TRNG I HC NNG NGHIP H NI VIN O TO SAU I HC KHOA CNG NGH SINH HC

Tiu lun:

CH PHM VI SINH VT CI TO T

GV hng dn:

TS. NGUYN VN GIANG NGUYN KHNH TON NGUYN NGC TUN CNSHA K20

Hc vin thc hin:

Lp:

H Ni, 2012
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M U t c tnh m v lc v vy c vai tr quan trng trong vic ngn chn s phn tn ca cc cht nhim. Tuy nhin vi s pht trin ca cng nghip ho hc v cc ngnh cng nghip nh: khai khong, ch to my, cng nghip sn... S pht tn ca cc cht nhim vt qu kh nng t cn bng ca t gy nn hin tng tch t v lm nh hng xu n h sinh thi. Trong s cc cht gy nhim t trng ngi ta quan tm nhiu n cc kim loi nng, cc thuc ho hc bo v thc vt hu c. Ti sinh t nhim bng phng php sinh hc khng ch gii quyt v mt mi trng m cn c tc dng nng cao nng sut v cht lng cy trng. Chng ta u bit, cc acid hu c c th ho tan v lm linh ng hn cc hp cht kim loi nng khng tan. Trong t nhin mt s vi sinh vt vng r cy trng c kh nng sn sinh ra cc acid hu c v tao phc vi kim loi nng hoc cc kim loi c hi vi cy trng (nhm, st...), mt s khc c kh nng phn hu hp cht ho hc ngun gc hu c. Cng ngh vi sinh vt trong ci to t b nhim l s dng cc loi vi sinh vt c kh nng phn gii hoc chuyn ho cc cht gy nhim trong t qua tao li cho t sc sng mi. Ngoi ra, cc vi sinh vt s dng cn c kh nng phn hu cc ph thi hu c cung cp cc cht dinh dng cho cy trng, ng thi gip cy chng li cc tc nhn gy bnh ngun gc t t, to ra cc cht kch thch sinh trng thc vt lm n nh cu trc t vng r cy trng. Cc vi sinh vt thng dng trong ci to t thoi ho, t c vn do nhim c th k n l nm r ni cng sinh (VAM-Vascular Abuscular Mycorhiza) v vi khun Pseudolnonas. Sn phm Agrobacter sn xut c t 2 loi vi sinh vt trn c nghin cu th nghim s dng nhiu ni trn th gii. Kt lun cho thy c th khi phc vng t phn mn, vng t b nhim kim loi nng hay cc vng ct ang b sa mac ho bng ch phm vi sinh ny. Nghin cu sn xut v ng dng cc ch phm vi sinh vt ti sinh phc hi t c vn v nng cao ph ca t ang c y mnh nhiu nc trn th gii, trong c Vit Nam.
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I. T V H VI SINH VT TRONG T 1. t: t l cc vt cht nm trn b mt Tri t, c hnh thnh qua nhiu qu trnh sinh hc, vt l v ho hc. C kh nng h tr s sinh trng ca thc vt ng vt v l mi trng sinh sng ca vi sinh vt. H sinh thi t l mt th thng nht bao gm cc nhm sinh vt sng trong t. C quan h tng h ln nhau di tc ng ca mi trng sng, c s trao i vt cht v nng lng. Trong h sinh thi t, vi sinh vt ng vai tr quan trng, chng chim i a s v thnh phn cng nh s lng so vi cc sinh vt khc. Trong t c cha: Cht hu c: l ngun thc n cho nhm sinh vt d dng. VD nhm vsv chuyn ho cc hp cht carbon, nit hu c. Cht v c: l ngun thc n cho nhm sinh vt t dng. VD nhm vsv chuyn ho Fe, S, P 2. H vi sinh vt trong t

T l trung bnh cc loi vi sinh vt trong t

vi khun x khun vi nm 1% 1% 8% to, ng sinh v

90%

T l ny thay i tu theo cc loi t khc nhau cng nh khu vc a l, tng t , ch canh tc II. HNG S PHN B VI NM TRONG T V CC YU T NH

Vi nm c kch thc rt nh, d dng pht tn nh gi, nc, v cc sinh vt khc. c bit l nhng vi nm c bo t, bo t ca chng c kh nng sng tim sinh trong cc iu kin kh khn. n khi gp iu kin thun li, bo t ny mm v pht trin.

