phat quang atp

1

anh gia s tng thich gia phng phap kim tra vi sinh vt truyn thng va phng phap o ATP quang sinh hoc trong qua trinh kim tra v sinh cua dy chuyn san xut bia

Vin Cng nghip thc phm

Trn Thi Hao

Trng i hoc Khoa hoc T nhin; Khoa Sinh hoc Chuyn ngnh: Vi sinh vt; M s: 604240 Ngi hng dn: TS. Phm Vn Thnh

Nm bo v: 2011

Abstract. Tng quan tinh hinh ng c thc phm Vit Nam, thc trng v sinh

mt s c s sn xut ch bin thc phm v vi sinh vt gy ng c thng gp

trong thc phm; cng ngh sn xut bia, nhng cng on cn thit phi kim tra

v sinh trong sn xut. Nghin cu cac phng phap kim tra v sinh trong sn

xut hin nay; gii thiu v phng phap ATP quang sinh. Kim tra qua trinh v

sinh trong dy chuyn cng ngh sn xut bia c s sn xut ca Vin Cng

nghip thc phm; cac v tr kim tra l cac v tr trong yu ca qua trinh nh cac

tec ln men v cac tec sn phm, may loc sn phm. Trinh by cac phng phap

nghin cu: phng phap kim tra vi sinh vt bng nui cy truyn thng; phng

phap xac nh sch bng ATP quang sinh. a ra kt qu v tho lun: phng

phap phn tch truyn thng; phng phap ATP quang sinh hoc. anh gia s tng

thch gia kim tra vi sinh vt bng phng phap truyn thng v o ATP quang

sinh. Kho sat ng dng phng phap to mu xac nh tng s Coliforms trn may

quang ph U-1900 ( UV-VIS spectrophotometre).

Keywords. Vi sinh vt; Sinh hoc; V sinh; Cng nghip thc phm; Ng c thc

phm

Content.

Thc phm l nhu cu thit yu hng ngy ca mi con ngi. An ton

thc phm tr thnh vn ton cu; trc nhng him hoa lun rinh rp,

e doa i sng con ngi bi cac thc phm khng an ton, i hi mi

quc gia phi c nhng quc sach m bo an ton v sinh thc phm.

2

Trn thc t, mt thc trng cn bc xc trong ngnh sn xut ch bin

thc phm nc ta l cht lng v sinh cn rt km nhiu c s sn xut.

Nhng yu cu nghim ngt v v sinh an ton thc phm trong sn xut cha

c quan tm v u t thch ang. Mt khac do li nhun m mt s t nh

cung cp s dng cac ha cht c hi ch bin, nui trng v sn xut

cac sn phm thc phm; gy nh hng khng tt ti sc khe ngi tiu

dng. Vic s dng cac thc phm khng m bo cht lng v sinh c th

gy ng c v gy ra cac bnh tiu ha cp tnh cho ngi s dng, nghim

trong hn c th dn ti t vong. V lu di, nu cac c t tch ly dn dn

ti mt ngng nht nh c th s phat sinh cac bnh nguy him, lm bin

i cu trc gen gy d tt, d dng cho cac th h tip theo.

Trc thc trng trn, ngoi vic khuyn khch cac c s sn xut thc

hin v sinh an ton thc phm Nh nc v chnh ph cn ch th v vic

tng cng cng tac bo m cht lng v sinh an ton thc phm tng

bc nng cao cht lng sn phm. Tuy nhin phng thc qun l kim tra

cht lng v sinh an ton thc phm da trn vic kim tra cht lng sn

phm cui cng khng m bo c cht lng v sinh sn phm trong khi

sn xut. iu ny ch c m bo khi c s kim soat ch ng v h

thng cac yu t cht lng v iu kin v sinh trong ton b qua trinh sn

xut, ch bin, cung ng mi c kh nng m bo an ton thc phm.

ap ng c yu cu ny khng gi khac l phi ap dng cac phng phap

kim tra nhanh mc v sinh trong cac dy chuyn sn xut, ch bin thc

phm .

Gp phn vo s kim soat v sinh an ton trong sn xut ung ni chung

v sn xut bia ni ring, chng ti tin hnh thc hin kim tra v sinh trong

dy chuyn sn xut bia, vi ti: anh gia s tng thch gia phng

phap kim tra vi sinh vt truyn thng v phng phap o ATP quang sinh

hoc trong qua trinh kim tra v sinh ca dy chuyn sn xut bia Vin cng

nghip thc phm.

