Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605

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Chemical Engineering Research and Design

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

Experimental and modeling of reactive separation

of protocatechuic acid

Biswajit S. De a, Kailas L. Wasewar a,∗, Vicky R. Dhongde a, Anjali A. Ingle a,

Himangshu Mondal b
a Advanced Separation and Analytical Laboratory (ASAL), Department of Chemical Engineering, Visvesvaraya
National Institute of Technology (VNIT), Nagpur, Maharashtra 440010, India
b Department of Chemical Engineering, Institute of Technology, Guru Ghasidas Vishwavidyalaya, Bilaspur,

Chhattisgarh 495009, India

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

Article history: Protocatechuic acid (PCA) possess antioxidant, anticancer, and antidiabetic activity. It is a
Received 15 September 2017 novel active substance against infectious disease bursal virus infection and avian influenza
Received in revised form 22 January virus H9N2 infection. The separation of PCA was experimentally investigated using tri-n-
2018 butyl phosphate (TBP) in natural non-toxic diluents (sunflower oil, soybean oil, sesame oil).
Accepted 29 January 2018 Dimerisation constant, partition coefficient, distribution coefficient, extraction efficiency,
Available online 8 February 2018 and loading ratio were determined. A single cycle achieved a maximum 90% separation.
The trend observed for separation in diluent was sunflower oil > soybean oil > sesame oil.
Keywords: Mass action law, relative basicity and Langmuir model were used to represent the reactive
Protocatechuic acid extraction equilibrium of protocatechuic acid using TBP. The toxic effects can be averted by
Reactive extraction evasion of direct contact of organisms with organic phase and by the utilization of natural
Equilibrium modelling non-toxic diluents. The diffusion coefficients of protocatechuic acid were estimated using
Wilke–Chang equation Wilke–Chang equation and Reddy–Doraiswamy equation. The number of theoretical stages
Reddy–Doraiswamy equation (NTS) for counter current extraction was found to be 5 using modified Kremser equation.
Column design The results can be employed to design reactive extraction column.
© 2018 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

1. Introduction
Protocatechuic acid is a phenolic compound chemically known as 3,4-
dihydroxybenzoic acid (3,4-DHBA). Phenolic compounds are considered
secondary metabolites and derived from phenylalanine via shikimic
acid pathway (Kakkar and Bais, 2014). Protocatechuic acid is detected
in many fruits such as gooseberries (Ribes uvacrispa L.) (Li et al., 1993),
grapes (Vitis vinifera) (Li et al., 1993), and plums (Prunus domestica L.)
(Miura et al., 2002). A variety of research has been carried out on pro-
tocatechuic acid and its derivatives and conforms (aldehydes, esters,
etc.) and has been reported for its potential action such as antiox-
idant activity (Li et al., 2011), antibacterial activity (Chao and Yin,
2009), antidiabetic activity (Scazzocchio et al., 2011), anticancer activ-
ity (Tanaka et al., 2011), antiulcer activity (Kore et al., 2011), antiageing
activity (Shi et al., 2006), antifibrotic activity (Li et al., 2012), and antiviral
activity (Zhou et al., 2007). Protocatechuic acid also exhibited antihy-
pertensive and cardioprotective effects (Pacheco-Palencia et al., 2008).
It has also proven its utility in preventing damage by agents causing
oxidative stress-mediated nephrotoxicity (Lee et al., 2009). Protocate-
chuic acid was also found to be a potent antiseptic, anti-inflammatory,
and analgesic (Lende et al., 2011). It possesses antiatherogenic effect by
virtue of its anti-inflammatory activity. Some recent studies identified
protocatechuic acid as a novel active substance against infectious dis-
ease bursal virus (IDBV) infection (Ou et al., 2012) and avian influenza
virus H9N2 infection (Ou et al., 2014). Apart from its pharmacological
properties, protocatechuic acid is a new building block for synthesizing
polymers and bioplastics which are produced from sugar by recom-
binant microorganisms (Okai et al., 2016). The composite polymer of
protocatechuic acid and aniline functions as an electrode with an excel-
lent electrochemical activity (Sun et al., 1998).

∗
Corresponding author.
E-mail address: klwasewar@che.vnit.ac.in (K.L. Wasewar).
https://doi.org/10.1016/j.cherd.2018.01.054
0263-8762/© 2018 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.
594 Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605

The direct extraction of protocatechuic acid from plant secondary

Iodine value

MW: molecular weight, BP = boilng point, MP = melting point, SWater = solubility in water,  = density, RI = refractive index,  = viscosity, ε = dielectric constant, log Kow = octanol–water partition coefficient, TBP = tri-
metabolites is difficult. Thus, biosynthesis of protocatechuic acid

125–140

128–143

103–116
(Chien et al., 2011; Mycroft et al., 2015; Okai et al., 2016; Sun et al.,
1998; Williams et al., 2012; Wilson et al., 2006) is more economical and
gaining importance to suffice its demand owing to its varied applica-

–
tions. Numerous suggested techniques for its recovery areadsorption
(Sarma and Mahiuddin, 2014), microbial degradation (Buchan et al.,

Saponification value
2000), ultrafiltration (Galanakis et al., 2010), Fenton (Rivas et al., 2005),
and O3 /UV or H2 O2 /UV (Benitez et al., 1996). However, these methods
sufferfrom few demerits such as less selectivity, ineffective for dilute
solutions, energy intensive, time-consuming, and generate toxic by-

188–194

189–195

188–195
products (Khin et al., 2012).
Reactive extraction is a promising technique for the recovery of car-

–
boxylic acids from dilute solutions or fermentation broth (Kertes and
King, 1986; Wardell and King, 1978; Wennersten, 1983). The selection

log Kow
of appropriate solvents is the key to achieve higher separation yield

2.5–4
0.86
(Kertes and King, 1986).

