Sản Xuất Chitin - Chitosan Bằng Phương Pháp Sinh Học

B GIO DC V O TO
TRNG I HC K THUT CNG NGH TP.HCM
SN XUT CHITIN CHITOSAN

BNG PHNG PHP SINH HC
KHOA MI TRNG & CNG NGH SINH HC

CHUYN NGNH CNG NGH SINH HC
TP. H Ch Minh, 2013
L TN PH
LI NI U
Nh chng ta bit v nhng c nhn, t chc hot ng v lnh vc
khoa hc k thut th cng hiu r hn v chitin chitosan, y l mt
polysacharite t nhin v chim sn lng rt ln, ch sau cellulose. Bn
cnh l nhng ng dng tuyt vi ca chitosan va hiu qu va rt an
ton v c bit l thn thin vi mi trng mt vn rt c quan tm
trn th gii hin nay. D rng chitin chitosan c sn lng ln, ng dng
hon ho nhng thin nhin vn d khng u i con ngi qu nhiu, mun
c c chitin chitosan th cn phi c mt quy trnh x l hiu qu, bt
ngun t i hi cc nh khoa hc bt u nghin cu tm ra nhiu
phng php c th sn xut c chitin chitosan. Trong sn xut chitin
chitosan bn cht chung ca qu trnh vn l kh cc thnh phn khong v
hm lng protein cn d tha trong ph liu v tm s cho ra chitin. Vy
qu trnh c thc thi ra sao? Dng tc nhn no kh khong, loi
protein thu chitin v deacetyl ha chitin thu sn phm chitosan. Vi
phng php ha hc ngi ta dng tc nhn ca ha cht x l, nhng
bn cnh xut hin thm nhiu vn khc cn x l nhc im ca
quy trnh. Ci tin hn ngi ta ngh ra phng php ha sinh, phng php
ny c nh gi gim thiu phm no lng ha cht dng trong quy trnh,
tuy nhin u cng cha gii quyt tn gc c vn , vn cn lng
ha cht dng trong qu trnh deacetyl ha chitin. V vy n vn cn nh
hng ln ti mi trng sng, sc khe cng nhn lao ng, ngi tiu
dngDo phi lm sao gii quyt cc nhc im , nng cao hiu
sut qu trnh, gim chi phkhng u khc hn l phng php sinh hc,
ch c sinh hc mi gn gi v gn lin vi s sng, s tn ti ca mun loi.
Tuy nhin n ch mi c manh nha trong suy ngh v ang c nghin
cu th nghim mt s phng th nghim. Trong chuyn ny s tng
quan hu ht cc phng php sn xut chitin chitosan bng ha hc, ha
L TN PH
sinh, bn c c th khi qut kin thc c bn v chitin chitosan. Bn

cnh c s cho php ca ban lnh o khoa Mi Trng & Cng Ngh
Sinh Hc trng i Hc K Thut Cng Ngh Tp.hcm, ng cho ti lm
vic ti phng th nghim ca trng, a ch 475A in Bin Ph, phng
15, qun Bnh Thnh, Tp.hcm. Cng vi s hng dn tn tm ca ging
vin TS. Nguyn Hoi Hng v vic thc hin th nghim sn xut chitin
chitosan t ph phm thy sn v tm s, nhm thc hin chuyn chn
thnh bit n v iu ny.
ii
L TN PH
MC LC
LI NI U ...........................................................................................................1
MC LC ................................................................................................................ iii
DANH SCH HNH .................................................................................................v
DANH SCH BNG .............................................................................................. vi
CHNG 1. TNG QUAN V CHITIN - CHITOSAN ..................................7
1.1.
Gii thiu ......................................................................................................7
1.1.1.
Chitin ......................................................................................................7
1.1.2.
Chitosan ..................................................................................................8
1.2.
Ngun gc thu nhn ...................................................................................10
1.3.
Tnh hnh nghin cu trong v ngoi nc .............................................10
1.3.1.
1.3.1.1.
Tnh hnh sn xut v nghin cu trong nc ...............................10
1.3.1.2.
Tnh hnh sn xut v nghin cu ngoi nc ...............................11
1.3.2.
1.4.
Chitin ....................................................................................................10
Chitosan ................................................................................................12
1.3.2.1.
Tnh hnh sn xut v nghin cu trong nc ...............................13
1.3.2.2.
Tnh hnh sn xut v nghin cu ngoi nc ...............................13
Cu trc ha hc ........................................................................................13
1.4.1.
Cu trc ha hc ca chitin ................................................................13
1.4.2.
Cu trc ha hc ca chitosan............................................................15
1.5.
Tnh cht ha l sinh hc ..........................................................................16
1.5.1.
Tnh cht l ha ...................................................................................16
1.5.2.
Tnh cht sinh hc ...............................................................................17
1.5.2.1.
Chitin ..............................................................................................17
1.5.2.2.
Chitosan .........................................................................................18
CHNG 2. CC PHNG PHP SN XUT CHITIN CHITOSAN .....21

2.1.
Phng php ha hc ................................................................................21
2.1.1.
S lc v phng php ......................................................................21
2.1.2.
Quy trnh k thut ................................................................................21
iii
L TN PH
2.1.3.
Thuyt minh quy trnh .........................................................................22
2.1.4.
u nhc im ....................................................................................24
2.2.
2.1.4.1.
u im ..........................................................................................24
2.1.4.2.
Nhc im ....................................................................................24
Phng php ha sinh kt hp .................................................................25
2.2.1.
S lc v phng php ......................................................................25
2.2.2.
Quy trnh k thut ................................................................................25
2.2.3.
2.2.4.
u nhc im ....................................................................................27
2.3.
2.2.4.1.
u im ..........................................................................................27
2.2.4.2.
Nhc im ....................................................................................27
Phng php sinh hc ...............................................................................28
2.3.1.
S loc v phng php ......................................................................28
2.3.2.
Quy trnh k thut ................................................................................29
2.3.3.
2.3.4.
Thit b ..................................................................................................35
2.3.5.
u im ................................................................................................37
2.3.6.
Nhc im ..........................................................................................38
CHNG 3. KT LUN V KIN NGH .........................................................40

3.1.
Kt lun ......................................................................................................40
Ti liu tham kho:...........................................................................................42

HT ....................................................................................................................44
iv
L TN PH
DANH SCH HNH

Hnh 1. 1 Deacetyl ha chintin thnh chitosan .........................................................10
Hnh 1. 2 Cu trc ha hc N-acetyl-D-glucosamine lin kt vi nhau bng cu ni
(1-4) glucoside .......................................................................................................14
Hnh 1. 3 Cu trc ha hc chitin .............................................................................14
Hnh 1. 4 cu trc chitosan .......................................................................................16
Hnh 2. 1 Qu trnh deacetyl ha bng phng php ha hc ................................24
Hnh 2. 2 Ni ln men c s dng trong quy trnh sn xut .................................36
Hnh 2. 3 np ni v cc chi tit thit b ...................................................................37
Hnh 2. 4 B iu khin .............................................................................................37
L TN PH
DANH SCH BNG

Bng 1. 1 Thnh phn chitin ca mt s ph liu thy sn:..................................8
Bng 2. 1 Cng thc phi trn nguyn liu ln men ............................................32
vi
L TN PH
CHNG 1. TNG QUAN V CHITIN - CHITOSAN

