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8/13/2019 kha lun - s dng phng php sc k lng

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Xc nh Rhodamine B trong thc phmbng k thut sc k lng hiu nng cao

HPLC s dng detector UV

Trn Th Thanh Nga

Trng i hc Khoa hc T nhin;Khoa Ha hcChuyn ngnh: Ha Phn tch; M s: 60 44 29.

Ngi hng dn: PGS.TS. Phm LunNm bo v: 2011

Abstract.Tng quan v mt s vn c bn ca Rhodamine B nh cng thccu to, tnh cht vt l, tnh cht sinh hc, ng dng, cc phng php xc nhRhodamine B. Chn c cc iu kin ph hp cho vic xc nh RhodamineB c trong cc mu thc phm bng k thut HPLC s dng detector UV- Vis.nh gi c phng php phn tch nh khong tuyn tnh, gii hn phthin, gii hn nh lng, lp li. Chn c quy trnh phn tch v kho stc cc dung mi chit tch i vi cc loi thc phm. tin hnh xc nhc hm lng Rhodamine B trong cc mu thc phm: ht da, bnh xu x,siro du, nc ngt hng du trn quy trnh ti u tm c.

Keywords. Phm mu cng nghip; Rhodamine B; Ha phn tch; Thcphm; K thut sc k

Content:

Khi x hi ngy cng pht trin th vn sc kho ca con ngi ngy

cng c ch trng, trong vn an ton thc phm v v sinh mi trng

c t ln hng u v n c nh hng trc tip n sc kho ca con ngi,

hn na cng l vn ng quan tm ca cc khu ch bin, sn xut v xut

nhp khu thc phm. S tn d ca cc cht c hi c trong thc phm ang l

vn ng lo ngi i vi ngi tiu dng. Ngy nay cng vi s pht trin ca

khoa hc k thut, nhiu k thut phn tch mi, hin i c p dng trong
http://hus.vnu.edu.vn/?portal=main&obj=donvi&id=125http://hus.vnu.edu.vn/?portal=main&obj=donvi&id=125http://hus.vnu.edu.vn/?portal=main&obj=donvi&id=125http://hus.vnu.edu.vn/?portal=main&obj=donvi&id=125http://hus.vnu.edu.vn/?portal=main&obj=donvi&id=125


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nhiu lnh vc khc nhau c bit trong nh gi, kim nh cc cht gy c

trong thc phm.

Trong qu trinh ch bin thc phm, to cho thc phm mu sc p,

bt mt, ngi ta s dng phm mu cng nghip. Phm mu cng nghip ni

chung, Rhodamine B ni ring u c hi, b cm s dng trong thc phm v

kh phn hu. Tu tng c th m nh hng n gan,thn hoc tn d lu ngy

gy c hi n c th con ngi, c bit c th gy ung th. Phm mu thc

phm v t nhin c bn km hn, li t hn phm mu cng nghip. Do

vy nhiu ngi kinh doanh lm dng phm mu cng nghip d cht ny t

lu b cm s dng. V vy vic nghin cu xc nh hm lng ca cc

Rhodamine B- mt thnh phn ca phm nhum trong thc phm l vn cnthit i vi sc kho cng ng.

Tuy nhin, ngoi s c mt ca Rhodamine B cn c cc thnh phn ho

hc khc c trong phm nhum nh Sudan- I, Sudan- IV,Phng php ti u

nht xc nh Rhodamine B l sc k lng hiu nng cao (HPLC). y l mt

phng php c ng dng nhiu nht trong nhiu nm gn y. N c p

dng tch nhn dng v xc nh hng lot cc hp cht m mt s phngphp trc y gp nhiu kh khn nh cc hp cht khng bn vi nhit, cc

hp cht c tnh cht ho hc tng t nhau,Phng php HPLC cng c

nhiu u im m cc phng php khc khng c nh: xc nh ng thi c

nhiu cht, tn t mu, thao tc n gin,

Trong phn tch bng phng php HPLC c hai loi ct tch thng sdng l ct trao i ion v ct tch pha o. Trong lun vn ny, chng ti s

tin hnh kho st nh hng ca cc yu t n qu trnh tch v xc nh hm

lng ca Rhodamine B trong mt s mu thc phm bng phng php sc k

lng hiu nng cao (HPLC) ghp ni detetor UV- VIS. Phng php ny c


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chn lc cao, nhy tt v c trang b nhiu c s kim nghim nc ta,

c tnh kh thiv tnh ng dng thc t cao. Phn tch mt s mu thc phm

nh ht da, bnh xu x, mt, v cc mu thc phm khc nhm nh gi hm

lng Rhodamine B trong cc mu thc phm ny. Da trn cc kt qu nghincu v phn tch hm lng Rhodamine B trongcc mu thc phm m c th

nh gi vn an ton thc phm ca cc c s sn xut.

