Flavonoids of carthamus tinctorius flowers

Article abstract of DOI:10.1007/s10600-014-0983-z

The known flavonoids luteolin, cinaroside, 5-O-methylluteolin, azaleatin (3,7,3′,4′-tetrahydroxy-5-methoxyflavone), and the new natural product 3,7,3′,4′-tetrahydroxy-5-methoxyflavone 7-O-β- Dglucopyranoside (safloroside) were isolated from Carthamus tinctorius flowers and characterized by PMR and UV spectroscopy and mass spectrometry.

Full text of DOI:10.1007/s10600-014-0983-z

Chemistry of Natural Compounds, Vol. 50, No. 3, July, 2014 [Russian original No. 3, May–June, 2014]
FLAVONOIDS OF Carthamus tinctorius FLOWERS
*
V. A. Kurkin and A. V. Kharisova
The known flavonoids luteolin, cinaroside, 5-O-methylluteolin, azaleatin (3,7,3ꢀ,4ꢀ-tetrahydroxy-5-
methoxyflavone), and the new natural product 3,7,3ꢀ,4ꢀ-tetrahydroxy-5-methoxyflavone 7-O-ꢁ-D-
glucopyranoside (safloroside) were isolated from Carthamus tinctorius flowers and characterized by PMR
and UV spectroscopy and mass spectrometry.
Keywords: dyer’s saffron, Carthamus tinctorius, flowers, flavonoids, luteolin, cinaroside, 5-O-methylluteolin, azaleatin
(
3,7,3ꢀ,4ꢀ-tetrahydroxy-5-methoxyflavone), safloroside (azaleatin 7-O-ꢁ-D-glucopyranoside).
Dyer’s saffron (Carthamus tinctorius L., Asteraceae) is a known oil crop, the seeds of which contain up to 40% fatty
oil that is used as a component in the production of drugs, biologically active additives (BAAs), and cosmetics [1, 2]. We
studied earlier the fatty-acid composition of fatty oil from seeds of C. tinctorius [3]. However, the flowers of this plant, which
have antioxidant, hepatoprotective, anti-inflammatory, neurotropic, and other properties, are also interesting as a source of
drugs [4]. In our opinion, the flavonoids are of interest with respect to the manifestation of antioxidant and hepatoprotective
activity by C. tinctorius flowers. According to the literature [4–7], flowers of this plant contain acacetin, luteolin, quercetin,
acacetin-7-O-ꢁ-D-glucuronide, acacetin-6-C-ꢁ-D-glucuronyl-8-C-ꢁ-D-glucuronide, luteolin-7-O-ꢁ-D-glucopyranoside,
luteolin-7-O-ꢁ-D-glucuronide, luteolin-7-O-(6ꢀꢀ-O-acetyl)-ꢁ-D-glucuronide, kaempferol-3-O-rutinoside, glycosides of
6
-hydroxykaempferol, quercetin-3-O-ꢁ-D-glucopyranoside, quercetin-7-O-ꢁ-D-glucuronide, quercetin-7-O-(6ꢀꢀ-O-acetyl)-ꢁ-
D-glucuronide, and rutin.
Herein we present results from a study of the flavonoid composition of C. tinctorius flowers that were cultivated in
Samara Region.
We studied flowers of C. tinctorius that was cultivated at Tulaikov Samara State Agricultural Research Institute,
Russian Academy of Agricultural Sciences (Bezenchuk, Samara Region). Chromatographic separation of the evaporated
aqueous EtOH extract of C. tinctorius flowers and subsequent rechromatography of the obtained fractions isolated five (1–5)
flavonoid compounds.
OH
OH
RO
O
O
OH
O
HO
OH
1: R =
HO
H
3
CO
OH
1
, 2
2: R = H
The PMR spectrum of 1 exhibited resonances for aromatic protons H-2ꢀ at 7.78 ppm (1H, d, J = 2.5 Hz), H-6ꢀ at 7.75
1H, dd, J = 2.5, 9 Hz), H-5ꢀ at 6.95 (1H, d, J = 9 Hz), H-8 at 6.86 (1H, d, J = 2.5 Hz), and H-6 at 6.43 ppm (1H, d, J = 2.5 Hz).
(
The observation of a 3H singlet at 3.83 ppm indicated that 1 contained a methoxy that was assigned to ring A based on mass
+
spectral data. A peak for an ion with m/z 168 corresponded to an [A + H] fragment. This conclusion was confirmed by
UV spectral data. The 5-OH was identified as the methylation site because the resonance of this group was missing in
PMR spectra of 1 and aglycon 2 that was prepared by acid hydrolysis and also isolated pure from C. tinctorius flowers. We
identified 2 as 3,7,3ꢀ,4ꢀ-tetrahydroxy-5-methoxyflavone (azaleatin) [8]. The ꢁ-configuration of the glycoside bond in 1 was
confirmed by a doublet at 5.05 ppm with SSCC 7 Hz in the PMR spectrum that belonged to the glucose anomeric proton.
Samara State Medical University, 443099, Samara, Ul. Chapaevskaya, 89, fax: 8 (846) 333 29 76, e-mail:
Kurkinvladimir@yandex.ru. Translated from Khimiya Prirodnykh Soedinenii, No. 3, May–June, 2014, pp. 386–388. Original
article submitted January 6, 2014.
446
0009-3130/14/5003-0446 ©2014 Springer Science+Business Media New York

