Tgp chi Khoa hgc Tru&ng Dgi hgc Cdn Tho Phdn A: Khoa hge Ty nhien, Cdng nghe vd Moi irudng: 26 (2013): 6-9
Tap chf Khoa hoc Tru'dng Oai hoc Can Thtf website: sj.ctu.edu.vn
N G H I E N C l J t J T H A N H P H A N H O A H Q C T R O N G C A O P E T R O L E U M E T H E R C U A R E C A Y M A M {A VICENNIA MARINA)
Le Thanh Phudc va Tir Minh Td'
' Khoa Khoa hoc Tu nhien. Trudng Dgi hoc Can Iha Thong tin chung:
Ngay nhgn: 14/09/2012 Ngdy chdp rdign: 19/06/2013 rale:
Study on the chemical components in petroleum ether extract from Avicennia marina root Tirkhda:
Re Mdm Avicennia marina, thdnh phdn hda hoc, taraxerone, betulin Keywords:
Avicennia marina roots, chemical components, taraxerone, betulin
ABSTRACT
Phytochemical investigation on the pneumatophores (aerial roots) of Avicennia marina (Forssk.) Vierh., collected in Dong Hai district, Bac Lieu province, resulted in the isolation of two triterpenoids, taraxerone (CSQHJ^) and betulin (C30H50O2). The structures of these compounds have been elucidated by modem spectroscopic methods: API-MS,' H-NMR. '^ C-NMR, DEPT NMR
T O M T A T
Nghien cuu hda thuc vat tren ri phdi cua cdy Avicennia maria (Forssk.) Vierh., du^c trong tgi huyen Dong Hdi, tinh Bac Lieu, kit qud phdn lap dugc hai triterpenoid Id taraxerone (Csf04^) vd betulin (C30H30O2). Cdu tnic hoa hgc cua cdc chdt ndy da dugc xdc dinh bdng cdc phuang phdp pho hien dgi: API-MS, 'H-NMR,
"C-NMR, DEPT NMR.
1 DAT VAN B E
Cdy Mam ed ten khoa hgc la Avicennia marina, thugc hg Cd roi ngua (Verbenaceae) (Pham Hodng Hg et al, 2003). Day Id lodi cay sinh tmdng vdi sd lugng ldn dugc ttdng khdp Viet Nam tap tnmg nhidu nhdt d cac khu rimg ngdp man ven bien vdi he thdng re phdt ttidn hdt siie da dang. Gan day nhidu nha nghien ciru dac biet ddnh gid cao ve cdy Mdm va xem nd nhu Id ngudn dugc lieu day tiem nang (Miles el al, 1998). Trong ddn gian vd cd;y Mdm duge su dung phd bidn dd chua mdt sd benh nhu: ddu miia, cae bdnh vd da, thap khdp.
Id loet (Fauvel et al, 1993; Bandaranayake, 2002). Trong nhimg nam ttd lai ddy da cd nhieu tdc gia tten thd gidi da quan tam den thdnh phan hda hgc va boat tfnh sinh hgc cua rd Mdm. 6 Viet Nam, Pham Thi Thuy Trang et ai, 2010, da phdn lap dugc bdn chdt tinh khiet ttong bg phan re ciia eay Mam thd hidn hoat
tinh khang ung thu cao. Vdi muc dich gdp phan tao co sd khoa hgc cho cac bai thudc ttong dan gian ndu tren ddng thdi tiep tuc khao sdt thanh phdn hda hgc cua vd rd Mdm ttdng tai Bac Lieu, ttong bdi bao nay, chiing tdi ttinh bay kdt qud phdn lap va nhdn danh cdu tnic hai hgp chdt tir cao pettoleum ether tten bd phdn rd phdi eua cdy Mdm.
2 PHlTONG TIEN VA PHU'ONG PHAP Nguydn lieu: Re cdy Mam dugc thu hai (phdn phfa ttdn mat ddt) tai huyen Ddng Hai, tinh Bac Lieu sau dd rita sach, Ioai bd phan sau, eat nhd, phai khd ddn khdi Iugng khdng ddi va xay nhd trudc khi su dung.
Phirong p h a p : Chidt hoat chdt: bgt re Mam dugc ngdm ttong cdn 96", phan dich chiet ndy duge loai dung mdi dudi dp sudt kem thu cao cdn. Sau dd ldy cao cdn chiet vdi dung mdi
Tgp ehi Khoa hoc Tru&ng Dai hoc Cdn Tha Phdn A • Khoa hgc Ty nhien, Cong nghi vd Moi tru&ng: 26 (2013). 6-9 petroleum ether (PE), dich chiet PE dugc loai
dung mdi dudi dp sudt kem thu dugc cao PE.
