• N g h i e n C L F U - K y t h u a t
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P h a n l a p v a x a c d j n h c a u t r u e n e r i a s i d v a a c i d o l e a n o l i c t i r l a t r u e d a o
Dat van de
Cay true dao cdn goi Id dao Id, gidp true ddo, tdn khoa hpc Id Nerium oleander L. thupc hp True ddo Apocynaceae. Ld loai cdy nho song ldu ndm, hoa mdu hong hay trdng, thudng dupe trong Idm canh. Vd thanh phan hoa hpc, cdy triic ddo chira mpt so glycosid dpc, ngodi ra cdn chira nhua, tannin, mpt so loai paraffin, vitamin C vd tinh dau. Cac bp phan cua cdy true ddo cd tdc dung bo tim, sdt trung, lpi tidu, thudng dirpc diing dd chua suy tdm thu, viem co tim ndo suy (loan nhip nhanh), tri b$nh ngodi da Id ngda, mun lodt, dung gidp | 1 , 2l Tuy nhidn, nhdng hpp chdt ndy cP thd gdy td vong d ngirdi, ddc biet Id
HCO 3
Le Anh Hao, Nguyen Tien Virng Vien Phdp y Quoc gia tre em. Dpc tinh cua triic dao rat manh, co nhieu thdng bao cho thdy trong mpt so truPng hpp chi can mpt lupng nho cung da dii gay hdu qua tu vong hay can kd tu vong. De dap ung giam dinh hod phdp vd cong tdc cap cdu nan nhan ngp dpc cdy triic ddo tai cdc trung tam chong dpc can phai tao cac chat chuan, phuc vu cho cdc phuong phdp phdn tich, phdt hidn chat dpc tu cdy triic dao..
Tu phdn Id cdy triic ddo chiing toi da phdn Idp vd xdc djnh dupe cdu triic ciia 2 hpp chat Id neriasid (1) vd acid oleanolic (2) (hinh 1). Chat 1 cd cau triic Id mpt cardenolid glycosid la nhom chat cP kha ndng gdy dpc rat manh.
25 " 13 17
26 1 1 4 . 1ft 'COOH
Hinh 1: Cdu true hda hoc cua hop chit 1 vd 2 Nguyen lieu va p h u a n g p h a p
Nguyen lieu
Mau Id cdy triic ddo dupe thu hdi tai Hd Npi vdo thdng 6/2010. Mdu cdy trdn dupe GS.Phan Ke LSc xdc djnh Id Nerium oleander L , thupc hp Triic ddo.
Hda chit thiit bj:
Sic ky Idp mdng (TLC): Sic ky Idp mong dupe thuc hidn trdn ban mong trdng sin DC- Alufolien 60 F254 (Merck 1,05715), RP 18
F254s (Merck). Phdt hidn chit bing ddn tu ngoai d hai budc sdng 254nm vd 368nm hodc diing thuoc thu Id dung djch H2S04 10% dupe phun deu ldn ban mdng, say khd rdi ho nPng tu tu din khi hidn mdu.
Sic ky cdt (CC): Sic ky cdt dupe tien hdnh vdi chit hip phu Id silica gel pha thudng vd pha dao. Sillicagel pha thudng cd cd hat Id 0,040- 0,063mm (240-430 mesh). Sillica gel pha dao YMC RP18 (30-50 um, FuJisilisa Chemical Ltd.).
TAPCHi DUQC HQC-5/2011 (SO 421 NAM 51) 39
• N g h i e n C L P U - K y t h u a t Phd cdng hwdng tw hat nhan (NMR): dirge ghi bing may Bruker AM500 FT-NMR Spectrometer voi chit chuan npi la TMS.
Phd khdi Iwong (ESI-MS): dirge do tren he may AGILENT 1200 series LC-MSD Ion Trap.
