)
[CH3 OH]x [OH− ]x
Because of mass balance, initial NaOH ([NaOH]Ⅰ , i ) is described
as Eq. (9) to consider the consumption OH− to form CH3 O− indi-
d
[H − FA] = kc ([H − FA]max − [H − FA] ) − k4 [H − FA][OH− ]I
cated by Eq. (7) [51]. dt
    (16)
[NaOH]I,i = CH3 O− I
+ OH−
I
(9)

Then Eq. (10) could be obtained substituting [CH3 O− ]Ⅰ from Eq.

(9) into Eq. (8). d d
[Na − FA]I or [FA− ]I
− dt dt
[NaOH]I,i − [OH ]I [CH3 OH]I
= KM (10) = k2 [CH3 − FA]I [OH− ]I + k3 ·3·[G(FA )3 ][OH− ]I + k4 [H − FA][OH− ]
[OH− ]I [H2 O]I
(17)
Because the concentrations of methanol (8–16 M) and water
(18–37 M) are both in excess amount in this study, the right side
of Eq. (10) can be approximately taken as a constant, a (define in d
[CH3 − FA]I = k1 [G(FA )3 ][CH3 O− ]I − k2 [CH3 − FA]I [OH− ]I (18)
Eq. (11) with their initial values). Thus, Eq. (10) may be rewritten dt
as Eq. (12) and the concentrations of hydroxide and methoxide an-
ions could be obtained as Eqs. (13) and (14), respectively. If we express lipid for both TG and H–FA in term of equiva-
lent -FA, and assume fraction (m) of lipid is TG and H–FA accounts
[CH3 OH]I, i for the rest, i.e. fraction of (1-m). Thus, these equations can be ob-
a= (11)
[H2 O]I,i tained as the following:

[NaOH]I, i − [OH− ]I [CH3 OH]I,i 3 · [G(FA )3 ] = m · [Lipid] (19)

∼ KM = KM a (12)
[OH− ]I [H2 O]I,i

  [NaOH]I,i [H − FA] = (1 − m ) · [Lipid] (20)

OH− = (13)
I 1 + KM a

  [NaOH]I,i KM a 3 · [G(FA )3 ]max = m · [Lipid]max (21)

CH3 O− = (14)
I 1 + KM a
In view of the discussion above, methoxide anions formation in
the hydroxide–methoxide equilibrium is a fast reaction. Reactions [H − FA]max = (1 − m ) · [Lipid]max (22)
in Fig. 4 can be further expressed as Fig. 5, all proceeding after
the methoxide anions formation. Therefore, rate equations of the With Eqs. (13) and (14) and the aid of Eqs. (19)–(22) for G(FA)3
saponification step could be written as Eqs. (15)–(18) together with and H–FA in Eqs. (15)–(18), the individual rate equations of reac-
mass transfer (cell disruption) of lipid based on the quantification tants and products can be obtained:
of -FA. In Eqs. (15) and (16) where kc is the mass transfer coeffi- d
cient of releasing lipid (TG and H–FA) into reaction solution. m · [Lipid] = kc (m · [Lipid]max − m · [Lipid] )
dt
d [NaOH]I.i KM a
3·[G(FA )3 ] = kc (3·[G(FA )3 ]max − 3 · [G(FA )3 ] ) −k1 · m · [Lipid]
dt 1 + KM a
−k1 ·3·[G(FA )3 ][CH3 O− ]I − k3 ·3·[G(FA )3 ][OH− ]I [NaOH]I,i
−k3 · m · [Lipid] (23)
(15) 1+KM a

Please cite this article as: Y.-R. Fang et al., A novel strategy of biodiesel production from wet microalgae by direct saponification–
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the rate equations in the esterification step may be expressed as

follows:
d
[Na − FA] = −k5 [Na − FA]II [HCl]II,i (36)
dt
Fig. 6. The route in the esterification step.