I. I TNG V PHNG PHP NGHIN CU

2.1. i tng nghin cu

Kim tra qua trinh v sinh trong dy chuyn cng ngh sn xut bia

c s sn xut ca Vin Cng nghip thc phm. Cac v tr kim tra l cac v

3

tr trong yu ca qua trinh nh cac tec ln men v cac tec sn phm, may loc

sn phm.

Ch tiu la chon kim tra l: tng nm men, nm mc, tng Coliform

v E.coli, tng vi sinh vt hiu kh

2.2. Trang thit b, dng c v phng php nghin cu

2.2.1. Dng c v thit b, ha cht

a. Dng c thit b

+ May o ATP HY_LITE 2

+ May quang ph UV-VIS U-1900

+ Ni kh trng

+ T m

+ T cy

+ May o pH

+ Pipet t ng, u ht

+ a petri, giy nhm, n cn, binh xt cn, a thy tinh,

b. Ha cht

+ Plate count agar ( Peptone t Carein 5,0g; cao nm men 2,5g; D+

gluco 1,0g; agar 14,0g)

Cach pha:

Ha tan 22,5 g PCA trong 1000 mL nc ct. Hp kh trng 121oC,

15pht. Sau khi kh trng pH: 7,0 0,2 25 oC.

+ Lactose broth ( Peptone 10g; lactose 10g; beef extract 6g; nc ct

1000 mL).

Cach pha:

Mi trng n: ha tan 13g lactose broth trong 1000mL nc ct.

Mi trng kp: ha tan 26g lactose broth trong 1000 mL nc ct.

Ha tan mi trng, sau em hp kh trng 121oC, 15pht. Sau khi kh

trng pH: 6,9 0,2 25 oC.

4

+ Brilliant-green lactose (bile) broth ( Peptone 10g; lactose 10g; OX

bile 20g; Brilliant green 13 mL; nc ct 1000 mL).

Cach pha:

Mi trng n: ha tan 40g trong 1000 mL nc ct.

Mi trng kp: ha tan 80g trong 1000 mL nc ct.

Ha tan mi trng, sau em hp kh trng 121oC, 15pht. Sau khi kh

trng pH: 7,2 0,2 25 oC.

Tryptone

Tryptone ( Peptone t casein ): 20g.

NaCl: 5g.

Nc ct 1000 mL.

Cach pha:

Ha tan mi trng, sau hp kh trng 121oC, 15pht. Sau khi kh trng

pH: 7,5 0.2 25 oC.

+ R2A agar (Yeast extract 0,5; proteose peptone 0,5; casein hydrolysat

0,5; glucose 0,5; starch soluble 0,5; natri pyruvat 0,3; K2HPO4 0,3; MgSO4

0,05; agar 12,0).

Cach pha:

Ha tan 15.2 g trong 1000 mL nc ct. em hp kh trng 121oC trong

vng 15 pht. Sau kh trng pH 7,2 0,2 25 oC.

+ Kovacs

+ Oxidase

+ Fluoro cult (Tryptone 5,0; NaCl 5,0; Sorbit 1,0; Tryptophan 1,0; 4-

brom-4chlor-3 indoly B-D- glactogyranoside (X-gal) 0,08; 4-

metylumbelliferyl -D-glucuronide (MGG) 0.05; 1-isopropyl -D-1-

thiogalactopyranoside 0.1 ).

Cach pha:

Mi trng n: cn 17g/1000mL nc ct

5

Mi trng kp: cn 34g/1000mL nc ct

Ha tan mi trng, sau hp kh trng 121oC, 15pht. Sau khi kh trng

pH: 6,8 0,2 25 oC.

2.2.2. Phng php nghin cu

2.2.2.1.Phng php kim tra vi sinh vt bng nui cy truyn thng

Phng phap truyn thng l phng phap ph bin nht trong cac

nghin cu vi sinh vt hoc. Trn mi trng c mi vi sinh vt s phat trin

thnh khun lc hoc s sinh kh v biu hin c ng nghim. Da trn

y c th kim tra s lng vi sinh vt trong mi n v th tch ca dch

nghin cu. Phng phap m khun lc thng c s dng kim tra s

lng nm men, nm mc.

