Optimization of cellulase mixture for efficient hydrolysis of steam-exploded corn stover by statistically designed experiments (original) (raw)
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Journal of the Japan Petroleum Institute, 2017
Bioethanol is currently employed as a renewable energy to replace gasoline and addresses the issue of fossil fuel depletion. The main feedstocks used to produce bioethanol are sugar and starchy materials, with corn, sugarcane, and cassava typically being employed in the USA, Brazil, and Indonesia, respectively, as these three materials contain sugar or starch compounds that can be fermented to produce ethanol after hydrolysis when necessary. However, the use of such sugary and starchy materials competes with the demand for food supply, resulting in increased prices and the threatening of food security. In this context, lignocellulosic materials from agricultural and forest residues, such as rice straw, sawdust, and palm oil empty fruit bunch, have attracted growing attention as alternative feedstocks for bioethanol production, as they are both abundant and unutilized 1). In the case of lignocellulosic materials, the cellulose present in these materials can be hydrolyzed to give glucose, which can then be employed as a substrate for ethanol fermentation. However, due to the presence of lignin and hemicellulose in the lignocellulosic structure, this potential feedstock must go through an appropriate pretreatment method prior to hydrolysis 2). For example, methods such as ammonia explosion, dilute acid treatment, lime pretreatment, and hydrothermal pretreatment are among a number of techniques reported to date 3). Following such pretreatment, hydrolysis must then be carried out to convert cellulose to glucose, and so this stage is particularly important when considering the production costs of bioethanol 2). For example, hydrolysis by enzymes has been identified as a "green" and environmentally friendly process 4) in which the cellulose polymer is degraded by cellulase to give the monomer glucose, which can in turn be naturally fermented by the yeast Saccharomyces cerevisiae to yield ethanol 5). As such, a number of synergistic studies have been conducted to decrease the enzyme costs for the commercial production of ethanol 6). One potential strategy is low-cost enzyme production and subsequent recovery and reuse of the enzyme. For example, Nojiri et al. 7) reported that alkaline-treated woody biomass can be used as a low-cost material for an enzyme production medium. In addition, Kobayashi et al. 8) employed the functional lignin-based material lignocresol, which was synthesized from hinoki wood meal, to produce immobilized cellulase. An alternative strategy involves the optimization of an enzyme cocktail for saccharification. Currently, several commercial cellulases have been identified to produce cellobiose and glucose as its main products. The composition of these cellulases can be divided into three categories of enzyme, namely endo-1,4-β-g l u c a n a s e s , c e l l o b i o h y d r o l a s e s , a n d 322
Enhanced glucose production from cellulose using coimmobilized cellulase and β-glucosidase
Applied Biochemistry and Biotechnology, 1989
~-Glucosidase was covalently immobilized alone and coimmobilized with cellulase using a hydrophilic polyurethane foam (Hypol | FHP 2002). Immobilization improved the functional properties of the enzymes. When immobiliTed alone, the Km for cellobiose of fl-glucosidase was decreased by 33% and the pH optimum shifted to a slightly more basic value, compared to the free enzyme. ImmobiliTed/~-glucosidase was extremely stable (95% of activity remained after 1000 h of continuous use). CoimraobfliTation of cellulase and/5-glucosidase produced a cellulose-hydrolyzing complex with a 2.5-fotd greater rate of glucose production for soluble cellulose and a four-fold greater increase for insoluble cellulose, compared to immobiliTed cellulase alone. The imrno-biliTed enzymes showed a broader acceptance of various types of insoluble cellulose substrates than did the free enzymes and showed a long-term (at least 24 h) linear rate of glucose production from microcrystalline cellulose. The pH optimum for the coimrnobiliTed enzymes was 6.0. This method for enzyme immobilization is fast, irreversible, and does not require harsh conditions. The enhanced glucose yields obtained indicate that this method may prove useful for commercial cellulose hydrolysis.
Agricultural crop residues viz. sugarcane bagasse, wheat straw, wheat bran, rice husk, banana peel and orange peel have great potential to produce bioethanol as they have immense quantity of carbon source. These agricultural crop residues were pretreated with NaOH for delignification and then subjected to solid state fermentation for the production of cellulase enzyme using Aspergillus niger. Cellulase production parameter such as pH, incubation period and temperature were optimized. Activity of cellulose was checked by FPase method which was found as 3.69 U/ml from wheat bran at pH 6 and at 28 0 C after 6 days of incubation and the cellulase was produced under these optimum conditions. Subsequently enzymatic hydrolysis was done as cellulase play a catalytic role in the hydrolysis of cellulose to glucose monomer units, so that it could be further converted into ethanol. After enzymatic hydrolysis, glucose released was inoculated with Sacchromyces cerevisiae (MCCB 0278) under optimum condition which converted glucose into ethanol. After this, fractional distillation was done for the purification of bio-ethanol produced and specific gravity was also checked.