Chng 1: TNG QUAN

1.1. Mt vi nt v Rhodamine B1.1.1.Cng thc cu to

Rhodamine B l mt hp cht ha hc, l mt thnh phn ca phm mucng nghip.

Cng thc phn t l C28H31ClN2O3

Phn t khi l 479,02g/mol.

Cng thc cu to ca Rhodamine B


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[9-(2-carboxyphenyl)-6-diethylamino-3-xanthenylidene]-diethylammonium chloride

Tnh cht vt l

Rhodamin B l nhng tinh th mu ti c nh xanh hay dng bt mu nu .

Nhit nng chy khong t 2100C n 2110C

Rhodamine B l mt thuc nhum lng tnh, c hi, tan tt trong

methanol, ethanol, nc (khong 50 g/l). ho tan trong 100 gam dung mi:

nc 0,78gam (260C),ru etylic 1,74gam. Dung dch nc v ru etylic c

mu nh xanh nht pht hunh quang mu mnh, c bit r trong cc

dung dch long. Dung dch nc hp th cc i vi nh sng c = 526 v 517 nm

1.1.2.Tnh cht sinh hcRhodamine B gy c cp v mn tnh. Qua tip xc, n gy d ng hoc

lm mn nga da, mt,... Qua ng h hp, n gy ho, nga c, kh th, au

ngc. Qua ng tiu ha, n gy nn ma, c hi cho gan v thn. Nu tch t

dn trong c th n gy nhiu tc hi i vi gan, thn, h sinh sn, h thn kinh

cng nh c th gy ung th. [19,23 ]. Thc nghim trn chut cho thy

Rhodamine B gy ung th vi liu lng 89,5mg/kg qua ng ung hoc timvo tnh mch [23], khi Rhodamine B i vo c th c th chuyn ha thnh

amin thm tng ng c phn c hi hn loi Rhodamine B thng, gy ung

th v pht trin khi u d dy, ti y Rhodamine B v dn xut ca n s tc

ng mnh m n cc qu trnh sinh ha ca t bo gy ung th gan, v gan l


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c quan tng u tin lc cht Rhodamine B [31]. Mt s thc nghim khc cho

thy Rhodamine B tc ng ph v cu trc ADN v nhim sc th khi a vo

nui cy t bo [26,21].

1.1.3.ng dngRhodamine B thng c s dng nh mt thuc nhum tracer trong

nc xc nh tc v hng ca dng chy vn chuyn[24].

c s dng rng ri trong cc ng dng cng ngh sinh hc nh knh

hin vi hunh quang, m t bo dng chy, quang ph hunh quang[24].

Rhodamine B ang c th nghim s dng nh mt bio maker trong

vacxin bnh di cho ng vt hoang d, nh gu trc, xc nh ng vthoang d c thuc phng nga bng cch cho Rhodamine B vo ru v rng

ca ng vt.[24]

N cng c trn vo thuc dit c. Ngoi ra Rhodamine B cn c s

dng to mu v nhum mu trong cng nghip si, nhum mu trong phng

th nghim, xt nghim t bo do tnh bn mu[25].

Rhodamine B c s dng trong sinh hc nh l mt thuc nhum

hunh quang. Tn dng c tnh pht quang ca Rhodamine B, ngi ta dng

chng gip kim sot lng thuc bo v thc vt phun ln cy t, cy ly

du, Rhodamine B c th thm vo t nu dnh du trong my p t, phi t trn

sn c sn cng c th gy ly nhim cht nhum trn. y ban Gia v

cn khuyn co khng ng cc ti ci nhum mu do nghi ngi cht nhum c

th thm lu vo sn phm[25]. Mt khc con ng thm nhp ha cht ny

vo cc sn phm cy trng hu nh khng ai n t trc n nay v nht

l chng li din ra nhiu nc ang pht trin. Ngoi ra, khng ch vi t bt

hay cc cht giav ni chung, cht to mu Rhodamine B c nguy c xut hin

trong hu ht cc sn phm lng thc, thc phm i t cy trng c dng phn

bn ha hc[25].


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1.2. Cc phng php xc nh Rhodamine B1.2.1.Cc phng php sc kPhng php sc k c in (phng phpsc k giy hay sc k bn mng- TLC)

Phng php ny kh n gin v khng yu cu thit b c bit, dng

kim tra nh gi s b cc cht phn tch. Phng php ny c tnh u vit, tinhnh nhiu mu song song trong mt lc rt tin li. Khi TLC c trang b

phn pht hin l mt my o quang c th phn tch nh tnh v nh lng

[27,28].