2.2.2.1.1. Phng phap xac nh tng s Coliform v E.coli

Cy vo 5 ng ng mi trng canh thang lng nng kp vi mt

lng 10 mL mu (hoc dung dch mu pha long), trong mi ng u th

ng duham ln ngc. Tip theo, cy vo 5 ng ng mi trng canh thang

lng nng n vi mt lng 1 mL mu (hoc dung dch mu pha long),

trong mi ng u th ng duham ln ngc. Cy vo 5 ng ng mi trng

canh thang lng nng n vi mt lng 0.1 mL mu (hoc dung dch

mu pha long), trong mi ng u th ng duham ln ngc.

cac ng sau khi cho mu vo nhit 35oC trong 24 hoc 48 h v

quan sat. Vi nhng ng c biu hin c v sinh kh, chuyn tip ln lt 10

mL , 1mL, 0.1mL vo 5 ng mi trng brilliant kp v 5; 5 ng mi trng

Brilliant n, nui tip 35oC sau 48 h. sau y quan sat v oc kt qu theo

bng kt qu trong ISO 9308-phn II. Mu c coliform vi biu hin c v

sinh kh ng.

Chuyn tip 10 mL, 1mL, 0.1 mL vo 5 ng mi trng tryptone kp

v 5; 5 ng mi trng Tryptone n, nui tip 44oC sau 24 h. Cho vo mi

ng vi giot Kovacs, nu biu hin mu hng trong ng, chng t s c mt

ca E.coli.

6

2.2.2.1.2. Tnh tng s nm men, nm mc

Chun b dch pha long v mi trng a thch. Nu tng loi mi

trng thch hp cho tng nhm vi sinh vt. Ha tan mi trng ri phn phi

vo cac lo, kh trng bng ni hp ap lc 121oC trong 15 pht. Ly 1mL

mu vi t l pha long nht nh cho vo cac hp a thch c kh

trng, mi mc pha long dng 4 a thch. Sau y rt vo mi hp a

khong 10 mL mi trng thch sau kh trng v ngui 40-50oC. y

np v xoay trn mi hp trn mt t cy, ngui n khi mt thch ng

li, ghi s mu v nng pha long ln mi ay hp petri v boc giy bao

ri em nui 35oC v oc kt qu sau 48h 3.

2.2.2.1.3. Tng vi sinh vt hiu kh

S dng mi trng R2A agar, ha tan vi lng 15,2g/1000 mL, tip

theo em hp kh trng 121oC trong 15 pht.

Ly 1 mL mu hoc mu pha long cho vo hp petri c kh

trng, rt vo khong 10 mL mi trng R2A agar kh trng, y np v

xoay trn u mi hp trn mt t cy, ngui, i khi mt thch ng li

hon ton sau y ghi tn mu v nng pha long ln ay hp petri, bao

giy ri em nui 35oC trong vng 48h 3. Khi thi gian, em ra oc

kt qu bng cach m cac khun lc trn a thch. Sau y bao cao s

khun lc trn mt n v th tch mu tng ng.

2.2.2.2. Phng php xc nh sch bng ATP quang sinh

nghin cu ng dng phng phap kim tra nhanh v sinh trong

dy chuyn sn xut bia bng o ATP quang sinh, cac bc cn phi tin

hnh nh sau:

- Nhn dng cac im kim tra: Cac im kim tra c coi l cac im kim

soat ti hn (CCP), i din cho hiu qu ca qua trinh v sinh. Kt qu thu

c s phn anh thc t hiu qu ca qua trinh v sinh.

Cac im kim tra ni chung c chia lm hai nhm:

B mt tip xc trc tip vi sn phm v nhm b mt tip xc gian tip vi

sn phm.

7

- Lp k hoch ly mu: Cn c s sn xut, xy dng k hoch ly mu,

tn sut ly mu cho ph hp tng v tr kim tra.

Thu thp d liu v xy dng gii hn thng quy: Ni dung ny gip nhn

din tinh trng v sinh hin ti ca cac v tr kim tra. Nhng d liu ny bao

gm ti cac v tr kim tra c trng thai bn v trng thai v sinh sch s.

- Xac nh cac gia tr gii hn: Sau khi c cac s liu v tinh hinh v sinh

tng v tr kim soat, kt hp vi theo di qua trinh sn xut v nhng kt qu

kim tra v sinh thng thng s tin hnh xy dng mc kim soat v sinh

bng ATP quang sinh.