Phan lap cae hgp chdt tir cao PE: thuc hien phuang phdp sdc k^ cdt, chdt hap phu la silica gel, theo ddi qua ttinh sdc ky cdt bdng sac ky ldp mdng (TLC). Giai ly cdt bdt ddu vdi PE sau dd tdng ddn do phdn cue bdng dung dich PE vd ethyl acetate (EtOAc) theo ty le thfch hgp. Thuoc thii hidn vdt sir dung la: dung dich sulfuric acid 10% ttong methanol va dung djch KMn04 trong NaOH 5%. Cdc phan doan thd hidn R/ gidng nhau tten TLC dugc gdp lai.
Tidp tyc tidn hanh sdc ky cdt cdc phdn doan giong nhau de phdn lap dugc chat sach.
Xac djnh edu tnic ciia cdc chdt da phan lap duoc: sir dung cac phuong phap phd nghidm:
MS, 'H-NMR, '^C-NMR, DEPT NMR va cac tai lidu lidn quan dd xac dinh edu tnic cac chdt phdn lap dugc. Pho NMR dugc do tten may Bmker Advance 500 MHz (Vidn Cdng nghe, sd 18 Hoang Qude Viet, Cdu Gidy, Ha Ngi).
Silica gel diing cho sdc ky cdt pha thudng cd hat 0.040-0.063 mm. Sde ky ldp mdng dugc thuc hidn tten ban mdng ttang sdn silica gel KG 60 F254. Cac Ioai hda chat tinh khiet khac cd xudt xir tir Trung Qudc.
3 K I : T QUA VA THAO L U ^ 3.1 K^t qua sdc ky cgt
Tir 5.037 g cao pettoleum ether tidn hanh
Bang 1: Pho NMR ciia PHUOCTO-03 so voi tai lieu tham khao Tian Minqing va c^ng sir, 6 ppm sde ky cdt thudng vdi he dung mdi PE va dung dich PE vdi EtOAc theo ty Id thfch hgp. Kdt qua d phan doan PE:EtOAc = 95:5 thu dugc mot hgp chdt tinh khiet (22.5 mg), cd dang tinh thd mau trdng hidn mgt vet ttdn mau tfm cd R/ = 0.46 (PE:EtOAc = 70:30). Hgp eh^t nay dugc ky hieu la PHUOCTO-03. 0 phan doan PE 100% thu duge mdt chdt tinh khidt khac (9 mg), cd dang tinh the mau trang hinh kim hidn vdt mau cam sau dd chuydn sang mau tfm cd R/ = 0.49 (PE:CH2Cl2 = 50:50). Hgp chdt nay ky hidu la PHUOCTO-04.
3.2 Ket qua dir lieu pho Hgp chdt PHUOCTO-03
Phd A P r - M S cho bidt [M+H]"" = 443 amu tuong irng vdi cdng thuc phdn tir C30H50O2.
Pho 'H-NMR (500 MHz, CDCI3), 6 (ppm), J (Hz): 0.76 (s, 3H, CH3-24), 0.82 (s, 3H, CH3-26), 0.97 (s, 3H, CH3-23), 0.98 (s, 3H, CH3-27), 1.02 (s, 3H, CH3-25), 1.68 (s, 3H, CH3-3O), 3.19 (dd, IH, J = 11.5, 5.0 Hz, H-3), 3.33 (d, 1 H , J = 11.5 Hz, H-28), 3.80 (dd, IH, J = T 1 . 5 , 3.5 Hz, H-28), 4.58 (t, IH, J = 1.5 Hz, H-29), 4.68 (d, IH, J - 1 . 5 Hz, H-29).
Phd DEPT NMR kdt hgp '^C NMR cho thdy cd 12 nhdm >CH2, 6 nhdm >CH-, 6 nhdm -CH3,6 C tii cdp.
