Sie sind auf Seite 1von 9

Biochemical Engineering Journal 151 (2019) 107298

Contents lists available at ScienceDirect

Biochemical Engineering Journal


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

Regular article

Orange peel waste valorisation through limonene extraction using bio-based T


solvents
Baranse Ozturka, James Winterburna, Maria Gonzalez-Miquela,b,

a
School of Chemical Engineering and Analytical Science, Faculty of Science and Engineering, The University of Manchester, Manchester, M13 9PL, United Kingdom
b
Departamento de Ingeniería Química Industrial y del Medioambiente, ETS Ingenieros Industriales, Universidad Politécnica de Madrid, C/José Gutiérrez Abascal 2, 28006,
Madrid, Spain

HIGHLIGHTS

• Green solvents are promising to develop sustainable citrus waste biorefineries.


• CPME and 2-MeTHF are efficient bio-based solvents to extract limonene from orange peels.
• Recycling and reuse of bio-based solvents for limonene extraction is feasible.
• Extraction bio-based solvents promote efficient structure biomass disruption.
• COSMO-RS is an effective tool to select green solvents for limonene extraction.

ARTICLE INFO ABSTRACT

Keywords: Orange peel waste (OPW) can be an effective feedstock for extraction of natural bioactive components such as
Orange peel waste limonene, a high value-added chemical broadly exploited for food, pharmaceutical, and cosmetic industrial
Limonene applications. Extraction of limonene from OPW has been conventionally performed via solvent extraction using
Solvent extraction hexane, a hazardous petrochemical solvent currently restricted under international regulations. In this work, we
Green solvents
have conducted a comparative assessment of the performance of a variety of green solvents for sustainable
COSMO-RS
valorisation of OPW through limonene extraction. In particular, cyclopentyl methyl ether (CPME), ethyl lactate
(EL), isopropyl alcohol (IPA), polyethylene glycol 300 (PEG 300), isopropyl acetate (IAc), dimethyl carbonate
(DMC), methyl ethyl ketone (MEK), 2-methyl-tetrahydrofuran (2-MeTHF) and ethyl acetate (EAc) have been
evaluated as hexane replacement for the recovery of limonene from OPW. Initially, a preliminary solvent
screening was carried out using the COnductor-like Screening MOdel for Real Solvents (COSMO-RS) to estimate
the solubility of limonene in the proposed solvents and rank their theoretical extraction performance.
Afterwards, experimental studies were performed to determine the limonene extraction yields and optimize the
operating conditions (temperature, time and solvent load) for limonene recovery from OPW using the various
solvents, as confirmed by gas chromatography mass spectrometry (GC–MS) analysis. Overall, results support that
CPME and 2-MeTHF bio-based solvents significantly outperform the benchmark petrochemical solvent hexane
by increasing limonene extraction yields up to 80% and 40% respectively at optimum operating conditions.
Moreover, recovery and reuse of these solvents in consecutive extraction cycles was successfully accomplished,
while scanning electron microscopy analysis (SEM) suggests that solvent effects on biomass structure disruption
could be beneficial for further bioprocessing.

1. Introduction 1.6 Gt, accounting for a carbon footprint of 3.3 Gt of CO2 equivalent and
a direct economic cost of USD 750 billion per year [1]. The citrus in-
Food waste has become a major problem worldwide, as roughly one dustry is a major contributor to food waste, as over 70 Mt of oranges are
third of all the food produced for human consumption is wasted every produced annually worldwide [2], with juice processing plants gen-
year. In particular, the global volume of food waste is estimated at erating peel residues up to 50–60% of the total fruit weight. However,


Corresponding author at: Departamento de Ingeniería Química Industrial y del Medioambiente, ETS Ingenieros Industriales, Universidad Politécnica de Madrid,
C/José Gutiérrez Abascal 2, 28006, Madrid, Spain
E-mail address: maria.gonzalezmiquel@upm.es (M. Gonzalez-Miquel).

https://doi.org/10.1016/j.bej.2019.107298
Received 7 March 2019; Received in revised form 19 June 2019; Accepted 10 July 2019
Available online 17 July 2019
1369-703X/ © 2019 Elsevier B.V. All rights reserved.
B. Ozturk, et al. Biochemical Engineering Journal 151 (2019) 107298

