From the fruits of Silybum marianum (L.) Gaertn. cultivated in the North of Vietnam, (±)2,6-bis(4′-hydroxy-3′-methoxy-phenyl)-3,7-dioxabicyclo[3.3.0]octane (pinoresinol) (1) and 3,4′,5,7-tetrahydroxy-3′-methoxyflavanone (2) were isolated. Their structures were elucidated by analyses of the NMR and mass spectra in comparison with the reported data. This is the first report of both 1 and 2 from Silybum species

Bang, Bui Thi

Braca, Alessandra

Diep, Trinh Thi

Dong, Nguyen Thuong

Kiem, Phan Van

Minh, Chau Van

Tung, Nguyen Huu

Vietnam Academy of Science and Technology: Journals Online

219
Journal of Chemistry, Vol. 45 (2), P. 219 - 222, 2007 
PINORESINOL AND 3,4’,5,7-TETRAHYDROXY-3’-
METHOXYFLAVANONE FROM THE FRUITS OF  
SILYBUM MARIANUM (L.) GAERTN. 
Received 19 April 2006 
TRINH THI DIEP1, PHAN VAN KIEM2, NGUYEN THUONG DONG1, NGUYEN HUU TUNG2,
BUI THI BANG1, CHAU VAN MINH2 AND ALESSANDRA BRACA3
1National Institute of Medicinal Materials, Ministry of Health 
2Institute of Natural Products Chemistry, VAST 
3Department of Bioorganic Chemistry and Biopharmaceutics University of Pisa, Italia 
 
SUMMARY 
From the fruits of Silybum marianum (L.) Gaertn. cultivated in the North of Vietnam, (±)2,6-
bis(4-hydroxy-3-methoxy-phenyl)-3,7-dioxabicyclo[3.3.0]octane (pinoresinol) (1) and 3,4,5,7-
tetrahydroxy-3-methoxyflavanone (2) were isolated. Their structures were elucidated by analyses 
of the NMR and mass spectra in comparison with the reported data. This is the first report of both 
1 and 2 from Silybum species.  
 
