nghien cuu ly trich curcumin tu cu nghe vang

8/7/2019 Nghien Cuu Ly Trich Curcumin Tu Cu Nghe Vang

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MC LC

TrangTrang ph ba ............................................................................................................i

Li cam oan .......................................................................................................... ii

Li cm n............................................................................................................. iii

Mc lc....................................................................................................................1

PHN MU......................................................................................................3

CHNG 1: C S L THUYT

1.1. Tm hiu v ngh ...........................................................................................5

1.1.1. M t thc vt ........................................................................................5

1.1.2. Thnh phn ha hc ...............................................................................6

1.1.3. Dc tnh...............................................................................................7

1.2. Tm hiu v curcumin....................................................................................9

1.3. Phng php tch bit v tinh ch hp cht hu c......................................111.3.1. Phng php chit................................................................................11

1.3.2. Phng php kt tinh li.......................................................................14

1.4. Tm hiu v HPLC (sc k lng hiu nng cao)............................................17

1.4.1. Gii thiu phng php HPLC.............................................................17

1.4.2. Tm hiu v h thng HPLC.................................................................17

CHNG 2: THC NGHIM

2.1. Chun b dng c, ha cht, nguyn liu.......................................................22

2.1.1. Dng c................................................................................................22

2.1.2. Ho cht...............................................................................................22

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2.1.3. Nguyn liu..........................................................................................22

2.2. Thc nghim.................................................................................................23

2.2.1. Chit curcumin .....................................................................................23

2.2.2. Kt tinh curcumin.................................................................................24

CHNG 3: KT QU V THO LUN

3.1. Kt tinh bng cn 900....................................................................................25

3.2. Kt tinh bng ietylete..................................................................................25

PHN KT LUN KIN NGH ........................................................................27

TI LIU THAM KHO......................................................................................28PH LC



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PHN MU

1. L do chn ti:

Nhiu cng trnh nghin cu th nghim cc nc pht trin trn th gii

khng nh t lu rng curcumin c tc dng hy dit t bo ung th vo loi mnh. TiM, i Loan, ngi ta tin hnh th lm sng dng curcumin iu tr ung th v

kt lun: curcumin c th kim hm s pht tc ca t bo ung th da, d dy, rut, vm

hng, curcumin cn l cht b cho d dy, rut, gan, mt, lc mu, lm sch mu,

iu tr vt thng, chng vim khp, dng, nm, chng vi khun c hiu lc.

Cch y 5000 nm c nghc bit n nh mt loi gia v, thuc gia truyn

cha c rt nhiu bnh, cha lin so, Tc dng kim hm s pht trin ca t bo

ung th v iu tr nhiu bnh ca ngh l do hot cht trong c ngh gi l curcumin.

Curcumin l thnh phn c bit v l hot cht chnh to nn mu vng c trng cho

c ngh. Trong lng curcumin ch chim khong 0,3 - 1% v khi lng c ngh.

Ch c curcumin t nhin trong c ngh mi c kh nng phng v chng li s pht

trin ca t bo ung th v cc bnh khc cao.

Vit Nam l mt nc nng nghip nhit i nng m vng ng nam Chu

, chnh iu kin trn rt ph hp vi loi cy ly c nh: gng, ngh, ti, hnh,

Trong cy nghc cy trng rng ri mt s nc ti Chu nhn ,

Trung Quc v Vit Nam cng khng ngoi l. nc ta ni chung v tnh ng Thp

ni ring th cy nghc trng rt ph bin vi nhiu chng loi a dng v phong

ph. Vi ngun nguyn liu di do v phong ph ni trn cng vi mc ch l vn

dng nhng kin thc hc cng thm nhng thao tc th nghim c hc trong

cc mn thc nghim ha hc. Do ti tin hnh ly trch curcumin t c ngh vng

vi tn ti kha lun tt nghip l: Nghin cu ly trch curcumin tc ngh vng.

2. Mc tiu nghin cu.

Nghin cu ly trch curcumin t c ngh vng ti thnh ph Cao Lnh tnh ng

Thp

3. Nhim v nghin cu.

Tm hiu thnh phn, ng dng ca c ngh v hot cht curcumin.



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Nghin cu ly trch curcumin.

Tm hiu v HPLC.

nh danh v nh lng curcumin bng HPLC.

4. Phng php nghin cu.Nghin cu cc ti liu c lin quan n ti.

T tm ti, t nghin cu.

Nghin cu thc nghim.

5. i tng nghin cu.

C ngh thu c t cy ngh vng trng ti a bn thnh ph Cao Lnh, tnh

ng Thp.

6. Lch snghin cu.Hin nay c rt nhiu ti nghin cu v curcumin cng nh tc dng ca

curcumin n n vic phng chng bnh ung th v cc bnh v tim mch khc,

Tuy nhin cha c ti no nghin cu ly trch curcumin t c ngh vng ti a bn

thnh ph Cao Lnh tnh ng Thp.

nc ngoi cng c mt s ti tng hp thnh cng curcumin t vanilin

nhng curcumin t nhin trong c ngh th c tc dng cao i vi sc khe con ngi.



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PHN NI DUNG

CHNG 1: CSL THUYT

1.1. Tm hiu v ngh.[1], [2], [4], [6]- Ngh cn c tn l ut kim, khng hong, safran des Indes.

- Tn khoa hc Curcuma longa L. (Curcuma domestica Lour).

- Thuc h Gng Zingiberaceae.

- Ta dng thn r gi l khng hong (Rhizoma Curcumae longae) v r c gi l ut

kim (Radix Curcumae longae).