–
Reactive extraction has been effectively employed for the recov-
ery/separation of various carboxylic acids from aqueous phase such as:

10.30

3.157

3.102
tartaric acid (Sharma et al., 2017), gallic acid (Rewatkar et al., 2016, 2017;

ε
Athankar et al., 2016), nicotinic acid (Waghmare et al., 2013), pheny-

3
–
lacetic acid (Athankar et al., 2017, 2013, 2015; Gaidhani et al., 2002),
caproic acid (Wasewar and Shende, 2010, 2011a,b), lactic acid (Wasewar

 (cP)

3.80

48.8

40.5

52.5
et al., 2002a,b, 2003, 2004a, b), itaconic acid (Wasewar et al., 2011), pro-

–
pionic acid (Keshav et al., 2008a,b, 2009; Keshav and Wasewar 2010),
acrylic acid (Keshav and Wasewar, 2009), levulinic acid (Uslu et al., 2009)

1.424

1.475

1.473

1.474
and benzoic acid (Datta et al., 2011).
RI

Researchers have performed several studies on the use of –

environmental-friendly organic phases such as natural diluents (Uslu
et al., 2014), ionic liquids (Blahušiak et al., 2013; Marták and Schlosser,
 (kg m−3 )

2007; Oliveira et al., 2012; Tonova et al., 2014) and novel extractants
1540

(Ijmker et al., 2014; Krzyzaniak et al., 2014; Reyhanitash et al., 2015).

975

919

917

914
The catalytic activity of micro-organisms is affected by the presence
of organic solvents which give rise to series of physical microbial and
biochemical effects. Toxicity of the extractant and organic solvent is a
SWater (g L−1 )

Immiscible

Immiscible

Immiscible
critical problem in fermentation and the degree of toxicity depends on
the combination of microbes and extractant solution. The toxic effects
12.4

can be substantially reduced by preventing direct contact of micro-

5.9

organisms with organic phase. The least toxicity can be expected from
solvents of low polarity in combination with high solvent molecular
MP (K)

weight as reported by Brink and Tramper (1985). Bar and Gainer (1987)
494

193

256

257

267
discussed the toxicity of solvents arising from the soluble portion of sol-
Table 1 – Physicochemical properties of chemicals used in present study.

vent and the presence of two phases. The use of a membrane to prevent
direct contact of solvent with a cell containing the broth has also been
BP (K)

investigated (Cho and Shuler, 1986; Frank and Sirkar, 1986; Matsumura
562

498

530

483

and Märkl, 1986). Tik et al. (2001) obtained a maximum total lactic acid
–

concentration (2.5 times that without extraction) when 15% Alamine

336 in oleyl alcohol together with immobilized cells with 15% sunflower
Molecular structure

oil was used. The sunflower oil can also extract Alamine 336 that dif-
fused into the gels and prevent the toxic effect of the solvent. Reactive
extraction of lactic acid with a combination of carriers like TOA and Ali-
quat 336 dissolved in either sunflower oil or its mixtures with TBP was
studied by Harington and Hossain (Harington and Hossain, 2008). Kar
et al. (2017) also reported that natural non-toxic diluents such as soy-
bean oil, sunflower oil, and rice bran oil can be successfully employed
–

in a reactive extraction of acrylic acid using TBP. In the present study,

although TBP is toxic to microbes, the diluents (sunflower oil, soybean
MW (g mol−1 )

oil, and sesame oil) are totally non-toxic in nature. Thus, these diluents
can be rationally employed in combination with TBP for the extraction
154.121

266.31

879.38

885.43

of protocatechuic acid.
848

In the present study, sunflower oil, soybean oil, and sesame oil were
used as a natural non-toxic diluent and tri-n-butyl phosphate (TBP) as
an extractant. The results were discussedregarding distribution coef-
Protocatechuic acid

n-butyl phosphate.

ficient, extraction efficiency, loading ratio, and extraction equilibrium

complexation constant. Three different models, namely, mass action
Sunflower oil

law, relative basicity, and Langmuir model were implemented to rep-

Soybean oil

Sesame oil
Chemicals

resent the reactive extraction equilibrium for protocatechuic acid –

TBP – diluent. This isthe first kind of study on reactive extraction of
TBP

protocatechuic acid from aqueous solution using natural diluents.

 Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605 595

2. Materials and experimental procedure
2.1. Materials
Protocatechuic acid (purity >98%) was obtained from
Sigma–Aldrich. Tri n-butyl phosphate (TBP, purity >99%)
was supplied by Spectrochem Pvt., Ltd., India. Soybeanoil,
sunflower oil, and sesame oil were procured from Gemini
Hyderabad, India. All chemicals were utilized without any
further treatment. The typical physicochemical properties of
chemicals used are enlisted in Table 1.
2.2. Experimental procedure
The solution of protocatechuic acid (0.001–0.01 mol L−1 ) was
prepared in deionised water. The concentration range of pro-
tocatechuic acid was considered based on cytotoxicity to
human cell lines and maximum concentration of acid in any
aqueous stream (Jung et al., 2000). The initial pH of protocat-
echuic acid in aqueous solution was low (∼3.34), which was
preferable as most unionized acid molecules were extractable
(pKA = 4.48). The initial concentration of TBP in natural dilu-
ents was varied from 0.37 to 2.94 mol L−1 . The aqueous phase
(protocatechuic acid) and organic phase were taken in a vol-
umetric ratio of 1:1 in 100 mL Erlenmeyer flask and shaken
for 6 h at 300.15 ± 1 K and atmospheric pressure in an orbital
shaking incubator (REMI S-24BL, India). Reactive extraction
of protocatechuic acid involves mass transfer with chemi-
cal reaction and kinetics of the process can be explained by
determining the reaction regimes (very slow reaction, slow
reaction, fast reaction, instantaneous reaction) as described
by Doraiswamy and Sharma (1984). The preliminary experi-
ments were conducted to ascertain the equilibrium time it
was observed that 2 h are sufficient to achieve the equilib-
rium. But for safer side and in order to avoid uncertainties, 6 h
was considered sufficient time to achieve equilibrium (Datta
et al., 2015; Gu et al., 1998). The two mixed phases were sep-
arated effectively by centrifugation (REMI R-4C) for 5 min at
3000 RPM. The initial and equilibrium concentration of pro-
tocatechuic acid were analysed by UV/VIS spectrophotometer
(Shimadzu 1800, Japan) at 260 nm. The organic phase concen-
tration of protocatechuic acid was evaluated by mass balance.
The reproducibility of results was ensured by repeating few
experiments and analysis under identical conditions and was
found reliable within ±2%.
3. Results and discussion
Removal of protocatechuic acid (0.001–0.01 mol L−1 ) in natu-
ral non-toxic diluents (sunflower oil, soybean oil, and sesame
oil) with TBP were studied for a better understanding of pro-
tocatechuic acid extraction and the results are summarised in
Tables 2–4.
The non-dissociated protocatechuic acid in aqueous phase
is transferred to the organic phase at pH less than pKa (∼4.48)
value as most of the non-dissociated acid molecules are
available for extraction. The extraction of protocatechuic acid
with was analysed by means of distribution ratio (KD ) as:

Table 2 – Chemicalequilibrium of protocatechuic acid with tri-n-butyl phosphate in sunflower oil at 300.15 ± 1 K.a
[CTBP ]org (mol L−1 ) [CPCA ]0 (mol L−1 ) [CPCA ]aq (mol L−1 ) [CPCA ]org (mol L−1 )
0
KD Avg. KD % Avg. % zPCA

0 0.001 0.0010 0.000013 0.01 0.05 1.35 4.70 –

0.003 0.0029 0.000091 0.03 3.03
0.005 0.0048 0.0002 0.05 4.80
0.007 0.0066 0.0004 0.07 6.14
0.010 0.0092 0.0008 0.09 8.16

0.37 0.001 0.0004 0.0006 1.42 2.49 58.67 70.08 0.0016

0.003 0.0009 0.0021 2.23 69.01 0.0056
0.005 0.0014 0.0036 2.48 71.24 0.0097
0.007 0.0019 0.0051 2.72 73.15 0.0139
0.01 0.0022 0.0078 3.61 78.30 0.0213

0.74 0.001 0.0003 0.0007 1.92 4.47 65.77 79.83 0.0009

0.003 0.0006 0.0024 3.90 79.60 0.0032
0.005 0.0008 0.0042 5.07 83.52 0.0057
0.007 0.0011 0.0059 5.42 84.42 0.0080
0.01 0.0014 0.0086 6.05 85.82 0.0117

1.47 0.001 0.0002 0.0008 3.10 6.69 75.64 85.59 0.0005

0.003 0.0004 0.0026 5.75 85.19 0.0017
0.005 0.0006 0.0044 7.35 88.02 0.0030
0.007 0.0008 0.0062 7.88 88.74 0.0042
0.01 0.0010 0.0090 9.36 90.35 0.0061

2.21 0.001 0.0002 0.0008 4.22 8.56 80.85 88.38 0.0004

0.003 0.0004 0.0026 6.85 87.25 0.0012
0.005 0.0005 0.0045 9.17 90.17 0.0020
0.007 0.0006 0.0064 10.44 91.26 0.0029
0.01 0.0008 0.0092 12.14 92.39 0.0042

2.94 0.001 0.0002 0.0008 5.55 9.88 84.73 90.03 0.0003

0.003 0.0003 0.0027 8.18 89.11 0.0009
0.005 0.0005 0.0045 10.07 90.97 0.0015
0.007 0.0006 0.0064 11.43 91.96 0.0022
0.01 0.0007 0.0093 14.16 93.40 0.0032

a
Standard uncertainties (u) are u(T) = ±1 K; u(HPC) = ±0.001 mol L−1 .
596 Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605

Table 3 – Chemical equilibrium of protocatechuic acid with tri-n-butyl phosphate in soybean oil at 300.15 ± 1 K.a
[CTBP ]org (mol L−1 ) [CPCA ]0 (mol L−1 ) [CPCA ]aq (mol L−1 ) [CPCA ]org (mol L−1 )
0
KD Avg. KD % Avg. % zPCA

0 0.001 0.0010 0.000027 0.03 0.07 2.74 6.56 –

0.003 0.0029 0.0001 0.04 3.96
0.005 0.0047 0.0003 0.06 5.68
0.007 0.0065 0.0005 0.08 7.37
0.010 0.0087 0.0013 0.15 13.06

0.37 0.001 0.0005 0.0005 1.19 2.21 54.27 67.39 0.0015

0.003 0.0010 0.0020 1.93 65.82 0.0054
0.005 0.0016 0.0034 2.20 68.76 0.0094
0.007 0.0019 0.0051 2.62 72.39 0.0138
0.010 0.0024 0.0076 3.12 75.73 0.0206

0.74 0.001 0.0004 0.0006 1.01 3.83 50.22 75.37 0.0008

0.003 0.0007 0.0023 3.55 78.04 0.0032
0.005 0.0009 0.0041 4.67 82.38 0.0056
0.007 0.0013 0.0057 4.53 81.92 0.0078
0.010 0.0016 0.0084 5.37 84.30 0.0115