1.1.
Gii thiu
1.1.1. Chitin
-
Chitin chitosan l mt polysacharite tn ti trong t nhin vi sn lng rt

ln (ng th hai sau cellulose). Chitin ln u tin c tm thy trong nm
bi nh khoa hc ngi Php Braconnot vo nm 1811, nm 1823, Odier phn
lp c mt cht t b cnh cng m ng gi l chitin hay chiton, ting Hy
lp c ngha l v gip, nhng ng khng pht hin ra s c mt ca nit trong
. Cui cng c Odier v Braconnot u i n kt lun chitin c dng cng
thc ging vi cellulose. Ngy nay chitin c pht hin c ng vt v thc
vt.
Trong ng vt, chitin l mt thnh phn cu trc quan trng ca cc v mt s

ng vt khng xng sng nh: cn trng, nhuyn th, gip xc v giun trn.
Trong ng vt bc cao monome ca chitin l mt thnh phn ch yu trong m
da n gip cho s ti to v gn lin cc vt thng da, c bit trong cc loi
thy sn c bit l trong v tm, cua, gh, hm lng chitin chim kh cao dao
ng t 14 35% so vi trng lng kh, c bit l tm. Theo c tnh ca T
chc Nng Lng Th Gii (FAO) sn lng tm nui trng trn th gii nm
2011 l 2.934.168 tn, ring Vit Nam 403.600 tn V vy v tm, cua, gh
l ngun nguyn liu chnh sn xut chitin. Cn trong thc vt chitin c
thnh t bo nm h zygenmyctes, cc sinh khi nm mc, mt s loi to
V c tnh sinh hc v cu trc ha hc chitin l polysacharite c m khng

c, c khi lng phn t ln.[13]
Phn b
-
Chitin hin din thc vt bc thp. Trong th gii thc vt, chitin ch gii hn
loi nm, to lc, a y, non khun.
tm, chitin hin din dng kt hp vi cc thnh phn khc (khong).
L TN PH
Chitin cng hin din ng vt bc thp. Chitin l thnh phn cu to nn

xng hu c chnh ng vt khng xng sng (trng t, tit tc, nhuyn
th, cn trng). i vi ngnh tit tc chitin ch l mt trong nhng thnh phn
cu to ca b xng ngoi. Trong thin nhin, dng chitin nguyn cht ch tn
ti trong nang ca loi mc ng Logigo.
thy sn, chitin tn ti rt nhiu, c bit l v tm, cua, gh, nang

mc,...hm lng cao. V vy, phn ln chitin c chit xut t v ca cc
loi gip xc nht l v tm, k l v cua, gh. v tm cng nh v cua,
chitin to phc h vi protein v thng cha thm mt t l ln CaCO3.
Chitin tn ti trong t nhin dng tinh th. Trong t nhin him thy chitin
tn ti dng t do, n lin kt di dng phc hp chitin vi protein, chitin
vi cc hp cht v c,...Khi tn ti nh th, chitin c kh nng khng i
vi cc cht thy phn ha hc v enzyme, gy kh khn cho vic tinh ch, tch
chit. Ty thuc vo c tnh c th v s thay i tng giai on sinh l m
trong cng mt loi, ngi ta c th thy c s thay i v lng v cht chitin.
Mc d gp rt nhiu kh khn trong vic chit tch chitin nhng ngi ta

khng nh v s hin din ca chitin thc vt ln ng vt da vo php o
quang hc, phn tch bng tia X v phn ct bi enzyme hay phn ng ho
hc.[1]
Bng 1. 1 Thnh phn chitin ca mt s ph liu thy sn:
Ph liu
Thnh phn chitin (%)
V hn
0,4
V c
1,24
V cua ng
23,8
V tm ng
30,0
V tm bin
33,1
1.1.2. Chitosan
L TN PH
Chitosan l mt polysaccharide tuyn tnh. Loi chitosan thng mi c MW t

100.000 n 1.000.000 Da v thng mang mt trong ba trng thi : v nh
hnh, bn tinh th, tinh th. Cc dng vi tinh th ca chitosan (MW: 10.000
300000 Da) c kh nng hp th ln nh mt gel phn tn hay dng bt mn to
ra phn t chitosan c cu trc v kch thc c th. Cc nhm amin ca
chitosan c pKa ~ 6.5 chng mang in v tan trong mi trng acid. Chitosan
rt c o n l polime hu c t nhin duy nht mang in tch dng do c
nhng nhm amino t do tch in dng khng tan trong nc, v vy chng
tng tc c vi phn t
mang in tch m nh acid bo, acid mt,
phospholipid, protein v polysaccharide, chnh iu ny to cho chitosan nhng

thuc tnh c bit nht v ng kinh ngc. Mui ca chitosan c th tan trong
nc ph thuc vo pH, nhit ca qu trnh acetyl ha.[1]
-
Do mt s c tnh vn c, l hp cht t nhin, khng c, c th b phn hy,

c kh nng tng tc vi nhiu phn t sinh hc nn c ng dng rng ri
trong thc t trong nhiu lnh vc nh: nng nghip, ha hc, cng ngh sinh
hc, cng nghip giy, m phm, x l nc, bo qun thc phm, ph gia thc
phm, dc phm
Chitosan c sn xut bng cch kh acetyl (deacetylation DD) ca chitin,

mc deacetylation (% DD) c th c xc nh bng ph NMR, v DD%
trong chitosans thng mi l trong khong 6 10%. Cht c kh acetyl t
chitin c khm ph bi Roughet vo nm 1859, sn phm kh acetyl
chitin c t tn l chitosan bi nh khoa hc ngi c Hoppe Seyler vo
nm 1894.
L TN PH
Hnh 1. 1 Deacetyl ha chintin thnh chitosan

1.2.
Ngun gc thu nhn
S tng hp chitin loi gip xc

-
Billard quan st trn loi gip xc Decapoda cho bit s tng hp ca

chitin theo sau giai on lt xc. Glycogen c s dng hon ton v
tch t khi lp v mi c hnh thnh. S tng hp chitin gip xc lin
quan n vic s dng glycogen v n phi chuyn sang dng n gin
hn l D-glucose.
S tng hp v phn hy chitin gip xc rt tch cc v phn ln lp v

b tiu hy ch c hnh thnh li sau mi ln lt xc. Trong giai on
ny s thiu thc n lm v mng hn cho thy kh nng s dng chitin
nh mt cht bin dng v d tr.
Tnh hnh nghin cu trong v ngoi nc
1.3.
1.3.1. Chitin
1.3.1.1.
-
Tnh hnh sn xut v nghin cu trong nc
Vic nghin cu, sn xut chitin v cc ng dng ca chng trong sn xut,

phc v i sng l mt vn tng i mi nc ta. Nm 1978, trng i
10
L TN PH
Hc Thy Sn bt u nghin cu chit tch chitin. Trc yu cu x l ph

liu thy sn ngy cng cp bch, trc nhng thng tin khoa hc, k thut mi
v chitin, cng nh tim nng th trng ca chng, thc y cc nh khoa
hc nc ta bt tay vo nghin cu hon thin quy trnh sn xut chitin bc
cao hn, ng thi nghin cu cc ng dng ca chng cc lnh vc khc
nhau.
-
Gn y, khi chitin tr thnh nhu cu trong nhiu ngnh cng nghip v c gi

tr th rt nhiu c quan nghin cu nh: trng i Hc Nng Lm TP.HCM,
i Hc Tng Hp TP. HCM, i Hc Thy Sn, i Hc Cn Th, tp
trung vo nghin cu, sn xut v ng dng cng ngh ny. Tuy nhin, cht
lng sn xut v nhng ng dng ca n cha c nh gi y .
pha Bc, Vin Khoa Hc Vit Nam kt hp vi X Nghip Thy c Sn

H Ni sn xut chitin v ng dng trong lnh vc nng nghip v c hiu qu
bc u.
pha Nam, Trung Tm Cng Ngh v Sinh Hc Thy Sn phi hp vi mt

s c quan khc nh: i Hc Y Dc TP. HCM, Phn Vin Khoa Hc Vit
Nam, Vin Khoa Hc Nng Nghip Min Nam v ang nghiu cu, sn xut
v ng dng chitin trong cc lnh vc: nng nghip, y dc v m phm.
1.3.1.2.
-
Tnh hnh sn xut v nghin cu ngoi nc
Nm 1972, hng Kyowa Oid and Fat ca Nht ln u tin a vo sn xut

cng nghip chitin.
Nm 1977, Vin k thut Masachusetts (M), khi tin hnh xc nh gi tr ca

chitin v protein trong v tm, cua, cho thy vic thu hi cc cht ny rt c
li nu s dng trong cng nghip, phn chitin thu c, c dng sn xut ra
cc dn xut c nhiu ng dng khc nhau.
Sn lng chitin 1990 trn th gii l 1200 tn. Nc s dng hng u l Nht
(600 tn/nm) v M (400 tn/nm). Ngoi ra cc nc nh Trung Quc, n
, Php cng ang trin khai thm cc c s sn xut qui m 50kg
chitin/ngy vi gi bn ra l 200 300France/kg. M, hng nm tng gi tr
11
L TN PH
v cc ch phm chitin chitosan s dng l 355 triu USD, trong 190 triu
thuc ngnh y t, sau nng nghip (54 triu) v m phm (50 triu). Theo
FAO, nhu cu chitin chitosan c th ln ti 36.700 tn/nm trong thp k ti.
-
Cc phng th nghim ca Malaysia v cng nghip lm ngt nc bin

thnh lp cng ty lin doanh Seafresh chitosan khai thc kh nng cung cp
thng phm cc cht dn xut ca tm. Cng ty ny c bit quan tm vi
chitin v polisaccharide t v tm thi b v hng hot ng chnh vo cc th
trng xut khu.
Theo tin s Arisol Alimuniar gim c k thut, ng hy vng sn xut

khong 150 180 triu sn phm chitin chitosan/nm gp phn cng cc
nc sn xut chnh khc nh Nht Bn, M, Na Uy, Canada v Nga. Gi c
ca cc loi sn phm chitin chitosan bin ng t 30 USD n 400 USD/kg
tu thuc vo cht lng sn phm, lng bn chitin v chitosan t khong 2
t USD.
Nhiu nc nh Nht, M, Anh, Hi chitin thuc cng ng Chu u