orange peel waste (OPW) is a chemically complex and highly biode- extraction from citrus waste has not been performed. Thus, in this work,
gradable residue, that contains a wide array of valuable compounds, we assess the possibility of using alternative, bio-based solvents to di-
including fermentable sugars, carbohydrate polymers, flavonoids, rectly extract limonene from orange waste to promote sustainable va-
polyphenols and essential oils, providing a unique opportunity for lorization processes of citrus by-products. This will lead to identify
feedstock valorisation as a renewable source of high value-added che- promising biorenewable solvents with better extraction peformance
micals and energy [3,4]. than conventional methods to develop citrus waste biorefinery schemes
Essential oils are mixtures of aromatic hydrocarbons found in the with improved overall process performance, lowering energy require-
vesicles located in the flavedo of citrus peel, possessing organoleptic, ments and solvent volumens while increasing product output and
antioxidant and therapeutical properties which are exploited in a quality. Herein, the initial solvent selection was accomplished by
variety of fine chemical, nutritional, and medical applications [5]. Li- carefully considering Environmental, Health and Safety (EHS) para-
monene, a highly lipophilic cyclic monoterpene, is the main constituent meters, as well as physico-chemical properties of a variety of green
of citrus essential oil (around 68–98% w/w) and one of the main solvents [26,27]. As a result, cyclopentyl methyl ether (CPME), ethyl
components of citrus peel waste (up to 4% w/w) [6]. Due to its anti- lactate (EL), isopropyl alcohol (IPA), Polyethylene glycol 300 (PEG
oxidant properties and fragrance character [7], limonene is widely used 300), isopropyl acetate (IAc), dimethyl carbonate (DMC), methyl ethyl
as preservative agent in food industry as a Generally Recognized As Safe ketone (MEK), 2-methyl-tetrahydrofuran (2-MeTHF) and ethyl acetate
("GRAS") additive in the Code of Federal Regulations [8]. In fact, li- (EAc) were selected as potential green solvents to replace hexane in the
monene plays a key role in the global flavour and fragrance market extraction of limonene from OPW. Solvents such as EAc, IAc and IPA
valued at over USD 18.6 billion, with increasing demand for industrial have been identified as less hazardous organic solvents in the Pfizer and
applications in nutraceutical, cosmetic and pharmaceutical formula- Sanofi selection guides for pharmaceutical applications [28]. Mean-
tions [9]. Moreover, limonene has been identified as an effective bio- while, EL is a bio-based, food grade solvent that has been previously
based substitute to volatile organic compounds (VOCs) in analytical used to extract phytonutrients and phenolic compounds from fruit and
chemistry, as a building block for manufacturing purposes and as an vegetable by-products [29,30]. In addition, EL, DMC and CPME have
extraction solvent for separation processes [10,11]. This is of special been proposed for the extraction of oils from yeast [31]. Specifically,
interest considering that the bio-based solvent market is expected to CPME and 2-MeTHF, which have been widely recognize as bio-based
reach USD 13.74 billion by 2024 due to current regulations in relation alternatives to ether solvents [28,32], have been proved to be especially
to limit VOC emissions from chemical industrial sectors, including ad- effective for hexane replacement in extraction of carotenoids and mi-
hesives, paints and coatings, pharmaceuticals, domestic and industrial croalgal lipids [33,34]. MEK is a non-halogenated solvent that has been
cleaners, and cosmetics among others [12]. Extraction of limonene suggested to replace chloroform to recover poly(3-hydroxybutyrate-co-
from citrus peel is also essential for bioconversion, as this molecule is 3-hydroxyvalerate) [(PHBV)] from bacterial cells [35] and for crude oil
strongly toxic for fermentative bacteria due to its antibacterial prop- dewaxing applications [36]. PEG 300 was previously suggested as an
erties, acting as a microbial growth inhibitor and hindering biofuel efficient limonene extractant [37], hence it was included in this study
production processes [13,14]. Furthermore, limonene has also been for the purpose of comparison.
used as green solvent for biomolecule extraction [15] as well as a As there is still lack of information regarding application of green
platform chemical to produce value added products through oxidation solvents for citrus waste valorization, the goal of this work is to provide
routes [16]. Hence, extraction of limonene from orange waste is an a comparative assessment of the performance of alternative solvents for
attractive biorefinery strategy for recovering valuable molecules while limonene recovery. To do so, a preliminary solvent screening was
promoting further biological biomass valorization processes. performed using the COnductor-like Screening MOdel for Real Solvents
The design of sustainable extraction processes from natural products (COSMO-RS) [38] as a quantum-chemical approach to estimate the
is nowadays receiving increasing attention to improve resource effi- solubility of limonene in the abovementioned list of preselected 10
ciency and cost-effective supply, while reducing the use and generation solvents (9 green solvents plus hexane as benchmark). Having com-
of hazardous substances. Following the principles set by Green pleted the computational screening to rank the theoretical solvent
Chemistry [17,18], sustainable extraction of natural products should performance, experimental studies were performed by means of solid-
encompass energy efficient techniques along the use of alternative liquid extraction to validate the predicted trends and further optimize
solvents and renewable feedstocks, while ensuring safe and high quality the operating parameters for limonene recovery from actual OPW using
product extracts [19]. Solid-liquid extraction has been proposed as an the various solvents. Experimental results were evaluated in terms of
effective alternative to hydro-distillation, an ancient technique for ex- limonene extraction yield and the effect of key variables including ex-
traction of essential oils that requires high energy consumption and traction temperature, time and solvent load were evaluated to de-
long operational times [20,21]. Conventional solvent extraction for li- termine the most favourable conditions for limonene extraction. Li-
monene recovery has been carried out using hexane as extractant due to monene is an unsaturated hydrocarbon prone to degradation under
its low boiling point and highly hydrophobic character [22]; however, influence of heat and light, hence extraction time and temperature are
hexane is a toxic, petrochemical solvent which causes significant en- key operating conditions to maximize extraction yield whilst avoiding
vironmental and health issues, which is currently restricted by different degradation. In addition, solvent recovery and recycling was success-
international regulations, such as REACH (EC 1907/2006), concerning fully accomplished. Lastly, scanning electron microscope (SEM) eva-
the Registration, Evaluation, Authorization and restrictions of Chemi- luation was performed to illustrate the solvent effects on the biomass
cals, or IPPC (96/61/EC) regarding Integrated Pollution Prevention and structure as this can affect further bioprocessing steps. Overall, this
Control. In this context, the use of benign alternatives to hazardous study offers a comprehensive evaluation of green solvents for citrus
volatile organic solvents in the chemical industry plays an important waste valorization through extraction of high-value limonene for
role, such as the so-called green solvents which are those preferentially market applications, while facilitating further biomass conversion
produced from biomass feedstock (i.e. bio-based solvents) and/or en- processes for sustainable citrus waste biorefinery development.
vironmentally friendly petrochemical solvents that are considered non-
toxic and biodegradable [23]. Our previous works have proved the 2. Materials and methods
possibility to valorize citrus waste using renewable deep eutectic sol-
vents (DESs) for purification of orange essential oils in downstream 2.1. Plant material
processes [24] and extraction of natural antioxidants from orange peel
residues [25]. However, to the best of our knowledge, a systematic OPW was provided by a local juice bar (Falafel Express, Oxford
study addressing the feasibility of using green solvents for limonene Road, Manchester, United Kingdom). The resulting dry orange peel

