TAP CHi KHOA HQC D^I HQC VAN LANG Trirffng Th4 Quang
HOAT TINH CHONG OXY HOA CUA SAPONIN TRONG CAO CHIET Ttr THAN RE CAY BAY LA MOT HOA (Paris polyphylla Sm.)
ANTIOXIDANT ACTIVITY OF SAPONIN EXTRACTION FROM RHIZOME OF SEVEN LEAVES ONE FLOWER TREE (Paris polyphylla Sm.)
TRVONG TH£ QUANG^*^
TOM TAT: Xdy dung quy trinh tach chiet vd uac luong hieu sudt thu nhgn cao ehiet tic thdn ri cay bdy Id mgt hoa la 22,03%. Phdn bo hdm lugng saponin todn phdn trong cay bdy Id mgt hoa nhiiu nhdt a thdn re 7,19%, a thdn Id 0,89% vd thdp nhdt & Id 0.49%. Hieu sudt cao chiit tic thdn re cdy bdy Id mgt hoa dgt 20,03%, dg dm cua cao chiit 18,02%, do tro eaa cao chiet 5,86%, hdm luong saponin todn phdn trong eao chiit 61,30%. Khdo sdt khd nang chong oxy hoa theo nong dg saponin vd u&c lugng hogt tinh chong oxy hoa caa saponin trong cao chiit bdng phuang phdp MDA duac ICS50 = 159,35 pg/ml Khdo sdt khd ndng chong oxy hoa theo nong dg ehdt chudn trolox vd u&c luong hogt tinh chong oxy hoa caa trolox bdng phuang phdp MDA duac ICT50 =-562,81 pg/ml. Hogt tinh chong oxy hoa cua saponin trong cao chiit mgnh han chdt chudn trolox 3,53 Idn.
Tit khoa: cdy bdy Id mot hoa; thdn ri; cao chiet; saponin; trolox.
ABSTRACT: The extraction process has been built and the extraction yield from the rhizome of Paris polyphylla is estimated at 22.03%. Distribution of total saponin content of Paris polyphylla in rhizomes is the highest with 7.19%, while the stem is 0.89% and in the lowest leaf is 0.49 %. Extractration yield of the rhizome of Paris polyphylla is 20.03%, the moisture content of extracts is 18.02%, the ash content of extracts is 5.86%, the total saponin content of extracts is 61.30 %. Examination of antioxidant capacity by saponin concentration and estimated antioxidant activity of saponin in extracts by MDA method is ICsso = 159.35 fig/mi Examination of antioxidant capacity by trolox standard concentration and antioxidant activity estimation of trolox by MDA method is ICTSO = 562.81 fig/ml The antioxidant activity of saponin in extracts is 3.53 times stronger than that of trolox standard.
Key words: Paris polyphylla; rhizome; extracts; saponin; trolox.
1. DAT VAN DE oxy hda. Vi v^y, viec nghien curu tao ra chat NbQng tac dong tir ben trong hoac ben ehdng oxy hda dl bd sung da va dang dugc ngoai CO th§ con ngudi gay ra su mix can bing cac nha khoa hoc tim kiSm va phat tnhn.
cua he thdng ehdng oxy hda. Nhilu can benh Nhung kham pha vl hogt tinh ehdng oxy hda hien nay dugc chiing minh do quy trinh stress cua mot sd loai dugc lieu va cac hop chat cd ngudn gdc tu nhien dugc uu tien hang diu vi
'' TS. Trudng Dai hpc VSn Lang, [email protected], Masd: TCKHl 1-12-2018 23
TAP CHi KHOA HQC DAI HQC VAN LANG So 11, Thang 9-2018 tinh nang va dg an toan, cung nhu hieu qua
trong viec ngan ngCra va chiia tri cac loai benh nhu ung thu, xa vGa dong mach,... Dac biet, hgp chat ehdng oxy hda d thuc vat chiem da s6 nhu saponin, carotenoid, flavonoid, vitamin C, vitamin E [9, tr.44-84].
Cay bay la mdt hoa {Paris polyphylla Sm.) va nhung loai thuc vat khac cung chi la nhung cay thuoc thuge dien quy hiem d Viet Nam dugc sir dung de chong khuan, chong nam, chiia ran can, cam mau,... trong dan gian [3, tr.90-92].
