固定化纤维素酶对预处理纤维素原料水解效果的影响

以壳聚糖为载体用交联法制备固定化纤维素酶,考察固定化纤维素酶对蒸爆、球磨、超声波、喷淋、高温预处理玉米秸秆纤维素原料的酶解效果.结果表明:物料经蒸爆预处理后酶水解效率最高可以达到95%,球磨预处理水解效率次之,达到60%.用电镜和FT-IR对处理前后秸秆结构进行表征分析,证明预处理对物料的物理结构及化学组成有一定的影响.蒸爆法和球磨法可以使物料致密的天然结构彻底破坏,从而增加物料的比表面积;蒸爆预处理可以使纤维素内部氢键和官能团改变,使物料更易于酶解.     

沙柳原料高浓度底物酶解发酵乙醇工艺

为了提高沙柳生物转化过程的经济可行性,考察了沙柳原料经过蒸爆、超微粉碎+稀酸、超微粉碎+稀碱预处理后高浓度底物补料酶解的效果,并对其高浓度水解糖液进行了乙醇发酵。结果表明:蒸爆处理法水解效果最好,通过补料酶解,底物质量分数可以达到30%,酶解液中总糖质量浓度达到132 g/L,葡萄糖质量浓度105 g/L;超微粉碎+稀酸预处理原料底物质量分数可以达到22%,酶解液中总糖质量浓度达到123 g/L,葡萄糖质量浓度73 g/L;超微粉碎+稀碱预处理原料底物质量分数可以达到22%,酶解液中总糖质量浓度133 g/L,葡萄糖质量浓度77 g/L。3种预处理使沙柳原料的酶解糖液都可以较好地被酿酒酵母利用发酵产乙醇,蒸爆处理原料的酶解糖液乙醇发酵效果最好,乙醇质量浓度达到47 g/L。     

预处理沙柳酶解液脱毒及其丁醇发酵

为了提高沙柳原料的丁醇发酵效果,考察沙柳原料经过蒸爆、超微粉碎+稀酸和超微粉碎+稀碱预处理后补料酶解的效果,优化了沙柳酶解液活性炭脱毒工艺参数,并对经过脱毒处理的酶解液进行了丁醇发酵研究,结果表明:预处理沙柳原料酶解底物质量浓度为200 g/m L时,3种预处理方法中蒸爆处理法水解效果最好,每克底物的滤纸酶酶加量15 U,酶解96 h后,酶解液总糖质量浓度达到57 g/L。活性炭脱毒处理的最优条件:p H 4.8,碳加量4%(质量分数)、温度70℃、1 h,该条件下的沙柳水解液脱色率达到97.4%、糖损失率3.1%。3种预处理沙柳原料的酶解液经活性炭脱毒后都可以被丁醇梭菌正常利用发酵产丁醇,发酵液总溶剂(ABE)质量浓度约为14 g/L。     

稀酸和稀碱预处理对四种不同生物质资源制备还原糖的影响

本论文探讨了不同浓度的稀H_2SO_4和稀NaOH预处理对大豆秸秆、水稻秸秆、象草和狼尾草四种不同生物质酶解制备还原糖的影响。结果表明,大豆秸秆、水稻秸秆、象草和狼尾草具有较高的纤维素和半纤维素含量,是制备还原糖的理想原料。与稀H_2SO_4预处理相比,经稀NaOH预处理后的样品表现出较好的酶解性能。通过使用4%的NaOH对大豆秸秆和狼尾草进行预处理,还原糖产量分别为145.8 mg/mL和319.2 mg/mL。此外,以1%NaOH预处理后的水稻秸秆和象草为原料,可以分别获得385.2 mg/mL和231.6 mg/mL还原糖产量。     

重组大肠杆菌利用废纸水解液发酵产L-乳酸

前期通过基因工程手段,构建了一株大肠杆菌工程菌E.coli WL204,该菌株可以有效利用木糖为底物发酵产L-乳酸。以废纸为发酵原料,研究该菌株利用木质纤维素发酵产乳酸的特性。原料以稀硫酸预处理后,经纤维素酶酶解,得到的水解液用Ca(OH)2脱毒后,接种E.coli WL204,在7L发酵罐中发酵72h,每100g废纸可以产生31g乳酸,糖酸转化率为79%。结果表明,E.coli WL204可以木质纤维素原料为底物发酵生产L-乳酸,具有一定的工业化开发潜力。     

