JOURNAL OF SCIENCE & TECHNOLOGY * No. 87 - 2012
STUDY ON FLAVONOID CONSTITUENTS O F HYMENOCALLIS LITTORAUS NGHIEN CUU THANH PHAN FLAVONOIT CAY BACH TRINH BIEN
{HYMENOCALLIS LISTTORALIS)
Dinh Thi Phuong Anh, Vu Dinh Hoang Tran Bach Duong Hanoi University of Science and Technolog}' Vietnam Institute of Industrial Chemistry
Received April 11, 2012; accepted April 25, 2012 ABSTRACT
3', 7-Dihydroxy-4'-methoxy-8-methylfiavan, 3\ 7-dihydroxy-4'-methoxyflavan and 5-hydroxy-7- methoxy-2-methyl-4H-chromen-4-one (eugenin) were isolated from the whole plant of Hymenocallis littoralis. 3', 7-Dihydroxy-4'-methoxy-8-methylfiavan. 3'. 7-dihydroxy-4''methoxynavan were found for the first time from the species. Structures of the compounds were elucidated by means of spectroscopic methods.
Keywords. Hymenocallis littoralis. Amaryllidaceae, flavonoid.
TOM T A T
Sa hpp chat fiavonoit da dugc phdn Idp t(r cdn flavonoit tdng cua cay Bach trinh bidn Hymenocallis littoralis Bdng cdc phuang phdp phd kdt hgp, ciu true eua ba hgp chit nay du-gc xdc dinh la 3'. 7-dihydroxy-4'-methoxy-8-methylfiavan (1). 3', 7-dihydroxy-4'-methoxyfiavan (2) vd 5- hydroxy-7-methoxy-2-methyl-4H-chromen-4-one (eugenin) (3). Day Id iin dau tien hai chdt 3'.7- dihydroxy-4'-methoxy-8-methylflavan (1) vd 3',7-dihydroxy-4'-methoxyfiavan (2) duge phdn lap tir loai Hymemocallis littoralis.
.. INTRODUCTION
Amaryllidaceae family consists of about 1600 species in the world. Hymenocallis genus is a large genus of this family with about 50 species [i],[2]. It is a good garden subject in warm climate, 70-90 cm high and has lovely white flowers. The Amaryllidaceae species are subjects to extensive study in the world and in Vietnam [3]-[5]. There are some studies on chemical constituents and biological activities of Hymenocallis littoralis [6]-[8] but this Is Ihe first report in Vietnam. The paper described the isolation and structure elucidation of three compounds from the plant namely as 3',7- Dihydroxy-4'-methoxy-8-methyinavan (1), 3',7- dihydroxy-4'-methoxyfiavan (2) and 5-hydroxy- 7-methoxy-2-meihyl-4H-chromen-4-one (eugenin) (3)
2. EXPERIMENTAL Instruments and chemicals
NMR experiments were performed on a Bruker AM500 FT-NMR Spectrometer using TMS as internal standard. Thin layer chromatography was carried out on precoated plates DC-Alufolien 60 F^.j and RP18 F:s4
(Merck-Germany) and visualized by UV light (254 nm) and sprayed with Vont's Reagent.
Column chromatography was carried out with normal phase (silica gel 90-240 mesh, 240-430 mesh, Merck).
Plant materials
The sample Hymenocallis littoralis was collected in Hanoi on December 2010 and identified by Dr. Vu Van Chuyen Institute of Ecology and Biological Resources, Vietnam Academy of Science and Technology. The sample was air-dried and powdered Extraction and isolation
The air-dried and powdered materials (18.5 kg) were extracted with methanol by Soxhlel extraction from 8h to lOh. The combined extracts were concentrated under vacuum to obtain crude residue (268.36 g) which was then acidified by 2% H2SO4 and ullrasoniced for getting homogenous mixture.
After removing sediment complex, the mixture was extracted by chlorofonn, gardienled with 5% Na:COi to pH<6 and concentrated lo gel total flavonoid residue (45.99 g) which was then dissolved by methanol lo give the soluble ,
JOURNAL OF ,SnF.NCE Si TECHNOLOGY * No, >7 - 2012 part in methanol (HLT, 20.9g), the insoluble
one was dissolved by chlorofonn (HLC, 25.09g).
The soluble part in chloroform (HLC) was chromalographed on a silica gel column (90-240 mesh) eluted with chloroform/methanol (49:1 v/v) to afford seven fractions HLCI-7. Fraction HLC7 (1.35g) was subjected to repeated column chromatography eluting with chloroform/methanol (49:1 and 19:1 v/v) to give three fractions. Then fraction HLC7.1 (0.39g) was chromatographed on a silica gel column (240-430 mesh) eluting with n-hexan/ethylacetate/methanol (3:9:1 v/v) to give six fractions HLC7.1.1-7.1.6. Compound 1 (20mg) was obtained from HLC7.1.1 (0.045g) with Rf being 0.4 at n- licxane/ethylacetate/methanol (3:10:0.1 v/v).