d
[H − FA]II = k5 [Na − FA]II [HCl]II,i − k6 [H − FA][CH3 OH]II,i
d dt
(1 − m ) · [Lipid] = kc ( (1 − m ) · [Lipid]max − (1 − m ) · [Lipid]) +k−6 [CH3 − FA][H2 O]II,i (37)
dt
[NaOH]I,i
−k4 (1 − m ) · [Lipid] (24)
1+KM a d
[CH3 − FA] = k6 [H − FA][CH3 OH]II,i − k−6 [CH3 − FA][H2 O]II,i
dt
d [NaOH]I,i [NaOH]I,i (38)
[Na − FA]I = k2 [CH3 − FA]I + k3 · m · [Lipid]
dt 1+KM a 1+KM a
The dilution effect due to the addition in the second step
[NaOH]I,i
+k4 (1 − m ) · [Lipid] (25) should be noted (Eq. (38)) and the corresponding initial conditions
1+KM a are given in Eqs. (40) and (41) for the esterification reaction.
VI
d [NaOH]I,i KM a [Na − FA]II (t = 0 min ) = [Na − FA]I (t = t f min ) × (39)
[CH3 − FA]I = k1 · m · [Lipid] VII
dt 1 + KM a
[NaOH]I,i [H − FA]II (t = 0 min ) = 0 (40)
−k2 [CH3 − FA]I (26)
1+KM a
VI
Finally, the kinetic model in the saponification step of DSEC can [CH3 − FA]II (t = 0 min ) = [Na − FA]I (t = t f min ) × (41)
be proposed when combining Eq. (23) combines with Eq. (24): VII

d 3.4. Determination of kinetic constants

[Lipid] = kc ([Lipid]max − [Lipid] ) − k1 [Lipid][NaOH]I,i
dt
−k3 [Lipid][NaOH]I,i (27) The kinetic models of the DSEC process were described by
three differential equations Eqs. (27)–(29) for the saponification
step and three differential equations Eqs. (36)–(38) for the esterifi-
d
[Na − FA]I = k2 [CH3 − FA]I [NaOH]I,i + k3 [Lipid][NaOH]I,i (28) cation step. For simulation, estimate of mass transfer coefficient kc
dt and [Lipid]max in Eq. (6) should be sought first. Then, two sets of
differential equations (Eqs. (27)–(29) and (36)–(38)) together with
d
[CH3 − FA]I = k1 [Lipid][NaOH]I,i − k2 [CH3 −FA]I [NaOH]I,i (29) proper initial conditions (Eqs. (33)–(35) and (39)–(41)) can be nu-
dt merically integrated with MATLABۚ (ODE45) and fitted against ex-
where the following rate constants relative to those in the reac- perimental data by method of least-squares.
tions of Fig. 5 are defined as following:
4. Results and discussion
k1 m KM a
k1 = (30)
1 + KM a 4.1. Extraction model (Kc )

k2
k2 = (31) During the saponification step (1.33 M NaOH at 100 °C), exper-
1 + KM a imental results of lipid release for various microalgae concentra-
tions ([C ]Ⅰ = 8.67, 17.33 and 24.67 g/L) are shown in Fig. 7. Accord-
k3 m + ( 1 − m )k4
k3 = (32) ing to Eq. (6), [Lipid]max were obtained as the saturation/constants
1 + KM a values in the figure and kc could be estimated as a constant value
That the initial concentrations of above are: (kc = 0.8 min−1 ) for various cell concentrations satisfactorily. The
results in Fig. 7 also indicated that release of lipid could be com-
[Lipid](t = 0 min ) = 0 mM (33) pleted in less than 10 mins during the saponification step and
reach its maximum lipid concentration, [Lipid]max. Please note kc
is an overall mass transfer coefficient which include cell disruption
[Na − FA]I (t = 0 min ) = 0 mM (34)
and releasing lipid into the solution. Therefore, one might expect
that NaOH/methanol concentration, temperature and so on would
[CH − FA]I (t = 0 min ) = 0 mM (35) have effect on this constant.

