本文選題：曲拉 + 乳酸桿菌�。� 參考：《鄭州大學(xué)》2017年博士論文

【摘要】：本研究以青海省不同地區(qū)藏族傳統(tǒng)手工制作的牦牛乳曲拉樣品為研究對象,對其營養(yǎng)特點(diǎn)、微生物菌群組成、乳桿菌的特征和種類進(jìn)行了系統(tǒng)的分析,并首次對曲拉在制作過程中各種微生物的動(dòng)態(tài)變化和乳酸菌種類的變化進(jìn)行了研究,首次對曲拉中的益生性乳酸桿菌進(jìn)行了體外篩選,并對篩出的優(yōu)良菌株進(jìn)行了傳代穩(wěn)定性的分析,同時(shí)結(jié)合小鼠試驗(yàn)對其免疫調(diào)節(jié)、抗氧化性進(jìn)行了較為全面的功能性評價(jià),通過研究發(fā)現(xiàn):(1)從青海省不同地區(qū)采集了 43份傳統(tǒng)手工制作牦牛乳曲拉樣品,根據(jù)地區(qū)不同,抽取11份代表樣品對其pH,水分、灰分、蛋白質(zhì)、脂肪含量進(jìn)行了分析,并將分析結(jié)果同其他類型的奶酪相比較,發(fā)現(xiàn)曲拉蛋白質(zhì)的含量高于其他奶酪;利用平板計(jì)數(shù)法對43份曲拉樣品的微生物菌群組成進(jìn)行了分析,通過對曲拉中的乳酸菌、大腸桿菌、霉菌、酵母菌、好氧性細(xì)菌、梭菌、芽孢桿菌分析發(fā)現(xiàn):總體來看,乳酸菌、酵母菌是曲拉中數(shù)量上的優(yōu)勢菌。(2)根據(jù)菌落形態(tài)和革蘭氏染色的結(jié)果,同時(shí)結(jié)合生理生化特征分析、API碳源發(fā)酵試驗(yàn)、16SrRNA基因序列分析和recA試驗(yàn),43份樣品中共分離出69株乳桿菌代表株:Lactobacillus sakei(2株)、Lactobacillus diolivorans(1株)、Lactobacillus casei(28株)、Lactobacillus plantarum(34株)、Lactobacillus kefiri(4 株),Lactobacillus plantarum為優(yōu)勢種,占49%,Lactobacilluscasei 為第二大種,占 41%。(3)通過對曲拉制作過程中微生物的動(dòng)態(tài)變化進(jìn)行監(jiān)測,發(fā)現(xiàn):在曲拉制作過程中,均能不同程度地檢測到乳酸菌、大腸桿菌、好氧性細(xì)菌、絲狀真菌以及酵母菌群;乳酸菌和酵母為參與曲拉發(fā)酵的優(yōu)勢菌群;參與曲拉發(fā)酵的乳酸菌種有:Leuconostoc mesenteriodes、Enterococcus hirae、Enterococcus mundtii、Lactococcus lactis、Lactobacillus plantarum、Lactobacillus sakei、Lactobacillus buchneri、Lactobacillus diolivorans、Lactobacillus casei。在曲拉制作過程中,Leuconostocmesenteriodes 一直為優(yōu)勢種。Leuconostocmesenteriodes、Lactobacillusplantarum、Lactobacillus sakei是原料乳中固生的乳酸菌種類,在曲拉的制作過程中,這三種乳酸菌一直起主導(dǎo)作用。(4)對分離到的69株乳桿菌進(jìn)行了益生性能的篩選,采用的篩選指標(biāo)有:耐酸、耐膽鹽性能;耐模擬人體胃腸液;抑菌性能;抗生素敏感性;細(xì)胞表面疏水性。經(jīng)過篩選,有7株菌益生性能表現(xiàn)良好:Lactobacillus/splantarum 1197、Lactobacillus pl ant arum 1141、Lactobacillus kefiri1059、Lactobacillus casei1138、Lactobacillus casei1089、Lactobacillus plantarum 1086-1、Lactobacilluscasei 1133。在這 7 株菌中,綜合各種指標(biāo),選擇益生性能較其他5株菌相對優(yōu)秀的L.plantarum1086-1、L.casei1133進(jìn)行下一步的評價(jià)。(5)對L.plantaruL.086-1、L.casei1133進(jìn)行了連續(xù)傳20代遺傳穩(wěn)定性考察,從表型特征看:兩株菌的細(xì)胞形態(tài)、活菌數(shù)、產(chǎn)乳酸活力、碳源發(fā)酵類型在傳代過程中均能穩(wěn)定遺傳;從發(fā)酵特性看:L.plantarum1086-1、L.casei1133發(fā)酵性能穩(wěn)定性均較差,L.plantarum1086-1從第15代開始,發(fā)酵性能明顯下降,L.casei1133從第10代開始,發(fā)酵性能明顯下降;從益生特性穩(wěn)定性看:兩株菌的耐酸-膽鹽性、對模擬人體胃腸液的耐受性、細(xì)胞表面疏水性、抗生素抗性在傳代過程中均能穩(wěn)定遺傳。(6)通過BALB/c小鼠體內(nèi)試驗(yàn)對L.plantarum1086-1、L.casei1133進(jìn)行了免疫調(diào)節(jié)功能的評價(jià),結(jié)果表明:不同劑量的L.plantarum1086-1和L.casei1133均能極顯著提高小鼠小腸派伊爾結(jié)巨噬細(xì)胞的吞噬活性(P0.01),前者無劑量依賴性,后者呈現(xiàn)劑量依賴性;高劑量的L.plantarum1086-1和L.casei1133菌液對小鼠IgA的分泌沒有影響,但能顯著促進(jìn)機(jī)體IgG、IgM的分泌(P0.05,P0.01);高劑量的L.plantarum1086-1 和 L.casei1133均能極顯著提高小鼠血清中IL-6,IK-10,IFN-γ和TNF-α的水平(P≤0.001);不同劑量的L.plantarum1086-1和L.casei1133均能極顯著上調(diào)小鼠肝臟IL-6,IL-10,IFN-γ 和 TNF-a mRNA 表達(dá)水平(P0.01)。(7)通過 BALB/c 小鼠體內(nèi)試驗(yàn)對 L.plantarum1086-1、L.casei1133 進(jìn)行了抗氧化功能的評價(jià),結(jié)果表明:高劑量的L.plantarum1086-1能極顯著提高血清抗氧化能力(P0.01),L.casei1133對血清的抗氧化能力的提高程度不太顯著,但有改善的趨勢;不同劑量的L.plantarum1086-1和L.casei1133均能顯著上調(diào)小鼠脾臟SOD、CAT、GSH-Px mRNA表達(dá)水平(P0.05,P0.01),各劑量組之間差異顯著(P0.05,P0.01),呈現(xiàn)劑量依賴性。
