T 51(6) 770-773 TAP CHI HOA HQC THANG 12 NAM 2013 KET QUA BAN DAU VE THANH PHAN HOA HOC CAO CHIET DICLOMETAN LA CUC QUY (TITHONIA DIVERSIFOLIA (HEMSL.) A GRAY) Tan Thi Thanh Thayl*, Tan Van Hieul, Vu Thi Thu Ha', Trinh Thi Thu?, Trill Van Sung2 Phong Thi nghiem diem Cong nghe loc, hoa dau, Vien Hoa hQc Cong nghiep Viet Nam Vien Hoa hQc, Vien Han lam Khoa hoc va COng nghe Viet Nam Den Tim soon 15-10-2013 Abstract Four compounds: myristyl myristate (1), hispidulin (2), eupafolin (3) and tagitinin A (4) were isolated from the dichlomethane leaf extract of Tithonia diversifolia (Hemsl.) A Gray Their structures were elucidated by analysis of IR, MS, and NMR spectra and comparison with the published data Compounds and were isolated for the time from this plant Keywords: Tithonia diversifolia, Asteraceae, hispidulin, eupafolin, tagitinin A DAT VAN DE Citc quy hay gqi di quy, sum quy bien diep, quy dai, huang ducmg dai, huang duang Mexico (Tithonia diversifolia) la,rnat loii thuc vat ho Cue (Asteraceae) co ngu8n g'Oc tir Mexico, hien duqc phan be rOng khip a cfic khu vuc can nhiet dai va nhi'et dai [1] Clic quS, thuc clang cay bui, cao 12 in, than cif) long, phan nhieu canh, la thuon, phien co thity, bia co ring Hoa a du ng9n, tren cueng dai Hoa a bia hinh m8i lep, yang ttrai; hoa gi6a hinh Ong; gift hoa co vay Cay hoa tir thing 12 den thing Viet Nam, cay cue quy hien da duqc to nhien hem, moc bin a eat nhieu nai, dac biet la a vimg Tay Nguyen [2] Trong y hQc co truyen Chau My, ntrac ep tir re" va cay duqc sir dung dieu tri fip-xe [3]; Mexico loai duqc uong de chi-1.a sot ret va dien hinh la chita cfic khiSi to mfiu va rut ca, bOt tir la kho tri vet tham, vet thuang va Wing da nhiem Dai Loan, dich chiet nuac ctia cay duqc sir dung de tri dfii thfio &rang (cluing dai nhieu va chung khfit nhieu) [4] va viec ling dung dm T diversifolia dieu tri dfii thao dul:mg cling di duqc ding ki bio 118 [5] Tren the giai, cay cue quy duqc quan tam nghien elm ye thinh phAn hoa h9c va hoat tinh nhieu nhAt cfic loai thuc vat thuc chi Tithonia [10] Cfic hq? chit sesquiterpen lacton vi ditecpen la thinh phan hda hoc chinh vi co nhieu hoat tinh thit vi cua 'oil [10] Tuy nhien, cho den nay, chtra co cong trinh nao ye thinh phin h6a hQc cita cay cfic quS, thu hfii tai Viet Nam duqc cong b6 Bai bar) ding be ket qui !mac du ye vi0c khao sat plan hoa hQc cua cao chiet diclometan tir la cay cuc quy thu hai tai Tay Nguyen THIJC NGHIEM 2.1 Phuong phi') nghien cum Sic 14r ban mong duce tien hanh tren ban silica gel Merck 60F254 Sa'c ky cOt sir dung silica gel c6 hat 0,04-0,063 mm Pho Ong huang tir hat nhan NMR duqc ghi teen may Bruker Avance 500 MHz, pill) khoi ESI—MS duce tren may Water LC/MS/MS-Xevo TQ Pho IR duqc ghi tren may Nicolet 6700 FT-IR 2.2 MAu thurc vat MAu thuc vat ducic thu hfii tai tinh Gia Lai vao thing nam 2012 Tieu ban duac Wu girt tai Phong Thi nghiem diem Cong nghe 19c hoa dau Mau lfi duqc rim sach, phai, sAy kho 23 Thuc ngh4m Li cue quy (400 g) di sAy kho, xay nh6 duqc chiet nhanh (khofing phut) bang diclometan (x 31An) a nhiet d8 phang theo qui trinh tai lieu [13] LQC dich chiet, loai dung moi dual fip suit gam CArt lai duqc him tan Me0H-H20 (7:3, v:v) va chi& phan lop lin Itrqt vii n-hexan va 770 Trein Thi Thanh Thiry va Ong sy- TCHH, T 51(6), 2013 diclometan Cit loai dung mei dual ip suit giam thu dtrqc can cac cao chiet Wang ling Cao chiet diclometan (15,7 g) duqc phan tach bang sic ky cet ten silica gel, dung mei rua giai la hon hop n-hexan-etyl axetat vii lam etyl axetat tang din, thu dtrqc 12 phan doan (Fl-F12) Phan doan Fl (320 mg) duqc rira va ket tinh lai tong metanol thu duqc chit 1, phan doan F6 duqc tinh the ten cet Sephadex LH-20 WA dung mei rim giai metanol thu duqc 35 mg (0,0087 %) chit 2.Chit (40 mg, 0,010 %) thu dtrqc tir phan doan F10 (100 mg) sau tinh cite tren cet Sephadex LH-20 Phan doan F5 duqc tinh the ten OA silica gel vii he dung re 3:7 thu dtrqc moi rim giai n-hexan:etyl axetat hqp chit sach (44 mg, 0,011 %) - Myristyl myristat (tetradecyl tetradecanoat, 1): bet matt tring, C28115602, ESI-MS (m/z): 425[M+H]'; ham lacing 0,08% so vii mau kho 'H NMR (500 MHz, CDC13, Sppm): 4,05 (2H, t, J= Hz, 0-CH2), 2,29 (2H, t, J = 7,5, CH2), 1,60 (6H, m), 1,28 (40H, brs), 0.88 (6H, t, J= Hz) '3C NMR (125 MHz, CDC13): 8, 174,0 (-COO), 64,4 (0-CH2), 22,7-34,5 (CH2), 14,1 (2CH3)- Hispidulin (4',5,7-trihydroxy-6-methoxyflavon, 2): tinh the mau yang nhat, C161-11206, ESI-MS (m/z): 301 [M+H]+; ham lucmg 0,009% IR (KBr) v*max (cm'): 3423, 1657, 1606 'H-NMR (500 MHz, CDCI3+CD3OD, Sppm): 7,83 (2H, d, J = Hz, 1-1-2', H-6'), 6,93 (2H, d, J=9 6,55 (1H, s, H-3), Hz, 11-3', H-5'), 6,57 (1H, s, 3,90 (3H, s, -OCH3) '3C NMR (125 MHz, CDC13+CD3OD, Sppm): 166,3 (C-2), 103,3 (C-3), 184,1 (C-4), 153,9 (C-5), 132,8 (C-6), 158,8 (C-7), 95,3 (C-8), 154,6 (C-9), 105,7 (C-10), 123,2 (C-1'), 129,3 (C-2' & C-6'), 116,9 (C-3' & C-5'), 162,6 (C-4'), 60,9 (6-OMe); - Eupafolin (3 ',4',5,7-tetrahydroxy-6-methoxyflavon, 3): bet yang nhat, CI6H1 207, ESI-MS (m/z): 316 [M+H]+; ham luqng 0,01 % IR (KBr) v*max (cm"): 3411, 1654, 1602 'H NMR (500 MHz, CD3OD, Sppm): 7,35 (1H, s, H-2'), 7,34 (1H, d, J = 8,5 Hz, 11-6'), 6,89 (1H, d, J = 8,5 Hz, H-5'), 6,51 (111, s, H-8), 6,50 (1H, s, 3,89 (3H, s, 0-Me) '3C-NMR (125 MHz, CD3OD, Sppm): 166,3 (C2), 103,3 (C-3), 184,1 (C-4), 153,9 (C-5), 132,9 (C6), 159,0 (C-7), 95,3 (C-8), 154,6 (C-9), 105,7 (C10), 123,6 (C-1'), 114,1 (C-2'), 147,0 (C-3'), 153,9 (C-4'), 116,7 (C-5'), 120,3 (C-6'), 60,9 (6-OMe) - Tagitinin A (3, 10-epoxy-germacrolide-6,7trans-lacton, 4): tinh the khong mau, C19H2807, ESIMS (m/z): 369 [M+H]+; ham lucmg 0,011 % IR (cm"): 3495, 2930, 1756, 1663, 1447 (KBr) 'H NMR (CDC13, 500 MHz, Sppm), 6,27 (1H, d, J = 3,5 Hz, H-13a), 5,58 (1H, ddd, J = 3,0, 5,5, 8,5 Hz, H-8a), 5,54 (1H, d, J= 3,5 Hz, H-13b), 4.57 (1H, ddd, J= 2, 6,5, 10,0 Hz, 11-613), 4,25 (111, dd, J = 7,5, 9,5 Hz, H-la), 4,08 (1H, ddd, J = 3,0, 6,5, 10,0 Hz, H-7a), 2,44 (1H, m, H-2A), 2,44 (1H, m, H-2'), 2,08 (1H, m, H-4P), 2,08 (1H, m, H-2b), 2,08 (1H, m, H-5b), 