Tgp chi Khoa hgc TntdngDgi hgc Cdn Tba PhdnB: Nong nghifp. Thuy sdn vd Cong nghi Sinh hgc: 33 (2014): 53-60
Tap chl' Khoa hoc TrUcJng Dai hoc Can Thd website: sj.ctu.etju.vn
DINH DANH VA XAC DINH MOT SO DAC TINH SINH HOA CUA CAC DONG VI K H U A N LACTIC TRONG SAN P H A M M A M CHUA CA SAC D6 Thi Tuyit Nhung', Nguygn Van Thanh" va Nguyin Hiiu Hiep^
' Trudng Cao ddng Kinh te - Ky thudt Cdn Tha
^ Vien Nghien cuu & Phdt trien Cdng nghe Sinh hoc, Trudng Dgi hoc Cdn Tha Thdng tin chung:
Ngaynhdn: 20/03/2014 Ngdy chdp nhgn: 28/08/2014 Title:
Determination and characterization of biochemical parameters of lactic acid bacteria in sour fermentedflsh
Tit khoa:
Cd sgc, len men, mam chua, vi khudn lactic, xuang mem Keywords:
Fermention, gourami, lactic acid bacteria, soft bone, sour fermented flsh
ABSTRACT
The iradilional product, named "Mdm chua cd sgc" or "Mdm tiiu xuang". is a sow- fermented soft bone flsh (Trichogaster trichopterus; Trichogaster microlepis). Fish was mixed with salt, sugar, ethanol, grilled rice powder, garlic, chilli and naturally fermented at 28-30°C within 30 days Fermented products had the harmonious combination of sweet, sour and salty taste; especially, they still remained the whole shape offish but their skeleton turned really soft that it was boneless in the sense of taste.
One of the factors effecting the quality oflhe product was the source oflc^tic acid bacteria presented to products. Ther^ore, it was necessaiy to isolate and determine lactic acid bacteria based on the characteristics of form, biochemistry and molecular biology techniques (DNA sequencing. These experiments were set up based on the basis of development of procedures for processing such as inoculating the bacteria into the products ...In this study, bacteria were isolated in fermented products on MRS medium with 6% of salt added As a result, four isolated strains of bacteria including Ll, L2, L3, L4 had whiter or milk-white colonies; l-2mm in length; bacilli, diplococci and cocci in shape. All these four strains were Gram positive bacteria Their reaction ofcatcdase, oxidase were negative and positive to amylase; and especially, Ll, L2 and L3 indicated positive results in protease. The results ofMSrRNA sequences ofLl, L2, L3. L4 showed thai they were Pediococcus acidilactici. Lactobacillus farciminis and Staphylococcus hominis respectively. Among them. Staphylococcus hominis M'OS not lactic acid bacteria
^ .TOM TAT
"Mdm chua cd sgc " hiy "Mdm tiiu xuong" Id ten ggi ddn gian cua dgng sdn phdm cd sgc lin men. Cd sgc dugc phdi trgn cimg vdi mudi, dudng, rugu, thinh, ^mg. tdi. dl vd tiin hdnh len men tu nhiin a nhiet dg 28-30'C trong thai gian khodng 30 ngdy. Mgt trong nhirng yiu Id dnh hudng din chdt lugng sdnplidm (miii, vi, cdu tnic,...) do chinh Id ngudn vi khudn lactic cd trong sdn phdm. Vi vdy vi^c phdn lap vd xdc dinh lodi vi khudn lactic dua trin dgc diim vi hinh thdi, dgc diem sinh hda vd sinh hoc phdn lu (gidi trinh tu gen) Id rdi cdn thiet di ldm ca sd cho cdc nghiin cuu cdi tiin quy Irinh che biin nhu chiing gidng vi sinh vdt vdo sdn phdm, ...lien hdnh phdn lap vi khudn hiin diin trong sdn phdm trin mdi tnrdng MRS cd bd simg 6% mudi Bdn ddng vi khudn Ll, L2, L3, L4 dugc phdn lap vdi cdc dgc diem khudn lgc mdu trdng hodc trdng sira. dudng kinh khodng 1-2 mm; ti bdo hinh que. hmh edu ddi hogc cdu dan Cd bdn ddng vi khudn ngy deu Id vi kliudn Gram duang, clio kit qud din tinh vdi enzyme a^alase, enzyme oxidase: duang tmh vdi enzyme amylase; ciu co ba ddng Ll. 12, L3 duang tinh vdi enzyme protease. Bdn ddng ndy eo mirc do luang ddng vi trmh tu gen 16S rRNA trin 99% vdi cdc ddng vi sinh vol: Pediococcus acidilactici, Lactobacillus farciminis vd Staphylococcus hominis. Trong do Staphylococcus hominis Ididng phdi Id vi khudn lactic.
