Nghien ^ i'u - Ky thuat
Giocli.i^riosid F, mot phenolic glycosid moi
w c. y bon bot {Glochldion eirocarpum Champ.)
Vu Kim Thu'''\ Phan Van Kiem', Chau Van Minh', Pham Hai Yen' Nguyen Xuan Cu-crng', Hoang Thanh Huong' Vien Hoa hoc cac hap chdt Men nhien, Vien Khoa hoc vd Cong nghe Viet Nam Truang dgi hoc Mo dia chdt. Tit Liem, Hd Noi
Mcr d a u
Cay bon bpt {Glochldion eriocarpum Champ.), eon gpi la bpt l e h , ehe bpt, 16 lao nhlng. Cay nay mpe hoang nhilu 6' viing nui nyo'c ta. La bon bpt eo vj nhat, tinh binh, eo tae dung Igi thIp, thanh nhiet, tieu doe, Igi tieu, giai dpc do di irng san. La bon bpt chu'a mam ngira, may day, eczema. Theo kinh nghiem dan gian, cay bon bpt diing de ehCra rIn e I n (gia l l y nyo'c ulng eon ba d i p len vet thyang), chira dj irng do san, la chay, ly tryc khuan. Tai eae benh vien d6ng y, ngyai ta dung la eay nay chu'a phu thung, ca phij do than, vo'i lilu 35g la kh6/ngay, s i c lly nyo'c ulng cho hieu qua dilu trj rIt t i t '^"^'.
Cae hgp chit lupenon, 3-epi-lupeol, gloehidon, gloehidonol, glochidiol va lup-20(29)-ene-ip,3|3- diol da dyge phan lap tir la eay, trong do hgp c h i t glochidiol co hoat tinh ire c h i manh sy phat trien eua eae dong t l bao ung thy MCF-7, NCI-H-460 va SF-268 trong thCr nghiem in vitro Tac dung ehIng 6x1 hoa va ehIng nhiim khuan eua eay bon bpt eung da dygc nghien ciru '"'. Trong chyang trinh sang Ipc tim kiem cae hoat c h i t chIng ung thy eo nguIn g l e thien nhien, chung t6i da tiln hanh thir nghiem hoat tinh ire c h i sy phat trien eua mot so dong te bao ung thy ngyo'i thyc nghiem eua djeh chiet methanol tir la eay bon bpt. K i t qua eho t h i y djeh chilt methanol cua phin tren mat d i t cua eay bon bpt u'C e h i manh sy phat then eiia eae t l bao ung thy nhy ung thy eo tir eung (Hela:
ICso = 6,17±0,88 yg/ml; SK0V3: IC50 = 7,26
±0,35 yg/ml), ung thy vii (MCF7: IC50 = 12,22±0,37 yg/ml; KPL4: IC50 5,87±0.66|j.g/ml), ung thy bieu m6 than chuyen dang (C0S7: IC50 = 15,70±1,48 |ig/ml).
Trong cae bai bao tryo'c day '®"^' ehiing t6i da th6ng bao qua trinh phan lap va xac djnh c l u triic hoa hpe eae hgp chit tritecpenoid saponin
[51
mo'i la glochierlosid A, glochierlosid B, glochierlosid C, glochierlosid D, glochierlosid E, glochieriol, eae hgp c h i t lupan nhy gloehidon, lup-20(29)-en-3a,23-diol, lup-20(29)-en-1 p,3p- diol, lup-20(29)-en-ip,3a-diol, lupeol, 3-epi- lupeol va gloehidonol, eac hgp c h i t flavan nhy 5,7,3',4'-tetra-0-methyl-enf-epieateehin 3-0-j3-D- glueopyranosid, 5,7,3',4'-tetra-0-methylepieateehin.
Trong s i do gloehieriosid A va glochierlosid B eho k i t qua r I t t i t ve gay dpc t l bao tren mot so dong t l bao ung thy ngyo'i nhy ung thy mau (HL-60), da day (HT-29), vu (MCF-7) va buing trirng (SK-OV-3) '®'. Bai bao nay thong bao k i t qua tach chilt va xac dinh e l u true hoa hpe cua 4 hgp c h i t benzyl 4-0-p-D-gIueopyranosyl-3,4- dihydroxy-benzoat (1), eateehin (2), gloehidionolaeton E (3) va benzyl-(3-D- glueopyranosid (4) tir la eay bon bpt.
T h y c n g h i e m v a Phu'O'ng p h a p M i u thyc vat
Phan tren mat d I t eua eay bon bpt {Glochldion eriocarpum Champ.) dyge thu tai Tam Dao, VTnh Phiie thang 12 nam 2006. Miu dyge TS Dyang Dwc Huyin, TS Ninh Khie Ban, TS Ha Van T u i , ThS Nguyin T h i Cyo'ng, V i l n Sinh thai va Tai nguyen sinh vat giam djnh.
M i u tieu ban dyge lyu giy tai Vien Hoa hpe eac hgp chit thien nhien.
