Flavonol glycosides from Cassia hirsuta

Article abstract of DOI:10.1021/np980195c

A new flavonol glycoside, kaempferol 3-O-α-L-rhamnopyranosyl (1→2)- α-L-rhamnopyranoside (1), was isolated from the flowers of Cassia hirsuta along with two known flavonol glycosides, kaempferol 3-O-rutinoside and rutin. The structure of compound 1 has been established on the basis of spectral data and by acid hydrolysis.

Full text of DOI:10.1021/np980195c

J . Nat. Prod. 1999, 62, 305-306
305
F la von ol Glycosid es fr om Ca ssia h ir su ta
K. V. Rao,† A. G. Damu, B. J ayaprakasam, and D. Gunasekar*
Natural Products Division, Department of Chemistry, Sri Venkateswara University, Tirupati - 517 502, India
Received May 12, 1998
A new flavonol glycoside, kaempferol 3-O-R-L-rhamnopyranosyl (1f2)-R-L-rhamnopyranoside (1), was
isolated from the flowers of Cassia hirsuta along with two known flavonol glycosides, kaempferol 3-O-
rutinoside and rutin. The structure of compound 1 has been established on the basis of spectral data and
by acid hydrolysis.
Cassia hirsuta L. (Leguminosae) is a diffuse shrub widely
distributed in the hilly tracts of Rayalaseema region of
South India.1 Previous phytochemical study on the seeds
of this plant revealed the presence of an anthraquinone.2
In the present paper, we describe the isolation and char-
acterization of a new flavonol glycoside (1), together with
kaempferol 3-O-rutinoside and rutin from the ethanol
extract of the flowers of C. hirsuta.
downfield (5.7 ppm) when compared with the chemical shift
(71.2 ppm) of the corresponding carbon atom (C-2′′′) of the
terminal rhamnose.4,6 The configuration of two anomeric
centers of the rhamnopyranosyl moieties in 1 was deter-
mined to be R from the presence of anomeric carbon signals
at δ 101.5 and 104.5 in its 13C NMR spectrum and also
from small coupling constants (J ) 1.5 Hz)7,8 for the
anomeric proton signals of the two rhamnosyl moieties in
its 1H NMR spectrum. Thus, the structure of 1 was
elucidated as kaempferol 3-O-R-L-rhamnopyranosyl(1f2)-
R-L-rhamnopyranoside.
Kaempferol 3-O-rutinoside and rutin were identified by
comparison of their spectral data (UV, IR, 1H NMR, 13C
NMR, MS) with the published results.9-11
Exp er im en ta l Section
Compound 1 was obtained as yellow needles and ap-
peared to be a kaempferol derivative with a substituted
3-hydroxyl group from its UV spectral analysis with
diagnostic reagents.3 Acid hydrolysis of 1 gave kaempferol
and L-rhamnose, confirmed by co-chromatography with
authentic samples. Comparison of the 13C NMR spectral
data of 1 with its aglycon, kaempferol, showed an upfield
shift of 1.7 ppm for C-3 signal and downfield shifts of 11.2
and 3.2 ppm for the C-2 and C-4 signals, respectively,
suggesting that the site of glycosylation was at position
C-3.4 The negative FABMS exhibited a quasimolecular ion
at m/z 577 [M - H]-, which is in accord with the molecular
Gen er a l Exp er im en ta l P r oced u r es. Melting points were
uncorrected. UV spectra were taken in MeOH on a Beckman
25 spectrophotometer. IR spectra were run in KBr. Optical
rotation was determined on a Perkin-Elmer model 241 pola-
rimeter. All NMR experiments were performed on a Nicolet
NT 300 WB or J EOL-FX-90Q spectrometer equipped with 5