2,08 (111, m, H-5a), 1,96 (11-1, dd, J= 6,0, 14,0 Hz, H-9b), 1,82 (1H, m, 9,0, 13,0, H-9a), 1,44 (3H, s, H-14), 1,11 (31-1, d, J = 6,5 Hz, H-15), 1,08 (311, d, J = 7,0 Hz, II-3), 1,06 (3H, d, J = 7,0 Hz, H-4') '3C NMR (CDC13, 125 MHz, Sppm): 176,34 (C-1'), 169,42 (C-12), 137,07 (C-11), 121,57 (C-13), 105,73 (C-3), 81,77 (C-6), 81,46 (C-10), 78,26 (C-1), 70,39 (C-8), 47,84 (C-7), 47,04 (C-2), 44,34 (C-4), 37,79 (C-5), 34,62 (C-9), 34,03 (C-2'), 24,99 (C-14), 19,18 (C-15), 18,72 (C-3'), 18,35 (C-4') OH 2: R = H 3: R = OH Ifinh 1: Cau truc ciia chAt 1-4 KET QUA VA THAO LUAN Ket hcrp thong tin tir phe 'H-, '3C-NMR va pic ion phan tir tai m/z 425 [M+H]+ ten phe ESI-MS dm la C28115602 Tren suy cling thirc phan pito 'H-NMR cua chat xuat hien tin hreu triplet tai 0,88 (J= Hz) viii tY le tich phan 6H ttrcmg (mg vii cac proton dm nhom metyl du mach Hai nhOrn metyl co ding de djch chuyenho,c cho thay hqp chat co ciu triic mach Wang doi can ming Tin II iOu triplet dm proton a 4,05 (J = 771 TCHH, T 51(6), 2013 Ket qua ban ddu ve phdn 7Hz) dac trung cho tin hieu cua nhom oxymetylen (0-CH2) ma ben canh nhom CH2 mot nhom CH2 khac Tung tv nhis very tin hieu triplet a 2,29 (.1 = Hz) ding dac trtmg cho nhom CH2 et gift men cacbon bac va met nhom CH2 Tren xuat hien cam tin hieu cua 46 proton a di) dich chuyin vung 1,25-1,62 ppm PhO DC-NMR bao gom tin hieu cila nhom cacbonyl este (COO) a 174 ppm, tin hieu CH2-0 a 64,4 ppm, tin hieu CH3 a 14,1 ppm va cum tin hieu cua cac CH2 vung 22,7-34,4 ppm Ket hop cac phuang phfip phO va so sfinh veri tai lieu tham khao [6], cong thirc ciu tao chit dugc xac dinh la tetradecyl tetradecanoat (myristyl myristat) Chat din tai dual dang bOt mau yang nhat PhO IR dm hqp chit cho thiy dai hap thu rang 3423 cm"' gqi y sir c6 mat ctia cac nhom hydroxyl Dai hap thv a 1657 cm"' dac trtmg cho nh6m carbonyl clang ot,ti-chtra b5o hoa Ph6 ESI-MS a the de) ion am cho pic ion phan tir [M-H]" tai m/z 299 tren ph8 ESI-MS the di) ion dtrcmg xuAt hien cac pic [M+Na]' va [M+H]+ tai cac gifi tri m/z ttrcmg Ung la 323 va 301 cac dir lieu ph6, cong thirc phan dm chat dugc dir doan C161-11206 Tuy nhien tren ph6 "C-NMR (lac trung cho ciu trim cua hqp chit khung flavon chi xuat hien 14 tin hieu Ong twang Dieu cho thay hop chat c6 cap CH twang dtrcrng tai 129,3 (C-2'/C-6') va 116,9 ppm (C-3'/C-5'), goi St rang nhan thorn & von B co nh6m the a C-4' Ngoai tren pito xuat hien hai tin hieu co di) dich chuyen h6a hoc & 8103,3 va 95,3 ppm cila nhom CH nhan thorn khfic Tin hieu Ong Wang cua nhom C=0 dac trung cua nhom cacbonyl xuAt hien tai 184,1 ppm Tin hieu a 60,9 ppm dugc gin cho nhom metoxy Tren ph8 'HNMR ton tai tin hieu dm cap proton nhan thorn duplet c6 ttrong tac ortho (J = 9,0 Hz) tai 7,83 va 6,93 ppm, ttrcmg ung vai tin hieu cong htrong cua H-2'/H-6' va H-3'/H-5' Du lieu hoan toan phu hcrp vbi dir lieu phO '3C-NMR Tin hieu don co cuang de) tich phan 3H a 3,90 dugc xac dinh la nhom metoxy Tir dir lieu phO pan tich a ten, ket hqp vbi so lieu phi') der cong bo [7], cong