Phu'O'ng phap nghien ciPu
Phwo'ng phap phan lap cac hg'p chat sic ky lop mong (TLC): S i c ky lo'p mong dyge thyc hien tren ban mong trang s i n DC- Alufolien 60 F254 (Merck 1,05715), RP^g F254S (Merck). Phat hien c h i t b i n g den tu' ngoai 6' hai byo'c song 254 nm va 365 nm hoac diing thulc thir la dung dich H2SO4 ^0% d y g e phun d i u ien ban mong, s l y kh6 r l i ha nong tren bep dien tir tir d i n khi hien mau.
N g h i e n
CLPU- Ky t h u a t
sic ky cot (CC): Sac ky cot dygc tiln hanh VO'I ehit h i p phu la Silica gel pha thyo'ng va pha dao. Silica gel pha thyang co eo' hat la 0,040-0,063 mm (240-430 mesh). Silica gel pha dao YMC (30-50 ym, Fujisilisa Chemical Ltd.).
Phu'cng phap xac djnti c l u true hoa hoc cac hgp chit
- Diem nong chay do tren may Kofler micro- hotstage
- Pho khIi lygng phun mil dien tCr (Electrospray Ionization mass spectra) dyge do tren may AGILENT 1100 LC-MSD Trap.
- Pho eong hyo'ng tir nhan NMR dyge do tren may Bruker AM500 FT-NMR Spectrometer.
Phan lap hg'p chat
Phin tren mat d i t eua cay bon bpt (30 kg) dyge bam nho, phai kh6 r l i va nghiln thanh bpt (7,0 kg) sau do dem chilt vai methanol thu dygc 500 g can c h i l t Hoa tan can chilt nay vao 3 lit nyo'c e l t r l i chilt l l n lygt vo'i n-hexan, eloroform va ethyl aeetat (m6i loai 3 lit) thu d y g c cac can chilt n-hexan (10 g), eloroform(113 g), ethyl aeetat (120 g) sau khi loai dung m6i va phin djeh nyac. Can chilt ethyl aeetat (120 g) dyge phan lap tren s l e ky cot vo'i c h i t h i p phu la silica gel pha thyo'ng va dung m6i rCra giai la cloroform/methanol (gradien tir 10/1 d i n 1/2, v/v) thu dygc 4 phan doan ky hieu la FBI (25 g), FE2 (30 g), FE 3 (45 g) va FE4 (20 g). Phan doan FE2 (30 g) dyge tilp tue phan lap tren s i c ky cot nhoi YMC RP18 vo'i dung moi rCra giai la methanol/nyo'c (4/1, v/v) thu dyge 5 phan doan nho han, ky hieu la SFE1-SFE5. Phan doan SFE3 (8 g) dyge tinh e h i " b i n g s l e ky cot nhli silica gel pha thyo'ng vo'i he dung moi rira giai la cloroform/methanol/nyo'c (7/3/0,1, v/v/v) thu dyge hgp c h i t 2 (15 mg) dyo'i dang e h i t rIn mau vang. Djeh nyo'c dyge phan lap tren s i c ky cot trao doi ion HP-20 vo'i dung m6i nyac/methanol (gradien: 100/0, 75/25, 50/50, 25/75, 0/100, v/v, m i l l l n 2 lit) thu d y g c 5 phan doan ky hieu l l n lygt la FN1-FN5. Phan doan FN4 (75% MeOH, 25 g) dyge phan lap tren s l e ky cot nhli silica gel pha thyo'ng vo'i he dung moi la cloroform/methanol 4/1 (v/v, 2 lit) thu dyge 5 phan doan nho han ky hieu la SFN1- SFN5. Phan doan SFN2 (7,5 g) dyge t i l p tue tinh c h i tren s l e ky cot RP18 vai he dung moi la methanol/nyo'c/aceton (5/1/1, v/v/v) thu dyge hgp c h i t 1 (12 mg) dyo'i dang tinh the hinh kim
kh6ng mau. Hgp chat 3 (8 mg) va 4 (11 mg) thu du'gc dyo'i dang c h i t bpt kh6ng mau sau khi tinh e h i phan doan SFN5 (10 g) tren s l e ky cot RP18 VO'I he dung m6i la methanol/nyo'c/aceton (6/1/0,5, v/v/v).
Benzyl 4-0-p-D-glucopyranosyl-3,4-dihydroxy -benzoate (1): Tinh the hinh kim kh6ng mau, nhiet dp nong chay 134 - 136°C.
ESI-MS: m/z 407 [M+H]^ (positive), 405 [M- H]" (negative), M=406, CTPT C20H22O9.
' H - N I M R (500 MHz, CD3OD) 8 (ppm): 7,30 (1H, 6, J = 9,0 Hz, H-3), 6,98 (1H, dd, J = 2,0, 9,0 Hz, H-4), 7,21 (1H, d, J = 2,0 Hz, H-6), 7,49 (2H, d, J = 7,0 Hz, H-2', H-6'), 7,40 (2H, {. J = 7,0 Hz, H-3', H-5'), 7,35 (1H, m, H-4'), 5,33 (1H, d,J = 12,5 Hz, Ha-7'), 5,39 (1H, d, J = 12,5 Hz, Hb-7'), 4,73 (1H, d, J = 7,5 Hz, H-1"), 3,46 (H- 2"), 3,40 (H-3"), 3,48 (H-4"), 3,38 (1H, m, H-5"), 3,71 (1H, dd, J = 5,0, 12,0 Hz, Ha-6"), 3,92 (1H, dd, J = 2,0, 12,0 Hz, Hb-6").
