1

PHN 1: M U

C ph c con ngi bit n khong hn 300 nm nay, mun hn rt

nhiu so vi nhiu cy lng thc, thc phm quan trng khc. Tuy nhin, ngy nay
cy c ph tr thnh mt loi cy cng nghip c gi tr kinh t cao, l mt mt
hng xut khu quan trng ca nhiu quc gia trn th gii. Sn phm ca n l mt
loi thc ung th v khng th thiu ca nhiu dn tc. Bn cnh , mt s mt hng
thc phm nh sa, bnh, ko,... c ch bin vi s c mt ca c ph s lm tng
thm hng v v gi tr dinh dng cho sn phm.
S d c ph c s dng ngy cng nhiu v trong ht c ph cha 0,8 3%
caffeine, mt hot cht c tc dng kch thch thn kinh, gip t bo no tng cng
kh nng lm vic, kh nng t duy v qua thc y hot ng ca h tun hon,
tng cng phn ng ca c bp,
Ngoi ra, trong ht c ph cn cha nhiu cht dinh dng cn thit cho c th
con ngi nh ng saccharose, ng kh, cc protein ha tan, c bit l cc
vitamin B1, B2, B6, B12 v PP c hm lng kh cao, l nhng cht cn thit cho
nhu cu sinh l ca c th chng ta. Do c ph gip nng cao sinh lc, chng mt
mi, gip con ngi lm vic sng sut v thoi mi hn.
S sng khoi v b dng m c ph mang li cho con ngi l do nhng
thnh phn ha tan trong ht. Trong k thut ch bin c ph hin nay c hai phng
php chnh l ch bin kh v ch bin t. Tuy nhin, x l c ph theo qui trnh
thng thng cha mang li hiu qu ti u cho vic trch ly cc thnh phn ha tan c
trong ht. Nguyn nhn ca mt hn ch ny l do s hin din ca pectin v cellulose
hai thnh phn ch yu trong c ph nhn. Vn t ra l s dng phng php ln
men nng cao ti a hiu sut trch ly ca c ph, t thu c lng cht ha tan
cao nht.
Cng vi s pht trin ca i sng v ca x hi, s pht trin ca cng ngh
sinh hc ni chung, cng ngh enzyme v cng ngh ln men ni ring ng mt vai
tr quan trng trong vic p ng nhu cu ngy cng cao ca con ngi, c bit trong
vn nng cao kh nng khai thc cc thnh phn ha tan ca ht c ph. Lun vn
ny khng nm ngoi khun kh , chng ti n lc nghin cu sn xut mt loi
 2

ch phm sinh hc (ch yu l pectinase v cellulase) vi tn gi Biocoffee-1, t

chng nm mc Aspergillus niger v Trichoderma reesei c hot tnh phn gii mnh
trn i tng c ph nhn, tc ng ch yu vo hai thnh phn chnh l pectin v
cellulose.
Vit Nam hin nay, c ba loi c ph nhn ch yu c bun bn v s
dng ph bin trn th trng l c ph Bi, c ph S v c ph Mokka. Mc ch ca
ti l x l ba loi c ph ny bng cng ngh ln men v cng ngh enzyme, nhm
thu c cht ha tan cao vi hiu sut trch ly cao nht.
Ni dung nghin cu ca ti bao gm 4 ni dung:
1. Xc nh cc thnh phn ch yu trong c ph nhn
2. To ch phm Biocoffee-1 vi hot tnh cao ca pectinase v cellulase
3. Kho st cc iu kin ti u cho qu trnh ln men c ph
4. Kho st s thay i trng lng v nhit ca khi
ti ca chng ti l nhng bc i u tin cho nhng nghin cu, kho st
tip theo trong vic pht trin sn phm c ph ln men. y l mt gii php nhiu u
im cho qu trnh ch bin c ph v l mt hng i mi gn gi vi thc t sn
xut cng nh thc t cuc sng, mang li nhiu li ch thit thc cho ngnh cng
ngh thc phm, cho nn nng nghip quc gia v cho nhu cu m thc ca ngi dn
Vit Nam chng ta.
 3

PHN 2: TNG QUAN TI LIU

Tng quan ti liu ca chng ti gm 4 phn chnh sau:

o Tm hiu v c ph
o Tm hiu v cellulose v pectin hai thnh phn ch yu trong c ph nhn
o Tm hiu v h enzyme cellulase v pectinase
o Tm hiu v nm si sinh tng hp hai h enzyme trn

TM HIU V C PH

Ht rang

Ht kh
Ht ang kh

Xay thnh bt

Ht ra sch

C ph pha Ht ln men
Hoa

Qu ang chn

Tht qu
Qu xanh Qu chn

Hnh 2.1: Qui trnh ca c ph t khi n hoa cho n thnh phm cui cng
 4

2.1. TM HIU CY C PH TRONG NC V TRN TH GII

2.1.1. Lch s cy c ph [20], [21]
Tn gi coffee xut pht t ting rp l kahwa, m lc u l mt t ng
trong th ca dng ch ru vang. Do o lut ca Hi gio nghim cm gio dn
ung ru, nn tn gi y c bin tng thnh coffee, v thng qua ting gi tng
ng ca Th Nh K l Kahweh tr thnh Caf (Php), Caffee (), Kaffee (c),
Koffie (H Lan), Coffee (Anh), v tn Latin l Coffea dng trong phn loi ging loi
thc vt.
Ring ti Vit Nam, vo nm 1888, thc dn Php mang c ph vo trng u
tin nc ta, ri t , tn gi c ph ca Vit Nam l do s Vit ha trong phin
m t s pht m Caf ca ngi Php m ra.
Theo truyn thuyt k li rng c ph c tm thy rt lu k t thi k
v nhng tc dng kch thch ca n c ghi nhn t xa xa, c l l do vic
quan st cc tc dng y qua trng thi biu l ca cc ng vt gm c sau khi chng
n nhng tri c ph hoang di. Ngun gc v a d v nng sn ban u ca c ph
l t Ethiopia.
T trc nm 1200, vic tiu th c ph lan rng t vng bin ti Aden,
Mecca v Cairo. Ti cc vng rp (Arabia) ny, cc bi cy c ph c trng
trt v c ti tiu trong hn mt ngn nm. Cho nn, ht c ph mi c tn gi l
Coffea arabica, mc nhin tha nhn rng n c canh tc v tiu th ph bin trong
cc nc rp, m qun i ci ngun ca th nc gii kht ny l x s Ethiopia.
Ti Vit Nam, cy c ph c a vo trng t nm 1857, trc ht l ti mt
s nh th Qung Bnh, Kontum, Song mi n u th k 20 tr i th cy c ph
mi c trng trn quy m tng i ln ca cc ch n in ngi Php ti Ph
Qu Ngh An v sau l c Lc v Lm ng, nhng tng din tch khng qu
vi ngn ha. Sau Cch mng thng Tm, din tch c ph min Bc c pht trin
thm ti mt s nng trng quc doanh v thi k c din tch cao nht l trn 10.000
ha vo nm 1963 1964. min Nam trc ngy gii phng, vo nm 1975 din tch
c ph c khong 10.000 ha. C ph trng min Bc trong nhng nm trc y ch
yu l c ph ch (Coffea arabica). Do iu kin sinh thi khng ph hp, c bit l
c mt ma ng gi lnh ko di, v vy cy c ph vi (Canephora robusta) kh c
 5

kh nng pht trin min Bc, nhiu vng trng c ph vi sau phi hy b v
km hiu qu.
Din tch trng c ph min Nam trc ngy gii phng ch yu l ging c
ph vi (Canephora robusta), mt s din tch nh c ph ch c trng Lm
ng. Nng sut c ph vi trong thi k ny thng t trn di 1 tn/ha, mt s
n in c qui m va v nh cng t c nng sut t 2 3 tn/ha. Ngy nay,
nh c p dng nhng tin b k thut, nng sut tng ln rt nhanh.
C ph Vit Nam lun lun l mt mt hng nng sn quan trng trn th
trng th gii v em v ngun ngoi t xng ng trong nn kinh t quc dn.
2.1.2. Tnh hnh sn xut, tiu th
Tnh hnh sn xut c ph trn th gii [11], [20], [36]
Trn th gii hin nay c trn 75 nc trng c ph vi din tch trn 10 triu
ha v sn lng hng nm bin ng trn di 6 triu tn. Nng sut bnh qun cha
vt qu 6 t c ph nhn/ha. Trong , chu Phi c 28 nc vi nng sut bnh
qun khng vt qu 4 t c ph nhn/ha, Nam M t di 6 t nhn/ha. Bn nc c
din tch c ph ln nht l: Brazil trn 3 triu ha chim 25% sn lng c ph th
gii, Cte Dlvoire (chu Phi) v Indonesia (chu ) mi nc khong 1 triu ha, quc
gia th t l Colombia c gn 1 triu ha vi sn lng hng nm t trn di 700
ngn tn.
Do p dng mt s tin b k thut mi nh ging mi v mt trng dy nn
c hng chc nc a nng sut bnh qun t trn 1 tn/ha. in hnh c Costa
Rica Trung M vi din tch c ph ch l 85.000 ha nhng t nng sut bnh
qun trn 1.400 kg/ha.
C ph ch hin nay vn chim 70% sn lng ca th gii. Din tch c ph
ch c trng tp trung ch yu Trung v Nam M, mt s nc ng Phi nh:
Kenya, Cameroon, Ethiopia, Tanzania v mt phn chu nh: Indonesia, n ,
Philippines.
My nm nay sn lng c ph Robusta trn th gii tng ln nhanh chng, v
2000/2001 t ti 44,8 triu bao tng ti 12,2 triu bao so vi v trc v chim ti
38% tng sn lng c ph.
 6

Theo kt qu thm d mi y nht, gi c ph trong nm 2005 s bin ng

mnh vi nhiu kh nng gi tng cao nu thi tit xu lm cho sn lng d kin
gim mnh. (Ngun: Reuters, 27/01/05)
o Th trng c ph Arabica th gii s thiu ht khong 5,4 triu bao c ph
nin v 2005/2006 sau khi thng d 2,5 triu bao ca nin v trc. Tng sn lng c
ph th gii c tnh t 111 triu bao nin v 2005/2006, gim so vi mc 117 triu
bao ca 2004/2005.
o i vi c ph Robusta, theo d kin s tng 84% t 759 USD/tn ca cui
nm 2004 ln 1.400 USD/tn nm 2005 nu gi c ph Arabica tng mnh. c tnh
s c khong 149 triu bao c ph nin v 2005/2006 gm c c ph tn kho cc
nc tiu th v sn xut.
S pht trin ca ngnh c ph Vit Nam [11], [20], [44], [46], [48], [49]
Vit Nam gia nhp T chc C ph Quc T (International Coffee
Organization, ICO) vo nm 1991.
Ngh trng c ph Vit Nam l mt ngun thu nhp cho mt nhm ng dn
c nng thn, trung du v min ni. Vi 500.000 ha c ph to vic lm cho hn
600.000 nng dn v s ngi c cuc sng lin quan n cy c ph ln ti trn 1
triu ngi.
Ngnh c ph Vit nam c nh gi l cn mi m, phi cnh tranh vi
ngnh c ph ca nhiu nc c truyn thng lu i hn, vn c ting tm v mt
cht lng v s bn vng. y l mt vn m ngnh c ph Vit Nam phi c
gng trn nhiu lnh vc t khu p dng nhng k thut sn xut nng nghip n
cng ngh ch bin tin tin, a ra th trng nhiu chng loi sn phm mi.
 7

Bng 2.1: Xut khu c ph ca Vit Nam qua cc v t 1994 n 2002

Sn lng xut khu Kim ngch xut khu
Nin v
(nghn tn) (triu USD)
1994/1995 192.379 51.291,649
1995/1996 221.496 401.948,791
1996/1997 336.242 402.817,916
1997/1998 395.418 601.430,778
1998/1999 404.206 554.974,838
1999/2000 653.678 573.976,994
2000/2001 874.676 381.883,542
2001/2002 712.735 263.259,766
(Ngun: Hip hi C ph Ca cao Vit Nam, 2005)

Bng 2.2: Kt qu xut khu c ph nhn sng ca Vit Nam nm 2004

Sn lng xut Gi bnh qun
Thng Tr gi (USD)
khu (tn) (USD/tn)
1 68.815 44.114.705 641,1
2 86.570 57.439.162 663,5
3 83.000 54.509.033 656,7
4 82.940 54.343.678 665,2
5 78.582 51.933.539 660,9
6 118.731 78.797.170 663,7
7 55.829 36.883.303 660,6
8 51.144 33.022.482 645,7
9 56.086 35.386.587 630,9
10 65.474 40.077.916 612,1
11 55.292 33.940.074 613,8
12 87.242 55.639.710 637,8
Tng 889.705 576.087.360 647,5
(Ngun: Hip hi C ph Ca cao Vit Nam, 2005)
 8

Khuynh hng pht trin c ph hin nay l thm canh tng nng sut c ph
Robusta, m rng din tch c ph Arabica trn vng cao min Bc nh Sn La,
Tuyn Quang, Lai Chu, Bc Cn, Thi Nguyn, Lng Sn, Ha Bnh, Ngh An vi
nhng dng chn lc nh Catimor. Catimor cng c a vo cao nguyn Lm
ng v c Lc thay cho mt s ging d nhim bnh r st.
Robusta l dng sn xut chnh v c c im d thch nghi, nng sut cao,
trung bnh 4 n 5 tn nhn/ha.
Hin nay, Vit Nam, c ph l mt hng nng sn xut xut khu c gi tr
ln ng th hai sau go (nhng con s thng k ca Hip hi C ph Ca cao Vit
Nam iu tra vo nm 2000 cho thy din tch c ph c nc ln n 520.000 ha
vi sn lng 900.000 tn, a Vit Nam ln v tr th 2 th gii v sn xut c ph).
C ph Vit Nam c xut sang trn 50 quc gia v vng lnh th. Gi tr c
ph xut khu thng chim gn 10% tng kim ngch xut khu hng nm. V
2000/2001 Vit Nam xut c ph i 61 nc, trong , 10 nc nhp khu c ph
ng u c trnh by trong bng 2.3 di y.
Bng 2.3: 10 nc nhp khu c ph hng u ca Vit Nam
S lng Tr gi T phn so vi tng
STT Tn nc
(tn) (USD) xut khu (%)
1 B 138.603 57.947.984 15,85
2 M 137.501 59.371.585 15,72
3 c 134.321 60.054.805 15,36
4 Ty Ban Nha 73.852 31.666.889 8,44
5 62.559 27.796.789 7,15
6 Php 45.998 20.147.381 5,26
7 Ba Lan 38.155 17.171.839 4,36
8 Anh 30.153 13.055.058 3,45
9 Nht 26.905 13.274.686 3,08
10 Hn Quc 26.288 11.310.104 3,01
(Ngun: Hip hi C ph Ca cao Vit Nam, 2005)
 9

Bng 2.4: Din bin din tch v sn lng c ph Vit Nam trong 9 nin v
t 1992 n 2001
Nin v Din tch (ha) Sn lng (tn)
1992/1993 140.000 140.400
1993/1994 150.000 181.200
1994/1995 215.000 211.920
1995/1996 295.000 236.280
1996/1997 350.000 342.300
1997/1998 410.000 413.580
1998/1999 460.000 404.206
1999/2000 520.000 700.000
2000/2001 500.000 900.000
(Ngun: Hip hi C ph Ca cao Vit Nam, 2005)

C ph l mt loi nc ung cao cp, nhu cu i hi ca ngi tiu dng vn

khng ngng tng ln, cha c nhng sn phm nhn to c chp nhn thay th
cho c ph, v vy vic trng, xut khu, nhp khu loi hng ha c bit ny vn c
mt ngha kinh t ln i vi nhiu nc.
2.1.3. Tiu chun cho c ph Vit Nam [48]
Trc y c ph Vit nam c bn vi 3 ch tiu cht lng n gin: thy
phn %, ht en v % v tp cht %. Mc d Vit Nam tng ban hnh Tiu chun
Nh nc v yu cu k thut i vi c ph nhn TCVN 4193 86. Tuy nhin vo
thi k m ca, ngnh c ph tip xc trc tip vi th trng th gii, trong bui ban
u cn c mt h thng tiu chun n gin v d thc hin hn nn ra i TCVN
4193 93 vi 3 ch tiu nh nu trn. Ngy hm nay, ngnh c ph Vit Nam
trng thnh hn mt bc v th trng i hi cht lng cao hn, cn c tiu chun
cp Nh nc ph hp vi tiu chun quc t. Do ngnh c ph Vit Nam c
Nh nc h tr xy dng v cho php ban hnh TCVN 4193 2001. C th coi y
l mt bc tin ng k ca ngnh c ph Vit Nam.
Ban K thut v Tiu chun c ph (TCVN/TC/F16) m Hip hi C ph Ca
cao Vit Nam l c quan ch tr hon thnh vic sot xt mt s tiu chun Nh
nc v c ph v c Tng cc Tiu chun o lng cht lng trnh ln B Khoa
 10

hc, Cng ngh v Mi trng xt duyt. Ngy 05/11/2001, B trng B Khoa hc,
Cng ngh v Mi trng c quyt nh s 57/2001/Q-KHCNMT ban hnh 5 bn
tiu chun v c ph.
Bng 2.5: 5 tiu chun v c ph do B Khoa hc, Cng ngh v Mi trng ban
hnh ngy 05/11/2001
C ph nhn yu cu k thut
1 TCVN 4193 : 2001
(Sot xt ln 3 Thay th TCVN 4193: 1993)
C ph v cc sn phm ca c ph Thut ng v
TCVN 4334 : 2001
2 nh ngha
(ISO 3509 1989)
(Sot xt ln 1 Thay th TCVN 4334 86)
C ph nhn Phng php xc nh c ht bng
TCVN 4807 : 2001
3 sng tay
(ISO 4150 1991)
(Sot xt ln 2 Thay th TCVN 4807 89)
TCVN 6928 : 2001 C ph nhn Xc nh s hao ht khi lng
4
(ISO 6673 1983) 105oC
TCVN 6929 : 2001 C ph nhn Hng dn phng php m t cc
5
(ISO 9116 1992) quy nh
(Ngun: Hip hi C ph Ca cao Vit Nam, 2005)

2.1.4. Mt s tin b k thut mi trong ngh trng v s dng c ph trn th

gii [20]
o S dng cc ging mi qua qu trnh chn lc, lai to mang c cc c
tnh tt v: kh nng khng su bnh, cht lng tt, cho nng sut cao.
o p dng phng php nhn ging v tnh bng k thut gim cnh, nui
cy m, to ra cc vn c ph thun chng, nng sut cao.
o p dng k thut trng dy, tng mt trng trn mt n v din tch, cho
php t nng sut cao, rt ngn thi gian, mang li hiu qu ln.
o Bn phn da trn kt qu chn on nhu cu dinh dng khong ca cy
trng thng qua bin php phn tch l v t.
o S dng nhng cy che bng mi, ph hp vi nhu cu sinh l v nh sng
ca cy c ph.
 11

o S dng cc loi ch phm bo v thc vt mi, t c i vi ngi, gim

s nhim i vi mi trng.
o S dng cc tin b k thut ca ngnh in t ch bin v phn loi c
ph, to ra nhiu dng thnh phm c kh nng bo qun gi cht lng c lu hn.
o S dng cc ch phm kch thch sinh trng lm cho qu trnh ra hoa, u
qu v chn tp trung.
o S dng nhng thnh tu ca cng ngh sinh hc tc ng vo qu trnh
sn xut v ch bin, to ra thnh phm a dng v cht lng hn.
2.1.5. M t cy c ph [2],[29], [35]

Hnh 2.2: Qu c ph Hnh 2.3: Hoa c ph

Cy nh, mc sum su, lun xanh, cao 6 10 m. Thn cnh hnh tr hoc hnh
vung, phn nhnh nhiu. L mc i, hnh bu dc hoc hnh trng, di 10 15 cm,
rng 5 6 cm, u nhn, mt trn xanh sm bng, mt di rt nht, gn ni r, mp
un ln; cung l di 0,7 1 cm; l km c gc rng, u nhn.
Cm hoa nh xim co gm 8 15 hoa nch l; hoa trng, rt thm; i ct;
trng hnh ng ngn, 5 cnh hoa u; nh 5 dnh hng trng, xen k vi cnh hoa;
bu 2 , mi cha mt non.
Qu hch xoan hi dt, lc cha chn c mu xanh lc, khi chn chuyn sang
mu , cha 2 ht.
Ma c ph n hoa v kt tri vo khong t thng 11 n thng 4.
 12

2.1.6. Phn loi [17], [21]

Cc ging c ph hin trng thuc:
B: c ph (Rubiales)
H: c ph (Rubiacea)
Loi: c ph (Coffea L.)
Cc ging c ph thng trng gm c:
2.1.6.1. C ph ch (Coffea arabica L.)
Ngun gc cao nguyn Jimma Ethiopia. C ph ch c trng lu i nht,
chim 70% sn lng c ph ton th gii.
Cy bi cao 3 4 m, iu kin thun li c th cao 6 7 m. Cnh mc tng
i i xng, cnh th cp nhiu, to vi cnh c bn mt mt phng ct ngang thn
cy. L mc i xng, hnh bu dc di, u nhn, ra l qun, xanh m, c 9 12
cp gn l, di 10 15 cm, rng 4 6 cm.
Hoa trng, hng thm du, mc thnh chm nch l. Qu hnh trng, khi
chn mu , c ng knh 10 18 mm. Ht c 2 lp v, lp ngoi cng gi l v
tru, lp trong mng bm st vo ht gi l v la. C ph ch c c tnh t hoa th
phn cho nn thun chng cao.
C ph ch c nhiu chng khc nhau:
Coffea arabica Var typica
Coffea arabica Var bourbon
Coffea arabica Var amarello
Coffea arabica Var managogype
Coffea arabica Var laurina
Coffea arabica Var mokka
Coffea arabica Var mundonovo
Coffea arabica Var catuai
Coffea arabica Var catimor
2.1.6.2. C ph vi (Coffea canephora pierre)
C ph vi mi c pht hin chu Phi vo u th k XX, vng rng thp
thuc chu th sng Cng g.
 13

