com Ochratoxin

LI M UNh ta u bit ,con ngi l sinh vt d dng nn cc nhu cu v dinh dng ca mnh thng c tha mn bng cc thc phm t ngun ng vt v thc vt.Vic chn la nhng loi ng vt v thc vt lm thc n xut pht t kh nng t kh nng cung cp bi mt mi trng sinh thi xc nh cng l cch ch bin tng hp to nn nt vn ha m thc cho cc c dn tng vng khc nhau .Tiu chun cho s la chn ny ,ngoi cht lng cm quan ,kh nng cung cp v bo qun th s vng mt cc c tnh c coi l yu t quyt nh .Trong qu trnh ny ,nhng sn phm tuy c li ch no v mt dinh dng nhng hm cha mt c tnh ni ti cao th thng b loi khi b su tp thc phm ca con ngi .Thc ra bn cnh cc cht dinh dng ,mt s thc phm thng dng lun cha nhng cht khng c gi tr dinh dng hoc thm ch c nhng cht gy nguy him cho c th . Trong i sng hng ngy ,con ngi lun i u vi nhiu hp cht t nhin hoc nhn to . Trong nhng iu kin nht nh ,s i mt ny c th l nguyn nhn dn n nhng hu qu tai hi cho sc khe : t nhng ri lon sinh hc ti thiu n nhng bnh nguy him .Do vy con ngi lun mong mun hiu bit cht c v phng nga c cc hiu ng c . y nhm em tm hiu chi tit v c t Ochratoxin . Theo T chc Nghin cu ung th quc t (International Agency for Research on Cancer), trong thc phm , c t Ochratoxin c th gy ung th cho ngi m trong qu trnh ch bin cng khng th loi b hon ton c.

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MC LCI/ TNG QUAN 1. Gii thiu chung .................................................................................................3 2. c t ochratoxin .............................................................................................. 4 2.1. S lc .................................................................................................4 2.2. Cu trc ..........................................................................................................4 2.3. Nm mc tng hp OTA................................................................................. 5 2.4. c tnh .......................................................................................................... 5 II/ PHNG PHP PHN TCH 1. Phn tch vi sinh vt ............................................................................................ 7 2. C s khoa hc ...................................................................................................7 2.1. Phm vi p dng ............................................................................................ 8 2.2. Nguyn tc.....................................................................................................8 2.3. Thuc th ...................................................................................................... 8 2.4. Chun b dung dch gc OTA......................................................................... 9 2.5. Dung dch chun OTA ................................................................................... 9 2.6. Dung dch hiu chun OTA............................................................................ 9 2.7. Cch tin hnh ............................................................................................... 9 2.7.1. Khi qut ........................................................................................... 10 2.7.2. Chun b mu th ................................................................................. 10 2.7.3. Chit OTA ra khi mu ........................................................................ 10 2.7.4. iu kin vn hnh HPLC .................................................................... 10 2.7.5. ng chun ........................................................................................ 11 2.8. nh tnh ..................................................................................................... 11 2.8.1. Nhn bit ............................................................................................. 11 2.8.2. Xc nh .............................................................................................. 11 2.8.3. Khng nh .......................................................................................... 12 2.9. nh lng .................................................................................................. 12 3. Phng php Kit Veratox .................................................................................. 13 3.1. Mc ch s dng ........................................................................................ 13 3.2. Th nghim ................................................................................................. 13 3.3. Phng php ............................................................................................... 13 3.4. Thng s k thut ......................................................................................... 13 3.5. Cc vt liu khuyn co ............................................................................... 14 3.6. Nhng vt liu Neogen khng cung cp ...................................................... 14 III/ XUT GII PHP 1. Hn ch ............................................................................................................. 14 2. Bin php .......................................................................................................... 15.

