Luận án Nghiên cứu quá trình thủy phân protein cá bằng enzyme protease từ B.subtilis S5

.49,3.50,3.51,3.52. Bang3.24.Anh hu'dngeuaty l~CPE de'nsrithayd6itr~ngthaiearnquaD euacaemiluthuyphiloeaeo'm Thai M§:u1 M§:u2 M§:u3 M§:u4 M§:u5 glan (gio) 0 H6n h<;Jpthitea H6n h<;Jpthitea H6n h<;Jpthitea H6n h<;Jpthitea H6n h<;Jpthttea mall tdng hoi mh tdng hoi mall tr~nghoi mall tr~nghoi mall tr~nghoi xam, eo mlii xam, eomlii xam, eo mlii xam, eo mlii xam, eo mlii tanhtv nhien, tanhtv nhien, tanhtv nhien, tanhtv nhien, tanhtv nhien, thitcadi;iedeo, thitcadi;iedeo, thiteadi;iedeo, thiteadi;iedeo, thitead(tedeo, hoil6nnh6n. hoil6nnh6n. hoil6nnh6n. hoil6nnh6n. hoil6nnh6n. 2 H6n h<;Jpthitea I H6n h<;Jpthitea H6n h<;Jpthitca H6n h<;Jpthitea H6n h<;Jpthitca mh tr~nghoi hoi fiat, mall fiat, mall tr~ng fiat nhuy~nhoi long nhuyen, xam, mlii I tdng hoi xam,I hoi xam, mlii I long, mh Imall hoi xam, tanh,thitcara'tI eonmliitanh. di;ie. hoitanh. tdng hoi xam,Ima'tmlii tanh. ma'tmlii tanh. 4 H6nh<;Jpthitca H6nh<;Jpthitca H6nh<;Jpthitea H6nh<;Jpthitca H6nh<;Jpthitca hoi nhiio,mall fiat, mall hoi fiatnhuy~nhoi long nhuy~n,d~ngra'tlong, tr~ngxam,mlii xam, ma'tmlii long, mall hoi mall hoi xam, mh hoi xam, bot tanh. tanh. xam, ma'tmlii Ima'tmlii tanh. tanh. ma'tmlii tanh. 6 H6n h<;Jpthitea H6n h<;Jpthitea H6n h<;Jpthitea H6n h<;Jpthitea H6n h<;Jpthttca hoi fiat, mall fiat nhuy~n, long nhuy~n, d~ngra't long, long nhu'nu'ae, tdng xam,ma't mall hoi xam, mh hoi xam, mall hoi xam, hoi philo tang, mlii tanh. eo mlii di;ieI eo mlii d(teI eo mlii di;ieI mall hoi xam, mlii d(tetru'ng.tru'ong. tru'ong. tru'ng. 110 H6n h<;1pthit ca oat, d~e,m~u s~m,mlii Mi nhy,hdieobQt khi. 8 H6n h<;1pthitea d<;lng long, m~uxam, mlii H6n h<;1pthit ea rtt long, m~u xam, mlii hdi Mi, hdieobQt khi. hdi hoi,hdi eo bQtkhi. H6n h<;1pthitca d<;lnglong nhu nude,hdi philn t~ng, m~u xam, mlii hdi hoi, hdi eo bQt khi. H6n h<;1pth~tea long nhu nude, phan thg m<;lnh, mh xam, mlii hdi hoi, hdi eo bQt khi thoatfa. H6nh<;1pthitea H6nh<;1pthitca H6nh<;1pthitca H6nh<;1pthitea H6nh<;1pthitea hdinhuyen,eo d<;lngrtt long, lOngnhunude, longnhunude, longnhunude, m~uxam,mlii m~uxam,mlii hdi philnt~ng, philn t~ng philn thg rtt hoi,eobQtkhi. Mi, eobQtkhi. m~uxam,mlii m<;lnh, mh m<;lnh, m~u hoi,eobQtkhi. xam, mlii Mi, xam, mlii hoi, 10 H6nh<;1pthitea H6nh<;1pthitea H6nh<;1pthitca eo d<;lnglong, lOngnhunude, longnhunuac, eo m~uxam, hdi philnt~ng, philn t~ng, mlii hdi th6i, m~uxam,mlii m~uxam,mlii eo bQt khi th6i, bQt khi th6i,nhi~ubQt thoatranhi~u. thoatranhi~u. khithoatra, . 12I c:: «:$ ;; ~ 200 0 :::I ...c:: '«<:$ c:: "0 '§ ~150 0 .D I-< ""'; 0.. "0 t)J) >- .§. ~100 .E ~ S '-' ,«:$ 50 ::r:: 250 eobQtkhi. H6n h<;1pth~tca long nhu nude, phan t~ng ra't m<;lnh, m~u xam, mlii th6i, bQtkhi thoatra nhi~u. y =0,2196x5-4,8587x4+41,525x3- 173,27x2+371,3x- 134,86 R2 =0,9998. eobQtkhi. H6n h<;1pth~tea long nhu nude, phan dog rtt m<;lnh, m~u xam, mlii th6i, bQtkhi thoatra nhi~u. y =-0,0852x4+ 1,3886x3- 6,8375x2+ 19,189x+86,229 R2 =0,9996 8 10 12 Thai gian thuy phan (gia) 0 MlIu 4 A MlIu 5 I 0 0 2 4 6 ? ffinh3.49.Anh hu'dngcuaty l~CPE de'nham1u'Q'ngprotein boa tan cua caemduthuyphan ca com (HamluQngproteinhoatanband~ula 34,13mg/gmallvaduQCtrlnhbay d bang3.1- ph\!h,lc3) :I(MlIu1 -Mhz . MlIu3 111 450 400 y = -O,145Ix4 +4,1462x'-43,746x2+ 224,91x - 86,186 R2 =0,9989 ~ 350-0 ::J- oR '~ 300 v c: ~ ro 25000.D c: 0"" S' '~200 S S; 150 'ro ~ :r:: '--" 100 50 0 0 y =O,2473x4-3,7548x'+ 19,146x2- 16,452x+ 100,6 R2 =0,9928 2 4 6 8 10 12 Thai gianthuyphan(gia) . Mh 2 . Miiu3 0 Mh 4 .. Miiu 5 II :t:Miiu1 ? Hlnh3.50.Anh hu'dngeuatyl~CPE de'nhamlu'gngpeptideuacae mfiuthuyphaneacom (Hamlucjngpeptidband<1ulaO,364mg/gm§:uvaducjctrinhbay (j bang3.2- ph\ll\lc 3) 300 y = 0,0242x4 - 1,6712x3+ 14,042x2+5,43x+82,871 R2=0,9986c:'s 250 ro :J , '<ro ';;: '0 200ro c: '0 ro ,<;::.D c: ;-6 15° 00 ;;. c: 0 S' '" 100 - ~8 '--",ro :r:: 50 y =0,0178x4- 0,7543x3+8,1958x2- 12,503x+ 105,06 R2=0,9999 0 0 2 4 6 8 10 12 Thai gian thuy phan (giO) :t:Miiu1 . Miiu2 . Miiu3 0 Miiu4 .. Miiu5 I Hlnh3.51.Anh hu'dngeuaty l~CPE de'nhamlu'gngNaaeuacaemfiu thuyphaneacom (HamlucjngNaaband<1ula O,296mg/gm§:uvaducjctrinhbay (j bang3.3- ph\ll\lc 3) 750 600 '" ~ ::r:: ;::Iz '<~ B ~450 '2 ] OJ) ,- I:: "0 '0' ;> ~ 0 300'" S 109,~ ,",' ::r:: '-' 150 0 112 y = 1,5591x4-22,912x3+112,87x2- 117,35x+126,59 R2=0,9995 y =0,4443x4-7,9563x3+50,746x2- 71,218x+128,11 R2= 1 0 4 8 10 12 Thai gian thuy phan (gia) 0 Mh 4 A M§:u5 I 2 6 )II:M§:u 1 -Mhz . M§:u3 Hlnh3.52.Anh hu'dngcuaty l~CPE de'nhamIu'QngNNH3 cua cacrn~u thuy phanca cdrn (HamluQngNNH3band~ula 0,023mg/gm~uvaduQCtrinhbay (5bang3.4- phl,lIl,lC3) Tli cacke'tquanghienCUu(5lIen chopheprutra mQts6nh~nxet: *V~tr~ngthai carnquaD:cangtangty l~che'ph§mproteaseB.subtilis55 ~ sungvaocacm~uthuyphanthlmucdQthuyphancangm<;lnhmethe'hi~nla iu cangnhanhchuye'nsangtr<;lngthai long.Tuy v~y,do thuyphan(5nhi~tdQ u'ongl<;likhongb6 sungmu6iande'uc che'visinhv~tgayth6inentu giai do<;ln giOthuyphantr(5di cac m~ud€u co hi~ntuQngca doi mu6ibie'uhi~nla cac iu thuyphanb~td~ucob9tkhi vabi n~ngmlii. Nhu v~ytoi thai die'mnaynen j sungthemmu6ianvam~u3(b6sung0,2%CPE) (5thaidie'm6 giGthuyphan I tr<;lngthaicamquailt6t. * V~rn~tboa hQc: Tli ke'tquatrinhbay(5cachlnh3.49,3.50va 3.51chotha'ytheothaigian uyphancacm~u2, 3,4 va 5 d€u co hamluQngproteinhoatan,hamluQng 113 peptidva hamlU<;5ngNaat~othanhtangnhanhhan sovoi m~u1.d thaidi~m12 giGthuyphancacm~u1,2, 3,4 va5 cohamlU<;5ngproteinhoatant~othanhtang tu'dngungsovoiband~ula: 157,1%,177,7%,220,1%,234,5%va242,7%;ham lu'Qngpeptidt~othanhtangtuangungso voi band~ula: 228,8%,339,1%, 401,2%,413,4%va 417,5%;hamlu<;5ngNaatangtuangung so voi band~ula: 203,1%, 250,1%, 260,6%;288,3%va 294,1%. Nhu v~ym~u3 (b6 sung0,2% CPE)co ham lu<;5ngproteinhoa tan, ham lu<;5ngpeptidva ham lu<;5ngNaat~o thanhcaohannhi~uso voi m~u1 va 2 nhungtha'phanme)tchtitsovoi m~u4 va 5.Trangkhi do, m~u3 l~ico hamlU<;5ngNNH3t~othanhtha'phan kha nhi~uso vdim~u4 va5 nhungcaohankhongdangk~sovoi m~u1va 2 (hinh3.52). Titcacphantick iJ trenchothaymt1u3 (b6sung0,2%che'phdmprotease B.subtilis55)cotrc;mgthaicamquant5tvahamlu(1ngproteinhoatan,peptid,Nuu t(lOthankkhacaocanhamlu(1ngNNH3t(LOthankla co thi chapnh(indu(1c.Vzv(iy tyl~proteaseCPE du(1ch9ndi b6sungvaoquatrinhthuyphancd carnla 0,2%. 