JOURNAL OF SCIENCE A TECHNui^ii^T«>o. ni-ivit
FURANOCOUMARINS AND FALCARINDIOL FROM ANGELICA DAHVRICA CkC HOP CHAT FURANOCOUMARIN VA FALCARINDIOL PHAN LAP
T O C A Y BACH CHi ANGELICA DAHURICA
Nguyen Hoai Nam", Phan Thi Thanh Huon^, Chau Ngoc Diep', Le Due Daf, Ninh Thi Ngoc", VuAnh Tu", Nguyen Phuong Thao", Nguyen Xuan Cuon^,
Nguyen The Cuon^, Tran Thu Huon^, Ninh Khac Ban", Phan Van Kiem", Chau Van Mink'
" In.ititute of Marine Biochemistry. VAST
* In.stltutc of Ecology and Biological Resoitrctw. VAST 'Hanoi University oJScience and Technology
Received February 20.2012; accepted April 25.2012
ABSTRACT
Three coumarins and a polyacetyienlc oxylipin were isolated fmm the roots of Angelica dahurica.
Analysis of their spectral data (ESI-MS. 'H-NMR. '^C-NMR. DEPT. HSQC and HMBC) and comparison with the literature data conTirmed their structures as imperatorin (1). isoimperattvin (2), xanthotoxol (3), and falcarindiol (4). The ^'C-NMR data at C-5 and C-8a of 2 were reassigned by 2D- NMR spectra. Among them, compounds 1 and 4 were main constituents of this plant. In the literatum data, these compounds showed various biological activities such as. Anti-HIV. anticonvulsant, anlh inflammatory, antimicrobial, analgesic, antimutagenic and antiproliferative activities.
T 6 M TAT
Sa hpp chdt furanocoumann vd mdt hgp chdt polyacetylen oxylipin di/gc phdn Idp lir rd cdy bach chi (Angelica dahurica) Bdng cdch kdt hgp cdc s6 lidu phd (phd cdng hwdng tir hat nhdn m^
chieu'H-NMR. '^C-NMR. DEPT vd hai chidu: HSQC vd HMBC, phd khdi lu^g: ESI-MS) so vdi lai lieu tham khdo. ciu true hda hge eua ede hr^rp chdt dugc xdc d/nh Id imperatorin (1). isoimperatonn (2).
xanthotoxol (3) vd falcarindiol (4). Sd lidu phd '^C-NMR if cdc v\ tri C-5 vd C-6a qua hgp chit 2 dugc gdn lai chinh x^e trdn ea s& kdt qud phd cdng hudmg tir hat nhdn hai chidu. Theo cdc tdi lidu tham khdo. ede hgp chdt 1, 4 id thdnh phdn chinh cua cdy bach chi vd thd hidn nhidu hoat tinh sinh hgc rdt thu V!, vi dl,! nhw: hoat tinh khdng HIV, chdng co gidt. khdng vidm. khdng vi sinh vdt, gidm dau. khdng dQt bidn vd tdng smh td bdo.
1. INTRODUCTION
Angelica L, is a large genus of the Apiaceae family. There are about 110 Angvlun species in the world. In Vietnam, 4 or 5 species were recorded to dale. Of ihesc. Agelica dahurica is perennial herb, I-I 5 m high [I).
The roots of this plant are used in Chinese traditional medicine as antipyretic and analgesic agents for the ircalmenl of common cold, headache, sinusitis, and toothache [2). In Vielnam, this plant is also used as antipyretic and analgesic agents for the treatment of common cold, petechial fever, headache.
toothache, and period pains. In addition,'this plant is used against arthritis, mastitis, gathering pimples, and traumatic injuries [1]
Previous investigations indicated Ihal coumann
derivatives an: main consiiiuenis of Ihis plani
[ii-pi
Furanocoumiarins from .4. dahurica roois were found lo ha\c aniimutagenic activity against the mutagenicit\ of 2-amino-3-methyl- iniida/o[4,5-nquinoline (IQ) in Salmonella tvphimuriuin, anii-inflammaiory activity, cNioloxic aetuii) and inhibitory effects against a number of cn/>nics [2]. Furanocourmarins also used as reference standards m the quality control of A. dahurica and its products.