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I / TNG QUAN V C T OCHRATOXIN: 1. Gii thiu chung : Mycotoxin l cc hp cht trao i bc hai c c tnh do mt s vi nm tng hp nn trong cc iu kin xc nh .Thut ng mycotoxin thng c p dng hn ch cho cc c t nm mc ngoi bo c c tnh i vi ng vt mu nng. S nhim nm mc trn nng sn thc phm gy mt mt ,tiu hao nguyn liu/ sn phm trong sut dy chuyn sn xut ra sn phm .c bit nghim trng hn ,s nhim nm mc c kh nng tng hp mycotoxin cc nng sn thc phm lm nhim c chui thc phm v gy ra cc vn nghim trng cho sc khe vt nui ,ngi tiu dng .Theo nh gi ca FAO nm 1996 ,hng nm c hn 25% nng sn b nhim nm mc .Hin nay cc bo co cho thy khng ch mt loi m ng thi nhiu loi mycotoxin nh aflatoxin B,fumonisin hay ochratoxin cng c pht hin trn mt nng sn thc phm. Mycotoxin c th gy c cc cp khc nhau : cp tnh hay trng din .S nhim c cp tnh do mycotoxin gy nn rt nhiu t dch bnh v km theo t l t vong rt cao . Trong chin tranh th gii th hai ,do tnh trng i km ,ngi Nga buc phi n go cn st li trn ng rung .Hng ngn ngi b nhim c ,c lng hon ton b xa s do nhim c t T-2 ca nm mc Fusarium sporotrichioides pht trin trn cc ht la t trn rung. Bn cnh s nhim c cp tnh ,mycotoxin cn gy nn cc bnh l rt khc nhau do n phi Hnh 1 . Nm mc Fusarium thc n c c t trong mt thi gian di .Ty vo bn cht ca tng loi ,mycotoxin c th gy c cho cc h thng khc nhau ca c th nh gan ,thn ,h thn kinh ,h tiu ha ca ng vt tiu th cc sn phm nhim cc c t nm mc. Nhiu mycotoxin nh aflatoxin ,ochratoxin,funomisin c th l cc cht gy bin d ,gy hi chng ung th v qui thai .Ngoi ra, mycotoxin cn c th gy ra s mt cn bng trong chuyn ha thc n v gim t l sinh sn ca vt nui .Bn cnh ,cc dn xut ca mycotoxin thng c i vi h thn kinh v do vy gy tn hi h thng c v gy rt nhiu hi chng khc nh bnh ngoi da ,chng tng sng ha ,tc ng gy ng c ,sy thai .Ngoi cc tc ng trc tip ,mycotoxin cn c kh nng gy cc hi chng gin tip nh c ch vitamin,n khng ngon ming ,hoc bin i cc cht khc thnh cc cht c .Tuy nhin,bnh do mycotoxin gy nn khng c tnh di truyn ,khng ly nhim ,khng gy nhim trng . Do c tnh cc mycotoxin l khc nhau ,v c th do cc nm mc khc nhau tng hp nn ,khng c mt l thuyt chung cho cc mycotoxin v kh nng nhim ,c tnh ,kh nng loi tr ,phng php phn tch V th Ochratoxin l mt c t in hnh ca mycotoxin.