^H-NIWR (500 MHz, DMSO-de) 5 (ppm): 7,05 (1H, d, J = 3,0 Hz, H-2), 7,14 (1H, d, J = 8,5 Hz, H-5), 6,90 (1H, dd, J = 3.0, 8,5 Hz, H-6), 7,49 (2H, d, J = 8,0 Hz, H-2', H-6'). 7,39 (2H, t, J = 8,0 Hz, H-3', H-5'), 7,48 (1H, t, J = 8,0 Hz, H-4'), 5,30 (2H, s, H-7'), 4,71 (1H, d, J = 7,5 Hz, H-1"), 3,25 (H-2"), 3,30 (H-3", H-5"), 3,16 (H-4"), 3,46 (1H, dd, J = 3,0, 10,0 Hz, Ha-6"), 3,69 (1H, dd, J
= 5,0, 10,0 Hz, Hb-6").
"C-NMR (125 MHz, CD3OD va DMSO-de) xem bang 1.
Catechin (2): C h i t r I n mau vang, nhiet dp nong chay 175-177°C.
ESI-MS: m/z 291[M+Hr (positive), 289 [M- H]" (negative), M= 290, CTPT C15H14OS.
^H-NMR (500 MHz, CD3OD) 5 (ppm): 4,59 (1H, 6,J = 7,5 Hz, H-2), 4,00 (1H, m, H-3), 2,53 (1H, dd, J = 8,0, 16,0 Hz, Ha-4), 2,87 (1H, dd, J
= 5,5, 16,0 Hz, Hb-4), 5,95 (1H, 6, J = 2,5 Hz, H- 6), 5,89 (1H, d,J = 2,5 Hz, H-8), 6,86 (1H, d, J = 1,5 Hz, H-2'), 6,74 (1H, dd, J = 1,5, 8,0 Hz, H- 5'), 6,78(1 H,d, J = 8,0 Hz, H-6').
"C-NMR (125 MHz, CD3OD) 5 (ppm):-82,87 (C-2), 68,84 (C-3), 28,53 (C-4), 156,93 (C-5), 96,35 (C-6), 157,85 (C-7), 95,56 (C-8), 157,60 (C-9), 100,87 (C-10), 132,24 (C-1'), 115,30 (C- 2'), 146,25 (C-3'), 146,25 (C-4'), 116,14 (C-5'), 120,08(0-6').
Glochidionolactone E (3): chat bpt khong mau, nhiet dp nong chay 125-126 °C.
Nghiert j £ - Ky thuat
ES! """ -^ •^ ' [ M + i f (positive), M=316, CTF' ,H2o".
i >HmF .500 MHz, CD3OD) 5 (ppm): 3,09 (1 5,62 (1H, d, J = 10,5 Hz, H-2), 6,08 (1H, ud, J = 1,0, 10,5 Hz, H-3), 4,43 (1H, m, H-4), 1,88 (1H, ddd, J = 3,0, 9,0, 11,0 Hz, Ha- 5), 2,57 (1H, m, Hb-5), 4,98 (1H, m, H-6), 2,32 (1H, dd, J = 3,0, 17,5 Hz, Ha-7), 2,89 (1H, dd, J
= 9,0, 17,5 Hz, Hb-7), 4,43 (1H, d, J = 7,5 Hz, H- 1'), 3,19 (1H, dd, J = 7,5, 9,0 Hz, H-2'), 3,31 (H- 3'), 3,30 (H-4'), 3,36 (H-5'), 3,69 (1H, dd, J = 5,5, 12,0 Hz, Ha-6'), 3,89 (1H, dd, J = 1,5, 12,0 Hz, Hb-6').
"C-NWIR (125 MHz, CD3OD) 5 (ppm): 35,79 (C-1), 128,79 (C-2), 132,45 (C-3), 70,93 (C-4), 32,02 (C-5), 79,32 (C-6), 36,36 (C-7), 179,00 (C-8), 103,48 (C-1'), 75,10 (C-2'), 77,98 (C-3'),
71,62 (C-4'), 78,07 (C-5'), 62,73 (C-6').
Benzyl-p-D-glucopyranoside (4): c h i t bpt kh6ng mSu, nhiet d6 nong chay 123-125°C, ESI- MS: m/z 271 [M+H]" (positive), M=270, CTPT CisHisOs.
^H-NIMR (500 MHz, CD3OD) 5 (ppm): 7,44 (2H, d,J = 7,5 Hz, H-2, H-6), 7,34 (2H, t, J = 7,5 Hz, H- 3, H-5), 7,28 (1H, t, J = 7,5 Hz, H-4), 4,70 (1H, d, J
= 12,0 Hz, Ha-7), 4,96 (1H, d, J = 12,0 Hz, Hb-7), 4,38 (1H, d, J = 7,5 Hz, H-1'), 3,25 -H 3,38 (4H, H- 2', H-3', H-4', H-5'), 3,71 (1H,dd, J = 12,0, 5,0 Hz, Ha-6'), 3,92 (1H, dd, J = 12,0, 2,5 Hz, Hb-6').