Vi nm thng tp trung tng mt v tng ny c nhiu oxy. Cng xung su, mt cng gim do hm lng oxy gim. Lng vi sinh vt trong t theo chiu su ( t canh tc) Chiu su t (cm) 3-8 20-25 35-40 65-75 135-145 Vi khun 9.750.000 2.179.000 570.000 11.000 1.400 X khun 2.080.000 245.000 49.000 5.000 Nm mc 119.000 50.000 14.000 6.000 3.000 Rong to 25.000 5.000 500 100

nhng t y cht dinh dng, thong kh tt, nhit , m, pH thch hp th vi nm pht trin nhiu v s lng v thnh phn. S lng v thnh phn vi nm trn mt t t do b mt t b mt tri chiu ri, lm cht v lm gim m cn thit cho s pht trin thun li ca vi nm. Khi chiu su t t 5-10 cm so vi b mt, tng t ny m thch hp, cc cht dinh dng tch lu nhiu, khng chu s tc ng ca mt tri nn nm pht trin nhanh, cc qu trnh chuyn ho quan trng cng din ra tng t ny. Nm cng nh cc loi vi sinh vt khc tp trung phn gn r v r cy thng xuyn tit ra cc cht hu c lm cht dinh dng cho vi sinh vt. R cy

lm cho t thong kh, gi c m. Ngoi ra, r cy cng cung cp mt lng cht hu c kh ln khi n cht i. III. LI CH CA NM

Bn cnh tc ng gy hi, mt s loi nm mc rt hu ch trong sn xut v i sng nh nm n, nm dc phm (nm linh chi, Penicillium notatum tng hp nn penicillin, Penicillium griseofulvum tng hp nn griseofulvin...), nm Aspergillus niger tng hp cc acit hu c nh acit citric, acit gluconic, nm Gibberella fujikuroi tng hp kch thch t gibberellin v mt s loi nm thuc nhm Phycomycetina hay Deuteromycetina c th k sinh trn cn trng gy hi qua c th dng lm thin ch dit cn trng. Ngoi ra, nhng loi nm sng cng sinh vi thc vt nh Nm r (Mycorrhizae), gip cho r cy ht c nhiu hn lng phn v c kh tan v cung cp cho nhu cu pht trin ca cy trng.Nm cn l i tng nghin cu v di truyn hc nh nm Neurospora crassa, nm Physarum polycephalum dng tng hp ADN v nhng nghin cu khc. Trong qu trnh phn gii cht hu c, nm mc pht trin mt h khun ti kh ln trong t. Khi nm mc cht i, vi khun phn hu chng thnh cc cht do c kh nng kt dnh cc ht t vi nhau. Vi nm sn sinh ra cc enzyme nh cellulose, protease phn gii cellulose v protein to thnh cc sn phm lin kt cc ht t vi nhau to nn cu trc t. Ngoi ra, nm mc c kh nng phn gii cc hp cht hu c nh cellulose, tinh bt, lignin, pectin. to thnh mn. Mn khng nhng l ni tch lu cht hu c lm nn ph nhiu ca t m cn l nhn t t nn cu trc t.

IV.

VAI TR CA VI NM TRONG CI TO T:

VSV sng vng r cy c kh nng sn sinh ra cc axit hu c v to phc vi kim lai nng hoc kim lai c hi vi cy trng ( nhm, st .. ), mt s vi sinh vt khc c kh nng phn hy hp cht ha hc c ngun gc hu c. Cc vi sinh vt c kh nng phn gii hoc chuyn ha cc cht gy nhim trong t, qua to li cho t sc sng mi. Ngai ra, cc vi sinh vt s dng cn c kh nng phn hy cc cht ph thi hu c, cung cp dinh dng cho cy trng, ng thi gip cy tng kh nng khng bnh do cc tc nhn trong t gy ra.
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Phn hy cc hp cht hu c: trong t thng tch t nhiu hp cht hu c dng polymer ch yu l cellulose, pectin, tinh bt Nu khng c qu trnh phn gii ca vi sinh vt th lng cht hu c khng l ny s trn ngp tri t.Vi sinh vt gp phn ng k vo s chuyn ha cc cht l h vi nm. 1. Thy phn cellulose: Nhiu loi vi sinh vt trong t c kh nng phn hy cellulose, trong s th h vi nm tit ra lng cellulase nhiu nht. Mi loi s tit ra mt loi enzyme trong h enzyme cellulase, chng phi hp nhau phn gii c cht. H enzyme cellulase bao gm 4 enzyme khc nhau. Enzyme C1 c tc dng ct t lin kt hydro, bin dng cellulose t nhin c cu hnh khng gian thnh dng cellulose v nh hnh, enzyme ny gi l cellobiohydrolase. Enzyme th hai l Endoglucanase c kh nng ct t cc lin kt - 1,4 bn trong phn t to thnh nhng chui di. Enzyme th 3 l Exo - gluconaza tin hnh phn gii cc chui trn thnh disaccarit gi l cellobiose. C hai loi enzym Endo v Exo gluconase c gi l Cx. Enzym th 4 l - glucosidase tin hnh thy phn cellobiose thnh glucose. i din cho h vi nm thy phn cellulose l Trichoderma, Aspergillus, Penicillium.