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2. c t ochratoxin : S lc : c t Ochratoxin, l mt sn phm chuyn ho th cp ca mt s loi nm . Ochratoxin c mt trong khp cc loi nng sn thc phm: ng cc, tho dc, bia, c ph... v c trong cc sn phm c ngun gc ng vt do b ly nhim trc. Ochratoxin c to ra t nm Aspergillus (v d: A. ochraceus) v mt vi loi Penicillium. c t nm mc thuc nhm ny gi l Ochratoxin A (= OTA hay OA). C cc phc hp khc trong nhm ny, nhng chng t c hn. Nm Aspergillus ochraceus (to ra c t OTA) thch hp trong iu kin kh hu nng, trong khi nm Penicillium verrucosum to ra c t OTA trong vng kh hu n i. V d A. ochraceus c tm thy Brazil, Chile, Egypt, Senegal, Tunisia, India v Indonesia. Penicillium verrucosum c quan st thy Canada, M, Chu u, v Nam M (theo ngun FAO). Hnh 2. c t nm mc ochratoxin Cc nghin cu ca Php v Chu u mi y cho thy rng cc mu thc phm cha OTA nng t nht vt qu 10 ppb,ln hn 10 ln ngng an ton .Mi y ,nghin cu ca cc nh khoa hc Php cho thy rng trong s 450 mu ru nho ca Chu u ,vang c pht hin nhim OTA nhiu hn so vi vang trng hay vang hng .Ngoi ra s nhim OTA ph thuc rt nhiu vo xut x a l ca nguyn liu . 2.2. Cu trc : Ochratoxin l mt c t rt ph bin bn cnh cc mycotoxin khc nh aflatoxin l cc ochratoxin A,B,v C v cc dn xut ca chng . V cu to ha hc OTA l hp cht ca izocoumarin lin kt vi mt nhm L-phenylalanin.c tnh ca cc ochratoxin khc nhau lin quan ti vic nhm hydroxyl phenol c tch ra kh hay d.

H

COOH

OH

O O CH3

CCNC H H H O

Cl Ochratoxin A4


H

COOH

OH

O O CH 3

CCNC H H H O

H Ochratoxin B H COOH OH O O CH 3

CCNC H H H O

Ochratoxin CCu trc ha hc OTA 2.3.Nm mc tng hp OTA : Cc chng nm mc c kh nng tng hp OTA cn cha c xc nh .Nhng cc nghin cu v OTA cho ti nay cho thy rng cc chng nm mc c th rt khc nhau trn cc i tng khc nhau ,v thuc vo cc ging nm mc ph bin Aspergillus v Penicillium .V d: cc chng gp rt ph bin trn lng thc v thc phm nh Aspergillus carbonarius v Aspergillus ochraceus .Cc nghin cu ch ra rng ,s pht trin ca cc nm mc sinh OTA khc nhau v vic hnh thnh OTA t chng ph thuc rt khc nhau vo nhit , m ,hot nc ca sn phm ,bn cht sn phm .Cc chng sinh OTA rt khc nhau theo khu vc a l ,kh hu v bn cht ca sn phm b nhim .Hn th na ,cc chng tng hp OTA cng c th tng hp ng thi nhiu loi mycotoxin nh axit penicilic hoc citrinin. 2.4. c tnh : Trong s cc ochratoxin , ochratoxin A ( OTA) c tnh c cao nht .y l hp cht khng mi, kt tinh, ha tan trong dung mi phn cc v trong dung dch bicabonat, ha tan hn ch trong nc.

