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Ultrasonics Sonochemistry 15 (2008) 308–313

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Ultrasound-assisted extraction of rutin and quercetin from

Euonymus alatus (Thunb.) Sieb
Yi Yang *, Fan Zhang
College of Science, Beijing University of Chemical Technology, Beijing 100029, China

Received 1 February 2007; received in revised form 17 April 2007; accepted 1 May 2007
Available online 13 May 2007

Abstract

An ultrasonic method for the extractions of rutin and quercetin from Euonymus alatus (Thunb.) Sieb was investigated. The influence
of four extraction variables on extraction yield of rutin and quercetin was discussed. The optimum extraction conditions found were: 70%
aqueous ethanol; solvent: sample ratio 40:1 (v/w); extraction time 3 · 30 min. The recovery of rutin and quercetin and the reproducibility
of the extraction method were determined. The optimized ultrasonic extraction conditions were applied to extract rutin and quercetin
from dried stalks of E. alatus (Thunb.) Sieb. The application of sonication method was shown to be highly efficient in the extraction
of rutin and quercetin from E. alatus (Thunb.) Sieb, compared with classical methods. The scanning electron microscopy (SEM) micro-
graphs provided evidence of more rapid opening of plant cells treated by UAE in contrast to maceration.
 2007 Elsevier B.V. All rights reserved.

Keywords: Ultrasound; Rutin; Quercetin; Euonymus alatus (Thunb.) Sieb; Extraction

1. Introduction the consumption of a large amount of solvent and the long

Euonymus alatus (Thunb.) Sieb is a plant which has been
used as Chinese folk remedies for over 2000 years. Rutin
and quercetin from the extracts is of great importance,
mainly because they can be used as an excellent source of
pharmaceutical products for phytotherapy. Its therapeutic
action is mainly related to the heart and kidney [1]. The
therapeutic effects of it such as antimicrobial and antihy-
pertensive have been reported [2]. Rutin and quercetin were
obtained in the stalks of E. alatus (Thunb.) Sieb, resulting
in identical therapeutic potency.
In conventional methods, the extraction of rutin and
quercetin is accomplished by heating, boiling or refluxing.
The disadvantage of this procedure is the loss of rutin
and quercetin due to ionization, hydrolysis and oxidation
during extraction [3–5]. Moreover, the methods brought
*
Corresponding author. Tel.: +86 10 64454599; fax: +86 10 64434898.
E-mail address: yangyi@mail.buct.edu.cn (Y. Yang).
extraction time [6–8].
The main improvements of sonication for the extractions
are related to the yield and shortening of the extraction time
[9,10]. Ultrasounds produce cell disruption, particle size
reduction and ultrasonic jet towards solid’s surfaces leading
to a greater contact area between solid and liquid phase,
better access of solvent to valuable components [11,12].
The enhancement of extraction yield of organic compounds
by ultrasound is attributed to the phenomenon called
acoustic cavitation. If the ultrasound intensity is sufficient,
the expansion cycle can create cavities or micro-bubbles in
the liquid. Once formed, these bubbles will absorb the
energy from the sound waves and grow during the expan-
sion cycles and recompress during the compression cycle.
The increase in pressure and temperature caused by the
compression leads to the collapse of the bubbles, which
causes shock wave that passes through the solvent, enhanc-
ing the mass transfer within the system [13–15].
Nowadays, sonication has been employed to extract
active compounds such as flavonoids [16], hydrocarbons