2
B. Ozturk, et al. Biochemical Engineering Journal 151 (2019) 107298

residue was obtained by initially chopping orange peels and pre- dissolved in methanol and then filtered through 0.45-μm filter to re-
treating with vacuum oven (Vacutherm, ThermoScientific) at 50 °C and move any remaining impurities before chromatographic analysis. All
150 mbar for 24 h to remove moisture. Afterwards, liquid nitrogen was experiments were carried out in triplicate.
applied to avoid degradation of the components present in the peel With the aim of optimizing the extraction conditions, the effect of
following the procedure described elsewhere [39]. Treated peel was important operating parameters including extraction time, temperature
then powdered using a blender and sieved to obtain a particle size of and solid to liquid ratio on limonene recovery from pre-treated OPW
1 mm (mm). was further assessed following a similar extraction procedure.
Extraction time was recorded at every 30 min up to 180 min to find the
2.2. Reagents and solvents point at which equilibrium is reached and solvent is fully saturated, as
beyond this time no more solute will be recovered and hence longer
The following solvents used in the extraction process were pur- extraction times will be nonsensical [22,40]. The effect of increasing
chased from Sigma Aldrich: hexane (purity > 97%), ethyl lactate (EL) the extraction temperature from 30 °C up to 50 °C, 70 °C and 90 °C on
(purity > 98%), isopropyl alcohol (IPA) (purity > 99.7%), cyclopentyl the limonene recovery yield was subsequently evaluated. Increasing
methyl ether (CPME) (purity > 99%), dimethyl carbonate (DMC) temperature can enhance solubility of a solute in a chosen solvent
(purity≥99%), 2-methyl-tetrahydrofuran (2-MeTHF) (purity > 99%), which increases extraction yield; however, a temperature limit of 90 °C
ethyl acetate (EAc) (purity > 99%); isopropyl acetate (IAc) was set as previous studies [41] have shown that degradation of the
(purity≥97%) was purchased from Alfa Aesar and both methyl ethyl product occurs at temperatures above this point. Furthermore, solid to
ketone (MEK) and polyethylene glycol 300 (PEG 300) were purchased liquid ratios of 1:5, 1:10 and 1:20 were studied to evaluate the effect of
from VWR International ltd. Limonene standard (purity > 98%) was solvent load on the extraction performance, as this is also an important
purchased from Fisher Scientific and methanol (purity = 99%) was also parameter to maximise extraction yield whilst avoiding wasting solvent
purchased from Alfa Aesar. Chemical structures of solvents and limo- and giving both economic and environmental benefits. Following the
nene are shown in Fig. 1. extraction, the solvent was recovered by vacuum distillation using ro-
tary evaporator and sample yield recorded. The % yield of limonene
2.3. Limonene extraction was calculated in terms of dry matter (DM) as described in Eq. (1):
weight of limonene obtained after extraction
Limonene was recovered from dry OPW by solid-liquid extraction Limonene yield (%) = × 100
weight of OPW (dry matter , DM )
using hexane (benchmark petroleum derived solvent) and green sol-
vents i.e. cyclopentyl methyl ether (CPME), ethyl lactate (EL), isopropyl (1)
alcohol (IPA), Polyethylene glycol 300 (PG 300), isopropyl acetate To evaluate the feasibility of solvent recycling, the recovered sol-
(IAc), dimethyl carbonate (DMC), methyl ethyl ketone (MEK), 2-me- vents were used without any further purification in subsequent ex-
thyl-tetrahydrofuran (2-MeTHF) and ethyl acetate (EAc), structures traction cycles to recover limonene from OPW following the extraction
shown in Fig. 1. To preliminary assess the feasibility of the solvents for procedure explained above. The overall extraction procedure followed
limonene extraction, the following solid-liquid extraction procedure is depicted in Fig. 2.
was performed at base case conditions [6,22]: pre-treated OPW was
mixed with each extraction solvent at 1:10 solid to liquid ratio in 15 ml 2.4. Analytical method for limonene identification
polypropylene centrifuge tubes and the mixture was mechanically
stirred using a shaking incubator (Labnet VorTemp™ 1550) for 120 min Gas chromatography mass spectrometry (GC − MS) analysis was
at the temperature of 30 °C at 900 rpm speed. The mixture was then performed in all the extracts using Agilent Technologies 5973 equipped
centrifuged at the same temperature for 10 min (Labnet Spectrafuge™ with Elite-5 ms (30 m x0.25 mm x0.25 μm) column following a method
6C Compact Research Centrifuge) at 5000 rpm to separate the super- proposed by Bourgou et al [42]. Helium was used as a carrier gas at a
natant. After achieving complete phase separation, the solvents were flow rate of 1.2 mL per minute (ml/min) and a split ratio of 1:60 using
evaporated under vacuum in a rotary evaporator. The residue was the following temperature program: rising from 50 °C to 240 °C at a rate

Fig. 1. Chemical structures of solvents and limonene.

3
B. Ozturk, et al. Biochemical Engineering Journal 151 (2019) 107298

screenings to evaluate the theoretical performance of novel solvents in


separation processes based on the solubility of target compounds in the
liquid phase. In this study, the calculation of the solubility of limonene
( x j ) in the proposed solvents have been performed using COSMO-RS
model as per Eq. (2) [44]:

log10(xj ) = log10[exp ((µpure


j µsolvent
j Gj, fusion)/RT )] (2)

where µjpure is the chemical potential of pure compound j, µjsolvent is the


chemical potential of j at infinite dilution and Gj, fusion is the free en-
ergy of fusion of j.
All COSMO-RS calculations were performed using COSMOthermX
software, version C30, release 1601 at the BVP86/TZVP/DGA1
quantum chemical level and (BP_TZVP_C30_1601) parametrization as
described elsewhere [45–47]. Previous works have also used COSMO-
RS to predict the solubility values of carotenoids and lipids in green
solvents [31,33,48] as well as to model the thermodynamic behaviour
of terpenic hydrocarbon mixtures [24].

3. Results and discussion

3.1. Preliminary assessment of solvent performance: COSMO-RS vs


experimental evaluation

COSMO-RS simulations were conducted to perform an initial as-


sessment of the capability of the proposed green solvents to replace the
petrochemically derived hexane to extract limonene from OPW, as
presented in Table 1.
Calculations have been performed in terms of the relative solubility,
log(xi), of limonene in the various solvents, setting to 0 the logarithm of
the solvent with the highest solubility value so all other solvents are
given values relative to this; afterwards, this algorithm is converted into
probabilities (%) [33]. Solvents that present log (xi) values close to 0 are
Fig. 2. Scheme for limonene extraction from orange peel waste (OPW) using
the most feasible solvents, hence have higher probability values. There-
bio-based solvents.
fore, these results provide a relative solubility ranking that represent
trends of the solubility of the target solute, i.e. limonene, in the proposed
of 5 °C/min. The injector and detector temperature were both main- solvents. As presented in Table 1, the probability of solubility of limo-
tained at 240 °C. Mass spectra were obtained by means of electron io- nene in the reference solvent hexane is 86%, while there are three sol-
nization at 70 eV in the range of m/z 40–300 with 1 s mass range. Li- vents with higher probability values, which are CPME (P = 100%), 2-
monene was identified as the major and most significant product in all MeTHF (P = 99%) and PEG (P = 92%). Therefore, considering the
the extracts using mass spectra in NIST mass spectroscopy library, global computational results, CPME followed by 2-MeTHF appear to be
comparing retention times and mass spectra against the standard promising bio-based solvents to replace hexane for limonene extraction
component. from citrus peel waste among the proposed green solvent systems. Sub-
sequently, all proposed solvents were experimentally employed to per-
2.5. Scanning electron microscopy form the actual extraction of limonene from citrus peel waste to validate
the computational trends predicted by COSMO-RS.
A ZEISS EVO 60 model microscope was used to obtain scanning The experimental results, which are graphically represented in Fig. 3,
electron microscope (SEM) images to visually observe changes occur- support that all the proposed solvent systems are capable of extracting
ring at the surface of the samples before and after the extraction.
Samples of dry OPW before and after the extraction were dried over-
night to eliminate any water/moisture, as this may adversely affect Table 1
microscopic analysis, and later coated with a thin layer of carbon to COSMO-RS relative solubility, log10(xi), and probability of solubility of limo-
improve the sample conductivity. 1 nm (nm) resolution power and 5 kV nene, P, in various solvents.a.
(kV) acceleration voltage was used to obtain final images.
Solvent log10(xi) P (%)