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2.4.1. i vi cc loi ng cc : Ngay t khi gieo trng, tt c cc loi ng cc u b nhim nm mc. Loi nm mc ny c tn l Fusarium. Trong qu trnh thu hoch v lu gi ti kho, ng cc li c nguy c cao b nhim mt loi nm mc c tn l Aspergillus Chnh nhng loi nm mc ny s to nn nhng c t nm mc( nh ochratoxin). c t nm mc l sn phm ph ca qu trnh trao i cht t nhin ca nm mc v gy nn mt s bnh cho gia sc cng nh con ngi . c t nm mc c tnh bn vng nhit cao, n 340oC m khng b phn hy. Hu ht cc loi nguyn liu thc n gia sc u l cc loi ht cc mang sn nm mc v cc loi c t, qu trnh ch bin nhit cng khng lm nh hng n s tn ti ca cc loi c t nm mc ny, y chnh l nguyn nhn lm gia sc mc bnh v gy nn nhng thit hi khng nh. Hnh 3 . c t nm mc ng cc 2.4.2. Tc ng i vi vt nui : OTA l c t nm mc lin quan ti bnh thn cp tnh ca ln, gy qui thai cho chut, v phi g. LD 50 cu OTA l 20mg/kg i vi chut v 3,6mg/kg i vi g con 10 ngy tui . * Trn heo, OTA gy tn thng thn. Thc ra, OTA thu ht cc protein huyt tng bao quanh n, to ra mt hp cht kh bn vng . Hp cht ny c tm thy tht heo v cc sn phm t tht. OTA cng nh hng n s sinh sn ca heo nc. N c truyn qua nhau thai, v c th nh hng n s pht trin ca bo thai. Hin tng cht hng lot heo con mi sinh i khi mt phn l do vi nhim c t OTA. Stoev v cng tc vin quan st thy c t ochratoxin A c trong thc n s lm tng nhy cm ca heo i vi bnh thng hn. Th nghim c thc hin trn 18 con heo, c s bc pht tc thi do S. choleraesuis trn ton b s heo cho n vi khu phn c cha 3 ppm ochratoxin A v trn 1/3 s heo cho n vi khu phn 1ppm ochratoxin. Trong cng th nghim, khng c heo no b mc bnh l i chng. S xut hin tc thi ca bnh thng hn c th gii thch rng nu quan st bng mt thng th tt c heo cc tri u kho mnh, nhng thc t th mi c th heo u c nhim S. cholerasuis. Trong mt th nghim khc, tt c cc heo u c tim vaccin chng li S. choleraesuis (nguyn nhn gy bnh hng l). Tuy nhin, c t nm mc li gy bng pht bnh do nh hng ca Serpulina hyodysenteriae v Campylobacter coli (Stoev v cng s, 2000). Mt s tc gi khc (Muller v cng s, 1999; Verma v Mathew, 1998; Baudrimont v cng s, 1994) trnh by v mt s nh hng khc ca Ochratoxin A trn s min dch ca th: lm gim t bo lympho (t bo min dch), tng apoptotic thc bo (lm cht cc bch cu trung tnh v i thc bo), tng lng bch cu a acid (t bo c lin quan n s p ng min dch), lm tng lng bch cu trung tnh, bch cu a baz v lm gim s thc bo (kh nng tiu dit vi khun ca c th th). 6


* g: chm pht trin, kh nng trao i ging gim, sn lng v cht lng trng gim, tiu chy. 2.4.3. Tc ng i vi c th ngi : i vi ngi, OTA gy chng bnh suy thn vng Bancang ngi .Nhng trng hp nhim c OTA cp tnh c th b t vong .T 1993, C quan nghin cu Quc t v ung th xp OTA vo cc hp cht nhm 2B l nhm cc hp cht c th gy ung th .OTA cn b nghi ng l cht c th gy nhim c thn kinh. Cc khuyn co cho rng ngng hp thu hng ngy cho php i vi OTA l 1,2 - 1,4 mg/kg/ngy. Ngng qui nh cho OTA trong mt s sn phm nh nc qu hoc vang nho c khuyn co l 0,2 1mg/ kg.

II / PHNG PHP PHN TCH : 1. Phn tch vi sinh vt : phn lp cc nm mc nhim trn thc phm, phng php phn lp trc tip hiu qu hn phng php gin tip i vi cc sn phm dng ht .Ht ng cc c t trc tip ln mi trng thch sau khi c lm sch trong dung dch ty clo 0,4% loi b cht bn trn b mt m khng nh hng ti nm mc. i vi cc sn phm dng bt, c th tin hnh ng ha hoc lc mu trong cc dung mi thch hp v s dng k thut pha long xc nh nm mc nhim trn sn phm . Vn c bn khi phn tch nm mc nhim trn sn phm l s pht trin rt nhanh chng ca 1 s loi nm nh Mucor, Rhizopus v cc loi Zygomycetes lm cho vic pht hin hoc nh lng cc nm mc khc rt kh khn. C th s dng cc mi trng ngho nh Potato Dextro agar hoc mi trng c cha cht c ch mt s nm mc nh dicloran, hoc s dng cc mi trng chn lc c ch mt s nm mc c kh nng sinh h si nhanh chng nh Rhizopus. 2. Theo c s khoa hc : Cc ochratoxin pht hunh quang v hp th UV cc i ti 365nm.Do vy c th phn tch OTA bng TLC, bng HPLC, pht hin mu di tia UV. Theo TCVN 7595-07 th c 2 phng php xc nh ochratoxin A trong ng cc v sn phm ng cc bng phng php sc k lng :