Cellulose

Monosaccharide
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2. Thy phn pectin: Pectin l hp cht polygalacturonic, cu to gm n acid galacturonic lin kt nhau bng lin kt 1,4 , c trong qu, c, ht, thni din l Mucor stolinifer 3. Thy phn tinh bt: Tinh bt ( c nhiu trong xc b thc vt : c, qu ) cu to t n glucose bng lin kt 1,4 v 1,6 glycoside. Nm Aspergillus niger, Asp.cadidus,Asp.oryzae c kh nng amylase phn ct tinh bt thnh cc n phn glucose, lm tng ph ca t. Ngoi ra h vi nm cn tham gia vo chu trnh chuyn ha cc v c trong t cho thc vt s dng v gp phn ci to t. - Chu trnh chuyn ha ln: Trong t, phosphor v c thng dng kh tan: ferrophosphate, b xng, apatite, phosphate bicalcit, phosphate st/ nhm . Vi sinh vt c kh nng chuyn ha cc hp cht trn thnh dng tan c d dng cho cy hp thu. i din l: Aspergillus, Pencillium. - S chuyn ha Mn, K: trong t nhin Mn tn ti 2 dng: Mn2+ / Mn3+ ( MnO2 ). Mn2+ tan trong nc c cy hp th. iu kin acid ( pH = 5,5 ): kh Mn3+ Mn2+ iu kin kim ( pH = 8,0 ): oxy ha Mn2+ Mn3+ i din cho vi nm l: Streptomyces

V. QUY TRNH CHUNG NHN GING V SN XUT VI NM CI TO T: Phn lp

Tuyn chn ging

Nhn ging

Ln men

Thnh phm - Phn lp: phn lp cc mu vi sinh vt thu c trong t t nhiu vng khc nhau. - Tuyn chn ging: chn nhng dng vi nm mong mun ( v d: vi nm tit ra cellulase c hot tnh cao, chuyn ha c P, Mn, K, tc sinh sn cao). Mt trong cc phng php tuyn chn l th hot tnh ca enzyme do vi nm tit ra - Nhn ging: chn mi trng thch hp v to iu kin cho vi nm pht trin - Ln men: VI. HIU QU CA CH PHM VI SINH VT I VI NGNH NNG NGHIP: - Cn bng h sinh vt trong mi trng sinh thi. - Tng ph nhiu cho t. - Ci to t, tng nng sut v cht lng nng sn. - C tc dng tiu dit su hi, cn trng. - C kh nng i khng, tng sc khng ca cy trng. - Phn hy, chuyn ha cc cht hu c bn vng, ph thi cng nghip lm sch mi trng.

VII. SN PHM C TH: 1. Danh mc phn bn c php sn xut, kinh doanh v s dng Vit Nam:[4] Ngun Tn thng Thnh phn v hm Nhm n gc S TT thng/Tn lng cc cht dinh phn v tnh phn nh thng mi dng chnh bn bn
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quyt

1 EM - MX

Cng ty Bacillus sp: 1x109 TNHH Rhotopseudomonas: sn xut Quyt nh s Phn 1x106 Lactobacillus sp: CFU/g v 40/2004/Qvi sinh 1x107 Sacchromyces thng BNN sp: 1x106 mi Mai Xun

Cng ty Bacillus sp: 1x109 TNHH Rhotopseudomonas: sn xut Quyt nh s Phn 2 Vi.EM - MX 1x106 Lactobacillus sp: CFU/g v 40/2004/Qvi sinh 1x107 Sacchromyces thng BNN sp: 1x106 mi Mai Xun Cng ty TNHH sn xut Quyt nh s Phn % v 40/2004/Qvi sinh thng BNN mi Mai Xun

3 Tricho-MX

m: 30

4 Tricho-MX

Cng ty TNHH Trichoderma sp: 1x109 sn xut Quyt nh s Phn Streptomyces sp: 1x107 CFU/g v 40/2004/Qvi sinh Bacillus sp: 1x 108 thng BNN mi Mai Xun

Vin C Azotobacter vinelandii: in 2x109; Bacillus subtilis Nng Trichodermin BS 16: 2x109; Quyt nh s nghip Phn 5 chuyn cho Trichoderma Cfu/g 40/2004/Qv Cng vi sinh la harsianum: 2x109; BNN ngh Azospirillum Sau thu brasilence: 2x109 hoch Trichodermin 6 chuyn cho la Vin C in Quyt nh s Nng Phn % 40/2004/Qnghip vi sinh BNN v Cng ngh
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HC: 25

Sau thu hoch (Azotobacter, VSV C N Azospirillum, 7 cho la Flavobacter, Bacillus) >5.106 Vin KHKT Quyt nh s Nng Phn % 40/2004/Qnghip vi sinh BNN Vit Nam

Hu sinh

Cng ty P2O5(hh): 2,8 Mn: 10 Ho Quyt nh s vi Axit Humic: 2 % Phn cht 40/2004/QVSV(N,P): 1.107 mi CFU/g vi sinh Qung BNN chng Bnh Vin C in Phn Nng Quyt nh s vi nghip v 67/2007/Qsinh Cng BNN vt ngh Sau thu hoch Vin C in Phn Nng Quyt nh s vi nghip v 67/2007/Qsinh Cng BNN vt ngh Sau thu hoch Vin C in Phn Nng Quyt nh s vi nghip & 84/2007/Qsinh Cng BNN vt ngh Sau thu hoch