STT 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
PHUOCTCMIS 'H(J,Hz)
3.19 (dd, 11.5, 5.0)
" C 38.7 27.4 79.0 38.9 55.3 18.3 34.3 40.9 50.4 37.2 20.9 25.2 37.3 42.7 27.1
Betulin
•'c 387 27.5 79.0 38.9 55.4 18.3 344 41.0 50.5 37.4 20.9 25.3 372 42.8 27.1
STT 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30
PHH0CTCMI3 'H(J,Hz;)
0.97 (s, 3H) 0.76 (s,3H) 1.02 (s,3H) 0.82 (s, 3H) 0.98 (s, 3H) 3.33 (d, 11.0) 3.80(dd, 11.0, 1.5) 4.58(1,1.5) 4.68(41.5)
1.68 (S.3H)
"c
29.2 29.7 48.8 47.8 150.5 29.8 34.0 28.0 15.4 16.1 16.0 14.8 60.6 109.7 19.1
Betulm
" C 29.3 47.9 48.9 47.8 150.5 29.9 34.0 28.0 15.3 16.1 16.0 14.8 60.6 109.7 19.1
Tgp chi Khoa hgc Tru&ng Dgi hgc Cdn Tho Phan A: Khoa hoc Ty nhien, Cong nghe vd Moi trudng- 26 (2013): 6-9 Tir cac du lieu tten vd so sanh vdi cdc ket
qua da dugc cdng bo (Seyed Abdolmajid Ayatollahi et al. 2009) hgp chdt P H U O C T O - 03 dugc nhdn danh Id betulin (Hinh 1).
sinh vat Streptococcus pyogene (Supaluk Prachayasittikul et al, 2010). NhGng nghien ciru gan day cho thdy betulin da dugc thii nghiem thanh cdng trong didu tti bdnh day simg quang hda (Sebastian Jager et al, 2008).
Hgp chat PHUOCTO-04
Phd APT-MS cho bidt [M-H]" = 423 amu tuong ling vdi cong thuc C3oH4ffO.
Phd 'H-NMR (500 MDHz, CDCI3), 5 (ppm), J (Hz): 0.83 (s, 3H, CH3-26), 0.91 (s, 3H, CH3- 30), 0.92 (s, 3H, CH3-28), 0.96 (s, 3H, CH3- 29), 1.03 (m, IH, H-18), 1.07 (s, 3H, CH3-24), 1.08 (s, 3H, CH3-25), 1.09 (s, 3H, CH3-23), Hinh 1: Cdng thiic cau tao hda hgc betulin I.14 (s, 3H, CH3-27), 5.56 (dd, IH, J = 8.5, 3.5 Betulin Id mgt pentacyclie ttiterpen cd " ^ ' ""IS),
nhieu ttong vd than, Id, rd cua nhidu loai thirc ph6 DEPT NMR kdt hgp '^C NMR cho vdt da dugc bidt ddn vdi kha ndng khang khdi thay co 10 nhdm >CH2, 4 nhdm >CH-, 8 u, ddc bidt la chdng khdi u dc tfnh va ung thu nhdm -CH3, 8 C tii cap.
da. Ngoai ra betulin cdn cd hoat tinh khang vi
Bang 2: Phd NMR cua PHUOCTO-04 so vol tai lieu tham khao A. K. Jamal va c^ng sy 6 ppm SFT -
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
PHUOCTO-04 'H(J,Hz) 1.40(ni, IH) 1.87 (m,lH) 2.33 (m, IH) 2.58 (m,lH)
5.56 (dd, 8.5, 3.5)
"c
38.4 34.2 217.5 47.6 55.8 20.0 35.1 38.9 48.7 35.8 17.5 37.7 37.6 157.7 117.2
Taraxerone
"c
38.5 34.4 217.0 47.8 56.0 202 35.3 39.1 48.9 37.9 17.7 36.0 37.8 157.8 117.4
STT - 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30
PHUOCT&04 'H(J,Hz)
1.09 (s. ,3H) 1.07 (s,3H) 1.08 (s.
0.83 (s.
1.14 (s.
0.92 (s.
0.96 (s.
0.91 (s.
,3H) ,3H) ,3H) ,3H) ,3H) ,3H)
" C 36.7 37.8 48.8 40.7 28.8 33.6 33.1 26.1 21.4 14.8 29.9 25.6 29.9 33.4 21.5
Taraxerone
" C 36.9 37.9 49.0 40.8 29.0 33.8 333 263 21.7 15.0 30.1 25.8 302 33.6 21.6 Tir nhirng dii kidn tren va so sanh vdi cdc
kdt qua da dugc cdng bd (A. K. Jamal et al, 2009) hgp chdt PHUOCTO-04 dugc nhan danh la taraxerone (Hinh 2).