mm 1H and 13C probes operating at 300.06 and 75.45 MHz or
90 and 22.5 MHz, respectively. Samples were run in acetone-
d6 or DMSO-d6 with TMS as internal standard. Mass spectra
were obtained in the EI mode at 70 eV. FABMS was obtained
on VG Micro Mass ZAB-HF mass spectrometer in the negative-
ion mode with glycerol as the matrix.
P la n t Ma ter ia l. C. hirsuta flowers were collected in J uly
1993 from the Horsely Hills, Madanapalle, Andhra Pradesh,
South India. A voucher specimen, DR 585, has been deposited
in the Herbarium of the Department of Botany, Sri Ven-
kateswara University, Tirupati.
formula C27H30O14 and was further corroborated by the 13
C
NMR spectrum (27 carbon atoms).
1
The H NMR spectrum of 1 showed the expected signals
of the B ring protons at δ 7.80 (2H, d, J ) 8.5 Hz) and 6.95
(2H, d, J ) 8.5 Hz) for H-2′, H-6′ and H-3′, H-5′, respec-
tively. Two meta-coupled doublets (J ) 2.5 Hz) at δ 6.42
and 6.80 were attributed to the C-6 and C-8 protons. A pair
of one-proton doublets (J ) 1.5 Hz) centered at δ 5.60 and
5.28 was assigned to two rhamnosyl anomeric protons.5 The
presence of a pair of three-proton doublets at δ 0.85 (J )
5.8 Hz) and 1.18 (J ) 6.0 Hz) indicated the presence of
two rhamnosyl methyl groups.
Extr a ction a n d Isola tion . The air-dried and powdered
flowers (500 g) of C. hirsuta were defatted with hexane and
extracted with 90% EtOH (4 × 2 L) under reflux. The combined
extracts were concentrated, and the residue (15 g) obtained
was dissolved in H2O (300 mL) and extracted with EtOAc (3
× 50 mL). The EtOAc-soluble part was subjected to column
chromatography over silica gel (E.Merck, 60-120 mesh) and
eluted with CHCl3, CHCl3-EtOAc, and EtOAc-MeOH. EtOAc
and EtOAc-MeOH (9:1) fractions on concentration gave a
yellow solid (0.16 g) that on crystallization from MeOH
furnished yellow needles (130 mg) of 1. Concentration of the
EtOAc-MeOH (1:1) fractions afforded a yellow residue (0.21
g), which on purification by preparative paper chromatography
employing n-BuOH-HOAc (40-10%)-H2O (4:1:5) gave
kaempferol 3-O-rutinoside (40 mg) and rutin (12 mg).
Com p ou n d 1: yellow needles (MeOH); mp 195-196 °C;
[R]D -105.2° (c 0.7, MeOH); UV (MeOH) λmax (log ꢀ) 264 (4.32),
320 sh (3.92), 343 (4.19) nm; +AlCl3 280, 400 nm; +AlCl3/HCl
275, 395 nm; +NaOAc 274, 385 nm; +NaOAc/H3BO3 264, 340
Acid hydrolysis of the permethylated glycoside 1 yielded
an aglycon, characterized as kaempferol 5,7,4′-tri-O-methyl
ether and two sugars identified as 3,4-di-O-methyl-L-
rhamnose and 2,3,4-tri-O-methyl-L-rhamnose. These re-
sults suggested that 1 is a kaempferol 3-O-dirhamnoside
with a 1f2 interglycosidic linkage, further evidenced by
the fact that the C-2′′ signal at 76.9 ppm was shifted
* To whom correspondence should be addressed. Tel: 0091-8574-22471.
Fax: 0091-8574-27499/25211. E-mail: pcjobs@mail.com.
† Present address: Biotechnology Division, Dr. Reddy’s Research Foun-
dation, Bollaram Road, Miyapur, Hyderabad 500 050, India.
nm; +NaOMe 278, 425 nm; IR (KBr) ν
3385, 2935, 1650,
max
1599, 1455, 1373, 1180, 1136, 1100, 1059, 1020, 974, 934 cm-1
;
10.1021/np980195c CCC: $18.00
© 1999 American Chemical Society and American Society of Pharmacognosy
Published on Web 01/28/1999