Trichodermin 9 9 chuyn cho la

HC: 25

Azotobacter vinelandii: 2x109; Bacillus subtilis BS Trichodermin 16: 2x109; 10 Cfu/g chuyn cho la Trichoderma harsianum: 2x109; Azospirillum brasilence: 2x109 Azotobacter vinelandii: 2x109; Phn bn vi Bacillus subtilis BS sinh a chng 16: 2x109; 11 Cfu/g R3 chuyn cho Trichoderma la harsianum: 2x109; Azospirillum brasilence: 2x109 12

Lactobacillus Cng ty Phn Quyt nh s sporengenes: 1x107 TNHH vi Maruzen cfu/ml 10/2007/QPhotosynthetic Thc n sinh BNN bacteria: 1x107 Thu sn vt
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Pseudomonas sp: 1x107 Streptomyces saraticus: 1x107

Vit Thng

2. Tam Nng Trichoderma:[3] Ch phm sinh hc nm i khng Trichoderma ngai tc dng sn xut phn bn hu c sinh hc, hay s dng nh mt lai thc BVTV th cn c tc dng x l phn chung, phn gia sc, v c ph, cht thi hu c nh rm, r, rc thi hu c rt hiu qu.

TAM NNG TRICHODERMA C.68 Mi gam ch phm ny c hai triu bo t ca loi Trichoderma konigii, c kh nng phn gii nhanh cht x trong rm r, xc b thc vt v phn chung. Ha tan 1 kg ch phm ny trong 600 lt nc x l 4 5 khi cht hu c. Ti u trn tng lp cht hu c dy 10 15cm, y kn trong vng t 1 2 tun, gi m 60% x khun phn gii hon ton cht hu c thnh cht mn.
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Tri rm u trn mt rung mi gt. Ha 1kg ch phm Tam Nng Trichoderma v 320 lt nc phun cho 1 ha. Khong 5 7 ngy sau khi x l, cho nc v ngp gc r, ngm trong thi gian 1 tun, sau lm t gieo s v mi. Ha tan 1 kg ch phm ny trong 600 lt nc x l 4 5 khi v c ph kh. Ti u trn tng lp v dy 10 15cm, y kn gi m 60% trong 1 2 tun. Sau phun nc va m, o trn u v tip tc kn cho n khi v c ph phn gii thnh cht mn. i vi loi xc b hu c cn ti nh thn bp, t ma, dy thanh long Rc ch phm theo tng lp xc b dy t 10 20cm, y kn v gi m cht hu c phn gii hon ton thnh cht mn. Dng 1kg ch phm x l 4 - 5 khi cht hu c. i vi phn chung kh: Ha tan 1 kg ch phm ny trong 600 lt nc x l 4 5 khi. Cch x l nh i vi v c ph. i vi phn chung ti: Rc ch phm ln tng lp phn dy 10 20cm, y kn trong vng 1 2 tun. Sau o trn, y kn v gi m cht hu c phn gii hon ton thnh cht mn.

3. Phn hu c truyn thng:[2]

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3.1 Thnh phn

Phn chung x l (g, b, ct...): 50%. Humic v cc axit hu c t mn: 25%. Trichoderma ging M8, M32, M35 B sung khong dinh dng: Ln, Kali, Canxi, Magi, St, Km, ng,

Bo... 3.2 Cng Dng - Thay th phn chung cha x l, km cht lng v cn nhiu vi sinh vt gy hi. - Cung cp v nui dng h vi sinh vt c ch: C nh m, phn gii ln, phn gii cht hu c. - c ch tuyn trng hi r, cc bnh nt thn, x m, cht nhanh, cht chm, vng l, vng cnh, ho dy... trn thanh long, cy n tri, cy cng
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nghip, rau mu, la... nh tp on nm Trichoderma c tuyn chn bng k thut ht nhn. Cung cp axit amin, axit humic gip h r pht trin mnh m, tng c kh nng hp thu dinh dng, tit kim phn bn, hn ch su bnh hi hiu qu nht tit kim chi ph sn xut. - B sung trung v vi lng cn i cho cy vng cnh, l xanh khe, hoa u, tri, ht bng p nng k. - Tng nng sut v cht lng nng sn t 15 - 30%, cy vng cnh, l xanh khe, hoa u, tri, ht bng p, nng k; cao su m nhiu, m cao. Lu : Bo qun ni kh ro, thong mt, trnh nh sng trc tip