ECCHIS v ang nghin cu mt cch c h thng v cp nhiu ni
dung khoa hc trong c vic ng dng chitin nh mt cht hp ph trao i
ion tinh ch nc gii kht.
Hin nay, c khong 10 cng ty ln, hu ht Nht, sn xut chitin trn th

gii. Cng ty Protan biopolymer, mt trong nhng cng ty ln trn th gii sn
xut chitin chitosan nghin cu ra nhiu sn phm c ngun gc chitin s
dng x l nc, kh cc ion kim loi c, bc ht v nhiu ng dng khc
trong nng nghip.
Ngy nay, ngi ta tp trung vo cc dn xut ca chitin v kh nng ng dng

ca nhng dn xut ny. Ton b qu trnh hot ng khoa hc ca R.A.A.
Muzzarelli (i Hc Y Khoa Ancona ) tp trung vo chitin v dn xut ca
n. Cho n nay, trn th gii c nhiu quy trnh sn xut chitin, vi nhiu
ngun nguyn liu khc nhau, nhng ch yu l v tm, cua, gh.
1.3.2. Chitosan
12
Tnh hnh sn xut v nghin cu trong nc
1.3.2.1.
-
L TN PH
Vit Nam, vic nghin cu v sn xut chitin, chitosan v ng dng ca

chng trong sn xut phc v i sng l mt vn tng i mi m nc
ta. Vo nhng nm 1978 1980, trng i hc Thy sn Nha Trang cng
b quy trnh sn xut chitosan ca tc gi Minh Phng m u bc
ngoc quan trng trong vic nghin cu tuy nhin cha c ng dng no thc
t trong sn xut.[4]
Tnh hnh sn xut v nghin cu ngoi nc
1.3.2.2.
-
Chitosan c chp nhn s dng nh mt ph gia thc phm hay thc phm
b sung, cc quc gia nh Nht, Anh, Hoa K, Italy, B o Nha, v Phn
Lan.
Nm 1992 chitosan c cng nhn l thc phm c li cho sc khe bi S y

t Nht Bn. H xc nh rng chitosan mang nhng thuc tnh cn thit trong
thc phm b sung, k c nga bnh, tng cng tui th, tng min dch, v
iu khin nhp sinh hc. FDA khng ph duyt vic s dng chitosan nh
thuc, n c dng trong thc phm b sung.[5]
Th trng chitosan c gi 5USD/kg cho sn phm nng nghip v 200USD/kg

cho sn phm cht lng cao, siu tinh khit dng trong chm sc sc khe.
mt ng dng quan trng ca chitosan l trong cng nghip thc phm dinh
dng.
1.4.
Cu trc ha hc
1.4.1. Cu trc ha hc ca chitin

-
Cu trc ca chitin l tp hp cc monosacharite (N-acetyl--D-glucosamine)

lin kt vi nhau bi cc cu ni (1-4) glucoside v hnh thnh mt mng cc
si c t chc.
13
L TN PH
Hnh 1. 2 Cu trc ha hc N-acetyl-D-glucosamine lin kt vi nhau

bng cu ni (1-4) glucoside
-
Nhiu cc monosacharit ni vi nhau to thnh chitin.
Hnh 1. 3 Cu trc ha hc chitin

-
Cng thc ho hc ca chitin (C8H13NO5)n, trong :

C = 47.29 %.
H = 6.45 %.
N = 6.89 %.
O = 39.37 %.
Phn t lng: MChitin =(203,09)n.
Bng phng php nhiu x tia X ngi ta c th chng minh c chitin tn

ti 3 cu hnh: chitin, chitin, chitin. Cc dng ny ch khc nhau v
hng sp xp ca mi mt xch (N-Acetyl-D-glucosamine) trong mch. V
ngi ta biu din mi mt xch ny bng mi tn ch nhm CH2OH, phn
14
L TN PH
ui ch nhm NHCOCH3, th cc cu trc chitin, chitin, chitin

c m t nh sau.
Chitin
-
Chitin
Chitin
Chitin c cu trc cc mch c xp xp ngc chiu nhau u n nn

ngoi lin kt hydro trong mt lp v h chui, n cn c lin kt hydro gia
cc lp do cc chui xp k nhau nn rt bn vng. Do mt xch xp xp o
chiu, xen k thun li v mt khng gian v nng lng. y cng l dng
nng lng ph bin trong t nhin.
Chitin a mt mt xch ghp vi nhau theo kiu song song, gia cc lp

khng c lin kt hydro. Dng chitin cng c th chuyn sang dng
chitin nh qu trnh acetyl ha cho cu trc tinh th bn vng hn.[1]
Chitin a mt mt xch ghp vi nhau theo kiu v hai song song mt ngc
chiu, gia cc lp khng c lin kt hydro.
Chitin c xem l dn sut ca cellulose, trong gc OH v tr C2 c

thay th bi gc acetamine NHCOCH3. im ging nhau ca lin kt glucoside
chitin v cellulose c th hin qua nhit lng ta ra tng ng nhau vo
khong 28Kcal/mol.[13]
1.4.2. Cu trc ha hc ca chitosan

-
Chitosan l dn xut acetyl ho ca chitin, trong nhm (-NH2) thay th

nhm (-COCH3) v tr C2. Ni mt cch tng th hn chitosan l s kt hp
ngu nhin gm -(1-4)-D glucosamine v N-acetyl-D glucosamine, do vy
chitosan c th gi l poly -(1-4)-2-amino-2-deoxi-D-glucoz hoc l poly (1-4)-D-glucozamin.
15
L TN PH
Hnh 1. 4 cu trc chitosan

1.5.
Tnh cht ha l sinh hc
1.5.1. Tnh cht l ha

-
Chitin c mu trng hay mu trng pht hng, dng vy hoc dng bt, khng
mi, khng v, khng tan trong nc, trong mi trng kim, axit long v cc
dung mi hu c nh ete, ru Nhng tan trong dung dch c nng ca
mui thioxianat liti (LiSCN) v thioxianat canxi (Ca(SCN)2) to thnh dung
dch keo, tan c trong h dimetylacetamid-LiCl 8%, tan trong hexafluoroisopropyl
alcohol
(CF3CHOHCF3)
hexafluoracetone
sesquihydrate
(CF3COCF3.H2O). C th tan trong Ordinethylactamine (DMA) c cha 8%

lithium choloride hoc axit m c, khng tan trong nc, axit long, st, cn
hoc cc dung mi hu c khc.[1]
-
Chitin c cu to tinh th bn vng, him thy trng thi t do m thng lin

kt vi protein bi ni cng ho tr di dng chitin protein phc hp.
Chitin khng tan trong alcol, trong dung dch acid v kim long hay m c,
v cc dung mi thng thng.
Chitin c th tan trong H2SO4 m c, H3PO4 78 97% v acid formide khan.
Trong dung dch HCl, chitin c trin quang thay i t -140 n +560. S
thay i ny chng t c s thy gii chitin.
phn tn ca dung dch keo chitin trong cc dung dch mui trung tnh ngm
nc tng i cao.[15]
Chitin c kh nng hp thu tia hng ngoi c bc sng 884 890 cm-1.
16
L TN PH
Chitin n nh vi cc cht oxy ho mnh nh thuc tm (KMnO4), oxy gi

(H2O2), nc javen (NaOCl NaCl) li dng tnh cht ny m ngi ta s
dng cc cht oxy ho trn kh mu cho chitin
Khi un nng trong dung dch NaOH m c (40 50%), nhit cao th
chitin s b kh mt gc acetyl to thnh chitosan.
-CH2OH
Chitin
-CH2OH
-CH2OH
-OH
Chitosan
-NHCOCH3
-
-OH
-NH2
Khi un nng trong axit HCl m c, nhit cao th chitin s b ct mch

thu c glucosamin.
-CH2OH
Chitin
-CH2OH
-CH2OH
-OH
Chitosan
-NHCOCH3
-OH
-NH2
Phn ng este ha
-
Chitin tc dng vi HNO3 m c cho sn phm chitin nitrat.
Chitin tc dng vi anhydrit sunfuric trong pyridin, dioxin v N,