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+ Phng php sc k lng hiu nng cao lm sch bng sillicagel + Phng php sc k lng hiu nng cao lm sch bng bicacbonat. y nhm em tm hiu v phng php sc k lng hiu nng cao lm sch bng sillicagel. 2.1. Phm vi p dng : Tiu chun ny qui nh phng php xc nh ochratoxin A vi cc hm lng ln hn 0,4 g/kg. Phng php ny c nh gi thnh cng trong hai nghin cu lin phng th nghim theo ISO 5725 : 1986, thc hin trn bt m c cha hm lng ochratoxin A t 0,4g/kg n 1,2 g/kg. Nguyn tc : Sau khi axit ha bng axit clohydric v nng ion c tng bng cch b sung magie clorua, dng toluene chit ochratoxin A (OTA ). Lm sch dch chit trn mt ct nh sillicagel v xc nh OTA bng sc k lng hiu nng cao HPLC ( High-Performance Liquid Chromatography ) trn ct pha o, nhn dng v sa i bng hunh quang. Kt qu c xc minh li bng dn xut vi bo triflorua trong dung dch metanol nu cn . Ch : OTA gy c n gan, thn v c th gy ung th . Phi tun th cc yu cu phng nga v an ton thch hp khi x l cc hp cht nh th v c bit l trnh x l chng di dng kh v bn cht tnh in hc c th dn n s phn tn v ht phi. Cc dng c thy tinh c th kh nhim bng dung dch natri hypoclorit 4%. 2.3. Thuc th : Trong sut qu trnh phn tch, ch s dng thuc th c cng nhn t cht lng tinh khit phn tch v ch s dng nc ct hoc nc loi 1 ca TCVN 4851 (ISO 3696 ), tr khi c qui nh khc. Dung mi phi t cht lng dng cho phn tch HPLC. 2.3.1. Natri sunfat, dng khan. 2.3.2. Axit acetic bng, ( CH3COOH) 98%. 2.3.3. Dung dch HCl = 2 mol/l 2.3.4. Dung dch MgCl2 = 0,4 mol/l 8


2.3.5. Axetonitril 2.3.6. Toluen. 2.3.7. n-Hexan. 2.3.8. Diclorometan. 2.3.9. Axeton. 2.3.10. Metanol. 2.3.11. Bo triflorua. 2.3.12. Ochratoxin A, dng tinh th hoc di dng vin nang. 2.4.Chun b dung dch gc OTA : Ha tan 1mg ochratoxin (tinh th ) hoc lng cha trong mt vin nang (nu OTA thu c di dng mng mng) vo trong hn hp dung mi I ( gm : toluen v axit acetic bng vi cc phn tng ng l 99 + 1 theo cc phn th tch ) c c dung dch cha khong t 20 g/ml n 30 g/ml OTA. 2.5.Dung dch chun OTA : Cho bay hi di dng kh nito cho n kh : 1 ml dung dch chun gc hoc mt phn tng ng vi mt lng chnh xc 100 g OTA n kh, ri dng pha ng ( trn 99 phn th tch axetonitril vi 99 phn th tch nc v 2 phn th tch axit acetic bng v kh kh dung dch ny trc khi s dng ) pha long n 100 ml. Dung dch ny c th bo qun 4 oC trong t lnh. Cn kim tra tnh n nh ca dung dch. 2.6. Dung dch hiu chun OTA : Dng pipet ly cc dung tch thch hp ca dung dch chun OTA, v d : 1ml ; 2,5ml; 4ml v 5ml cho vo bnh nh mc 100ml v dng pha ng pha long n vch mc. Nng OTA trong cc dung dch hiu chun cn nm trong dy 0,2 ng n 1,0 ng trong 20 l th tch bm.

