Sản Xuất Chitin – Chitosan Bằng Phương Pháp Sinh Học

7/23/2019 Sn Xut Chitin Chitosan Bng Phng Php Sinh Hc

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B GIO DC V O TO

TRNG I HC K THUT CNG NGH TP.HCM

SN XUT CHITINCHITOSAN

BNG PHNG PHP SINH HC

KHOA MI TRNG & CNG NGHSINH HC

CHUYN NGNH CNG NGHSINH HC

TP. HCh Minh, 2013


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SN XUT CHITINCHITOSAN L TN PH

i

LI NI U

Nh chng ta bit v nhng c nhn, tchc 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 quva rt an

ton v c bit l thn thin vi mi trngmt vn rt c quan tm

trn thgii hin nay. D rng chitinchitosan 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 thsn xut c chitinchitosan. Trong sn xut chitin

chitosan bn cht chung ca qu trnh vn l khcc thnh phn khong v

hm lng protein cn d tha trong phliu v tm scho ra chitin. Vy

qu trnh c thc thi ra sao? Dng tc nhn no khkhong, loi

protein thu chitin v deacetyl ha chitin thu sn phm chitosan. Viphng php ha hc ngi ta dng tc nhn ca ha cht xl, nhng

bn cnh xut hin thm nhiu vn khc cn xl 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,

chc sinh hc mi gn gi v gn lin vi ssng, stn ti ca mun loi.

Tuy nhin n chmi c manh nha trong suy ngh v ang c nghin

cu thnghim mt sphng th nghim. Trong chuyn ny stng

quan hu ht cc phng php sn xut chitinchitosan bng ha hc, ha


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ii

sinh, bn c c thkhi qut kin thc c bn vchitinchitosan. Bn

cnh c s cho php ca ban lnh o khoa Mi Trng & Cng Ngh

Sinh Hc trng i Hc KThut Cng NghTp.hcm, ng cho ti lm

vic ti phng th nghim ca trng, a ch475A in Bin Ph, phng

15, qun Bnh Thnh, Tp.hcm. Cng vi s hng dn tn tm ca ging

vin TS. Nguyn Hoi Hng vvic thc hin thnghim sn xut chitin

chitosan t phphm thy sn vtm s, nhm thc hin chuyn chn

thnh bit n v iu ny.


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iii

MC LC

LI NI U ........................................................................................................... 1

MC LC ................................................................................................................ iii

DANH SCH HNH ................................................................................................. v

DANH SCH BNG.............................................................................................. vi

CHNG 1. TNG QUAN VCHITIN - 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. Chitin.................................................................................................... 10

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. Chitosan................................................................................................ 12

1.3.2.1. Tnh hnh sn xut v nghin cu trong nc............................... 131.3.2.2. Tnh hnh sn xut v nghin cu ngoi nc ............................... 13

1.4. Cu trc ha hc ........................................................................................ 13

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

1.4.2. Cu trc ha hc ca chi tosan............................................................ 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 CHITINCHITOSAN..... 21

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

2.1.1. S lc vphng php ...................................................................... 21

2.1.2. Quy trnh kthut................................................................................ 21


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iv

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

2.1.4. u nhc im.................................................................................... 24

2.1.4.1. u im .......................................................................................... 24

2.1.4.2. Nhc im .................................................................................... 24

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

2.2.1. S lc vphng php ...................................................................... 25

2.2.2. Quy trnh kthut................................................................................ 25


2.2.4. u nhc im.................................................................................... 27

2.2.4.1. u im .......................................................................................... 27

2.2.4.2. Nhc im .................................................................................... 27

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

2.3.1. S loc vphng php ...................................................................... 28

2.3.2. Quy trnh kthut................................................................................ 29


2.3.4. Thit b.................................................................................................. 352.3.5. 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


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v

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 ....................................................................................................... 14Hnh 1. 3 Cu trc ha hc chitin ............................................................................. 14Hnh 1. 4 cu trc chitosan ....................................................................................... 16

Hnh 2. 1 Qu trnh deacetyl ha bng phng php ha hc................................ 24Hnh 2. 2 Ni ln men c sdng trong quy trnh sn xut................................. 36Hnh 2. 3 np ni v cc chi tit thit b ................................................................... 37Hnh 2. 4 Biu khin ............................................................................................. 37
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DANH SCH BNG

Bng 1. 1 Thnh phn chitin ca mt sphliu thy sn: .................................. 8

Bng 2. 1 Cng thc phi trn nguyn liu ln men ............................................ 32


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7

CHNG 1. TNG QUAN VCHITIN - CHITOSAN

1.1. Gii thiu

1.1.1.Chitin

- Chitin chitosan l mt polysacharite tn ti trong tnhin vi sn lng rt

ln (ng thhai 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 tbcnh cng m ng gi l chitin hay chiton, ting Hy

lp c ngha l vgip, nhng ng khng pht hin ra sc mt ca nit trong. Cui cng cOdier v Braconnot u i n kt lun chitin c dng cng

thc ging vi cellulose. Ngy nay chitin c pht hin cng vt v thc

vt.

