HOI NGH! KHOA HQC C O N G NGHE SINH HOC T O A N Q U O C 2013
NANG CAO HOAT TJNH SINH T 6 N G H(7P NISIN COA CHONG LACTOCOCCUS LACTIS SUBSP. LACTIS PD14.1 BANG PHUONG PHAP DAP CfNG BE MAT
Hoa Thi Minh Tii', Tran Thanh Thuy, Phjm Thj Thu PhiFong, Nguyen Kim Thoa, Trin Dinh Min, L6 Thanh Blnh
Vi$n Cdng ngh^ sinh hgc. Vi$n Han lam Khoa hQC vS Cdng ngh$ Vi^t NamT6M TAT
Chiing Lactoeoccus lactis subsp. lactis PDJ4.1 tfing hop nisin ducc phan lap hi siia b6 tuoi 6 Vift Nam c6 khi ning khang nisin o ning aO 6000 lU/ral. Cac thanh ph4n mfli tnr&ng nhir saccaroza, pepton KHjPO^ v i stta gay khi nghien c6utic d^ng riSng re da cho thiy chung c6 inh hudog manh dSn qui trinh sinh t^ng hgp nisin ciia chiing niy. Do vSy, viec nghien cihi toi uu h6a thanh phan m6i tiutag theo phuong phip dap iing be mat sii dung plwn mem Design - Expert 7,1 iK dupc tien hanh nhim nfing cao ho?t tinh sinh tSng hiTp nism ciia chiing. Trong s^ nhung Ihinh phan moi tnrcmg chin§ h^n nhir saccaroza, pepton, KH2PO4 sua gay thi cic bi&i s5 quan tr^ng li pepton v i KH2PO4 c6 inh huong m^nh nhit dSn sinh tong h ^ nisin. Sir dung phuong phap quy ho^ch thvc nghiem lap l^i t^i tam (central conqjosite design (CCD)) \(n kSt qua thu dirge ciia 30 thi nghiem trong mS hinh, kiem tia dp tin cjy va sv thich ung cua m6 hinh voi 3 lin thvc nghi§m l|p I?i. da dira ra phuong trinh bac 2 tim ra m6i tuong quan giira sinh t6ng hgrp nisin vi thinh phin mSi tnrong. Kit qui ho?t tinh nisin cao nhit d^t 7960 lU/ml sau 12 gio nufli cay * nhiet &q 30 "C khuiy 100 v/p trong moi truong t5i iru, cao gip 6 iJn so voi mSi trudng co so. Thanh phin moi truing toi uu bao g5m (gfl): CB5 thu (10), cao men (5), pepton (15), saccaroza (20). CHjCOONa 3HiO (8,3), KH1PO4 (10). amonicitrat (2), MgSp4.7H20 (0.2), MnS04 (0.04), Tween 80 (Iral), sira giy 25 %, pH = 6.
OAT VAN o e
Nisiti \k chit bio quin thi/c phSm c6 nguin g6c sinh hpc ban chit Id m^t protein hay mOt peptit trpng lii'ip'ng phSn til khodng 3,45 kDa bao g«n 34 acid amin (aa), flu^c tfing hiyp tren ribosome cua lodi Ladococcus lactis subsp lactis (Reunanen J,2007). Hi#n nay nisin Id bacleriocln duy nhit flu-pc it chiic Y te The gi*! (WHO) vd co quan quan ly ThuSc va Thg-c phim Hoa ky FDA khuyin cdo si> dgng. Ph?m vi ij-ng dyng cOa nisin khd rOrig. tO cdc sdn phim tuoi sOng nhif sOa, thjt, cd, rau qua ain cac loai thi/c phim ISn men, d6ng hOp, binh mj'-, nif6c u6ng .Si> dung nisin trong bdo quan thu-ong khSng Idm thay doi cdm quan cung nhu- chit lu'png, khflng d i lai du tifpng trong th^rc phim (Anh, 2010;
Reunanen Jetal, 2007; Kwaadstenlet de M et al, 2008; Fernanda Mozzi et al, 2010).