"C-NMR (125 MHz, CD3OD) 5 (ppm):
139,10 (C-1), 129,27 (C-2), 129,20 (C-3), 128,69 (C-4), 129,20 (C-5), 129,27 (C-6), 71,77 (C-7), 103,33 ( C - r ) , 75,17 (C-2'), 78,05 (C-3'), 71,75 (C-4'), 78,14 (C-5'), 62,85 (C-6').
OH 2
Hinh 1: Cau triic hda hoe cua 1-4 K § t q u a v a t h a o l u a n
Hgp chit 1 dygc phan lap dyo'i dang tinh the hinh kim, khong mau, nhiet dp nong chay 134 - 136°C. Tren pho ESI-MS x u l t hien pie ion vo'i cyo'ng dp cao tai m/z 407 [M+Hf, 405 [M- H]", phu hgp vo'i eong thirc phan tCr C20H22O9.
Pho ^H-NMR eua 1 (do trong DMSO-dg) x u l t hien cae tin hieu dae tryng cho 8 proton eua hai vong benzen 5 tir 6,90 -^ 7,49 ppm. Trong do eo 3 pie cong hyang tai 5 7,39 (2H, d, J = 8,0 Hz) va 7,49 (2H, [, J = 8,0 Hz), m i l pie tyang irng vo'i 2 proton va tin hieu triplet tai 5 7,48 (1H, t, J
= 8,0 Hz) ehirng to sy t i n tai eua mot vong benzen t h i mono. Ba tin hieu eong hyo'ng tai 5 7,05 (1H, d, J = 3,0 Hz, H-2), 7,14 (1H, d. J = 8,5 Hz, H-5), 6,90 (1H, dd, J = 3,0, 8,5 Hz, H-6)
khang djnh sy co mat cua mot vong benzen the 1,3,4. Ngoai ra, hai tin hieu tai 5 5,30 (2H, s, H- 7'), eung vo'i k i t qua phan tich pho DEPT va pho HSQC eho b i l t sy ton tai cua mot nhom oximethylen. Sy co mat cua mot phan ty dyo'ng glucose cung dyge xac djnh bo'i cae tin hieu tai 5 4,71 (1H, d, J = 7,5 Hz, H-1"), 3,25 (H-2"), 3,30 (H-3", H-5"), 3,16 (H-4"), 3,46 (1H, dd, J = 3,0, 10,0 Hz, Ha-6") va 3,69 (1H, dd, J = 5,0, 10,0 Hz, Hb-6"). Tren pho HSQC, hai tin hieu tai 5 3,46 va 3,69 d i u eo tyang tae vo'i carbon 5 60,81 ehirng to day la cae tin hieu cua nhom oximetylen. Tin hieu tai 5 4,71 vo'i hIng s i tyang tae J = 7,5 Hz eung ehirng to sy co mat eua lien k i t p-glyeosit. Tren pho "C-NWIR va DEPT cua 1 (do trong DMSO-dg) co 18 tin hieu
• N g h i e n
CLFU- Ky t h u a t
cua carbon, vo'i 2 tin hieu CH eo cyo'ng dp manh tai 5 = 127,94 (x 3) v^ 128,37 (x 2) 5 ehirng to vong benzen the mono. Ngoai eae tin hieu eua hai vong benzen [121,97 (C-1), 115,94 (C-2), 151,90 (C-3), 149,20 (C-4), 118,93 (C-5), 120,16 (C-6), 136,10 (C-1'), 127,94 (C-2', C-4', C-6'), 128,37 (C-3', 5')] va 6 tin hieu eua mot phan tii' dyo'ng glucose [5 102,41 (C-1"), 73,42 (C-2"), 77,09 (C-3"), 69,72 (C-4"), 76,39 (C-5"), 60,81 (C-6")] eon eo tin hieu eua carbon caboxylat tai 6 165,75 va carbon methylen mang oxi tai 5 66,19. Gia trj eao (5 66,19) ehirng to nhom oximetylen nay phai n i l vo'i carbon earbonyl qua e l u n i l este. De xac djnh vj tri eua eac nhom t h i tren vong benzoyl, pho HSQC va HMBC da dygc thyc hien. Tren pho HMBC, tyang tac cua proton cua nhom oximetylen tai 5 5,30 vai carbon tai 5 127,94 (CH x 2)1 136,10 (C)/165,75 (COO) khIng djnh nhom benzyl n i l vo'i vong benzoyl qua e l u este. Tren vong benzoyl, tin hieu doublet tai 5 7,05 (J = 3,0 Hz) tyang tac vo'i carbon tai 5 165,75 (C=O)/149,20 (C-4)/120,16 (C-6), tin hieu tai 5 6,90 (dd, J = 3,0, 8,5 Hz) tyang tac vo'i carbon tai 5 115,94 (C- 2)/149,20 (C-4) nhyng khong eo tyang tae vo'i C- 3 (5 151,90), trong khi do proton H-1" cua phan tir dyo'ng lai eo tyang tae HMBC vo'i C-4 (5 149,20) ehirng to vj tri eua hai carbon n l i vo'i oxi la C-3 va C-4, ding tho'i phan tir dyo-ng n l i vao C-4 eua vong benzoyl. Cae tyang tae HMBC khac eung dyge ehi ra tren hinh 2. Nhy vay, hgp chit 1 dygc xac djnh la benzyl 4-0-p-D- glucopyranosyl-3,4-dihydroxy-benzoat, day la chit mo'i dyge dat ten la gloehieriosid F. Hgp chit nay eo dang g i n gilng vo'i trichoearpin ''^', tuy nhien sy khac biet v l vj tri nhom the da lam thay doi kha manh dp djeh chuyen hoa hpe eua vong benzoyl. Trong tryang hgp eua trichoearpin (la), phan ty dyang noi vao C-2 bing lien k i t ete thi gia trj dp djeh chuyen hoa hpe eua C-1 thIp han nhilu (5 116,94) so vo'i hgp chat 1 (5
121,97 khi do trong DMSO-dg, 5 123,19 khi do trong CD3OD), dong tho'i gia trj carbon tyang irng vo'i proton co tin hieu doublet (J = 3,0 Hz) tren pho HSQC eua hgp chit 1 lai eo gia trj dp djeh ehuyin hoa hoc thIp han nhilu (5 115,94 khi do trong DMSO-dg, 5 117,62 khi do trong CD3OD) so vo'i gia tri eua carbon tyang irng eua hgp chit trichoearpin (5 120,26) '^^'.