- Trong ng vt, chitin l mt thnh phn cu trc quan trng ca cc vmt 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 chyu trong m

da n gip cho sti to v gn lin cc vt thng da, c bit trong cc loi

thy sn c bit l trong vtm, cua, gh, hm lng chitin chim kh cao dao

ng t1435% so vi trng lng kh, c bit l tm. Theo c tnh ca T

chc Nng Lng ThGii (FAO) sn lng tm nui trng trn thgii nm

2011 l 2.934.168 tn, ring Vit Nam 403.600 tnV vy vtm, cua, gh

l ngun nguyn liu chnh sn xut chitin. Cn trong thc vt chitin c

thnh tbo nm hzygenmyctes, cc sinh khi nm mc, mt sloi to

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

c, c khi lng phn tln.[13]

Phn b

- Chitin hin din thc vt bc thp. Trong thgii thc vt, chitin chgii hn

loi nm, to lc, a y, non khun.

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


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8

- 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 chl mt trong nhng thnh phn

cu to ca bxng ngoi. Trong thin nhin, dng chitin nguyn cht chtn

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 vtm, k l vcua, gh. vtm cng nh vcua,

chitin to phc hvi protein v thng cha thm mt tlln CaCO3.

- Chitin tn ti trong tnhin dng tinh th. Trong tnhin him thy chitin

tn ti dng tdo, n lin kt di dng phc hp chitin vi protein, chitin

vi cc hp cht v c,...Khi tn ti nh th, chitin c khnng 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 thv s thay i tng giai on sinh l m

trong cng mt loi, ngi ta c ththy c sthay i vlng v cht chitin.

-

Mc d gp rt nhiu kh khn trong vic chit tch chitin nhng ngi ta khng nh vshin 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 sphliu thy sn:

Phliu Thnh phn chitin (%)

Vhn 0,4

Vc 1,24

Vcua ng 23,8

Vtm ng 30,0

Vtm bin 33,1

1.1.2.Chitosan


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9

- 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 thca chitosan (MW: 10.000

300000 Da) c khnng hp thln 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 tnhin duy nht mang in tch dng do c

nhng nhm amino tdo 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 thtan trong

nc phthuc vo pH, nhit ca qu trnh acetyl ha.[1]

- Do mt sc tnh vn c, l hp cht tnhin, khng c, c thbphn hy,

c khnng tng tc vi nhiu phn tsinh hc nn c ng dng rng ri

trong thc t trong nhiu lnh vc nh: nng nghip, ha hc, cng nghsinh

hc, cng nghip giy, mphm, xl nc, bo qun thc phm, phgia thcphm, dc phm

- Chitosan c sn xut bng cch khacetyl (deacetylation DD) ca chitin,

mc deacetylation (% DD) c thc xc nh bng phNMR, v DD%

trong chitosans thng mi l trong khong 610%. Cht c khacetyl 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.


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10

1.2. Ngun gc thu nhn

Stng 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 vtch tkhi lp vmi c hnh thnh. Stng hp chitin gip xc lin

quan n vic sdng glycogen v n phi chuyn sang dng n gin

hn l D-glucose.

- Stng hp v phn hy chitin gip xc rt tch cc v phn ln lp v

btiu hy chc hnh thnh li sau mi ln lt xc. Trong giai on

ny sthiu thc n lm vmng hn cho thy khnng sdng chitinnh mt cht bin dng v dtr.

1.3. Tnh hnh nghin cu trong v ngoi nc

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 vi sng l mt vn tng i mi nc ta. Nm 1978, trng i

Hnh 1. 1 Deacetyl ha chintin thnh chitosan


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11

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, kthut mi

vchitin, cng nh tim nng thtrng 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 trthnh nhu cu trong nhiu ngnh cng nghip v c gi

trth 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 Nghv Sinh Hc Thy Sn phi hp vi mt

sc quan khc nh: i Hc Y Dc TP. HCM, Phn Vin Khoa Hc VitNam, 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 mphm.

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 kthut Masachusetts (M), khi tin hnh xc nh gi trca

chitin v protein trong vtm, cua, cho thy vic thu hi cc cht ny rt c

li nu sdng 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 thgii l 1200 tn. Nc sdng 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 200300France/kg. M, hng nm tng gi tr


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vcc chphm chitinchitosan sdng l 355 triu USD, trong 190 triu

thuc ngnh y t, sau nng nghip (54 triu) v mphm (50 triu). Theo

FAO, nhu cu chitinchitosan c thln ti 36.700 tn/nm trong thp kti.

- Cc phng th nghim ca Malaysia v cng nghip lm ngt nc bin

thnh lp cng ty lin doanh Seafresh chitosan khai thc khnng cung cp

thng phm cc cht dn xut ca tm. Cng ty ny c bit quan tm vi

chitin v polisaccharide tvtm thi bv 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 chitinchitosan bin ng t30 USD n 400 USD/kg

tuthuc vo cht lng sn phm, lng bn chitin v chitosan t khong 2

tUSD.