thirc ciu tao cua hqp chit dugc xac dinh la 4',5,7trihydroxy-6:methoxyflavon (hispidulin) Hispidulin dugc biet den hop chat co hoat tinh chong ung thu, chOng virus ding chit St 18, 10] Chit dugc tach tir cay cue quSt dm An DO va mot vai loci thirc vat khac [9, 10] Chat thu dugc dual dang bOt mau yang Twang tir vbi hap chat 2, phi!) IR dm hop chat ding cho thay cac dai hap thu & 3411 cm' va 1654 cm-' dac trtrng cho sir c6 mat twang ung cua cac nhom hydroxyl va carbonyl dang a,/3-chtra bao hoer PhO ESI-MS a che de) ghi ion am va &rang lAn Itigt cho cac pic [M-H]" tai m/z 315, [M+Na]+ va [M+H]+ tai cac gia tri m/z twang 1rng la 339 va 317 Tir dir lieu khOi, ket hqp vai ph6 NMR, cong thirc phan tir dm chit dugc xac dinh la C16111207 Ara c6 nhieu han mot nguyen tir oxi so veri chit Tren "CNMR xuAt hien 16 tin hieu cong hue:mg dac trung cho cAu trim dm hop chit khung flavon bao g6m tin hieu C=0 (8 184,1 ppm), tin hieu C bac va tin hien CH nhan tham (95,3-123,7 ppm) va tin hieu cua nhom OCH3 (8 60,9 ppm) Ket hqp dir lieu NMR vai dir lieu MS cho phep nhan dinh so vai 2, hop chit , c6 cAu trite twang tir nhtmg co them mot nhomthe 0H Tren ph Al H-NMR, proton co de dich chuyen 6,51 va 6,50 ppm twang ter tren hop chit dugc gin cho H-8 va H-3 Tin hien dm proton nhan thorn c6 ttrcmg tic ortho (d, J = 8,5 Hz) tai 7,34 va 6,89 ppm, twang ung vai tin hieu cong htrang dm H-6' va H-5' Tren ph8 proton khong xuAt hiOn twang tac meta vi the tin hieu dan cua proton tai 87,35 (Iliac gin cho H-2' Tin hieu clan vai cueing de) tich phan la 3H a 3,90 la dm nhom methoxy Ket hqp dn lieu va so sfinh phi') veri chit 2, cAu true dm chit dtrqc xac dinh la 3%4%5,7tefrahydroxy-6-methoxyflavon (eupafolin) Hqp chit cling dugc cong bo co hoat tinh gay dac to bao tren clang to bao ung thisbieu m8 KB [7] Him chat c6 cong thirc phan tier C19112807, dtrqc dua dva vao viec ket hap cac ph8 ESI-MS, 'HNMR va '3C-NMR PhO IR dm hqp chit (lac trung cho khung sesquiterpen lacton chtra bao Ma yeti cac tin hieu a so song 3495, 1756, 1663, 1598, 1447 cm' Tin hieu a vung 1756 cm'' thuang dtrqc gap nhom carbonyl dm cac hqp chit y-lacton [11] Tren ph6 'H-NMR xuAt hien tin hieu dm nhom metyl co dO dich chuyen Ma hoc et 1,44-1,08 ppm Cap tin hieu duplet a di) dich chuyen hoa hoc 6,27 va 5,54 ppm veil J = 3,5 Hz la dau hieu nhan dang cho nhom exocylic metylen cua khung sesquiterpen lacton [11] Ph6 '3C-NMR cho thiy str c6 mat dm 19 tin hieu cacbon, bao Om: 4xCH3, x d6 CO nhom cacbonyl CH2, 6xCH, Phan tich phi) va so sfinh veri tai lieu tham khao cho phep xac dinh cAu trim hap chit la 3,10-epoxygermacrolide-6,7-trans-lacton (tagitinin A) [9] TaFitinin A dugc ding 1)6 hqp chit co hoat tinh chong tieu dtrang thong qua &rang hoat hoa thy the tang sinh peroxisome [12] KET LUAN Tim dich chiet diclometan cua la cay cue qujt (Tithonia diversifolia) TA' phan lap dugc hqp chat 772 Tran Thi Thanh Thtiy va Ong sir TCHH, T 51(6), 2013 F D Gunstone et al Chem Phys Lipids, 17, 1(1976) S M Kupchan, C W Sigel, J R Knox, M S Udayamurthy