Hinh 2: Mot sd twong tac HMBC (H-C) chu yiu cua 1
Hgp chit 2 thu dyge dyo'i dang c h i t rIn mau vang, co nhiet d6 nong chay 175-177°C.
Pho ESI-MS xult hien pie ion vo'i cyo'ng dp cao tai m/z 291 [M+H]" (positive) va 289 [M-H]"
(negative) tyang irng vo'i c6ng thirc phan tir C15H14O6. Pho MS ciing vo'i pho NMR co dang pho cua mot hgp chit flavon dae tryng. Pho ' ' H - NMR xult hien 5 tin hieu trong vung 5 5,89-^6,78 tyang irng vai tin hieu dae tryng ciia 5 proton eua vong benzen. Trong do co 3 proton cua mot vong benzen tyang tac theo he ABX tai 5 6,86 (d, J = 1,5 Hz), 6,74 (dd, J = 1,5, 8,0 Hz) va 6,78 (d, J = 8,0 Hz) dien hinh cho vong B the 1,3,4.
Hai tin hieu tai 5 5,95 va 5,89 {J = 2,5 Hz) phii hgp VO'I hai proton tai C-6 va C-8 eua vong A.
Ngoai ra, co hai tin hieu eua 2 proton tai 5 4,59 (d, J = 7,5 Hz) va 4,00 (dd, J = 7,5, 13,0) irng vo'i hai proton g i n vo'i carbon mang oxi; hai proton tai 5 2,35 (dd, J = 8,0, 16,0 Hz) va 2,87 (dd, J = 5,5, 16,0 Hz) thupc vao nhom -CH2- dyge xac dinh bing pho HSQC. Tren pho " C - NMR eua 2 x u l t hien tin hieu eua 15 carbon, trong do eo 12 carbon vo'i gia tri 5 tir 96,35 -^
157,85 tyang irng vo'i hai vong benzen. Tir eac dir kien pho tren eho phep ta xac djnh hgp chit 2 la mot flavan. Trong do, vong A eo 2 nhom t h i - O H tai vj tri C-5 va C-7 (5 156,93 va 157,85);
vong B eung co 2 nhom the - O H tai vj tri C-3' va C-4' (5 146,25 vS 146,25); vong C chira nh6m the - O H tai vj tri C-3 vo'i gia tri 5 la 68,84 ppm.
Cae dir kien pho NMR eua 2 hoan toan phii hgp VO'I eac dir kien da e6ng b l cho hgp chit (+)- eateehin •^"•''^••'^i. Gia trj dp djeh chuyen hoa hpe eua proton va hing so tyang tae eua eae proton tai vj tri C-2, C-3, va C-4 eua hgp ehit 2 [4,59 (d, J = 7,5 Hz, H-2), 4,00 (1H, m, H-3), 2,53 (dd, J = 8,0, 16,0 Hz, Ha-4), 2,87 (dd, J = 5,5, 16,0 Hz, Hb-4)] hoan toan phii hgp vai cac gia trj tyang irng da eong b l eho (+)-eateehin [4,55 (d, J = 6,7 Hz, H-2), 3,99 (m, H-3), 2,53 (dd, J = 8,4, 16,1 Hz, Ha-4), 2,91 (dd, J = 5,5, 16,1 Hz, Hb-4) '"'; 4,59 (d, J = 8,5 Hz, H-2), 4,02 (m, H-
Nghien c u - Ky thuat
3), 2,56 (6^: J= "' 5, 16,0 Hz, Ha-4), 2,91 (dd, J = 5,7, lo, iZ, Hb \ D i l u do ehirng to proton H-2 va H-3 nam 6- vj tri trans vo'i nhau. Trong tryo-ng h; • -'-2 va H-3 n I m 6' vj tri cis vo'i nhau nhy epicaiccnin thi proton H-2 eo gia trj 8 eao han (4,88 ppm) va lai x u l t hien dyai dang singlet, ding tho'i hIng s i tyang tae JH-3/H-4 CO
Bang 1 : S6 lieu phd NMR ciia 1-4
gia tri 4,6 Hz va 3,2 Hz, t h i p han nhieu so vc.
