基于响应面法对秸秆纤维素乙醇生产工艺参数的优化

文章在对里氏木霉T12菌株产纤维素酶的培养条件进行单因素优化的基础上,以滤纸酶活力(FPA)为响应值,通过Plackett-Burman设计法筛选出对产酶影响最显著的3个因素,依次为麦麸>温度>氯化钙。响应面优化结果为当麦麸、温度、氯化钙分别为6.27 g/L,31℃,0.709 g/L时,纤维素酶理论最大FPA酶活为62281.3 U/m L。在优化后的培养条件下纤维素酶粗酶液的实际FPA酶活为60 126.5±16.0 U/m L。将纤维素酶粗酶液以10%添加量加入秸秆一步转化乙醇的5 L发酵罐中,经过144 h的发酵,乙醇产量(v/v)可达到7.05%±0.18%。     

混合纤维素酶对杨木酶解的研究

以风干的杨木为原料,通过蒸汽爆破处理后,用混合纤维素酶降解。采用响应面分析法,对酶解温度、pH、β-葡萄糖苷酶/滤纸酶的比进行研究。研究表明,酶解杨木的最佳工艺参数:酶解温度为50.04℃、pH值为4.99、β-葡萄糖苷酶/滤纸酶的比值为1.39。在最优的条件下,底物质量分数为5%时,使用里氏木霉RutC-30和爪哇正青霉ZN-205制备的混合纤维素酶(15FPU/g底物,β-葡萄糖苷酶/滤纸酶比值为1.39),酶解48h,可产生还原糖为25.54g/l,糖的转化率为78.184%。     

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。     

4种工业微生物产纤维素酶酶学特性的比较研究

文章对康宁木霉、绿色木霉、黑曲霉和里氏木霉表达纤维素酶的酶学特性进行了初步研究,结果表明,纤维素酶在pH低于5.0以及温度高于60℃条件下的稳定性较差;Fe2+和Mn2+的存在能够显著提高酶活性;Pb2+和Zn2+的存在严重抑制酶活性。黑曲霉表达纤维素酶活性高,是发酵产纤维素酶的较理想菌株。     

一株高活力纤维素酶产生菌——绿色木霉T2产酶研究

以绿色木霉(Trychoderma viride)T2为实验材料,麸皮和稻草粉为培养基,采用固体发酵法,对其产纤维素酶情况进行了研究。在优化条件下,其羧甲基纤维素酶(CMCase)活力为94.52IU,滤纸酶活力(FPA)为57.4IU,β-葡萄糖苷酶(β-glucosidase)活力为28.93IU。     

酸法-酶法处理麦秆木质纤维素的工艺研究

试验研究了稀硫酸常压预处理对纤维素酶水解麦秆的影响.试验结果表明:麦秆在温度为80℃、稀硫酸质量分数为4%、固液比(质量体积比,g/ml,下同)为1:25的条件下水解4h,再在温度为50℃、pH值为5.2、酶量/干物质为25 FPU/g、MgSO4/干物质为0.1 mg/g条件下水解12 h,葡萄糖得率为34.5%.经酸法-酶法处理的麦秆比未经酸处理直接酶解的麦秆的葡萄糖得率提高50%.     

酶法水解低聚木糖生产废渣工艺的研究

以低聚木糖生产废渣为酶法水解工艺的研究对象,考察了底物浓度和酶用量对酶解效率和葡萄糖产率的影响。研究结果表明,当纤维素酶Celluclast 1.5L用量在10FPIU/g,底物质量浓度50g/L条件下水解48h,酶解得率达到74.9%,葡萄糖得率为35.1%。单酶水解反应体系寡糖浓度水平较高,添加纤维二糖酶可有效解除可溶性寡糖累积引起的反馈抑制作用。当纤维二糖酶Novozyme 188添加量为30CBIU/g时,相同条件下水解48h酶解得率上升到98.1%,而葡萄糖得率达到78.2%。     

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。     

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。     

高效纤维素降解菌的筛选及其系统发育分析

从刚果红平板上筛选到S41、Z3b、Z28和Z41共4株纤维素酶高产菌株,通过形态观察,生理生化特性分析以及系统进化分析等,鉴定出S41和Z41属于木霉属(Trichoderma sp.),Z3b属于曲霉属(Aspergillus sp.),Z28属于肉座菌属(Hypocrea sp.);4株菌的CMC酶活,FPA酶活和β-葡萄糖苷酶活几乎均高于绿色木霉(Trichderma viride)AS3.2774,葡萄糖对4株菌的β-葡萄糖苷酶均有一定的反馈抑制作用,对Z28的p葡萄糖苷酶抑制作用较低.     

