红曲菌利用生物柴油废液生产红曲色素

对生物柴油废液作简单处理,利用红曲茵发酵生物柴油废液中副产物甘油生产红曲色素。通过响应面方法确定最佳发酵培养基为：甘油48．49g/L,蛋白胨3．12g／L,K2HPO4·3H202．01g/L,MgSO4 0．48g／L,ZnSO4·7H2O 0．04g／L,MnSO4·H2O 0．03g／L,玉米浆13mL／L,植物油10mL／L,起始pH为6。发酵结果表明：在接种量6％（v／v）,转速140r／min,35℃的条件下发酵培养6d,红曲色素最高产量到达204U／mL。说明用生物柴油废液中的粗甘油为原料生产红曲色素是基本可行的。可望为生物柴油废液的资源化提供一条环境友好型的途径。     

戊糖乳杆菌发酵培养基氮源条件的优化

对戊糖乳杆菌发酵培养基的氮源条件进行了优化。通过单因素实验及响应面分析优化利用木糖高产乳酸的戊糖乳杆菌发酵培养基的不同氮源组合。优化得到的牛肉膏与柠檬酸氢二铵复合的最佳组成为牛肉膏17.72 g/L,柠檬酸氢二铵1.91 g/L,得到乳酸实际最大产量42.37 g/L。添加玉米浆与酵母粉和无机氮源复合的最佳组成为玉米浆46.54 g/L,酵母粉21.95 g/L,柠檬酸氢二铵9.95 g/L,可得到乳酸最大产量41.06 g/L。通过响应面优化减少了有机氮源的种类。牛肉膏与柠檬酸氢二铵的复合得到了更高的乳酸产量,且减少了有机氮源用量,节约了成本。玉米浆与酵母粉的复合解决了单一玉米浆造成的木糖利用速率过低的问题,同样得到较高浓度的乳酸。     

响应面分析优化产木聚糖酶毕赤酵母基因工程菌培养条件

研究不同碳源、氮源和无机盐对毕赤酵母AX181菌株产木聚糖酶的影响。实验表明,分别采用葡萄糖和玉米浆干粉为碳源和氮源可以明显提高木聚糖酶的产量。无机盐单因子优化实验显示添加适量的（NH4）2SO4、KH2PO4、MnSO4·H2O、FeSO4·7H2O也可以部分提高木聚糖酶产量。在此基础上利用响应面法优化毕赤酵母产木聚糖酶培养基,利用12次实验的Plackett—Burman设计实验筛选出影响产木聚糖酶的3个主要因素,即玉米浆干粉、MnSO4·H2O和FeSO4·7H20。并进一步通过最陡爬坡路径逼近最大响应区域,采用中心组合实验设计确定最佳条件。优化后的产木聚糖酶培养基组分为（g/L）：葡萄糖40．00,玉米浆干粉80．84,（NH4）2SO46．25,KH2PO41．25、MnSO4·H2O0．35,FeS04-7H2O1．31。培养基优化后,实际产酶2883．86u／mL,是优化前YPD培养基产酶的2．51倍。     

产酸克雷伯氏杆菌发酵产2，3-丁二醇的培养基优化

采用不同设计方法相结合的策略对耐高糖产酸克雷伯氏杆菌（Klebsiella oxytoca）ME—UD-3-4发酵产2,3-丁二醇的培养基进行优化。首先在单因素实验的基础上采用Plackett—Burrnan设计法对影响ME—UD-3-4发酵产2,3-丁二醇的相关因素进行研究,筛选到3种有显著效应的因素（P〈0．05）：葡萄糖、玉米浆和MgSO4·7H2O。然后利用响应曲面法（Response Surface Methodology,RSM）对这3种因素的最佳水平范围进一步探讨;对得到的回归模型进行分析,得最佳条件（g／L）：葡萄糖220、玉米浆19和MgSO4·7H2O 0．4;在最佳条件下,发酵80h,2,3-丁二醇产量从原来的57．3 g／L提高到86．1 g／L,生产强度由0．72g／（L·h）提高到1．08g/（L·h）。     

Perfusion culture process plus H2O2 stimulation for efficient astaxanthin production by Xanthophyllomyces dendrorhous

