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Isolation of flavonoids from onion skin and their effects on K562

cell viability
Guo-Qing Shi1, Jing Yang1, Jiang Liu2, Sheng-Nan Liu1, Han-Xue Song1, Wen-En Zhao3
and Yan-Qi Liu1
1
School of Food and Bioengineering, Zhengzhou University of Light Industry, Henan, Zhengzhou 450 002,
P. R. China; 2National Engineering Laboratory for Further Processing of Wheat and Corn, Henan University of
Technology, Henan, Zhengzhou 450 000, P. R. China; 3School of Chemical Engineering and Energy, Zhengzhou
University, Henan, Zhengzhou 450 052, P. R. China.

Article Info Abstract

Received: 16 January 2016 To investigate the anti-proliferative activity of flavonoids from onion skins,
Accepted: 4 February 2016 extraction by 50% ethanol (v/v), soxhlet polar fractionation, pH gradient
Available Online: 19 February 2016 separation, thin-layer chromatography, and recrystallization methods were
DOI: 10.3329/bjp.v11iS1.26419 used to isolate and purify flavonoids from dry onion skins. Anti-proliferative
activities of some flavonoids obtained on leukemia K562 cell line were deter-
mined by MTT assay. Results showed that flavonoids of onion skins were
mainly in form of quercetin, kaempferol, isorhamnetin, apigenin-7-O- -D-
glucopyranoside, quercetin-3-O- -D-glucopyranoside, kaempferol-7-O- -D-
glucopyranoside and rutin. Quercetin and kaempferol decreased K562 cell
Cite this article:
viability, and quercetin had stronger effect. However, isorhamnetin and rutin
Shi GQ, Yang J, Liu J, Liu Sn, Song
Hx, Zhao WE, Liu Yq. Isolation of exhibited certain proliferation-promoting effects. It suggests that ortho
flavonoids from onion skin and their hydroxyl groups on B ring of onion flavonoids might be the key structural
effects on K562 cell viability. elements of their cytotoxic effects on K562 cells, and hydroxyl groups in
Bangladesh J Pharmacol. 2016; 11: S18 position 3 or carbonyl groups in position 4 might be one of the structural
-S25. effect elements.

Introduction their structures, because flavonoids with different

structures exhibit a variety of different biological
Onion (Allium cepa L.) is considered to be one of the activities (Kefalas et al., 2006).
world’s oldest cultivated vegetable, and contains high
level of dietary flavonoids (Slimestad et al., 2007), The isolation and identification of flavonoids from
which are present in much higher concentrations in the onion skin would not only enhance our understanding
onion skin than they are in the fleshy bulb (Kim and of the material basis for their biological activities, but
Kim, 2006; Yao et al., 2004; Sellappan and Akoh, 2002). could also provide a theoretical basis for the utilization
There is a growing body of evidence indicating that of these compounds in a number of other areas of
flavonoids may exhibit health-promoting effects (Gri- research.
ffiths et al., 2002).
Significant research efforts have been directed towards
The results of several studies have shown that the the characterization of onion and onion skin flavonoids
physiological functions of flavonoids are dependent on during the last decade (Soltoft et al., 2009; Pérez-
This paper was presented in the 3rd International Conference on Gregorio et al., 2010; Kiassos, et al., 2009; Jin, et al.,
Biomedicine and Pharmaceutics in Zhuhai, China, on December 2011) which has led to a significant increase in our
11-13, 2015. understanding of these compounds. Despite these
 Bangladesh J Pharmacol 2016; 11: S18-S25 S19

studies, information pertaining to the systematic phase was sequentially extracted with 5% (w/v)
isolation and identification of flavonols from discarded Na2CO3 and 1% (w/v) NaOH solutions to give the
onion skin remains scarce. corresponding diethyl ether phase components. The
ethyl acetate phase was subjected to the same
separation method to give the NaHCO3, Na2CO3, and
Materials and Methods NaOH components of the ethyl acetate phase.

