TAP c i i CBNG TItfONG
KHAO SAT HAM LI/0NG
POLYPHENOL TONG, SAPONIN TONG VA HOAT TINH KHANG OXY HOA
Tl/ CAO CHIET CAY DINH LANG (POLYSCIAS FRUTICOSA (L.) HARMS)
• NGUYEN DLTC DO - TH! LINH DUYEN - HUYNH THANH DUY - IRAN NHAN DUNG
TOM T.\T:
Cao chie't dinh ldng difdc dung de khdo sdt ham Ifldng polyphenol, saponin tdng s6v^ khd ndng khdng o\\ hda didng qua phfldng phdp khfl DPPH vd khiV peroxide. Kd't qua cho tha'y, hdm Ifldng polyphenol long cao nha'l d cao chie't Id - methanol vdi 72,003 mg GAE/g. Dinh Ifldng saponin tong cho thd'y ham Ifldng cao nha't trong cao chie't Id - ethanol vdi 118,44 mg/g. Kha ndng oxy hda cua cao chie'i Id - ethanol cho hieu qua cao nhd'l vdi gid tri IC50 dd'i vdi DPPH vd peroxide lan lifdt Id 266.3 pg/mL va 226,13 |ag/mL.
Tiif khda: Dinh ldng, DPPH. khfl gdc peroxide, polyphenol tdng, saponin tdng.
l.Ddtvdnde
Cdc gdc phdn flng chfla o\\ (ROS) \a cdc gd'c phan flng chfla nitd(R.NS) dfldc taora trong cd the con ngifdi. Chiing cd the ga> ra nhflng ton thfldng o\\ hda lien quan de'n mot so' benh nhif xd vfla ddng mach. benh mach vdnh. parkinson, lieu dfldng, thoai hda than kinh (Alzheimer), lao hda
\a ung thfl. \1 \ a \ . viec can bdng sif sdn xud't gdc tfl do \ a mflc do chd'ng o\\ hoa la rat can ihie't cho
•-dc khde (Rungthip Kaw aree \ a Sombal Chow wanapoonpoh.2009)
Pol\ phenol difdc biel den la lac nhan chd'ng oxy hda, CAC nghien cflu da chi ra dan xud't polyphenol cd the bdo \e protein vi cac \at lieu di iruven khdi
346 So 8 - Thdng 4/2020
sfl phd huy cua gdc tfl do {Gharass, 2009). Saponin cd nhieu hoal tinh sinh hgc trong dd cd ldm tan mdu, bao ve gan. giam suy nhfldc, khdng virus, chdng kl sinh trung va khdng viem (Rahimi et al., 2009). Hdp chd't nay cd the dfldc lim tha'y Ifldng ldn irong cdc ioai thflc vat ho Araliacea, nhfl dinh ldng.
O Viet Nam, nguon cay dinh lang [Polyscias fruticosa (L.) Harm) doi ddo, gia thanh va gid trj thdp. Cdy dinh lang cdn di/dc gni la cay goi cd va dfldc trong phd bie'n de lam cdnh. lam ihudc va gia vi (Dd Ta't Ldi. 1995). Thay diffic tiem nang tan dung nguon nguyen lieu, mang bi cac gid tri sflc khde cho con ngfldi. nghien LLVU difdc thflc hien de khao sat khd ndng chdng nxv hda, ham Ifldng
HOA IOC-CONBNEHElifePIAM
ntJ^tibenol va saponin cua cdc chie't xudt bdng 4ing moi khde nhau d Id va ra dinh ldng. Tfl dd. cd yijnp i^n dung ngudn dinh lang doi dao vdo ihifttngmaihda.
;. Dfli tfli^ng va phifdng phap nghian ciJu Hda ch^t: ethanol (EtOH) (Vidt Nam), methanol (MeOH) (Viet Nam), H2SO4 dam dac (Trung (Juoc), acid gallic (Trung Quo'c), thud'c thu"
Fbhn - Ciocalteu (Dflc), Na2C03 (Trung Qudc).
NaH,P04.2H2O (Trung Qudc), Na2HP04.12H20 (Trang Qudc), H2O2 (Trung Quoc), vitamin C (Trang Qudc). 2,2-diphenyl-1 -picrylhydrazyl (DPPH) (My).