- o ATP quang sinh bng thit b o HY_LITE 2.

c tnh k thut ca my:

+ Khong tuyn tnh 0-99000 RLU

+ C th lu hn 2000 kt qu o

+ Tn hiu nn < 10 RLU

+ Cac dng kim tra: HACCP, Test and store, Test only

+ u d: photomultiplier (PMT) loi cc nhy c chc nng bo v qua ti

Trong phng phap xac nh bng ATP quang sinh c hai dng dng c ly

mu c s dng

+ Que ly mu b mt:

Bi ly mu: Rt que ly mu ra khi dng c Clean-trace, bi ln b mt

kim tra vi din tch khong 10cm2

Hot ha/lc: Cn thn t que ly mu vo dng c Clean-trace, n xung v

lc hot ha mu trong vng 10 giy.

oc kt qu: t dng c Clean-trace vo b phn nhn mu ca may v i

oc kt qu khong 10 giy sau y.

+ Que ly mu nc:

Bi que ly mu: Rt que ly mu khi dng c Aqua- trace, nhng que ly

mu vo dung dch kim tra

8

Hot ha/lc: Cn thn t que ly mu vo dng c Aqua-trace, lc hot

ha mu trong vng 10 giy.

oc kt qu: t dng c Aqua-trace vo b phn nhn mu ca may v i

oc kt qu trong vng 10 giy sau y.

2.2.2.3. ng dng my o quang UV-VIS U-1900

c tnh k thut

H quang: Ratio beam

Bc sng: 190-1100nm

rng di quang ph: 4 nm

Stray light: trong 0.5 % (220 nm NaI; 340 nm NaNO2)

chnh xac bc sng: 0.5 nm

tai lp bc sng: 0.3 nm

Dng o: Photometry, Wavelength scan, Time scan, Multiple, Wavelength

measurement

Photometric range: Abs (-3.000 ti 3.000)

% T (0 ti 300%T)

Nng (0.000 ti 9999)

Photometric Accuracy : 0.002 Abs ( 0 ti 0.5 Abs)

0.004 Abs (0.5 ti 1.0 Abs)

% 0.3 T

Photometric repeatability : 0.001 Abs ( 0 ti 0.5 Abs)

0.002 Abs (0.5 ti 1.0 Abs)

% 0.15 T

Tc scan bc sng: 10, 100, 200, 400, 800, 1200, 2400, 3600 nm/pht

Baseline stability ( tri ): 0.0004 Abs/h (500nm, 2 ting sau khi bt may)

Baseline flatness: 0.002 Abs (200 ti 950 nm)

Baseline level: 0.00015 Abs (500nm)

9

Ngun sang: WI v n D2

Detector: Silicon photodiode

Tin hnh: Cac mu mi trng phat quang c chun b. S dng

cuvet 20 mm cho php o. Chon di qut bc sng t 190 nm - 1100 nm.

II. KT QU V THO LUN

Chng ti thc hin kim tra v sinh mt s im trong yu trong dy

chuyn sn xut bia nh:

+ Cac tec ln men

+ May loc sn phm

+ Cac tec sn phm

3.1. Phng php phn tch truyn thng

3.1.1. Kim tra cc tec ln men

Tec ln men 1

Bng 2: Kt qu kim tra v sinh tec ln men 1

Ch tiu phn tch Ra ln 1 Ra ln 2 Ra ln 3 Ra ln 4

Tng vi sinh vt hiu kh 5.4 x106 5.8 x106 5.2 x104 1.5 x103

Tng nm men, nm mc 4.2 x 103 3.7 x10

3 3.2 x10

3 1.0 x 10

2

Tng E.coli (MPN) 0 0 0 0

Tng Coliform (MPN) 4 2,0 2,0

10


Tng vi sinh vt hiu kh 4.2 x106 3.7 x106 2.1 x104 1.0 x103

Tng nm men, nm mc 3.6 x103 3.7x10

3 1.9 x10

3 1.0 x10

2


Tng Coliform (MPN) 2,0 4,0 2,0 2,0

Tec ln men 3

Bng 4: Kt qu kim tra v sinh tec ln men 3


Tng vi sinh vt hiu kh 5.7 x106 5.5 x10

6 4.7 x10

4 1.8 x10

3

Tng nm men, nm mc 3.4 x103 3.3x10

3 2.6 x10

3 0.5 x 10

2


Tng Coliform (MPN) 4,0 2,0

11

Bng 6: Kt qu kim tra v sinh may loc sn phm t 2



3 2.0 x10

2 5 x10

Tng nm men, nm mc 4.7 x102 4.8 x10

2 3.0 x10

2 12


Tng Coliform (MPN) 4,0

12

Bng 9: Kt qu kim tra v sinh ca tec sn phm 2



3 8.0 x10

2 12

Tng nm men, nm mc 5.7 x102 5.4 x10

2 6.2 x10 14


Tng Coliform (MPN) 2,0

13

4 V sinh ln 4 295 297 285

Khi v sinh cac tec ln men ln ra th 1 v th 2 nc cn bn,

cac tec o c u khong 6000 RLU. T ln 3 tr i mc v sinh

kha hn, cac tec o c khong 500 RLU. Kt thc qua trinh kim tra (ln

4) mu o c t mc di 300 RLU c 3 tec

3.2.2. Kim tra my lc sn phm

Mu ly may loc sn phm c tin hnh sau khi x thao vi loc, x

b cn ln u tin trn cac bn loc.

Bng 12: Kt qu kim tra bng phng phap ATP quang sinh

may loc sn phm

S th t Tn mu t 1 (RLU) t 2 (RLU) t 3 (RLU)

1 V sinh ln 1 790 795 787

2 V sinh ln 2 792 620 690

3 V sinh ln 3 254 265 235

4 V sinh ln 4 150 120 137

hai ln v sinh u, may loc sn phm vn cha c sch, sau ln

th 3 v sinh kt qu RLU c ba t u nh hn 300 RLU. may loc sn

phm ch sau 3 ln c th t yu cu v sinh.

3.2.3. Kim tra cc tec sn phm

Bng 13: Kt qu kim tra bng phng phap ATP quang sinh

cac tec sn phm

S th t Tn mu Tec 1 (RLU) Tec 2 (RLU) Tec 3 (RLU)

1 V sinh ln 1 870 734 674

2 V sinh ln 2 723 627 580

3 V sinh ln 3 225 243 235

14

4 V sinh ln 4 131 127 130

Sau ln 1 v ln 2, qua trinh v sinh vn khng thay i ang k. n

ln 3, tt c cac tec u t kt qu di 300 RLU. Cng nh may loc sn

phm, tec sn phm cng t yu cu sau 3 ln v sinh.

3.3. nh gi s tng thch gia kim tra vi sinh vt bng phng php

truyn thng v o ATP quang sinh

3.3.1. Mu bit trc nng

so sanh s tng thch, chng ti chun b cac mu th bit trc

nng . Mu c chun b t nm men Saccharomyces carlbergensis dng

trong sn xut bia, vi nng ban u l 3.5 x 106 t bo/mL Cac mu c

chun b vi nhiu mc nng khac nhau vi h s pha long l 10-1, 10-2,

10-3

, 10-4

kt qu c trinh by bng di y.

Bng 14: Kt qu phn tch mu bit trc nng bng c

phng phap truyn thng v phng phap ATP quang sinh.

Tn mu Tng vi sinh vt

(CFU) o ATP (RLU)

Mu ban u 3.5 x 106 2302

Mu 10-1 3.5 x 105 747

Mu 10-2 3.5 x 104 243

Mu 10-3 3.5 x 103 158

Mu 10-4 3.5 x 102 132

Qua bng kt qu ta thy kt qu o c t phng phap ATP quang

sinh v tng s t bo nm men m c c s tng thch, s n v RLU

tng dn khi s lng t bo tng dn. giai on pha long gia nng

mu 10-4 v mu 10-3, kt qu thay i khng nhiu, nhng c s nhy vot

gia mu 10-1 v 10-2 v c bit l t mu ban u so vi mu pha long

10 ln ( Mu 10-1).

15

3.3.2. Mu trn dy truyn sn xut bia

a. Kim tra tec ln men

Tec 1

Bng 15: Kt qu phn tch tec ln men 1 bng phng phap truyn

thng v phng phap ATP quang sinh


hiu kh (CFU) o ATP (RLU)

Ra ln 1 5.8 x 106 6608

Ra ln 2 5.4 x106 5460

Ra ln 3 5.2 x 104 520

Ra ln 4 1.5 x 103 295

Tec 2

Bng 16: Kt qu phn tch tec ln men 2 bng phng phap

truyn thng v phng phap ATP quang sinh



Ra ln 1 4.2 x 106 5500

Ra ln 2 3.7 x106 4673

Ra ln 3 2.1 x 104 510

Ra ln 4 1.0 x 103 297

Tec 3

Bng 17: Kt qu phn tch tec ln men 3 bng phng phap




16

Ra ln 1 5.7 x 106 6740

Ra ln 2 5.5 x106 6580

Ra ln 3 4.7 x 104 495

Ra ln 4 1.8 x 103 285

Nhn xt:

Qua trinh v sinh, ra ln 1 v ln 2 cha c s thay i nhiu th hin

c phng phap xac nh truyn thng v phng phap ATP quang sinh. T

ln ra th 3, cac tec ln men mi thy biu hin r rt s thay i v sinh,

v n ln ra cui (ln 4) cng thi s m bo v sinh mi c hon tt.