Trong phng php ny, ngi ta ha tan Rhodamine B chun trong

ethanol tuyt i thu c dung dch c nng khong 10g/ml. Ri tin

hnh xc nh nh tnh trong iu kin sc k s dng bn mng silicagel60F254, hot ha 1100C trong 30 pht. Pha ng c s dng gm hai h:

H 1: CHCl3- MeOH- H2O (65: 35: 10), lylp di;

H 2: EA- MeOH- H2O (100: 17: 13)

Pht hin vt bng cch quan st vt nh sng thng hoc soi di n t

ngoi, bc sng 366nm. So snh v tr v mu sc ca cc vt trn sc k ca

dung dch th vi vt mu ca Rhodamine B trn sc k ca dung dch chun

nh gi kt qu[28].

1.2.1.2. Phng php sc k lng hiu nng cao (HPLC)

Trong nhng nm gn y, phng php HPLC ng vai tr v cng

quan trng trong vic tch v phn tch cc cht trong mi lnh vc khc nhau,

nht l cc lnh vc ca ho dc, sinh ho, ho thc phm, nng ho, ho du,

ho hc hp cht thin nhin, cc loi cht c tc dng c hi, phn tch mitrng,c bit l tch v phn tch lng vt cc cht.

Trn th gii, phng php HPLC c s dng rng ri xc nh

Rhodamine B trong thc phm trong cc loi mu khc nhau, kh u th so vi

cc phng php khc v c chnh xc, nhy v lp li cao


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Detector ghp ni trong my HPLC cho php pht hin s xut hin cht

sau ra gii. Ngy nay c rt nhiu loi detector c s dng cho mc ch ny

m rng kh nng pht hin c rt nhiu loi cht bng phng php

HPLC i vi phn tch d lng th ngi ta hay s dng detector khi ph(MSD) nht l tch v phn tch cht trong cc i tng phc tp. Cn thng

dng ngi ta dng detector UV-Vis hay detector hunh quang. Dng detector

UV-Vis th xc nh c nhiu loi cht, nhng detector hunh quang thng

nhy hn, chn lc hn v t hn cc tng tc do cc hp cht c trong nn

mu. Ngoi ra cn dng mt s detector khc nh detector diode array (DAD),

detector in ho,[18] cc detector ny cng thng c ng dng phn

tch cc cht c trong phm nhum.

1.2.1.3. Nguyn tc chung v trang b ca phng php HPLC

Sc k lng l mt k thut tch cht da trn s t hp ca nhiu qu

trnh va c tnh cht ho hc li va c c tnh cht l hc. N l nhng cn

bng ng xy ra trong ct sc k gia pha tnh v pha ng, l s vn chuyn

v phn b li lin tc ca cc cht tan (hn hp mu phn tch) theo tng lp

cht trongct (pha tnh) t u ct tch n cui ct tch. Trong qu trnh

cht tan lun lun c phn b li gia hai pha, trong khi pha ng chy lin

tc qua ct tch vi mt thnh phn pha ng nht nh, hay gradient. Ngha l

i vi mt phn t cht tan, th trong qu trnh sc k, n lun chuyn t pha

ny sang pha kia nhiu ln t du ct n cui ct sc k. Mt khc, cng v cu

trc v tnh cht ca mi phn t ca cht tan l khc nhau nn tc dch

chuyn trung bnh ca mi cht tan l khc nhau. Khi trong pha ng, n

chuyn dch theo tc ca dng pha ng, cn khi trn pha tnh n li khng

dch chuyn, m b pha tnh gi li. Nh vy l c mt khong thi gian nht

nh cht tan bgi li trong ct tch sc k, thi gian ny ph thuc vo bn


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cht sc k ca ct pha tnh, cng nh tnh cht v cu trc ca mi cht tan

khc nhau ng thi cng ph thuc vo bn cht ca thnh phn pha ng dng

ra gii cht tan, c cht tan t b lu gi, iu dn n kt qu l, c qu

trnh tch ca cc cht xy ra trong ct sc k [6].

Qu trnh tch cht c th xy ra theo ba c ch chnh nh sau: Tng tc hp th Tng tc trao i ion Tng tc theo c ch ry phn tTng ng vi ba c ch trn c ba phng php tin hnh tch khc nhau.