2.6. Computational methodology Hexane −0.0657 86


Cyclopentyl methyl ether (CPME) 0.0000 100
Ethyl lactate (EL) −0.2881 52
COnductor-like Screening MOdel for Real Solvents (COSMO-RS) is a
Isopropyl alcohol (IPA) −0.5730 27
quantum-chemical method to estimate thermodynamic properties and Isopropyl acetate (IAc) −0.1048 79
phase equilibrium of pure fluids and its mixtures based on the predic- Dimethyl carbonate (DMC) −0.4480 36
tion of the chemical potential of the compounds [38]. This approach Methyl ethyl ketone (MEK) −0.1869 65
combines quantum chemical considerations (COSMO) with statistical 2-methyltetrahydrofuran (2-MeTHF) −0.0029 99
Ethyl acetate (EAc) −0.1687 68
thermodynamics (RS) for prediction of thermophysical properties Polyethylene glycol (PEG) −0.0376 92
without the need of experimental data, as calculations are based on
information relative to the chemical structure of the compounds [43]. a
Bold = Reference; Bold/Italic = Higher than reference; Italic = Lower
Hence, this tool results particularly useful to perform preliminary than reference.

4
B. Ozturk, et al. Biochemical Engineering Journal 151 (2019) 107298

Fig. 3. Limonene yield (%) from OPW (dry matter, DM) at base case experi- Fig. 4. Limonene yield (%) from OPW (dry matter, DM) as a function of the
mental operating conditions (t = 120 min, T = 30 °C, S/L = 1:10) using various extraction time (at T = 30 °C and S/L = 1:10) using various extraction solvents.
extraction solvents.

[50]. Additionally, due to the nature of limonene, prolonged extraction


limonene from OPW; however, solvent performance to replace hexane times could lead to degradation of this unsaturated compound with the
showed variations. The limonene extraction performance under bench- influence of light, heat and air. In this study, from 30 min to 150 min,
mark conditions (t = 120 min, T = 30 °C, S/L = 1:10) using the various the limonene yield using hexane increased from 0.31% to 0.65%, which
extraction solvents is as follows: CPME > 2-MeTHF > PEG represents over two-fold extracted limonene; and with the best two
300 > IAc > Hexane > EAc > MEK > EL > DMC > IPA. In parti- solvents, the limonene extraction yield has changed from 0.63% to
cular, CPME has significantly outperformed the benchmark solvent 1.14% when using CPME and from 0.49% to 0.84% when using 2-
(hexane) by providing 0.81% limonene yield, this is 1.5 fold increase in MeTHF, representing increases of 81% and 71% respectively. Mean-
comparison with the amount extracted by hexane (0.53% limonene while, between 150 and 180 min, the limonene recovery only increased
yield). CPME is an aprotic dipolar solvent [26], which means that pro- by 1.7% with hexane, 2.5% with CPME and 6% with 2-MeTHF.
motes the dissolution of any compound with an alkane chemical struc- Therefore, due to the minimal increase in limonene yield from 150 to
ture such as limonene; in fact, previous studies have shown that CPME 180 min and to avoid degradation of product, 150 min was chosen as
performs particularly well for the extraction of carotenoids, components the optimal extraction time.
that exhibit alkane-based structure [33]. Additionally, 2-MeTHF yielded It is worth noting that stirring rate was set to a high value in all the
0.64% of limonene, which represents 1.2 fold increase in comparison experiments to favour contact between biomass and solvent phases in
with the solute recovery yield provided by the benchmark solvent order to enhance mass transfer and reducing extraction times. However,
hexane. 2-MeTHF is also an aprotic dipolar solvent which has been ef- if lower stirring rates are used, longer extraction times may be required,
fective in extracting vegetable oils from food crops [49] and lipids from which could promote product degradation.
microalgae [34] due to the favourable solvent extraction kinetics, i.e.
starting accessibility and effective diffusivity, enabling solvation of
higher amount of solute at the surface with faster extraction rate than 3.2.2. Extraction temperature
hexane [33]. Note that aprotic highly dipolar solvents like DMC, with Extraction of limonene from OPW with proposed solvents via solid-
moderate hydrogen bonding strength, or amphiprotic solvents including liquid extraction was examined under the temperatures of 30, 50, 70
EL or IPA, with higher hydrogen-bond donor/acceptor ability, have a and 90 °C, by keeping other operating conditions at base case
lower performance than hexane for limonene extraction; this is due to (t = 120 min and S/L = 1:10), to understand the influence of heat on
the increased polarity of those solvents, which are not capable of fa- the extraction ability of the solvents but also to find a limit to avoid
vourably interacting with the non-polar limonene solute; therefore, upon degradation of the target limonene product. Generally, as we can see
mixing, terpenic hydrocarbon molecules will stay associated through from the results in Fig. 5, increasing the operating temperature up to
London dispersion forces and neat solvent molecules will interact 70 °C boosted the extraction efficiency of the process, which could be
through hydrogen bonding, hence limiting effective solute-solvent mo-
lecular interactions, resulting in low extraction yields.
Overall, considering the theoretical limonene solubility trends
computed using COSMO-RS method and the experimental data ob-
tained for actual extraction of limonene from OPW, it can be concluded
that COSMO-RS predictions were consistent with the experimental
observations, thus validating this computational tool for screening of
novel solvents for limonene extraction.

3.2. Optimisation of limonene extraction operating conditions

3.2.1. Extraction time


The change in limonene extraction yield was evaluated as a function
of the extraction time starting from 30 min up to 180 min keeping other
operating conditions at base case (T = 30 °C and S/L = 1:10). Results in
Fig. 4 show that the limonene extraction yields progressively increase
up to 150 min, and afterwards the change in the extraction yield is
significantly less noticeable. This could be explained in terms of the Fig. 5. Limonene yield (%) from OPW (dry matter, DM) as a function of the
Fick’s second law of diffusion, as mass transfer of the solute from bio- extraction temperature (at t = 120 min and S/L = 1:10) using various extrac-
mass to solvent phase only occurs until the system reaches equilibrium tion solvents.