Nu dng phng phng kim tra nhanh bng ATP quang sinh thi tec ln

men hon thnh qua trinh v sinh khi ch s RLU nh hn 300.

b. Kim tra may loc sn phm

t 1

Bng 18: Kt qu phn tch may loc sn phm t 1 bng phng

phap truyn thng v phng phap ATP quang sinh



Ra ln 1 6.2 x 103 790

Ra ln 2 5.9 x103 792

Ra ln 3 3.0 x 102 254

Ra ln 4 0.5 x 102 150

t 2



Tn mu Tng vi sinh vt o ATP (RLU)

17

hiu kh (CFU)

Ra ln 1 6.1 x 103 795

Ra ln 2 6.0 x103 620

Ra ln 3 2.0 x 102 265

Ra ln 4 0.4 x 102 120

t 3





Ra ln 1 5.9 x 103 787

Ra ln 2 5.7 x103 690

Ra ln 3 1.0 x 102 235

Ra ln 4 2.0 x 10 117

Nhn xt: may loc sn phm t sau ln ra th ba s m bo v sinh

c hon tt, c ba t ch s RLU o c u nh hn 300.

a. Kim tra cac tec sn phm

b. Tec 1

Bng 21: Kt qu phn tch tec sn phm 1 bng phng phap




Ra ln 1 6.3 x 103 870

18

Ra ln 2 6.2 x103 723

Ra ln 3 10 x 102 225

Ra ln 4 2.0 x 10 131

Tec 2



Tn mu Tng vi sinh vt hiu

kh (CFU) o ATP (RLU)

Ra ln 1 6.0 x 103 734

Ra ln 2 6.1 x103 627

Ra ln 3 8.0 x 102 243

Ra ln 4 1.2 x 10 127

Tec 3





Ra ln 1 5.4 x 103 674

Ra ln 2 4.8 x103 580

Ra ln 3 9.0 x 102 235

Ra ln 4 1.7 x 10 130

Nhn xt: cac tec ln men sau ln ra v sinh th 3, cac tec t yu cu

v v sinh, cac ch s RLU o c c 3 tec sau ln ra th 3 u nh hn

300.

19

3.4. Kho st ng dng phng php to mu xc nh tng s Coliform

trn my quang ph U-1900 ( UV-VIS spectrophotometre)

S dng nc thi sau v sinh ca xng sn xut kim tra

Coliforms. T ngun nc thi xac nh c Coliforms tng s (k hiu

M) em pha long ra cac nng khac nhau M x 5 -1; M x 10 -1; M x 10 -2

c cac mu c cha Coliforms khac nhau nui cy, sau em o trn

quang ph UV-VIS.

Chng ti tin hnh scan bc sng ba mc nng ( M x 5 -1; M

x 10 -1

; M x 10 -2

). t bc sng 190nm -1100nm, tc 100nm/pht. Kt qu

thu c nh sau:

Nng M x 10-2mL

Bng 24: Kt qu scan mu c nng pha long 10-2 bc sng

190 1100nm

Peak Valley

Th t WL Abs WL Abs

1 967 0.467 1078 0.265

2 655 0.930 908 0.322

3 614 0.957 644 0.897

4 284 3.587 559 0.830

5 243 3.776 277 3.551

Nng M x 10-1


190 1100nm

Peak Valley

Th t WL Abs WL Abs

1 972 0.409 1074 0.220

20

2 292 3.646 907 0.258

3 242 3.753 273 3.591

Nng M x 5-1


190 1100nm

Peak Valley

Th t WL Abs WL Abs

1 970 0.447 910 0.306

2 655 0.951 644 0.914

3 614 0.979 558 0.840

4 292 3.637 276 3.587

5 241 3.793

c 3 mc nng , hp th t cao nht bc sng 241nm 243nm.