A semicontinuous perfusion culture process (repeated medium renewal with cell retention) was evaluated together with batch and repeated fed-batch processes for astaxanthin production in shake-flask cultures of Xanthophyllomyces dendrorhous. The perfusion process with 25% medium renewal every 12 h for 10 days achieved a biomass density of 65.6 g/L, a volumetric astaxanthin yield of 52.5 mg/L, and an astaxanthin productivity of 4.38 mg/L-d, which were 8.4-fold, 5.6-fold, and 2.3-fold of those in the batch process, 7.8 g/L, 9.4 mg/L, and 1.88 mg/L-d, respectively. The incorporation of hydrogen peroxide (H(2)O(2)) stimulation of astaxanthin biosynthesis into the perfusion process further increased the astaxanthin yield to 58.3 mg/L and the productivity to 4.86 mg/L-d. The repeated fed-batch process with 8 g/L glucose and 4 g/L corn steep liquor fed every 12 h achieved 42.2 g/L biomass density, 36.5 mg/L astaxanthin yield, and 3.04 mg/L-d astaxanthin productivity. The lower biomass and astaxanthin productivity in the repeated fed-batch than in the perfusion process may be mostly attributed to the accumulation of inhibitory metabolites such as ethanol and acetic acid in the culture. The study shows that perfusion process plus H(2)O(2) stimulation is an effective strategy for enhanced astaxanthin production in X. dendrorhous cultures.     

Production of mycelial biomass and exo-polymer by <Emphasis Type="Italic">Hericium erinaceus</Emphasis> CZ-2: Optimization of nutrients levels using response surface methodology

The Doehlert experimental design was used to optimize the production of mycelial biomass and exopolymer from Hericium erinaceus CZ-2 in this study. Statistical analysis showed that the linear and quadric terms of 3 variables: corn flour, yeast extract, and corn steep liquor had significant effects. The optimized combination of these 3 variables was confirmed through validation experiments. The optimal conditions for higher production of mycelial biomass (19.92 g/L) were estimated when the media composition concentrations were set as: 30.85 g/L, corn flour; 2.81 g/L, yeast extract; 16.9 mL/L, corn steep liquor; 10 g/L, glucose; 1 g/L, KH₂PO₄; and 0.5 g/L, MgSO₄·7H₂O; while a maximal exo-polymer yield (1.653 g/L) could be achieved when setting concentrations of: 32.71 g/L, corn flour; 2.35 g/L, Yeast extract; 14.42 mL/L, Corn steep liquor; 10 g/L, glucose; 1 g/L, KH₂PO₄; and 0.5 g/L, MgSO₄·7H₂O. The upscale production was also investigated using a 15 L fermentor using the optimized medium.     

A new strategy to enhance glutathione production by multiple H2O2 induced oxidative stresses in Candida utilis

Effect of H(2)O(2)-induced oxidative stress on glutathione (GSH) production in Candida utilis was investigated. Based on the results that H(2)O(2) can effectively stimulate GSH accumulation but inhibit cell growth simultaneously, a novel strategy of multiple H(2)O(2) stresses with different concentrations (1 mmol/L at 4h, 2 mmol/L at 8h, and 4 mmol/L at 12h) were developed to maximize GSH production. As a result, a maximal GSH yield of 218 mg/L was achieved and a corresponding intracellular GSH content was 2.15%, which were 54.6% and 58.1% higher than the control. By further applying this strategy to 7 L fermentor, GSH yield and intracellular GSH content were 328 mg/L and 2.30%. Moreover, increased activities of catalase (CAT) and GSH reductase (GR) indicated that GSH and CAT were directly involved in protecting cell against oxidative stress by H(2)O(2).     

Production of Gluconobacter oxydans cells from low-cost culture medium for conversion of glycerol to dihydroxyacetone

Gluconobacter oxydans could be immobilized as a biocatalyst for the conversion of glycerol to dihydroxyacetone. To reduce the production cost, the cells were produced from agricultural byproducts. Corn meal hydrolysate and corn steep liquor were employed to replace of sorbitol and yeast extract as medium for G. oxydans cell production. The optimal medium contained 80 g/L reducing sugar, 25 g/L corn steep liquor, and 10 g/L glycerol. The cell mass was about 4.22 g/L and the glycerol dehydrogenase activity was about 5.23 U/mL. For comparison, the cell mass was about 4.0 g/L and the glycerol dehydrogenase activity was about 5.35 U/mL cultured in sorbitol and yeast extract medium. These studies shown the corn meal hydrolysate and corn steep liquor medium was similar in performance to a nutrient-rich medium, but the cost of production was only 15% of that cultured in sorbitol and yeast extract medium. It was an economical process for the production of G. oxydans cells as biocatalyst for the conversion of glycerol to dihydroxyacetone in industry.     

