Bioconversion Of Lignocellulosic Biomass Research Articles

A novel thermostable GH10 xylanase with activities on a wide variety of cellulosic substrates from a xylanolytic Bacillus strain exhibiting significant synergy with commercial Celluclast 1.5 L in pretreated corn stover hydrolysis

BackgroundCellulose and hemicellulose are the two largest components in lignocellulosic biomass. Enzymes with activities towards cellulose and xylan have attracted great interest in the bioconversion of lignocellulosic biomass, since they have potential in improving the hydrolytic performance and reducing the enzyme costs. Exploring glycoside hydrolases (GHs) with good thermostability and activities on xylan and cellulose would be beneficial to the industrial production of biofuels and bio-based chemicals.ResultsA novel GH10 enzyme (XynA) identified from a xylanolytic strain Bacillus sp. KW1 was cloned and expressed. Its optimal pH and temperature were determined to be pH 6.0 and 65 °C. Stability analyses revealed that XynA was stable over a broad pH range (pH 6.0–11.0) after being incubated at 25 °C for 24 h. Moreover, XynA retained over 95% activity after heat treatment at 60 °C for 60 h, and its half-lives at 65 °C and 70 °C were about 12 h and 1.5 h, respectively. More importantly, in terms of substrate specificity, XynA exhibits hydrolytic activities towards xylans, microcrystalline cellulose (filter paper and Avicel), carboxymethyl cellulose (CMC), cellobiose, p-nitrophenyl-β-d-cellobioside (pNPC), and p-nitrophenyl-β-d-glucopyranoside (pNPG). Furthermore, the addition of XynA into commercial cellulase in the hydrolysis of pretreated corn stover resulted in remarkable increases (the relative increases may up to 90%) in the release of reducing sugars. Finally, it is worth mentioning that XynA only shows high amino acid sequence identity (88%) with rXynAHJ14, a GH10 xylanase with no activity on CMC. The similarities with other characterized GH10 enzymes, including xylanases and bifunctional xylanase/cellulase enzymes, are no more than 30%.ConclusionsXynA is a novel thermostable GH10 xylanase with a wide substrate spectrum. It displays good stability in a broad range of pH and high temperatures, and exhibits activities towards xylans and a wide variety of cellulosic substrates, which are not found in other GH10 enzymes. The enzyme also has high capacity in saccharification of pretreated corn stover. These characteristics make XynA a good candidate not only for assisting cellulase in lignocellulosic biomass hydrolysis, but also for the research on structure–function relationship of bifunctional xylanase/cellulase.

Overcoming biomass recalcitrance by synergistic pretreatment of mechanical activation and metal salt for enhancing enzymatic conversion of lignocellulose

BackgroundDue to biomass recalcitrance, including complexity of lignocellulosic matrix, crystallinity of cellulose, and inhibition of lignin, the bioconversion of lignocellulosic biomass is difficult and inefficient. The aim of this study is to investigate an effective and green pretreatment method for overcoming biomass recalcitrance of lignocellulose.ResultsAn effective mechanical activation (MA) + metal salt (MAMS) technology was applied to pretreat sugarcane bagasse (SCB), a typical kind of lignocellulosic biomass, in a stirring ball mill. Chlorides and nitrates of Al and Fe showed better synergistic effect with MA, especially AlCl3, ascribing to the interaction between metal salt and oxygen-containing groups induced by MA. Comparative studies showed that MAMS pretreatment effectively changed the recalcitrant structural characteristics of lignocellulosic matrix and reduced the inhibitory action of lignin on enzymatic conversion of SCB. The increase in hydroxyl and carboxyl groups of lignin induced by MAMS pretreatment led to the increase of its hydrophilicity, which could weaken the binding force between cellulase and lignin and reduce the nonproductive binding of cellulase enzymes to lignin.ConclusionsMAMS pretreatment significantly enhanced the enzymatic digestibility of polysaccharides substrate by overcoming biomass recalcitrance without the removal of lignin from enzymatic hydrolysis system.