Dry red onion skins were collected from Zhengzhou Isolation

North Central Vegetable Wholesale Market, China. The NaOH components of the diethyl ether and ethyl
Onion skin powder was obtained through the sequen- acetate phases were discarded because they contained
tial cleaning, drying and grounding of the onion skins. very low levels of flavonoids. The flavonoids in the
Extraction NaHCO3 and Na2CO3 components of the diethyl ether
and ethyl acetate phases were isolated by preparative
Two hundred grams of onion skin powder was extrac- TLC (developing solvents: toluene/ethyl, formate/
ted with 6 L of 50% aqueous ethanol at 72°C for 2 hours formic acid, and toluene/ethyl formate/ethanol/formic
in a water bath. The resulting extract was then cooled to acid) and recrystallized from chloroform and methanol
ambient temperature and filtered through filter paper. to give seven flavonoids (Table I).
This procedure was repeated two times and the combi-
ned extracts were concentrated under vacuum on a Identification reactions
rotary evaporator near to cream, which was absorbed HCl-Mg reaction was used for the identification of the
onto diatomaceous earth and dried in the air. flavonoids. Briefly, a few milligrams of Mg powder
Soxhlet polar fractionation were added to 1 mL ethanol solutions of the samples
(0.1 mg/mL). A few drops of concentrated hydrochloric
The dried diatomaceous earth samples were sequen-
were added to the solutions, and the resulting mixtures
tially extracted with petroleum ether, diethyl ether, and
were heated in a water bath when necessary. The
ethyl acetate using a soxhlet extractor. Each solvent
formation of a red or purple color indicated the
extraction procedure was conducted for 12 hours. The
occurrence of a positive reaction.
petroleum ether phase was discarded, whereas the
diethyl ether and ethyl acetate extracts were collected Molisch reaction was used for the identification of
for further processing. glycosidic units. Solution I was made as follows: -
naphthol (1 g) was accurately weighed into a volumet-
pH gradient separation
ric flask and dissolved and diluted to 10 mL with 75%
The diethyl ether phase was extracted three times with ethanol. Solution II was concentrated sulfuric acid. Two
2% (w/v) NaHCO3 solution, and the pH of the combi- or three drops of solution I were added to 1 mL of
ned aqueous extracts was adjusted with concentrated ethanol or an aqueous solution of the flavonoid sample,
hydrochloric acid to pH 2. The acidified aqueous was and the resulting solution was thoroughly mixed. A
then extracted with diethyl ether before being evapora- small amount of solution II was then slowly added to
ted to dryness under vacuum to give the NaHCO3 the sample mixture along the wall of the tube. Notably,
components of the diethyl ether phase. The organic the interface between the samples solution and solution

Table I

Flavonoids isolated from onion skins

Source Number Compound name
The NaHCO3 components of diethyl ether phase Compound 1 Isorhamnetin
Compound 2 Quercetin
Compound 3 Kaempferol
The Na2CO3 components of diethyl ether phase Compound 5 Quercetin-3-O- -D-glucopyranoside
The NaHCO3 components of ethyl acetate phase Compound 7 Rutin
Compound 2 Quercetin
The Na2CO3 components of ethyl acetate phase Compound 4 Apigenin-7-O- -D-glucopyranoside
Compound 6 Kaempferol-7-O- -D-glucopyranoside
S20 Bangladesh J Pharmacol 2016; 11: S18-S25