Sc k hp ph (hp ph pha thng NP- HPLC v hp ph pha ngcRP- HPLC)

Sc k trao i ion (EX- HPLC) Sc k ry phn t (Gel- HPLC)Vy phng php sc k lng hiu nng cao (HPLC) l mt k thut tch

cht trong xy ra qu trnh cc cht tan chuyn dch trong ct tch c cha

cc cht nhi kch thc nh, cht tan chuyn dch vi vn tc khc nhau ph

thuc vo h s phn b ca n. Cc cht nhi ct c kch thc nh p ng hiu qutch sc k tt. Thnh phn pha ng c th thay i t c

lc ra gii ph hp nht. Sau khi cht tan chuyn ti cui ct tch c chuyn

ti detector pht hin. Tu thuc vo bn cht ca cht tan m dng cc loi

detector khc nhau [6].

1.2.2.Cc kt qu nghin cu v Rhodamine B bng phng php HPLCTrn th gii c mt s cng trnh nghin cu xc nh Rhodamine B.

Theo Carcinogen v Pesticide Branch [15], phng th nghim ha phn tch

OSHA, thnh ph Salt Lake, Utah, lm th nghim tch Rhodamine B bng

phng php HPLC, s dng detector hunh quang vi cc iu kin nh sau:


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Ct tch: hypersil ODS, 100mm x 2,1mm, 5m.

Nhit : 400C

Pha ng: 85% axetonitrile, 15% nc vi 0,005M axit 1- heptansunfonic, c

iu chnh pH ti 3,5 bng axit H3PO4.Tc dng: 0,2ml/pht

Bc sng: 556nm

Vng mu: 1,0l.

Vi cc iu kin nh trnmu Rhodamine B c pht hin 4,5 pht.

Theo R.W. Mason v L.R.Edwards[22], phn tch Rhodamine B trong

huyt tng th v ngi, s dng my sc k lng hiu nng cao Shimadzu

DR-3 vi detector hunh quang vi cc iu kin:

Ct tch: Bondapak CN (25mm x 4,6mm)

Pha ng: axetonitril v nc (35: 65 hoc 40: 60) cha 0,1% axit

orthophotphoric.

Tc dng: 1,8ml/pht.

Nhit ct: 18 20C

Mu huyt tng: 0,5ml huyt tng c pha long trong 0,5ml ca 0,05M

Kalidihidrogen photphat, pH= 5,5, c chit trong 5ml etylaxetat.

Kt qu phn tch: Rhodamine B c pht hin 9,7 pht. Mu huyt tng

em phn tch c cha t 25 n 50 ng/ml [27].

Rhodamine B trong m phm cng c phn tch bi L. Gagliardi, D. De

Orsi, G. Multari, D. Tonelli [18]. Cc iu kin phn tch c thc hin nh sau:

Ct tch: C- 18

Pha ng: Axetonitril v nc cha 0,1M natri perclorat vi thnh phn thay i

t l t 50: 50n 70: 30.

Mu phn tch c pha trong metanol v nc vi t l 8: 2


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Kt qu phn tch mu thc cho thy dung dch cha 0,3g/ml Rhodamine B, pic

xut hin 9,15 pht [18].

Tc gi J.W.Hofstraat v cng s ng dng k thut chit pha rn v sc k

lng hiu nng cao vi detector hunh quang xc nh Rhodamine B trong

nc b mt. Gii hn pht hin ca phng php l 10 pg/l [18].Ti Vit Nam, vin kim nghim thuc trung ng [1] tin hnh phn

tch Rhodamine B trn cc mu dc liu. Cc iu kin sc k c thc

hin nh sau:

Ct tch: RP- C18 (5m, 4,6mm x 250mm)

Pha ng: 50% Axetonitril- 50% m kalidihidrophotphat 20mM- trietylamin

(100: 0,3), iu chnh pH ti 3,0 bng axit photphoric.

Tc dng: 1,4ml/pht

Detector UV-Vis

Bc sng 525nm.

Lng tim: 20l.

Kt qu phn tch cho thy trong dc liu c cha Rhodamine B[1].

Thng 4 nm 2011, Vit Nam cng xut TCVN 8670-2011 v vic

xc nh Rhodamine B bng HPLC [11] trn c s ca vin kim nghim an

ton v sinh thc phm quc gia xy dng v thc nghim cho mt s loi

thc phm c nhum mu.

1.2.3.Phng php UV- Vis xc nh Rhodamine B xc nh Rhodamine B, ngi ta cn s dng phng php UV- Vis.

Ly mu cht em ho tan trong dung mi thch hp, lc, rung siu m v chitly dung dch. em dung dch chit c o bng my UV- Vis, da vo cc i

hp thta c th xc nh trong mu cht c cha Rhodamine B hay khng

[13, 17].