斑茅酶解转化可发酵单糖的液氨预处理及参数优化

斑茅(Saccharum arundinaceum Retz.)的生物产量高,对土壤条件要求低,可作为纤维素乙醇生产的原料作物在我国南方地区广泛种植.实验以斑茅为原料,采用液氨预处理法克服其水解顽抗性,并添加纤维素酶进行酶解,运用高效液相色谱(HPLC)测定了酶解液中的单糖含量.实验结果表明在纤维素酶添加量为15FPU/(g当量葡聚糖)、预处理原料含水率为80％、预处理温度为130℃、预处理驻留时间为10 min、液氨与生物质的质量比例为2∶1时,葡聚糖和木聚糖的总转化率分别为69.34％和82.60％,相比于未作预处理的原料分别提高了573％和1 056％,单糖产量提高8倍.实验结果表明液氨预处理对斑茅是一种有效的预处理方式,并优于稀酸或湿爆法预处理,与酸预处理和氨爆法(AFEX)处理效果接近.     

稻草生物质的预处理及其发酵产酶与酶解效果研究

采用稀酸、稀碱、高温稀碱、亚硫酸盐法(SPORL法)和稀酸-亚硫酸盐法(稀酸SPORL法)对粉碎稻草秸秆预处理,考察不同预处理方法对稻草基质多菌发酵产纤维素酶的影响,分析预处理前后稻草基质主要成分的变化,酶水解液中糖组分的含量。结果表明:稀酸SPORL法处理的稻草粉在固态发酵产酶和酶解糖化都具有较好的效果,所得羧甲基纤维素酶(CMCase酶)和β-葡萄糖苷酶(β-G)比酶活分别达到21 511.22和51 508.41 U/g,同时酶水解率达到84.99%。除SPORL法外,其他预处理方式所得酶活均出现了不同程度的下降。稀酸预处理对稻草基质中的半纤维素去除效果较好,含量由20.77%下降到7.34%;稀碱高温处理对木质素脱除效果较好,Klason木质素含量由12.47%下降到7.58%。通过酶解糖化实验发现,未处理稻草粉酶水解率仅为17.82%,稀碱高温法效果最好,稻酶水解率达到91.66%。稀酸和稀酸SPORL法处理后,稻草粉基质的酶解糖化液中,戊聚糖占总糖相对含量较低,分别为7.38%和6.92%。     

乙醇预处理麦秆的酶解性能研究

采用H2SO4催化和自催化乙醇法对麦秆进行预处理,比较预处理后麦秆的主要化学组成、纤维素酶解性能和半同步糖化发酵生产乙醇特性,并进行物料衡算。结果表明:H2SO4催化和自催化乙醇预处理过程中纤维素固体回收率大于90%。添加非离子表面活性剂吐温20和吐温80没有显著提高H2SO4催化乙醇预处理后纤维素的酶解葡萄糖得率及半同步糖化发酵过程中乙醇的产量,而对自催化乙醇处理后麦秆的酶解和半同步糖化发酵过程有一定程度的促进作用,相应的酶解葡聚糖转化率由72.7%提高到85.0%,而半同步糖化发酵过程中乙醇质量浓度提高了11.4%。物料衡算结果表明:酸催化和自催化乙醇预处理后葡聚糖回收率分别为91.0%和95.4%;半同步糖化发酵生产乙醇的得率分别为10.4和11.6 g(按100 g原料计)。     

稻壳作为缓释碳源及载体的改性研究

稻壳可作为废水处理的外加碳源, 通过适当改性处理可提高其应用性能。为探索稻壳的改性条件, 以不同浓度的NaOH、Ca(OH)2、NaClO为改性试剂对稻壳进行改性处理, 并研究了改性后稻壳的表面结构、芽孢杆菌吸附量、静态释碳量、可生化性以及成分含量变化。结果表明: 6% NaOH、0.9% Ca(OH)2和3% NaClO处理对稻壳表面糙化、芽孢杆菌吸附性和静态释碳能力有良好的提升效果。在此三组中, 6% NaOH处理后稻壳可生化效果最佳, CD600增长率为其他处理组的4倍; 纤维素含量增加了16.03%, 灰分含量显著降低, 仅剩4.9%; 且结构改性效果最为明显, 适用于稻壳改性优化。     