I - INTRODUCTION 
In previous papers, we have reported the 
isolation and structural elucidation of 
flavonolignan and flavonoid compositions of the 
fruits of Silybum marianum (L.) Gaertn. 
(Asteraceae) cultivated the North of Vietnam 
and used in the folk medicines [1 - 3]. This 
paper deals with further investigation of the 
bioactive compositions from S. marianum. By 
various chromatography methods, (±)2,6-bis(4-
hydroxy-3-methoxy-phenyl)-3,7-
dioxabicyclo[3.3.0]octane (Pinoresinol) (1) and 
3,4,5,7-tetrahydroxy-3-methoxyflavanone (2)
were isolated from the methanolic extract of the 
fruits from this plant. The structures were 
elucidated by analyses of the NMR and ESI 
mass spectra in comparison with the reported 
data. This is the first report of both 1 and 2 from 
Silybum species. To the best of our knowledge, 
no 13C-NMR data of 2 has been reported 
previously. 
II - EXPERIMENTAL 
1. General experimental procedures 
The Electronspray Ionization (ESI) mass 
spectrum was obtained using a AGILENT 1100 
LC-MSD Trap spectrometer. The 1H-NMR (500 
MHz) and 13C-NMR (125 MHz) spectra were 
recorded on a Bruker AM500 FT-NMR 
spectrometer using TMS as the internal 
standard. Column chromatography (CC) was 
performed on silica gel (Kieselgel 60, 70-230 
mesh and 230-400 mesh, Merck) or YMC RP-
18 resins (30-50 µm, Fujisilisa Chemical Ltd). 
Thin layer chromatography (TLC) was 
performed on DC-Alufolien 60 F254 (Merck 
1.05715) or RP18 F254s (Merck) plates. 
2. Plant material  
The fruits of Silybum marianum (L.) Gaertn. 
were collected in Sapa, Lao Cai Province in 
June, 2004 and were identified by Dr Nguyen 
Tap, Department of Botany and Medicinal Plant 
220
Resources, National Institute of Medicinal 
Materials. 
3. Extraction and isolation  
Air-dried and powdered fruits of Silybum 
marianum (L.) Gaertn. (200 g) were defatted 
with petroleum and then extracted with 
methanol. After removal of the solvent under 
vacuum, the residue (5.8 g) was obtained, which 
was then repeated chromatographed on silica gel 
columns to get 1 (5.2 mg) as colorless 
amorphous and 2 (6.5 mg) as yellow crystals. 
III - RESULTS AND DISSCUSSION 
Compound 1 was isolated as colorless 
amorphous substance, mp. 120 - 121oC. The 1H-
NMR spectrum showed three peaks of the 
aromatic protons at  6.89 (1H, d, J = 1.5 Hz), 
6.82 (1H, dd, J = 8.0, 1.5 Hz) and 6.88 (1H, d, J
= 8.0 Hz) suggesting that the aromatic ring has 
three substituted positions. Oxymethine proton 
was evident at  4.73 ppm as a doublet and the 
methylene groups bearing oxygen atom at 
4.24 and 3.88 ppm as two doublet of doublet. A 
methine proton was observed at  3.10 ppm 
(multiplet) and methoxy group was confirmed at 
3.90 ppm (singlet). 
The 13C-NMR spectrum of 1 exhibited 
signals of 10 carbons at  146.73, 145.28,
132.96, 118.99, 114.30, 108.65, 85.90, 71.69, 
55.99 and 54.20 as listed in table 1. The partial 
structures were deduced from the HOMOCOSY 
and HMQC spectra, and were connected based 
on long-range correlations in the HMBC 
spectrum of 1. In the HOMOCOSY, cross peaks 
between the proton at  3.10 (H-5) and protons 
at  4.73 (H-6)/ 4.24 (Ha-4)/ 3.88 (Hb-6), and 
between proton at  6.88 (H-5) and proton at 
6.82 (H-6) were observed. In addition, H-C 
long-range correlations between the proton at 
3.10 (H-5) and the carbon at  85.90 (C-6)/
71.70 (C-4)/132.9 (C-1), between the proton 
at  4.73 (H-6) and carbons at  132.90 (C-1)/
118.99 (C-6)/ 108.65 (C-2) confirming the 
lignan skeleton of 1. The ESI mass spectra of 1
showed an ion peak at m/z 341 [M-H2O+H]
+
with strongest indensity corresponding to the 
molecular formula of C20H22O6. These findings 
suggested the structure of 1 as shown in Fig. 1. 
All NMR assignments of 1 were confirmed by 
detailed analyses of HSQC and HMBC spectra 
(table 1), which are in good agreement with 
those reported in the literature [4]. Thus, 
compound 1 was determined to be (±)2,6-bis(4-
hydroxy-3-methoxy-phenyl)-3,7-
dioxabicyclo[3.3.0]octane (Pinoresinol), the 
compound was isolated from Festuca argentina 
[4]. However, this is the first report of 1 from 
Asteraceae family. 
Compound 2 was isolated as yellow crystals 
suggesting that 2 is a flavonoid. The ESI mass 
spectrum showed an ion peak at m/z 319 
corresponding to the the molecular formula of 
C16H14O7. The 
13C-NMR and DEPT spectra of 2
confirmed the presence of 16 carbon atoms 
including 8C, 7CH and 1CH3 of the flavanon  
 
O
O
OH
OCH3
OCH3
HO
HH 1
2
3
4
5
6
7
8
1 '
2 '
3 '
4 '
5 '
6 '
1"
2"
3"
4"
5"
6"
 