Thdnh phin mfli tnjdng ching hgn nhir nguin cacbon, nitp', cdc dieu ki?n nuISi ciy da c6 dnh hu&ng m?nh den sinh trudng vd t6ng hpp nisin. Vi#c tim ra yiu to ndq c6 Snh hudng m?nh din sinh trying vd sinh tSng top nisin tir d6 t6i uu mai tn/dng nuOi ciy la vi#c lam rit khd khan. Phuong phdp fmyen thing trudc ddy khi ngWfin ci>u dnh hudng cOa thdnh ph^n mdi towng den ho^t tinh nisin thudng xdc flinh riSng r§ hjng ylu to mgt, do do khdng chl mit nhiiu Ihdi gian md cdn cho kit qud khdng tit vd khdng chinh xdc (Chan Li et al, 2002).
Hi$n nay. phuong phdp ddp ung b i mat (Response surface methodology, RSM ) d i gidi quyit dug-c cac m$t han chi cOa cdc phuong phdp cidiin. RSM dupc thg'c hi#n vdi si/trgr giup cua phin mem Design - Expert 7.1, cho ph4p thilt k l cdc thl nghiem, xdy ct^ng md hlnh, nghiSn cdu tdc ddng ding thdi cdc ylu t6 cOng nhu tim ra dupc dieu kl#n tii uu cho m6i myc trSu eg thi hudng tdi (Chan Li et al., 2002 ; VO Hong Son et al., 2009; Wei-Iiang Guo et at, 2010) L lactis subsp lactis cd nhu ciu dinh dudng phdc tap. Diiu nay khdng nhung Idm lang chi phi san suit ma cdn phat sinh nhO'ng khd khdn trong qud trinh thu hii vd linh s^ch sin phim. Cdc mdi trudng khdc nhau thudng dupc su di,ing trong nghidn nhu CM', SM8, M17, M17S va MRS (Chan Li et ai., 2002) Tit ca cdc mdi tn/dng ndy diu tdt cho qua trinh sinh trudng vd trung hda axit lactic nhung chua dupc quan tdm v i khA nang ting nisin bdi trong cdc mdi trudng ndy diu chua nguin nito d^c bidt la ngudn protein hay peptit cao, din din nhO'ng khd khan trong qua trinh tinh sach nisin.
Vdi mgc tiSu xdc dinh dupc thanh phin mfli trudng til uu cho ting hpp nisin cao, \dm tSng nang suit nisin cOng nhu thuin lpi cho qud trinh thu hii va tinh s^ch. Trong nghiSn cCru trudc ddy, chOng tdi da xdc dinh duoc cac ylu t i ridng re dnh hudng din sinh trudng vd sinh nisin cua chdng L lactis subsp lactis PD14.1 bao gim saccaroza, pepton, KH2PO4, BO'S giy (skim milk). Kit qud ndy tuong dong vdi cdng b i trudc d3y cOa mdt s l tdc gia trong nude cung nhu trdn t h i gidi (Hoa e( a/., 2001; Hoa, 2003. Chan Li e( a/., 2002 , Wei-Iiang Guo e( a/., 2010). Cdc ylu te ndy dupc thilt l$p trong md hinh tii uu hda dap irng b l mat (RSM) vdi sg- trp giiip cua phin mem Design - Expert 7.1.
NGUVeN U?U VA PHU-ONG PHAP Chung giing vd mdi trudng
Chiing L lactis subsp lactis PD14.1 sinh nisin phdn Igp tii' sO-a bd tuoi d Vi^t Nam cd khd ndng khdng nisin d ning dd 6000 lU/ml, thudc bd suu t^p giing cita Phdng Cdng nghg V|t lieu sinh hgc, Vidn Cdng nghg sinh hgc, Vi§n Hdn Ldm KHSiCN Vi$t Nam. ChOng kilm i^nh Lactobacillus plantarum JCM 1149 do Bao tdng gidng vi sinh vgt Nhgt Ban (JCM) cung d p , Nhung chOng ndy dupc nudi tren mdi trudng MRS (Man Rogosa Sharpe).