Tgp chi Khoa hgc Tntang Dgi bgc Cdn Tho
GIOI THIEU
Cd len men Id radt ttong nhimg sdn phdm lln men truyen thdng lau ddi vd rat dac trung d cae qudc gia Ddng Nam A. Cd lln men cd vai ttd quan ttgng ttong biia an ciia ngudi ddn nhdt Id ngudi dan ngheo do cd gia thinh re Iai cung cip tfalm acid amin dugc thiiy phin tii protein ca vd cd huong vi dgc dao (Hall, 2011). (3 nude ta sdn pham ca lln men dugc ggi la "mam". Hau hit nhiing sin phim mam len men tir ca nhat Id san pham len men tir cac logi ca nhd nhu cd sac, cd tten,... cdc logi mam nay thudng dugc diing chu yeu dl nau lay huang vi nliu nau lau, kho, chung,.. it khi dugc dimg phd bien dl "in sdng" (khdng qua ndu) vi thit it, xuang nhilu vi ciing, thai gian san xuat ra tiianh pham kha ddi (tii vdi tfadng ttd lln).
"Mdm chua cd sdc" ciing li dgng sin pham len raen ci. Net ddc trung cua sdn phim niy la ed vin giii dugc hinh dgng nguyen ven nfaung xuang rat mem tao cim giic nhu khdng cd xuong khi in; mui vi tham ngon, hii hda han cdc sdn phim mam thdng thudng khac nin khdng cdn phdi chi lai trudc kfai an va rit tfaich hgp ding "an sdng", dac biet han la thdi gian len men ngdn khoang 30 ngdy.
Day cung la dang sin phara len men lactic (Do Thi Tuyet Nhung et al. 2011).
Pban B: Nong nghifp, Thiiy sdn vd Cdng nghf Sinh hgc- 33 (2014)' 53-60
dd ed nhiing djnfa faudng mang tinfa khoa fagc cho vile toi uu fada quy trinh che biin vd cd the ling dyng vdo san xudt di cho ra san phdm cd chat lugng dn dinh, khdng phy thudc nhiiu vio dieu kiln tu nhien.
Hinh 1: San pfadm mam cfaua cd sac Chup ngdy 17/10/2010
Tuy nfailn, bi quyit lira sin phim ndy dang din that ttuyen do mang tinh chdt gia truyin ciia gia dinh. Ddng thdi, niueu yiu td tdc d^ng lln san phara chua dugc nghien ciiu va xdc dinh rd ring ddn den chit lugng san phdm khdng on dinh. Ddi vdi san phdm len men. vi sinfa vat li mgt trong nhiing nhan td quan ttgng ed dnh hudng rit Idn din qud ttinli phdn giai sdn phira. Vi sinh vdt tham gia chu yeu vao qud ttinh phdn giii san pham mam cd li vi khuin vdi cic gidng chii yiu: Pediococcus, Lactobacillus, Streptococcus, Staphylococcus,....
Den thdi diem hien nay chua co nhiiu cdng ttinh khoa hgc cdng bd nfaiing vin dl cd liln quan den sin pham mam chua cd sac. Vi vdy, vile phan lap cdc gidiig vi kfauin thara gia vdo qud trinh phdn gidi la rit can thiet nham cung cip thira thdng tin klioa hgc vl sin pham mim cua Viet Nam, phuc vu cho ngfailn ciiu, gidng dgy. Ddng thdi, trin ca sd
2 PHU'ONG PHAP NGHIEN CUtJ 2.1 Vgt lilu
Nguyin lieu: ca sgc loai cd kich tiiudc viia (khoang 15-20g/con), dugc tiiu mua d Long MJ - Hiu Giang.
Hda chdt: mdi truang de Man Rogosa and Sharsp (MRS agar va broth) cua An Dg, Sodium cloride (NaCl > 99,5yo) eiia cdng ty Guangdong- Trung Qudc, Tettamethyl - p - phenylenediamirffii;
dihydrochloride 1% vd mdi ttudng Luria-Bertaiu"^
(LB) ciia Merck, Tfaermo Scientific GeneRuler' I OObp plus DNA - ladder - USA.