The preparation and application of crude cellulase for cellulose-hydrogen production by anaerobic fermentation

Strategies were adopted to cost-efficiently produce cellulose-hydrogen by anaerobic fermentation in this paper. First, cellulase used for hydrolyzing cellulose was prepared by solid-state fermentation (SSF) on cheap biomass from Trichoderma viride. Several cultural conditions for cellulase production on cheap biomass such as moisture content, inoculum size and culture time were studied. And the components of solid-state medium were optimized using statistical methods to further improve cellulase capability. Second, the crude cellulase was applied to cellulose-hydrogen process directly. The maximal hydrogen yield of 122 ml/g-TVS was obtained at the substrate concentration of 20 g/L and cultured time of 53 h. The value was about 45-fold than that of raw corn stalk wastes. The hydrogen content in the biogas was 44–57%(v/v) and there was no significant methane gas observed.     

Cellulase production using biomass feed stock and its application in lignocellulose saccharification for bio-ethanol production

A major constraint in the enzymatic saccharification of biomass for ethanol production is the cost of cellulase enzymes. Production cost of cellulases may be brought down by multifaceted approaches which include the use of cheap lignocellulosic substrates for fermentation production of the enzyme, and the use of cost efficient fermentation strategies like solid state fermentation (SSF). In the present study, cellulolytic enzymes for biomass hydrolysis were produced using solid state fermentation on wheat bran as substrate. Crude cellulase and a relatively glucose tolerant BGL were produced using fungi Trichoderma reesei RUT C30 and Aspergillus niger MTCC 7956, respectively. Saccharification of three different feed stock, i.e. sugar cane bagasse, rice straw and water hyacinth biomass was studied using the enzymes. Saccharification was performed with 50 FPU of cellulase and 10 U of β-glucosidase per gram of pretreated biomass. Highest yield of reducing sugars (26.3 g/L) was obtained from rice straw followed by sugar cane bagasse (17.79 g/L). The enzymatic hydrolysate of rice straw was used as substrate for ethanol production by Saccharomyces cerevisiae. The yield of ethanol was 0.093 g per gram of pretreated rice straw.     

Cellulolytic enzymes produced by a newly isolated soil fungus Penicillium sp. TG2 with potential for use in cellulosic ethanol production

A newly isolated soil fungus, Penicillium sp. TG2, had cellulase activities that were comparable to those of Trichoderma reesei RUT-C30, a common commercial strain used for cellulase production. The maximal and specific activities were 1.27 U/mL and 2.28 U/mg for endoglucanase, 0.31 U/mL and 0.56 U/mg for exoglucanase, 0.54 U/mL and 1.03 U/mg for β-glucosidase, and 0.45 U/mL and 0.81 U/mg for filter paper cellulase (FPase), respectively. Optimal FPase activity was at pH 5.0 and 50 °C. We used a simultaneous saccharification and fermentation (SSF) process, which employed the yeast Kluyveromyces marxianus and Penicillium sp. TG2 cellulolytic enzymes, to produce ethanol from empty palm fruit bunches (EFBs), a waste product from the palm oil industry. The present findings indicate that Penicillium sp. TG2 has great potential as an alternative source of enzymes for saccharification of lignocellulosic biomass.     

