本文選題：丁酸梭菌 + 培養(yǎng)工藝　； 參考：《湖北工業(yè)大學(xué)》2017年碩士論文

【摘要】：丁酸梭菌,即丁酸梭狀芽孢桿菌,又名酪酸菌、宮入菌,是存在于人和畜禽腸道的一種厭氧益生菌,可以改善宿主腸道細(xì)菌生態(tài)系統(tǒng)的平衡,對(duì)治療因各種原因所致的腸道疾病有很好的效果。除此之外,丁酸梭菌對(duì)提高豬、雞、牛、羊等肉質(zhì)也有一定的效果,因此被作為微生態(tài)制劑廣泛應(yīng)用于醫(yī)藥領(lǐng)域和飼料行業(yè)當(dāng)中。丁酸梭菌在使用過程中主要以活菌制劑狀態(tài)存在,因此本文以提高丁酸梭菌菌體濃度、芽孢生成率為主要目標(biāo),利用傳統(tǒng)發(fā)酵培養(yǎng)方法以及結(jié)合數(shù)學(xué)分析軟件優(yōu)化丁酸梭菌的發(fā)酵培養(yǎng)工藝。由于丁酸梭菌屬于嚴(yán)格厭氧菌,所以在此過程中還探究了一種利用Fe-C原電池除氧方法來培養(yǎng)丁酸梭菌。1.采用數(shù)學(xué)分析方法對(duì)提高丁酸梭菌生物量的發(fā)酵工藝進(jìn)行了優(yōu)化。在單因素實(shí)驗(yàn)的基礎(chǔ)上利用Plackett-Burman實(shí)驗(yàn)設(shè)計(jì)篩選出影響菌體濃度的顯著性因素。之后,運(yùn)用最陡爬坡實(shí)驗(yàn)找出中心組合實(shí)驗(yàn)(Central Compose Design,CCD)的中心點(diǎn),最后通過響應(yīng)面法分析獲得最佳發(fā)酵培養(yǎng)基組成成份。結(jié)果表明:酵母浸粉、FeSO_4和K_2HPO_4為影響菌體量的3個(gè)顯著性因素。最終確定可溶性淀粉2%、酵母浸粉6%、FeSO_4 1.74%、K_2HPO_4 0.37%、NaCl 0.2%、MgSO_4 0.024%為最佳培養(yǎng)基組分。最佳培養(yǎng)條件為:溫度34℃,pH7.0,接種量為4%。優(yōu)化后的菌體數(shù)可達(dá)1.01×109個(gè)/m L,與優(yōu)化前(2.3×108個(gè)/mL)相比,提高了4.39倍。2.在提高芽孢率過程中發(fā)現(xiàn)乳糖為最佳碳源,并以芽孢中2,6-吡啶二羧酸(2,6-Pyridine Dicarboxylic Acid,DPA)的含量為檢測指標(biāo)來測定芽孢的數(shù)量。在此基礎(chǔ)上,利用Plackett-Burman實(shí)驗(yàn)設(shè)計(jì)篩選出乳糖、MgSO_4為影響芽孢生成率的兩個(gè)顯著因素,用最陡爬坡路徑逼近最大DPA濃度區(qū)域后,利用響應(yīng)面中心組合設(shè)計(jì)對(duì)顯著因素進(jìn)行優(yōu)化,結(jié)果表明乳糖、MgSO_4最佳添加量分別為0.3%、0.035%。優(yōu)化后培養(yǎng)基組分為乳糖0.3%、MgSO_4 0.035%、酵母浸粉1.0%、NaHCO_30.2%、K_2HPO_4 0.3%、KH_2PO_4 0.1%。最佳培養(yǎng)條件為溫度43℃、pH7.0,接種量為2%。優(yōu)化后的DPA含量為15.16μM,與優(yōu)化前(2.02μM)相比提高了7.5倍,換算成芽孢數(shù)后為6.31×107個(gè)/mL,芽孢轉(zhuǎn)化率為60%。3.探究了Fe-C原電池系統(tǒng)對(duì)丁酸梭菌生長的影響。首先確定了Fe-C原電池系統(tǒng)的最佳組成成份:Fe粉0.5%,C粉0.3%。結(jié)果表明:Fe-C原電池系統(tǒng)可以維持丁酸梭菌培養(yǎng)液長時(shí)間處于一個(gè)較低的的氧化還原電位環(huán)境下。與對(duì)照相比(無Fe-C系統(tǒng)),丁酸梭菌的最大比生長速率從(h-1)0.23提高到0.43,細(xì)胞得率(g cell/g glucose)從0.10提高到0.14,最終生物量提高從0.71g/L提高到了1.31g/L。相反,有機(jī)酸(丁酸和乙酸)的比生產(chǎn)率都有一定的下降:丁酸比生產(chǎn)速率(g/g glucose·h)從0.15下降到0.10;乙酸比生產(chǎn)速率(g/g glucose·h)從0.28下降到0.18。
[Abstract]:Clostridium butyrate, also known as Clostridium butyricum, is a kind of anaerobic probiotics that exists in human and animal intestines, which can improve the balance of intestinal bacterial ecosystem. It has a good effect on the treatment of intestinal diseases caused by various causes. In addition, Clostridium butyrate can improve meat quality of pigs, chickens, cattle and sheep, so it is widely used in medicine and feed industry. Clostridium butyrate mainly exists in the form of living bacteria in the process of use. Therefore, the main aim of this paper is to increase the concentration of Clostridium butyrate and the sporulation rate. The fermentation culture technology of Clostridium butyrate was optimized by traditional fermentation method and mathematical analysis software. Because Clostridium butyrate belongs to strict anaerobes, a method of deoxidizing Clostridium butyrate by Fe-C primary battery has been explored in this process to culture Clostridium butyrate. The fermentation process for improving the biomass of Clostridium butyrate was optimized by mathematical analysis. On the basis of single factor experiment, Plackett-Burman experiment was used to screen the significant factors affecting the bacterial concentration. After that, the center point of the Central composition Design CCD was found out by the steepest climbing experiment, and the optimum composition of the fermentation medium was obtained by response surface analysis. The results showed that FeSOs _ 4 and K _ 2HPOs _ 4 were the three significant factors affecting the biomass. Finally, soluble starch 2 was determined, and yeast extract powder 6 / FeSOs 41.74 + K2HPO4 0.37 + NaCl 0.2% MgSO4 0.024% was the best medium component. The optimum culture conditions were as follows: temperature 34 鈩，

本文編號(hào)：2054121