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2.7. Cch tin hnh : 2.7.1. Khi qut : Ton b qui trnh phn tch cn c thc hin trong ngy. Nu mt s mu c chun b cng mt lc th ton b cc mu cn c phn tch cng mt thi im trong sut c m bng cch s dng b bm mu t ng. 2.7.2. Chun b mu th : Dng my nghin phng th nghim nghin nh mu cho n khi lt qua sng v trn k . i vi bt m c c ht ti a l 250 m th khng cn thit phi nghin. 2.7.3. Chit OTA ra khi mu : - Cn 20g ( mo ) mu chun b chnh xc n 0,1g cho vo ng li tm. Tip theo , cho 30 ml dung dch axit clohydric, 50 ml dung dch MgCl2 , dng a thy tinh khuy v thm 100 ml toluen ( V1). - Lc trong 60 pht v sau cho li tm huyn ph. Thi gian li tm ph thuc vo hiu qu ca my li tm, vic lm lnh trnh tht thot toluen. Ly ra 50 ml (V2) ra khi lp toluen pha trn v np vo ct pha rn loi nh s dng mt ln c chun b v ct ny c ni vi xiranh lm ni cha dung mi. - Ra ct hai ln, mi ln bng 10 ml n-hexan v ra tip 2 ln, mi ln bng 10 ml hn hp dung mi II ( axeton v toluen vi cc phn tng ng l 5 + 95 ). Tip theo, ra bng 5 ml toluen. Loi b tt c nc ra. - Ra gii OTA bng 2 phn hn hp dung mi III ( toluen v axit acetic bng vi cc phn tng ng l 90 + 10 ) mi phn 15 ml cho vo bnh qu l 50 ml. Cho bay hi dch ra gii di p sut gim cho n kh, ch khng nhit vt qu 40 oC. Ly phn cn ra, dng pipet ht 1 ml (V3 ) pha ng cho vo bnh hnh qu l v lc qua b lc mng, vo l. 2.7.4. iu kin vn hnh HPLC : S dng ct tch HPLC pha o phn tch th ci t nh sau : + Tc dng : 1 ml/ pht

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+ Pht hin hunh quang : Bc sng kch thch : 330 nm Bc sng pht x + Th tch bm 2.7.5. ng chun : Chun b ng chun ngay t khi bt u phn tch v bt c khi no thay i cc iu kin sc k . Bm t nht bn dung dch hiu chun c cc nng thch hp khc nhau. V th cc gi tr pht hunh quang ca cc dung dch hiu chun OTA da theo cc nng khi lng OTA tnh bng nanogam. 2.8. nh tnh : 2.8.1. Nhn bit : Nhn bit OTA bng cch so snh thi gian lu ca mu vi thi gian lu ca cht chun. i khi c th cn phi nhn bit pic ca OTA bng cch bm ng thi dung dch mu th v dung dch chun . 2.8.2. Xc nh : Chy sc k ngay i vi mu th. Tin hnh xc nh bng phng php ngoi chun , tch phn in tch pic hoc xc nh chiu cao pic v so snh cc kt qu thu c vi cc gi tr ca cht chun vi din tch pic/ chiu cao pic gn nht, hoc s dng ng chun. Trong trng hp ng chun th cc dung dch b sung c cc nng nm trong di tuyn tnh c th c chun b cho ng chun. Bm cc th tch bng nhau ca dung dch mu th v dung dch chun c dng cho ng chun . c khi lng OTA (m1 ) ,tnh bng nanogam, tng ng vi hunh quang ca dung dch mu th t ng chun. Nu hm lng OTA ca mu nm ngoi ng chun, th iu chnh lng mu bm bng cch c c hoc pha long dung dch mu th. 11 : 20 l ( V4 ) : 460 nm