5
B. Ozturk, et al. Biochemical Engineering Journal 151 (2019) 107298

due to the increased solubility and diffusion coefficients of the target


compound in the solvents [51] along the reduced surface tension and
viscosity of the solvent systems, promoting mass transfer of the solute
from solid to liquid phase. Furthermore, at a higher temperature, speed
of the molecular movements is faster, which causes the extracting agent
to diffuse more quickly into the sample and similarly, solute to diffuse
faster from biomass into the solvent. However, at 90 °C, limonene de-
gradation starts occurring as evidenced from the product extraction
yield decreasing gradually. Previous researchers [41] suggested that
70 °C is the highest appropriate temperature for terpene extraction to
avoid degradation of products.
Results have revealed that with hexane, limonene extraction yield
was about 0.99% at 70 °C but decreased to 0.91% limonene yield at
90 °C. Additionally, the limonene extraction yield provided by CPME
was 1.69% at 70 °C, and reduced to 1.51% at 90 °C. Likewise, the li-
Fig. 7. Limonene yield (%) from OPW (dry matter, DM) at optimum experi-
monene recovery yield provided by 2-MeTHF was reduced from 1.34% mental conditions (t = 150 min, T = 70 °C, S/L = 1:10) using various extrac-
at 70 °C to 1.28% 90 °C. According to previous studies, 25% terpene tion solvents.
degradation occurs at 30 min of heating time at 100 °C [52], and 50%
terpene degradation occurs at 24 h of heating time at 120 °C [53].
efficiency of the limonene extraction process for the reasons explained
Therefore, an extraction temperature of 70 °C is deemed appropriate to
above.
enhance the limonene recovery yield while minimizing product de-
gradation.
3.2.4. Optimal extraction conditions
Selecting appropriate operating conditions is vital to enhance li-
3.2.3. Solid/liquid ratio monene recovery from OPW while reducing time, costs and environ-
Operating and capital costs are significantly affected by the solvent mental impact of the extraction process. In this study, extraction time,
load used in the extraction plant, hence solid/liquid (S/L) ratio is an temperature and solid/liquid ratio have been evaluated to optimize the
important parameter to consider for process profitability. Different S/L operating parameters for orange waste valorisation through limonene
ratios of 1:5, 1:10 and 1:20 were studied whilst keeping other condi- recovery, with results supporting 150 min of extraction at 70 °C and
tions at base case (t = 120 min, T = 30 °C) to evaluate the amount of using solid/liquid ratio of 1:10 as favourable conditions for such pur-
solvent required to enhance the limonene recovery yield without pose. Accordingly, limonene extraction essays were conducted for all
compromising the cost factor. As seen in Fig. 6, the limonene yield with solvents under the aforementioned optimal operating conditions, as
all solvents progressively increases with the amount of solvent used, as presented in Fig. 7. Based on these results, it is possible to conclude that
this promotes contact between both phases, which enhance extraction both bio-based solvents CPME and 2-MeTHF significantly outperform
until the equilibrium is reached. Moreover, higher S/L ratios increase hexane by providing limonene extraction yields of 1.78% and 1.37%
the concentration gradient and diffusion rates promoting the extraction respectively, while hexane yielded 0.99% of limonene. This means that
ability of the solvent. CPM increased the limonene recovery yield nearly 80% whereas 2-
Although using a S/L ratio of 1:20 provides the highest extraction MeTHF increased the limonene recovery to around 40% in comparison
yield, operating with large quantities of solvent will increase energy to the benchmark organic solvent hexane. Additionally, PEG 300 was
requirements for heating and pumping, material and equipment cost, as able to slightly enhance the limonene recovery, yielding 1.17%; while
well as effluent disposal while providing less concentrated extracts, IAc, MEK and EAc provided limonene yields of 0.99%, 0.94% and
which may limit the profitability of the plant. Increasing the S/L ratio 0.98% respectively, which are similar to those obtained with hexane.
from 1:5 to 1:10 enhanced the limonene extraction yield from 0.29% to Remaining solvents, i.e. EL, DMC and IPA, were not able to reach the
0.53% for hexane, from 0.66% to 0.81% for CPME, and from 0.49% to limonene extraction yields provided by hexane as a reference solvent.
0.64% for 2-MeTHF. Moreover, when using S/L ratios of 1:10, CPME Furthermore, when comparing limonene extraction results at optimal
and 2-MeTHF outperformed hexane by providing higher limonene re- conditions (Fig. 7) to base case conditions (Fig. 3), it can be noted that
covery. Therefore, 1:10 was chosen as a suitable ratio to enhance the by setting more favourable extraction parameters the limonene re-
covery efficiency was significantly enhanced, increasing over two-fold
for the best solvent candidates, i.e. CPME and 2-MeTHF.
When comparing these results to previous works, it can be con-
firmed that the limonene extraction yield provided by hexane in this
work (0.99% w/w) is similar to the limonene extraction yield reported
for hexane at similar operating conditions (0.91% w/w) [54]. Fur-
thermore, bio-based solvents proposed in this work provide comparable
limonene recovery from orange peels than conventional techniques
such as hydrodistillation (nearly 2% w/w) [6].

3.3. Biomass surface characterization

The cell wall disruption during the extraction process was explored
through Scanning Electron Microscopy (SEM) analysis on dried OPW
biomass before and after the extraction, employing hexane as bench-
mark solvent along the best two green solvents (CPME and 2-MeTHF)
Fig. 6. Limonene yield (%) from OPW (dry matter, DM) as a function of the under optimal extraction conditions (t = 150 min, T = 70 °C and S/
solid/liquid ratio (at T = 30 °C and t = 120 min) using various extraction sol- L = 1:10). As shown in Fig. 8, biomass cells before the extraction pre-
vents. sent uniform and well-defined shapes with large particles due to the

6
B. Ozturk, et al. Biochemical Engineering Journal 151 (2019) 107298

Fig. 8. SEM images of dry OPW biomass (A) before solvent extraction and after the solvent extraction with (B) hexane, (C) CPME and 2-MeTHF (D) under optimal
extraction conditions (t = 150 min, T = 70 °C and S/L = 1:10).

