1

Cng ngh sn xut vang

Chng V
Chu K Sn
05/2014
TRNG I HC BCH KHOA H NI
VIN CNG NGH SINH HC V CNG NGH THC PHM
B MN CNG NGH THC PHM
Chng V: Vi khun lactic
v qu trnh ln men malolactic
2
Th trng v tim nng ca ch phm malolactic
Sn lng v tiu th ru vang nm 2004 (ONIVINS, 05/2005).
Sn lng vang
( 10
3
hL)
T l (%)
Tiu th vang
( 10
3
hL)
Th gii 290.000 - 230.000
EU (15 nc) 173.776 59,9 127.759
Php 55.733 19,2 33.100
50.962 17,6 28.300
Ty Ban Nha 42.136 14,5 13.900
c 10.500 3,6 20.250
M 19.510 6,7 24.664
Achentina 15.464 5,3 11.112
Chil 6.301 2,2 2.700
Nam Phi 9.135 3,2 3.507
New Zealand 1.250 0,4 675
c 14.712 5,1 4.361
Liu lng s dng trung bnh ch phm malolactic khong 1 g/hL
Cn 290 tn /nm
Gi khong 1400 /kg
Th trng tim nng cc ch phm malolactic 400 triu euro/nm
C hai hng ln ang phn chia th trng: Chr. Hansen v
Lallemand SA
Tim nng cc ch phm malolactic
3
5.1.1 c tnh chung
5.1 Vi khun lactic
Vi khun lactic c s dng rt nhiu trong cng nghip thc
phm:
- cc sn phm sa (sa chua, phomt, sa ln men)
- cc sn phm tht
- cc loi ung (vang, nc qu ln men, bia)
Vi khun lactic l mt nhm khng ng nht c kh nng ln
men gluxit to ra ch yu l axit lactic
Tnh cht vi khun lactic: Gram dng, bt ng, khng to
bo t, khng sinh tng hp c enzim catalase, nitrate
reductase v cytochrome oxidase, c yu cu phc tp v dinh
dng: axit amin, vitamin, gluxit ln men c.
Mt s sn phm c s dng vi khun lactic
4
Vi khun lactic gm cc ging sau:
- Lactococcus, Lactobacillus, Leuconostoc, Oenococcus,
Pediococcus v Streptococcus, Vagococcus, Aerococcus,
Tetragenococcus, v hai ging Bifidobacterium v Carnobacterium
c ngun gc t Lactobacillus
S phn loi ca vi khun da vo cc tnh cht hnh thi, ha
sinh, sinh l
v sinh hc phn t: lai ha DNA/DNA, DNA/RNA
gii trnh t gen (sequencing) 16S ca
rRNA
V.1.2 Chuyn ha ng ca vi khun lactic
Ln men homolactic: axit lactic l sn phm trao i cht ch
yu ca qu trnh ln men ng
Ln men heterolactic: sn phm qu trnh trao i cht gm c
axit lactic, CO
2
, axit acetic, ethanol
Homofermentation
5
(Fugelsang v Edwards, 2007)
(Fugelsang v Edwards, 2007)
6
(Fugelsang v Edwards, 2007)
7
V.1.3 Vi khun lactic trong sn xut vang
C mt t nhin trn b mt qu nho, l nho, dch ng, trong
mi trng sn xut vang
sn phm qu trnh trao i cht gm c axit lactic, CO
2
, axit
axetic, ethanol
Mt s loi vi khun lactic c trong dch ng v vang
Ging Hnh thi Kiu ln men Loi
Oenococcus Cu khun Heterolactic
Oenococcus oeni
Pediococcus Cu khun Homolactic Pediococcus damnosus
Pediococcus pentosaceus
Pediococcus parvulus
Leuconostoc Cu khun Heterolactic
Leuconostoc mesenteroides
Lactobacillus Trc khun Heterolactic ty tin
Heterolactic bt
buc
Lactobacillus casei
Lactobacillus plantarum
Lactobacillus brevis
Lactobacillus hilgardii
V.2 c trng ca Oenococcus oeni
C tn c l Leuconostoc oenos (i tn thnh Oenococcus oeni
vo nm 1995)
O. oeni c mt trn b mt nho, l nho v trong mi trng sn
xut vang
L cu khun, Gram dng, bt ng v khng to bo t. Cc
cu khun tn ti thnh tng cp hoc thnh tng chui
L vi khun ym kh ty tin, chu c axit cao (pH sinh
trng ti u l 4,8)
8
Mi trng vang:
- Nhit thp 12 18C
- Nng ethanol cao 10 -14% v/v
- pH thp 3,0 3,5
- nng SO
2
cao (> 10 mg/L)
- mi trng ngho cht dinh dng
Qu trnh sinh trng ca O. oeni i hi mi trng phc tp:
Yu cu bt buc vi mt s axit amin (arginine, axit glutamic,
isoleucine, tyrosine, valine v cysteine)
Sinh trng chm: khun lc pht trin sau 5 ngy, thi gian
th h khong 4h trn mi trng tng hp FT80, pH 5,3, 30C.
Chuyn ha ng theo c ch ln men d hnh
(heterofermentative: hexose chuyn ha thnh axit D-lactic,
CO
2
, axit axetic, ethanol)
C kh nng to ra axit L-lactic t axit L-malic nh enzim
malolactic (ln men malolactic)
O. oeni l vi khun thch hp nht cho qu trnh ln men
malolactic:
- Thch nghi tt nht vi mi trng vang
- khng gy ra nhng tc ng xu ti vang
9
5.3 Qu trnh pht trin ca vi khun lactic trong sn
xut vang
0
1
2
3
4
5
6
7
8
0 5 10 15 20 25 30 35 40 45 50
Temps (jours)
P
o
p
u
l
a
t
i
o
n