2.8.3. Khng nh : Nu cn khng nh vic nhn bit bng cch lm bin mt pic ti thi gian lu i vi OTA v cho xut hin pic mi ti cng thi gian lu nh thi gian lu ca este metyl chun ca OTA. Ly 500 l dch chit chun b, chuyn sang bnh hnh qu l v cho bay hi n kh trong b ct quay. Cho 1 ml diclorometan v thm 2 ml dung dch bo triflorua metanol vo phn cn cn li. y cht np bnh v un trn ni cch thy trong 15 pht 50-60oC. Sau khi ngui, chuyn dung dch sang phu chit 50 ml c cha 30 ml nc , lc 3 ln, mi ln 30 giy sau khi thm 10 ml diclorometan . Gp cc pha hu c vo phu chit 50 ml th hai, thm 20 ml nc ra v lc trong 30 giy. Tip theo, lc pha diclorometan qua natri sunfat cho vo bnh hnh qu l, cho bay hi n kh, ri cho thm 500 l pha ng v dung dch ny dng tch sc k di cc iu kin m t trong 2.6.4. Kt thc vic to dn xut ny c th kim tra c t sc ph. C th p dng qui trnh ny xc nh hm lng OTA khng nh hn 0,4 g/kg.. Cn x l mt dung dch chun ring bit kim tra cc thi gian lu ca este metyl ca OTA v kt thc to dn xut.

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2.9. nh lng : Tnh phn khi lng wOTA ca OTA bng g/kg, s dng phng php ngoi chun : wOTA= V1 * V3 * m1 / ( V2 *V4 * m0 ) Trong : V1 : th tch dung mi dng chit ( ml ) , y l 100 ml V2 : th tch li tm (th tch ca toluen) (ml) , y l 50 ml V3 :tng th tch ca dung dch mu th (ml) , y l 1 ml V4 : th tch bm , (ml) m 1 : khi lng ca OTA tng ng vi din tch pic o c hoc chiu cao pic c c t ng chun (ng ,nanogam ) m0 : khi lng ca phn mu th ( g). Ghi li kt qu theo qui tc hin hnh sau khi lm trn n hai ch s thp phn. 12


3. Phng php KIT VERATOX ( nh lng Ochratyoxin ) Hng sn xut: NEOGEN - M (www.neogen.com) 3.1.Mc ch s dng Kit Veratox c s dng phn tch nh lng Ochratoxin trong ng cc, cafe xanh v cc loi hoa qu sy kh. 3.2. Th nghim Th nghim ny dng phng php cnh tranh Elisa xc nh nng chnh xc ca Fumonisin nng c phn t (ppb). Phng php ELISA c s dng rt thng dng trong cc phn tch s b mycotoxin. Phng php ny bao gm vic chit ochratoxin t thc phm, pha long hoc c c mu trn ct, thc hin phn ng ELISA v nh gi kt qu da theo s thay i mu ca c cht trn phn ng.Phng php ny t tin, ch ph hp khi cn phn tch rt nhiu mu cng lc. Ochratoxin t do trong mu v mu so snh cnh tranh vi c t gn enzyme lin kt (tip hp) v pha lin kt khng th. Sau khi ra, cht nn phn ng vi cht tip hp enzyme lin kt to thnh sn phm mu xanh l. Bng phng php quang hc xc nh chnh xc nng ca Ochratoxin. 3.3. Phng php Cn phi ly mu chit trc khi th: Thm 100 l cht tip hp vo mi ging trn Thm 100 l cht so snh v mu th vo mi ging trn. Trn u. Chuyn 100 l vo mi ging khng th. trong khong 10 pht Trt cht lng t mi ging khng th ra. Ra sch hon ton cc ging bng nc loi ion Thm nc bng giy thm. Chuyn 100 l cht nn t cc phn ng ti cc ging khng th s dng 12-u pipet. trong 10 pht. - Chuyn 100 l cht km hm mu t cc phn ng ti mi ging khng th. - S dng b c bn ging vi knh lc 650nm c kt qu.

