Tap chi Khoa hgc Truong Dai hgc Cdn Tha Phdn A" Khoa hoc Ty nhiin, Cong nghi vd Mdi Iru&ng: 26 (2013): 1-5
Tap chf Khoa hpc Trifcing Da'\ hoc Can Thd website: sj.ctu.edu.vn
G O P P H A N K H A O S A T T H A N H P H A N H O A H O C C U A V O C A Y M A M 6 I {AVICENNIA MARINA)
Le Thanh Phudc va Ldm Thiiy Phuong' ' Khoa Khoa hgc Tu nhien, Trudng Dgi hoc Cdn Tho Thdng tin chung:
Ngdy nhgn: 14/09/2012 Ngdy chdp nhan: 19/06/2013
Title:
Contribution to the study on the chemical components ofAvicennia marina bark
Tit khda:
Vo cdy Mdm oi Avicennia marina.
thdnh phdn hda hgc, taraxerone, taraxerol, betulin
Keywords:
Avicennia marina bark, chemical components, taraxerone, taraxerol, betulin
ABSTRACT
From the petroleum ether extracts of the bark ofAvicennia Marina, collected in the coast of Bac Lieu province, three compounds have isolated: taraxerol (CsoHsoO), taraxerone (CsoHjgO), betulin (CsoHsoO^) The structures of these compounds have been elucidated by modem spectroscopic methods such as- H NMR, C NMR, DEPT NMR and by comparison with those of previously reported data.
T O M T A T
Tie dich chiet petroleum ether cita vo cdy Mam oi, thu hdi tgi ven bien tinh Bac Lieu, da co Igp dugc ba hgp chdt Id.- taraxerol (C30H50O), taraxerone (C^oHjsO), betulin (C30HS0O2). Cdu true hda hoc cua cdc hap chdt ndy dd dugc xdc dfnh bdng cdc phucmg phdp phd hiin dgi 'H-NMR, '^C-NMR, DEPT NMR va du(?c so sdnh vdi tdi lieu da cong bo
1 D A T V A N D E
Cay Mam di cd tdn khoa hgc la Avicennia marina, thudc hg Cd roi ngua (Verbenaceae) (Pham Hoang Hd, 2003). Cay dugc ttdng hoac mgc hoang d viing nude man hay nude lg, gap d ca hai mien nude ta. Theo mdt sd tai lieu vd y hgc nhdn gian tten the gidi, cdy Mam di la ngudn dugc lieu cd gia tti chiia benh. Vd cdy ciia loai ndy da dugc sii dung ttong y hgc cd tmyen d Ai Cap dd didu tri cdc benh ve da, thdp khdp, bdnh ddu miia, loet. (W. M.
Bandaranayake, 2002). Ngodi ra, nhimg kdt qua nghien cuu vd boat tfnh sinh hgc eiia cay Avicennia marina da khang dinh boat tfnh chong sdt ret, ddc td bao, ddc biet la hoat tinh gay ddc te bao ung thu va chdng khdi u cung da dugc ghi nhdn (M. Sharaf e? al, 2000). Mac
dii, d nude ta cay Mam di da duge sir dyng rdng rai d nhieu nai vdi myc dich chiia bdnh nhung chua cd nghien cim ndo vd thanh phdn hda hgc va hoat tfnh sinh hgc ciia loai cdy ndy.
Dd dd, chiing tdi da thuc hien de tai: "Khao sat thanh phdn hda hge ciia vd cdy Mdm di {Avicennia marina)". Trong bai bdo nay chung tdi da phan lap vd xdc dinh duge cau tnic hda hge ciia cdc hgp chat taraxerol, tarexerone va betulin.
2 PHU'ONG T I | : N V A PHU'ONG P H A P Nguyen lieu: Vd eay Mam di dugc thu hai tai xa Dinh Thanh, huyen Ddng Hai, tinh Bac Lieu, sau do riia sach, cdt nhd, phoi kho.
Phuong p h a p : Chiet boat chdt: Vd cdy Mam dugc ngdm ttong cdn ethanol 96'*, phdn
Tgp chi Khoa hgc Tru&ng Bgi hge Cdn Tha Phdn A: Khoa hgc Ty nhiin, Cong nghi vd Mdi irudng: 26(2013)-1-5 dich chidt cd quay loai dung mdi thu dugc cao
con. Sau dd lay eao cdn chidt vdi dung mdi pettoleum ether (PE) cd quay loai dung mdi thu dugc eao PE.