Thanh phan cua chiing chiia nhieu hgp cbSt cd kha nang chong oxy hda nhu s^ronin dugc ling dung trong Imh vuc y hpc.
• SipD01EtlllcllXTJJld(ill(')
^ DaiMsuna T. CnmUliiu
1 SCDOUUI
1 Spnowlni 5 SDlxudui 6 Ciutnictlac
Hinh 1. S a d o phan loai saponin [5, tr. 3-15]
Saponin la glycoside phan b6 kha rpng trong thuc vat. Dua vao e t o true aglyeon, nguW ta chia saponin thanh hai nh6m 16n \k saponin steroid va saponin ttiterpenoid.
Rieng saponin triterpenoid gdm c6 pentacyclic (nhom olean, nhom ursan, nh6m hopan,...) va tetracyclic saponin (nhom dammaran, nhom cucurbitan,...) [4, tr.126-127]. Saponin c6 hojt tinh khang khuan, khang njm, khang viem. Saponin nhom spirostan c6 nhiSu ehdt c6 boat tinh khang ung thu. Cac glycoside spirostanol CO tren b6n dan vi dudng thi thdy c6 tac dung ch6ng ung thu ro r^t. Tac dung huong
dye, tac dyng lien he t h t a kinh, tac dung len dOng v$t mau lanh va con trimg dupe ling dung de chong m6i, lam liet giun,...
Saponin steroid dupe dung dS t6ng hpp cic noi tiet to steroid nhu testosterone, progesteron, cortison [5, tr. 3-15].
Nghien cihi hoat tinh ch6ng oxy hoa (HTCO) cua saponin trong cao chi^t lix than ri cay bay la mpt hoa 1^ can thiet, nhSm gop phli l^m sang to thanh ph§n hoa hpc ciing nhu gia trj ve dupe lieu chiet xuat tu than re cSy nay.
Hinh 2. Than re cSy bay la mOt hoa (Paris polyphylla Sm.) [7]
Trong nghien cuu nay, FTTCO ciia saponin trong cao chi6t dupe djnh nghia la n6ng dp saponin ICsso (ng^ml) ung vai 50 % kha nSng bit g6c ttr do DPPH (1,1 - diphenyl- 2 - picrylhydrazyl) hoac ling vol 50 % kha nSng uc che peroxy hoa lipid t l bao bang phuong phap MDA (Malonyl Dialdehyd).
Kha nang bat g6c tu do DPPH hoac kha nang lie chS peroxy hoa lipid tS bjo dupe gpi la kha nang ch6ng oxy hoa va ky hi$u la KNCO (%).
TAP CHI KHOA HQC DAI HQC VAN LANG TriroTig The Quang O Viet Nam, cac de tai nghien ciiu ve
thanh phan hda hgc va tac dung dugc ly cay bay la mdt hoa van cdn ban chL Hien nay, Vien Dugc lieu cd \\m trii mdt sd cdng trinh nghien ciiu ve cay bay la mgt hoa nhung da sd la tai ligu nude ngoai, chii y8u la cac kk qua khao sat thuc trang va bao ve ngudn gene cua loai cay nay tai Viet Nam [10].
Nam 2007, viec tim kiem cac san pham tu nhien boat tinh sinh hgc tir than rl Paris polyphylla ciia Dekota K. P. va c^c cdng s\r dan den su cd l£ip ciia bdn hgp chit saponin. Cau true bda hgc ciia chiing dugc xac dmh bang each so sanh quang phd vdi cac sd lieu da bao cao. Cac chat chiet xuat tu chloroform, ethyl acetate va butanol loai cay nay dugc tim thay cd tiem nSng lic ch8 tu nhe den trung binh ehdng lai enzyme tyrosinase va antileishmanial [2].
Nam 2010, Wang. G. X. va cdng su da bao cao ve tiem nang kiem soat boat dgng ciia san Dactylogyrus ixx cac hgp chat cd lap dugc trong than re Paris polyphylla. Ca bai hgp ch4t dioscin va polyphyllin D deu the hien kha nSng khang dang ke vdi Dactylogyrus [ 11, tr. 1102-1105].