b
a
c
t

r
i
e
n
n
e

(
l
o
g

C
F
U
.
m
L
-
1
)
0
1
2
3
4
5
C
o
n
c
e
n
t
r
a
t
i
o
n

a
c
i
d
e

L
-
m
a
l
i
q
u
e

(
g
.
L
-
1
)
Ln men ru cn
Ln men malolactic
Oenococcus oeni
Lactobacillus
Pediococcus
Leuconostoc
Oenococcus
Lactobacillus
Oenococcus
Ban u, vi khun lactic c trn b mt qu nho vi t l 10
2

10
4
CFU/mL (Lactobacillus plantarum, Lactobacillus casei,
Lactobacillus hilgardii, Lactobacillus brevis, Leuconostoc
mesenteroides, Pediococcus damnosus v Pediococcus
pentosaceus, ch c mt t l nh O. oeni
Khi kt thc qu trnh ln men ru cn, O. oeni chim u th
v thc hin qu trnh ln men malolactic (10
6 -
10
7
UFC/mL)
10
5.4 Qu trnh ln men malolactic
5.4.1 C ch ln men malolactic
HOOC-CHOH-CH
2
-COOH HOOC-CHOH-CH
3
+ CO
2
EML
NAD, Mn
2+
11
5.4.2 Qu trnh sinh nng lng nh ln men malolactic
a) S vn chuyn axit malic bi enzim malate permease
Oenococcus oeni
Hmal
-
Hmal
-
H
+
Lac
-
CO
2
Kim
-
+
axit
nH
+
?
Lactococcus lactis
Mal
2- Mal
2-
H
+
CO
2
Kim
-
+
axit
?
Lac
-
Lac
-
Lac
-
nH
+
ADP + Pi
ATP
nH
+
H
2
-Malate (si pH < 3,5)
nH
+ CO
2
H
+
H Malate
-1
H Malate
-1
Lactate
-1
Lactate
-1
NAD
+
Mn
2+
?

