重组Pichia pastori高密度培养和表达过程的溶氧调控

在重组人血清白蛋白（recombinant human serum albumin，rHSA）Pichia pastori表达系统中，研究了高密度培养时细胞生长、改变碳源和诱导表达时溶氧的变化和调控规律，溶氧与细胞生长和重组蛋白表达密切相关。发酵前期的细胞生长阶段，细胞需氧降低表明碳源甘油的消耗，根据溶氧水平适时流加碳源甘油；改变碳源前，应停止流加甘油碳源，并以溶氧指标判断甘油是否消耗完全；重组蛋白诱导时流加甲醇的速度应缓慢增大，并保持细胞一定的需氧水平。高密度培养时通过流加底物速度、搅拌转速、通气量和富氧培养来关联控制发酵液溶氧水平。     

重组复合干扰素摇瓶发酵条件的研究

目的对表达重组复合干扰素(con-IFN)的工程菌E.coli DH5α的摇瓶发酵条件进行研究。方法利用单因素比较实验及正交试验等方法考查工程菌摇瓶发酵条件,探讨发酵条件对工程菌的生长和con-IFN表达的影响。结果优化后条件为培养基含玉米粉6%、牛肉膏5.5%、谷氨酸钠0.5%,发酵液初始pH 7.2,30℃培养7 h后升温至42℃诱导表达6 h,转速220r/min,装样量10%,接种量2%。结论优化后平均蛋白质表达量达到36.39%,和LB培养基相比,目的蛋白质表达量提高了4.8%,为工程菌的大规模发酵提供参考。     

产腺苷蛋氨酸酵母菌株的选育

目的筛选产腺苷蛋氨酸(SAM)的酵母菌并进行诱变育种。方法以选择性培养基筛选酵母菌株并用高效液相色谱检测SAM。利用紫外线和γ-射线处理对菌株进行诱变。结果从34份土样中筛选到1株产SAM的酵母菌Q95菌株,经过5轮紫外线诱变和1轮γ-射线诱变,筛选到1株SAM产量显著提高的正突变株Q6-13。摇瓶发酵27 h后,其SAM产量达到1860μg/mL,与Q95相比提高了86.9%。在15 L外循环气升式生物反应器中补料发酵22 h后,Q6-13的SAM产量达到2540μg/mL。结论通过筛选和诱变,得到了1株高产SAM的酵母菌突变株,为SAM的微生物发酵法规模化生产奠定了基础。     

瓜子金发酵总皂苷的抗炎镇痛活性

目的:研究瓜子金发酵液中所含总皂苷的抗炎镇痛活性。方法:提取瓜子金发酵液中的总皂苷,采用二甲苯诱导小鼠耳廓肿胀试验模型和腹腔毛细血管通透性试验观察抗炎作用;采用小鼠扭体和热板试验观察镇痛作用;采用最大耐受剂量法进行急性毒性试验,观察小鼠出现的毒性反应和剂量毒性效应的关系。结果:瓜子金发酵总皂苷对二甲苯致小鼠耳廓肿胀、冰醋酸致腹腔毛细血管通透性增高均有明显的抑制作用;能减少冰醋酸刺激致痛小鼠的扭体次数及提高热板致痛小鼠的痛阈值;小鼠灌胃最大耐受剂量13.3g.kg-1,相当于临床用量的100倍。结论:瓜子金发酵总皂苷具有显著的抗炎、镇痛作用,且毒性较低。     

Effect of Colonic Lactulose Availability on the Timing of Drug Release Onset in Vivo from a Unique Colon-Specific Drug Delivery System (CODES™)

