Thông tin tài liệu
• Nghien civu - K9 thOt M.E Aulton, Editor., Churchill Livingstone Press p 336 — 342 Gowthamarajan K., G T Kulkami, and M N Kumar (2004), "Pop the pills without bitterness", Resonance, 9(12), p 25-32 4.Ding Van Giap (2002), Thief ke va t6i UV hoa cling thtic va quy trinh, TP F1$ Chi Minh, Nha xuat ban Y hcc BO Y to (2009), Dopy di4n Vitt Nam IV, Nha xuat ban Y hcc (NA/ nhOn bak 14/03/2017 - Ngay phan bien: 03/04/2017 - Ngay duyat dang: 05/05/2017) Len men too natto tir d4u tirong Vi0 Nam, tach chitt va danh gia hoot tinh tieu fibrin ciia nattokinase Nguyen Thi 1,41*, Nguyen Thi Loan', Phum Thi NguyOt Truenig Dai hoc Dupe Ha Nei Vien Duey lieu E-mail: lapnt@hup.edu.vn Summary Natto, a traditional Japanese food produced by fermentation of soybeans with Bacillus subtilis natto, was made from Vietnamese soybeans in the same fermentation way, i.e with very Bacillus subtilis natto Further, the natural proteolytic enzyme nattokinase of natto was isolated, purified and determined the enzymatic activity Extracted from the Natto made from small, green soybean, the highest Nattokinase yield reached 0.18 g and 0.45 g per 150 g of wet and dried Natto, respectively Fibrinolytic effect of the Nattokinase extracts was also studied in comparison with that of the standard specimens, showing the same significant action on thrombolysis and anticoagulation as the standard did It suggested the Vietnamese soybean would be a promising material for natto fermentation in view of nattokinase production Keywords: Natto, Nattokinase, Bacilus subtilis natto, fermentation, thrombolysis Oat Van de Benh tim mach la nguyen nhan gay tin vong hang dau dei vol the giai nOi chung, dac biet 16 dOi vol cac nvac dang phat then nhv Viet Nam Vi vay viec san xuet ye phe bien thyc pham chVc nang china cac yeu to tieu huyet khei nham phong ngira va dieu tri cac benh 19 tim mach nguy hiem de- va dang duvc quan tam MOt nhCrrig 19 quan khien ngutri Nhat Ban song lau va song khOe nhat the gidei le ho sir dyng hang mOt bpi thCrc an phi) bien, da chinh la natto, &vac lam tin dau twang len men vol vi khuen Bacilus subtilis natto Nam 1980, GS Hiroyuki Sumi da phat hien thine pham tv nhien china enzym Mu fibrin, lam tan huyet khei manh me, Ong d5 dot ten enzym la nattokinase tst Nattokinase c6 hieu Ivc phan giai fibrin (fibrinolytic) c6 chine nang giong plasmin, n9hia la trvc tier) giang hoa fibrin cut mau (long bang cach hoot hOa flan urokinase urokinase va hopt h6a qua trinh tong hap t-PA huyet twang Do d6 sir dyng natto cs tdc dyng ngan ngira va dieu trj cac chirng banh lien quan den tim mach Tren the da c6 nhieu kat qua nghien ciru chiet xuet va tinh the enzym nattokinase, chung minh tac dyng tieu fibrin nhem lam thuOc va thvc pham chin nang (1,61 Hien nay, nhieu nirac nhu' Trung QuOc, Thai Lan da tim hieu phirang phap san xuet, the bien that pham chine nang chira natto nhir thOi quen an Wong cho ngvai dan TAP CHI DU'VC HQC - 5/2017 (SO 493 NAM 57) lam tin chinh MU twang cua nutec minh Tuy nhien cac san pham thuec va