【摘要】：本研究對(duì)從東北地區(qū)自行采集的菌株進(jìn)行了分離、純化、培育,并進(jìn)行了相關(guān)的分子生物學(xué)技術(shù)鑒定和相關(guān)酶活力變化規(guī)律的研究。利用自行研制的智控益生菌液體發(fā)酵器和智控固體生態(tài)培養(yǎng).干燥一體機(jī)對(duì)菌株進(jìn)行了液體發(fā)酵與固體生態(tài)培養(yǎng)及干燥一體化研究,探尋了其相關(guān)指標(biāo)動(dòng)態(tài)變化規(guī)律,測(cè)試智控固體生態(tài)培養(yǎng).干燥一體機(jī)的運(yùn)行效果,為其工廠規(guī)模化生產(chǎn)應(yīng)用提供了必要的技術(shù)參考,具有較好的理論應(yīng)用價(jià)值。研究結(jié)果如下：(1)對(duì)自行采集分離純化的M-01菌株進(jìn)行了形態(tài)學(xué)研究和生理生化測(cè)定,并采用16SrDNA測(cè)序以及gyrA基因序列系統(tǒng)發(fā)育樹(shù)分析技術(shù)對(duì)菌株進(jìn)行了分子生物學(xué)鑒定,鑒定結(jié)果：M-01菌株為解淀粉芽孢桿菌(M-01, Bacillus amyloliquefaciens subsp.plantarum),屬于好氧菌。(2)在18℃、pH 7.2條件下,對(duì)M-01菌株進(jìn)行分泌淀粉酶、蛋白酶、脂肪酶酶活力動(dòng)態(tài)變化規(guī)律的研究,結(jié)果表明：M-01菌淀粉酶活力68h時(shí),達(dá)到較高水平13.02U/dL；蛋白酶活力在56h后趨于穩(wěn)定,至74h時(shí)為8.04U/dL：脂肪酶活力在56h時(shí)達(dá)到峰值,為1.159U/L.(3)利用自行研制的智控益生菌液體發(fā)酵器對(duì)M-01菌進(jìn)行液體發(fā)酵培養(yǎng),在適宜的液體發(fā)酵營(yíng)養(yǎng)底物組成和接種量條件下,合理有效控制發(fā)酵液溫度35±0.2℃,初始pH值7.2±0.1以及氣體攪拌方式等發(fā)酵條件,結(jié)果表明：M-01菌經(jīng)24h液體發(fā)酵,活菌數(shù)為146.675±4.05×108CFU/m L,芽孢率為91.76±1.50%；在整個(gè)發(fā)酵過(guò)程中,發(fā)酵液pH值呈下降趨勢(shì),發(fā)酵液中還原糖含量呈下降趨勢(shì),至24h時(shí),還原糖含量為5.857±0.314mg/mL;在發(fā)酵前期0-4h,發(fā)酵液中氨基氮含量增高,4-24h期間,發(fā)酵液中氨基氮含量逐漸降低,與此同時(shí),在整個(gè)發(fā)酵過(guò)程中微生物總蛋白(MCP)含量呈現(xiàn)持續(xù)上升趨勢(shì)。(4)通過(guò)L9(33)正交試驗(yàn)設(shè)計(jì),研究培養(yǎng)溫度、菌料濕度、作用時(shí)間3因素對(duì)固體生態(tài)培養(yǎng)28h后的M-01菌芽孢形成率的影響,結(jié)果表明：各因素對(duì)M-01菌芽孢形成率影響的主次順序?yàn)椋号囵B(yǎng)溫度菌料濕度培養(yǎng)時(shí)間。故在M-01菌固體發(fā)酵培養(yǎng)28h后,促進(jìn)芽孢率形成的影響因素的最佳組合為A1B3C2,即培養(yǎng)溫度45℃,菌料濕度55%,培養(yǎng)時(shí)間2h。(5)采用自行研制的智控固體生態(tài)培養(yǎng).干燥一體機(jī)對(duì)M-01菌進(jìn)行固體生態(tài)培養(yǎng)及溫敏干燥效果的研究,自動(dòng)控制發(fā)酵溫度35℃±1.0℃,菌料濕度55%±5%,并調(diào)節(jié)菌料初始pH 7.2±0.1,根據(jù)正交設(shè)計(jì)結(jié)果,采用分階段固體發(fā)酵培養(yǎng),第一階段0-28h固體培養(yǎng),控制發(fā)酵溫度35℃±1.0℃,菌料濕度55%±5%,發(fā)酵培養(yǎng)28h,以獲得較高的活菌數(shù)；第二階段28-30h固體培養(yǎng),提高發(fā)酵溫度至45℃,維持培養(yǎng)2h,以刺激芽孢的形成,以獲得較高芽孢數(shù)。結(jié)果表明：至30h時(shí),菌質(zhì)中活菌數(shù)為224.25±4.47×108CFU/g,芽孢數(shù)為203.63±4.51×108CFU/g,芽孢形成率為90.8±0.73%。在自行研制的智控固體生態(tài)培養(yǎng).干燥一體機(jī)中,培養(yǎng)終止后即刻進(jìn)行43℃±2℃溫敏脫水干燥,能有效保持菌體活性并能同時(shí)達(dá)到干燥效果,檢測(cè)到干燥菌料中芽孢數(shù)210.05±4.96×1O8CFU/g,含水率5.53±0.12%,雜菌率0.681±0.003%。
[Abstract]:In this study, the self-collected strains of the Northeast China were isolated, purified and cultivated, and the related molecular biological identification and the change of the activity of the related enzymes were carried out. The self-developed intelligent control probiotics liquid fermentation device and the intelligent control solid ecological culture are utilized. The dry all-in-one machine has carried on the research of liquid fermentation and solid ecological culture and dry integration of the strain, and the dynamic changes of relevant indexes of the strain are explored, and the intelligent control solid ecological culture is tested. the operation effect of the drying all-in-one machine provides the necessary technical reference for the large-scale production and application of the factory, and has better theoretical application value. The results of the study were as follows: (1) The M-01 strain isolated and purified by self-collection was determined by morphological study and physiological and biochemical test, and the sequence of 16SrDNA and the phylogenetic tree of gyrA gene were used to identify and identify the strain. The M-01 strain is Bacillus amyloliquefaciens (M-01, Bacillus amyloliquefaciens subsp.planarum), and belongs to aerobic bacteria. (2) The activity of amylase, protease and lipase in M-01 strain was studied under the conditions of 18 鈩，

本文編號(hào)：2419483