Nghiên Cứu Vai Trò Của Alginat Trong Quá Trình Tạo Nguyên Liệu Probiotics Chứa Lactobacillus Acidophilus

7/23/2019 Nghin Cu Vai Tr Ca Alginat Trong Qu Trnh To Nguyn Liu Probiotics Cha Lactobacillus Acidophilus

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B GIO DC V O TO B Y T

TRNG I HC DC H NI

NINH TH KIM THU

NGHIN CU VAI TR CA ALGINAT

TRONG QU TRNH TO NGUYNLIU PROBIOTICS CHALactobacil lus acidophi lus

LUN VN THC S DC HC

H NI 2013


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B GIO DC V O TO B Y T

TRNG I HC DC H NI

NINH TH KIM THU

NGHIN CU VAI TR CA ALGINATTRONG QU TRNH TO NGUYN

LIU PROBIOTICS CHALactobacil lus acidophi lus

CHUYN NGNH CNG NGH DC PHM V BO CHM S60720402

Ngi hng dn khoa hc:

1. TS. m Thanh Xun

2. ThS. Nguyn Th Trinh Lan

H NI 2013


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MC LC

DANH MC Trang

T VN 1

Chng 1: TNG QUAN 2

1.1. i cng v probiotics 2

1.1.1. Khi nim 2

1.1.2. Cc vi sinh vt c s dng trong ch phm probiotics 2

1.1.3. C ch tc dng 4

1.1.3.1. C chtc dng 5

1.1.3.2. ng dng caprobiotics 5

1.2. Tnh hnh nghin cu v sdng probiotics trn thgii v Vit

Nam

5

1.2.1. Tnh hnh nghin cu sn xut v s dng cc ch phm

probiotics trn thgii

7

1.2.2. Tnh hnh nghin cu sn xut v s dng cc ch phm probiotics

Vit Nam

7

1.3. Lactobacillus acidophilus 10

1.3.1. c im hnh thi, sinh l 10

1.3.2. Kh nng chuyn ha carbohydrat 11

1.3.3. Cc dng ch phm probiotics cha L. acidophilus 11

1.4.

Alginat 12

1.4.1.

Tnh cht 12

1.4.2.ng dng 13

1.4.3.Mt s nghin cu s dng alginate trong bo ch probiotics 13

1.5. Phng php ng kh 15


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1.5.1. u nhc im ca phng php ng kh 16

1.5.2. ng dng 16

1.5.3.Cc t dc bo v thng dng trong ng kh vi sinh vt 16

1.6.

Nang cng 19

1.6.1.c im dng thuc nang cng 19

1.6.2.nh gi cht lng nang cng 20

Chng 2. NGUYN LIU, THIT B V PHNG PHP NGHIN

CU

21

2.1. Nguyn vt liu v thit b 21

2.1.1. Nguyn vt liu s dng 21

2.1.2. Mi trng s dng trong nghin cu22

2.1.3. Cc dung dch v hn dch s dng trong nghin cu22

2.1.4. Thit b22

2.2. Phng php nghin cu 23

2.2.1. Phng php nhn ging 232.2.2.Phng php nui cy thu sinh khi 23

2.2.3.Phng php ng kh 24

2.2.4.Phng php tit khun Tyndall 25

2.2.5.Phng php to thuc bt cha L. acidophilus 25

2.2.6.Phng php to nang cng cha L. acidophilus 25

2.2.7.

Phng php xc nh hm m 25

2.2.8.Phng php xc nh s lng VSV theo nguyn tc pha long

in tc

26

2.2.9.Phng php nh gi nh hng ca alginate n khnng sng 27


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st ca Lactobacillus acidophilus trong qu trnh ng thuc bt

2.2.10.Phng php nh gi nh hng ca alginate n kh nng

sng st ca Lactobacillus acidophilus trong qu trnh ng thuc nang

cng

28

Chng 3. THC NGHIM, KT QU V BN LUN 30

3.1. Nghin cu nh hng ca alginate n khnng sng st ca vi

khun Lactobacillus acidophilus trong qu trnh ng kh

30

3.1.1.nh gi th cht ca cc nguyn liu cha Lactobacillus

acidophilus to thnh sau khi ng kh

30

3.1.2.nh gi m v tc ht m ca cc mu ng kh vi sinh

vt vi t dc bo v l sa gy v alginate

33

3.1.3.Kho st s lng vi khun Lactobacillus acidophilus sng st

sau ng kh

36

3.1.4.So snh kh nng sng st ca vi sinh vt trong nguyn liu ng

kh c v khng c kt hp alginate v sa gy

39

3.2. Nghin cu nh hng ca alginate n khnng sng st ca vi

khun Lactobacillus acidophilus trong qu trnh ng thuc bt v ng

nang cng

42

3.2.1.Nghin cu nh hng ca alginate n kh nng sng st ca vi

khun L. acidophilus trong qu trnh ng thuc bt

43

3.2.2.Nghin cu nh hng ca alginate n kh nng sng st ca vi

khun L. acidophilus trong qu trnh to nang cng

45

Chng 4: BN LUN 48

KT LUN V XUT

55


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DANH MC CC CH VIT TT

B. subtilis :Bacillus subtilis

Cfu (Colony- Forming Units) : S n v khun lc

FAO (Food and Agriculture Organization) : T chc nng lng th gii

Glass : mt trng thi nhit ng qu bo ha khng bn vi nht cao

HDL (High density lipoprotein) : Lipoprotein t trng cao

HDSK : Hn dch sinh khi

IDF (Internation Dairy Federation) : Lin on b sa th gii

Kl/kl : Khi lng/khi lng

Kl/tt : Khi lng/th tch

L. acidophilus :Lactobacillus acidophilus

LAB (Lactic acid bacteria) : Nhm vi khun lactic

LDL (Low density lipoprotein) : Lipoprotein t trng thp

MRS (de Man, Rogosa, Sharpe) : Mi trng nui cy vi khunMT : Mi trng

PPI (Proton Pump Inhibitor) : c ch bm proton

S. boulardii : Saccharomyces boulardii

Tt/kl : Th tch/khi lng

VSV : Vi sinh vt

WHO (World Health Organization) : T chc Y t th gii


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DANH MC CC BNG

STT Tn bng Trang

Bng 1.1 Cc th h bo ch ca ch phm cha probiotics 7

Bng 1.2 Mt s ch phm probiotics trn th trng Vit Nam 9

Bng 1.3 C ch bo v ca t dc trong qu trnh ng kh 17

Bng 2.1 Cc ha cht dng trong nghin cu 21

Bng 2.2 Cc thit b dng trong nghin cu 22

Bng 3.1 Th cht ca cc mu ng kh L. acidophilus vi cc t

dc bo v ti thi im ngay sau khi ng kh

31

Bng 3.2 Bin thin hm m (%) ca mt s mu ng kh L.

acidophilus vi t dc bo v khc nhau theo thi gian

34

Bng 3.3 S lng vi khun sng st trong 4 mu sau ng kh 37

Bng 3.4 T l VSV sng st sau ng kh so vi mu ban u 37

Bng 3.5 Hm m ca cc mu ng kh vi t dc bo v l hn

hp sa gy vi alginate cc nng

40

Bng 3.6 S lng vi sinh vt sng st tnh trn 1g bt sau ng kh

trong cc mu ng kh s dng kt hp alginate v sa

gy.

41

Bng 3.7 S lng vi sinh vt sng st tnh trn 1g bt ng kh 44

Bng 3.8 S lng vi sinh vt sng st trong 1 vin nang trong mi

trng acid HCl pH 1,2 vi tc khuy 50 v/pht

45

Bng 3.9 T l vi sinh vt sng st ca mu alginate so vi mu tinh

bt trong mi trng pH 1,2

46


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DANH MC CC HNH V BIU

STT Tn bng Trang

Hnh 1.1 Hnh nh L. acidophilus trn knh hin vi in t. 11

Hnh 1.2 Cng thc cu to ca acid - L- guluronic v acid - D-

mannuronic

12

Hnh 3.1 Hnh nh mu ng kh vi nc ct. 32

Hnh 3.2 th biu th hm m trung bnh ca cc mu ngay sau

ng kh v trong thi gian bo qun.

34

Hnh 3.3 Hnh nh mu ng kh vi alginate ngay sau khi tho khi

my v sau 3 pht ngoi mi trng.

35

Hnh 3.4 th biu din s lng vi khun L. acidophilus sng st

trong cc mu sau ng kh

38

Hnh 3.5 Hm m ca cc mu ng kh vi t dc bo v l hn

hp sa gy vi alginate cc nng

40

Hnh 3.6 S lng vi sinh vt sng st tnh trn 1g bt sau ng kh

trong cc mu ng kh s dng kt hp alginate v sa gy.

42

Hnh 3.7 Biu th hin s lng vi sinh vt sng st sau khi th

trong mi trng acid HCl pH 1,2

44

Hnh 3.8 S lng vi sinh vt sng st trong 1 vin nang trong mi


46


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LI CM N

Vi s knh trng v long bit n su sc ti xingi li c n n TS. m

Thanh Xunv ThS. Nguyn Th Trinh Lan,nhng ngi th tn tnh hng n

v ch bo cho ti t nhng bc u tin cho n khi ti hon thin lunvn n

ng thi, ti xin gi li c n tiDs. L Ngc Khnhcng cc th c gio,

cc anh ch k thut vin trong b n Cng Nghip Dc to i iu kin gip

ti trong sut qu trnh thc hin ti, ch bo ti trong thi gian l thc nghi.

hn p n ti cng xin gi li c n n an gi hiu cng ton th cc

th c gio trng i hc c i v to i iu kin thun li cho

ti trong thi gian ti hc tp ti trng

V cui cng l li c n ti gi ti M ca ti, gia nh, ngi thn v bn b

ng vin, gip ti trong sut qu trnh hc tp v hon thnh lunvntt nghip

o thi gian l thc nghi cng nh kin thc ca bn thn c hn, kha lun

n cn c nhiu thiu st i rt ong nhn c s gp ca cc th c, bn b

kha lun c hon thin hn

i xin chn thnh c n

i, ng 30, thng , n

cvin

s inh hKim Thu


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T VN

Probiotics thuc nhm cc vi sinh vt sng khng th thiu i vi sckhe con ngi.Nhng vi sinhvt nya vo c th khi cn sng vimt

lng y s c li cho sc khe ca vtch.Chng thngc b sung

km vi ch n hoc qua cc ch phm cha probiotics.

Vi sinh vt hay c s dng nht trong cc ch phm probiotics l

nhm vi sinh vt sinh acid lactic, m trong in hnh l Lactobacillus

acidophilus.

Tuy nhin, probiotics km n nh dobt li ca iu kin bo qun

cng nh hng ro sinh hc ca vt ch. Do vy, cc nghin cu trong nc

cng nh trn th gii hin nay u c thc hinvi mc tiu gip n nh

vi sinh vt. Trong , alginat l t dc c nghin cu nhiu nht trn th

gii. Nhng trong nc cc nghin cu v t dc ny cn rt hn ch.

Xut pht t thc t trn, chng ti thc hin ti:

N a alginat o q o y l

pobos Lactobacil lus acidophil us.

Vi cc mc tiu:

alginat

.

alginat
http://vi.wikipedia.org/w/index.php?title=K%C3%BD_ch%E1%BB%A7&action=edit&redlink=1http://vi.wikipedia.org/w/index.php?title=K%C3%BD_ch%E1%BB%A7&action=edit&redlink=1http://vi.wikipedia.org/w/index.php?title=K%C3%BD_ch%E1%BB%A7&action=edit&redlink=1http://vi.wikipedia.org/w/index.php?title=K%C3%BD_ch%E1%BB%A7&action=edit&redlink=1http://vi.wikipedia.org/w/index.php?title=K%C3%BD_ch%E1%BB%A7&action=edit&redlink=1


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C 1. TNG QUAN

1.1. probiotics

1.1.1.

Khi nim

Probiotics bt ngun tting Hy Lp c ngha l ssng.Khi nim

probiotics u tin xut pht t nh khoa hc Eli Metchnikoff, trong cun

sch Ko di ssng ca ng a ra 1908. ng cho rng nhng ngi nng

dn Bulgary sng lu l v h thng xuyn s dng sa chua c cha vi

khun lactic, cc vi khun ny c li cho vi sinh vt ng rut. [55]

Thut ngprobiotics c gii thiu ln u tin vo nm 1953 biKollath. Probiotics c nh ngha l cc yu tc ngun gc tvi khun,

kch thch spht trin ca cc vi khun khc. [55]

Nm 1974, Parker pht trin nh ngha ny, ng cho rng probiotics

l nhng vi sinh vt v nhng c cht gip cn bng h vi khun ng

rut. Sau n nm 1989, Fuller thay i nh ngha ny, ng cho rng

probiotics l thc phm bsung mt svi sinh vt sng c ch cho vt chnhm cn bng hvi sinh vt ng rut ca n. nh ngha ny nhn mnh

vai tr kh nng sng ca vi sinh vt. Nm 1998, Salminen nh ngha

probiotics l nhng thc phm cha vi khun sng nh hng c li cho sc

kho. Nm 2002 Marteau nh ngha probiotics hon chnh hn

probiotics l nhng chphm cha tbo vi sinh vt hay nhng thnh phn

ca tbo vi sinh vt m nh hng c li cho sc kho. [23], [55]

Nm 2002, Tchc Y t thgii (WHO) v Tchc Lng thc th

gii (FAO) a ra nh ngha probiotics c coi l hon chnh nht hin nay:

Probiotics l nhng vi sinh vt sng m khi sdng vi lng ph hp s

em li li ch cho sc khe ca sinh vt ch. [23], [55]

1.1.2. Cc vi sinh v c sdng trong chphm probiotics

Probiotics c chia lm 3 nhm chnh sau [6]:


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- Nhm vi khun lactic (chim phn ln): gm 2 chi ph bin l chi

Lactobacillus vBifidobacterium.