Nguyen !i6u: Re va Id cay dinh ldng (Poly.scias jnuicosa (L.| Harm khong bi sau benh va tdn ihiWng,(lif^c thu hdi vao sdng sdm cf iinh An Giang.
Sau d6 rifa s^ch vd di rdo nfldc d nhiet dp phdng.
Miu \i va rl dfldc xay nhuyen vdi dung mdi (ty le mau: dung mdi la 1:5) kdt hdp vdi xu" ly sdng sieu am i20W trong 60 phdt vd chia thdnh 4 nghiem ihtfc, m6i nghiem thflc gdm 500 gram mdu (la hoae re)du'0c ly trich vdi 2500 mL (methanol 96% hodc eihanol 96%). Mau dfldc de yen trong 24 gid, rdi Uen h&nh loc, cd quay chan khdng de thu dfldc 4 loai cao chie't: la - ethanol (LET), la - methanol (LME). re-ethanol (RET) va rl - methanol (RME).
Djnh lmh cac h^p chd't thi^c vat bang phifdng phap do quang: Thflc hien theo quy trinh chuan cilaHarbome(1973).
Dinh Iflt^g ham Wdng polyphenol tdng: Thflc hien dtfa theo mo td cua Yadav vd Agarwala (2011) c6hi$u chlnh.
Cho 0,5 mL dung dich cao chie't (ndng dp 0,5 mg/mL) hodc dung dich gallic acid (20,40,60, 80, iOO, 120pg/mL) vao d'ng nghiem. Sau dd. them I.--"' mL thud'c thfl Folin-Ciocalieu, de phan flng
•rong 5 phul. Tie'p de'n them vao 1 mL dung dich Na,CO, 29!-. Sau 45 phut phan flng d nhiet dp phing. lie'n hanh xdc dinh dp ha'p thu bdng mdy J" 4"ang phd d bfldc sdng 765 nm. Dfldng chuan I* dung djch gallic acid. Gid tri OD du'cfc ghi nhan
^i ti&n hanh ve dfldng thang hieu chuan de xac
^oh hSm Iflcfng polyphenol tdng trong cdc mdu ao chiet.
Him lifdng polyphenol long dfldc tinh bang each
dfla vao phfldng trinh > = ax + b ciia difdng chua'n acid gallic.
Ham lifdng polyphenol tdng: C = c \ V/m Trong dd: C: hdm Ifldng polyphenol tdng (mg GAE/g chie't xuat I; c: gia in \ ti^ difdng chua'n \di acid gallic (pg/mL); V: the tich dich chie't (mL): m:
khdi Ifldng cao chiet cd trong the lich V (g).
Dinh Iifdng ham Ifldng saponin tdng: Thflc hien iheo md td cua Hiai vd cpng su^l 1976) cd hieu chinh. Chat chua'n dfldc sfl dung la Rb 1. Rg 1, Rg3 va cdc nghiem thflc cao chie't du'de hda tan bdi dung mdi methanol. Thud'c thfl Id vanilin 8% dfldc hda tan bdi dung mdi ethanol va dung djch H-,S04 72'.;.
Lan lifdt cho 500 pL chat chuan vdi ndng dp (10 - 60 pg/mL) hodc cao chie't vao dng nghiem. Tidp theo. them vao iOO pL dung djch vanillin 8%. Sau do, them vao 2 mL HiSOj 72% va trpn deu. Hdn hdp dfldc u irong be d 60"C trong 10 phut, sau dd lam lanh trong 15 phut. Tie'n hanh ghi nhdn ket qua dp hd'p thu cua mdu lai bfldc sdng 550nm. Mau ddi chflng du'de thflc hien chi bd sung hdn hdp vanillin 8% vd dung dich HiSOj 12%. Hani Ifldng saponin dfldc tinh bdng cdch dfla vao phfldng fllnh y = ax + b cua dfldng chuan lai cac dp hd'p thu cua chat chuan.
(D-b) Saponin =
axV - x N / W
Trong dd: D: Do ha'p thu cua mdu; a.b: gid tri ifl dfldng chuan; V: the lich cua dich chiet (mL); N: dd pha lodng; W: khd'i lifdng cao chie't cd trong the lich V(mg).