II became purple in color for a positive reaction. Results

Cell culture and drug treatment Structures of isolated compounds
K562 cells were cultured in RPMI 1640 medium Seven flavonoids were isolated from the dry onion
supplemented with decomplemented fetal bovine skins evaluated in the current study (Figure 1). The
serum (10%, v/v), penicillin (100 IU/mL), and strepto- structures of these compounds were elucidated as
mycin (100 μg/mL) in a humidified incubator under 5% follows:
CO2 and 95% air at 37°C.
Compound 1: Physical form, yellow powder; hydro-
The flavonoids were added to the cells using dimethyl chloric acid-Mg reaction, positive; Molisch reaction,
sulfoxide (DMSO) as a solvent. The concentrations of negative. These results suggested that compound 1 was
the DMSO solutions were adjusted to be the same in all a flavonoid without any glycosidic groups. ESI-MS
of experiments, and the final concentrations were never (Agilent 1100 LC-MSD-Trap-XCT, Agilent, Agilent
greater than 0.5% (v/v). The control group received the Technologies Inc. SantaClara, California, USA) in the
same amount of DMSO without any flavonoids. negative ion mode displayed a pseudomolecular ion
Cell viability was measured using an MTT assay. peaks with an m/z value of 315, corresponding to [M–H]
–. Elemental analysis (Flash EA1112 elemental analyzer,
Briefly, the K562 cells were seeded into 96-well culture
plates and cultured with various concentrations of the Thermo Electron SPA Company, USA) showed that the
different flavonoids for 72 hours at 37°C. After the amounts of C and H in compound 1 were 60.44 and
treatment, each cell was treated with an MTT solution 3.83%, respectively. Taken together with the NMR
to a final concentration 500 μg/mL, and the plates were results (Avance-300 NMR spectrometer, Bruker, Bruker
then incubated for 4 hours at 37°C. At the end of the Optics, Germany), the molecular formula of compound
incubation period, the media was carefully removed 1 was determined to be C16H12O7. The Fourier transform
from each well and replaced with 100 μL of DMSO. The infrared (FT-IR) spectrum (Nexus470 Fourier transform
resulting mixtures were then gently agitated to allow infrared spectrometer, Nicolet Instrument Company,
for the solubilization of the precipitated formazan USA) spectrum of compound 1 contained a broad peak
crystals, and the absorbance values of the cells were at 3238 cm-1, as well as a much narrower C-O peak in
then measured at 570 nm using a Bio-Rad model 680 the range of 1250–1050 cm-1, which were consistent with
microplate reader (Richmond, CA, USA). The absor- the presence of hydroxyl groups. Another peak was
bance values of the treated cells were compared with observed at 1656 cm-1, which was attributed to the
those of the controls, where the cells were considered to stretching vibration of a carbonyl group. Several other
be 100% viable. The rate of inhibition could then be peaks were observed at 1616, 1560, 1509 and 1446 cm -1,
calculated using the following equation. together with broader peaks over 3000 cm-1, which were
attributed to an entitative phenyl ring structure. The
Inhibition rate (%) = (1 - absorbance of treatment peaks at 1616, 1509 and 1383 cm-1 could also be
groups / absorbance of control groups) × 100%. indicative of the presence of an oxygen heterocycle.

Basic flavonoid nucleus Isorhamnetin Quercetin Kaempferol

Apigenin-7-O-β-D-glucopyranoside Quercetin-3-O-β-D-glucopyranoside Kaempferol-7-O-β-D-glucopyranoside Rutin

Figure 1: Structures of the flavonoids obtained from onion skins

 Bangladesh J Pharmacol 2016; 11: S18-S25 S21

NMR data: 1H-NMR (300 MHz, DMSO-d6) δ: 12.48 (1H, that the amounts of C and H in compound 3 were 61.91
s, 5-OH), 10.74 (1H, s, 7-OH),9.75 (2H, s, -OH), 7.76 (1H, and 3.63%, respectively. Taken together with the NMR
d, J = 1.7 Hz, H-2 ), 7.69 (1H, dd, J = 1.7, 8.1 Hz, H-6 ), results, these data suggested that the molecular formula
6.95 (1H, d, J = 8.5 Hz, H-5 ), 6.49 (1H, d, J = 2.0 Hz, H- of compound 3 was C15H10O6. The FT-IR spectrum of
8), 6.20 (1H, d, J = 1.7 Hz, H-6), 3.85 (3H, s, -OCH3); 13C- compound 3 contained peaks at 3318 and 1250–1050 cm-
NMR (75 MHz, MeOD) : 175.8 (C-4), 164.1 (C-7)）, 1, which were consistent with the existence of hydroxyl

