本文關(guān)鍵詞： 保拉迪酵母菌 流加補(bǔ)料培養(yǎng) 動(dòng)力學(xué)模型 凍干保護(hù)劑 抗熱保護(hù)劑　出處：《陜西科技大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：保拉迪酵母是一種益生菌,其在37℃(動(dòng)物體溫)下生長(zhǎng)良好,而普通酵母菌生長(zhǎng)的最適溫度30℃左右,保拉迪酵母具有獨(dú)特的生物活性,是適合用于人和動(dòng)物的益生菌。目前,保拉迪酵母普遍用于治療嬰幼兒腹瀉,國(guó)內(nèi)保拉迪酵母制劑主要依賴進(jìn)口,有關(guān)培養(yǎng)及制備研究的報(bào)道甚少。本研究以實(shí)驗(yàn)室保藏的保拉迪酵母為試驗(yàn)菌株,研究其培養(yǎng)基組成和發(fā)酵工藝條件,進(jìn)而對(duì)比研究冷凍干燥和噴霧干燥法制備菌粉的技術(shù)工藝、菌粉活性及穩(wěn)定性,獲得以下研究結(jié)論。1.單因素試驗(yàn)和響應(yīng)面法優(yōu)化確定了保拉迪酵母菌最適培養(yǎng)條件為:裝液量52 mL/250 mL、轉(zhuǎn)速181 r/min、接種量3.1%、培養(yǎng)基初始pH值6.4。此條件下培養(yǎng)液中的活菌數(shù)達(dá)到1.86×108 CFU/mL,比優(yōu)化前提高了14.1%。2.上述培養(yǎng)條件下,單因素和Plackett-Burman試驗(yàn)設(shè)計(jì)篩選出影響保拉迪酵母菌生長(zhǎng)的主要因子為:葡萄糖、酵母浸粉、低聚果糖、磷酸氫二鈉、氯化鈣和蛋白胨,其中葡萄糖、酵母浸粉和低聚果糖為主要影響因素,爬坡試驗(yàn)和Box-Behnken試驗(yàn)設(shè)計(jì)獲得優(yōu)化后的培養(yǎng)基組成為:葡萄糖4.83%、酵母浸粉0.18%、低聚果糖0.18%、磷酸氫二鈉0.45%、氯化鈣0.2%、蛋白胨2%。該條件下培養(yǎng)保拉迪酵母菌,活菌數(shù)達(dá)到3.91×108 CFU/mL,菌體干重5.1911 g/L,比對(duì)照組分別提高了2.1×108 CFU/mL和2.0811 g/L。3.保拉迪酵母菌在5 L通風(fēng)發(fā)酵罐中分批培養(yǎng)和流加補(bǔ)料培養(yǎng)結(jié)果表明,流加補(bǔ)料培養(yǎng)能夠促進(jìn)保拉迪酵母菌的生長(zhǎng),活菌數(shù)和菌體干重最高可達(dá)到7.3×108 CFU/mL和12.7644 g/L。較發(fā)酵罐中分批培養(yǎng),活菌數(shù)(4.8×108 CFU/mL)提高了1.5倍,菌體干重(6.5612 g/L)提高了近2倍。采用Logistic和DoseResp模型分別建立了分批培養(yǎng)過程中菌體生長(zhǎng)和底物消耗的非線性擬合動(dòng)力學(xué)模型為:。所得模型與試驗(yàn)數(shù)據(jù)擬合良好。4.響應(yīng)面法優(yōu)化確定了保拉迪酵母菌復(fù)合凍干保護(hù)劑配方為:菊糖0.6%、低聚木糖0.4%、碳酸氫鈉0.3%、硫酸鎂0.75%、乳糖21.41%、海藻糖21.98%、谷氨酸鈉4.02%、脫脂乳22%,磷酸緩沖液添加量與菌泥重之比為1.2:1。保拉迪酵母菌凍干存活率和單位菌粉活菌數(shù)分別為72.55%和1.32×1010 CFU/g。水浴模擬試驗(yàn)優(yōu)化確定了保拉迪酵母菌抗熱保護(hù)劑配方為:脫脂乳15.12%、明膠1.8%、海藻糖9.72%。經(jīng)驗(yàn)證測(cè)得菌體存活率為17.82%,與預(yù)測(cè)值無顯著性差異。該條件下噴霧干燥制備的保拉迪酵母菌粉,菌體存活率達(dá)到26.12%,單位菌粉活菌數(shù)達(dá)到1.018×109 CFU/g。5.加速試驗(yàn)結(jié)果表明,保拉迪酵母菌凍干菌粉和噴霧干燥菌粉于-18℃(凍藏溫度)下貯存的失活速率常數(shù)分別為:k-18F=8.04×10-6和k-18S=1.04×10-5,凍干菌粉的穩(wěn)定性優(yōu)于噴干菌粉。加速試驗(yàn)測(cè)得凍干菌粉制備的保拉迪酵母菌羊奶粉在4℃和25℃時(shí)下失活速率常數(shù)分別為:k4=4.48×10-4和k25=9.72×10-3,穩(wěn)定性較好。研究確定的保拉迪酵母菌優(yōu)化培養(yǎng)基和流加補(bǔ)料培養(yǎng)條件能顯著提高菌體濃度;研究確定的凍干保護(hù)劑和抗熱保護(hù)劑用于制備保拉迪酵母凍干菌粉和噴霧干燥菌粉,能顯著提高菌粉的存活率,為保拉迪酵母菌制劑的開發(fā)提供了參考依據(jù)及技術(shù)支持。
[Abstract]:Paradi is a probiotic yeast, in 37 degrees (animal body temperature) grow well, and common yeast optimum growth temperature of 30 degrees Celsius, Paradi yeast has a unique biological activity, is suitable for probiotics for human and animal. At present, Paradi is widely used in the treatment of infantile diarrhea in yeast, yeast preparation mainly domestic Paradi the study on Preparation of dependence on imports, culture and system are rarely reported. In this study, Paradi yeast preserved in our laboratory as the test strains, the medium composition and fermentation conditions, technology and comparative study of freeze drying and spray drying method for preparing bacterial powder, powder bacteria activity and stability, and obtain the following conclusions.1. response single factor test method optimized the optimum culture conditions for yeast Paradi: Volume 52 mL/250 mL, speed of 181 r/min, 3.1% inoculation quantity, initial pH value of medium 6.4 The number of live bacteria. Up to 1.86 * 108 CFU/mL cultured under these conditions, improve the cultivation conditions of 14.1%.2. than before optimization, the single