- Nhm khng phi vi khun lactic:+ ChiPropionibacterium: P. freudenreichii, P. cyclohexanicum,...

+ ChiBacillus: B. subtilis, B. clausii,...

+ ChiBrevibacillus: B. laterosporus,

+ Chi Sporolactobacillus: S. laevolacticus,

+ ChiEscherichia: E. coli,...

- Nhm nm men: chyu l chi Saccharomyces cerevisiae.

Vic la chn cc vi sinh vt trn dng lm nguyn liu probiotics u

phi da vo cc tiu ch sau:

C khnng bm dnh vo nim mc ng tiu ha ca vt ch

Chu c pH thp ddy v acid mt rut non

Khng sinh c tv khng gy bnh cho vt ch

C khnng sinh enzym, cht khng sinh hoc cc sn phm cui cng

m vt chc thsdng c

C khnng sng v c tr trong rut

Dnui cy v c khnng tn ti c lp trong mt thi gian di

L. acidophilusl trc khun chim tlchyu trong scc vi sinh vt c

ch c tr on trn ca ng tiu ha. N c khnng lm gim s lng

cc vi sinh vt hoc nm c hi rut non; c khnng sinh lactase - mt

enzym quan trng trong chuyn ha sa; c lin quan n qu trnh sn xut

mt s vitamin nhm B nh: niacin, acid folic, pyridoxin; tng cng chc

nng ca hthng min dch; gim stiu dit vi sinh vt c ch do sdng

khng sinh di ngy.[6]

Vi cc c im trn,L. acidophilusl vi sinh vt c la chn u

tin sn xut cc chphm probiotics.


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1.1.3.C tc dng ca probiotics

1.1.3.1. C tc dng [3],[4],[6],[7]

- Cnh tranh nng lng, v tr bm:Probiotics cnh tranh cht dinhdng v nng lng vi cc vi khun gy bnh duy tr v pht trin. Bng

cch chim lnh v bm cht vo thnh rut, probiotics ngn nga cc vi

khun c hi tn cng v pht trin.

- Tng cng chc nng chng ca nim mc rut:Probiotics sng

ti ng rut cng ng thi tng cng chc nng chng ca nim mc

rut v gim thiu sdi chuyn ca cc vi khun v khng nguyn trut vo

mch mu. Chc nng ny gip gim snhim khun v dng i vi cc

khng nguyn c trong thc phm.

- Sn sinh ra cc cht c ch:Mt sprobiotics sn sinh ra cc khng

sinh nh bacteriocins gip ngn nga v tiu dit mm bnh. Trong qu trnh

ln men ng, cc thnh phn khc nhau ca probiotics sn sinh ra acid

lactic gip lm gim pH ca rut v ngn cn spht trin ca cc vi khun

khng cn thit. Mt s sn phm sinh ra trong qu trnh trao i cht, qu

trnh ln men chuyn ha carbohydrat nhbutyrat v acid butyric c khnng

chng ung th.

- Kch thch p ng min dch: Probiotics c kh nng lm tng c

phn ng min dch c hiu v khng c hiu bng cch kch thch cc t

bo phn ng min dch (macrophases, lymphocytes) v tng cng s sn

xut ra cc thnh phn min nhim (cytolines, immunoglobulins, interferon).

- Tng cng chc nng tiu ha:Probiotics trong rut non gip gim

lng mt trong c thv gim cholesterol. Probiotics sn sinh ra cc enzym

khc phc qu trnh tiu hy carbohydrate v lm thun tin hn qu trnh

rut hp thnng lng tcc cht dinh dng. Probiotics cng ng thi ln


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men nhng carbohydrat khng tiu ha c trong rut non v sn xut ra

vitamin B, K.

1.1.3.2. ng dng ca probiotics:Probiotics c ng dng:

Trong cc bnh tiu ha [16], [20], [27]

- Tng khnng tiu ha lactose v hot ng ca cc enzym khc

- Htrtrong iu trtiu chy do sdng khng sinh

- Tc dng lnHelicobacterpylori

Tng cng min dnh [22], [39]- Kch thch min dch nim mc

- Tng cng min dch, gim phn ng dng

- Gim nguy c nhim vi khun, nm

- Gim nguy c mc ung th

Chng tng huyt p, gim cholesterol mu [14], [26]

ng dng trong nui trng thy sn

1.2.Tnh hnh nghin cu v sdng probiotics trn thgii v Vit Nam

1.2.1. Tnh hnh nghin cu sn xut vsdng cc chphm probioti cs

trn thgii

Vic s dng thc phm c probiotics (nh 1 thnh phn t nhin ca

thc phm hoc thc phm ln men) c bit n t lu, nhng vic

nghin cu h vi sinh vt ng rut v s dng probiotics mi thc s phttrin t nhng nm 80 ca th k 20. Nhng nghin cu v c im phn

loi v qun th vi sinh vt ng rut ngi v ng vt c tin hnh

bi Apajalahti v cs (1998) [13]; Vander Wielen v cng s (2000) [49]

cho thy nu nh trong rut non ca ngi Bacteroides v Bifidobacterium

chim u th th g l Ruminococcusv Streptococcus. Bng k thut gen,

cc nh nghin cu ch ra rng ch c khong 20 n 50% s loi vi sinh


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vt ng rut ng vt c phn lp, nui cy nh ngun probiotics.

Netherwood v cs (1999) [38]; Gong v cs (2002) [23]; Zhu v cs (2002) [54]

s dng k thut gen nghin cu s thay i cu trc qun th v cim sinh hc ca h vi sinh vt ng rut ng vt di tc ng ca

probiotics. Tuy nhin, cho n nay nhng nhn t no gp phn to nn mt

h vi sinh vt cn bng hoc lm ri lon s cn bng ca h vi sinh vt

ng rut cngcha c hiu bit y . c rt nhiu nghin cu v

vai tr ca probiotics i vi i sng ng vt nh tc ng ca probiotics

i vi h thng min dch nim mc rut (Schat v Mayer, 1991)[46];

(Hersbberg v Mayer, 2000) [25] i vi s thay i ca nim mc rut non

vt nui (McCracken v Lorenz, 2001). [37]

Nhng nh hng c li ca probiotics th hin nhiu kha cnh khc

nhau nhng nhng hiu bit ca con ngi v c ch tc ng ca probiotics

cn rt hn ch. C mt s tc gi cho rng hiu qu ca probiotics trong vic

c ch s pht trin ca cc vi khun gy bnh trong ng tiu ha ca ng

vt c ngha rt quan trng. S km hm c thc hin theo nhng cch

sau: cnh tranh cht dinh dng, sn xut c t v cc sn phm trao i

(cc acid bo bay hi, cc cht ging khng sinh...), cnh tranh v tr bm dnh

nim mc rut v kch thch h thng min dch rut. [22], [40]

a phn cc sn phm probiotics s dng theo ng ung, chng phi

chu tc dng ca dch v, cng nh ca acid mt v th vi sinh vt b cht rt

nhiu v khng n rut c hoc n rut vi s lng rt t khng gy

tc dng. Vic m bo kh nng sng st ca vi sinh vt probiotics trong sn

xut, bo qun, lu hnh, v t l sng st cao khi nrut khng n gin, do

vy c nhiu hng nghin cu khc nhau nh ci tin phng php ng

kh to bt[32], [33], p dng phng php ln men 2 bc, phng php vi

nang ha [15], [21], phng php s dng kt hp vi t dc bo v [41],


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[42]. Vic bo v cc vi sinh vt probiotics chng sng st v pht huy tc

dng khi n rut l mt vn quan trng v ang c gii quyt.

1.2 T x m V am

nc ta, cc ch phm probiotics chim lnh th trng hn 10

nm; tuy nhin vic nghin cu sn xut probiotics phc v cho i sng cn

rt mi m v bt u c quan tm trong khong mt thp k gn y. n

nay, trn th trng nc ta c mt cc sn phm probiotics ca bn thh

bo ch nh sau: [6]

B 1.1: C bo pm pobos

T m VSV m N m

T 1

(Non-

coated)

VSV sng (trong

sa chua, phomat,

kim chi, )

Gi r s dng

nhiu.

VSV gn nh khng

sng st khi qua d dy

v dch mt.

Bo t VSV. Thun li cho nui

cy, sn xut, lu

hnh.

Mt nhiu thi gian

pht trin thnh VSV

chm tc dng, hn ch

kh nng cnh tranh vi

VSV gy bnh.

Probiotics ng

kh.

T l sng nhit

thng cao hn

T l VSV b cht vn

cao.

T 2

Enteric-

coated

VSV a vo di

dng vin nn,

vin nang c lp

bao ngoi tan

trong rut.

Bo v VSV khi i

qua d dy v dch

mt.

Mt thi gian mng bao

tan r gii phng

VSV.


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T 3

Micro-

encapsulated

Dng bao vi nang. Hon thin tng

ca th h 2.

Cha khc phc c

nhc im ca th h

2: kh nng phng thchti rut cha tt, tan

chm trong rut.

T 4

Dual

coated

Lp bn trong l

protein, lp ngoi

l polysaccharid.

VSV c t l sng

cao. Chu c

acid dch v v

mui mt. Lp bao

bn trong c tnh

thc y phng

thch VSV ti ch.

i hi cng ngh cao,

k thut bo ch hin

i, trang thit b t

tin.

Gi thnh cao.

Mt sc sc sn xut nguyn liu probiotics nc ta l Cng ty

TNHH mt thnh vin Pasteur Lt, Cng ty TNHH mt thnh vin vaccin

v sinh phm Nha Trang, vi mt schphm quen thuc trn thtrng nh

Enzym biosub,Biosubtyl DL, Viabiovit, Vivac bio, Healthy liver. Tuy nhin,

nguyn liu chyu trong cc chphm probiotics ca Cng ty TNHH mt

thnh vin Pasteur Lt cng nh cng ty TNHH mt thnh vin vaccin v

sinh phm Nha Trang u l chng vi sinh vtBacillus subtilis. V mi y

xut hin thm cng ty ANABIO R$D, c nh my sn xut nguyn liu

probiotics hin i khng km so vi cc nc trn thgii. Ngoi vic sn

xut nguyn liu probiotics cin trong nc nh nguyn liu cha chng vi

sinh vtBacillus subtilis, ANABIO cn c thm nguyn liu cha cc chng

vi sinh vt lactic. Do vy, thtrng chphm probiotics khng chln vs

lng m cn a dng vchng loi vi sinh vt, ng thi gi thnh sn phm

cc chphm probiotics c hnhiu ln, ngi sdng c thsdng


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sn phm cht lng cao m gi thnh li r. Tuy nhin, sn phm y vn

chyu thuc thh1 nn khnng bo vVSV vn cn rt thp (Bng 1.2.)