纤维素酶水解稻草生产单细胞蛋白（SCP）：1.纤维素酶水解稻草的研究

对不同预处理方法与底物得糖率,纤维素转化率的关系进行了研究,以1％NaOH在100℃煮沸1h效果最好。研究了稻草酶解的适宜条件,在pH5、50℃、底物浓度7％、酶解48h的条件下,底物得糖率达53.8％,含糖量5.0％,全纤维素转化率80.2％。     

Kinetics of the enzymatic hydrolysis of cellulose: 2. A mathematical model for the process in a plug-flow column reactor

The kinetics of enzymatic cellulose hydrolysis in a plug-flow column reactor catalysed by cellulases [see 1,4-(1,3;1,4)-β-d-glucan 4-glucanohydrolase, EC 3.2.1.4] from Trichoderma longibrachiatum adsorbed on cellulose surface have been studied. The maximum substrate conversion achieved was 90–94%. The possibility of enzyme recovery for a reactor of this type is discussed. A mathematical model for enzymatic cellulose hydrolysis in a plug-flow column reactor has been developed. The model allows for the component composition of the cellulase complex, adsorption of cellulases on the substrate surface, inhibition by reaction products, changes in cellulose reactivity and the inactivation of enzymes in the course of hydrolysis. The model affords a reliable prediction of the kinetics of d-glucose and cellobiose formation from cellulose in a column reactor as well as the degree of substrate conversion and reactor productivity with various amounts of adsorbed enzymes and at various flow rates.     

The effects of four different pretreatments on enzymatic hydrolysis of sweet sorghum bagasse

Four pretreatment processes including ionic liquids, steam explosion, lime, and dilute acid were used for enzymatic hydrolysis of sweet sorghum bagasse. Compared with the other three pretreatment approaches, steam-explosion pretreatment showed the greatest improvement on enzymatic hydrolysis of the bagasse. The maximum conversion of cellulose and the concentration of glucose obtained from enzymatic hydrolysis of steam explosion bagasse reached 70% and 25 g/L, respectively, which were both 2.5 times higher than those of the control (27% and 11 g/L). The results based on the analysis of SEM photos, FTIR, XRD and NMR detection suggested that both the reduction of crystallite size of cellulose and cellulose degradation from the Iα and Iβ to the Fibril surface cellulose and amorphous cellulose were critical for enzymatic hydrolysis. These pretreatments disrupted the crystal structure of cellulose and increased the available surface area, which made the cellulose better accessible for enzymatic hydrolysis.     

Enhanced rates of enzymatic saccharification and catalytic synthesis of biofuel substrates in gelatinized cellulose generated by trifluoroacetic acid

Background

The crystallinity of cellulose is a principal factor limiting the efficient hydrolysis of biomass to fermentable sugars or direct catalytic conversion to biofuel components. We evaluated the impact of TFA-induced gelatinization of crystalline cellulose on enhancement of enzymatic digestion and catalytic conversion to biofuel substrates.

Results

Low-temperature swelling of cotton linter cellulose in TFA at subzero temperatures followed by gentle heating to 55 °C dissolves the microfibril structure and forms composites of crystalline and amorphous gels upon addition of ethanol. The extent of gelatinization of crystalline cellulose was determined by reduction of birefringence in darkfield microscopy, loss of X-ray diffractability, and loss of resistance to acid hydrolysis. Upon freeze-drying, an additional degree of crystallinity returned as mostly cellulose II. Both enzymatic digestion with a commercial cellulase cocktail and maleic acid/AlCl₃-catalyzed conversion to 5-hydroxymethylfurfural and levulinic acid were markedly enhanced with the low-temperature swollen cellulose. Only small improvements in rates and extent of hydrolysis and catalytic conversion were achieved upon heating to fully dissolve cellulose.

Conclusions

Low-temperature swelling of cellulose in TFA substantially reduces recalcitrance of crystalline cellulose to both enzymatic digestion and catalytic conversion. In a closed system to prevent loss of fluorohydrocarbons, the relative ease of recovery and regeneration of TFA by distillation makes it a potentially useful agent in large-scale deconstruction of biomass, not only for enzymatic depolymerization but also for enhancing rates of catalytic conversion to biofuel components and useful bio-products.