Cy cao 8 12 m, c nhiu thn. Cnh di, cnh th cp t. L hnh bu, hoc

hnh mi mc, u l nhn, phin l gn sng mnh. Hoa mc thnh chm nch l,
gm 1 3 cm, c 15 30 hoa. Trng hoa trng c 5 7 cnh hoa.
Qu hnh trn hoc hnh trng, trn qu c nhiu gn dc, khi chn c mu .
Ht mu xm xanh c hoc ng vng ty theo ging. Ht nh hn c ph ch, t l
nhn/qu ti cao hn c ph ch.
C ph vi gm cc chng chnh sau:
Coffea canephora Var robusta
Coffea canephora Var kouilou
Coffea canephora Var niaouli
2.1.6.3. C ph mt (Coffea excelsa Chev)
Pht hin u tin nm 1902 x Ubangui Chari nn thng c gi l c
ph Chari.
Cy cao 15 20 m. L to, chiu di ti a ti 25 40 cm, hnh trng, hoc mi
mc, u l c mi ngn, bt, c 6 9 i gn l. Hoa mc thnh chm nch l gm
1 5 xim, mi xim c 2 4 hoa.
Qu hnh trng, hi dt, nm qu li, khi chn c mu sm, tht qu gin, t
ngt. Ht mu xanh ng vng, v la bm cht vo ht kh lm trc ht.
Hm lng caffeine thp: 1,02 1,15% trong khi c ph ch l 1,8 2% v c
ph vi l 2,5 3%.
Cy sinh trng khe, t su bnh, chu nng hn kh, t kn t. ng ch l
hoa qu pht trin c trn cc mt ng ca cnh gi, k c trn thn cy, ni phn cnh.
Phm cht c ph mt km, v chua, hng km hoc khng c hng.
2.1.6.4. C ph mt du da (Coffea liberia Bull. In Hirn)
Ngun gc Liberia.
Cy cao 15 18 m, cnh khe bn, l to hnh bu dc, di 15 35 cm, rng 8
15 cm, phin xanh m, bng, c 8 12 i gn ni r di. Hoa mc thnh cm,
trng hoa mu trng. Qu tha, to ging qu du da, tht qu dy nhiu x, nhn mu
vng, phm cht km.
Ngoi ra cn c loi Arabusta v Catimor, kt qu ca s tp giao nhn to
v t nhin gia 2 loi Arabica Robusta.
 14

Arabusta c hm lng caffeine thp, cho cht lng c ph tt hn Robusta v

sc khng bnh cao hn so vi Arabica.
Hin nay Vit Nam c 3 loi c ph chnh l: Arabica, Robusta v Chari.
Trong , Arabica v Robusta l 2 ging ph bin.
2.2. THNH PHN CA C PH NHN
2.2.1. Cu to ht c ph [40]

Hnh 2.4: Cu to ht c ph
2.2.2. Thnh phn ha hc trong ht c ph [2], [11], [28]
Ht c ph l i tng thu hoch ch yu, v cng l mt hng nng sn xut
khu quan trng. Cc bin php canh tc, thu hoch v ch bin ng vai tr quan
trng trong vic to ra c ph nhn sng t tiu chun xut khu.
Ht c ph xanh giu glucid v lipid, glucid chim hn 50%, phn ln l cc
polysaccharide. Lipid (10 15%), protein (10 15%) v acid hu c, c bit l cc
acid cafeylquinic hay acid chlorogenic.
Hm lng caffeine thay i, t 0,5 n 1,8%, mt phn ca caffeine thng
kt hp vi acid chlorogenic.
C ph rang ln c nhng cht thm, gi chung l cafeol (acid cafeic, oleic,
linoleic, palmitic), khong 0,05%, nhng ng thi li to ra mt yu t phc hp c
l cafeotoxin (0,07%).
Thnh phn ha hc trong ht c ph c nh hng rt ln trong qu trnh nh
gi cht lng th nm cm quan.
 15

Bng 2.6: Thnh phn ha hc ht c ph

Hm lng
Thnh phn
g/100 g mg/100 g
Nc 8 12
Cht du 4 18
m 1,8 2,5
Proteine 9 16
Caffeine 1 (Arabica), 2 (Robusta)
Chlorogenic acid 2
Trigonelline 1
Tannin 2
Caffetanic acid 89
Caffeic acid 1
Pentosane 5
Tinh bt 5 23
Saccharose 5 10
Cellulose 10 20
Hemicelllulose 20
Lignine 4
Calcium 85 100
Phosphur 130 165
St 3 10
Sodium 4
Manganese 1 45
(Ngun: L Quang Hng, 1999)
Caffeine l mt alkaloid cha m, cng thc l C8H10O2N4.H2O, c Runge
v Robiquet tm ra nm 1820. Caffeine kt tinh vi mt phn t nc nhng tr nn
khan khi nung nng n nhit 80 100oC. Caffeine khan cha 28,8% N.
Caffeine khan dng tinh th trng hnh kim, tan trong nc v chloroform.
Caffeine (1,3,7-trimethylxanthine) l purine chnh trong ht c ph. Ngoi ra
cn c theobromine (3,7-dimethylxanthine) v rt t theophylline (1,3-
dimethylxanthine).
 16

Phn b hm lng caffeine theo b phn ca cy nh sau

Bng 2.7: Phn b hm lng caffeine trn cy 2 tui theo t l (%)

B phn
L Thn R
Ging

Arabica 75,89 14,23 9,88

Robusta 85,29 8,33 6,38
(Ngun: L Quang Hng, 1999)
2.2.3. Cng dng ca c ph [7], [17]
C ph l mt loi nc ung cao cp c s dng rng ri hu ht cc nc
trn th gii. C ph l sn phm ca cc nc nhit i, nhng th trng tiu dng
ch yu li cc nc vng n i.
C ph c dng lm ung di dng chit hm bng nc si t bt ht c
ph rang, hoc di dng c ph ha tan. Ung c ph lm mt bun ng, gip tinh
thn tnh to, gim cm gic mt mi.
Hot cht ch yu cha trong ht c ph l caffeine. Ngoi ra, trong ht c ph
cn cha nhiu cht dinh dng cn thit cho c th con ngi nh ng saccharose,
ng kh, protein ha tan, cc vitamin nhm B (B1, B2, B6,) v vitamin PP l
nhng vitamin cn thit ca c th con ngi.
ng thi, c ph c mt hng v c o. Bng phng php phn tch
quang ph, ngi ta xc nh gn 300 hp cht phc tp khc nhau cu thnh nn
hng thm ca c ph. Do , tp tc ung c ph tr thnh nhu cu v thc ung
thng dng ca nhn dn nhiu nc trn th gii, c bit l cc nc pht trin Ty
u, Bc u, Nam M,
Ngoi ra, ph phm ca c ph nh tht qu cn l nguyn liu ch bin ru,
nc gii kht; v qu c ph lm phn bn; b ch bin lm thc n gia sc. Mt s
thuc cha bnh cng c ch to t c ph.
2.2.4. Dc tnh ca c ph [2], [28], [29], [37]
C ph c v ng, c tc dng kch thch thn kinh, lm tng hot ng ca
tim, co mch trung ng, co mch ngoi vi (mch phi, mi, thn, vnh).
 17

C ph cn c tc dng h st, li tiu, lm khoan khoi, kch thch tiu ha.

Cy c c nhng ch khi dng vi liu cao v thi gian ko di, gy bn chn, mt
ng, au dy thn kinh, trm cm, c t l cafeotoxin. Nhng nu ung c ph un
si th cafeotoxin s b tiu hy.
Trong nhn dn, ht c ph sng gi nt ngm vi ru ung cha t thp, st
rt. C ph rang ung c tc dng tiu m, tiu c ru v thuc phin. L c ph sc
ung lm chng tiu thc n, tiu nc cha ph thng.
Trong y hc, c ph thng c dng tr suy nhc, mt sc do bnh
nhim trng, mt trng lc d dy. Ngi ta c th pha bt c ph ung, dng
caffeine dng vin hoc dng thuc tim di da.
Tuy nhin, chng ch nh cho mt s trng hp: lon thn kinh, vim c tim
tin trin.
Tc dng dc l ca caffeine
i vi h thn kinh trung ng, caffeine c tc dng rt mnh, ngi dng
caffeine hoc ung c cha caffeine cm thy tnh to, mt bun ng, t mt mi v
suy t minh mn.
Caffeine dng vi liu ln th kch thch mnh h thn kinh trung ng, xut
hin trng thi bn chn, lo lng, mt ng, nhng ngi quen dng c ph, nhng
triu chng ny nh hn. Liu cao lm xut hin co git cc b hoc ton thn.
Caffeine cn c tc dng kch thch h hp. i vi h tim mch, caffeine c
tc dng lm gim tr khng ngoi vi, kch thch tim. Liu 250 350 mg caffeine gy
nn nhp tim nhanh, huyt p tng nh c huyt p tm thu v huyt p tm trng,
nhng cng vi liu trn i vi nhng ngi dng c ph th cc triu chng trn
khng xut hin. nng cao hn, caffeine c tc dng lm tng nhp tim r rt,
nhng ngi mn cm c th xut hin lon nhp tim (hin tng ny cng gp
nhng ngi dng qu nhiu ung c caffeine).
i vi c trn, caffeine c tc dng lm gin c trn kh ph qun c bit i
vi kh ph qun b co tht nh nhng bnh nhn hen suyn trn lm sng hoc trong
cc m hnh gy co tht kh ph qun thc nghim.
i vi c vn, caffeine c tc dng tng cng kh nng lm vic ca c bp
ngi. Caffeine vi liu 6 mg/kg lm tng kh nng chy ua ca nhng vn ng
vin trt tuyt.
 18

i vi h tit niu, caffeine tng cng s hnh thnh nc tiu, gia tng tc
lc ca cu thn v tng lng mu qua thn.
Alcaloid caffeine v cc dng mui nh caffeine citrate v caffeine natri
benzoate c dng trong trng hp cp cu ng c opioid v cc thuc gy ng
khc.
Mt cng trnh nghin cu ca trng i hc Gifu min Trung Nht Bn
cho bit: ung mt tch c ph/ngy c th ngn chn pht trin ung th rut kt ph
n. Tuy y ch l kt qu nghin cu mang tnh thng k, cha l gii c c ch
tc ng ca c ph ti ung th rut kt, nhng iu ny cng phn no cho chng ta
thy c tc dng dc l ca c ph. (Ngun: bo The Mainichi Shimbun)
T nm 1992, nhm nghin cu di s lnh o ca gio s Hiroyuki Shimizu
tin hnh theo di ch n ung ca 30.224 c dn vng Gifu, c tui t 35 tr
ln v khng mc bnh ung th rut kt thi im bt u nghin cu.
Nhm nghin cu chia tt c nhng ngi tham gia kho st thnh 3 loi:
khng ung c ph, ung di 1 tch c ph/ngy, v ung 1 hoc trn 1 tch c
ph/ngy.
Sau khi iu chnh cc d liu v thi quen ht thuc, ung ru, mc bo
ph, tui tc v cc yu t khc, nhm nghin cu tnh ton ri ro pht trin ung th
rut kt h. Kt qu cho thy, nu nguy c ung th rut kt nhng ngi khng
ung c ph l 1 th nguy c ung th rut kt nhng ph n ung t 1 tch c
ph/ngy tr ln ch l 0,49.
2.3. CC PHNG PHP X L C PH HIN NAY [6], [10], [11], [20], [21]
Sau qu trnh ch bin c ph nhn l cc qu trnh ch bin tinh khit hn nh
ch bin c ph rang, c ph bt th, c ph ha tan,hoc cc sn phm khc c phi
ch nh c ph sa, cc loi bnh ko c ph, Phm cht v hng v c ph ph
thuc vo mt s yu t chnh sau y: ging, kh hu, cao so vi mt bin, ph
ca t v ch bn phn, chn khi thu hi, k thut ch bin v ch bo qun.
Trong k thut ch bin c ph nhn, c 2 phng php chnh:
Phng php ch bin kh
Phng php ch bin t
Ngoi ra, cn c phng php trung gian l na kh na t. C ph em xt
ti cn ln c v qu em phi khng qua cng on ln men v ngm ra.
 19

Khi thu hoch ngi ta ch hi nhng qu chn, khng hi qu xanh v qu cn

non. C ph hi ngy no nn c ch bin ngay trong ngy hm , s lng cn li
khng nn thnh ng lu qu 24 gi. c c ph nhn phc v cho xut khu, t
l qu chn khi thu hi phi t trn 95%.
C ph l mt sn phm dng ch ra loi nc ung cao cp. cu to thnh
nhn ca c ph, theo nhng kt qu phn tch ngi ta thy c ti 670 hp cht hu
c hp thnh trong c nhiu hp cht to nn hng thm ca c ph. Do vy, cc
khu thu hi, ch bin, phi sy, bo qun c ph c mt v tr ht sc quan trng. Cc
cng on trong thu hoch, ch bin, phi sy v bo qun c mi lin kt khng kht
vi nhau.
2.3.1. Phng php ch bin kh
y l phng php ch bin lu i v n gin hn so vi phng php ch
bin t, thng c p dng trong ch bin c ph Robusta.
Trc tin, ngi ta phi hay sy nhm lm gim m ca tri c ph ti ,
trong khong 1 3 tun l.
Phng php phi nng c ph lm tn nhiu cng sc do phi thng xuyn
co o khi phi, phi che ph cho c ph vo ban m v khi tri ma. Trong nhng
iu kin thi tit thun li, cng vic phi nng cho c ph hon tt sau 10 14 ngy.
Vic phi c ph trn cc sn phi bng t s lm cho thnh phm c ph v
sau khi ung s c mi ca t. Ngoi ra, do s khng ng u v m ca khi ht
khi phi nng nn s to iu kin cho vic b nhim cc vi sinh vt, kt qu l lm
gim mi v ca c ph khi ung.
p dng cng ngh sy cho c ph s khc phc c cc khuyt im ca vic
phi nng nh vo cc thit b sy in hnh hin nay nh:
o My sy tnh c kt hp vi vic o trn
o My sy thng quay, vi tc quay ca trng trong khong 1100 2500
vng/pht
o My sy thp
o My sy tng si
Theo kt qu ca mt s cng trnh nghin cu, nhit khng kh sy cho c
ph nn trong khong 50 55oC.
 20

Sau khi hon tt cng on phi/sy (v c ph ht c mu nu m v gin),

c ph ht c thu gom li v chuyn sang giai on bc v thu c c ph nhn
v em i bo qun.
2.3.2. Phng php ch bin t
Phng php ch bin t tn km v phc tp hn so vi phng php ch
bin kh. Tuy nhin, phng php ny sn xut ra c ph nhn sch v phm cht tt
hn. Phng php ny tiu th mt lng nc ng k, v c ng dng rng ri
nhiu ni trong sn xut Arabica, c bit Kenya v Colombia. i khi phng php
ny cng c p dng trong sn xut ging c ph lai Arabusta.
Phng php ch bin t nhm bin i tri c ph ti ban u thnh dng c
ph thc, bao gm cc cng on sau:
o Thu nhn
o Xt v
o Tch lp nhy (ln men)
o Ra v phn loi theo t trng dng nc
o Phi/sy
o Tch v
2.3.2.1. Thu nhn
C ph tri sau khi thu hoch c em v a im tip nhn v cho ngay vo
b y nc c trang b ng siphon phn loi, nhm mc ch phn loi: tri no
nng s chm xung b (em i ch bin t), tri no chn kh s nh, ni ln trn
(c em i ch bin kh). Ngoi ra, cnh, l, tp cht, ni ln trn mt nc cng
c loi ra khi khi tri.
2.3.2.2. Xt v
y l giai on c xem l quan trng nht trong phng php ch bin c
ph t. iu cn thit l vic xt v cn thc hin cng sm cng tt, nhm trnh tin
trnh ln men ngoi mun v pht sinh cc v l trong ht c ph.
Giai on ny nn hon tt trong vng 12 24 gi sau khi hi tri, nhng nu v
l do no khng th thc hin ngay c, trong khi ch em i xt v, khi tri nn
c bo qun trong b nc.
Xt v l s tc ng c gii vo tri c ph trong dng nc.
Thit b xt v thng dng c 3 dng sau y:
 21

o My xt a
o My xt hnh trng vi trc nm ngang
o My xt hnh trng vi trc thng ng
Yu cu quan trng cho cc thit b xt v l phi thc hin chnh xc cng vic
tch v bn ngoi m khng lm tn thng n v thc v ht c ph bn trong.
Trong phng php ch bin t, nguy c nhim vi sinh vt d dng xy ra mt
khi trn v thc v ht c xut hin cc thng tn, kt qu l s lm nh hng n
phm cht c ph v sau.
Lp tht tri c ph sau khi c tch ra thng c s dng lm phn
xanh. Ngoi ra, ngi ta cn c th trch ly chng bng nc hoc cc dung mi hu
c thu c caffeine v cc hp cht khc, phn b cn li c th ch bin lm thc
n cho gia sc.
2.3.2.3. Tch lp nhy (ln men)
B dy ca lp nhy trong tri c ph khong 1,5 mm nhng thay i ty theo
ging c ph. Lp nhy khng ha tan trong nc, khng c cu trc t bo r rt
nhng th keo v nh hnh, n kt dnh rt cht vo ht c ph. Lp nhy c ph
cha khong 85% nc lin kt lng v 15% cht rn dng khng ha tan.
Bng 2.8: Thnh phn ha hc ca lp nhy c ph
Thnh phn Phn trm
Nc 84,2
Protein 8,9
ng 4,1
Pectic acid 0,9
Tro 0,7
Tng s 98,8
(Ngun: Phm Tr Thng, 1999)
Ngoi ra, cn c mt s cc enzyme c to ra trong lp nhy v trong lp tht
bao quanh lp nhy nh protopectinase, pectinase, pectinesterase v pectase. Cc
enzyme ny b gy cc phn t ln bng cch ct t cc lin kt polymer, dn dn to
thnh cc chui ngn hn cho ti khi cn cc acid n phn t v cc ester.
Khi tri c ph c tin hnh xt v, v s cc vi sinh vt nh nm men, nm
mc, vi khun, trong mi trng chung quanh khi ht tm thy y mt mi
 22

trng v cng thun li cho s pht trin ca chng. Trong qu trnh pht trin,
chng t sn xut ra cc enzyme v tc ng ln lp nhy ca khi c ph xt v.
Tin trnh ln men thng thng do cc vi sinh vt, u tin l cc loi nm
men tm thy cc iu kin thch hp pht trin v trong qu trnh pht trin, chng
lm gia tng hm lng cn t cc cht ng trong khi . Sau , cc vi khun s
dng cc cht cn ny lm thc n, to thnh cc acid lactic, acid butyric v cc
acid carboxylic do s oxi ha trong qu trnh ln men. Khi acid butyric bt u hnh
thnh cng l lc v acid pht trin trong ht c ph.
Sau khi xt v, c ph c tin hnh tch lp nhy (giu pectin) cn dnh vo
v thc. Trong k thut ch bin c ph theo phng php t, ngi ta c nm bin
php tch lp nhy c ph nh sau:
o Ln men t nhin
o Bin php sinh ha, s dng enzyme
o Bin php ha hc, vi xt (NaOH) hoc sodium carbonate (Na2CO3)
o Nc m
o Ma st
a. Ln men t nhin
Tin hnh ln men khi c ph trong cc b ln men bng xi mng hoc thng
g. Hin nay c hai phng php ln men t nhin l ln men kh (x ht nc tch
dch ln men) v ln men t (ln men khi c ph trong khi nc).
b. Bin php sinh ha
Nhm thc y nhanh tin trnh phn hy lp nhy, ngi ta cn s dng cc
enzyme b sung vo khi c ph nh pectase, protopectinase, pectinase v
pectinesterase.
c. Bin php ha hc
tch lp nhy nhanh, ngi ta cn nghin cu dng sodium hydroxide
(NaOH), sodium carbonate hoc vi. Khi c ph xt v c cho vo b cha bng
b tng. Sau , x ht nc ra v cho hn hp kim pha long vo b va ph
kn mt khi ht. Khi lp nhy tan chy ht th tin hnh ra khi ht nh thng l.
d. Nc m
Mt phng php n gin tch nhanh lp nhy l tin hnh trn mt khi
lng bng nhau gia khi c ph v nc sinh hot, ri un nng hn hp ny n
 23