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Chng 2:

I TNG V PHNG PHP NGHIN CU2.1. i tng, mc tiu v nhim v nghin cu

2.1.1. i tng v mc tiu nghin cu

2.2. Phng php nghin cu

2.2.2. Phn tch nh lng bng HPLC

Trong iu kin phn tch chn, i lng c trng cho mt cht l

thi gian lu tRica cht trn ct tch. Chng ta c th da vo thi gian luny nh tnh c cht thng qua mu chun. Sau da vo cc tn hiu

phn tch thu c (chiu cao pic hoc din tch pic) nh lng cc cht.

Thng thng trong phng php HPLC ngi ta biu din quan h nng

cht ph thuc vo chiu cao pic hoc din tch.

H= k.Cb

S= k. C

b

Trong :

H l chiu cao pic sc k ca cht

S l din tch pic sc k ca cht

k l hng s ca iu kin thc nghim tch sc k


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b- l hng s bn cht, n nhn gi tr trong vng: 0


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phi tin hnh kho st iu kin chn ra c bc sng ph hp nht cho

phn tch cht.

Tin hnh ghi ph hp th quang ca Rhodamine B trong vng Vis trn

my UV-Vis 8453 thu c cc i hp th 526 nm v 572 nm.Chng ti cng tin hnh ghi ph hp th nh sng trong vng Vis trn my UV-

Vis 8453 vi dung dch chun Rhodamine B c pha trong cc thnh phn

dung mi khc nhau:

Rhodamine B 1ppm trong 100% nc3.2. Chn pha tnh

Pha tnh l mt yu t quan trng quyt nh ti hiu qu tch. Bn chtpha tnh quyt nh c ch tch v kh nng lu gi ca cht tan. Ty theo bn

cht ca cht phn tch m chn loi pha tnh, kch thc ht nhi, chiu di ct

cho ph hp qu trnh sc k.

H pha ngc c ng dng ph bin do n nh, lp li v kh

nng tch c nhiu loi cht. Ngoi ra dungmi khi s dng cho pha ngc

c tnh kinh t hn. nghin cu tch v xc nh hm lng Rhodamine B, lcht c tnh phn cc do ch yu cc cng trnh nghin cu c cng b u

s dng ct tch cha cht nhi pha o nh RP- C18, RP- C8,

3.3. Ti u ha pha ng

3.3.1. nh hng ca thnh phn pha ng ti kh nng tch sc k

T l thnh phn dung mi to ra pha ng c nh hng n qu trnh

ra gii cc cht mu ra khi ct tch. Khi t l thnh phn pha ng thay i th

lc ra gii ca pha ng thay i, tc l lm thay i thi gian lu ca cht

phn tch, v do lm thay i h s dung lng ca cht phn tch.


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Do , c c mt thnh phn pha ng ph hp th cn tin hnh

kho st cc t l khc nhau vi cc thnh phn pha ng la chn gm:

Pha ng th nht:

Dung dch m c pH=3 v dung mi hu c methanol (MeOH), thay i

t l pha ng: 60%MeOH- 40% H2O; 65%MeOH- 35% H2O; 70%MeOH- 30%H2O; 75%MeOH- 25% H2O; 80%MeOH- 20% H2O; 85%MeOH- 15% H2O;

90%MeOH- 10% H2O; 95%MeOH- 5% H2O; 100%MeOH- 0% H2O.

3.3.1.1. Pha ng th nht

3.3.1.2. Pha ng th hai

3.3.2.1. Pha ng gm 70% MeOH- 30% m3.3.3.nh hng ca cc cht ph3.3.3.1. nh hng ca trietylamin i vi h pha ng gm MeOH v m3.3.4.Kho st tc pha ng

Cng vi yu t thnh phn pha ng, th tc pha ng khi chy sc k

cng nh hng khng t n kt qu tch sc k. Tc pha ng cng l mt

yu t quyt nh n qu trnh ra gii cc cht trong ct sc k v n nh

hng n qu trnh thit lp cn bng ca cht tan gia hai pha tnh v pha

ng. Tc pha ng qu nh s gy ra hin tng dong pc, thi gian ra

gii cc cht ln lm gim tnh kinh t ca phng php. Nhng tc pha ng

ln qu c th lm cho cc cht trong mu khng kp tch ra khi nhau, dn n

hin tng dong pic. V vy cn la chn c tc pha ph hp.

3.4. nh gi phng php phn tch3.4.1. Tng kt cc iu kin chn

3.4.2. Kho st lp ng chun trong khong nng 0,01- 2,00ppm


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so snh, chng ti tin hnh kho st khong tuyn tnh ca php o

vi cc iu kin nh sau

3.4.3. Gii hn pht hin (limit of detection- LOD)

Gii hn pht hin l nng thp nht ca cht phn tch m phngphp phn tch cn cho tn hiu phn tch khcc ngha vi tn hiu ca mu

trng hay tn hiu nn, hay pc sc k ca cht phn tch phi c chiu cao H tho mn:

H 3 x n

Trong :

H l chiu cao pic

nl lch chun ca tn hiu nn.