Modeling of Xanthophyllomyces dendrorhous growth on glucose and overflow metabolism in batch and fed-batch cultures for astaxanthin production

An astaxanthin-producing yeast Xanthophyllomyces dendrorhous ENM5 was cultivated in a liquid medium containing 50 g/L glucose as the major carbon source in stirred fermentors (1.5-L working volume) in fully aerobic conditions. Ethanol was produced during the exponential growth phase as a result of overflow metabolism or fermentative catabolism of glucose by yeast cells. After accumulating to a peak of 3.5 g/L, the ethanol was consumed by yeast cells as a carbon source when glucose in the culture was nearly exhausted. High initial glucose concentrations and ethanol accumulation in the culture had inhibitory effects on cell growth. Astaxanthin production was partially associated with cell growth. Based on these culture characteristics, we constructed a modified Monod kinetic model incorporating substrate (glucose) and product (ethanol) inhibition to describe the relationship of cell growth rate with glucose and ethanol concentrations. This kinetic model, coupled with the Luedeking-Piret equation for the astaxanthin production, gave satisfactory prediction of the biomass production, glucose consumption, ethanol formation and consumption, and astaxanthin production in batch cultures over 25-75 g/L glucose concentration ranges. The model was also applied to fed-batch cultures to predict the optimum feeding scheme (feeding glucose and corn steep liquor) for astaxanthin production, leading to a high volumetric yield (28.6 mg/L) and a high productivity (5.36 mg/L/day).     

"Oxidative stress" response in submerged cultures of a recombinant Aspergillus niger (B1-D)

A recombinant strain of Aspergillus niger (B1-D), engineered to produce the marker protein hen egg white lysozyme, was investigated with regard to its susceptibility to "oxidative stress" in submerged culture in bioreactor systems. The culture response to oxidative stress, produced either by addition of exogenous hydrogen peroxide or by high-dissolved oxygen tensions, was examined in terms of the activities of two key defensive enzymes: catalase (CAT) and superoxide dismutase (SOD). Batch cultures in the bioreactor were generally found to have maximum specific activities of CAT and SOD (Umg x protein(-1)) in the stationary/early-decline phase. Continuous addition of H2O2 (16 mmole L(-1) h(-1)), starting in the early exponential phase, induced CAT but did not increase SOD significantly. Gassing an early exponential-phase culture with O2 enriched (25 vol%) air resulted in increased activities of both SOD and CAT relative to control processes gassed continuously with air, while gassing the culture with 25 vol% O2 enriched air throughout the experiment, although inducing a higher base level of enzyme activities, did not increase the maximum SOD activity obtained relative to control processes gassed continuously with air. The profile of the specific activity of SOD (U mg CDW(-1)) appeared to correlate with dissolved oxygen levels in processes where no H2O2 addition occurred. These findings indicate that it is unsound to use the term "oxidative stress" to encompass a stress response produced by addition of a chemical (H2O2) or by elevated dissolved oxygen levels because the response to each might be quite different.     

桦褐孔菌液体深层培养研究

研究了不同碳源、氮源、生长因子和无机盐对桦褐孔菌液体深层培养菌丝体产量的影响。采用“二次回归正交组合设计”对培养基进行优化,结果表明,当培养基配方为小麦淀粉水解糖100.83g/L(还原糖含量为24%)、酵母粉18.70g/L、玉米浆11.01g/L、KH2PO44.59g/L、MgSO4·7H2O1.5g/L、pH6.0,在30℃、160r/min的恒温培养振荡器上振荡培养120h,生物量可达19.80g/L。     

共聚焦显微技术研究ABA诱导蚕豆气孔保卫细胞H2O2的产生（简报）

The methods of confocal laser scanning microscopy (CLSM) and microinjection were used to study ABA-induced H2O2 in guard cells (Vicia faba), which were labeled with H2O2 specific probe-2, 7-dichlorofluorescin diacetate(H2DCFDA). The results indicated 100 U/mL catalase (CAT) could inhibit partly stomatal closure induced by ABA. 10(-3) mmol/L ABA could significantly induce H2O2 production in chloroplast in guard cells of Vicia faba following microinjection, and 100 U/mL CAT could partly abolish the effects following simultaneous microinjection of ABA and CAT. These suggest that H2O2 is possibly involved in ABA signaling leading to stomatal closure.     