ext
int
+
+
+
-
-
-
nH
+
nH
+
EML
pH
pHe 3,0 5,5
pHi
5,5 6,4
nH
+
a. a Axit amin
nH
+
b) Ngun nng lng cho O. oeni v qu trnh iu chnh pH ni bo
12
5.4.3 Li ch ca qu trnh ln men malolactic
a) Gim chua ca vang
Nng axit malic c trong dch ng t 2 -10 g/L
Sau khi ln men malolactic, pH tng ln 0,1- 0,4 n v
Lm cho vang du hn v lm gim cht ca tanin.
b) Thay i tnh cht cm quan
Lm cho vang m du, tinh t hn
Cc tnh cht cm quan thay i ty theo chng v rt kh lp
li nhng s thay i ny
Nhiu phn t cht thm c to ra lm thay i tnh cht
cm quan ca vang: qu trnh s dng axit citric (sn phm
ch yu l diacetyl ngoi ra cn c axetat, ethanol)
Qu trnh s dng ng cng nh hng n tnh cht cm
quan trong vang (ru bc cao, ester (ethyl lactate)
Qu trnh ng ha axit amin (arginin v methionin) nh
hng n tnh cht cm quan trong vang
nh hng xu n tnh cht cm quan: to ra biogenic amin,
carbanate ethyl
Cc nghin cu c s dng cc phng php cm quan kt hp
vi gas chromatography (GC) olfactometry v GC mass
spectrometry c s dng nghin cu su hn v cc nh
hng ny n gi tr cm quan vang
LMML c nh hng n mi vang, b sung s a dng cho mi
ca vang
nh hng ca LMML n mi v vang ph thuc vo vi khun
lactic s dng v ty thuc vo tng loi vang.
Theo Henick-Kling, LMML lm tng mi hoa qu (fruity aroma) v
mi b (buttery note) ca vang nhng lm gim mi cy c
ti.
b) Thay i tnh cht cm quan
13
b) Thay i tnh cht cm quan
Mi hoa qu c tng cng do s to thnh cc esters. Cc
esters (ethyl acetat, ester ca ru etylic vi cc axit bo C4-
C10). Cc esters ny c to thnh ch yu trong qu trnh
ln men ru cn di tc ng ca nm men, tuy nhin vi
khun lactic trong vang cng sinh ra mt lng nht nh cc
ester ny.
Mi b (buttery note) c tng cng do s to thnh diacetyl
t ln men axit citric bi vi khun lactic
Axit citric l mt trong ba axit hu c chnh trong nho v trong
vang sau axit malic v axit tartric
Ln men axit citric to thnh axit lactic, acetic, diacetyl, acetoin
v 2,3-butanediol.
Diacetyl: (CH
3
COCOCH
3
): 2,3-butanedione, biacetyl,
Chng ln men
malolactic
Cc chng O. oeni khc nhau c kh nng sinh tng
hp diacetyl khac nhau
Loi vang
Vang cho nhiu diacetyl hn vang trng
Lng vi khun
malolactic
Lng vi khun malolactic thp (10
4
-10
6
UFC/mL to
nhiu diacetyl
Tip xc vi nm
men sng
Gim lng diacetyl
Vang tip xc vi oxi
trong LMML
Tng lng diacetyl do oxi ha alpha-acetolactate
thnh diacetyl
Sulfite
Sulfite phn ng vi diacetyl lm mt gi tr cm quan
ca diacetyl
B sung sulfite lm c ch nm men/vi khun c tc
dng lm n nh lng diacetyl ti thi im b sung
Hm lng axit citric
T l thun vi lng diacetyl, tuy nhin to ra nhiu
axit bay hi (axit acetic)
pH khi LMML
pH thp tng cng qu trnh to diacetyl
Nhit LMML
18-25C l thch hp cho qu trnh to diacetyl
Mt s tc nhn nh hng n hm lng diacetyl trong vang
14
S gim mi c ti (vegetative, green/grassy aroma) l do s
ng ha cc aldehit ca vi khun lactic trong vang to thnh
ethanol v axit axetic
Aldehit ch yu trong vang l acetaldehit c to thnh trong
qu trnh ln men ru di tc dng ca nm men
gim lng aldehit c th b sung mt lng nht nh SO
2
(tc dng vi cc hp cht carbonyl (aldehit hoc cetone) to thnh
sulfonate)
Ngoi ra, LMML cn to ra mt s mi c trng khc nh: hoa
(floral), hnh nhn (nutty), nm men (yeasty), g si (oaky), m
hi (sweaty), gia v (spicy), rti (roasted), nng (toasty), vani
(vanilla), khi (smoky), t (earthy), ng (bitter), mt (honey).
Cn nhiu nghin cu hn c th lin h s thay i mi n s
to thnh hoc phn hy cc hp cht c trong vang bi vi khun
lactic
b) Thay i tnh cht cm quan
b) Thay i tnh cht cm quan
Bn cnh nh hng n mi, LMML cn lm tng hu v ca vang
do s to thnh cc polyols (glycerol v mannitol) v cc
polysaccharides ca vi khun lactic
Cc monoterpen l cc hp cht thm quan trng c trong nho v