     

Reduced inhibition of enzymatic hydrolysis of steam-pretreated softwood

Softwood constitutes the main source of lignocellulosic material in Sweden which can be used for ethanol production from renewable resources. To make the biomass-to-ethanol process more economically feasible, it is preferable to include the sugar-rich prehydrolysate, i.e. the liquid obtained after the pretreatment step, in the enzymatic hydrolysis of the solid fraction. This study shows that the prehydrolysate inhibits cellulose conversion in the enzymatic hydrolysis step. When the prehydrolysate was included in the enzymatic hydrolysis, the cellulose conversion was reduced by up to 36%. However, this inhibition can be overcome by fermentation of the prehydrolysate prior to enzymatic hydrolysis.     

Bioconversion of wheat straw to ethanol: Chemical modification,enzymatic hydrolysis,and fermentation

Native wheat straw (WS) was pretreated with various concentrations of H₂SO₄ and NaOH followed by secondary treatments with ethylene diamine (EDA) and NH₄OH prior to enzymatic saccharification. Conversion of the cellulosic component to sugar varied with the chemical modification steps. Treatment solely with alkali yield 51–75% conversion, depending on temperature. Acid treatment at elevated tempeatures showed a substantial decrease in the hemicellulose component, whereas EDA-treated WS (acid pretreated) showed a 69–75% decrease in the lignin component. Acid-pretreated EDA-treated straw yielded a 98% conversion rate, followed by 83% for alkali–NH₄OH treated straws. In other experiments, WS was pretreated with varying concentration of H₂SO₄ or NaOh followed by NH₄OH treatment prior to enzymatic hydrolysis. Pretreatment of straw with 2% NaOH for 4 h coupled to enzymatic hydrolysis yield a 76% conversion of the cellulosic component. Acid–base combination pretreatment yielded only 43% conversions. A reactor column was subsequently used to measure modification–saccharification–fermentation for wheat straw conversion on a larger scale. Thirty percent conversions of wheat straw cellulosics to sugar were observed with subsequent fermentation to alcohol. The crude cellulase preparation yielded considerable quantities of xylose in addition to the glucose. Saccharified materials were fermented directly with actively proliferating proliferating yeast cells without concentration of the sugars.     

Modelling ethanol production from cellulose: separate hydrolysis and fermentation versus simultaneous saccharification and fermentation

In ethanol production from cellulose, enzymatic hydrolysis, and fermentative conversion may be performed sequentially (separate hydrolysis and fermentation, SHF) or in a single reaction vessel (simultaneous saccharification and fermentation, SSF). Opting for either is essentially a trade-off between optimal temperatures and inhibitory glucose concentrations on the one hand (SHF) vs. sub-optimal temperatures and ethanol-inhibited cellulolysis on the other (SSF). Although the impact of ethanol on cellobiose hydrolysis was found to be negligible, formation of glucose and cellobiose from cellulose were found to be significantly inhibited by ethanol. A previous model for the kinetics of enzymatic cellulose hydrolysis was, therefore, extended with enzyme inhibition by ethanol, thus allowing a rational evaluation of SSF and SHF. The model predicted SSF processing to be superior. The superiority of SSF over SHF (separate hydrolysis and fermentation) was confirmed experimentally, both with respect to ethanol yield on glucose (0.41 g g^?1 for SSF vs. 0.35 g g^?1 for SHF) and ethanol production rate, being 30% higher for an SSF type process. High conversion rates were found to be difficult to achieve since at a conversion rate of 52% in a SSF process the reaction rate dropped to 5% of its initial value. The model, extended with the impact of ethanol on the cellulase complex proved to predict reaction progress accurately.     

