产木聚糖酶菌株的筛选及产酶条件优化

通过透明圈法从土壤中筛选到两株产木聚糖酶菌株CS-1和CS-2,CS-2产木聚糖酶的酶活力优于CS-1。CS-2产木聚糖酶最佳发酵培养条件:碳源(甘蔗渣)3.0 g/ml,氮源(NH4Cl)0.5 g/ml,接种量为10%,250 ml三角瓶装液量50 ml,pH=8.0,温度为55℃,转速为160 r/min,发酵时间为3 d。在此发酵条件下,CS-2产木聚糖酶的酶活力达到7.980 U/ml。     

绿色木霉的选育及固态发酵产纤维素酶的研究

从纤维素酶产生菌绿色木霉TY-2出发,通过紫外诱变技术选育出1株遗传稳定性良好的高产菌株H-28,其产纤维素酶的滤纸酶活力稳定在2.67 U/g左右,较出发菌株提高58.08%。以麸皮和蔗渣为主要原料对变异株H-28进行固态发酵研究,单因素优化了培养基、培养条件和表面活性剂对菌株H-28发酵产纤维素酶的影响,最后选取影响产酶较大的4个因素:发酵时间、Mandels营养盐液、蛋白胨、吐温-80做4因素3水平正交试验。最终优化后突变菌株H-28的产酶能力最高值为6.79 U/g,是出发菌株的4.07倍。     

耐酒精半纤维素降解菌的筛选、鉴定及产酶分析

从石油污染严重地区的贫瘠土壤中筛选得到的一株高产木聚糖酶且能够耐受一定浓度酒精的细菌,经生理生化实验及16SrDNA鉴定为短小芽孢杆菌(Bacillus pumilus),命名为DT83。对其产木聚糖酶的发酵条件进行优化,结果表明:最适碳源为木聚糖,最适氮源为蛋白胨,产酶最适温度为28℃,最佳初始pH值为8.5。SDS-PAGE分析表明,木聚糖酶分子量约为23kD。DT83和E.coli BL21的酒精耐受实验结果比较表明,DT83具有较高酒精耐受度,可达到8%～10%。     

优化产酶培养基用于稻草发酵产乳酸的研究

对利用稻草为碳源发酵产纤维酶的过程从菌种选择、培养基的选择、碳源的结构、含水率进行了优化。以黑曲霉及里氏木霉的混合菌株为产酶菌种,在稻草粉和麦麸以3:1的比例混合作碳源,培养基中含水率为70%时发酵产出的纤维素酶酶活达到最高,最高的Cx酶活为1568.47U/g,滤纸酶活为489.3U/g。以该条件下产的纤维素酶分别用短乳杆菌和米根霉进行乳酸发酵实验,产乳酸结果为:短乳杆菌:10.8g/L;米根霉:9.2g/L。     

β-葡萄糖苷酶的异源表达及与纤维素酶协同酶解竹纤维

在毕赤酵母GS115中表达东方肉座菌EU7-22的β-葡萄糖苷酶基因（bgl?）,获得基因工程菌株BP17。优化BP17发酵产酶条件后,重组β-葡萄糖苷酶活力达121 IU/mL。酶学性质研究表明,该酶最适反应温度为70℃,在60℃以下有较好的热稳定性;最适催化pH为5.0,在pH 3.0 ~ 8.0之间有较好的稳定性。将异源表达的β-葡萄糖苷酶添加到东方肉座菌的纤维素酶液中协同降解经过预处理的竹纤维,当纤维素酶添加量为FPA 20 IU/g底物,β-葡萄糖苷酶添加量为BG 6 IU/g底物时,纤维二糖浓度显著下降,酶解得率达到83.03%,表明重组β-葡萄糖苷酶的加入更有利于纤维素的酶解糖化。     

纤维素乙醇木聚糖酶的固体发酵工艺研究

该研究立足于河南天冠企业集团纤维素乙醇项目,以酶解糖化工艺对木聚糖酶的需求为出发点,利用黑曲霉X06作为产酶菌株,采用固体发酵工艺,通过正交设计试验,优化了培养基配方和发酵控制工艺,最优方案:麸皮∶玉米芯=6∶4、硝酸铵4%、尿素1%、磷酸二氢钾0.4%、硫酸镁0.2%、初始含水量65%、初始pH=4.0、温度28℃、环境相对湿度70%、72 h酶活达到10 096.74 IU/g。这一方案在生产中得到进一步放大和优化,所生产的固体木聚糖酶应用在秸秆酶解工艺中,酶解液中木糖含量提高了66.9%。     