[ I l l
tryo-ng hgp eua (+)-eatechin (8,5 va 6,7 Hz) Catechin dyge biet d i n nhy mot thanh phan then c h i t cua tannin eo trong che xanh, sy co mat cua catechin da d y g c ehirng minh la mang lai tae dung chIng oxi hoa hieu qua eiia ehe xanh.
c
1 2 3 4 5 6 7 8 9 10 C=0
r
2' 3' 4' 5' 6' 7' 1"
2"
3"
4"
5"
6"
la Sc*
116,94 152,18 120,26 122,60 150,18 121,38
168,19 136,10 128,59 129,08 128,88 129,08 128,59 67,47 105,53 75,47 77,80 71,33 78,04 61,52
1 5c''' 123,19 117,62 154,08 151,88 121,56 121,87
167,87 137,39 129,28 129,62 129,32 129,62 129,28 68,04 105,32
75,05 78,43 71,39 77,60 62,70
8c''' 121,97 115,94 151,90 149,20 118,93 120,16
165,75 136,10 127,94 128,37 127,94 128,37 127,94 66,19 102,41
73,42 77,09 69,72 76,39 60,81
5c®
82,0 67,9 28,1 156,3
96,7 156,9
95,7 156,7 100,9
131,5 115,6 145,4 145,5 116,6 120,3
2 Sc''"
82,87 68,84 28,53 156,93
96,35 157,85
95,56 157,60 100,87
132,24 115,30 146,25 146,25 116,14 120,08
Sc' 35,6 128,8 132,5 71,0 32,1 79,3 36,4 179,0
103,5 75,2 78,0 71,7 78,2 62,8
3 Sc''' 35,79 128,79 132,45 70,93 32,02 79,32 36,36 179,00
103,48 75,10 77,98 71,62 78,07 62,73
5c 138,0 129,6 129,5 129,2 129,5 129,6 72,2
102,5 74,4 77,2 70,9 77,2 62,0
4 Sc''' 139,10 129,27 129,20 128,69 129,20 129,27 71,77
103,33 75,17 78,05 71,75 78,14 62,85
"Do trong CD3OD, "125 MHz, "Do trong DMSO-de, ® 5c ciia catechin [do trong (CD3)2CO)'''°'], *5c cua trichoearpin [do trong (CD3)2CO)'^^'], *8c ciia gloehidionolaeton E (CDsOD)'^"', * 5c ciia benzyl-j3-D-glucopyranosid (do trong CD3OD) '^^'.
proton ggi y trong phan ty eiia 3 co mot phan tCr dyo'ng. Ngoai ra eo hai gia trj cong hyo'ng cua 2 proton tai 5 4,43 (m) va 4,98 (m) cho bilt trong elu tao cua 3 CO 2 nhom oximetin (CH-0). Tren pho
"C-NMR va DEPT cho bilt sy eo mat cua 14C trong do co 1 C bae 4, 10 carbon CH va 3 carbon CH2. Gia trj eong hyo'ng ciia carbon bae 4 tai 6 170 ppm eho phep khang djnh sy co mat eua mot nhom earbonyl C=0. Hai nhom CH vo'i 5 la 128,79 va 132,45 ggi y sy ton tai cua mpt lien k i t doi Hgp chit 3 phan lap dygc dyo'i dang chit bpt
mau trIng, nhiet dp nong chay 125-126 °C. Tren pho khIi lygng ESI-MS xult hien pie ion co cyo'ng dp lo'n tai gia trj m/z 317 [M+H]* (positive), phii hgp VO'I c6ng thirc phan tCr la Ci4H2o08- Tren pho ' H - N M R eiia 3 xult hien tin hieu eong hyo'ng eua 2 proton olefin tai 5 5,62 (1H, d, J = 10,5 Hz), 6,08 (1H, dd, J = 1,0, 10,5 Hz) va 1 proton anome tai 4,43 (d, J - 7,5 Hz). Vo'i eae gia trj eong hyo'ng 5 trong viing tir 3,30 ^ 4,43 ppm tyang irng vo'i 7
• N g h i e n
CLFU- Ky t h u a t
CH=CH. Ngoai ra, 6 tin hieu carbon eong hyo'ng tir 5 62,73 d i n 103,48 ppm cijng vo'i sy g i n k i t VO'I cac proton dae tryng eua dyo'ng (dygc khing dinh thong qua pho HSQC) cho ta k i t luan phan tir 3 CO g i n mot dyo'ng p-D-glueoza. Phan tich eae pho ^ H - N M R , ' ^ C - N M R va DEPT khing djnh dyge CO 2 nhom oximetin CH-0 tai gia trj 5 70,93 va 79,32. Tir viec phan tich cae dii' kien pho neu tren ta CO the ghep n l i eae manh e l u true de xay dyng nen elu true eua 3 (Hinh 1) va dygc kiem chirng bing p h i HMBC. Tren pho HMBC, proton H-1' (5 4,43, d, J = 7,5) tyang tae vo'i C-4 (5 70,93) va C-5' (8 78,07); hai proton H-7 (8 2,32 va 2,89) d i u tyang tac VO'I C-1 (35,79), C-2 (128,79), C-6 (79,32) va C- 8 (179,00); proton H ^ (8 4,43, m) tyang tae vo'i C-2 (128,79), C-3 (132,45), C-6 (79,32) va C-1' (103,48). NhCrng tyang tae neu tren hoan toan phu hgp VO'I elu triic da dy doan eua hgp chit 3. T i t ea eac gia trj pho NMR cua 3 deu phii hgp vo'i cae gia trj tyang irng ciia chit gloehidionolaeton E, mot hgp chit da bilt tir chi Glochldion '^'''.