Cdc hda chit dting trong thi rglii^m Id cac hda chit dupc mua tir hang Sigma vd.Merck cd dO hnh khilt cao, sua g^y (skim milk) cd xuit xu- Australia. Nisin ttiuong phim ciia hang Sigma dupc su- dyng d i di/ng dudng chuin.
Xac djnh hogt tinh nisin
Xdc dinh hogt tinh nisin bing don \\ lU (International Unit) theo phuong phap cua Fowler vd cpng si/ 1975 (Fowler ef
a ,1975). Nisin chuin ciia hang Sigma dupc pha b ddi cdc ning dd khdc nhau. X^c ^nh hogt tinh ciia nhu-ng maundy
bing cdch do dudng kinh vimg u-c chi ddi vdi L. plantanim JCM1149, l|p phuong trinh dudng chuin tuong quan giCi-a
ning dd nisin (tinh bing lU/ml, mg) vd dudng kfnh vdng i>c chl Id phuong trinh cd dgng ham s6 mu: Y= 23.105e" " ^ "
- H Q l NOI I! Kl lOA I IQC Ct^Nb IsibHt;: blNI i I IQC TOAN QUOC 2013
b i n g p h i n m i m Excel cOa Microsoft, hg s6 h i i quy R ' = Q.9699. Trorvg cdng thirc n d y x = n - d (mm) n- la d i i m i q - __kinh-vdng-vdJd3uin.^m)fdHd-audng-kinhio-ktToarr(mmj. T : r i o a t t i n n ctiS phdm. Vdi d u d n g cong c h u i n da di/ng,
bang cdch ddi c h i i u se xdc dinh d u p c hoat tinh cac mau c i n t i m . P h i r a ^ g phap quy hogch t h u c nghigm
Xdc djnh y l u t i dnh hudng t d i k h i ndng sinh t r u d n g va sinh t i n g hpp nisin cOa chung L factfs. subsp lactis P D 1 4 J b i n g cdch s i i dgng phuong phap qiiy hogch thyc n g h i | m igp Igi tgi ^ m cho 4 y l u t i : saccaroza. pepton, KHzP04. s u a g i y . S d dyng p h i n m i m Design - Expert" software DX7. M i i y l u td d u p c tien hdnh tgi 5 m d c (-a. - 1 . 0, + 1 , +a) n h u Bdng 1. v d i a =1.25. Quy hogch t h y c nghigm g i m 30 t h i nghigm trong dd cd 6 thi nghiem l | p Igi tgi tdm, vdi 1 hdm mgc ti§u hogt tinh (lU/ml). XO Ij s i li^u t h y c nghigm b i n g p h i n m i m t h i n g kd Design - Expert 7.1 d i phdn tich cdc h f s i h i i quy, b i mgt d i p dng vd tdi u u hda v d i thugt toan hdm mong d p i (Stat - Ease, Inc. 2007)
Bdng 1- GIS trj m i h6a vk cio yeu t i thyv nghiem
Biin
Saccaroza Pepton- KHjPO, S i j a g i y
sd Kihi^u
A B C D
Ekmvj
g/i gfl g/I
%
- 0 7.46 3.73 0 13.73
10 5 1 15
Muc
20 10 ,5 . 20 '
+1 30 15 10 - 25
* a 32.54 16.27 11,14 26.27
K £ T Q U A V A T H A O LUAN T h i l t l$p m d hlnh
Khdo sdt i n h hudng cOa c^c Ipgr mdi trudng: MRS. C M . M17 d i n kha nang sinh t i n g hpp nisin cho thay chung L lactis subsp/actfs PD14.1 cd hogt tfnh cao n h i t (dgt 1350 l U / m l ) trong mdi tnrong MRS chda sacanaza sau I Z g i d n u d t d n h i ^ t dd 30 "C cd t i c dd k h u i y 100 v/p. Theo mdt s i tai li#u da cdng b i . cdc y l u t6: saccanaza, pepton. KFEP04, sOa g i y Snh hudng ro r^ d i n qud trinh sinh t i n g hpp nisin ciia L lactis subsp ISctis.
