Journal of Supercritical Fluids 22 (2002) 87 – 92
www.elsevier.com/locate/supflu
Phase equilibrium for
capsaicin+water + ethanol+supercritical carbon dioxide
Catarina M.M. Duarte *, Marcelo Crew 1, Teresa Casimiro,
Ana Aguiar-Ricardo, Manuel Nunes da Ponte
Departamento de Quı́mica, Centro de Quı́mica Fina e Biotecnologia, Faculdade de Ciências e Tecnologia,
Uni6ersidade No6a de Lisboa, 2829 -516 Caparica, Lisboa, Portugal
Received 21 February 2001; received in revised form 18 July 2001; accepted 17 August 2001
Abstract
The possibility of extracting capsaicin with supercritical carbon dioxide from a hydroalcholic mixture containing
the alkaloid was investigated. High-pressure phase equilibrium on the quaternary system CO2 + ethanol+water+
capsaicin was measured in order to obtain the separation factor of the natural product from hydro-alcoholic model
mixtures. Experiments on phase equilibrium behavior were performed at several pressures (12, 15 and 18 MPa) and
temperatures of 40 and 50 °C. The separation factors for three mixtures of different composition— 0.40 water + 0.60
ethanol mass fraction and 0.9 water + 0.10 ethanol mass fraction, containing 0.02% of capsaicin and 0.40 water mass
fraction containing 0.04% of capsaicin—are compared. The effect of the water content in the selectivity of the
extraction process is discussed. © 2002 Elsevier Science B.V. All rights reserved.
Keywords: Capsaicin; Alkaloids; Hydroalcoholic mixtures; Vapor– liquid equilibrium; Supercritical fluid extraction
1. Introduction
Extraction with supercritical carbon dioxide has
found many applications in the processing of
natural products [1,2]. Most of these applications
involve extraction from solid materials in large
high-pressure vessels, in batch or semi-batch
mode. Fractionation of liquids in countercurrent
* Corresponding author. Tel.: + 351-212948500; fax: +
351-212948385.
E-mail address: [email protected] (C.M.M. Duarte).
1
On leave from the Department of Chemistry, University of
Nottingham, UK.
extraction columns is a more advantageous process, as it may be performed continuously and
using much smaller volumes under pressure.
However, in most cases the target substances to
be recovered from natural products are trapped in
solid material. Fractionation can only be used as
a secondary step, after a primary extraction with
a liquid solvent.
Supercritical carbon dioxide is perceived as a
non-toxic solvent. If it is to be used only in a
second step of a process, the first extraction
should use one of the few other solvents that are
also considered suitable for contact with products
for human consumption. Hydroalcoholic mixtures
0896-8446/02/$ - see front matter © 2002 Elsevier Science B.V. All rights reserved.
PII: S 0 8 9 6 - 8 4 4 6 ( 0 1 ) 0 0 1 1 4 - 0
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C.M.M. Duarte et al. / J. of Supercritical Fluids 22 (2002) 87–92
are in this category and they are currently used to
extract many different substances from plants.
Moreover, phase equilibrium in the ternary system water+ethanol +carbon dioxide has been
extensively studied [3]. In order to examine the
feasibility of the secondary extraction, the partition coefficient of the substance of interest between high-pressure carbon dioxide and
water+ ethanol mixtures must be determined.
This work is part of a study designed to provide
these data for a series of substances that are
commonly extracted from plants, in particular
alkaloids. Stahl and collaborators [4] were the first
to study the solubility of these substances in supercritical carbon dioxide. Atropine– benzeacetic
acid,
a-(hydroxylmethyl)-8-methyl-8-azabicyclo
[3.2.1] oct-3-yl ester endo (+/ − )-and capsaicin
(8-methyl-N-vanillyil-6-nonenamide) are among
the most soluble. Atropine was the subject of a
previous report [5].
Capsaicin is the ‘hot’ component of paprika
and chili peppers and has found numerous applications [6]. It can be extracted from those plants
using different solvents, including supercritical
carbon dioxide [7] and water+ ethanol mixtures
[8]. Zeljko and Steiner [9] reported solubilities of
capsaicin in dense CO2 at several experimental
conditions.
Phase equilibrium in the ternary system water + ethanol+carbon dioxide has been extensively studied [3]. However, the design of the
supercritical extraction process to carry out the
second stage separation requires a detailed knowledge of the vapor– liquid equilibrium compositions
of
the
quaternary
mixtures
CO2 +ethanol+water +natural product.
The best optimized conditions for the extraction from the solid botanical source with a hydroalcoholic solvent were found to be 0.40
water + 0.60 ethanol mass fraction. After the primary extraction, the composition of the resulting
hydroalcoholic mixture can be changed by evaporation. As the solution is evaporated it becomes
more viscous. The composition of 0.90 water+
0.10 ethanol mass fraction corresponds to the
limit of acceptable viscosity for the second stage
extraction with carbon dioxide in a continuous
mode.
High-pressure phase equilibrium experiments
on quaternary mixtures CO2 + ethanol+ water+
capsaicin were performed at several pressures (12,
15 and 18 MPa) and at the temperatures of 40
and 50 °C. The separation factors for three mixtures of different composition— 0.40 water+0.60
ethanol mass fraction and 0.90 water+ 0.10 ethanol mass fraction, containing 0.02% of capsaicin
and 0.40 water mass fraction containing 0.04% of
capsaicin— are compared. The effect of the water
content in the selectivity of the second stage extraction process is discussed.
2. Experimental
2.1. Materials
The substances used in this work were: 98%
pure capsaicin supplied by Sigma [404-86-4] CAS;
99.8% ethanol from Merck; 99.8% methanol from
Merck; 99.998 mol% carbon dioxide from Air
Liquide. Doubly distilled water was also used.
2.2. Equilibrium determination
The phase equilibrium apparatus is built
around a sapphire tube with the following dimensions, height, 15 cm; internal diameter, 1.9 cm;
external diameter, 3.2 cm. This type of equilibrium cell was described in detail by Pereira et
al. [10] and a similar apparatus was recently described by Gourgouillon and Nunes da Ponte [11].
In this work, modifications were introduced in
the cell design to allow for an improved resistance
to leakage at low pressures, where the unsupported area type seals used on both ends of the
sapphire tube might give problems. Fig. 1 shows
the modified version of the equilibrium cell. The
Teflon seal is extruded against sapphire by means
of two metal pieces tightened one against the
other.
The cell is initially loaded to approximately half
of its height with a mixture of water+ethanol+
capsaicin of known composition, and then purged
with a flow of pure CO2, at low pressure. The cell
is closed, and the CO2 is introduced up to the
desired pressure using a manual pressure genera-
C.M.M. Duarte et al. / J. of Supercritical Fluids 22 (2002) 87–92
tor. The pressure inside the sapphire-cylinder is
measured with a pressure transducer (Omega
PX931-5KSV calibrated between 0 and 343 MPa
(precision, 0.1%; accuracy, 0.15%). The temperature is measured with a platinum-resistance
(HART 5616 RTD) in thermal contact with sapphire cylinder and connected to a PID controller
(HART 2100). The temperature is assumed to be
homogeneous inside the insulated air-bath, heated
by means of a 150 W resistance connected to the
PID controller and a fan. The typical temperature
stability during experiments is 90.01 K.
At fixed temperature, a typical equilibration
time is 1 h. A magnetic bar activated by a magnet
performs stirring inside the cell. Samples are withdrawn from both phases through two high-pressure six-port switching valves, located on the top
and the bottom of the cell, into sample loops. The
gas in the samples is then expanded into calibrated volumes and the amount of CO2 in each
sample is calculated from the measurement of the
pressure increase at the working temperature. The
sample loops are later flushed with methanol to
collect the solute precipitated during the large
pressure drop that occurred with the expansion.
2.3. Analytical aspects
The samples collected were diluted in 10 ml of
methanol. The resulting methanolic solutions of
capsaicin were analyzed by Supercritical Fluid
Chromatography with APCI-MS detection, using
89
the protocol developed by Davidson and collaborators [12]. The measurements were carried out at
the Department of Chemistry of the University of
Nottingham, in a Gilson SF3 system, with a 821
regulation valve interfacing into a VG Trio 2000
single quadrupole mass spectrometer (VG Biotech, Altrincham, UK). The column was a standard HPLC column with cyano propyl stationary
phase, and the eluent phase a 90% CO2 + 10%
methanol mixture. Calibration was obtained via
use of standard samples.
3. Results and discussion
In this work, two water+ ethanol mixtures of
different composition, 0.40 water+ 0.60 ethanol
mass fraction and 0.90 water+ 0.10 ethanol mass
fraction-were used, in order to study the effect of
the water content in the selectivity of the extraction process. Capsaicin was added to these mixtures until a composition of 2× 10 − 4 mass ratio
was obtained. For the 0.40 water mixture, an
additional mixture with 4× 10 − 4 in capsaicin was
also prepared. These concentrations are of the
order of magnitude of those obtained in hydroalcoholic extracts from plants.
Experiments were carried out at pressures of 12,
15 and 18 MPa, and temperatures of 40 and
50 °C. Samples from the vapor (CO2-rich) phase
and liquid (water+ethanol-rich) phase were analyzed for capsaicin content. The compositions of
Fig. 1. Seal detail of the high-pressure equilibrium cell.
C.M.M. Duarte et al. / J. of Supercritical Fluids 22 (2002) 87–92
90
Table 1
Phase equilibrium compositions of water, ethanol and CO2, on the quaternary system CO2+ethanol+water+capsaicin
T (°C)
p (MPa)
Liquid phase
Solubility in CO2
(×10−6 g/g)
xCO2
ywater
yethanol wt % yCO2
0.02% capsaicin in 40% water+60% ethanol
40
12
37.1
44.0
40
15
37.1
44.0
40
18
37.1
43.9
50
12
37.5
44.5
50
15
37.0
43.9
50
18
36.6
43.3
18.9
18.9
19.0
18.0
19.1
20.1
0.8
0.9
1.1
0.7
1.0
1.3
6.3
7.6
8.8
7.0
8.3
9.6
92.9
91.5
90.1
92.3
90.7
89.1
0.04% capsaicin in 40% water+60% ethanol
40
12
37.1
44.0
40
18
37.1
43.9
18.9
19.0
0.8
1.1
6.3
8.8
92.9
90.1
7.6
18.3
0.02% capsaicin in 90% water+10% ethanol
40
12
87.4
7.7
40
15
86.9
7.6
40
18
86.4
7.6
50
12
88.2
7.7
50
15
87.7
7.7
50
18
87.2
7.6
4.9
5.5
6.0
4.1
4.6
5.2
0.2
0.4
0.5
0.2
0.4
0.6
0.8
1.4
1.9
1.6
1.9
2.1
98.9
98.3
97.6
98.1
97.7
97.3
31.5
31.0
24.0
19.8
12.9
14.4
xwater
xethanol wt. %
Gaseous phase
0.51
0.54
0.69
0.16
0.29
0.57
h ?cap
0.51
0.54
0.69
0.16
0.29
0.57
0.4
0.67
83
754
687
59
292
162
Solubility of capsaicin, expressed in terms of mass of capsaicin per mass of carbon dioxide in the gaseous phase, and the separation
factors of capsaicin between the gaseous and liquid phases.
both phases in carbon dioxide, water and ethanol
were calculated from mass balances, on the basis
of the correlation of Duarte et al. [3] of phase
equilibrium data of several authors for the CO2 +
water + ethanol system. It was assumed that the
presence of capsaicin, due to the very small concentrations in both phases, did not significantly
affect the equilibrium ratios for the other
components.
Mass balances of total capsaicin in the cell
(gas + liquid phases) showed that, in the case of
the initial mixture richer in water, 0.90 water+
0.10 ethanol mass fraction, some precipitation of
solid capsaicin occurred when CO2 was added to
the cell. Carbon dioxide acted as anti solvent
upon dissolution in the aqueous mixture.
Table 1 summarizes the experimental vapor–
liquid equilibrium data. This table shows the equilibrium compositions of water and ethanol in the
liquid and gaseous phases, the solubility of capsaicin-expressed in terms of mass of capsaicin per
mass of carbon dioxide in the gas phase-and the
separation factors of the alkaloid between the
gaseous and liquid phases. These later results are
expressed as:
h=
(wtcapsaicin/wtwater + ethanol)gas
(wtcapsaicin/wtwater + ethanol)liquid
In Figs. 2 and 3, the separation factors for the
studied mixtures are plotted as a function of
pressure for the all studied mixtures.
Fig. 2 shows the separation factors obtained for
the mixture of 0.40 water+ 0.60 ethanol mass
fraction containing 0.02% of capsaicin, at 40 and
C.M.M. Duarte et al. / J. of Supercritical Fluids 22 (2002) 87–92
Fig. 2. Separations factors of capsaicin between the gaseous
and liquid phases, as function of pressure. Results obtained for
the mixture of 0.40 water + 0.60 ethanol mass fraction containing 0.02 % of capsaicin, at 40 °C (") and 50 °C (
), and
the results for the mixture containing 0.04 % of capsaicin at
40 °C for the 0.4 water ratio mixture(“).
50 °C, and the results for the mixture containing
0.04% of capsaicin at 40 °C. The three curves are
similar and they show the increasing trend of the
separation factor with increasing pressure and
decreasing temperature. This behavior follows the
density variations of carbon dioxide.
In Fig. 3, the separation factors, obtained for
the mixture of 0.90 water+ 0.10 ethanol mass
fraction containing 0.02% of capsaicin, are plotted
as a function of pressure at 40 and 50 °C. The
separation factor increases with pressure, between
12 and 15 MPa, and shows a slight trend to
decrease when pressure increases up to 18 MPa.
This behavior suggests that the extraction pressure of the natural product from the hydroalco-
91
holic mixture should be performed close to 15
MPa. At higher pressures the small decrease of
the separation factor might be due to the higher
solvent power of carbon dioxide with lose of
selectivity.
The effect of the water content in the separation factor of capsaicin is obviously very large.
When the initial hydroalcoholic mixture is richer
in water, lower concentrations in capsaicin are
allowed in the equilibrium liquid phase due to the
less solute–solvent affinity. Therefore, higher ratios of capsaicin in the gaseous phase are obtained. If the experiments are performed with
hydroalcoholic mixtures richer in ethanol, lower
separation factors are obtained, showing that ethanol prevents the extraction of capsaicin to the
gaseous phase. Ethanol acts as co-solvent to both
water and carbon dioxide, increasing capsaicin
solubility in liquid and gaseous phases. However,
this effect is more significant in the aqueous
phase, and lower equilibrium concentrations of
capsaicin are attained when extracting with carbon dioxide from ethanol richer mixtures.
It may be concluded from these results that
capsaicin should be easily extracted from waterrich mixtures and that a low content in ethanol is
crucial for the success of processing. This means
that supercritical fluid extraction of capsaicin
should be performed from mixtures containing
the maximum quantity of water allowed in the
initial hydroalcholic mixture before the alkaloid
starts to precipitate with carbon dioxide.
Acknowledgements
Fig. 3. Separation factor of capsaicin between the gaseous and
liquid phases, as function of pressure, obtained for the mixture
of 0.90 water +0.10 ethanol mass fraction containing 0.02 %
of capsaicin, at 40 °C (") and 50 °C (
).
The authors are grateful for financial support
from the European Commission through Contract
ERBFAIRCT962003. Marcelo Crew thanks for
financial support from the European Commission
through SOCRATES/ERASMUS 98/99. The authors thanks Keith Helliwell and Derek Petri
from WR&S (UK), George Davidson from University of Nottingham (UK), Bernard Marty from
Microlithe (France) and Erol Drevici from Genex
(Germany) for helpful discussions.
92
C.M.M. Duarte et al. / J. of Supercritical Fluids 22 (2002) 87–92
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