1.47 0.001 0.0003 0.0007 2.58 5.65 72.04 83.32 0.0005

0.003 0.0005 0.0025 4.78 82.71 0.0017
0.005 0.0007 0.0043 5.87 85.45 0.0029
0.007 0.0009 0.0061 6.94 87.41 0.0042
0.010 0.0011 0.0089 8.09 89.00 0.0061

2.21 0.001 0.0002 0.0008 4.11 7.38 80.42 87.18 0.0004

0.003 0.0004 0.0026 6.18 86.08 0.0012
0.005 0.0006 0.0044 7.87 88.73 0.0020
0.007 0.0007 0.0063 8.73 89.72 0.0028
0.010 0.0009 0.0091 10.03 90.93 0.0041

2.94 0.001 0.0002 0.0008 4.68 8.79 82.40 88.78 0.0002

0.003 0.0004 0.0026 7.05 87.58 0.0009
0.005 0.0005 0.0045 8.67 89.66 0.0015
0.007 0.0006 0.0064 10.92 91.61 0.0022
0.010 0.0007 0.0093 12.61 92.65 0.0031

a
Standard uncertainties (u) are u(T) = ±1 K; u(HPC) = ±0.001 mol L−1 .

[CPCA ]org MATLAB R2013a for non-linear regression (Kertes and King,
KD = (1)
[CPCA ]aq 1986):

Also, the extraction efficiency (%) can be defined as the
ratio of protocatechuic acid concentration in extract phase to
initial protocatechuic acid concentration in aqueous solution
at equilibrium as:
[CPCA ]0aq − [CPCA ]aq
% = × 100
[CPCA ]0aq
where [CPCA ]0 aq is the initial protocatechuic acid concen-
tration in aqueous phase.
A schematic representation of the mechanism for extrac-
tion of protocatechuic acid without extractant is depicted in
Fig. 1. The distribution coefficient and extraction efficiency of
protocatechuic acid in natural non-toxic diluents without TBP
(physical extraction) was found to follow the trend as: soy-
beanoil (0.081, 7.35%) > sesame oil (0.067, 6.19%) > sunflower
oil (0.05, 4.7%). The distribution coefficient increased with
increasing protocatechuic acid concentration. It is observed
that the distribution coefficient is higher in soybean oil than
other natural non-toxic diluents (sunflower oil, and sesame
oil). Soybean oilfavours extraction process by providing better
solvation through hydrogen bonding and limits the formation
of dimers in organic phase.
The values of partition coefficient (P) and dimerisation con-
stant (D) were evaluated by using the curve fitting toolbox
KD = P + 2 P2 D [CPCA ]aq (3)
The curve of KD versus [CPCA ]aq without extractant was
plotted and the values of P and D were obtained from the
non-linear regression. The values of partition coefficient for
protocatechuic acid were estimated as 0.0049, 0.0055, and
(2) 0.0062 in sunflower oil, soybean oil, and sesame oil respec-
tively. Further, dimerisation constant was calculated to be
189,400, 227,200, and 184,200 for sunflower oil, soybean oil,
and sesame oil respectively.
It is observed that distribution coefficient and extraction
efficiency for protocatechuic acid in natural non-toxic diluents
without TBP are not sufficiently high. The extent of hydration
of the acid and the energy of bond to water molecule are the
two main controlling factors for extractability of carboxylic
acid. These low values of distribution coefficients (<1) and
extraction efficiency may attribute to higher affinity of proto-
catechuic acid for water which makes it difficult to separate.
The conventional extraction technique was not successful
for removal of protocatechuic acid in dilute aqueous stream.
Reactive extraction can offer better possibility to increase the
efficiency by employing organophosphorus compound such as
TBP.
The reactive extraction involves a chemical reaction
between protocatechuic acid and TBP. TBP is chemically stable
and can serve an important role as an efficient extractant with
 Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605 597

Table 4 – Chemical equilibrium of protocatechuic acid with tri-n-butyl phosphate in sesame oil at 300.15 ± 1 K.a
[CTBP ]org (mol L−1 ) [CPCA ]0 (mol L−1 ) [CPCA ]aq (mol L−1 ) [CPCA ]org (mol L−1 )
0
KD Avg. KD % Avg. % zPCA

0 0.001 0.0010 0.000013 0.01 0.07 1.26 6.17 –

0.003 0.0029 0.000077 0.03 2.57
0.005 0.0047 0.0003 0.05 5.06
0.007 0.0064 0.0006 0.09 8.39
0.010 0.0086 0.0014 0.16 13.58

0.37 0.001 0.0007 0.0003 0.53 1.58 34.59 58.46 0.0009

0.003 0.0013 0.0017 1.37 57.79 0.0047
0.005 0.0018 0.0032 1.86 64.99 0.0088
0.007 0.0024 0.0046 1.96 66.21 0.0126
0.010 0.0031 0.0069 2.20 68.70 0.0187

0.74 0.001 0.0005 0.0005 0.89 2.44 47.08 68.19 0.0006

0.003 0.0009 0.0021 2.26 69.33 0.0028
0.005 0.0013 0.0037 2.77 73.45 0.0050
0.007 0.0019 0.0051 2.68 72.80 0.0069
0.010 0.0022 0.0078 3.61 78.31 0.0107

1.47 0.001 0.0003 0.0007 1.92 4.45 65.79 79.62 0.0004

0.003 0.0006 0.0024 3.90 79.59 0.0016
0.005 0.0009 0.0041 4.45 81.66 0.0028
0.007 0.0011 0.0059 5.32 84.17 0.0040
0.010 0.0013 0.0087 6.63 86.90 0.0059

2.21 0.001 0.0003 0.0007 2.68 4.92 72.82 81.70 0.0003

0.003 0.0006 0.0024 3.77 79.02 0.0011
0.005 0.0008 0.0042 4.91 83.07 0.0019
0.007 0.0010 0.0060 6.03 85.78 0.0027
0.010 0.0012 0.0088 7.21 87.82 0.0040

2.94 0.001 0.0003 0.0007 2.79 5.81 73.64 83.89 0.0003

0.003 0.0005 0.0025 4.94 83.15 0.0008
0.005 0.0007 0.0043 6.28 86.27 0.0015
0.007 0.0009 0.0061 6.94 87.40 0.0021
0.010 0.0011 0.0089 8.09 89.00 0.0030

a
Standard uncertainties (u) are u(T) = ±1 K; u(HPC) = ±0.001 mol L−1 .