161.1 (C-9), 156.2 (C-5), 148.4 (C-3 ), 147.6 (C-2), 146.8 (C groups. A peak was also observed at 1661 cm-1, which
-4 ), 135.8 (C-3), 122.2 (C-1 ), 120.8 (C-6 ), 115.6 (C-2 ), was attributed to the stretching vibration of a carbonyl
111.8 (C-5 ), 102.8 (C-10), 98.8 (C-6), 94.0 (C-8), 55.2 (3- group. The vibrational absorption peaks observed at
OCH3). These data were found to be consistent with 1613, 1569, 1508 and 1438 cm-1 were consistent with the
those reported in the literature for isorhamnetin (Park presence of a phenyl ring skeleton, and the peaks at
and Lee, 1996; Bonaccorsi et al., 2005; Ning, 2000; 1611, 1522 and 1408 cm-1 indicated the presence of an
Fossen and Abdersen, 2006; Markham and Geiger, 1994; oxygen-containing heterocycle.
Stochmal et al., 2001), and so compound 1 was NMR data: 1H-NMR (300 MHz, DMSO-d6) δ: 12.50 (1H,
identified as isorhamnetin. s, 5-OH), 10.83 (1H, s, 7-OH-7), 9.38 (3H, s, -OH), 7.67
Compound 2: Physical form, yellow powder; hydro- (1H, d, J = 2.1 Hz, H-2 ), 7.53 (1H, dd, J = 7.2, 1.9 Hz, H-
chloric acid-Mg reaction, positive; Molisch reaction, 6 ), 6.88 (1H, d, J = 8.5 Hz, H-5 ), 6.40 (1H, d, J = 1.7 Hz,
negative. The results suggested that compound 2 could H-8), 6.18 (1H, d, J = 1.7 Hz, H-6); 13C-NMR (75 MHz,
be a flavonoid without any glycoside functionality. ESI- MeOD) δ: 175.9 (C-4), 164.2 (C-7), 161.1 (C-9), 156.8 (C-
MS analysis of compound 2 in the negative ion mode 5), 147.4 (C-2), 146.6 (C-4 ), 144.8 (C-3 ), 135.7 (C-3), 122.7
revealed a pseudomolecular ion peak with an m/z value (C-1 ), 120.3 (C-6 ), 114.8 (C-5 ), 114.5 (C-2 ), 103.1 (C-10),
of 301, corresponding to [M+H]–. Elemental analysis 97.8 (C-6), 93.0 (C-8). These data were found to be
showed that the amounts of C and H in compound 2 consistent with those reported in the literature for
were 59.61 and 3.32%, respectively. Taken together with kaempferol (Ning, 2000; Fossen and Andersen, 2006;
the NMR results, these data suggested that the Markham and Geiger, 1994; Stochmal et al., 2001), and
molecular formula of compound 2 was C15H10O7. FT-IR so compound 3 was identified as kaempferol.
analysis revealed peaks at 3318 and 1250–1050 cm-1, Compound 4: Physical appearance, yellow powder;
which were consistent with the presence of hydroxyl hydrochloric acid-Mg reaction, positive; Molisch reac-
groups. The peak at 1663 cm-1 was attributed to the tion, positive. These results suggested that compound 4
stretching vibration of a carbonyl group. Several other was a flavonoid glycoside.
peaks were observed at 1611, 1561, 1522 and 1450 cm -1,
which were consistent with the presence of a phenyl NMR data: 1H-NMR (300 MHz, DMSO-d6) δ: 7.97 (2H,
ring skeleton. The peaks observed at 1611, 1522 and d, J = 9.4 Hz, H-2 ,6 ), 6.95 (2H, d, J = 8.3 Hz, H-3 ,5 ),
1408 cm-1 also suggested the presence of an oxygen- 6.88 (1H, s, H-3), 6.83 (1H, s, H-8), 6.44 (1H, s, H-6), 5.09
containing heterocycle. (2H, d, J = 6.9 Hz, H-1 ); 13C-NMR (75 MHz, DMSO-d6)
δ: 182.1 (C-4), 164.4 (C-2), 163.0 (C-7), 161.5 (C-5), 161.2
NMR data: 1H-NMR (300 MHz, DMSO-d6) δ: 12.50 (1H,
(C-4 ), 157.0 (C-9), 128.7 (C-6 ), 128.7 (C-2 ), 121.1 (C-1 ),
s, 5-OH), 10.83 (1H, s, 7-OH), 9.38 (3H, s, -OH), 7.67
116.1 (C-3 ), 116.1 (C-5 ), 105.4 (C-10), 103.2 (C-3), 100.0
(1H, d, J = 2.1 Hz, H-2 ), 7.53 (1H, dd, J = 7.2, 1.9 Hz, H-
(C-1 ), 99.6 (C-6), 94.9 (C-8), 77.3 (C-3 ), 76.4 (C-5 ), 73.2
6 ), 6.88 (1H, d, J = 8.5 Hz, H-5 ), 6.40 (1H, d, J = 1.7 Hz,
(C-2 ), 69.6 (C-4 ), 60.7 (C-6 ). These data were
H-8), 6.18 (1H, d, J = 1.7 Hz, H-6); 13C-NMR (75 MHz,
discovered to be consistent with those reported in the
MeOD) δ: 175.9 (C-4), 164.2 (C-7), 161.1 (C-9), 156.8 (C-
literature of apigenin-7-O- -D-glucopyranoside (Ning,
5), 147.4 (C-2), 146.6 (C-4 ), 144.8 (C-3 ), 135.7 (C-3), 122.7
2000; Fossen and Andersen, 2006; Markham and Geiger,
(C-1 ), 120.3 (C-6 ), 114.8 (C-5 ), 114.5 (C-2 ), 103.1 (C-10),
1994; Stochmal et al., 2001), and so compound 4 was
97.8 (C-6), 93.0 (C-8). These data were consistent with
identified as apigenin-7-O- -D-glucopyranoside.
those reported in the literature for quercetin (Rhodes
and Price, 1996; Fossen et al., 1998; Lee et al., 2008; Ly et Compound 5: Physical form, yellow powder; hydro-
al., 2005; Ning, 2000; Fossen and Andersen, 2006; chloric acid-Mg reaction, positive; Molisch reaction,
Markham and Geiger, 1994; Stochmal et al., 2001), and positive. These results suggested that compound 5
so compound 2 was identified as quercetin. could be a flavonoid glycoside.
Compound 3: Physical form, yellow powder; hydro- NMR data: 1H-NMR (300 MHz, DMSO-d6) δ: 12.65 (1H,
chloric acid-Mg reaction, positive; Molisch reaction, s, 5-OH-5), 9.0–11.0 (3H, br, -OH), 7.59 (2H, d, J = 5.9
negative. These results suggested that compound 3 was Hz, H-2 , 6 ), 6.85 (1H, d, J = 8.9 Hz, H-5 ), 6.41 (1H, s, H-
a flavonoid without any glycoside functionality. ESI-MS 8), 6.20 (1H, d, J = 1.4 Hz, H-6), 5.48 (1H, d, J = 6.9 Hz, H
analysis in the negative ion mode revealed a -1 ); 13C-NMR (75 MHz, DMSO-d6) δ:177.5 (C-4), 164.3
pseudomolecular ion peaks with anm/z value of 285, (C-7), 161.3 (C-5), 156.4 (C-2), 156.3 (C-9), 148.6 (C-4 ),
corresponding to [M–H]–. Elemental analysis showed 144.9 (C-3 ), 133.4 (C-3), 121.7 (C-6 ), 121.3 (C-1 ), 116.3
S22 Bangladesh J Pharmacol 2016; 11: S18-S25

