【作者】 王永生；

【导师】 袁其朋；

【作者基本信息】 北京化工大学 ， 生物化工， 2003， 硕士

【摘要】 番茄红素,类胡萝卜素的一种。在自然界中,它以全反式结构存在,并有 11 个碳碳共轭的双键。正是基于这种特殊的结构,番茄红素具有淬灭单线态氧、捕获自由基、诱导细胞间信息传递等卓越的生理功能,并具有防癌、抗癌、防治心血管疾病等功效。近年来,番茄红素的制备工艺,特别是发酵法生产,受到研究人员的重视。本文研究了利用三孢布拉氏霉菌生产番茄红素的工艺,主要包括:培养基成分的确定,培养条件的优化,阻断剂的选择,小分子效应物及菌体形态对发酵的影响以及新型色谱检测番茄红素方法的建立等五个方面。首先研究了培养基成分和培养条件对β-胡萝卜素生产的影响,确定了三孢布拉氏霉菌的种子培养基:葡萄糖 10g/L,玉米淀粉 30g/L,玉米浆 60g/L,磷酸二氢钾 0.5g/L,硫酸镁 0.25g/L, VB10.5mg/L;得到了最优的发酵培养基:玉米淀粉 40g/L,玉米浆 40g/L,棉籽油 50ml/L,磷酸二氢钾 1g/L,硫酸镁 0.25g/L,VB10.5mg/L;并优化得到了适宜的培养条件:28℃,250 转/分,装液量 40ml/250ml,pH6.5,接种量 10%,正负菌接种比例 1:10。在研究前体物和促进剂对β-胡萝卜素生产影响时发现:添加β-紫罗酮可提高产量 3.6 倍,复合前体物添加可以提高产率 4.3 倍,β-胡萝卜素产量可达到 180mg/L。该菌经物理化 I<WP=5>学诱变后,β-胡萝卜素的产量达到 830mg/L。在三孢布拉氏霉菌发酵生产β-胡萝卜素的过程中添加含氮类杂环化合物,阻断番茄红素环化酶环化反应,细胞中可积累大量番茄红素。研究了不同阻断剂的阻断效果,结果表明:吡啶衍生物的阻断效果明显高于吡啶,烟草废弃物可阻断番茄红素向β-胡萝卜素的转化,当添加 1g/L 烟草废弃物时,番茄红素的产量达到 400mg/L。研究了不同小分子效应物对三孢布拉氏霉菌合成番茄红素的影响,结果表明:trinton-X100 对菌体中番茄红素含量影响较小,span-20、β-紫罗酮和异烟肼可以提高番茄红素的产量,其中添加 span-20 可使番茄红素产量提高 3 倍。其主要原因可能是:添加煤油、trinton-X100时形成较大的菌丝球,不利于物质的传递,细胞合成番茄红素的能力受限制,而添加 span-20 则形成分散型菌丝体,有利于氧及营养物质的传递,促进了番茄红素的合成。建立了以苏丹 I 代替番茄红素作标准品进行色谱检测的方法。更多还原

【Abstract】 Lycopene is one of the important caronenoids. It is found primarily inthe all-trans configuration in the nature, which has 11 conjugatedcarbon-carbon double bonds. Lycopene has predominantly functions ofquenching singlet oxygen, scavenging of free radicals and inducing thecommunication between cells, so it could act great roles on preventingcancer and atherosclerosis. At the same time, the producing technologies,especially fermentation, have been studied. Fermentation is one of potential methods for producing lycopeneindustrially by Blakeslea trispora . The culture medium, the conditions offermentation, the choices of inhibitor, the influence of modulators andmycelial morphology have been investigated. The results showed that the best basal medium was listed as fellow:glucose10g/L, corn starch30g/L, corn steep liquor60g/L, potassiumdihydrogen phosphate 0.5g/L, magnesium sulfate0.25g/L and thiaminehydrochloride0.5mg/L. The optimized fermentation mediums was cornstarch40g/L, corn steep liquor40g/L, cottonseed oil 50ml/L, potassiumdihydrogen phosphate 0.5g/L, magnesium sulfate0.25g/L and thiaminehydrochloride 0.5mg/L . The flask cultures were incubated at 28℃ for120h under the condition of pH6.5, 250rpm,column of incubation 10%, III<WP=7>and the ratio between plus and minus 1:10. The yield of β-carotene couldreach 180mg/L when adding β -ionone , isonicotinoylhydrazine andsuccinmide as the inductors of the biosynthesis of β -carotene.Furthermore, The yield ofβ-carotene was up to 830mg/L through thefermentation by mutants which was attained through physical and chemicalmethods. Some heterocyclis substances such as pyridine and its ramificationtotally inhited the cyclization reaction of the lycopene cyclase but inducedthe production of very important quantities of lycopene. The study showedthat the role of inhibitor of pyridine’s ramification was more effective thanof pyridine itself, and found that the lycopene could attain 400mg/L whenadding 1g/L tobacco dust. The modulator could make different effect on fermentation.triton-X100 has little effect on fermentation, but span-20 and β-iononecould enhance the productivity of lycopene significantly. The content oflycopene in the medium increased 3 times of the control experiments byadding 1g/L span-20. Mycelial morphology was exploited a great influenceto the metabolites of fungi. The effect of the mycelial morphology onproducing lycopene of Blakeslea tripora was also investigated. The resultsindicated that pellets has little role on fermentation when adding keroseneand triton-x100,However, The content of lycopene in the medium couldenhance 3 times of the control experiments by adding 1g/L span-20, IV<WP=8>forming dispersed mycelia with optimizing various external factors at themacroscopic. It is essential to establish a accurate and effective method ondetermining lycopene by HPLC. The relation of content between lycopeneand sudan I was been made.更多还原