2.2 Pfammg p h i p
Chuan bi mdu: xOr Iy nguyen lieu ca sac (edt b6 viy, ndi tgng, ngdm ttong nudc, sau do cha vay va dk rio vdi tdng thdi gian xii 1^ Id 25 gid); phdi tr^n cimg vdi 10% muoi (cd do dm 1-1,5%), 15%
duang (cd do am 3-4,5%), 1% edn thuc phim (99,5"). 5% tiiinfa (cd do dm 1-1,5%), 10% (gimg, tdi, dt); cho vdo hd, d^y kin; u d nhilt dg phdng tii 28-30°C. Sau 10, 20, 30 ngdy u, phin lap gidng vi khudn ttin mdi trudng MRS agar (ed bo sung 6%
NaCl) d nhiet dO 37°C ttong 48-72 gid. Cac khuin Igc sau khi phan lap rdng dugc nhan diln dk xac djnh loai tren ca so:
- Xdc dfnh hinh thdi ciia cdc ddng vl khudn ; phdn lap: dua vao _d§c diim hinh thai khuin Igc;- ,'i die diim hinh thdi t l bdo; nhudm Gram
Khdo sdt khd ndng sinh enzyme cAa cdc ddng vi khudn phdn lap:
Kha ning sinh enzyme'protease: sir dung mdi tiirdng Luria-Bertani cd bd sung Skim milk di phit hiln kha ndng sinh en^me protease ngogi bio ciia cdc ddng vi khudn pfaanlgp (Pailin et al, 2001).
Khd ndng sinh enzyme amylase: khio sat kha ndng phin giii tinfa bgt dua vdo pfaan iing mdu vdi iodine (Nguyin Due Luqng vd ctv., 2006).
Kha ndng sinh enzyme catalase: dua vao klii ning tao bgt faay kfadng tao bgt ctia cdc dong vi kfauin kfai cho kfaudn lac vio dung dich H2O2 3% ] (Karen, 2010). ' \
Khd ndng sinh enzyme oxidase: dua vdo klid ndng lira tiiay ddi radu Tettametiiyl-n- phenylenediamine dihydrochloride ctia cdc ddne\d khuin (Patricia and Laura, 2010).
Tgp chi Khoa hgc Tnrdng Dgi hgc Cdn Tho Phdn B: Nong nghiep. Thuy sdn vd Cong nghi Sinb hgc: 33 (2014): 53-60 - Xdc dinh lodi ciia cdc ddng vi khudn phdn
lap bang ky thugt sinh hgc phdn tu (Sambrook and Russell, 200J): nhdn di?n cic ddng vi sinh vdt phdn lap dugc bing ky tiiudt gidi trinfa tu ttin he thdng mdy ABI 3130XL. Pfaan tich kit qui bing phdn raera sequencing analysis 5.3 vi so sinh kit qud ttin ngan faing Gen de tira loii cd quan fae gan.
3 KET QUA VA THAO LU4.N 3.1 Bac diem ve fainh tfadi cua cac ddng vi kbuan phan l^p
Kit qud cd 4 ddng vi khuan Gram duong (ky Bang 1: Dgc diim hinfa tfaii ciia 4 ddng vi khuan phan lap
hieu Ll, L2, L3, L4) vdi dgc diem rdt ddc tnmg tfaudng dugc diing de nfagn dang cdc ddng vi khuin sinfa acid lactic da dugc Salminen et al., (2004) md ta nhu: vi khudn Gram duong; hinh que hogc hinh cau; tten mdi trudng MRS, khuan lac mau ttdng hodc trdng siia, dudng b'nh kfaodng 1-2 nmi. Cic ddc trung ndy vl hinh thai khuan lgc va te bdo ciing da dugc nhieu tie gid su dung de nhan dgng cdc ddng vi khuin LAB phan lap tur cd va tdm (Parvathy and Puthuvallil, 200S), tur nguyin lieu sira (Mahankumar andMurugalalha. 2011),...
Dong Hinh dang va dac diim kfauan lac Hinh dang va dac diem t l bao
Khuin lgc trdn, bdng, do ndi md, miu trdng stta, bia nguyin, dudng kinh 1,5-2 mm.