脂肪酶产生菌种的筛选鉴定及产酶条件优化

从富油土壤中分离筛选到一株脂肪酶产生菌M-4,通过对M-4进行形态和r DNA ITS序列鉴定,确定其属于酵母菌Sporidiobolus salmonicolor strain。通过正交试验设计对该菌株进行产酶条件探索,得出其摇瓶培养最适产酶条件为:种子培养时间为90 h,最佳发酵培养基碳源为葡萄糖2%+橄榄油3%,氮源为5%酵母膏,初始p H值为7.0,发酵温度为30℃,在此条件下的产酶可达12.2 U/m L。     

Brønsted酸性离子液体合成及催化制备生物柴油

合成4种成功能化酸性离子液体,采用红外光谱、热重分析等分析法进行表征验证,并用其催化菜籽油酯交换制备生物柴油,考察醇/油物质的量之比、反应温度、反应时间、离子液体用量和水含量对转化率的影响。结果表明,4种离子液体都有较强酸性,与浓硫酸酸性相当;带—SO₃H基团的离子液体表现出更好的催化活性,且随着烷基链的增加,催化活性提高;在（n甲醇）∶n（菜籽油）=12∶1,反应温度130 ℃,反应时间3 h,离子液体（[BSO₃HMIM][HSO₄]）用量为菜籽油质量2%（质量分数）条件下,生物柴油转化率可达99%以上。在反应体系中,水会破坏离子液体的结构并导致其失活,而升高反应温度,可缓解水对离子液体的结构破坏,在130 ℃条件下,即使水分含量为5%时,生物柴油转化率仍可保持在约85%。     

利用农业废弃物制取纤维素酶的优化试验研究

以绿色木霉突变株为出发菌株,农业废弃物小麦秸秆和麸皮为主要原料,利用固体发酵法制取纤维素酶并对其发酵条件进行优化。首先通过正交试验设计,考察了秸秆与麸皮质量比、料水比、初始pH值、表面活性剂含量、硫酸铵含量、磷酸氢二钾含量、硫酸镁含量等7个因素对产酶的影响;利用正交试验数据,建立了BP神经网络模型并运用遗传算法对模型优化,得到了优化的固体发酵条件:秸秆与麸皮质量比为3.7:1.3,原料与水质量比为1:2.1,初始pH为4.5,表面活性剂为1.90%,(NH_4)_2SO_4 1.50%,K_2HPO_40.89%,MgSO_4·7H_2O 0.12%(以上均为原料质量分数)。经试验验证,所产纤维素酶酶活达到108.643U/g,相比初始固体发酵条件酶活提高了约38.6%。     

混合固态发酵降解甘蔗渣产糖产乙醇研究

以碱处理甘蔗渣为原料,比较里氏木霉(Trichoderma reesei CICC40359)和斜卧青霉(Penicillium decumbens LSM-1)单菌和混菌固态发酵及转化乙醇效果,研究发酵过程菌体生长、产酶产糖和乙醇转化情况。结果表明:混菌发酵效果优于单菌,在接种量8%,发酵温度30℃,混菌固态发酵(SSF)72h后总糖和还原糖产量最大值为20.21 g/L和12.47 g/L;β-葡萄糖苷酶活力和菌体生物量在144 h后分别达到0.48 IU/m L和0.21 g/g DM;对发酵3 d后底物(包括生成糖、合成酶及未降解基质)接种酵母进行乙醇同步糖化发酵,乙醇浓度在发酵24 h时达到5.83 g/L,发酵效率达到理论值的40.84%。利用多菌混合固态发酵转化底物产乙醇能避免传统乙醇生产过程高成本纤维素酶的应用,为纤维乙醇生产提供一条经济有效的新途径。     

蒸汽爆破预处理玉米芯及其酶解工艺研究

以蒸汽爆破预处理后的玉米芯为原料,进行了玉米芯酶解工艺条件的研究。粉碎后的玉米芯在压力2.8 MPa、保压时间240 s条件下蒸汽爆破预处理,在初始固形物含量为14%(w/v),pH 5.0的条件下,分别添加纤维素酶15 FPA/g(底物)、木聚糖酶225 IU/g(底物),同时添加环境因子MgSO40.005 g/g(底物)、Tween-800.001 g/g(底物),糖化48 h后,还原糖浓度达到71.81 g/L,糖化率达到80.85%。试验结果表明,蒸汽爆破预处理及添加适量环境因子对玉米芯的糖化效果影响显著。     

Enzymatic depolymerization of Ricinus communis, a potential lignocellulosic for improved saccharification

Residual lignocellulosics left to decay in fields and forest has a huge potential to serve as a low cost feedstock for production of bioethanol. In Indian subcontinent Ricinus communis is a major lignocellulosics growing in arid conditions containing 42% cellulose and 19.8% lignin. In the present study, Response Surface Methodology (RSM) based on Central Composite Design (CCD) has been used to explore the effects of pH, temperature, solid to liquid ratio (w/v), enzyme concentration and incubation time on enzymatic depolymerization of R. communis. The maximum delignification obtained was 85.69%. In case of lignified R. communis the optimum reducing sugar produced was about 288.83 mg/g dry substrate, whereas, in case of delignified R. communis the optimum reducing sugar produced was about 775.17 mg/g dry delignified substrate. After delignification reducing sugar yield was increased to about 2.68 fold.     