Bng 1.2:Mt schphm probiotics trn thtrngVit Nam

STT T

B d T p d bo N s x

1 1 L-Bio-3D 300mg vi khun: L. acidophilus,

B. longum, L. rhamnosus

Me-Auspharm

Vit Nam

2 1 Antibio Gi bt ung 1g cha 108 cfu L.

acidophilus

OrganonM

3 1 Biosubtyl

LD

Gi 1g cha 107 - 108vi khunB.

subtilisvL. acidophilus

Vin vaccin

LtVit

Nam

4 1 Biolacto Vin nang cha t nht 10 vi

khunL. acidophilusvL. bul

garicus

UnionM

5 1 Lactomed Vin nang 320 mg cha:

L. bifidus2 106vi khun

L. acidophilus2 106vi khun

Streptococcus faecalis2 106vi

khun

II Dong - Hn

Quc

6 1 Ultraflore Vin nang 0,2 g cha 2 109nm

men Saccharomyces cerevisiae

SobioPhp

7 1 Antibio

-philus

Vin nang cha 2 1082 109

vi khunL. acidophilus

Lyocentre

Php

8 1 Probio Gi bt ng kh cha 109 L. Imexpharm


19/7210

acidophilus Vit Nam

9 2 Abiiogran Thuc cm, 1g cha 10 cfu L.

acidophilus

Dae Han New

PharmHnQuc

10 3 Lactomin

plus

Hn hp Probiotics bao vi nang

chaL. acidophilus, B. longum, L.

rhamnosus 300 mg

Mebipharm-

Austrapharm

Vit Nam

11 4 Laclean

gold

Bt cm pha hndchchaLactic

cid Bacteria1,25 x 109,

L.acidophilus(100 t cfu/g);L.

rhamnosus(100 t cfu/g);B.

longum(50 t cfu/g);B. bifidum

(50 t cfu/g);Enterococcus

aecium(100 t cfu/g)

Cell-biotech

Hn Quc

12 4 Kidlac Bt cm pha hndch ung:

Lactic Acid Bacteria 0,5 x 109,L.

acidophilus (100 t cfu/g),B.

longum (50 t cfu/g),B. breve (50

t cfu/g), Enterococcus faecium

(100 t cfu/g)

Cell-biotech

Hn Quc

1.3. Lactobaci ll us acidophi lus

1.3.1. m hnh thi, sinh l

L. acidophilus thuc nhm vi sinh vt sinh acid lactic, l trc khun

Gram dng, kch thc t0,60,9 1,56,0 m, mc n hoc i hoc

to chui ngn, khng c bo t, khng di ng, kkh khng bt buc, phn


20/7211

ng catalase m tnh, pht trin ti u 370C, khng pht trin 20220C

hoc nhit cao hn 480C [4], [12], [44].

Hnh 1.1: Hnh nh L. acidophilus trn knh hin vi in t.

1.3.2. Kh yn ha carbohydrat

L. acidophilus chuyn ha glucose, lactose, sucrose sinh acid lactic,

nhng khng sinh kh; khng chuyn ha glycerol, erythritol, D-arabinose, L-

arabinose, D-ribose, D-xylose. [6], [12]

1.3.3. Cc dng chphm probiotics cha L. acidophil us

Trn th trng dc phm hin nay, cc chphm cha L. acidophilus

chyu l dc phm v thc phm chc nngbo chdi dng bt, cm,

nang cng.

Khc vi chphm khc, chphm probiotics phi duy tr slng VSV

sng st nht nh trong thi hn sdng cng nh trong h thng tiu ha

ca ngi s dng. Do , phng php sn xut v thnh phn ch phm

probiotics c ci bin khc so vi chphm thng thng.

V thnh phn, chphm probiotics thng c thm cc t dc bo v

nh cht x, ng,

Vphng php sn xut nguyn liu, hin nay c rt nhiu phng php

sn xut nhm tng kh nng bo v VSV. in hnh l phng php by

bng cch to vi nang [21], phng php phun sy [15],Trong , mt

trong nhng phng php t ra hiu quv c s dng nhiu l phng


21/7212

php ng kh.Phng php ng kh gip cho VSV n nh v gip VSV

c tlsng st cao trong thi gian bo qun. [17]

1.4.

Alginate1.4.1.Tnh cht

Alginate c ngun gc ch yu l t to nu, rong m. Alginate l tn

ch chung acid alginic v 1 s mui ca n nh natri alginate, kali alginate.

Acid alginic l mt heteropolyme saccharid mch thng cu to t hai

gc uronic l acid - L- guluronic (G) v acid - D- mannuronic (M).[43]

Acid LGuluronic Acid DMannuronic

Hnh 1.2:Cng thc cu to ca acid - L-guluronic v acid - D-

mannuronic

Ty theo ngun gc ca alginate m di trung bnh ca mch phn

t, di ca mi block, t l v trnh t kt hp ca chng vi nhau c khc

nhau. iu ny lm cho tnh cht ca alginatebin i trong mt di rng.

Mui natri ca acid alginic dht m, tuy nhin kh n nh nu c

bo qun trong iu kin mt m v m tng i thp.Dung dch Natri

alginate n nh nht pH 410, pH < 3 acid alginic b kt ta li, hp tit

khun c th lm gim nht ca dung dch [24], [43], [50].


22/7213

Thc t natri alginate khng tan trong ethanol (95%), ether,

chloroform, v hn hp ethanol nc c cha hn 30% ethanol. Khng tan

trong cc dung mi hu c khc v trong dung dch acid c pH


23/7214

nc, natri alginate c s dng trong mt s thuc dng ngn cn tro

ngc d dy- thc qun, gim chng hi [43].

1.4.3.

M alginate sTrong nhng nm gn y vic nghin cu tm ra phng php v t

dc thch hp nhm ci thin kh nng sng st ca vi khun probiotics

ngy cng c quan tm. Mt snghin cu gny v vinang ha vi sinh

vt bng vic kt hp dung dch alginate vi dung dch CaCl2 lm gia tng

s lng VSV sng st trong cc iu kin bt li nh: nhit cao, acid thp

d dy, mui mt... [21], [31], [43]. Latha Sabikhi thc hin nghin cu

nm 2010 v nh gi kh nng bo v L. acidophilus trong ch phm

probiotics bng phng php to vi nang vi alginate. Vi nguyn liu ban

u cha khong 109cfu/g, mu khng c bo v u c s lng sngst

sau 1h, 2h trong iu kin pH 1 gim xung cn khong 105 106 cfu/g (gim

khong 1000 ln), cn viphng php to vi nang th sau 1h trong pH 1 s

lng Lactobacillus acidophilus LA1 gim t 107 cfu/g lc ban u xung

cn khong 106cfu/g (ngha l gim khong 10 ln), sau 2h trong iu kin

acid pH 1, s lng VSV ca mu bao vi nang cn 105cfu/g (gim 100 ln so

vi ban u). Cn trong th nghim c s dng alginate lm t dc n th

sau 1h trong mi trng acid HCl pH 1,2 s lng vi sinh vtgim t 1,7 x

109cfu/g xung cn khong 1,9 x 107cfu/g (tng ng khong xp x 100

ln), cn ti thi im sau 2h lng vi sinh vt cn li l khong 1,1 x 107

cfu/g (gim khong 155 ln so vi ban u)[31].

Mt s nghin cukhc licho thy victo nang cng vicc mui

alginate ca cc kim loi ha tr mt (mui natri alginate hoc kali alginate)

cng cho tc dng bo v vi sinh vt. Randolph Stanley Porubcan nghin cu

to nang cng probiotics cha natri alginate vi hm lng bin thin t 10%

n 99%. Kt qu ca nghin cu cho thy t l phi hp ti u ny l 20


24/7215

60%. Cng theo nghin cu ca Randolph Stanley Porubcan, s lng VSV

b cht trong mi trng acid pH 1,6 khng nhiu, c th trc khi th trong

mi trng acid pH 1,6 th s lng cfu/ nang l 9,0x10

9

, sau khi th trongmi trng acid pH 1,6 trong 90 pht s lng cfu/ nang khng cha alginate

nh hn 104, cn s lng cfu/ nang cha alginate l 4x108. Kt qu nghin

cu cng cho thy khi s dng alginate lm t dc n trong nang cng

lm gia tng t l VSV sng st trong mi trng acid pH thp[42].

1.5. P pp k

ngkh cnh ngha nh l mt quy trnh to s n nh m trong

vt cn ng kh c lm ng lnh trc, tip theo lng dung mi c

trong vt (dung mi thng l nc) c loi i nh qu trnh thng hoa, sau

l qu trnh kh m lin kt cn li n gi tr m s khng th khin VSV

pht trin hoc xc tc cc phn ng ha hc [17], [32].

ng kh gm c 3 giai on chnh: tin ng, thng hoa v lm kh

G o

Tin ngl giai on u tin ca qu trnh ng kh, thng c tin

hnh nhit -700C n -800C. giai on ny phnln nc c tch

ra khi dc cht v t dc, h tch thnh nhiu pha. Kt thc giai on

ny, c th c trng thi kt tinh, v nh hnh hoc kt tinh kt hp vi v

nh hnh.[17], [11]

G o o

giai on ny vt c t trong bung sy, tin hnh ht chn khng.

Nc cchuyn trc tip t trng thi rn sang trng thihi m khng

qua giai on lng. Trong giai on ny nhit v psutbn trong bung

sy c duy tr mcdi nhit ng bng. [17], [11]

Giai o lm k


25/7216

giai on ny nhit tng nhanh, m trong vt l m lin kt v

trng thi lng. Qu trnh sy trong giai on ny ging nh qu trnh sy

trong cc thit b sy chn khng bnh thng.Nhit mi cht trongbung sy lc ny cng cao hn nhit giai on thng hoa. [11]

1.5.1. m

ngkh c rt nhiuu im so vi cc phng php sy khc nh:

Gi nguyn mu sc, cu trc, hng v; Gi c hot tnh sinh hc; Bo v

nguyn vn cc vitamin nh lc cn ti, khng lm bin cht albumin Gi

nguyn c th tch ban u ca vt sy nhng c cu trc xp nn d hp

th nc tr li trng thi ban u. Tuy nhin, phng phpng kh cng

c nhng nhc im ring khin n cha c s dng rng ri: gi thnh

thit b cao; vn hnh phc tp, i hi ngi thao tc phi c trnh k

thut cao;tiu hao in nng ln lm gi thnh sn phm tng cao [11].

1.5.2.

Do nhng u nhc im trn nnphng phpng kh ch c p

dng trn cc sn phm c yu cu cht lng cao, ng thi km bn vi

nhit cao. Trong ngnh Dc c ng dng sn xut cc bt ng kh

pha tim (v d: khng sinh penicillin, cephalexin), lm kh cc ch

phm sinh hc nh albumin, vi sinh vt. Ngoi ra ng kh cng c s

dng nh mt phng php bo qun trong ngnh cng nghip thc phm

bo qun 1 s sn phm khc.

1.5.3.C

Qu trnh ng kh vi sinh vt tuy c nhiu u im nhng n cng l

1 trong nhng nguyn nhn lm gim s lng vi sinh vt sng st trong qu

trnh to nguyn liu v ch phm probiotics. c bit giai on tin ng l

nguyn nhn gy ln p lc cho thnh t bo vi khun. ng lnh lm cho lp

lipid mng t bo d b tn thng.Mt nghin cu khc li cho rng s thay


26/7217

i hnh thi t nhin ca lipid mng v s thay i cu trcca cc protein

nhy cm l nguyn nhn chnh gy ra s suy gim s lng tbo trong qu

trnh ng kh [29]. Mt s ti liu li cho rng chnh vic hnh thnh tinh th ni phn t c th l nguyn nhn gy ph v mng t bo. Cc t dc

bo v c thm vo trong thnh phn nguyn liu em ng kh nhm: bo

v t bo trong qu trnh ng kh v m bo sc chu ng ca vi sinh vt

trong qu trnh lm kh [32].

Mt trong nhng phng php c s dng gia tng s sng st

ca vi sinh vt trongqu trnh ny l b sung vo nguyn liu ng kh cc

tc nhn bo v. Cc tc nhn ny nhm bo v t bo vi sinh vt trong qu

trnh ng kh. Cc t dc chia thnh cc nhm chnh sau: nhm

carbohydrate (surcrose, lactose, trehalose, maltose, glucose, galactose,

raffinose), nhm polymer (gelatin, xanthan gum, maltodextrans, dextran),

nhm protein (sa gy, trypsin, pepton, trytone, cao tht,), polyols

(mannitol, glycerol, sorbitol, butanol,), nhm amino acid (Na glutamate,

proline, glutamate,), nhmchng oxy ha (acid ascorbic, Na thiosulfate,)

v mt s t dc bo v khc nh mt ong, Na acetate, Ca chloride,[17].

C ch bo v ca cc nhm t dc ny trong qu trnh ngkh c tm

tt trong bng 1.3[17]:

B 3:C ch bo v ca t dc trong qu trnh ng kh

STT Nhm C bo

1 Carbohydrate Thay th lin kt hydro trong lp mng t bo

lipid kp

2 Polymer Lm gim t l lin kt vi nc

3 Protein Cung cp hm lng acid amin, enzym phn gii


27/7218

protein thnh acid amin, vitamin, lm tng kh

nng m

4 Polyols Thay th nc bay hi t mng t bo lipid kp5 Amino acid n nh protein

6 Chng oxy ha Dn gc t do

7 Mt s t dc bo

v khc

Tng hot tnh enzym, thay th lin kt hydro

trong lp mng lipid kp,

i vi qu trnh ng kh t bo vi sinh vt, nhm carbohydrate,

nhm protein, nhm amino acid v nhm polymer l cc nhm hay c s

dng nht bo v vi sinh vt trong qu trnh ng kh. Trong khng th

khng k n nhm t dc bo v rt hay c s dng lprotein vi t

dc in hnh: sa gy.

S y

Sa gy hay cn gi l skim milk l cc sn phm sa m trongthnh phn c cha khng qu 0,5 % cht bo (trong khi sa nguyn cht

cha hm lng cht bo khong 3,5%, sa tch b-butter milk cha khong

1% cht bo).