1.1.1. M t thc vt.

Ngh l mt loi c cao 0.60m n 1m. Thn r thnh c hnh tr hi dt, khi b

hoc ct ngang c mu vng cam sm. L hnh tri xoan thon nhn hai u, hai mt

u nhn di ti 45cm, rng ti 18cm. Cun l c b, cm hoa mc t gia cc l ln,

thnh hnh nn tha, l bc hu th khum hnh mng rng, u trn mu xanh lc nht,

l bc bt th hp hn, mu hi tm nht. Trng c phin, cnh hoa ngoi mu xanh lc

vng nht, chia thnh ba thy, thy trn to hn, phin cnh hoa trong cng chia thnh

ba thy, 2 thy hai bn ng v phng, thy di hm thnh mng su. Qu nang 3

ngn, mbng 3 van. Ht c o ht.

c trng khp ni trong nc ta lm gia v v lm thuc ngoi ra cn mc v

c trng cc nc n , Innxia, Campuchia, Lo, Trung Quc, v cc nc

nhit i.

Ngh thng c thu hoch vo ma thu.



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Hnh 1.1: Thn, hoa, c ngh.

1.1.2. Thnh phn ha hc.

Trong ngh, ngi ta phn tch c:

- Cht mu curcumin 0,3%- 1%, tinh th nu nh tm, khng tan trong nc,

tan trong ru, ete, clorofoc, dung dch c hunh quang mu xanh lc. Cng thc

curcumin c xc nh nh sau:

+ Cc cht mu vng gi chung l curcumin. Vo u th k XIX ngi ta

chit c curcumin tinh th khng tan trong nc, tan trong cn, ete, du bo. Nhng

nm 1953 Srinivasan K. R chng minh bng sc k ct silic rng l hn hp:



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+ Curcumin chnh thc (cn gi l curcumin I) chim 60% y l mt

ixeton i xng khng no c th coi nh l diferuloyl-metan (axit ferulic l axit

hydroxy-4-metoxy-3-xinamic).

+ Curcumin II hay demetoxy-curcumin chim 24% v curcumin III hay bis-demetoxy-curcumin chim 14% trong 1 hay 2 hydroxyxinamic thay cho axit ferulic.

Nu dng sc k trn giy s thy cc cht curcumin khc na nhng vi lng rt nh.

+ Nm 1977, Nguyn Khang (i hc Dc H Ni) chit t bt c ngh

sau khi ct ly ht tinh du bng bezen, sau thu hi dung mi trong p lc gim

v kt tinh bng cn etylic cho ti khi c chy khng thay i v mt vt trn sc k

lp mng thu c 0,76 1,1% curcumin I tinh khit, chy 182 -183oC. Nu

chn c ngh thu vo thng 1, thng 2 c tht ti 1,5% curcumin.- Tinh du 1 5% c mu vng nht, thm. Trong tinh du gm c 25% cacbua

tecpenic, ch yu l zingiberen v 65% xeton sespuitecpenic, cc cht tumeron,

curcumen C15H24 mt cacbon khng no.

Tumeron

- Ngoi ra cn tinh bt, canxi oxalat, cht bo. C ngh cha 8 10% nc, 6

8% cht v c, 40 50% tinh bt nha.

1.1.3. Dc tnh.

C nghc bit n vi nhiu cng dng khc nhau, c bit c n v

nhiu nc, c phng ng ln phng Ty, s dng nh mt loi dc liu tr bchbnh.

Theo hi ng nghin cu Trung ng, c ngh c th cha c nhiu bnh. C

ngh c th cha bnh hen suyn, ho, tr cm, nghin ru, mn v cc bnh ngoi da,

c ngh c th gim vim nhim, tr to gan v nhim trng bng quang, ri lon kinh



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nguyt, tng cng sc khe cho tim. C ngh ngm vi nc v mt ong gip li tiu,

hoc nghin ngh vi b qua lc sch c th cha hiu qu bnh tiu ng.

Ngoi ra c ngh cn gip trau rng v nga su rng, gim au bao t, gip tiu

ha v to cm gic thm n. Nh nc ngh un si vo mi gip cha au u vchng mt ng.

C ngh khng ch c cng dng gip lin so nh nhiu ngi bit, m cn

mang li rt nhiu tc dng hu ch, c bit i vi sc khe con ngi.

Gip gim cn, lu thng v lc mu; gip c th chng li cc vi khun sng k

sinh trong rut, c bit tt cho h tiu ha; mi y ngi ta chng minh c rng

c th s dng ngh chng ung th v ngh c kh nng khng vim, gim nguy c

nhim trng; c th dng ngh kh trng v lm mau lnh vt thng.Khi gp rc ri v tiu ha, nghin cu cho thy, ngh c th kch thch tiu ha v

gii phng cc enzim tiu ha, ph v lin kt cacbonhydrat v cc cht bo. Chnh v

th, trong trng hp bau bng, mt cc tr ngh s gip ch rt nhiu.

Cht curcumin c t nhin trong c ngh tng c cc nh khoa hc chng minh l

mt cht chng oxy ha cc mnh c li cho sc khe, c li cho tim mch, chng

cholesterol v ung th.

Ngh c th lm gim hm lng cholesterol c hi trong mu v c kh nngchng li chng x va ng mch, curcumin c trong ngh cho php gim nguy c

mc bnh nhi mu ctim, c bit l nhng ngi hay ngy ng. Nu cht curcumin

c ng dng thnh cng i vi con ngi, n s mra mt hng i mi cho cch

phng v iu tr bnh nhi mu ctim ng thi bo v sc khe tim mch ca chng

ta. Khc vi hu ht cc hp cht t nhin khc vi hiu qu hn ch, cht curcumin c

tc dng trc tip ln nhn t bo bng cch nga nga vic sn sinh qu nhiu protein

bt thng. Tuy nhin, cc chuyn gia cng khuyn nn n nghmc va phi v

c n qu nhiu cng khng gip tng hiu qu ca tr bnh ca ngh.

