本文選題：醋糟 + 地衣芽孢桿菌��； 參考：《河北大學》2015年碩士論文

【摘要】：地衣芽胞桿菌(Bacillus licheniformis)和枯草芽孢桿菌(Bacillus subtilis)是兩種常用的飼料添加劑生產(chǎn)用菌株。目前,地衣芽孢桿菌和枯草芽孢桿菌發(fā)酵生產(chǎn)的原料主要取材于玉米粉、豆粕以及麩皮等等,隨著人口壓力增大,糧食供給越來越緊張的情況下,材料取材越來越難,難以實現(xiàn)長期生產(chǎn)發(fā)展。醋糟作為一種纖維素原料,取材方便且儲備量較大,對其進行開發(fā)利用,生產(chǎn)各類益生菌,不失為一種長遠之策。本實驗首先將醋糟進行預先處理工藝以及利用纖維素酶酶解醋糟的工藝優(yōu)化,其次,探討了采用醋糟酶解液作為碳源代替?zhèn)鹘y(tǒng)培養(yǎng)基發(fā)酵生產(chǎn)地衣芽孢桿菌和枯草芽孢桿菌的可行性,然后通過正交實驗、不同醋糟初始糖濃度實驗,優(yōu)化了醋糟酶解液發(fā)酵生產(chǎn)枯草芽孢桿菌的發(fā)酵培養(yǎng)基,最后,在優(yōu)化培養(yǎng)基的基礎上,進一步考察了芽孢桿菌的7 L發(fā)酵罐補料分批發(fā)酵實驗,并且初步考察了其發(fā)酵參數(shù)。進一步分析了利用醋糟酶解液發(fā)酵生產(chǎn)丙酸的可行性,為丙酸的生產(chǎn)提供了一定技術支持。本論文的主要研究成果如下:(1)首先,我們采用水熱處理技術對醋糟進行初步的預處理,經(jīng)機械粉碎,纖維素酶酶解,得到葡萄糖含量為20.00 g/L的醋糟酶解液;其次,對酶解醋糟的纖維素酶酶解工藝進行了探索,主要研究了固液比、纖維素酶酶量、p H以及酶解時間對醋糟酶解的影響,最終確定了酶解醋糟的最佳條件為:固液比為2.0:10、460 U/g醋糟、p H4.8、纖維素酶酶解時間為48 h,最后獲得葡萄糖濃度為27.00 g/L的醋糟酶解。(2)通過與葡萄糖發(fā)酵培養(yǎng)基的對比,對醋糟酶解液發(fā)酵生產(chǎn)芽孢桿菌的可行性進行了探討,結果顯示,醋糟酶解液可以作為碳源,替代傳統(tǒng)發(fā)酵培養(yǎng)基發(fā)酵生產(chǎn)芽孢桿菌。在葡萄糖濃度相當?shù)那闆r,醋糟酶解液發(fā)酵周期40 h,枯草芽孢桿菌的活菌數(shù)可達到4.64×1010 cfu/m L,與傳統(tǒng)葡萄糖培養(yǎng)基(3.56×1010 cfu/m L)相比,提高了30.34%。發(fā)酵周期28 h,地衣芽孢桿菌的活菌數(shù)達到2.03×109 cfu/m L,與傳統(tǒng)葡萄糖培養(yǎng)基(8.07×108 cfu/m L)相比,提高了151.55%。(3)采用正交實驗L9(43)對枯草芽孢桿菌醋糟酶解液發(fā)酵培養(yǎng)基進行了優(yōu)化,結果表明,醋糟酶解液初始糖濃度為29.30 g/L,最佳發(fā)酵培養(yǎng)基配方為玉米漿20.0 g/L、磷酸鹽1.0 g/L、硫酸銨5.0 g/L,枯草芽孢桿菌活菌數(shù)可達到5.26×1010 cfu/m L。然后考察了不同初始糖濃度的醋糟酶解液對枯草芽孢桿菌發(fā)酵的影響,結果表明,初始糖濃度為50.00 g/L時,枯草芽孢桿菌TS-02活菌數(shù)可達到5.76×1010 cfu/m L,同時芽孢生成率在85%以上。7 L發(fā)酵罐結果表明,發(fā)酵周期28 h,枯草芽孢桿菌的活菌數(shù)可達到6.16×1010cfu/m L,比搖瓶發(fā)酵培養(yǎng)提高了32.76%。7 L發(fā)酵罐分批補料發(fā)酵結果顯示,發(fā)酵周期為18 h,活菌數(shù)達到8.25×109 cfu/m L,芽孢產(chǎn)率為83%以上,比搖瓶培養(yǎng)提高了2.23%。(4)初步探討了醋糟酶解液發(fā)酵生產(chǎn)丙酸的可行性,結果顯示,醋糟酶解液可以作為碳源發(fā)酵生產(chǎn)丙酸,發(fā)酵周期132 h時,丙酸產(chǎn)量達24.35 g/L�？疾炝舜自銤饪s液發(fā)酵生產(chǎn)丙酸的情況,初步探索高糖濃縮液對丙酸發(fā)酵的影響情況。結果表明,發(fā)酵周期132 h,丙酸產(chǎn)量達18.8 g/L。丙酸產(chǎn)量較酶解液低,可能是由于高糖濃度醋糟酶解液抑制了產(chǎn)酸丙酸桿菌的生長或其中產(chǎn)生了某些抑制性的物質。
[Abstract]:Bacillus licheniformis (Bacillus licheniformis) and Bacillus subtilis (Bacillus subtilis) are two commonly used production strains for feed additives. At present, the raw materials produced by Bacillus licheniformis and Bacillus subtilis are mainly derived from corn flour, soybean meal and bran and so on. With the increase of population pressure, the supply of grain is getting tighter and tighter. It is difficult to obtain material for long term production. Vinegar Grains, as a kind of cellulose material, can be used as a kind of cellulose material, which is convenient and has a large reserve. It is a long-term strategy to develop and utilize all kinds of probiotics. In this experiment, vinegar grains were first treated and cellulase was used to hydrolyse vinegar grains. Process optimization, secondly, the feasibility of producing Bacillus licheniformis and Bacillus subtilis by using vinegar residue as carbon source instead of traditional culture medium was discussed. Then through orthogonal experiment, the experiment of the initial sugar concentration of different vinegar grains was carried out, and the fermentation medium of Bacillus subtilis fermentation was optimized by fermentation of Vinegar Grains. Finally, the optimization culture was carried out. On the basis of the nutrient base, the experiment of feed batch fermentation in the 7 L fermenting tank of bacillus was further investigated, and its fermentation parameters were preliminarily investigated. The feasibility of producing propionic acid by fermentation of vinegar grains was further analyzed, and some technical support was provided for the production of propionic acid. The main research results of this article are as follows: (1) first