3.4.2.2.Xac dinhnhi~tdQthichh«1pchoquatrinh thuy phanca cdm f)~chQnnhi~tde)thichh<;5pcho quatrinhthuyphanca comb~ngprotease B.subtilis55, tie'nhanh 6 m~uthi nghi~m:m~u1: m~ud6i chungthuyphand nhi~tde)thuang,m~u2: m~ucob6sungproteasethuyphand nhi~tde)thuang; m~u3: m~ud6i chungthuyphand nhi~tde)50°C,m~u4: m~uco b6 sung proteasethuyphand nhi~tde)50DC;m~u5:m~ud6ichungthuyphand nhi~tde) 50°Ctrong2 giad~usaudotie'nhanhd nhi~tde)phong,m~u6:m~ucob6sung proteasethuyphand nhi~tde)50°Ctrong2 giad~usaudotie'nhanhd nhi~tde) phong.Cacm~uthinghi~md~ud~usud\lng5kgcacomxay,b6sung0,2%CPE vathuyphand pH tv nhiencuaca.Saudola'ym~utheothaigianthuyphand~ danhgiacamquailvaphantichhoahQc,ke'tquadu<;5ctrinhbayd bang3.25va cachinh3.53,3.54,3.55,3.56. 114 Bang3.25.Anh hu'dngcuanhi~tdQde'nsrithayd6itr~ngthaiearnquaD cuacaerniluthuyphancacOmb~ngproteaseB.subtilis85 ThC1i Mhl Mh2 M~u3 M~u4 M~u5 M&u6 glan (giC1) ;0 H6n hQpth~tI H6nhQpth~tI H6nhQpth~tI H6nhQpth~tI H6nhQpth~tI H6nhQpth~t ca tr~ng mh l ea hdi tdng mffu l ea hdi tdng mh l ca hdi tdng mh l ca hdi tr~ng mall I ca mffu hdi I trang hdi xam,comlii I xam,comliiIxam,comlii I xam,comlii I xam,comliiI xam,comlii tanh tt! tanh tt! nhien, th~t nhien, thit ca d~cdeo, ca d~cdeo, hdi 16n hdi 16n tanh tt! tanh tt! tanh tt! tanh tt! nhien, thit nhien, th~t nhien, thit nhien, th~t ca d~cMo, ca d~cdeo, ca d~cdeo, ca d~cdeo, hdi 16n hdi 16n hdi 16n hdi 16n nh6n. nh6n. nh6n. nh6n. nh6n. nh6n. 2 H6nhQpthit H6nhQpth~tH6nhQpthit H6nhQpthit H6nhQpthit H6nhQpthit ca mffu ca fiat ca mffuhdi ca fiat ca mh hdi ca fiat tr~ng hdi nhuy~nhdi xam, mlii nhuy~nhdi xam, mlii nhuy~nhdi xam, mlii long, mffu tanh,thitca long, mffu tanh,thitca long, mffu tanh,thitca tr~ng hdi nitd~c. hdi xam, riftd~c. hdi xam, riftd~c. xam, milt milt mlii milt mlii mlii tanh. tanh,comlii cahilp. tanh,comlii cahilp. 4 H6nhQpthit H6nhQpthit H6nhQpthit H6nhQpthit H6nhQpthit H6nhQpthit cahdinhao, ca long ca mh hdi ca d~lllgca mffuhdi ca d<;lng mffu hdi nhuy~n, xam, mlii dich long, xam, mlii dich long, xam, mlii mh hdi tanh,thitca hdi phan tanh,th~tca mffu hdi bottanh. xam, milt hdi fiat tffng, mffu hdi fiat xam,comlii mlii tanh. nhuy~n. hdi xam,co nhuy~n. d~ctning. mlii d~c tning. 6 H6nhQpthit H6nhQpthit H6nhQpthit H6nhQpthit H6nhQpthit H6nhQpthit ca hdi fiat, ca d<;lngrift ca fiat ca d<;lngca fiat ca d<;lng mffu hdi long, mh nhuy~n, dich rift nhuy~n, dich long, xam, milt hdi xam,co mffu hdi long, phan mffu hdi hdi phan mliitanh. mlii d~c xam,comlii thg, mffu xam,comlii tffng, mh tning. d~ctning. hdi xam,co d~ctning. hdi xam,co mlii d~c mlii d~c tning tning H6n h<;fpthit H6n h<;fpthit ca mH, d~e, ea d<;lng mh sftm, long nhu mlli hoi nude, hdi nh~, hdi co phan t~ng, btkhi. mh xam, mlli hdi hoi, hdi co bt khi. H6n h<;fpthit ea long nhu nude, philn t~ng m<;lnh, mall xam, mlli h6i, hdi co btkhi. 8 10 H6n h<;fpthit ca hdi nhuy~n,co mh xam, mlli h6i,co bt khi thoatfa. 12 H6n h<;fpthit ea co d<;lng long, co m~u xam, mlli hdi th6i, co bt khi thoatra nhi~u. t:: «J..... 500 H6n h<;fpthit ea d<;lng dieh ra't long, philn thg m<;lnh, hdi n~ng mlIi, hdi co bt khi thoatfa. H6n h<;fpthit H6n h<;fpthit H6nh<;fpthit ea d<;lngea long ea d<;lng dieh ra't nhuy~n, dieh ra't long, phan mh xam, long, phan thg m<;lnh,co mlli h6i, t~ng m<;lnh, mlli Mi, co co btkhi mlli h6i, co bt khi thoatfa. bt khi thoatfa. thoatfa. H6n h<;fpthit H6n h<;fpthit H6n h<;fpthit H6n h<;fpthit H6nh<;fpthit ea longnhu .ea d<;lngea d<;lngea d<;lngca d<;lng nude, phan dieh long, dieh ra't dieh long, dieh ra't t~ng ra't mh xam, long, philn m~u xam, long, phan ffi<;lnh,m~u mlli h6i thg m<;lnh,mlli Mi t~ng m<;lnh, xam, mlli th6i,btkhi mlli h6i th6i, btkhi mlli Mi th6i, btkhi thoat ra th6i,btkhi thoat ra th6i, btkhi thoat ra nhi~u. thoat ra nhi~u. thoat ra nhi~u. nhi~u. nhi~u. 115 H6n h<;fpthit ea long 'nhuyen, m~u xam, hdi n~ng filIi, hdi co bt khi thoatfa. H6n h<;fpthit H6n h<;fpthit ca long ca d<;lng nhuy~n, dieh long, mh hdi philn t~ng, xam, hdi hdi n~ng n~ng mlIi, filIi, hdi co hdi co btbt khi khi thoatfa. thoatfa. H6n h<;fpthit ea long nhuyen, m~u xam, mlli h6i, co bt khi thoatfa. '~ :3 400 ..c:: '«<J t:: "0 t::2 «J 300 0 .D ...... 0.. "0 I:>J);> 200 t:: 0 :g. [/)- ~ 100E '-" ,«J :I: 0 I .M~u1 y=O,1591x4-2,0232x3+8,0333x2+49,273x+43,986 R2=0,9993 y =-O,0852x4+ 1,3886x3-6,8375x2+ 19,189x+86,229 R2=0,9996 0 ? mnh 3.53.Anh hudngcua nhi~tdQde'nhamluqng protein boa tan cua cae mftuthuy philo ca cdmbftngproteaseB.subtilis55 (HamluQngproteinboatanband§u la 34,14mg/gm~uvaduQctrlnhbay d bang3.5- ph1;ll1;lc3) 2 4 6 8 10 12 Thai gian thuyphiln (giO) Mu 2 8Mh3 0 Mu 4 AMh5 6. Mu 6 I 116 1000 800 "0 :3 'R '~ &§600 bO.D d "... ~' '~ 400S 55 ,~ ~ ::r: '-" 200 y=-I,O364x4+9,1152x3- O,5X2+38,72x+58,314 R2=0,9965 0 4 ~] , y=O,2473x- 3,7548x'+19,146x-- 16,452x+100,6 R2_O992 0 I +Mhl mnh3.54.Anh hu'dngcuanhi~tdQde'nhamlu'qngpeptidcuacaeman thuyphilocacdmbAngproteaseB.subtilis55 (HamluQngpeptidbandfiulaO,366mg/gm~uvaduQctrlnhbay 0 bang3.6- phl;}ll;}c3) 700 d 's 600 ~ :3 '(~ '~ "0 500~ d '0 ~.D '0 ~B 400 bO ;;. -6, 55 300':3- ~ s '-" 200,~ ::r: y =-I,161x4+14,945x]- 55,904x2+ 148,02x- 4,7714 R2=O,9991 0 y =O,Ol78x4- O,7543x]+8,l958x2- 12,503x+ 105,06 R2=0,9999 100 0 2 4 6 8 10 12 Thai gianthuyphiln(gio) ~Mftu5 I!.Mftu6 II +Mftu1 Mh2 . Mftu3 0 Mftu4 mnh3.55.Anh hu'dngcuanhi~tdQtOihamlu'qngNaacuacaeman thuyphilocacdmbAngproteaseB.subtilisS5 (HamluQngNaabandfiula O,295mg/gm~uvaduQCtrlnhbay 0 bang3.7- phl;}ll;}c3) 2 4 6 8 10 12 Thai gianthuyphiln(gia) Mh2 . Mftu3 0 Mftu4 Mftu5 