thvc pham chin nang c6 nguen g6c tin natto hpn the tai thj trvang Viet Nam, CO le nguyen nhan chinh china tu' san xuat duvc nen gia nguyen lieu nhep khau cao Xuat phat tin nhOng l9 neu ten, chung to' thy'c hien nghien dru vai muc tieu: Len men too dvqc natto tin dau Wang Viet Nam, tech chiet enzym nattokinase va danh gia hoot tinh lieu fibrin dm the pham D6i tuvng va plarang phap nghien CCM Nguy8n li8u va hoa chAt Hat dau nanh da phai kho, Iva chon loci nguyen lieu Qt.' nanh thvang CO san ten thj trutyng (Hoang Su Phi, Ha Giang): MU yang ram, to va dau yang xanh, nho Nattokinase NSK-SD, chat dOi chieu duvc san xuet bai CT Japan Bio Science Laboratory Co., Ltd., (Nh5t Ban) Bacillus subtilis natto va natto (lyric cung cap boi CT Taishi Foods (Nhat Ban) Ca chat dac hieu cOa nattokinase S-2251 (H-D-valyl-L-leucyl-L-lysin-pnitroanilid dihydrochlorid) duvc cung cep bai SigmaAldrich (DUO Cac h6a chat sir dung nghien cOu deu dot tieu chart phan tich Phwang phap nghien cu'u Plurong pluip len men too natto Xuat phat kr 19 thuyet va kinh nghiem len men natto theo phvang phap truyen thong cila nguai Nhat, nghien dm ben hanh len men dau Wang nhu sau: • Nghien civu - Ky thulat Hat deu kW) duvc rim such va ngam nt.r6c veri t9 le dau/nuac:1/3 Thai gian ngam dOu khodng 12 gib vao mem he ye 20 gier vao mem dang nhiet de phang cho hat dau hap thy de nverc Deu da ngam &Par luOc gib hoec hap 121°C 30 phut Deu de chin duvc lam nguei den khoang 60°C, sau d6 ten dgu yeri chung vi khuan Bacillus subtilis natto, 37°C khoang 48 gib de to natto Natto duvc bao quan nhiet dO khoang 5°C ngan can sy, kher amin Met kg dau twang san xuet duvc khoang kg natto a Phuong phtip djnh luting protein Nang de nattokinase duvc djnh luting bang bang k9 thuet Gomal dkra ten nguyen tic dm phan irng Biuret, quang bverc sang 540 nm Xay dung bieu da mau de djnh luting protein, dung dung djch protein da biet neng de (dung djch casein 1%) lam dung djch mau Chief tacit nattokinase Tech chi& va tinh the nattokinase tir deu twang len men (natto) theo phvang phdp dm Peng, et al., (2003) [4) duvc cai tan, m6 to tarn tit nhu, sau: Natto &yr ngam gib NaCI 0,9% chira CaCl2 mM Ly tam 6, tec de 4000 yang/phut 10 phut, thu djch Them bet (NH4)2SO4 de thu duvc dung dich c6 neng de 19 g/100 ml, khuay k9, de Ong gier, ly tam tec de 10.000 vang/phet 15 phut, thu djch Sau da digu chinh ye pH = 8,6 va them bOt (NH4)2SO4 de thu duvc dung djch 66 neng dO 45 8/100 ml, khuay k9, de long gier, ly tam t6c de 10.000 vang/phut 15 phut, thu djch Djch duvc tharn tich dung djch 50 mM Tris HCI (pH 7,0) gib, cir sau gitt thay dich mai Djch sau tham tich duvc dua tinh the qua cat sec k9 Sephadex G75 Djch thu duvc &Tic lam clang kh6 de thu duvc bet nattokinase Danh gia hoattinh phangial fibrin coa nattokinase Xdc djnh host tinh tieu fibrin theo phucmg phap to fibrin barn ten dau dila they tinh [1]• Mau tuvi cua ba kheie manh duvc chung clang bang dung dich natri citrat 3,8%, li tam roc de 1500 yang/ phut phut de thu duvc huyet twang ngheo lieu ceu Pha ml huyet twang 50 ml