of all, we pick up the production of propionic acid. The process of enzymatic hydrolysis of Vinegar Grains with glucose content of 20 g/L was obtained by mechanical pulverization and enzymatic hydrolysis. Secondly, the enzymatic hydrolysis process of enzymatic hydrolysis of vinegar grains was explored. The effects of solid-liquid ratio, cellulose enzyme amount, P H and enzyme hydrolysis time on the hydrolysis of vinegar grains were mainly studied. The optimum conditions for enzymatic hydrolysis of vinegar residue were as follows: the ratio of solid to liquid was 2.0:10460 U/g Vinegar Grains, P H4.8, the enzymatic hydrolysis time of cellulase was 48 h, and the enzymatic hydrolysis of Vinegar Grains with glucose concentration of 27 g/L was obtained. (2) the feasibility of producing Bacillus by fermentation with glucose fermentation medium was discussed by comparison with glucose fermentation medium. The enzymatic hydrolysate of vinegar grains can be used as a carbon source instead of the traditional fermentation medium to produce Bacillus spore. In the case of equal glucose concentration, the fermentation period of acetic acid hydrolysate is 40 h, and the number of Bacillus subtilis can reach 4.64 x 1010 cfu/m L. Compared with the traditional glucose medium (3.56 x 1010 cfu/m L), the fermentation period of 30.34%. is increased by 28. H, the living bacteria of Bacillus licheniformis reached 2.03 * 109 cfu/m L, compared with the traditional glucose medium (8.07 x 108 cfu/m L), improved 151.55%. (3) using orthogonal experiment L9 (43) to optimize the fermentation medium of Bacillus subtilis vinegar residue enzyme solution fermentation medium. The results showed that the initial sugar concentration of the hydrolysate of vinegar grains was 29.30 g/L and the optimum fermentation medium. The formula was corn pulp 20 g/L, phosphate 1 g/L, ammonium sulfate 5 g/L, the number of Bacillus subtilis viable bacteria can reach 5.26 x 1010 cfu/m L., and then the effect of different initial sugar concentration of acetic acid hydrolysate on Bacillus subtilis fermentation was investigated. The results showed that the number of Bacillus subtilis TS-02 live bacteria could reach 5.76 * 1 when the initial sugar concentration was 50 g /L. 010 cfu/m L, and the result of the spore formation rate of more than 85%.7 L fermentor showed that the fermentation period was 28 h, the living bacteria of Bacillus subtilis could reach 6.16 x 1010cfu/m L, which showed that the fermentation period of the 32.76%.7 L fermenting tank was 18 h and the number of living bacteria reached 8.25 x 109 cfu/m L, and the spore yield was 83% more than that of the shake flask fermentation. The feasibility of producing propionic acid by enzymatic hydrolysis of vinegar grains was preliminarily studied by 2.23%. (4). The results showed that the hydrolysate of vinegar grains could produce propionic acid as a carbon source. When the fermentation period was 132 h, the yield of propionic acid was 24.35 g/L. and the condition of propionic acid produced by the fermentation of Vinegar Grains concentrated solution was investigated and the high sugar concentrate was initially explored to propionic acid. The results showed that the fermentation period was 132 h, the yield of propionic acid was 18.8 g/L. and the yield of propionic acid was lower than that of the hydrolysate, which may be due to the inhibition of the growth of propionic acid by the high glucose concentration of acetic acid enzyme solution or some inhibitory substances.

【學位授予單位】：河北大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：S816.3;TQ920.6

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