Hgp chit 4 thu dyge la tinh the hinh kim, khong mau, nhiet dp nong chay 123-125°C, pho' ESI-MS CO pie m/z 271 [M+H]* (positive) tyang irng vo'i eong thirc phan tCr dsHisOs. Pho ^H- N M R xult hien eae tin hieu trong vijng 8 7,28 -^
7,44 (5H), trong do eo 2 tin hieu co cyo'ng dp manh tyang irng vo'i 2 cap proton doi xirng, dieu nay khing djnh s y eo mat eua 1 vong benzen t h i mono. Ngoai ra con eo sy eong hyo'ng ciia eae proton eua mot phan tir dyo'ng (8 3,25 + 4,38, 7H), vo'i tin dae tryng tai 8 4,38 (1H, d, J = 7,5 Hz) eua proton anome. Hai proton con lai vo'i 8 4,70 (1H, d, J = 12,0 Hz) va 8 4,96 (1H, d, J = 12,0 Hz) tyang irng vo'i mot nhom oximetilen. Phan tich pho " C - N M R va DEPT eho thiy eo 11 tin hieu eong hyang eua carbon, trong do eo 2 tin hieu eua CH vo'i cyang dp manh tai 8 129,27 va 129,20 xac nhan cho 2 cap carbon d l i xirng cua vong benzen tai C-2, C-6 va C-3, C-5. Mot tin hieu eong hyo'ng eua CH tai 8 128,69 tyang irng vo'i C-4, tin hieu eon lai eua 1 carbon bae b i n 8 139,10 la eua C-1 ciia vong benzen co g i n nhom t h i . Sy co mat eua 1 nhom CH vai dp djeh chuyen hoa hpe 8 103,33 va 1 nhom -CH2- tai 8 62,85, eiing vo'i 4 nhom CH tir 75,17 -^ 78,14, va k i t hgp vo'i cac gia trj proton dae tryng eho vung dyo'ng noi tren eho phep khing djnh trong phan tir eua 4 eo g i n mot dyo'ng j3-D-glueose. Tin hieu djeh ehuyin manh v l phia tryo'ng t h i p han eiia
nhom oximetylen tai 8c = 71,77 ppm, cung vo'i tin eae hieu 8H 4,70 (1H, d, J = 12,0 Hz) va 8 4,96 (1H, d, J = 12,0 Hz) eho phep khing djnh sy CO mat eua nhom oximethyl va lien k i t ether dyge tao nen giCra nhom nay vo'i nhom benzyl.
Tir eac dii' kien pho neu tren, hgp c h i t 4 dyge xac nhan la benzyl-/3-D-glueopyranosid''^' mot hgp c h i t da bilt tir loai Epimedium grandiflorum var. thunbergianum.
K§t luan
Bing eae phyang phap s i c ky k i t hgp, b i n hgp c h i t benzyl 4-0-P-D-glucopyranosyl-3,4- dihydroxy-benzoat (gloehieriosid F, 1), eateehin (2), gloehidionolaeton E (3), benzyl-p-D- glueopyranosid (4) da dyge phan lap tir la eay bon bpt (Glochldion eriocarpum). C i u triic hoa hpe eua ehiing dyge xac djnh b i n g cae phyang phap pho khIi lygng, pho cong hyo'ng tir hat nhan mot ehilu va hai ehilu. Trong do, hgp c h i t 4-0-P-D-glueopyranosyl-3,4-dihydroxy-benzoat (1) la mot chit mo'i l l n d i u tien d y g e phan lap tir thien nhien. Cae hgp c h i t gloehidionolaeton E va benzyl-(3-D-glueopyranosid d y g c phan lap l l n d i u tien tir eay bon bpt
L&l cam an: Cac tac gia chan thanh cam on TS Dwong Dire Huyin, TS Ninh Khie Ban, TS Ha Van Tui, ThS Nguyen Thi Cwong, Vien Sinh thai va Tai nguyen sinh da giup xac dinh ten khoa hoe eiia eay.
Summary
From the aerial parts of Glochldion eriocarpum, a new phenolic glycoside (gloehierioside F, 1) and three known compounds were isolated by various chromatography methods. Their structures were identified as benzyl 4-0-P-D-glucopyranosyi-3,4- dihydroxy-benzoate (1), catechin (2), glochidionolactone E (3), benzyl-^D-glucopyranoside (4) from the analysis of ESI-MS, 1D- and 2D-NMR spectra in comparison with the literature values.