4. Ch phm sinh hc BIMA (Trichoderma)[1]

4.1. Thnh phn: * Cc chng nm Trichoderma: 5106 bo t/gam * Hu c: 50%; m < 30%.
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4.2. Cng dng: Cha nm i khng Trichoderma c kh nng tiu dit v khng ch ngn nga cc loi nm bnh hi cy trng gy bnh x m, vng l thi r, cht yu, ho r nh: Rhizoctonia solani, Fusarium, Pythium, Phytophthora sp., Sclerotium rolfsii, To iu kin tt cho vi sinh vt c nh m pht trin sng trong t trng. Kch thch s tng trng v phc hi b r cy trng. Phn gii tt cc cht x, chitin, lignin, pectin trong ph thi hu c thnh cc n cht dinh dng, gip cho cy hp thu c d dng. Kt hp vi phn hu c c tc dng ci to t xp hn, cht mn nhiu hn, tng mt cn trng c ch v gi c ph ca t. 4.3. Quy trnh phn chung, xc b thc vt C 34 kg ch phm BIMA; 20 30 kg super ln trn u vi 1 tn phn chung, xc b thc vt. Phun dung dch ur (1 kg ur/100 lt nc ) vo ng cho t u, m t 5055% (dng tay vt cht hn hp trn, thy nc rn ra l c) o trn v y bt, sau 45 ngy, nhit s ln khong 60oC. Tin hnh o trn. Nu thy kh, phun nc vo to m. Sau 25 30 ngy, o li 1 ln, phun nc m bo m 50 55%. Nu phn cha hoai, tip n 30 ngy sau th phn hoai hon ton, c th em s dng. Sn phm phn hu c thu c c th trn vi phn NPK, ur, super ln, kali v cc lai tro tru.

VIII.

TIU CHUN NH GI:

TIU CHUN VIT NAM TCVN 6168 : 1996 Phn bn vi sinh vt gii xenluloza Cellulose-degraing microbial fertilizer 1. Phm vi p dng Tiu chun ny p dng cho cc li phn bn cc chng vi sinh vt sng c kh nng phn gii xenlluloza v quy nh cc yu cu k thut, phng php kim tra nh gi i vi phn bn vi sinh vt phn gii xenluloza. 2. Tiu chun trch dn
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TCVN 4833-89 (iso 4833-1978) Hng dn chung m vi sinh vt k thut m khun lc 300C TCVN 4881-89 (ISO 6887-1983 )Hng dn chung v cch pha ch dung dch pha long kim nghim vi sinh vt TCVN 5814 -1994 Phn bn hn hp NPK. Phng php th TCVN 6169-1996 Phn bn vi sinh vt. Thut ng 3. Yu cu k thut 3.1 Phn bn vi sinh vt phn gii xenluloza phi cha mt hoc nhiu chng vi sinh vt c kh nng pht trin trn mi trng cha ngun cacbon duy nht l xenluloza t nhin 3.2 Phn bn vi sinh vt phn gii xenluloza c mt vi sinh vt sng ph hp vi quy nh trong bng 1 Bng1 - Mt vi sinh vt Mt vi sinh vt CFU * /g ( hay ml) phn bn Tn ch tiu Cht mang thanh Cht mang trng khng thanh trng Khi Cui K Cui xut xng hn bo hi xut hn bo hnh xng hnh 1.Vi sinh vt phn gii 1.0 9 xenluloza, khng nh hn 10 2. Vi sinh vt tp, khng ln hn 106 1,0 106 1,0 108 1,0 0 107
-

1,

1,0 106

CFU*: n v hnh thnh khun lc 3.3 Phn vi sinh vt phn gii xenluloza phi c tc dng tt i vi t v cy trng. Hiu qu ca phn bn phi t cc ch tiu cht lng ghi trn nhn v c xc nh ti phng th nghim c cng nhn hoc ch nh 3.4 an ton ca cc chng vi sinh vt cha trong phn phi c xc nh v cng nhn ti cc phng th nghim c cng nhn hoc ch nh 3.5 Thi gian boat hnh ca phn bn vi sinh vt phn gii xenluloza khng t hn 6 thng k t ngy xut xng 3.6 Hm lng cc cht dinh dng v m ca phn bn vi sinh vt phn gii xenluloza phi c ng k ti cc c qun qun l nh nc v cht lng v c xc nh nh gi ti cc phng th nghim c cng nhn hoc ch nh
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4 Ly mu 4.1 Quy nh chung 4.1.1 Vic ly mu c tin hnh sao cho mu kim tran phi l mu i din cho c l hng. Ngi ly mu phi c hun luyn v c kinh nghim trong vic ly mu 4.1.2 Trong qu trnh ly mu, vn chuyn v x l mu, phi m bo trnh s ly nhim t bn ngoi v phi bo m l mu c nguyn trng nh ban wuf cho ti khi em phn tch trong phng th nghim 4. 1.3 Khng c b xung thm bt c mt tc nhn bo qun, dit khun hoc dit nm vo mu kim tra. 4.1.4 Mu c ly phi l cc bao nguyn gi. 4.1.5 Phi tin hnh ly mu nhng ni khng c hi nc nng, ho cht c hi, khng c nh nng gay gt hoc bi v c a ngay vo cc dng c cha mu. 4.1.6 Cc dng c ly mu v cha mu phi sch s v v trng 4.2 Chun b dng c ly v cha mu 4.2.1 Dng c ly mu phi l loi c lm t thp khng r hoc bng thu tinh 4.2.2 Cc dng c ly v cha mu sch s v v trng bng cch sy trong t sy nhit 1700C trong thi gian khng t hn 1gi hoc trong ni hp nhit 1210 C trong thi gian khng t hn 30 min v c bo qun trong cc dng c thch hp, m bo trnh ly nhim t bn ngoi. 4.3 S lng mu 4.3.1 L hng bao gm cc bao ( ti ) sn phm phn bn vi sinh vt phn gii xenluloza c sn xut cng mt t vi cng mt ngun nguyn liu. 4.3.2 S lng bao ( ti) cn ly kim tra i vi mi l hng ph thuc v ln ca l hng v ph hp vi quy nh trong bng 2. Bng 2- S lng bao ( ti ) cn ly kim tra C l hng ( bao, ti) n 100 t 101 n 1000 t 1001 n 10000 ln hn 10000 S lng mu cn ly ( bao, ti) 7 11 15 19