Chiit va phan lap: Phan la cua cay true dao sau khi thu hai ve dugc phoi kho trong bong ram, xay nho (3,5 kg) va ngam chiit voi methanol (10 I x 3 lan) bing thiet bj sieu am Ultrasonic 2010 d nhiet do 40-50°C (3 x 60 phut). Dich chiit sau do dugc co ddc bing may cat quay voi ap suit giam thu dugc 300g can chiet methanol. Cdn chiet nay dugc hda vdi nugc roi chiet phan doan lin lugt vdi n-hexan, clorofom va ethyl acetat thu dugc cac cdn chiit n-hexan (60g), clorofom (100g) va ethylaxetat (50g) tuang uhg. Tien hanh sic ky cot silica gel doi vai can chiet clorofom rua giai bing he dung moi gradient n-hexan-aceton (100:1v/v —>100%
aceton) thu dugc 6 phan doan ky hieu la C1—>
C6. Tu phan doan C3 tien hanh sic ky cdt silica gel (n-hexan- aceton 3:1) thu dugc hgp chit 2 (12 mg).
Phin cdn chiit EtOAc dugc tien hanh sic ky cot YMC RP18 vai he dung moi rua giai MeOH- nugc 1:3 thu dugc 3 phan doan ky hidu la E1 -»
E3. Tien hanh phan ldp phdn doan E1 (10 g) tren sic ky cot silica gel, hd dung mdi CHCI3- MeOH-nugc 5:1:0.1 thu dugc hgp chit 1 (8mg).
Hgp chit 1: Neriasid
Dang bdt rin mdu tring, CTPT C3oH4608, KLPT M=534, ESI-MS m/z 557 [M+Na], 1H-NMR (500 MHz, CDCI3), 5 (ppm): 5.82 (1H, d, J=1,5, H-22), 4,69 (1H, dd, J=1,5, 17,5, H-21a), 4,79 (1H, dd, 2,0, 17,5, H-21B), 4,48 (1H, dd, J=2,0, 10,0, H-1'), 4,13 br s (1H, H-3), 3,69 brs (1H, H- 4'), 3,42 (1H, q, J=6,5, H-5') 3,40s (3H, 3'- OCH3), 3,32 (1H, ddd, J=2,0, 5,0, 12,0, H-3'), 2,86 (1H, t, J=9,0, H-17), 1,34 (3H, d, J=6,5, H- 6'), 1,00s (3H, H-19), 0,84s (3H, H-18), "C-NMR (125 MHz, CDCI3), 8 (ppm): 30,8 (C-1), 27,2 (C- 2), 72,6 (C-3), 30,3 (C-4), 36,1 (C-5), 29,5 (C-6), 32,0 (C-7), 216,3 (C-8), 51,4 (C-9), 42,5 (C-10), 17,5 (C-11), 34,7 (C-12), 50,9 (C-13), 77,9 (C- 14), 37,8 (C-15), 28,3 (C-16), 45,8 (C-17), 16,7 (C-18), 23,9 (C-19), 171,4 (C-20), 73,5 (C-21), 116,7 (C-22), 173,9 (C-23), 98,49 (C-1'), 32,0 (C-2'), 78,0 (C-3'), 67,1 (C-4'), 70,5 (C-5'), 16,8 (C-61), 55,7 (C-3'-OMe).
Hgp chit 2: Acid oleanolic
Dgng tinh the, mdu triing, nhidt do ndng chay 306-308°C, CTPT C30H48O3, KLPT M=456, ESI- MS m/z 457 [M+Hf. 1H-NMR (500 MHz, CDCI3), 8 (ppm): 5,23 (1H, m, H-12), 3,15 (1H, m, H-3), 0,77, 0,81, 0,90, 0,93, 0,94, 0,97 va 1,15 (3H cho mdi vj tri, s, H-24, 25, 26,27,29,30), 1 3C- NMR (125 MHz, CDCI3), 8 (ppm): 39,8 (C-1), 27,8 (C-2), 79,7 (C-3), 38,1 (C-4), 56,7 (C-5), 19,5 (C-6), 33,8 (C-7), 40,5 (C-8), 49,8 (C-9), 38,1 (C-10), 24,0 (C-11), 123,6 (C-12), 145,2 (C-13), 42,7 (C-14), 28,8 (C-15), 24,5 (C-16), 47,6 (C-17), 42,9 (C-18), 47,2 (C-19), 31,6 (C- 20), 34,9 (C-21), 34,0 (C-22), 28,7 (C-23), 16,0 (C-24), 15,8 (C-25), 17,7 (C-26), 26,4 (C-27), 181,8 (C-28), 33,5 (C-29), 23,9 (C-30).