nhit 50 3oC. Do nc m c kh nng b gy cc cu trc ca pectin tn ti th

keo nn lp nhy d dng tch ra.
e. Ma st
Nhm mc ch tch lp nhy, cc bin php v c kh cng c p dng nh
dng my Raoeng hay my Aquapulpa.
2.3.2.4. Ra
Mc ch ca cng on ra l tch cho trit cht nhy cn dnh bm trn v
thc. V ch vi mt t cht nhy cn st li s b cc vi sinh vt tn cng v lm cho
ht c ph b ln men ngoi mun trong cc cng on ch bin tip theo, v dn n
hu qu l lm tn hi n phm cht v v c ph sau ny.
Cng on ra c th thc hin ngay trong cc b ln men trc bng cch
cho nc vo b v khuy trn ht bng th cng.
Ngoi ra, cng vic ra c th thc hin bng c gii nh vo cc my ra.
2.3.2.5. Phi / Sy
Cng vic phi/sy cho c ph trong phng php ch bin t n gin hn so
vi trong phng php ch bin kh bi v:
o m ban u ca c ph t rt ng u
o Lng nc cn lm cho bc hi t
o C th sy ti cc nhit sy v tc sy cao
Hin nay, c ba phng php sy c ph t sau y:
o Sy t nhin
o Sy nhn to
o Sy kt hp (t nhin v nhn to)
a. Sy t nhin
Sy t nhin hay cn gi l phi nng thch hp cho nhng ni no c y nh
nng mt tri.
Sau khi phi, thnh phm c ph t thu c gi l c ph thc v c khong
10 kg c ph t sau khi phi s cn li khong 5 kg c ph thc.
b. Sy nhn to
Cc my sy dng trong sy c ph t v c bn cng tng t nh cc my
sy cho c ph ch bin theo phng php kh.
 24

c. Sy kt hp
C ph c th c p dng sy nhn to trong giai on u ri sau em i
phi nng h m ht n gi tr mong mun.
2.3.2.6. Tch v
C ph d ch bin theo phng php no, khi mun thu c c ph nhn u
phi tri qua cng on tch v. i vi c ph ht th cn tch lp v kh bao bc ht
c ph, cn i vi c ph thc th cn tch lp v thc.
C mt s my tch v ch dng ring cho c ph t hoc kh. Tuy nhin,
ngy nay vic tch v cho cc loi c ph c th dng chung mt my vi nhng
iu chnh cn thit cho ph hp vi tng loi c ph.
Mt im cn lu l cho d l loi c ph no i chng na th khi em vo
tch v, c ph phi hon ton kh ro, nu khng s xy ra hin tng tc nghn v
qu ti cho my.
2.3.2.7. Cc cng on khc
Trong cng ngh ch bin c ph nhn cn qua thm mt s cng on ph tr
khc, bao gm nh bng, sng lc v phn loi.
a. nh bng l cng on nhm tch lp v bc.
b. Sng lc c mc ch tch c ph c khuyt im ra khi khi c ph nhn
hon ton tt, cng nh loi b cc tp cht khc.
c. Vic phn loi c ph nhn vi mc ch tch ra c cc ht c ph thng
phm ho hng. Hin nay, trong cng ngh sn xut c ph c nhiu h thng phn
loi v ty theo yu cu ca thnh phm sau cng m ngi ta s dng cc phng
php phn loi khc nhau:
o Phn loi theo kch thc
o Phn loi theo trng lng
o Phn loi theo mu sc
2.3.3. Bo qun v vn chuyn c ph nhn
C ph nhn thng c bo qun tm mt thi gian trong cc kho cha trc
khi c chuyn ch n cc nc tiu th. Cc bao b (ay) (ng khong 60 70
kg) vn cn c s dng rng ri d vic vn chuyn c ph ri trong cc thng hng
(containers) ngy cng ph bin. Ti kho cha, cc bao c ph nhn c xp trn
nhng gi g. Nn tin hnh thng thong kho thng xuyn trnh tch t m.
 25

Ch bin kh Ch bin t
Tri ti

Thu nhn/B nc
Tp cht nh

Phi/sy Xt v
V

C ph tch v

Tch lp nhy
C ph ht Lp nhy

Ra v phn loi

C ph thc t

Phi/Sy

C ph thc kh

Lm sch s b
Bi bm v tp cht

Sy li (nu cn thit)

Tch v ht/v thc V ht, v thc

Phn loi theo t trng V, ht lp, mnh v

Phn loi theo mu sc Ht en, nht mu, khuyt im khc

ng gi/cn C PH THNG PHM

S 2.1: S ch bin c ph nhn

 26

TM HIU V PECTIN V CELLULOSE

Phin gia

Mng nguyn
sinh

Vch plasma

Hnh 2.5: Cu trc thnh t bo thc vt

2.4. PECTIN [13], [39], [43]

Hnh 2.6: M hnh cu trc pectin trong thnh t bo thc vt

2.4.1. nh ngha
Pectin l mt phn t tng t nh tinh bt, khc ch n v tun hon ca
pectin l acid galacturonic thay v glucose tinh bt. Pectin l hp cht cao phn t
mch thng c cu to t s kt hp ca cc acid galacturonic qua cc lin kt -1,4-
glucoside.
 27

Trong thc vt, pectin c chc nng t nhin nh mt cht keo gi thnh t
bo cng nh cc t bo li vi nhau. Ty thuc vo ngun pectin m pectin c khi
lng phn t t 80.000 200.000 Da.
Pectin khng ha tan trong ru v cc dung mi hu c khc. Pectin tan trong
nc, NH3, dung dch kim, Na2CO3 v trong glycerin nng.
2.4.2. Cu trc

Hnh 2.7: Cu trc ha hc ca phn t pectin

2.4.3. Phn loi
Trong thc vt, pectin tn ti di 3 dng:
o Pectin ha tan: l ester methylic ca acid polygalacturonic.
o Pectinic acid: l polygalacturonic acid c mt phn nh cc nhm carboxyl
c ester ha bng methanol.
o Protopectin: to cng cho qu xanh, khng tan trong nc. Trong thnh
phn c cc phn t pectin, cc phn t cellulose v cc ion Ca2+, Mg2+, cc gc
phosphoric acid, acetic acid v ng. Protopectin khi b thy phn bng acid th gii
phng pectin ha tan.
2.5. CELLULOSE [12], [13], [23], [30]
Mi nm c khong 232 t tn cht hu c c thc vt tng hp nh qu
trnh quang hp. Trong s ny c 172 t tn c to thnh trn t lin v 60 t tn
 28

c to thnh trong cc i dng. Trong s ny c n 30% l mng t bo thc

vt m thnh phn ch yu l cellulose. Cellulose chim n 89% trong bng v 40
50% trong g.
Ligno-cellulose l thnh phn cu trc chnh ca cy g v cy thn mm (c,
rm r,) gm cellulose, hemicellulose v lignin.
2.5.1. nh ngha
Cellulose l polymer thng ca cc n v -D-glucose c ni vi nhau qua
lin kt -D-1,4-glucan. Nh phng php phn tch bng tia Rnghen ngi ta bit
rng cellulose c cu to dng si. Cc si ny lin kt li thnh nhng b nh gi l
cc microfibril (vi si) c cu trc khng ng nht, chng c nhng phn c (phn
kt tinh) v nhng phn xp hn (phn v nh hnh).
2.5.2. Cu trc
Cc phn t cellulose kt hp vi nhau bng lc ht Vanderwall v lin kt
hydro. Chng to nn si s cp, si s cp kt hp vi nhau to thnh vi si.

Hnh 2.8: Cu trc cellulose trong t bo thc vt

2.5.3. Tnh cht
Cellulose c trng lng phn t t 50.000 n 2.500.000 Da..
Cellulose thng cha 10.000 14.000 gc ng.
 29

Cellulose l mt trong nhng hp cht t nhin kh bn vng. N khng tan

trong nc m ch c th b phng ln do hp thu nc, b phn hy khi un nng vi
acid hoc kim nng kh cao. Cellulose b thy phn nhit bnh thng hoc
nhit 40 50oC nh cc enzyme thy phn cellulose c gi chung l
cellulase.
Trong iu kin t nhin, cellulose b phn hy bi vi sinh vt c trong iu
kin hiu kh ln ym kh. Cc loi vi sinh vt thay phin nhau phn hy cellulose n
sn phm cui cng l glucose. Tuy nhin, s phn gii cellulose trong iu kin t
nhin thng rt chm v thng khng trit .
Trong t bo thc vt, cellulose lin kt cht ch vi hemicellulose (chim 20
40% trng lng kh) l mt loi heteropolymer cha nhiu loi monosaccharide nh
galactose, mannose, glucose, xylose, arabinose v cc nhm acetyl. Do bn cht khng
kt tinh nn hemicellulose tng i d b thy phn. Cellulose cn lin kt cht ch
vi lignin (10 25% trng lng kh). y l thnh phn nh hng rt nhiu n s
thy phn cellulose ca enzyme. Ch trong mt s trng hp (v d trong si bng)
cellulose tn ti trng thi mt polymer gn tinh khit.
Vic s dng c cht lignocellulose sn xut cc enzyme thy phn c cht
ny c tim nng ng dng trong nhiu ngnh cng nghip nh ha cht, nhin liu,
thc phm, ru bia, thc n gia sc, vi si, bt git, giy, bt giy,
2.5.4. V tr
Cellulose l ngun hu c nhiu nht trong thc phm, cht t v ha cht.
Cellulose tn ti mt lng rt ln trong thc vt, khng u cc c quan
khc nhau, t nht l v nhiu nht l thn cy.
S hin din ca cellulose ng vai tr l mt vt liu ph bin ca vch t bo
thc vt c pht hin ln u tin bi Anselm Payen vo nm 1838. N tn ti hu
ht nhng hnh thc nguyn trong si cotton v trong t hp vi nhng vt liu khc,
nh lignin v hemicellulose, trong g, l v thn Mc d, nhn chung c xem nh
vt liu thc vt nhng cellulose cng c sn xut bi mt s vi khun.
Cellulose l nn tng b xng ca thnh t bo thc vt. Ngoi ra ngi ta cn
thy chng c nhiu t bo mt s loi vi sinh vt (cellulose vi sinh vt thuc loi
v nh hnh, vic phn hy thng d thc hin). Cellulose khng c trong t bo
ng vt.
 30

TM HIU V H ENZYME PECTINASE V CELLULASE

2.6. ENZYME [12], [13]

Ch enzyme c bt ngun t ting Hy Lp c ngha l cht trong nm
men.
Enzyme c cc c th sinh vt tng hp nn v tham gia cc phn ng ha
hc trong c th. Enzyme l mt cht hu c, trong khi cc cht xc tc ha hc
thng l cht v c. Sau ny, cc nh khoa hc xc nh chng l protein.
Nh vy enzyme l mt protein c kh nng tham gia xc tc cc phn ng ha
hc trong v ngoi c th.
Pavlov ni: Hot ng ca enzyme l biu hin u tin ca hot ng
sng. Khng c s sng no li khng c qu trnh enzyme. Cc enzyme xc tc cho
hu ht cc phn ng ho hc xy ra trong c th sng, bo m cho cc qu trnh
chuyn ha cc cht trong c th sng tin hnh vi tc nhp nhng cn i, theo
nhng chiu hng xc nh. Enzyme bo m cho s trao i thng xuyn gia c
th sng v mi trng ngoi, ngha l bo m iu kin tin quyt cho s tn ti ca
c th sng.
Enzyme khng nhng hot ng xc tc trong t bo m cn ngoi t bo sinh
vt, chnh iu ny c ngha quan trng trong vic ng dng enzyme vo nhiu lnh
vc khc ca cng nghip v i sng .
Enzyme c hiu sut xc tc cc k ln, hot xc tc ca enzyme ln gp
hng trm, hng nghn hoc hng triu ln cc cht xc tc v c v hu c khc. V
d nh trong phn ng thy phn saccharose nu dng saccharase lm xc tc tc
phn ng tng nhanh gp 2 1012 ln so vi khi dng acid lm xc tc.
iu quan trng l enzyme c th thc hin hot ng xc tc trong iu kin
nh nhng, p sut v nhit bnh thng ca c th, pH mi trng gn pH sinh
l. Hn na enzyme li c kh nng la chn cao i vi kiu phn ng m n xc tc
cng nh i vi cht m n tc dng.
V enzyme l nhng cht khng th ch bin c bng phng php tng hp
ha hc nn ngi ta thng thu chng t cc ngun sinh hc. Enzyme c trong tt c
cc c quan, m ca ng thc vt cng nh trong t bo vi sinh vt.
 31

Enzyme c thu nhn t nhiu ngun khc nhau: ng vt (pepsin t d dy,

trypsin t ty tng.), thc vt (amylase t thc ny mm, bromelin t thm). Tuy
nhin khng th dng hai ngun ny lm nguyn liu sn xut cc ch phm
enzyme vi qui m cng nghip v nhng l do nh kh tm, tc sinh sn chm, t
tin, Mc d vy, ngi ta c th tn dng ph phm t hai ngun ny lm mi
trng v thc n nui cy vi sinh vt cho sn xut enzyme.
Trong cc ngun nguyn liu sinh hc th ngun nguyn liu vi sinh vt (nm
men, nm mc, vi khun) l di do v y ha hn. Vi sinh vt l gii sinh vt
c xem nh nhiu nht v c kh nng chuyn ha vt cht trong thin nhin mnh
nht. Hin nay ngi ta khai thc enzyme nhiu t vi sinh vt v c ng dng rt
nhiu trong i sng, sn xut.
H enzyme vi sinh vt v cng phong ph. Vi sinh vt c th ng ha bt
k cht no trong thin nhin, trong khi c nhiu cht m ng vt v thc vt khng
th ng ho c.
Enzyme ca vi sinh vt c hot tnh rt mnh, vt xa cc sinh vt khc. Do
vy ch cn s dng mt lng nh enzyme c th chuyn ho mt lng ln c cht.
Ngi ta tnh rng trong 24 gi, vi sinh vt c th chuyn ha mt lng thc n gp
30 40 ln so vi trng lng c th ca chng.
Vi sinh vt sinh sn vi tc cc k nhanh chng, tuy khi lng nh,
kch thc b, nhng t l enzyme trong t bo tng i ln nn qu trnh sn xut
ch phm enzyme kh d dng, thao tc thun li, hiu sut thu hi cao. Trong mt
thi gian ngn, ta c th thu nhn c mt khi lng enzyme rt ln. i vi mt s
trng hp c th dng 100% sinh khi vi sinh vt lm ngun enzyme.
Phn ln thc n nui vi sinh vt li d kim v r tin. a s vi sinh vt
cho enzyme thng c kh nng pht trin trn mi trng n gin r tin nh cc
ph ph liu, ph phm ca cc ngnh sn xut.
u im ln ca vic thu nhn enzyme t vi sinh vt l c kh nng tng
cng sinh tng hp cc enzyme nh chn ging khi to c nhng bin chng c
hot lc cao. Vi sinh vt rt nhy cm vi tc ng mi trng, c kh nng thch ng
vi ngun dinh dng. V vy, khi thay i iu kin sng ca vi sinh vt hoc tc
ng ln chng bng cc tc nhn khc nhau c th thay i d dng h enzyme cng
 32

nh hot tnh ca chng v gia tng ti a qu trnh sinh tng hp. iu cho php ta
c th to c nhng enzyme theo mun, d dng thu nhn c enzyme c
thun khit cao.
Ngy nay vi s pht trin khng ngng ca khoa hc, c bit trong lnh vc
sinh hc ni chung v cng ngh sinh hc ni ring, cng vi nhu cu enzyme trong
cc ngnh cng nghip ngy cng cao, phng php sinh tng hp enzyme ngy cng
c p dng rng ri, v c hiu qu trong nhiu nc trn th gii. Nht Bn l mt
trong nhng quc gia i u trong lnh vc ny, Nht c hn 20 cng ty sn xut
ch phm enzyme. Mi nm Nht sn xut khong 9850 tn amilase, 8960 tn
protease, 2200 tn glucoamilase, 100 tn lipase, 200 tn enzyme khc cc nc
pht trin nh c, Hungari, Nga, Php, Anh, an Mch, Trung Qucngnh cng
nghip sn xut enzyme cng c ch v pht trin kh mnh.
2.7. PECTINASE [12], [13], [39]
2.7.1. nh ngha
Enzyme pectinase l enzyme xc tc s phn hy ca cc polymer pectin. S
phn hy pectin trong t nhin thng xy ra khi tri cy chn. Nhng enzyme ny c
mt vai tr ht sc quan trng trong qu trnh bo qun tri cy v rau qu.
Enzyme pectinase cng c ng dng nhiu trong ch bin thc phm, c
bit l kh nng lm trong nc qu.
2.7.2. ng dng
Pectinase c to ra trong sut qu trnh chn t nhin ca mt s qu (cng
vi cellulase), chng gip lm mm thnh t bo ca qu.
2.7.3. Phn loi
Enzyme pectinase c th c phn loi theo c ch tc dng ca chng
o Pectinesterase (PE): xc tc s thy phn ca cc nhm methyl ester.
o Polygalacturonase: cn c tn gi l poly -1,4-
galacturoniglucanohydrolase, xc tc s phn ct cc mi lin kt -1,4-glycoside.
Polymethylgalacturonase: hay cn gi l -1,4-galacturonite-
methylesglucanohydrolase, tc dng trn polygalacturonic acid c methoxyl ha.
Enzyme ny li c phn thnh 2 nhm nh l endo-glucosidase-
polymethylgalacturonase kiu I v exo-glucosidase-polymethylgalacturonase kiu III.
 33

Polygalacturonase: enzyme tc dng trn pectic acid (hoc pectinic),

cng c chia thnh 2 nhm nh l endo-glucosidase-polygalacturonase kiu II v
exo-glucosidase-polygalacturonase kiu IV.
o Pectate lyase (PEL): xc tc s phn ct cc n v galacturonate khng b
ester ha. Cc enzyme vi sinh vt ngoi bo ny ng mt vai tr rt quan trng trong
qu trnh gy bnh thc vt, gy ra s phn hy m ca thnh t bo, lm mm v
lm mc m thc vt.
Ngoi ra cn c:
o Pectin-transeliminase
o Polygalacturonate-transeliminase
o Pectin lyase (PNL)
2.7.4. Chc nng
Pectinase lm t trng thi gn cht gia nhng phn t acid galacturonic
lm ngn chui pectin t nhng on ln thnh nhng on nh hn.
C ch xc tc ca pectinase s dng nc, v th pectinase c xem l mt
enzyme thy phn. N ct t phn t nc, gn H vo mt C v gn OH vo mt C
khc.

Hnh 2.9: Hot ng ca pectinase

2.7.5. Cc vi sinh vt tng hp pectinase
o Nm mc
Aspergillus awamori, Aspergillus foetidus, Aspergillus niger,
Aspergillus terrus, Aspergillus saitoi
Fusarium moniliforme
 34

Penicillium glaucum, Penicillium ehrlichii, Penicillium chrysogenum,

Penicillium expanam, Penicillium cilrrimim,
o Nm men
Saccharomyces fragilis
o Vi khun
Bacillus polymyxa
Flavobacterium pectinovorum
Klebsiella aerogenes,
Ngoi ra, pectinase cng c tit ra bi mt s mm bnh cy trng nh:
o Nm Monilinia fructigena
o Vi khun gy thi ra Enwinia carotovora
2.8. CELLULASE [12], [13], [26], [34]
2.8.1. nh ngha
Cellulase l mt phc h hydrolase gm t cellulase C1 n cellulase Cx v
-glucosidase. Chng s phn hy ln lt cellulose cui cng to ra sn phm
ng glucose.
2.8.2. Tnh cht
Hot tnh ca h enzyme cellulase t cao nht nhit nm trong khong 40
60oC, pH nm trong khong 4 7.
Cellulase b c ch bi nhng sn phm phn ng ca n nh glucose,
cellobiose.
Hg c ch hon ton cellulase, trong khi cc ion nh Mn2+, Ag+, Cu2+ v Zn2+
ch c ch nh.
Hot tnh ca ch phm cellulase b mt hon ton sau 10 15 pht 80oC.
2.8.3. ng dng
Hin nay, cellulase t vi sinh vt c sn xut vi quy m cng nghip v
c ng dng vo nhiu mc ch khc nhau. Cc ng dng quan trng nht c th
k n l:
Trong cng nghip thc phm, cellulase c s dng sn xut glucose,
mt ng t nguyn liu cha nhiu cellulose nh rm, r, g vn, mt ca,
Trong cng nghip sn xut ethanol t cc ph liu c cha cellulose.
 35

Trong chn nui ng vt n c: vic b sung cellulase vo thc n s tng

cng s tiu ha hp th thc n cc loi ny.
Trong sn xut dc liu c ngun gc thc vt: cellulase ph v thnh t
bo thc vt gip cho vic trch ly c d dng.
2.8.4. Phn loi
Enzyme cellulase c phn chia theo chc nng thnh 3 nhm sau :
Endocellulase
Exocellulase
-glucosidase
Theo danh php quc t th c phn chia thnh 3 nhm:
1,4- -D-glucan cellobiohyrolase
1,4- -D-glucan-4-glucanohydrolase
-D-glucoside glucohydrolase
2.8.5. Chc nng
Cellulase Cx cn c gi l endocellulase hay endoglucanase, tn khc l
1,4- -D-glucan-4-glucanohydrolase.
Thy phn lin kt -1,4-glucosid v tr ngu nhin trong chui cellulose,
chui cellodextrin v cc dn xut cellulose nh Carboxyl Methyl Cellulose (CMC) v
Hydroxyethyl Cellulose (HEC).
Chng tham gia tc ng mnh n cellulose v nh hnh, tc ng yu n
cellulose kt tinh.
S thy phn ca endocellulase to ra nhng chui cellodextrin c u khng
kh.
Cellulase C1 cn c gi l exocellulase hay exobiohydrolase, tn khc l
1,4- -D-glucan cellobiohyrolase.
Thy phn c hiu lin kt -1,4-glucosid u khng kh ca chui
cellulose hoc cc cellodextrin gii phng cellobiose (cu trc gm 2 phn t D-
glucose ni vi nhau bng lin kt -1,4-glucosid).
Enzyme ny khng c kh nng phn gii cellulose dng kt tinh m ch lm
thay i tnh cht ha l, gip cho enzyme endocellulase phn gii chng.
 36

-glucosidase (cellobiase), tn khc l -D-glucoside glucohydrolase.