Fig. 1 – Reaction mechanism for extraction of protocatechuic acid in natural non-toxic diluents.

good separation effect with solutions containing chemically TBP (0.37–2.94 mol L−1 ) in different natural non-toxic diluents
similar elements. The value of various parameters for reactive are summarized in Tables 2–4. Fig. 2(a–e) shows chemical equi-
extraction of protocatechuic acid (0.001–0.01 mol L−1 ) using librium isotherms for extraction of protocatechuic acid using
598 Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605

Fig. 2 – Equilibrium isotherms for reactive extraction of protocatechuic acid (0.001–0.01 mol L−1 ) with various concentration
of TBP (0.36–2.94 mol L−1 ) in (a) sunflower oil, (b) soybean oil, (c) sesame oil at 300.15 ± 1 K.

TBP in sunflower oil, soybean oil, and sesame oil respectively.

Extraction of protocatechuic acid involving TBP with natural
non-toxic diluents shows enhanced removal (upto 93.40%) as
compared to physical extraction (maximum 7.35%).
The values of distribution coefficient and extraction effi-
ciency estimated for chemical extraction by TBP in natural
non-toxic diluents for various protocatechuic acid concentra-
tion (0.001–0.01 mol L−1 ) were found to be much higher than
pure diluents signifying the advantage of chemical extrac-
tion over physical extraction. The increase in distribution
coefficient and extraction efficiency with increase in initial
protocatechuic acid concentration was observed. This may be
due to alower concentration range of protocatechuic acid in
initial aqueous solution, for higher initial protocatechuic acid
concentration it may decrease with increasing acid concen-
tration.
The diluents are used in association with TBP to lower its
viscosity, decrease the surface tension at interface, and to
Fig. 3 – Effect of TBP concentration (0.36–2.94 mol L−1 ) on
improve its physical properties. The effect of diluents depends
average distribution coefficient (KD ) for reactive extraction
mainly on its ability to solvate polar ion-pair organic species
of protocatechuic acid in different non-toxic diluents
through hydrogen bonding or dipole–dipole interaction thus
(sunflower oil, soybean oil, and sesame oil) at 300.15 ± 1 K.
favouring the formation of one or more acid-extractant com-
plexes. TBP is used because of low water co-extraction (mass
fraction, 4.67%) and very low solubility in aqueous phase (mass tion of extractant in organic phase (Kertes and King, 1986).
fraction, 0.039%) (Keshav et al., 2008a,b) and does not add sub- Figs. 3 and 4 demonstrate the effect of TBP concentration
stantially to COD of aqueous raffinate. (0.37–2.94 mol L−1 ) on distribution coefficient and extraction
The distribution coefficient and extraction efficiency efficiency of protocatechuic acid. The average distribution
vary with the type of solvent and resulting concentra- coefficients increased from 2.49 to 9.88, 2.21 to 8.79, and 1.58
to 5.81 whereas average extraction efficiency (%) increased
 Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605
Fig. 4 – Effect of TBP concentration (0.36–2.94 mol L−1 ) on
average extraction efficiency (%) for reactive extraction of
protocatechuic acid in different non-toxic diluents
(sunflower oil, soybean oil, and sesame oil) at 300.15 ± 1 K.
from 70.08 to 90.03, 67.39 to 88.78, and 58.48 to 83.89 with
TBP concentration (0.37–2.94 mol L−1 ) in sunflower oil, soy-
bean oil, and sesame oil respectively. It can be observed that
considerable amount of protocatechuic acid was removed
from aqueous solution using TBP. The increase in distribution
coefficient and extraction efficiency were observed at higher
TBP concentration at fixed protocatechuic acid concentration.
This can be attributed to the fact that it contains phospho-
ryl group which is a stronger Lewis base than carbonyl group
and contains PO4 − group which acts as both electron donor
and electron acceptor and undergoes specific interactions like
self-association and molecular complex formation with dilu-
ents and other solutes (Wilke and Chang, 1955). The average
distribution coefficient and average extraction efficiency of
protocatechuic acid follow the trend with different natural
non-toxic diluentsas: sunflower oil > soybean oil > sesame oil.
It may be due to better solvation of sunflower oil with TBP
and presence of various percentages of polyunsaturated fatty
acids having double bonds which favour protocatechuic acid
extraction.
The loading ratio (zPCA ) signifies the extent to which the
organic phase (TBP + diluents) may be loaded with acid and is
defined as the ratio of total acid concentration in organic phase
to the total extractant concentration as (Kertes and King, 1986):
[CPCA ]org
zPCA = (4)
[CTBP ]0org
]0
where [CTBP org is the initial concentration of TBP in
organic phase. Fig. 5 shows the effect of TBP in different nat-
ural non-toxic diluents on loading ratio. It is observed that
loading ratio decreases with increase in TBP concentration for
all concentrations of protocatechuic acid in different diluents
and higher loading were obtained at a low concentration of
TBP and higher protocatechuic acid concentration. The stoi-
chiometry of the overall extraction reaction is based on the
loading ratio in the organic phase. The organic phase in the
extraction of protocatechuic acid is not highly concentrated,
i.e., zPCA < 0.5 for all concentrations of TBP and protocatechuic
acid. This indicates no overloading and formation of only 1:1
complexation of protocatechuic acid–TBP as represented in
Fig. 6. The reversible chemical complexation reaction illus-
599
Fig. 5 – Variation in loading factor (z) with concentration of
TBP (0.36–2.94 mol L−1 ) for reactive extraction of
protocatechuic acid in different natural non-toxic diluents
(sunflower oil, soybean oil, and sesame oil) at 300.15 ± 1 K.
trated in Fig. 6 is an integral part of the reactive extraction
scheme. The production of protocatechuic acid by biosynthe-
sis requires its separation from fermentation broth. The acidic
functional group in solute is capable of complexation and
promote selectivity and enhance uptake capacity. The ben-
efit of transferring protocatechuic acid from aqueous phase to
organic phase is low solute concentration in aqueous phase
which offers a high driving force of free association sites
on complexing agent which is very favourable for extraction
unlike other separation processes. The organic phase can be
regenerated by temperature-swing regeneration for recycling
complexing agent and recovery of protocatechuic acid (Keshav
and Wasewar, 2010). The protocatechuic acid is extracted at
low temperature where the equilibrium favours solvent phase
followed by regeneration at a higher temperature where the
equilibrium favours transfer of protocatechuic acid to aqueous
phase which can then be utilized for various pharmaceutical
applications (Hong et al., 2001).
The value of loading ratio depends on extractability of acid
and its concentration in aqueous phase. The stoichiometry of
overall extraction equilibrium relies on loading ratio in organic
phase. The organic phase for extraction of protocatechuic acid
is not highly concentrated as the values of zPCA < 0.5, indicating
the formation of 1:1 protocatechuic acid–TBP complex and the
following equation holds (Kertes and King, 1986):
zPCA
= K11 [CPCA ]aq (5)
1 − zPCA
where K11 is the experimental extraction equilibrium com-
plexation constant. For a particular diluent, a higher value of
K11 indicates that it can extract protocatechuic acid better than
other diluents along with TBP. The experimental values of K11
for protocatechuic acid in different diluents with TBP follows
the order: sunflower oil > soybean oil > sesame oil. The differ-
ences in values of equilibrium complexation constant for the
same acid in different diluents indicates solvation by diluent
is a critical factor in extraction of protocatechuic acid and poor
solvation of the polar complexes result in the lower values of
K11 and 1:1 complex formation.
The mass action law (Tamada et al., 1990) for describing
the equilibrium for the reactive extraction of protocatechuic
600 Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605
Fig. 6 – Protocatechuic acid–TBP complex (1:1) in different natural non-toxic diluents (sunflower oil, soybean oil, and sesame
oil) at 300.15 ± 1 K.
acid by TBP in various diluents (sunflower oil, soybean oil, and
sesame oil) can be represented as:
11K
CPCA,aq + ˛Corg ↔ [CPCA .C˛ ]org
where ˛ is the solvation number of TBP. The protocatechuic
acid–TBP complex is rapidly extracted into the organic phase
as it is formed at the interphase. The law of mass action is
applied to compute the extraction equilibrium constant (K11 )
and the number of reacting molecules of extractant. It is the
ratio between concentrations of reactant molecules and the
concentration of the product molecules as:
[CPCA .C˛ ]org
K11 =  ˛
[CPCA ]aq Corg
where [CPCA ]aq , [C]org , and [CPCA .C˛ ]org indicate acid, extrac-
tants, and complex concentration in the respective phases.
The effect of acid dissociation for carboxylic acids is negligi-
bly small, hence Eq. (7) can be modified and then solved to
get:
log (KD ) = log (K11 ) + ˛ log[Corg ]
where [C]org can be expressed as:
   