(C-5 ), 115.3 (C-2 ), 104.1 (C-10), 100.9 (C-1 ), 98.8 (C-6), MHz, DMSO-d6) δ: 177.5 (C-4), 164.3 (C-7), 161.4 (C-5),
93.6 (C-8), 77.7 (C-3 ), 76.6 (C-5 ), 74.2(C-2 ), 70.0 (C-4 ), 156.8 (C-9), 156.8 (C-4 ), 156.6(C-2), 144.9 (C-3 ), 133.5 (C
61.1 (C-6 ). These data were found to be consistent with -3), 121.8 (C-6 ), 121.3 (C-1 ), 116.4 (C-5 ), 115.4 (C-2 ),
those reported in the literature for quercetin 3-O- -D- 104.1 (C-10), 101.3 (C-1 ), 100.9 (C-1‴), 98.9 (C-6), 93.8 (C
glucopyranoside (Park and Lee, 1996; Bonaccorsi et al., -8), 76.6 (C-3 ), 76.1 (C-5 ), 74.2 (C-2 ), 72.0 (C-4‴), 70.7
2005; Lee et al., 2008; Ning, 2000; Fossen and Andersen, (C-3 ), 70.5(C-3‴), 70.2 (C-2‴), 68.4 (C-5‴), 67.2 (C-6 ),
2006; Markham and Geiger, 1994; Stochmal et al., 2001), 17.9 (C-6 ). These data were found to be consistent with
and so compound 5 was identified as quercetin 3-O- -D those reported in the literature of rutin (Park and Lee,
-glucopyranoside. 1996; Ning, 2000; Fossen and Andersen, 2006; Markham
et al., 1994; Stochmal et al., 2001), and so compound 7
Compound 6: Physical form, yellow powder; hydro-
was identified as rutin.
chloric acid-Mg reaction, positive; Molisch reaction,
positive. These results suggested that compound 6 Isorhamnetin (17 mg), quercetin (64 mg), kaempferol
could be a flavonoid glycoside. (14 mg), apigenin-7-O- -D-glucopyranoside (3 mg),
quercetin 3-O- -D-glucopyranoside (3 mg), kaempferol-
NMR data: 1H-NMR (300 MHz, DMSO-d6) δ: 12.51 (1H,
7-O- -D-glucopyranoside (2 mg) and rutin (11 mg)
s, -OH), 10.16 (1H, s, -OH), 9.56 (1H, s, -OH), 8.09 (2H,
were obtained from 200 g of dry onion skins using the
d, J = 8.8 Hz, H-2 ,6 ), 6.95 (2H, d, J = 8.8 Hz, H-3 ,5 ),
extraction and isolation methods described above.
6.82 (1H, d, J = 1.5 Hz, 8-H), 6.44 (1H, d, J = 1.7 Hz, H-
6), 5.08 (1H, d, J = 6.3 Hz, H-1 ); 13C-NMR (75 MHz, Inhibitory effects of the flavonoids from onion skins
DMSO-d6) 176.1 (C-4), 162.8 (C-7), 160.4 (C-5), 159.4 (C- towards the proliferation of K562 cells
4 ), 155.8 (C-9), 147.6 (C-2), 136.1 (C-3), 129.7 (C-2 ), 129.7
Although a wide range of flavonoids have recently
(C-6 ), 115.5 (C-3 ), 115.5 (C-5 ), 104.8 (C-10), 100.9 (C-1 ),
reported to exhibit antitumor activities, inhibit the
100.0 (C-1 ), 98.8 (C-6), 94.5 (C-8), 77.2 (C-3 ), 76.5 (C-5 ),
proliferation of leukemia cells, and induce cell apop-
73.2(C-2 ), 69.7 (C-4 ), 60.7 (C-6 ). These data were
tosis, none of these flavonoids have been reported to be
found to be consistent with those reported in the litera-
nontoxic or even weakly toxic to normal human cells
ture for kaempferol-7-O- -D-glucopyranoside (Ning,
(Cárdenas et al., 2006; Shen et al., 2007). To evaluate
2000; Fossen and Andersen, 2006; Markham and Geiger,
the anti-proliferative activities of the flavonoids isolated
1994; Stochmal et al., 2001), and so compound 6 was
from onion skins in the current study, we investigated
identified as kaempferol-7-O- -D-glucopyranoside.
the effects of isorhamnetin, rutin, quercetin, and
Compound 7: Physical form, yellow powder; hydro- kaempferol on the viability of chronic myelogenous
chloric acid-Mg reaction, positive; Molisch reaction, leukemia K562 cells.
positive. These results therefore suggested that
As shown in Figure 2, quercetin and kaempferol led to
compound 7 could be a flavonoid glycoside. ESI-MS
a significant reduction in the proliferation of the K562
analysis of compound 7 in the negative ion mode
cells. Furthermore, the anti-proliferative activity of
revealed a pseudomolecular ion peak with anm/z value
quercetin was stronger than that of kaempferol when
of 609, corresponding to [M–H]–. Elemental analysis
they were administered at the same concentration.
showed that the amounts of C and H in compound 7
Interestingly, however, isorhamnetin and rutin led to
were 53.22 and 4.86%, respectively. Taken together with
an increase in viability of the K562 cells.
the NMR results, these data suggested that the
molecular formula of compound 7 was C27H30O16. The
FT-IR spectrum of compound 7 contained a broad peak
at 3423 cm-1 and a second peak at 1250–1050 cm-1, which Discussion
indicated the presence of hydroxyl groups. A peak was
also observed at 1656 cm-1, which was attributed to the In this study, seven flavonoids were isolated from dry
stretching vibration of a carbonyl group. Several other onion skins and evaluated in terms of their anti-
vibrational absorption peaks were observed at 1601, proliferative activity towards K562 cells. Although
1574, 1505 and 1456 cm-1, together with a shoulder peak isorhamnetin and rutin exhibited certain proliferation-
at 3000 cm-1, which were consistent with an entitative promoting effects, quercetin and kaempferol led to a
phenyl ring skeleton. The peaks at 1601 and 1505 cm-1 significant decrease in the viability of K562 cells, with
were indicative of the presence of an oxygen-containing quercetin exhibiting the stronger effect of the two
heterocycle. The peak at 1363 cm-1 was attributed to the compounds. In terms of the structural features of these
stretching vibration of a methyl group. flavonols, the presence of an ortho hydroxyl group on
the B ring or hydroxyl and carbonyl groups at the 3-
NMR data: 1H-NMR (300 MHz, DMSO-d6) δ: 12.61 (1H, and 4-positions, respectively, were critical to the anti-
s, 5-OH), 8.5–11.5 (1H, br, -OH), 7.54 (2H, d, J = 7.5 Hz, proliferative activity towards K562 cells.
H-2 ,6 ), 6.84 (1H, d, J = 8.4 Hz, H-5 ), 6.38 (1H, s, H-8),
6.19 (1H, s, H-6), 5.34 (2H, d, J = 7.1 Hz, H-1 ), 4.38 (1H, As mentioned above, the mono- and diglucosides of
s, H-1‴), 0.99 (3H, d, J = 6.1 Hz, H-6‴); 13C-NMR (75 quercetin account for up to 80% of the total flavonol
 Bangladesh J Pharmacol 2016; 11: S18-S25 S23