Curcumin c tc dng khng ung th, c lp v tiu hy t bo ung th. Curcumin

l mt thnh phn c bit lm nn mu vng c trng ca ngh c kh nng tiu dit

hai loi protein trong cc t bo ung th, cc protein ny chnh l ngun duy tr s tn



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ti ca chng. Ngh cng gip ngn chn khng cho t bo ung th lan trn i khp ni

trong cth (chng di cn). Ngoi ra ngh cng rt an ton v khng c phn ng ph.

Cc bc s bnh vin University of Texas MD Anderson Cancer Center, mt bnh

vin chuyn v cha tr cc bnh ung th vo bc ln nht th gii, sau khi b ranhiu nm nghin cu v th nghim, u ng thanh xc nhn rng: thuc bo ch

bng c ngh c tc dng trc tip git cht cc t bo ung th, ng thi c ngh cng

l mt loi chng oxi ha cc mnh rt cng hiu chng li s ph hoi ca gc t

do v gip tng cng h thng min nhim cho nn h khuyn bnh nhn ung th

nn dng ngh hng ngy.

Tuy nhin, khng nn xem y l thn dc, v n ch c tc dng khi ung u n

v va phi trong mt thi gian di.1.2. Tm hiu v curcumin. [8], [9], [10], [11]

- Tn IUPAC: (1E, 6E) -1,7-bis (4-hydroxy-3-metoxyphenyl) -1,6-heptadien-

3,5-dion.

- Cng thc phn t: C21H20O6

- Phn t khi: 368,38 g / mol.

- Nhit nng chy: 183C (361 K).

- Curcumin l tinh th mu nu , l hot cht c chit ra t ca ngh vngthuc h gng. Hin ti ngi ta tm thy curcumin tn ti 4 dng hp cht:

+ Curcumin l hp cht chnh chim 60%:

Curcumin

+ Demetoxy-curcumin chim 24% c cng thc cu to sau:



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Demetoxy curcumin

+ Bis-demetoxy-curcumin chim 14%:

Bis-demetoxy curcumin+ V mt hp cht mi pht hin l xiclocurcumin chim khong 1%:

Xiclocurcumin

- Curcumin l mt polyphenol v l sc t to nn mu vng c trng ca cngh.

- Curcumin c th phn ng c vi axit boric to nn hp cht c mu cam

nn c ng dng dng nhn bit mui ca nguyn t bo.

- Chnh v curcumin l sc t to nn mu vng sng nn curcumin c dng

lm cht ph gia thc phm. Trong cht ph gia thc phm curcumin c k hiu di

m s E100.

- Dc tnh:

+ Curcumin l cht hy dit ung th vo loi mnh nht theo cch hy

dit tng bc cc t bo c tnh. Chng lm v hiu ha t bo ung th v ngn chn

khng cho hnh thnh cc t bo ung th mi. Trong khi , cc t bo lnh tnh khng

bnh hng. Curcumin c coi l cht tiu biu nht cho th h mi cc cht chng



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ung th va rt hiu lc, va an ton, khng gy tc dng ph. Curcumin c kh nng

loi b cc loi men gy ung th nh COX-1, COX-2 c trong thc n, nc ung, v

hiu ha cc gc t do hnh thnh trong qu trnh t v ca c th, do bc xc hi

cng nh do cc loi sc thn kinh, th lc, cc c t ha hc (dioxin, furan).+ Curcumin c kh nng mnh m gii c v bo v gan, bo v v lm

tng hng cu, loi b cholesterol xu, iu ha huyt p, h mmu, ngn chn bo

ph, xa b tn nhang, i mi, trng c chng rng tc gip mau chng mc tc, lm

cho da d hng ho, tng cng sc p, sc lc v c tui th

+ Curcumin l mt trong nhng cht chng vim, chng xi ha in

hnh. N khng chiu trc lc cho cc bnh ung th, lot d dy, hnh t trng, i

trng, yu gan mt, vim gan B, C, sgan c chngm cn iu tr va nh nhngva hiu qu cao cc bnh ri lon h min dch nh vim ton thn, vim a khp,

vim li cu khp, bnh a scng, bnh cng b, long xng, vim c, vy nn, ban

h thng, au h tiu ha, ri lon tuyn gip, u mu, suy gim tr nh,h triu

tr bnh Parkison, nhn no.

+ Curcumin c kh nng khng nm, khng khun nh virt HP, vim

gan B, C rt cao.

+ Curcumin nhiu nc trn th gii c coi nh va l thuc va lthc phm iu tr gn 20 loi ung th khc nhau. Ring i vi ung th mu cc nh

khoa hc cho bit curcumin c tc dng tng hng cu, chng suy kit sc lc

1.3. Phng php tch bit v tinh ch hp cht hu c. [3]

1.3.1. Phng php chit.

1.3.1.1. nh ngha.

Phng php chit dng tch cht ra khi hn hp cht rn hoc dung dch

hoc huyn ph bng dung mi thch hp nhit phng hoc nhit si ca dung

mi. Csvt l ca phng php l da vo nh lut phn b Nernst:

Khi thm mt cu t th ba vo h dung dch c hai cu t khng tan hon ton

vo nhau hoc tan c gii hn th s ho tan ca cu t ny vo hai cu t theo mt t l

nht nh nhit khng i, gi l hng s phn b Nernst K:



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2

1

2

1

C

C

S

SK ==

C1, C2 l nng ca cc cu t trong dung mi.

S1, S2 l tan ca hai cu t.

K cng ln khi2

1

S

Scng ln th vic ly cht rn ra rt kh khn, phi dng dung

mi chit nhiu ln.

Cng mt lng dung mi chit, cn phi chia nhiu ln chit. C th tnh

c lng cht cn li sau ln chit th n da vo hng s Nernst:

n

nSkV

kVGG

+= 0

Gn l lng cht cn li sau n ln chit.