Phdn lap.chdttir cao PE: tbuc hidn qud trinh sdc ky cdt, chat hap phy la silica gel, dung mdi giai Iy egt bdt ddu tir PE sau dd tang do phan cue bang dung dich PE vdi ethyl acetate (EtOAc) theo ty le thich hgp. Theo ddi qua ttinh sac ky cdt bdng sdc ky Idp mdng (TLC).
Thudc thii hien vit Id dung dich sulfuric acid 10% ttong medianol va dung dich KMn04 ttong NaOH 5% vd sdy bdn mdng d 110 °C.
Cac phdn doan thd hidn Rf gidng nhau ttdn TLC dugc gom lai. Tien hanh sde ky cgl tidp tyc vdi cdc phdn doan gidng nhau de phdn lap duge chat sach.
Xdc dinh cdu tnic ciia chat da phdn lap dugc: sur dung cac phuong phap phd nghidm;
Bang 1: Ket qua sac kf cdt silica gel ciia cao petroleum ether
^H-NMR, '^C-NMR, DEPT NMR va cac tai lieu lien quan de xac dinh cdu true cac chat phan lap dugc. Phd NMR dugc do tren may Bruker Advance 500 MHz (Vien Hda hgc, Vidn Khoa hge va Cdng nghe Viet Nam, sd 18 Hoang Qudc Viet, Cdu Gidy, Ha Ngi).
Silica gel diing cho sdc ky cdt pha thudng CO hat 0,040 - 0,063 mm. Sdc ky ldp mdng dugc thye hidn tten ban iridng ttdng sdn silica gel KG 60 F254. Cdc hda chdt tinh khiet khdc cd xudt xii tir Tmng Qudc.
3 ICET QUA VA THAO LUAN 3.1 Ket qua sac ky cgt cao PE tu vo cay
Mam
Kdt qud sdc ky cgt silica gel Ci 6.615 g eao PE cho 9 phdn doan dugc trinh bay d Bdng 1 sau day:
Phan doan 1 2 3 4 5 6 7 8 9
HI dung mdi giai ly PE
PE:EtOAc = 99:l PE:EtOAc = 95:5 PE:EtOAc = 9:l PE:EtOAc = 8:2 PE:EtOAc = 7:3 PE:EtOAc=l:l EtOAc EtOAc:MeOH=l:l
Kh5i Iinrng
(s)"
0,526 2^0 2,098
0.45 0,195 0,189 0,03 0,247 0,318
Kdt qud TLC Nhidu vet Co hai vdt chinh Nhieu vet Co hai vet chinh
Co 1 vdt chinh mau tim va 2 vet md Nhieu vet cd I vet chinh Nhieu vet
Nhieu vet Nhieu vet Nhidu vet Nhidu vdt
Ghi chn Khao sdt Khao sat Khao sat
Phdn dogn 1: dugc tdch tten cdt silica gel, rira giai bdng hd dung mdi PE 100%, PE:EtOAc = 99:1 tbu dugc chat rdn, tinb thd hinh kim mau ttdng. Kidm tta trdn TLC vdi he dung mdi w-hexane:CHCl3 = 6:4 va phdt hidn bdng H2SO4 10% ttong MeOH cho vdt ttdn mau cam sau dd chuyen sang tfm cd R/= 0,39.
Ky hieu hgp chdt nay Id PHUOC-PH-02 (khodng 3.2 mg, khdng thu gom hdt).
Phdn dogn 2: xudt hidn kdt tua mau ttdng, tinh chd bdng sdc ky cgt thudng vdi cdc he dung mdi gidi Iy cdt PE 100%, PE:EtqAc = 99:1, PE:EtOAc = 98:2, thu duge tinh thd hinh kim mau ttdng, hidn vdt mdu do tun cd
R/= 0,34 (PE:EtOAc - 8:2) tten TLC khi diing thudc thu la H7SO4 10% ttong MeOH. Ky hieu hgp chdt nay la PHUOC-PH-01 (173 mg).