T xc nh gii hn pht hin theo hai phng php sau:

3.4.4.Gii hn nh lng (limit of quanlity- LOQ)Gii hn nh lng l nng thp nht (xQ) ca cht phn tch m h

thng phn tch nh lng c vi tn hiu phn tch (yQ) khc c ngha nh

lng vi tn hiu ca mu trng hay tn hiu nn (blank or background).

LOQ = 10 x SB/b

Trong :

b l h s gc ca phng trnh hi quy

SBl lch chun ca mu trng, cng c xc nh theo phng trnh

hi quy.

Nh vy theo phng trnh hi quy ta c:

LOQ= 0,0502 g/ml

3.4.5. ng ca php o


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chnh xc ca php o c nh gi thng qua phn trm sai s.

Chn cc mu phn tch c nng ti im u, cui v gia khong tuyn tnh

kho st. Chng ti tin hnh chun b 3 mu chun c nng ln lt

l 0,1ppm; 0,5ppm; 1,0ppm ri tin hnh chy sc k, mi mu chy lp 8 ln.Phn trm sai s c tnh theo cng thc sau:

% 100i t

t

S SX x

S

Trong : Sil din tch tnh t ng chun

Stl din tch theo sc

Tin hnh kho st cc mu chun vi cng iu kin sc k vi cc nng

khc nhau

Pha tnh: RP- C8 (4,6x150 mm, 5m)

Pha ng: 85% ACN- 15% m (HCOOH- 0,007M natri heptansunfonat), pH=3

Tc pha ng: 0,8 ml/pht

Nhit ct tch: 300C

Th tch vng mu: 20l

Detector: UV-Vis 550 nm

Kt qu thu c i vi mu chun Rhodamine B 0,1ppm c th hin

bng 3.13. v hnh 3.26.

3.4.6. lp li ca php oMt phng php phn tch tt ngoi vic c sai s nh cn yu cu c

3.5. Phn tch mu thc phm, quy trnh x l v kt qu phn tch3.5.1. Kho st dung mi chit ly Rhodamine B

3.5.1.1. X l s b mu phn tch

p dng cc iu kin ti u c phn tch bn i tng mu l ht

da, bnh xu x, nc ngt v tng t.


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Tng s mu phn tch l 3 mu/i tng x 4 i tng (ht da, bnh xu

x, nc ngt v tng t). C ba n v mu ca mt i tng thu thp c

trn vi nhau thnh mu phc hp phn tch.

3.5.1.2. Chn dung mi chitDung mi chit l mt yu t v cng quan trng quyt nh hiu sut ca

qu trnh x l mu. Cc dung mi chit chng ti s dng y l ACN -

Axeton, Etanol- nc, Nc- KCl, ACN- Axeton- nc, nc, etanol.

Chng ti tin hnh x l s b mu ht da ri chit vi cc dung mi

chit khc nhau la chn dung mi chit tt nht.

3.5.2. Phn tch mu thc t dung dch chit

3.5.2.1. Mu ht da

Sau khi tm c cc iu kin ti u cho qu trnh x l mu, chng ti

tin hnh phn tch mu thc th trong cc i tng mu: ht da, bnh xu x,

nc ngt, tng t

i vi mu ht da,quy trnh chit nh sau: mu ht da c cn chnh

xc lng 0,50,002 g trn cn phn tch ri chuyn vo bnh nh mc 25ml,

thm 10 ml hn hp etanol v nc theo t l 40:60, lc u, siu m 60 pht.

Sau khi chit, ly 2,0ml dung dch chit chuyn vo bnh nh mc 10ml v nh

mc bng pha ng, lc u lc qua giy lc thng, ri lc qua mng lc

Whatman 0,45m, ly 2ml dch chit bm vo ct sc k HPLC vi cc iu

kin sc k nh chn

3.5.2.2. Mu bnh xu xGing nh mu ht da trn, chng ti tin hnh chit mu v phn tch

theo quy trnh tng t nhng khng pha long.

i vi mu bnh xu x,quy trnh chit nh sau: mu bnh xu x c cn

chnh xc lng 0,10300,00054 g trn cn phn tch ri chuyn vo bnh nh


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mc 25ml, thm 10 ml hn hp etanol v nc theo t l 40:60, lc u, siu m