Khao sat kha nang khang oxy hda bdng DPPH: Thflc hien theo md id cua Blois (1958) cd hieu chinh. Hoa ian cao chie't. vitamin C (dimg lam dfldng chuan) va thud'c thfl DPPH (2.2-diphenyl-l- picr>'lhydrazyl) 0.1 mM bdng dung mdi methanol.
Lan Ifldt cho 1 mL vitamin C (nong dp 2.4; 6; 8; 10:
12 pg/mL): hodc cdc nghiem thflc cao chie't va 2 niL DPPH 0.1 mM vao d'ng nghiem. Tiep Uieo. u trong ldi d nhiet phong trong 30 phut. Sau dd. tie'n hanh do dd hd'p thu d bfldc sdng 517 nm. Mau control dfldc thflc hien tfldng tfl nhflng Uiay the cao chidt bdng MeOH. Kha ndng flc chd DPPH difdc tinh theo cdng thflc sau-
Phdn trdm flc chd DPPH =-'^-"^- ^ IOO
So 8-Thang4/2020 347
TAP cij C i K riirtfNG
Trong Jd .\; dd ha'p thu cua mdu ddi chung (khdng chfla cao chiei i. -\ dp hd'p thy cua mau.
Nu\ duni; dfldng chuan \ ^ a\+b vdi phan triim flc che' DPPH d cdc ndng Jo khac nhau. Tfl dd. tinh gid in IC50 cija vitamin C ha\ cao chiei.
Khao sat kha nSng khuT gdc peroxide bdng H2()>: Thflc hien theo md ta cua Rahaie va cdng Hfi2or^).
\fl Iy sd li^u: So lieu thi nghiem dfldc xfl ly bdng phan mem Excel va phan iich thd'ng ke bdng ehfldng uinh \Iinii.ih 16, phdn tich ANON* A. cdc gia tri trung hinh dfldc so sdnh bdng phep thfl Tuke\.
3. Ket qua va thao luan
3.1. Djnh tinh mgt sdhcfp chd't thUc vgt Sfl hien dien cua cdc hdp cha'i hda hpc thflc vat dfla vdo dd ha'p thu ciia hdp chd'l dd tai cdc diem bu'dc sdng. Dp hd'p thy cang cao cho ihd'y ham Ifldng cdc hdp chat hien dien nhieu trong mau.
Ke't qua dinh tinh (Hinh 1) cho Iha'y trong dinh langcd hien dien cdc hop cha'l hda hpc thflc \di phd bien nhfl steroids, trilerpenoids. phenolics.
quinones, tannins. Ilavonoid, carolenoids. saponins
\d alkaloids. Trong dd, sfl hien dien cua cdc hdp chat trong chiel xua't bang dung mdi ethanol cho
ham Ifldng cdc hdp chat cao hdn so \di chiet xuat bdn2 dung mdi methanol. Ethanol 95'i la dung mdi dfldc si:^ d~ung phd bien cho L'\' vi hau het cdc hdp cha'l dd hda tan ffong nci.
Nhif va\. trong cdc nghiem thflc cao chiel dinh ldng da phat hien dfldc h^u het cdc hdp chd't hda hpc thflc vat quan d-ong. trong dd cao chiel dfldc ly trich bdi dung mdi ethanol cho ham Ifldng cdc hdp cha't cao hdn dung mdi methanol.
3.2. Dinh lUgng polyphenol vd saponin tdng Gid tri mg GAE/g chie't xua'l (ham Ifldng polyphenol tdng), mg/g chie't xuat (ham Ifldng saponin tdng) cang cao thi ham Ifldng chung cd trong cao chie'i cang cao vd ngifdc lai.
Hdm Ifldng polyphenol tong trong cdc nghiem thflc cao chie't dinh lang dao dpng trong khodng 20,71 ± 0.271 de'n 71.4656 ± 1.437 mg GAE/g chie't xua't. Cdc nghiem thflc cua Id dinh ldng cho thd'y hieu qua chie't xua't cdc hdp chd't polyphenol tdng cao hdn cdc nghiem thflc cua re. Theo Lu vd Foo (1995), trong qud tnnh quang hdp, thflc vdt sdn xud't ra cdc hdp cha't phenolic de chd'ng lai cdc gd'c tfl do oxy hda (ROS), nen cd sfl chenh lech gifla ham Ifldng phenolic d Id va re dinh ldng.