Tl bio cd dang hinh cau ddi, it cdu don, dudng kinh klioang 1-2 [im. Gram duong.
Khuan lac ttdn bong, mau trang ngd, bia nguyen, dudng kinli khodng 0,6-1,0 mra.
'':$mm
Te bao Ii dgng hinh que dii, dudng kinh khoang (1-1,5) x (5-8) )jm. Gram duang.
Khuan lgc bdng. dd ndi md, rpau trang nga, bia rang cua. duong kinh 1-1.5 mm.
Te bio hinh que ngan, dudng kinh khoang (I 1.5) x (4-6) \im. Gram duong.
Tgp chi Khoa hpc Tnrong Dgi hgc Cdn Tho Pbdn B: Ndng nghifp, Thity sdn vd Cong nghf Sinb hgc: 33 (2014)- 53-60
L 4
K h u a n lgc trdn bdng, d o ndi md, mdu ttang
siia, bia nguyen, d u d n g kinh 0,5-1 ram. T i bdo cd dgng hinh c i u don, d u d n g kinh kfaodng Ipra, Gram duong.
Hinh chip ngay 29/5/2013
3.2 Kfaa n i n g sinh e n z y m e ciia cdc d d n g vi k h u a n p h a n Igp
- Kit qud kiim tra khd nang sinh enzyme oxidase vd catalase: cd 4 d d n g vi kfauin phdn ldp
d u g c d i u cd tinh c h i t sinh h d a rdt ddc trung cila vi k h u i n lactic: oxidase vd catalase am tinh. K i t qud b i n g hinfa dnfa d u g c ghi nhdn qua Hinh 2 va Huifa 3.
- 1 b . D o n g L 2 c . D o n g Ld d. U o n g i H i n h 2: H i n h a n h v e k i t q u a thir k h a n a n g sinfa e n z y m e oxidase eiia 4 d d n g vi k h u i n Bon dong Ll. L2, L3, L4 khdng ldm thay doi mdu ihudc thit Telramelhyl-p-phenylenediamine dihydrochloride cho kit qud Ihit: dm linh (Chup ngdy 29/5/2013)
a. Dong LI b. Dong L2 c. Dong L3 d. Ddng L4
Hinh 3 : Hinh anh ve ket qua thir kha nang sinh e n ^ m e catalase ciia 4 dong vi k h u i n Khi cho khudfilgc cm bdn dong Lf, L2, L3. L4 vdo dung dich H2O2 3%, khdng ifidy xudt hi4n hien iugng lao bgt: dm tinb (Hinh chup ngdy 29/5/2013)
Kit qud kiem tra khd ndng sinh enzyme protease: cd 3 ddng L l . L2 va L3 cho ket qud
d u o n g tfnh vdi protease, cdn lgi ddng L4 cho k i t qud dm tinh (Hinh 4).
a. Ddng L1 ( d u a n g tinh) b. D d n g L2 ( d u a n g tinh) c. Ddng L3 ( d u o i ^ tinh) d. D d n g L4 (dm tinh) H i n h 4 : H i n h d n h v i k i t qud t h i i k h a n i n g sinfa e n z y m e p r o t e a s e c u a 4 d d n g vi k h u i n Cdc dong vi khudn tgo vong sdng khi nuoi cdy trin mdi trudng Luria-Bertani co bd sung Skim milk: duang linh (Hinh chup ngdy 30/5/2013)
Tgp chi Khoa bgc Tntang Dgi bgc Cdn Tho Phdn B: Ndng nghiip. Thity sdn vd Cong nghf Sinh hgc: 33 (2014)- 53-6
- Kit gud kiem tra khd ndng sinh enzyme amylase: bdn ddng vi khuan phin lip dugc deu cbo
Am tinh
kit qui duong tinh vdi amylase (Hinh 5).
Hinh 5: Ket qua thir amylase eiia 4 ddng vi kfaudn phan Idp Ll, L2, L3, L4 Cdc ddng khi chiing vdo moi tnrdng (Hinh chup ngdy 30/5/2013)
Tdng hgp kit qud kfaao sdt mgt sd dgc tinfa sinfa hda ciia bdn ddng vi khuin pfaan ldp dugc thi hiln o Bing 2.
