一株运动发酵单胞菌Zy-1快速生产乙醇

经多次实验优化，得到运动发酵单胞菌Zy发酵葡萄糖生产乙醇较合适的条件．Zy的诱变菌株Zy-1在该条件下发酵葡萄糖生产乙醇比原始菌株更有较大优势．当葡萄糖浓度为200gL^-1时，发酵48h，乙醇浓度96．5gL^-1，残糖2．3gL^-1，发酵效率为94．42％．Zy-1发酵天然原料米粉、木薯、红薯干等，发酵时间44h，乙醇浓度达95gL^-1以上，发酵效率92％以上．发酵液用DNS法测定，还原糖约2gL^-1，残总糖因原料种类不同，其值有所差异（5～20gL^-1）．经薄层层析分析，发酵液无葡萄糖，而是二糖、三糖等低聚糖．图2表5参12     

米曲霉产氨基酰化酶条件及部分酶学性质研究

通过单因子和多因子摇瓶正交优化试验,确定了米曲霉液态发酵产氨基酰化酶的最佳发酵条件.优化发酵培养基组成(ρ/gL-1):葡萄糖40,蔗糖10,可溶性淀粉20,蛋白胨2.5,马铃薯液1000mL,pH自然.培养基装量50mL/250mL三角瓶,接种量4%.培养温度30℃,转速100r/min,发酵时间42h.每50mL培养物的总酶活由优化前的2627u提高到7338u,是优化前的2.79倍.研究了米曲霉氨基酰化酶的部分酶学性质.该酶催化反应的最适pH为7.0,最适温度为40℃,低浓度的Co2 (5×10-4mol/L)对酶活激活作用显著.图5表2参8     

棒状链霉菌突变株CCRC11518-Ⅲ50克拉维酸发酵条件的研究

通过单因子和多因子摇瓶正交优化试验,确定了克拉维酸产生菌棒状链霉菌突变株Streptomyces clavuligerusCCRC11518-Ⅲ50的三角瓶发酵条件.发酵培养基组成(ρ/g L-1):甘油30,大豆蛋白胨60,麦芽浸膏7.5,K2HPO4·3H2O 1.0,MgSO4·7H2O 2.0,FeSO4-7H2O 1.0;培养基初始pH 6.5;接种量15％;培养基装量20 mL/250 mL三角瓶;培养温度25℃;发酵时间72 h.克拉维酸效价由优化前的834.8 μg mL-1提高到1 082.615 μg mL-1,提高了29.7％.还在1 600 mL发酵罐中进行了初步放大试验.当接种量15％,通气量1:0.5,转速450 r min-1,25℃发酵60h克拉维酸效价达到高峰1 025 μg mL-1.图12表1参12     

一株烟草内生拮抗放线菌发酵条件优化

为提高烟草内生放线菌Y12发酵时产生抑菌物质的产量,通过摇瓶正交优化试验,确定了Y12的三角瓶发酵条件.发酵培养基组成为:黄豆粉1.5%,蛋白胨1.5%,葡萄糖1.5%,可溶性淀粉1.5%,MgSO4.7H2O 0.05%,(NH4)SO40.25%,NaCl 0.4%,KH2PO4 0.1%,CaCO3 0.5%;培养基初始pH 7.5,培养温度30℃,发酵时间72 h.经过优化发酵液的抑菌圈直径增加了44.9%.     

不同谷氨酸浓度下钝顶螺旋藻的生长反应

通过在Zarrouk培养基中添加不同浓度的谷氨酸（Glu）对螺旋藻进行混合营养培养，研究了不同浓度的Glu对螺旋藻的生长、叶绿素、蛋白质含量以及光合作用的影响，结果表明，Glu浓度小于1．0gL^-1时，藻生长状态、生理活性都明显优于Z氏培养基，其中以在0．8gL^-1浓度时生长最好；当Glu浓度为1．0gL^-1时，藻的生长状态与Z氏培养基中极为相似；Glu浓度大于1．0gL^-1时，对藻的生长产生一定抑制作用，Glu的添加对藻液体系的pH值有一定的缓冲作用，图4表2参7     