Ket qua va t h a o l u a n
Pho 1H-NMR cua 1 xuit hien tin hidu cua mdt proton olefin duy nhat dudi dang dan bdi tai 8 5,88 chung to su cd mdt cua mot noi ddi thi 3 vi tri, ngodi cdc tin hidu dien hinh cua mdt phdn tu dudng tai 8 tgi 8 4,48 dd (1H, dd, J=2,0, 10,0 Hz, H-1'), 1,94 (1H, H-2'a), 1,69 (1H, H-2'p), 3,34 (1H, ddd, J=3,0, 5,0, 12,0 Hz, H-3'), 3,69 (1H, br s, H-4'), 3,42 (1H, q, J=6,5 Hz, H-5'), 1,34 (3H, d, J=6,5 Hz, H-6'), 3,40 (3H, s, 3'- OCH3) cdn xuit hien tin hidu cua 2 nhdm methyl singlet (8 0,84, 1,00), mdt nhdm methylen noi vdi oxy d 8 4,70 (1H, dd, J=1,0, 17,5, H-21<x) vd 8 4,88 (1H, dd, J=2,0, 17,5 Hz, H-2ip).
Pho 13C-NMR vd DEPT cua 1 xuit hidn tin hidu cua 30 vach tin hidu gom 3 methyl, 1 methoxy, 11 methylen, 10 methyl vd 5 nhdm carbon bde 4. Cdc tin hidu pho tai 8 98,49 (C-1'), 32,0 (C-2'), 78,0 (C-3'), 67,1 (C-4'), 70,5 (C-5'), 16,8 (C-6'), 55,7 (OMe-C-31) cua nhdm dudng phii hgp vdi so lidu cua dudng B-D- diginopyranosid rit pho bien trong cdc lodi thuQc chi Nerium | 3l Trong so 23 tin hidu cdn lai, cdc tin hidu gom mOt nhdm carbonyl d 8 216,6, mQt hd carboxyl a, (3 khdng no tgi 8 173,9 (CO), 171.4(C) vd 116,7(CH), hai carbon noi vdi oxi tai 72,6 (C-3) vd 77,9(C-14) cho phdp du dodn hgp chit 1 cd dang khung cardenolides vdi vide lidn ket giua C8 vd C14 bi dut[ 4 ). So sdnh cdc du kidn ndy vd kit hgp vdi pho khoi lugng ESI-MS vdi su cd mdt ciia pic m/z 557 [M+Na] tuang ung vdi khoi lugng phdn tu M=534 vd CTPT C3oH4608 dd khdng dinh hgp chit 1 Id
40 TAP CHI DUQC HQC-5/2011 (S6421 NAM 51)
• N g h i e n C U H J - K y t h u a t neriagenin-3-0-/?-D-diginopyranosid, hay ten khac la neriasid '4 , S |.
Ngoai ra m$t hpp chat triterpen khac la acid oleanolic (2) cung 6a dupe phan ldp. Tren pho
1H-NMR cua 2 xuat hidn 7 tin hieu don bpi cua 7 nhPm methyl tai 8 0,77, 0,81, 0,90, 0,93, 0,94, 0,97 va 1,15. Pho 13C-NMR va DEPT cua 2 cho thay su1 cP mdt ciia 30 tin hieu gom 7 methyl, 10 methylen, 5 methin va 8 nhPm carbon bde 4, dieu nay cho phep khing dinh day la mpt triterpen 5 vPng cP dang khung olean. Pho khoi lupng ESI-MS vdi sy cP mat cua pic m/z 457 [M+H]+ chung to 2 cP khoi lupng phan tu la 456.