Nam 2012, Xia Wu va cdng su da phan Mp dugc mudi mgt saponin steroid tir than rl ciia Paris polyphylla, trong dd cd bay saponin steroid dugc bik dk\. Cac hgp chat cd l§p da dugc thu nghiem cd kha nang gay ddc te bao ung thu bigu md vdm hgng va hien thi tac dung lie cb6 manh hon cisplatin [13,tr.33-143].
Nam 2014, Yan-Shi Wen va cdng sir da phan lap dugc chin saponin steroid tii than rl cua Paris polyphylla, ben canh bay hgp chdt da dugc biet den, cd hai saponin chua timg dugc bao cao trudc dd la dianchonglouosides A va B. Ngoai ra.
nhdm tac gia cung chiing minh rSng cac hgp chdt dugc cd lap ed dgc tinh kha manh vdi hai ddng t§ bao ung thu d ngudi HEK293 va HepG2 [14, tr.31-34].
2. MVC TIEU VA NOI DUNG NGHIEN CUU 2.1. Muc tieu nghien cii'u
Khao sat KNCO theo ndng do cua saponin trong cao chiSt tu than rl cay bay Id mdt hoa, udc lugng HTCO ciia saponm trong cao chiet va ddi chiing vdi chat chuan trolox.
2.2. Ngi dung nghiSn cuu
Xay dyng quy trinh chi^t xuit cao cdn tii than re cay bay la mgt hoa, udc lugng mgt sd chi tieu hda ly ciia saponin trong cao chiet, khao sat kha nang ehdng oxy hda ciia saponin trong cao chiet bing phuang phap bat gdc tu do DPPH (1,1 - dipbenyl - 2 - picrylhydrazyl), khao sat kha nang ehdng oxy hda cua saponin trong cao chiet qua kha nang lic che peroxy hda lipid te bao bang phuang phap MDA (Malonyl Dialdehyd), udc lugng HTCO ciia saponin trong cao chiet so vdi chat chuan trolox.
3. PHUONG PHAP NGHIEN CUU 3.1. Quy trinh chiit cao tir thSn re cSy bay la mdt hoa
Than re cay bay la mdt hoa dem phai nang cho khd, sau dd nghien va ray 3mm de cd bgt min, sau dd trich ly bang dung mdi cdn 80 do theo ty le 1:6 va cd dac chan khdng thu dugc cao chiet cdn ggi tat la cao chiet.
3.2. Phutmg phap dinh lugtig saponin tong s6 Ham lugng saponin tdng sd trong cao chiet dugc udc lugng bang phuong phap khdi lugng. Can chinh xac 0,5 g cao chiit, hoa vdi 10 ml nude cat va lac loai tap vdi diethyl ether, thu idy pha nude va I5c tiep vdi n-butanol bao hda nude.
TAP CHi KHOA HQC DAI HQC V A N LANG So 11, Thang 9-2018 A. 100
M(IOO-D) (1)
Trong dd: X: Ham lugng saporm traig m3u caodiia(%); A: Khdi lugng cin saponin thu dugc (g); D: Do dm ciia mau cao chiet (%);
M: Khdi lugng mau cao cbi§t (g).
Cd bay hoi n-butanol thu duge cin saponin. Sdy khd cin saponin d 105^C den khdi lugng khdng ddi va can [1, tr.390-396].
Ham lugng saponin theo nguyen li?u khd tuyet doi dugc tinh theo cdng thiic (1).
3.3. iTo'c lirgng kha nang chong oxy hda cua saponin b^ng phu-ong phdp DPPH
DPPH la ehSt mau tim cd gdc tu do nhd vao dien tu chua ghep ddi. Saponin la chat chong oxy hda cd kha nang bat gdc ty do cua DPPH lam giam cudng do mau. Vi the, nong do saponin trong mlu cang cao thi cudng dg mau cua DPPH cang giam, dugc xac dinh bang each do do hip thu A tren may quang phd hip thu phan tu UV- VIS d budc sdng 517 nm [1, tr.390-396].
Kha nang ehdng oxy hda KNCO (%) dugc tinh theo cdng thiic (2).
H]
100 (2)Trong do: Ay: Dp hap thu eua mtu thir;
Ao:Dph^thucuaniluliSng(chungdungm6i).