Sa gy c trong cc ch phm ng kh probiotic v n c tc dng

bo v tt probiotic trong qu trnh ng kh. Theo nhiu tng kt nhiu

nghin cu trong v ngoi nc t trc ti nay th sa gy s dng nng khong 10% chot l sng st cao nht. Tc dng bo v ny c th do n

cha t l dinh protein v carbohydrat- nhng cht m c kh nng bo v t

bo. Thnh phndinh dng chnh trong sa gy gm 32- 35,7 % protein v

48,4- 54,1 % lactose.

Lactose c trong sa gy cng nh mt s disaccharid khc c th thm

qua thnh t bo, sau chng hnh thnh lin kt hydro gia ng - protein.


28/7219

Cc lin kt ny gip duy tr cu trc ca protein mng khi nc b loi i.

Protein gip hnh thnh trng thi glass xung quanh v bn trong t bo to ra

mt mi trng nht bn trong v ngoi t bo gip ngn cn s bin ica t bo mc thp nht [17].

Theo mt nghin cu khc th protein trong sa gy c th to thnh

lp o bo v trn thnh t bo, ng thi sa gy cn cung cp nhng cht

m tan trong nc (mui phosphat, mui citrat) gip n nh pH trong qu

trnh ng kh [32].

Nguyn liu ng kh vi sa gy va to cu trc xp gip qu trnh

b nc d dng, nhng ng thi cng lm cho nguyn liu ng kh ht m

kh nhanh nu khng c bo qun tt.

1.6. Nang cng

1.6.1. m dng thuc nang cng

Nang cng gelatin l mt dng thuc phn liu gm mt v rng

ng thuc (bng gelatin), gn lin vi thuc v a vo c th cng vi

thuc. Sau khi tan r gii phng thuc,v ng c tiu ha trong c th

[2]. Hin nay, nang cng l dng bo ch ph bin nht vi cc ch phm

probiotics sau dng tinh bt.

i vi ch phm probiotics, vic to dng thuc nang cng gip bo

v vi sinh vt trnhmt phn tc ng bt li ca ngoi mi nh m, nh sng

do vy lm tng t lsng st ca vi sinh vt. Thnh phn t dc chyu

trong nang cng l t dc n, vi in hnh l tinh bt, cc loi ng n,

ng i nh saccarose, lactose, glucose,... Ngoi ra, c thm mt sthnh

phn khc nh t dc trn, t dc mu, t dc iu hng iu v,

i vi nang cng cha probiotics, thnh phn cn c thm cc t dc

gip bo v vi sinh vt trong iu kin bo qun, iu kin ngoi mi, nh

sng, m, hay bo vvi sinh vt bn vng qua acid ddy. in hnh l


29/7220

nhm cht x Fructooligosaccharide, Vegetable cream, inulin,l nhm t

dc hay c sdng nht.

1.6.2 ng nang cngMun hon thin cng thc v quy trnh sn sut nang cng, ng thi

m bo thu c cc nang cng c cht lng ng nht qua cc l mkhc

nhau cn phi nh gi cht lng nang cng da trn cc chtiu cht lng.

Mt schtiu cht lng ca nang cng cn nh gi l: [2]

ng u vhm lng

ng u vkhi lng

r: nang cng phi r trong vng 30 pht

Trc nghim ha tan: Thha tan gip nh gi tc gii phng

dc cht, gip cho cc nh sn sut la chn, sng lc cng thc v cc

thng skthut trong qu trnh sn xut, ng thi y cng l mt chtiu

nh gi cht lng sn phm.

Cc th nghim khc: Xc nh hm lng dc cht, mc ng

nht vhm lng dc cht, tin hnh theo nh chdn trong cc chuyn

lun cthca Dc in i vi tng loi chphm cth.


30/7221

C 2. NGUYN LIU, THIT B

V PHNG PHP NGHIN CU

2.1. Ny l b2.1.1.y

Chng vi sinh vt:Lactobacillus acidophilusATCC 4653

Nguyn liu :

- Alginat:

S dng dng mui Natri alginat.

Ngun gc: to nu

- V nang gelatin:

C nang: c s 1

Mu sc: mu xanh

- Di y l bng cc t dc c s dng trong nghin cu

B 2.1: C d o

T N T Glucose Trung Quc TC NSX

Pepton Merk-c; n TC NSX

Cao tht Merk- c TC NSX

Cao nm men Merk- c TC NSX

K2HPO4 Trung Quc TC NSX

MgSO4.7H2O Trung Quc TC NSX

Calci clorid Trung Quc TC NSX

Sa gy Trung Quc TC NSX

Alginate Trung Quc TC NSX

Triamoni citrat Trung Quc TC NSX


31/7222

Acetat natri Trung Quc TC NSX

MnSO4.4H2O Trung Quc TC NSX

HCl Trung Quc TC NSX

Tinh bt Vit Nam TC NSX

V nang gelatin Vit Nam TC NSX

2.1.2.M

MRS lng (MT1).

Glucose 20g Acetat natri 5g

Pepton 10g K2HPO4 2g

Cao tht 10g MgSO4.7H2O 0,2g

Cao nm men 5g MnSO4.4H2O 0,05g

Triamoni citrat 2g Nc my v 1000ml

pH 6,8 7,0

Mi trng MRS thch (MT2).

MT2= MT1 + thch agar (22g/1000ml mi trng).

2.1.3. C

Hn dch sa gy 10%, 20%

Dung dch alginate 1%, 2%, 5%

Dung dch HCl pH 1,2 (pha theo DVN4)

2.1.4. T

B C b d o

T b N

T cy v trng(Sanyo) Nht


32/7223

T m CO2 (Sanyo) Nht

T lnh su c

T lnh LG Hn Quc

Thit b ng kh c

My li tm c

Ni hp tit khun ALP -Nht

Cn phn tch (Satorious) c

Cn k thut(Satorious) c

My o hm mPressica Thy in

My khuy tWisd Hn Quc

My lc (Vortex) Trung Quc

Hthng thha tan ERWEKA DT 60 c

a petri, ng nghim, u cn,

2.2. P pp

2.2.1.P

Cn, ong cc thnh phn trong cng thc mi trng MRS lng (cng

thc nu trong mc 2.1.2). Ha tan ht cc thnh phn. Phn chia mi trng

vo cc ng nghim sch bng phu thy tinh, mi ng nghim 10ml MRS

lng. y kn bng nt bng. Hp tit khun mi trng 115

o

C / 20 phtrong ni hp tit khun. Sau khi tit khun ngui xung khong 37

40oC. Cy ging vo ng nghim trn (thao tc cy phi c thc hin trong

t cy v trng). cc ng nghim c cy ging VSV trong t m 5%

CO2 trong 24h 37 1oC [9].


33/7224

2.2.2.P y

Cn, ong cc thnh phn trong cng thc mi trng MRS lng (theo

cng thc nu trong mc 2.1.2). Ha tan cc thnh phn trong cng thc trongcc c m. Chuyn 100ml mi trng MRS lng vo bnh nn dung tch thch

hp. y kn ming bnh bng nt bng.Hp tit khun 115oC /20ph trong

ni hp t ng.Ly ra, ngui xung khong 37 40 oC. Cy ging vo cc

bnh nn trn trong t cy v trng, mi bnh 1 ng ging 10ml. trong t

m CO25% trong 24h nhit 371oC.

2.2.3.P Nui cy VSV trong cc bnh nn cha mi trng MRS iu kin

37oC trong 24h trong t m CO25% (nh phng php nu trong mc 2.3.3).

ng nht dch nui cy bng cch lc bnh cha dch nui cy v khuy t

trong 10 pht, ong vo cc ng li tm mi ng 10ml hn dch sinh khi

(HDSK) trn, li tm cc ng ny vi tc 4000 vng/ph. Ly cc ng li tm

khi my, loi b phn dch trong, gi li phn cn sinh khi pha di, cho t

dc n chun b sn v c hp tit khun, ngui, lc trn my lc

(Vortex) cho sinh khi thu c phn tn u trong dung dch t dc n.

hn dch cc mu ny ra a petri c lm sch v hp tit khun t

trc, y kn bng giy nhm, trong t lnh su -80oC cho ng rn hon

ton, sau tin hnh giai on lm kh trong my ng kh. To cc mu

ng kh vi thnh phn nh sau:

Mu 1: SK + 10 ml nc ct.

Mu 2: SK + 10 ml dch li tm

Mu 3: SK + 10 ml sa gy 10%.

Mu 4: SK + 10 ml alginate 2%.

Tin ng: Cc mu c lm ng lnh trong t lnh su -80oC cho

tikhi mu ng rn hon ton.Tin ng trong thi gian 24h.


34/7225

Lm kh: Cc mu c lm kh trong khng gian lnh ca my ng

kh nhit khong -54oC, 0,055mbar. Lm kh trong thi gian 24h.

Kt thc qu trnh lm kh, ly mu ra khi thit b chuyn vo baopolyme c kha, xc nh s vi sinh vt sng st hoc y kn ri bo qun

trong t lnh nhit 4oC.

2.2.4.P pp k Tydll

Cho nguyn liu cn tit khun vo trong cc c m, y kn. cc

70 80oC trong 1h. Ly ra, cc ng nguyn liu vo t m 37oC trong

24h. Sau 24h, ly cc ng nguyn liu ra, lp li nh trn 3 ln.

2.2.5.P pp o b L. acidophilus

Chun b t dc: Tinh bt, Natri alginate c tit khun theo phng

php Tyndall

To thuc bt cha L. acidophilus: Tin hnh trong t cy v trng,

trn t dc v bt ng kh theo t l 1:1 to thnh khi bt ng nht,

thu c bt thuc cha tinh bt v bt thuc cha Natri alginate.

2.2.6. P pp o nang cng cha L . acidophilus

Chun b cc t dc: Tinh bt, Natri alginat c tit khun theo

phng php Tyndall

To nang cng cha L. acidophilus: Sau khi c hp tit trng, t

dc v bt ng khtrn ng lng to thnh khi ng nht. Sau em

ng thnhnang cngbng my to nang th cng.

Bo qun mu trong l nt kn nhit phng.

2.2.7.P pp x m m

Tin hnh o trn thit b o hm m Prescisa.

Lm nh mu cn o hm m. Cho ln mt a ca thit b khong 1g

mu, dn u mu ra mt a, y np, chy my.Ch my gia nhit trong


35/7226

khong 5 pht hoc n khi my hin kt qu.c, ghi li s liu hin trn

mn hnh (a1). Hm m ca mu l hiu s ca (100- a1) %.

2.2.8.

P pp x s l VSV eo y c pha longlin

Cn, ong chnh xc cc thnh phn theo cng thc mi trng MRS

c (nu trong mc 2.1.2). Ha tan cc thnh phn tan trong nc, phn tn

thch vo dung dch ng trong bnh nn thch hp. y kn bng nt bng.

Hp tit khun 115oC/ 20 pht trong ni hp tit khun. Ly bnh ng mi

trng ra khi ni hp, phn phi mi trng ln cc a petri c ra

sch, hp tit khun v sy kh (b dy lp thch trn cc a khong 2mm)

sao cho mt thch nhn, phng. Ch cho thch ngui v ng rn li, y kn.

Chun b cc ng nghim sch mi ng cha 9ml nc ct, y kn,

hp tit khun 115oC / 20 pht, ngui xung nhit khong 37 40oC.

Xc nh s lng VSV trong 1ml dch nui cy ban u.

Ht chnh xc 1ml hn dch sinh khi pha vo ng nghim th nht, lc

u. Ht chnh xc 1ml dch c ng nht trong ng nghim th nht

pha long sang ng nghim th 2 (cha 9ml nc ct c hp tit khun

115oC / 20 pht), lc u. C tip tc pha long nh vy cho ti nng

pha long cui cng (trong th nghim ny l 10-7, 10-8, 10-9). Cy tri ln mi

a petri 0,5ml dch trong cc ng phalong trn (lm trn 3 nng cui,

mi nng cy trn 2 a).

Mu xc nh s lng VSV cn sng sau ng kh

Lm nh nguyn liu, cn chnh xc khi lng ca mu, sau pha

vo bnh nn cha 100ml nc ct c hp tit khun 115 oC / 20 pht.

Phn tn u mu trong H2O bng khuy t, sau ht chnh xc 1ml dch t

bnh nn pha vo ng nghim cha 9ml nc ct, ri tip tc pha long, cy

nh i vi mu l cht lng nh trn.


36/7227

Mu xc nhs lngVSVsng st trong iu kin mi trng pHthp:

+ M b:Cn chnh xc khong 1,0g mu cn th, cho vo bnh nn

cha 100ml acid HCl pH 1,2 (pha theo Ph lc 2) c hp tit khun 115oC / 20 pht. Khuy t vi tc 50 vng/ph trong 1h, 2h. Ly mu ti

cc thi im 0h, 1h, 2h, tin hnh pha long lin tc nh cc bc nu

trn n (nng pha long 10-4, 10-5; 10-6; 10-7), cy tri dch trong 3 nng

pha long cui cng ln mt thch MRS, mi a 0,5ml, dn u dch trn

b mt thch bng que chang c tit khun.