Phdn dogn 3: thdy cd kdt tua mdu ttdng, tach lay ket hia tmh che bdng each cho rira phdn doan bdng pettoleum ether va kdt tinh lai nhidu lan phan khdng tan ttong CHiCL thu duge tinh the hinh kim, mau ttdng.
Kiem tta bang TLC vdi he dung mdi giai ly PE:EtOAc - 7:3 eho vdt mdu tfm cd 1^= 0,32 hidn hinh bdng thudc thii la H2SO4 10% trong MeOH. Ky hieu hgp chdt nay Id PHUOC-PH- 03 (71 mg).
Tap chi Khoa hoe Tnrmg Bgi hoc CSn Tha Phm A: Khoa hoc Tir nhim, Cong ngfie va mi Irwmg: 26 (2013): 1-5 3.2 Ket qua dir lieu pho
Hffp chat PHCOC-PH-01:
Ph6 "H-NMR (500 MHz, CDCI3), 6 (ppm):
5,53 (IH, dd, J = 8 va 3 Hz, H-15); 5 3,19 (IH, dd, y = U va 5 Hz, H-3). Ngoai ra, tin hieu ph5 con cho thay co 8 mui don 6 cac vi tri 5:
0,98 (3H, s, H-23); 0,93 (3H, s, H-24); 0,80 (3H. s, H-25); 1,09 (3H, s, H-26); 0,82 (3H, s, H-28); 0,91 (3H, s, H-27); 0,95 (3H, s, H-29);
0,91 (3H, s, H-30).
Ph6 " C - N M R (125,8 MHz, CDCI3), 5 (ppra): cho th^y co hai tin hi^u 158,1; 116,9 ppm lln lugt thu6c ve lien kk doi tai cac vi tri Cu; Ci5 va tin hieu c6ng huang ctia mot nhom hydroxy methine a 79,1 ppm. Ph6 DEPT NMR cho thSy hop chat PHUOC-PH-01 co 30 carbon trong do co: 10 nhom methylene (-CH2-), 5 nhom methine (-CH=), 8 nhom methyl (-CH3) va 7 carbon hi cap.
Cac pha 'H-NMR va "C-NMR cho thiy PHUOC-PH-01 la mot triterpenoid nam vong thu0c khung taraxeran cung vol rapt li§n kk doi va mpt nhom hydroxy trong phan ttr. Cac h§ng so tuong tac cua H-3 ( J = 11 Hz va 5 Hz) cho thay nhom hydroxy 6 C-3 co cau hinh ,8.
Tii' cac dir kien tren nhan danh dupe PHUOC-PH-Ol la taraxerol (Hinh 1). Ket qua nay cung phu hpp vol ket qua cua Nguyen Quyet Chi6n et ai, 2004.
Hinh 1: Cdng thih: c4u tao ciia taraxerol Taraxerol la mpt triterpen co hoat tinh khang vi sinh vat, chQng viem va ch6ng kh6i u. Trong do hoat tinh khang khuin voi cac nong dp irc che nho nhat (MIC: Minimum
Inhibitory Concentration) 0,04; 0,016; 0,63 va 0,31 mg/mL tuong iing voi cac loai vi khuin Staphylococcus aureus, Enterococcus faecalis, Pseudomonas aeruginosa va Escherichia coli.
Ngoai ra no co kha nang uc chl dang ki sir tang t n i t o g cua dong t l bao ung thu ph6i a nguoi H I S ? (J. 0 . Famakin, 2002).
Hg-p chat PHUOC-PH-02:
Phd 'H-NMR (500 MHz, CDCI3), 5 (ppm):
CO 8 tin hieu ciia 8 nhom methyl: 0,83 (s, 3H, H-26); 0,9i (s, 3H, H-30); 0,92 (s, 3H, H-28);
0,96 (s, 3H, H-29); 1,07 (s, 3H, H-24); 1,08 (s, 3H, H-25); 1,09 (s, 3H, H-23); 1,14 (s, 3H, H-27). Dp dich chuyin hoa hpc o 5,56 (IH, dd, J ^ 8 va 3,5 Hz) la ciia proton lien ket tgi carbon cf lien kit doi (nhom =CHtai C15).