N-dimetylanilin cho sn phm chitin sunfonat.
Chitosan phn ng vi acid m c to mui kh tan, tc dng vi I2 trong mi

trng H2SO4 cho phn ng mu tm, chitosan d phn ng hn chitin.[13],[16]
Trong phn t chitin/chitosan v mt s dn xut ca chitin c cha cc nhm

chc m trong c cc nguyn t O2 v N2 ca nhm chc cn cp electron
cha s dng, do chng c kh nng to phc, phi tr vi hu ht cc kim
loi nng v kim loi chuyn tip nh: Hg2+, Cd+2, Zn2+, Cu2+, Ni2+, Co2+ Ty
nhm chc trn mch polymer m thnh phn v cu trc ca phc khc nhau.
1.5.2. Tnh cht sinh hc

1.5.2.1.
-
Chitin
L mt vt th mu trng v nh hnh khng mi.
17
L TN PH
Trong t nhin chitin tn ti ph bin trong v ngoi ca cc lai nm khun

thc vt cp thp, cc lai ng vt gip xc nh tm, cua, cn trng v trong
mng t bo ca cc ng vt cao cp v.v
y l mt lai amylose tuyn tnh cao phn t, tc l mt loi amylose keo

trung tnh, cht chitin khng hot ng trn ha hc, khng thay i trong dch
c th, khng phn ng o thi vi c th, khng c t, c tnh khng huyt
khi, c im chu kh c nhit cao.
1.5.2.2.
Chitosan
Vt liu chitosan c ngun gc t nhin, khng c, dng an ton cho ngi.
Chng c tnh ha hp sinh hc cao vi c th, c kh nng t phn hu sinh

hc.
Chitosan c nhiu tc dng sinh hc a dng nh: c kh nng ht nc, gi

m, tnh khng nm, tnh khng khun vi nhiu chng loi khc nhau, kch
thch s pht trin tng sinh ca t bo, c kh nng nui dng t bo trong
iu kin ngho dinh dng, tc dng cm mu, chng sng u.
Chitosan khng nhng c ch cc vi khun gram dng, gram m m c nm

men v nm mc. Kh nng khng khun ca chitosan ph thuc mt vi yu t
nh loi chitosan s dng ( deacetyl, khi lng phn t), pH mi trng,
nhit , s c mt ca mt s thnh phn thc phm. Kh nng khng khun
ca chitosan v dn xut ca n c nghin cu bi mt s tc gi, trong
c ch khng khun cng c gii thch trong mt s trng hp. Mc d
cha c mt gii thch y cho kh nng khng khun i vi tt c cc i
tng vi sinh vt, nhng hu ht u cho rng kh nng khng khun lin quan
n mc hp ph chitosan ln b mt t bo. Trong , chitosan hp ph ln
b mt vi khun gram m tt hn vi khun gram dng.
Mt s c ch khng khun ca chitosan c gii thch nh sau:

-
Nh tc dng ca nhng nhm NH3+ trong chitosan ln cc v tr mang in m

trn mng t bo vi sinh vt, dn ti s thay i tnh thm ca mng t bo
lm cho qu trnh trao i cht qua mng t bo b nh hng. Lc ny vi sinh
18
L TN PH
vt khng th nhn cc cht dinh dng c bn cho s pht trin bnh thng
nh glucose dn n mt cn bng gia bn trong v bn ngoi mng t bo v
cui cng dn n s cht ca t bo.
-
Chitosan c th ngn cn s pht trin ca vi khun do c kh nng ly i cc

ion kim loi quan trng nh Cu2+, Co2+, Cd+... ca t bo vi khun nh hot
ng ca cc nhm amino trong chitosan c th tc dng vi cc nhm anion
ca b mt thnh t bo. Nh vy, vi sinh vt s b c ch pht trin do s mt
cn bng lin quan n cc ion quan trng.
in tch dng ca nhng nhm NH3+ ca glucosamine monomer pH < 6,3

tc ng ln cc in tch m thnh t bo ca vi khun, dn n s r r cc
phn t bn trong mng t bo. ng thi gy ra s tng tc gia sn phm
ca qu trnh thu phn c kh nng khuch tn bn trong t bo vi sinh vt vi
AND dn n s c ch mARN v s tng hp protein t bo.
Chitosan c kh nng ph hu mng t bo thng qua tng tc ca nhng

nhm NH3+ vi nhng nhm phosphoryl ca thnh phn phospholipid ca
mng t bo vi khun.
Ngoi ra, chitosan cn c tc dng lm gim cholesterol v lipid mu, lm to vi

ng mch v h huyt p, iu tr thn mn tnh, chng ri lon ni tit.
Chitosan l cht thn m c kh nng hp th du m rt cao c th hp thu n

gp 6 8 ln trng lng ca n. Chitosan nh phn t c in tch dng nn
c kh nng gn kt vi in tch m ca lipid v acid mt to thnh nhng cht
c phn t ln khng b tc dng bi cc men tiu ha v do khng b hp
th vo c th m c thi ra ngoi theo phn qua lm gim mc
cholesterol nht l LDL-cholesterol, acid uric trong mu nn c th gip ta
trnh cc nguy c bnh tim mch, bnh gt, kim sot c tng huyt p v
gim cn.
Vi kh nng thc y hot ng ca cc peptide-insulin, kch thch vic tit ra

insulin tuyn ty nn chitosan dng iu tr bnh tiu ng. Nhiu
cng trnh cng b kh nng khng t bin, kch thch lm tng cng h
19
L TN PH
thng min dch c th, khi phc bch cu, hn ch s pht trin cc t bo u,
ung th, HIV/AIDS.
-
Chitosan chng tia t ngoi, chng nga.[14]
c tnh
dng trong thc phm v y t phi m bo khc khe v tnh an ton ca
vt cht, nn c nhiu cng trnh nghin cu v c tnh ca chitosan v a
ra cc kt lun sau. [15]
-
Chitosan hu nh khng c, khng gy c trn xc vt thc nghim v ngi,

khng gy c tnh trng din.
Chitosan l vt liu ho hp sinh hc cao, n l cht mang l tng trong h

thng vn ti thuc, khng nhng s dng cho ng ung, tim tnh mch,
tim bp, tim di da, m cn ng dng an ton trong ghp m.
Chitosan vi trng lng phn t thp tim tnh mch, khng thy c tch
ly gan. Loi chitosan c DD = 50 %, c kh nng phn hu sinh hc cao, sau
khi tim vo bng chut, n c thi tr d dng, nhanh chng qua thn v
nc tiu, chitosan khng phn b ti gan v l lch.
Nhng li im ca chitosan: tnh cht c hc tt, khng c, d to mng, c

th t phn hy sinh hc, ho hp sinh hc khng nhng i vi ng vt m
cn i vi cc m thc vt, l vt liu y sinh tt lm mau lin vt thng.
Chitosan khng c hoc c tnh rt thp trn xc vt thc nghim v n c

th c s dng an ton trn c th ngi.[13],[15]
20
L TN PH
CHNG 2. CC PHNG PHP SN XUT CHITIN

CHITOSAN
2.1.
Phng php ha hc
2.1.1. S lc v phng php

-
Qu trnh tch chit v lm sch chitin c tin hnh theo phng php ha
hc thng thng c p dng t trc ti nay trong nhiu phng th nghim
trn th gii cng nh Vit Nam thu nhn chitin lm nguyn liu sn xut
chitosan t ph phm thy sn, qu trnh gm 3 giai on chnh l loi khong,
loi protein v kh mu. V sn xut chitosan ngi ta tin hnh thm bc
deacetyl ha bng dung dch kim.[4]
2.1.2. Quy trnh k thut
21
L TN PH
Ph phm u tm,
v cua(cc loi
gip xc)
HCl 5%, V/W = 20,
nhit phng, trong
Loi canxi
12 24h.
Ra trung tnh
NaOH 4%, 100oC,
thi gian 2 gi
Loi protein
Ra trung tnh
Kh mu bng H2O2 5%, thi
gian 15 30 pht. Bng NaOCl
5, thi gian 15 30 pht.
Bng KmnO4 0,1% v C2H2O4.
Kh mu
Chitin
Dung dch NaOH 35 50%,
90 100oC, thi gian 2 3 gi
Deacetyl ha
Chitosan
-
S quy trnh sn xut chitin.
2.1.3. Thuyt minh quy trnh

Bc 1: X l nguyn liu vi acid
-
Trong v tm, thnh phn khong ch yu l mui CaCO3 v rt t Ca3(PO4)2.