60 pht. Sau khi chit, ly 2,0ml dung dch chit chuyn vo bnh nh mc

10ml v nh mc bng pha ng, lc u lc qua giy lc thng, ri lc qua

mng lc Whatman 0,45m, ly 2ml dch chit bm vo ct sc k HPLC vi

cc iu kin sc k nh chn (mc 3.4.1).3.5.2.4. Mu siro du

Qu trnh chit mu v phn tch cng c tin hnh tng t mu ht

da v mu bnh xu x, khng pha long.

i vi mu siro du,quy trnh chit nh sau: mu siro duc ly chnh

xc0,50,001ml bng pipet menri chuyn vo bnh nh mc 25ml, thm 10

ml hn hp etanol v nc theo t l 40:60, lc u, siu m 60 pht. Sau khi

chit, ly 2,0ml dung dch chit chuyn vo bnh nh mc 10ml v nh mc

bng pha ng, lc u lc qua giy lc thng, ri lc qua mng lc Whatman

0,45m, ly 2ml dch chit bm vo ct sc k HPLC vi cc iu kin sc k

nh chn (mc 3.4.1).

Kt qu phn tch mu Rhodamine B trong mu siro du c trnh by

trong bng 3.21.

Bng 3.21. Kt qu phn tch mu siro du

TT Lng mu thc

(ml)

Lng Rhodamine

B chun thm vo

(ppm)

Din tch pc (mAu.s)

1 0,5 0,00 28047

2 0,5 0,01 61781

3 0,5 0,02 96730

Kt qu c tnh ton theo phn mm thng k Origin 7.5 nh sau:

(a) (b)


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Hnh 3.33. ng chun (a) v sc (b)khi thm chun i vi mu siro du.

Ta c:

Cx= a/b = 0,0081(ppm)

Theo cng thc tnh hm lng cht phn tch trong mu ban u, tnhc

mx= 8,1.10-5(mg) hay 0,162mg/l

Hiu sut thu hi ca cht phn tch trong mu l

3.5.2.5. Mu nc ngt hng du

Qu trnh chit mu v phn tch nc ngt hng du cng c tin

hnh tng t mu siro.i vi mu nc ngt hng du,quy trnh chit nh sau: mu nc ngt

c ong chnh xc 10,001 ml bng pipet menri chuyn vo bnh nh mc

25ml, thm 10 ml hn hp etanol v nc theo t l 40:60, lc u, siu m 60

pht. Sau khi chit, ly 2,0ml dung dch chit chuyn vo bnh nh mc 10ml

v nh mc bng pha ng, lc u lc qua giy lc thng, ri lc qua mng

lc Whatman 0,45m, ly 2ml dch chit bm vo ct sc k HPLC vi cc iu

kin sc k nh chn (mc 3.4.1).

Kt qu phn tch cng cho thy trong mu nc ngt c phn tch c

cha 0,126ppm (1,26 mg/l) Rhodamine B .


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KT LUNTrn c s nghin cu cc iu kin thc nghim, nhm ng dng k thut

phn tch HPLC s dng detector UV-Vis xc nh hm lng Rhodamine B

trong thc phm, chng ti thu c mt s kt qu sau y:

1. chn c cc iu kin ph hp cho vic xc nh hm lngRhodamine B c trong cc mu thc phm bng k thut HPLC s dng

detetor UV-Vis:

Pha tnh: RP- C8 (4,6 x 150 mm, 5m)

Pha ng: 85% ACN- 15% m (HCOOH- 0,007mM natri

heptansunfonat), pH=3

Tc pha ng: 0,8 ml/pht

Nhit ct tch: 300C

Th tch vng mu: 20l

Detector: UV-Vis 550 nm

2. nh gi phng php phn tch:Khong tuyn tnh ca Rhodamine B: 0,01- 2ppmGii hn pht hin:

+ Theo phng php phn tch trc tip l 1ppb

+ Theo phng php ng chun l 15ppb

Gii hn nh lng:


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+ Theo phng php phn tch trc tip l 3,33ppb

+ Theo phng php ng chun l 50,2ppb

3. Kho st mu thc chn c quy trnh phn tch v kho st c cc dung mi chit

tch i vi cc loi thc phm l 60% nc - 40% etanol.Trn c s

quy trnh ti u tm c tin hnh xc nh c hm lng

Rhodamine B trong cc mu thc phm gm mu ht da, mu bnh

xu x, mu siro du, mu nc ngt hng du vi lp li tt.

Kt qu xc nh cc mu thc cho thy: trong cc mu thc phmang c lu hnh trn th trng tin hnh phn tch, hm lng

Rhodamine B xc nh c u nm trong gii hn pht hin v gii

hn nh lng ca phng php. Chng t mc d b cm s dng

trong ch bin v bo qun thc phm nhng Rhodamine B vn ang

c s dng kh ph bin trong cc loi thc phm c lu hnh

trn th trng.