[Abstract]:In this study, the samples of yak lactotra, which were traditionally made by Tibetans in different regions of Qinghai Province, were studied. The nutritional characteristics, the composition of microbial flora, the characteristics and types of lactobacillus were systematically analyzed. The dynamic changes of various microbes and the variety of lactic acid bacteria were studied for the first time in the process of making tra. For the first time, the probiotic Lactobacillus in tra was screened in vitro, and the stability of the screened strains was analyzed. At the same time, a more comprehensive functional evaluation was carried out on the immunoregulation and antioxidation in mice. Through the study, it was found that (1) 43 traditional handcrafts were collected from different regions of Qinghai province. The samples of yak lactotra were selected from 11 representative samples to analyze their pH, moisture, ash, protein, and fat content. Compared with other types of cheese, the content of tra protein was higher than that of other cheeses, and the composition of microbial flora of 43 tra samples was made by flat plate counting method. Analysis, through the analysis of lactic acid bacteria, Escherichia coli, moulds, yeasts, aerobic bacteria, Clostridium, bacillus and bacillus, we found that lactic acid bacteria and Saccharomyces cerevisiae were the dominant bacteria in the number of tra. (2) according to the colony shape and the fruit of Gram stain, and the analysis of physiological and biochemical characteristics, API carbon source fermentation test, 16SrRN A gene sequence analysis and recA test showed that 69 strains of lactobacillus were isolated from 43 samples: Lactobacillus sakei (2 strains), Lactobacillus diolivorans (1 strains), Lactobacillus casei (28 strains), Lactobacillus plantarum (34 strains), Lactobacillus kefiri (4), accounting for 49% and second Large species, accounting for 41%. (3) monitoring the dynamic changes of microbes during the production of TRA, found that lactic acid bacteria, Escherichia coli, aerobic bacteria, filamentous fungi and yeast groups can be detected to varying degrees in the production process of TRA, and lactic acid bacteria and yeast are the dominant groups involved in tra fermentation, and lactic acid fermentation is involved in lactic acid. Leuconostoc mesenteriodes, Enterococcus hirae, Enterococcus mundtii, Lactococcus lactis, Lactobacillus plantarum, Lactobacillus sakei. Es, Lactobacillusplantarum, Lactobacillus sakei are the species of Lactobacillus in raw milk. In the process of making Tra, the three kinds of lactic acid bacteria have been playing the leading role. (4) screening for the probiotic properties of 69 isolated Lactobacillus strains, the screening indexes are acid resistance, bile