1350-4177/$ - see front matter  2007 Elsevier B.V. All rights reserved.
doi:10.1016/j.ultsonch.2007.05.001
 Y. Yang, F. Zhang / Ultrasonics Sonochemistry 15 (2008) 308–313
[17], fatty acid esters [18–20], antioxidants [21], steroids [22]
and anthraquinones [23,24] from plant materials. So far,
for the extraction of rutin and quercetin from dried stalks
of E. alatus (Thunb.) Sieb using ultrasound, no such work
has been reported in the literature. Hence, it is of interest to
investigate the effect of ultrasound on the extraction of
rutin and quercetin from these materials. In this paper,
the extraction conditions for extracting rutin and quercetin
from dried stalks of E. alatus (Thunb.) Sieb under ultra-
sonic have been studied and optimum condition for ultra-
sonic-assisted extraction established.
2. Materials and methods
2.1. Plant materials
The stalks of E. alatus (Thunb.) Sieb were collected in
Liaoning Province, China. The samples were dried in an
oven (24 h at 100 C) and ground (60 meshs). The ground
samples were kept in a dry place before use.
2.2. Maceration
Maceration was performed with 1.0 g of dried stalks and
100 mL ethanol. The mixture was left at ambient tempera-
ture for 6 h, with sporadic shaking with a glass agitator.
Then, the extract was filtered (0.45 lm (millipore)) for
HPLC analysis.
2.3. Ultrasound-assisted extraction
For the ultrasound-assisted extraction experiments, an
ultrasonic bath was used as an ultrasound source. The bath
(KQ-100DB, Kunshan Ultrasound Co. Ltd., China) was a
rectangular container (23.5 · 13.3 · 10.2 cm), to which
50 kHz transducers were annealed at the bottom. The bath
power rating was 100 W on the scale of 0–10. The extrac-
tion of rutin and quercetin was performed by adding
0.5 g of ground dried stalks into 20 mL of solvent in a
25 mL flask. The flask was then partially immersed into
Fig. 1. The liquid chromatograms of standard substances and samples: (a) standard substances; (b) dried stalks extract.
309
the ultrasonic bath, which contains 2.0 L of water. Water
in the ultrasonic bath was circulated and regulated at con-
stant desired temperature to avoid the water temperature
rise, caused by ultrasonic exposure.
2.4. Instrumentation and analytical conditions
The HPLC system (Model 1100, Agilent, USA) com-
prising the following components: system controller,
G1311A pump, and G1314A multi-wavelength detector
were used for the determination of rutin and quercetin.
The column used was an Eclipse XDB-C18 (Agilent,
USA) 5 lm, 150 · 4.6 mm.
For HPLC analysis, CH3OH–H2O–CH3COOH (45:55:
0.5, v/v) was used as the mobile phase, flow rate was
1 mL/min, the column temperature was room temperature
and sample volume injected was 20 lL. The absorbance
was measured at a wavelength of 254 nm for the detection
of rutin and quercetin. Calibration curve method was used
for the HPLC analysis of rutin and quercetin in E. alatus
(Thunb.) Sieb.
2.5. Scanning electron micrographs
In order to elucidate each extraction procedure and to
understand the extraction mechanism, samples were taken
at the end of the extraction processes for scanning electron
microscopy (SEM) analysis. The sectioned particles were
fixed on a specimen holder with aluminum tape and then
sputtered with a thin coating of gold using an Emitech
K500X sputter–coater. All the specimens were examined
using a Cambridge S250MK3 SEM under high vacuum
condition and an accelerating voltage of 20 kV.
3. Results and discussion
3.1. Chromatographic results
The chromatograms of standard and ultrasonically
extracted dried stalks of E. alatus (Thunb.) Sieb are given
310 Y. Yang, F. Zhang / Ultrasonics Sonochemistry 15 (2008) 308–313

in Fig. 1. Fig. 1a shows the standard substances with reten-

tion times of 4.5 and 13.5 min for rutin and quercetin,
respectively. Fig. 1b is the chromatogram of ultrasonically
extracted dried stalks of E. alatus (Thunb.) Sieb.

3.2. Effect of the solvent volume/sample weight ratio

In general, a larger solvent volume can dissolve constit-

uents more effectively leading to an enhancement of the
extraction yield. The influence of solvent volume on the
extraction yield of rutin and quercetin in dried stalks of
E. alatus (Thunb.) Sieb was evaluated (Fig. 2). The results
indicated that an increase of extraction yield of rutin and
quercetin could be observed with the increase of the solvent
volume/sample weight ratio, especially when this ratio was
increased from 20 to 40. Hence, the solvent volume/sample
weight ratio of 40 was selected. Fig. 3. Effect of ethanol–water compositions on extraction yield of rutin
and quercetin from stalks of Euonymus alatus (Thunb.) Sieb.
3.3. Effect of ethanol–water compositions

The effects of ethanol–water compositions on the extrac-
tion yield of rutin and quercetin obtained after 30 min of
ultrasound-assisted extraction at ambient temperature are
shown in Fig. 3. The extraction yields of rutin and querce-
tin increased with the increasing percentage of ethanol up
to 70% ethanol. This was probably due to the relative
polarity and the increase in effective swelling of the plant
by water, which helped increase the surface area for sol-
ute–solvent contact [25]. Thus, 70% aqueous ethanol was
chosen as the extraction solvent in the following studies.
3.4. Effect of extraction time
The influence of extraction time on the extraction yield
of rutin and quercetin from dried stalks of E. alatus
(Thunb.) Sieb is shown in Fig. 4. The results indicate that
in both cases, the extraction yield is highly time-dependent.
Under sonication, the rutin and quercetin extraction yield
increases rapidly with the extraction time for the first
30 min and slowly in the next 90 min.
As shown in Fig. 5, for a fixed total time of 90 min, the
extraction yield of rutin and quercetin is the highest under
sonication when the sample is extracted three times for
30 min each using fresh solvent. When the sample is
extracted twice for 45 min, the yield is also higher than that
the extraction was performed once for 90 min. It is con-
cluded that at a fixed total time of 90 min, three-step
extraction is suitable to achieve good extraction yield of
rutin and quercetin from dried stalks of E. alatus (Thunb.)
Sieb by sonication.
Considering the above results, the following experimen-
tal conditions were optimized: aqueous ethanol (70%);