Hinh 2: Cong thii-c cau tao cua taraxerone
Tgp ehi Khoa hgc Tru&ng Dgi hgc Cdn Tha Phdn .4: Khoa hgc Ty nhien. Cong nghe vd Mdi tru&ng: 26 (2013): 6-9 Taraxerone Id mgt ttong cac ttiterpen phd
bidn va tdn tai d mdt sd lodi thue vat trong tu nhidn. Thii boat tfnh sinh hgc ciia chdt nay da ghi nhdn dugc nhidu ket qua hap dan khi thii nghidm thanh cdng kha ndng khang ndm, khang khudn nhu: Bacillus subtilis, Escherichia coli, Candida albicans, Candida glabrata (Muhammad Imran anjum et al., 2007), khdng khdi u va khang Leishmania (Biswas Moulisha et al, 2009).
4 KET LUAN
Da phdn lap va nhdn danh edu tnic hai chat tinh khiet taraxerone va bemlin tir cao petroleum ether cua re Mam. Ket qud tten gdp phdn md rgng hon nghien cuu vd thanh phan hda hgc ciia re Mdm lai tinh Bac Lieu (Viet Nam) dd dugc cdng bd ttong thdi gian gdn ddy (Pham Thj Thuy Trang et al, 2010). Day la hai hgp chat thudc hg triterpenoid da dugc nghien eiiu vd nhung kdt qud khdo sdt cho thdy hoat tinh sinh hgc cua chiing kha cao. Dieu ndy lam CO sd giai thfch tai sao ngudi ta thudng sir dung rd Mdm nhu mdt Ioai dugc lieu de ehiia benh ttong dan gian.
TAI LIEU THAM KHAO
1. A.K. Jamal. W.A- Yaacob and I^ily B. Din, 2009. Triterpenes from the Root Bark of Phyllcmthus Columnaris, Australian Joumai of Basic and Applied Sciences, 3(2): 1428-1431.
2. Bandaranayake W.M, 2002. Bioactivities, bioactive compounds and chemical constituents of mangrove plants. Wetlands Ecology and Management, 10:421-452.
3. Biswas Moulisha, Mandal Nirup Bikash, Palit Peirtha, Ghosh Ashoke Kumar, Banneqee Sukdeb and Haldar Pallab.Kanti, 2009. In vitro Anti-Leishmanial and Anti-Tumor Activities of a Pentacyclie Triterpenoid Compound Isolated frdm the Fruits of Dregea volubilis Benlh Asclepiadaceae, Tropical Joumai of Pharmaceutical Research, 8(2): 127-131.
4. Fauvel, M.T., M. Bousquet, A. MouHs, C J . Gleye and S.R. Jensen, 1993. Iridoid glycosides from Avicennia germinans.
Phylocbemistry, 38:893-894.
5. Miles DH, Kokpol U, Chittawong V. Tip- Pyang S, TunsuwanK&Ngpyen C, 1998.
Mangrove foresis-Theimportance of conservation as a bioresource for ecosystem diversity and utilization as a source of chemical constituents with potential medicinal and agricultural value. 1999 lUPAC 70 (11):
1-9.
6. Muhammad Imran anjum, Ejaz Ahmed, Abdul Jabbar, Abdul Malik, Muhammad Ashraf, Muhammad Moazzam and Muhammad Azam Rasool, 2007. Antimicrobial Constituents from Fagonia cretica, Jour.Chem.SocPak. Vol.29, No., 6, P 634-639.
7. Phgm Hoang Hg, 2003. Cay cd Viet Nam, Nxb Tre, Thdnh phd Hd Chi Mnh. T.H: tr. 845 8. Phgm Th| Thiiy Trang vd Le Thanh Phudc,
2010. Khdo sdt thdnhphdn hda hgc ciia re cdy Mdm (Avicennia marina). Tap chi Khoa hpc, Tnrdng D?i hoc Cdn Tho S6 15b, 9-14.
9. Supaluk Prachayasittikul, Puttirat Saraban, Rungrot Cherdttakulkiat,Somsak Ruchirawat, Virapong Prachayasittikul, 2010. New Bioactive Triterpenoids and Antimalarial Activity ofDiospyros Rubralec, EXCLI Joumai, 9:1-10 ISSN 1611-2156.
10. Sebastian Jager, Melanie N. Laszczyk and Armin Scheffler, 2008. A Preliminary Pharmacokinetic Study of Betulin, the Main Pentacyclie Triterpene from Extract of Outer Bark of Birch {Betulae alba cortex), Molecules 2008, 13. 3224-3235.
11. Tian Minqing, Dai Haofli, Li Xiaoming, Wang Bingui, 2009. Chemical constituents of marine medicinal mangrove plant Sonneratia caseolaris, Chinese Joumai of Oceanology and Limnology Vol. 27 No. 2, P. 288-296.