Cytotoxic flavones from Eupatorium species, Tetrahedron, 25, 1603-1615 (1969) D Osei-Safo, M A Chama, I Addae-Menshah, R Waibel Hispidulin and other constituents of Scoparia dulcis Linn, Journal of Science and Technology, 29, 8-15 (2009) Narayan C Baruah, Jadab C Sarma, Nabin C Barua, Soneswar Sarma, Ram P Sharma Germination and growth inhibitory sesquiterpene lactones and a flavone from Tithonia diversifolia, Phytochemistry, 36(1), 29-36 (1994) 10.Dictionary of Natural Products, version 18:1, Copyright©1982-2009 Chapman & Hall/CRC 2009 11.M Iranshahi, S A Emami, M M Soltani Detection of Sesquiterpene Lactones in Ten Artemisia species Population of Khorasan Provinces, Iranian Journal of Basic Medical Sciences, 10(3), 183-188 (2007) 12.H R Lin Sesquiterpene lactones from Tithonia diversifolia act as peroxisome proliferator-activated receptor agonists, Bioorganic & Medicinal Chemistry Letters, 22, 2954-2958 (2012) 13.S R Ambrosio, Y.Oki, V C G Heleno, J S Chaves, P G B D Nascimento, J E Lichston, M G Constantino, E M Varanda, F B Da Costa Constituents of glandular trichomes of Tithonia diversifolia: Relationships to herbivoty and antifeedant activity, Phytochemistry, 69, 2052-2060 (2008) 1-4 Bang cac phuang phfip ph6 IR, MS, NMR va so cac dir lieu da ding 1)6, cAu triic dm sfinh cac hop chat &roc xfic dinh la: myristyl myristat (1),, hispidulin (2), eupafolin (3) va tagitinin A (4) Chat va Ian du duce tim thay loai cue quSt Liri cam cm: Car lac gia xin than cam on BO Cong thwang cid tai try kinh phi cho cac nghien ceru thong qua dd tai, ma so 043-2013/HD- DTCNHD TAI Lieu THAM KHAO M Heinrich, M Robles, J E West, B R de Montellano, E Rodriguez Ethnopharmacology of Mexican Asteraceae (Compositae), Ann Rev Pharmacol Toxicol, 38, 539 (1998) Cay có Viet Nam, tap III, trang Pham Hoang 275, Nxb Tre (2003) Jatem-Lasser, M S Ricardi, G Adamo Herbal traditional medicine of Venezuelan Andes: An ethnopharmacological study, Phytother Res., 12, S53 (1998) T Miura, K Nosaka, H Ishii, T Ishida Antidiabetic effect of Nitobegiku, the herb Tithonia diversifolia, in KK-Ay diabetic mice, Biol Pharm Bull 28, 2152 (2005) M Takahashi Compositions for curing diabetes mellitus, processes for the preparation of same, and usage of same, US Pat 5,773,004 (Fukuoka 1998, 814-01) Lien he: Trill Thi Thanh Thu), Phong Thi nghi'e'm ditm Cong nghe loc, boa dau S6 Pham NO Lao, Ha N6i Email: tranthithanhthuygl@gmail.com 773 ... Science and Technology, 29, 8-15 (200 9) Narayan C Baruah, Jadab C Sarma, Nabin C Barua, Soneswar Sarma, Ram P Sharma Germination and growth inhibitory sesquiterpene lactones and a flavone from... hydroxyl va carbonyl dang a, /3-chtra bao hoer PhO ESI-MS a che de) ghi ion am va &rang lAn Itigt cho cac pic [M-H]" tai m/z 315, [M+Na]+ va [M+H]+ tai cac gia tri m/z twang 1rng la 339 va 317 Tir...Trein Thi Thanh Thiry va Ong sy- TCHH, T 51( 6), 2013 diclometan Cit loai dung mei dual ip suit giam thu dtrqc can cac cao chiet Wang ling Cao chiet diclometan (15,7 g) duqc phan tach bang sic ky