Tuy nhin dung dch to mu c mu xanh lc, l vng nhin thy nm trong

di ph t 400 nm- 800nm. Do vy chng ti tin hnh kho vng bc

sng t 400 nm 800 nm. Kt qu thu c nh sau:

Nng ban u M

Bng 27: Kt qu scan mu M bc sng 400 800nm

Peak Valley

Th t WL Abs WL Abs

1 613.5 0.928 569.0 0.825

Nng M x 5 -1

21

Bng 28: Kt qu scan mu M x 5-1 bc sng 400 800nm

Peak Valley

Th t WL Abs WL Abs

1 615 0.992 568 0.886

Nng M x 10 -1


Peak Valley

Th t WL Abs WL Abs

1 615 0.909 588.5 0.775

Nng M x 10 -2


Peak Valley

Th t WL Abs WL Abs

1 656 1.301 640.5 1.272

2 617 1.301 565.5 1.089

Nng M x 10 -3


Peak Valley

Th t WL Abs WL Abs

1 657 0.972 636.5 0.960

2 617.5 0.985 561 0.823

22

c 5 mc nng , scan t bc sng 400nm 800nm, chng ti thu c

hp th cao nht bc sng 617 nm. Chng ti tin hnh o mu nc

thi ly b cha nc thi sau khi v sinh bc sng 617 nm, thu c kt

qu nh sau:

Bng 32: Kt qu phn tch mu ly b cha nc thi sau khi v

sinh bng phng phap truyn thng v o bng may UV-VIS 1900

ca Hitachi.

Tn mu ABS MPN

(coliforms)

M 1.634 13 x 105

M x 10-1

1.442 80 x 104

M x 10-2

1.266 50 x 103

M x 10-3

1.223 40 x 102

M x 10-4

0.852 23 x 10

M x 10-5

0.074 40

Nhn xt:

Nhin vo bng kt qu trn chng ta thy c s tng thch gia hp

thu o c trn may quang ph UV-VIS 1900 v phng phap xac nh

coliform truyn thng (Phng phap nhiu ng xac nh coliform: ISO 9308-

2). S khac nhau v hp th ABS gia cac mc nng khac nhau rt r.

Qua , cho thy ban u c th dng thit b UV VIS 1900 ca Hitachi vo

xac nh s c mt ca coliform.

III. KT LUN V KIN NGH

T kt qu nghin cu c ti cho chng ta mt s kt lun sau:

1. Phng phap nui cy vi sinh vt truyn thng v phng phap o ATP

quang sinh hoc c s tng thch nht nh.

23

2. Phng phap ATP quang sinh c nhiu u im, d thc hin v cho kt

qu nhanh, tng i chnh xac cho cac php th nhanh v v sinh an

ton thc phm.

T kt qu nghin cu cho thy, khi o bng phng phap ATP quang

sinh c kt qu nh hn 300 RLU, thi mt s khu vc v sinh trong dy

truyn sn xut bia t yu cu v v sinh.

Phng phap ATP quang sinh c nhng u im nh sau:

+ L phng phap o nhanh, n gin, d thc hin, d mang i hin

trng.

+ T kt qu o c bng ATP quang sinh khng ch phat hin c s

lng vi sinh vt sng m cn kim tra c hm lng tn d cac cht

dinh dng lm mi trng cho vi sinh vt phat trin.

+ Nh vy gip cac nh may, c s sn xut v ch bin thc phm khc

phc c cac s c v sinh tc thi.

Tuy nhin phng phap ny cng c mt s hn ch:

+ Phng phap ch kim tra c tng s vi sinh vt m cha c phn loi

c tng loi vi sinh vt.

+ Gia thnh ca mi que th ly mu cng tng i t v mi que ly

mu ch dng c mt ln duy nht, nu thao tac ly mu khng ng s

kha tn km khi ly i, ly li nhiu ln.

3. trong nghin cu ny, chng ti tin hnh th ng dng ca may quang

ph UV-VIS 1900 Hitachi v thy c s tng thch kha cht ch gia

hp th v tng s Coliform trong mu. Bc u c th ng dng may o

UV-VIS 1900 Hitachi vo vic xac nh mc c mt ca coliform mt

cach tng i.

Kin ngh:

1. Phng phap o ATP quang sinh cn l phng phap mi, c th em

ra ng dng rng ri v c hiu qu hn, chng ta cn phi u t thi

gian nghin cu v tham kho nhiu hn na.

C nhng nghin cu su hn v ng dng ca may quang ph UV-VIS vo

vic xac nh tng s vi sinh vt ni chung cng nh tng s coliforms. Cng

24

c thm mi tng quan gia hp th v nng vi sinh vt, t c th

cho ra mt kt qu tng ng gia n v hp th ABS v n v ca vi

sinh vt tng s cng nh tng s coliforms

References :

Ting Vit

1. Kiu Hu nh (2010), Gio trnh vi sinh vt hc thc phm, Nh xut bn

giao dc Vit Nam.