Biochemical conversion of sugarcane bagasse into bioproducts

The ground sugarcane bagasse conversions were examined through chemical treatment methods employing soaking in aqueous ammonia (SAA), and ethyl-hydro-oxides (EHOs). To characterize a chemical treatment method, both generated solvent based extract and pulp were examined. The generated pulps were evaluated through chemical composition and enzymatic saccharification. The enzyme mixtures were investigated including Trichoderma reesei Rut C-30 originated cellulase, T. reesei Rut C-30 originated cellulase with external added β-glucosidase, Accellerase^® 1500, and Cellic^® CTec2. The physiochemical effects of chemical treatments on the structural-chemical properties of treated-bagasse were also analyzed at high substrate enzymatic saccharification. The substrate loadings (using both SAA-treated and EHOs-treated bagasse) of 125, 150, 175, 200, and 225 g L⁻¹ were examined during enzymatic saccharification process. The generated phenolic compounds were characterized based on density, antioxidant activity, and anticancer activity. All findings are discussed in relation to developing a self-sustainable integrated biorefinery.     

Aerobic and anaerobic sequential culture fermentation (AASF) to produce bio-hydrogen from steam-exploded cornstalk

A novel aerobic and anaerobic sequential culture fermentation (AASF) method was designed to improve the conversion efficiency of steam-exploded cornstalk during bio-hydrogen production. The enzyme activities of cellulase and β-glucosidase produced by Trichoderma viride ACCC 30169 were 76.79 FPU g⁻¹ dry weight and 45.23 IU g⁻¹ dry weight after 6-days steam-exploded cornstalk fermentation, respectively. The aerobic fermentation residue was used as the substrate for bio-hydrogen production by Clostridium butyricum AS1.209 anaerobic fermentation. The optimum solid-to-liquid ratio of the anaerobic fermentation substrate was 1:5. The maximum bio-hydrogen yield was attained on the medium with addition of 0.1 g g⁻¹ substrate urea after 2 days of aerobic fermentation. Compared with simultaneous saccharification and fermentation (SSF), AASF for bio-hydrogen production could shorten the fermentation period by at least 66% and the hydrogen yield reached 83% of the total volume after 24 h of anaerobic fermentation. AASF from steam-exploded cornstalk was an effective way for bio-hydrogen production without additional commercial cellulase.     

Optimization of concomitant production of cellulase and xylanase from Rhizopus oryzae SN5 through EVOP-factorial design technique and application in Sorghum Stover based bioethanol production

Current study deals with the production of cellulases and xylanases from the Rhizopus oryzae SN5 isolated from composed soil of Himalayan pine forest, in order to meet the challenges of lignocellulosic biomass based biorefineries. Culture parameters for concomitant production of cellulase and xylanase were optimized through EVOP-factorial design technique under solid state fermentation. And maximum yield of cellulase and xylanase were obtained 437.54 U/gds and 273.83 U/gds, respectively at 30 °C and pH 6.0 after 5 days of incubation. On applying these enzymes for the saccharification of the dilute acid pretreated Sorghum Stover (SS), 0.407 g/g sugar was yielded. This hydrolysate on fermentation, yielded 0.411 g/g ehanol with Saccharomyces cerevisiae (NCIM 3288), which could be considered a good conversion. Therefore, Rhizopus oryzae SN5 was found as potent strain for the production of the cocktail of lignocellulosic biomasss hydrolytic enzymes and would be promising tool in the area of lignocellulose based bio-refineries.     

Enzymatic saccharification of pretreated rice straw by cellulase produced from Bacillus carboniphilus CAS 3 utilizing lignocellulosic wastes through statistical optimization

A marine bacterium, Bacillus carboniphilus CAS 3 was subjected to optimization for cellulase production utilizing cellulosic waste through response surface methodology. Plackett – Burman and Central composite design was employed and the optimal medium constituents for maximum cellulase production (4040.45 U/mL) were determined as rice bran, yeast extract, MgSO₄·7H₂O and KH₂PO₄ at 6.27, 2.52, 0.57 and 0.39 g/L, respectively. The cellulase produced was purified to the specific activity of 434.94 U/mg and 11.46% of recovery with the molecular weight of 56 kDa. The optimum temperature, pH and NaCl for enzyme activity was determined as 50 °C, 9 and 30% and more than 70% of its original activity was retained even at 80 °C, 12 and 35% respectively. Further, enzymatic saccharification of pretreated rice straw yielded about 15.56 g/L of reducing sugar at 96 h, suggesting that the purified cellulase could be useful for production of reducing sugars from cellulosic biomass into ethanol.     