content of onion skins (Rhodes and Price, 1996). Fossen minor flavonoid components. Furthermore, only trace
et al. (1998) reported the isolation quercetin, quercetin amounts of free quercetin and isorhamnetin 3-glucoside
3,7,4 -triglucopyranoside, quercetin 4 -O-b- were detected during this particular study. Although
glucopyranoside and quercetin 3,4 -O-b-digluco- no quercetin diglucosides or quercetin triglu-cosides
pyranoside from the pigmented scales of red onion. were isolatedfrom the onion skins used in the current
Seven flavonols were identified by high-performance study, a large amount of free quercetin was obtained, as
liquid chromatography (HPLC) diode array detector well as a minor amount of quercetin-3-O- -D-
coupled with ESI-MS from southern Italian red onions glucopyranoside. Suh et al. (1999) reported similar
(Bonaccorsi et al., 2005). In this case, quercetin-4 - results to those of the current study, when they
glucoside and quercetin-3,4 -diglucoside were found to identified quercetin and quercetin 4 -glucoside from
be the most abundant components, whereas quercetin-3 onion skins by fast atom bombardment. Suh et al. (1999)
-glucoside, quercetin-7,4 -digluco-side, quercetin-3,7,4 - also reported the isolation of substantial amounts of
triglucoside, isorhamnetin 4 -gluco-side and isorhamnetin, kaempferol and rutin from onion skins,
isorhamnetin 3,4 -diglucoside were identified as the but only managed to isolate a trace amount of