顺丁烯二酸异构酶工程菌发酵条件优化

为提高重组大肠杆菌中顺丁烯二酸异构酶表达量,通过正交试验设计,对工程菌的生长条件和目的蛋白可溶表达条件进行优化.采用250 mL三角瓶中装有50 mL(Amp 100 mg L-1)的培养基,分别研究培养基中葡萄糖、蛋白胨、酵母浸粉的浓度,培养基pH值以及摇床转速、装液量、接种量等对蛋白可溶表达量的影响.确定顺丁烯二酸异构酶工程菌最优化培养基为:蛋白胨20 g L-1、酵母浸粉2.5 g L-1、K2HPO4·3H2O 3.0 g L-1、KH2PO4 1.5 g L-1、NaCl 6 g L-1、MgSO43 g L-1,培养基pH调至6.5.确定顺丁烯二酸异构酶工程菌可溶性表达最优条件为:37℃下培养至D600 nm值为1.0时,添加终浓度为0.05 mmol L-1的IPTG进行诱导,诱导温度37℃,摇床转速220 r min-1,装液量20%,接种量5%,诱导时长为6 h.利用BioFlo 415发酵罐以最优化的培养基和发酵条件对该工程菌进行了3批发酵实验,与摇瓶实验相比,顺丁烯二酸异构酶的表达量提高了近1.5倍,单位发酵液的酶活力由46 U mL-1发酵液提高到78 U mL-1发酵液.以上数据为顺丁烯二酸异构酶重组工程菌的中试发酵奠定了基础.图7表2参17     

产耐温蛋白酶苏云金芽孢杆菌FS140液体发酵条件优化

应用快速有效的数学统计方法对苏云金芽孢菌FS140耐温蛋白酶的发酵条件进行优化.首先采用二水平Plackett-Burman设计对影响产酶的8因素进行筛选,获得培养基成分中3个重要影响的因子:黄豆饼粉、酵母粉、葡萄糖;再利用响应面分析法对该3个因素进行3水平的优化,获得它们的最佳组合:黄豆饼粉1.8%、酵母粉0.36%、葡萄糖0.14%.优化后产酶水平达到837.71U mL-1,与响应面数学模型的预测值只有1.34%的误差.同时进行了发酵温度、初始pH、摇瓶装量与接种量等发酵条件的优化,FS140最终发酵产酶水平达918.91U mL-1.图2表5参14     

环境和营养条件对煤油生物降解的影响

研究了南极假丝酵母（Candida antarctica)JWSH-112降解煤油烃的影响因素。利用正交试验方法，确证了最优条件为煤油14（ψ/%），NaNO32.0gL61,NaH2PO40.1gL^-1，蛋白胨2.0gL^-1，初始pH6.5，其中蛋白胨是影响JWSH－112降解煤油烃的重要因素。采用优化后的条件进行验证实验，煤油烃的生物降解率（49．2％）和细胞生长质量浓度（1.9gL^-1)均加高于正交试验中的最高生物降解率（47．9％）和细胞生长质量浓度（1.5gL^-1)。通过气相色谱定性分析发现，生物降解后煤油中的部分组份消失或含量明显降低。图6表5参13     

流加发酵及添加L-半胱氨酸对产朊假丝酵母高密度培养合成谷胱甘肽的影响

通过7L罐考察了不同葡萄糖流加速率以及添加L-半胱氨酸对产朊假丝酵母(Candida utilis)WSH02-08高密度发酵生产谷胱甘肽(GSH)的影响.结果表明,在恒定速度(5.5g L-1h-1)流加葡萄糖30h的基础上,发酵45h后细胞干重最高达到73g L-1,此时向发酵罐中一次性添加L-半胱氨酸40mmol L-1,最终GSH产量和胞内GSH含量分别达到了1458mg L-1和2.26%.图4表1参11     

好氧脱氮微生物的混合培养条件

从土壤和水中筛选分离到混合脱氮微生物菌群，能在好氧条件下将NH4^ 一步转化为N2排放，整个过程无NO3^-的积累，混合脱氮微生物菌群培养的最佳碳源为NaHCO3和CH3COONa的混合物，质量浓度均为0.25gL^-1;（NH4）2SO4为氮源的最适质量浓度为0.2gL^-1；最适pH7-10；温度30℃；在混合脱氮微生物菌群的最适培养条件下，30h内氨氮去除率达98%以上，细胞生长质量浓度达2.9gL^-1，采用分批补料策略补加（NH4）2SO4使菌浓提高了31.0%，图6表4参11     