Mgc dich ci^a qud trinh ndy la tim s y tuong tdc d i n g thdTcua cdc thdnh p h i n m5i t n / d n g sacaroza. pepton. KH2PO4. si>a g i y i n h hudng t d i qud trinh sinh t i n g hpp nisin d i l y a chpn m ^ trudng t i i u u nudi d y chiJng Lactoeoccus lactis subsp.
lactis PD14.1cho hogt dd nisin cao n h i t . Bdng 2. Bdng qui hogch thyc nghiem cila tha^t todn toi uu
STN 1 2 3 4 ' 5
e 7 8
• 9 10 . .. 11 -
12 13 15 16 17 19 20
23
30
Saccaroza (A) 100 10 0 20.0 20 0 20.0 7.5 30 0 10 0 30.0 30 0 20 0 20 0 30.0 30 0 20.0 30.0 20.0 10.0 30.0 20.0
20.0 20.0 20 0
Pepton (B) 15.0 5.0 ' 10.0 10.0 10.0 10.0 15.0 15.0 15.0 5.0 10.0 • 10.0 15.0 10.0 5.0 100 15 0 15.0 16.3 15.0 5.0 5 0 10.0 10.0 5.0 5.0 10.0 3.7 5.0 - 10.0
KHiP04 (C) 1.0 1.0 5.5 11.1 5 5 5.5 1.0 1.0 1.0 10.0
•- - 5.5 5.5 1D.0 5.5 1.0 -0.1 10.0 10.0 5.5 10.0 10.0 10.0 5.5 5.5 1.0 10.0 5 5 5 5 i.e 5.5
Sua g i y (D) 15.0 15.0 20.0 20.0 26.3 20.0 25.0 25.0 15.0 25.0 - . - 20-0-- 20.0 15.0 20.0 25.0 20.0 25 0 15.0 20.0 25.0 25.0 15.0 20.0 20.0 25.0 15.0 13.7 20.0 . 1 5 0 20.0
Ho?l tinh Nisin (lU/r 1920.0 1920.0 2101.0 6117.0 1920.0 1920.0 1920.0 1255.0 1437.0 1920.0 1920.0 960.0 5441.0 960.0 1328.0 2435.0 76S0.0 6730.0 3680.0 7680 0 . , 1920 0 • '
320.0 1920.0 1920.0 960.0 1296.0 960.0 480.0 960.0 1920.0
nl)
K i t q u i thu d u p c trong mfl hinh t h y c nghi$m ( B i n g 2). si> dgng p h i n m i m Design - Expert 7.1, phdn tich h i i quy vd Wim tra s y cd nghia cua cdc hd sd, tinh thlch Ong cOa m d hlnh (Bdng 3). cho t h i y C h u i n F ciia md hinh Id 99.16 h o a n b i n cd * nghTa t h i n g k§ v d i dd Un c$y 99.99 % (p < 0,0001). Vdi cdc gid tri P< 0.05 cho b i l t cdc h | s i h i i quy cd nghia.
C h u i n F • cho s y khdng t u o n g thfch" Yht Id 0.28 (p = 0.9584). h$ s o t u o n g quan h i i quy {R2) ciia mS hlnh bdng 0.9893 cho thdy mfl hlnh hodn todn t u o n g thlch vd m d phdng ddng v d i ttiyc nghigm.