Highly efficient conversion of xylose to ethanol without glucose repression by newly isolated thermotolerant Spathaspora passalidarum CMUWF1\u20132

BackgroundEfficient bioconversion of lignocellulosic biomass to bioethanol is one of key challenges in the situation of increasing bioethanol demand. The ethanologenic microbes for such conversion are required to possess abilities of utilization of various sugars including xylose and arabinose in lignocellulosic biomass. As required additional characteristics, there are a weak or no glucose repression that allows cells to simultaneously utilize various sugars together with glucose and thermotolerance for fermentation at high temperatures, which has several advantages including reduction of cooling cost. Spathaspora passalidarum ATCC MYA-4345, a type strains, isolated previously have mainly of these abilities or characteristics but its thermotolerance is not so strong and its glucose repression on xylose utilization is revealed.ResultsNewly isolated S. passalidarum CMUWF1–2 was found to have a high ability to produce ethanol from various sugars included in lignocellulosic biomass at high temperatures. The strain achieved ethanol yields of 0.43 g, 0.40 g and 0.20 g ethanol/g xylose at 30 °C, 37 °C and 40 °C, respectively. Interestingly, no significant glucose repression was observed in experiments with mixed sugars, being consistent with the strong resistance to 2-deoxyglucose, and antimycin A showed no effect on its growth in xylose medium. Moreover, the strain was tolerant to glucose and ethanol at concentrations up to 35.0% (w/v) and 8.0% (v/v), respectively.ConclusionsS. passalidarum CMUWF1–2 was shown to achieve efficient production of ethanol from various sugars and a high ethanol yield from xylose with little accumulation of xylitol. The strain also exhibited stress-resistance including thermotolerance and no detectable glucose repression as beneficial characteristics. Therefore, S. passalidarum CMUWF1–2 has remarkable potential for conversion of lignocellulosic biomass to bioethanol.

Methane production through anaerobic digestion: Participation and digestion characteristics of cellulose, hemicellulose and lignin

Lignocellulosic biomass is the most abundant natural resource with high biomethane potential. However, complex structure of lignocellulosic biomass has hampered the efficient utilization of this bioresource. Previous studies have investigated the overall anaerobic digestion performance of lignocellulosic biomass, but the individual participation of each lignocellulosic component during anaerobic digestion remained unclear. Thus, this study investigated the methane production characteristics of cellulose, hemicellulose, lignin and their mixtures along with the microbial communities involved in anaerobic digestion. The results showed that the biomethane potential of cellulose was higher than that of hemicellulose; however, hemicellulose was hydrolysed more quickly than cellulose, while lignin was very difficult to be digested. The higher concentrations of acetic, n-butyric and n-valeric acids hydrolysed from the hemicellulose resulted in a lower pH and more severe inhibition on methane production than that of cellulose, and the methanogenesis gradually recovered after pH adjustment. The co-digestion of cellulose and hemicellulose increased the methane yield and biodegradability compared to mono-digestions. The addition of lignin to cellulose brought more significant decrease in the methane yield of cellulose than that of hemicellulose. Substrate-related bacteria such as Clostridium sensu stricto, Lutaonella, Cloacibacillus and Christensenella showed higher relative abundance in cellulose digestate, and sugar-fermenting bacteria such as Saccharofermentans, Petrimonas and Levilinea were more rich in the digestate of hemicellulose. Moreover, methanogenic Methanospirillum and Methanothrix likely contributed to the methane production of cellulose, while aciduric methanogens from Methanobrevibacter, Methanomassiliicoccus, Methanobacterium and Methanoculleus contributed to that of hemicellulose. This study provides a deeper understanding of the mechanism in the bioconversion of lignocellulosic biomass during anaerobic digestion.

Ethanol production using vegetable peels medium and the effective role of cellulolytic bacterial (Bacillus subtilis) pre-treatment.

Background: The requirement of an alternative clean energy source is increasing with the elevating energy demand of modern age. Bioethanol is considered as an excellent candidate to satiate this demand. Methods: Yeast isolates were used for the production of bioethanol using cellulosic vegetable wastes as substrate. Efficient bioconversion of lignocellulosic biomass into ethanol was achieved by the action of cellulolytic bacteria ( Bacillus subtilis). After proper isolation, identification and characterization of stress tolerances (thermo-, ethanol-, pH-, osmo- & sugar tolerance), optimization of physiochemical parameters for ethanol production by the yeast isolates was assessed. Very inexpensive and easily available raw materials (vegetable peels) were used as fermentation media. Fermentation was optimized with respect to temperature, reducing sugar concentration and pH. Results: It was observed that temperatures of 30°C and pH 6.0 were optimum for fermentation with a maximum yield of ethanol. The results indicated an overall increase in yields upon the pretreatment of Bacillus subtilis; maximum ethanol percentages for isolate SC1 obtained after 48-hour incubation under pretreated substrate was 14.17% in contrast to untreated media which yielded 6.21% after the same period. Isolate with the highest ethanol production capability was identified as members of the ethanol-producing Saccharomyces species after stress tolerance studies and biochemical characterization using Analytical Profile Index (API) ® 20C AUX and nitrate broth test. Introduction of Bacillus subtilis increased the alcohol production rate from the fermentation of cellulosic materials. Conclusions: The study suggested that the kitchen waste can serve as an excellent raw material in ethanol fermentation.