Nn ngi ta thng dng cc loi acid nh HCl, H2SO4 kh khong. Khi
kh khong, nu dng HCl th cho hiu qu cao hn. Nu dng H2SO4 s to
22
L TN PH
mui kh tan khi lu chuyn nc ngm chm. Phn ng ca HCl kh

khong photphat caxni v cacbonate canxi nh sau:
CaCO2 + HCl
CaCl2 + CO2 + H2O.
Ca3(PO4)2 + 6HCl
3 CaCl2 + 2 H2PO4
Cho mt lng va dung dch acid HCl 2,74 N vo nguyn liu c ra

s b bng nc my nhm loi b bt tht, ngm trong 12 24 gi. Sau ra
bng nc my ti khi pH trung tnh, cng on ny c tc dng ra tri ht
lng CaCl2 to thnh, loi b HCl d bi v lng HCl nu cn n s trung
ho lm hao tn lng NaOH cng on kh protein hay deacetyl. Trong qu
trnh ra cng c phn no sc t b ra tri.
Bc 2: X l vi kim
-
Ngm sn phm va thu c vi dung dch NaOH 4% nhit phng trong

thi gian ngn ( 15 30 pht), un si 1 2 gi ri ra sch bng nc my
n pH trung tnh.
Bc 3: Kh mu
-
Cc cht mu trong sn phm chitin th c ty trng u tin bng cch dng

dung dch thuc tm KMnO4 nng 0,1%, ngm khong 15 pht v vi
C2H2O4 khong 1 2 gi cho n khi sn phm c mu trng th ly ra ra sch
bng nc my, phi kh hoc sy di 50oC. Ta thu c chitin sch.
Bc 4: Deacetyl ho
-
Chitosan l dn xut quan trong nht ca chitin. Chitosan c iu ch bng

cch tch gc acetyl khi nhm acetamid v tr C2 v hnh thnh nhm amino
nh phn ng deacetyl ha chitin.
Dng xt m c 35 50% deacetyl ha. Mc deacetyl ha ph thuc

vo: Nng xt 40%, un trc tip nhit 90 120oC, thi gian 5 12 gi.
Thu c chitosan, c im nhn bit chitosan c tnh cht dai hn chitin, mu
sc trng c. [1],[4]
Phng trnh deacetyl ha.
23
L TN PH
Hnh 2. 1 Qu trnh deacetyl ha bng phng php ha hc

2.1.4. u nhc im
2.1.4.1.
-
u im
Thi gian to ra sn phm rt nhanh ch trong 1 ngy, tit kim c thi gian
v sc lao ng.
S dng nhng ha cht tng i r tin v thng dng. T chi ph cho qu

trnh khng cao.
Quy trnh kh n gin, khng i hi k thut cao.
Trang thit b thng dng, c th dng dng c ni tr. Do khng tn km

nhiu trong u t trang thit b.
2.1.4.2.
-
Nhc im
iu quan trong nht c th k n l cht thi ca phng php ha hc ny

c kh nng gy nhim mi trng cao, c hi i vi con ngi vt nui,
ng vt thy sinh.
Hiu qu trch ly khng cao di 10%.
24
L TN PH
Cht lng sn phm km, cht lng khng ng iu gia cc m trch ly.
Kh khn trong vic kim sot qu trnh. [10]
2.2.
Phng php ha sinh kt hp
2.2.1. S lc v phng php

-
Nh chng ta c bit nc ta da c trng rt nhiu, v ngoi cc mc

ch s dng qu da th trong qu da cn cha hm lng enzyme bromeline
rt cao enzyme thy phn protein. Hm lng acid hu c cng cao, bng
chng l nc da hay dch p b da rt chua v nu ta b sung dch ny vo
khi nu tht th tht mn rt nhanh.
Tn dng hai tnh cht tt ny ca qu da thay th HCl v NaOH cho vic loi
b protein v cc khong cht trong nguyn liu ph thi. Ton b thi gian
thc hin quy trnh ny ko di 8 ngy.
25
L TN PH
Ph phm u tm, v
cua(cc loi gip
xc)
Ra loi tht
Nc p b da. T l
300ml/100g nguyn
liu
Loi khong, protein
Ra sch thu cn
NaOH 12% ,90oC,
thi gian 45 pht
Loi protein ln 2
Ra trung tnh
Kh mu bng H2O2 5%, thi gian
15 - 30 pht. Bng NaOCl 5, thi
gian 15 30 pht. Bng KMnO4
0,1% v C2H2O4
Kh mu
Chitin
Dung dch NaOH 35 50%,

90 100oC, thi gian 2 3 gi
Deacetyl ha
Chitosan
S quy trnh sn xut chitosan bng phng php ha sinh kt hp.

Bc 1: X l nguyn liu vi enzyme bromalin
26
L TN PH
Nguyn liu c ra s b loi bt tht ri cho enzyme vo x l loi

khong v thy phn protein, mt ngy thay enzyme 2 ln cch sau mi 12 gi
ng h. C x l nh vy 6 7 ngy, thu cn v ra sch.
Bc 2: X l vi kim
-
Protein c thy phn trong mi trng acid nn cn hin tng ta nn cn

x l vi NaOH 1 2% ,90oC, thi gian 45 pht nhm mc ch ho tan protein
v tin hnh ra s tng hiu qu ca giai oan loi protein hn. T cho sn
phm sch hn.
Bc 3 kh mu, bc 4 deacety ha
-
Do y l phng php kt hp ha sinh nn cc hai bc ny hon ton ging

nh phng php ha hc.
2.2.4. u nhc im
2.2.4.1.
-
u im
Hiu sut thu hi sn phm bng phng php enzyme cao hn so vi phng
php ha hc. Tng ng khong 7,45% v 10,4%.
Cht lng sn phm thu c bng phng php enzyme cng tt hn so vi

phng php ha hc.
Lng ha cht ch yu trong qu trnh tch chit chitin l HCl v NaOH c

s dng t hn, t tit kim c ho cht v lng nc ra sau x l acid
v kim. Mc khc, trong qu trnh loi protein trong phng php ha sinh kt
hp ny hon ton dng enzyme protease t dch p v da, m khng cn s
dng HCl trc . Lng NaOH tiu tn cn gim i nhiu ln ( t 24 37g
gim xung 4,2g/100g nguyn liu) v nng NaOH cng gim i 1% hoc
2% l t yu cu.
t gy nhim mi trng t cht thi.
Thit b s dng t hn, iu ny mang li ngha kinh t ln i vi quy trnh

cng ngh khi pht trin to ra sn phm thng mi.
2.2.4.2.
Nhc im
27
L TN PH
Hn ch ca phng php l thi gian thu nhn sn phm lu hn 7 8 ngy,

trong khi phng php ha hc ch 1 ngy l cho ra sn phm. Qu trnh t
hn nhng tiu tn nhiu cng lao ng hn.[8],[9]
2.3.
Tiu tn nhiu cng lao ng hn, do c nhiu tin trnh hn.

Phng php sinh hc
2.3.1. S loc v phng php

-
Qua tt c cc cng trnh nghin cu v chitin chitosan, d thy r rng

nguyn tc chung ca sn xut chitin chitosan kh khong bng acid v c,
kh protein bng kim. Trong phng php sinh hc ny s tin hnh thay tc
nhn kh khong bng acid hu c sinh ra qua qu trnh sng ca vi khun
lactic, v chnh qu trnh sng s l tc nhn thy phn protein trong nguyn
liu v tm.
Chng vi khun c s dng trong phng php ny l Lactobacillus

acidophilus ly t ngun dc phm. ANTIBIO ca Hanwha Pharm CoKOREA, mang chng vi sinh L. acidophillus, mt 109 cfu/g.
S lc v Lactobacillus acidophillus
-
Phn loi:
Gii: Bacteria
Ngnh : Firmicutes
Lp: Bacilli
H: Lactobacillaceae
Chi: Lactobacillus
Loi: Lactobacillus acidophillus
im c trng ph bin phn bit chng so vi phn ln cc chi khc l dng

hnh que v kh nng sinh acid lactic nh mt sn phm cui cng ch yu.
Ngoi ra, Lactobacillus cn l vi khun Gram dng, khng hnh thanh bo t
v him khi di ng. Ngi ta thy rng t bo Lactobacillus in hnh c dng
hnh que, vi kch thc bin i trong khong (0,5 1)(1 10)m, i khi
28
L TN PH
gn ging dng hnh cu trong iu kin no v thng hnh thnh dng