Chng khng c kh nng phn hy cellulose nguyn thy m tham gia phn
hy cellobiose, cellodextrin to thnh D-glucose.
2.8.6. H vi sinh vt c kh nng tng hp enzyme cellulase
H vi sinh vt c kh nng sinh tng hp cellulase kh phong ph, thuc 3
nhm: nm si, x khun v vi khun. Trong cng nghip sn xut cellulase hin nay,
ngi ta ch yu nui cy nm si v x khun, cn vi khun t c s dng.
o Nm si
Ch yu l nm mc v nm qu th, a s c kh nng sinh tng hp hai hoc
nhiu exocellulase, mt vi endocellulase v mt t -glucosidase. Cc enzyme ny
u l enzyme ngoi bo (c nm tit ra ngoi mi trng nui cy).
Aspergillus fumigatus, Aspergillus flavus, Aspergillus niger, Aspergillus
oryzae, Aspergillus sydovii, Aspergillus terrus
Trichoderma koningi, Trichoderma lignorum, Trichoderma roseum,
Trichoderma reesei
Penicillium spp
Altenaria tenuis
Fusarium culmorum, Fusarium oxysporum, Fusarium solani,
o X khun
Actinomyces spp,
Chim u th l loi x khun chu nhit Thermonospora cruvata c
phn lp t phn rc .
o Vi khun
H vi sinh vt trong d dy ca ng vt n c, v d: Cillobacterium
cellulosolvens, Ruminococcus albus, Ruminobacter parum,
Celluvibrio gilvus
Acetobacter xylinum,
 37

Trong , Trichoderma reesei c bit n nh mt vi sinh vt sn xut

cellulase rt tt. Mt s chng t bin ca Trichoderma reesei c th sn xut 160
250 UI/g cellulase.

Hnh 2.10: Hot ng ca cellulase

Ta c th tm tt ton b qu trnh thy phn cellulose nh sau

 38

S 2.2: Qu trnh thy phn cellulose

 39

Vng kt tinh u kh
u khng kh
Vng v nh hnh

Hnh 2.11: M hnh s phn hy cellulose tinh th

 40

TM HIU V NM SI

2.9. NM SI (NM MC) [5], [33]

Nm mc (molds, moulds) l tn chung ch tt c cc nhm nm khng phi
l nm men m cng khng phi l cc nm ln c m nm (nh nm rm, nm
g,).
Nm mc bao gm nhiu loi nm thuc cc nhm phn loi khc nhau. Nm
mc phn b rng ri trong t nhin (trong t, trong phn chung, trong nc, trong
khng kh, trn cc loi nguyn vt liu, lng thc, thc phm, hoa qu,).
Nm mc gp phn quan trng trong vic m bo cc vng tun hon vt cht
trong t nhin. Chng c kh nng phn gii mnh m cc hp cht hu c phc tp
(nh pectin, cellulose, protein, lipid,).
T lu nhn dn ta bit s dng cc qu trnh ln men truyn thng nh nm
mc, v d nh ch to tng. Trong nhng nm gn y, nm mc c s dng
rng ri trong cng nghip ln men hu ht cc nc trn th gii.
Nhiu loi nm mc c dng trong cng nghip sn xut cc ch phm men
(amylase, protease, lipase, pectinase, cellulase,) hoc c dng ch bin cc loi
thc phm (ng ha tinh bt, ch to nc chm, thc n gia sc,). Nhiu loi
nm mc c s dng tng hp quy m cng nghip cc loi acid hu c (acid
citric, acid lactic, acid oxalic, acid malic,). Cc loi acid ny u l nhng mt hng
c gi tr trong cng nghip ha cht, cng nghip thc phm, cng nghip in nhum,
cng nghip dc phm,
Mt s nm mc c kh nng tch ly vitamin (B2, -caroten) hoc cc cht
sinh trng (gibberelin, heterauxin,). Mt s nm mc c kh nng sn sinh nhiu
loi alkaloid c gi tr cha bnh cao (amanitin, bufofein, muscarin,).
Nm mc cn c s dng nh nhng tc nhn ha hc chuyn ha mt
cch tinh vi v chnh xc tng gc hp cht xc nh trong phn t cc cht thuc loi
steroid. Nh tc dng ny m cng ngy ngi ta cng c thm nhiu dc phm qu
gi.
Gn y, nhiu loi nm mc c nghin cu s dng sn xut mt s
acid amin hoc mt s nucleotide v nucleozide (dng trong cng nghip thc phm
 41

hoc trong y dc). C loi nm mc ang c nghin cu tng hp c cao su

nhn to.
Mt s nm mc c s dng nh nhng vi sinh vt ch th trong vic nh
lng vitamin, nh lng acid amin, nh lng nguyn t vi lng, kim tra hm
lng N v P d tiu trong t, v nht l kim tra hiu qu tc dng ca cc cht
chng ung th.
Nm mc cn l ngun ca hng lot cht khng sinh c gi tr nh Penicillin,
Cephalosporin, Fuzidin, Eniatin, Fungisporin, Verucarin,
C th ni kh nng ng dng nm mc ang tng ln hng ngy, hng gi trn
th gii. Tuy nhin, bn cnh nhng li ch nhiu mt, nm mc cng l nguyn nhn
gy ra nhiu tn tht ht sc to ln cho vic bo v ma mng, bo qun lng thc,
thc phm, hng ha, vi vc, phim nh, sch v,
Nhiu loi nm mc cn gy nn nhng bnh kh ph bin v kh iu tr
ngi, gia sc v cy trng.
Cc bnh nm ngi (nh hc lo, nm vy rng, nm k chn, nm tc, nm
ton thn,) rt hay gp nc ta v gy ra nhng nh hng khng nh i vi
sc khe v nng sut lao ng.
Nm mc c cu to hnh si phn nhnh, nhng si ny sinh trng nh
v pht trin rt nhanh, to thnh mt m chng cht cc si. Tng si c gi l
khun ti hay si nm. Cn c m si th c gi l khun ti th hay h si nm.
Nm mc l loi thc vt khng c dip lc, do khng c kh nng tin hnh
quang hp nh cc loi cy xanh. Chng sng nh cc loi thc n hu c c sn bng
kh nng hp th ca khun ti. l cc c th k sinh hay hoi sinh.
Mt s khun ti pht trin su vo c cht, gip nm mc bm cht vo c cht
v hp th cc loi thc n cha trong (khun ti c cht hoc khun ti dinh dng).
Mt s khun ti khc pht trin trn b mt c cht (khun ti kh sinh). Mi on
khun ti khi ri vo mi trng thch hp s pht trin nhanh chng thnh mt m
nm mc mi.
mt s loi, khun ti c phn ha thnh nhng si bm trng ging nh r
cy, chng c gi l r gi.
 42

Hai loi c quan dinh dng thng gp l th m v hch nm.

Th m trng ging nh ci m gh, cu to bi nhiu khun ti kt cht li
vi nhau. C quan sinh sn thng c sinh ra bn trn hoc bn trong th m.
Hch nm thng c hnh hi trn hoc hnh khng u, bn ngoi thng c
mu ti, bn trong l t chc si xp hn v thng c mu trng. Kch thc ca
hch nm thay i ty theo tng loi nm (t vi phn ca mm n vi cm, thm ch
mt s nm nhit i, hch nm c ng knh ln n vi dm).
Hch nm c th gip cho c th nm sng qua nhng iu kin ngoi cnh bt
li trong mt thi gian kh di. Khi iu kin sng tr li thun li, hch nm s ny
mm v pht trin thnh nhng khun ti hoc nhng th sinh sn mi.
Nm mc c th sinh sn theo nhiu hnh thc khc nhau:
Pht trin bng khun ti: mt on khun ti ring r pht trin thnh
mt khun ti th khun ti v t bi theo gi bay khp ni gp iu kin thun
li pht trin thnh c th mi.
Mt s loi nm mc c kh nng hnh thnh nhng bo t mng dy gi
l bo t o.
Mt s c th pht trin bng cc hch nm.
Nhng cch trn c gi l sinh sn sinh dng.
V sinh sn v tnh ca nm mc, ngi ta thy c 2 hnh thc:
Bo t kn c sinh ra trong cc nang nang v cc bo t kn
c phng thch ra ngoi, mi bo t li pht trin thnh mt khun ti th mi.
Sinh sn v tnh bng cc bo t trn, cc bo t trn do cc t bo sinh
bo t l cc th bnh to thnh.
Ngoi ra, nhiu loi nm mc cn c th sinh sn theo hnh thc hu tnh. Cng
ging nh cc sinh vt bc cao, sinh sn hu tnh nm cng xy ra qu trnh cht
giao, nhn giao v qu trnh phn bo gim nhim.
 43

2.10. ASPERGILLUS NIGER [33], [34], [42]

V tr phn loi ca Aspergillus niger c xp nh sau:
Lp: Deuteromyces
B: Moniliales
H: Moniliaceace
Ging: Aspergillus
Nhm: Aspergillus niger

Hnh 2.12: Aspergillus niger di knh hin vi ( phng i x400)

Khun lc Aspergillus niger trn mi trng cao nm men Czapek (CYA) c
dng lng nhung, khi cn non c mu trng, khi gi c mu nu en, mt tri ca petri
thng c mu vng nht n vng sng.
Cung sinh bo t c sinh ra t khun ti kh sinh, c chiu di 1 3 mm, c
vch t bo trn bng v trong sut. nh cung phnh ra c dng hnh cu c gi l
bng, c ng knh t 50 75 m, sinh ra 2 lp th bnh: lp th nht hnh tam gic
cn ngc; lp th hai hnh chai, sinh ra bo t nh hnh cu, bo t nh c ng
knh 4 5 m, c mu nu en, vch bo t nh c dng x x.
c tnh sinh l:
Aspergillus niger sinh trng nhit ti thiu 6 8oC, ti a 45 47oC, ti
u 35 37oC (Panasenko, 1967).
c t:
Aspergillus niger c xem l nm si khng sinh c t v c s dng rng
ri trong ch bin thc phm. Cc enzyme thy phn thu nhn c t chng ny
khng gy t bin vi khun v cc m ca chut.
 44

Sinh thi:
Aspergillus niger chim u th vng kh hu nhit i. Bo t nh mu en
nn khng b nh hng bi tia mt tri v tia cc tm. Aspergillus niger thng c
phn lp t cc sn phm phi nng: c, cy gia v,
Aspergillus niger lm thi ra tri cy (to, cam, qut du,) v hoa mu
(hnh, ti, c chua,)
2.11. TRICHODERMA REESEI [4], [26], [33], [41]
Trichoderma reesei c phn loi nh sau:
Lp: Deuteromycetes
B: Moniliales
H: Moniliaceae
Ging: Trichoderma
Nhm: Trichoderma reesei

Hnh 2.13: Trichoderma reesei di knh hin vi ( phng i x400)

Khun lc ban u trong sut, sau c mu xanh lc do hnh thnh bo t.
Cung sinh bo t c dng chm, phn nhnh lin tc, sinh ra cc th bnh c
dng hnh tam gic, mi nhnh kt thc bi mt th bnh. nh bo t hnh cu, ng
knh 3,6 4,5 m, c mu xanh lc, c vch x x.
c tnh sinh l:
Sinh trng nhit ti thiu 0oC, ti a 30 37oC, ti u 20 28oC.
c t:
Trichoderma reesei khng sinh ra cc cht khng sinh v khng gy bnh cho
ng thc vt, ch phm enzyme th t Trichoderma reesei khng gy t bin vi
khun v cc m, cng nh khng gy qui thai cho chut.
 45

Sinh thi:
Trichoderma reesei phn b vng cc Bc, vng ni, vng nhit i.
Trichoderma reesei l nm si sinh sn v tnh, phn lp c trn cc o
Solomon trong chin tranh Th gii th II t cc mnh vi cotton. Chng ny c kh
nng sinh ra mt lng ln cellulase v l t tin ca nhiu bin chng c s dng
hin nay.
Trichoderma reesei c kh nng phn gii hon ton cellulose t nhin vi hiu
sut cao v h enzyme cellulase ca Trichoderma reesei y 3 thnh phn
(endocellulase, exocellulase v -glucosidase) v c tit ra mi trng nui cy vi
mt lng ln (c th t 40 g/l). V vy, y l loi nm c nhiu tim nng trong
cng nghip sn xut enzyme cellulase.
Cellulase ca Trichoderma reesei c pH ti thch l 5,0 v nhit ti thch l
50oC.
Sinh tng hp cellulase t Trichoderma reesei cng nh t cc chng vi sinh
vt khc i hi phi b sung cht cm ng vo mi trng nui cy nh cm, rm, b
ma, mn ca,
 46

PHN 3: VT LIU V PHNG PHP NGHIN CU

Hnh 3.1: C ph Bi Hnh 3.2: C ph S

mua t th trng mua t th trng

Hnh 3.3: C ph Mokka

mua t th trng
 47

3.1. THI GIAN V A IM

3.1.1. Thi gian
T 1 3 2005 n 1 8 2005.
3.1.2. a im
o Phng th nghim B mn Cng ngh Sinh hc trng i hc Bch Khoa
Thnh ph H Ch Minh.
a ch: 268 L Thng Kit Qun 10 Thnh ph H Ch Minh.
3.2. VT LIU
3.2.1. C ph nhn
3.2.1.1. Ging
Chng ti tin hnh th nghim trn 3 loi c ph vi tn gi trn th trng l:
o C ph Bi
o C ph S
o C ph Mokka
3.2.1.2. Ngun gc
Ngun mua t ca hng mua bn c ph ti Thun Hip
a ch: 36A Nguyn Th Nh Phng 14 Qun 5 Thnh ph H Ch
Minh.
3.2.2. Nm si
Cc chng c s dng l Aspergillus niger v Trichoderma reesei nhn t B
mn Cng ngh sinh hc thuc khoa Cng ngh Ha hc v Du kh trng i
hc Bch Khoa Thnh ph H Ch Minh.
3.2.3. C cht
o Malt
o Cm go
o Bt c rt
o Tru
o Khoai ty
o Bt m
o Bt sn
o CMC (Sodium carboxymethyl cellulose)
 48

3.2.4. Mi trng s dng

3.2.4.1. Mi trng nui cy nm mc
S dng mi trng thch malt.
3.2.4.2. Mi trng nhn ging
S dng mi trng cm go.
3.2.4.3. Mi trng gi ging
S dng mi trng PGA.
3.2.5. Ha cht
3.2.5.1. Ha cht s dng cho mi trng
o (NH4)2SO4
o Glucose
o Agar
o Nc ct
3.2.5.2. Ha cht s dng cho phng php xc nh hot tnh
o Thuc th DNS (3,5-dinitrosalicylic acid) C7H4N2O7
DNS
NaOH
Mui potassium sodium tartrate tetrahydrate C4H4O6KNa
o Thuc th antron 0.2% (gm antron v H2SO4 m c)
o Dung dch CMC 1%
CMC
Dung dch CH3COOH 1M
Dung dch NaOH 1N
o Dung dch pectin 1%
o Dung dch NH3 10%
o Dung dch ZnSO4 15%
o Glucose, lactose
o Nc ct
3.2.5.3. Ha cht s dng cho cc phng php xc nh thnh phn
o Dung dch HNO3 m c
o Dung dch H2SO4 m c
o Dung dch CH3COOH m c
 49

o Dung dch H2O2 30%

o Dung dch H2SO4 0,1N
o Dung dch NaOH 0,1N
o Dung dch NaOH 5%
o Dung dch CH3COOH 1N
o Dung dch HCl 5%
o Dung dch ch acetate 10%
o Dung dch Na2HPO4 bo ha
o Dung dch CaCl2 2N
o Dung dch AgNO3 1%
o Acid perchloric (HClO4)
o Cn 90o, cn 80o
o Ru etylic 96%
o Ete etylic
o Thuc th DNS, antron
o Phenolphtalein
o Glucose
o Nc ct
3.2.6. Thit b - Dng c
Thit b
o My o quang ph SPECTRO 2000RS (LaboMed,inc.)
o My o pH
o My ly tm
o My xay
o My ct m bn t ng GERHARDT (c)
o Dng c o Brix Hand-held Refractometer (ATAGO)
o Ni hp, t sy, t m, t lnh
o T ht
o ng lm lnh hi lu
o Cn in t, cn phn tch
o Nhit k
 50

Dng c
o a petri
o ng nghim
o Erlen 250ml, 500ml
o Becher 100ml, 500ml, 1000ml
o ng ong cc loi
o Bnh nh mc cc loi
o Pipette cc loi
o n cn, que cy, a thy tinh
o Phu lc, giy lc
o Bp in, cho rang
o R nha, vi mng
3.3. PHNG PHP NGHIN CU
3.3.1. Phng php vi sinh vt [22]
3.3.1.1. Phng php cy chuyn v gi ging (trong iu kin v trng)
a. Phng php cy chuyn
Pha ch mi trng nui cy (mi trng thch malt)

Cho vo 1/3 ng nghim

Thanh trng 121oC trong 15 pht

nghing 45o n khi thch ng cng li ng ging

Dng que cy vng thanh trng cy ria theo ng zic zc

Cho vo t m
b. Phng php gi ging
Cy chuyn gi ging nm si t mi trng thch malt sang mi trng gi ging
(mi trng PGA).
 51

3.3.1.2. Phng php nhn ging (trong iu kin v trng)

ng nghim ging

Cho vo 10 ml nc ct hp v trng

Khuy

Cho vo mi trng cm go

o u

Cho vo t m
3.3.1.3. Phng php cy ging t mi trng nhn ging sang mi trng
ch phm
Dng a thy tinh v trng trn u mi trng nhn ging

Cho tt c vo mi trng ch phm

Trn u

Cho vo t m
3.3.1.4. Phng php ln men
C ph sau khi ngm nc t m cn thit, trn u vi ch phm, cho
vo r nha, ph vi mng ln v (t l ch phm, thi gian ty ch tiu kho st).
3.3.2. Phng php ha l
3.3.2.1. Phng php xc nh m
Nguyn tc
Da trn nguyn l lm kh mu nhit 100 105oC n trng lng
khng i. Lng mu mt khi sy mu n trng lng khng i l lng nc c
trong mu. Phng php ny loi tr c cc sai s do cc phn ng xy ra khi sy
mu nhit cao.
 52

Cch thc hin

Mu cn xc nh m

Nghin nh

Cn chnh xc mt lng nht nh (5 10 g)

Cho vo chn ng mu (c sy kh n trng lng khng i)

Cho vo t sy (100 105oC trong 6 8 gi n khi trng lng khng i)

Lm ngui trong bnh ht m (20 30 pht)

em cn

Tnh kt qu

m1 m2
W(%) 100
m
Trong :
W: m ca mu (%)
m1: Trng lng chn v trng lng mu trc khi sy (g)
m2: Trng lng chn v trng lng mu sau khi sy (g)
m: Trng lng mu phn tch (g)
 53

3.3.2.2. Phng php trch cht ha tan

Phng php trch cht ha tan trong c ph sau khi ln men:

Cn 10 g c ph bt

B sung 30 ml nc si

Thu ly dch lc

Thm 20 ml nc si

Thu ly dch lc

Cho vo becher xc nh trng lng

un trn bp
Sy kh

Thu cht kh l cht ha tan

cho vo t sy

em sy nhit 100 105oC

Thu cht kh

Cn trng lng
o pH
o ha tan

3.3.2.3. Phng php o kch thc

Dng thc o k thut o chiu ngang, chiu di v b dy ca ht c ph.
 54

3.3.2.4. Phng php nghin

Dng my xay nghin c ph (c ph ti v c ph ln men) khi cn lng
c ph bt nghin cu.
3.3.2.5. Phng php ray
Dng li ray ray nhng nguyn vt liu cn tp cht hoc khi cn thu lng
bt mn.
3.3.2.6. Phng php m
Dng mt thng m s ht c ph, thu nht nhng ht h, hng v tp
cht.
3.3.2.7. Phng php ngm
Dng nc ngm c ph, mc ch l ht c ph t c m cn thit.
3.3.3. Phng php ha sinh
3.3.3.1. Cc phng php xc nh hot tnh enzyme
a. Xc nh hot tnh cellulase [14], [25]
Nguyn tc
Phng php ny da trn s thy phn CMC (Sodium carboxymethyl
cellulose) bng enzyme nhit 40oC v pH 5,0.
Sau phn ng thy phn s to ra mt lng ng kh, lng ng kh ny
s phn ng vi thuc th DNS (3,5-dinitrosalicylic acid).
Sau tin hnh o hp thu OD ca dung dch ng bng my o mt
quang bc sng 540 nm.
 55

Cch thc hin

Phn ng enzyme Phn ng th khng

1 ml dung dch mu n nhit 1 ml dung dch mu n nhit

40 0.1oC trong 5 pht 40 0.1oC trong 5 pht
C cht CMC n nhit C cht CMC n nhit
tng t tng t

Thm 1 ml dung dch CMC vo dung Thm 4 ml dung dch lactose DNS
dch mu vo dung dch mu