一株花生根际促生菌的筛选鉴定及其特性研究

从种植于红壤的健康花生根际,筛选出7株产吲哚乙酸(IAA)菌株,以菌株L4合成IAA的能力最强,培养24 h时IAA产生量达135.67μg.mL-1,且菌株L4具有解磷能力。通过菌株形态、生理生化特征测定及16SrRNA的保守序列鉴定,初步确定菌株L4为氯酚节杆菌(Arthrobacter chlorophenolicus),其GenBank登录号为JQ277449。菌株生长和发酵条件试验结果表明,菌株L4生长和分泌IAA的最佳培养条件并不完全一致,既能促进菌株生长又能合成较多IAA的最佳培养条件是初始pH值为5～6,装液量为50 mL.(250 mL)-1,30℃摇床培养24 h;促进菌株生长的最佳碳、氮源分别是麦芽糖和酵母粉,而提高IAA产生量的最佳碳源是木糖,最佳氮源是KNO3。     

固定化细胞生产L-苯丙氨酸研究

用聚乙烯醇（ＰＶＡ）凝胶包埋固定假单胞菌Ｅ４－１０６细胞,最佳ρ（ＰＶＡ）为８０ｇ／Ｌ;最适湿细胞包埋量为每ｍｌ凝胶０．３ｇ;转氨酶活力回收率为７８％,连续反应２１批次后,固定化细胞酶活力为初始最高活力的９１％;用固定化细胞进行间歇式批次反应,结果表明其酶活性半寿期为４个月;制得的固定化细胞为直径约３ｍｍ的珠状颗粒,机械强度良好,间歇式批次反应１０个月,固定化细胞凝胶颗粒无溃破     

深黄伞形霉(Umbelopsis isabellina)华2-1产油发酵培养条件优化

为开发微生物油脂资源,从富含油脂的土样中分离获得一株产油脂微生物——深黄伞形霉(Umbelopsisisabellina)华2-1,油脂含量达48.60%.采用单因素和正交实验方法,对深黄伞形霉华2-1的发酵培养条件进行优化研究.优化的培养条件为:葡萄糖100 g/L,酵母粉3 g/L,接种量为20%,初始pH值为5.0～6.0,MgSO4.7H2O和KH2PO4的添加浓度分别为0.5 g/L和2 g/L,培养温度为31℃,最佳发酵培养时间为168 h.华2-1在优化发酵条件下可获得菌体生物量、油脂产量和油脂含量分别为45.86 g/L、24.47 g/L和59.53%,较优化前分别提高了21.48%、33.42%和22.49%.因此,深黄伞形霉华2-1作为微生物油脂生产的新资源具有广阔的应用前景.     

由一株青霉菌产生的聚乙烯醇降解酶

从纺织污水活性污泥中筛选得到一株新型聚乙烯醇(PVA)降解酶产生菌,根据形态学特征鉴定该菌属于青霉属(Penicillium sp.),实验室编号WStt02-21.这是由霉菌产生PVA降解酶的首例报道.在考察了菌株WSH02-21基本生长和产酶特性的基础上,研究了营养条件对PVA降解酶合成的影响.通过对营养条件的单因素考察和正交试验,确定了最优培养条件为PVA 40 g L-1、葡萄糖3.0 g L-1、NH4Cl 8.0 g L-1、KH2PO4 2.0 g L-1、酵母膏1.0 g L-1、:MgSO40.5 g L-1。、CaCl2 1.0 g L-1、NaCl 0.02 g L-1、FeSO4·7H2O 0.02 g L-1,初始pH 6.4.其中PVA浓度是影响Penicilliumsp.WSH02-21合成PVA降解酶的最重要因素.采用最优化条件进行验证试验,PVA降解酶酶活(4.4 U mL-1)略高于正交试验中的的最高酶活(4.3 U mL-1).图7表2参11     