4.3.3 Cc bao ( ti ) mu c la chn ngu nhin theo TCVN 1694-75 Tin hnh ly mu trung bnh t mu chung l tp hp cc mu ban u trong l hng kim tra. Chia mu trung bnh lm 2 phn bng nhau ri bao gi
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ph hp vi yu cu ca sn phm. Mt phn dng kim tra v mt phn lu v bo qun trong iu kin qui nh m mi loi sn phm yu cu dng khi phn tch trng ti. Trn mi gi mu phi c nhn ghi r: tn mu v i tng cy trng c s dng; tn c s sn xut; thi gian sn xut; thi gian v a im ly mu; tn ngi ly mu , c quan ly mu. 5. Tin hnh kim tra v xc nh 5.1 Chun b kim tra 5.2.1.1 Trang thit b , dng c kim tra Trang thit b ca phng kim nghim vi sinh vt thng thng bao gm: thit b kh trng kh ( t sy ) hoc kh trng hi nc, (ni hp) t m, c th iu chnh c 30 0 C 1 0C; hp lng thu tinh hoc cht do, ng knh t 90 n 100 mm; pipet thu tinh c dung tch 1,0 ml; 5,0 ml; 10,0 ml; my m khun lc c y c chiu sng vi nn ti c gn mt thu knh phng i , phng i 1,5 ln v mt dng c m c hc hoc hin s in t; my o pH c chnh xc n 0,1 n v o pH ni cch thu n nhit; ng nghim thu tinh 18 x 18 mm; bnh tam gic c dung tch thch hp; ng ong v cc dng c thu tinh khc Cc dng c dng trong xc nh vi sinh vt phi kh trng bng cch: gi t 170 n 1750 C khng t hn 1 h trong t sy, hoc gi p sut 1,0 atmotphe ( 1210 C) khng t hn 30 min trong ni hp Cc thit b pha trn my trn quay c tn s quay t 8 000 n 45 000 vng / min, c bnh cha bng kim loi hoc thu tinh, chu c cc iu kin tit trng; dng c trn nhu ng ( stomacher) vi cc ti cht do v trng; dng c trn : c th trn t 1 ml n 2 ml mu th; cn k thut c chnh xc ti 0,01 g.

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5.2.1.2 Dch pha long c dng l dung dch mui n ( NaCl) 0,85 %, khng cha cc hp cht cacbon, c pH l 7,0 sau khi kh trng 25 0C. Ly 90 ml dch pha long vo cc bnh cu hoc l bng cht do ( cho dch huyn ph ban u ), hoc Ly 9 ml dch pha long vo ng nghim ( cho ccs dch pha long thp phn) Nt li bng bng m, kh trng trong ni hp ap lc 1210C trong 30 min. Nu cha s dng ngay, dch pha long cn c bo qun trong t lnh nhit t 0 n 50 C, thi gian bo qn khng qu 1 thng k t ngy chun b. Ch thch - trnh lm nh hng n cc vi sinh vt do thay i nhit t ngt, nn diu chnh nhit ca dch pha long n nhit phng th nghim 5.2.1.3 Chun b mi trng kim tra Mi trng dng kim tra phn vi sinh vt phn gii xenluloza c s dng l mi trng cha ngun cacbon duy nht l xenluloza t nhin, thnh phn mi trng ph thuc vo chng loi vi sinh vt m nh sn xut s dng. Nu khng c yu cu ca nh sn xut, khi kim tra s dng mi trng theo ph lc A. Mi trng c pha ch theo th t cc ho cht trong thnh phn cho v chia vo cc dng c thu tinh chun bih trc ri kh trng iu kin 1 atmotphe ( 1210 C) trong 30 min. Phan chia mi trn vo cc hp lng kh trng iu kin v trng. Kim tra sch ca mi trng sau 2 ngy nhit t 28 n 300C. Ch s dng cc hp lng cha mi trng nui cy vi sinh vt khng pht hin thy tp nhim. Ch thch- i vi phn vi sinh vt phn gii xenluloza cha cc vi sinh vt di dng tim sinh, trc khi kim tra cn phi hot ho. 5.2.1.4 Chun b dch huyn ph xenluloza. Cn 30 g giy lc loi nh lng ( th d Whatman 1 ) x thnh mnh nh cho vo bnh thu tinh c dung tch 4 lit, thm mt lit nc ct v mt t vin bi thu tinh sao cho ngp giy. Lp bnh vo my lc, lc c hc vi tc 74 ln/ min trong khong 30 min, lng sau 24 h , cht ly dung dch huyn ph xenluloza. 5.2.1.5 Chun b dung dch ch th Benedict: Thnh phn dung dch Benedict; Natri xitrat 17, 3 g Na2 CO3 10, 0 g CuSO4. 7H2O 1, 73 G ho tan natri xitrat v Na2CO3 vo 80 ml nc m. B xung nc ct va t 85 ml ( dung dch A)
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ho tan CuSO4 vo 10 ml nc ( dung dch B) trn u dung dch A v dung dch B; b xung nc va t 100 ml. 5.2.2 Tin hnh kim tra 5.2.2.1 Pha long mu a) i vi mu dng lng: dng pipet v trng c nt bng phn nt ly ra 10 ml mu trn u bng lc tay hay thit b lc c hc v a vo 90 ml dch pha long theo 6.2.1.2. Ch trnh chm pipet vo dch pha long, trn cn thn mu chun b bng cch ht- th li 10 ln vi mt pipet c nt bng phn ht v trng khc hoc bng dng c trn c hc trong 5 10 s, nhp quay ca dng c ny c chn sao cho mu trn nh cun xoy dng ln cch mp l cha khong 2 n 3 cm. Dung dch to ra c gi l dung dch huyn ph ban u. b) i vi mu dng c bit: cn 10 g mu c chnh xc ti 0,01 g v cho vo bnh cha v trng, thm 90 ml dch pha long trong 6.1.1.2. t bnh vo my trn trong 2 n 5 min sao cho c c mt dung dch c phn b ng u, lng c phn t nng trong khong 15 min, gn c dung dch huyn ph ban u. c) dng mt pipet v trng ly 1 ml dch huyn ph ban u ( a hoc b) cho vo ng nghim cha 9 ml dch pha long ( 6.2.1.2 ) chun b sn nhit phng, trnh chm pipet vo dch pha lang. Trn k bng cch ht - th khong 10 ln vi mt pipet khc c nt bng u ht v trng, c dch pha long mu c nng l 10-2. Qa trnh ny c lpp li lin tc c dch mu c nng pha lang theo quy nh sau: i vi phn vi sinh vt trn nn cht mang thanh trng s dng nng pha long l 10 -7. i vi phn vi sinh vt trn nn cht mang khng thanh trng s dng nng pha long l 10-5. 5.2.2.2 Cy mu Dng pipet v trng ring cho tng pha long ring ly ra t dch mu c nng pha long 10-5. 10-6; 10-7 i vi mu phn vi sinh vt trn nn cht mang thanh trng; 10-3; 10-4; 10-5 i vi mu phn vi sinh vt trn nn cht mang khng thanh trng, mt lng dch mu 0,05 ml, cy vo 1 hp lng cha mi trng chun b sn theo 6.2.1.3. Mi mu c cy lp li trn hai hp lng. Nhc nh hp lng dch mu dn tri trn b mt thch , ch khng dch mu dnh vo thnh hp lng, i b mt thch kh, p ngc hp lng v a vo t m, iu chnh nhit t m cho ph hp vi yu cu ca tng loi vi sinh vt. Thi gian nui l 48 -72 h. 5.2.2.3 Pht hin vng phn gii
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Ly 1 ml huyn ph xenluloza v 1 ml dung dch ch th Benedict nh ln b mt thch cy mu ( 6.2.2.2) nhit 30 n 350 C trong 10 n 15 min. Quan st vng phn gii trong sut bao quanh cc khun lc phn gii xenluloza 5.2.3 c kt qu` Vi sinh vt phn gii xenluloza c tnh l s khun lc trong hp lng to vang phn gii ( vng trn trong sut) bao quanh khun lc. Vi sinh vt tp l tt c cc khun lc khng c c diim c trng trn 5.2.4 Cch tnh mt vi sinh vt trn mt dn v kim tr ( gam hay mililit) Mt vi sinh vt trong n v kim tra ( A) c tnh theo cng thc: a.20 A = d trong a - s khun lc theo yu cu c trong hp lng d - nng dch pha long Ch thch : 1) s lng khun lc trung bnh c tnh l trung bnh cng s khun lc ca cc hp lng c cy t cng mt pha long, tong ch tnh cc hp lng cha t 5 n 50 khun lc. 2) s lng khun lc trung bnh cng c th c tnh t trung bnh cng s khun lc ca cc hp lng c cy t hai pha long k tip nhau bng cch tnh s khun lc trung bnh ca mi pha long , trong s khun lc pha long cao hn c nhn vi 10, sau ly trung bnh cng ca hai gi tr nu trn nu t s gia gi tr ln v gi tr nh khng ln hn 2. Nu t s ny ln hn 2 th ly gi tr n lm kt qu. 3) Mt vi sinh vt trn mt n v kim tra c biu th bng mt s gia 1,00 v 9,99 nhn vi 10n, n l s m thch hp. 5.3 .Xc nh hm lng cc cht dinh dng 5.3.1 Xc nh hm lng nit ( N) Tin hnh theo TCVN 5815 -1994. 5.3.2 Xc nh hm lng photpho ( P2O5 hu hiu) Tin hnh theo TCVN 5815-1994 5.3.3 Xc nh hm lng kali ( K2O) Tin hnh theo TCVN 5815-1994. 5.3.4 Xc nh hm lng cc cht hu c v vi lng
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Tin hnh theo cc phng php v uy nh hin hnh 5.3.5 Xc nh m Tin hnh theo TCVN 5815-1994 5.4 Bo co kt qu kim tra Bo co kt qu kim tra phi m t li tnh trng mu trc khi tin hnh kim tra( cc chi tit cn v xc nh mu), cc phng php kim tra v kt qu t c. Bo co cng phi nu tt c cc iu kin thao tc khng quy nh trong tiu chun ny hoc c coi l tu chn, cng nh bt k tnh hung no c th nh hng n kt qu. 6. Yu cu bao gi, ghi nhn, bo qun v hng dn s dng 6.1 Phn bn vi sinh vt phn gii xenluloza phi c bao gi bng cc cht liu bo m khng gy c hi ti vi sinh vt, ngi, ng thc vt v mi trng sinh thi, ng thi m bo thi hn bo hnh ca phn bn trc cc iu kin bt l t bn ngoi. 6.2 Trn mi bao ( gi) sn phm phn vi sinh vt phn gii xenluloza phi c ghi nhn vi y cc ni dung sau: tn c s sn xut; tn sn phm v tn khoa hc ca loi vi sinh vt s dng; thnh phn dinh dng v m; cng dng ngy sn xut v thi gian bo hnh khi lng tnh s dng k cht lng 6.3 Sn phm phi c bn hng dn s dng km theo ( in trn bao b hoc in ring). Ni dung hng dn phi ghi liu lng v qui trnh s dng cng nh hiu qu ca phn bn i vi cy trng hay kh nng thay th cc loi phn bn khc. PH LC A ( Quy nh ) A.1 Mi trng nui cy kim tra nm phn gii xenluloza Glucoza 10,0 g Pepton 5,0 g KH2PO4 1,0 g MgSO4. 7H2O 0,5 g
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Rose bengal Streptomycin sunphar Aga Nc ct va pH