Lc u Lc u

n nhit 40 0.1oC trong 10 pht

Thm 4 ml dung dch lactose DNS Thm 1 ml dung dch CMC

Lc tht k Lc u

Dng nylon bt kn ming ng nghim Dng nylon bt kn ming ng nghim

t trong nc si 15 pht t trong nc si 15 pht

Lm ngui bng nc lnh Lm ngui bng nc lnh

Tin hnh o OD ( = 540 nm) Tin hnh o OD ( = 540 nm)

(dng nc ct lm i chng) (dng nc ct lm i chng)

AT AB
 56

Dng ng chun glucose

Pha dung dch glucose 1 mg/ml bng cch ha tan 100 mg glucose
monohydrate vi 80 ml nc ct v chuyn vo bnh nh mc 100 ml, thm nc ct
n vch v lc u. Thc hin mt lot 7 ng nghim theo bng di y:
Bng 3.1: Dng ng chun glucose
ng s 0 1 2 3 4 5 6
Nng dung dch glucose
0 0,1 0,2 0,3 0,4 0,5 0,6
chun (mg/ml)
- ml dung dch glucose 1 mg/ml 0 0,1 0,2 0,3 0,4 0,5 0,6

- ml dung dch CMC 1% 1 1 1 1 1 1 1

- ml dung dch lactose-DNS 4 4 4 4 4 4 4

- ml nc ct 1 0,9 0,8 0,7 0,6 0,5 0,4

Lc u cc ng nghim ny

Dn nylon ln ming ng nghim

un si cch thy trong 15 pht

Lm lnh v nhit phng

(bng mt chu nc lnh)

o hp th OD bc sng 540 nm AG
ng s 0 l ng i chng AW
Tnh kt qu
nh ngha n v hot tnh
Mt n v hot tnh CMCase l lng enzyme cn thit gii phng ra
ng kh (nh glucose) khi thy phn CMC vi vn tc 1 mol/pht di cc iu
kin phn ng.
 57

Tnh ton
Tnh h s glucose
o Hiu chun hp thu OD ca dung dch glucose chun bng hiu s
AG - AW
o Dng th ng chun glucose vi trc tung l mt quang (OD), trc
honh l nng d glucose (mg/ml).
H s glucose c tnh bng cng thc:
CG
F
AG AW

Trong :
F: H s glucose
CG: Nng dung dch glucose chun (mg/ml)
AG: hp thu OD ca dung dch glucose chun
AW: hp thu OD ca phn ng vi nc ct
o Chia nng (mg/ml) cho hp thu OD ca mi dung dch glucose chun
thu c gi tr h s glucose, sau tnh gi tr h s trung bnh.
Xc nh hot tnh enzyme
Hot tnh CMCase c tnh theo cng thc sau:

1000 1 1 1
CMCase (UI/g) (AT AB ) F
180 10 phut 1 ml C

Trong :
AT: hp thu OD ca dung dch phn ng enzyme
AB: hp thu OD ca phn ng th khng
F: H s glucose (mg/ml)
1000: Chuyn t mg sang g
180: Trng lng phn t ca glucose, i t g sang mol
10 pht: Thi gian phn ng
1 ml: Th tch dung dch enzyme
C: Nng dung dch mu (g/ml)
 58

b. Xc nh hot tnh pectinase [14]

S dng phng php so mu, l phng php phn tch da trn vic so snh
cng mu ca dung dch tiu chun c nng xc nh.
Phng php so mu cho php sai s 5%.
Nguyn tc
nh lng acid galacturonic, l sn phm ca qu trnh thy phn pectin di
tc dng ca pectinase khng kt ta bi ZnSO4.
Cch thc hin
20 ml dung dch pectin

Cho vo becher 100 ml

Thm 10 ml dung dch enzyme 1%

Lc u

nhit 30oC (nhit phng) trong 60 pht

Thm 2 ml dung dch ZnSO4 15%

Lc qua giy lc

Thu dch lc

Pha long 5 ln

Ta c dung dch pha long gi l dung dch (*)

 59

5 ml dung dch antron cho vo ng nghim

Thm 2,5 ml dung dch (*)

Lc mnh 10 pht

un cch thy 70oC trong 12 pht

Lm ngui

Tin hnh o OD bc sng 584 nm

Tnh kt qu
Mt n v hot tnh pectinase l lng enzyme trong iu kin tiu chun c
kh nng xc tc 1 g pectin thnh acid galacturonic trong 60 pht.
iu kin tiu chun:
Nhit : 30oC, thi gian: 60 pht
pH 3,9 4,1
Nng pectin: 0,66%
Mc thy phn pectin: 30%

0,34 OD 0,0104
Hot tnh pectinase (UI)
V
Trong :
OD: Mt quang ca dung dch sau khi phn ng vi antron
V: Lng enzyme ly tin hnh phn ng (g hoc ml)
3.3.3.2. Cc phng php xc nh thnh phn ch yu trong c ph
a. Xc nh hm lng cellulose [16]
Nguyn tc
Phng php nh lng cellulose da vo tnh cht bn vng ca cellulose
trc tc dng ca acid mnh, kim mnh, khng b thy phn di tc dng ca acid
yu. Trong khi , cc cht khc thng i km theo vi cellulose nh hemicellulose,
lignin, t bn hn i vi tc dng ca acid v kim, nn b oxi ha, phn gii v tan
 60

vo dung dch khi x l nguyn liu bng dung dch kim hoc bng hn hp acid
nitric vi acid acetic.
Cch thc hin
Cn 2 g mu c ph nghin nh

Sy kh n trng lng khng i (100 105oC)

Cho vo bnh tam gic 250 ml

Thm vo bnh 16,5 ml hn hp 1,5 ml HNO3 m c

15 ml CH3COOH m c

un si 30 pht (c lm lnh hi lu) Giy lc

ngui, pha long bng nc nng Sy kh n trng lng khng i

Lc

Ra kt ta cellulose 3 ln (10 15 ml nc ct nng/ln)

Ra bng ru etylic 96% 2 ln (10 15 ml/ln)

Ra bng ete etylic 1 ln (15 20 ml)

Sy giy lc c cha cellulose n trng lng khng i

(100 105oC trong 2 3 gi)
 61

a 100
Tnh kt qu X
w
Trong :
X: Hm lng cellulose tnh bng %
a: Trng lng cellulose (g)
w: Trng lng mu th nghim (g)
100: H s chuyn i thnh %
b. Xc nh hm lng pectin (S dng phng php Canxi pectat) [9], [16]
Nguyn tc
Phng php da trn c s thu nhn mui canxi pectat dng kt ta.
Cch thc hin
Cn 0,15 g mu c ph

Thm vo 100 ml dung dch NaOH 0,1N

hn hp qua m x phng ha hon ton pectin thnh acid pectic

Thm 50 ml dung dch CH3COOH 1N

5 pht
Thm vo 50 ml dung dch CaCl2 2N Giy lc khng tro
1 gi
un si 5 pht Sy kh n trng lng khng i

Lc

Ra kt ta canxi pectate bng nc ct nng n khi khng cn ion Clo na

(th nc ra khng c kt ta trng bng dung dch AgNO3 1%)

Sy giy lc n trng lng khng i (100 105oC)

Tnh kt qu
Hm lng canxi pectate c tnh bng hiu s ca trng lng giy lc c kt
ta v giy lc khng.
 62

Hm lng pectin c tnh theo cng thc sau:

w 0,92 100
P
B
Trong :
P: Hm lng pectin (%)
W: Trng lng kt ta canxi pectat (g)
B: Lng pectin ly em x phng ha (g)
(khong 0,15 g)
0,92: H s chuyn tr hm lng canxi trong kt ta
(pectin chim 92% trng lng canxi pectat)
100: H s chuyn i thnh %
c. Xc nh hm lng ng tng s ha tan [1], [9]
Nguyn tc
S nh phn ny cn bn da trn s phn ng mu c trng cho bi ng
v nhiu cht hu c vi s hin din ca acid sulfuric (H2SO4).
S chnh xc ca kt qu ph thuc vo:
o sch cc dng c
o tinh khit ca thuc th, nht l H2SO4
o Nhit phi c nh trong sut thi gian un
 63

Cch thc hin

Trch ng
Cn 1 g mu c ph nghin nh

Cho vo becher 100 ml v thm vo 10 ml cn 90o

un cch thy cho si 3 ln

Khuy u, ngui

Lc qua giy lc khng tro (gi cn, khng cn ln giy lc)

Thm 10 ml cn 80o vo cc cha cn, khuy u

un cch thy cho si 2 ln

Khuy, ngui, lc

Lp li 2 ln

a cn ln giy lc v ra sch 2 3 ln bng cn 80o nng

un nh dch lc trn ni cch thy cn bay hi ht

Pha long cn thu c vi nc ct, nh mc 50 ml

lng
 64

Thc hin phn ng mu

Ta c th to phn ng mu ca dung dch ng vi mt trong nhng thuc
th sau y: phenol, orcinol hay antron. Trong ti ny, chng ti chn thuc th l
antron.
Thc hin theo bng di y:
Bng 3.2: Bng thc hin phn ng mu vi antron

ng s 1 2 3 4 5 6 7 8 9

Glucose mu 0,01% (ml) 0 0 1 2 3 4 5

Nc ct (ml) 5 5 4 3 2 1 0

Dung dch ng nghin cu (ml) 5 5

Nng ng mi ng (mg/ml) 0 0 0,1 0,2 0,3 0,4 0,5 x x

tt c cc ng vo mt ni nc

Thm tht chm 10 ml thuc th antronvo mi ng (chy theo thnh ng nghim)

Dng a thy tinh khuy chm

un si cch thy ng 7,5 pht

Lm lnh trong nc

Khi ngui, o OD bc sng 630 nm

Tnh kt qu
Tr s mt quang ca nhng ng mu sau khi tr i tr s ca ng th
khng s xc nh c ng chun.
Vi dung dch ng cn nh nng , ta cng ly tr s mt quang tr i
tr s mt quang ca ng th khng ri chiu vo ng chun suy ra nng x,
t tnh ra % lng ng trong mu.
 65

d. Xc nh hm lng ng kh [16]
S dng phng php Acid dinitrosalicylic (DNS).
Nguyn tc
Phng php da trn c s phn ng to mu gia ng kh vi thuc th
DNS. Cng mu ca hn hp phn ng t l thun vi nng ng kh trong
mt phm vi nht nh.
Da theo th ng chun ca glucose tinh khit vi thuc th DNS s tnh
c hm lng ng kh ca mu nghin cu.
Cch thc hin
Dng ng chun glucose
Pha dy glucose chun t 0,1 0,8 mg/ml

Cho 1 ml dung dch glucose chun vo 8 ng nghim

Thm 4 ml thuc th DNS

un si 15 pht

Lm lnh v nhit phng

o mt quang bc sng 540 nm (i chng l nc ct)

Dng ng chun glucose

(vi trc tung l mt quang, trc honh l nng glucose)

Xc nh lng ng kh
Dung dch ng cn phn tch cng c chun b nh phn trch ng ca
phng php xc nh ng tng s.
Hn hp phn ng gia glucose vi thuc th DNS c tin hnh nh phn
dng ng chun glucose (thay 1 ml dung dch glucose chun bng 1 ml dung dch
ng cn phn tch).
 66

Sau khi o c mt quang (OD) bc sng 540 nm, da vo ng

chun glucose suy ra lng ng kh trong mu.
e. Xc nh hm lng tinh bt [16]
(S dng phng php thy phn bng acid)
Nguyn tc
Di tc dng ca acid, tinh bt b thy phn to thnh glucose. Xc nh lng
glucose to thnh ri nhn vi h s 0,9 ta c hm lng tinh bt.
(C6H10O5)n + nH2O nC6H12O6
162,1 18,02 180,12
162,1
H s F 0,9
180,12

Cch tin hnh

Cn 200 250 mg mu ( nghin nh v bit r m) cho vo bnh cu 100 ml

Thm 50 ml nc ct, lc u, gi yn 30 45 pht

Lc b nc lc loi b ng tan

Ra tinh bt bng nc ct 2 3 ln, chc thng giy lc v dng 25 ml

dung dch HCl 5% chuyn tinh bt vo bnh cu

Lp ng lm lnh hi lu, un cch thy hn hp 3 5 gi

Kim tra s thy phn hon ton
ca tinh bt bng dung dch Iod
Lm ngui, trung ha hn hp bng dung dch NaOH 5% n pH = 5,6 6,0

Chuyn hn hp vo bnh nh mc 100 ml, kt ta protein bng dung dch

ch acetate 10%, dng Na2HPO4 bo ha loi b ch acetate

Thm nc ct ti vch mc, lc u v lc

nh lng ng glucose theo phng php DNS

 67

f. nh lng Nit tng v protein th [24]

(S dng phng php Kjeldahl)
Nguyn tc
Di tc dng ca H2SO4 m c nhit cao, cc hp cht c cha nit b
phn hy v b oxi ha n CO2 v H2O, cn nit chuyn thnh NH3 v tip tc kt
hp vi H2SO4 to thnh mui (NH4)2SO4 tan trong dung dch. ui NH3 ra khi dung
dch bng NaOH, ng thi ct v thu NH3 bng mt lng d H2SO4 0,1N. nh
phn lng H2SO4 cn li bng dung dch NaOH 0,1N chun, sau nh lng nit
c trong mu th nghim.
Cch tin hnh
Bc 1: V c ha mu (tin hnh trong t ht trnh kh c SO2, CO2)
Ly 0,5 g mu xay nh

Thm cht xc tc acid perchloric HClO4

un nh hn hp trnh si tro
(ch un mnh khi hn hp chuyn hon ton sang lng)
Thnh thong lc nh v trng kho
khng cn mu st li trn thnh bnh
un n khi dung dch trong bnh hon ton trng

Bc 2: Ct m (tin hnh trong my ct m bn t ng GERHARDT)

Chuyn ton b dung dch sau khi v c ha xong bnh Kjeldahl vo bnh
nh mc 100 ml

Thm nc ct n vch (ch phi lm lnh v lc tht u)

Cho vo erlen 10 ml dung dch H2SO4 0,1N v lp vo my

Ly 10 ml dung dch th nghim t bnh nh mc

cho vo ng phn ng v lp vo h thng
 68

Bc 3: nh phn
Ly erlen ra khi my sau khi trng nc ct ly ht mu bm trn ng.
Cho 10 git phenolphtalein vo bnh v nh phn bng NaOH 0,1N.
Bc 4: nh h s iu chnh nng NaOH 0,1N
Ly 10 ml H2SO4 0,1N vo erlen, nh phn bng NaOH 0,1N.
Tnh nng thc t v nng tnh ton ca NaOH.
Tnh kt qu
Lng m c trong mu c ph c tnh theo cng thc:
(a bT) 0,0014 100 100
x
10 0,5
Trong :
x: Phn trm nit c trong mu nghin cu
a: S ml H2SO4 0,1N em hp th NH3
b: S ml NaOH 0,1N tiu tn cho chun
0,5: Lng mu c ph em th nghim
0,0014: Lng nit ng vi 1 ml H2SO4 0,1N
T: H s iu chnh nng NaOH 0,1N
Tnh hm lng protein th
Nit trong vt liu ch yu l nit protein, ngoi ra cn c mt lng nh nit
trong cc thnh phn khc gi l nit phi protein.
Nit tng = nit protein + nit phi protein
Hm lng nit trong cc vt liu sinh hc c tnh bng cch nhn hm
lng nit protein vi mt h s chuyn i xc nh (do hm lng nit c t l n
nh t 15 18% protein), i a s l 16%.
Trong cc nguyn liu sinh hc, do hm lng nit phi protein nh v vic tch
ring rt phc tp nn theo qui c, ngi ta tnh hm lng protein theo nit tng v
gi l protein tng (hay protein th).
Cng thc tnh hm lng phn trm protein th c trong i a s cc nguyn
liu l:

Protein (%) = Nit (%) x 6,25

Ring ng cc v u c h s chuyn i l 5,7.
Sa c h s chuyn i l 6,38.
 69

g. Xc nh hm lng tro v tng hu c [1], [9]

Tro ha v xc nh tng lng tro
Cn chnh xc 4 g mu c ph

Cho vo chn s bit trng lng kh tuyt i

Thm vo 5 git HNO3 m c v vi git H2O2 30%

Cho vo l nung 550oC n khi nguyn liu bin thnh tro trng nh tn thuc l

Cho ngay chn s vo bnh ht m, ngui v cn chnh xc

Lp li n khi trng lng khng i

Tng lng tro c tnh nh sau:

a 100
Tng lng tro = (%)
b
Trong :
a: Trng lng tro (g)
b: Trng lng mu kh (g)
100: H s chuyn i thnh %
Xc nh tng hu c
Cu trc hu c ca sinh vt d b ph hy bi cc tc nhn kim hay acid
mnh c tnh oxi ha v nhit cao.
Cc cht hu c b oxi ha hon ton chuyn thnh CO2 v H2O bay ra khi
chn s, ch cn li mt t khong cht trong chn s.
V vy, cch tnh tng hu c nh sau:

Tng hu c (%) = 100% tng lng tro (%)

 70

3.3.4. Phng php ly trch enzyme

Cn 1 g ch phm

Nghin nt vi thy tinh v

Ly trch bng 6 ml dung dch NaCl 0,3%

Ly tm (5000 6000 vng trong 10 pht)

Thu c dch trch enzyme th

Pha long vi dung dch NaCl 0,3%

nh mc 100 ml

Thu dch enzyme 1%

3.3.5. Phng php thit k th nghim

3.3.5.1. Kho st nh hng ca t l c cht (bt m v bt sn) n qu
trnh sinh tng hp hai loi enzyme pectinase v cellulase
C nh thi gian nui cy l 48 gi.
Sau khi chuyn mi trng cm go vo mi trng ch phm, nui cy trong
48 gi, sau xc nh hot tnh hai enzyme pectinase v cellulase chn ra t l ti
u nht.
Chng ti kho st vi 7 t l (bt m : bt sn).
 71

Bng 3.3: Nghim thc t l c cht trong ch phm

T l bt m : bt sn %bt m : %bt sn
Mu 1 0:1 0 : 100
Mu 2 1:3 25 : 75
Mu 3 1:2 33 : 67 (xp x)
Mu 4 1:1 50 : 50
Mu 5 2:1 67 : 33 (xp x)
Mu 6 3:1 75 : 25
Mu 7 1:0 100 : 0

a. Kho st nh hng ca t l trn n sinh tng hp pectinase

Ly trch enzyme t 7 mu ch phm, sau tin hnh xc nh hot tnh
pectinase, chn mu ti u.
b. Kho st nh hng ca t l trn n sinh tng hp cellulase
Ly trch enzyme t 7 mu ch phm, sau tin hnh xc nh hot tnh
cellulase, chn mu ti u.
3.3.5.2. Kho st nh hng ca thi gian nui nm si trong mi trng ch
phm n qu trnh sinh tng hp hai loi enzyme pectinase v cellulase
Sau khi chn c t l c cht bt m : bt sn (cho mi trng ch phm) ti
u cho sinh tng hp 2 loi enzyme pectinase v cellulase, chng ti c nh t l ny.
Bc tip theo l kho st thi gian nui nm si thch hp nht thu c pectinase
v cellulase c hot tnh cao nht.
Chng ti kho st 11 mu vi 11 khong thi gian nui khc nhau (cch u
nhau 6 gi).
 72

Bng 3.4: Nghim thc thi gian nui nm mc to ch phm

Thi gian nui cy
Mu 1 12 gi
Mu 2 18 gi
Mu 3 24 gi
Mu 4 30 gi
Mu 5 36 gi
Mu 6 42 gi
Mu 7 48 gi
Mu 8 54 gi
Mu 9 60 gi
Mu 10 66 gi
Mu 11 72 gi

a. Kho st nh hng ca thi gian nui n sinh tng hp pectinase

Ly trch enzyme t 11 mu v xc nh hot tnh pectinase, chn mu ti u.
b. Kho st nh hng ca thi gian nui n sinh tng hp cellulase
Ly trch enzyme t 11 mu v xc nh hot tnh cellulase, chn mu ti u.
3.3.5.3. Kho st nh hng ca thi gian ln men
Sau khi chn c t l c cht (bt m : bt sn) v thi gian nui cy ti u,
chng ti to 1 mu ch phm ti u v s dng ln men c ph.
Chng ti c nh hm lng ch phm l 0,1%, tin hnh ln men c ph vi 6
mu th nghim (ln men 6 khong thi gian khc nhau, cch u 4 gi).
Bng 3.5: Nghim thc thi gian ln men c ph
Thi gian ln men
Mu 1 12 gi
Mu 2 16 gi
Mu 3 20 gi
Mu 4 24 gi
Mu 5 28 gi
Mu 6 32 gi
 73