Bang 2: Bang tong fagp kit qua kiem tra kfaa nang sinfa enzyme ciia 4 ddng vi sinh vat phan lap
D6ng Gram Ll + L2 + L3 + L4 +
Kfaa nang sinh enzyme Catalase Oxidase Amylase Protease
+ + + + ,-f- +
+ Tfaeo Nongpanga et al. (2008), tiiu chi quan ttgng ve tinh chat sinh hda ttong viec xdc dinh vi khuin lactic: te bdo Gram duang; tfaii boat tinh enzyme catalase, oxidase cho kit qud dm tinh. Bdn ddng vi khudn pfain ldp deu tfada man tiiu cfai niy.
Nfaiiu tfai ngfaiem pfain lip vi khudn lactic tir san pham lln men da dung tiiu chi ndy dl budc diu nli|in dgng vi kfauin lactic nhu phdn lap LAB tii siia ddng vd dua leo (Mahantesk et al, 2010), phan lap Lactobacillus delbreukii tir siia lln men (Morami et al.,2013),...
Ngodi ra, mara cfaua cd sac Id sdn phdm cd len men lactic. Do do, trong qua trinh len raen ddi fadi phdi cd vi sinfa vdt tfaam gia qui ttinh chuyin hda
carbohydrate thanh acid lactic, ddng thdi ciing phdi cd vi sinh vdt phan giii protein thanh acid amin.
Nhu viy, ea 4 ddng vi sinh vit ttin deu cd khi ndng su- dung ngudn carbohydrate ttong san phdm, tuy nhien clii cd 3 ddng Ll. L2. L3 cd khi nang phin giai thit cd. Diy la co sd quan ttgng cfao vile cfagn gidng vi kfauan de chiing vdo sin pfaim giiip qua ttinh len men tdt ban.
Mac dii, phucmg phdp xdc djnfa LAB hiln nay vin dya tten viec xdc dinh hinh tiiii, tuy nliien, phuang phap nay klidng phdi Iiic nao ciing dam bdo do chinh xac mi cd thi dan den viec xdc dinh sai. Ngiy nay, viec ip dung kJ thuat PCR va gidi trinfa tu gen 16S rRNA eho phep xac dinfa nhanh chdng mdt lugng ldn vi sinfa vdt (Rantsiou et ai, 2006 va Cocolin and Rantsiou, 2012).
5.3 Nfaan dien cdc ddng vl khuin phan lap bdng ky thudt sinfa hgc phan tir
Kit qua cho thiy bd gen ciia vl khuin dd dugc ttich ly thinh cdng vdi sdn phdm PCR thu dugc cd chdt lugng tdt, bang ro duy nhdt cho m6i ddng vi khuin ttin gel diln di vdi kich thudc doan gen khoang 1500 bp (Hinh 6). Giii ttinh ty gen
16S rRNA ciia bdn ddng vi kfauin cfao kit qua d Bang 3.
Sii dung cap mdi (William et al. 1991) 1492R 5'-TACGGTTACCTTGTTACGACT-3' 27F 5'-AGAGTTTGATCCTGGCTC-3' de kfauech dai ttinh tu viing bao tdn ttin 16S rRNA ciia bdn ddng vi kfauan pfadn lap.
Hinfa 6: Phd diln di ciia gen 16S rRNA ciia 4 ddng vi kfauan va Ladder lOObp plus
Tap chi Khoa hoc TrudngDai hoc Cdn Thcr Fhdn B: Ndng nghiep. Thiiy sdn vd Cdng ngh4 Sinh hoc' 53 (2014): 33-60