3.4. Thng s k thut ca sn phm Gii hn d thp nht Khong nh lng Nng mu so snh Thi gian th Khng th phn ng S test/kit 1ppb 2 ppb-25 ppb 0, 2, 5, 10, 25 ppb 20 pht 100% Ochratoxin , 18% OchratoxinB ti 38

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3.5. Cc vt liu khuyn co nhng khng cung cp cng vi Kit. C th t t Neogen: 1. Dung dch methanol 70% tinh khit. 2. ng ong chia 3. Cc ng mu dung tch 125 mL 4. Syringe lc Neogen, giy lc Whateman s 01 hoc tng ng 5. ng ng mu 6. My nghin Agri-Grind, hoc tng ng. 7. Cn c th cn vi trng lng 5g 25g 8. B c Mircowell vi knh lc 650 nm 9. Pipet 100 l, pipt 12 knh v u tp. 10. Gi ging 11. ng h bm gi 12. L ra 13. 02 thuyn thuc th cho pipt 12 u. 3.6. Nhng vt liu Neogen khng cung cp: 1. 2. 3. 4. Giy lm kh hoc vt thm nc L cha mu bng nha dung tch gallon (2,27 lt) cha cht thi. Nc ct hoc nc loi ion. Bt ghi bn vi nc.

III / XUT GII PHP : 1 . Hn ch : Cc hiu bit hin ti v OTA cn rt hn ch so vi aflatoxin. Cc nghin cu cho rng c th dng kh quyn iu ha 30% CO 2 c ch hon ton s to thnh OTA. OTA rt bn vng vi cc x l nhit v ha cht. Cc th nghim cho thy ch c 76% hm lng OTA gim i khi gia nhit 250 oC trong 40 pht . So vi aflatoxin,OTA rt bn vng trong mi trng m. Dung dch OTA trong methanol cng khng b phn hy khi b chiu x ( ti 7,5 Mrad ) Cho ti nay, cha c mt bin php hiu qu phn hy c t nm mc loi ny. V vy, mi bin php gp phn lm gim v hn ch vic hnh thnh OTA trong cc sn phm u cn c quan tm v p dng. 2. Bin php : Nm mc v c t nm mc rt ph bin v rt bn, chng ta khng c bin php g hn ch chng trong t nhin cng nh trong qu trnh ch bin thc n gia sc ( v tn km v khng mang li hiu qu kinh t ).

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Gii php duy nht hin nay cho vic hn ch tc hi ca c t nm mc l vic s dng cc cht hp ph. Trn th gii hin nay s dng nhiu loi cht hp ph nhng ch yu tp trung vo 3 nhm chnh : c ngun gc t cht khong, enzym hay nm men. Tuy nhin, s dng cht hp ph c ngun gc cht khong hin c xem l c hiu qu hn c. Vic s dng cc cht hp ph hin nay l ph bin trn th gii, nht l ti cc nc c nn chn nui tin tin. iu kin bo qun trong kho bi cng gip hn ch s pht trin ca nm mc v c t nm mc rt nhiu .

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TI LIU THAM KHO

[ 1 ] L Ngc T, c t hc v an ton thc phm,2006. [ 2 ] Tiu chun quc gia, TCVN 7595-1 : 2007, ISO 15141-1 : 1998, H Ni, 2007 [3]http://www.tihe.org.vn/web/?idx=hoatdongnghiencuu_detail&mod=hoatdon gnghiencuu&act=detail&id=83&ngay=2006-09-27

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