Sau khi ngng qu trnh ln men c ph ln lt c ra sch, rang v xay

mn. Sau , tin hnh trch cht ho tan v kho st cc ch tiu.
Trong kha lun ny, khi lng cht tan c hiu l khi lng cc cht ha
tan c trong 10 g c ph bt sau khi trch ly, c cn v sy kh. ha tan c
hiu l lng cht tan trong 50 ml nc v c trch t 10 g c ph bt.
a. Kho st nh hng ca thi gian n kh nng trch ly c ph
Chn mu ln men vi thi gian ln men thch hp nht, bng cch chn mu
c kh nng trch ly cht ha tan cao nht (thng qua khi lng cht tan cao nht).
b. Kho st nh hng ca thi gian n ha tan ca c ph ln men
Chng ti kho st ha tan bng dng c o Brix.
c. Kho st nh hng ca thi gian n pH ca dung dch cht tan
trch t c ph ln men
o pH ca dung dch cht tan trch t c ph ln men (bng my o pH) trc
khi em sy kh.
3.3.5.4. Kho st nh hng ca hm lng ch phm
Sau khi chn c thi gian ln men ti u, chng ti c nh thi gian v
tin hnh kho st hm lng ch phm cho vo khi c ph ln men, vi 6 nghim
thc (cch u nhau 0,04%).
Bng 3.6: Nghim thc hm lng ch phm s dng ln men c ph
Hm lng ch phm (%)
Mu 1 0,04
Mu 2 0,08
Mu 3 0,12
Mu 4 0,16
Mu 5 0,20
Mu 6 0,24

a. Kho st nh hng ca hm lng ch phm n kh nng trch ly c

ph
Chn mu ln men vi hm lng ch phm thch hp nht, bng cch chn
mu c kh nng trch ly cht ha tan cao nht (thng qua khi lng cht tan cao
nht).
 74

b. Kho st nh hng ca hm lng ch phm n kh nng tng ha

tan cht trch c ph
o ha tan bng dng c o Brix.
c. Kho st nh hng ca hm lng ch phm n pH ca dung dch
cht tan trch t c ph ln men
o pH ca dung dch cht tan trch t c ph ln men (trc khi sy).
3.3.5.5. Kho st nh hng m ht c ph trc khi ln men ln kh nng
trch ly cht ha tan
Trc khi ln men, chng ti ngm ht c ph trong nc vi nhng khong
thi gian khc nhau (cch nhau 0,5 gi). Sau , tin hnh ln men v trch cht ha
tan.
Nhng khong thi gian ngm nc kho st:
0,5 gi 1 gi 1,5 gi 2 gi 2,5 gi 3 gi 3,5 gi 4 gi
3.3.5.6. Kho st s thay i trng lng khi c ph trc v sau khi ln men
Vi mi ch tiu kho st, chng ti u cn trng lng khi c ph trc v
sau khi ln men.
Mi m ln men, chng ti u s dng 100 g c ph ln men (trng lng
khi c ph trc khi ln men l 100 g).
3.3.5.7. Kho st nhit khi theo thi gian ln men
C nh thi gian ln men v hm lng ch phm ti u, tin hnh ln men 1
m c ph v kho st nhit khi theo thi gian (cch nhau 2 gi).
3.3.6. Phng php x l s liu
Chng ti tin hnh x l s liu theo phng php trung bnh, ngha l tin
hnh lp li th nghim nhiu ln (t nht l 3 ln lp li) n khi thu c cc kt qu
kh ging nhau.
Sau tin hnh cng tt c cc s liu ca cc ln th nghim chia cho s ln
tin hnh th nghim c c kt qu trung bnh trnh by trong kha lun.
 75

PHN 4 : KT QU V THO LUN

Hnh 4.1 v 4.2: C ph ang c ln men

Hnh 4.3: C ph ln men Hnh 4.4: Thnh phm c ph ln men

c rang c rang v xay thnh bt
 76

4.1. KT QU KHO ST MT S C IM CA BA LOI C PH

4.1.1. Hnh dng ht
Bi: ht nh nht trong 3 loi c ph, hi trn, b mt nhn, mu xanh lc.
S: ht to hn c ph Bi nhng nh hn c ph Mokka, ht di v dp hn
c ph Bi, b mt ht nhn, mu xanh lc.
Mokka: ht to v do nht trong 3 loi, mu hi en, hi sn si.
4.1.2. S ht c ph trong 100 g
Cc s liu c ly trung bnh ca 3 ln m.
Bi: 675 ht
S: 591 ht
Mokka: 610 ht
4.1.3. S ht en, hng trong 100 g
Cc s liu c ly trung bnh ca 3 ln m.
Bi: 41 ht
S: 50 ht
Mokka: 128 ht
4.1.4. Kch thc trung bnh ht
Chn ngu nhin 30 ht c kch thc tng ng v ph bin, o cc kch
thc bng thc o k thut, sau ly kt qu trung bnh.
4.1.4.1. Chiu ngang trung bnh
Bi: 6,60 mm
S: 7,31 mm
Mokka: 8,32 mm
4.1.4.2. Chiu di trung bnh
Bi: 8,45 mm
S: 9,45 mm
Mokka: 10,92 mm
4.1.4.3. B dy trung bnh
Bi: 5,34 mm
S: 4,47 mm
Mokka: 4,80 mm
 77

Hnh 4.5: o chiu di ht c ph Bi

Hnh 4.6: o chiu ngang ht c ph S

Hnh 4.7: o b dy ht c ph Mokka

 78

4.2. XC NH CC THNH PHN CH YU TRONG C PH NHN

Bng 4.1: Cc thnh phn ch yu c trong c ph nhn

T l phn trm (%) so vi khi lng
Thnh phn
C ph Bi C ph S C ph Mokka
1. ng tng s ha tan 20,95 21,10 25,65
2. Hm lng ng kh 2,95 3,00 4,17
3. Hm lng pectine 54,71 52,62 55,14
4. Hm lng cellulose 15,29 16,80 17,04
5. Hm lng tinh bt - - 11,29
6. Hm lng nit - - 1,96
7. Hm lng protein th - - 12,22
8. Hm lng tro 5,00 5,00 5,50
9. Tng hu c 95,00 95,00 94,50
10. m 7,86 7,26 10,60
(Ngun t thc nghim)
Nhn xt:
T cc s liu thu c, chng ti nhn thy cht hu c l thnh phn ch yu
trong c ph nhn. Trong hai thnh phn pectin v cellulose chim mt t l kh ln
v chng l nhng thnh phn rt cng, bn v kh phn gii. Chnh hai thnh phn
kh phn gii ny lm hn ch kh nng trch ly cc thnh phn ha tan ca c ph. V
l do ny m chng ti t vn phi tm ra gii php bng cch ng dng cng ngh
ln men v cng ngh enzyme gia tng ti a hiu sut trch ly ca c ph nhn, t
thu c cht ha tan cao nht. Gii php ny m ra mt tng lai y ha hn cho
ngnh cng nghip ch bin c ph ha tan v cc sn phm khc t c ph, gp phn
lm pht trin ngnh cng ngh sinh hc ni chung v cng ngh sinh hc trong thc
phm ni ring ca Vit Nam chng ta.
 79

4.3. XC NH MI TRNG CH PHM TI U CHO SINH TNG HP

PECTINASE V CELLULASE
4.3.1. Xc nh nh hng ca t l c cht (bt m v bt sn) n qu trnh
sinh tng hp hai loi enzyme pectinase v cellulase
C nh thi gian nui cy l 48 gi, kho st 7 mu vi 7 t l khc nhau.
4.3.1.1. Xc nh nh hng ca t l trn n sinh tng hp pectinase
Bng 4.2: Mi tng quan gia t l bt m : bt sn v hot tnh pectinase
Mu s 1 2 3 4 5 6 7
T l
0:1 1:3 1:2 1:1 2:1 3:1 1:0
m:sn
OD 0,925 0,932 0,950 1,060 1,110 1,035 0,915
Hot tnh
304,10 306,48 312,60 350,00 367,00 341,50 300,70
(UI/g CT)
Trong , mt quang OD l s liu c ly trung bnh.

390

370 367.00
Hot tnh enzyme pectinase

350.00
350
(UI/g CT)

341.50

330

312.60

310 306.48
304.10 300.70

290
1 2 3 4 5 6 7
Mu (t l bt m : bt sn)

th 4.1: Hot tnh pectinase theo t l bt m : bt sn

 80

Nhn xt:
Kt qu t th 4.1 cho thy, hot tnh pectinase tng dn cng vi s gia tng
ca t l bt m trong hn hp, nhng hot tnh pectinase ch tng dn cc mu t
0% n 67% bt m, sau th gim dn. Hot tnh pectinase t cao nht mu ch
phm s 5, vi t l bt m : bt sn l 2 : 1 (tng ng vi khong 67% bt m v
33% bt sn).
Tuy nhin, khi mi trng ch phm qu nhiu bt m hoc ch ton bt m th
s khng c thong kh lm cho nm mc khng pht trin tt c. Nguyn nhn
l trong bt m c gluten, lm cho bt m vn li khi b sung nc vo, t mi
trng s c m qu cao, khng c thong kh v cc loi nm men s c iu
kin pht trin, lm nh hng n hot tnh enzyme. Trong khi , bt sn vi tnh
cht kh xp s gip cho mi trng ch phm c thong kh. Do vy, mi trng
ch phm c s pha trn 67% bt m v 33% bt sn l thch hp cho nm mc sinh
tng hp nhiu pectinase nht (hot tnh pectinase l 367,00 UI/g CT). y l mi
trng va c m thch hp va c thong kh thun li cho qu trnh sinh tng
hp pectinase ca nm mc.
4.3.1.2. Xc nh nh hng ca t l trn n sinh tng hp cellulase
Bng 4.3: Mi tng quan gia t l bt m : bt sn v hot tnh cellulase
Mu s 1 2 3 4 5 6 7
T l m:sn 0:1 1:3 1:2 1:1 2:1 3:1 1:0
OD 0,028 0,061 0,069 0,070 0,075 0,072 0,071
Hot tnh cellulase
0,924 2,013 2,277 2,310 2,475 2,376 2,343
(UI/g CT)

Trong , OD = AT AB v l s liu c ly trung bnh.

 81

2.475
2.5 2.376 2.343
Hot tnh enzyme cellulase 2.277 2.310

2.013
2.0
(UI/g CT)

1.5

1.0
0.924

0.5
1 2 3 4 5 6 7
Mu (t l bt m : bt sn)

th 4.2: Hot tnh cellulase theo t l bt m : bt sn

Nhn xt:
Kt qu th 4.2 cho thy, hot tnh cellulase cng tng dn cc mu t
mu 1 n mu 5, v gim dn t mu 5 n mu 7. Nh vy tng t vi mi tng
quan gia t l bt m : bt sn v pectinase, hot tnh enzyme cellulase cng tng dn
cng vi s gia tng ca t l bt m. Hot tnh cellulase t cao nht mu ch phm
s 5, vi t l bt m : bt sn l 2 : 1 (tng ng vi khong 67% bt m v 33%
bt sn).
Ngoi ra, t mu 3 n mu 7 (tng ng vi t l bt m t 33% n 100%),
hot tnh cellulase thay i khng nhiu.
Vy, mi trng ch phm vi t l tinh bt l 2 phn bt m v 1 phn bt sn
(tng ng vi 67% bt m v 33% bt sn) l mi trng thch hp nht cho qu
trnh sinh tng hp hai loi enzyme pectinase v cellulase ca nm mc.
T l ny c s dng cho nhng nghin cu tip theo.
4.3.2. Xc nh nh hng ca thi gian nui nm si trong mi trng ch
phm n qu trnh sinh tng hp hai loi enzyme pectinase v cellulase
Sau khi xc nh c t l ti u ca c cht tinh bt trong ch phm, chng
ti c nh t l ny (2 phn bt m : 1 phn bt sn), sau kho st 11 mu vi 11
khong thi gian nui khc nhau.
 82

4.3.2.1. Xc nh nh hng ca thi gian nui n sinh tng hp pectinase

Bng 4.4: Mi tng quan gia thi gian v hot tnh pectinase
Thi gian (gi) OD Hot tnh pectinase (UI/g CT)
12 0,863 283,02
18 0,882 289,48
24 0,917 301,38
30 1,141 377,54
36 0,950 312,60
42 0,905 297,30
48 0,902 296,28
54 0,880 288,80
60 0,851 278,94
66 0,806 263,64
72 0,785 256,50

Trong , mt quang OD l s liu c ly trung bnh.

390
377.54

370

350
Hot tnh enzyme pectinase

330
(UI/g CT)

312.60
310
301.38
297.30 296.28
289.48 288.80
290
278.94
283.02

270 263.64
256.50

250
12 18 24 30 36 42 48 54 60 66 72

Thi gian (gi)

th 4.3: Hot tnh pectinase theo thi gian nui cy

 83

Nhn xt:
Kt qu t th 4.3 cho thy, trong giai on u nm mc cha thch ng vi
mi trng ch phm nn hot tnh pectinase thp (283,02 n 301,38 UI/g CT). Sau
, hot tnh pectinase tng v t cao nht mu ch phm 30 gi (377,54 UI/g CT).
Trong nhng thi gian tip theo, hot tnh pectinase gim dn t mu 36 gi n mu
ch phm 72 gi (gim t 312,60 n 256,50 UI/g CT) .
4.3.2.2. Xc nh nh hng ca thi gian nui n sinh tng hp cellulase
Bng 4.5: Mi tng quan gia thi gian v hot tnh cellulase
Thi gian (gi) OD Hot tnh cellulase (UI/g CT)
12 0,042 1,386
18 0,047 1,551
24 0,059 1,947
30 0,092 3,036
36 0,070 2,310
42 0,065 2,145
48 0,060 1,980
54 0,057 1,881
60 0,034 1,122
66 0,031 1,023
72 0,026 0,858

Trong , OD = AT AB v l s liu c ly trung bnh.

 84

3.5

3.036
3.0
Hot tnh cellulase (UI/g CT)

2.5 2.310
2.145
1.947 1.980
2.0 1.881

1.551
1.5
1.386 1.122
1.023
1.0 0.858

0.5
12 18 24 30 36 42 48 54 60 66 72
Thi gian (gi)

th 4.4: Hot tnh cellulase theo thi gian nui cy

Nhn xt:
th 4.4 cho thy hot tnh enzyme cellulase t cao nht khi nui cy nm
mc c 30 gi (hot tnh l 3,036 UI/g CT). Thi gian cng tng (t 36 n 72 gi)
th hot tnh cellulase cng gim (2,310 UI/g CT 36 gi v gim n 0,858 UI/g CT
72 gi). Tng t nh mi lin h gia thi gian nui cy vi hot tnh pectinase,
thi gian 12 gi, 18 gi v 24 gi, hot tnh enzyme cellulase cng khng cao do nm
mc cha thch ng v pht trin c trong mi trng ch phm. Thi gian 30 gi l
khong thi gian thch hp nht cho nm mc pht trin v sn sinh cellulase.
Vy 30 gi l khong thi gian ti u cho qu trnh sinh tng hp ca nm mc
i vi 2 loi enzyme pectinase v cellulase (vi t l c cht trong ch phm l 2 phn
bt m : 1 phn bt sn).
 85

Hnh 4.8: Mu ch phm ti u v t l vi 2 phn bt m : 1 phn bt sn

Hnh 4.9: Mu ch phm ti u v t l (2 phn bt m : 1 phn bt sn)

v ti u v thi gian nui cy (30 gi)
 86

4.4. XC NH NH HNG CA THI GIAN LN MEN

To mt mu ch phm ti u, vi t l c cht l 2 phn bt m : 1 phn bt sn
v thi gian nui cy l 30 gi.
C nh thi gian ngm ht trong nc l 1 gi (c xc nh trong mc 4.6).
Tip theo, c nh hm lng ch phm s dng l 0,01%, chng ti tin hnh ln men
6 mu c ph vi 6 khong thi gian khc nhau.
Sau , chng ti xc nh nh hng ca thi gian ln men ln cc yu t nh
kh nng trch ly, ha tan v pH ca dch trch.
Mu i chng l c ph nhn cha ln men, khng qua ngm v cng c
rang, nghin vi cng mc .
Trng lng mi m c ph em ln men l 100 g (vi 0,01 g ch phm).
4.4.1. Xc nh nh hng ca thi gian n kh nng trch ly c ph
Kh nng trch ly ca c ph th hin qua khi lng cht tan thu c.
Bng 4.6: Mi tng quan gia thi gian ln men v khi lng cht tan
Thi gian (gi) i chng 12 16 20 24 28 32

Khi Bi 1,67 1,77 2,02 1,91 1,87 1,86 1,86

lng cht S 1,76 1,82 2,49 2,42 2,41 2,38 2,38
tan (g) Mokka 1,60 1,79 2,04 1,86 1,82 1,82 1,82

3
Khi lng cht tan (g))

2.5

2.02
2 1.91 1.87 1.86 1.86

1.77

1.5

1
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.5: Khi lng cht tan ca c ph Bi theo thi gian ln men

 87

Nhn xt: Theo th 4.5, i vi c ph Bi, khi lng cht tan thu c nhiu nht
thi gian ln men l 16 gi (t 2,02 g).

2.49
Khi lng cht tan (g))

2.5 2.42 2.41 2.38 2.38

2
1.82

1.5

1
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.6: Khi lng cht tan ca c ph S theo thi gian ln men

Nhn xt: Theo th 4.6, sau khi ln men 16 gi, khi lng cht tan ca c ph S
cng t cao nht vi 2,49 g.

3
Khi lng cht tan (g))

2.5

2.04
2 1.86 1.82
1.82 1.82
1.79

1.5

1
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.7: Khi lng cht tan ca c ph Mokka theo thi gian ln men
Nhn xt: Theo th 4.7, khi lng cht tan ca Mokka cng nhiu nht sau khi ln
men 16 gi (t 2,04 g), tng t vi c ph Bi v S.
 88

3 Bi S Mokka

Khi lng cht tan (g)

2.5

1.5

1
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.8: Khi lng cht tan 3 loi c ph theo thi gian ln men
Nhn xt:
Kt qu tng hp th 4.8 cho thy, khi lng cht tan ca cc loi c ph
(Bi, S, Mokka) t cao nht 16 gi, sau gim nh v khng thay i cc
khong thi gian sau. thi gian 12 gi, nm mc cha pht trin nhiu nn khi
lng cht tan khng cao. Sau , nm mc pht trin mnh, sau khi s dng ht
nhng cht hu c n gin trong c ph, chng s phn gii pectin v cellulose thnh
nhng tiu phn lm thc n. Chnh iu ny gip gia tng lng cht tan trong
dung dch, bi v khi pectin v cellulose (l hai thnh phn kh phn hy lm cn tr
kh nng trch ly) b phn gii th s gia tng hiu sut trch ly ca c ph. Thi gian
16 gi l thi im qu trnh phn gii ca chng din ra mnh nht v trit nht.
Khi , khi lng cht tan thu c cao nht. Sau 16 gi, nm mc bt u s dng
nhng sn phm va phn gii c, do khi lng cht tan gim nh.
4.4.2. Xc nh nh hng ca thi gian n ha tan
Bng 4.7: Mi tng quan gia thi gian v ha tan
Thi gian (gi) i chng 12 16 20 24 28 32
Bi 6,4 6,6 8,2 7,8 7,4 7,0 7,0
Brix S 5,2 5,2 6,0 5,8 5,6 5,0 5,0
Mokka 6,2 6,4 7,8 7,6 7,2 7,0 7,0
 89

9.0
8.2

8.0 7.8

ha tan ( Brix))
7.4
7.0 7.0
7.0
6.6

6.0

5.0

4.0
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.9: ha tan ca c ph Bi theo thi gian ln men

Nhn xt: Theo th 4.9, ha tan ca c ph Bi t cao nht l 8,2 ( Brix)
mu 16 gi.

9.0

8.0
ha tan ( Brix))

7.0

6.0
6.0 5.8
5.6

5.2 5.0 5.0

5.0

4.0
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.10: ha tan ca c ph S theo thi gian ln men

Nhn xt: Theo th 4.10 th 6,0 ( Brix) l ha tan cao nht ti 16 gi ca c
ph S.
 90

9.0

8.0 7.8
7.6

ha tan ( Brix))
7.2
7.0 7.0
7.0

6.4
6.0

5.0

4.0
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.11: ha tan ca c ph Mokka theo thi gian ln men

Nhn xt: Tng t vi c ph Bi v c ph S, kt qu t th 4.11 cho thy thi
gian 16 gi cng l mu c ha tan cao nht i vi c ph Mokka (7,8 Brix).

9 Bi S Mokka

8
ha tan ( Brix)

4
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.12: ha tan ca 3 loi c ph theo thi gian ln men

Nhn xt:
ha tan ca c ph ln men tng ng vi khi lng cht tan thu c, hay
ni cch khc l ha tan t l thun vi khi lng cht tan, cng nh t l thun vi
kh nng trch ly. Kt qu t th tng hp 4.12 cho thy, ha tan ca c ph Bi,
S v Mokka cng t gi tr cao nht mu ln men 16 gi. Sau th ha tan
 91

cng gim dn do vic nm mc s dng ngc li nhng sn phm n gin m

chng va phn gii c t pectin v cellulose.
4.4.3. Xc nh nh hng ca thi gian n pH ca dung dch cht tan
trch t c ph ln men
Bng 4.8: Mi tng quan gia thi gian ln men v pH dung dch cht tan
Thi gian (gi) i chng 12 16 20 24 28 32
Bi 5,68 6,15 5,92 5,87 5,73 5,69 5,60
pH S 5,69 6,02 5,86 5,83 5,71 5,66 5,65
Mokka 5,29 5,74 5,65 5,61 5,53 5,48 5.39

6.5

6.15

6.0 5.92
5.87

5.73
pH

5.69
5.60

5.5

5.0
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.13: pH dung dch cht tan trch t c ph Bi theo thi gian ln men
Nhn xt: Theo th 4.13, pH ca dung dch cht tan trch t c ph Bi ln men gim
dn theo thi gian ln men.
 92

6.5

6.02
6.0
5.86 5.83
pH

5.71
5.66 5.65

5.5

5.0
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.14: pH dung dch cht tan trch t c ph S theo thi gian ln men
Nhn xt: Theo th 4.14, pH ca dung dch cht tan trch t c ph S cng gim
dn theo thi gian ln men.