HOI NGHI KHOA HOC C O N G NGHg SINH HQC T O A N Q U 6 C 2013
Bang 3. K i t qua phdn tich phuong sai, hoi quy ANOVA cda mo hinh t6l uu nisin ciia chung L. lactis subsp lacHs PD14.1
Ngu6n goc M6 Wnti A-SUCROSE B-PEP C-KH2P04 D-SKIMMILK AB AC AD BC BD . CD A2 B2 C2 D^
Khdngtutmg thich
Phuong sai 1.234E+008 7.859E+005 3.936E*007 3.505E+007 1.783E+006 13282.56 2.151E+005 1.405E+006 2.681 E+007 1.182E-HX)5 2 392E*006 6.008E*005 52694 52 1.352E+007 6.0088*1X5 4.79aE*005
Ctiuin F 99.16 8.84 442.94 394.41 20.07 0.15 2.42 15.81 301,68 1.33 26.92
6.76 0.69 152.72
6,76 0,28
Gia tri P Kh5 n&ig > F
<0OO01 0 0095
< 0.0001
< 0.0001 0.0004 0.7045 0.1406 0.0012
< 0.0001 0.2669 0.0001 0.0201 0 4532
< 0.0001 0.0201 0.9584
L i n lupt xet dnh h u d n g cda t d n g y l u t i (khi cdc y l u t i khdc g i u d mire trung lam) doi v d i sinh t i n g hop nisin cho thay, cdc y i u t i cd anh h u d n g manh n h i t Id: (B) Pepton vd sau dd d i n (C) KH2P04, sau dd den (D) s u a gay, n g u i n cacbon A (saccaroza) cd dnh h u d n g r i t it h i u n h u khflng ddng k l d i n sinh tong hpp nisin (Hinh 1) Dieu do d u p c mmh hoa ro hon khi nghiSn cdu b l m j t ddp i m g cQa hdm Y(ht) Hinh 1.
Hlnh 1. Anh huong cua cac yeu to trong mo hinh t o u u s i n h Nisin ciJa chiing L /ac(/s subsp lactis PD14
Hinh 2. B& mdt Odp irng cua timg d p cac yeu to dnh'huo'ng d i n sinh tong h y p nisin ciia chiing Uactls subsp lactis PD14.1:
Saccaroza - KH1PO4 (A); pepton - KHzPO. (B), pepton - sua Qay(C) vd KH1PO4 - sira giy(D).
Qua Hinh 2, nhJn t h i y smh t i n g hgp nisin tdng d i n t u m i u xanh d | m d i n mdu vang vd cao n h i t d vimg mdu dd s ^ m . K h i o sdt tdc ddng ddng thdi cdc yeu l i d i n qud trinh sinh t i n g h p p Nisin cho t^,iy, n i n g dd pepton - KH2PO4 c 6 tac
" H O I N G H I K H O A H Q C C d N G N G H E S I N H H O C T O A N Q U O C 2 0 1 3
Annrj rAl l d n t d j Qud trinh sinh t i n g hpp nisin, sau dd la s i j a gay, cdn-ndng dp saccaroza-Jt anh hucmg h o n . Khi-ddnli gid l(|[ hpp tac dflng ddng thdi cua~haryeu td saccaroza - pepton, saccaroza - KH2PU4, saccaroza - skim milk, pepton - KH2P04, pepton - sua g i y , KH2P04 - sua g i y cho &iay ca 6 nhdm tren d i u cd anh hudng it hay nhieu d i n sinh tdng hpp nisin cOa chiing L lactis subsp lacts P D 1 4 1 . Tuy nhiSn, chi 2 y l u td pepton - KHaP04 (BC) trong md hlnh (Hlnh 2B) cd anh t i u d n g Idn n h i t Sau khi xet cdc y l u t i dnh h u o n g ciia tOng c§p cdc y l u t i anh h u d n g , d y a vdo k i t qua cdc y i u l i trfin bdng quy hogch thyc nghigm vdi s y ho trp cua phan mem DX7.1 t h i l t I|lp d u p c p h u o n g Irlnh t i i u u cho sinh t i n g h p p nisin n h u sau:
Y hogt llnh(IU/ml) = 780.27749 - 4.82072 ' A- 184.29464* B - 1 3 7 5 . 6 5 9 6 r C + 315.84030 * D+ 0.57625 * A " B -2.57639
• A * C + 5.92625 * A * D+ 57.53056 * B " C + 3.43750 * B * D+ 17.18611' C * D- 3.13892" A2 +3.71850 " B2 + 73.53003 ' C2 -12.55569 ' D2.