G0 l lng cht ban u c trong th tch V.

S l s ml th tch dung mi cho vo.

Nh vy, mun Gn cng nh th n phi ln v S phi nh, ngha l cht nh

chit ra cn li trong nh chit ra cn li trong dung dch cng nh th cng mt lng

dung mi cn phi chit nhiu ln.

1.3.1.2. La chn dung mi khi chit.Dung mi chit phi m bo cc yu cu sau:

- Dung mi chit phi ha tan cht nh chit ln hn dung mi c.

- Khng trn ln vi dung mi c, ngha l c t khi khc nhiu vi dung mi

c.

- Dung mi ny d tch ra khi tinh ch li v t c kh nng to nh tng v t

c.

Khi la chn dung mi chit phi ch n tan ca cht vo dung mi. tan ca cht ph thuc vo bn cht ca cht tan, bn cht ca dung mi, nhit v b

mt tip xc gia cht tan v dung mi. V vy, khi chit ngi ta thng lc k. Tuy

nhin, nu c hin tng to huyn ph, nh tng khi lc th phi ph s to huyn

ph.



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1.3.1.3. Kthut chit cht lng.

Dng c chit l phu chit.

Trc khi chit phi kim tra li kha v bi vazlin vo kha phu. dung

dch vo phu chit, thm dn dung mi vo sao cho th tch ch chim khong 2/3 thtch ca phu. Lng dung mi cho vo khong 1/5 n 1/3 th tch dung dch. y nt,

mt tay gi nt v phu, mt tay gi kha phu, cn thn lc nh v dc ln dc xung

phu nhiu ln. Khi lc thng lm tng p sut trong phu, do phi ngc phu,

mkha phu cho cn bng vi p sut bn ngoi ri ng kha phu, lc mnh tip

khong 1 2pht. Lc xong, cp phu vo gi yn mt lc cho phn lp hai cht

lng. Sau mkha phu v tch ly cc phn khc nhau tu thuc vo t khi ca

dung dch. Nu lp di l dung dch cn ly th li mt t trong phu, nu ly lptrn th cho chy qu mt t cht lng.

Khi chit nhng cht d to thnh nh tng phi ch lc nh. Nu nh tng

to thnh do mt lng kt ta to thnh trn b mt phn chia hai pha lng th phi lc,

nu do sc cng b mt th thng thm ru etylic ph sc cng b mt phn chia

gia hai pha.

Nu do s khc nhau v t khi ca hai cht lng khng ln lm th thng thm

dung dch NaCl bo ha tng t khi ca dung dch nc.Tt nht l yn lng trong mt thi gian lu.

Khi chit nhng cht tan trong nc nhiu hn tan trong dung mi hu cc th

dng chit lin tc trn dng c chit cht vi t khi khc nhau so vi nc.

Nu chit vi cht rt t, c th dng ng nghim hi nhn u ri thm dung

mi vo bng mt pipet nh. yn cho tch lp, dng pipet sch tch ly lp dung

dch ha tan cht nh chit.

1. 3.1.4. Chit cc cht rn.

Phng php n gin l ngm chit cht rn trong dung mi hoc ha tan trong

dung mi nhit thng hay nhit si ca dung mi, sau lc hoc gn ly

dung dch. Mun ly cht t dung dch th ct ui dung mi my c quay chn

khng hoc bng cc phng php thng thng. S chit cht trong hn hp rn ph



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thuc nhiu vo ha tan ca cc cht vo dung mi la chn v b mt tip xc ca

cht rn vi dung mi v nhit . tng kh nng chit, ngi ta thng phi nghin

nh cht rn ri ngm chit nhit thng hoc nhit si ca dung mi.

Trong phng th nghim thng chit bng b chit Soklet chit lin tc. Chtrn nh chit nghin nhc gi trong giy lc (ti vi) t vo phn hnh tr trn

bnh cu ca my chit. Cho dung mi vo bnh cu (ty thuc vo lng cht chit m

cho lng dung mi khong 1/2 th tch bnh cu), lp ng sinh hn hi lu trn ri

un cho si dung mi. Hi dung mi bay ln v ha tan cht rn trong bc giy lc ri

xung bnh cu trong ng dn hi ngng t. C nh vy nng ca cht tan trong

dung mi tng dn theo thi gian un hi lu. Nu cht tinh ch ha tan vo dung mi

thu c dung dch cht tinh ch trong bnh cu, c quay ui dung mi ri kt ta licht rn. Nu trong hn hp cht rn, cc cht bn ha tan vo dung mi th phn cht

rn sc l cht sch. Nu cht tan l cht ph v cht tinh ch s cn li trong bnh

chit ch vic ly cht rn trong giy lc ra, lm kh s thu c cht sch.

1.3.2. Phng php kt tinh li.

1.3.2.1. nh ngha.

y l phng php quan trng nht tinh ch cc cht rn. Csl thuyt ca

phng php l da vo s khc nhau v tan ca cc cht trong mt dung mi hayh dung mi cc nhit khc nhau, cng nh s khc nhau v tan vo dung mi

ca cht tinh ch v cht bn cng mt nhit . Qu trnh chung l ha tan cht rn

thnh dung dch bo ha nhit si ca dung mi v khi lnh th cht rn kt tinh

li di dng tinh khit.

1.3.2.2. Chn dung mi.

Ngi ta phi chn dung mi hay h dung mi thch hp ha tan cht tinh ch

nhit si v khng ha tan hoc ha tan t nhit lnh, cn tp cht th hin

tng ngc li. Sau khi lc nng loi b tp cht th cht rn s kt tinh li sch hn.