Purification and characterization of novel bi-functional GH3 family β-xylosidase/β-glucosidase from Aspergillus niger ADH-11

β-Xylosidase plays an important role in xylan degradation by relieving the end product inhibition of endo-xylanase caused by xylo-oligosaccharides. β-Xylosidase has a wide range of applications in food, feed, paper and pulp, pharmaceutical industries and in bioconversion of lignocellulosic biomass. Hence, in the present study focused on purification, biochemical characterization and partial sequencing of purified β-xylosidase from xylanolytic strain Aspergillus niger ADH-11. Acetone precipitation followed by GPC using Sephacryl S-200 yielded 20.59-fold purified β-xylosidase with 58.30% recovery. SDS-PAGE analysis of purified β-xylosidase relieved a monomeric subunit with a molecular weight 120.48kDa. Kinetic parameters of purified β-xylosidase viz Km, Vmax, Kcat and catalytic efficiency were assessed. Purified β-xylosidase was additionally active on p-nitrophenyl-β-d-glucopyranoside substrate also. Moreover, peptide mass fingerprinting analysis support our biochemical studies and showed that the purified protein is a novel β-xylosidase with β-glucosidase activity and belongs to the bi-functional GH3 superfamily. Besides, tolerance of purified β-xylosidase towards glucose and xylose was also assessed.

Assessment of alkaline peroxide-assisted wet air oxidation pretreatment for rice straw and its effect on enzymatic hydrolysis

Biomass recalcitrance is considered to be one of the impediments in bioconversion of lignocellulosic biomass (LCB) to sugars. Rice straw, a potential lignocellulosic waste, owing to the surplus availability, renewability and high carbohydrate content was used as a model LCB in the present study. The alkaline hydrogen peroxide-assisted wet air oxidation (APWAO) was evaluated as plausible pretreatment for rice straw based on the 23—factorial experimental design with a goal to reduce biomass recalcitrance by enhancing the cellulose recovery, hemicellulose solubilization, lignin removal and concomitantly generating limited degradation products. APWAO resulted in an overall cellulose recovery ranging from 80.54 to 93.02%, hemicellulose solubilization of 36.44–82.08% and lignin removal of 65.95–81.11% (all on w/w basis) respectively and absence of potent inhibitors viz. furfural and 5-hydroxymethylfurfural. The statistically significant pretreatment factors that affected each of these responses were assessed and optimum pretreatment conditions were determined to be biomass soaking in 0.5% H2O2 for 14 h succeeded by wet air oxidation (WAO) at 190 °C, 6 bar, 20 min by multi-objective numerical optimization. Further, the morphological and structural changes occurring as a result of pretreatment were scrutinized using SEM and FT-IR. APWAO further ensued in enhanced cellulose accessibility during enzymatic saccharification indicated by the glucose yield ranging from 113.97 to 200.34 g/kg untreated rice straw. Thus, the combined pretreatment (APWAO) i.e. pre-soaking in alkaline H2O2 followed by WAO was shown to enhance glucose yields owing to significant delignification.

Characterization of natural co-cultures of Piromyces with Methanobrevibacter ruminantium from yaks grazing on the Qinghai-Tibetan Plateau: a microbial consortium with high potential in plant biomass degradation

Anaerobic fungi reside in the gut of herbivore and synergize with associated methanogenic archaea to decompose ingested plant biomass. Despite their potential for use in bioconversion industry, only a few natural fungus–methanogen co-cultures have been isolated and characterized. In this study we identified three co-cultures of Piromyces with Methanobrevibacter ruminantium from the rumen of yaks grazing on the Qinghai Tibetan Plateau. The representative co-culture, namely (Piromyces + M. ruminantium) Yak-G18, showed remarkable polysaccharide hydrolase production, especially xylanase. Consequently, it was able to degrade various lignocellulose substrates with a biodegrading capability superior to most previously identified fungus or fungus–methanogen co-culture isolates. End-product profiling analysis validated the beneficial metabolic impact of associated methanogen on fungus as revealed by high-yield production of methane and acetate and sustained growth on lignocellulose. Together, our data demonstrated a great potential of (Piromyces + M. ruminantium) Yak-G18 co-culture for use in industrial bioconversion of lignocellulosic biomass.