With the second addition of HCl/methanol after the saponifica-
tion step, HCl would first neutralize NaOH which is a fast reaction
and, consequently, convert soap into free fatty acids (H–FA). As a
result, based on the proposed mechanisms in Fig. 2, the reactions
in the esterification step is shown in Fig. 6, where k5 is the rate
constant for H–FA formation, and k6 and k-6 are the rate constants
for the reversible reaction of esterification. It is noted that the
initial concentrations of methanol (18.58 M), water (11.47 M) and
HCl (0.45 M) are much higher than the initial concentration of free
fatty acid (0.0 03–0.0 09 M) in the 2nd step. Under these conditions,
4.2. The kinetic model in DSEC
The kinetic parameters were estimated by fitting the experi-
mental data and the obtained results are listed in Table 2. The
simulation with the corresponding kinetic parameters and the ex-
perimental results are depicted in Fig. 8 for the saponification step
and Fig. 9 for the esterification step. These results were satisfac-
tory, especially in view of very complicated composition in reac-
tion broth including various cell constituents as well as different
carbon length of fatty acids.

Please cite this article as: Y.-R. Fang et al., A novel strategy of biodiesel production from wet microalgae by direct saponification–
esterification conversion (DSEC), Journal of the Taiwan Institute of Chemical Engineers (2017), https://doi.org/10.1016/j.jtice.2017.12.001
JID: JTICE
ARTICLE IN PRESS [m5G;December 15, 2017;20:5]

Y.-R. Fang et al. / Journal of the Taiwan Institute of Chemical Engineers 000 (2017) 1–9 7

30
[C]I =8.67 g/L
25 [C]I =17.33 g/L
[C]I =24.67 g/L
20
R2 = 0.86
[Lipid] (mM)

15

10 R2 = 0.85

5 R2 = 0.86

0
0 5 10 15 20 25 30
time (min)
Fig. 7. The experimental data and prediction of extraction model for the lipid pro-
files for different wet microalgae concentrations, and NaOH = 1.33 M at 100 °C (◦:
[C ]Ⅰ = 8.67 g/L, : [C ]Ⅰ = 17.33 g/L, : [C ]Ⅰ = 24.67 g/L).

Table 2
Parameters of the kinetic model developed for
biodiesel production from microalgae by DSEC.

Parameters Value Unit

kc 0.8 min−1
k 1 1.5 × 10−2 mM–1 min–1
k 2 6 × 10−3 mM–1 min–1
k 3 1 × 10−4 mM–1 min–1
k5 10.1 min−1
k6 9.67 × 10−5 mM–1 min–1
k-6 6.9 × 10−6 mM–1 min–1

As shown in Fig. 8 for the saponification step, lipid re-

leased from microalgae cell by NaOH/methanol that was a non-
mechanical cell disruption and then converted into CH3 –FA or Na–
FA quickly through the routes shown in Fig. 5. The first and the
second route in Fig. 5 involving transesterification and saponifica-
tion for G(FA)3 , the kinetic constants of k1  and k3  (Table 2) indi-
cated that transesterification rate was faster than that of saponi-
fication. That is, kinetic constants indicated that TG of lipid was
mainly consumed into CH3 –FA in transesterification since k1  > k3  .
However, k1  , k2  and k3  would depend on initial methanol/water
ratio (Eq. (11)) due to the hydroxide–methoxide equilibrium. On
the other hand in the esterification step, the kinetic constants in-
dicated that k5 > k6 > k-6 in the acid solution. The result provided
a good agreement that neutralization of Na–FA followed by esteri-
fication of H–FA as the fourth route in Fig. 6.
For the esterification step where HCl was used as the acid cata-
lyst, comparison between simulation (kinetic constants in Table 2)
and experimental data were depicted in Fig. 9. The simulation re-
sults showed a good agreement with proposed kinetic model by
assuming esterification a reversible reaction. Na–FA firstly converts
into H–FA by neutralization and then H–FA undergoes esterifica-
tion to produce CH3 –FA eventually. As the kinetic curves shown in
Fig. 9, it indicated that lipid containing TG almost converted into Fig. 8. Experimental data and simulation of the kinetic model in the saponification
Na–FA and therefore there is no TG remained in the following es- step for various cell concentrations, and NaOH concentration 1.33 M at 100 °C (:
Lipid, : CH3 -FA, ◦: Na-FA). microalgae concentration (a) [C ]Ⅰ = 8.67 g/L (b)17.33 g/L
terification step. Therefore, the reaction rate in the esterification
(c) 24.67 g/L.
step of DSEC would be enhanced by avoiding transesterification