NaCI 0,9% va 0,8 ml CaCl2 mM, khuay k9 bang dila they tinh (met deu mai rap), 37°C gib Sau d6 them 20 ml dung djch NaCI 0,9% de cyc dang fibrin tech ra, barn vao dua they tinh met mang mong, res mang fibrin barn len dau dim they tinh bang dung djch NaCI 0,9% Nattokinase (0,1%) duvc pha bang dung djch dem pH 8, 40°C Xac djnh kha nang tigu fibrin cea nattokinase bang phvang phap quan sat thai gian lieu fibrin ten dila they tinh chuan bj nhu, ten Chuan bj cdc eng nghiem nha, cho vao moi eng 10 ml the pham nattokinase th6 nang 650,1% va the pham chuan ding dieu kien 40°C Treo dim they tinh cho toan be du c6 fibrin ngep eng nghiem va kh&i dOng Clang h6 barn giay, theo doi thai gian cho den fibrin tan hoan toan Phtrong phtfp xac dint) hoot dpi peptidase Hoot de peptidase duvc bang phvang phap quang ph6 UV-VIS sir dyng co, chat dec hiOu cea nattokinase, S-2251 (H-D-valyl-L-leucyl-L-lysin-pnitroanilid dihydrochlorid) [31, tam tat nhu' sau: Hon hop phan Ong (1 ml) chipa dung djch nattokinase, x 10-4 M ca chat va 0,1 M dOm phosphat (pH 7,4) Sau u 37°C phOt, ham luting san pham phan Ong gidi phOng la p-nitroanilin duvc xac djnh de hap thy bverc song 405 nm Met den vj hoot de (1U) duvc bleu dien bang se nanomol ca chat bj they phan met phut met ml enzym Moi gid tri &vac tinh trung binh cea Ian thi nghiem cung diau Hen K6t qua va ban lOn Len men to natto kfa oilcan It* ddu twang co khd nang sinh nattokinase cao nhdt San xuet natto chiu anh twang cea nhieu yeu to nguen gOc, chet luting hat dOu twang, nhiet de em, theri gian len men, thvc khan the cea Bacillus subtilis natto Trong nghien c,Cru chung tai too natto tCr bon mai, nguyen lieu sau: - Mau 1: dau yang ram, to, nguyen hat - Mau 2: Mu vang xanh, nha, nguyen hat - Mau 3: dOu yang ram, to, nghien tuft len men - Mau 4: deu yang xanh, nho, nghien tnyerc len men Binh thverng sau len men natto xuet hien mei amoniac nhe, sau cho vao to lanh mei se met Khi mei qua mpnh, da la elk hieu me natto de bj qua lau khien cho cac acid amin bj phan giai tier) too nhieu amoniac, lam natto met MO A Do da thed digm de hoar' me natto la rat quan Quan sat chat luting natto thu duvc (de nhat va mui dec trung) giCip Iva chon duvc thai gian len men thick hap la 16 gier tai nhiet de 27°C De em la met yeu to quan tong can dvor kiem salt chat the vi lien quan den sv kh6 be met hat deu, anh hu,6,ng de dinh nhat cua natto Dec biet vai deu twang de nghien nha thi sv kh6 be met de )(ay hen Sau da, natto to duvc bao quan nhiet de &rad 5°C ngan can sv khe, amin Trong qua trinh bao guar', nhieu tinh thg ding, tang hai yang, nhe hinh tren be met natto Thanh phan cCia tinh the la tyrosin va hoec struvite (ammonium magnesium sulfat) Ve cam quan cho they natto to to c6 mau sac, de nhert va MCJi vj gen twang to nhu, san pham natto thvang mai nhap khau te, Nhat Ban Sau len men thu natto, tack chi& enzym, xac dinh thai gian tieu fibrin cea c,dc mau thu duvc Kat qua duvc trinh bay de bang Bang Ham luting nattokinase va thdti gian tieu fibrin cua bon mare au tuong Mau a Ham krcmg nattokinase (9/150 g natio) 0,10 0,13 0,15 0,18 INN gian tieu fibrin (phut) 213 193 115 92 Ghl chti: Mau 1: au yang rum, to, nguyen hat; Mhu 2: au yang xanh, nhd, nguyen hat; MAu 3: dau yang tom, to, nghien Crud len men; MAu 4: au yang xanh, nhd, nghien auto len men TAP CHI DU6C HQC - 5/2017 (SO 493 NAM 57) • Nghien ciru - K9 thOt Ket qua a bang cho they ca mau deu thu duvc ham luring nattokinase (tip 0,1-0,18 g) Trong do, met, cO ham luring nattokiase thu duvc cao nhet (0,18 g) va thai gian tieu fibrin thep nhet (92 phut), chirng tO hoot tinh tieu fibrin manh nhet bOn met] Dieu cO the hat deu nhO han, duvc nghien trtyac thi cO the hep thu nhieu nu,ac va de chin han Ben canh dO qua trinh len men la hieu nen lopi hat nhO han se tao moi twang hieu tot han TO, kat qua chi:mg tai chon loaf deu bran hat nhO, vO yang xanh, nghien tube cho cac nghien ciru tiep theo DOi vdri loai dau twang hat nh6, vo yang xanh hieu suet thu nattokinase dat duvc la 0,18 g bet nattokinase /150 g natto vat Neu tinh cho 150 g rneu kho thi hieu suet chi& xuet nattokinase ca the dat mac 0,45 g bet nattokinase /150 g natto kho, Wang &rang vbi kat qua nghien curu dm Peng va CS [4] dm 100 g the phem Sa be danh gia chet natto lam tip deu Krung hat nha, vo yang xanh, nghien tuft CO ham luring protein la 15,53 % twang &yang vdri ham luvng protein natto thiyang mai Tech chiet nattokinase va danh gia hoot tinh tieu fibrin Len men 100 g CIOU twang loai hat nhO, vo yang xanh, de nghien trivi&c thu duvc 234,97 g natto Tien hanh song song cOng dieu kin de tech chiet nattokinase fir mau natto tty tao (rneu 3) va met., natto twang mai nhep khan theo phiyang phap de trinh bay phen Phtyang phap nghien ciru Danh gia kha nang tieu fibrin dm mew Nattokinase thu duvc tiv natto ty' tao, natto Hwang mai va nattokianase chuen diya tren sty theo dOi theri gian tieu fibrin Thi nghiem duvc tien hanh 10 [nett, mOi meu chira 0,1 ml dung dich the phem cO nOng TO 1% dem pH = 7,4 Ket qua duvc the hien b hinh 120 100 18° 60 Vs 40 20 Mau Maui Hinh Thai gian tieu fibrin cua nattokinase Ghi che: Mgu 1: Nattokinase tech chgt to natto ty, tao; Mk 2: mgu nattokinase chugn; Mho 3: Natokinase tech chit ter natto thuung mai nhep ter Nhet Elan Ket qua iy hinh cho they cOng dieu kin nghien ciru, thai gian tett fibrin trung binh cOa nattokinase rnAu 1, tech chiet tip natto tu, tao (98,3 phut), gen twang &rang vai thai gian tett fibrin trung binh cua nattokinase meu 3, tech chiet tiv natto twang mai nhep tit Nhet Ban (92,5 phut) Ket qua TAP CHI DUVC HQC - 5/2017 (SO 493 NAM 57) kheng dinh san phem natto tty tao duvc c6 hoat tinh Nu fibrin cao twang &rang san phem nhep kheu tO, Nhet Ban Tuy nhien, thai gian tieu fibrin trung binh cue nattokinase tech chiet duvc tir met, natto ty' tao va natto twang mai cao hart so vai thtri gian Nu fibrin trung binh cue nattokinase chuen (78,8 phtit) (p < 0,05) g 70 V 60 € 50 r • 40 i* 30 i 20 10 60.5 MaU1 mitt Hinh Hogt tinh peptidase MAu 1: Nattokinase tech chi& ter natto ttr tao; Mho 2: rdu nattokinase chuen Xac dinh hoot tinh peptidase