Reactor design for minimizing product inhibition during enzymatic lignocellulose hydrolysis: II. Quantification of inhibition and suitability of membrane reactors

Product inhibition of cellulolytic enzymes affects the efficiency of the biocatalytic conversion of lignocellulosic biomass to ethanol and other valuable products. New strategies that focus on reactor designs encompassing product removal, notably glucose removal, during enzymatic cellulose conversion are required for alleviation of glucose product inhibition. Supported by numerous calculations this review assesses the quantitative aspects of glucose product inhibition on enzyme-catalyzed cellulose degradation rates. The significance of glucose product inhibition on dimensioning of different ideal reactor types, i.e. batch, continuous stirred, and plug-flow, is illustrated quantitatively by modeling different extents of cellulose conversion at different reaction conditions. The main operational challenges of membrane reactors for lignocellulose conversion are highlighted. Key membrane reactor features, including system set-up, dilution rate, glucose output profile, and the problem of cellobiose are examined to illustrate the quantitative significance of the glucose product inhibition and the total glucose concentration on the cellulolytic conversion rate. Comprehensive overviews of the available literature data for glucose removal by membranes and for cellulose enzyme stability in membrane reactors are given. The treatise clearly shows that membrane reactors allowing continuous, complete, glucose removal during enzymatic cellulose hydrolysis, can provide for both higher cellulose hydrolysis rates and higher enzyme usage efficiency (kg_product/kg_enzyme). Current membrane reactor designs are however not feasible for large scale operations. The report emphasizes that the industrial realization of cellulosic ethanol requires more focus on the operational feasibility within the different hydrolysis reactor designs, notably for membrane reactors, to achieve efficient enzyme-catalyzed cellulose degradation.     

氨基磺酸应用于甘蔗渣预处理的研究

本研究尝试将氨基磺酸应用于甘蔗渣预处理,探究其作为酸预处理试剂对甘蔗渣成分和酶解的影响。氨基磺酸预处理最优条件为浓度3%,温度121℃,预处理1 h。在该条件下,甘蔗渣的固体回收率为64.45%,半纤维素和木质素去除率分别为70.81%和25.10%,纤维素损失率仅7.56%。与硫酸、盐酸预处理相比,氨基磺酸的半纤维素和木质素去除率不如硫酸、盐酸预处理,但固体回收率更高,纤维素损失率低,能保留更多纤维素有效成分。进一步酶解显示,氨基磺酸预处理的纤维素转化率高于硫酸、盐酸预处理。氨基磺酸作为一种新的酸预处理试剂,在木质纤维素降解上有良好应用前景。     

磁性Fe3O4微粒对葵花籽壳酶水解的影响研究

将磁化后的Fe3O4微粒添加于葵花籽壳酶水解过程中, 分析在不同的Fe3O4添加量和不同的添加方法下, 葵花籽壳酶水解过程中纤维素酶酶活﹑纤维素转化率及还原糖浓度的变化特征, 研究磁性Fe3O4微粒对纤维素酶水解葵花籽壳的影响。并通过考察酶水解反应前后水解液的表面张力值和pH值的变化, 探讨和分析磁性Fe3O4微粒作用下纤维素酶的磁效应机制。结果表明, 磁性Fe3O4添加量为0.5 g/L~2.0 g/L时, 对纤维素酶酶活的提高﹑还原糖浓度的增加和纤维素的转化在48 h后表现出较明显的促进作用。磁性Fe     

Changes in the enzymatic hydrolysis rate of Avicel cellulose with conversion

The slow down in enzymatic hydrolysis of cellulose with conversion has often been attributed to declining reactivity of the substrate as the more easily reacted material is thought to be consumed preferentially. To better understand the cause of this phenomenon, the enzymatic reaction of the nearly pure cellulose in Avicel was interrupted over the course of nearly complete hydrolysis. Then, the solids were treated with proteinase to degrade the cellulase enzymes remaining on the solid surface, followed by proteinase inhibitors to inactive the proteinase and successive washing with water, 1.0 M NaCl solution, and water. Next, fresh cellulase and buffer were added to the solids to restart hydrolysis. The rate of cellulose hydrolysis, expressed as a percent of substrate remaining at that time, was approximately constant over a wide range of conversions for restart experiments but declined continually with conversion for uninterrupted hydrolysis. Furthermore, the cellulose hydrolysis rate per adsorbed enzyme was approximately constant for the restart procedure but declined with conversion when enzymes were left to react. Thus, the drop off in reaction rate for uninterrupted cellulose digestion by enzymes could not be attributed to changes in substrate reactivity, suggesting that other effects such as enzymes getting "stuck" or otherwise slowing down may be responsible.