+ M :Ti cc thi im 15ph, 30ph, 1h, 2h, ht ly 1 ml dch

mu th, pha long lin tc n nng 10-3, 10-4, sau cy tri dch trong 3

nng pha long cui cng ln mt thch MRS, mi a 0,5 ml, dn u

dch ln trn b mt thch bng que chang c tit khun

cc a ny trong t CO2 5%, nhit 37oC.

Tin hnh xc nh s lng VSV trn cc mu:

- 1ml dch nui cy sau 24h.

- Mu to thnh sau ng kh.

- Mu sau khi trong dch acid pH 1,2.

Sau 48h c kt qu

S lng VSV c trong 1g nguyn liu hoc 1 ml dch nui cy ban u

c tnh theo cng thc:

Trong :

m: khi lng mu em tin hnh xc nh s lng.

X: s VSV c trong 1 g hoc 1 ml mu.

An: S khun lc mc trn cc a petri c cy cc nng pha long n.


37/7228

2.2.9.P pp ng ca alginate n kh sng

st ca L . acidophiluso q thuc bt

Nui cy vi sinh vt trong bnh nn cha 200ml mi trng MRS lngtrong 24h (theo phng php nu trong mc 2.2.2).

Thu gn b mt phn dch trong, gi li khong 60ml hn dch sinh

khi pha y, to mu tin ng gm: hn dch sinh khi trnng lng

vi hn dchsa gy 20%.

ng kh cc mu (theo phng php nu trong mc 2.2.3)

m s lng vi sinh vt c trong 1g nguyn liubt ng kh theophng php pha long nu trong mc 2.2.8

Cho tng mu vi khi lng 1g vo bnh nn cha 100ml dung dch

HCl pH 1,2 . Khuy t vi tc 50 vng/ pht. Cc mu c thnh phn nh

sau:

Mu a: Bt ng kh L. acidophilus vi hn dch sa gy10% (Bt

ng kh).

Mu b: Bt ng kh + tinh bt.

Mu c: Bt ng kh + alginate.

Trong cc mu bt ng kh phi t tiu chun trn v hm m,

th cht v s lng VSV.

Sau 1h, ng nht ha mu trong bnh. Nhanh chng ht chnh xc 1 ml

dch trong bnh tin hnh pha long lin tc theo mc 2.2.8.

trong t CO25%, sau 48h tin hnh m s lng khun lc. Tnh s

lng vi sinh vt cn sng st (theo cng thc nu trong phn 2.2.8.)

2.2.10. P pp ng ca alginate n kh

sng st ca L. acidophilustrong qu trnh cng

Sdng my thha tan ERWEKA DT60 nh gi slngL.

acidophiluscn sng sau khi thtrong iu kin mi trng acid pH 1,2


38/7229

iu kin th:

- Thit b: giquay.

-Tc quay: 501 vng/pht.

- Mi trng th: 200 ml mi trng pH 1,2.

- Nhit mi trng th: 37 0,5oC.

- Thi gian ly mu: 15 ph, 30ph, 1h, 2h.

Tin hnh:

To bt ng kh t tiu chun v hm m, th cht v s lng VSV.

Phn tn nh bt ng kh. embt ng khng nang vi t dc

alginate v tinh bt vi t l 1:1. Cc mu c thnh phn nh sau:

Mu 1: Bt ng kh + tinh bt.

Mu 2: Bt ng kh + alginate.

em mi nang cho vo trong mt cc th ha tan n mi trng

acid pH 1,2 t 370C. Sau tin hnh khuy vi tc khuy 50 vng/pht.

Ti cc thi im 15 pht, 30 pht, 1h, 2h, ht 1 ml dch v tin hnh m

VSV theo phng php pha long lin tc mc 2.2.8.


39/7230

C 3. THC NGHIM, KT QUV BN LUN

3.1. Nghin cu ng ca alginate n kh sng st ca vi

khun L. acidophiluso q k

Trong nhng nm gn y alginate c nghin cu s dng kh nhiu

trong cc ch phm probiotics v cho thy kh nng bo v vi sinh vt khi

nhiu iu kin bt li [15], [41], [42]. Cc nghin cu di y c tinhnh nhmnh gi tc dng ca alginat trong qu trnh ng kh vi sinh vt

v so snh tc dng ca n vi mt nguyn liu c dng ph bin trong

ng kh l sa gy.

Trong sn xut, bt c nguyn liu no cng cn phi c tiu chun

lm mc nh gi cht lng, gip sn phm cht lng tt. i vi nguyn

liu probiotics, theo tiu chun ca IDF, s lng VSV trong 1 gam ch phmt 108-109 cfu/g. Mt khc, i vi nguyn liu ng kh, hm m l mt

tiu chun quan trng, v hm m cao lm cho nguyn liu d hp th nc

tr li [11], kh m bo t l VSV sng cao. Mt tiu chna cn quan tm

l ti xp ca nguyn liu, ti xp ln s gip nguyn liu d phn tn,

ng u hm lng s tt hn. Do vy, nguyn liu sau khi ng kh cn

t c tiu chun nu ln di y:

Th cht ca nguyn liu to thnh sau khi ng kh: ti xp

Hm m sau ng kh vi natri alginate: 108cfu/g


40/7231

Cc nghin cu di y nhm nh gi tc dng ca alginate trong

qu trnh ng kh v so snh vi alginate nhng u da trn mc tiu t

c tiu chun trn.3.1.1. y

Mt trong nhng vai tr quan trng ca t dc s dng trong qu

trnh ng kh l gi cho ch phm cth cht n nh, trnh b ht mtrong

qu trnh bo qun. Mt khc, y l ch phm probiotics nn t dc cn c

thm mt vai tr quan trng na l bo v VSV, gi cho VSV c t l sng

st cao sau khi ng kh. Theo cc nghin cu cng b th sa gy 10%

va bo v VSV tt, va gi cho ch phm c th cht n nh [32], [33]. Do

vy, trong th nghim ny, mt trong nhng t dc dng so snh vi

alginate l sa gy 10%, v ch tiu so snh l s n nh ca th cht mu

sau khi ng kh.

Trong cc nghin cu to ch phm vi nang hay bt phun sy cha

probiotics, t dc natri alginateu c s dng vi nng 2%[15], [21].

Do vy, t dc alginate c kho st trong nghin cu nycng c s

dngvi nng 2%.

Mc tiu:

Kho st th cht ca bt ng kh cha VSV vi t dc alginate

nng 2%, ng thi so snh vi sa gy 10% v dch ly tm.

Kt qu:

Th chtca cc mu thu c c th hin trong bng 3.1.

B 3: Th cht ca cc mu ng kh L. acidophilus vi cc t dc bo

v ti thi im ngay sau khi ng kh

T d T


41/7232

bo

Nc ct Sn phm thu c l 1 lp rt mng, dnh a. Rt kh

ly ra ht khi dng c cha.Dch li

tm

Sn phm thu c l 1 lp rt mng, dnh a. Rt kh

ly ra ht khi dng ccha

Sa gy

10%

Th cht ng nht, kh, xp, d lm ti, d dng ly ra khi

dng c cha

Alginate

2%

Th cht ng nht, nh, xp, kt thnh mng, rt kh lm nh.

D ly ra khi dng c cha nu thc hin thao tc ny trckhi mu b ht m

Nhn xt:

Khi khng dng t dc n, cc mu ng kh L. acidophilusc

to thnh l mtlp mng bm vo dng c cha, kh ly ra, ng thi cc

mu ng kh ny cng nhanh chng ht m tr li.

Hnh 3.1: Hnh nh mu ng kh vi nc ct.


42/7233

Cc mu ng khL. acidophilusvi t dc l hn dch sa gy10%

v dung dch alginate 2% u cho th cht c xp ln. Tuy nhin, mu

ng kh vi alginate c th chtxp hn nhng li bkt thnh mng, nhanhb ht m v kh phn tn nh.

Tm li, s dng dung dch alginate lm t dc trong ng kh gip

thu c mt mu nguyn liu c th cht xp, d dng ly ra khi dng c

cha hn so vi cc mu khng s dng t dc. Tuy nhin s dng alginate

lm t dc trong ng kh s to ra nguyn liu nhanh b ht m, kh phn

tn nh thnh dng bt mn, vic ny dn n kh ng nht gias lng

VSV probiotics cc l m trong qu trnh bo ch to sn phm tip theo.

3.1.2. m m m

y alginate

Hm m l mt ch sgy nh hng ln n kh nng sng st ca vi

sinh vt trong qu trnh bo qun. Hm m ln l nguyn nhn gy gim s

lng vi sinh vt sng st. Trong phng php ng kh, tiu chunhm m

ca cc mu ngay sau ng kh l khng nh hn 5%.V vy, cc th nghim

di y c thc hin nhm nh gi hm m ca cc mu cc thi im

ngay sau ng kh, v trong qu trnh bo quntrong ti polymer kn nhit

4oC.

Tuy nhin, khi thc hin nh gi th cht v t l VSV sng st cc

mu mc 3.1.1 nhn thy: th cht ca cc mu s dng nc (mu 1) v

dch ln men (mu 2) khng thch hp to nguyn liu nn vic theo di

hm m ca mu 1 v mu 2 theo thi gian khng cn nhiu ngha. Do

chng ti thc hin th nghim nh gi hm m v tc ht m theo thi

gian ca2 mu c th cht t ti xp, lmu ng kh vi sa gy10%

v mu ng kh vi alginate 2%.


43/7234

Mc tiu:Xc nh hm m ca cc mu ngay sau khi ng khv tc

ht m ca cc mu trongthi gian bo qun.

Kt qu: Hm m ca cc mu c th hin trong bng 3.2.

B 3.Bin thinhm m (%) ca mt s mu ng kh L. acidophilus

vi t dc bo v khc nhau theo thi gian

T bo qHm m (%)

M s y

10%

M

natri alginate 2%

Ny s k 2,8 + 0,3 6,5 + 1,6

Sau 2 ngy 3,4 + 0,7 10,8 + 2,9

S 1 3,5 + 0,7 12,5 + 3,3

S 2 3,5 + 0,6 13,3 + 3,0

To : Mu cha sa gy 10%: mu 3 trong mc 3.1.1


44/7235

Mu cha alginate2%: mu 4 trong mc 3.1.1

Hnh 3.2: th biu th hm m trung bnh cacc mu ngay sau ng kh

v trong thi gian bo qun.

Nhn xt:

Kt qu a ra bng 3.2cho thy hm m ca mu ng kh vi sa

gy 10% ngay sau ng kh l 2,8% t tiu chun hm m ca cc ch phm

thuc bt (nh hn 5%). Sau 2 tun bo qun, hm m ca mu ng kh vi

sa gythay i khng ng k v kt qu o hm m ca mu vn t gi tr

nh hn 5%. Trong khi , mu ng khL. acidophilusvi alginate 2% li

c hm m kh cao v bin i kh ln gia cc mu khc nhau khi ng kh

trong cng mt iu kin, cng thi gian nh nhau, hm m thu c ca mu

4 l 6,5% ngay sau ng kh.

ng thi, tc ht m ca mu ng kh vi sa gy cng chm

hn so vi mu ng kh vi alginate. Sau 2 tun bo qun, hm m ca mu

ng kh s dng alginate 2% c hm m kh cao, t 13,3+ 3,0, trong khi

mu ng kh vi sa gy vn t gi tr nh hn 5%.

Hnh 3.3.Hnh nh mu ng kh vi alginate ngay sau khi tho khi my

v sau 3 pht ngoi mi trng.


45/7236

Trong qu trnh tin hnh th nghim, khi tho sn phm ra khi my

ng kh nu khng nhanh chng bo qun trong bao b kn th nguyn liu

ng kh vi alginate s ht m rt nhanh v d b bt dnh vo dng c chahoc chy nc.

Tm li, ngay sau ng kh v trong qu trnhbo qun, mu ng kh

viNatri alginate 2% u c hm m v tc ht m cao hn so vi mu

ng kh vi sa gy 10%. Khi s dng t dc bov l sa gy, hm m

ca mu ng kh kh n nh theo thi gian, trong khi hm m ca mu

ng kh vi alginate li thay i kh nhiu, mi ch sau 2 tun bo qun

trong bao polymer kn hm m ca mu ln ti khong 13%. Nh vy mu

ng kh vi alginate 2% kh c th m bo yu cu v hm m.

3.1.3.K

nh gi tnh kh thi ca vic s dng ca bt k t dc no lm t

dc bo v trong ng kh Lactobacillus acidophilus l nh gi s lng

v t l vi sinh vt sng st sau khi ng kh. c coi l c tc dng bo

v trongng kh th trc tin t dc phi cho thy kh nng ci thin

c s lng cng nh t l vi sinh vt sng st trong qu trnh ng kh, t

nht l sovi cc mu ng kh khng s dng t dc bo v. V vy , th

nghim tip theo c tin hnh vi tiu ch nhm xc nh s lng v t l

vi sinh vt sng st ca cc mus dng t dc alginate sau qu trnh ng

kh.