Tai lieu tham khao
1. Do Huy Bich va cong sy. Cay thulc va dong vat lam thuoe a Viet Nam, Nha xuit ban Khoa hoc va Ky thuat, Tap 1, tr. 224-225, (2004)
2. Do Tat Lai. NhOng cay thulc va vi thulc Viet Nam, WXeV hoc, (2001)
3. V5 Van Chi. Tir dien cay thuoe Viet Nam. NXB Yhoc, Ha Noi, Xr. 47:, (1999)
4. Le Kieu Nhl, Nguyen Van Dau, Phan Tong San,. Gop phan nghien ciru hoa hoc cay bon bot {Glochldion eriocarpum Champ., Euphorbiaceae), Tap chi Dwac hoc, T. 39, S6 12, tr. 9-10 (1999)
(Xem tiep trang 14)
• Nghien c t u - Ky thuat
khau va thIp hen i o vai mire biln dpng chi s i hang hoa tieu duug (CPI).
K § t l u a n
Trong khoang thai gian tiF thang 5 nam 2005 d i n thang 2 nam 2007, gia thulc san x u l t trong nyac tieu thu tai cac nha thulc a Ha Noi tyang d l i on djnh. K l tir thang 3 nam 2007 d i n thang 5 nam 2008 gia thulc san x u l t trong nyac eo sy biln dpng theo ehilu hyang tang len vai bien dp dao dpng khoang 2,%. Tuy nhien, s y biln dpng do vSn thIp han s y biln dpng cua chi s i hang hoa tieu dung.
S u m m a r y
This study was conducted in 10 pharmacies in the area of Hanoi from June 2005 to June 2008.
Data is collected, updated daily at 10 pharmacies.
Research analyst price fluctuation of some drugs produced domestically and analysis process by using statistical methods and then compared with the consumer price index (CPI).
In the period from May 2005 to March 2007, the price increases domestic production sold in pharmacies in Hanoi is relatively stable. Drugs
in 2007 domestic production is highly vanable tended to increase with the amplitude venation
of about 3%. 2008 price increases domestic production is highly variable increases. Although
the price increases domestic production is highly vanable with the increasing but still lower than the fluctuation of the consumer price index.
T a i lieu t h a m k h a o
1. Nguyin Thanh Binh, Vu Nang Thoa, Phan tich sy bien dpng gia thuoe nhap khau tren dja ban Ha NQI, Tap chi Diegc hoc, so l i /2009, (2009)
2. Cue quan ly Du'ac - Bo Y te. Hoi nghi tong kgt eong tae 6\yac nam 2008 va 6 thang d§u nam 2005 - Then khai eae djnh hirang chien lu'ac phat trien nganh dygc Viet Nam giai doan din 2010 va tarn nhin 2015 (2009)
3. Cue quan ly Dygc - Bo Y t l , Bao eao t6ng kgt Du'gc nam 2008 trien khai ke hoach nam 2009, ngay 23/6/2008 (2008)
4. Tryang Quoc Cyang, Danh gia nang lye eac doanh nghiep san xuat Dygc pham trong nu'ac hien nay, Thdng tin dwac so 1/2005, (2005)
5. Tir Minh Koong, Nguyin Thanh Binh, Sy bien dong v l gia ciia thuSe nhap ngoai tren thj tryang va nhij'ng ylu t l anh hyang. Tap chi Thdng tin Y Dwac, s6 11/2004 (2004)
6. Vien Khoa hpe Thong ke. Mot s6 vIn de phu'ang phap luan thong ke, Nha xuit ban Khoa hoc Thong ke (2005)
Glochierlosid F... (TiSp trang 55)
5. Puapairoj P., Naengehomnong W., Kijjoa A., Pinto M. M., Pedro M., Nascimento M. S., Silva A. M., Herz W., Planta Med., Vol. 71, 208-213 (2005)
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Phannaceutical Bulletin, Vol. 57 (1), 102-105, (2009) 7. Vu Kim Thu, Phan Van Kiem, Pham Hai Yen, Nguyen Xuan Nhiem, Nguyen Huu Tung, Nguyen Xuan Cuong, Chau Van Minh, Hoang Thanh Huong, Thnh Van Lau, Ngo Thi Thuan, and Young Ho Kim' Triterpenes and triterpenoid saponins from aerial parts of Glochldion eriocarpum. Natural Product Communications, 5(3) (dang in) (2010)
8. Vu Kim Thu', Phan Van Kiem, Pham Hai Yin, Hoang Thanh Hu'ang, Hoang Thi My Nhung, Nguyen Thj Quy, Nguyin Xuan Cu'ang, Nguyen Hoai Nam, Chau Van Minh. Cac hgp ch§t lupan tir cay bon bot {Glochldion eriocarpum), Tap chi Dwac hoc, S6 405, Tap50,tr. 39-43, (2010)
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Huong. A new flavan glucoside from Glochldion eriocarpum. Tap chi Hoa hoc (dang in) (2010)
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14. Hideaki Otsuka, Eiji Hirata, Anki Takushi, Takakazu Shinzato, Yoshio Takeda, Masahiko Bando and Masaru Kido, Glochidionolaetones A-F : Butenolide glueosides from leaves of Glochldion zeylanicum (GAERTN) A. JUSS, Chem. Phami. Bull., Vol. 48(4), 547-551,(2000)
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