So high purii} preparation of them is of great interest. As a part of out imcstigations on this plant, the preseni paper deals with the isolation and structural elucidation of three coumarins, imperatorin (1), isoimpciaiorin (2) and xanthotoxol (3), and one pol\,i^et\lenic
JOURNAL OF SCIENCE & TECHNOLOGY * No. 87 - 2012 oxylipin, falcarindiol (4), ftx>m methanol extract
of A. dahurica roots,
Fig. I Structures of 1-4 2. MATERIAL AND METHODS General experimental procedures
All NMR spectra were recorded on a Bruker AM500 FT-NMR spectrometer. The ESI mass spectra were obtained on an Agilent 1100 Series LC-MSD Ion Trap spectrometer.
Colunm chromatography (CC) was performed on silica gel 230-^400 mesh (0.040 -H 0.063 mm, Merck) or YMC RP-18 resins (30^50 ^m, Fujisilisa Chemical Ltd.). Thin layer chromatography was performed on DC- Alufolien 60 F2S4 (Merck 1.05715) or RPis ?25A.
(Merck) plates. Compounds were visualized by spraying with aqueous 10% H2SO4 and heating for 5 minutes.
Plant material
Samples of Agelica dahurica (Fisch.ex Hofftn.) Benth. et Hook.f., were collected in Tam Dao botanic park, Vinh Phuc, Vietnam, during December 2011 and identified by MSc Nguyen The Cuong. A voucher specimen was deposited at the Herbarium of Institute of Ecology and Biological Resources, VAST.
Extraction and isolation
Air-dried and powdered roots of A.
dahurica (5 kg) were exhaustively extracted (three times, each 60 min) with MeOH (40 - 50°C) under ultrasonic condition. The combined extract was evaporated under vacuum to obtain 200 g MeOH residue. This was suspended in water (3 1) and partitioned successively with n- hexane ( 3 x 3 1) and CHCI3 ( 3 x 3 1), giving 75 and 35.5 g of corresponding residues.
The chloroform extract (35,5 g) was subjected to a silica gel CC using stepwise
elution of n-hexane-acetone (from 100/1 to 1/10) to give seven fractions, C1-C7. Fraction C3 (2.5 g) was crystallized and followed by YMC CC with acetone-water (2/1, v/v) as eluent to furnish compound 2 (8 mg, white powder).
Compound 1(15 mg. white powder), 3(10 mg, white powder) and 4 (200 mg, pale yellow oil) were purified from fraction C4 (3.5 g) by silica gel CC with n-hexane-acetone (4/1) followed by YMC CC with MeOH-water (2/1).
Imperatorin (1): White powder; C16H14O4, M = 270; ESI-MS: m/z 271 [M+Hf; 'H-NMR (CDCI3, 500 MHz) and "C-NMR (CDClj, 125 MHz) see Table 1.
Isoimperatorin (2): White powder;
C16H14O4, M = 270; ESI-MS: m/z 271 [M+H]*;
'H-NMR (CDCI3, 500 MHz) and '^C-NMR (CDClj, 125 MHz) see Table 1.
Xanthotoxol (3): White powder; C11H6O4, M = 202; ESI-MS: m/z 203 [M+H]*; 'H-NMR (CDCI3 + CD3OD, 500 MHz) and "C-NMR (CDClj + CD3OD, 125 MHZ) see Table I.
Falcarindiol (4): Pale yellow oil; [ajo" -130 (c 0.5, CHCI3); C17H24O2, M = 260; ESI-MS: m/z 261 [M+H]*; 'H-NMR (CDCI3, 500 MHz): 8H (ppm) 5.26 (IH, br d, J = 10.0 Hz, H,-l), 5.48 (IH, br d, / = 17.5 Hz, Ht-l), 5.94 (IH, m, H- 2). 4.95 (IH, d, J = 5.5 Hz, H-3), 5.21 (IH, d, 7
= 8.5 Hz, H-8), 5.52 (IH, br d, J = 8.5 Hz, H-9), 5.61 (IH, m, H-10),2.10(2H, dd,J = 7.5, 15.5 Hz, H-11), 1.38 (2H, m, H-12), 1.28 (8H, m, H- 13 to H-16),and 0.88 (3H,t,y = 6.5 Hz, H-17);
"C-NMR (CDCI3, 125 MHz) 6c (ppm) 117.31 (C-l), 135.79 (C-2), 63.44 (C-3), 78.25 (C-4), 70.26 (C-5), 68.68 (C-6), 79.85 (C-7), 58.55 (C-8), 127.65 (C-9), 134.64 (C-10), 27.68 (C- 11), 29.26 (C-12), 29.14 (C-13), 29.09 (C-14), 31.77 (C-15), 22.62 (C-l 6), and 14.08 (C-17).
3. RESULTS AND DISCUSSION Compound 1 was isolated as a white powder. The 'H-NMR spectrum showed signals of two pairs of coupled olefinic protons at 5H 6.37 (1H, d,J= 9.5 Hz, H-3)/7.76 (I H, d, J = 9.5 Hz, H-4) and 6.81 (IH, d, J = 2.0 Hz, H- 9)/7.69 (IH. d, J = 2.0 Hz, H-10) and one isolated aromatic proton at 5 7.36 (IH, s, H-5).
The small coupling constant of one pair of the olefinic protons (J - 2.0 Hz) suggesting for a
JOURNAL OF .SCIENCE « TECni;
furanocoumarin, the main constituent of A.
rfa*uWca[l]-[3].
Fig. 2 Key HMBC correlations of I Tile presence of a prenyloxy moiety was suggested by proton signals at 5 5.01 (2H, d, J
= 7.0 Hz, H-l'), 5.61 (IH, t.J- 7,0 Hz, H-2'), 1.74 (3H, s, H4'), and 1.72 (3H, s, H-5'), which was further confirmed by corresponding carbon
Table I. NMR (500 MHi. CDCIO data of 1-3 and reported compounds signals at 8 70.17 (C-l'), 119.78 (C-2'). 139-75 (C-3'), 25.81 (C-4'), and 18.12 (C-S'). The C- NMR spectrum of 1 revealed sixteen carbon signals of a furanocoumarin having a prenyloxy moiety. All protons of 1 were assigned with relevant carbons by an HSQC spectrum. The NMR data of 1 were limilar lo those of imperatorin (table I) [4], one of the main compounds of A. dahurica (IH31. The structure of 1 was confirmed by HMBC experiment. The HMBC correlation between H- r (8 5.01) and C-8 (8 131.70) indicated the attachment of the prenyloxy moiety at C-8.
Similarly, detailed analyses of the other HMBC cross peaks indicated the structure of I as 9-(3- methyl-2-butenyloxy)-7-oxofiirot34- gjchromene with common name imperatorin.
c
2 3 4 4a 5 6 7 8 Sa 9 10 f 2' 3' 4' 5'
1 '6c 160.6 114.7 144.3 116.1 113.1 125.9 148.6 131.7 143.9 106.7 146.6 70.2 119.8 139.8 25.8 18.1
8c 160.52 114.72 144.33 116.S0 113.14 125.86 148.64 131.70 143.85 106.71 146.62 70.17 119.78 13975 25.81 18.12
SH mult. (J = Hz) 6.37 d (9.5) 776 d (9.5) 7.36 s
6.81 d (2.0) 769 d (2.0) 5.01 d (7.0) 5.611(7.0) 1.74 s 1.72 s
2 '6c 161.1 112.4 139.6 107.4 152 f 114.0 158.0 94.1 HS.f
105.0 144.8 69.4 119.0 1398 25.8 18.2
8r 161.29 112.52 139.57 10748 US 94 114.16 158.11 94.18 152.64 105.04 144.87 69.71 119.08 13981 25.81 18.22
8H mult. U - Hz)
-
6.27 d( 10.0) 8.15d(10.0)7.15 s 6.% d (2.0) 7.59 d (2.0) 4.92 d (7.0) 5 54 1 (7.0) 1.80 s 1.70 s
i' '8c 161)7 114.2 145.8 116.8 110.5 125.9 145.8 130.7 1403 107.5 147 7
V
161 99 1 n 4f) 145.77 116.04 109.80 125.94 145.68 130.54 139.28 106 57 146.79
8»
mulL U • Hz)
-
6.36dd(1.0.9J) 7.76 dd( 1.0.9J) 7.25 s6.82 d (2.0) 7.72 d (2.0)
'recorded in CDCb + CDjOD, ""Sc of imperatorin [4], %• of isoimperatorin [5], % of xanthotoxol [6],
^ihese iwo values must be reversed.
The 'H- and "C-NMR spectra of 2 are similar lo those of 1. indicate that 2 was also a furanocoumarin. The ' H - N M R spectrum of 2 still revealed the presence of two pairs of coupled olefinic protons, one isolated aromatic proton and protons of a prenyloxy moiety. The difference of the proton signals between the two compounds is only in the chemical shifts (table 1) suggesting 2 is an isomer of 1. The proton H- 4 was shifted down-field lo 8 8.15 suggesting the placement of the prenyloxy moiety at C-5.
which was further confirmed by HMBC correlations between H-l' (6 4.92) and C-5 (6 148.94) and between H-4 (5 8.15) and C-5 (5 148.94). Thus, compound 2 was identified as isoimperatorin [5]. The '^C-NMR chemical shifts reported for C-5 and C-8a (table 1) in the literature were incorrect and must be reversed.
The NMR data of compound 3 were similar to those of I, except for ihe absence of the prenyloxy moiety. Detailed analyses of the
JOURNAL OF SCIENCE & TECHNOLOGY • No. 87 - 2012 ID, 2D-NMR and ESI-MS data in comparison
with the literature values led to the elucidation of 3 as xanthotoxol [6].
OH QH
Fig. 3 Key HMBC correlations of 4 Compound 4 was isolated as a pale yellow oil. Signals in the "C-NMR spectrum for four quaternary carbons at 5 79.85 (C-7), 78.25 (C-4), 70.26 (C-5) and 68.68 (C-6) were indicative of a polyacetylene natural product possessing two triple bonds. Further signals in the 'H and '^C-NMR spectra included a cis- double bond [5H 5.52 (IH, br d, 7 = 8.5 Hz, H- 9)/5.61 (IH, m, H-10)], two oxymethine [5H 4.95 (IH, d,J= 5.5 Hz, H-3)/5c 63.44 (C-3) and 5H 5.21 (IH, d, J = 8.5 Hz, H-8)/5c 58.55 (C-8)], an exo-cyclic methylene [6H 5.26 (IH, b r d , J = 10.0Hz,H,-l)/5.48(lH,brd,J- 17.5
Hz, Hh-I) and 8c 117.31 (C-l)], one terminal methyl [8H 0.88 (3H, t, / = 6.5 Hz, H-17)/8c 14.08 (C-17)], and five aliphatic methylenes.
The above evidence, a good agreement of the NMR data of 4 with the literature values and HMBC results (Fig. 3) indicated compound 4 to be falcarindiol [7].
Among isolates, 1 and 4 were main constituents of Ihis plant. In the literature data, these two compounds showed various biological activities (such as: Anti-HIV, anticonvulsant, anti-inflammatory, antimicrobial, analgesic, antimutagenic and antiproliferative activities; cytokine release and DNA topoisomerase inhibitors) [1-3]
suggesting for further application investigations to make high-value medicinal products.
Acknowledgments
This work was financially supported by VAST (CTG.03/11-12). The authors would like to thank MSc. Dang Vu Luong (Institute of Chemistry) for recording the NMR spectra.
REFERENCES
1. Bich D. H., Chung D. Q., Chuong B. X., Dong N. T., Dam D. T . Hien P. V., Lo V. N., Mai P. D., Man P. K., Nhu D. T., Tap N., Toan T; Medicinal Plants and Animals of Vietnam; pp 127-131, Hanoi Science and Technology Publisher, Vol. I, 2004.
i . Tang W., Eisenbrand G.; Handbook of Chinese Medicinal Plants - Chemistry, pharmacology, toxicology; pp 127-138, Wiley-VCH publisher. Vol. 1, 2011.
3. Dictionary of Natural Products on DVD, version 18.1, Copyright® 1982-2009 CRC Press.
4. Masuda T , Takasugi M., Anetai M.; Psoralen and other linear furanocoumarins as phytoalexins in Glehnialittoralis. Phytochemistry, 47, 13-16 (1998).
5. Yang F., Zhang T., Liu Q., Xu G., Zhang Y., Zhang S, Ito Y.; Preparative isolation and purification of notopterol and isoimperatorin from Notopterygium forbessi Boiss (Chinese traditional medicinal herb) by high-speed counter-current chromatography. Joumal of Chromatography A, 883, 67-73 (2000).
6. Breitmaier E., Voelter W.; Carbon-13 NMR Spectroscopy; p. 441, VCH. Weinheim, 1986.
7. Lechner D., Stavri M., Oluwatuyi M., Pereda-Miranda R., Gibbons S.; The anti-staphylococcal activity of Angelica dahurica (Bai Zhi). Phytochemistry, 65, 331-335 (2004).
Author's address: Nguyen Hoai Nam - Tel. (+84) 912260146, email: [email protected] Institute of Marine Biochemistry, Vietnam Academy of Science and Technology No. 18, Hoang Quoc Viet road, Ha Noi, Viet Nam