Mc ch

nh gi nh hng cadung dch alginate 2% ln s lng v t l

sng st ca vi sinh vt trong qu trnh ng kh, ng thiso snh vi mu

ng kh khng s dng t dc v mu ng kh s dng hn dch sa

gy10%.


46/7237

Kt qu v nhn xt

S lng vi sinh vt Lactobacillus acidophilusc trong 1ml dch nui

cy trc ng kh l khong 1,09 x 10

9

VSV/ 1ml.

B 33S lng vi khun sng st trong 4 mu sau ng kh

(Tnh theo khi lng tng ng 1ml dch nui cy ban u)

S l s M 1

()

M 2

(dh ln men)

M 3

(s y 10%)

M 4

(alginate 2%)

Ln 1 2,9 x 106 7,9 x 105 5,1 x 108 1,7 x 107

Ln 2 2,5 x 106 8,5 x 105 7,1 x 108 2,2 x 107

Ln 3 2,3 x 106 7,7 x 105 4,6 x 108 2,3 x 107

Trung

bnh2,57 x 106 8,03 x 105 5,6 x 108 2,07 x 107

RSD 0,31 x 106 0,42 x 105 1,32 x 108 0,32 x 107

B 34:T l VSV sng st sau ng kh so vi mu ban u

M 1

()

M 2

(d l me)

M 3

(s y 10%)

M 4

(alginate 2%)

T l VSV

sng st0,24% 0,07% 51,38% 1,9%


47/7238

Trong : Mu 1: ng khL. acidophilusvi dch li tm.

Mu 2: ng khL. acidophilusvi nc ct.

Mu 3: ng khL. acidophilusvi sa gy10%.

Mu 4: ng khL. acidophilusvi alginate 2%.

Hnh 3.4. th biu din s lng vi khun L. acidophilus sngst

trong cc mu sau ng kh

Kt qu trongbng 3.3 cho thy ng kh vi t dc l dung dch

alginate 2% cho t l vi sinh vt sng st thp hn nhiu so vi t dcdng

ph bin l sa gy10%. Tuy vy, s liu cng cho thyalginate 2% c tc

dng bo v khi s dng lm t dc ng kh vi t l sng st ca mu ny

cao gp 27,1ln so vi mu ng kh khng c t dc bo v.

Nh vy, alginate c tc dng bo v Lactobacillus acidophilus trong

qu trnh ng kh. Khi ngkh vi t dc l alginate 2% cho t l sng st

caL. acidophilus l 1,90% cao gp 27,1 ln so vi mu chng ng kh vi

nc ct, v cao gp 7,91 ln so vi mu ng kh vi dch li tm. Tuy nhin


48/7239

t l sng ca L. acidophilus trong mu ng kh vi alginate cn km sa

gy nhng vn t s lng vi sinh vt l 107.

3.1.4.

S y alginate y

T kt qu th nghim 3.1.3 cho thy alginate c tc dng bo v vi

sinh vt trong qu trnh ng kh, tuy nhin nu s dng mt mnh lm t

dc ng kh th tc ht m ca nguyn liu to thnh ln lm cho

nguyn liu kh m bo yu cu v hm m. Mt khc nguyn liu to thnh

sau khi ng khL. acidophilus vi alginate 2% cho t l vi sinh vt sng st

thp hn so vi khi ng kh vi sa gy 10%. M tiu chun s lng VSV

sng st l tiu chun quan trng nht ca ch phm probiotics nn trong qu

trnh ng kh phi kt hp vi sa gy 10% t c tiu chun trn.Do

chng ti thit kth nghim tip theo: ng kh vi sinh vt s dng kt

hp t dc l dung dch alginate cc nng 1%, 2%, 5% v sa gy 10%

nhm ci thin th cht cng nh t l sng st VSV ca mu ng kh vi

alginate.

Mc tiu:

S b so snh t l sng st cavi sinh vt ca mu s dng tdc

n l sa gy 10% v mu s dng sa gy c kt hp thm dung dch

alginate.

Kt qu v nhn xt

V th cht

C 4 mu u to ra bt nguyn liu c th cht ng nht, ti xp,

kh, mu m dn theo nng alginate c trong mu. Nh vy vic thay

i nng alginate t 1 5% khng lm nh hng n th cht ca cc

mu.

Hm m


49/7240

B 35:Hm m ca cc mu ng kh vi t dc bo v l hn hp sa

gy vi alginate cc nng

T ianHm m (%)

0% alginate 1% alginate 2% alginate 5% alginate

0h 2,8 + 0,3 3,1 + 0,4 3,6 + 0,5 3,7 + 0,5

Sau 2 ngy 3,4 + 0,7 3,4 + 0,5 4,0 + 0,6 4,1 + 0,6

Sau 1 tun 3,5 + 0,7 3,7 + 0,6 4,3 + 0,6 4,6 + 0,6

Sau 2 tun 3,6 + 0,7 4,0 + 0,7 4,4 + 0,6 4,7 + 0,7

Hnh 3.5:Hm m ca cc mu ng kh vi t dc bo v l hn hp sa

gy vi alginate cc nng

Hm m ca cc mu ngay sau ng kh c kt hp sa gy 20% v

alginate 1%, 2%, 5% t t: 2,8% 3,7%, gi tr ny gn tng ng vi

hm m ca mu ng kh vi sinh vt vi t dc l sa gy 10% (hm m

2,8%), v t tiu chun hm m ca dng thuc bt [2]. Hm m sau thi


50/7241

gian bo qun trong ti polyme kn ca cc mu ny vo khong 3,4 4,7%

trong khi mu ng kh vi sa gy 10% c hm m 3,4 3,6%. Nh vy xt

v th cht v hm m th khi kt hp alginate v sa gy 10% th nguyn liuto thnh c th cht v hm m tng ng vi mu ng kh ch s dng

sa gy, nhng t l alginate cng cao th ch phm c mu cng m v hm

m cng ln hn so vi mu c hm lng alginate thp (tuy nhin s thay

i v hm m gia 3 mu ny l khng ng k).

V s lng vi sinh vt Lactobacillus acidophilussng stsau ng kh

S lng vi sinh vt sng st c nu trong bng 3.6 cho thy tuy mu

kt hp alginate 2% vi hn dch sa gy10% cho t l sng cao nht 26,1 x

108 cfu/g so vi 20,2 x 108 cfu/g ca mu ng kh vi sa gy v 21,2 x

108cfu/g; 24,3 x 108cfu/g ca 2 mu cha 1% v 5% alginate. Tuy nhin s

lng sng st trong mung kh khng c s khc nhau nhiu (c 4 mu

u cha khong 109VSV/1g).

S lng VSV sng st c th hin trong bng sau:

B 36. S lng vi sinh vt sng st tnh trn 1g bt sau ng kh trong

cc mu ng kh s dng kt hp alginate v sa gy.

S l s ( x108 )

0%

alginate

1%

alginate

2%

alginate

5%

alginate

Ln 1 8,51

14,50 29,40 23,92Ln 2 21,89 9,05 34,52 27,84

Ln 3 20,42 29,91 4,90 1,29

Trung

bnh20,2 21,2 26,1 24,3


51/7242

Hnh 3.6:Slng vi sinh vt sng st tnh trn 1g bt sau ng kh trong

cc mu ng kh s dng kt hp alginate v sa gy.

Nh vy khi kt hp vi sa gy 10% ng kh, natri alginate th

hin rt t kh nng gia tng t l sng st trn Lactobacillus acidophilus.

Nguyn nhn c th do t l natri alginate s dng kh thp, hn na nh thnghim 3.1.3 ta thy tc dng bo v vi sinh vt ca alginate nh hn kh

nhiu so vi sa gy (khong 27ln). Do vy khi kt hp alginate vi sa gy

to hn hp t dc khi ng kh ta s khng thy c s thayi ln trn

t l sng st. Mt khc,s kt hp t dc s lm cho qu trnh bo ch tr

ln phc tp hn, nguyn liu to thnh vi cc mu c nng alginate cao

c mu sc khng p, ht m cng cao hn tuy khng nhiu.

Tuy nhin v c th cht, hm m, cng nh s lng vi sinh vt sng

st trong mu phi hp alginate vi sa gy u t gi tr tt hn so vi mu

ng kh s dng ring alginate.


khun Lactobacil lus acidophil uso q thuc b

nang cng


52/7243

Cc kt qu nghin cu nu trong mc 3.1.2cho thy alginate c tc

dng lm gia tng t l sng st ca vi sinh vt trong qu trnh ng kh. Tuy

nhin vic s dng n nh mt t dc bo v trong ng kh vi sinh vt cnnhiu hn ch v sn phm to thnh c th cht v hm m khng thch hp.

Do vy ccth nghimtip theo c thit knhm kho st kh nng bo v

ca natri alginate khi s dng lm t dc n trong cc ch phm cha vi

sinh vt. Mt trong nhng phng php th tc dng bo v ph bin nht

hin nay l th tc dng bo v ca cc t dc s dng ln t l sng st ca

vi sinh vt khi tip xc vi iu kin acid d dy. Th nghim ny tin hnh

th nghim trn cc mu: nguyn liu ng kh vi sa gy 10% v khi c

tin hnh trn ng lng bt ng kh ny vi t dc n l natri alginate,

tinh bt - mt t dc n c dng ph bin hin nay c s dng i

chng.

3.2.1.N alginate k s s

kL. acidophiluso q b

Mc tiu:

Nghin cu nh hng caalginate n kh nng sng st ca vikhun

L. acidophilusc s dng lm nguyn liu thuc bt

Kt qu, nhn xt

Mu s dng alginate lm t dc n cho t l vi sinh vt sng st sau

1h v 2h cao hn khong 37ln so vi khi s dng t dc n l tinh bt v

mu khng s dng t dc l mu bt ng kh. Mu s dng alginate cho

s lng VSV sng st vo khong 107, trong khi mu s dng tinh btv

mukhng s dng t dc (mubt ng kh)cho s lng VSV sng st

khong 105 n 106.


53/7244

B 37. S lng vi sinh vt sng st tnh trn 1g bt ng kh

trong mi trng acid HCl pH 1,2

S l s

B Sau 1h Sau 2h

M M b M M M b M

L 1 1,0 x 109 9,8 x 106 6,6 x 105 6,3x 105 4,1x 106 2,0x 105 1,9 x 105

L 2 1,9 x 109 2,2 x 107 3,1 x 105 2,9x 105 1,3x 107 2,6x 105 2,6 x 105

L 3 2,2 x 109 2,4 x 107 4,6 x 105 4,7x 105 1,6x 107 4,0x 105 4,3 x 105

SLTB 1,7 x 109 1,9 x 107 4,8 x 105 4,6x 105 1,1x 107 2,9x 105 2,9 x 105

SD 0,6 x 109 5,6 x 107 1,7 x 105 1,7 x 105 2,2 x 107 1,0 x 105 1,2 x 105

Trong : Mu a: Bt ng khL. acidophilusvi hn dch sa gy10%.

Mu b: Bt ng kh + tinh bt.

Mu c: Bt ng kh + alginate.

Hnh 3.7. Biu th hin s lng vi sinh vt sng st sau khi th trong mi

trng acid HCl pH 1,2


54/7245

Hai mu b v a cho trong cc ln cng th nghim th cho s lng

VSV sng st ngang nhau, t 4,8x105cfu/g sau 1h v 2,9x105sau 2h. iu

ny chng t vic b sung thm t dc n l tinh bt khng h gip cithin s lng VSV sng st trong iu kin acid d dy.