Please cite this article as: Y.-R. Fang et al., A novel strategy of biodiesel production from wet microalgae by direct saponification–
esterification conversion (DSEC), Journal of the Taiwan Institute of Chemical Engineers (2017), https://doi.org/10.1016/j.jtice.2017.12.001
JID: JTICE
ARTICLE IN PRESS
8 Y.-R. Fang et al. / Journal of the Taiwan Institute of Chemical Engineers 000 (2017) 1–9
Fig. 9. Experimental data and simulation of the kinetic model in the esterification
step with different microalgae concentrations, and HCl = 0.45 M (after neutraliza-
tion with NaOH) at 100 °C (: Na–FA, : H–FA, ◦: CH3 –FA). Different microalgae
concentration (a) [C ]Ⅱ = 5.14 g/L (b) 10.28 g/L (c) 15.43 g/L.
Please cite this article as: Y.-R. Fang et al., A novel strategy of biodiesel production from wet microalgae by direct saponification–
esterification conversion (DSEC), Journal of the Taiwan Institute of Chemical Engineers (2017), https://doi.org/10.1016/j.jtice.2017.12.001
[m5G;December 15, 2017;20:5]
of TG compared to a typical acid-catalyzed transesterification. In
other words, the reason that biodiesel could be produced without
the limitation of transesterification was attributed nearly no TG in
the final step. However, H–FA perhaps did not completely convert
into CH3 –FA in esterification step which indicated by a “reversible”
esterification. Hence, we can calculate the conversion of H–FA into
CH3 –FA in esterification by Eq. (42).
[H − FA]II,i − [H − FA]II, f
XH−FA to CH3−FA = (42)
[H − FA]II,i
where [H–FA]Ⅱ , i is the initial concentration of H–FA obtained
in neutralization before esterification, [H–FA]Ⅱ , f is the final con-
centration of H–FA after undergoing esterification when reaction
reaches equilibrium. It is estimated that conversion in Fig. 9 for
various microalgae concentrations by Eq. (42) was around 0.96.
With average molecular weight of biodiesel of 284 (measured by
GC), the oil content of microalgae was estimated as 0.164 g/g-cell
while biodiesel yield as 0.166 g/g-cell.
Although conventional transesterification by the acid catalyst
can overcome the restriction of water and free fatty acid content
in feedstock and both TG and FFA can convert into biodiesel si-
multaneously, slow reaction and low biodiesel yield are the draw-
backs [28,29]. According to Suwannakarn et al. [29], a comparison
of the reaction rate between transesterification and esterification
with methanol at 130 °C for Tricaprylin (TG, C8:0) via transesteri-
fication and the mixture of 0–25 wt% Lauric acid (FFA, C12:0) and
Tricaprylin via esterification, it was found a higher reaction rate
and a higher biodiesel conversion when Lauric acid in the mix-
ture changed from 0 to 25 wt%. Also, the kinetic model by Hi-
dalgo et al. [54] supported that the rate constant of esterification
(k = 0.034 min−1 ) was apparently higher than the rate constant of
transesterification (k = 0.0030 min−1 ). Our DSEC work pointed out
that biodiesel was primarily produced via esterification and thus
higher conversion and reaction rate would be obtained. The equi-
librium constant of esterification was estimated as 14 at 100 °C as
estimated from Table 2 (k6 /k-6 ). The values obtained in this work
are of the similar order to some studies of the kinetics of esterifica-
tion [28,52–54]. For example, Pisarello et al. [28] reported that the
rate constant of forward, backward reaction and the equilibrium
constant in esterification at 60 °C is 0.031 min−1 , 0.0011 min−1 ,
and 28, respectively for the pseudo first-order kinetic model.
As pointed out in introduction, some direct alkali-acid con-
version similar to DSEC in this work were conducted by a cou-
ple of studies [35,36] and a better biodiesel yield were revealed
previously. However, without investigating the detail mechanism,