306 J ournal of Natural Products, 1999, Vol. 62, No. 2
Notes
1H NMR δ 12.30 (1H, brs, HO-5), 7.80 (2H, d, J ) 8.5 Hz, H-2′,
H-6′), 6.95 (2H, d, J ) 8.5 Hz, H-3′, H-5′), 6.80 (1H, d, J ) 2.5
Hz, H-8), 6.42 (1H, d, J ) 2.5 Hz, H-6), 5.60 (1H, d, J ) 1.5
Hz, rhamnosyl H-1′′), 5.28 (1H, d, J ) 1.5 Hz, rhamnosyl
H-1′′′), 3.18-4.00 (8H, m, rhamnosyl protons), 1.18 (3H, d, J
) 6.0 Hz, rhamnosyl CH3), 0.85 (3H, d, J ) 5.8 Hz, rhamnosyl
CH3); 13C NMR δ 179.1 (C-4), 164.7 (C-7), 161.2 (C-5),159.8
(C-4′), 158.0 (C-2), 157.3 (C-9), 133.9 (C-3), 131.4 (C2′ and C-6′),
122.3 (C-1′), 115.9 (C-3′ and C-5′), 104.7 (C-10), 104.5 (C-1′′′),
101.5 (C-1′′), 98.4 (C-6), 94.2 (C-8), 76.9 (C-2′′), 74.1 (C-4′′),
73.7 (C-4′′′), 72.7 (C-5′′), 72.3 (C-3′′′), 72.2 (C-3′′), 71.2 (C-2′′′),
71.0 (C-5′′′), 18.0 (C-6′′), 17.6 (C-6′′′); FABMS (negative ion)
600 [M + Na - H]-, 577 [M - H]- (100), 431 (M - rhamnosyl),
285 (M - rhamnosyl - rhamnose).
Acid Hyd r olysis of th e P er m eth yl Eth er of 1. The
permethylated derivative of 1 (20 mg) on acid hydrolysis with
7% aqueous alcoholic H2SO4 (6 mL) at 100 °C for 3 h and the
usual workup gave long pale-yellow needles (9 mg) of kaempfer-
ol 5,7,4′-trimethyl ether, mp 134 °C, and a mixture of two
sugars identified as 3,4-di-O-methyl-L-rhamnose (Rf 1.02) and
2,3,4-tri-O-methyl-L-rhamnose (Rf 0.84) by co-paper chroma-
tography using 2,3,4,6-tetra-O-methyl-D-glucose as reference
sugar.13
Ack n ow led gm en t. The authors thank Dr. A. V. Rama
Rao, Former Director of the Indian Institute of Chemical
Technology, Hyderabad, India, for providing the spectral data
and Dr. D. Rangachari, Department of Botany, Sri Ven-
kateswara University, Tirupati, for collection and identification
of plant material.
Acid Hyd r olysis of 1. An EtOH solution of 1 (10 mg) in
7% H2SO4 (3 mL) heated at 100 °C for 2 h gave kaempferol
(spectral data compared with pure compound) and L-rhamnose
(co-paper chromatography with an authentic sample).
Refer en ces a n d Notes
(1) Gamble, J . S. Flora of the Presidency of Madras; Botanical Survey of
India: Calcutta, 1956; Vol. 1, pp 282-284.
(2) Singh, J .; Singh, J . Phytochemistry 1986, 25, 1985-1987.
(3) Mabry, T. J .; Markham, K. R.; Thomas, M. B. The Systematic
Identification of Flavonoids; Springer-Verlag: New York, 1970; p 44.
(4) Markham, K. R.; Turner, B.; Stanley, R.; Geiger, H.; Mabry, T. J .
Tetrahedron 1978, 34, 1389-1397.
(5) Waterman, P. G.; Khalid, S. A. Phytochemistry 1980, 19, 909-915.
(6) Mizuno, M.; Iinuma, M.; Tanaka, T.; Sakakivara, N.; Fuzikawa, T.;
Hanioka, S.; Ishida, Y.; Lu, X. S.; Murata, H. Phytochemistry 1988,
27, 3645-3647.
P a r tia l Acid Hyd r olysis of 1. Compound 1 (20 mg) was
dissolved in 5 mL of 0.5% H2SO4 at room temperature, kept
aside for 5 days, and then extracted with EtOAc. The EtOAc
extract was evaporated to yield a yellow solid, which on
crystallization from MeOH gave yellow needles (12 mg), mp
172-174 °C. It was characterized as kaempferol 3-O-R-L-
rhamnoside by UV, 1H NMR, 13C NMR, and mass spectral
studies.12 The aqueous layer on usual workup gave a sugar,
identified as rhamnose by co-paper chromatography with an
authentic sample.
P er m eth yla tion of 1. Compound 1 (50 mg) was treated
with a mixture of MeI (3 mL) and NaH (6 mL) in DMF (10
mL) under anhydrous conditions in an inert atmosphere of
nitrogen. After the mixture was kept in the dark for 1 h at 32
°C, the workup was accomplished by partition between CHCl3
and H2O. The permethylated derivative was purified on TLC
silica gel using EtOAc (Rf 0.14, 22 mg): EIMS m/z 690 [M]+
(1), 485 (4), 453 (2), 384 (16), 351 (10), 328 (100), 189 (15), 188
(21), 157 (11), 142 (10), 135 (12), 101 (12), 88 (54), 71 (25), 59
(19).
(7) Mabry, T. J .; Markham, K. R.; Thomas, M. B. The Systematic
Identification of Flavonoids; Springer-Verlag: New York, 1970; p 269.
(8) Ito, Y.; Hirayama, F.; Suto, K.; Sagara, K.; Yoshida, T. Phytochemistry
1988, 27, 911-913.
(9) Adinarayana, D.; Gunasekar, D.; Ramachandriah, P.; Seligmann, O.;
Wagner, H. Phytochemistry 1980, 19, 478-480.
(10) Adinarayana, D.; Ramachandraiah, P.; Syamsundar, K. V. Leather
Sci. 1980, 17, 268-270.
(11) Chourasia, N.; Wichtl, M. Planta Med. 1987, 53, 432-434.
(12) Sarada, M.; Adinarayana, D. J . Indian Chem. Soc. 1984, 7, 649-
650.
(13) Lederer, E.; Lederer, M. Chromatography; Elsevier: Amsterdam,
1957; p 251.
NP980195C