P h i " C - N M R (125,8 MHz, CDCI3), 5 (ppm): cho thiy nguyen tii' carbon o trang thai lai hoa sp' thu6c nhom ^ C H co dp dich chuyen hoa hpc a 117,2 ppm. Nguyen tix carbon bac 4 ciing a trang thai lai hoa sp" tham gia vao lien ket doi vol nguyen tu nay co dp dich chuyen hoa hpc o 157,6 ppm. Ngoai ra, pho '^C-NMR con cho m6t tin hieu of trucmg rit yeu tuong U'ng vol dp chuyen hoa hpc 217,5 ppm dac trung cho carbon trong nhom carbonyl tai vi tri C-3.
Ph5 DEPT NMR cho thiy co 10 nh6m -CH2, 4 nhom -CH=, 8 nhom -CH3, 8 carbon ttr cap. Nhu vay co the kit luSn ring hpp chat nay la mot triterpen thupc khung olean.
Tir nhiing du kien teen PHUOC-PH-02 dupe nhan danh la taraxerone (Hinh 2). K i t qu^ nay ciing ph£i hgp voi kit qu^ ciia A.K.
Jamale/aZ. 2009.
Hioh 2: Cong thl^c ciu tao h6a hQC taraxerone
Tgp ehi Khoa hgc Trudng Dgi hge Cdn Tha Phdn A: Khoa hge Ty nhien, Cong nghe vd Moi trudng: 26 (2013): 1-5 Taraxerone Id mdt triterpen, hgp chat nay
da dugc nghien curu in vitto cho thay hoat tfnh chdng benh sdt den do ky sinh triing Leishmania donovani (AG 83) gdy benh va boat tinh chdng khdi u ddi vdi ddng te bao bach cdu K562. (Biswas Moulisha et al, 2009). Ngoai ra, taraxerone cdn cd boat tfnh chdng Iai the boat ddng cua k^ sinh trung Giardia lamblia cao hon so vdi taraxerol va seopoletin (ICso 11.33 pg/mL). (Ignacio Hemandez-Chavez et al, 2012)
Hgp ch^t PHUOC-PH-03:
Phd 'H-NMR (500 MHz, CDCI3), 6 (ppm):
0,79 (3H, s,H-24); 0,84 (3H, s, H-26); 0,98 (3H, s, H-23); 0,99 (3H, s, H-27), 1,05 (3H, s, H- 25); 1,68 (3H, s, H-30); 2,39 (IH, dt, / - 11 vd 5,5 Hz, H-19); 3,19 (IH, dd, / - 11 va 4,5 Hz, H-3); 3,33 (IH, dd, J = 11 vd 5 Hz, H-28);
3,81 (IH, dd, J= 11,5 va 3,5 Hz, H-28); 4,58 (IH, br s, H-29); 4,68 (IH, br s, H-29).
Tren phd '^C-NMR eiing dd dang nhdn thdy tin hieu cdng hudng eua mgt lien kdt ddi d 109,7 va 150,5 ppm mong ung vdi carbon d cdc vi tri C29 va C20, mdt nhdm hydrojQ'methine (5c 79; C3) va mdt nhdm hydroxymethylene (6c 60,6; C28). Pho '^C-NMR kdt hgp vdi DEPT cho thdy PHUOC-PH-03 la mdt triterpen thugc khung lupan, cd 30 tfn hieu carbon, trong dd cd 6 nhdm methyl, 12 nhdm methylene, 6 nhdm methine, 6 carbon tii cdp.
Tir nhimg du kien ttdn PHUOC-PH-03 dugc nhdn danh Id betulin (Hinh 3). Ket qua nay cung phii hgp vdi kdt qud cua Seyed Abdolmajid Ayatollahi et al. 2009.