vang
Anthocyan l hp cht mu quan trng ca nho v vang .
Monoterpen v anthocyan thng lin kt vi ng bng cc lin
kt glycoside
Monoterpen lin kt vi ng l hp cht khng mi v khng
bay hi. Di tc dung ca glycosidase thy phn lin kt glycoside
gii phng monoterpen to mi cho vang
15
c) n nh vi sinh vt
Ln men malolactic lm ngho cht dinh dng v cc ngun
carbon thun li cho qu trnh pht trin ca vi khun nh:
axit citric, axit malic, ng ln men cn st (pentose, glucose
v fructose): hn ch s pht trin ca vi khun c hi.
C s tng hp cc cht khng khun trong qu trnh ln men
malolactic
CH : khng nn thc hin qu trnh ln men malolactic i
vi vang c pH cao (to iu kin thun li cho s pht trin
ca vi khun c hi)
V.4.4 Kim sot qu trnh ln men malolactic (LMML)
I) Cc yu t nh hng n qu trnh ln men malolactic
Cc yu t cng ngh
Cc bc cng ngh sn xut ru vang nh hng n ln
men malolactic: Qu trnh lc nh hng khng tt n s
pht trin ca vi khun lactic (loi b h vi sinh vt ni ti,
gim lng cht dinh dng)
Qu trnh chuyn thng khi kt thc ln men ru cn loi b
nm men (ngun dinh dng khi nm men t phn)
Khng nn b sung sulfite khi kt thc qu trnh ln men ru
cn
Thay thng g bng thng inox cng lm hn ch ln men s
pht trin ca h vi sinh vt ni ti
16
Cc yu t vi sinh vt
Tng tc gia cc nhm vi sinh vt cng tn ti trong vang:
vi khun - vi khun, vi khun - nm men: cnh tranh cht
dinh dng.
Trong ln men ru cn : tng tc vi khun - nm men
mang tnh cnh tranh, sau c tnh h tr
Tng tc vi khun vi khun: Pediococcus,
Lactobacillus, Leuconostoc v Oenococcus: canh tranh dinh
dng, mt s loi vi khun sinh tng hp bacteriocin
S tn cng ca bacteriophage
Cc yu t ha l
a) pH:
pH c nh hng ln nht n s pht trin ca vi khun
lactic
pH ti u ca vi khun lactic gn pH trung tnh
pH cng thp th thi gian gia ln men ru cn v ln men
malolactic cng lu
pH < 3,0, s sinh trng ca vi khun lactic rt kh thm
ch khng th xy ra
axit cao lm gim pH ni bo v nh hng n hot ng
ca cc enzim khc nhau
pH nh hng n nhiu n sinh tng hp ATP, RNA, protein
lm v hot protein.
pH nh hng n qu trnh chuyn ha ng v axit malic
pH 3,0, vi khun khng th chuyn ha ng
pH nh hng n mng t bo cytoplasmic
17
Cc yu t ha l
b) Nng ru ethylic:
Ru ethylic l sn phm ca qu trnh ln men ca nm men
(10 16% v/v)
Kh nng chng chu ru ethanol ph thuc vo cc iu
kin khc ca mi trng: kh nng ny gim khi nhit
tng v pH ca vang gim
Nng ethanol thp (khong 3% v/v) kch thch s pht
trin ca O. oeni
Khi nng t 8%, ethanol bt u c ch v c th tiu
dit O. oeni
Nu ethanol c b sung t t th s lm gim s c ch i
vi O. oeni
Cc yu t ha l
nh hng ca ethanol:
Mt n nh cu trc mng t bo: tng lng mng t bo,
mt cn bng mng t bo (mt kh nng bn thm)
nh hng n qu trnh nhn bn DNA, hot enzim ni
bo, s vn chuyn c cht (axit amin, ng, axit hu c)
Bin tnh protein
18
Cc yu t ha l
c) Nhit :
Nhit nh hng ln ti cht lng ru vang.
Nhit nh hng ti tc sinh trng ca vi sinh vt ni
chung
Nhit ti u ca vi khun l 25-30C
Nhit thch hp LMML l 18-22C
15C, LMML rt chm v hu nh khng c s pht trin
ca O. oeni
Nhit thp nh hng ti cu trc DNA, RNA, lm cng
ha mng t bo
Nhit l yu t d dng iu chnh nht (so vi nng
ethanol v axit)
Cc yu t ha l
d) Sulfite:
Sulfite c th c b sung vo trong qu trnh sn xut vang
di dng mui hoc kh, sulfite c tc dng chng oxi ha v
khng khun
Sulfite c th c sinh ra bi qu trnh chuyn ha ca nm
men
Sulfite cng c kh nng khng khun khi pH cng thp
Nng 1 - 10 mg L
-1
de SO
2
t do v t 50 - 100 mg L
-1
SO
2
kt hp (100 - 150 mg L
-1
SO
2
tng s) c th c ch s pht
trin ca vi khun lactic
Sulfite nh hng n hot mt s enzim: enzim
malolactic, enzim ATPase
19
H
2
O + SO
2
H
+
+ (HSO
3
)
-
2H
+
+ SO
3
=
0
10
20
30
40
50
60
70
80
90
100
1,0 2,0 3,0 4,0 5,0 6,0 7,0 8,0 9,0
pH
T