the process was improperly categorized as direct transesterification
(DT). In fact, our study indicated that this DSEC mainly undergo
saponification followed by esterification into biodiesel. And be-
cause of the confirmation of the reaction mechanism, it explained
that the existence of free fatty acid or water should not be a con-
cern as the conventional biodiesel production processes.
5. Conclusion
Biodiesel production from wet microalgae with water content
higher than 90 wt% by DSEC was studied in this work. Instead of
the conventional feedstock drying and lipid extraction, DSEC is a
simple process that significant amount effort and cost could be
avoided by direct converting wet microalgae to biodiesel. Also, the
proposed kinetic model with the experimental results indicated
that the mechanism is mainly via saponification/esterification con-
version, which primarily converted lipid released from microalgae
cell into soap in the saponification step followed by the esterifica-
tion into biodiesel in the next step. In the study, high molar ra-
tio of methanol to lipid in the saponification step and the high
molar ratio of methanol to free fatty acid were adopted to verify
JID: JTICE
ARTICLE IN PRESS
Y.-R. Fang et al. / Journal of the Taiwan Institute of Chemical Engineers 000 (2017) 1–9
the mechanism as well as to simplify the kinetic model develop-
ment. Future optimization of these ratios and recovery are needed
in large-scale production by DSEC. However, this work showed that
DSEC could completely eliminate water inhibition and maintain
a stable biodiesel yield of 0.18 (g/g-dried-cell) when water con-
tent was higher than 90 wt%, which an alkali-catalyzed transes-
terification would not be possible. While compared with an acid-
catalyzed transesterification, DSEC would take less than 15 min
(shown as Figs. 8 and 9) to accomplish overall reaction and over-
came the problem in lower productivity of biodiesel. Furthermore,
the proposed kinetic model indeed verified DSEC mechanism and
simulate well the corresponding concentration profiles in each step
of DSEC. Based on these, it is suggested that DSEC would be a fea-
sible process for biodiesel production directly from wet microalgae.
Acknowledgment
Financial support from Ministry of Science and Technology, Tai-
wan is greatly appreciated.
References
[1] Singh SP, Singh D. Biodiesel production through the use of different sources
and characterization of oils and their esters as the substitute of diesel: a re-
view. Renew Sustain Energy Rev 2010;14(1):200–16.
[2] Balat M. Potential alternatives to edible oils for biodiesel production – a review
of current work. Energy Convers Manag 2011;52(2):1479–92.
[3] Talebian-Kiakalaieh A, Amin NAS, Mazaheri H. A review on novel processes of
biodiesel production from waste cooking oil. Appl Energy 2013;104:683–710.
[4] Cherubini F. The biorefinery concept: using biomass instead of oil for produc-
ing energy and chemicals. Energy Conver Manag 2010;51(7):1412–21.
[5] Jain S, Sharma MP. Prospects of biodiesel from Jatropha in India: a review.
Renew Sustain Energy Rev 2010;14(2):763–71.
[6] Huang G, Chen F, Wei D, Zhang X, Chen G. Biodiesel production by microalgal
biotechnology. Appl Energy 2010;87(1):38–46.
[7] Singh J, Gu S. Commercialization potential of microalgae for biofuels produc-
tion. Renew Sustain Energy Rev 2010;14(9):2596–610.
[8] Ratha SK, Prasanna R, Prasad RBN, Sarika C, Dhar DW, Saxena AK. Modulating
lipid accumulation and composition in microalgae by biphasic nitrogen sup-
plementation. Aquaculture 2013;392–395:69–76.
[9] Campbell PK, Beer T, Batten D. Life cycle assessment of biodiesel production
from microalgae in ponds. Bioresour Technol 2011;102(1):50–6.