Cac du kidn pho nay hoan toan trung khdp vdi so li$u da cPng bo cua acid oleanolic[6].
Ket luan
Nhu v$y tu la true dao chung tdi da phan ldp dupe 2 chit Ciu true cua 2 chit dupe xac djnh Id neriasid vd acid oleanolic dua trdn co sd phdn tich cdc pho 1H-NMR, 13C-NMR, HMBC, COSY, MS, trdn co sd so sdnh vdi cdc du lidu dd cdng bo.
S u m m a r y
From the leaves of Nerium oleander growing in Vietnam, a cardenolide glycoside and an
olean triterpene were isolated and spectroscopically identified to be neriaside (1) and oleanolic acid (2). Of these, neriaside (1) belongs to the toxic group of cardenolides.
Key words: Nerium oleander, cardenolid, neriasid, acid oleanolic
Tai lieu t h a m khao
1. Do Tat Lei. Nhdng cay thudc va vi thuoc Viet Nam, Nha xuat ban Y hoc, 2000, trang 586.
2. V6 Van Chi, Td dien cay thuoc Viet Nam, Nha xuat ban Y hoc, 2000, trang 1276.
3. Liming Bai, Ming Zhao, Asami Toki, Jun-ichi Sakai, Xiao-yang Yang, Yuhua Bai, Mariko Ando, Katsutoshi Hirose, and Masayoshi Ando. Three New Cardenolides from Methanol Extract of Stems and Twigs of Nerium oleander. Chem. Pharm. Bull. (2010) 58(8) 1088-1092.
4. Fumiko Abe and Tatsuo Yamauchi.
Cardenolide triosides of oleander leaves.
Phytochemistry, Vol. 31 (1992), No. 7.pp. 2459-2463.
5. Tatsuo Yamauchi and Fumiko Abe. Neriaside, A 8,14-seco-cardenolide in Nerium odorum.
Tetrahedron Letters No. 1978, 21, pp. 1825-1828.
6. M. Maillard, C. O. Adewunmi and K.
Hostettmann. A triterpene glycoside from the fruits of Tetrapleura tetraptera. Phytochemistry, 1992, Vol. 31, No. 4, pp. 1321-1323.
N g h i e n c i r u t o n g h o p t h u o c d i e u t r j u n g t h u * a l t r e t a m i n s i r d u n g x i i c t a c z e o l i t
v a k y t h u a t v i s o n g Dinh Thi Thanh Hai, Dinh Thj Hai, Tran Tuan Hiep
Bo mdn Hod hiru co, Trudng Dgi hoc Duqc Hd Noi Dat van de
Altretamin (Hexamethylmelamin) Id thuoc chong ung thu dd dupe su dgng trong Y hpc de diiu trj ung thu phoi, ung thu buong trung, ung thu vu, Id cdc loai ung thu phi biin nhit. Loai thuic ndy hidn dang dupe su dgng trong dieu tri ung thu vdi cdc bidt dupe nhu Hexastat (Phdp), Hexinawas (Tdy Ban Nha), Altretamin (My), Hexalen (US Bioscience - My) v.v...'1'
Tiep tgc hudng nghidn cuu tong hpp altretamin Idm thuoc dieu trj ung thu. Dua trdn kit qua cdng b i cua nhieu cdng trinh nghidn cuu trdn the gidi vd trong nude ve ung dgng r$ng rdi cua xiic tdc zeolit trong tong hpp huu
co I 2'. Tiip theo nhdng thdng bdo trudc ddy cua
nhdm chiing tdi ve kit qua nghidn cdu tong hpp thuic dieu trj ung thu altretamin13,41, trong cdng trinh ndy chiing tdi cdng b6 kit qua nghidn