Corg = C0org − ˛[CPCA ]org
The value of ˛ for protocatechuic acid can be taken as
one as the solvation number of carboxyl groups on each acid
were same as the aliphatic carboxylic acids (Niitsu and Sekine,
1978). The model values are in good accordance with the
experimental values. All data are within ±10% data points as
indicated in the combined parity plot in Fig. 7. This model can
be used to describe the reactive extraction of protocatechuic
acid using TBP in different diluents.
The overall equilibrium complexation constant of 1:1
acid–extractant complex is related to relative basicity by rela-
tive basicity model as proposed by Shan (2003). The significant
factors which govern extraction equilibrium are namely acid
hydrophobicity (log P), dissociated equilibrium constant of
acid (pKa ), and apparent basicity of extractant to hydrochloric
acid (pKb ). The relative basicity of extractant represents the
nature of solute, diluent, and extractant, and solvating power
(6)
(7)
Fig. 7 – Parity plot for mass action law model predicted K11
for extraction of protocatechuic acid using TBP in various
natural non-toxic diluents.
if the basicity of the extractant mixture is relative to the solute.
The model is expressed as:
log K11 = [1 (pKb − pKa ) + log (2 P)] (10)
(8) where  1 and  2 are constants. The values of K11 obtained
from the model for the reactive extraction of protocatechuic
acid using TBP in sunflower oil, soyabean oil, and sesame oil
are listed in Table 5. The model values can be seen to be in
good accordance with the experimental values. The equilib-
(9)
rium complexation constant K11 was observed to be in the
range of 6.8–3.3 in sunflower, 6–3 in soybean, and 4.3–1.9 in
sesame for 0.37–2.94 mol L−1 of TBP in organic phase respec-
tively. Sunflower oil exhibited higher K11 value than soybean
and sesame, hence, sunflower oil is more preferable for the
extraction of protocatechuic acid using TBP. The parity plot
for relative basicity model predicted K11 is shown in Fig. 8.
All the data points are within ±10%, hence, relative basic-
ity model can be used to describe the reactive extraction of
protocatechuic acid using TBP in natural non-toxic diluents.
The equilibrium behaviour was modelled by postulating
the formation of various stoichiometric complexes of acid and
extractant as the chemical interaction between the compo-
nents of the acid–extractant complex are strong compared
to the physical interactions in the system. A Langmuir type
of isotherm (Bauer et al., 1989) model is simple but practi-
cally useful to interpret the equilibrium data. The individual
complexes present in the organic phase were identified and
 Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605 601