c

d

n
g

(
%
)
Dng bisulfite
(HSO3-)
Dng phn t
(SO2)
Dng sulfite
(SO3--)
Cn bng gia cc dng dioxit lu hunh cc pH khc nhau
Cc yu t ha l
e) S thiu ht cht dinh dng:
Vang non l mi trng ngho cht dinh dng,
Thnh phn ha hoc ca vang thay i ph thuc vo nhiu
yu t khc nhau: thi tit, chn ca nho, chng nm men.
Cui qu trnh ln men: ng st; glucose, fructose, pentose
vi nng t 0,1 10 g/L vi khun pht trin
Cc ngun nit (protein, peptide, axit amin) bi thiu ht nht.
Ngun ny c b sung bi s t phn nm men
O. oeni c kh nng sinh tng hp cc enzim exoprotease v
aminopeptidase phn ct cc hp cht cao phn t sinh ra
t qu trnh t phn nm men.
20
Cc yu t ha l
f) Mt s yu t c trng ca vang:
Cc hp cht polyphenol v anthocyane c th c ch hoc
knh thch s pht trin ca vi khun.
Cc phn t axit bo cng nh hng n s pht trin ca vi
khun:axit decanoic v dodecanoic l sn phm qu trnh trao
i cht ca nm men: nng thp, chng kch thch hot
enzim malolactic, nng cao, chng lm gim hot
enzim malolactic.
Trong mt s trng hp, axit oleic c tc dng kch thch bo
tn s sng cho vi khun
S thch ng ca vi khun
Protein b bin tnh
Protein b phn hy Protein hot ng
Proteases
+ ATP
Chaperonnes
+ ATP
1 2
Nguyn tc hot ng ca HSP (Heat Shock Protein)
21
Mt s stress (ethanol, nhit , axit, sc thm thu, thiu
dinh dng, stress oxi ha) sinh tng hp protein sc nhit HSP.
Mt s loi protein c vai tr chaperon nh (GroES, DnaK) v
protein thuc h Clp (Caseinolytic protein) c cu tr li chung
vi cc stress khc nhau
Chaperon phn t GroES v DnaK c vai tr quan trng vi n
hn ch s bin tnh ca protein, cho php protein tr li trnh
thi hot dng
Nhm Clp l nhm cc enzim ATPase (ClpX, ClpL) v protease
(ClpP) thy phn protein b bin tnh bt thun nghch qua
cc stress khc nhau.
Ngoi ra cn mt s protein c trng cho cc stress khc
nhau cng c sinh tng hp trong cc qu trnh chng chu
stress v thch nghi vi mi trng
ASP (Acid Shock Protein): chng chu stress axit: sa cha
protein bi hng trong cc stress axit
CSP (Cold Shock Proten): chng chu stress nhit thp:
lm n nh RNAm trong qu trinh dch m
GSP (General Shock Proten): chng chu s thiu cht dinh
dng
22
Kt lun chung:
Vang l mi trng rt phc tp (multi-stress).
Nhiu yu t c th nh hng n s sng st cng nh pht
trin ca vi sinh vt (cng hng ha trit tiu ln nhau).
Cc chng O. oeni khc nhau th kh nng chng chu stress
cng hon ton khc nhau.
S thay i ca tt c cc yu t cng ngh, vi sinh vt ha l,