Nattokinase la met loai protease kh6ng chi CO tac dung Wu fibrin ma cO tac dung ten cac ca chet plasmin 1251 Do vey, tru diem not bet ctia nattokinase so yen cac loaf thuoc lam tan huyet khac la CO the sir dung &rang uOng de uOng dieu trj clty phong Khi sir dung duang uOng, nattokinase duvc hep thu qua &rang ruet vao mau lam tieu fibrin va hoat haa yeu to hoat hoa plasminogen m6 (t-PA) Do do, tie') theo se chang nattokinase Lech minh de dec hieu va hoat tinh chiet duvc bang cach xac clinh hoat de peptidase sir dung ca chet plasmin dec hieu cCia nattokinase, S-2251 (H-D-valyl-L-leucyl-L-lysin-p-nitroanilid dihydrochlorid), tien hanh song song vdri nattokinase chuen Ket qua a hinh cho they cung dieu kien nghien ciru, hoat tinh peptidase cua nattokinase mkt thir (natto tty tao) 16 60,5 (nmol/ml/phut), gen twang &rang vbi thai gian tieu fibrin trung binh cua nattokinase chuen (68,8 (nmol/ml/phitt)) (p < 0,01) Ket qua kheng dinh san phem nattokinase thu du,cyc tir natto by tao CO tinh dec Neu cao vai ca chet plasmin, hopt tinh peptidase twang tty nhu, nattokinase chuen Nhu, vele, kat qua tit, hinh va the hien san phem nattokinase tech chi& &vac tit, natto to tao CO the diva dat duvc de tinh khiet nhu, san phem nattokinase chuen, cern phai cO nhOng bulac cai tien quy trinh tech chi& cac nghien ciru tiep theo Mac du Vey, CO the biyac dau kheng dinh chung t6i de °Ong viec len men tao natto tech chiet, tinh the tao nattokinase CO de tinh khiet va hoat tinh tieu fibrin twang dal cao Viet kheng dinh haat tinh tat fibrin dm the phem natto nghien ciru c6 met j( ngifia quan trong viec cung cap them bang chirng de chirng minh san phem natto tao sir dung deu nanh cO nguOn goo tong rook c6 kha nang lam tan huyet khoi twang tiy san phem nhep kink Phiyang phap to natto nhu, da trinh bay & ten, kha clan glen de lam tai cac gia dinh, nguOn nguyen lieu • Nghien -1(9 thu#t dau nenti lai de kiem, re tien gitip lbm gram gia san pham Vi vby, nghien CCM (la me, Men vong viec sir dung va ph6 bien thei quen dung natto doi vbi nguai dan Viet Nam a pheng ngt:ra va h6 lir/ dieu trl cac berth ye tim mach nhu' cao huyet ap, nh6i mau ca tim, tai bien mach mau nao Ket luen Da ding viec len men tea san phem natto tie phu,ang phap len men dm nguti dan Nhbt Ben, SO dung Tau twang dia Viet Nam, chon duvc loci dbu twang mau xanh, hat nhe cho ham lu'c'ng nattokinase nhieu nhat va heat tint) tot nhat Kat qua ding cho they nattokinase thu duvc sau tech chiet tie meu natto len men tee dbu twang Viet Nam ce kha nang lieu fibrin gen twang ti,' mau natto thu,ang mai nhap khau tir Nhat Ben Mbt khac, nattokinase thu duvc ding CO db dbc hibu cao v61 ca chat plasmin va hoat tinh peptidase gen tuyyng tv so vial nattokinase chuhn • Tai lieu tham khao icy Do Trung Phan (2009), thuat xet nghiam buy& hoc va Owen mau (mg dung bong lam sang, Man Huyat hoc va truyen mau trung 'fang, Nha xuet ban Y hoc, Ha N6i, tr 81, 83 Dubey R., Kumar J., Agrawala D., Char T and Pusp P (2011), "Isolation, production, purification, assay and characterization of