Fraction HLC7.3 (0.25g) was chromatographed on a silica gel column (90-240 mesh) eluting with n-hexane/ethylacetate/methanol (3:10:0.1) 10 yield two fractions HLC7.3.1-7.3.2.
Compound 2 (17mg) was purified from HLC7.3,1 (0.033g) by a silica gel column eluting with n-hexane/ethylacetate/methanol (10:10:0.1 v/v). R, compound 2 is 0.4 at n- hexane/ethylacetate/methanol (3:10:0.1 v/v).
Similarly, the soluble part in methanol (HLT) was chromatographed on a silica gel column (90-240 mesh) eluting with n- hexane/ethylacetate/methanol (20:10:0.1 v/v) to give ih-.t fractions HLTl-3. Fraction HLT2 (2.43g) was subjected to repeated column chromatography eluting with n- hexan/ethylacetate/methanol (20:10:0.1 v/v) to yield ten fractions HLT2.1-10. Compound 3 (32mg) was crystallized from HLT2 10 (0.21g);
Ri of 3 is 0.43 at n-hexane/ethyl acelatc/mclhanol (3:10:0.1 v/\)
3',7-Dihydroxy-4'-mcthoxy-8- mcthylflavan: Colourless oil; 'H NMR (500 MHz, CDCl,), 6 (ppm): 7.02 (IH, </ J = 2Hz, H-2'), 6.94 (IH, M. J = 8.5; 2Hz. H-6'), 6.87 (IH, </ J = 8Hz, H-5'), 6.78 (IH, cl. J 8Hz.
H-5), 6 39 (IH. d J - 8.5Hz, H-6), 4.98 (IH, till. .1 - 10; 2Hz. H-2), 3.90 (3H. ,, H-4'), 2.90 (2H, m. H-4). 2 71 (2H, cM. y = 16; 5. 411/. H- 4), 2.17 (211. »,. H-3), 2.14 (3H, .v, C'-,S). 1.99 (211, m. C-j). '-'C NMR (I25MH7. ClX'l,). « (ppm) 153.6 (v. C-9), 152.727 (.s-. C-71, 146 1)
(.(, C-4'), 145,6 (j, C-3'), 135.6 (.'. C-l'), 126,5 (rf, C-5), 113.9 (.V, C-IO), 112.3 (rf, C-2'), 111,5 (.V, C-8), 107,3 (rf, C-6), 77.2 (rf, C-2), 56.0 (?, C-4'), 30.0 (;, C-3), 24.8 (/, C-4), 8.1 (q. C-8).
3',7-Dlhydroxy-4'-iiiethoxynavaii:
Colourless oil; 'H NMR (SOOMHz; CDCl,), 6 (ppm): 6.95 (IH, .,. H-l' ), 6.89 (IH, rf ,; = 8Hz, H-5 ), 6.88 (IH,.,, H-6'), 6.87 (IH, t. H- 5'), 6.39 (IH, rfrf 7 = 8; 2,5Hz, H-6). 6.37 (IH, rf y « 2,5Hz, H-8 ), 4.92 (IH, rfrf J -/O;
2,5Hz, H-2), 3.88 (3H, >. H-4'), 2.86 (2H, tn. H- 4), 2.02-2.06 (2H, m. H-3); " C NMR (125MHz, CDCl,), 8 (ppm): 155 7 (,. C-9), 155.5 (.1. C-7), 146.7 (j, C-4'), 145.7 («. C-3'), 134.6 (,. C-l'), 129.8 (rfC-5), 117.5 (rf C-6'), 113.0(.V. C-10), l l 2 . 8 ( r f C - 2 l , lll,0(rfC-5'), 107.8 (rf C-6), 103.0 (rf C-81, 77.4 (rf C-2).
55.7 (ry. C-4'), 29.7 il. C-3), 24.1 (/. C-4).
, 5-Hydroxy-7-niethoxy-2-methyl-4fl"- chroincn-4-one (Eugenin): Needle white crystal, melting poim: 119-I20°C; 'H NMR (500 MHz, CDCl,), 6 (ppm): 6.35 (IH, rf 7 - 2,5Hz, H-8), 6.33 (IH, rf V = 2,5Hz, H-6), 6.03 (IH, i, H-3), 3.85 (3H, i, H-7), : 3 5 ( 3 H , s , H - 2); " C NMR (125MHz, CDCl,), 8 (ppm):
182.5 (s. C-4), 166.8 (s, C-2), 165.3 (s, C-7), 162.2 (.^, C-5), 158.1 (i. C-9). 108.8 (rf, C-3), 105.2 (.t, C-10), 97.9 (rf, C-6), 92.5 (rf, C-8), 55.7 (q. C-7-OCH,), 20.5 (</. C-2-CH,).
3, RESULTS AND DISCUSSION Compound 1 was isolated as colorless oil. The ' H NMR spectrum of 1 showed proton signals ofan ABX spin system at S,, 7.02 (IH, rf J " 2Hz, 11-2). 6.94 ('iH, rfrf J = 8.5; 2Hz, H-6'), 6.87 (IH. rf J 8Hz. H-5'), two doublets and t\\o multiplets of aromatic protons at 8,1 6.78 (IH, rf rf = 8Hz. H-5), 6.39 (IH, rf, J - 8.5Hz, H-6), 2.90 (2H, m. H-4), 2.17 (2H, m, H-3). In addition, an oximethme and one melhoxy group were also observed at 4.98 (IH, rfrf. rf = 10; 2Hz, H-2). 3.90 (3H, ftr.s, 4'-OCH5).
The "C NMR, DEPT 90 and DEPT 135 spectra of I revealed seventeen carbon signals including Iwo CH,, two CH2, six CH and seven quaternary carbon atoms. One methine, one methoxy groups and three saturated carbons in which one carbon responded with oxide appeared at S, 8.1 (C-8), 56.04 (C-4) and 29.99 (C-3). 24.78 (C-4), 77.21 (C-2) together with H • -'"."•••^ .•„!,„„, f S ,nn .- 150 28
JOURNAL OF SCIENCE & TFXHNOLOGY • No. 87 - 2012 suggesting Ihe fiavan of 1. From this data in
comparison with the literature, compound 1 was determined to be 3',7-dihydroxy-4'-methoxy-8- methylflavan [9].
Compound 2 was isolated as colorless oil. The NRM data of compound 2 were similar to those of 1, except for the absence of the methine at SH 2.14 and 5c 8.10. Thus, compound 2 was identified as 3',7-dihydroxy- 4'-methoxyflavan [10].
Compound 3 was isolated as needles white crystal. The '^C NMR, DEPT 90 and DEPT 135 spectra of I revealed eleven carbon signals including two CHi, three CH and six quaternary carbon atoms. The "C NMR showed two singlets and two doublets of aromatic
carbons at 162.2 (.y, C-5), 165.3 (s, C-l), and 97.9 (d, C-6), 92,5 (d, C-8). Two olefinic carbons were recognized from 5c 90 to 170. In addition, a methine, a methoxy and a carbonyl carbon appeared at 5c 20.5 (q, 2-CH3), 55.7 {q, 7-OCH3), and 182.5 (,v, C-4) suggesting for a chromenone. The ' H NMR spectrum of 3 showed a singlet of OH group at 5n 12.7 (1H, s, H-5). Two doublet protons were observed at Su 6.35 (IH, d,J = 2,5Hz, H-8), 6.33 (IH, dJ = 2,5Hz, H-6). Two resonance singlet protons of a methine and a methoxy group were confirmed ay 2.35 (3H, .f, H-2), 3,85 (3H, s, H-7). From this data in comparison with the literature, compound 3 was determined to be 5-hydroxy- 7-methoxy-2-methyl-4H-chromen-4-one (eugenin) [11].
(1) (2)
Fig I. Structure of compounds I. 2, 3 isolated form the flavonoid residue of Hymenocallis littoralis
4. CONCLUSION
From the total flavonoid residue of the Hymenocallis littoralis, 3',7-dihydroxy-4'- methoxy-8-methylflavan; 3',7-dihydroxy-4'- methoxyflavan and 5-hydroxy-7-melhoxy-2- methyl-4//-chromen-4-one (eugenin) were isolated. The two flavonoids 3',7-Dihydroxy-4'- methoxy-8-methylflavan and 3',7-dihydroxy-4'-
melhoxyflavan were isolated for the first time from Heminocallis littoralis. These structures were determined by means of spectroscopic methods and comparison with the literature data.
Acknowledgments
This work was financially supported by Ministerial Project B2010-01-317.
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Author's address: Vu Dinh Hoang - Tel. (i S4) 914661299, Email: hoangvu1964'(/>ahoo.com School of Chemical Engineering
Hanoi University of Science and Technology No.l Dai Co Viel Sir.. Ha Noi. Vicl Nam