A 120
Quercetin

100 Kaempferol
a
a
Cell viability (% of control)

80 a a
b
60
b
b

40 b
b
b
20

0
0 20 40 60 80 100
Concentration (µM)

Isorhamnetin

Rutin
B 120

100
Cell viability (% of control)

80

60

40

20

0
0 20 40 60 80 100
Concentration (µM)

Figure 2: Cytotoxicity of quercetin, kaempferol, isorhamnetin, and rutin on K562 cells

K562 cells were treated with quercetin, kaempferol, isorhamnetin, and rutin at various concentrations for 72 hours, and the cell viability was then
determined using an MTT assay. Data are shown as the mean ± SD (n = 6). ap<0.05, bp<0.01 compared with the control
S24 Bangladesh J Pharmacol 2016; 11: S18-S25

kaempferol-7-O- -D-glucopyranoside. Fur-thermore, Conclusion

no diglucosides or triglucosides of any other flavonols
were isolated from onion skins during this study. Taken Onion is a suitable source of flavonoids, its quercetin
together, these results suggest that dry onion skins are and kaempferol have good anti-proliferative activities
an abundant source of free flavonols, as well as on leukemia K562 cell line.
containing small amounts of flavonol mono-glucosides,
and tracequantities of flavonol di- and triglucosides.
Rutin was separated from red Spirit onions by References
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Author Info
Wen-En Zhao (Principal contact)
e-mail: [email protected]
Yan-Qi Liu (Principal contact)
e-mail: [email protected]
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