Isolation and screening of cellulolytic fungi from Salacca zalacca leaf litter

Sari SLA, Setyaningsih R, Wibowo NFA. 2017. Isolation and screening of cellulolytic fungi from Salacca zalacca leaf litter. Biodiversitas 18: 1282-1288. Cellulases are the main enzymes in a bioconversion of lignocellulosic biomass. Fungal cellulases have been proven to be better candidates for this process than other microbial cellulases since they secrete free cellulase complex including endoglucanases, cellobiohydrolases, and β-glucosidase. This study was conducted to isolate and screen of cellulolytic fungi from Salacca leaf litter. There were about 12 fungal isolates which were obtained in this research and 8 isolates showed cellulolytic activity. Based on morphological characters, these cellulolytic fungi were identified as belonging to 4 genera, i.e. Penicillium, Aspergillus, Paecilomyces, and Thielaviopsis. Based on clear zone formation, isolate SLL03, SLL06 and SLL10 showed highest cellulolytic activity. Whereas, based on 18S rRNA gene sequence, these isolates were identified as Aspergillums flavus (SLL03), Penicillium sp. (SLL06) Thielaviopsis ethacetica (SLL10). Therefore, these isolates would be good candidates for cellulase producer.

Overexpression of Escherichia coli Acetyl Esterase Using a Strategy of Multi-copy Promoters

Acetyl esterase (AE) plays an important role in bioconversion of lignocellulosic biomass. However, the expression level of AE from wild type strains is relatively low. The aim of the study was to enhance the activity of AE using the strategy of multi-copy promoters. A commercial strain E. coli JM110 was used as the host of expression plasmid. pKK223-3-ybaC (pK-ybaC) is a recombinant plasmid, and it carries ybaC gene (encoding AE) that was derived from E. coli RB3. E. coli JM110 (pK-ybaC; EcKy) was a recombinant strain that contains pK-ybaC. Fragment P3 that contains three copies of the core-tac-promoter and fragment P5 that contains five copies of the core-tac-promoter were synthesized. Plasmid pK-ybaC was extracted from EcKy and was linearized by EcoR I and BamH I to remove its wild promoter. Then fragment P3 was subcloned into linearized pK-ybaC to generate pKTH-ybaC and fragment P5 was inserted into pK-ybaC to generate pKFI-ybaC. In-Fusion kit (Takara) was used to construct recombinant plasmids according to the instruction of the manufacturer. The two plasmids were then transformed into E. coli JM110 to get the recombinant strain E. coli JM110 (pKTH-ybaC; EcTH) and E. coli JM110 (pKFI-ybaC; EcFI). The highest activity of AE from EcTH was 3.04 U mL−1. It was 7.8 times of that from the parental strain E. coli RB3. The peak value of AE activity from EcFI was 35.66 U mL−1, which was 91.4 times of that from E. coli RB3. EcFI showed the highest transcription level of ybaC gene, and its value was 4239 times of that in E. coli RB3. The K m value was 3.49 mmol L−1 and the V max value was 43.29 µmol s−1 mg−1 for AE from EcFI. The K cat/K m value was around fivefold of that from the parental strain E.coli RB3. Two engineered bacterial strains that contain three copies and five copies of the core-tac-promoter were constructed. The activity of AE from the two engineered strains was enhanced significantly compared to that from the parental strain. The results indicated the feasibility of enhancing AE activity using the strategy of multi-copy promoters.

Development of Escherichia coli Strain with Enhanced Enzymatic Activity of Acetyl Esterase Using a Strategy of Tandem Repetitive Promoters

Purpose Acetyl esterase (AE) plays an important role in bioconversion of lignocellulosic biomass. However, the expression of AE from wild type strains is at relatively low level. The study aims to construct an engineered Escherichia coli strain with enhanced enzymatic activity of AE by adopting the strategy of tandem repetitive promoters.

Production, Partial Purification and Characterization of Enzyme Cocktail from Trichoderma citrinoviride AUKAR04 Through Solid-State Fermentation