chui hoc tn ti c lp.
-
Nhit pht trin ti u khong 30 40oC nhng chng c th sinh trng

trong phm vi 5 53oC.
c tnh quan trng nht ca chng vi sinh ny l kh nng sinh acid lactic
mnh, v chu c mi trng c pH thp. Khong pH ti u cho chng pht
trin l 5,5 5,8, nhng nhn chung chng c th sinh trng pH < 5.
Ngoi ra ngi ta c th da trn cc sn phm cui ca qu trnh bin dng

phn loi cc loi Lactobacillus thnh 2 loi. Cc loi ln men ng hnh v
cc loi ln men d hnh. Cc loi thuc nhm ln men ng hnh to ra 85%
acid lactic t glucose, trong khi loi ln men d hnh ch to ra 50% acid lactic
v mt lng ng k carbon dioxide, acetate v ethanol. L. acidophilus l mt
chng ln men ng hnh c trng nht.
Lactobacillus acidophilus: L. acidophilus l thnh vin ca mt trong tm chi

chnh ca vi khun axit lactic. Mi chi v cc loi c c im khc nhau nhng
ni chung chng l nhng cu khun dng que, Gram (+) , khng di ng , s
dng ng glucose, aesculin, cellobiose, galactose, lactoza, mantoza, salicin
v sucrose, vi khun khng hnh thnh bo t, ym kh, sn xut axit lactic l
mt sn phm ch yu hoc duy nht ca s trao i cht ln men lactose v s
dng lm ngun carbon sn xut nng lng . Ngoi ra qu trnh ng ha
cn to ra lipase, amylase, protese v c bit l acidophilin. L. acidophilus c
th sng trong mi trng c tnh axit cao pH 4 5 hoc thp hn v c m
t nh l mt v d loi ln men ng hnh sn xut axit lactic l sn phm duy
nht (Vijayakumar et al., 2008).
29
L TN PH
10,5% ng.
0,875% CH3COONa
V tm s
Ra s b
0.35% KH2PO4
Phi trn nguyn liu
T l nc : v tm
2,5:1
Chnh pH 6.5
Gia nhit 80oC, 15p
Cy ging trn 15%
Ging Lactobacillus
acidophilus.
Ln men 72h 96h,

37oC. ym kh, khuy
Ra
Ty mu bng H2O2
Sy kh 50 65oC
Chitin
-
Quy trnh kh khong v kh protein trn v tm s sn xut chitin ngun

nguyn liu sn xut ra chitosan.

Bc 1: Ra s b v tm
30
L TN PH
Mc ch
-
Loi b bt mt phn vi sinh gy thi c sn trong v tm, loi b tp cht nh

rc, t, ctv mt s tht tm b thi ra (gim bt mi).
Tin hnh
-
Cn mt lng xc nh v tm, mang i ra trc tip bng nc sinh hot bnh

thng, ra qua mt ln l .
Rng vt v tm ln ro nc.
Ch
-
Khng ngm v tm qu lu trong nc.
Khng ra nhiu ln gy hao tn nc m cn ra tri ht phn tht xt li trn

v tm, phn tht ny l ngun cung cp nit cho vi khun duy nht trong qu
trnh. Nu thiu th qu trnh kh thnh cng.
Nn dng thao chu sch ra, s gim thiu ti a ngun nhim vi sinh vt
gy thi.
V tm ra xong cn ni kh ro hoc cho trc tip vo ni ln men, xa

ngun nhim bn.
Bc 2: Phi trn nguyn liu

-
Qua tm hiu cc bi bo khoa hc, v cc ti nghin cu ca kha trc.

Tuy nhin hu nh cn mt s vn cha gii quyt tt nn hiu sut qu
trnh cha cao. Da trn nn tng ti tin hnh thc nghim sn xut vi mt
s ci bin t cng thc gc nh sau.
Mc ch
-
y l mt bc kh quan trng, c th ni n quyt nh qu trnh x l c tt

hay khng, lng acid sinh ra cng nhiu th hiu qu x l cng cao.
Mc nh ca bc ny l nhm to mt mi trng ti u nht cho vi khun

pht trin, sinh ra lng acid ln nht, pH gim thp nht c th.
B sung cc thnh phn khc m c th c ch cc vi sinh vt khc pht trin,

gim thiu cnh tranh mi trng sng cho chng vi khun ta cy ging.
Tin hnh
31
L TN PH
Cn mt lng nguyn liu v cc cht theo bng sau:

Bng 2. 1 Cng thc phi trn nguyn liu ln men
Quy v % so vi v tm
Nc
250%
Sucrose
10,5%
CH3COONa
0,875%
KH2PO4
0,35%
Ch
-
Ngun nc s dng trong quy trnh phi l ngun nc sinh hot hoc tt hn.
Dng cc ngun nc khc s khng kim sot c cht lng nc, nu nc
b nhim gy nh hng qu trnh.
Cn thn l iu cn thit, nn cn thn ly v cn ng ha cht.
Bc 3: Xay v tm
Mc ch
-
Nhm ng ha cc cht trong nguyn liu.
V tm c xay nh s tng b mt tip sc, phn b u trong mi trng.
Tin hnh
-
Dng my xay bnh thng hoc c th bm bng tay, tuy nhin cn t yu

cu l v tm cng nhuyn cng tt, ti thiu 2cm.
Bc 4: Gia nhit
Mc ch
-
Gim ti thiu lng vi sinh vt tp nhim c trong mi trng ln men.
i vi vi sinh vt sinh bo t th lm cho chng chm pht trin li.
To mi trng tng i sch, u th cho L.acidophilus.
Tin hnh
-
Ci t thit b hot ng gia nhit 80oC trong 15 pht.
Ch
-
Cn m bo thi gian v nhit gia nhit.
32
L TN PH
Cn kim tra bo tr thit b trc khi vn hnh m bo an ton v hiu qu.
Bc 4: Cy ging
Mc ch
-
Cung cp sinh khi ging khi ng cho qu trnh tng trng vi khun.
To qun th vi sinh vt pht trin cnh tranh v c ch vi sinh tp khc.
L tc nhn sinh acid v thy phn protein trong qu trnh sng.
Tin hnh
-
Ging c tng sinh 24h 37oC trn mi trng MRS. C mt trn 3,5105
cfu/ml.
Cy ging trc tip vo mi trng ln men. 15% th tch ging (so vi khi
lng v tm).
Ch
-
Phi m bo mt v th tch cn cy.
Thao tc cy ging v trng.
Bc 5: Ln men
Mc ch
-
Duy tr cc iu kin nh nhit , iu kin ym khto iu kin cho vi

sinh pht trin trong sut thi gian cho ti khi kt thc.
Tin hnh
-
Sau khi cy ging to iu kin ym kh, nhit 37oC duy tr 72 96h, c

khuy o theo chu k 5pht khuy 10 pht ngh.
Ch
-
Kim tra thit b trc khi ln men.
Kim tra h thng c hot ng u trong sut thi gian ln men. V quan st
24h u c sinh kh (kh ny c th do vi sinh gy thi sinh ra, v sau khi x b
kh ny i th cho ti lc kt thc ln men khng thy sinh kh na). Nu 24h
u sinh kh nhanh, x kh t th.
Nu chy th chc chn b nhim vi sinh gy thi sinh kh CH4 ch thm vi

gi na xem chng vi sinh ta cy vo c c ch c khng ( thng l khng
33
L TN PH
chng sinh CH4 ny kh mnh) v nu cn sinh kh v t chy na th nn b

ngay m ln men, tip tc s rt thi. Nu khng cn kh sinh ra th chng t
chng vi khun ta cy vo c ch c, cho ln men tip.
-
Nu khng chy, th ch thm vi gi xem cn sinh kh khng, nu cn cng

tin hnh loi b m ln men, nu khng th chng t chng vi khun ta cy vo
c ch c v tip tc ln men.
Quyt nh cho ln men tip th canh thi im 40 48h sau ln men tin hnh
o pH thy c gim th cho ln men tip, nu tng th tin hnh loi b m ln
men.
Bc 6: Ty mu
- Sau ln men tin hnh vt phn v tm ra khi dch ln men, tt hn nn ra
nc cng on ny ri mi ty mu, ty mu xong s ra sch, tuy nhin cn
tit kim ti a chi ph, vy nn khng ra m i tin hnh ty mu trc tip. C
hai nguyn nhn th nht l tit kim chi ph s dng nc, th hai trn v tm
cn nhiu sinh khi vi sinh vt, mc d y l vi sinh vt ng rut an ton
vi ngi v vt nui, tuy nhin nn c bin php loi tr vn an ton hn. V
vy khi cho vo ty mu vi H2O2 l mt cht oxy ha mnh, kh nng dit
khun rt cao.
Mc ch
-
V tm sau ln men s c mu sc vng nht, nh lm gim gi tr cm