presence of complex linkage formed of long cellulose, hemicellulose,


pectin and lignin polymer chains. Meanwhile, biomass treated with
hexane shows smaller particles with less defined shapes due to the
chemical disruption after the biomass contacted with the extracting
solvent. These modifications in cell wall structure, such as shrinking
and wrinkling with smaller particles, are more noticeable for CPME and
2-MeTHF due to their greater ability to break the complex linkages and
enhance the limonene extraction yields. This is in agreement with
previous studies suggesting that extraction conditions and solvent
nature can greatly affect biomass disruption and recovery of target
compounds [34,40,55]. In fact, CPME and 2-MeTHF, bio-based solvents
that provided higher limonene yield than hexane, also display more
efficient chemical disruption which could be beneficial for further
biomass processing towards promoting integrated bio-refineries for ci-
Fig. 9. Limonene yield (%) from OPW (dry matter, DM) using hexane, CPME
trus waste valorisation.
and 2-MeTHF in three consecutive extraction cycles.

3.4. Solvent recyclability


proposed solvents, CPME and 2-MeTHF, is more efficient than that of
Recyclability of the extraction solvent plays an important role in hexane; in particular, the limonene extraction yields provided by CPME
developing sustainable biomass valorisation processes considering and 2-MeTHF in the third extraction cycle were 0.97% and 0.59% re-
economic and environmental factors. To evaluate the possibility to re- spectively, in comparison to 0.25% obtained with hexane. Overall, this
cover the solvent after the extraction and reuse it within the process, further supports the suitability of replacing hexane with the proposed
three consecutive extraction cycles were carried out under optimal bio-based solvents to develop sustainable processes for citrus waste
conditions using hexane as benchmark solvent along the best two green valorisation.
solvents, CPME and 2-MeTHF, for comparison purposes. As seen in
Fig. 9, the decrease in the limonene extraction yield from cycle 1 to 4. Conclusions
cycle 2 was about 42% when using hexane, whereas such reduction was
around 20% for CPME and 25% for 2-MeTHF. In addition, when going This study demonstrates that green solvents such as bio-based CPME
from cycle 2 to cycle 3, the magnitude of the decrease in the limonene and 2-MeTHF are promising alternatives for replacing hexane in the
extraction yield grows to 55%, 32% and 40% for hexane, CPME and 2- extraction of limonene from orange peel residues to develop sustainable
MeTHF respectively. These results confirm that the recyclability of the bio-refineries for citrus waste valorisation. At optimum extraction