Fig. 2. Effect of the solvent volume/sample weight ratio on extraction Fig. 4. Effect of the extraction time under sonication on extraction yield of
yield of rutin and quercetin from stalks of Euonymus alatus (Thunb.) Sieb. rutin and quercetin from stalks of Euonymus alatus (Thunb.) Sieb.
 Y. Yang, F. Zhang / Ultrasonics Sonochemistry 15 (2008) 308–313 311

Table 2
Comparison between maceration and ultrasonic-assisted extraction
Comparison Maceration Ultrasonic-assisted
extraction
Extraction time (min) 360 3 · 30
Extractant volume (mL) 100 20
Rutin extraction yield (mg/g) 0.157 ± 0.015 0.299 ± 0.022
Quercetin extraction yield 0.009 ± 0.002 0.013 ± 0.001
(mg/g)

maceration. On extraction yield, ultrasonic-assisted extrac-

tion is suitable for sample preparation prior to active com-
ponents analysis.

3.7. Structural changes after extraction

Fig. 5. Effect of the extraction times on extraction yield of rutin and
quercetin from stalks of Euonymus alatus (Thunb.) Sieb.
solvent/sample weight ratio 40/1 (v/w); extraction time
3 · 30 min.
3.5. Recovery and reproducibility
Under the above optimized conditions, three samples of
the stalks with added rutin and quercetin as shown in Table
1 were analyzed. The recovery of rutin and quercetin from
dried stalks of E. alatus (Thunb.) Sieb is 99.5% and 100.3%,
respectively.
To study the reproducibility of the ultrasound-assisted
extraction method for rutin and quercetin from dried stalks
of E. alatus (Thunb.) Sieb, five samples were processed
under the optimized conditions. The extraction yields of
rutin and quercetin showed a good reproducibility of
experiments.
3.6. Comparison between maceration and ultrasonic-assisted
extraction
The comparison results of extraction yields of macera-
tion and ultrasonic-assisted extraction are listed in Table
2. Ultrasonic-assisted extraction gives significantly higher
values, while maceration yielded lower values. On extrac-
tion time, ultrasonic-assisted extraction was faster than
The various extraction methods produced distinguish-
able physical changes in E. alatus (Thunb.) Sieb. Fig. 6a
revealed an SEM micrograph of the untreated stalks, which
could be compared with the structures of the treated sam-
ples presented in Fig. 6b (maceration), Fig. 6c and d (ultra-
sound-assisted extraction). In the case of maceration
(Fig. 6b), puny damage was observed on the external sur-
face of stalks. Fig. 6c and d showed the typical structure
of the stalk surface after ultrasound-assisted extraction
and a number of finestras were observed on the plant
surface.
Extraction from dried materials consists in a two-stage
process involving: (i) steeping vegetable materials in solvent
to facilitate the swelling and the hydration processes; and
(ii) the mass transfer of soluble constituents from the mate-
rial to solvent by diffusion and osmotic processes [26]. This
indicated that the ultrasounds induced a subsequent
change in the surface tension of the cellulose, and a number
of pits appeared on the surface of plant (Fig. 6). The
change of the cellulose could cause the plant to crumble
or rupture more readily. What is more, the pits could be
propitious to diffusion and osmotic process.
4. Conclusions
Two extraction techniques for rutin and quercetin in the
dried stalks of E. alatus (Thunb.) Sieb were investigated
and compared. Ultrasound-assisted extraction shows a
high yield and low solvent consumption. It is important