[10] Yen HW, Ho SH, Chen CY, Chang JS. CO2 , NOx and SOx removal from flue gas
via microalgae cultivation: a critical review. Biotechnol J 2015;10(6):829–39.
[11] Harrlngton K, D’Arcy-Evans C. Transesterification in situ of sunflower seed oil.
Ind Eng Chem Prod Res Dev 1985;24:314–18.
[12] Lepage G, Roy C. Direct transesterification of all classes of lipids in a one-step
reaction. J Lipid Res 1986;27:114–20.
[13] Balat M, Balat H. Progress in biodiesel processing. Appl Energy
2010;87(6):1815–35.
[14] Harrlngton K, D’Arcy-Evans C. Comparison of conventional and in situ methods
of transesterification of seed oil from a series of sunflower cultivars. J Am Oil
Chem Soc 1985;62:1009–13.
[15] Hincapié G, Mondragón F, López D. Conventional and in situ transesterification
of castor seed oil for biodiesel production. Fuel 2011;90(4):1618–23.
[16] Haas MJ, Wagner K. Simplifying biodiesel production: the direct or
in situ transesterification of algal biomass. Eur J Lipid Sci Technol
2011;113(10):1219–29.
[17] Srivastava A, Prasad R. Triglycerides-based diesel fuels. Renew Sustain Energy
Rev 20 0 0;4:111–33.
[18] Marchetti JM, Miguel VU, Errazu AF. Possible methods for biodiesel production.
Renew Sustain Energy Rev 20 07;11(6):130 0–11.
[19] Ma F, Hanna M. Biodiesel production: a review. Bioresour Technol
1999;70:1–15.
[20] Wright H, Segur J, Clark H, Coburn S. A report on ester interchange. Oil Soap
1944;21:145–8.
[21] Liu K. Preparation of fatty acid methyl esters for gas-chromatographic analysis
of lipids in biological materials. J Am Oil Chem Soc 1994;71:1179–87.
[22] Zhang Y. Biodiesel production from waste cooking oil: 1. Process design and
technological assessment. Bioresour Technol 2003;89(1):1–16.
[23] Vicente G, Martinez M, Aracil J. Integrated biodiesel production: a com-
parison of different homogeneous catalysts systems. Bioresour Technol
2004;92(3):297–305.
[24] Naik M, Meher L, S N, L D. Production of biodiesel from high free fatty acid
Karanja (Pongamia pinnata) oil. Biomass Bioenergy 2008;32(4):354–7.
Please cite this article as: Y.-R. Fang et al., A novel strategy of biodiesel production from wet microalgae by direct saponification–
esterification conversion (DSEC), Journal of the Taiwan Institute of Chemical Engineers (2017), https://doi.org/10.1016/j.jtice.2017.12.001
[m5G;December 15, 2017;20:5]
9
[25] Freedman B, Pryde E, Mounts T. Variables affecting the yields of fatty esters
from transesterified vegetable oils. J Amn Oil Chem Soc 1984;61:1638–43.
[26] Hidalgo P, Ciudad G, Schober S, Mittelbach M, Navia R. Improving the FAME
yield of in situ transesterification from microalgal biomass through particle
size reduction and cosolvent incorporation. Energy Fuels 2015:823–32.
[27] Freedman B, Butterfield R, Pryde E. Transesterification Kinetics of Soybean Oil.
J Am Oil Chem Soc 1986;63:1375–80.
[28] Pisarello M, Dalla C, Mendow G, Querini C. Esterification with ethanol to
produce biodiesel from high acidity raw materials. Fuel Process Technol
2010;91(9):1005–14.
[29] Suwannakarn K, Lotero E, Ngaosuwan K, Goodwin J. Simultaneous free fatty
acid esterification and triglyceride transesterification using a solid acid cata-
lyst with in situ removal of water and unreacted methanol. Ind Eng Chem Res
2009;48:2810–18.
[30] Shiu P, Gunawan S, Hsieh W, Kasim N, Ju Y. Biodiesel production from rice
bran by a two-step in-situ process. Bioresour Technol 2010;101(3):984–9.
[31] Hayyan A, Alam M, Mirghani M, Kabbashi N, Hakimi N, Siran Y, et al. Sludge
palm oil as a renewable raw material for biodiesel production by two-step pro-
cesses. Bioresour Technol 2010;101(20):7804–11.
[32] Hayyan A, Mjalli F, Hashim M, Hayyan M, AlNashef I, Al-Zahrani S,
et al. Ethanesulfonic acid-based esterification of industrial acidic crude palm