7
B. Ozturk, et al. Biochemical Engineering Journal 151 (2019) 107298

conditions (T = 70 °C, t = 150 min, S/L = 1:10), CPME and 2-MeTHF limonene by bacteria, fungi, yeasts, and plants, Appl. Microbiol. Biotechnol. 61
respectively increased limonene extraction yields up to 80% and 40% in (2003) 269–277, https://doi.org/10.1007/s00253-003-1221-y.
[17] P.T. Anastas, J.C. Warner, Green Chemistry: Theory and Practice, Oxford University
comparison with the benchmark organic solvent hexane. Moreover, Press, Oxford, 1998.
recyclability of these solvents within the process was successfully ac- [18] J.H. Clark, D. Macquarri, Handbook of Green Chemistry and Technology, Wiley-
complished, with CPME and 2-MeTHF providing 4-fold and 2-fold li- Blackwell Publishing, 2002.
[19] F. Chemat, M.A. Vian, G. Cravotto, Green extraction of natural products: concept
monene extraction yields in comparison to that provided by hexane and principles, Int. J. Mol. Sci. 13 (2012) 8615–8627, https://doi.org/10.3390/
after three consecutive extraction cycles. Additional microscopy ana- ijms13078615.
lysis suggests that both solvents promote biomass structure disruption, [20] M. Lahlou, Methods to study the phytochemistry and bioactivity of essential oils,
Phytother. Res. 18 (2004) 435–448.
which could be beneficial for further bioprocessing. Furthermore, [21] C. Galanakis, Nutraceutical and Functional Food Components: Effects of Innovative
COSMO-RS method was proved an effective tool to conduct preliminary Processing Techniques, Academic Press, London, United Kingdom, 2017.
solvent screenings and rank the theoretical performance of green sol- [22] C. Lopresto G, F. Petrillo, A. Casazza A, B. Aliakbarian, P. Perego, V. Calabrò, A non-
conventional method to extract D -limonene from waste lemon peels and compar-
vents for limonene extraction.
ison with traditional Soxhlet extraction, Sep. Purif. Technol. 137 (2014) 13–20,
https://doi.org/10.1016/j.seppur.2014.09.015.
Acknowledgements [23] E. Yara-Varón, A. Selka, A. Fabiano-Tixier, M. Balcells, R. Canela-Garayoa, A. Bily,
M. Touaibia, F. Chemat, Solvent from forestry biomass. Pinane a stable terpene
derived from pine tree byproducts to substitute n-hexane for the extraction of
Baranse Ozturk would like to thank EPSRC (Engineering and bioactive compounds, Green Chem. 18 (2016) 6596–6608, https://doi.org/10.
Physical Sciences Research Council) for the Ph.D. scholarship provided. 1039/c6gc02191c.
[24] B. Ozturk, J. Esteban, M. Gonzalez-Miquel, Deterpenation of citrus essential oils
using glycerol-based deep eutectic solvents, J. Chem. Eng. Data 63 (2018)
Appendix A. Supplementary data 2384–2393, https://doi.org/10.1021/acs.jced.7b00944.
[25] B. Ozturk, C. Parkinson, M. Gonzalez-Miquel, Extraction of polyphenolic anti-
Supplementary material related to this article can be found, in the oxidants from orange peel waste using deep eutectic solvents, Sep. Purif. Technol.
206 (2018) 1–13, https://doi.org/10.1016/j.seppur.2018.05.052.
online version, at doi:https://doi.org/10.1016/j.bej.2019.107298. [26] L. Moity, M. Durand, A. Benazzouz, C. Pierlot, V. Molinier, J.M. Aubry, Panorama of
sustainable solvents using the COSMO-RS approach, Green Chem. 14 (2012) 1132,
References https://doi.org/10.1039/c2gc16515e.
[27] P.G. Jessop, Searching for green solvents, Green Chem. 13 (2011) 1391–1398.
[28] F.P. Byrne, S. Jin, G. Paggiola, T.H.M. Petchey, J.H. Clark, T.J. Farmer, A.J. Hunt,
[1] Food and Agriculture Organization of the United Nations (FAO), Food Wastage C.R. Mcelroy, J. Sherwood, Tools and techniques for solvent selection: green solvent
Footprint - Impacts on Natural Resources – Summary Report, Available at: http:// selection guides, Sustain. Chem. Process. (2016) 1–24, https://doi.org/10.1186/
www.fao.org/sustainable-food-value-chains/library/detalles/es/c/266219/ s40508-016-0051-z.
(Accessed on 12 February 2019) (2013). [29] Y. Leng, S. Gan, A. Morris, H. Kiat, Ethyl lactate as a potential green solvent to
[2] Food and Agriculture Organisation of the United Nations, (FAO), Citrus Fruit extract hydrophilic (polar) and lipophilic (non-polar) phytonutrients simulta-
Statistics, Available at: http://www.fao.org/3/a-i5558e.pdf (Accessed on 12 neously from fruit and vegetable by-products, Sustain. Chem. Pharm. 4 (2016)
February 2019) (2016). 21–31, https://doi.org/10.1016/j.scp.2016.07.003.
[3] K. Rezzadori, S. Benedetti, E.R. Amante, Proposals for the residues recovery: orange [30] M. Lores, M. Pájaro, M. Álvarez-casas, J. Domínguez, C. García-jares, Use of ethyl
waste as raw material for new products, Food Bioprod. Process. 90 (4) (2012) lactate to extract bioactive compounds from Cytisus scoparius: comparison of
606–614, https://doi.org/10.1016/j.fbp.2012.06.002. pressurized liquid extraction and medium scale ambient temperature systems,
[4] J. Esteban, M. Ladero, Food waste as a source of value-added chemicals and ma- Talanta 140 (2015) 134–142, https://doi.org/10.1016/j.talanta.2015.03.034.
terials: a biorefinery perspective, Int. J. Food Sci. Technol. 53 (2018) 1–14, https:// [31] C. Breil, A. Meullemiestre, M. Vian, F. Chemat, Bio-based solvents for green ex-
doi.org/10.1111/ijfs.13726. traction of lipids from oleaginous yeast biomass for sustainable aviation biofuel,
[5] B. Satari, K. Karimi, Citrus processing wastes: environmental impacts, recent ad- Molecules (2016) 1–14, https://doi.org/10.3390/molecules21020196.
vances, and future perspectives in total valorization, Resour. Conserv. Recycl. 129 [32] K. Watanabe, N. Yamagiwa, Y. Torisawa, Cyclopentyl methyl ether as a new and
(2018) 153–167, https://doi.org/10.1016/j.resconrec.2017.10.032. alternative process solvent, Org. Process Res. Dev. 11 (2007) 1–6, https://doi.org/
[6] V. Negro, G. Mancini, B. Ruggeri, D. Fino, Citrus waste as feedstock for bio-based 10.1021/op0680136.
products recovery: review on limonene case study and energy valorization, [33] E. Yara-Varón, S. Fabiano-Tixier, M. Balcells, R. Canela-Garayoa, A. Bily, F. Chemat,
Bioresour. Technol. 214 (2016) 806–815, https://doi.org/10.1016/j.biortech.2016. Is it possible to substitute hexane with green solvents for extraction of carotenoids?
05.006. A theoretical versus experimental solubility study, RSC Adv. 6 (2016)
[7] B. Ali, N. Al-wabel, S. Shams, A. Ahamad, S.A. Khan, F. Anwar, Essential oils used in 27750–27759, https://doi.org/10.1039/C6RA03016E.
aromatherapy: a systemic review, Asian Pac. J. Trop. Biomed. 5 (2015) 601–611, [34] W.M.A.W. Mahmood, M. Gonzalez-Miquel, C. Theodoropoulos, Enhanced micro-
https://doi.org/10.1016/j.apjtb.2015.05.007. algal lipid extraction using bio- based solvents for sustainable biofuel production,
[8] CFR (Code of Federal Regulations), (2015) Retrieved from https://www.accessdata. Green Chem. (2017) 5723–5733, https://doi.org/10.1039/c7gc02735d.
fda.gov/scripts/cdrh/cfdocs/cfcfr/cfrsearch.cfm (Accessed on 10 January 2019). [35] Y. Yang, J. Jeon, D.H. Yi, J. Kim, H. Seo, C. Rha, A.J. Sinskey, C.J. Brigham,
[9] Grand View Research, Flavors And Fragrances Market Analysis By Product (Natural, Application of a non-halogenated solvent, methyl ethyl ketone (MEK) for recovery
Aroma), By Application (Flavors, Fragrances), By Region (North America, Europe, of poly (3-hydroxybutyrate-co-3-hydroxyvalerate) [P(HB-co-HV)] from bacterial
APAC, MEA, Central & South America), & Segment Forecasts, 2018–2025, (2019), cells, Biotechnol. Bioprocess Eng. 20 (2015) 291–297, https://doi.org/10.1007/
pp. 2018–2025. Available at: https://www.grandviewresearch.com/industry- s12257-014-0546-y.
analysis/flavors-fragrances-market . (Accessed on 10 January 2019). [36] A. Am, Y. Am, O. Eo, Solvent dewaxing of heavy crude oil with methyl ethyl ketone,
[10] Z. Chemat-djenni, M.A. Ferhat, V. Tomao, Carotenoid extraction from tomato using J. Pet. Environ. Biotechnol. 6 (2015), https://doi.org/10.4172/2157-7463.
a green solvent resulting from orange processing waste, J. Essent. Oil Bear. Plants 1000213.
13 (2013) 139–147, https://doi.org/10.1080/0972060X.2010.10643803. [37] P.S. Kulkarni, C. Brazinha, A.M. Afonso, G. Crespo, Selective extraction of natural
[11] M. Lohrasbi, M. Pourbafrani, C. Niklasson, M.J. Taherzadeh, Process design and products with benign solvents and recovery by organophilic pervaporation: frac-
economic analysis of a citrus waste biorefinery with biofuels and limonene as tionation of D -limonene from orange peels, Green Chem. 12 (2010) 1990–1994,
products, Bioresour. Technol. 101 (2010) 7382–7388, https://doi.org/10.1016/j. https://doi.org/10.1039/c0gc00339e.
biortech.2010.04.078. [38] A. Klamt, F. Eckert, COSMO-RS: a novel and efficient method for the a priori pre-
[12] Grand View Research, Green And Bio – Based Solvents Market Trend Analysis diction of thermophysical data of liquids, Fluid Phase Equilib. 172 (2000) 43–72.
Byproduct (Bio – Alcohols, Bio – Glycols, Bio – Diols, Lactate Esters, Methyl Soyate), [39] K. Ghasemi, Y. Ghasemi, M.A. Ebrahimzadeh, Antioxidant activity, phenol and
By Application (Paints, Industrial & Domestic Cleaners, Adhesives, Printing Inks, flavonoid contents of 13 citrus species peels and tissues, Pak. J. Pharm. Sci. 22
Cosmetics) And Segment, (2019), pp. 1–7. Available at: https://www. (2009) 277–281.
grandviewresearch.com/industry-analysis/green-bio-based-solvents-market . [40] V. Negro, B. Ruggeri, D. Fino, Recovery of D-limonene through moderate tem-
(Accessed on 10 January 2019). perature extraction and pyrolytic products from orange peels, J. Chem. Technol.
[13] B. Ruiz, X. Flotats, Citrus essential oils and their influence on the anaerobic di- Biotechnol. 93 (2017) 1186–1191, https://doi.org/10.1002/jctb.5107.
gestion process: an overview, Waste Manag. 34 (2014) 2063–2079, https://doi.org/ [41] F. Temelli, J.P.O. Connell, C.S. Chen, R.J. Braddock, Thermodynamic analysis of
10.1016/j.wasman.2014.06.026. supercritical carbon dioxide extraction of terpenes from cold-pressed orange oil,
[14] K. Fisher, C. Phillips, Potential antimicrobial uses of essential oils in food: is citrus Ind. Eng. Chem. Res. 29 (1990) 618–624, https://doi.org/10.1021/ie00100a019.
the answer? Trends Food Sci. Technol. 19 (2008) 156–164, https://doi.org/10. [42] S. Bourgou, F.Z. Rahali, I. Ourghemmi, M.S. Tounsi, Changes of peel essential oil
1016/j.tifs.2007.11.006. composition of four Tunisian citrus during fruit maturation, Sci. World J. 12 (2012)
[15] C.D. Tanzi, M.A. Vian, C. Ginies, M. Elmaataoui, F. Chemat, Terpenes as green 1–10, https://doi.org/10.1100/2012/528593.
solvents for extraction of oil from microalgae, Molecules 17 (2012) 8196–8205, [43] A. Klamt, Conductor-like screening model for real solvents: a new approach to the
https://doi.org/10.3390/molecules17078196. quantitative calculation of solvation phenomena, J. Phys. Chem. 97 (1995)
[16] W.A. Duetz, H. Bouwmeester, J.B. Van Beilen, B. Witholt, Biotransformation of 2224–2235.