Bang 3: Ket qua giai trinh tir gen 16S rRNA cQa bon dong vi Itliuan Dong Trinh tg gen 16S rRNA
GCAAGTCGAACQAACTTCCGTTAATTGATTATQACGTGCTTGCACTGAATQAGATTT TAACACGAAGTGAQTGGCGGACGGGTGAGTAACACGTGGGTAACCTOCCCAGAAGC AGGGGATAACACCTGGAAACAGATGCTAATACCGTATAACAGAGAAAACCGCCTGG TTTTCTTTTAAAAGATGGCTCTGCTATCACTTCTGGATGGACCCGCGGCGCATTAGC L1 TAGTTGGTGAGGTAACGGCTCACCAAGGCGATGATGCGTAGCCGACCTGAGAGGGT
AATCGGCCACATTGGGACTOAGACACGGCCCAGACTCCTACGGGAGOCAGCAGTAG OGAATCTTCCACAATGGACGCAAGTCTGATGGAGCAACGCCGCGTGAGTGAAGAAG GGTTTCGGCTCGTAAAGCTCTGrrGTTAAAGAAGAACGTGGGTGAGAGTAACTGTTC ACCCAGTGACGGTATTTAACCAGAAAGCCACGGCTAACTACGTG
GCAAGTCOAACGAACCAAACTGTTGATTAAAGCTTGCTrTATGATTCAGACCTTGGT GAGTGGCGGACGGGTGAGTAACACGTGGGTAACCTGCCCAAAAGTGGGGGATAAC AnTGGAAACAAGTGCTAATACCGCATAACAACTACTTTCACATGATCGTAGCTTGA AAGATGGCTCTGCTATCACnTTGGATGGACCCGCGGCGTATTAGCTAGTTGGTGAG L2 GTAATAGCTCACCAAGGCAATGATACGTAGCCGACCTGAGAGGGTAATCGGCCACA
TTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGCAGTAGGGAATCTrCCA CAATGGGCGAAAGCCTGATGGAGCAATGCCOCGTGAGTGAAGAAGGTTTTCGGATC GTAAAACTCTGTTGrTGAAGAAGAACATGCGTGAGAGTAACTGTTCACGTACTGAC GGTATTCAACCAGAAAOCCACGGCTAACTACOTO
GCAAGTCGAACGAACCATCCTGTAGATTGAAGCTTGCTTCATGATTCAGATnTGGT GAGTQGCGGACGGGTGAGTAACACGTGGGTAACCTGCCCAAAAGTGGGGGATAAC ATTTGGAAACAAGTGCTAATACCGCATAACAACTACTTTCACATGATCGTAGCTTGA AAGATOGCTCTGCTATCQCnTTGGATGGACCCGCGGCGTATTAGCTAGTTGGTGAG L3 GTAATAGCTCACCAAGGCAATGATACGTAGCCGACCTGAGAGGGTAATCGGCCACA
TTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGCAOTAGGGAATCTTCCA CAATGGGCGAAAGCCTGATGGAGCAATGCCGCOTGAGTGAAGAAGGmTCGGATC GTAAAACTCTGTTGTTGAAGAAGAACATGCGTGAGAGTAACTGTTCACGTACTGAC GGTATTCAACCAGAAAGCCACGGCTAACTACGTGC
CGTCAGACGTGCACAGTTACTTACACGTTTGTTCTTCCCTAATAACAGAOTTTTACG ATCCGAAGACCTTCATCACTCACGCGGCGTTGCTCCGTCAGGCTITCGCCCATTGCG GAAGATTCCCTACTGCTGCCTCCCGTAGGAGTCTGGACCOTGTCTCAGTTCCAGTGT GGCCGATCACCCTCTCAGGTCGGCTACGTATCGTTGCCTTGGTAAGCCGTTACCTTA L4 CCAACTAGCTAATACGGCGCAGGTCCATCTATAAGTGATAGCAAAGCCATCTTTCAC
TATCGAACCATGCGGTTCGAAATATrATCCGGTATTAG_CTCCGGTTTCCCGAAGTTA TCCCAGTCTTATAGGTAGGTTACCTACGTGTTACTCACCCGTCCGCCGCTAACGTCA AAGGAGCAAGCTCCTCGTCTGTTCGCTCGACTTGCATGTATTAGGCACGCCGCCAGC QTTCATCCTGAGCCAG
So sanh trinh tir gen cua bon khuan
(wwwncbi.nlm.nhi.gov) dg xdc djnh cac loai c6 phim lap "tren ngin hang gen fl"^ he gan. Ket qua duoc die h.en a Bang 4.