Table 5 – Equilibrium complexation constant K11 for reactive extraction of protocatechuic acid using TBP in various
natural non-toxic diluents.a
[CTBP ]org (mol L−1 )
0
Solvent K11 (Exp.) K11 (MAL) K11 (RBM) K11 (LM)

Sunflower oil 0.37 6.86 8.26 6.84 7.13

0.74 6.12 7.45 6.09 6.99
1.47 4.56 5.53 4.55 5.45
2.21 3.9 4.72 3.89 3.59
2.94 3.36 4.03 3.36 3.30

Soybean oil 0.37 6.08 7.43 6.08 6.81

0.74 5.34 6.52 5.25 5.50
1.47 3.85 4.72 3.86 3.48
2.21 3.35 4.01 3.35 3.38
2.94 3 3.61 2.99 2.23

Sesame oil 0.37 4.35 5.46 4.31 4.82

0.74 3.34 4.12 3.29 3.34
1.47 3.03 3.72 3.03 3.31
2.21 2.23 2.71 2.23 2.39
2.94 1.97 2.4 1.97 1.58

a
Exp: experimental, MAL: mass action law, RBM: relative basicity model, LM: Langmuir model.

Fig. 8 – Parity plot for modified Langmuir model predicted Fig. 9 – Parity plot for relative basicity model predicted K11
K11 for extraction of protocatechuic acid using TBP in for extraction of protocatechuic acid using TBP in various
various natural non-toxic diluents. natural non-toxic diluents.

their contributions to the overall extraction were determined

as a function of relevant process parameters represented as:
 n
[CPCA ]org K11 [CPCA ]aq
=   (11)
[CPCA ]max
org 1 + K11 [CPCA ]aq n
where n = [CTBP ]max 0
org /[CTBP ]org is assumed to be constant in
the concentration range considered. The model values of equi-
librium constant K11 for sunflower, soybean, and sesame are
in good agreement with the experimental values of K11 . The
parity plot for the combined data shown in Fig. 9 for Langmuir
model is within ±10% (except for 2–3 data points). Hence, this
model can be used to describe the reactive extraction of proto-
catechuic acid using TBP in various natural non-toxic diluents.
The equilibrium behaviour for the reactive extraction of
protocatechuic acid using TBP in various natural non-toxic
solvents was studied using different models (relative basic-
ity model, mass action law model, and Langmuir model). The
comparison of model predicted and experimental values of K11
are shown in Table 5. The trend of accuracy of model based on
the K11 values are relative basicity model > mass action law
model > Langmuir model. The same can be observed from the
parity plots. Hence, the relative basicity model is abest-suited
model to explain the reactive extraction of protocatechuic acid
using TBP in sunflower oil, soybean oil, and sesame oil respec-
tively.
The variation in values of average distribution coefficient,
average extraction efficiency, loading ratio, and extraction
equilibrium complexation constant for different natural non-
toxic diluents is possibly due to its typical composition of
various fatty acids. The typical composition range of fatty
acids for natural non-toxic diluents is presented in Table 6. The
presence of free fatty acids in natural oil may raise additional
concern on the recovery of protocatechuic acid. This compli-
cation was circumvented by pre-treating with distilled water
at 353.15 K to remove free fatty acids by back-extraction (Hong
et al., 2001) from natural oils. Moreover, no formation of third
phase other than aqueous and organic phase was observed
after the extraction of protocatechuic acid into the organic
phase which is otherwise unfavourable. Sunflower oil exhib-
ited better extraction may be due tolower concentrations of
palmitic acid, stearic acid, oleic acid and higher concentra-
tions of linoleic acid, arachidic acid and behenic acid.
Dielectric constant is a measure of solvent polarity and
its ability to insulate charges from each other. Higher
dielectric constant signifies higher polarity and acts as a
602 Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605

Table 6 – Typical composition of fatty acids (%) innatural non-toxic diluents.a

Fatty acid Structure Sunflower oil Soybean oil Sesame oil

Palmitic acid 3–6 7–11 7–9

Stearic acid 1–3 2–6 4–5

Oleic acid 14–35 22–34 40–50

Linolenic acid – 5–11 –

Linoleic acid 44–75 43–56 35–45

Arachidic acid 0.6–4 – 0.4–1

Behenic acid 0.8 – –

a
Typical values adopted from https://www.chempro.in/fattyacid.htm, Accessed May 09, 2017.