33 mg (10%) 3 ml 20,0 g 1000 ml 6,5-7,0

A.2 Mi trng nui cy kim tra vi khun, x khun phn gii xenluloza Peton 5,0 g Cao tht b 3,0 g NaCl 5,0 g Aga 20,0g Nc ct va 1000 ml pH 6,8 7,0 A.3 Mi trng nui cy kim tra vi khun, x khun phn gii xenluloza Glucoza 1,0 g K2HPO4 0,5 g Aga 20,0 g Nc chit t* 100 ml Nc ct va 900 ml Ph 6,5 -7,0 *Chun b nc chit t: - 1 kg t vn ti + 1 l nc sch; - hp kh trng 30 pht trong ni hp; - b xung 1 g CaCO3 ri lc ly nc trong. IX. TI LIU THAM KHO: 1. http://www.hcmbiotech.com.vn/production_business_detail.php?id=9 2. http://www.bannhanongvn.com/index.aspx?pid=42&lang=V 3. http://www.bannhanongvn.com/index.aspx?pid=35&lang=V 4. http://www.cuctrongtrot.gov.vn/ctt/ChuyenTrang/DMPhanbon.aspx?idn hom=19

PH LC
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I. T V H VI SINH VT TRONG T 1. t 2. H vi sinh vt trong t II. S PHN B VI NM TRONG T V CC YU T NH HNG III. LI CH CA NM IV. VAI TR CA NM TRONG CI TO T 1. Thy phn cellulose 2. Thy phn pectin 3. Thy phn tinh bt V. QUY TRNH CHUNG CA NHN GING V SN XUT VI NM CI TO T VI. HIU QU CA CH PHM VI SINH VT I VI NGNH NNG NGHIP: VII. SN PHM C TH 1. Danh mc phn bn c php sn xut, kinh doanh v s dng Vit Nam: 2. Tam Nng Trichoderma 3. Phn hu c truyn thng 4. Ch phm sinh hc BIMA (Trichoderma) VIII. TIU CHUN NH GI IX. TI LIU THAM KHO:

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