8
B. Ozturk, et al. Biochemical Engineering Journal 151 (2019) 107298

[44] COSMOthermX User Guide, COSMOlogic GmbH & Co. KG., 2016. N.B. Perry, Extraction of phenolics and essential oil from dried sage (Salvia offici-
[45] J.F.B. Pereira, A. Magri, M.V. Quental, M. Gonzalez-Miquel, M.G. Freire, nalis) using ethanol-water mixtures, Food Chem. 101 (2007) 1417–1424, https://
J.A.P. Coutinho, Alkaloids as alternative probes to characterize the relative hy- doi.org/10.1016/j.foodchem.2006.03.050.
drophobicity of aqueous biphasic systems, ACS Sustain. Chem. Eng. 4 (2016) [51] M. Dent, V. Dragovi, M. Peni, M. Brnčić, T. Bosiljkov, B. Levaj, The effect of ex-
1512–1520, https://doi.org/10.1021/acssuschemeng.5b01466. traction solvents, temperature and time on the composition and mass fraction of
[46] E.J. González, I. Díaz, M. Gonzalez-Miquel, M. Rodríguez, A. Sueiras, On the be- polyphenols in dalmatian wild sage (Salvia officinalis L.) extracts, Food Technol.
havior of imidazolium versus pyrrolidinium ionic liquids as extractants of phenolic Biotechnol. 51 (2013) 84–91, https://doi.org/10.1186/2251-6581-13-11.
compounds from water: experimental and computational analysis, Sep. Purif. [52] Y. Yang, B. Kayan, N. Bozer, B. Pate, C. Baker, A.M. Gizir, Terpene degradation and
Technol. 201 (2018) 214–222, https://doi.org/10.1016/j.seppur.2018.03.006. extraction from basil and oregano leaves using subcritical water, J. Chromatogr. A
[47] J. Esteban, M. Gonzalez-Miquel, Thermodynamic insights on the viscometric and 1152 (2007) 262–267, https://doi.org/10.1016/j.chroma.2006.11.037.
volumetric properties of binary mixtures of ketals and polyols, J. Mol. Liq. 263 [53] R.W. Hemingway, L.L. Ingram, C.S. Canady, W.B. Mcgraw, Thermal degradation of
(2018) 125–138, https://doi.org/10.1016/j.molliq.2018.04.133. terpenes: camphene, Δ3-carene, limonene, and α-terpinene, Environ. Sci. Technol.
[48] A. Filly, A.S. Fabiano-tixier, X. Fernandez, F. Chemat, Alternative solvents for ex- 33 (1999) 4029–4033, https://doi.org/10.1021/es9810641.
traction of food aromas. Experimental and COSMO-RS study, LWT 61 (2015) 33–40, [54] T.M. Attard, B. Watterson, V.L. Budarin, J.H. Clark, A.J. Hunt, Microwave assisted
https://doi.org/10.1016/j.lwt.2014.11.021. extraction as an important technology for valorising orange waste, New J. Chem. 38
[49] A. Sicaire, M. Vian, F. Fine, F. Joffre, P. Carré, S. Tostain, F. Chemat, Alternative (2014) 2278–2283, https://doi.org/10.1039/c4nj00043a.
bio-based solvents for extraction of fat and oils: solubility prediction, global yield, [55] K. Hs, Comparison of two isolation methods for essential oils from orange peel
extraction kinetics, chemical composition and cost of manufacturing, Int. J. Mol. (Citrus auranticum L) as a growth promoter for fish: microwave steam distillation
Sci. 16 (2015) 8430–8453, https://doi.org/10.3390/ijms16048430. and conventional steam distillation, J. Aquac. Res. Dev. 7 (2016), https://doi.org/
[50] N.E. Durling, O.J. Catchpole, J.B. Grey, R.F. Webby, K.A. Mitchell, L.Y. Foo, 10.4172/2155-9546.1000409.