Optimization of acidic hydrolysis conditions of rice husk for fermentable sugar production

Lignocellulosic biomass (LSB) is the most abundantly available renewable source in the world. Rice husk (RH) is also one of the LSB. In this study, the optimization of dilute acid hydrolysis conditions of RH by using one-factor-at-a-time method was performed. The optimum hydrolysis conditions of RH were determined as 131.04°C, 1:10.28 (w/v) of solid:liquid ratio, 1.47% (w/v) of acid ratio and 24.05 min, which yielded as 25.52 g/L of fermentable sugar concentration. Furthermore, the chemical composition of hydrolysate was also examined and the levels of phenolics, 5-hydroxymethylfurfural (HMF), acetate, glucose, and fructose+xylose were found as 0.89, 0, 3.27, 2.10, and 13.80 g/L, respectively. Accordingly, RH can be utilized as a favorable feedstock for the production of value-added products.     

Hydrolysis of pretreated rice straw by an enzyme cocktail comprising acidic xylanase from Aspergillus sp. for bioethanol production

The most crucial enzyme involved in xylan hydrolysis is endoxylanase which cleaves the internal glycosidic bonds of xylan. The aim of this work was to study the production of extracellular xylanase by a locally isolated strain of Aspergillus sp. under solid-state fermentation (SSF) and to evaluate the potential of the enzyme in enzymatic hydrolysis of pretreated rice straw. Xylanase production reached maximum with incubation period (96 h), moisture level (80%), inoculum size (3 × 10⁶ spores/mL), pH (4.8), temperature (25 °C), carbon source (wheat bran) and nitrogen source (yeast extract). Under optimized conditions, xylanase production reached to 5059 IU/gds. Crude xylanase was used for supplementing the enzyme cocktail comprising cellulases (Zytex, India), β-glucosidase (In-house) and xylanase (In-house) for the saccharification of alkali-pretreated rice straw to get the maximum reducing sugar production. The cocktail containing the three enzymes resulted a maximum of 574.8 mg/g of total reducing sugars in comparison to 430.2 mg/g sugars by the cocktail without xylanase. These results proved that the crude xylanase preparation from Aspergillus sp. could be a potent candidate for the enzyme cocktail preparation for biomass hydrolysis in lignocellulosic bioethanol program.     

有机负荷对杂交狼尾草和牛粪混合发酵系统内微生物群落的影响

选取杂交狼尾草和牛粪为原料,研究有机负荷和原料配比对混合原料发酵系统内微生物群落组成的影响。研究结果表明,属水平上系统内优势细菌有梭菌属（Clostridum）、普雷沃菌属（Prevotella）、互养棍状菌属（Syntrophorhabdus）、假单胞菌属（Pseudomonas）和噬蛋白质菌属（Proteiniphilum）等;优势产甲烷古菌有甲烷丝菌属（Methanothrix）和甲烷蝇菌属（Methanolinea）等。当系统有机负荷低于4.5 g VS/(L∙d) （GC31）或5.0 g VS/(L∙d)（GC11）时,贡献系统内丙酸和丁酸累积的梭菌属等细菌菌属平均相对丰度为0.53%,而参与丙酸/丁酸转化细菌菌属的平均相对丰度为1.27%,有助于丙酸和丁酸转化;同时,假单胞菌属和噬蛋白质菌属等贡献系统氨氮积累的菌属丰度较低,此时甲烷丝菌属是优势产甲烷菌属,乙酸型产甲烷途径是主要的转化途径。随着有机负荷升至5.0 g VS/(L∙d) 或5.5 g VS/(L∙d) 时,贡献丙酸和丁酸累积的细菌菌属的相对丰度升高了240%,而参与丙酸/丁酸转化细菌菌属丰度降低了90%,导致系统内丙酸和丁酸浓度增加;此时与系统中氨氮浓度升高有关的假单胞菌属和噬蛋白质菌属丰度增加了210%;同时乙酸营养型产甲烷菌属丰度降低,与系统中乙酸浓度增加有关。由此可见,原料配比和有机负荷可影响系统内微生物群落组成结构,进而影响系统产气性能。     