24 23
Hinh 3: Cdng thuc cau tao hoa hoc betulin
Bemlin la mdt triterpen ed mat ttong nhidu Ioai thue vdt thugc cdc hg khdc nhau. Betulin dugc ly tti'eh tir cay Betula utilis chiia betulin ldn den 12% trgng lugng cua nd. Do dd, bemlin dugc dung Idm nguyen lieu ban dau de chuydn hda thanh axit betulinic cd boat tinh sinh hgc cao. (K. M. Nadakami, 1976). Ngoai ra, betulin cdn thd hien hoat tinh gay ddc te bao tten hai ddng HeLa va Hep-2 vdi ciing gid tri ICso la 40 pg/mL. Betulin cung thd hidn hoat tinh chdng HIV vdi gia tti IC50 Id 6,1 pg/mL. (K. S. El Deeb ei al., 2003). Cae nghien ctiu ciia Miura cdn cho thdy betulin cd tdc dung bdo ve gan vd Idm giam kha nang gay ddc cua CdCl? d ndng do thap 0,1 pg/mL. Co che ed the Id do betulin thiic ddy su tong hgp cac protein cd tac dung bao vd cac td bao khdi dnh hudng cua Cdcia. (N. Miura et al., 1999) 4 K E T L U ^ N
Bdng cac phuong phdp sdc ky kdt hgp vdi cac phuang phap phd hidn dai da phdn lap va nhdn dang dugc 3 hgp chdt triterpen tir cao pettoleum ether eua vd cay Mdm di la:
taraxerol, taraxerone vd bemlin. Ddy Id ldn ddu tidn 3 ttitetpenoid dugc phan lap tir bg phan vd cua cdy Mam oi va nhung hgp chat nay ddu cd hoat tfnh sinh hgc cao.
TAI LIEU THAM KHAO
1. AK. Janal, W.A. Yaacob and Laily B. Din, 2009. Triterpenes from the Root Baric of Phyllanthus Columnaris, Austtalian Joumnal of Basic and Applied Sciences 3(2): 1428- 1431.
2. Biswas Moulisha, Mandal Nimp Bikash, Palit Partha, Ghosh Ashoke Kumar, Banneijee Sukdeb and Haldar Pallab-Kanti, 2009. In vitto Anti-Leishmanial and Anti-Tumour Activities of aPentacyclic Triterpenoid Compound Isolated from the Fruits of Dregea volubilis Benth Asclepiadaceae, Tropical Joumai of Pharmaceutical Research 8 (2): 127-131.
3. J. O. Famakin, 2002. Investigation of antibacterial compounds present in Combretum woodii duemmer. MSc Thesis.
University of Pretoria, South Africa (unpublished).
4. Seyed Abdolmajid Ayatollahi, Asie Shojaii, Farzard Kobarfard, MittaNori, Mohammad
Tgp chi Khoa hgc Trudng Dai hgc Cdn Tha Phdn A: Khoa hgc Ty nhiin. Cdng nghi vd Moi truong- 26 (2013): 1-5 9. N. Muira, Y. Matsumoto, S. Miyairi, S.
Nishiyama and A. Naganuma, 1999. Protective Effects of Triterpeod'Compounds. Against the Cytotoxicity of Cadmium m HepG2 Cells, Molecular Pharmacology, 56,1324-1328.
10. Nguyin Quydt Chidn, Nguyin Van Hiing, Trdn Van Sung, 2004. Nghidn ciiu thanh phdn hoa hoccay Kydia Glabrescens, Tap chi Hda hoc, T. 42(1), tt. 71-75.
11. Pham Hoang Hg, 2003. Cay c6 Viet Nam, Quydn II, NXB Tre, TP H6 Chi Minh, 844 - 845.
12. W.M. Bandaranayake, 2002. Bioactivities, bioactive compounds and chemical constituents of magrove plants, Wetlands Ecology and Management 10: 421-452.
Fathi and Mohammad Iqbal Choudhari, 2009.
Terpens from aerial parts of Euphorbia splendida, Joumai of Medicinal Plants Research Vol. 3(9), pp. 660-665.
5. Ignacio Hernandez-Chavez, Luis W. Torres- Tapia, Paulino Sima-Polanco, Roberto Cedillo- Rivera, Rosa Moo-Puc and Sergio R. Peraza- Sanchez, 2012. Antigiardial Activity of Cupania dentata Bark and its Constituents, J.
Mex. Chem. Soc, 56(2), 105-108.
6. K.M. Nadakami, 1976. Betula utilis D.Don, Indian Mater. Med., 1,198-1296.
7. K. S. EI Deed, R. A. Al-Haidari, J. S. Mossa and A. Abdel Monem, 2003. Phylochemical and Pharmacological studies of Maytenus Forsskaoliana, Saudi Pharraaceuticai Joumai, 11(4), 184-191.
8. M. Sharaf, M. A EI-AnSari and N. A Saleh, 2000. New flavonoids from Avicennia marina, Fitoterapia, 71(3), 271-277.