Tren ciing mpt bp phan (Id hodc re), cdc cao chiet
Hinh 1: Quang pho UV-Vis cua cdc nghiem thCfc cao chiet Dinh Idng
1.2 -.
1 2-0.8
^ ^ ^ ^ ^ ^ ^ ^
- L M E -LET - R M E -RET
205 255 305 355 405 455 505 555 Birdc song (nm)
348 So 8 - Thong 4/2020
IDA IOC CONG NGHE l i n e P I K M
Bang 1: Ham Wang p o l y p h e n o l t d n g v d mxxiln l^ng trong c a c c o o ctiiet dinti l o n g
HSmluiWPolypt'S''"'' I * * " iSig(mg GAE/g ( * «
xu«)(1)
IME LET RME RET
t t a m IUdng s a p o n i n long (mg/g chiei
xua)(2)
77,16±6,62»
'GIti chti: (I): cdc gid Irj trong cdt ndy dtfdc xdc djnh dtfa vdo pltitditg Irinh ducfng chuan cda galltc ccid (y = 0.0I26X - 0.0329: r2 = 0,9994). (2): cdc gii trj Irong CQI ndy dttpc xdc dinh dua veto phUcfng ttinh dudng chudn cua Rbl. Rgl. Rg3: y = 0.0!99x + 0,iJS9; fl = 0.9968). Trong cung mot cor, cdc so tnmg binh theo sou hdi mgl hoae nhihig chd cdi giAtg nhau Ihi khde biel khong co y nghTa Ihong ke dtntk^ nghia 1 % bang phep Ihii: Tukey.
dlUfc ly trich vdi dung m6i metlianol co liam lifdng pttenolic long niiieu httn cac cao ctiie't di/dc tricti tiing dung moi ellianol (Bang 1). So sanli vdi ham ll/i;ng phenolic long Irong dich trich melhanol la dinh liing tro (Polyscias guilfoylei) la 10,36 mg GAE/g chiel xuii'l (Sundu va cpng sir, 2015), thap hdn khodng 7 lan so vdi doi lirpng khao sal trong nghien ci?u niiy.
H&m Iifdng saponin Id'ng cd Irong cac nghiem thifc cao chie'i tit 40,38 ± 1,26 de'n 118,44 ± 3,44 mg/g chi6't xuat. Kel qua cho tha'y, ham iifpng saponin u-ong cac nghiem Ihifc la dinh lang cao iKJn c5c nghiem thd'c rh va khac biel cd y nghia ihimg ke
Nhif v4y, vdi dung mdi melhanol cd the ly trich iJiWc nhieu ham Iifpng polyphenol tdng dien hinh d nghifm thifc LME, dung mdi ethanol cd kha nang chiet xua't nhieu ham Iifdng saponin tdng, the hien S nghiem thUc LET. Trdng ca 2 thi nghiem, ham li^ng polyphenol long va saponin tdng trong la cao lK*isovdii^,
3.3. Khi ndng khdng oxy hda cua cao chiel dinh ldng
Hoat tinh chdng o\\ hda ciia cac cao chie't dinh lang difpc danh gia qua kha nang khif gdc lif do DPPH (2,2-diphen\l-l-picr>lhydrazyl) \i loai bo chat oxy hoa hydrogen peroxide (H,0,), khii nilng khang oxy ciia cao chie'i difpc the hiiin qua gia tri nong dp Clia man ma Iai dd cd the ifc che .SO^ gdc lir do. Giii tri IC^n cang Uiii'p mau sc cd hoat Iinh chd'ng oxy hda cang cao va ngifdc lai,
Bdng 2: Gid trj ICjo c i j a c a c nghidm ttiCfc c o o chid't dinh Idng vdi ttiuoc thijf DPPH v d HjO;
Nghi$m thtic
Nong d$ cao chlgtijic Nong d$ cao chiet Lie che50%DPPH(IC5o) cheSD'AHAtlCso)
(|ig/mL) (Ma/mL) LME ' 573 + 26,8=
LET I 266122,9"
RME 1519±4L7' RET I 1220±59,9«
iVilaminC 6,0C1±25,8'
23017,99' 22611.1"
23812,71"
24912,24' 12210,61'
*Ghi cltu: Trong cung tngl col. ctic so'trung binh theo sau bdi mot hodc nhiing chd cdi gidng nhau lhi khde bill khong cd y tiglud tliong kc d mt)c y ttghJd I % bdng phep tlni Tukey.
