本文關鍵詞： 惡臭假單胞菌 培養(yǎng)基 冷風干燥 石油降解菌 協(xié)同降解　出處：《華中農(nóng)業(yè)大學》2015年碩士論文　論文類型：學位論文

【摘要】：石油污染的生物修復是當前的研究熱點,石油降解菌大部分屬于革蘭氏陰性菌。然而,革蘭氏陰性菌普遍存在不耐干燥,不耐保藏的問題,這限制了革蘭氏陰性菌菌劑的產(chǎn)品化及其應用。假單胞菌屬于革蘭氏陰性菌,在生物修復、生物轉化、生物防治等領域有廣闊的應用前景。本文以一株能降解石油的惡臭假單胞菌(Pseudomonas putida)J7為研究對象,分別從培養(yǎng)基優(yōu)化、干燥方法的選擇、冷風干燥制備活菌粉工藝的優(yōu)化、協(xié)同菌株的選擇及其對模擬含油湖水修復等方面展開研究。具體研究結果如下:(1)研究完成了惡臭假單胞菌J7培養(yǎng)基成分的優(yōu)化。分別對碳源、氮源、無機鹽采用單因素及正交試驗,優(yōu)化得到惡臭假單胞菌的培養(yǎng)基配方為:葡萄糖6g/L、酵母粉12 g/L、Mg SO4×7H20 3.5 g/L、KH2PO4 2.5 g/L、Ca Cl2 1.5 g/L,培養(yǎng)后OD600為10.71,活菌數(shù)達到2.15×1010 CFU/m L,達到國內最高培養(yǎng)水平。(2)研究了冷風干燥制備惡臭假單胞菌菌粉的工藝。分別研究了不同載體與濃縮菌液混合比下的冷風干燥曲線,以及成品菌粉含水率、載體、載體預處理、保護劑對冷風干燥制備惡臭假單胞菌菌粉的干燥存活率及保藏穩(wěn)定性的影響。優(yōu)化后的冷風干燥制備工藝為:復合載體為硅藻土和堿處理玉米芯粉,質量比為1:2,載體與濃縮菌液混合比為1:2,保護因子(W/W)為甘露醇7%、谷氨酸鈉5%、甘油1%,成品含水率控制在7%以下。干燥菌粉活菌數(shù)達1.03×1011CFU/g,干燥存活率74.63%,顯著高于噴霧干燥(23.95%),說明冷風干燥是一種適合惡臭假單胞菌干燥的技術。優(yōu)化工藝制備的冷風干燥菌粉,保藏穩(wěn)定性顯著提高。室溫保藏150d活菌數(shù)為2.13×109 CFU/g,而未優(yōu)化的對照組菌粉,室溫保藏90 d活菌數(shù)僅為3.26×108 CFU/g;4℃保藏條件下,對照組菌粉活菌數(shù)為1.84×1010 CFU/g,優(yōu)化組活菌數(shù)為3.36×1010CFU/g,提高將近1倍。(3)初步研究石油降解菌劑的應用。篩選出與惡臭假單胞菌J7協(xié)同降解的菌株為短小芽孢桿菌Y11(Bacillus pumilus Y11)、地衣芽孢桿菌B36(Baclicus lincheniformis B36)。在含油湖水搖瓶體系中,對比單一菌株,混合菌株石油降解率提高13.0%。添加有機營養(yǎng)劑比無機營養(yǎng)劑更利于石油微生物降解,降解率提高15.2%,且降解體系C/N為20:1時較適。靜態(tài)模擬的含油湖水體系中,投加混合菌株及有機營養(yǎng)劑能使石油降解效率從20.1%提高到56.6%。且投加制備的石油降解菌菌粉的處理組,依然保持相當?shù)氖腿コ?54.7%)。
[Abstract]:Bioremediation of petroleum pollution is a hot research topic at present. Most of the petroleum degrading bacteria belong to Gram-negative bacteria. However, Gram-negative bacteria are generally intolerant to drying and preservation. This limits the production of Gram-negative bacteria and their application. Pseudomonas belong to Gram-negative bacteria, bioremediation, biotransformation, In this paper, a strain of Pseudomonas putida)J7, which can degrade petroleum, was used as the research object. The optimization of culture medium, the selection of drying methods and the optimization of the technology of preparing live bacteria powder by cold air drying were studied in this paper. The selection of synergistic strains and their remediation of simulated oil-bearing lake water were studied. The results are as follows: 1) the optimization of the composition of the culture medium of Pseudomonas oxysporum J7 was carried out. The carbon sources and nitrogen sources were obtained, respectively. Inorganic salts were tested by single factor and orthogonal test. The optimized medium formula of Pseudomonas odour was: glucose 6g / L, yeast powder 12g / L mg SO4 脳 7H203.5 g / L KH2PO4 2.5 g / L KH2PO4 2.5 g / L Ca Cl2 1.5 g / L, OD600 10.71, viable bacteria 2.15 脳 1010 CFU/m / L, which reached the highest culture level in China. The technology of Pseudomonas odoratum powder. The drying curves of cold air under the mixing ratio of different carrier and concentrated bacteria solution were studied. And the moisture content of finished product powder, carrier, carrier pretreatment, The effect of protective agent on the survival rate and preservation stability of the preparation of Pseudomonas odour powder by cold air drying. The optimized preparation technology of cold air drying is as follows: the composite carrier is diatomite and the corn cob powder is treated with alkali. The mass ratio is 1: 2, the mixture ratio of carrier to concentrated bacteria is 1: 2, the protection factor W / W is mannitol 7, sodium glutamate 5, glycerol 1, the moisture content of finished product is controlled below 7%. The number of viable bacteria in dried powder is 1.03 脳 10 11 CFU / g, and the drying survival rate is 74.63, which is significantly higher than that in spray drying. It shows that cold air drying is a suitable drying technology for Pseudomonas odour. The number of live bacteria stored at room temperature for 150 days was 2.13 脳 109 CFU / g, while that of control group was only 3.26 脳 10 ~ 8 CFU / g at 4 鈩，

本文編號：1503850