oil for biodiesel production. Bioresour Technol 2011;102(20):9564–70.
[33] Chen L, Liu T, Zhang W, Chen X, Wang J. Biodiesel production from algae oil
high in free fatty acids by two-step catalytic conversion. Bioresour Technol
2012;111:208–14.
[34] Dong T, Wang J, Miao C, Zheng Y, Chen S. Two-step in situ biodiesel pro-
duction from microalgae with high free fatty acid content. Bioresour Technol
2013;136:8–15.
[35] Griffiths M, Hille R, Harrison S. Selection of direct transesterification as
the preferred method for assay of fatty acid content of microalgae. Lipids
2010;45(11):1053–60.
[36] Kumar V, Muthuraj M, Palabhanvi B, Ghoshal A, Das D. Evaluation and
optimization of two stage sequential in situ transesterification process
for fatty acid methyl ester quantification from microalgae. Renew Energy
2014;68:560–9.
[37] Stein J. Handbook of Phycological Methods. Culture methods and growth mea-
surements 1973:448.
[38] Ehimen E, Sun Z, Carrington C. Variables affecting the in situ transesterification
of microalgae lipids. Fuel 2010;89(3):677–84.
[39] Park J, Park M, Lee Y, Yang J. Advances in direct transesterification of algal oils
from wet biomass. Bioresour Technol 2015;184:267–75.
[40] Meher L, Vidyasagar D, Naik S. Technical aspects of biodiesel production by
transesterification—a review. Renew Sustain Energy Rev 2006;10(3):248–68.
[41] Vicente G, Martínez M, Aracil J, Esteban A. Kinetics of sunflower oil methanol-
ysis. Ind Eng Chem Res 2005;44:5447–54.
[42] Bambase M, Nakamura N, Tanaka J, Matsumura M. Kinetics of hydroxide-cat-
alyzed methanolysis of crude sunflower oil for the production of fuel-grade
methyl esters. J Chem Technol Biotechnol 2007;82(3):273–80.
[43] Eze V, Harvey A, Phan A. Determination of the kinetics of biodiesel saponifica-
tion in alcoholic hydroxide solutions. Fuel 2015;140:724–30.
[44] Formo M. Ester reactions of fatty materials. J Am Oil Chem Soc
1954;31:548–59.
[45] Vauchel P, Leroux K, Kaas R, Arhaliass A, Baron R, Legrand J. Kinetics model-
ing of alginate alkaline extraction from Laminaria digitata. Bioresour Technol
20 09;10 0(3):1291–6.
[46] Halim R, Gladman B, Danquah M, Webley P. Oil extraction from microalgae for
biodiesel production. Bioresour Technol 2011;102(1):178–85.
[47] Halim R, Rupasinghe T, Tull D, Webley P. Modelling the kinetics of lipid ex-
traction from wet microalgal concentrate: a novel perspective on a classical
process. Chem Eng J 2014;242:234–53.
[48] Bambase M, Nakamura N, Tanaka J, Matsumura M. Kinetics of hydroxide-cat-
alyzed methanolysis of crude sunflower oil for the production of fuel-grade
methyl esters. J Chem Technol Biotechnol 2007;82(3):273–80.
[49] Komers K, Skopal F, Stloukal R, Machek J. Kinetics and mechanism of the KOH
— catalyzed methanolysis of rapeseed oil for biodiesel production. Eur J Lipid
Sci Technol 2002;104(11):728–37.
[50] Reyero I, Arzamendi G, Zabala S, Gandía L. Kinetics of the NaOH-catalyzed
transesterification of sunflower oil with ethanol to produce biodiesel. Fuel Pro-
cess Technol 2015;129:147–55.
[51] Eze V, Harvey A, Phan A. Determination of the kinetics of biodiesel saponifica-
tion in alcoholic hydroxide solutions. Fuel 2015;140:724–30.
[52] Aranda D, Santos R, Tapanes N, Ramos A, Antunes O. Acid-catalyzed homo-
geneous esterification reaction for biodiesel production from palm fatty acids.
Catal Lett 2008;122(1–2):20–5.
[53] Farag H, Azza E, Taha N. Kinetic study of used vegetable oil for esterification
and transesterification process of biodiesel production. Int J Chem Biochem Sci
2013;3:1–8.
[54] Hidalgo P, Ciudad G, Mittelbach M, Navia R. Biodiesel production by direct
conversion of Botryococcus braunii lipids: reaction kinetics modeling and op-
timization. Fuel 2015;153:544–51.