UV spectral data indicated that the 7-OH was glycosylated. Thus, a bathochromic shift of the short-wavelength band was not
observed in the electronic spectrum in the presence of NaOAc [9] whereas this phenomenon did appear in aglycon 2.
The PMR spectrum of 3, in contrast with that of 2, contained at 6.04 ppm an additional 1H singlet for the C-3 proton.
This together with UV and mass spectral data, which showed peaks for ions with m/z 300 and 168 that corresponded to the
+
molecular ion and an [A + H] fragment, allowed 3 to be identified as 5-O-methylluteolin [10].
Compounds 4 and 5 were identified as 5,7,3ꢀ,4ꢀ-tetrahydroxyflavone (luteolin) and 5,7,3ꢀ,4ꢀ-tetrahydroxyflavone
7
-O-ꢁ-D-glucopyranoside (cinaroside) [11], which were isolated previously from C. tinctorius flowers [4].
Thus, five flavonoids were isolated from C. tinctorius flowers and characterized by PMR and UV spectroscopy and
mass spectrometry. Among these, azaleatin (3,7,3ꢀ,4ꢀ-tetrahydroxy-5-methoxyflavone) and 5-O-methylluteolin were described
for the first time from this plant and azaleatin 7-O-ꢁ-D-glucopyranoside, which we called safloroside, was a new natural
product.
EXPERIMENTAL
PMR spectra were taken on Bruker AM 300 (300 MHz) instruments. Mass spectra were recorded on a Kratos MS-30
mass spectrometer. UV spectra were recorded using a Specord 40 spectrophotometer (Analytik Jena).
Extraction and Isolation. Air-dried C. tinctorius flowers (100 g) that were collected in August 2013 were extracted
with EtOH (70%) first twice at room temperature for 24 h and then with heating on a boiling-water bath for 30 min. The
combined aqueous EtOH extract was evaporated in vacuo to 50 mL, mixed with L 40/100 silica gel (30 g), and dried. The
dried powder (dry extract + silica gel) was placed on a layer of silica gel (8 cm diameter, 5 cm high) formed from a suspension
in CHCl . The chromatography column was eluted by CHCl and CHCl –EtOH (99:1, 98:2, 97:3, 95:5, 93:7, 90:10, 85:15,
3
3
3
8
0:20, 70:30, 60:40, and 50:50). The separation of the compounds was monitored by TLC on PTSKh-AF-A-UF Sorbfil plates
using CHCl –EtOH (9:1), CHCl –EtOH–H O (26:16:3), and n-BuOH–AcOH(glacial)–H O (4:1:2).
3
3
2
2
Fractions containing dominant 1 were combined. The resulting precipitate was separated and recrystallized from
EtOH to afford 1 in 0.1 mass% yield (of air-dried raw material). Fractions containing 2–5 were placed on Woelm polyamide
for further purification. Dry powder (extract + polyamide) was placed onto a chromatography column (5 cm high, 4 cm
diameter) and eluted with H O and aqueous EtOH (20%, 40, 70, 96) to afford 2 and 3 (eluent 96% EtOH), 4 (eluent 70%
2
EtOH), and 5 (eluent 40% EtOH), which were additionally purified by recrystallization from aqueous EtOH.
Acid hydrolysis of 1 used HCl (2%) at 100°C for 30 min. Cleavage of glycoside 5 used more forcing conditions
(
