酵母对木质纤维素酸解物中抑制物的应答及菌株开发

木质纤维素稀酸预处理过程中产生的抑制物会干扰酵母细胞的生长和发酵。根据酵母对抑制物应答的特点,开发那些能够对抑制物原位脱毒的高耐受性菌株,是生物质乙醇转化工业可持续发展的关键。综述了木质纤维素稀酸预处理过程中抑制物的产生、分类、对酵母的影响以及酵母对其应答的特点,结合系统生物学和基因工程方法从酵母耐受的角度探讨了耐受性优势酵母菌株的开发。     

大肠杆菌对木质纤维素水解液抑制物的胁迫耐受性

木质纤维素类生物质是前景广阔的化石原料替代品，其生物炼制可生产生物能源、生物基化学品和生物材料等多种产品，可降低碳排放，有助于实现“双碳”目标，因此受到越来越多的关注。然而，木质纤维素生物炼制需要经过预处理、微生物发酵和产物纯化等多个步骤，其中，预处理过程产生的多种化合物抑制微生物的细胞生长和发酵性能，是制约生物转化效率的瓶颈之一。大肠杆菌是木质纤维素生物炼制常用的宿主，被广泛应用于多种化合物的生产，研究其对木质纤维素水解液中抑制物的耐受性，对于提高木质纤维素生物炼制效率具有重要意义。本文首先介绍了木质纤维素的主要成分和基本结构，对木质纤维素的预处理方法以及预处理后水解液中的主要抑制物种类进行了简单阐述；随后，总结了木质纤维素水解液中几类主要抑制物呋喃类、羧酸类和酚类对大肠杆菌细胞的毒性，以及大肠杆菌对上述抑制物的胁迫响应机制和基于机制的菌株改造靶点；最后，综述了提高大肠杆菌对上述抑制物的胁迫耐受性的菌株改造策略，包括随机突变、实验室适应性进化和组学辅助的理性设计等，为利用代谢工程构建用于木质纤维素生物炼制的高效大肠杆菌菌株提供参考。     

过程工程在木质纤维素发酵抑制物解除中的应用

发酵抑制物对宿主细胞产生毒害作用,是木质纤维素生物炼制的主要瓶颈之一。减少抑制物含量、解除抑制作用是提高发酵效率的重要环节。本文讨论了木质纤维素发酵抑制物的来源、组成、特点以及相应的解除方法,提出了"源头降低抑制物—纤维素木质素分级转化"炼制模式和"发酵促进剂设计技术",为木质纤维素发酵抑制物的解除及木质纤维素开发利用提供了全新的技术路线。     

纤维素乙醇生物加工过程中的抑制物对酿酒酵母的影响及应对措施

以木质纤维素为原料生产乙醇,预处理是必需的环节,这一过程中不可避免产生了多种对微生物有抑制作用的化合物,这些抑制物主要有3大类:弱酸、呋喃醛类和酚类化合物。这些化合物影响后续乙醇发酵微生物酿酒酵母(Saccharomyces cerevisiae)的生长及发酵性能,降低了乙醇的得率和产量,是木质纤维素原料大规模生产乙醇的一个主要障碍。以下介绍了3类抑制物的形成及作用机制,并介绍了应对抑制物作用、提高酵母发酵能力的措施及研究进展,包括发酵前预处理原料脱毒、通过进化工程驯化菌种或通过对抑制物耐受性相关基因的代谢工程操作提高酿酒酵母耐受性,及通过发酵过程控制减少抑制物影响等。     

利用木质纤维素生产丁醇的研究进展

随着化石燃料的逐年减少,以生物质为原料的生物能源研究近年来成为能源领域的研究热点,充分利用可再生生物质为发展经济的生物燃料生产工艺提供了一个极好的机会。与燃料乙醇和生物柴油相比,生物丁醇更具有优越性,以可再生木质纤维素生物质为原料进行发酵生产丁醇在近年来被广泛的研究。对于利用可再生生物质为原料生产丁醇,需要解决原料的选择、产品收率低、抑制物对生产菌株毒性等问题。本文对以木质纤维素生物质为原料进行生物丁醇发酵过程中的原料预处理、抑制物对丁醇生产菌的影响,以及水解液的脱毒和耐抑制物菌株的选育等方面进行综述,并对以木质纤维素生产燃料丁醇所面临的机遇与问题进行了简要评述。     

木质纤维素预处理液膜分离脱毒技术研究进展

木质纤维素预处理会产生各种抑制物,严重影响后续的酶水解和微生物发酵。因此,对木质纤维素预处理液进行脱毒处理成为木质纤维素高效经济生物转化的前提条件之一。重点介绍了膜分离技术应用于木质纤维素预处理液的脱毒过程所取得的研究进展。这些膜分离技术包括膜萃取、膜吸附、纳滤、反渗透、电渗析、电去离子、膜电容脱盐、渗透汽化和膜蒸馏等。膜分离技术在木质纤维素预处理液脱毒领域具有十分广阔的应用前景。     

木质纤维素生物质预处理研究现状

预处理是木质纤维素生物质转化为燃料乙醇的关键步骤,综述了现有常见预处理技术的国内外研究现状,同时分析比较了各处理技术的优缺点,并对今后木质纤维素生物质预处理的主要研究方向进行了展望,以期为木质纤维素生物质转化条件的优化提供参考。     

丝状真菌Amorphotheca resinae ZN1的糠醛降解代谢分析

木质纤维素在预处理过程产生的降解产物对后续的酶水解和微生物发酵过程产生了强烈的抑制。因此,这些抑制物的脱除即所谓的"脱毒"步骤是正常进行后续酶解和发酵的前提条件。我们对本实验室筛选的丝状真菌Amorphotheca resinae ZN1的糠醛的代谢路径进行了研究。丝状真菌A.resinae ZN1转化糠醛的降解代谢途径可以简述为:糠醛首先快速地转化为毒性较低的糠醇;在有氧条件下,糠醇又再度生成不致对微生物产生危害的低浓度糠醛,糠醛继续氧化为糠酸。推测糠酸可能继续进入TCA循环,进而完成糠醛的完全降解。研究结果为将来加快丝状真菌A.resinae ZN1生物脱毒速率、改善木质纤维素生物转化的限速步骤提供了重要的实验依据。     

微生物利用木质纤维素的研究进展

木质纤维素原料是世界上最为丰富的资源之一,可用作微生物发酵生产高附加值生物化学品的原料。与传统用于微生物发酵的可食用生物质原料相比,目前微生物利用木质纤维素还存在以下几个关键问题:开发经济有效的木质纤维素预处理工艺、提高微生物对木质纤维素水解液中第二大单糖木糖的有效利用水平、增强微生物对木质纤维素水解液中混糖的综合利用能力以及提高微生物对木质纤维素水解液中糠醛、乙酸等发酵抑制物的耐受能力。综述了近年来国内外针对这几个关键问题的最新研究成果。为今后微生物大规模利用木质纤维素进行商业生产提出了展望和建议。     

预处理对木质纤维素生物质细胞壁超微结构的影响

预处理是提高木质纤维素生物质向生物燃料转化的有效途径,但生物质的天然抗降解屏障严重阻碍了这一转化的进行,因此全面了解预处理过程中植物细胞壁的微观结构及区域化学变化是实现农林生物质高效转化的基础。本文总结了多种预处理方法对植物细胞壁超微结构影响的研究进展,对生物质科学研究可能有一定的促进和指导作用。     

Molecular mechanisms of yeast tolerance and in situ detoxification of lignocellulose hydrolysates

Pretreatment of lignocellulose biomass for biofuel production generates inhibitory compounds that interfere with microbial growth and subsequent fermentation. Remediation of the inhibitors by current physical, chemical, and biological abatement means is economically impractical, and overcoming the inhibitory effects of lignocellulose hydrolysate poses a significant technical challenge for lower-cost cellulosic ethanol production. Development of tolerant ethanologenic yeast strains has demonstrated the potential of in situ detoxification for numerous aldehyde inhibitors derived from lignocellulose biomass pretreatment and conversion. In the last decade, significant progress has been made in understanding mechanisms of yeast tolerance for tolerant strain development. Enriched genetic backgrounds, enhanced expression, interplays, and global integration of many key genes enable yeast tolerance. Reprogrammed pathways support yeast functions to withstand the inhibitor stress, detoxify the toxic compounds, maintain energy and redox balance, and complete active metabolism for ethanol fermentation. Complex gene interactions and regulatory networks as well as co-regulation are well recognized as involved in yeast adaptation and tolerance. This review presents our current knowledge on mechanisms of the inhibitor detoxification based on molecular studies and genomic-based approaches. Our improved understanding of yeast tolerance and in situ detoxification provide insight into phenotype-genotype relationships, dissection of tolerance mechanisms, and strategies for more tolerant strain development for biofuels applications.     

木质纤维素预处理及高值化技术研究进展

木质纤维素生物质是地球上最丰富的可再生生物资源.随着化石能源的消耗及环境的污染,以取代石化燃料为目标的由生物质向生物燃料的转化受到了广泛的关注.木质纤维素有很强的天然抗降解屏障,需先通过物理、化学及微生物等手段进行预处理,进而以更低的成本和更高的效率转化为生物燃料及其他高附加值产品.本文在总结酸碱等传统预处理方法优缺点...     

Soluble inhibitors/deactivators of cellulase enzymes from lignocellulosic biomass

Liquid hot water, steam explosion, and dilute acid pretreatments of lignocellulose generate soluble inhibitors which hamper enzymatic hydrolysis as well as fermentation of sugars to ethanol. Toxic and inhibitory compounds will vary with pretreatment and include soluble sugars, furan derivatives (hydroxymethyl fulfural, furfural), organic acids (acetic, formic and, levulinic acid), and phenolic compounds. Their effect is seen when an increase in the concentration of pretreated biomass in a hydrolysis slurry results in decreased cellulose conversion, even though the ratio of enzyme to cellulose is kept constant. We used lignin-free cellulose, Solka Floc, combined with mixtures of soluble components released during pretreatment of wood, to prove that the decrease in the rate and extent of cellulose hydrolysis is due to a combination of enzyme inhibition and deactivation. The causative agents were extracted from wood pretreatment liquid using PEG surfactant, activated charcoal or ethyl acetate and then desorbed, recovered, and added back to a mixture of enzyme and cellulose. At enzyme loadings of either 1 or 25mg protein/g glucan, the most inhibitory components, later identified as phenolics, decreased the rate and extent of cellulose hydrolysis by half due to both inhibition and precipitation of the enzymes. Full enzyme activity occurred when the phenols were removed. Hence detoxification of pretreated woods through phenol removal is expected to reduce enzyme loadings, and therefore reduce enzyme costs, for a given level of cellulose conversion.     

Comparison of different pretreatment methods for lignocellulosic materials. Part I: conversion of rye straw to valuable products

The conversion of lignocellulose to valuable products requires I: a fractionation of the major components hemicellulose, cellulose, and lignin, II: an efficient method to process these components to higher valued products. The present work compares liquid hot water (LHW) pretreatment to the soda pulping process and to the ethanol organosolv pretreatment using rye straw as a single lignocellulosic material. The organosolv pretreated rye straw was shown to require the lowest enzyme loading in order to achieve a complete saccharification of cellulose to glucose. At biomass loadings of up to 15% (w/w) cellulose conversion of LHW and organosolv pretreated lignocellulose was found to be almost equal. The soda pulping process shows lower carbohydrate and lignin recoveries compared to the other two processes. In combination with a detailed analysis of the different lignins obtained from the three pretreatment methods, this work gives an overview of the potential products from different pretreatment processes.     

Detoxification of Lignocellulose Hydrolysates: Biochemical and Metabolic Engineering Toward White Biotechnology

Chemical hydrolysis of lignocellulosic biomass (LB) produces a number of inhibitors in addition to sugars. These inhibitors include lignin-derived phenolics, carbohydrate-derived furans, and weak acids that have shown a marked effect on the productivities of various metabolites and the growth of biocatalysts in the fermentative reaction. In the past, a number of physicochemical and biological approaches have been proposed to overcome these fermentation inhibitors, including modified fermentative strategies. Additionally, the timely intervention of genetic engineering has provided an impetus to develop suitable technologies for the detoxification of lignocellulosics in biorefineries. However, the improvements in detoxification strategies for lignocellulose hydrolysates have resulted in significant loss of sugars after detoxification. Hydrolysis of myco-LB (LB after fungal pretreatment) has been recognized as a promising approach to avoid fermentation inhibitors and improve total sugar recovery. Biotechnological inventions have also made it possible to widen the range of suitable biocatalysts for biorefineries by microbial-routed induction of enzymatic expression for the elimination of inhibitors, eventually improving ethanol production from acid hydrolysates. This article aims to highlight the strategies that have been adopted to detoxify lignocellulosic hydrolysates and their effects on the chemical composition of the hydrolysates to improve the fermentability of lignocellulosics. In addition, genetic manipulation could widen the availability and variety of substrates and modify the metabolic routes to produce bioethanol or other value-added compounds in an efficient manner.     

By-products resulting from lignocellulose pretreatment and their inhibitory effect on fermentations for (bio)chemicals and fuels

Lignocellulose might become an important feedstock for the future development of the biobased economy. Although up to 75 % of the lignocellulose dry weight consists of sugar, it is present in a polymerized state and cannot be used directly in most fermentation processes for the production of chemicals and fuels. Several methods have been developed to depolymerize the sugars present in lignocellulose, making the sugars available for fermentation. In this review, we describe five different pretreatment methods and their effect on the sugar and non-sugar fraction of lignocellulose. For several pretreatment methods and different types of lignocellulosic biomass, an overview is given of by-products formed. Most unwanted by-products present after pretreatment are dehydrated sugar monomers (furans), degraded lignin polymers (phenols) and small organic acids. Qualitative and quantitative effects of these by-products on fermentation processes have been studied. We conclude this review by giving an overview of techniques and methods to decrease inhibitory effects of unwanted by-products.     

A review of biological delignification and detoxification methods for lignocellulosic bioethanol production

Future biorefineries will integrate biomass conversion processes to produce fuels, power, heat and value-added chemicals. Due to its low price and wide distribution, lignocellulosic biomass is expected to play an important role toward this goal. Regarding renewable biofuel production, bioethanol from lignocellulosic feedstocks is considered the most feasible option for fossil fuels replacement since these raw materials do not compete with food or feed crops. In the overall process, lignin, the natural barrier of the lignocellulosic biomass, represents an important limiting factor in biomass digestibility. In order to reduce the recalcitrant structure of lignocellulose, biological pretreatments have been promoted as sustainable and environmentally friendly alternatives to traditional physico-chemical technologies, which are expensive and pollute the environment. These approaches include the use of diverse white-rot fungi and/or ligninolytic enzymes, which disrupt lignin polymers and facilitate the bioconversion of the sugar fraction into ethanol. As there is still no suitable biological pretreatment technology ready to scale up in an industrial context, white-rot fungi and/or ligninolytic enzymes have also been proposed to overcome, in a separated or in situ biodetoxification step, the effect of the inhibitors produced by non-biological pretreatments. The present work reviews the latest studies regarding the application of different microorganisms or enzymes as useful and environmentally friendly delignification and detoxification technologies for lignocellulosic biofuel production. This review also points out the main challenges and possible ways to make these technologies a reality for the bioethanol industry.     

溶解性多糖单加氧酶的研究进展

木质纤维素是地球上储藏量最为丰富的可再生生物资源。将木质纤维素酶解成寡糖或单糖是生物质利用的关键。然而,传统的糖苷水解酶很难对其进行有效降解。溶解性多糖单加氧酶是一种全新的生物质降解酶,丰富了生物质降解的模式。它以氧化方式作用于糖链,产生更多的还原端以便糖苷水解酶能进一步进行催化。本文综述了LPMO的发现历史、分类、作用机制与活性测定方法,并讨论了LPMO在饲料添加剂、功能性食品与生物能源等领域的应用前景。