salt resistance, simulated human gastrointestinal fluid and bacteriostasis. Lactobacillus/splantarum 1197, Lactobacillus pl ant Arum 1141, Lactobacillus kefiri1059, Lactobacillus casei1138, Lactobacillus casei1089, Lactobacillus 1086-1, 1133. in the 7 strains. In order to synthesize various indexes, L.plantarum1086-1, L.casei1133, which was better than the other 5 strains, was selected for the next step. (5) the genetic stability of L.plantaruL.086-1 and L.casei1133 was examined for 20 generations of continuous transmission. From the phenotypic characteristics, the morphology of the two strains, the number of living bacteria, the activity of lactic acid, and the type of carbon source fermentation were found. The fermenting performance of L.plantarum1086-1, L.casei1133 was poor, and the fermentation performance decreased obviously from the fifteenth generation. The fermentation performance of L.casei1133 decreased obviously from the tenth generation. From the probiotic stability, the acid resistance of two strains of bacteria and the simulated human body were in the simulated human body. The tolerance, surface hydrophobicity and antibiotic resistance of the gastrointestinal tract were stable in the process of passage. (6) the immunoregulation function of L.plantarum1086-1 and L.casei1133 was evaluated by BALB/c mice in vivo. The results showed that the different doses of L.plantarum1086-1 and L.casei1133 could significantly improve the small intestinal pary of mice. The phagocytic activity of macrophages (P0.01), the former has no dose dependence, the latter is dose-dependent, and the high dose of L.plantarum1086-1 and L.casei1133 bacteria have no effect on the secretion of IgA in mice, but can significantly promote the secretion of IgG, IgM (P0.05, P0.01), and the high dose of L.plantarum1086-1 and L.casei1133 can greatly improve the small amount of L.plantarum1086-1 and L.casei1133. The levels of IL-6, IK-10, IFN- gamma and TNF- alpha in rat serum (P < 0.001), and L.plantarum1086-1 and L.casei1133 in different doses all significantly increased the level of IL-6, IL-10, IFN- gamma and TNF-a mRNA in mice. (7) the evaluation of antioxidative functions was carried out through the experiment in vivo. The results showed that high dose of L.plantarum1086-1 could significantly increase serum antioxidant capacity (P0.01), and the increase of L.casei1133 on serum antioxidant capacity was not significant, but there was a tendency to improve; L.plantarum1086-1 and L.casei1133 at different doses could significantly increase the mRNA expression of SOD, CAT, GSH-Px (P0.05, P0.01) in the spleen of rats. The difference between the dose groups was significant (P0.05, P0.01), showing a dose-dependent manner.
【學(xué)位授予單位】：鄭州大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：TS252.54

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本文編號：2015496