HCl, 10%, 100°C, 2 h). Precipitates obtained after cooling the reaction mixture were rinsed with purified H O and recrystallized
2
from aqueous EtOH. Glycosides 1 and 5 afforded aglycons 2 and 4.
3
,7,3ꢀ,4ꢀ-Tetrahydroxy-5-methoxyflavone 7-O-ꢁ-D-glucopyranoside (1), yellow crystalline compound, C H O ,
2
2 22 12
+
mp 230–233ꢂÑ (aqueous EtOH). Mass spectrum (70 eV, 200ꢂÑ, m/z, %): 316 (Ì of the aglycon, 100%), 168 (2), 153 (12),
1
4
37 (8). UV spectrum (ꢃ , nm): 260, 274 sh, 371; + NaOAc 260, 274 sh, 371; + NaOAc + H BO 260, 378; +À1Ñ1 274,
max 3 3 3
1
32. Í NMR spectrum (300 MHz, DMSO-d , ꢄ, ppm, J/Hz): 7.78 (d, J = 2.5, Í-2ꢀ), 7.75 (1Í, dd, J = 2.5, 9, Í-6ꢀ), 6.95 (1Í,
6
d, J = 9, Í-5ꢀ), 6.86 (1Í, d, J = 2.5, Í-8), 6.43 (1Í, d, J = 2.5, Í-6), 5.05 (d, J = 7, Í-1ꢀꢀ Glc), 3.83 (3Í, s, ÑÍ Î), 3.1–3.8 (6Í,
3
m, Glc).
3
,7,3ꢀ,4ꢀ-Tetrahydroxy-5-methoxyflavone (2), yellow crystalline compound, C H O , mp 282–285ꢂÑ (aqueous
16 12 7
+
EtOH). Mass spectrum (70 eV, 200ꢂÑ, m/z, %): 316 (Ì , 100 %), 168 (8), 167 (25), 137 (34). UV spectrum (ꢃmax, nm): 260,
74 sh., 371; + NaOAc 265, 277 sh, 372; + NaOAc + H BO 260, 378; +À1Ñ1 274, 432. Í NMR spectrum (300 MHz,
1
2
3
3
3
DMSO-d , ꢄ, ppm, J/Hz): 7.78 (d, J = 2.5, Í-2ꢀ), 7.75 (1Í, dd, J = 2.5, 9, Í-6ꢀ), 6.95 (1Í, d, J = 9, Í-5ꢀ), 6.86 (1Í, d, J = 2.5,
6
Í-8), 6.43 (1Í, d, J = 2.5, Í-6), 3.82 (3Í, s, ÑÍ Î).
3
7
,3ꢀ,4ꢀ-Trihydroxy-5-methoxyflavone (3), yellow amorphous compound, C H O . UV spectrum (ꢃ , nm):
56, 266 sh, 350; +NàÎÀñ 260, 268 sh, 390; +AlCl , 278, 330, 355, 400. Mass spectrum (70 eV, 200ꢂÑ, m/z, %): 300 (Ì ,
00%), 168 (4), 167 (7), 153 (14), 137 (23). Í NMR spectrum (300 MHz, DMSO-d , ꢄ, ppm, J/Hz): 7.60 (d, J = 2, Í-2ꢀ), 7.55
16 12 6 max
+
2
1
3
1
6
(
dd, J = 9, 2, Í-6ꢀ), 7.14 (d, J = 9, Í-5ꢀ), 6.82 (s, Í-3), 6.80 (d, J = 2, Í-8), 6.70 (d, J = 2, Í-6), 3.83 (3Í, s, ÑÍ Î).
3
5
,7,3ꢀ,4ꢀ-Tetrahydroxyflavone (4), yellow crystals, C H O , mp 227–230ꢂÑ (aqueous EtOH). UV spectrum (EtOH,
15 10 6
ꢃmax, nm): 256, 266 sh, 358; +NàÎÀñ 259, 268 sh, 390; +AlCl 278, 330, 355, 404. Mass spectrum (70 eV, 200ꢂÑ, m/z, %):
3
+
1
2
86 (Ì , 100%), 153 (17), 137 (30). Í NMR spectrum (300 MHz, DMSO-d , ꢄ, ppm, J/Hz): 12.60 (1H, s, 5-ÎÍ), 7.40 (dd,
6
J = 9, 2, Í-6ꢀ), 7.37 (d, J = 2, Í-2ꢀ), 6.88 (d, J = 9, Í-5ꢀ), 6.65 (s, Í-3), 6.42 (d, J = 2, Í-8), 6.18 (d, J = 2, Í-6).
,7,3ꢀ,4ꢀ-Tetrahydroxyflavone 7-O-ꢁ-D-glucopyranoside (5), light-yellow crystals, C H O , mp 232–234ꢂÑ
5
2
1 20 11
(
aqueous EtOH). UV spectrum (EtOH, ꢃmax, nm): 257, 266 sh, 352; +NàÎÀñ 258, 268 sh, 380; +AlCl 276, 330, 350, 394.
3
447

+
1
Mass spectrum (70 eV, 200ꢂÑ, m/z, %): 286 (Ì of the aglycon, 100%), 153 (52), 137 (46). Í NMR spectrum (300 MHz,
DMSO-d , ꢄ, ppm, J/Hz): 12.98 (1H, s, 5-ÎÍ), 7.45 (dd, J = 9, 2, Í-6ꢀ), 7.41 (d, J = 2, Í-2ꢀ), 6.91 (d, J = 9, Í-5ꢀ), 6.78 (d,
6
J = 2, Í-8), 6.73 (d, J = 2, Í-6), 6.42 (s, Í-3), 5.03 (d, J = 7.2, Í-1ꢀ), 3.1–3.9 (6Í, m, Glc).
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