factor experimental design and Plackett-Burman screening of the main factors affecting the growth of yeast Paradi: glucose, yeast extract powder, fructo oligosaccharides, two sodium hydrogen phosphate, calcium chloride and peptone. The protein of glucose, yeast extract and fructose as the main factors, climbing test and Box-Behnken test design to obtain the optimized medium was: glucose 4.83%, yeast extract 0.18%, fructose 0.18%, two sodium hydrogen phosphate 0.45%, calcium chloride 0.2%, peptone yeast culture Paradi 2%. in the condition of live bacteria the number is up to 3.91 * 108 CFU/mL dry cell weight 5.1911 g/L, than in the control group were increased by 2.1 * 108 CFU/mL and 2.0811 g/L.3. Paradi 5 L yeast in the fermentation tank ventilation in batch culture and fed batch culture. The result showed that the feeding culture can promote Paradi yeast growth, the number of viable cells and cell dry weight can reach 7.3 * 108 CFU/mL and 12.7644 g/L. fermentor batch culture, the number of live bacteria (4.8 x 108 CFU/mL) was 1.5 times higher than that of dry cell weight (6.5612 g/L) increased nearly 2 times were established. The batch culture nonlinear fitting kinetic models of cell growth and substrate consumption in the process of using Logistic and DoseResp model. The optimized surface method: Paradi yeast formula of cryoprotectants for the response model and test data fitting good.4.: inulin 0.6%, xylo oligosaccharide 0.4%, sodium bicarbonate 0.3%, Magnesium Sulfate 0.75% 21.41%, lactose, trehalose 21.98%, monosodium glutamate 4.02%, skim milk 22%, phosphate buffer and adding bacterial mud weight ratio and survival rate of freeze-dried bacteria powder unit living bacteria number 1.2:1. Paradi yeast were 72.55% and 1.32 * 1010 CFU/g. water bath simulation test of the optimized Paradi yeast protective agents formula: 15.12% skim milk, 1.8% gelatin, trehalose 9.72%. verified the measured cell survival rate was 17.82%, no significant difference with the predicted value. The conditions of Paradi yeast powder prepared by spray drying, the cell survival rate reached 26.12%. The number of viable bacteria powder units up to 1.018 * 109 CFU/g.5. accelerated test results show that the Paradi yeast freeze-dried powder and spray drying powder at -18 DEG C (freezing temperature) under the deactivation rate constant storage were: k-18F=8.04 * 10-6 and k-18S=1.04 * 10-5, stability is better than spraying freeze-dried powder dry powder accelerated test. The measured freeze-dried powder preparation of Paradi yeast sheep milk in 4 degrees and 25 degrees the inactivation rate constants were k4=4.48 * 10-4 and k25=9.72 * 10-3, good stability. Study on yeast and determined Paradi Culture medium and feeding culture conditions can significantly improve the cell concentration; of cryoprotector and thermal protective agent for the preparation of Paradi yeast freeze-dried powder and spray drying powder, can significantly improve the survival rate of bacteria powder, to provide a reference and technical support for the development of Paradi yeast preparation.

【學(xué)位授予單位】：陜西科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：TQ926

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