- 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 nidung khoa hc trong c vic ng dng chitin nh mt cht hp phtrao i

ion tinh chnc 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 thgii sn

xut chitinchitosan nghin cu ra nhiu sn phm c ngun gc chitin s

dng xl nc, khcc 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 khnng 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 thgii c nhiu quy trnh sn xut chitin, vi nhiu

ngun nguyn liu khc nhau, nhng chyu l vtm, cua, gh.

1.3.2.Chitosan


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13

1.3.2.1. Tnh hnh sn xut v nghin cu trong nc

- Vit Nam, vic nghin cu v sn xut chitin, chitosan v ng dng ca

chng trong sn xut phc vi sng l mt vn tng i mi mnc

ta. Vo nhng nm 19781980, 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

ttrong sn xut.[4]

1.3.2.2.

Tnh hnh sn xut v nghin cu ngoi nc

- Chitosan c chp nhn sdng nh mt phgia thc phm hay thc phm

b sung, cc quc gia nh Nht, Anh, Hoa K, Italy, Bo Nha, v Phn

Lan.

- Nm 1992 chitosan c cng nhn l thc phm c li cho sc khe bi Sy

tNht Bn. Hxc nh rng chitosan mang nhng thuc tnh cn thit trong

thc phm bsung, kcnga 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 bsung.[5]- Thtrng 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 tchc.


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14

- Nhiu cc monosacharit ni vi nhau to thnh chitin.

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

C = 47.29 %.

H = 6.45 %.

N = 6.89 %.O = 39.37 %.

- Phn tlng: MChitin=(203,09)n.

Bng phng php nhiu xtia X ngi ta c thchng minh c chitin tn

ti 3 cu hnh: chitin, chitin, chitin. Cc dng ny chkhc 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 chnhm CH2OH, phn

Hnh 1. 2 Cu trc ha hc N-acetyl-D-glucosamine lin kt vi nhaubng cu ni (1-4) glucoside

Hnh 1. 3 Cu trc ha hc chitin


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15

ui ch nhm NHCOCH3, th cc cu trc chitin, chitin, chitin

c m tnh 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 hchui, n cn c lin kt hydro gia

cc lp do cc chui xp knhau nn rt bn vng. Do mt xch xp xp o

chiu, xen k thun li vmt khng gian v nng lng. y cng l dng

nng lng phbin trong tnhin.

- 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 nhqu trnh acetyl ha cho cu trc tinh thbn 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 gcOH vtr C2 c

thay thbi gc acetamine NHCOCH3. im ging nhau ca lin kt glucoside

chitin v cellulose c thhin qua nhit lng ta ra tng ng nhau vo

khong 28Kcal/mol.[13]

1.4.2.Cu trc ha hc ca chi tosan

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

nhm (-COCH3) vtr C2. Ni mt cch tng thhn chitosan l skt hp

ngu nhin gm -(1-4)-D glucosamine v N-acetyl-D glucosamine, do vy

chitosan c thgi l poly -(1-4)-2-amino-2-deoxi-D-glucoz hoc l poly -

(1-4)-D-glucozamin.


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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 hexafluoro-

isopropyl alcohol (CF3CHOHCF3) v 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 thbn vng, him thy trng thi tdo m thng lin

kt vi protein bi ni cng ho trdi dng chitinprotein 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 thtan trong H2SO4m c, H3PO47897% v acid formide khan.

- Trong dung dch HCl, chitin c trin quang thay i t -140 n +560. S

thay i ny chng tc sthy gii chitin.

- phn tn ca dung dch keo chitin trong cc dung dch mui trung tnh ngm

nc tng i cao.[15]

- Chitin c khnng hp thu tia hng ngoi c bc sng 884890 cm-1.

Hnh 1. 4 cu trc chitosan


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- 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 khmu cho chitin

- Khi un nng trong dung dch NaOH m c (4050%), nhit cao th

chitin sbkhmt gc acetyl to thnh chitosan.

Chitin

-CH2OH

-OH

-NHCOCH3

-CH2OH

Chitosan

-CH2OH

-OH

-NH2

- Khi un nng trong axit HCl m c, nhit cao th chitin sbct mch

thu c glucosamin.

Chitin

-CH2OH

-OH

-NHCOCH3

-CH2OH

Chitosan

-CH2OH

-OH

-NH2

Phn ng este ha

- Chitin tc dng vi HNO3m 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 dphn ng hn chitin.[13],[16]

- Trong phn tchitin/chitosan v mt sdn xut ca chitin c cha cc nhm

chc m trong c cc nguyn tO2v N2ca nhm chc cn cp electron

cha sdng, do chng c khnng 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 thmu trng v nh hnh khng mi.


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- Trong tnhin chitin tn ti phbin trong vngoi ca cc lai nm khun

thc vt cp thp, cc lai ng vt gip xc nh tm, cua, cn trng v trong

mng tbo 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 khc nhit cao.

1.5.2.2.

Chitosan

- Vt liu chitosan c ngun gc tnhin, khng c, dng an ton cho ngi.

- Chng c tnh ha hp sinh hc cao vi c th, c khnng tphn husinh

hc.

- Chitosan c nhiu tc dng sinh hc a dng nh: c khnng ht nc, gi

m, tnh khng nm, tnh khng khun vi nhiu chng loi khc nhau, kch

thch spht trin tng sinh ca tbo, c khnng nui dng tbo trong

iu kin ngho dinh dng, tc dng cm mu, chng sng u.

-

Chitosan khng nhng c chcc vi khun gram dng, gram m m cnmmen v nm mc. Khnng khng khun ca chitosan phthuc mt vi yu t

nh loi chitosan s dng (deacetyl, khi lng phn t), pH mi trng,

nhit , sc mt ca mt sthnh phn thc phm. Khnng khng khun

ca chitosan v dn xut ca n c nghin cu bi mt stc gi, trong

c chkhng khun cng c gii thch trong mt strng hp. Mc d

cha c mt gii thch y cho khnng khng khun i vi tt ccc i

tng vi sinh vt, nhng hu ht u cho rng khnng khng khun lin quan

n mc hp phchitosan ln bmt tbo. Trong , chitosan hp phln

bmt vi khun gram m tt hn vi khun gram dng.

Mt sc chkhng khun ca chitosan c gii thch nh sau:

- Nhtc dng ca nhng nhm NH3+ trong chitosan ln cc vtr mang in m

trn mng tbo vi sinh vt, dn ti sthay i tnh thm ca mng tbo

lm cho qu trnh trao i cht qua mng tbo bnh hng. Lc ny vi sinh


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vt khng thnhn cc cht dinh dng c bn cho spht trin bnh thng

nh glucose dn n mt cn bng gia bn trong v bn ngoi mng tbo v

cui cng dn n scht ca tbo.

- Chitosan c thngn cn spht trin ca vi khun do c khnng 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 bmt thnh tbo. Nh vy, vi sinh vt sbc chpht trin do smt

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 tbo ca vi khun, dn n sr rcc

phn tbn trong mng tbo. ng thi gy ra stng tc gia sn phm

ca qu trnh thuphn c khnng khuch tn bn trong tbo vi sinh vt vi

AND dn n sc chmARN v stng hp protein tbo.

- Chitosan c kh nng ph hu mng tbo thng qua tng tc ca nhng

nhm NH3+ vi nhng nhm phosphoryl ca thnh phn phospholipid ca

mng tbo vi khun.- Ngoi ra, chitosan cn c tc dng lm gim cholesterol v lipid mu, lm to vi

ng mch v hhuyt p, iu trthn mn tnh, chng ri lon ni tit.

- Chitosan l cht thn mc khnng hp thdu mrt cao c thhp thu n

gp 68 ln trng lng ca n. Chitosan nhphn tc in tch dng nn

c khnng gn kt vi in tch m ca lipid v acid mt to thnh nhng cht

c phn tln khng btc dng bi cc men tiu ha v do khng bhp

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 khnng 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 bkhnng khng t bin, kch thch lm tng cng h


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thng min dch c th, khi phc bch cu, hn chspht trin cc tbo u,

ung th, HIV/AIDS.

-

Chitosan chng tia tngoi, chng nga.[14]

c tnh

dng trong thc phm v y tphi m bo khc khe v tnh an ton ca

vt cht, nn c nhiu cng trnh nghin cu vc 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 khnng phn husinh hc cao, sau

khi tim vo bng chut, n c thi trddng, nhanh chng qua thn v

nc tiu, chitosan khng phn bti gan v l lch.- Nhng li im ca chitosan: tnh cht c hc tt, khng c, dto mng, c

thtphn 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

thc sdng an ton trn c thngi.[13],[15]


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CHNG 2. CC PHNG PHP SN XUT CHITIN

CHITOSAN

2.1.

Phng php ha hc

2.1.1.S lc vphng php

- Qu trnh tch chit v lm sch chitin c tin hnh theo phng php ha

hc thng thng c p dng ttrc ti nay trong nhiu phng th nghim

trn thgii cng nh Vit Nam thu nhn chitin lm nguyn liu sn xut

chitosan tphphm thy sn, qu trnh gm 3 giai on chnh l loi khong,

loi protein v khmu. V sn xut chitosan ngi ta tin hnh thm bc

deacetyl ha bng dung dch kim.[4]

2.1.2.Quy trnh kthut


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- S quy trnh sn xut chitin.

2.1.3.Thuyt minh quy trnh

Bc 1: Xl nguyn liu vi acid

- Trong vtm, thnh phn khong chyu l mui CaCO3v rt t Ca3(PO4)2.

Nn ngi ta thng dng cc loi acid nh HCl, H2SO4khkhong. Khi

khkhong, nu dng HCl th cho hiu qucao hn. Nu dng H2SO4s to

Deacetyl haDung dch NaOH 3550%,90100oC, thi gian 23 gi

HCl 5%, V/W = 20, nhit phng, trong

1224h.

Kh mu bng H2O2 5%, thigian 15 30 pht. Bng NaOCl5, thi gian 15 30 pht.

Bng KmnO40,1% v C2H2O4.

Ra trung tnh

Loi protein

Loi canxi

Phphm u tm,vcua(cc loi

gip xc)

Ra trung tnh

NaOH 4%, 100oC,thi gian 2 gi

Khmu

Chitin

Chitosan


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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 bbng nc my nhm loi bbt tht, ngm trong 1224 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 bHCl d bi v lng HCl nu cn n s trung

ho lm hao tn lng NaOH cng on khprotein hay deacetyl. Trong qu

trnh ra cng c phn no sc tbra tri.

Bc 2: Xl 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 giri ra sch bng nc my

n pH trung tnh.

Bc 3: Khmu

-

Cc cht mu trong sn phm chitin th c ty trng u tin bng cch dngdung dch thuc tm KMnO4 nng 0,1%, ngm khong 15 pht v vi

C2H2O4khong 12 gicho 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 chbng

cch tch gc acetyl khi nhm acetamid vtr C2 v hnh thnh nhm amino

nhphn ng deacetyl ha chitin.

- Dng xt m c 35 50% deacetyl ha. Mc deacetyl ha phthuc

vo: Nng xt 40%, un trc tip nhit 90120oC, thi gian 512 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.


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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 chtrong 1 ngy, tit kim c thi gian

v sc lao ng.

- Sdng nhng ha cht tng i rtin v thng dng. T chi ph cho qu

trnh khng cao.

-

Quy trnh kh n gin, khng i hi kthut cao.- Trang thit bthng dng, c thdng dng cni tr. Do khng tn km

nhiu trong u t trang thit b.

2.1.4.2. Nhc im

- iu quan trong nht c thkn l cht thi ca phng php ha hc ny

c khnng gy nhim mi trng cao, c hi i vi con ngi vt nui,

ng vt thy sinh.

-

Hiu qutrch ly khng cao di 10%.


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- Cht lng sn phm km, cht lng khng ng iu gia cc mtrch ly.

- Kh khn trong vic kim sot qu trnh. [10]

2.2.

Phng php ha sinh kt hp

2.2.1.S lc vphng php

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

ch sdng quda th trong quda 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 bsung dch ny vo

khi nu tht th tht mn rt nhanh.

- Tn dng hai tnh cht tt ny ca quda thay thHCl 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.



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- S quy trnh sn xut chitosan bngphng php ha sinh kt hp.


Bc 1: Xl nguyn liu vi enzyme bromalin

Chitosan

Nc p bda. Tl300ml/100g nguyn

liu

Khmu bng H2O25%, thi gian

15 - 30 pht. Bng NaOCl 5, thigian 15 30 pht. Bng KMnO40,1% v C2H2O4

Ra sch thu cn

Loi protein ln 2

Loi khong, protein

Phphm u tm, vcua(cc loi gip

xc)

Ra trung tnh

NaOH 12% ,90oC,thi gian 45 pht

Khmu

Chitin

Deacetyl haDung dch NaOH 3550%,90100oC, thi gian 23 gi

Ra loi tht


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- 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. Cxl nh vy 67 ngy, thu cn v ra sch.

Bc 2: Xl vi kim

- Protein c thy phn trong mi trng acid nn cn hin tng ta nn cn

xl vi NaOH 12% ,90oC, thi gian 45 pht nhm mc ch ho tan protein

v tin hnh ra stng hiu quca giai oan loi protein hn. T cho sn

phm sch hn.

Bc 3 khmu, 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 viphng php ha hc.

- Lng ha cht chyu trong qu trnh tch chit chitin l HCl v NaOH c

sdng t hn, t tit kim c ho cht v lng nc ra sau xl acid

v kim. Mc khc, trong qu trnh loi protein trong phng php ha sinh kt

hp ny hon ton dng enzyme protease tdch p vda, m khng cn s

dng HCl trc . Lng NaOH tiu tn cn gim i nhiu ln ( t24 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 tcht thi.

- Thit bsdng t hn, iu ny mang li ngha kinh tln i vi quy trnh

cng nghkhi pht trin to ra sn phm thng mi.

2.2.4.2.

Nhc im


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- Hn chca phng php l thi gian thu nhn sn phm lu hn 7 8 ngy,

trong khi phng php ha hc ch1 ngy l cho ra sn phm. Qu trnh t

hn nhng tiu tn nhiu cng lao ng hn.[8],[9]

- Tiu tn nhiu cng lao ng hn, do c nhiu tin trnh hn.

2.3. Phng php sinh hc

2.3.1.S loc vphng php

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

nguyn tc chung ca sn xut chitin chitosan khkhong bng acid v c,

khprotein bng kim. Trong phng php sinh hc ny stin 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 sl tc nhn thy phn protein trong nguyn

liu vtm.

- Chng vi khun c s dng trong phng php ny l Lactobacillus

acidophilus ly t ngun dc phm. ANTIBIO ca Hanwha Pharm Co-

KOREA, mang chng vi sinhL. acidophillus, mt 109cfu/g.

S lc vLactobacillus acidophillus- Phn loi:

Gii: Bacteria

Ngnh : Firmicutes

Lp: Bacilli

H: Lactobacillaceae

Chi:Lactobacillus

Loi:Lactobacillus acidophillus

- im c trng phbin 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, Lactobacilluscn l vi khun Gram dng, khng hnh thanh bo t

v him khi di ng. Ngi ta thy rng tboLactobacillusin hnh c dng

hnh que, vi kch thc bin i trong khong (0,51)(1 10)m, i khi


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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 thsinh trng

trong phm vi 553oC.

- 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 5,55,8, nhng nhn chung chng c thsinh trng pH < 5.

- Ngoi ra ngi ta c thda trn cc sn phm cui ca qu trnh bin dng

phn loi cc loiLactobacillusthnh 2 loi. Cc loi ln men ng hnh v

cc loi ln men dhnh. Cc loi thuc nhm ln men ng hnh to ra 85%

acid lactic tglucose, trong khi loi ln men dhnh chto ra 50% acid lactic

v mt lng ng kcarbon dioxide, acetate v ethanol.L. acidophilusl 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 , sdng 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 chyu hoc duy nht ca strao 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

thsng trong mi trng c tnh axit cao pH 4 5 hoc thp hn v c m

tnh l mt v dloi ln men ng hnh sn xut axit lactic l sn phm duy

nht (Vijayakumar et al., 2008).



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- Quy trnh khkhong v khprotein trn v tm s sn xut chitin ngun

nguyn liu sn xut ra chitosan.


Bc 1: Ra s bvtm

Vtm s

Ra s b Phi trn nguyn liu Tlnc : vtm2 5:1

10,5% ng.

0,875% CH3COONa0.35% KH2PO4

Chitin

GingLactobacillusacidophilus.

Chnh pH 6.5

Gia nhit 80oC, 15p

Cy ging trn 15%

Ln men 72h 96h,37oC. ym kh, khuy

Tymu bng H2O2

Ra

Sy kh 5065oC


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31

Mc ch

- Loi bbt mt phn vi sinh gy thi c sn trong vtm, loi btp cht nh

rc, t, ctv mt stht tm bthi ra (gim bt mi).

Tin hnh

- Cn mt lng xc nh vtm, mang i ra trc tip bng nc sinh hot bnh

thng, ra qua mt ln l .

- Rng vt vtm ln ro nc.

Ch

- Khng ngm vtm qu lu trong nc.

- Khng ra nhiu ln gy hao tn nc m cn ra tri ht phn tht xt li trn

vtm, 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, sgim 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

sci bin tcng thc gc nh sau.

Mc ch

-

y l mt bc kh quan trng, c thni n quyt nh qu trnh xl c tt

hay khng, lng acid sinh ra cng nhiu th hiu quxl 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.

- Bsung cc thnh phn khc m c thc chcc vi sinh vt khc pht trin,

gim thiu cnh tranh mi trng sng cho chng vi khun ta cy ging.

Tin hnh


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- 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 vtm

Nc 250%

Sucrose 10,5%

CH3COONa 0,875%

KH2PO4 0,35%

Ch - Ngun nc sdng trong quy trnh phi l ngun nc sinh hot hoc tt hn.

Dng cc ngun nc khc skhng 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 vtm

Mc ch

-

Nhm ng ha cc cht trong nguyn liu.- Vtm c xay nhstng bmt tip sc, phn bu trong mi trng.

Tin hnh

- Dng my xay bnh thng hoc c thbm bng tay, tuy nhin cn t yu

cu l vtm 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 tth lm cho chng chm pht trin li.

- To mi trng tng i sch, u thchoL.acidophilus.

Tin hnh

- Ci t thit bhot ng gia nhit 80oC trong 15 pht.

Ch

-

Cn m bo thi gian v nhit gia nhit.


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- Cn kim tra bo tr thit btrc 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 thvi 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 vtm).

Ch

- Phi m bo mt v thtch 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 k5pht khuy 10 pht ngh.

Ch

-

Kim tra thit btrc khi ln men.

- Kim tra hthng c hot ng u trong sut thi gian ln men. V quan st

24h u c sinh kh (kh ny c thdo vi sinh gy thi sinh ra, v sau khi xb

kh ny i th cho ti lc kt thc ln men khng thy sinh kh na). Nu 24h

u sinh kh nhanh, xkh t th.

-

Nu chy th chc chn bnhim vi sinh gy thi sinh kh CH4chthm vi

gina xem chng vi sinh ta cy vo c c chc khng ( thng l khng


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chng sinh CH4ny kh mnh) v nu cn sinh kh v t chy na th nn b

ngay mln men, tip tc srt thi. Nu khng cn kh sinh ra th chng t

chng vi khun ta cy vo c chc, cho ln men tip.

- Nu khng chy, th ch thm vi gixem cn sinh kh khng, nu cn cng

tin hnh loi bmln men, nu khng th chng tchng vi khun ta cy vo

c chc v tip tc ln men.

- Quyt nh cho ln men tip th canh thi im 4048h sau ln men tin hnh

o pH thy c gim th cho ln men tip, nu tng th tin hnh loi bmln

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 sra sch, tuy nhin cn

tit kim ti a chiph, vy nn khng ra m i tin hnh ty mu trc tip. C

hai nguyn nhn thnht l tit kim chi ph sdng nc, thhai trn vtm

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 trvn an ton hn. Vvy khi cho vo ty mu vi H2O2 l mt cht oxy ha mnh, khnng 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 trcm 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 vtm.

- Tin hnh khuy o sau mi 15 pht ngm.

- Thu hi lng H2O2c thti sdng li.

Ch


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35

- H2O2l cht oxy ha mnh, nguy him i vi ngi sdng, khng tip xc

da tay trc tip.

-

Trang bcn thn bo hlao ng khi lm vic.

Bc 7: Ra

Mc ch

- Sau ln men cc thnh phn khong lin kt trn vtm c tch ra, cc ion

khong ny chyu l canxi kt hp vi acid lactic to thnh canxiumlactate,

thnh phn na l protein bthy phn snm 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 bbm li trn vtm. 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,

khnng n mn da tay rt nhanh, cn thc hin cc bin php an ton khi tipxc.

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 tsy, ci t nhit

dao ng t5065oC, trong vng 46h.

- Nu thi tit thun li nhiu nng, c thphi 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, slm xu mu sn phm.

2.3.4.Thit b


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36

Cc thit bphc vcho quy trnh ln men c bn nh sau:

- Mt sthit bthng dng ti phng th nghim nh pipet, cn, my o pH

-

ng vai tr quan trng hn l thit bln men, c tnh ca thit bc thhp

tit trng nh mt autoclave, giv to iu kin v trng, ym kh, hiu kh,

gin nh mt nhit cn thit.

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

thm hthng trao i nhit nhm rt ngn thi gian lm ngui sau gia nhit v

ng c khuy c th ty chnh chkhuy o theo chu kcn thit, gip

mi trng c khuy u lin tc to iu kin cho vi khun sng tt hn.

Hnh 2. 2 Ni ln men c sdng trong quy trnh sn xut


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- Thit bhot ng da trn mt biu khin, cho php bn tng ha mt

phn quy trnh sn xut.

2.3.5.u im

Hnh 2. 4 Biu khin

Hnh 2. 3 np ni v cc chi tit thit b


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38

- Trc tin phi ni n l thn thin mi trng gn gi cuc sng nhgim

thiu tc nhn gy nhim mi trng thng qua gim cc ha cht sdng

trong phng php sn xut truyn thng.

- Vic khng sdng ha cht cng ko theo chi ph cho sn xut gim xung,

tng li nhun kinh t.

- Xl c tc nhn nhim tphthi vtm, vtm khng c xl 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 sdng vi khunLactobacillus acidophilus, l mt vi sinh ng rut t

san ton hn cho cng nhn thc hnh sn xut. To ra sn phm cng an ton

hn cho ngi sdng.

- Sdng mt tc nhn an ton hn l vi sinh nhng gii quyt c hai vn

l khkhong v khprotein.

- Dch sau ln men c cha rt nhiu thnh phn dinh dng, nn co bin php

thu hi c thlm ngun thc n chn nui giu dinh dng.

-

Bqua nhiu bc gia nhit stit kim c ngun nng lng kh ln so viphng php ha hc.

- Vtm sau xl trng p hn mu ng ca cc phng php khc.

2.3.6.Nhc im

- Quy trnh tin hnh c nhiu bc i hi kthut chuyn mn hn, v dk

thut v trng, hoc cc kthut lm gim thiu ngun nhim bn tngoi 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 chmt 1 ngy.

- Tlthnh cng ca qu trnh ln men phthuc nhiu yu t, vy nn vn c

mln men tht bi gy thi. Trong khi phng php ha hc th thng lun

thnh cng.


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- Cht lng sn phm khng ng du, v n phthuc nhiu vo ngun nguyn

liu vtm ban u.

-

Phi theo di thng xuyn trong qu trnh ln men, nhm pht hin sctp

nhim gy h hng.


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40

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

- Chitinchitosan 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 khnng khng khun Nnc 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 khprotein 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 scho kt qutt hn na.