Role and significance of beta-glucosidases in the hydrolysis of cellulose for bioethanol production
Bioresource Technology, 2013
One of the major challenges in the bioconversion of lignocellulosic biomass into liquid biofuels includes the search for a glucose tolerant beta-gulucosidase. Beta-glucosidase is the key enzyme component present in cellulase and completes the final step during cellulose hydrolysis by converting the cellobiose to glucose. This reaction is always under control as it gets inhibited by its product glucose. It is a major bottleneck in the efficient biomass conversion by cellulase. To circumvent this problem several strategies have been adopted which we have discussed in the article along with its production strategies and general properties. It plays a very significant role in bioethanol production from biomass through enzymatic route. Hence several amendments took place in the commercial preparation of cellulase for biomass hydrolysis, which contains higher and improved beta-glucosidase for efficient biomass conversion. This article presents beta-glucosidase as the key component for bioethanol from biomass through enzymatic route.
Development of cellulolytic strain by genetic engineering approach for enhanced cellulase production
Genetic and Metabolic Engineering for Improved Biofuel Production from Lignocellulosic Biomass , 2020
Recently, the global attention has been shifted toward improving second-generation biofuel, which seems to be the best alternative in solving the challenges of feedstock for bioethanol production as the demand for food is increasing daily due to growing human population. However, finding economically viable hydrolytic enzymes that can degrade lignocellulosic biomass with higher specific activity, better stability, lower susceptibility to inhibition, and improved physicochemical properties has been a bottleneck to researchers. These limitations therefore provide a possibility for strain improvement through genetic and metabolic engineering technologies. This chapter critically examines the classification of cellulolytic enzymes (cellulases and xylanases), methods for hydrolysis, strain improvement strategies through mutagenesis, genetic and metabolic engineering, and directed evolution, epigenetic, promoter, gene deletion approaches, and artificial chimera. Economic outlook and future prospect of cellulolytic enzymes were also examined.
Journal of Biotechnology, 2012
Cellulase, an enzymatic complex that synergically promotes the degradation of cellulose to glucose and cellobiose, free or adsorbed onto Si/SiO 2 wafers at 60 • C has been employed as catalyst in the hydrolysis of microcrystalline cellulose (Avicel), microcrystalline cellulose pre-treated with hot phosphoric acid (CP), cotton cellulose (CC) and eucalyptus cellulose (EC). The physical characteristics such as index of crystallinity (I C), degree of polymerization (DP) and water sorption values were determined for all samples. The largest conversion rates of cellulose into the above-mentioned products using free cellulase were observed for samples with the largest water sorption values; conversion rates showed no correlation with either I C or DP of the biopolymer. Cellulose with large water sorption value possesses large pore volumes, hence higher accessibility. The catalytic efficiency of immobilized cellulase could not be correlated with the physical characteristics of cellulose samples. The hydrolysis rates of the same cellulose samples with immobilized cellulase were lower than those by the free enzyme, due to the diffusion barrier (biopolymer chains approaching to the immobilized enzyme) and less effective contact between the enzyme active site and its substrate. Immobilized cellulase, unlike its free counterpart, can be recycled at least six times without loss of catalytic activity, leading to higher overall cellulose conversion.
Cellulases: From Lignocellulosic Biomass to Improved Production
Energies
Cellulases are enzymes that are attracting worldwide attention because of their ability to degrade cellulose in the lignocellulosic biomass and transform it into highly demanded bioethanol. The enzymatic hydrolysis of cellulose by cellulases into fermentable sugars is a crucial step in biofuel production, given the complex structure of lignocellulose. Due to cellulases’ unique ability to hydrolyze the very recaltricant nature of lignocellulosic biomass, the cellulase market demand is rapidly growing. Although cellulases have been used in industrial applications for decades, constant effort is being made in the field of enzyme innovation to develop cellulase mixtures/cocktails with improved performance. Given that the main producers of cellulases are of microbial origin, there is a constant need to isolate new microorganisms as potential producers of enzymes important for biofuel production. This review provides insight into current research on improving microbial cellulase productio...