3.4. iTo'c lu-cmg kha nang chong oxy hoa cua saponin bSng phirong phip IMDA
Ucrc lupng kha nang chSng oxy hoa cua saponin trong cao chi& qua kha nang lie ch6 peroxy hoa lipid mang tk bao bjng viec xac dinh ham lupng MDA la san phta ciia qua trinh peroxy hoa lipid.
Trong do: AT: Bp hSp thu cua raau thu;
Arf Dph2plhucijamluti3ng(chungdungm60.
MDA CO kha nang phan ung voi acid thiobarbituric dS tao thanh phiic hpp trimethin mau hong c6 buac song hip thu cue dai 532nra [6, tr. 243-256]. Kha nSng ch6ng oxy hoa KNCO (%) dupe tinh theo eong thiic (3).
3.5. Ho^t tinh chong oxy h6a ciia saponin so \6i ch^t chuSn trolox
Trolox (&Ji>dnixy.2A7,84eliamehyldiiomai>
2-carboxylic acid) la chit chong oxy hoa nhu vitamin E. Hoat tinh chong oxy hoa ciia saponin trong cao chiet dupe tinh theo hoat tinh chong oxy hoa cua chat chudn trolox tuang duong, dupe do bing don vi trolox gpi la Trplox Unit (TU).
Uac lupng phuong trinh KNCO = f(x), trong do KNCO (%) la kha nang lie chS peroxy h6a lipid mang te bao theo nong dg X cua saponin (hoac trolox). Sau do, npi suy ra gia tri HTCO eua saponin (hoSe trolox) X50 = IC50 ling vai kha nang iic che peroxy hoa lipid KNCO = 50 %, gia tri IC50 cang nho tuang img vai HTCO cang Ion. HTCO cua saponin ICsso se dupe tinh theo don vj TU dua vao ICTSO ciia chat chuin trolox.
'•[-i]
100 (3)XV^O-^J OH
Hinh 3. Cdu tao hoa hoc ciia trolox [12]
TAP CHi KHOA HQC BAI HQC VAN LANG Truwig The Quang
4.2, Ham lugng saponin trong cao chiet tir than rl cSy bay I^ mgt hoa
Hieu sudt cao chi^t tCr than vi cay bay la mpt hoa dat 20,03 %, do dm trung binh cua cao chiit 18,02 %, do tro trung binh ciia cao chi6t 5,86 %, ham lugng saponin toan phan trong cao chiet 61,30 %.
4.3, Uoc lirgng KNCO cua saponin trong cao chiet bling phu-ong phdp DPPH
Phan tich phirong sai vdi miie tin cay 0,05 eho thay kha nang cbdng oxy hda ciia saponin trong eao chiet tSng ty le thuan theo ndng do saponin. Kha nang ehdng oxy hda cao nhat 78,36 % d ndng dg saponin 2000 |j.g/ml, kha nang chong oxy hda thap nhat 19,89 % d ndng do saponin 50 (xg/ml, xem Bang 2.
Bang 2. Kha nang chong oxy hoa cua saponin trong cao chiet theo phirang phdp DPPH [8]
4. KET QUA VA THAO L U . ^ 4.1. Phan bo hhm lumig saponin toan phln trong cay b§y Id mgt hoa
Phan tich phuong sai vdi miic tin cay 0,05 ham lugng saponin toan phin cd trong cay bay la mgt hoa nhilu nhdt d than re 7,19 %, cdn d thSn la 0,89 % va thdp nhit d ia 0,49 % (Bang I). Do dd, dung nguyen lieu 1^ than re cay bay Id mgt hoa di chiit xuat cao cdn cd chiia saponin.
Bang 1. Phan bo h£km lugng saponin trong cay bay Id mOt hoa [8]
Ham lirgng saponin (%) Than r l
7,19'
Than | La
0,89» 1 0,49"
Nong d9 saponin x (^g/^1l) KNCO (%)
2000 78,36"
1000 50,15"
500 41,10'
250 32,68'
100 29,06=
50 1
19,89' 1 4.4. if&c lu-ong KNCO cua saponin trong theo ndng do khao sat. Kha nang chong oxy cao chiet bdng phirong phap MDA hda cao nhat la 89,23 % d ndng do saponin Phan tich phuang sai vdi muc tin cay 2000 p-g/ml, kha nang ehdng oxy hda thdp 0,05 cho thdy kha nang ehdng oxy hda cua nhdt la 27,25 % d ndng do saponin 50 saponin trong cao chiet tang ty 1? thuan |ig/ml, xem Bang 3.