6.5

6.0
pH

5.74
5.65
5.61
5.53
5.48
5.5 5.39

5.0
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.15: pH dch cht tan trch t c ph Mokka theo thi gian ln men
Nhn xt: Theo th 4.15, tng t nh c ph Bi v c ph S, pH ca dung dch
cht tan trch t c ph Mokka cng gim dn theo thi gian ln men.
 93

Bi S Mokka

6.5

6
pH

5.5

5
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.16: pH dung dch cht tan trch t 3 loi c ph theo thi gian
Nhn xt:
Kt qu tng hp th 4.16 cho thy, pH ca dung dch cht tan trch t 3
loi c ph Bi, S, Mokka gim dn theo thi gian ln men. Nguyn nhn l do qu
trnh nm mc phn gii pectin thnh cc n phn ca n l acid galacturonic, ng
thi to iu kin gii phng cc thnh phn tan trong nc c trong ht c ph, trong
c nhng acid. Theo thi gian, nm mc pht trin to ra trong dch c ph ngy
cng nhiu acid lm cho pH ca dung dch ngy cng gim.
4.5. XC NH NH HNG CA HM LNG CH PHM
Sau khi chn c thi gian ln men ti u l 16 gi, chng ti c nh thi
gian v tin hnh kho st hm lng ch phm cho vo khi c ph ln men.
Thc hin 6 mu th nghim vi 6 hm lng ch phm khc nhau.
C nh thi gian ngm ht trong nc l 1 gi (c xc nh trong mc 4.6).
Mu i chng l c ph nhn cha ln men, khng qua ngm v cng c
rang, nghin vi cng mc .
Trng lng mi m c ph em ln men l 100 g.
 94

4.5.1. Xc nh nh hng ca hm lng ch phm n kh nng trch ly c

ph
Bng 4.9: Mi tng quan gia hm lng ch phm v khi lng cht tan
i
T l ch phm (%) 0,04 0,08 0,12 0,16 0,20 0,24
chng

Khi Bi 1,67 2,01 2,09 2,12 2,25 2,15 2,08

lng cht S 1,76 1,97 2,07 2,09 2,19 2,17 2,11
tan (g) Mokka 1,60 1,80 1,82 1,90 1,98 1,92 1,86

2.3
2.25

2.2
2.15
Khi lng cht tan (g))

2.12
2.09 2.08
2.1
2.01
2.0

1.9

1.8

1.7
0.04 0.08 0.12 0.16 0.2 0.24
T l ch phm (%)

th 4.17: Khi lng cht tan ca c ph Bi theo hm lng ch phm

Nhn xt: Theo th 4.17, khi lng cht tan ca c ph Bi t cao nht (2,25 g)
mu c hm lng ch phm 0,16%.

2.3

2.19
2.2 2.17
Khi lng cht tan (g))

2.11
2.07 2.09
2.1

2.0 1.97

1.9

1.8

1.7
0.04 0.08 0.12 0.16 0.2 0.24
T l ch phm (%)

th 4.18: Khi lng cht tan ca c ph S theo hm lng ch phm

 95

Nhn xt: Theo th 4.18, c ph S, khi lng cht tan t 2,19 g mu s dng
0,16% ch phm, y cng l gi tr cao nht.

2.3

2.2
Khi lng cht tan (g))

2.1

2.0 1.98
1.92
1.90
1.9 1.86
1.82
1.80
1.8

1.7
0.04 0.08 0.12 0.16 0.2 0.24
T l ch phm (%)

th 4.19: Khi lng cht tan ca c ph Mokka theo hm lng ch phm

Nhn xt: Theo th 4.19, trong trng hp ca c ph Mokka cng tng t nh c
ph Bi v S, khi lng cht tan thu c cao nht (1,98 g) mu s dng 0,16% ch
phm.

Bi S Mokka

2.3

2.2
Khi lng cht tan (g)

2.1

2.0

1.9

1.8

1.7
0.04 0.08 0.12 0.16 0.20 0.24
T l ch phm (%)

th 4.20: Khi lng cht tan ca 3 loi c ph theo hm lng ch phm

 96

Nhn xt:
Kt qu tng hp t th 4.20 cho thy, khi lng cht tan tng dn cng vi
s gia tng ca hm lng ch phm. T mu c hm lng ch phm 0,04%, khi
lng cht tan thu c tng dn n mu c hm lng ch phm 0,16% th t gi
tr cao nht, sau th gim xung. Khi hm lng ch phm cng ln th lng nm
mc cng nhiu, do qu trnh thy phn pectin v cellulose ca chng cng mnh,
to ra cng nhiu cc sn phm l cc cht tan. Bn cnh , khi qu trnh phn gii
pectin v cellulose tng s ng thi gia tng s gii phng cc thnh phn l cc cht
tan c bn trong ht c ph.
Nhng khi s lng nm mc qu nhiu th chng s s dng nhiu c cht l
cht tan, khi s dng ht nhng cht n gin, chng s s dng n cc sn phm
cht tan m chng va to ra. Do , hm lng ch phm 0,20% v 0,24% l qu ln,
nn khi lng cht tan gim xung.
4.5.2. Xc nh nh hng ca hm lng ch phm n ha tan
Bng 4.10: Mi tng quan gia hm lng ch phm v ha tan
i
T l ch phm (%) 0,04 0,08 0,12 0,16 0,20 0,24
chng
Bi 6,4 7,0 7,6 7,8 8,0 7,8 7,4
Brix S 5,2 7,0 7,2 7,2 8,0 7,8 7,6
Mokka 6,2 6,8 7,0 7,0 7,4 7,2 7,0

8.0
8.0
7.8 7.8
7.6
7.4
ha tan ( Brix))

7.0
7.0

6.0
0.04 0.08 0.12 0.16 0.20 0.24
T l ch phm (%)

th 4.21: ha tan ca c ph Bi theo hm lng ch phm

 97

Nhn xt: Theo th 4.21, ha tan ca c ph Bi cao nht l 8,0 mu s dng

hm lng ch phm 0,16%.

8.0
8.0
7.8
7.6
ha tan ( Brix))

7.2 7.2
7.0
7.0

6.0
0.04 0.08 0.12 0.16 0.20 0.24
T l ch phm (%)

th 4.22: ha tan ca c ph S theo hm lng ch phm

Nhn xt: Theo th 4.22, ha tan ca c ph S cng t cao nht l 8,0 mu s
dng hm lng ch phm 0,16%.

8.0

7.4
ha tan ( Brix)

7.2
7.0 7.0 7.0
7.0
6.8

6.0
0.04 0.08 0.12 0.16 0.20 0.24
T l ch phm (%)

th 4.23: ha tan ca Mokka theo hm lng ch phm

Nhn xt: Theo th 4.23, ha tan ca Mokka t cao nht mu 0,16% vi gi
tr 7,4.
 98

Bi S Mokka
8.0
ha tan ( Brix))

7.0

6.0
0.04 0.08 0.12 0.16 0.20 0.24
T l ch phm (%)

th 4.24: ha tan ca 3 loi c ph theo hm lng ch phm

Nhn xt:
Kt qu tng hp t th 4.24 cho thy, ha tan t l thun vi khi lng
cht tan, v cng t l thun vi kh nng trch ly ca c ph ln men. ha tan ca
c ph Bi, c ph S v c ph Mokka t gi tr cao nht mu c hm lng ch
phm 0,16%.
Vy 0,16% l hm lng thch hp c ph ln men thu c lng cht tan
nhiu nht, t ha tan cao nht v cng l hm lng c ph ln men c kh
nng trch ly cao nht.
4.5.3. Kho st nh hng ca hm lng ch phm n pH ca dung dch
cht tan trch t c ph ln men
Bng 4.11: Mi tng quan gia hm lng ch phm v pH dch cht tan
i
T l ch phm (%) 0,04 0,08 0,12 0,16 0,20 0,24
chng
Bi 5,68 5,48 5,47 5,45 5,43 5,39 5,36
pH S 5,69 5,85 5,80 5,73 5,63 5,50 5,43
Moka 5,29 5,51 5,48 5,44 5,39 5,38 5,35
 99

5.9

pH 5.7

5.48 5.47
5.5 5.45
5.43
5.39
5.36

5.3
0.04 0.08 0.12 0.16 0.20 0.24
T l ch phm (%)

th 4.25: pH dch cht tan trch t c ph Bi theo hm lng ch phm

Nhn xt: Theo th 4.25, pH gim dn tng ng vi hm lng ch phm tng
dn.

5.9
5.85
5.80
5.73

5.7
5.63
pH

5.50
5.5
5.43

5.3
0.04 0.08 0.12 0.16 0.20 0.24
T l ch phm (%)

th 4.26: pH dch cht tan trch t c ph S theo hm lng ch phm

Nhn xt: Theo th 4.26, pH dung dch gim dn t hm lng 0,04% n hm
lng 0,24%.
 100

5.9

pH 5.7

5.51
5.48
5.5 5.44
5.39 5.38
5.35

5.3
0.04 0.08 0.12 0.16 0.20 0.24
T l ch phm (%)

th 4.27: pH dung dch cht tan trch t Mokka theo hm lng ch phm
Nhn xt: Tng t nh dung dch cht tan trch t c ph Bi v c ph S, th 4.27
cho thy dung dch cht tan ca c ph Mokka cng c pH gim dn khi hm lng
ch phm s dng tng dn.

Bi S Mokka

5.9

5.7
pH

5.5

5.3
0.04 0.08 0.12 0.16 0.20 0.24
T l ch phm (%)

th 4.28: pH dch cht tan trch t 3 loi c ph theo hm lng ch phm

Nhn xt:
Kt qu tng hp t th 4.28 cho thy, pH dung dch cht tan ca c 3 loi c
ph Bi, S, Mokka u gim dn theo hm lng ch phm tng dn. Ni cch khc l
 101

pH ca dch trch cht tan t l nghch vi hm lng ch phm s dng. Nm mc

trong ch phm khi pht trin s phn gii pectin v cellulose to ra cc n phn l
acid, cng nh gii phng cc acid c trong thnh phn ca ht c ph. Khi s dng
hm lng ch phm cng nhiu, lng nm mc cng nhiu v pht trin mnh s to
ra nhiu acid. Do lm cho pH ca dung dch ngy cng gim.
4.6. XC NH NH HNG M HT C PH TRC LN MEN LN
KH NNG TRCH LY CHT HA TAN
Trng lng ban u mi khi c ph em ngm nc l 100 g. Mu i chng
l c ph nhn khng ngm nc v khng ln men.
Nhng khong thi gian ngm nc kho st:
0,5 gi 1 gi 1,5 gi 2 gi 2,5 gi 3 gi 3,5 gi 4 gi
Bng 4.12: Mi tng quan gia thi gian ngm nc v m ht c ph
Thi gian ngm (gi) 0.5 1 1.5 2 2.5 3 3.5 4
m Bi 22,50 29,22 32,56 34,83 36,28 41,51 41,73 41,82
ht S 21,52 26,35 33,18 34,36 35,95 42,98 43,59 43,70
(W%) Mokka 20,93 26,02 29,15 35,32 41,43 44,76 44,92 45,24

Bi S Mokka

50

45

40
m ht (W%))

35

30

25

20

15
0.5 1 1.5 2 2.5 3 3.5 4
Thi gian ngm (gi)

th 4.29: m ht theo thi gian ngm nc

 102

Nhn xt:
m ht c ph nguyn liu trc khi ln men l 7,86% (i vi c ph Bi),
7,26% (i vi c ph S) v 10,60% (i vi c ph Mokka). Vi hm lng nc
thp nh vy, khng thun li cho s pht trin ca nm mc. Do phi ngm ht
trong nc trc khi b sung ch phm.
Kt qu t th 4.29 cho thy, thi gian ngm ht cng lu th m ca ht
cng tng. T 0,5 gi n 3 gi th ht ht mt lng nc kh nhiu nn m tng
nhiu. Tuy nhin, t thi gian ngm 3 gi tr i, ht gn t bo ha v nc
nn lng nc ht vo khng nhiu, do m tng khng ng k.
4.6.1. Xc nh nh hng m ht ln kh nng trch ly
Bng 4.13: Mi tng quan gia thi gian ngm nc v khi lng cht tan
Thi gian ngm (gi) C 0.5 1 1.5 2 2.5 3 3.5 4
Bi 1,67 1,85 2,01 1,89 1,87 1,86 1,82 1,82 1,81
Khi lng
S 1,76 1,87 2,12 2.03 1,95 1,91 1,88 1,86 1,86
cht tan (g)
Mokka 1,60 1,90 1,96 1,89 1,88 1,85 1,83 1,82 1,79

Bi S Mokka

2.5
Khi lng cht tan (g)

2.0

1.5
0.5 1 1.5 2 2.5 3 3.5 4
Thi gian ngm (gi)

th 4.30: Khi lng cht tan 3 loi c ph theo thi gian ngm nc
 103

Nhn xt:
Kt qu t th 4.30 cho thy, khi lng cht tan thu c cao nht khi ngm
ht 1 gi. Khi ngm ht 0,5 gi, c ph khng m cho s pht trin ca nm
mc, do khi lng cht tan cha cao. m ht tng ln to iu kin thun li
cho nm mc pht trin, tit ra enzyme thy phn pectin v cellulose thnh nhng hp
cht d tan, ng thi gii phng cc thnh phn tan trong nc c bn trong ht c
ph. V vy khi lng cht tan s tng. Tuy nhin, khi ngm ht lu hn, ht trng
n lm cho qu trnh phn gii petin v cellulose xy ra qu mnh, to ra nhng l
hng trn b mt ht c ph. Do , khi tin hnh ra ht sau khi , s xut hin ca
nhng l hng ny to iu kin cho s mt mt nhng thnh phn ha tan ca ht d
dng hn.
4.6.2. Xc nh nh hng m ht ln ha tan
Bng 4.14: Mi tng quan gia thi gian ngm nc v ha tan
Thi gian ngm (gi) C 0.5 1 1.5 2 2.5 3 3.5 4
ha Bi 6,4 6,8 7,6 7,0 6,8 6,8 6,6 6,4 6,0
tan ( S 5,2 6,2 6,8 6,0 6,0 5,8 5,6 5,6 5,2
Brix) Mokka 6,2 6,4 7,2 6,8 6,4 6,4 6,2 6,0 5,6

Bi S Mokka

8.0
Khi lng cht tan (g)

7.0

6.0

5.0
0.5 1 1.5 2 2.5 3 3.5 4
Thi gian ngm (gi)

th 4.31: ha tan 3 loi c ph theo thi gian ngm nc

 104

Nhn xt:
Kh nng trch ly ca c ph ln men khng ch th hin qua khi lng cht
tan, m bn cnh n cn th hin qua ha tan o c. V ha tan ca c ph
lun lun t l thun vi khi lng cht tan. V vy, kt qu t th 4.31 cho thy,
ha tan ca c ph t gi tr cao nht cng mu c thi gian ngm nc l 1 gi.
Khi khi lng cht tan gim th ha tan cng gim.
4.6.3. Xc nh nh hng m ht ln pH ca dung dch cht tan
Bng 4.15: Mi tng quan gia thi gian ngm v pH dch trch cht tan
Thi gian ngm (gi) C 0.5 1 1.5 2 2.5 3 3.5 4
Bi 5,68 5,62 5,54 5,73 5,69 5,68 5,65 5,62 5,61
pH S 5,69 5,76 5,71 5,84 5,79 5,79 5,78 5,76 5,73
Mokka 5,29 5,40 5,38 5,72 5,64 5,62 5,61 5,60 5,59

Bi S Mokka

6.0
pH

5.0
0.5 1 1.5 2 2.5 3 3.5 4
Thi gian ngm (gi)

th 4.32: pH dch trch cht tan ca 3 loi c ph theo thi gian ngm nc
Nhn xt:
Kt qu th 4.32 cho thy, nhn chung pH ca dch chit gim theo chiu
tng ca m. Khi thi gian ngm l 1,5 gi th pH tng vt, sau li gim xung.
Nh cp trong phn 4.6.1, khi m ht tng ln, nm mc pht trin thun li
s tit nhiu enzyme phn gii pectin v cellulose, va to ra cc n phn ca pectin
 105

l acid galacturonic, va gip gii phng cc thnh phn tan trong nc bn trong ht
(trong c cc acid). Bn cnh thi gian ngm nc cng lu s to ra m cng
cao v lm ht c ph trng n nhiu, iu ny to thun li cho qu trnh ln men
ph to acid ca mt s loi vi sinh vt (nm men, nm mc, vi khun) c sn trn b
mt ht c ph cng nh nhim t khng kh vo trong qu trnh ln men. Chnh v
nhng l do ny lm cho pH ca dch trch gim xung.
Tuy nhin, khi thi gian ngm l 1,5 gi th pH tng vt cho thy s ra tri cc
thnh phn ca ht c ph l rt mnh, nhng acid trong ht b tan theo nc, do pH
tng ln.
4.7. XC NH S THAY I TRNG LNG KHI C PH TRC V
SAU KHI LN MEN
Trng lng mi khi c ph ca mi m trc khi ln men u l 100 g. Sau
qu trnh ln men, chng ti cn mt ln na trc khi em ra.
4.7.1. Xc nh s thay i trng lng khi c ph theo thi gian ln men
Bng 4.16: Mi tng quan gia thi gian ln men v trng lng
khi c ph ln men
Thi gian (gi) 12 16 20 24 28 32
Bi 145,23 141,75 140,64 140,97 141,31 141,51
Trng
S 147,05 140,32 132,74 133,23 133,86 134,01
lng
Moka 136,15 133,86 131,52 132,08 132,32 132,73

150
Trng lng khi ln men (g)))

145.23
145

141.75 141.51
140.64 140.97 141.31

140

135

130
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.33: S thay i trng lng c ph Bi theo thi gian ln men

 106

Nhn xt: Theo th 4.33, trng lng khi c ph gim dn theo thi gian ln men,
sau 20 gi th bt u tng nh.

150
Trng lng khi ln men (g)))

147.05

145

140.32
140

135 134.01
133.23 133.86
132.74

130
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.34: S thay i trng lng c ph S theo thi gian ln men

Nhn xt: Theo th 4.34, trng lng ca khi c ph S cng gim t mu 12 gi
n mu 20 gi v sau cng tng nh.

150
Trng lng khi ln men (g)))

145

140

136.15
135 133.86 132.73
131.52 132.08 132.32

130
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.35: S thay i trng lng c ph Mokka theo thi gian ln men

Nhn xt: Theo th 4.35, c ph Mokka cng c trng lng khi c ph sau ln
men gim n mu 20 gi v sau cng tng nh.
 107

Bi S Mokka

150
Trng lng khi ln men (g)

145

140

135

130
12 16 20 24 28 32
Thi gian ln men (gi)

th 4.36: S thay i trng lng 3 loi c ph theo thi gian ln men

Nhn xt:
Kt qu tng hp t th 4.36 cho thy, cc mu 12, 16 v 20 gi, trng
lng khi ln men gim dn.
Nguyn nhn th nht l do nm mc ngy cng pht trin, s dng ngy cng
nhiu c cht lm thc n v chuyn ha nhng cht hu c phc tp c trng lng
phn t ln thnh nhng cht n gin c trng lng phn t nh.
Nguyn nhn th hai l trong qu trnh ln men, c mt lng nc bc hi vo
khng kh. Ngoi ra, khi nm mc thc hin qu trnh thy phn pectin v cellulose s
to ra mt s cht bay hi, chng hn nh CO2 v H2O, y l nguyn nhn th ba.
Sau thi gian 20 gi, trng lng khi ln men bt u tng ln nhng ch l s
tng nh. Nguyn nhn ca s tng trng lng ny trc ht l do khi thi gian cng
lu, nm mc pht trin ngy cng nhiu v sinh khi ca chng s tng ln rt ng
k. Bn cnh nm mc trong ch phm m chng ti b sung vo, cn c s c mt
ca cc loi nm mc, nm men v vi khun c sn trn b mt ht c ph cng nh
nhim t khng kh vo trong qu trnh ln men. Chnh s gia tng sinh khi ca tt c
cc vi sinh vt ny lm gia tng trng lng khi c ph ln men.
 108

4.7.2. Xc nh s thay i trng lng khi theo hm lng ch phm

Bng 4.17: Mi tng quan gia hm lng ch phm v trng lng khi
T l ch phm
0,04 0,08 0,12 0,16 0,20 0,24
(%)
Bi 138,67 137,06 136,82 134,96 134,26 133,77
Trng
S 140,30 138,61 136,95 136,27 135,92 135,35
lng
Moka 133,58 133,55 130,80 130,63 130,21 129,75

142

140
Trng lng khi ln men(g))

138.67
138
137.06 136.82

136
134.96
134.26
133.77
134

132

130

128
0.04 0.08 0.12 0.16 0.20 0.24

T l ch phm (%)

th 4.37: S thay i trng lng ca c ph Bi theo hm lng ch phm

Nhn xt: Theo th 4.37, trng lng ca c ph Bi gim dn khi hm lng ch
phm tng dn.

142

140.30
140
Trng lng khi ln men(g))

138.60
138
136.95
136.27
135.92
136 135.35

134

132

130

128
0.04 0.08 0.12 0.16 0.20 0.24

T l ch phm (%)

th 4.38: S thay i trng lng ca c ph S theo hm lng ch phm

 109

Nhn xt: Theo th 4.38, s thay i trng lng ca khi c ph S ln men cng
theo chiu hng gim dn ng vi s tng dn ca hm lng ch phm.