fibrinolytic enzymes (Nattokinase, Streptokinase and Urokinase) from bacterial sources", African J Biotech, 10, 1408-1420 Fujita M., Nomura K., Hong K., Ito Y., Asada A., and Nishimuro S (1993), "Purification and characterization of a strong fibrinolyitc enzyme (Nattokinase) in the vegetable cheese Natto, a popular soybean fermented food in Japan", Biochem Biophys Res.Commun., 197(3), 1340-1347 Peng Y., Huang Q., Zhang R., Zhang Y (2003), "Purification and characterization of a fibrinolytic enzyme produced by Bacillus amyloliquefaciens DC-4 screened from douchi, a traditional Chinese soybean food", Comp Biochem Physiol Part B., 134, 45-52 Sumi H., Hamada H., Tsushima H., Mihara H., & Muriki H (1987), "A novel fibrinolytic enzyme (Nattokinase) in the vegetable cheese Natto: a typical and popular soybean food in the Japanese diet", Experientia, 43, 1110-1111 Sumi H., Yatagai C., Naito S., Ohsugi T & Saito J (2009), "Purification, crystallization and preliminary X-ray diffraction experiment of nattokinase from Bacillus subtilis natto", J Thromb Haemost., 7, Suppl 2, PP-WE-148 (Ngay nhOn bei: 24/03/2017 - Ngay phan biOn: 24/04/2017 - Ngay dup)t clang: 05/05/2017) Nghien aril dOc tinh cap, tic dung bao ve gan, chong oxy hoi ciia hon hop cao chiet tir actiso, rau ding dit va bim bim bitc Plum Thi Nguyet Hang*, Le Viet Dung Vien Dupr lieu 'E- mail: pnhang2004@yahoo.com Summary Acute toxicity, hepatoprotective and antioxidant activity of the combined extracts from Cynara scolymus L., Glinus oppositifolius (L.) A.DC and Pharbitis nil (L.) Choisy were evaluated At doses of 280 mg/kg and 560 mg/kg demonstrated antioxidant and hepatoprotective effects on acute liver damage model caused by paracetamol at dose 220 mg/kg body weight of white mice through showing a reduction in AST, ALT activity and MDA level The acute toxicity of the extracts was so low that the LD was undeterminable up to the maximum dose of 28.8 g/kg b w of mice Keywords: Extract, Cynara scolymus L., Glini oppositifolius (L.) A.DC, Pharbitis nil (L.) Choisy, acute toxicity, antioxidant, hepatoprotective Det yen de Gan lb ca quan chuyen hea phti,c tap va Ian nhat ca the, tham gia vao sy' trao d6i chat dm qua trinh tong hap sinh hoc va du, trO, [1] Gan cOng chlu tech nhiern do' v61 qua trinh giei deo va can bang chuyen hea nOi mo m Vi CO hoat tfnh chuyen hem, la ca quan co, ben cua chuyen hem sinh hoc Nib trao doi chat, nen gan chlu anh hu,ang bbl cac gOc oxy tie (ROS), lb san pharn dia qua trinh trao doi chat va giai duvc tao to bbo gan Theo de, gan CO ca the chong oxy hea de bao v0 to bao chong lai ROS Tuy nhien, viec san xuat ROS lien tuc va qua mt:fc ding v61 sv met can bang cac co, the chong oxy hod nOi sinh gay stress oxy hoe Su, stress den den ten thu,ang met se phan ter sinh hoc bao gem lipid, protein va ADN, va qua thei gian, gop phan lam ton thu,ang gan Met se cluvc lieu nhu, actis6, rau Clang dat va bim bim biec duvc sir dung lam thu6c y hoc co truyen TAP CHI DOC HQC - 5/2017 (S6 493 NAM 57)
Ngày đăng: 05/07/2023, 11:34
Xem thêm:
