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.     

乙醇预处理麦秆的酶解性能研究简

采用H2 SO4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：H2 SO4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H2 SO4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）。     

乙酸分级预处理甘蔗渣对纤维素酶解性能的影响

为提高甘蔗渣的纤维素酶解性能,采用乙酸脱木素结合碱脱乙酰基的预处理工艺 (Acetoline工艺) 对甘蔗渣进行预处理,考察了乙酸脱木素过程中若干因素对预处理结果的影响,并对预处理后甘蔗渣的纤维素酶解性能进行了研究。结果表明,经过Acetoline预处理后甘蔗渣在7.5％固体含量、15 FPU+10 CBU/g固体的纤维素酶和β-葡萄糖苷酶用量下酶解48 h,酶解聚糖转化率接近80％。与稀酸预处理相比,Acetoline预处理可以得到更高的酶解聚糖转化率。实验结果表明Acetoline工艺是一种可有效提高甘蔗渣纤维素酶解性能的预处理方法。     

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%。     

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。     

中和剂对稀酸蒸爆玉米秸秆酶解效果的影响

以稀酸蒸爆的玉米秸秆为研究对象,考察直接水洗、Ca(OH)2、NaOH、氨水中和物料至pH 5,在固液比1∶10、酶添加量为每克纤维素14 U(滤纸酶活)的酶解条件下对纤维素转化率的影响。结果表明:水洗、Ca(OH)2、NaOH、氨水中和物料酶解72 h后,纤维素转化率分别为91.7%、80.7%、83.1%及81.7%。同时对影响纤维素酶解效率的各种因素进行了探讨。从综合成本及后续发酵过程考虑,用氨水中和稀酸蒸爆物料更适合于工业化生产。     

北美鹅掌楸原生质体的分离与培养

以鹅掌楸属植物北美鹅掌楸的悬浮细胞和组培苗叶片为材料,对北美鹅掌楸原生质体分离、纯化与培养条件进行研究.结果表明:叶片和悬浮细胞用含有0.1％2-吗啉乙磺酸(MES)和0.6 mol/L甘露醇的Cell ProtoplastWash(60M-CPW)溶液25℃预处理lh效果最好;悬浮细胞最佳酶解液为60M-CPW+ 1％纤维素酶+1％半纤维素酶+0.2％果胶酶Y-23+0.1％ MES,每克材料25℃酶解6h有效原生质体产量可以达到3×106个;叶片最佳酶解液为60M-CPW+2％纤维素酶+1％半纤维素酶+0.2％果胶酶Y-23+0.1％ MES,每克材料25℃酶解10 h有效原生质体产量可以达到11×106个;悬浮细胞原生质体易于培养,在KM8p+1.0 mg/L 2,4-D+0.5 mg/L 6-BA培养基中培养25 d可形成肉眼可见的愈伤组织.     

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。     

玉米秸秆不同部位的组成及其对预处理的影响

不同玉米秸秆部位的成分组成及分布对预处理和酶解影响显著。研究表明:韧皮部与髓芯的成分相近,但叶子的差异较大,其木聚糖和总糖的质量分数最高,分别为29.48%和66.15%,而木质素的质量分数最低,因而叶子更容易预处理。玉米秸秆在稀酸预处理过程中可回收96.9%葡聚糖和50.0%～70.0%木聚糖,其中50.0%～60.0%木聚糖水解成木糖溶出;不同部位的木聚糖损失率与初始的木聚糖含量正相关;经稀酸预处理后,叶子中葡聚糖的质量分数最高,达72.40%,叶子和髓芯易于被纤维素酶水解生成葡萄糖,而韧皮部困难。不同部位的酶解得率与自身的葡聚糖含量正相关,与酸不溶木质素含量负相关,同时受原料的物理结构、葡聚糖和木质素大分子的化学组成等影响。     

Optimization of ammonia fiber expansion (AFEX) pretreatment and enzymatic hydrolysis of Miscanthus x giganteus to fermentable sugars

Miscanthus x giganteus is a tall perennial grass whose suitability as an energy crop is presently being appraised. There is very little information on the effect of pretreatment and enzymatic saccharification of Miscanthus to produce fermentable sugars. This paper reports sugar yields during enzymatic hydrolysis from ammonia fiber expansion (AFEX) pretreated Miscanthus. Pretreatment conditions including temperature, moisture, ammonia loading, residence time, and enzyme loadings are varied to maximize hydrolysis yields. In addition, further treatments such as soaking the biomass prior to AFEX as well as washing the pretreated material were also attempted to improve sugar yields. The optimal AFEX conditions determined were 160 degrees C, 2:1 (w/w) ammonia to biomass loading, 233% moisture (dry weight basis), and 5 min reaction time for water-soaked Miscanthus. Approximately 96% glucan and 81% xylan conversions were achieved after 168 h enzymatic hydrolysis at 1% glucan loading using 15 FPU/(g of glucan) of cellulase and 64 p-NPGU/(g of glucan) of beta-glucosidase along with xylanase and tween-80 supplementation. A mass balance for the AFEX pretreatment and enzymatic hydrolysis process is presented.     

Enzymatic digestion of liquid hot water pretreated hybrid poplar

Liquid hot (LHW) water pretreatment (LHW) of lignocellulosic material enhances enzymatic conversion of cellulose to glucose by solubilizing hemicellulose fraction of the biomass, while leaving the cellulose more reactive and accessible to cellulase enzymes. Within the range of pretreatment conditions tested in this study, the optimized LHW pretreatment conditions for a 15% (wt/vol) slurry of hybrid poplar were found to be 200^oC, 10 min, which resulted in the highest fermentable sugar yield with minimal formation of sugar decomposition products during the pretreatment. The LHW pretreatment solubilized 62% of hemicellulose as soluble oligomers. Hot‐washing of the pretreated poplar slurry increased the efficiency of hydrolysis by doubling the yield of glucose for a given enzyme dose. The 15% (wt/vol) slurry of hybrid poplar, pretreated at the optimal conditions and hot‐washed, resulted in 54% glucose yield by 15 FPU cellulase per gram glucan after 120 h. The hydrolysate contained 56 g/L glucose and 12 g/L xylose. The effect of cellulase loading on the enzymatic digestibility of the pretreated poplar is also reported. Total monomeric sugar yield (glucose and xylose) reached 67% after 72 h of hydrolysis when 40 FPU cellulase per gram glucan were used. An overall mass balance of the poplar‐to‐ethanol process was established based on the experimentally determined composition and hydrolysis efficiencies of the liquid hot water pretreated poplar. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009     

Statistical optimization of sulfite pretreatment of corncob residues for high concentration ethanol production

In this study, a central composite design of response surface method was used to optimize sulfite pretreatment of corncob residues, in respect to sulfite charge (5-10%), treatment time (1-2h), liquid/solid (l/s) ratio (6:1-10:1) and temperature (150-180°C) for maximizing glucose production in enzymatic hydrolysis process. The relative optimum condition was obtained as follows: sulfite charge 7.1%, l/s ratio 7.6:1, temperature 156°C for 1.4h, corresponding to 79.3% total glucan converted to glucose+cellobiose. In the subsequent simultaneous saccharification and fermentation (SSF) experiments using 15% glucan substrates pretreated under this kind of conditions, 60.8 g ethanol l(-1) with 72.2% theoretical yield was obtained.     

Low temperature alkali pretreatment for improving enzymatic digestibility of sweet sorghum bagasse for ethanol production

A low temperature alkali pretreatment method was proposed for improving the enzymatic hydrolysis efficiency of lignocellulosic biomass for ethanol production. The effects of the pretreatment on the composition, structure and enzymatic digestibility of sweet sorghum bagasse were investigated. The mechanisms involved in the digestibility improvement were discussed with regard to the major factors contributing to the biomass recalcitrance. The pretreatment caused slight glucan loss but significantly reduced the lignin and xylan contents of the bagasse. Changes in cellulose crystal structure occurred under certain treatment conditions. The pretreated bagasse exhibited greatly improved enzymatic digestibility, with 24-h glucan saccharification yield reaching as high as 98% using commercially available cellulase and β-glucosidase. The digestibility improvement was largely attributed to the disruption of the lignin-carbohydrate matrix. The bagasse from a brown midrib (BMR) mutant was more susceptible to the pretreatment than a non-BMR variety tested, and consequently gave higher efficiency of enzymatic hydrolysis.     

Novel Single-step Pretreatment of Steam Explosion and Choline Chloride to De-lignify Corn Stover for Enhancing Enzymatic Edibility

It is important to develop efficient and economically feasible pretreatment methods for lignocellulosic biomass, to increase annual biomass production. A number of pretreatment methods were introduced to promote subsequent enzymatic hydrolysis of biomass for green energy processes. Pretreatment with steam explosion removes the only xylan at high severity but increases lignin content. In this study, corn stover soaked in choline chloride solution before the steam explosion is economically feasible as it reduced cost. Enzymatic hydrolysis of de-lignified corn stover is enhanced by combinatorial pretreatments of steam explosion and choline chloride. Corn stover pretreated with choline chloride at the ratio of 1:2.2 (w/w), 1.0 MPa, 184 °C, for 15 min efficiently expelled 84.7% lignin and 78.9% xylan. The residual solid comprised of 74.59% glucan and 7.51% xylan was changed to 84.2% glucose and 78.3% xylose with enzyme stacking of 10FPU/g. This single-step pretreatment had ∼ 4.5 and 6.4 times higher glucose yield than SE-pretreated and untreated corn stover, respectively. Furthermore, SEM, XRD and FTIR indicated the porosity, crystalline changes, methoxy bond-cleavage respectively due to the lignin and hemicellulose expulsion. Thus, the released acetic acid during this process introduced this novel strategy, which significantly builds the viability of biomass in short pretreatment time.     

Optimization of enzymatic hydrolysis and ethanol fermentation from AFEX-treated rice straw

An abundant agricultural residue, rice straw (RS) was pretreated using ammonia fiber expansion (AFEX) process with less than 3% sugar loss. Along with commercial cellulase (Spezyme® CP) at 15 filter paper unit/g of glucan, the addition of Multifect® Xylanase at 2.67 mg protein/g glucan and Multifect® Pectinase at 3.65 mg protein/g glucan was optimized to greatly increase sugar conversion of AFEX-treated RS. During enzymatic hydrolysis even at 6% glucan loading (equivalent to 17.8% solid loading), about 80.6% of glucan and 89.6% of xylan conversions (including monomeric and oligomeric sugars) were achieved. However, oligomeric glucose and xylose accounted for 12.3% of the total glucose and 37.0% of the total xylose, respectively. Comparison among the three ethanologenic strains revealed Saccharomyces cerevisiae 424A(LNH-ST) to be a promising candidate for RS hydrolysate with maximum ethanol metabolic yield of 95.3% and ethanol volumetric productivity of 0.26 g/L/h. The final concentration of ethanol at 37.0 g/L was obtained by S. cerevisiae 424A(LNH-ST) even with low cell density inoculum. A biorefinery combining AFEX pretreatment with S. cerevisiae 424A(LNH-ST) in separate hydrolysis and fermentation could achieve 175.6 g EtOH/kg untreated rice straw at low initial cell density (0.28 g dw/L) without washing pretreated biomass, detoxification, or nutrient supplementation.     

Mild pretreatment of yellow poplar biomass using sequential dilute acid and enzymatically-generated peracetic acid to enhance cellulase accessibility

Biomass contains cellulose, xylan and lignin in a complex interwoven structure that hinders enzymatic hydrolysis of the cellulose. To separate these components in yellow poplar biomass, we sequentially pretreated with dilute sulfuric acid and enzymatically-generated peracetic acid. In the first step, the dilute acid with microwave heating (140°C, 5 min) hydrolyzed 90% of xylan. The xylose yield in hydrolysate after dilute acid pretreatment was 83.1%. In the second step, peracetic acid (60°C, 6 h) removed up to 80% of lignin. This sequential pretreatment fractionated biomass into xylan and lignin, leaving a solid residue enriched in cellulose (~80%). The sequential pretreatment enhanced enzymatic digestibility of the cellulase by removal of the other components in biomass. The glucose yield after enzymatic hydrolysis was 90.5% at a low cellulase loading (5 FPU/g of glucan), which is 1.6 and 18 times higher than for dilute acid-pretreated biomass and raw biomass, respectively. This novel sequential pretreatment with dilute acid and peracetic acid efficiently separates the three major components of yellow poplar biomass, and reduces the amount of cellulase needed.     

Whole slurry fermentation of maleic acid-pretreated oil palm empty fruit bunches for ethanol production not necessitating a detoxification process

The yield of ethanol from oil palm empty fruit bunches (EFB) was increased on exploiting maleic acid pretreatment combined with fermentation of the pretreated whole slurry. The optimized conditions for pretreatment were to expose EFB to a high temperature (190 °C) with 1 % (w/v) maleic acid for a short time duration (3 min ramping to the set temperature with no holding) in a microwave digester. An enzymatic digestibility of 60.9 % (based on theoretical glucose yield) was exhibited using pretreated and washed EFB after 48 h of hydrolysis. Simultaneous saccharification and fermentation (SSF) of the whole slurry of pretreated EFB for 48 h resulted in 61.3 % theoretical yield of ethanol based on the initial amount of glucan in untreated EFB. These results indicate that maleic acid is a suitable catalyst not requiring detoxification steps for whole slurry fermentation of EFB for ethanol production, thus improving the process economics. Also, the whole slurry fermentation can significantly increase the biomass utilization by converting sugar from both solid and liquid phases of the pretreated slurry.     

Two-stage pretreatment of rice straw using aqueous ammonia and dilute acid

Liberation of fermentable sugars from recalcitrant lignocellulosic biomass is one of the key challenges in production of cellulosic ethanol. Here we developed a two-stage pretreatment process using aqueous ammonia and dilute sulfuric acid in a percolation mode to improve production of fermentable sugars from rice straw. Aqueous NH? was used in the first stage which removed lignin selectively but left most of cellulose (97%) and hemicellulose (77%). Dilute acid was applied in the second stage which removed most of hemicellulose, partially disrupted the crystalline structure of cellulose, and thus enhanced enzymatic digestibility of cellulose in the solids remaining. Under the optimal pretreatment conditions, the enzymatic hydrolysis yields of the two-stage treated samples were 96.9% and 90.8% with enzyme loadings of 60 and 15FPU/g of glucan, respectively. The overall sugar conversions of cellulose and hemicellulose into glucose and xylose by enzymatic and acid hydrolysis reached 89.0% and 71.7%, respectively.     

Sugar yields from dilute sulfuric acid and sulfur dioxide pretreatments and subsequent enzymatic hydrolysis of switchgrass

Dacotah switchgrass was pretreated with sulfuric acid concentrations of 0.5, 1.0, and 2.0 wt.% at 140, 160, and 180 °C and with 1 and 3 wt.% sulfur dioxide at 180 °C over a range of times. Sulfur dioxide loadings of 0%, 1%, 3%, 5%, and 10%wt.% of dry biomass were also tested at 180 °C for 10 min. Sugar yields were tracked for pretreatment and subsequent enzymatic hydrolysis to identify conditions for the highest total sugar yields. Pretreatment with 1 wt.% dilute sulfuric acid at 140 °C for 40 min followed by enzymatic hydrolysis with 48.6 mg enzyme/g initial glucan in raw biomass resulted in ～86% of theoretical yield for glucose and xylose combined. For sulfur dioxide pretreatment, the highest total sugar yield of about 87% occurred at 5% SO? for 10 min and 180 °C. However, xylose yields were higher at shorter times and glucose yields at longer times.