lm cho qu trnh LMML t nhin cng kh khn hn
C nhiu bin php kim sot qu trnh LMML
II) Kim sot v ti u qu trnh LMML
LMML t nhin din ra rt kh khn trong mi trng vang:
thng xy ra hin tng LMML khi ng chm hoc thm
ch khng khi ng c
Trong nhiu trng hp qu trnh LMML din ra lu cng nh
hng n cht lng vang
Nhiu phng n c s dng kim sot qu trnh LMML:
+ kch thch LMML t nhin
+ s dng lng sinh khi vi khun c nh
+ b sung sinh khi vo vang non LMML
23
II) Kim sot v ti u qu trnh LMML
a) Kch thch LMML t nhin
+ nhit hm: gi nhit trn 18C (y l yu t d iu
chnh nht nhng gi thnh cao)
+ pH: i vi mt s vang qu chua (pH qu thp) c th gim
axit bng cch b sung base yu vo trung ha (natri
tartrate, natri bicarbonate)
+ Sulfite ha: dng lng va , ph thuc vo nng sulfite
v pH vang
+ Sc kh trong ln men ru cn c th kch thch s pht trin
ca vi khun
+ Ln men p lc (gi mt phn CO
2
cui ln men ru cn
cng c th kch thch pht trin ca vi khun
+ Trn cc thng LMML tt vi cc thng LMML cha tt
II) Kim sot v ti u qu trnh LMML
b) Phng php c nh vi khun
+ t bo vi khun c c nh trong gel alginate, qu trnh ln
men din ra trong cc thit b ln men (bioreactor): phng
ny t c s dng do hot enzim malolactic thp do
lng t bo vi khun bi gim trong mi trng c ch nh
vang, nh hng xu n cc tnh cht cm quan ca vang
24
II) Kim sot v ti u qu trnh LMML
c) B sung cc ch phm malolactic vo vang
+ mt s loi vi khun khc nhau c s dng lm cc ch
phm malolactic (cc loi thuc ging Lactobacillus).
+ O. oeni l chng c s dng nhiu nht v c kh nng thch
nghi vi mi trng vang tt nht LMML
+ B sung cc ch phm malolactic cho php kim sot c mt
t bo, thi gian v hiu qu ca qu trnh LMML.
d) B sung nm men kt hp vi vi khun
+ B sung vi khun sau khi b sung nm men 24 h.
+ Vi khun s thch nghi dn vi mi trng khc nghit nh
vang (nghin cu su tng tc gia vi khun v nm men)
+ Ch : cn kim sot s pht trin ca vi khun trc qu
trnh LMML (to v chua cho vang)
B sung cc ch phm malolactic cn qu trnh hot ha
+ Ch phm malolactic th h u tin:
Cc ch phm ny cn qu trnh hot ha (a vi khun tr li
mi trng lng) v thch nghi vi khun vi mi trng vang vi
mc ch tng t l sng st ca t bo.
25
Phng php hot ha ch phm malolactic lnh ng.
Phng php 1
Vang : 50 %
Nc : 50 %
Hot ha: 4
gL
-1
Vi khun:
Pha 1
Vang
22 C
LMML

T l cy
0,5 %
(v/v)
Phng php 2
Vang : 50 %
Nc : 50 %
Hot ha: 4
gL
-1
Vi khun:
Pha 1
Pha 2
Vang
22 C
LMML
+ 50 %
vang
22 C
LMML