Tm li, kt qu th nghim cho thy mu nguyn liu s dng Natri

alginate lm t dc n cho t l sng st trong mi trng acid pH 1,2 cao

hn so vi mu th vi bt ng kh v mu bt ng kh c s dng thm

t dc n l tinh bt. T c th s b kt lun natri alginate khi s dng

lm t dc n trong ch phm vi sinh vi t l cao (trong th nghim ny l

50%) cho hiu qu lm gia tng t l sng st caL. acidophilus ATCC 4653.


kL. acidophiluso q o

Mc tiu:Nghin cu nh hng ca alginate n kh nng sng st

ca vi khunL. acidophilus trong qu trnh tonang cng

Kt qu:

B 3.8: S lng vi sinh vt sng st trong 1 vin nang trong mi trng

acid HCl pH 1,2 vi tc khuy 50 v/pht

S l s

B Sau 15 pht Sau 30 pht Sau 1h Sau 2h

M 1 M 2 M 1 M 2 M 1 M 2 M 1 M

L 1 1,5x108

1,2x104

1,5x106

1,0x103

1,1x105

1,0x102

3,0x104

- -

L 2 2,7x108 1,6x104 1,9x106 1,3x103 1,5x105 1,5x102 3,9x104 - -

L 3 3,3x108 1,8x104 2,1x106 1,6x103 1,8x105 1,9x102 4,1x104 - -

SLTB 2,5x108 1,5x104 1,8x106 1,3x103 1,5x105 1,5x102 3,7x104 - -

SD 0,9x108 0,3x104 0,3x106 0,3x103 0,4x105 0,5x102 0,6x104 - -

Ch thch:(-):khng thy xut hinkhun lc


55/7246

Bng 3.9:Tlvi sinh vt sng st ca mu alginate

so vi mu tinh bt trong mi trng pH 1,2

M 1 M 2

T l VSV sng sau 15 pht 1 120

T l VSV sng sau 30 pht 1 115

T l VSV sng sau 1h 1 240

T l VSV sng sau 2h - -

Hnh 3.8:S lng vi sinh vt sng st trong 1 vin nang trong mi


Nhn xt:

Kt qu bngcho thy mu nang c s dng alginate lm t dc n

cho t l sng st VSV trong mi trng acid pH 1,2 ln hn so vi mu

nang c s dng tinh bt lm t dc n.Trong th nghimny, t l VSV

sng st ca mu s dng alginate cao gp hn100 lnso vi mu s dng

tinh bt.Ngay sau 15 pht, s lng VSV sng st ca mu nang cha tinh


56/7247

bt gim mnh xung 1,5x104 cfu/nang, trong khi mu nang cha

alginate vn gi c 1,8x106 cfu/nang. Sau 1,5 gi, s lng VSV sng st

trong mu nang alginate gim xung cn 3,7x10

4

, trong khi mu nang tinhbt s lng VSV l 1,5x102. Nh vy, i vi mu nang tinh bt, lng

VSV sng st b gim mnh ngay trong 15 pht u, cn nang alginate lng

VSV sng st b gim mnh sau 1,5 gi.

Tuy nhin, sau 2 gi trong mi trng acid pH 1,2 hu nh khng cn

VSV sng st. Do vy, cn tm thm cc bin phpci thin khc, lm gia

tng t l sng st ca VSV trong mi trng acid thp.


57/7248

C 4: BN LUN

4.1.

Nghin cu ng ca alginate n kh sng st ca vikhun L. acidophiluso q k

4.1.1. c ngy

Khi nh gi th cht ca nguyn liu to thnh sau khi ng kh, mu

ng kh vi dch sa gy 10% v mu ng kh vi dung dch alginate 2%

c th cht xp hn so vi mu ng kh vi nc ct v dch ly tm. Tuy

nhin, mu ng kh vi alginate li b kt thnh mng, kh lm nh. Vic

mu ng kh vi dung dch natri alginate to thnh dng nguyn liu kh

nghin nh thnh dng bt mn c th c gii thch nh sau: alginate thc

cht l mt heteropolyme saccharid mch thng cu to t hai gc uronic l

acid - L- guluronic (G) v acid - D- mannuronic (M) [43]. Chnh cu trc

chui polymer ca alginate lm cho nguyn liu to thnh sau ng kh c

cu trc dng cc mng. Trong qu trnhhp tit khun do tc ng ca nhit

cao (> 70oC) nn cc chui polymer ny mt phn b polymer ha lm

gim nht ca dung dch polymer trc khi kt hp vi sinh khi v cng

l l do cc mng nguyn liu ng kh to thnh khi ng kh vi alginate

khng c cu trc mng di m ch l cc cu trc dng mng xp, ngn (vi

mm). V chnh cu trc dng chui polymer ca alginate khin cho nguyn

liu to thnh kh c th c lm nh thnh dng bt mn.

Khc vi vic s dng alginate trong ng kh, khi s dng alginate

lm t dc vi nang ha, sn phm probiotics c to ra c th cht tt

hn. C th, trong th nghim ca Asma Sohail v cng s [15], vi nang cha

probiotics c to thnh bi alginate 2% v CaCl2bngphng php phun

sy c kch thc phn b ng u t 10 40 m v hnh dng trn u p.


58/7249

Tm li, s dng dung dch alginate lm t dc trong ng kh gip

thu c mt mu nguyn liu c th cht xp, d dng ly ra khi dng c

cha hn so vi cc mu khng s dng t dc. Tuy nhin s dng alginatelm t dc trong ng kh s to ra nguyn liu nhanh b ht m, kh phn

tn nh thnh dng bt mn, vic ny dn n kh ng nht gias lng

VSV probiotics cc l m trong qu trnh bo ch to sn phm tip theo.

4.1.2. m m m

y alginate

Hm m v tc ht m ca mu ng kh vi alginate 2% ngay sau

ng kh v trong thi gian bo qun 2 tun u ln hn so vi mu ng

kh vi sa gy 10%, c bit ch trong 2 tun bo qun hm m ca mu

ng kh vi alginate 2% ln ti khong 13%, trong khi hm m ca

mu ng kh vi sa gy 10% vn t tiu chun nh hn 5% . Trong hnh

3.3 cho thy mu ng vi alginate ht m rt nhanh, ngay sau 3 pht

chuyn t dng ti xp sang dng do. iu ny c th gii thch l do natri

alginate l mui ca acid alginic, alginic l acid c cu to bi cc gc

uronic. Gc uronic cha nhiu nhm phn cc, a nc [43], do vy tc

ht m ca alginate ngoi mi trng rt nhanh. Hm m 13% sau thi gian

bo qun 2 tun l qu ln, khng m bo c tiu chun ca nguyn liu,

do vy khng s dng ring alginate 2% lm t dc trong qu trnh ng kh

c, c th kt hp b sungvi sa gy hoc cc t dc khc.

V tiu ch hm m nguyn liu, sa gy vn t. Do vy, sa gy l

mt trong nhng la chn u tin lm t dc bo v VSV trong khi ng

kh. Cn theo nghin cu ca Lim Chi Ming, sa gy c tc dng bo v

VSV trong qu trnh ng kh l do protein trong sa gy to thnh lp o

bo v trn thnh t bo v cung cp cht m gip n nh pH trong sut qu

trnh ng kh [32]. Trn thc t, trong cc ch phm ng kh, sa gy


59/7250

thng c s dng km thm vi cc nhm t dc khc nh carbohydrat,

polymer, polyols, aminoacid, tng cng tc dng bo v VSV[17].

4.1.3.

K . Kt qu kho st t l sng st caL. acidophilussau ng kh cho thy

khi s dng cht bo v lhn dch sa gy10% cho s lng vi sinh vt sng

st cao nht (khong 108), tip theo l mu ng kh vi dung dch alginate

(khong 107), mu ng kh vi dch ln men (106), mu ng kh vi nc

ct (105). Kt qu ny tng t vi kt qu ca Lim Chi Minhv cng s

thc hin nm 2008 [32]. Theo kt qu cng b ca Lim Chi Minh, t l

sng st ca VSV sau ng kh ca sa gy (13,03%) cng cao hn so vi

nc ct (0,07%).c bit, theo Lim Chi Minh, t l sng stca VSV sau

khi ng kh vi nc (0,07%) tng ng vi kt qu nghin cu trn

chng vi sinh vt khc cng thuc chi Lactobacillus l Lactobacillus

salivarius vi nc(0,08%).

Tc dng bo v vi sinh vt trong qu trnh ng kh c th c gii

thch nh sau:

Natri alginate c cu trcpolyme nn s trng n trong nc, to ra

c xung quanh v bn trong t boVSV mt mi trng c nht tng

t gip gim thiu bin i cu trc phn t mc thp nht [32].

Vi sa gy c thnh phn ch yu gm lactose v protein chnh l

nhng yu t quyt nh kh nng bo v ca chng. Protein c trong sa to

ra mi trng c nht cao gip bo v cu trc t bo vi sinh vt, trong khi

thnh phn lactose s to lin kt hydro gia ng - protein, lin kt ny

gip n nh protein t bo, gip duy tr cu trc ca protein mng khi nc

b loi i. S n nh mng t bo v protein cu trc bng lin kt hydro ca

cc disaccharid cn c gi l l thuyt thay th nc [17]. Cn theo

nghin cu khc th protein trongsa gy c th to nn mt lp o bo v


60/7251

trn thnh t bo. Trong sa gy cn cha nhiu cht tan nh mui phosphat,

citrat c kh nng m gip n nh pH mu khi m dung mi mt dn trong

qu trnh ng kh [32].Nh vy, alginate c tc dng bo v Lactobacillus acidophilus trong

qu trnh ng kh. Khi ng kh vi t dc l alginate 2% cho t l sng st

caL. acidophilus l 1,90% cao gp 27,1 ln so vi mu chng ng kh vi

nc ct, v cao gp 7,91 ln so vi mu ng kh vi dch li tm. Tuy nh in

t l sng ca L. acidophilus trong mu ng kh vi alginate cn km sa

gy nhng vn t s lng vi sinh vt l 107.

4.1.4.S y

alginate y

Khi kt hp sa gyvi alginate vi t l tng dn n 5%, khng thy

c s gia tng v t l sng st ca VSV. Mt khc, vic kt hp ny lm cho

qu trnh bo ch phc tp hn nn vic kt hp vi alginate vi t l nh

trong th nghim ny l khng kh thi.

Dianawati Dianawati v cng scng thc hin nghin cunm 2012

[19] v phi hp cc t dc bo v trong qu trnh ng kh, lnh gi

tc dng bo vca vic phi hp 5 loi protein v 3 loi ng i vi B.

longum trong qu trnh ng kh. 5 loi protein c nghin cu l natri

caseinate 12%, whey protein 12%, natri caseinate:whey protein 6%:6%, sa

gy 12%, du u nnh 12% c phi hp ln lt vi 3 loi ng l

glycerol (3% kl/tt), mannitol (3% kl/tt), maltodextrin (3% kl/tt) to ra 15 dch

nh tng. LngB. longum v nh tng c a vo khi ng kh theo

t l 1:4. Kt qu cho thy nh tng phi hp gia whey protein natri

casenate v Glycerol gip VSV c t l sng st cao sau ng kh, t 99,2%.

Cn i vi nh tng phi hp gia natri casemate v manitol, t l VSV

sng st sau ng kh, sau qu trnh tip xc vi acid v sau qu trnh tip


61/7252

xc vi mui mt ln lt l 97,4%, 81,6% v 99,3%. Kt qu cui cng cho

thy sa gy c tc dng bo v vt tri so vi du u nnh v loi ng

c nhiu nhm OH bo v tt hn maltodextrin[19]. iu ny li khng nhmt ln na tc dng bo v VSV vt tri ca sa gy trong qu trnh ng

kh.


khun L. acidophilustrong qu trnh thuc b cng


k L. acidophiluso q o b

Theo kt qu nghin cu cho thy alginate c tc dng bo v VSV tt

hn so vi tinh bt trong mi trngpH 1,2. Tc dng bo v ca alginate

trn cc vi sinh vt trong iu kin acid pH thp ca d dy c th gii thch

nh sau: khi tip xc vi dung dch acid pH thp mui natri alginate chuyn

thnh dng acid alginic khng tan. Acid alginic trng n to ra mt lp

mng bao quanh bt ng kh cha vi sinh vt, cch li vi sinh vt vi iu

kin bt li ca mi trng xung quanh [30].

So snh kt qu th nghim ny vi kt qu trong nghin cu ca Latha

Sabikhi nm 2010 khi nh gi kh nng bo v L. acidophilus trong ch

phm probioticsbng phng php to vi nang vi alginate. Vi nguyn liu

ban u cha khong 109cfu/g, mu khng c bo v u c s lng sng

st sau 1h, 2h trong iu kin pH 1 gim xung cn khong 10 5 106 cfu/g

(gim khong 1000 ln) [31]. nghin cu ca Latha Sabikhi vi phng

php to vi nang bo v th sau 1h trong pH 1 s lng Lactobacillus

acidophilusLA1 gim t 107 cfu/g lc ban u xung cn khong 106cfu/g

(ngha l gim khong 10 ln), sau 2h trong iu kin acid pH 1, s lng

VSV ca mu bao vi nang cn 105cfu/g (gim 100 ln so vi ban u). Cn

trong th nghim c s dng alginate lm t dc n th sau 1h trong mi


62/7253

trng acid HCl pH 1,2 s lng vi sinh vt gim t 1,7 x 109cfu/g xung

cn khong 1,9 x 107cfu/g (tng ng khong xp x 100 ln), cn ti thi

im sau 2h lng vi sinh vt cn li l khong 1,1 x 10

7

cfu/g (gim khong155 ln so vi ban u)[31].