氯化二丁基锡对小鼠外周血液中睾丸酮和雌二醇含量的影响

用放射免疫法分别测定了DBTCl对妊娠d6雌小鼠和成年雄小鼠外周血液中雌二醇和睾丸酮水平的影响.每天按小鼠体重一次腹腔注射0.025～0.400μg/kgd-1DBTCl，共染毒7d.实验条件为(22±2)℃和光暗=12h12h.结果表明，处理组睾丸酮和雌二醇的含量随处理剂量的增加而增加，当w（DBTCl）≥0.05μg/kg时，二者呈明显的剂量-效应关系.雌二醇含量的增加比睾丸酮更为显著CK雄性小鼠睾丸酮浓度为（2.88±0.72）ng/mL，在0.40μg/kg剂量组达到(9.95±2.5)μg/mL时，约为CK的3.5倍；而CK和0.40ng/kg剂量组妊娠小鼠雌二醇浓度分别为(30.32±5.43)ng/mL和(287.57±51.13)ng/mL，相差约9.5倍.文中还就DBTCl对小鼠血液中激素含量的影响机理进行了初步探讨.图1表2参20     

五氟苯甲酰氯衍生色谱法分析水体中痕量羟基多溴联苯醚

以五氟苯甲酰氯(PFBCl)作为衍生化试剂,气相色谱-电子捕获检测器(Gas chromatography-electron capture detector,GC-ECD)为检测手段,建立了水体中痕量羟基多溴联苯醚(Hydroxylated polybrominated diphenyl ethers,简称OH-PBDEs)的酰基化衍生-GC/ECD检测方法.通过实验讨论了衍生化缓冲溶液、衍生化时间以及衍生后萃取溶液异辛烷的用量对检测结果的影响,确定了最佳的衍生化条件.在最佳的衍生化条件下,4-OH-BDE 42、4’-OH-BDE 49、6-OH-BDE 47、2’-OH-BDE 68和6’-OH-BDE 99的衍生化产物均获得了良好的分离度和色谱响应值.本方法的仪器检出限为0.02—0.14 pg,方法检出限为0.61—4.67 pg.L-1,标准曲线线性相关系数的范围为0.9973—0.9996,相对标准偏差均小于10%.3种采自大连市不同地区环境水样的加标回收率检测结果表明,所建立的分析方法适于水体中痕量OH-PBDEs的检测.     

Recovery of Ni(II) from real electroplating wastewater using fixed-bed resin adsorption and subsequent electrodeposition

Resin adsorption and subsequent electrodeposition were used for nickel recovery. Treated wastewater can meet the Electroplating Pollutant Discharge Standard. The spent resin is completely regenerated by 3 BV of 4% HCl solution. 95.6% of nickel in concentrated eluent was recovered by electrodeposition. Effective recovery of high-value heavy metals from electroplating wastewater is of great significance, but recovering nickel ions from real electroplating wastewater as nickel sheet has not been reported. In this study, the pilot-scale fixed-bed resin adsorption was conducted to recover Ni(II) ions from real nickel plating wastewater, and then the concentrated Ni(II) ions in the regenerated solution were reduced to nickel sheet via electrodeposition. A commercial cation-exchange resin was selected and the optimal resin adsorption and regeneration conditions were investigated. The resin exhibited an adsorption capacity of 63 mg/g for Ni(II) ions, and the average amount of treated water was 84.6 bed volumes (BV) in the pilot-scale experiments. After the adsorption by two ion-exchange resin columns in series and one chelating resin column, the concentrations of Ni(II) in the treated wastewater were below 0.1 mg/L. After the regeneration of the spent resin using 3 BV of 4% (w/w) HCl solution, 1.5 BV of concentrated neutral nickel solution (>30 g/L) was obtained and used in the subsequent electrodeposition process. Using the aeration method, alkali and water required in resin activation process were greatly reduced to 2 BV and 3 BV, respectively. Under the optimal electrodeposition conditions, 95.6% of Ni(II) in desorption eluent could be recovered as the elemental nickel on the cathode. The total treatment cost for the resin adsorption and regeneration as well as the electrodeposition was calculated.     