- Tn dng gn nh hon ton sn phm sau ln men tdch 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 thLactobacillus acidophillus bng mt chng vi sinh

khc ti u hn.

- Nghin cu tm ra cc thnh phn khong bsung vo nguyn liu tng hiu

ququ trnh ln men. Tm ra nng thch hp nht bsung.


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- Hin nay vn cha deacetyl ha chitin cho ra chitosan bng sinh hc, y l mt

bc tng gi trsdng tngun ph thi v tm, nn rt mong sc cng

trnh nghin cu cho vn ny.

- Nghin cu thm quy trnh thu hi caxium lacatate trong canh trng ln men.

V thu hi cphn canh trng ln men lm thc n gia sc rt giu m.


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42

Ti liu tham kho:Ting vit:

1.

Phm l Dng, Trnh Bnh, Li Thu HIn cng cc cng s(1997). Vt liusinh hc t chitin, Vin Ha Hc-Vin Cng Ngh Sinh Hoc, Trung tm

khoa hc v Cng Nghquc 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 Thnh 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 chbin mt ssn phm

cng nghip tphliu vtm, 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 nghenzyme,Nh xut bn i Hc Quc

Gia Tp.HCM.

7. Nguyn Tin Thng (2003). Mt sk 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 xl enzyme, Tp ch

Khoa hc - Cng nghThy sn, 1, 3-9.

9.

Trang S Trung, V Ngc Bi, Phm Than Phng (2007). Nghin cukt hp enzym protease trong cng nghsn xut chitin tph liu u v

tm, Tp ch Khoa hc - Cng nghThy sn, 3, 11-17.

10.Trang S Trung, Phm Th an Phng (2012). Tnh cht ca chitin v

chitosan tv tm thchn trng, khprotein bng phng php ha hc

v sinh hc,Tp ch khoa hc-cng nghthy sn, 3.


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43

11.Trn Thi Trung (2005). Nghin cu cc yu t nh hng n qu trnh

deacetyl v ct mch chitin iu chglucosamine, 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, 233242.

16.

Stevens, W. F (2001). Production of chitin and chitosan: Refi nement andsustainability 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-625-

chitosan.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/107-

chitosan-tng-quan-nghien-cu-ng-dng.html
http://www.chitosan.com.vn/http://www.chitosan.com.vn/http://www.webmd.com/vitamins-supplements/ingredientmono-625-chitosan.aspx?activeIngredientId=625&activeIngredientName=chitosanhttp://www.webmd.com/vitamins-supplements/ingredientmono-625-chitosan.aspx?activeIngredientId=625&activeIngredientName=chitosanhttp://www.webmd.com/vitamins-supplements/ingredientmono-625-chitosan.aspx?activeIngredientId=625&activeIngredientName=chitosanhttp://www.webmd.com/vitamins-supplements/ingredientmono-625-chitosan.aspx?activeIngredientId=625&activeIngredientName=chitosanhttp://doan.edu.vn/do-an/de-tai-tim-hieu-ve-chitin-chitosan-5929/http://doan.edu.vn/do-an/de-tai-tim-hieu-ve-chitin-chitosan-5929/http://www.cyberchemvn.com/cyberchem/technology-and-apply/107-chitosan-tng-quan-nghien-cu-ng-dng.htmlhttp://www.cyberchemvn.com/cyberchem/technology-and-apply/107-chitosan-tng-quan-nghien-cu-ng-dng.htmlhttp://www.cyberchemvn.com/cyberchem/technology-and-apply/107-chitosan-tng-quan-nghien-cu-ng-dng.htmlhttp://www.cyberchemvn.com/cyberchem/technology-and-apply/107-chitosan-tng-quan-nghien-cu-ng-dng.htmlhttp://www.cyberchemvn.com/cyberchem/technology-and-apply/107-chitosan-tng-quan-nghien-cu-ng-dng.htmlhttp://www.cyberchemvn.com/cyberchem/technology-and-apply/107-chitosan-tng-quan-nghien-cu-ng-dng.htmlhttp://doan.edu.vn/do-an/de-tai-tim-hieu-ve-chitin-chitosan-5929/http://www.webmd.com/vitamins-supplements/ingredientmono-625-chitosan.aspx?activeIngredientId=625&activeIngredientName=chitosanhttp://www.webmd.com/vitamins-supplements/ingredientmono-625-chitosan.aspx?activeIngredientId=625&activeIngredientName=chitosanhttp://www.chitosan.com.vn/


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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%20IJBAS-

IJENS.pdf

32.http://www.plantphysiol.org/content/66/2/205.short

33.

http://www.tandfonline.com/doi/abs/10.1080/0890543970954992034.http://www.sciencedirect.com/science/article/pii/S1381514800000389

35.http://www.foodproductiondaily.com/Packaging/Researchers-develop-eco-

friendly-packaging-using-chitosan-from-shrimp-shells

HT
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