底物质量浓度对玉米秸秆同步糖化生物制氢的影响

利用HAU-M1光合细菌对玉米秸秆同步糖化生物制氢工艺进行实验研究,提出了同步糖化生物制氢工艺中玉米秸秆底物质量浓度与pH值、还原糖质量浓度、氢气体积分数和累积产氢量等因素之间的相关关系,探讨了底物质量浓度对玉米秸秆同步糖化生物制氢工艺的影响规律。实验结果表明:当玉米秸秆底物质量浓度为25g/L时,玉米秸秆同步糖化生物制氢工艺的累积产氢量达到最高,为186mL;当玉米秸秆底物质量浓度为15g/L时,玉米秸秆同步糖化生物制氢工艺的氢气体积分数达到最高,为48%;玉米秸秆同步糖化制氢工艺的产氢高峰期为12~48h,48h后逐渐停止产氢,可为进一步优化和完善以生物质为基质的同步糖化生物制氢工艺理论与技术提供科学参考。     

Conversion of paper sludge to ethanol by separate hydrolysis and fermentation (SHF) using Saccharomyces cerevisiae

The conversion of ethanol from paper sludge using the separate hydrolysis and fermentation (SHF) process with cellulase and Saccharomyces cerevisiae GIM-2 were investigated in this paper. Optimization strategy based on statistical experimental designs was employed to enhance degree of saccharification by enzymatic hydrolysis of paper sludge. Based on the Plackett-Burman design, hydrolysis time, substrate concentration and cellulase dosage were selected as the most significant variable on the degree of saccharification. Subsequently, the optimum combination of the selected factors was investigated by a Box-Behnken approach. A mathematical model was developed to show the effects of each factor and their combinatorial interactions on the degree of saccharification. The optimal conditions were hydrolysis time 82.7 h, substrate concentration 40.8 g L⁻¹ and cellulase dosage 18.1 FPU g⁻¹ substrate, and a degree of saccharification of 82.1% can be achieved. When hydrolysate was further fermented with S. cerevisiae GIM-2, the conversion rate of sugar to ethanol was 34.2% and the ethanol yield was 190 g kg⁻¹ of dry paper sludge, corresponding to an overall conversion yield of 56.3% of the available carbohydrates on the initial substrate. The results derived from this study indicate that the response surface methodology is a useful tool for optimizing the hydrolysis conditions to converse paper sludge to ethanol.     

Biomass to bio-ethanol: The evaluation of hybrid Pennisetum used as raw material for bio-ethanol production compared with corn stalk by steam explosion joint use of mild chemicals

The second generation biomass to bio-ethanol production is of growing interest. Energy crop were becoming important for second generation biomass to bio-ethanol production for their growth advantages. Hybrid Pennisetum as a new hybrid energy crop was selected as a model to compare with corn stalk. As pre-treatment methods, steam explosion and its combined action with dilute sulfuric acid, bisulfite, and mixed dilute acid and bisulfite were selected. The enzymatic hydrolysis demonstrated that the cellulose conversion is a strong function of the pre-treatment method applied, with corn stalk providing slightly better results. With dilute acid steam explosion (DA-SE), conversions were 67.6% and 54.5% for corn stalk and pennisetum, respectively. This can be attributed to the higher Cr. I of pennisetum (65.03%) than of corn stalk (54.05%). The cell lumen of pretreated pennisetum was smaller than for corn stalk as shown in SEM photos, meaning there was a substantially higher enzyme accessible surface and porosity in pennisetum, thus responsible for the higher cellulase adsorption of pretreated pennisetum. DA-SE was the most effective pre-treatment method, but the inhibitors' concentration was higher than in other methods. Combined dilute acid and bisulfite can moderately remove hemicelluloses and lignin. Cr. I values, lignin content, accessible surface and porosity were supplied the energy crop evaluation standards for bio-ethanol production.