Antioxidant enzymes in the digestive gland of the common musselMytilus edulis

Antioxidant enzymes function to remove deleterious reactive oxygen species, including the superoxide anion radical and H₂O₂. Subcellular distributions and optimal and other properties of catalase (EC. 1.11.1.6), superoxide dismutase (SOD; EC. 1.15.1.1), selenium-dependent glutathione peroxidase (Se-GPX; EC. 1.11.1.9) and total glutathione peroxidase (GPX) activities were determined in the digestive gland of the common musselMytilus edulis L. by spectrophotometric and cytochemical/electron microscopic (catalase) techniques. Assay conditions for Se-GPX and total GPX activities were determined which optimized the difference between the non-enzymic and enzymic rates of reaction. General peroxidase activity (guaiacol as substrate) (EC. 1.11.1.7) was not detectable in any subcellular fraction. Catalase was largely, if not totally, peroxisomal, whereas SOD and GPX activities were mainly cytosolic. Distinct mitochondrial (Mn-SOD) and cytosolic (CuZn-SOD) SOD forms were indicated. Catalase properties were consistent with a catalase, rather than a catalase-peroxidase. The pH-dependence and temperature-dependence of GPX activity were different with H₂O₂ or CHP as substrate, and these and other observations indicate the existence of a distinct Se-GPX. Under saturating or optimal (GPX) assay conditions, the apparent Michaelis constantsK _m (mM) were: catalase, 48 to 68 (substrate, H₂O₂); Se-GPX, 0.11 (H₂O₂) and 2.0 (glutathione); and total GPX, 2.2 (eumene hydroperoxide) and 1.2 (glutathione). Calculated catalase activity was 2 to 4 orders of magnitude greater than Se-GPX activity over an [H₂O₂] of 1 to 1000 M. The results are discussed in relation to theoretical calculations of in vivo oxyradical production and phylogenetic differences in antioxidant enzyme activities.     

Optimization and modeling of coagulation-flocculation to remove algae and organic matter from surface water by response surface methodology

Charge neutralization and sweep flocculation were the major mechanisms. Effect of process parameters was investigated. Optimal coagulation conditions were studied by response surface methodology. ANN models presented more robust and accurate prediction than RSM. Seasonal algal blooms of Lake Yangcheng highlight the necessity to develop an effective and optimal water treatment process to enhance the removal of algae and dissolved organic matter (DOM). In the present study, the coagulation performance for the removal of algae, turbidity, dissolved organic carbon (DOC) and ultraviolet absorbance at 254 nm (UV₂₅₄) was investigated systematically by central composite design (CCD) using response surface methodology (RSM). The regression models were developed to illustrate the relationships between coagulation performance and experimental variables. Analysis of variance (ANOVA) was performed to test the significance of the response surface models. It can be concluded that the major mechanisms of coagulation to remove algae and DOM were charge neutralization and sweep flocculation at a pH range of 4.66–6.34. The optimal coagulation conditions with coagulant dosage of 7.57 mg Al/L, pH of 5.42 and initial algal cell density of 3.83 × 10⁶ cell/mL led to removal of 96.76%, 97.64%, 40.23% and 30.12% in term of cell density, turbidity, DOC and UV₂₅₄ absorbance, respectively, which were in good agreement with the validation experimental results. A comparison between the modeling results derived through both ANOVA and artificial neural networks (ANN) based on experimental data showed a high correlation coefficient, which indicated that the models were significant and fitted well with experimental results. The results proposed a valuable reference for the treatment of algae-laden surface water in practical application by the optimal coagulation-flocculation process.     

Optimization of fermentation process for saccharide-based fuel ethanol production by Saccharomyces cerevisiae北大核心CSCD

低成本、高产量的发酵工艺是实现工业燃料乙醇经济和环境可持续性发展的关键,而不需要重大基础设施改变或投资.为获得酿酒酵母(Saccharomyces cerevisiae)利用甘蔗汁生产燃料乙醇的最优发酵工艺,首先对发酵体系的氮源条件进行优化;其次,在单因素试验基础上,以乙醇发酵效率为响应值,通过响应面法优化了燃料乙醇生产的发酵工艺,并通过补料分批发酵技术在5 L发酵罐中进一步扩大发酵.结果表明,以1.0 g/L (NH)SO和1.0 g/L酵母提取物作为发酵氮源,乙醇发酵效率和得率比对照可分别提高4.80%、9.52%.响应面设计获得的最优发酵工艺条件为在总糖浓度150.0 g/L、酵母提取物浓度2.0 g/L、发酵时间24.5 h、pH5.0、外加(NH)SO浓度1.0 g/L时,最高乙醇发酵效率可达到91.10%.在5 L发酵罐中采用补料分批发酵获得的最终乙醇浓度达到98.92 g/L,发酵效率维持在90%左右,乙醇生产力最高达到3.81 g Lh.本研究获得了一种高效生产糖质燃料乙醇的发酵工艺,可在较短时间内获得高浓度乙醇且消耗较少氮源,结果可为进一步利用糖质原料进行高效生物炼制及高浓度乙醇工业化生产提供参考.(图6表6参30)