毛栓菌产漆酶条件优化及该酶对合成染料脱色的特性

【目的】提高菌株Trametes hirsuta SYBC-L19漆酶产量,并研究该酶对合成染料脱色的性质。【方法】通过单因素和响应面设计,对产漆酶培养基进行优化。【结果】最优培养基为:玉米粉20.0 g/L、马铃薯淀粉32.4 g/L、酒石酸铵2.9 g/L、吐温80 0.5 g/L、CuSO4.5H2O 2.0 mmol/L、香兰素0.54 mmol/L、NaH2PO4.2H2O 2.0 g/L、MgSO4.7H2O0.5 g/L、MnSO4.H2O 0.1 g/L;最佳培养条件为:培养温度30°C,初始pH 6.0,装液量40 mL/250 mL,接种量8%。【结论】培养8 d酶活达35 U/mL,是优化前的39倍。对漆酶催化合成染料脱色进行了考察,发现该酶在60°C下对偶氮类染料AR1和RB5能迅速脱色,5 min内即可完成。     

氧化胁迫对Alkalibacterium sp. F26防御酶和辅因子的影响

将一株能够高产过氧化氢酶的低度嗜盐嗜碱茵Alkalibacterium sp.F26作为模式微生物,采用高效液相色谱技术测定胞内代谢物浓度,研究氧化胁迫对其防御酶活性和辅因子的影响.研究结果表明:相比低浓度H2O2(＜1 mmol/L)胁迫,此菌株在高浓度H2O2(＞1 mmol/L)胁迫下的应答表现曼为明显:经3 mmol/L H2O2胁迫后胞内CAT酶活为106.54 U/mg protein,是对照产量的1.76倍;ATP浓度则从对照浓度20.55 μmol/L下降到17.80 μmol/L;NAD 浓度自对照样品的69.89 μmol/L减少至31.77 μmol/L.由于ATP和NAD 浓度的减少,相比未经过H2O2胁迫菌体.细胞能荷值EC从0.77降低至0.68,NADH/NAD 则从0.08增加至0.41.然而,这种应答机制在细胞受到低浓度H2O2的胁迫后并不明显:除发现100 μmol/L H2O2能够导致细胞防御机制的激活而使胞内ATP浓度相比对照有所增加的情况外,经50 μmol/L和500 μmol/L H2O2胁迫后胞内ATP水平从对照的22.69 μmol/L只下降到22.38 μmol/L和13.70 μmol/L;并且此种胁迫条件下NADH浓度变化也不显著.     

共聚焦显微技术研究ABA诱导蚕豆气孔保卫细胞H_2O_2的产生(简报)

逆境下,植物细胞内ABA含量急剧增加,同时植物也可通过一些酶代谢反应积累活性氧,如H_2O_2,O_2~-。ABA作为逆境信号对气孔运动的显著调节作用已被诸多实验所证实,但关于其对气孔运动调节的细节还知之甚少。H_2O_2作为氧化信号分子在植物抗病信号转导中已得到广泛研究,但H_2O_2是否介导保卫细胞的气孔运动还缺乏直接的证据。我们已初步发现H_2O_2可参与外源ABA诱     

Copper induction of lignin-modifying enzymes in the white-rot fungus Trametes trogii

Trametes trogii, a white rot basidiomycete involved in wood decay worldwide, produces several ligninolytic enzymes, laccase being the dominant one, with higher titers than those reported for most other white rot fungi studied up to date. The effect of copper on in vitro production of extracellular ligninolytic activities was studied. CuSO(4)·5H(2)O concentrations from 1.6 μM to 1.5 mM were tested in a synthetic medium with glucose 20 g/L and asparagine 3 g/L. The addition of copper (up to 1 mM) did not affect growth but strongly stimulated ligninolytic enzyme production; faster decolorization of the polymeric dye Poly R-478 was observed as well. Maximal production of manganese peroxidase, laccase, and glyoxal oxidase [1.28 U/mL, 93.8 U/mL (with a specific activity of 720 U/mg protein), and 0.46 U/mL respectively] was attained with 1 mM CuSO(4)·5H(2)O. However, higher copper concentrations inhibited growth and notably decreased manganese peroxidase production, although they did not affect laccase secretion. Laccase activity in the culture filtrate was maximal at 50 C and pH 3.4, and the enzyme was completely stable at pH 4.4 and above, and at 30 C for up to 5 d. Denaturing polyacrylamide gel electrophoresis of extracellular culture fluids showed two laccase activity bands (mol wt 38 and 60 kDa respectively). The pattern of isoenzyme production was not affected by medium composition but differed with culture age.     