I!.M§u6 I 117 700 600 y=1,6489x4-2 ,223x3+ 119,36x2- 136,61x+ 144,37 R2=0,9934:£'§' 500 Z '~ :g c 'c 15 400 bI) .,..., C '"0 '0- ~ .E ~ 300 S ~,<':I '-' ::r: 200 y=0,4443x4-7,9563x3+50,746x2- 71,218x+ 128,II R2= I 100 0 0 2 4 6 8 10 12 Thdi gian thuy phan (gid) I . M~u1 ~ M~u2 8Mh3 0 M~u 4 &Mhs l!.M~U61 Hinh 3.56.Anh hu'dngciia nhi~td(}de'nham lu'qngNNH3ciia cac rnfiu thiiy phan ca cOmbiingproteaseB.subtilis55 (HamluQngNNH3bandftula O,O22mg/gm~uva duQetrlnhbay d bang3.8- phl,lll,le3) Tucaek€t quanghieneUudtrenehophepnitramQts6nh~nxetnhusau: * V~tr~ngthai carnquaD:caem~uthuyphancob6sungenzymed€u co illU'cdQehuyentr(;lngthaitud~esanglongnhanhhdnm~ud6ichung.M~u4va6 comuedQthayd6itr(;lngthaitud~esanglongnhanhhdntfiteacaem~uconl(;li. TllYv~y,doquatrlnhthuyphankhongb6sungmu6iandeueeh€ gayth6inen tu8giothuyphantrddicaem~uthuyphancohi~ntuQngeadoimu6ituelahi~n tuQngm~uthuyphancobieuhi~ncobQtkhi vabi n~ngfilii. ChinhVI v~y,toi thaidiem6gioeftnb6sungthemmu6ianvaocaem~uthuyphan. * V~rn~'tboahQc: Tucaek€t quatrlnhbaytrongcaehlnh3.53,3.54va3.55ehothfiycaem~u thinghi~mcob6sungproteaseB.subtilisS5dli thuyphand nhi~tdQnaod€u co 118 hamlu'<;fngproteinboatan,hamlu'<;fngpeptid,hamlu'<;fngNaat':!othanhtangtheo thaigianvatangcaDhonsovoicacm~ud6ichungtu'ongling.d thai di€m sau 12giathuyphan,cacm~u1,2, 3,4, 5 va 6 co hamlu'<;fngproteinboatant':!o thanhtangtu'ongung so voi bandfiula: 157,1%, 234,5%,240,3%,381,3%, 238,7%va470,5%;hamhi<;fngpeptidt':!othanhtangtu'ongungsovoibandfiula: 228,8%,413,4%,372,4%,843,4%,397,8%va 948,6%;hamlU<;fngNaat':!othanh tangWongling so voi bandfiula: 203,1%,288,3%,272,1%,632,4%,260,4%va 569,9%.Nhu v~yhai m~u4 va 6 luan co hamlU<;fngproteinboatan,hamlU<;fng peptid,hamlu<;fngNaat':!othanhcaDhoncacm~u1,2,3 va5. Ngoai fa, m~u4 co hamlu'<;fngproteinboatanvahamlU<;fngpeptidt':!othanhcaDhonmQtchlitsovoi m~u6 nhungl,:!icohamlU<;fngNaat':!othanhthc1phonmQtit sovoi m~u6. Khi phantichhamlU<;fngNNH3t':!othanhtheothaigianthuyphan(hlnh3.56) chothc1ycac m~uco b6 sungproteaseB.subtilis55 d~uco hamlU<;fngNNH3t':!o thanhcaDhonmQtchlitit sovoim~ud6ichungkhangcoenzyme.K€t quanay coth€ giaithichdoquatrlnhthuyphanchuab6sungmu6iand€ lic ch€ vi sinh v~tgayth6inencacm~ucoquatrlnhthuyphanm,:!nhmeclingco hamlU<;fng NNH3t':!othanhcaDhonchlitit.Ngoaifa,hamlU<;fngNNH3t':!othanhd m~u6caD hdnmQtchlitsovoi m~u4. Tit nhilngphdn tick (j tren cho the/ykhi thityphdn ca C{fmbdngprotease B.subtilis55 (j nhi~tdq 50°Cho(ic(j nhi~tdq50°Ctrong2 giGddusaudo (j nhi~t af)thu:angthi cacmduthityphdnco hamlU:<;1ngproteinhoatan,hamlU:<;1ngpeptid vahamlU:<;1ngNaatqo thank caDhan nhi~uso WJi mduthityphdn (j nhi~tdq thu:ang. Tuyv(iyne'uthityphdn lien tl;lC(j nhi~tdq 50°C trong thai gian dai thi nit kho khan aOIWJiddu ki~nsanxudtth1!Cte'tqi Vi~tNamvalamchiphi gia thanksecaDhan. Conne'uthityphdn(j nhi~tdq 50°Ctrong2 giGddusaudo (j nhi~tdq thu:angthi hamlU:<;1ngprotein hoa tan, hamlU:<;1ngpeptid va ham lU:<;1ngNaat(1OthankcungcaD 119 harznhiJuso vaimduthayphana nhiftdQthuiJng;nhltngniu so sanhvaiqua trinhthayphana nhift dQ50oethihamlu(Jngproteinhoatanvahamlu(lngpeptid U;1Othanha mdunaycaohonchutit conhamlu(JngNaatr;1OthanhIt;lithaphon chungtoquatdnhthayphankhacbi~tkhongnhdu.Hon nilakhi thuyphan(ynhi~t d(j50°etrong2 gilJ saudo thayphana nhift dQthuiJngthi trongth1!Cti It;lidl ap dl,mgbaisanxudtnuacmiimco thi kef:h(lpvaiphai niingdi t<:ind1:lngniingb,t(lng m(ittrlJi ma cac tlnhmiJn TrungthuiJngco s6 gilJ niingcao. Vi v<:iychi dQthuy phiin(ynhi~tdQ50oetrang2 gilJ saudo thuyphan(ynhift dQthulJng11!achQndi titpt1:lCnghiencau. 3.4.2.3.Xac dinhche'dQb6 sungmu6ian choquatrinh thuyphanca cdmbAngproteaseB.subtilis85 Tie'nh~mhcaethinghi~mthuyphaneaedmb~ngproteaseB.subtilis55d nhi~tdQ50DCtrong2 giod~usaild6d nhi~tdQthu'ong.M6i mftuthuyphansa dt,mg5 kg eaedmxay,b6 sung0,2%CPE tuB.subtilis55,thuyphand pH tv nhieneuathiteacontyl~mu6ianb6sungthlthayd6inhu'sau:mftu1:hie6gio b6sung1%mu6iansovoinguyenli~usaild6ell6gioehothem1%mu6ianeho toikhid(;ltdu 25%mu6i;mftu2:hie6 giob6 sung2%mu6iansovoi nguyenli~u saild6 ell 6 gio ehothem2% mu6ianehotoi khi d(;ltdu 25%mu6i;mftu3:hIe6 giOb6sung3%mu6iansovoinguyenli~usaild6ell6gioehothem3%mu6ian chotoikhid(;ltdu25%mu6i;mftu4:hie6giob6sung5%mu6iansovoinguyen li~usaild6ell6 gioehothem5%mu6ianehotoikhid(;ltdu25%mu6i;mftu5: . hie6giob6sung10%mu6iansovoinguyenli~usaild6ell6 gioehothem10% mu6ianehotoi khi d(;ltdu25%mu6ivamftu6:hie6 giob6 sungdu25%. Sail caekhmlngthaigian0, 2,...,168giOIffymftudanhgia earnquaDva phantichhoahQe.Ke'tquadu'Qetrlnhbayd bang3.26va caehlnh3.57,3.58, 3.59.3.60. 120 Bang3.26.Anh hu'dngeiia rnu6ian de'nsrI thayd6i tr~ngthai earnquaD eiia caernftuthiiy phan eaedmbiingproteaseB.subtilisSS Thai M~u1 M~u2 Mh3 Mh4 M~u5 M~u6 glaD (gia) 0 H6n h<;1pthit I H6n h<;1pthit I H6n h<;1pthjt I H6n h<;1pthjt I H6n h<;1pthjt I H6nh<;1pthjt ca tnlng mh l ca hdi tra:ng m~u I ca hdi tra:ng mh l ca hdi tra:ng mh I ca hdi tra:ng m~u I ca hdi tra:ng m~u hdi xam,comui xam,comui xam,comui xam,comui xam,comui xam,comui tanh t1,l'tanh t1,l'tanh t1,l'tanh t1,l' tanh t1,l'tanh t1,l' nhien, thjt nhien, thjt nhien, thjt nhien, thjt nhien, thjt nhien, thjt cad~ccleo, cad~ccleo, cad~ccleo,cad~ccleo, cad~ccleo, cad~ccleo, hdi 16n hdi 16n hdi 16n hdi 16n hdi 16n hdi 16n nh6n. nh6n. nh6n. nh6n. nh6n. nh6n. 2 H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt ca. fiat ca fiat ca fiat ca fiat ca fiat ca fiat nhuyenhdi nhuyenhdi nhuyenhdi nhuyenhdi nhuyen hdi nhuyenhdi long, mh long, mh long, mh long, m~u long, mh long, m~u hdi xam, hdi xam, hdi xam, hdi xam, hdi xam, hdi xam, ma't mui I ma't mui I ma't muiI ma't mui I ma't miii I ma't miii 4 tanh,comiii tanh,comiii tanh,comiii tanh,comiii tanh,comiii tanh,comiii caha'p. caha'p. caha'p. caha'p. caha'p. caha'p. H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt ca d,.lOgca d;:lngca d;:lngca d;:lng ca d;:lngca d;:lng djch long, djch long, djch long, djch long, djch long, djch long, mh hdi m~u hdi m~u hdi mh hdi mh hdi mh hdi xam,comuiI xam,comuiI xam,comiii I xam,comiii I xam,comiii I xam,comiii d~ctntng. d~ctntng. d~ctntng. d~ctntng. d~ctntng. d~ctning. 