oil, it is considerably low. This finding can be correlated with

Table 7 – Diffusion coefficients (DPCA ) of protocatechuic
acid into organic phase from Wilke–Chang equation and average extraction efficiencies of protocatechuic acid in vari-
Reddy–Doraiswamy equation at 300.15 K. ous diluents with TBP. The average extraction efficiencies of
sunflower oil and soyabean oil are nearly same (∼90% for sun-
Organic phase DPCA,W -C DPCA,R-D
(diluent + TBP) (m2 s−1 ) × 10−10 (m2 s−1 ) × 10−10 flower oil and ∼88% for soybean oil) as compared to that of
sesame oil (∼84%). This can be rationalized by their respective
Sunflower oil + TBP 0.10–1.40 4.34–6.95 diffusivities.
Soybean oil + TBP 1.20–1.66 5.23–8.26
The feasibility of extraction for the recovery of protocate-
Sesame oil + TBP 0.90–1.28 4.01–6.46
chuic acid was assessed by calculating S/F ratio and a number
of theoretical stages (Krzyzaniak et al., 2014).
hydrogen bond donor. Sunflower oil exhibited higher values
of average distribution coefficient and average extraction effi- S xin − xout
ciency as compared to soybean oil and sesame oil possibly due = (14)
F min KD xin − yin
to its higher dielectric constant (Table 1).
The reactive extraction of protocatechuic acid involves the
where xin and xout are the concentration of protocatechuic
transfer of species from one phase into another, i.e., transport
acid in feed and raffinate, and yin is the initial concentration
of solute from bulk of the phases to and from the interphase is
of protocatechuic acid in extract phase. The (S/F)min ratio was
an important factor to be taken into consideration. Thus dif-
calculated for the recovery of protocatechuic acid in different
fusion process can be rate determining in extraction process.
diluents with TBP and shown in Table 8. The thumb rule of an
The diffusion coefficients (DPCA ) or diffusivity of protocate-
extraction process with a finite number of extraction stages,
chuic acid into organic phase having avarious concentration of
the S/F ratio is 1.5 times (S/F)min (Seader et al., 1998). The num-
TBP were estimated using Wilke–Chang equation (Wilke and
ber of theoretical stages (NTS) for a counter-current extraction
Chang, 1955) and Reddy–Doraiswamy equation (Reddy and
processes were found using the modified Kremser equation
Doraiswamy, 1967) using Eqs. (12) and (13) respectively as:
with extraction factor Ex (Krzyzaniak et al., 2014) as:
√
T M   
DPCA,W−C = 7.4 × 10−12  0.6 (12) ln
xin −yin /KD
(1 − 1/Ex ) + 1
xout − yin /KD Ex
 V acid NTS = (15)
ln Ex
√
T M
DPCA,R−D = 10−11  1/3 (13) S
Ex = KD (16)
 V diluent V acid F

where  denotes the diluents association factor, V signi-
fies the molar volume of the component, T is the temperature
(K), M is the molecular weight (g mol−1 ) and ␩ is the viscos-
ity (cP) of the diluents. The values of diffusion coefficient are
enlisted in Table 7. The diffusivities of protocatechuic acid into
sunflower oil and soyabean oil are nearly same, but in sesame
These correlations consider significant assumptions and
an attempt was made to extend it to non-ideal mixtures. The
values of NTS showed that at maximum, approximately 5
theoretical stages would be sufficient to achieve the desired
extraction efficiency of protocatechuic acid in a continuous
extraction column.
 Chemical Engineering Research and Design 1 3 2 ( 2 0 1 8 ) 593–605 603

Table 8 – Minimum solvent – to – feed (S/F) ratio and number of theoretical stages (NTS) for the recovery of protocatechuic
acid.
Diluent xin xout KD (S/F)min (S/F)actual NTS

Sunflower oil 0.001 0.0002 5.55 0.14 0.21 2.80

0.003 0.0003 8.18 0.11 0.16 4.01
0.005 0.0005 10.07 0.09 0.13 4.01
0.007 0.0006 11.43 0.08 0.11 4.30
0.01 0.0007 14.16 0.06 0.09 4.68

Soybean oil 0.001 0.0002 4.68 0.17 0.25 2.80

0.003 0.0004 7.05 0.12 0.18 3.50
0.005 0.0005 8.67 0.10 0.15 4.01
0.007 0.0006 10.92 0.08 0.12 4.30
0.01 0.0007 12.61 0.07 0.11 4.68

Sesame oil 0.001 0.0003 2.79 0.25 0.37 2.15

0.003 0.0005 4.94 0.16 0.25 3.10
0.005 0.0007 6.28 0.13 0.20 3.40
0.007 0.0009 6.94 0.12 0.18 3.55
0.01 0.0011 8.09 0.11 0.16 3.83

4. Conclusion
The isothermal equilibrium distribution of protocatechuic
acid onto an aqueous-organic biphasic system containing TBP
as a reactive extractant has been explicated. The simultane-
ous effects of chemical and physical interactions were closely
related to the nature of diluents. Inexpensive and natural
non-toxic solvents with less environmental impact such as
sunflower oil, soybean oil, and sesame oil were used alone and
in combination with TBP for extraction of protocatechuic acid.
The results are expressed in terms of different parameters
such as distribution coefficient, extraction efficiency, loading
ratio, and equilibrium complexation constant. The aver-
age distribution coefficient and average extraction efficiency
was found to follow the trend as:sunflower oil > soybean
oil > sesame oil. The highest achievable protocatechuic acid
removal efficiency is 93.40% and distribution coefficient 14.16
with TBP (2.94 mol L−1 ) in sunflower oil. The loading ratio was
less than 0.5, no overloading was obtained and only 1:1 pro-
tocatechuic acid-TBP complex was observed. The equilibrium
complexation constants (K11 ) for reactive extraction of pro-
tocatechuic acid in TBP (0.37–2.94 mol L−1 ) were estimated by
different models like mass action law, relative basicity, and
Langmuir model. Relative basicity is the best-suited model for
reactive extraction of protocatechuic acid with sunflower oil
showing highest K11 values. The diffusion coefficients (DPCA )
of protocatechuic acid were estimated using Wilke–Chang
equation and Reddy–Doraiswamy equation. The number of
theoretical stages (NTS) for counter current extraction was
found to be 5 using modified Kremser equation. The use
of these natural non-toxic diluents can reduce the toxicity
upto certain levels. Sunflower oil is found to be more prefer-
ableoversoybean oil, and sesame oil.
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