【摘要】：微生態(tài)制劑的固態(tài)發(fā)酵比液體發(fā)酵能提供更為豐富、活性更高的酶系,其所有發(fā)酵產(chǎn)物均可作為制劑組分,且具有能耗低、無廢液排放等優(yōu)勢。因此本文首先對飼用微生態(tài)制劑進行菌株的篩選和菌株種子液培養(yǎng)基的優(yōu)化,然后初步建立了一套飼用微生態(tài)制劑的固態(tài)發(fā)酵工藝,并對菌株種子液和制劑的安全性進行驗證。其結果如下:(1)通過對干酪乳桿菌M9(Lactobacillus casei M9)進行耐酸、耐豬膽鹽和耐高溫馴化及對產(chǎn)朊假絲酵母Y5(Candida utilis Y5)進行耐高溫馴化以提高兩株供試菌株的耐受性并得到新的菌株:干酪乳桿菌M9c(Lactobacillus casei M9c)和產(chǎn)朊假絲酵母Y5a(Candida utilis Y5a);然后對枯草芽孢桿菌R5(Bacillus subtilis R5)、干酪乳桿菌M9c、產(chǎn)朊假絲酵母Y5a進行菌株間拮抗作用確定各自之間不存在拮抗作用,得到可以研制飼用微生態(tài)制劑的優(yōu)良菌株。(2)通過單因素試驗及正交試驗,得到三種菌的最優(yōu)培養(yǎng)基及培養(yǎng)條件:B.subtilis R5:葡萄糖2%,蛋白胨1%,NH4Cl 0.6%,MgSO4 0.6%,NaCl 0.5%,pH 6.5。接種量為5%,34℃,180rpm搖床培養(yǎng)16h。L.casei M9c:酵母膏1%,蛋白胨2%,紅糖2%,NH4Cl 0.6%,KH2PO4 0.4%,MgSO4 0.2%。pH 6,接種量為2.5%,37℃靜置培養(yǎng)16h。C.utilis Y5a:去皮馬鈴薯20%,葡萄糖1%,酵母膏0.5%,KH2PO4 0.05%,ZnSO40.01%,MnSO4 0.075%。pH為6,接種量為5%,32℃,180rpm搖床培養(yǎng)16h。(3)通過PB試驗挑選出培養(yǎng)基組分的三個關鍵因素,采用最陡爬坡實驗確定其最佳濃度區(qū)域,利用中心旋轉組合試驗確定固態(tài)發(fā)酵培養(yǎng)基的配比:麥麩18g,玉米皮粉7.5g,豆粕4.5g,麥芽糖0.45g,紅糖0.74g,氯化銨0.3g,硫酸銨0.3g,碳酸鈣0.15g,磷酸二氫鉀0.29g,硫酸錳0.08,硫酸鎂0.15。由單因素試驗確定了固態(tài)發(fā)酵條件為:含水量(即干物料質量:水的體積)為1:0.8,B.subtilis R5、L.casei M9c、C.utilis Y5a的接種量分別為5%、2.5%、5%,整個發(fā)酵過程中溫度控制在30℃~34℃,發(fā)酵44h。(4)通過對發(fā)酵產(chǎn)物進行一系列處理,初步得到一套發(fā)酵后預處理工藝及保存方式:將發(fā)酵產(chǎn)物置于60℃真空干燥箱烘至含水量為10%,置于18×25全新PET/PE復合加工而成真空袋中4℃真空保藏,180d后活菌數(shù)依然可達1.033×1011 cfu/g,高于國家標準。(5)用篩選的菌株種子液和固態(tài)發(fā)酵的微生態(tài)制劑灌喂小白鼠14~30天,小白鼠均未出現(xiàn)死亡情況,并且小白鼠飲食、精神面貌等臨床現(xiàn)象都正常。解剖觀察小白鼠的肝臟和腎臟,未發(fā)現(xiàn)有病變等不良現(xiàn)象,并且制劑可有效增加小白鼠的體重,降低腹瀉率。
[Abstract]:The solid-state fermentation of microecological agents can provide more enzymes with higher activity than liquid fermentation. All the fermentation products can be used as preparation components and have the advantages of low energy consumption and no waste liquid emission. Therefore, in this paper, the strains were screened and the medium of seed liquid was optimized, then a set of solid-state fermentation technology was established, and the safety of the seed liquid and preparation was verified. The results are as follows: (1) the acid tolerance of Lactobacillus casei M9 (Lactobacillus casei M9 was studied. High temperature tolerance of Candida utilis Y5 (Candida utilis Y5) in order to improve the tolerance of the two strains and to obtain new strains: Lactobacillus casei M9c (Lactobacillus casei M9c) and Candida utilis Y5a (Candida utilis Y5a). Bacillus subtilis R5 (Bacillus subtilis R5), Lactobacillus casei M9c, Candida prion Y5a were used for the antagonism between strains. The optimum culture medium and culture conditions of the three strains were obtained by single factor test and orthogonal test. The optimum culture conditions were: glucose 2, peptone 1 + NH 4Cl 0.6 + MgSO 4 0. 6% NaCl 0.5 + pH 6. 5. The optimum medium and culture conditions of the three strains were obtained by single factor test and orthogonal experiment. The optimum culture medium and culture conditions were as follows: 1% B. subtilis R5: glucose 2%, peptone 1% NH 4 Cl 0. 6 and MgSO 4 0. 6%. The inoculation amount was 5 ~ 34 鈩，

本文編號：2202127