6 H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt H6nh<;1pthjt ca d;:lngca d;:lngca d;:lngca d;:lngca d;:lngca d;:lng djch long, djch long, djch long, djch long, djch long, djch long, hdi phan hdi phan hdi philo hdi phan hdi philo hdi philo ;:, ;:,;:, ;:,;:, ;:,;:, ;:,;:, ;:,;:, ;:, tang, mall tang, mall tang, mall tang, mall tang, mall tang, mall hdi xam,co hdi xam,cohdi xam,co hdi xam,co h~ixam,co hdixam,co mui d~c miii d~c miii d~c miii d~c miii d~c miii d~c tning tntng tru'ng tning tntng trung 121 H6n h<,1pthit H6n h<,1pthit H6n h<,1pthit H6n h<,1pthit H6n hQpthit H6n h<,1pthit ea d,.wg ca d,.wg ea d<:lngea d<:lngca d<:lngca d<:lng long nhu long nhu lOng, hdi long, hdi long, hdi long, hdi nude, phan nude, phan phan thg, phan t<1ng,phan t<1ng,phan t<1ng, 8 dug, m<1,~ I t<1,ng, mm xam, m<1uImh miiiI miii l 110 I xam, d~etrung.d~etrung. trung. xam, I m<1u d~e miii xam, I m<1u d~eI miii trung. xam,I mh xam, d~eI miii d~e trung. H6n h<,1pthit H6n h<,1pthit H6n h<,1pthit H6n h<,1pthit H6n h<,1pthit H6n hQpthit ea long nhu ea long nhu ca d<:lngea d<:lngra't ea d<:lngea d<:lng nude, phan nude, phan long nhu long, phan long, hdi long, hdi dng m<:lnh,thg m<:lnh,nude, phan thg, m<1uphan t<1ng,phan t<1ng, Idp Ph:a I t<1,ng, mauxam,tren Ph:a l Id:mati tren yang miii nh<:lt,I d~etrung hdi nh<:lt, I yang hdi miii n6ng. n6ng. trung. mh I xam, miii I d~etrung. miii I m<1u miii trung. xam, I m<1u d~e miii xam, d~e trung. H6n h<,1pthit H6n h<,1pthit H6nh<,1pthit H6n h<,1pthit H6n h<,1pthit H6n h<,1pthit ea lOng nhu ea long nhu ea long nhu ea d<:lngea d<:lngca d<:lng nude, phan nude, phan nude, phan long nhu long, phan long, hdi t<1ng ra't thg ra't. t<1ngm<:lnh,nude, phan dug, m<1uphan t<1ng, 12 m<:lnh, phia Idp I ldP tren tren Idp I m<:lnh, tren phia mh yang I mh miii nh<:lt, Ya~~ I Ya,~gmm mmnh<:lt, m~mnhy. n6ng.m~mnhy. Ph:a I t<1,ng, mati xam, mh I xam, miii I d~etrung. nh<:lt,I d~etrung hdi miii I m<1u xam, miii d~e trung. H6n h<,1pthit H6n h<,1pthit H6n h<,1pthit H<3nh<,1pthit H<3nh<,1pthit H6n h<,1pthit ca longnhu ea longnhu ea longnhu ea longnhu ea d<:lngea d<:lng nude, phan nude, phan nude, phan nude, phan lOng nhu long, phan t<1ng ra't thg ra't t<1ng ra't t<1ngm<:lnh,nude, phan dug, m<1u 36 m<:lnh, phia Idp I m<:lnh, tren phia Idp I m<:lnh, tren phia m<1u Yang, I m<1uyang4 I m<1u eomiiinudeI eomiiinudeI nh<:lt, mall. m~m. m~mnhyo Idp jIdp phia I dug, trenI tren matiI xam, Ya~~ I Ya,~g mm mm nh<:lt,I d~etrung hdi n6ng. m<1uI xam, miii miii I d~etrung. 122 60 H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht ea longnhu ea lOngnhu ea longnhu ea longnhu ea longnhu ca dng nuoe, phan nuoe, phan nuoe, phan nuoe, phan nuoe, phan long, phan tng rtt tng rift thg rift thg rift tng mnh, dug, i>mall mnh, lOp mnh, lOp mnh, lOp mnh, lOp lOp phia xam, mlii phla tren phia tren phia tren phia tren tren i> de trung.mall i> Yang, i> Yang, mu Yang, i> yang yang nht,mall mall mall mlii nuoe mlii nuac eo mlii nht, mlii mlii hdi miimr6rt$t. miimr6rt. miim. miimnhy. i> nong. 84 H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht ca longnhu ea longnhu ea lOngnhu ea longnhu ca longnhu ca dng nuoe, phan nuoe, phan nuoe, phan nuoe, phan nuoe, phan long nhu tng rtt thg rift tng rift tng rift tng rift nuoe, phan mnh, lOp mnh, lOp mnh, lOp mnh, lOp mnh, lOp thg, i>mall phia tren phia tren phla tren phia tren phia tren xam, mlii i> Yang, i> Yang, i> Yang, i> Yang, i> yang de trungmall mall mall mall mall mlii de mlii de mlii nuac eo mlii nht, mlii trung ? trung ? miim r6 rt. J miimnhy.eua eua maID.. nuoemiim. nuoemiim. 108 H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht ea longnhu ea longnhu ca longnhu ca longnhu ea longnhu ca longnhu nuoe, phan nuoe, phan nuoe, phan nuoe, phan nuoe, phan nuoe, phan tng rift thg rift dng rift tng rtt thg rift tng mnh, mnh, lOp mnh, lOp mnh, lOp mnh, lOp mnh, lOp lOp phia phia tren phia tren phia tren phia tren phia tren tren i> mall i> Yang, i> Yang, mu Yang, i> Yang, i> Yang, yang nht,mall mall mall mall mlii de mlii de mlii de mlii nuoe co mlii mlii hdi trung eua trung eua trung eua miimr6rt. maID. i> nong. nuoemiim. nuoemiim. nuoemiim. 132 H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht H6n h<;1ptht ea longnhu ca longnhu ea longnhu ea longnhu ca longnhu ca longnhu nuoe, phan nuoe, phan nuoe, phan nuoe, phan nuoe, phan nuac, phan thg rift thg rift tng rift thg rift tng rift tng rtt mnh, lOp mnh, lOp mnh, lOp mnh, lOp mnh, lOp mnh, lOp phia tren phia tren phia tren phia tren phla tren phia tren i> Yang, i> Yang, mh Yang, i> Yang, i> Yang, i> yangmall mall mall mall mall mlii de mlii de mlii de mlii de mlii nuoe nht, mlii trung eua trung ? trung ? trung eua miimr6 rt. miimnhy.eua eua nuoemiim. nuoemiim. nuoemiim. nuacmiim. 123 156 H6n h<;1pthit H6n h<;1pthit H6n h<;1pthit H6n h<;1pthit H6n h<;1pthit H6n h<;1pthit ea longnhu ea longnhu ea longnhu ea longnhu ea longnhu ea lOngnhu nude, phan nude, phan nude, phan nude, phan nude, phan nude, phan t~ng ra't thg ra't t~ng ra't t~ng ra't thg ra't t~ng ra't m~nh, lOp m~nh, lOp m~nh, lOp m~nh, lOp m~nh, lOp m~nh, lOp phia tren phia tren phia tren phia tren phia tren phia tren mffu mlii Yang, I mffu di;iemlii Yang, I m~u di;iemlii Yang, I m~u di;iemlii Yang, I m~u di;iemlii Yang, I m~u di;ie eo Yang, mlii tru~g J eua I tru~gJ eua I tru~gJ eua I tru~g J ciia I tru~g J euaImam.nudemam. nudemam. nudemam. nudemam. nudemam. 168 H6n h<;1pthit H6n h<;1pthit H6nh<;1pthit H6n h<;1pthit H6n h<;1pthit H6nh<;1pthit ea longnhu ea longnhu ea longnhu ea longnhu ea longnhu ea longnhu nude, phan nude, phan nude, phan nude, phan nude, phan nude, phan tftng ra't thg ra't t~ng ra't thg ra't thg ra't t~ng ra't m~nh, lop m~nh, lop m~nh, lOp m~nh, lOp m~nh, lOp m~nh, lOp phia tren phia tren phia tren phia tren phia tren phia tren mffu mlii Yang, I mffu di;iemlii Yang, I mffu di;iemlii Yang, I m~u di;iemlii Yang, I mffu di;iemlii Yang, I m~u di;ie mlii Yang, nude trung eua I trung eua I trung eua I trung eua I trung euaImamro. nudemiim. nudemiim. nudemiim. nudemiim. nudemiim. 550 500 ~ )I(~-)I(--",~ ~-)I(~ ~ ~--)I( c ~ 450 ,C;I :;-400 O,<c;I ..c '0 350c c '2 .8 3000 .- 15.'~ 250 b.O 0 g. '" 200 :E! ~ E '-" 150 ,C;I ::r: -+-M~u1 --M~u2 --4- M~u3 50 0 -0- M~u4 -)1(- M~u5 -t:r- M~u6 100.; 0 2 4 6 8 10 122 4 36 48 60 72 849 6 108 120132144 156168 Thai gian thuyphan(giO) ? mnh 3.57.Anh hlidng cuamu6ian de'nhamllic;ingprotein boa tan cua cae miu thuy phan ca cdmbiingproteaseB.subtilisSS (Hamlu'<Jngproteinboatanbandc1ula 34,1mg/gm~uva du'<JCtrlnhbay (j bang3.9- ph\ll\lc 3) 124 900 800 -+-Mau 1 --Mau2 -e-Mau 3 -o-Mau 4 -)1(- Mau5 -tr-Mau 6 c: ~ .D ~'6 ~ 0'" 700 600 ~ :;; 8500 ':;: '(~ 8' '0 400 0- gf 300 '0' ~ 200s ,~ :r: 100.: 0 0 2 4 6 8 10 12 24 36 48 60 72 84 96 108 120 132 144 156 168 Thai gian thuyphan (gia) mnh3.58.Anh hu'dngcuamu6iande'nhamhi<jngpeptidcuacaemdu thuyphancacOmb~ngproteaseB.subtilis55 (HamluQngpeptidbandftula 0,36mg/gm~uva duQCtrlnhbay dbang3.10- ph\ll\lc 3) 600 -+-Mau 1 -- Mau2 -e- Mau3 -0- Mau4 -:I(-Mau5 -tr- Mau6 :1(-:1(-:1(-:1(-:1(-:1( :1(-:1(-:1( :1(-:1( v :1(-:1(;;::0=~ 500c '8 ~ (Ij ;::I ,':::,~ 400 :x: c (Ij (Ij '0 .D "2 ~'6 300 bIJ ;;. c 0 B' '" ~ 200 S '-' ,(Ij ::c 10:1 0 2 4 6 8 101 2 24 36 48 60 72 84 96 108120132144156168 Thai gian thuyphan(gio) mnh3.59.Anh hu'dngcuamu6iande'nhamlu'<jngNaacuacaemdu thuyphancacdmb~ngproteaseB.subtilis55 (HamluQngNaabandftula0,41mg/gm~uvaduQCtrlnhbay dbang3.11- ph\ll\lc3) 125 1500 -:':-M~u5 -l:r- M~u 6 -:.:-:.:-:.:-:.:-:.:-:.:-:.:-:.:-:.:-:.:-:.:-:.:-:.:-:.:-+ . . . '" 1200 ~,g '0 "0 .'.:: I:: I:: ~ 900 OJ) .- I:: "0 '0' .. .:= 0 s :; 600 ,oj ,"' ::t: '-" -+- M~u1 ~ M~u2 M~u3 -0- M~u4 300 --lX~ 0 0 2 4 6 8 10 12 24 36 48 60 72 84 96 108120132144156168 Thai gianthuyphan(gia) Hinh3.60.Anh hlidng euarnu6ian de'nham lliqng NNH3euacaern§u thuy phan eaedrnb~ngproteaseB.subtilisSS (HamIU<;5ngNNH3band~ula 0,023mg/gm~uvadu<;5etrlnhbay d bang3.12- phl;lll;le3) Til caeke"tquanghieneUud trenehophepnit ra mQts6nh~nxetnhusau: *V~tr~ngthai earnquaD:m~uthuyphanb6 sungmu6ieangit thl muedQ thuyphaneangnhanhbi~uhi~nIa theothaigianm~ueangnhanhe6hi~ntu<;5ng phant~ngvah6nh<;5pthuyphaneangnhanhehuy~nsangtq.ngthailong. * V~rn~tboahQe: Til ke"tquatrlnhbaytrenhlnh3.57va 3.58ehoth§ytheothaigianthuy phancaem~ub6sungmu6ianband~uvoity l~eangth§pmuedQthuyphanthit eelrangm(;1nhnenhamIU<;5ngproteinhoatan,hamlu<;5ngpeptidt(;10thanheang nhanh,nhi€u vathaigianmahamIU<;5ngcaeeh§tnayd(;1ttoigiatri qie d(;1ieang som.Ch£ngh(;1nthaigianmahamlu<;5ngproteinhoatand(;1tqted(;1idcaem~uI, 2,3,4,5 va6 la: 12,12,12,24,60va 144giavaeaog§ptudnglingsovoiban d~ula:5,51;5,36;5,29;5,28;5,18va5,111~n;thaigianmahamIU<;5ngpeptidd(;1t el/ed(;1idcaem~u1,2,3,4,5 va61a:12,12,12,24,132va 156giOvaeaotudng lingg§p:8,40;8,37,8,34,8,32,8,00va 7,81I~nso voi band~u.Saud6ham 126 lu<;1ngproteinboatan,hamhlQngpeptidcuacaem~unayb~tdgugiamm(;lnhva m~uco thai gian d(;ltCl,l'cd(;licangsamthl muc dQgiamcangnhanh.Nhuv?y, m~u4 (b6sung5% mu6ian)co hamluQngproteinboatanva hamluQngpeptid t~othanhkhi d(;ltCl,l'cd(;lichI tha'pchutit so voi caem~u1, 2, 3 nhungcaobon nhi€usovoi caem~u5 va6. K€t phantichhamluQngNaatheothai gianthuyphan(hlnh3.59)chotha'y caem~uthuyphancacomb~ngproteaseB.subtilisS5d€u cohamluQngNaatang theothaigianthuyphanva m~ub6 sungmu6ian19ndgucangtha'pmucdQthuy phant(;lOthanhNaacang nhanh.d thai di€m sail 168gia thuy phan ham luQngNaa t~othanhcuacaem~u1,2, 3,4,5 va6 tangtuonglingsovoi bandgula: 590,5%; 585,3%;561,8%;488,0%;450,0%va 350%.Nhu V?ym~u4 (b6sung19ndgu5% mu6ian) co hamluQngNaat(;lOthanhtha'pboncaem~u1,2, 3 va caobonnhi€u sovoim~u5 va 6. K€t quaphantichs1;1'bi€n d6i cuahamluQngNNH3(hlnh3.60)chotha'ycae m~uthuyphanco ty l~mu6ian b6 sung19ndgucangtha'pthl hamluQngNNH3 t~othanhtheothai gian cangm(;lnh.Ngoai fa, k€t quanghienCUllcon chotha'y khity l~mu6ianb6 sungvaom~uthuyphand(;ltdu 25%thl hamluQngNNH3t(;lO thanhcuam~uthuyphanb~tdgugiamm(;lnh.Caem~ub6 sungmu6ian19ndgu yoity l~tha'pd€u co hamluQngNNH3t(;lOthanh(d clingmQtthai di€m) caobon caem~ub6 sungmu6ian 19ndguvoi ty l~cao.d thai di€m 168 giO thuy phan hamluQngNNH3cuacaem~u1,2, 3,4,5 va 6 tangtuongling so voi bandgula: 1422,4%;1100,0%;573,0%;301,3%;255,9%va 207,0%.Nhu V?ym~u4 coham lU<;1ngNNH3t(;lOthanhtha'pbon ra'tnhi€u so voi m~u1, 2, 3 va cao bon khong dangk€ sovoi m~u5 va6. Titcaephantick iJ trenchothaymdu4 (b6sungmw5'ian Iandau5%)coham lu(Jngproteinhoa tan,hamIu(lngpeptid,hamIu(lngNaat(,lOthanktheothaigian thayphanthaphanm(Jtchutsovelicaemdu1,2,3nhungcaohannhi~usovelicae mdu5 va6.M(itkhachamIu(lngNNH3t(,lOthankcuamdu4 I(,lithaphannhi~uso valcaemdu1,2,3 nhungcaohankhongnhdu socaemdu5 va6.Ran mia,s6'Ian 127 b6sungmuolddivaimJu4 (b6sungmudianIdnddu5%)khongnhi~unentrong rangngh~che'biitnnuacmiimth1!Cte'dl th1!Chi~n.Do v(lYthinghi~mch(Jnche'dQ b6sungmudianVaGh6nh(lpthayphancdcamthuang(5toiephoruscommersonii) blingproteaseB.subtilis55 Ia Iuc6 gia chomuo'iIdndduvai tYI~5%. 3.4.3.ThO'nghi~msan xua'tmioc miim ca co'mb~ngproteaseB.subtilis ss 3.4.3.1.So sanh kha Dangthuy philo ca com trong san xmlt nu'oc m~mgh1aproteaseB.subtilisSSvOimQts6proteasekhac D€ so sanhkha nangthuyphanca cdmtrongsanxu:1tmtocmaIDcua proteaseB.subtilis55voi cacmQts6lo(;liproteasekhacnhu':NFD cuaCanada, che'phffmNeutraseva FlavourzymecuaNovo- DanM(;lch,thinghi~mdfftie'n hilllh6m~u,rn6im~u20kgcacdmthuyphantheocacche'dQkhacnhau: M~u1:m~ud6i chungtrQncacdmxayvoi 25%mu6ian. M~u2: b6 sung0,2%CPE tuB.subtilis55VaGh6nh<;5pca cdmxayva sail6 giothuyphanb6 sung5% mu6ian,saild6cu6 giGb6 sungtie'p5% mu6iancho toikhidu25%. M~u3: b6 sung0,2%CPE tu B.subtilis55 VaGh6nh<;5pca nguyenconthuy phansail6 giGthl b6 sung5% mu6iansaild6 cu 6 giGb6 sungtie'p5% mu6ian chotoi khi du 25%. M~u4: b6 sung0,2%che'phffmproteaseNFD cuaCanadaVaGh6nh<;5pca comxay,che'dQchomu6itu'dngtV. M~u5: b6 sung0,2% che'phffmproteaseNeutrasecua Novo - Dan M(;lch VaGh6nh<;5pcacdmxay,che'dQchomu6itu'dngtV. M~u6: b6 sung0,2%che'phffmproteaseFlavourzymecuaNovoVaGh6n h<;lpcacdmxay,che'dQchomu6itu'dngtV. Sailcackhoangthaigianthuyphan0,2,4,...,168giGl:1ym~udanhgiaearn quanvaphantichmQts6chitieuhoahQc;ke'tquadu'<;5ctrlnhbayd bang3.27va cachlnh3.61,3.62,3.63,3.64. 128 Bang 3.27.811bie'nd6i tqmg thai earnquaDcua caemilu thuy phan ca co'mb~ngm()ts6lo~iproteasekhac nhau Thai M~u1 glan (gia) M~u2 M~u3 ca mallI ca H6n h<;1pthit I H6n h<;1pthit I Kh6i mhlmh trangI ca ca I H6nh<;1pthit I H6n h<;1pthit I H6n h<;1pthit mall r 0 f tdng hcliI tr~ng hcliI hclixam,co I tr~ng M~u4 mall I ca hcliI tdng M~u5 M~u6 mallI ca hcliI tr~ng hcli tanh hI I tanh xam,comlii Ixam,comlii Imlii tanhtvIxam,comliiI xam,comliiI xam,comlii nhien, thitI nhien, tvI nhien. tanh thit nhien, ca di,icdca,I cadi,icdca,I cadi,icdca, 16nhcli 16nI hcli ca di,icdca, I cadi,icdCa, hcli nh6n. nh6n. 16n nh6n. tv I tanh thitI nhien, 16nI hcli nh6n. tv I tanh tv thitthitInhien, 16nI hcli nh6n. ca16nnh6n,I ca H6n h<;1pthit I H6n h<;1pthit I Kh6icahcliIH6n h<;1pthit I H6n h<;1pthit I H6n h<;1pthit2 mall fiat I b~ bvng,I ca caI nhuyenhcliI nhuyenhcliI nhuy~nhcli mall hcliI nhuyenhcliI mtoc xam,comlii I long, tanh. hcli ma't mhlmh hcli lOng, ma't hclixam,I vang, mlii I mliitanh. ma't tanh,comlii I tanh,comlii I tanh,comlii caha'p. tanh,comlii caha'p.caha'p. fiat I ca mall I long, xam,I hcli mlii I ma't cahtlp. fiatI ca fiat mall I long, xam,I hcli xam, mlii I ma't mlii ca con 16nI ca H6n h<;1pthit I H6n h<;1pthit I Kh6icahcliI H6n h<;1pthit I H6n h<;1pthit I H6n h<;1pthit4 nh6n, mh I dich hclixam,coImh mliitanh. d~ngIfiat,nttocca I ca long,Imh hcliI nhuyen, hcliI vang, ma'tImall di,ictrttng. xam,comlii Imliitanh. di,ictrttng. xam,comlii I xam,comlii I xam,comlii di,ictrttng. longI ca nhuyen, hcliI mall di,ictrttng. longI ca long nhuyen, hcliI mall hcli 129 H6nh<Jpthit H6nh<Jpthit Kh6i ca hdi H6nh<Jpthit H6nh<Jpthit H6nh<Jpthit ca hdi oat, ca di;lng oat,nudcca ca di;lng ca di;lng ca di;lng m~u xam, dich long, mh hdi dich long, long, m~u long, m~u mliitanh. hdi phan Yang, ma't m~u hdi hdi xam,co hdixam,co t~ng, m~u mliitanh. xam,comlii mlii d<)cmlii d<)c 6 I I i I ~ I 24 thdm xam,comlii d<)c thdm tIling. tIling d<)cI tIling tIling H6nh<Jpthit H6nh<Jpthit Kh6i ca hdi H6nh<Jpthit H6nh<Jpthit H6nh<,1pthit ca oat ca longnhu oat,nu<kca ca di;lng ca di;lng ca di;lng nhuy~n, nudc, phan m~u hdi dich long, dich long, dich long, m~u xam, thg mi;lnh,Yang, ma't hdi phan hdi phan hdi phan co mlii hdi I ldp tanh. tIeD yang mlii d<)ctIling. phia I mliitanh. mh t~ng, xam,comlii Ixam,comlii I xam,comlii d<)cnhi;lt, thdm thdm tIling. m~uI t~ng, d<)cI thdm tIling. m~uI t~ng, m~u d<)cI thdm tIling. H6nh<Jpthit H6nh<Jpthit Kh6i ca oat H6nh<Jpthit H6nh<Jpthit H6nh<Jpthit ca long ca longnhu nhuy~ncon ca di;lng ca di;lng ca di;lng nhuy~n, nudc, phan mQt chUt dich long, dich long, dich long, m~u xam, t~ng ra't thit dinh phan thg phan t~ng phan thg ma't mlii mi;lnh, ldp vao xudng, mi;lnh, ldp mi;lnh, ldp mi;lnh, ldp 48 tanh. phia mh tIeD i comliinu<kI phia yangI m~m. m~u nhi;lt,comlii maID. Yang, mlii maID. tIeD I phia hdi I m~u co I Yang, mlii maID. tIeD I phia hdi I m~u tIeD hdi co I Yang, co mlii maID. H6nh<Jpthit H6nh<Jpthit Kh6i ca oat H6nh<Jpthit H6nh<Jpthit H6nh<Jpthit ca long ca longnhu long tach ca di;lng ca di;lng ca di;lng nhuy~n, nudc, phan rieng khoi dich long, dich long, dich long, mh xam, t~ng ra't xudng, co phan t~ng phan t~ng phan t~ng 72 ma't mlii I mi;lnh, tanh. phia m~u ldp I mlii tren m~m. nudc I mi;lnh, phia Yang, m~u maID. comliinu<k mlii maID. Yang, ldp I mi;lnh, tIeD phia hdi I m~u coI Yang, mliim~m. ldp I mi;lnh, tIeD phia ldp tIeD hdi I m~u hdi coI Yang, co mliim~m. 120 H6n h<;Jpthit ea d~ng long, mffu xam, mtt mliitanh. 168 H6n h<;Jpthit ea d~ng lOng, mffu xam, mtt mliitanh. 600 i:: 500 <:13.... '<:13~ 0 :::! ..c::'«:13400 i:: '0., i:: (!) <:13 '0 .D S. ~~300 01) ;> i:: 0 :0' en lJ ~ 200 8 '-' '<:13 ::I: 1°: j ~ ~ 130 H6nh<;Jpthit Kh6i ca fiat H6nh<;Jpthit H6nh<;Jpthit H6nh<;Jpthit ca longnhu long nhu ca d~ng ca d~ng ca d~ng nude, phan nude, phan dieh long dieh long dieh long tffng rtt tffng, mlii nhu nude, nhu nude, nhu nude, m~nh, ldp d~e trung phan thg phan tffng phan thg phia tren eua nude rtt m~nh, rtt m~nh, rtt m~nh, mffu v~lllg,m~m. ldp phia ldp phia ldp phia mlii d~e tren mffu tren mffu tren mffu trung eua nudem~m. hdi V~lllg,eo I hdi Yang,eo Ihdi Yang,eo mliiroam. mliiroam. mliiroam. H6nh<;Jpthit Kh6i ea fiat H6nh<;Jpthit H6nh<;Jpthit H6nh<;Jpthit ea longnhu long nhu ea longnhu ea longnhu ea longnhu nude, phan nude, phan nude, phan nude, phan nude, phan thg rtt. tffng, mlii tffng rtt tffng rtt thg rtt m~nh, ldp d~e trung m~nh, ldp m~nh, ldp m~nh, ldp phia tren eua nude phia tren phia tren phia tren mffu Yang,Im~m. mlii d~e trung eua nudem~m. mffu mlii Yang, I mffu d~emlii Yang, d~e Yang, I mall d~e mlii trung eua I trung eua I trung eua nudem~m. nudem~m. nudem~m. --+- M~uI -0- M~u2 --- M~u3 -0- M~u4 -;1(- M~u5 -(r- M~u6 J. % "'~ ~~ ~ ,,% ",'Y '-~,," ". ,) Thaigianthuyphan(gio) ,,'Y IDnh3.61.811bie'nd6ihamlu'c;tngproteinboatancuacaemiluthuyphan caco'mb~ngmQts6lo~iproteasekhacnhau (HamhiQngproteinhoatanband~u34,22mg/gmfiuvadu'Qctrlnhbay (jbang3.13- phl,lll,lc3) 131 900 --+-M~u I -0- M~u 2 -+-M~u3 --M~u4 -)K- M~u5 ~ 500 -l:r- M~u6 :J '~ 800 c:: 2 700 ~-S ;> 600 0'" :9 400 0.. & 300 OJ) - c:: ], 200 E ,o:j ::r:: 100.: 0 - 0 2 4 6 8 10 12 24 36 48 60 72 84 96 108 120 132 144 156 168 Thai gian thuy phan (gia) IDnh3.62.811bie'nd6ihamlu'(jngpeptidcuacaemftuthuyphancacom biingmQts6lo~iproteasekhacnhau (HamluQngpeptidband~uO,35mg/gm~uva duQctrlnhbay (5bang3.14- phvIvc 3) 500 -+-M~u I -0- M~u2 -+-M~u 3 --M~u4 -)K- M~u5 -l:r- Mau6 ,S 400 E ~ o:j ;::I ~ '<o:j , >< "0 o:j c:: 300 "0 o:j~ .D '2 ~-s OJ) ;> :6, S; 200-=- ~ E '-' ,o:j ::r:: 100. 0 0 2 4 6 8 10 12 24 36 48 60 72 84 96 108120132144156168 Thai gian thuy phan(gia) ffinh3.63.811bie'nd6ihamlu'(jngNaacuacaemftuthuyphancacdm biingmQts6lo~iproteasekhacnhau (HamluQngNNH3band~uO,39mg/gm~uva duQctrlnhbay (5bang3.15- phvIvc 3) 132 0 0 2 4 6 8 10 12 24 36 48 60 72 84 96 108120132144156168 Thai gian thuy phan(giG) ffinh3.64.St1bie'nd6ihamh.i(jngNNH3euacaemiluthuyphaneaedm biingmQts6lo~iproteasekhaenhau (HamluQngNNH3bandatiO,O21mg/gm~uva duQCtrinhbay 0 bang3.16- ph\ll\lc 3) Tli cacke'tquanghienCUua trenrhopheprutramQts6nh~nxet: *V~tr~ngthaiearnquaD:ke'tquadanhgiaramquail0bang3.27rhoth:1y caem~uthuyphancob6sungcaclo;:tiproteasekhacnhaud~ucotr;:tngthairam quailthayd6itheothaigiannhanhhanm~ud6ichungkhongsud\lngenzyme. M~uthuyphancacomxayb~ngproteaseB.subtilis55cosVthayd6itr;:tngthai earnquailnhanhhancacm~ukhacbi€u hi~nla theothaigianm~unaynhanh chuy€nsangtr;:tngthailongvacohi~ntuQngphantangnhanhhall. * V~m~thoa-llQe Ke'tquaxacdinhsv bie'nd6ihamluQngproteinhoatan0 hlnh3.61eho th:1ycaem~ub6sungcacenzymekhacnhaud~ucohamluQngproteinhoatan t~othanhtangnhanhvam;:tnhanm~ud6ichungsaudod;:ttcvcd;:ti;eh£ngh;:tn m~u2 co hamluQngproteinhoatand;:ttcvcd;:ti527,6%sovoi bandau0 thai di€msau24giathuyphan;cacm~u3,4,5, 6rungcohamluQngproteinhoatan d~tcvcd;:ti0 thaidi€m sau36giathuyphanvacaotuangungsovoibandaula: 600 -+-Mu I-;- Mu 2'<0:1'0 .:: 500 -+- Mu 30:1 .Q --Mu 4 <6 -)1(- Mu 5;> 0 400 Mu 6'" :£ 300 Z B 'a 200-IOJ) .:: '0' .E S 100.:,0:1 ::r:: 133 18,6%520,7%,498,7%va 521,5%.Saudo hamluQngproteinboatanghlm ~nh,d€n thaidi~m168giOthuyphancaem~u1,2, 3, 4, 5, 6 co hamluQng oleinboatangiamchI contuongungso vdi band~ula: 227,4%,130,5%, 16,0%,145,6%,150,2%,139,8%.Nhuv~y,m~u2 (b6sungproteaseB.subtilis - dethuyphancacomxay)cohamluQngproteinboatangiamtha'pnha't.M~u ~lchungc~mUC.dOt:0 th~nhp~oteinbOatan~UC,d~u nh"va tangd€~danth:O I~lglanthuyphandenthOIdIemsau168gIOhamluQngproteInboatancua r§unayv~ncondangtang.K€t quanaycoth~giaithichla dudilacdQngcua pcproteaseb6sungproteinthitcabi thuyphangiaiphongdudid~ngboatanr6i audo l~iti€p tvc bi thuyphant~othanhcaepeptidva axil aminoCon m~u thongb6sungproteasethlquatrlnhthuyphanchuy€u doht%proteasenQit~ida: 'imu6ianucch€ lamgiammQtph~nho~tdQnent6cdQthuyphanch~m.VI th€ ranxua'tnudem~mtheophuongphaptruy€nth6ngthaigiansanxua'tphaikeo laitu6thangd€n canam. Khi phantichhamluQngpeptidcuacaem~uthuyphan(hlnh3.62)clingcho ~e'tquatuongh!Ia caem~uthuyphancob6sungcaeenzymed€u cohamluQng Jeptidt~othanhgiaido~nd~utangnhanhhonra'tnhi€u sovdi m~ud6ichung ;audod~tc1!cd~ir6igiamm~nhonsovdicaem~ud6ichung. K€t quaphantichhamluQngNaa(hlnh3.63)chotha'ycaem~ucob6sung ?roteased€u co hamluQngNaat~othanhtangd€u va caohonm~ud6ichung. M~u2 cohamluQngNaat~othanhtheethaigianthuyphantangd€u vanhanh honta'tca caem~ucon l~i.d thai di~m 168 giO thuy phan ham luQng Naat~o thanhtrongcaem~u1,2, 3,4, 5,6 tangtuongungsovdi band~ula: 217,6%, 491,2%,341,3%,416,3%,423,6%,432,7%. K€t quaphantichhamluQngNNH3(jhlnh3.64chotha'y(jgiaido~nd~ucua quatrlnhthuyphanhamluQngNNH3t~othanhtrongcaem~ucob6sungenzyme caohonm~ud6ichungnguyennhancothela dohIe'd~uchuab6sungdumu6i annenvi sinhv~tho~tdQngm~nhVIv~ymacaem~uthuyphancangm~nhthl mucdQt~othanhNNH3cangnhanh.Saukhib6sungdumu6ithlhamluQngNNH3 134 t(;10thanh(j cacm§:ub6 sungenzymel<;1ig amd~nr6i tha'pngangvdi hamluQng NNH3t<;10thanhtrongm§:ud6ichung. TiccaephantichiJ trenchothdymt1u2 (thayphancacamxayblingprotease B.subtilisS5) co hamlll(1ngproteinhoatan,hamlll(1ngpeptidva hamlll(1ngNuu t(LOthankcaDhancaemt1uthayphankhac.Caemt1uthayphanmc;mhanluc ddu seeohamlll(1ngNNH3eaohannhllngiJ giai doc;zncu(Jithihamlll(1ngNNH]lc;zigidm xuonggdnnhllnhau.Nhll V(lYsadT;tngproteaseB.subtilisS5di thayphancaeam sanxuatnllacmdmse co toedo thayphanmc;znhhancaeche'phdmenzymekhae mathinghi~mnaysadT;tng. 3.4.3.2.Tho.-nghit$msanxua'tm.iocmdmca com Thi nghi<$mti6nhanhthasanxua'tnudcm:lmb~ngphuongphapsadvng cae10<;1iproteasekhacnhaud€ thuyphancacom(jquimonhovdicom§:u20kg, caethongso"khactuongt\1'nhucaek6tquadffHmduQC(j tren.Sau21ngaythuy phaneacomb~ngproteaseB.subtilisS5vacaeeh6ph:1mproteasekhae,thudieh nu'dem:lmeacom.K6t quathaphantichthanhph~naxitamintrongnudcm:lm cacomsanxua'theophuongphaptruy€nth6ngkeDrutmQtl~nsau3thangva cac10<;1inudcm:lmcacomsanxua'tb~ngphuongphapsadvngcac10<;1iprotease khacnhaukeDrutmQtl~nsau21ngayduQcth€ hi<$ntrenbang3.28. K6t quaphantich hamluQngaxit amin(j bang3.28cho tha'ykhi dungcac 10<;1iproteasekhac nhaud€ sanxua'tnudcm:lmthl ham luQngaxit amintrong nudcm:lmthuduQcclingkhacnhauva luaucaDhonm§:utruy€n th6ng.M§:usan xua'tnudcm:lmsa dvngca ca comxay thuyphanb~ngproteaseB.subtilisS5 co hamluQngaxit aminthu duQccaDnha't.N6u x6p thu t\1'cha'tluQngnudcm:lm theohamluQngaxitaminthuduQcthlm§:u3>m§:u7 >m§:u5>m§:u9 >m§:u6 >m§:u10>m§:u3 >m§:u8 >m§:u2 >m§:u1.Nhu v~ysanxua'tnudcm:lmtheo phuongphaptruy€n th6ngvila lau mahamluQngaxit aminthuduQctrongnudc m:lml<;1ikhong caD;nudc m:lm san xua'ttheophuongphap sa dvng protease B.subtilisS5co hamluQngaxitamincaDhonm§:ukhongsadungenzymevacaD honcacm§:uthii'nghi<$msii'd\lngmQts610<;1ienzymekhac;ca comxaytrudckhi thuyphanchonudcm:lmcohamluQngaxitamincaDhonkhongxay. 135 Bang3.28.ThanhphftnaxitamiDtrongm/oemtimeaedmsanxua'theo phlidngphaptruy~nth6ngvacaephlidngphapsll d1.lngenzyme(f.lg/ml) Ghi ehu: - Mfiu1:nuoem~meaearnsanxua'theophuongphaptruy€nth6ng, - Mfiu2:nuoem~meaearnxaysanxua'theophuongphaptruy€nth6ng. - Mfiu 3:nuoem~meaearnxaysanxua'theophuongphapdungprotease 1.subtilis55. SIT Ten axit Hamhl(.1ngaxitamin(Ilg/ml) amin Mu I Mu 2 Mu 3 Mu4 Mu 5 Mu6 Mu 7 Mu8 Mu9 Mu 10 I Alanine 128,7 119,9 221,5 136,3 179,4 214,6 248,5 124,5 201,2 74,2 Glycine 265,4 660,5 468,7 461,9 306,8 385,3 502,3 283,0 498,4 218,1 Aminobu- 99,7 201,1 142,2 191,9 142,3 111,1 132,5 125,8 254,2 99,5 tyricaxit Valine 249,2 256,9 375,5 431,7 389,5 345,2 342,1 245,5 342,3 298,0 Leucine 107,6 180,7 228,3 400,9 174,2 166,5 233,2 206,3 212,6 107,5 Isoleucine 161,0 243,6 391,0 178,7 297,8 249,1 373,7 228,4 262,3 200,4 Norleucine 49,5 120 153,6 91,6 121,9 130,I 243,5 87,9 129,9 142,4 Threonine 281,6 297,9 623,1 379,0 289,0 181,6 415,6 368,9 348,7 200,7 Serine 54,4 62,2 255,6 183,6 158,5 152,4 163,3 153,4 213,4 154,1 () Aspartic 88,6 45,4 143,2 33,5 70,3 99,2 140,4 54,6 46,7 59,6 axit 1 Methionine 114,2 93,5 361,2 151,8 201,7 227,2 512,8 147,9 339,5 147,8 t2 Hydroxy 157,5 129,5 224,7 157,3 258,3 114,1 242,3 199,2 157,3 130,8 I Proline b Phenyl 204,5 106,1 282,3 114,5 271,3 125,7 264,5 244,1 241,5 85,5 -alanine 14 Glutamic 171,2 185,4 87,1 172,2 267,6 307,6 176,1 202,8 114,1 384,2 axit 15 Ornithine 38,4 33,5 45,5 36,6 42,8 46,9 52,9 41,4 48,8 38,8 16 Lysine 196,8 118,4 1328,2 408,7 759,7 623,7 703,6 571,4 782,6 414,6 17 Histidine 665,4 530,1 3126,2 1175,7 2382,2 2483,4 2353,1 564,8 1957,7 1100,2 18 Tyrosine 373,1 215,3 587,4 169,3 595,6 552,1 445,4 285,8 478,3 478,4 9 Tryptophan 155,2 167,4 958,7 133,4 402,5 367,2 638,6 498,1 352,4 328,4 TangcQng 3562,0 3767,4 10004,0 5008,6 7169,1 6882,9 8175,4 4633,8 6981,9 4672,2 136 - M~u4: mtdcm~mca cdmnguyenconsanxuc1ttheophudngphapdung )roteaseB.subtilisS5. - M~u5:nudcm~mcacdmxaysanxuc1ttheophudngphapdungNFD. - M~u6:nudcm~mcacdmxaysanxuc1ttheophudngphapdungNeutrase. - M~u 7: nudc m~m ca cdm xay san xuc1ttheo phudng phap dung <lavourzyme. - M~u8:nudcm~mcacdmxaysanxuc1ttheophudngphapdungprotease LoryzaeS4. - M~u9: nudcm~mca cdmxay sanxuc1ttheophudngphapdungh6nhQp lroteaseB.subtilisS5vaproteaseAoryzaeS4. - M~u10:nudcm~mcacdmxaysanxuc1ttheophudngphapdungbromelain lrdua. D€ minhchungthemchocacktt quanghiencUutren,titn hanhthttnghi~m t'td\lngproteaseB.subtilisS5trongsanxuc1tnudcm~mvdi qui mom~u1.000kg ~iXi nghi~pThuy sanNha Trang- Congty Thuy sanKhu vl1cII d~tt'.li584Le 16ngPhongNha Trang, cunghie titn hanh 3 m~uthi nghi~mva lc1yktt qua :ungbinh.M~uthi nghi~mva m~ud6i chungd€u sttd\lngmQtlO'.lica cdmco ha'tluQngnhunhau.M~uthttnghi~mduQctitp nhi~tb~ngcachphdin~ngktt 9Pvdi danhdaDm6i ngay 3 l~n,keDrut lien t\lC,con cac thongso'khacgiG' lingnhuktt quatill duQCtrongphongthinghi~m.M~ud6i chungsanxuc1tnudc 1~mtheophudngphapgai nenkeDruttruy€n th6ngchomu6imQtl~nvdi ty l~ 5%so vdi ca, titp nhi~tb~ngphudngphaptl1nhien,keDrut lien Wc. Ktt qua anhgia nudcm~msan xuc1theohai phudngphapduQctrinh bay trongbang .29vabang3.30. Bang 3.29.Di~mtrung blnhearnquaDmioemiim STT Tenchitieu Mu thttnghim Mu d6ichung 1 Mu sc 4,2 5,0 2 DQtrong 5,0 5,0 3 Mui 3,6 5,0 4 V 5,0 4,2 137 Bang3.30.Sosanhke'tquasanxua'tmiocmiim Thongs6ky thu~t Kh6ihiQngea(kg) So'm§u Tyl~mu6ib6 sungbandffu(% sovoi ca) Thaigianehornu6i Tyl~eh€ phcimproteaseB.subtilisS5 (%sovoi eanguyenli~u) Nhi~tdQthuyphan Thaigianthuyphan(ngay) Thai gian eua rnQtehu ky eh€ bi€n (ngay) Lu<,1ngmioee6t(lit/l000kgea) Lu<,1ngmioe m~m thu phcim (Ht/lOOOkgea) Hamlu'Qngcae lo"ti d(;lmtrongnu'oe mc1m(tinh trungblnh eho t1t ea cae lo<;linu'oem~m): - Niwt6ngs6(gN/lit) - Niw axitamin(gN/lit) - NNH3(gN/lit) Tr(;lngthai earnquail euanu'oem~m c6t M§u thii'nghi~m 3.000 3 5 Lue6gia 0,2 Nhi~tdQthu'ang va ti€p nhi~tb~ng phdin~ng 21:t2 35:t5 630:t20 470:t20 23:t 0,15 15,26:t0,15 4,18:t0,10 Trong su6t, mffu eanh dan, mlii nu'oem~mvila, vt ngQtco h~u M§u d6ichung 2.000 2 25 Ngaybandffu 0 Nhi~tdQthu'ang va ti€p nhi~tb~ng phdin~ng 120:t7 180:t 15 530:t20 540:t20 18,28:t 0,15 12,86:t 0,15 3,88:t 0,10 Trong su6t, mffu eanh dan, mlii nu'oem~mr1td~e tIling, vt ngQtco h~u DQm~neuanu'oern~m(gNaCl/llt) Hamlu'Qngaxit trongnu'oem~me6t (tinhtheog axitaeetie/lit) Til k€t quanghieneUu0 trenehoth1ym§u thii'nghi~msii'dl,mgeh€ phcim proteaseB.subtilisS5 de sanxu1tnu'oem~meaedmco thai gianeh€ bi€n ng~n 265:t 5 4,67:t0,15 265:t 5 4,45:t0,15 138 honva 111<;5ngm/Dema:me6tthudU<;5eeaoga'p1,18l§n so vDi phuongphilpsan xua'tnUDema:mtruy~nth6ng.Ke'tquanay eo th~giai thich la do taedQngeua proteasebe)sunglam thiteabi thuyphannhanhva tri~td~bon.Ngoaira ke'tqua danhgia earnquail nUDema:meonehotha'ym~unUDema:mthli'nghi~meo mui cuanUDema:mkernnUDema:msanxua'theophuongphilptruy~nth6ng.VI v~y dn phaitie'ptl;lenghieneUueai thi~nmui euanUDema:msanxua'ttheophuong phapnay. 3.4.3.3.D~xua"tqui trinh sanxua"tmiocm~m Tli eaeke'tquanghieneUud trenehophepd~xua'tquitrlnhsanxua'tnUDe ma:mnga:ngaytlieaearnnhusau: Muoian Nudemuoi22°Be Nguyenlit%u Xaynho Thuyphan ProteaseB.subtilis55 Keo rut,lQe Nudecot Bil 1 Diehehie't1 Bil 2 Phabil Xaem~m Phanbon Hlnh3.65.Quytrinhsanxua'tmiocm~mcacom 139 . Thuye'tminhquytrlnh: -Nguyenli~u:SITdl,mgcacomtuoi,saudorITas;:lchvaxaynhc>. - Mu6i an SITdl;1ngtrongche'bie'nIa lo;:limu6ikho, tr~ngs;:lchdffduQcbaa quankhoang3+4 thangtrudckhi SITdl;1ng. I - Thuy phan: b6 sung che'ph~mproteasetu B.subtilis 55 san xua'ttheo huongphapnuoi banr~n,vdi ty l~0,2%so vdi ca r6i daDtrQnd~uva choVaG 'thungchuQpco n~pd~yd~khoang6 giOthl b~td~ucho mu6i l~nd~uvdi ty l~ '5%sovdi nguyenli~uca. Phoi n~ngva danhdaDca m6i ngay3 l~nva Cll6 giO ~chothem5%mu6ianchotdikhi dumu6ithlchoVaGb~IQcdffd~plu san. - KeorutvaIQc:chuQpchinkeDrutnudcm~md~thunudem~me6t.Nude m~me6tcoth~choeh;:lyquacaethimgehuQpehe'bie'ntheophuongphaptruy~n th6ngho~ephada'uvdinudem~me6ttruy~nth6ngd~lamtangmliivi euanude m~m.Bii conl;:litie'ptl;1ephabii gi6ngnhuphuongphaptruy~nth6ngdfftrlnh baytrongph~nt6ngquan.Hinhanhv~nudem~meacomsanxua'theophuong phapSITdl;1ngproteaseB.subtilis55duQeth~hi~n(jhlnh3.66. AA NuOc mam c3((II. slid,!,,!!'" ~~ 111""'n~ SOOmI 1Ii'~ sanphiimdu<!CsanSujl'~ D!sch1>\ 'liCOJnsan xua't IDeo ptllraJ( 'dw1gprotease B.Subtilis S5 ~NU)c MAM ngon Ttnhkh" \~llIioh1tnh12thang.. u" UIII"Xjnghi~pChtblt".1Ii~ S646~H6ngPho".~- ~'" IDnh3.66.Nli~cmAmcacdmsanKuiltbiingphlidngphapsO'd"ngprotease B.subtilisSS