Vic la chn dung mi kt tinh rt quan trng. Dung mi kt tinh phi khng

tng tc ha hc vi cht kt tinh nhit thng cng nhnhit si. Dung

mi phi c nhit si thp hn nhit nng chy ca cht tinh ch t nht l 100C



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v li phi gii phng khi lc cng nh khi ra. Vic la chn dung mi hay h dung

mi phi da vo mi quan h cu to phn t ca cht kt tinh v dung mi, thng

thng cht phn cc th ha tan tan vo dung mi phn cc v ngc li. Khi cht kt

tinh cha r cu to th phi th ha tan trong dung mi t khng phn cc n dungmi phn cc. Cch lm nh sau: ly mt vi tinh th vo ng nghim, nh mt t dung

mi vo ri un si, quan st tan cho n khi tinh th tan ht, nu m bo yu cu

trn th cht s kt tinh li khi lnh. Dung mi c xem l tt nu c 0,1g cht kt tinh

tan trong 1ml dung mi nng.

Khi khng chn c dung mi th bt buc phi chn h dung mi. Nguyn tc:

Ly mt dung mi ha tan cht kt tinh ngay nhit thng, sau chn mt dung

mi khng ha tan hay km ha tan cht tinh ch nhng phi tan trong dung mi trn.Cch lm nh sau: Ly mt vi tinh th vo ng nghim, thm dung mi cho n khi

tan ht, ghi ly th tch dung mi. Nh t t dung mi khng tan cht tinh ch vo hn

hp trn cho n khi vn c, ghi ly th tch dung mi, un hn hp cho tan ht, lc

nng ngui, cht rn s kt tinh li. T l hai th tch o c coi l t l th tch

dung mi chn. Thng chn h dung mi etanol nc, etanol benzen, axeton

ete du ha, axit axetic nc,

Nu c nhiu cht c tan khc nhau nhit khc nhau vo mt dung dch,ngi ta dng phng php kt tinh phn on. mt nhit xc nh, mt cht no

tan qu bo ha s kt tinh li khi lnh, cn cht kia cha bo ha sli trong

dung dch.

Nu dung mi ha tan cht kt tinh, cn cht bn khng tan, ngi ta s lc nng

loi b cht bn, cn cht kt tinh li trong dung dch s kt tinh khi lnh.

1.3.2.3. Cc thao tc khi kt tinh.

- Chun b dung dch kt tinh hay dung dch nc ci.

Cho mt lng cht cn kt tinh vo bnh cu hai c c lp ng sinh hn hi lu

v phu chit ng dung mi hoc vo bnh tam gic c sinh hn hi lu, thm bt

ri cho dung mi vo t hn mt lng t theo lng tnh ton, un si. Nu si m



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16

cha tan ht th thm mt t dung mi vo cho n khi cht tan hon ton, cht bn s

khng tan.

Nu dng hn hp dung mi th cho dung mi tan tt vo trc cho n khi cht

rn tan hon ton ri thm dn dung mi ha tan km vo cho n khi cht rn kt tari tan nhit si. Nu dung dch c mu, phi thm cht ty mu nh than hot

tnh, than xng hay silicagel vi t l 1/20 hay 1/50. Khi cho cht kh mu phi

dung dch lnh, khng c thm cht kh mu khi dung dch ang nng trnh dung

dch b tro ra ngoi. Sau , un si li dung dch trong 2 3 pht.

- Lc nng dung dch.

Lc dung dch ang nng loi b cht bn, cht ph khng tan ra khi dung

dch. Cn lc nng bng phu lc nng, lc nhanh trnh cht kt tinh trn phu.- Thc hin kt tinh cht.

Dung dch nc ci c y ming khng nt cht, ngui hay lm lnh bng

nc lnh hay nc th tinh th cht s kt tinh.

Ch nu dung dch cha kt tinh, cha t n bo ha th phi c ui bt

dung mi ri mi lm lnh. Kch thc ca tinh th ph thuc vo tc kt tinh v

lm lnh. Nu lm lnh nhanh th thu c tinh th nh s hp th dung mi v cht

bn. Tinh th cng ln khi lm lnh kt tinh chm.Nu khng thy kt tinh th c th gy mm kt tinh bng cch cho vo dung

dch mt vi tinh th ca cht tinh ch hoc ly a thy tinh c vo thnh bnh.

Nhng cht c nhit nng chy thp thng tch ra dng du nn phi lm

lnh chm v su tinh th kt tinh chm.

- Tch lc tinh th.

Tinh th tinh khit c lc nhanh trn phu Bucsne p sut thp. Khi lc cn

ra li khi kt ta bng dung mi lnh. Mt s trng hp ngi ta s gn, lng. Trong

trng hp cht ho nc v d b oxi ha th dng phng php ring.

- Lm kh tinh th.



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C th lm kh trong khng kh hay trong t sy thng hoc t sy chn khng.

Ch lm kh trong t sy cn nhit thp hn nhit nng chy ca cht

khong 200C.

Cng c th lm kh tinh th bng gi ca my sy tc.- Xc nh nhit nng chy ca cht thu c.

Khi xc inh nhit nng chy thy cnh hoc sai khc t hn hoc bng

10C th coi nh cht sch.

1.4. Tm hiu v HPLC (sc k lng hiu nng cao).[5], [7], [8]

1.4.1. Slc vHPLC.

HPLC l ch vit tt ca 4 ch ci u bng ting Anh ca phng php sc k

lng hiu nng cao (High Performance Liquid Chromatography), trc kia gi lphng php sc k lng cao p (High Pressure Liquid Chromatography).

Phng php ny ra i nm 1967 1968 trn c spht trin v ci tin t

phng php sc k ct cin. Hin nay phng php HPLC ngy cng pht trin v

hin i ha cao nhs pht trin nhanh chng ca ngnh ch to my phn tch. Hin

nay n p dng rt ln trong nhiu ngnh kim nghim c bit l ng dng cho cc

ngnh kim nghim thuc v n hin l cng cc lc trong phn tch cc thuc a

thnh phn cho php nh tnh v nh lng.Khi nim: Sc k lng hiu nng cao l mt phng php chia tch trong

pha ng l cht lng v pha tnh cha trong ct l cht rn c phn chia di

dng tiu phn hoc mt cht lng ph ln mt cht mang rn, hay mt cht mang

c bin i bng lin kt ha hc vi cc nhm chc hu c. Qu trnh sc k lng

da trn cch hp ph, phn b, trao i ion hay phn loi theo kch c(ry phn t).

1.4.2. Tm hiu vhthng HPLC:

My HPLC gm cc b phn cbn tm tt trn s sau:



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Hnh 1.2: S thit b HPLC.

Trong :

1: Bnh cha dung mi pha ng.

2: B phn kh kh.

3: Bm cao p.4: B phn tim mu (bng tay hay Autosample)

5: Ct sc k (pha tnh).

6: Detector (nhn tn hiu).

7: H thng my tnh gn phn mm nhn tn hiu, x l d liu v iu khin h

thng HPLC.

8: In d liu.

1. Bnh ng dung mi.

- Hin ti my HPLC thng c 4 ng dung mi vo u bm cao p cho

php chng ta s dng 4 bnh cha dung mi cng mt ln ra gii theo t l mong

mun v tng t l dung mi ca 4 ng l 100%.

Degasse PumpColumn

Timmu

Detector

12

3

4

5

6

78



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19

- Tuy nhin theo kinh nghim th chng ta t khi s dng 4 ng dung mi cng

mt lc m chng ta ch s dng ti a l 3 v 2 ng cho h pha ng lun c

pha trn ng nht hn, h pha ng n gin hn qu trnh ra gii n nh.

- Hin 4 ng dung mi phc v ch yu cho vic ra gii Gradial dung mitheo thi gian v cng tc xy dng tiu chun.

Lu : + Tt c dung mi dng cho HPLC u phi l dung mi tinh khit v c

ghi r trn nhn l dng cho HPLC hay dung mi tinh khit phn tch.

+ Tt c cc ha cht dng pha mu v pha hm phi c s dng

l ha cht tinh khit phn tch.

S dng ha cht tinh khit nhm mc ch trnh hng ct sc k hay nhiu

ng nn, to nn cc peak tp trong qu trnh phn tch.2. B kh kh Degasse.

Mc ch ca b kh kh nhm loi tr cc bt nh cn st li trong dung mi

pha ng.

Nu nh trong qu trnh phn tch m dung mi pha ng cn st cc bt kh th

mt s hin tng sau y s xy ra:

+ T l pha ng ca cc ng dung mi khng ng s l cho thi gian

lu ca Peak thay i.+ Trong trng hp bt qu nhiu b kh kh khng th loi tr ht c

th c th bm cao p s khng ht c dung mi khi p sut khng ln v my sc

k s ngng hot ng.

Trong bt c trng hp no nu trn cng cho kt qu phn tch sai.

3. Bm cao p.

Mc ch bm pha ng vo ct thc hin qu trnh chia tch sc k. Pump

phi to c p sut cao khong 250 at 500 at (1at = 0,98 bar) v pump phi to

dng lin tc. Lu lng bm t 0,1 9,999 ml/pht.

My sc k lng hin nay thng c p sut ti a 412bar (khong 420at). Tc

dng 0,1 9,999 ml/pht.



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20

Tc bm l hng nh theo thng s c ci t. Hin ti bm c 2

Pisstone thay phin nhau y dung mi lin tc.

4. B phn tim mu.

a mu vo ct phn tch theo phng php khng ngng dng chy vidung tch 5 -100l.

C 2 cch ly mu vo ct: bng tim mu th cng v tim mu t ng

(Autosample).

5. Ct sc k.

Ct cha pha tnh c coi l tri tim ca h thng sc k lng hiu nng cao.

Ct pha tnh thng thng lm bng thp khng r, chiu di ct khong 10

30cm, ng knh trong 1 10mm, ht cht nhi ct c = 5 10m, ngoi ra cn mts trng hp c bit v kch thc v kch cht,

Vi cht nhi ct c = 1,8 5m c th dng ct ngn (3 10cm) v nh

(ng knh trong t 1 4,6mm) loi ct ny c hiu nng tch cao.

Cht nhi ct ty theo loi ct v kiu sc k.

Thng thng cht nhi ct l Silicagel (pha thun) hoc l Silicagel c

Silan ha hoc c bao mt lp mng hu c (pha o), ngoi ra ngi ta cn dng

cc loi ht khc nhau nh: Nhm oxit, Polyme xp, cht trao i ion.

* i vi mt s phng php phn tch i hi phi c nhit cao hoc thp hn

nhit phng th ct c t trong b phn iu nhit.

6. Detector.

L b phn pht hin cc cht khi chng ra khi ct v cho cc tn hiu ghi trn

sc c thnh tnh v nh lng. Ty theo tnh cht ca cc cht cn phn tch

m ngi ta s dng loi Detector thch hp v phi tha mn iu kin trong mt cng

nng nht nh ca cht phn tch.

A = k.C

Trong : A l tn hiu o c.

C l nng cht phn tch.



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K l hng s thc nghim ca Detector chn. Tn hiu ny c th l:

hp th quang; cng pht x, cng in th, dn in; dn nhit, chit

sut

Trn cs ngi ta ch to cc loi Detector sau:- Detector quang ph t ngoi 200 380 nm pht hin UV.

- Detector quang ph t ngoi kh kin (UV VIS): 190 900nm pht hin

cc cht hp th quang. y l loi thng dng nht.