Theo Bang 2. tii't ca cao chie't deu cd kha niing chdng dxy hda va xu hifdng khang oxy hda lifdng lif nhau qua 2 Ihiir nghiem, Vdi gia tri IC,,, nho nhal lan lifdl la 266 ± 22.9 va 226 ± 1,1 ng/mL 6 2 thi nghiem khang oxy vdi diudc Ihct DPPH va H,0,, nghiem thifc LET cho kba nang khang oxy hda manh nhii'i. Cung vdi la Salacca wallichiana khi chiel xua'l vdi dung mdi methanol va ethanol, bang Ihif nehiem khiJ gd'c uf do DPPH cho giii tri IC,,, lan lifpi l"a 86,03 ± 0,08 va 28,36 ± 0,20 pg/mL (Deepa vaNalini,2013)
Theo Babu et al. (2013) vii Balasundram el aL (2006). kha nang khang iix.v hda lot cua cic chii-I xua't thifc vat la do sit hien dien ciia c^c nhdm chat
So 8 - Thdng 4/2020 349
TAP c i i CONC TI^QiNS
nhu" phenol va tannin, flavonoid. alk.iloid.
gl> coside va saponin nhd vdo so' li/dng va \ j tri c u a nhom h y d r o \ \ l lien quan den nhom carbon chi?c ndng. K t i qua na> phu hdp vdi ket qua dinh tinh.
dinh li/dng ham li/dng pol>phenoi long va --aponin long infdc do Hon nffa. hoat dpng chong o \ \ hoa khong chi dya vao cac hdp chat polyphenol, ma c5n CO su" tde dpng cua cdc thdnh phan khac nhtf carotenoid. vitamin vd c a c khoang chat (Ratnam et ai.. 2006). Do dd. co sd khac biet \ e moi ttfdng quan giffa ham Iffdng polyphenol \ a saponin den hoai tinh khang oxy hoa.
6 dich inch methanol la dinh lang trd (Polyscias guilfoylei). Sundu va c p n g SI/{2015) da bdo c a o rdng hoal d o n g k h a n g oxy hda c u a dich trich nay c h o n g lai DPPH va H . O , vdi gia tri IC^Q lan Iffdl la 1880 \ a 1320 pg/mL. Dieu ndy cho tha'y cl loai dinh ldng nay c6 hieu qua khdng oxy
hda y e u hdn do'i tifdng dinh ldng dffdc k h d o sdt irona n a h i e n cffu.
N h f f \ a v . dun2 mdi ethanol d chiet xuat id dmh ldng cho hieu qua khdng oxv toi nha't so vdi cdc n g h i e m Ihffc con lai. v di n g h i e m thffc L E T cd kha n a n g cho'ng o\> hoa m a n h nhat.
4. K e t l u a n
Trong c a o chie't dinh ldng cd hien didn cdc hdp cha'l hda hgc Ihffc vai phd b i e n nhff steroids.
trilerpenoids, phenolics. q u i n o n e s , tannins.
flavonoid. carolenoids, saponins vd alkaloids. Hdm lffdng polyphenol tdng c a o nha't d n g h i d m thffc Id - methanol ( L M E ) va n g h i e m thffc la - edianol (LET) cho hdm Iffdng saponin tdng c a o nhat. C d c nghiem thffc c a o chie't la vd re dinh Iang d e u cd khd ndng khdng oxy h d a (qua thff n g h i e m khdng oxy hda vdi DPPH va p e r o x i d e ) vdi c a o chiet L E T d a i khd ndng khdng hidu qua nha't •
T A I I.IKU T H . \ M K H A O :
1. Do Tat LiJi (1995). MiiTng cdy thude va Vvi ihudc Viel Nam. Nlid xua't bdn Khoa hpc vd Ky thuat, trang 479-482.
2. Balasundram. N.. Sundrani. K. and Samman. S. (2006), Phenolic coinpound.s in plants and agri-industrial by-prodiieli: Antioxidanl aclivity. occurrence, and potential use.w Food Chemistry 99: 191-203.