T l cy
1 %
(v/v)
Qu trnh b sung cn c thc hin vo cui qu trnh ln men ru cn
Cht hot ha: 50% glucose + 50% cao nm men
Phng php ny hiu qu nhng mt nhiu thi gian v cng sc
B sung trc tip cc ch phm malolactic
+ Ch phm malolactic th h th hai:
S dng cc ch phm ny di dng lnh ng, m c ready
to use
+ Nguyn tc: b sung mt lng ln vi khun c thch nghi
thc hin qu trnh LMML m khng cn n qu trnh hot ha
II) Kim sot v ti u qu trnh LMML
d) S dng cc chng nm men ti t hp c kh nng
thc hin qu trnh LMML
26
III) Chun b cc ch phm malolactic
a) Mc ch:
Lm cho qu trnh LMML n gin, nhanh v hiu qu i vi qu
i vi ngui sn xut vang
Tableau 5 : Critres de slection pour les souches de O. oeni destines la production de prparations malolactiques.
Tiu ch
Chng chu stress C kh nng sinh trng trong mi trng vang, tc
sinh trng nhanh
Chu c pH thp (3,0)
Chu c nng cn ethanol cao (14 % v/v)
Chu doc SO
2
Sinh trng c nhit thp (14C)
Hot enzim malolactic cao
Chng chu c bacteriophages
Chu c qu trnh lnh ng
Cc tnh cht cm quan Khng sinh ra amines biogene v cc hp cht c
(carbamate ethyle)
To cht thm (diacetyle) v tng mi thm qu
Khng sinh ra axit bay hi
b) Tiu ch tuyn chn chng O. oeni cho LMML:
n nh cc tnh cht di truyn
27
b) Thch nghi vi khun vi mi trng vang:
Nguyn tc: vi khun c nui cy hoc sinh trng trong mi
trng vi cc stress va phi c kh nng chng chu cc stress
vi cng cao hn
Cn nui cy vi khun trong mi trng vi cc stress va phi
vi khun c thi gian hnh thnh v pht trin cc c ch thch nghi
vi mi trng khc nghit
Nguyn tc bo v cho (cross protection): vi khun c thch
nghi vi mt loi stress c th chu c nhiu stress c bn cht
khc
Vi khun trong pha n nh pht trin c ch chung cho cc stress
khc nhau (GSP)
b) Thch nghi vi khun O. oeni vi mi trng vang:
Thch nghi vi mi trng c cha ethanol vd: 8% (v/v)
Thch nghi vi mi trng c pH thp vd pH 3,5
Thch nghi vi nhit cao
Thch nghi ngn hn hoc thch nghi di hn
28
Chn v bo qun chng ging O. oeni - 80 C
Nhn ging
To canh trng v thch ng vi khun trong cc fermentor:
cc yu t khng ch ty theo phng php
Thu hi v c c sinh khi bng phng php ly tm
B sung cht chu lnh ( cryo-protectant)
Lnh ng ch phm - 80 C
Sy ng kh ch phm
ng gi v bo qun
Quy trnh sn xut v bo qun cc ch phm malolactic
IV) Qu trnh thch nghi vi cc stress O. oeni
STRESS
Mng t bo
Tip nhn thng tin
GENOME
TR LI
HSP
lng mng t bo
H
+
ATPases
L-Lac
MLE
L-Mal
permeases
L-Mal
L-Lac
fpm
29
Cc c ch thch nghi
Ln men malolactic: decarboxyl ha axit malic thnh axit lactic
to ra nng lng trong iu kin thiu cht dinh dng
Hot ha enzim ATPase: bm (y) H
+
ra khi t bo trong iu
kin mi trng axit. Qu trnh tiu tn nng lng bng cch thy
phn cc hp cht cao nng ATP
Sinh tng hp stress protein: HSP, ASP, CSP, GSP: thy phn
protein b bin tnh hoc a protein tr v trng thi hot ng
Thay i lng mng t bo, thay i thnh phn mng t bo
thay i kh nng bn thm ca mng t bo trong cc iu
kin c ch
Protein b bin tnh
Protein b phn hy Protein hot ng
Proteases
+ ATP
Chaperonnes
+ ATP
1 2
Nguyn tc hot ng ca HSP (Heat Shock Protein)