1
O
OH
OR2
OR1
O
HO
OH
2
3
45
6
7
8
9
10
1'
2'
3'
4'
5'
6'
 
2 R1 = H, R2 = CH3; 3 R1 = H, R2 = CH3 , 2,3 
4 R1 = H, R2 = H
Figure 1: The structures of 1 - 4
221
Table 1: The NMR data of 1
C C* Ca,b Ha,c (J, Hz) HMBC (H to C) 
1, 5 54.2 54.20 3.10 (2H, m) C-2, 4, 1
2, 6 85.9 85.90 4.73 (2H, d, 5.0) C-1, 5, 8, 1, 2, 6
4, 8 71.7 71.70 
4.24 (2H, dd, 7.0, 9.5) 
3.88 (2H, dd, 4.0, 9.5) 
C-1, 2 
C-1, 2 
1, 1 132.9 132.90 -  
2, 2 108.7 108.65 6.89 (2H, d, 1.5) C-2, 1, 3, 4, 6
3, 3 146.7 146.73 -  
4, 4 145.3 145.28 -  
5, 5 114.3 114.30 6.88 (2H, d, 8.0) C-1, 3, 4, 6
6, 6 118.9 118.99 6.82 (2H, dd, 1.5, 8.0)  C-2, 1, 2, 4
3, 3-OCH3 56.0  55.99 3.90 (3H, s) C-3
*C of pinoresinol [4],  aMeasured in CDCl3, b125 MHz, c500 MHz. 
Table 2: The NMR data of 2 - 4
C C of 3 [5] C of 4 [6] Ca,b of 2 Ha,c (J, Hz) 
2 147.1 84.3    85.2  5.01 (1H, d, 6.5) 
3 136.1 73.0 73.6 4.59 (1H, d, 6.5) 
4 176.3 198.0 198.4  - 
5 161.2 164.8 165.3  - 
6 98.6 97.0   97.4  5.94 (1H, d, 1.5) 
7 164.4 167.8 168.9  - 
8 93.9 96.0 96.3  5.91 (1H, d, 1.5) 
9 156.8 164.0 164.5  - 
10 103.5 101.4 101.8  - 
1 122.6 129.6 129.8  - 
2 112.7 115.7 112.5  7.13 (1H, d, 1.5) 
3 149.4 145.6 148.9  - 
4 147.9 146.4 148.3  - 
5 116.0 115.7 115.9  6.85 (1H, d, 8.0) 
6 122.2 120.8 122.2  7.98 (1H, dd, 1.5, 8.0) 
OCH3 56.2    56.5  3.89 (3H, s) 
aMeasured in CDCl3,
b125 MHz, c500 MHz. 
222
skeleton. The spin-coupling of three protons of 
ring C suggested the hydroxyl and methoxyl at 
C-4 and C-3. The chemical shifts at  85.2 
(CH), 73.6 (CH) and 198.4 (C=O) were very 
similar to that of the flavanone reported in the 
literature [6]. The methoxyl group evidenced at 
C 56.5 (q) and H 3.89 (3H, s). The NMR 
assignments of ring C of 2 were deduced from 
those of 3 [5], and the NMR assignments of ring 
A were compared with that of 4 [6]. All these 
data led to determine the structure of 2 as 
3,4,5,7-tetrahydroxy-3-methoxyflavanone, 
which is first isolated from Silybum species. 
Acknowledgements: The authors wish to thank 
to Mr Dang Vu Luong, the NMR Lab., Institute 
of Chemistry, Vietnamese Academy of Science 
and Technology for performing the NMR, and 
Dr. Nguyen Tap, Department of Botany and 
Medicinal Plant Resources, National Institute 
of Medicinal Materials for the plant 
identification. This study was supported by a 
grant from the Vietnam-Italia international 
cooperation project. 
REFERENCES 
1. V. V. Chi. Vietnamese Medical Plant 
Dictionary, Ha Noi Medicine Pub. (1997). 
2. T. T. Diep, N. T. Dong, B. T. Bang, C. V. 
Minh, P. V. Kiem. Journal of Medicinal 
Materials, Vol. 10(6), 175 - 178 (2005). 
3. T. T. Diep, N. T. Dong, B. T. Bang, C. V. 
Minh, P. V. Kiem. Journal of Medicinal 
Materials, Vol. 11(1), 9 - 13 (2006). 
4. B. Vermes, O. Seligmann and H. Wagner. 
Phytochemistry, 30(9), 3087 - 3089 (1991). 
5. G. N. Kumari, L. J. M. Rao and N. S. P. 
Rao, Proc. Indian Acad. Sciences, Vol. 97, 
171 (1986). 
6. E. Kiehlmann and E. P. M. Li. Journal of 
Natural Products, Vol. 58(3), 450 - 455 
(1995).
 


Pinoresinol and 3,4',5,7-tetrahydroxy-3'-methoxyflavanone from the fruits of Silybum marianum (L.) Gaertn

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Pinoresinol and 3,4',5,7-tetrahydroxy-3'-methoxyflavanone from the fruits of Silybum marianum (L.) Gaertn

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