- Detector hunh quang pht hin cc cht hu ccha hunh quang t nhin

cng nh cc dn xut c hunh quang, l loi Detector c chn lc cao nht.

- Loi hin i hn c Detector Diod Array (Detector tn x bay hi) cc

Detector ny c kh nng qut chng phnh tnh cc cht theo hp th cc ica cc cht.

Ngoi ra cn mt s loi Detector khc l:

+ Detector in ha: o dng, cc ph, dn, in lng

+ etector chit sut vi sai: Detector khc x (thng thng dng cho o cc

cht ng)

+ Detector o dn nhit, hiu ng nhit,

7. B phn ghi tn hiu. ghi tn hiu pht hin pht hin do Detector truyn sang.

- Trong cc my th h c th s dng my ghi n gin c th v sc k , thi

gian lu, din tch ca Peak, chiu cao.

- Cc my th h mi du dng phn mm chy trn my tnh n c th lu tt

c cc thng s, ph v cc thng s ca Peak nh tnh i xng, h s phn gii

trong qu trnh phn tch ng thi x l, tnh ton cc thng s theo yu cu ca ngi

s dng.

8. In kt qu.

Sau khi phn tch xong cc mu ta s in kt qu do phn mm tnh ton ra

giy hon thin h s.



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22

CHNG 2. THC NGHIM

2.1. Chun b nguyn liu, dng c, ha cht.

2.1.1. Dng c.

- T sy- B chit Soklet.

- Dng c phng th nghim thng thng: cc thy tinh, eclen, a thy

tinh, phu, giy lc,

- Cn in t.

- My li tm.

- My o HPLC.

2.1.2. Ha cht.

- Nc ct 1 ln.

- Etylaxetat

- Axeton

- itylete

- Cn 900.

2.1.3. Nguyn liu:

- Chn ngh gi, thn l vng, kh ri, khng cn xanh ti, thu mua trn a

bn thnh ph Cao Lnh. Ly c, chn nhng c tt khe, khng b ng, thi.

- Lt b lp v o bn ngoi, ra sch bng nc, kh, ct lt nh sau em

sy 800C n khi ngh mt nc hon ton. Dng ci, chy gi ngh sy thnh bt

mn.

Bng 1: Khi lng nguyn liu kh ng vi nguyn liu ti.

Khi lng nguyn liu ti (kg) Khi lng nguyn liu kh (kg)

2,7 0,4



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2.2. Thc nghim.

2.2.1. Chit curcumin.

- Cn 100g bt ngh sy kh (c mu vng sng) cho vo ti vi, cho vo

phn hnh tr ca b chit soklet.- Pha hn hp dung mi: pha 50ml axeton vo 250ml etylaxetat (t l 1: 5), dng

a thy tinh khuy nh hn hp dung mi cho vo bnh cu ca b chit. Lp ng sinh

hn hi lu (hnh 2.1) v tin hnh un 800C.

Hnh 2.1: B chit Soklet

- Khi un hi hn hp dung mi bay ln ha tan curcumin c gi trong ti vi

ri xung bnh cu qua ng dn hi ngng t. un trong khong thi gian 10h ta c

dch chit c mu nu .

Bt ngh

Dch chit



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- Ly dch chit ra khi bnh cu v thu hi dung mi bng phng php un

hon lu v thu hi li dung mi.

- Chit tng t nh trn cho 400g ngh kh cho 1 ln kt tinh.

2.2.2. Kt tinh curcumin.2.2.2.1. Kt tinh bng cn 900

- Ha tan cao curcumin sau khi ui dung mi trong 50ml cn 900 v lc k

cho cao curcumin ha tan ht trong cn ri tin hnh to mm phn ng bng cch

dng a thy tinh c vo thnh bnh.

- Sau khi ha tan vo cn cho vo t lnh nhit t 2 70C v tip tc

theo di. Sau 2 tun theo di khng thy hin tng kt tinh.

2.2.2.2. Kt tinh bng ietylete.- Ha tan cao curcumin sau khi ui dung mi trong 30ml ietyl ete v lc k

cho cao curcumin ha tan ht trong ietyl ete ri tin hnh to mm phn ng bng

cch dng a thy tinh c vo thnh bnh.

- Sau khi ha tan vo cao vo ietyl ete ri cho vo t lnh nhit t 2

70C v tip tc theo di.

- Sau khong 2 tun curcumin kt tinh, ly ra khi t lnh v tin hnh ly tm

sn phm kt tinh, gn b phn dung dch gi li phn tinh th. Sau sy kh thuc tinh th curcumin c mu nu .

- em cn lng curcumin va kt tinh c khi lng 2,08g v em nh danh

v nh lng curcumin bng sc k lng hiu nng cao (HPLC) ti Trung tm dch v

Phn tch th nghim, a ch: S 02 Nguyn Vn Th, phng akao, qun 1, thnh

ph H Ch Minh.



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CHNG 3: KT QU - THO LUN

3.1. Kt tinh bng cn 900

.Vi 400g nguyn liu (b ngh kh) em chit v dng hn hp dung mi

axeton : etylaxetat (1 : 5), phng php nh nu mc 2.2.1. Sau thi gian chit l

10h thu c dch chit mu nu .

Sau khi thu hi dung mi ra khi dch chit v tin hnh kt tinh vi cn 900 th

kt qu l kt tinh khng thnh cng.

Nhn xt: Nguyn nhn kt tinh khng thnh cng l do khi chit curcumin v

ui dung mi th cao curcumin cn li mt lng nha v tinh du, chnh nha v tinh

du ny lm cn trqu trnh kt tinh curcumin trong cn 900.