T kt qu thu c, chng ti thy phng php HPLC s dng detector

UV-Vis c nhy cao, thch hp cho vic xc nh hm lng Rhodamine B c

trong cc loi thc phm vi cch x l mu thch hp.

Chng ti hy vng nhng nghin cu trn s gp phn vo vic ng dng

k thut HPLC- UV-Vis ni ring v cc k thut HPLC ni chung xc nh

Rodamine B trong cc i tng mu thc phm, nhm phc v c lc cho cc

ngnh khoa hc v c bit trong lnh vc v sinh an ton thc phm gip bo v

sc kho con ngi.

References :


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Thi Bnh (18/ 11/2009), Rhodamine B c trong v thuc ng y l cht

gy ung th, bo Sc kho v i sng.

1.Nguyn Thc Ct, T Vng Nghi, o Hu Vinh (1980) Cs k thuytho hc phn tch, nh xut bn i hc v trung hc chuyn nghip, H Ni.

2.Nguyn Xun Dng, T Vng Nghi, Phm Lun (1986) Cc phngphp tch- Sc k lng cao p, i hc Tng hp Amsterdam, H Ni.

3. Trn T Hiu, T Vng Nghi, Nguyn Vn Ri, Nguyn Xun Trung(2003), Ho hc phn tch- Phn II- Cc phng php phn tch cng

c, HQG H Ni.

4. Nguyn c Kin (2009- 2010), Nghin cu shnh thnh v tch lyc tafltoxin trong bo qun thc n thy sn, lun vn thc s khoa hc-

i hc khoa hc t nhin, i hc Quc gia, H Ni.

5. Phm Lun (1999), C s l thuyt phn tch sc k lng hiu nng cao,i hc Tng hp H Ni.

6. Bi Th Ngoan, Trn Thng, o T Uyn, Phm Vn Hoan (2009), Xcnh Sudan I trong mt s loi giav bng k thut sc k lng hiu nng

cao (HPLC), tp ch y hc thc hnh, s 1 (641+642), trang 58-60.

7. Vn Qun (2007), Xc nh cc hp cht Sudan bng phng phpsc k lng c phn gii cao. Lun vn thc s khoa hc, i hc

Quc gia H Ni.

8. TCVN 8670-2011 v vicxc nh Rhodamine B bng HPLC9. An activity of asean committee on science and technology federation of

institutes of food science and technology in asean (2005), identification

of prohibited colorants in cosmetic products by TLC and HPLC, ACMSIN 02, pp 1-6.

10.Brian Stuart and M.Walker (2006) Analysis of illegal Dyes in ChiliPowder by LC- UV, Statutory analysis government chemist: Programme

ad hoc project 1, pp 1-11.


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11.C.Minier (1996) Rhodamine B accumulation and MXR proteinexpression in musscle blood cells: effects of exposure to vicristine

Marine ecology progress series vol 142 pp 165-173.

12.Carcinogen, Pesticide Branch, (2/1989), Rhodamine B, OSHA analyticalLaboratory- Salt Lake city- Utah.

13.Geertruida Sihombing (2001), AnExploratory Study on three SyntheticColouring Matters Commonly Used as Food colours in Jakarta, Master

Theses from JKPKBPPK.

14.Hu- sheng cheng (2007) Indentification of Rhodamine B 6g andRhodamine B dyes present in ballpoint pen ink using high performance

liquidchromatography and UV vis spectro mettry, Frorensic science

journal pp21-37.

15.L.Gagliardi, D.De Orsi, G.Cavazzutti, G.Multari, D. Tonelli, (6/1996),HPLC determination of rhodamine B (C.I. 45170) in cosmetic products,

Chromatographia Vol.43.ultari

16.Noureddine Barka and CS(2008) Factors influencing the photocatalyticdegradation of Rhodamine B by TiO2- coated non- woven paper journalof photochemistry and photobiology A: Chemistry 195, pp 346-351.

17.Giao Xun, (1/2/2010), Chili powder maker suspended for Rhodamine Bcontaminnation health news, bo Sc kho v i sng.

18.Petr botek, Jan Poustka (2007). Determination of banned dyes in spicesby liquid chromatography- Mass spectrometry, Czech J. Food Sci, vol.25,

No.1, pp 17- 24.19.R.W. Mason v L.R.Edwards (1989), High-performance liquid

chromatographic determination of rhodamine B in rabbit and human

plasma, Journal of Chromatography, 491 page 468- 472.


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