Table 1
The recovery of rutin and quercetin from dried stalks of Euonymus alatus (Thunb.) Sieb (n = 5)
Sample 102E (mg) 102A (mg) 102F (mg) Recovery (%)
a b a b
1 32.0 1.30 10.0 0.50 41.6a 1.82b 99.1a 101.1b
2 32.0a 1.30b 30.0a 1.50b 61.7a 2.83b 99.5a 101.1b
3 32.0a 1.30b 60.0a 3.00b 91.8a 4.27b 99.8a 99.3b
E: rutin or quercetin content of the sample determined; A: the amount of added rutin or quercetin standard; F: the amount of the sample determined with
added rutin or quercetin standard.
a
Rutin.
b
Quercetin.
312 Y. Yang, F. Zhang / Ultrasonics Sonochemistry 15 (2008) 308–313
Fig. 6. Electron micrograph of stalks: untreated (a); after maceration (b); after ultrasound-assisted extraction (c) and (d).
to note that using ultrasound, the extraction time is signif-
icantly shortened. The ultrasonic extraction technique was
shown to be very efficient in the extraction of rutin and
quercetin from E. alatus (Thunb.) Sieb. From SEM micro-
graph observations, both methods seemed to bring differ-
ent changes on the plant surfaces. Samples irradiated
with ultrasound revealed that a number of pits appeared
on the surface of plant, confirming that ultrasonic caused
the plants to crumble or rupture more readily.
References
[1] T. Fujita, K. Ohra, K. Miyatake, et al., Chem. Pharm. Bull. 43 (1995)
920.
[2] T. Vetrichelran, Biol. Pharm. Bull. 25 (2002) 526.
[3] M. Ohmishi, H. Morishita, H. Iwahashi, Phytochemistry 36 (1994)
579–583.
[4] H. Sakuma, S. Matsushima, S. Munakata, S. Sugawara, Agric. Biol.
Chem. 46 (1982) 1311–1318.
[5] G. Paganga, N. Miller, C.A. Rice-Evans, Free Radical Res. 30 (1999)
153–162.
[6] X. Yan, M. Suzuki, M. Ohnishi-Kameyama, Y. Sada, T. Nakanishi,
T. Nagata, J. Agric. Food Chem. 47 (1999) 4711–4713.
[7] J. Yu, T. Vasanthan, F. Temelli, J. Agric. Food Chem. 49 (2001)
4352–4358.
[8] T. Deyama, T. Ikawa, S. Kigagawa, Chem. Pham. Bull. 35 (1987)
1785–1789.
[9] M. Salisova, S. Toma, T.J. Mason, Ultrason. Sonochem. 4 (1997)
131–134.
[10] P. Valachovic, A. Pechova, T.J. Mason, Ultrason. Sonochem. 8
(2001) 111–117.
[11] T.J. Mason, E.D. Cordemans, Trans. Inst. Chem. Eng. 74 (1996) 511–
516.
[12] B. Sun, M.J. Peng, X.Y. Yang, Chem. Ind. Forest Prod. 19 (1999)
67–70.
[13] M. Toma, M. Vinatoru, L. Paniwnyk, T.J. Mason, Ultrason.
Sonochem. 8 (2001) 137–142.
[14] J. Wu, L. Lin, F. Chau, Ultrason. Sonochem. 8 (4) (2001) 347–
352.
[15] M. Palma, C.G. Barroso, Anal. Chim. Acta 458 (2002) 119–130.
[16] L. Paniwnyk, E. Beaufoy, J.P. Lorimer, T.J. Mason, Ultrason.
Sonochem. 8 (2001) 299–301.
[17] R.A. Jacques, L.S. Freitas, V.F. Perez, C. Dariva, A.P. de Oliveira,
J.V. de Oliveira, E.B. Caramao, Ultrason. Sonochem. 14 (2007) 6–12.
[18] C. Stavarache, M. Vinatoru, R. Nishimura, Y. Maeda, Ultrason.
Sonochem. 12 (2005) 367–372.
[19] C. Stavarache, M. Vinatoru, Y. Maeda, Ultrason. Sonochem. 13
(2006) 401–407.
[20] C. Stavarache, M. Vinatoru, Y. Maeda, Ultrason. Sonochem. 14
(2007) 380–386.
[21] S. Albu, E. Joyce, L. Paniwnyk, J.P. Lorimer, T.J. Mason, Ultrason.
Sonochem. 14 (2004) 261–265.
 Y. Yang, F. Zhang / Ultrasonics Sonochemistry 15 (2008) 308–313 313

[22] E.C. Schinor, M.J. Salvador, I.C.C. Turatti, O.L.A.D. Zucchi, D.A. [24] Y. Yang, X.Y. Hou, Z.K. Guo, Chem. J. Chinese Univ. 26 (2006)
Dias, Ultrason. Sonochem. 11 (2004) 415–421. 1627–1630.
[23] S. Hemwimol, P. Pavasant, A. Shotipruk, Ultrason. Sonochem. 13 [25] H. Li, L. Pordesimo, J. Weiss, Food Res. Int. 37 (7) (2004) 731–738.
(2006) 543–548. [26] M. Vinatoru, Ultrason. Sonochem. 8 (2001) 303–317.