2. Nguyn Ln Dng, Nguyn ng c, ng Hng Min (1976), Mt s

phng php nghin cu vi sinh vt hc, Nh xut bn khoa hoc k

thut, tp3.

3. Nguyn Ln Dng (1971), Kim tra s lng v phn lp cc nhm vi sinh

vt, Nh xut bn i hoc tng hp, tp 1

4. Nguyn Vn t (1974), Phn tch lng thc thc phm, Nh xut bn

khoa hoc k thut.

5. Nguyn Quang Ho (1980), Thc hnh vi sinh vt hc, Nh xut bn giao

dc.

6. Lu Lm, Nguyn Yn, Trn Quang (1969), V sinh thc phm, V k

thut.

7. Hong Tch Mch, H Huy Khi (1977), V sinh dinh dng v v sinh

thc phm, Nh xut bn Y hoc.

8. V Vn Ng (1987), K thut xt nghim vi sinh vt hc, Nh xut bn Y

hoc.

9. o Th Nguyn (1995), Vi sinh vt y hc, Nh xut bn Y hoc.

10. Lng c Phm (2000), Vi sinh vt hoc v an ton v sinh thc phm,

Nh xut bn Nng nghip.

11. Phm Vn S, Bi Th Nh Thun, Nguyn Phng Tin (1975), V sinh

thc phm, Nh xut bn Y hoc.

25

12. Phm Vn Thnh (2002), Nghin cu ng dng phng php phn tch

nhanh kim tra v sinh an ton thc phm. Bao cao ti nm

2002. B Cng nghip

13. Vn Tinh (1983), Kim tra cht lng sn phm nng nghip, Nh xut

bn Thnh ph H Ch Minh.

14. Bi Nh Thun, Nguyn Phng Tin, Bi Minh c (1991), Kim nghim

cht lng v thanh tra v sinh an ton thc phm, Nh xut bn Y

hoc.

15. ng dng cc phng php phn tch cng c trong kim sot mi trng

v thc phm (1999), Hi tho khoa hoc H ni.

16. ng dng cc phng php phn tch cng c hin i trong mi trng

v an ton lng thc thc phm (1994), Hi tho khoa hoc H ni.

17. Phm Thu Yn (2001), Kim tra qu trnh v sinh trong sn xut thc

phm, nh gi s tng thch gia phng php kim tra truyn

thng v phng php o ATP quang sinh hc, kha lun tt nghip.

Ting Anh

18. Adam M.R anh Moss M.O (1995), Food Microbiology, The Royal

Society Of Chemistry.

19. Claude Moreau (1974), Mousissures toxique dans Ladimentation

edition, Pari.

20. Harry H (1962), Practical food Microbiology and Technology Wesport,

Connecticut.

21. R.D. Gonzalez. L.M. Tamagnini, P.D. Olmos, G.B. de Sousa (2003),

Evalution of a chromogenic medium for total Coliformss and

Escherichia coli determination in ready-to-eat foods, Food

Microbiology, 20, pp. 601-604.

22. Denis Byamukama, Frank Kansiime, Robert L. Mach, and Andreas H.

Farnleitner (200), Determination of Escherichia coli Cotamination

with Chromocult Coliforms Agar Showed a High Level of

Discrimination Efficiency for Differing Fecal Pollution Levels in

26

Tropical Waters of Kampala, Uganda , Applied and Environmental

Microbiology, pp. 864-868

23. K. Geissler, M. Manafi, I. Amoros and J.L. Alonso (2000), Quantitative

detarmination of total Coliforms and Escherichia coli in marine waters

with chromogenic and flourogenic media, Joural of Applied

Microbiology, 88, pp. 280-285.

24. ICMSF (2005), Microorganisms in food, Second Edition, kluwer

Academic, Plenum Publishers

25. Jay, J.M. (2000), Modern food microbiology, Sixth edition, Aspen

Pushlisher

26. Manning, S.D., (2004), Escherichia coli infections, Chelsea House

Publishers.

27. Standard Method the Examination of Water and Wastewater 21th

edition,

2005, 9215B.

28. Standard Method the Examination of Water and Wastewater 21th

edition,

2005, 9215D

29. ISO 9308-2: First edition 1990-10-01

phat quang atp

Documents