Tir phuong trinh t i i uu ndy da xdc l^ip d u p c thanh phan mdi t r u d n g cita t i t ca cdc y l u t i dnh h u d n g d i n hogt tfnh nisin theo tinh todn ly thuyit g i m 48 phucmg dn (dng 4). Trong dd phuong dn sinh t i n g hpp nisin cao n h i t thuQc v i phucmg dn 12, igi dd nfing dd cCia cdc c h i t sacaroza 20 g/1, pepton 15 g/I, KHzP04 10 g/I vd sO-a g i y 25%. Hogt Hnh dgt d u p c theo m d hlnh t i i uu Id 7960 iU/ml, tang g i p 6 l i n khi sir .dung mdi tiudng MRS saccaroza blnh thudng khdng qua m d hlnh t i i u u .
Bang 4. C d & p h u o n g an m d p h d n g hoat t i n h nisin t h e o ly t h u y i t t h e o m d hlnh t i l u u
TT Sucrose Pepton KH;PO. SOa g i y Sacrose Pepton KH2PO4
g^v 10
5.5 10 1 10 1 10 10 1 10 10 10 1 1 1 1 1 8.506 9 9694 88489 6 7429 5.284 7.9786 2.9449
25 20 15 15 15
• 15 25 15 25 15 25 25 15 15 25 25 . 25 21445 16.935 16 828 15.353 21.081 17.028 24.469
2090.254 1854.384 1471.016 1071.777 5525.997 1944.664 1988.176 183.6877 1360.903 6698.076 7674.235 7960.064 1895.355 1236.336 1329.515 967.8433 1837.824
• 1314 307 1297 503 4291.815 3005.207 1401.493 1674.95 1131.12
19 924 18.45 22194 28.482 13.228 21794 16,386 28.248 12.642 14.878 25.368 11.944 27.568 21.416 11.674 27.996 21.808 12.64 28.524 28 59 26.094 19-844 12-272 12.272
9.402 12.087 9.37 5.485 14.348 9.555 ' 9.491 5.332 14.782 9.326 7.865 1448 13.171 7.405 7,242 11867 10.36 8.492 14 375
10.76 6.822 7 224 1127 11.27
8 9488 1 6327 9.3844 6.6799 2.3977 4.9294 9.5401 4.8628 8.9641 5.8276 8.7382 4.4029 4 2661 1.8901 8.839 3.9268 4 3057 3.2185 9 8884 8.4106 8 0866 2.0656 6 7474 6.7474
15 72 16.287 21.589 16.658 18,091 16 551 15.163 24.092 18 624 17 081 15.839 21.805 17.64 20.056 16.483- 20.434 20,869 . 15.879 15.999 17.463 20.702 24.428 20 579 20 579
2745.143- 1802.744 3918 064 162,1824 2080 179 1192.78 3264.119 345 9394 6059 899 1578.001 1550.65 2582.4 1726 248 1522.94 2D56.72 1656.797 1674.312 1165.237 5535,899 2735.137 1571 087 1188.466 2813.03 2813.03 18 28 878
19 26 326 • 20 11.05 1 21 25 244 1 22 16186 23 18.41 24 15.526 •
Cuoi cirng tim d u p c vimg t i i u u cOa t i t cd cdc yeu t i dnh hCFong, g i m 48 phuong dn, (Bdng 4) trong dd phuong dn tdi uu n h i t Id n i n g dd saccaroza 20 g/I. Pepton 15 g/I, KHzPO* lOg/l va su-a g i y 25%. Hogt tinh dgt d u p c theo md hinh tdi uuld-7960 lU/ml. V3y, mdi t r u d n g tdi u u d l nuQi c i y sinh t i n g hpp Nisin cd thdnh p h i n n h u s a u ( g / l ) : C a o thjt (10), cao men-(5), pepton (15), sacanaza (20), CH3COONa.3H20 (8,3). K H 2 P 0 4 (10), amonicitrat (2), M g S 0 4 . 7 H 2 0 (0.2), MnS04 (0.04), Tween BO (1ml). s&a g i y 2 5 % . pH = 6.8. B i sung n u d e c a t c h o dil 1 lit. ^^ _
K i l m t r a t i n h t u o n g t h l c h c i i a m d hinh v a t h y c ng'hl^m
D i k i l m tra l^i tinh dOng d i n cOa phuong trinh t i i u u , t i l n hdnh thf nghiem kiem chirng tgi 5 d i l m toi uu thu d u p c b i t kl (cdc d i i m t i i u u ndy cd cdc thflng s i cOa cdc bien .vd k i t qua v i hoat d d thu d u p c theo I'? t h u y i t ) K i t qua d i / p c t h l hidn trong Bang 5.