Melanin content and hydroperoxide metabolism in human melanoma cells

Human melanoma cells were grown to exponential and stationary phases showing melanin contents of 4.2 +/- 0.3 and 11.3 +/- 0.6 micrograms/10(6) cells, respectively. The cells were separated in four subpopulations by a Percoll gradient; the subpopulation of density 1.07 (g/ml) was the most enriched in pigmented cells and produced 28 and 58% of the cells in exponential and stationary phases, respectively. Melanoma cells had similar superoxide dismutase and glutathione peroxidase activities in exponential and stationary phases. Moreover melanoma cells exhibited a higher catalase activity in the stationary phase: whole homogenate and cytosol activities were 7.0 +/- 0.3 and 10.8 +/- 0.6 U/mg protein, whereas in exponential phase the activities were 4.9 +/- 0.1 and 7.6 +/- 0.3 U/mg protein for whole homogenate and cytosol, respectively. The intracellular H2O2 steady-state concentration was 3.3 +/- 0.2 and 2.1 +/- 0.2 microM H2O2 for exponential and stationary phases, respectively. The spontaneous chemiluminescence of the two culture phases was 169 +/- 27 cps/10(6) cells (exponential) and 78 +/- 24 cps/10(6) cells (stationary). The cytotoxicity of H2O2 generated extracellularly by glucose oxidase was determined after 60 min of exposure. IC50 values for exponential and stationary cell cultures were 0.9 and 2.4 mU/ml of glucose oxidase, respectively. The increased catalase activities in the stationary phase as compared with the exponential phase are consistent with the decreased intracellular H2O2, with the decreased spontaneous chemiluminescence, and with the increased resistance to exogenous H2O2.     

响应面法优化多杀菌素发酵培养基的研究

采用响应面分析方法,对刺糖多孢茵（Saccharopolyspora spinosa）H-2产多杀菌素的发酵培养基进行优化研究。运用单因子试验筛选出葡萄糖和棉籽粉为最适碳源和氮源,通过Plack—ett—Burman设计试验,对影响发酵培养基的8个相关因子进行评估并筛选出具有显著效应的4个因子：葡萄糖、棉籽粉、黄豆饼粉及玉米浆。通过最陡爬坡实验逼近以上4个因子的最大响应区域后,采用Box-Behnken响应面分析法,确定发酵产多杀菌素最佳培养基为葡萄糖64．5g,麦芽糖20g,玉米浆2g,大豆油40g,棉籽粉25g,黄豆饼粉2．4g,蛋白胨25g,CaCO35g,定容至1L,pH7．0。培养基优化后多杀菌素产量由278．1mg／L提高到508．7mg／L,比初始多杀茵素产量提高了1．83倍。     

碳源和氮源对5-酮基-葡萄糖酸生成的影响

氧化葡萄糖杆菌Gluconobacter oxydans可以将葡萄糖氧化成葡萄糖酸,并进一步氧化成2-酮基-葡萄糖酸(2KGA)和5-酮基-葡萄糖酸(5KGA),其中5KGA在催化剂的作用下能够转化为L(+)-酒石酸。为了提高5-酮基-葡萄糖酸产量,以仅生成5KGA的氧化葡萄糖杆菌Gluconobacter oxydans HGI-1为出发菌株,研究不同碳源(蔗糖、乳糖、麦芽糖、淀粉、葡萄糖)和有机氮源(酵母浸粉、鱼粉、玉米浆、黄豆饼粉、棉籽饼粉)对5KGA产量的影响。500 mL摇瓶试验结果表明,当葡萄糖浓度为100 g/L时,5KGA产量最高为98.20 g/L;当有机氮源为酵母浸粉、鱼粉和玉米浆,其添加量的蛋白含量为1.60%时,5KGA产量分别为100.20 g/L、109.10 g/L和99.83 g/L,其中,使用鱼粉的5KGA产量最高,使用玉米浆的5KGA产量比酵母浸粉略低。出于经济考虑,文中选择玉米浆作有机氮源,并在5 L发酵罐中进行分批发酵放大试验,5KGA的产量为93.80 g/L,最大生成速率为3.48 g/(L·h),平均生成速率为1.56 g/(L·h)。结果表明,葡萄糖和玉米浆分别为Gluconobacter oxydans HGI-1规模化生产5KGA的最适碳源和氮源,可利用葡萄糖几乎全部(85.93%)转化为5KGA。     

侧孢短芽胞杆菌产生线虫侵染性蛋白酶的优化

采用单因素试验确定侧孢短芽胞杆菌G4产线虫侵染性蛋白酶的最佳碳氮源,通过Placket-Burman设计筛选影响蛋白酶活力的主效因子,最陡坡试验和Box-Behnken设计获得主效因子的最佳水平,建立线虫侵染性蛋白酶的最佳生产体系:葡萄糖9.78 g/L、牛肉膏16.65 g/L、磷酸氢二钾0.75 g/L、可溶性淀粉12.5 g/L、氯化钠0.75 g/L、硫酸镁0.5 g/L、初始pH值自然、装液量50 mL,37℃摇瓶培养32 h,蛋白酶活力可达12 379.41 U/mL,较优化前的2 476.3 U/mL提高了4倍。