3. Deepa Babu. Prema Gurumunhy, Sai Krishna Borra and K M, Cherian (2013). Antioxidant and free radical -.cn cnuiiig aciivny of iriphala deiemiined b\ using different in vitro models. Joumal of Medicinal Plant Research. Vol.
7(39).pp.2S9S-2905.
4. Gharass E. H.. 2009. Polyphenols: food sources, properties and applications - A review. Internalional Journal of Food Se-ienee and Technology. 44' 2512-2518.
5. Hiai S. Oura H. Nakajima T. (1976) Mar Color reaction of.some sapogcnins and saponins with vanillin and sulfuric (iffV/. Plama Med. 29(2): 116-22
6 Rahimi. R.; Gbiasi. .S . Azimi. H ; Fakhari. S. & Abdollabi, M. (2009). A review of Ihe herbal phosphodiesterase inhibitors: hiture perspective of new drugs. Cytokine. Vol, 49, No. 2, (November 2CX)9), pp. 123-129. ISSN: 1043-4666.
7. Rauiam. D \'., Ankola. D. D.. Bhardwaj. V., Sahana. D. K. and Ravi Kumar. M. N. V. (2006). Role of antioxidants in propln la\is and therapy: A pharmaceutical prospective. Journal of Conti-cjlled Releci.se 113:189- 207.
8. RekM Sundu. \Vithawai \ling\anish. Enos Tangke Arung. Harlinda Kuspradini, Kriangsak Khownium (2015).
\niio.xidani and aniim'icrobial activities of crude meihanolic extract of polyscias guilfoylei leaves. Pure and Applied Chemisin.' Intemauonal Conference 2015 (PACCON 2015).
9. Rungthip Kawaree and Sombai Chowwanapoonpoh (2()()y] Stability of Chemical Componenis and Antioxidanl Activii> of \'olalile OiK from Some McJi.indl Plants in Thjilan. Chiang Mai University Joumal „; \a„iral Sciences Vol.Slh
350 So 8-Thang 4/2020
SgaynhSn Mi: 25/3/2020
ferity phan bi^n danh gia va sii'a chffa: 6/4/2020 NgJydiS'pnhandSngbai: 15/4/2020
Thdttg lilt llii: gid:
TS.NGUYlNDt?CDQ CN. THI LINH DUYEN CN. HUYNH THANH DUY PGS.TS. TRAN NHAN DUNG
Vifn Nghi6n Clhl va Phat triS'n Cdng nghe Sinh hpc, TrUtog DaihQcCanThtf
IDA IOC-CDNG NGHE M C PIAM
A STUDY ON POLYPHENOL, SAPONIN CONTENT AND ANTIOXm ANT ACTIVITIES OF POLYSCIAS FRUTICOSA (L.) HARMS EXTRACT
• Ph.D NGUYEN DUC DO
Biotechnology Research a n d Development Institute, Can Tho University
• THI UNH DUYEN
Biotechnology Research a n d Development Institute, Can Tho University
• HUYNH THANH DUY
Biotechnology Research and Development Institute, Can Tho University
• Assoc.Prof.Ph.D TRAN NHAN DUNG Biotechnology Research and Development Institute,
Can Tho University
ABSTRACT:
Polyscias fmlicosa (L.) Harms extracts were investigated on total polyphenol, .saponin coiilem and antioxidant activity by using DPPH and peroxide reduction methods. The result showed that leaf-methanol extracts had the highest polyphenol content with 72.003 mg GAE/g whereas highest saponin content belonged to leaf-ethanol extract with 118.44 mg/g.
Ual-ethanol extract showed the best efficiency widi ICjo value on of DPPH and peroxide reduction methods were 266.3 |jg/mL and 226.13 jig/mL, respectively.
Keywords: DPPH, hydrogen peroxide scavenging capacity, Polysciasfruticosa (L.) Harms, total polyphenol, total saponin.
So 8 - Thdng 4/2020 351