皇竹草厌氧发酵特性及稀土元素溶出研究

为探讨稀土尾矿区修复用皇竹草的厌氧发酵特性和发酵过程稀土元素溶出情况,开展不同发酵浓度（总固体浓度分别为2%、4%、6%、8%、10%）的中温（37±1℃）批式厌氧发酵实验。结果表明,总固体（TS）浓度为4%条件下皇竹草产甲烷性能最佳,日产甲烷率和累积产甲烷率分别为25.56 mL/g和197.33 mL/g。采用修正Gompertz方程能较好地模拟不同TS下皇竹草发酵累积产甲烷率的变化。分析发酵液中稀土元素浓度变化,结果表明,皇竹草内的稀土元素在厌氧发酵过程中只发生小部分溶出,主要包括La、Ce、Nd、Sc、Y等,各处理发酵液中稀土元素浓度并未超出文献报道的抑制浓度。相关性分析结果表明,稀土元素溶出率与发酵体系的TS浓度、化学需氧量（COD）呈负相关,与pH呈正相关。研究可为稀土矿区皇竹草沼气工程应用和发酵剩余物的肥料化利用提供指导。     

玉米秸秆的渗滤式稀酸预处理

对玉米秸秆的渗滤式稀酸预处理进行了考察,通过对温度、酸浓度、渗滤速度、液固比等影响因素的实验分析,得到了优化的工艺条件:反应温度170℃,硫酸浓度0.25%,液固比为10:1,渗滤速度150mL/min,70%水解液排出后,渗滤速度降为100mL/min。水解液中木糖浓度达到22g/L,糖得率达到约80%。经过稀酸预处理后的玉米秸秆进行酶水解,纤维素转化率达到80%。     

Comparative study on the production of biohydrogen from rice mill wastewater

This study presents the production of biohydrogen from rice mill wastewater. The acid hydrolysis and enzymatic hydrolysis operating conditions were optimized, for better reducing sugar production. The effect of pH and fermentation time on biohydrogen production from acid and enzymatic hydrolyzed rice mill wastewater was investigated, using Enterobacter aerogenes and Citrobacter ferundii. The enzymatic hydrolysis produced the maximum reducing sugar (15.8 g/L) compared to acid hydrolysis (14.2 g/L). The growth data obtained for E. aerogenes and C. ferundii, fitted well with the Logistic equation. The hydrogen yields of 1.74 mol H₂/mol reducing sugar, and 1.40 mol H₂/mol reducing sugar, were obtained from the hydrolyzate obtained from enzymatic and acid hydrolysis, respectively. The maximum hydrogen yield was obtained from E. aerogenes compared to C. ferundii, and the optimum pH for better hydrogen production was found to be in the range from 6.5 to 7.0. The chemical oxygen demand (COD) reduction obtained was around 71.8% after 60 h of fermentation.     

Enzymatic saccharification of dilute acid pretreated saline crops for fermentable sugar production

Four saline crops [athel (Tamarix aphylla L), eucalyptus (Eucalyptus camaldulensis), Jose Tall Wheatgrass (Agropyron elongatum), and Creeping Wild Ryegrass (Leymus triticoides)] that are used in farms for salt uptake from soil and drainage irrigation water have the potential for fuel ethanol production because they don’t take a large number of arable lands. Dilute sulfuric acid pretreatment and enzymatic hydrolysis were conducted to select the optimum pretreatment conditions and the best saline crop for further enzymatic hydrolysis research. The optimum dilute acid pretreatment conditions included T = 165 °C, t = 8 min, and sulfuric acid concentration = 1.4% (w/w). Creeping Wild Ryegrass was decided to be the best saline crop. Solid loading, cellulase and β-glucosidase concentrations had significant effects on the enzymatic hydrolysis of dilute acid pretreated Creeping Wild Ryegrass. Glucose concentration increased by 36 mg/mL and enzymatic digestibility decreased by 20% when the solid loading increased from 4 to 12%. With 8% solid loading, enzymatic digestibility increased by over 30% with the increase of cellulase concentration from 5 to 15 FPU/g-cellulose. Under given cellulase concentration of 15 FPU/g-cellulose, 60% increase of enzymatic digestibility of pretreated Creeping Wild Ryegrass was obtained with the increase of β-glucosidase concentration up to 15 CBU/g-cellulose. With a high solid loading of 10%, fed-batch operation generated 12% and 18% higher enzymatic digestibility and glucose concentration, respectively, than batch process.