quan, gim cht lng chitosan khi dng lng chitin ny lm nguyn liu sn
xut. Do cn ty mu cho ra chitin trng hn, to gi tr cm quan tt hn.
Tin hnh
-
Dng H2O2 nng cao, cho v tm vo ngm trc tip 2 3h, lng s
dng sao cho ngp u v tm.
Tin hnh khuy o sau mi 15 pht ngm.
Thu hi lng H2O2 c th ti s dng li.
Ch
34
L TN PH
H2O2 l cht oxy ha mnh, nguy him i vi ngi s dng, khng tip xc
da tay trc tip.
Trang b cn thn bo h lao ng khi lm vic.
Bc 7: Ra
Mc ch
-
Sau ln men cc thnh phn khong lin kt trn v tm c tch ra, cc ion
khong ny ch yu l canxi kt hp vi acid lactic to thnh canxiumlactate,
thnh phn na l protein b thy phn s nm trong phn canh trng ln men
v ln nht l sinh khi vi khun sau ln men. Do chitin c tinh sch ta
cn tin hnh ra sch bng nc.
Tin hnh
-
Dng nc sinh hot bnh thng ra tri ht dch b bm li trn v tm. Ra

khong 3 ln l tng i sch.
Ch
-
V trc ta c bc ty mu vi H2O2, y l mt cht oxy ha rt mnh,

kh nng n mn da tay rt nhanh, cn thc hin cc bin php an ton khi tip
xc.
Bc 8: Sy kh
Mc ch
-
Lm kh chitin ti mt m nht nh, thng trn di 10%, nhm c ch

cc hot ng ca nm hay vi khun gy gim, gy h hng sn phm
Tin hnh
-
Chitin sau khi ra sch ro nc, mang cho vo trong t sy, ci t nhit
dao ng t 50 65oC, trong vng 4 6h.
Nu thi tit thun li nhiu nng, c th phi trc tip di nng.
Tin hnh ly mu kim tra m, t iu kin yu cu th thu sn phm.
Ch
-
Khng sy nhit qu cao, s lm xu mu sn phm.
2.3.4. Thit b
35
L TN PH
Cc thit b phc v cho quy trnh ln men c bn nh sau:

-
Mt s thit b thng dng ti phng th nghim nh pipet, cn, my o pH
ng vai tr quan trng hn l thit b ln men, c tnh ca thit b c th hp

tit trng nh mt autoclave, gi v to iu kin v trng, ym kh, hiu kh,
gi n nh mt nhit cn thit.
Hnh 2. 2 Ni ln men c s dng trong quy trnh sn xut

-
c thit k bng thp khng g, chu c acid hay baz cao, c b tr lp

thm h thng trao i nhit nhm rt ngn thi gian lm ngui sau gia nhit v
ng c khuy c th ty chnh ch khuy o theo chu k cn thit, gip
mi trng c khuy u lin tc to iu kin cho vi khun sng tt hn.
36
L TN PH
Hnh 2. 3 np ni v cc chi tit thit b

Thit b hot ng da trn mt b iu khin, cho php bn t ng ha mt
phn quy trnh sn xut.
Hnh 2. 4 B iu khin
2.3.5. u im
37
L TN PH
Trc tin phi ni n l thn thin mi trng gn gi cuc sng nh gim

thiu tc nhn gy nhim mi trng thng qua gim cc ha cht s dng
trong phng php sn xut truyn thng.
Vic khng s dng ha cht cng ko theo chi ph cho sn xut gim xung,
tng li nhun kinh t.
X l c tc nhn nhim t ph thi v tm, v tm khng c x l s

gy thi v l mi trng cho cc vi sinh vt gy bnh pht trin, cc ng vt
k sinh nh rui, chutchng ly mm bnh vo cng ng.
Do s dng vi khun Lactobacillus acidophilus, l mt vi sinh ng rut t

s an ton hn cho cng nhn thc hnh sn xut. To ra sn phm cng an ton
hn cho ngi s dng.
S dng mt tc nhn an ton hn l vi sinh nhng gii quyt c hai vn

l kh khong v kh protein.
Dch sau ln men c cha rt nhiu thnh phn dinh dng, nn co bin php
thu hi c th lm ngun thc n chn nui giu dinh dng.
B qua nhiu bc gia nhit s tit kim c ngun nng lng kh ln so vi

phng php ha hc.
V tm sau x l trng p hn mu ng ca cc phng php khc.
2.3.6. Nhc im
-
Quy trnh tin hnh c nhiu bc i hi k thut chuyn mn hn, v d k

thut v trng, hoc cc k thut lm gim thiu ngun nhim bn t ngoi vo
nguyn liu.
Mt nhiu thi gian hn, t khu chun b ging khi ng, ti bc ra

nguyn liu cho ti thi gian ln men x l. Thng th mt 4 ngy trong khi
phng php ha hc ch mt 1 ngy.
T l thnh cng ca qu trnh ln men ph thuc nhiu yu t, vy nn vn c

m ln men tht bi gy thi. Trong khi phng php ha hc th thng lun
thnh cng.
38
L TN PH
Cht lng sn phm khng ng du, v n ph thuc nhiu vo ngun nguyn

liu v tm ban u.
Phi theo di thng xuyn trong qu trnh ln men, nhm pht hin s c tp
nhim gy h hng.
39
L TN PH
CHNG 3. KT LUN V KIN NGH

3.1.
Kt lun
- Qua tm hiu t nhiu ngun ti liu ng tin cy v thc hnh sn xut th

nghim chitin minh chng rng
-
Chitin chitosan l mt polymer sinh hc cao phn t, l thnh phn cu to v

ngoi ca mt s ng thc vt, rt an ton, c kh nng t hy, mang in
dng, c kh nng khng khun Nn c ng dng hiu qu trong nhiu
lnh vc.
Chitin c nhiu trong lp v ngoi ca cc loi gip xc nh tm, cua,

ghChitosan l sn phm c sn xut bng vic deacetyl ha chitin, gc
acetyl (y l mt lin kt kh chc chn nhng a con kh bo ca ha
sinh) nn ti ngy nay cng vic ny vn cn tiu tn kh nhiu nng lng v
ha cht thc hin deacetyl ha chitin.
Vic sn xut chitin thc hin qua hai qu trnh chnh l kh khong v kh
protein xa nay ngi ta vn dng phng php ha hc v phng php sinh
hc cng hon ton lm c iu , cn cho hiu qu cao hn. Ti nay
phng php sinh hc vn cha ti u nn tng lai s cho kt qu tt hn na.
Tn dng gn nh hon ton sn phm sau ln men t dch ln men ti phn v

tm.
3.2.
-
Kin ngh
Sn xut chitin chitosan bng phng php sinh hc cn rt nhiu b ng
cha nghin cu tm hiu r. Cn tin hnh nghin cu nhm hon thin quy
trnh sinh hc.
C th chn lc thay th Lactobacillus acidophillus bng mt chng vi sinh

khc ti u hn.
Nghin cu tm ra cc thnh phn khong b sung vo nguyn liu tng hiu

qu qu trnh ln men. Tm ra nng thch hp nht b sung.
40
L TN PH
Hin nay vn cha deacetyl ha chitin cho ra chitosan bng sinh hc, y l mt
bc tng gi tr s dng t ngun ph thi v tm, nn rt mong s c cng
trnh nghin cu cho vn ny.
Nghin cu thm quy trnh thu hi caxium lacatate trong canh trng ln men.
V thu hi c phn canh trng ln men lm thc n gia sc rt giu m.
41
L TN PH
Ti liu tham kho:

Ting vit:
1. Phm l Dng, Trnh Bnh, Li Thu HIn cng cc cng s (1997). Vt liu
sinh hc t chitin, Vin Ha Hc-Vin Cng Ngh Sinh Hoc, Trung tm
khoa hc v Cng Ngh quc gia H Ni.
2. Nguyn Hoi Hng (2009). Bi ging thc hnh ha sinh, Trng i hc
K Thut-Cng ngh, Tp.HCM.
3. Phm Th nh Hng (2003). K thut ha sinh, Nh xut bn i Hc Quc
Gia Tp.HCM.
4. Trn Th Luyn v Hunh Nguyn Duy Bo v mt s cng s (2000).
Hon thin quy trnh sn xut chitin-chitosan v ch bin mt s sn phm
cng nghip t ph liu v tm, cua, Bo co khoa hc, ti cp b, Nha
Trang.
5. Trn Th Luyn; Minh Phng; Nguyn Anh Tun (2000). Sn xut cc
ch phm k thut v y dc t ph liu thy sn, NXB Nng Nghip.
6. Nguyn c Lng (2004). Cng ngh enzyme, Nh xut bn i Hc Quc
Gia Tp.HCM.
7. Nguyn Tin Thng (2003). Mt s k thut phng th nghim sinh hc, T
sch Vin Sinh Hc Nhit i.
8. Trang S Trung (2009). nh gi cht lng sn phm v hiu qu mi
trng ca quy trnh sn xut chitin ci tin kt hp x l enzyme, Tp ch
Khoa hc - Cng ngh Thy sn, 1, 3-9.
9. Trang S Trung, V Ngc Bi, Phm Th an Phng (2007). Nghin cu
kt hp enzym protease trong cng ngh sn xut chitin t ph liu u v
tm, Tp ch Khoa hc - Cng ngh Thy sn, 3, 11-17.
10. Trang S Trung, Phm Th an Phng (2012). Tnh cht ca chitin v
chitosan t v tm th chn trng, kh protein bng phng php ha hc
v sinh hc, Tp ch khoa hc-cng ngh thy sn, 3.
42
L TN PH
11. Trn Thi Trung (2005). Nghin cu cc yu t nh hng n qu trnh

deacetyl v ct mch chitin iu ch glucosamine, tp ch khoa hc, i
hc Hu, 27.
Ting anh:
12. Rao, M. S., Munoz, J., & Stevens, W. F. (2000). Critical factors in chitin
production by fermentation of shrimp bio waste, Appl Microbio Biotechnol,
54, 808-813.
13. Rao, M. S., & Stevens, W. F (2005). Chitin production by Lactobacillus
fermentation of shrimp biowaste in a drum r eactor and its chemical
conversion to chitosan, J Chem Technol Biotechnol, 80, 1080-1087.
14. Rinaudo M (2006). Chitin and chitosan: Properties and application, Sci, 31,
603-632.
15. Sini TK , Santhosh S , et (2007) al. Research on chitin and chitosan
produced from shrimp shell by using Bacillus subtilis fermentation
Carbohydr , Res . 342, 233 242.
16. Stevens, W. F (2001). Production of chitin and chitosan: Re nement and
sustainability of chemical and biological process-ing. Chitin and chitosan
in Life Science, Proceedings 8th International, 34, 203-206.
17. Guoying Zhou and, et (2010) al. Identification of a chitin deacetylase
producing bacteria isolated from soil and its fermentation optimization,
African Journal of Microbiology Research, 4, 2597-2603.
Ngun internet:
18. http://www.chitosan.com.vn/
19. http://www.webmd.com/vitamins-supplements/ingredientmono-625chitosan.aspx?activeIngredientId=625&activeIngredientName=chitosan
20. http://doan.edu.vn/do-an/de-tai-tim-hieu-ve-chitin-chitosan-5929/
21. http://www.cyberchemvn.com/cyberchem/technology-and-apply/107chitosan-tng-quan-nghien-cu-ng-dng.html
43
L TN PH
22. http://www.banglajol.info/index.php/BJSIR/article/view/7330
23. http://www.faqs.org/patents/app/20090275745
24. http://www.ncbi.nlm.nih.gov/pubmed/21790136
25. http://www.france-chitine.com/fab.e.html
26. http://www.jstor.org/discover/10.2307/20107794?uid=3739320&uid=2&uid
=4&sid=21102453108017
27. http://www.hindawi.com/journals/er/2012/421683/
28. http://www.sigmaaldrich.com/catalog/product/sigma/c3646?lang=en&region
=VN
29. http://www.ncbi.nlm.nih.gov/pubmed/17707781
30. http://fishfarming.com/shrimp.html?gclid=CLzphbfngLgCFYgn4god63MAy
A
31. http://www.ijens.org/Vol%2011%20I%2001/110201-8484%20IJBASIJENS.pdf
32. http://www.plantphysiol.org/content/66/2/205.short
33. http://www.tandfonline.com/doi/abs/10.1080/08905439709549920
34. http://www.sciencedirect.com/science/article/pii/S1381514800000389
35. http://www.foodproductiondaily.com/Packaging/Researchers-develop-ecofriendly-packaging-using-chitosan-from-shrimp-shells
HT
44

Sản Xuất Chitin - Chitosan Bằng Phương Pháp Sinh Học

Uploaded by

Document Information

Original Title

Copyright

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Sản Xuất Chitin - Chitosan Bằng Phương Pháp Sinh Học

Uploaded by

Copyright:

B GIO DC V O TO

TRNG I HC K THUT CNG NGH TP.HCM

SN XUT CHITIN CHITOSAN

KHOA MI TRNG & CNG NGH SINH HC

TP. H Ch Minh, 2013

SN XUT CHITIN CHITOSAN

SN XUT CHITIN CHITOSAN

sinh, bn c c th khi qut kin thc c bn v chitin chitosan. Bn

SN XUT CHITIN CHITOSAN

Gii thiu ......................................................................................................7

Ngun gc thu nhn ...................................................................................10

Tnh hnh nghin cu trong v ngoi nc .............................................10

Tnh hnh sn xut v nghin cu trong nc ...............................10

Tnh hnh sn xut v nghin cu ngoi nc ...............................11

Tnh hnh sn xut v nghin cu trong nc ...............................13

Tnh hnh sn xut v nghin cu ngoi nc ...............................13

Cu trc ha hc ca chitin ................................................................13

Tnh cht ha l sinh hc ..........................................................................16

Tnh cht l ha ...................................................................................16

Tnh cht sinh hc ...............................................................................17

CHNG 2. CC PHNG PHP SN XUT CHITIN CHITOSAN .....21

Phng php ha hc ................................................................................21

S lc v phng php ......................................................................21

Quy trnh k thut ................................................................................21

SN XUT CHITIN CHITOSAN

Thuyt minh quy trnh .........................................................................22

Phng php ha sinh kt hp .................................................................25

S lc v phng php ......................................................................25

Quy trnh k thut ................................................................................25

Thuyt minh quy trnh .........................................................................26

Phng php sinh hc ...............................................................................28

S loc v phng php ......................................................................28

Quy trnh k thut ................................................................................29

Thuyt minh quy trnh .........................................................................30

CHNG 3. KT LUN V KIN NGH .........................................................40

Ti liu tham kho:...........................................................................................42

SN XUT CHITIN CHITOSAN

DANH SCH HNH

SN XUT CHITIN CHITOSAN

DANH SCH BNG

SN XUT CHITIN CHITOSAN

CHNG 1. TNG QUAN V CHITIN - CHITOSAN

Chitin chitosan l mt polysacharite tn ti trong t nhin vi sn lng rt

Trong ng vt, chitin l mt thnh phn cu trc quan trng ca cc v mt s

V c tnh sinh hc v cu trc ha hc chitin l polysacharite c m khng

tm, chitin hin din dng kt hp vi cc thnh phn khc (khong).

SN XUT CHITIN CHITOSAN

Chitin cng hin din ng vt bc thp. Chitin l thnh phn cu to nn

thy sn, chitin tn ti rt nhiu, c bit l v tm, cua, gh, nang

Mc d gp rt nhiu kh khn trong vic chit tch chitin nhng ngi ta

Thnh phn chitin (%)

SN XUT CHITIN CHITOSAN

Chitosan l mt polysaccharide tuyn tnh. Loi chitosan thng mi c MW t

mang in tch m nh acid bo, acid mt,

phospholipid, protein v polysaccharide, chnh iu ny to cho chitosan nhng

Do mt s c tnh vn c, l hp cht t nhin, khng c, c th b phn hy,

Chitosan c sn xut bng cch kh acetyl (deacetylation DD) ca chitin,

SN XUT CHITIN CHITOSAN

Hnh 1. 1 Deacetyl ha chintin thnh chitosan

Ngun gc thu nhn

S tng hp chitin loi gip xc

Billard quan st trn loi gip xc Decapoda cho bit s tng hp ca

S tng hp v phn hy chitin gip xc rt tch cc v phn ln lp v

Tnh hnh sn xut v nghin cu trong nc

Vic nghin cu, sn xut chitin v cc ng dng ca chng trong sn xut,

SN XUT CHITIN CHITOSAN