Bdng 5. K i t qud k i l m chirng s y t u o n g thich cOa m6 hinh theo ly Oiuyet v o i k i t qud thyc nghiem S i T T
5
Saccaroza(g/l) 10 12.6
20 28.5
30
pBplon{gfl) 15 14.8 14.3- 15
KHjPO.!
10 - 9 10 10 10
m
sua gay(%) 15 18.6 25 15.99925
Hoat do ly ^ u y l t (IU/ml) 6698.076 6059.899 7960.642 5535-899 7674-235
Hogt Jo thyc E4(IUftnl) 6680± 30 6040 t i o 7960 ±10 5500 ±10 7650± 20 Ghi chif t sal s6 giOa cAc thf nghl$m, 's6 thl nghi$m thi/c hi^n la 3.
HQI NGH1K^^A HOC CONG H&^^^ HQC TOAN QlH^^jm
K i t qua Bang 5 cho I h l y , t r o n g ^ thi i ^ i # m s y sai k h ^ h o ^ g ^ ^ ^ , ^ ^ , ^ ^ 1 , v i f c ^ , , v # . „ ^ ^ chdng tuono ddi nho (< 50 IU/ml). N h u v | y , k a qua t o i u u cua m f l h l n h ^ c h i f ^ j ^ ^ ^ ' J ^ j . "^^m R I ^ . ! . v a •„
K E T L U A N
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ENHANCE OF NISIN BIOSYNTHESIS OF U^OTOCOCCUS LAQfl^ SUBSP I LACTIS PD14.1 BY RESPONSE SURFACi METHODOLOGY " I
Hoa T i l l M i n h Tu", T r a n T h a n h T h u y , P h a m T h i T h u P h u w i g , NgMy«r) K i m T h o a , T r a n O m - j\ji5„^ j-S Thanh B i i * ^ InsOtute of Biotechnology, Vietnam Academy of Sicence and T^^f^^g^/ • •
S U > I > L \ R \
In Ais Study, optinial medium components for enhancing nisin pre^ti^m gf iactococais ladti Siift^p ft,^^ rtdl4 I st carried oul by Re^»nse Surface Methodology using the siatistieal gfRware paclcage Design * h^tM 1 I (SiatO- Minneapolis. USA) Among those component such as saccharose, psfl^e^ JiH-PO* and skim mitfe ^ Vrtost ilttnlfion^
influencing nisin production were peptone and KHjPO^. A 30-ruH i^}ri\ \wnposite design (CC1&| ^i^^ WWlwed io-W describing the relationship between nisin production and themediurft WK^l»m, a mathematical m ^ ^ \ y S l r t b y the "
second - order polynomial equation lo fit data in RSM,of CCD. Sut^:a^,^ily. the reliability of thQ 9il^k SmUrt rtcrf, prediction were checked by verification experiment in triplicate.The ^ ^ ^ yfyto ptoduction was yicll^%^ ^ ^ ^^j^l ^ u l inoculation in optinial medium which was 6 limes higher than that in b ^ W«ii\im. The optimal DWKe^if, Votvkl^tra rfC^""^
extract (10). yeast extract (5). peptone(15), saccharose (20), CHjCOQ?SiJrt.018,3), KHiPO,{10^ A W « * ^ t r t l W
(02),MnSO4(0.04),Twcen80(lmI),Skimmilk25%. '^nmc™K^i^w Keywords: Biosynthesis, enhance, Lactoeoccus lactis subsp. lactis, ni^i^ f^f^.Y.sc surfece melhodoli^
*Autfio(-ftircofresporcfence;Tel 84-437567103, EmaJ: hoathiminntu@g?ft^^ift.