Bang 3. Kha nSng chong oxy h6a cua saponin trong cao chiit theo phuang phap MDA [8]
Ndng d$ saponin X (^g/ml) KNCO C%)
2000 89,23'
1000 87,25'
500 78,24'
250 59.78'
100 38,02'
50 27,25'
4.5. Hoat tinh chong oxy hda ciia saponin trong cao chi^t tinh theo don vi trolox
Kha nang cbdng oxy hda cua trolox theo phuong phap MDA dugc neu trong
Bang 4. Kha nang chong oxy hoa cua trolox theo phucmg phap MDA [8]
bang 4, phan tich phuang sai vdi miic tin cay 0,05 eho thay KNCO phu thuge vao ndng do trolox.
Nong dO trolox x (Hg/ml) KNCO (%)
2000
77,70' 1000
63,49' 500 44,08'
250 29,96'
100 14,86'
50 5,32'
TAP CHi KHOA HQC S A I HOC VAN LANG So 1 1 , T h a n g 9-2018
Tit s6 heu Bang 3, udc lupng dupe phuong h-inh KNCO cua saponin trong cao chia, phuong trinh (4) voi binh phuong he s6 tuang quan R^ = 0,961 = 1.
KNCO = 18,397.ln(x) - 43,293 (4) HTCO cua saponin trong cao chiSt ICsso
= 159,35 |jg/ml ijmg vcri KNCO = 50 %.
K.NC0=lS.3971ii(i).43J9i R- = 0.9(1 iaTO(%) ICsso»159,35 re'ri
0 :50 500 "SO IMO i:5fl 1500 n50 20OO Nong <]9 saponin (^g'mi)
Hinh 4. Dfl thi bieu diln tuang quan gifta KNCO {%) va nong do (ng/ml) ciia saponin trong cao chiet [8]
Tir so lieu Bang 4, uoc lupng dupe phupng trinh KNCO cua trolox, phuang trinh (5) voi binh phuong he so tuong quan R^ = 0,987 a 1.
KNCO = 19,964.1nCx) - 76,431 (5) HTCO ciia trolox ICTSO = 562,81 ng/ml ung voi KNCO = 50 %.
K.\CO=19,9611n(s) .76431 R- = 0,9S) K.NCO(%) lCT<0 = 56;»liigml
0 250 500 750 1000 1250 15m 1750 2000 N6ngdotioIox({iEni])
Hinh 5. Do thj bieu dien tuang quan giira KNCO (%) va n6ng d$ (ng/ml) ciia trolox [8]
HTCOs ciia saponin trong cao chiet tir than re cay bay la mpt hoa cao hon trolox 3,53 Ian. HTCOs ciia saponin trong cao chiet tu than re cay bay la,mpt hoa tinh theo don vj TU:
HTCOs = ^ ^ = 3,53 (TU) ICsso
5. KET LUAN VA KIEN NGHI S.l. Ket luan
Xay d\rng quy trinh tach chiet va ucrc lupng hieu suat thu nhSn cao chiSt tCr than rl cay bay la mot hoa la 22,03%.
Phiin bd ham lupng saponin toan phan trong cay bay la mpt hpa nhieu nhat a than rl 7,19 %, eon a than la 0,89 % va thip nhit a la 0,49 %.
Hieu suit cao chiet tu than re cay bay la mot hoa dat 20,03 %, dp im ciia cao chiit 18,02 %, dp tro cua cao chiit 5,86 %, ham lupng saponin toan phan trong cao chiit 61,30%.
Khao sat va uoc lupng kha nang chong oxy h6a theo nong d6 saponin trong cao chiet bang phuang phap DPPH cho thay kha nang chong oxy hoa ty le vai nong dfi saponin. Phan tich phuong sai vol mirc tin cay 0,05 cho thay, kha nang ch6ng oxy hoa cao nhit 78,36% 6 n6ng dp saponin 2000 Hg/ml, kha nang ch6ng oxy hoa thip nhit 19,89 % o nflng dp saponin 50 ng/ml. Khao sat kha nang chfing oxy hoa theo n5ng dp saponin va uoc lupng hoat tinh ching oxy hoa ciia saponin ttong cao chiit bSng phuong phap MDA dupe ICsso = 159,35 ng/ml. Khao sat kha nang chong oxy hoa theo n6ng dp chit chuan trolox va uac lupng hoat tinh ching oxy hoa ciia frolox bing phuang phap MDA dupe ICTSO = 562,81 ng/ml.