Bang 4: So sdnh kit qua giai trinh t^ tren ngan hdng gen
Ddng Ma gen Cac ddng so sdnfa Miic dp tuong dong (%) Loai duge xdc dinfa JN836485.1 Pediococcus acidilactici strain 100 Pediococcus
- O mls-1 I6S ribosomal RNA acidilactici HM130540.1 Lactobacillus farciminis strain .f.,, Lactobacillus
Lf-10 16S rRNA gene ^"" farciminis L3 AB626064.1 L-ctobacillm farciminis gcnz 99 Laelobacillus
for 168 rRNA. partial se farciminis AM779066.1 Staphylococcus hominis •pa'ct.\a{ 100 Staphylococcus
16S rRNA gene, strain AK... hominis
Tgp chi Khoa hgc Tnrdng Dgi hgc Cdn Tha Phdn B: Nong nghifp, Thdy sdn vd Cong nghe Sinb hgc: 33 (2014): 53-60
Nfau vdy, bdn ddng vi sinh vat phan lap da dugc itiiin diln bang ky tfauat sinfa hgc phin tu chi cd 3 ddng (Ll, L2, L3) li thugc gidng vi khudn lactic, ddng L4 khdng phii li vi khudn lactic. Kit qud phan Igp ttin ciing pfaii fagp vdi kit qui ngfailn ciiu ciia Tanasupawat and Komagata (1995) khi pfaan laj) cic ddi^ vi khuin tir sdn phdm len men truyin thdng (cd, thit, rau qud,..) cua Thii Lan:
Lactobacillus spp. va Pediococcus acidilactici la cdc gidng thudng phan lap dugc vd tfainfa tfaoing cung phan lap dugc gidng Staphylococcus.
Cac ddng vi khuan lactic pfadn ldp dugc tii sdn phdm mara cfaua ca sac cung la cdc ddng vi khuin da duge nhgn diln ttong san pham cd len men ciia Thai Lan nhu Pla - ra, Pla - som, tning cd len men (Mura-kfaai-pla),... Cdng tfadng tin tryc tuyen NBRC cua Tfadi Lan dd cdng bd cdng khai 221 cfaiing vi khuan lactic cd ngudn goc tir tfayc phara lln men ciia Thdi Lan. Theo kit qui nay thi Lactobacillus farciminis va Pediococcus acidilactici la 2 ddng vi khuan lactic rdt phd biln ttong sin phara ca lln men, trong dd Lactobacillus farciminis chilm ti 11 74/221 va Pediococcus acidilactici chilm ti le 7/221 ttong tdng sd 121 chiing da dugc nhgn dien. (http://www.nbrc.nite.
go.jp/ e/thailact-e.html truy cap ngdy 20/5/2012).
Klyohara et ai (2012) khi nghiin ciiu vi quan thi vi sinfa vit hiln diln ttong sin phdm len men tii cd mudi vd com (narezushi) tfai nlign tiiay vi khuin sinh acid lactic chiera uu thi nhung trong dd mat sd vi khuan Lactobacillus gia tang ding kl ttong sudt qui ttinh lln men, ttong khi do mat sd cua nfaiing vi khuan khdc gia tdng khdng ding kl. Theo nghiin ciiu ciia Antonio et al. (2008) ve viec xac -dinh ddng hgc ciia qudn thi vi sinh vat hiln diln ' ttong san pfaam phd mai tir sira cim len men (cd ten ggi Id Cueva de la Magafaa) d giai doan ii chin. Kit qua ghi nhin Lactobacillus paracasei Id lodi chiem uu thi, vd cdc lodi Lactobacillus khdc dugc tim thdy d giai dogn ddu cua qui ttinh li chin. Cic ddng khdng phii vi khuan acid lactic nhu Kocuria v i Staphylococcus tiii duge phit hien d giai doan cudi cua qui ttinfa ii chin.
4 KET LUAN VA Dfe XUAT 4.1 Kit tudn
Bon ddng vi khuin dugc phin lap tir sin phim cd sic lln raen cfaua, xuong mim cd cae dgc diira ve fainli thii: khudn lgc mau ttdng hodc ttdng siia, dudng kinh khoing 1-2 mm; ti bao hinli que. hinh ciu ddi ho^c hinfa cau don.
Cd bdn ddng vi kfaudn pfadn lap dugc diu Id vi khudn Gram duong, cho kit qua im tinfa vdi enzyme catalase, enzyme oxidase; duong tinh vdi erayme amylase vi chi cd 3 ddng H , L2, L3 cho kit qui duang ti'nfa vdi en^me protease.
Bdn ddng ndy cd miic dg tuang ddng ve trinh tu gen 16S rRNA ttin 99% vdi cdc ddng vi sinfa vdt: Pediococcus acidilactici, Lactobacillus farciminis va Staphylococcus hominis.
4.2 Dl xuat
Khio sat kha nang sii dyng dudng glucose, duong succrose, tinh bdt cita cac ddng vi kfauan tten dl ldm co sd cho vile chgn gidng vi khuin chiing vio san phdm.
Phdn lap va klido sat vai ttd cua cdc ddng vi sinh vat khae (nim raen, nim mdc,..) cd ttong sdn phim.
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