Tm li, kt qu th nghim cho thy mu nguyn liu s dng Natri

alginate lm t dc n chot l sng st trong mi trng acid pH 1,2 cao

hn so vi mu th vi bt ng kh v mu bt ng kh c s dng thm

t dc n l tinh bt. T c th s b kt lun natri alginate khi s dng

lm t dc n trong ch phm vi sinh vi t l cao (trong th nghim ny l

50%) cho hiu qu lm gia tng t l sng st caL. acidophilus ATCC 4653.


k L. acidophiluso q o

Kt qu th nghim cho thy t l VSV sng st trong mi trng acid

pH 1,2 ca mu nang cng cha alginate gp 40 ln so vi mu nang cng

cha tinh bt. iu ny c th gii thch nh sau: khi tip xc vi dung dch

acid pH thp mui natri alginate chuyn thnh dng acid alginic khng tan.

Acid alginic trng n to ra mt lp mng bao quanh bt ng kh cha vi

sinh vt, cch li vi sinh vt vi iu kin bt li ca mi trng xung quanh

[30]. Do vy, t l VSV sng st ca mu s dng alginate ln hn so vi

mu s dng tinh bt.

Mt khc, t l sng st gia 2 mu alginate v tinh bt trong mi

trng acid pH 1,2 nang cng ln hn so vi thuc bt. C th, i vi

thuc bt, t l ny l 40, trong khi i vi thuc nang cng t l ny l

120. Nguyn nhn l do acid alginic trng n to lp kt dnh vi v gelatin,

trong khi mu thuc bt khng c s kt dnh ny nn v nang nhanh b v

hn so vi mu alginate. S kt hp ny lm cho t l sng st VSV trong

mu alginate cao hn nhiu ln so vi mu tinh bt ch phm nang cng.


63/7254

Kt quny cng tng t nh kt qu ca Randolph Stanley Porubcan

thc hin nm 2006[42]. Trong nghin cu ca Randolph Stanley Porubcan,

ng cng to nang cng probiotics cha Natri alginate vi hm lng t 10%n 99%. Kt qu ca nghin cu cho thy s lng VSV b cht trong mi

trng acid pH 1,6 khng nhiu, c th trc khi th trong mi trng acid

pH 1,6 th s lng cfu/ nang l 9,0x109, sau khi th trong mi trng acid

pH 1,6 trong 90 pht s lng cfu/ nang khng cha alginate nh hn 104,

cn s lng cfu/ nang cha alginate l 4x108 [42]. Kt qu nghin cu cng

cho thy khi s dng alginate lm t dc n trong nang cng lm gia

tng t l VSV sng st trong mi trng acid pH thp.

Tuy nhin, sau 2 gi trong mi trng acid pH 1,2 cho kt qu hu nh

khng cn VSV sng st. Do vy, cn phi c bin php phi hp vi cc t

dc bo v khc hoc thay i dng bo ch tng t l VSV sng st.


64/7255

KT LUN V XUT

I. KT LUN

ti thu c cc kt qu sau:1. alginate k s s

k L. acidophiluso q k:

Khi s dngdung dch natri alginate 2% lm t dc bo v trong qu

trnh ng kh lm tng t lsng st caL. acidophilus so vi mu ng

kh vi nc ct (khong 27ln).

Th cht ca sn phm thu c sau ng kh vi alginate 2%: kh

lm nh v nhanh b ht m tr li.

Khi kt hp sa gy vi natri alginate cc nng 1%, 2%, 5% to ra

c nguyn liu c th cht tt tng ng vi dng ring t dc lsa

gy. Nhng t l sng st li khng c thy s khc bit ln gia cc mu , t

khong 109cfu/g.

2. nghin cu ng ca alginate n kh sng st ca vi

khun L. acidophiluso q thuc bt v thuc nang cng

nh gi c tc dng bo v vi sinh vt ca alginate khi s dng

lm t dc n cho dng thuc bt probiotic trong iu kin tip xc vi mi

trng acid HCl pH 1,2 cho s lng vi sinh vt sng st khong (10 6 107

cfu/g) cao hn khong 37 ln so vi cc mu khng s dng thm t dc

n v mu bt ng kh c kt hp t dc n thng dng l tinh bt.

nh gi c tc dng bo v vi sinh vt ca alginate khi s dng

lm t dc n cho dng thuc nang cng: trong iu kin tip xc vi mi

trng acid pH 1,2 vi tc khuy 50 v/pht, mu nang cng vi alginate

lm t dc n cho t l VSV sng st cao hn 100 ln so vi mu nang

cng vi tinh bt lm t dc n, v t l ny cao hn so vi mu thuc bt.


65/7256

II. XUT

Do thi gian c hn nn ti cha cp c ht cc vn c lin

quan, chng ti xut mt s hng nghin cu nh sau: Nghin cu vn nh ca nang v thuc bt vi thi gian di ti 1

nm.

Nghin cu cc bin php kt hp t dc alginat vi cc t dc bo

v khc trong qu trnh to thuc bt v ng nang cng lm gia tng s

lng VSV sng st trong mi trng acid pH thp

Nghin cu mt s bin php bo ch khc ca t dc alginat vi

VSV nh phng php vi nang ha, phng php bao pellet,


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TI LIU THAM KHO

Ti liu ting Vit

1. B mn Cng nghip Dc, Trng i hc Dc H Ni (2009),K thut sn

xut Dc phm, Nh xut bn Y hc, tp III.

2. B y t (2010), Dc in Vit Nam IV, Nh xut bn Y hc.

3. Nguyn Ln Dng (2012), Vi sinh vt hc, Nh xut bn khoa hc v k thut

H Ni.

4.Nguyn Ln Dng, Nguyn ng c, ng Hng Min, Nguyn Vnh Phc,

Nguyn nh Quyn, Nguyn Phng Tin, Phm Vn Ty (1976), Mt s phng

php nghin cu vi sinh vt hc, NXB KHKT, tp 2, tr 68, 69, 128.

5. o Th Hnh (2012), Nghin cu bo ch cm probiotic cha vi khun

Lactobacillus acidophilus, Lun vn thc s, i hc Dc H Ni, trang 7-12.

6. Nguyn Ngc Hip, Bi Tng Hip (2008), Vi sinh vt trong sn phm

probiotic, Tp ch Dc hc s 390.

7. L Gia Hy (2006), Vi sinh vt hc i cng(tp 2), Vin cng ngh sinh hc,

Vin khoa hc v cng ngh Vit Nam, H Ni.

8.Ng ng Ngha (1999), Ti u ha quy trnh cng ngh sn xut alginat natri

t rong m Vit Nam v mt s ng dng ca n trong mt s lnh vc sn xut,

Lun n tin s, Trng i hc Thy Sn.

9. L Xun Phng (2008), Th nghim vi sinh vt hc, i hc Nng, 33-65.

10.Nguyn Vn Thanh (2009), Cng ngh sinh hc Dc, Nh xut bn Gio dc

Vit Nam.

11. Trng i hc Dc H Ni, Qu trnh thit b(2010).

12. Phm Vn Ty, V Nguyn Thnh (2006), Cng ngh sinh hc, NXB Gio dc,

tp 5, tr. 129 154.


67/72

Ti liu ting Anh

13. Apajalahti. J. H. A, L. K. Sarkilabti, B. R. E. Maki, J. P. Heikkinen, P. H.

Nurminen and W. E. Holben (1998), Effective recovery of bacteria DNA and

percent-guanine-plus-cytosin-based analysis of community structure in the

gastrointestinal tract of broiler chickens, Appl Environ. Microbiol, 64, pp. 4084 -

4088.

14. A. Sibel Akalin, Siddik Gn, Selmin Dzel (1997), Influence of Yogurt and

Acidophilus Yogurt on Serum Cholesterol Levels in Mice, Journal of Dairy

ScienceVolume 80, (Issue 11), Pages 27212725.

15. Asma Sohail, Mark S. Turner, Allan Coombes, Thor Bostrom, Bhesh Bhandari

(2011), Survivability of probiotic encapsulated in alginat gel microbeads using a

novel impinging earosols method, International Journal of Food Microbiology

145, pages 162-168.

16. C Felley, P Michetti (2003), Probiotics and Helicobacter pylori, Best

Practice & Research Clinical Gastroenterology, Volume 17, Issue 5, Pages 785-791.

17. C Morgan, G Vesey (2009), Freeze-Drying of Microorganisms, Elsevier,

Australia.

18. Coates D, Richardson G. (1974), A note on the production of sterile solutions

of sodium alginat. Can J Pharm Sci9: 6061.

19. Dianawati Dianawati, Vijay Mishra, Nagendra P. Shah (2013), Survival of

Bifidobacterium longum 1941 microencapsulated with proteins and sugars after

freezing and freeze drying,Food Research International 51, pages 503509

20. Drahoslava Lesbros-Pantoflickova, Ire`ne Corthesy-Theulaz, and Andre L.

Blum (2007), Helicobacter pylori and Probiotics, The Journal of Nutrition

Volume 137 no 3, 812-818.
http://www.sciencedirect.com/science/journal/00220302http://www.sciencedirect.com/science/journal/00220302http://www.sciencedirect.com/science/journal/00220302/80/11http://www.sciencedirect.com/science/journal/00220302/80/11http://www.sciencedirect.com/science/journal/00220302/80/11http://www.sciencedirect.com/science/journal/00220302http://www.sciencedirect.com/science/journal/00220302


68/72

21. Filomena Nazzaro, Florinda Fratianni, Raffaele Coppola, Alfonso Sada,

Pierangelo Orlando, (2009), Fermentative ability of alginat-prebiotic encapsulated

Lactobacillus acidophilus and survival under simulated gastrointestinal

conditions,Journal of Functional Food1, 319-323.

22. Gibson. G. R and Fuller. R. (2000),Aspect of in vitro and in vivo research

approaches directed toward identifying probiotics and prebiotics for human use,

J. Nut, 130, pp. 391-395.

23. Gong. J, Forster. R. J., Yu. H., Chamber. J. R., Sabour. P. M., Wheatcroft. R.

and Chen. S. (2002), Diversity and phylogenetic analysis of bacteria in the

muscosa of chicken ceca and comparison with bacteria in the cecal lumen, FEMS.

Microbiol. Lett, 208, pp. 1 - 7.

24. Hartman AW et al. Viscosities of acacia and sodium alginat after sterilization

by cobalt-60.Journal of Pharmaceutical Sciences1975; 64: 802805.

25. Hershberg. R. M. and L. F. Mayer (2000),Antigen processing and presentation

by intestinal epithelial cells polarity and complexity, Immunol. Today 21, pp.

123128.

26. James W. Anderson, MD and Stanley E. Gilliland, PhD, (1999), Effect of

Fermented Milk (Yogurt) Containing Lactobacillus acidophilus L1 on Serum

Cholesterol in Hypercholesterolemic Humans, Journal of The American College

of NutritionVolume 18 no 1, 43-50.

27. Jose A. Medeiros, MD, PhD and Marta-Isabel Pereira, MD (2013), The Use

of Probiotics in Helicobacter pylori Eradication Therapy, Journal of ClinicalGastroenterology Volume 47 - Issue 1,pages 1-5.

28. Kaila Kailasapathy and James Chin (2000), Survival and therapeutic potential

of probiotic organisms with reference to Lactobacillus acidophilus and

Bifidobacterium spp,Immunology and Cell Biology78, 8088.
http://journals.lww.com/jcge/toc/2013/01000http://journals.lww.com/jcge/toc/2013/01000http://journals.lww.com/jcge/toc/2013/01000


69/72

29. Karla Bigetti Guergoletto, Dried Probiotics for Use in Functional Food

Applications,Food Industrial ProcessesMethods and Equipment.

30. Kumar M, Nagpal R, Kumar R, Hemalatha R, Verma V, Kumar A,

Chakraborty C, Singh B, Marotta F, Jain S, Yadav H (2012), Cholesterol-

lowering probiotics aspotential biotherapeutics for metabolic diseases, PMC.

31. Latha Sabikhi, R. Dabu, D.K. Thompkinson, Suman Kapila (2010),

Resistance of Microencapsulated Lactobacillus acidophilus LA1 to Processing

Treatments and Simulated Gut Conditions, Food Bioprocess Technol 3, pages

586-593.

32. Lim Chi Ming, Raha Abd Rahim, Ho Yin Wan, Arbakariya B. Ariff (2009),Formulation of Protective Agents for Improvement ofLactobacillus salivariusI

24 Survival Rate Subjected to Freeze Drying for Production of Live Cells in

Powderized Form,Food Bioprocess Technol2, pages 431436.

33. M. Abadias, A. Benabarre, N. Teixido, J. Usall, I. Vinas (2001), Effect of

freeze drying and protectants on viability of the biocontrol yeast Candida sake,

International Journal of Food Microbiology65, 173-182.

34. Manoj Kumar, Ashok Kumar, Ravinder Nagpal, Dheeraj Mohania, Pradip

Behare1, Vinod Verma, Pramod Kumar, Dev Poddar, P. K. Aggarwal, C. J. K.

Henry, Shalini Jain & Hariom Yadav (2010), Cancer-preventing attributes of

probiotics: an update, Internationat Journal Food Sciences and Nutrition;

61(5):473-96.

35. Martha I. Alvarez-Olmos and Richard A. Oberhelman (2001), Probiotic

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