28
TAP CHi KHOA HQC DAI HOC VAN LANG TrtroTig The Quang Hoat tinh cbdng oxy hda ciia saponin 5.2, Kien nghi
trong cao chiet manh hon chat cbuin trolox Ti6p tue nghien ciiu quy trinh tach 3,53 Ian. Nhu vay, n6u tinh theo don vi chi^t, cd l$p va xac dinh ciu tciic hda hgc trolox thi HTCO cua saponin trong cao tung loai hgp chdt saponin tii than re cay chik bang 3,53 TU. bay la mdt hoa.
Nghien ciiu ung dung ciia cao chiet tii' than re cay bay la mdt hoa de phong va chua b?nh.
TAI LIEU THAM KHAO
[I] Boonchai Pongnaravane, Motonobu Goto, Mitsuru Sasaki, Thitipom Anekpankul, Prasert Pavasant, Artiwan Shotipruk (2006), Extraction of anthraquinones from roots of Morinda citrifolia by pressurized hot water: Antioxidant activity of extracts Original Research Articlee, The Journal of Supercritical Fluids, Vol 37{3).
[2] Devkota K. P., Khan M. T., Ranjit R., Lannang A. M., Samreen, Choudhary M. 1. (2007), Tyrosinase inhibitory and antileishmanial constituents from the rhizomes of Paris polyphylla.
[3] Dd Tat Lgi (2015), Nhirng cdy thuSe vd vi thuSc Viet Nam, Nxb Hdng Diic.
[4] Nguyen Kim Phi Phung (2007), Phuong phdp c6 lap hgp chdt hieu ca, Nxb Dai hgc Qude gia Thanh phd Hd Chi Minh.
[5] Ngd Van Thu (1990), Hda hgc saponin, Nxb Y hgc Ha Ndi.
[6] Stroev E. A., Makarova V. G. (1998), Determination of lipid peroxidation rate in tissue homogenate laboratory. Manual in Biochemistry, Moscow.
[7] Thuoc vudn nha (2012), Cdy bdy Id mgt hoa cd tdc di^ng gi?, http://www.tbuocvuonnhacom/c/cay-bay-la-mot-hoa-co-tac-dung-gi/hoi-dap, Truy cap 16/05/2015.
[8] Truang The Quang, Tran Thi Nggc Anh (2016), Nghiin cicu hogt tinh chSng oxy hoa caa cao chiet tic thdn re cdy bdy Id mot hoa (Paris polyphylla Sm.), Trudng Dai hgc Van Lang.
[9] Valko M., et al (2007), Free radicals and antioxidants in normal physiological functions and human disease. The International Journal of Biochemistry & Cell Biology, 39.
[10] Vien Dugc lieu (2016), Danh muc cdc cong trinh nghien cuu vi cdy bdy Id mgt hoa (Paris polyphylla var. yunnanensis), Trung tam Thdng tin Thu vien.
[II] Wang G. X., Han J., Zhao L. W., Jiang D. X., Liu Y. T., Liu X. L. (2010), Anthelmintic activity of steroidal saponins from Paris polyphylla, Phytomedieine.
[12] Wikipedia (2017), Trolox, https://en.wikipedia.org/wiki/rroIox, Truy cap ngay 19/05/2018.
[13] Xia Wu, Lei Wang, Hui Wang, Yi Dai, Wen-Cai Ye, Yao-Lan Li (2012), Steroidal saponins from Paris polyphylla var. yunnanensis, Phytochemistry.
[14] Yan-Shi Wena, Wei Ni, Xu-Jie Qin, Huan Yan, Chang-Xiang Chen, Yan Hua, Yung- Chi Cheng, Li He, Hai-Yang Liu (2014), Steroidal saponins with cytotoxic activity from the rhizomes of Paris polyphylla var. yunnanensis, Phytochemistry Letters.
Ngay nhan bai: 21-5-2018. Ngay bien tap xong: 28-5-2018. Duyet dang: 24-9-2018