3.2. Kt tinh bng ietyl ete.

Vi phng php chit v kt tinh nh nu mc 2.2.1 v dung hn hp

dung mi chit l axeton : etylaxetat (1:5) v dung dung mi kt tinh l ietyl ete sau

thi gian kt tinh khong 2 tun th thu c 2,08g tinh th curcumin mu nu ,

khng tan trong nc v kt qu phn tich bng my HPLC (Shimazu 10AVP, No:

1601, kt ni vi mt my tnh IBM PC, u d UV (model 481).

Sau khi phn tch mu bng HPLC thu c cc kt qu sau (vi sc kc

nu phn ph lc).

Bng 2: Kt qu kim nghim curcumin bng phng php HPLC.

Ch tiu kim nghim n v tnh Kt qu Phng php

Curcumin % 69,5HPLC HPLC Liquid

Chromatography



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Bng 3: So snh sc k HPLC ca curcumin chun v sn phm kt tinh.

Sn phm kt tinh Curcumin chun

Thi gian lu (pht) 4,811 4,818

T bng so snh ta thy cng mt iu kin phn tch th thi gian lu ca sn

phm kt tinh sai lch khng nhiu so vi thi gian lu ca curcumin chun. Do , kt

lun l trong sn phm kt tinh c curcumin vi hm lng 69,5% (bng 2).

Nhn xt: - Vi dung mi ietylete th kt qu kt tinh c curcumin tt

hn. Nguyn nhn l do ietylete c th tch nha v tinh du tt hn cn 900 v chnh

v iu lm cho vic kt tinh curcumin bng dung mi ietylete tt hn khi kt tinhcurcumin bng dung mi cn 900.

- Tinh th thu c khi kt tinh th lng curcumin ch chim

69,5% cn li l mt s tp cht nh: tinh du, nha, cht bo

- Sdcn mt s tp cht nh nu l do trong c ngh lng

nha chim 40 50%, cn lng tinh du chim khong 3 5%. Chnh lng nha v

tinh du tng i nhiu hn rt nhiu so vi lng curcumin trong c ngh nh th

nn trong qu trnh loi b dung mi ra khi dch chit khng trnh khi lng nha

cn li trong cao. Do , khi kt tinh th tinh th cn li mt lng nha v tinh du

tng i nhiu chim khong 30% trong tinh th.



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PHN KT LUN KIN NGH

Bng phng php chng ct lin tc chng ti ly trch c curcumin t

ngh vng trng trn a bn thnh ph Cao Lnh. y l phng php chng ct n

gin c th ly trch c curcumin t c ngh vng.Curcumin nhit thng l tinh th mu nu c mi thm hc t nhin

c trng ca ngh.

Curcumin c tc dng gip tng bi tit dch mt tiu ha cht bo, bo v gan,

c tnh khng sinh rt mnh. Curcumin c tc dng rt tt i vi d dy, chng li s

lo ha, lm mau lnh vt thng, mau lin so.

Curcumin c tc dng khng ung th, c lp v tiu hy t bo ung th.

Curcumin l mt thnh phn c bit lm nn mu vng c trng ca ngh c kh

nng tiu dit hai loi protein trong cc t bo ung th, cc protein ny chnh l ngun

duy tr s tn ti ca chng.

Lng curcumin ly trch c vi hm lng l 69,5%. Hm lng curcumin v

hiu sut chit cn ph thuc vo hm lng nc trong c ngh. Do c trng v th

nhng ca vng ng bng sng Cu Long l vng trng c nhiu nc v a bn

thnh ph Cao Lnh cng vy nn hm lng nc trong c ngh tng i nhiu.

Thi gian chit l 10h, hn hp dung mi l axeton:etylaxetat t l 1:5, thi gian

kt tinh l khong 2 tun dng dung mi l ietylete. Lng ngh kh cn chit cho

mt ln kt tinh l 400g.

Cc kin ngh cho cc nghin cu tip theo:

+ Khi kt tinh c curcumin th lng nha v tinh du cn nhiu v

bm vo tinh th curcumin nn cn phi tm ra phng php loi nha v tinh du ra

khi dch chit trc khi kt tinh.

+ Tip tc nghin cu kho st tm ra iu kin kt tinh, dung mi kt

tinh, nhit v thi gian kt tinh ti u hn.

+ Tin hnh ly trch vi ngh vng cc vng khc trong tnh ng

Thp sau so snh hm lng curcumin cc a bn.



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TI LIU THAM KHO

1. K Anh (2008), Tc dng thn k ca c gng & ngh phng & tr bnh, Nxb

Nng, H Ch Minh.

2. V Vn Chi (2005), Cy thuc An Giang, Nxb Y hc, H Ch Minh.3. L Anh o, ng Vn Liu (2005), Thc hnh ha hc hu c, Nxb i hc S

phm.

4. Tt Li (2006),Nhng cy thuc v v thuc Vit Nam, Nxb Y dc.

5. Phan Hin Lng, Kthut HPLC, trung tm dch v phn tch th nghim.

6. Nguyn Cng T (2005), Cy thuc v v thuc phng ng, Nxb Tng hp thnh

ph H Ch Minh.

7. o Hu Vinh, Nguyn Xun Dng, Trn Th M Linh, Phm Hng Vit (1985),

Cc phng php sc k, Nxb Khoa hc kthut.

8. www.google.com.vn.

9. http://books.google.com.vn/books?id=P2ykHQi6RvMC&pg=PA243&lpg=PA244 &

ots=OSN4lbYeR-&dq=isolation+curcumin&output=html&sig=AcfU 3U

2BIA154 tiMhOSxi-q5oPEDnILcfQ

10.http://www3.interscience.wiley.com/user/accessdenied?ID=112600014&Act=2138

&Code=4719&Page=/cgi-bin/fulltext/112600014/PDFSTART

11. http://www.phcog.net/phcogmag/issue11/4.pdf

nghien cuu ly trich curcumin tu cu nghe vang

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