Purpose. To test the hypothesis that the onset of drug release in vivo from a unique colon-specific drug delivery system (CODES) would depend on the colonic availability rate of lactulose. The site specificity of drug release in canine GI tract was also estimated. Methods. CODES tablets were prepared by tableting the granulation of acetaminophen and lactulose, followed with film coating. The pharmacokinetic performance of different CODES formulations was evaluated in six beagle dogs under fasted conditions. The release of acetaminophen and lactulose was also characterized in vitro. Results. The onset of acetaminophen release in beagle dogs was found to be dependent on the coating level of Eudragit E and lactulose loading in the core tablet. At Eudragit E coating levels of 4%, 8%, and 12% (coating weight gain), the onset of in vivo drug release occurred 5.5 (±1.9) h, 4.8 (±1.0) h. and 7.5 (±1.0) h, respectively, after dosing. A similar trend was observed when the loading of lactulose in the core tablet decreased from 78% to 58% and 38%. However, the rate and extent of acetaminophen absorption did not vary significantly in each situation based on the values of AUC and C_max. Conclusions. The onset of drug release in vivo from CODES tablets is predominantly dependent on colonic availability rate of lactulose because drug release from this system is triggered by localized drop of colonic pH from the fermentation of lactulose.     

L-缬氨酸高产菌选育及营养需求研究

以黄色短杆菌 (Brevibacteriumflavum)L 缬氨酸产生菌ZQ -2为出发菌株 ,经硫酸二乙酯(DES)和亚硝基胍 (NTG)诱变处理 ,氨基酸结构类似物定向筛选 ,获得一株L 缬氨酸高产菌XQ -6 (LeulAHVrα ABhr2 TAhr)。在以 1 4 %葡萄糖为碳源、5 %硫酸铵为氮源的培养基中直接发酵72h,产酸达 5 8～ 6 2g/L。同时添加Fe2 + 和Mn2 + 比单独添加Fe2 + 更有效。还进行了氨基酸、核酸、维生素和其它营养对L 缬氨酸发酵影响的试验。     

城市生活垃圾焚烧前堆酵脱水研究进展

分析了垃圾含水率及其与低位热值的关系,依据低位热值对垃圾焚烧工艺的影响,概述了垃圾焚烧前堆酵脱水的机理、研究现状,探讨了控制温度和接种微生物在加速堆酵脱水中应用的可行性,并对今后该领域的研究重点提出建议。     

超滤膜分离技术在植酸酶浓缩中的应用

目的 利用超滤法在低温下操作,生物活性物质不易失活的特点,探讨超滤法代替薄膜蒸发法浓缩植酸酶发醇液的技术可行性,以降低能耗。方法 采用截留分子量为２００００的ＰＵ－２０Ｋ－ＰＳ管式超滤膜系统浓缩１～５批次的植酸酶发醇液,测定浓缩倍数、浓缩收率和截留率并观察超滤过程的膜通量及其变化。结果 植酸酶的浓缩倍数为６．５３倍,浓缩收率为９９．６９％,截留率为９９．９３％,系统可以在植酸酶的工艺条件下连续浓缩〉１０ｈ,且经过简单清洗后,膜通量基本恢复。结论 用ＰＵ－２０Ｋ－ＰＳ管式超滤膜系统缩植酸酶发醇液在技术上可行。     

贵州豆豉发酵前后大豆异黄酮含量测定

目的：建立一种测定豆豉中大豆异黄酮类成分含量的分析方法,并对黄豆及其发酵后豆豉中的大豆异黄酮含量进行比较。方法：采用反相高效液相色谱法,色谱柱Diamonsil-C18（250 mm×4.6 mm,5μm）,保护柱Phenomenex—C1s（4.6mm×3 mm,5μm）,流动相乙腈-0.2％磷酸(36∶64),流速...     

Production of Clostridium difficile toxin in a medium totally free of both animal and dairy proteins or digests

In the hope of developing a vaccine against Clostridium difficile based on its toxin(s), we have developed a fermentation medium for the bacterium that results in the formation of Toxin A and contains no meat or dairy products, thus obviating the problem of possible prion diseases. Particular preparations of hydrolyzed soy proteins, especially Soy Peptone A3, have been found to replace both the meat/dairy product tryptone in the preparation of working cell banks and seed media, and NZ-Soy BL4 does the same in the fermentation medium. These replacements yield even higher toxin titers.