The 9th International Chemical Engineering Congress & Exhibition (IChEC 2015)

Shiraz, Iran, 26-28 December, 2015

Effects of Operating Conditions On Supercritical Extraction of

Glycyrrhizic Acid from Glycyrrhiza Glabra L. Using Methanol As
Modifier

Ali Hedayati, Seyyed Mohammad Ghoreishi*

Department of Chemical Engineering, Isfahan University of Technology, Isfahan 84156-83111, Iran
*Corresponding Author: e-mail: ghoreshi@cc.iut.ac.ir

Abstract
The extraction of Glycyrrhizic acid (GA) from Glycyrrhiza glabra root was investigated by
Soxhlet extraction and modified supercritical CO2 with methanol as co-solvents, 30 min of static
extraction time and 1.4 ml/min of CO 2 flow rate. Design of experiment was carried out with
response surface methodology (RSM) using Mini Tab software 17. The operating temperature (48-
81°C), pressure (12-32 MPa) and dynamic extraction time (47-113 min) were considered as the
range of operating variables. Response surface analysis verified that R2 and modified R2 of the
model were 96.51% and 93.37%, respectively. The RSM modeling predicted the optimal operating
conditions to be the pressure of 28.8 MPa, temperature of 66.5 ◦C and dynamic extraction time of
101 min in which the maximum recovery of 47.8% was obtained.

Keywords: Supercritical extraction; Glycyrrhizic acid (GA); response surface design;

optimization

Introduction
Glycyrrhiza glabra a leguminous shrub having a height of 70-200 cm occurs mainly in sub-
tropical regions where it grows wild and also under cultivation. Chemical constituents of the
root include several bioactive compounds, such as glycyrrhizin, different sugars (up to 18%),
flavonoids, saponins, sterols, starches, amino acids, gums and essential oils. GA has anti-
inflammatory, anti-ulcer, anti-oxidant, anti-hepatotoxic and anti-virus activities. Therefore
GA extraction from licorice root and its characterization is considered to be an important
research area.Technology for the extraction of GA from licorice has been the subject of recent
reports[1]. Solvent extraction is not affordable due to the high consumption of energy and
solvent, also complete solvent recovery is not possible which is very important in food and
drugs application. Disadvantages of these methods could be eliminated by replacing the toxic
liquid solvent with supercritical CO2, because CO2 is inexpensive, nontoxic, nonflammable
with low critical temperature and pressure and environmentally benign[2,3]. Attentions to the
supercritical technology are growing due to the unique properties of supercritical fluids, such
as high selectivity, liquid-like densities, gas-like viscosities and low surface tension. Co-
solvents can be used to change the polarity of SC-CO2 and increase its solvation power to
desired analyte[4]. The main objective of this research was GA extraction from licorice by
modified SC-CO2 in periodic static-dynamic procedure for pharmaceutical application. The
optimization was carried out by response surface methodology (RSM)[5].
The 9th International Chemical Engineering Congress & Exhibition (IChEC 2015)
Shiraz, Iran, 26-28 December, 2015

Experimental
The supercritical extraction system shown in Fig. 1 was used in this study. To increase the
purity of the CO2, which is stored in a CO2 cylinder (1), it was passed through a column of
molecular sieve and metal porous filter (2). Then, CO2 is cooled down in a chiller and pump
head cooler (3, 4) in the range of −10 to -5 ◦C. Then liquefied CO2 is charged by a feed pump
(5) through the needle valve (6) and is fed to the oven that controlled the temperature (298–
523 ± 0.5 K) with proportional–integral–derivative controller (11). Two vessels (modifier
storage column and extraction column) were placed in the oven. The first one (8) is filled with
methanol as modifier (10). CO2 is preheated using spring coil preheater (7) before entering the
modifier vessel (8) which is inside the oven. Then CO2 was first charged into the entrainer
vessel (8) and saturated with methanol/water mixture. Then it was preheated using a spring
coil preheater (9) before entering the extraction column. This column (10) was charged with 2
g of licorice root. Subsequently, modified SC-CO2 was passed through the second vessel and
carried out the extraction of GA from licorice root. In this process, the pressure of the
extraction vessel is controlled via needle valve (12) and back pressure regulator (BPR) valve,
in which the pressure fluctuation can be controlled within 0.2 MPa. (13). Finally, the modified
SC-CO2 is expanded via passage through the BPR valve and the extracted material in all
experiments were collected in bottles containing 10 ml of methanol that were placed in a
vessel containing ice, salt, and acetone (14).

Fig. 1: Schematic diagram of experimental setup for the supercritical extraction system
The GA recovery and extraction yield were calculated using the following equations:
recovery of GA=(mass of GA)/(total of extractable GA)×100 (1)
yield of extraction=(mass of GA)/(mass of licorice root) (2)
For Soxhlet extraction, 2 g of ground licorice root was weighted and set in a Soxhlet apparatus
and then continuously extracted for 8 hours using methanol as solvent. Then the amount of
GA was determined by HPLC method to provided a yield of 138 mg GA/g licorice root
which was considered as the total extractable GA content while calculating the GA recovery
by SC-CO2 extraction.
The Venusil MP C18 column was used (length = 250 mm, internal diameter = 4.6 mm and
particle characteristic= 5µm, 100Å, 360m2/g ultra-pure (99.999%) silica) for HPLC alalysis.
Mobile phase, mobile phase velocity, temperature, detector and injected volume were mixture
of 41% methanol and 59% water (3% acetic acid), 1 ml/min, ambient temperature, UV
detector with 248 nm wavelength and 20 µL, respectively.

Results and discussion

The shown experimental data in Table 1 were used to calculate the coefficients of the second-
order polynomial equation by the least squares technique based on RSM. These regression
coefficients were applied in equation (3) and the significance of each coefficient was
determined.
Table 1. Experimental data for the yield of galegine obtained from CCD.
The 9th International Chemical Engineering Congress & Exhibition (IChEC 2015)
Shiraz, Iran, 26-28 December, 2015
Run Coded variable Process variable Extraction
NO. Recovery(%)
Pressure (P) Time (t) Temperature (T) P t T
1 1 -1 -1 28 60 55 37.7
2 0 0 0 22 80 65 43.4
3 0 0 0 22 80 65 45.6
4 -1 -1 -1 16 60 55 30.4
5 0 -1.68 0 22 47 65 34.7
6 0 1.68 0 22 113 65 44.6
7 1.68 0 0 32 80 65 46.3
8 -1 1 1 16 100 75 39.2
9 1 1 -1 28 100 55 46.2
10 1 1 1 28 100 75 46.6
11 0 0 -1.68 22 80 48 34.6
12 0 0 0 22 80 65 44.7
13 0 0 1.68 22 80 81 38
14 1 -1 1 28 60 75 39.6
15 0 0 0 22 80 65 46.4
16 -1 1 -1 16 100 75 38.2
17 -1 -1 1 16 60 55 30.1
18 0 0 0 22 80 65 41.9
19 -1.68 0 0 12 80 65 32.3
20 0 0 0 22 80 65 45.4

The second-order polynomial model was obtained for GA recovery (R) as a function of
independent variables and based on analyse of variances as follows:
Recovery = 44.553 + 4.060P + 3.614t - 1.774 P2 - 1.650 t2 - 2.834 T2 (3)
Fig. 2 shows the effects of pressure and temperature on GA recovery at the fixed dynamic
extraction time (80 min) and flow rate of SC-CO2 (1.4 ml/min). The SC-CO2 density increases
with increasing pressure that leads to improved GA solubility and therefore causes higher
recovery. On the other hand, increasing the pressure decreased the GA diffusivity, convective
mass transfer coefficient, and, resulting in a lower GA recovery. The trend of Fig. 2-a and 2-b
illustrates that increasing operating pressure increases the extraction recovery because of
overwhelming positive effect of increased SC-CO2 density. But with further increasing of
pressure, the effect of decreasing permeability and mass transfer coefficient prevails the
increasing effect of density and GA recovery decreased. Also as temperature increases, a dual
counter effect is observed. On one hand, increasing the temperature leads to lower SC-CO2
density and, therefore, reduces the solubility of GA and causes lower recovery. On the other
hand, higher temperature increases the GA vapor pressure, resulting in enhanced recovery.
Based on which of these two effects is dominant, the effect of temperature on the extraction
will be different. As illustrated in Fig. 2-c and 2-d , increasing temperature leads to more
extraction recovery but further increasing temperature, the positive effect of temperature with
enhanced mass transfer coefficient and diffusivity is more dominant than the negative effect
of temperature with lower SCF density. The maximum GA recovery of 47.8% was predicted
by the RSM model at the optimal operating conditions of 8.8 MPa, 66oC and 101 min
(dynamic time).
The 9th International Chemical Engineering Congress & Exhibition (IChEC 2015)
Shiraz, Iran, 26-28 December, 2015

(a) (c)

(d)
(b)

Fig. 2: The effects of extraction dynamic time, pressure and temperature on GA extraction recovery (a) contour plot of t and P
(b) surface plot of t and P (a) contour plot of t and T (b) surface plot of t and T.

Conclusions
In this study, optimal conditions for the SFE of Glycyrrhizic acid from licorice root were
investigated using modified SC-CO2 with polar entrainer. Response surface analysis verified
that the data were adequately fitted to second-order polynomial model. The predicted and
experimental results of this study could be utilized by process engineers for design and scale
up of SCF of GA in the pharmaceutical industry.

Acknowledgements
The financial support provided by Isfahan University of Technology is gratefully acknowledged.

References
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