142

140
Trng lng khi ln men(g))

138

136

134 133.58
132.55
132
130.80 130.63
130.21
129.75
130

128
0.04 0.08 0.12 0.16 0.20 0.24

T l ch phm (%)

th 4.39: S thay i trng lng c ph Mokka theo hm lng ch phm

Nhn xt: Theo th 4.39 th tng t vi c ph Bi v c ph S, trng lng khi
c ph ln men ca Mokka cng t l nghch vi hm lng ch phm.

Bi S Mokka
142
Trng lng khi ln men (g))

140

138

136

134

132

130

128
0.04 0.08 0.12 0.16 0.20 0.24
T l ch phm (%)

th 4.40: S thay i trng lng 3 loi c ph theo hm lng ch phm

Nhn xt:
Kt qu tng hp t th 4.40 cho thy, trng lng khi c ph ln men gim
dn, t l nghch vi s tng dn ca hm lng ch phm.
 110

Tuy hm lng ch phm a vo khi ln men c khc nhau nhng trong

trng hp ny khng c s gia tng trng lng khi ln men. V cc mu ln men
trong cng mt thi gian l 16 gi nn s gia tng trng lng sinh khi ca nm mc
khng ln bng s thay i ca qu trnh phn gii.
Khi hm lng ch phm cng ln, s lng nm mc cng nhiu, chng s
dng mt lng ln c cht sinh trng v pht trin. Ngoi ra, cng nhiu nm
mc th qu trnh phn gii cng mnh, bn cnh vic to ra cng nhiu sn phm bay
hi (nh CO2 v H2O), chng cn lm tng nhit ca khi , lm tng lng nc
bc hi vo khng kh.
4.8. XC NH NHIT KHI THEO THI GIAN LN MEN
Trong th nghim ny, chng ti kho st nhit ca khi trong thi gian ln
men l 16 gi v t l ch phm s dng l 0,16% (y l thi gian v t l ti u
c xc nh). Sau 2 gi th chng ti o nhit mt ln bng nhit k. Trng lng
ca khi c ph em ln men l 100 g.

Hnh 4.10: o nhit khi c ph Hnh 4.11: Nhit khi c ph

ln men theo thi gian ln men sau 2 gi (27,3oC)

Bng 4.18: Mi tng quan gia thi gian ln men v nhit khi
Thi gian (gi) 0 2 4 6 8 10 12 14 16
Nhit (oC) 26,8 27,3 27,5 27,7 28,0 27,0 26,5 26,3 26,2
 111

30.0

29.0

28.0
28.0 27.7
27.5
Nhit (oC)

27.3

27.0
26.8
27.0
26.5
26.3 26.2

26.0

25.0
0 2 4 6 8 10 12 14 16
Thi gian (gi)

th 4.41: Nhit khi theo thi gian ln men

Nhn xt:
Kt qu th 4.41 cho thy, nhit ca khi tng dn t lc bt u ln
men cho n sau 8 gi, sau th nhit ca khi gim dn. Nhit ca khi
ln cao nht l sau khi ln men c 8 gi (y cng l thi im gia ca qu trnh
ln men). Nguyn nhn l do nm mc pht trin, phn gii c cht v ta nhit. Ngoi
ra, do nc bc hi dn nn nhit tng. Sau , nhit li gim xung do lng
nc sinh ra t qu trnh phn gii c cht ca nm mc.
 112

PHN 5: KT LUN V NGH

Qua nhng kt qu nghin cu trn, chng ti rt ra nhng kt lun sau:

1. T l c cht ti u cho ch phm l 2 phn bt m v 1 phn bt sn.
2. Thi gian ch phm 30 gi l ti u cho sinh tng hp pectinase v
cellulase.
3. Ch phm Biocoffee-1 c hiu qu tt trong ln men ba loi c ph ph bin
Vit Nam (c ph Bi, S v Mokka). C ph sau ln men c ha tan v khi
lng cht tan thu c tng r rt so vi c ph khng c ln men.
4. Ngm ht c ph trc khi ln men trong 1 gi l thi gian ngm thch hp
gip thu c nhiu cht ha tan nht.
5. Thi gian ln men ti u l 16 gi.
6. T l ch phm s dng l 0,16% th hiu sut trch ly t ti a, thu c
lng cht ha tan cao nht.
7. pH dch trch cht tan gim dn theo thi gian ln men v cng gim dn
theo s gia tng ca hm lng ch phm.
8. Trng lng khi gim dn theo s gia tng ca hm lng ch phm.
Theo thi gian ln men th trng lng khi gim dn, v sau li tng nh.
9. Nhit khi t cao nht sau khi ln men 8 gi.

V gii hn v thi gian v mt s iu kin, ti ca chng ti cha kho st

c mt s kha cnh c lin quan nn chng ti c mt s ngh sau:
1. To loi ch phm sinh hc mi vi thnh phn l mt s ngun tinh bt
khc v vi t l phi trn khc.
2. To loi ch phm sinh hc mi t cc ngun vi sinh vt khc.
3. Kho st cc thnh phn trong ht c ph sau ln men.
4. Tin hnh cm quan c ph thnh phm sau ln men.
5. Kho st qu trnh ln men i vi mt s loi c ph khc.
6. S dng enzyme tinh khit cho qu trnh ln men c ph.
 113

PHN 6: TI LIU THAM KHO

TING VIT
[1] Lm Th Kim Chu, Vn c Chnh v Ng i Nghip, 2004. Thc tp ln Sinh
ha. Nh xut bn i hc Quc gia, thnh ph H Ch Minh, trang 3 87.

[2] V Vn Chi, 1997. T in cy thuc Vit Nam. Nh xut bn Y hc, mc C 21.

[3] Vit Chng, 2000. K thut trng c ph. Nh xut bn Thnh ph H Ch Minh,
trang 73 76.

[4] Nguyn Ln Dng, 1979. Mt s phng php nghin cu vi sinh vt, tp 2. Nh

xut bn Khoa hc v K thut, H Ni.

[5] Nguyn Ln Dng, Phm Vn Ty v Dng c Tin, 1979. Vi sinh vt hc tp

1. Nh xut bn i hc v Trung hc chuyn nghip, H Ni, trang 171 190.

[6] Bi Th t v V Khc Nhng, 1998. K thut gieo trng, ch bin ch v c

ph. Nh xut bn Nng nghip, H Ni, trang 47 95.

[7] Nguyn Kh Ha, 1994. Ln vi cy c ph ch. Nh xut bn Nng nghip, H

Ni, trang 11 16.

[8] Phm Hong H, 1972. Cy c min Nam Vit Nam, tp 2. B Gio Dc Trung
tm hc liu, trang 73.

[9] Phm Th nh Hng, 2003. K thut sinh ha. Nh xut bn i hc Quc Gia,
thnh ph H Ch Minh, trang 5 115, 152 180.

[10] Trng Hng, 2001. K thut trng c ph mt dy. Nh xut bn Nng

nghip, H Ni, trang 3 23, 96 98.

[11] L Quang Hng, 1999. K thut trng v thu hoch c ph xut khu. Nh xut
bn Gio dc, trang 1 170.

[12] Nguyn c Lng, 2002. Cng ngh vi sinh, tp 2 Vi sinh vt hc cng

nghip. Nh xut bn i hc Quc Gia, thnh ph H Ch Minh, trang 187 272.

[13] Nguyn c Lng, Cao Cng, Nguyn nh Tuyt, L Th Thy Tin, T Thu
Hng, Hunh Ngc Oanh, Nguyn Thy Hng v Phan Th Huyn, 2004. Cng
ngh enzyme. Nh xut bn i hc Quc Gia, thnh ph H Ch Minh, trang 7
59, 288 526.
 114

[14] Nguyn c Lng v Cao Cng, 2003. Th nghim Cng ngh sinh hc, tp 1
Th nghim Ha Sinh hc. Nh xut bn i hc Quc Gia, thnh ph H Ch
Minh.

[15] Nguyn c Lng, Phan Th Huyn v Nguyn nh Tuyt, 2003. Th nghim

Cng ngh sinh hc, tp 2 Th nghim vi sinh vt hc. Nh xut bn i hc
Quc Gia, thnh ph H Ch Minh, trang 108 126.

[16] Nguyn Vn Mi, 2001. Thc hnh Ha Sinh hc. Nh xut bn Khoa hc v K
thut, H Ni, trang 42 48.

[17] on Th Thanh Nhn, trng i hc Nng nghip H Ni, 1996. Gio trnh
cy cng nghip. Nh xut bn Nng nghip, H Ni, trang 203 206.

[18] Trn Thnh Phong, 2004. Kho st kh nng sinh tng hp enzyme cellulase t
Trichoderma reesei v Aspergillus niger trn mi trng ln men bn rn. Lun
vn Thc s Sinh hc, chuyn ngnh Vi sinh. Trng i hc Khoa hc T nhin,
thnh ph H Ch Minh, Vit Nam.

[19] L Hng Ph, 2003. Nghin cu sinh tng hp enzyme pectinase v cellulase t
Aspergillus niger v ng dng x l v c ph trong sn xut phn hu c. Lun
vn Thc s Sinh hc, chuyn ngnh Ha sinh. Trng i hc Khoa hc T nhin,
thnh ph H Ch Minh, Vit Nam.

[20] Phan Quc Sng, 1995. K thut trng, chm sc, ch bin c ph. Nh xut bn
Nng nghip, thnh ph H Ch Minh, trang 5 9, 68 70.

[21] Phm Tr Thng, 1999. Gio trnh Bo qun Ch bin c ph. B mn Bo

qun - Ch bin Nng sn thc phm. Trng i hc Nng Lm, thnh ph H
Ch Minh, trang 1 50.

[22] Trn Thanh Thy, 1999. Hng dn thc hnh vi sinh vt hc. Nh xut bn
Gio dc, thnh ph H Ch Minh, trang 37 38, 48 52.

[23] L Ngc T, La Vn Ch, Phm Trn Chu v Nguyn Ln Dng, 1982. Enzyme
vi sinh vt, tp 2. Nh xut bn Khoa hc v K thut, H Ni.

[24] Trng i hc Bch Khoa, B mn Cng Ngh Thc phm, 2003. Th nghim
Ha Sinh. Khoa K thut Ha hc v Du kh, trng i hc Bch Khoa, thnh
ph H Ch Minh.

[25] Trng i hc Khoa hc T nhin, B mn Sinh ha, 2004. Gio trnh thc tp
Cng ngh enzyme v protein. Khoa Sinh hc, trng i hc Khoa hc T nhin,
thnh ph H Ch Minh, trang 4 7.
 115

[26] Trng i hc Khoa hc T nhin, B mn Cng ngh sinh hc, Khoa Sinh
hc, 2003. Gio trnh thc tp ln Chuyn ha sinh hc. T sch trng i hc
Khoa hc T nhin, thnh ph H Ch Minh, trang 66 68.

[27] Trng i hc Nng Lm, B mn Cng ngh sinh hc, 2004. Qui ch thc tp
tt nghip, thc hin kha lun v bo co tt nghip. Trng i hc Nng Lm,
thnh ph H Ch Minh, trang 1 12.

[28] Trng Trung hc Dc, 1997. Dc liu hc. Nh xut bn Y hc, H Ni,
trang 126.

[29] Vin dc liu, 2003. Cy thuc v ng vt lm thuc Vit Nam, tp 1. Nh

xut bn Y hc, H Ni, trang 297 300.

TING NC NGOI
[30] Carlo N.Hamelinck, 2003. Prospects for ethanol from lignocellulosic biomass:
techno-economic performance as development progresses. Universiteit Utrecht
Copernicus Institute, Science Technology Society.

[31] Clarke R.J. and Macrae R., 1988. Coffee, volume 1 Chemistry. Elsevier applied
science London and New York.

[32] Clarke R.J. and Macrae R., 1988. Coffee, volume 2 Technology. Elsevier
applied science London and New York.

[33] Pitt J.I. and Hocking A.D., 1997. Fungi and food spoilage. Blackie Academic and
Professional An Imprint of Chapman and Hall.

[34] Uhlig Helmut, Elfriede M.Linsmaier Bednar, 1998. Industrial enzymes and
their applications. Awiley Interscience Publication, John Wiley and Sons, Inc.

INTERNET
[35] www.agroviet.gov.vn/en/stories/TinTiengAnh/VietnamAgricultureIn2001.asp

[36] www.agroviet.gov.vn/loadasp/hang/spec-good.asp?type=coffee

[37] www.agroviet.gov.vn/tapchi/khcn/2003/Noidung/So2-02.asp

[38] www.angelfire.com/pa2/hartmann/pgs/coff_proc.html

[39] www.enzymes.co.uk/answer24_pectinase.htm

[40] www.gardfoods.com/coffee/coffee.plant.htm

[41] www.images.google.com.vn/imgres?imgurl
 116

[42] www.images.google.com.vn/imgres?imgurl=http://www.ttuhsc.edu/SOM/Microb
iology/mainweb/aiaq/Pictures/Aspergillus%2520niger.jpg&imgrefurl

[43] www.sci-toys.com/ingredients/pectin.html

[44] www.trade.hochiminhcity.gov.vn/data/ttth/2002/20021121-in-xkcpvn.htm

[45] www.vicofa.org.vn/pls/news/Cate$.htnoidung?v_cata_code=1201&v_item_id=5
7586

[46] www.vicofa.org.vn/vicofa/cafevn.shtml

[47] www.vicofa.org.vn/vicofa/cfvn_sx.shtml

[48] www.vicofa.org.vn/vicofa/cfvn_xk.shtml

[49] www.vicofa.org.vn/vicofa/tintuc.shtml
 117

PHN 7: PH LC

Hnh 7.1: Nm mc trong Hnh 7.2: Nm mc trong

mi trng thch malt mi trng gi ging PGA

Hnh 7.3: Nm mc c nui trong mi trng nhn ging

(mi trng cm go)
 118

Ph lc 1: Thnh phn mi trng s dng

Ph lc 1.1: Mi trng thch malt (nui cy nm mc)
Malt 200g
Agar 18o/oo
Nc ct cho 1000ml
Ph lc 1.2: Mi trng PGA (gi ging)
Khoai ty 200g
ng glucose 20g
Agar 16 18g
Nc ct cho 1000ml
Ph lc 1.3: Mi trng cm go (nhn ging)
Cm go 75%
Bt c rt 19%
Tru 5%
(NH4)2SO4 1%

Ph lc 2: Cc ng chun
Ph lc 2.1: ng chun glucose trong xc nh hot tnh cellulase
Bng 7.1: Mi tng quan gia nng glucose v mt quang
trong xc nh hot tnh cellulase
ng s 0 1 2 3 4 5 6
Nng glucose
0 0,1 0,2 0,3 0,4 0,5 0,6
(mg/ml)
OD 1,835 2,016 2,156 2,340 2,492 2,690 2,833
OD 0 0,181 0,321 0,505 0,657 0,855 0,998

Trong :
ng s 0 l ng i chng.
OD = tr s mt quang ca ng mu tr tr s mt quang ca ng i
chng (AG AW).
 119

1.0
0.998

0.8 0.855

0.657
0.6
OD

0.505
y = 1.6788x
0.4 R2 = 0.999

0.321

0.2
0.181

0.0
0 0.1 0.2 0.3 0.4 0.5 0.6
Nng glucose (mg/ml)

th 7.1: ng chun glucose trong xc nh hot tnh cellulase

Ph lc 2.2: ng chun glucose trong xc nh ng tng s ca c ph

Hnh 7.4: Kt qu phn ng mu vi antron khi dng ng chun glucose

trong xc nh ng tng s
 120

Bng 7.2: Mi tng quan gia nng glucose v mt quang

trong xc nh ng tng s
ng s 1, 2 3 4 5 6 7
Nng glucose
0 0.1 0.2 0.3 0.4 0.5
(mg/ml)
OD 0.063 0.352 0.581 0.894 1.118 1.396
OD 0 0.289 0.518 0.831 1.055 1.333

Trong :
ng s 1 v s 2 l ng i chng, ly tr s trung bnh mt quang ca 2
ng ta c mt quang 0,063.
OD = tr s mt quang ca ng mu tr tr s mt quang ca ng i
chng.

1.6

1.2 1.333

1.055
OD

0.8
0.831

0.518 y = 2.6733x
0.4
R2 = 0.9985
0.289

0
0 0.1 0.2 0.3 0.4 0.5
Nng glucose (mg/ml)

th 7.2: ng chun glucose trong xc nh ng tng s

 121

Ph lc 2.3: ng chun glucose trong xc nh ng kh ca c ph

Bng 7.3: Mi tng quan gia nng glucose v mt quang
trong xc nh ng kh
ng s 0 1 2 3 4 5 6 7 8
Nng glucose
0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8
(mg/ml)
OD 0 0.294 0.511 0.723 0.991 1.197 1.433 1.603 1.793

Trong :
ng s 0 l nc ct, l ng i chng.
OD = tr s mt quang ca ng mu tr tr s mt quang ca ng i
chng.

1.793
1.6
1.603
1.433
1.2
1.197
OD

0.991
0.8
0.723 y = 2.3332x
2
R = 0.9941
0.4 0.511
0.294

0
0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8
Nng glucose (mg/ml)

th 7.3: ng chun glucose trong xc nh ng kh

 122

Ph lc 3: Cc s liu t thc nghim

Ph lc 3.1: Cc s liu trong xc nh hm lng cellulose
Bng 7.4: Cc s liu trong xc nh hm lng cellulose
Trng lng giy lc Trng lng giy Trng lng
cha cellulose (g) lc (g) cellulose (g)
Bi 1,0384 0,7556 0,2828
S 1,0610 0,7469 0,3141
Mokka 1,0612 0,7476 0,3136

Ph lc 3.2: Cc s liu trong xc nh hm lng pectin

Bng 7.5: Cc s liu trong xc nh hm lng pectin
Trng lng giy lc Trng lng giy Trng lng kt ta
cha kt ta (g) lc (g) canxi pectat (g)
Bi 0,8618 0,7726 0,0892
S 0,8625 0,7767 0,0858
Mokka 0,8654 0,7755 0,0899

Ph lc 3.3: Cc s liu trong xc nh hm lng ng tng s

Sau khi trch ng t 1 g mu, dung dch thu c em pha long 10 ln trc
khi xc nh ng tng s.
Bng 7.6: Cc s liu mt quang trong xc nh hm lng ng tng s
OD1 OD2 OD3 ODTB ODi chng OD
Bi 1,192 1,182 1,175 1,183 0,063 1,120
S 1,202 1,185 1,190 1,192 0,063 1,129
Mokka 1,386 1,394 1,389 1,390 0,063 1,327

Bng 7.7: Cc s liu tnh ton trong xc nh hm lng ng tng s

Nng ng x (mg/ml) Trng lng ng (mg)
Bi 0,419 209,5
S 0,422 211,0
Mokka 0,513 256,5
 123

Ph lc 3.4: Xc nh hm lng ng kh
Sau khi trch ng t 1 g mu, dung dch thu c em pha long 2 ln trc
khi xc nh hm lng ng kh.
Bng 7.8: Cc s liu mt quang trong xc nh hm lng ng kh
OD1 OD2 OD3 ODTB ODi chng OD
Bi 0,690 0,687 0,689 0,689 0 0,689
S 0,706 0,692 0,706 0,701 0 0,701
Mokka 0,971 0,969 0,978 0,973 0 0,973

Bng 7.9: Cc s liu tnh ton trong xc nh hm lng ng kh

Nng ng (mg/ml) Lng ng kh trong mu (mg)
Bi 0,295 29,5
S 0,300 30,0
Mokka 0,417 41,7

Ph lc 3.5: Cc s liu trong xc nh hm lng tro

Bng 7.10: Cc s liu trong xc nh hm lng tro
Trng lng Trng lng Trng lng Trng lng tro
mu (g) chn (g) chn v tro (g) (g)
Bi 4,00 30,00 30,20 0,20
S 4,00 30,00 30,20 0,20
Mokka 4,00 30,00 30,22 0,22

Ph lc 3.6: m ca nguyn liu c ph

Bng 7.11: Cc s liu khi xc nh m ca nguyn liu c ph
C ph m1 m2 (g) m (g) W (%)
Bi 0,393 5 7,86
S 0,363 5 7,26
Mokka 0,530 5 10,60
Trong , hiu s m1 - m2 l cc s liu c ly trung bnh.
 124

Ph lc 3.7: m ca vt liu
Bng 7.12: Cc s liu khi xc nh m ca cc vt liu
Vt liu m1 m2 (g) m (g) W (%)
Bt m 0,810 10 8,10
Bt sn 0,977 10 9,77
Bt c rt 0,407 10 4,07
Cm go 0,717 10 7,17
Tru 0,410 5 8,20
Trong , hiu s m1 - m2 l cc s liu c ly trung bnh.

Ph lc 4: m yu cu cho s pht trin ti u ca nm mc

Bng 7.13: m yu cu cho s pht trin ti u ca nm mc
W (%)
Mi trng bn rn (cm go) 56
Mi trng ch phm 45
Khi c ph ln men 56

Ph lc 5: Thnh phn ch phm khi kho st t l tinh bt ti u

Trong mi mu ch phm:
Trng lng mi trng cm go chim 20% (132g)
Trng lng tinh bt chim 80% (528g)
Bng 7.14: Thnh phn ch phm khi kho st t l tinh bt ti u
Trng lng bt m Trng lng bt sn
Mu %bt m : % bt sn
(g) (g)
1 0 : 100 0 528
2 25 : 75 132 396
3 33 : 67 176 352
4 50 : 50 264 264
5 67 : 33 352 176
6 75 : 25 396 132
7 100 : 0 528 0
125