A strain of Trichoderma citrinoviride AUKAR04 was identified on the basis of morphological and 5.8S ribosomal RNA sequencing [GenBank: KF698728]. It produces cocktail of enzymes such as xylanase (55,000 IU gds−1), CMCase (375 IU gds−1) and β-1,3-glucanase (695 IU gds−1) after 72 h under solid-state fermentation. These enzymes were partially purified by a three-phase partitioning method, which recovered the maximum activities of xylanase (99.8 %) with 5.7-fold, CMCase (96.5 %) with 5.5-fold and β-1,3-glucanase (98.4 %) with 5.6-fold purification. The maximum activity of xylanase was observed at pH 5.0, CMCase at pH 5.0–6.0 and β-1,3-glucanase at pH 6.0. Optimum temperature of xylanase and β-1,3-glucanase was found to be at 50 °C, while for CMCase was at 60 °C. The activities of these enzymes were enhanced by Mg2+ and Mn2+ ions. Eucalyptus pulp fiber was incubated for 14 h with the enzyme cocktail. Xylanase hydrolyzed the pulp to yield arabinose (475 mg L−1) and xylose (1795 mg L−1), CMCase and β-1,3-glucanase released glucose (18763 mg L−1). The length of fiber was reduced from 0.881 to 0.056 mm. This is indicative of the potential application on bioconversion of lignocellulosic biomass into fermentable sugars by the enzyme cocktail produced from T. citrinoviride AUKAR04 for sustainable production of bioethanol.

Bioconversion of Cellulose to Biofuels

Major focus of Biotechnology in the early 1980s was in understanding the Life Science. But to meet societal energy challenge, understanding the capabilities of Life Sciences is at different stage today. Increase in industrialisation and motorisation has led to steep rise of demand for petroleum-based fuels. The sources of these fossil fuels are becoming exhausted, and now it is time to think about an alternative source. In this context bioconversion of biomass into biofuels seems to be a promising, sustainable and an eco-friendly technology. Lot of research has been done to convert most abundant source of biomass, cellulose to produce biofuels. Strain improvement methods, recombinant DNA technologies and selection of strains by breeding method are some of the methods used for enhancing biofuels production at industrial-scale. This review mainly focuses on bioconversion of Lignocellulosic biomass into biofuels.

Investigation of a submerged membrane reactor for continuous biomass hydrolysis

Enzymatic hydrolysis of cellulose is one of the most costly steps in the bioconversion of lignocellulosic biomass. Use of a submerged membrane reactor has been investigated for continuous enzymatic hydrolysis of cellulose thus allowing for greater use of the enzyme compared to a batch process. The submerged 0.65μm polyethersulfone microfiltration membrane avoids the need to pump a cellulose slurry through an external loop. Permeate containing glucose is withdrawn at pressures slightly below atmospheric pressure. The membrane rejects cellulose particles and cellulase enzyme bound to cellulose. Here proof-of-concept experiments have been conducted using a modified, commercially available membrane filtration cell under low fluxes around 75L/(m2h). The operating flux is determined by the rate of glucose production. Maximizing the rate of glucose production involves optimizing mixing, reactor holding time, and the time the feed is held in the reactor prior to commencement of membrane filtration and continuous operation. Maximizing glucose production rates will require operating at low glucose concentration in order to minimize the adverse effects of product inhibition. Consequently practical submerged membrane systems will require a combined sugar concentration step in order to concentrate the product sugar stream prior to fermentation.

Biological pre-treatment: Enhancing biogas production using the highly cellulolytic fungus Trichoderma viride

With regard to renewable sources of energy, bioconversion of lignocellulosic biomass has long been recognized as a desirable endeavor. However, the highly heterogeneous structure of lignocellulose restricts the exploitation of its promising potential in biogas plants. Hence, effective pre-treatment methods are decisive prerequisites to overcome these challenges in order to improve the utilization ratio of (ligno) cellulosic substrates during fermentation. In the present study, the application of Trichoderma viride in an aerobic upstream process prior to anaerobic digestion led up to a threefold increase in the yield of methane and total gas in a lab-scale investigation. Due to its highly efficient cellulolytic activities, T. viride seemed to be responsible for an improved nutrient availability that positively influenced the anaerobic microbiocenosis. Aerobic pre-treatment of organic matter with T. viride is therefore a promising solution to achieve higher methane yields and degradation performances without any additional energy demand, nor undesired by-product inhibition.

Isolation and Characterisation of Lignolytic Bacteria for Bioethanol Production

In the present century, usage of non-renewable hydrocarbon fuels for sufficing energy demands are not considered environmentally and ecologically sustainable. Second generation biofuel production as lignocellulosic bioethanol is considered a sustainable way for generating fuel. Bioethanol production is considered as an option among other thermo-chemical processes for providing energy in form of ethanol, preserving the ecosystem by reducing the pollutants emitted during fossil fuel combustion. The release of fermentable sugars had been a challenging issue due to the presence of lignin in plant cell walls. This paper focused its study on isolation of novel lignin degrading enzymes from bacteria present in the soil of Western Ghats of India as the region is unhabituated followed by its application for biodegradation of lignocellulosic biomass for bioethanol production. Much has been done for lignin degrading fungi but still there is not a single method of choice which can be applicable on industrial scale successfully. Hence, the present study focuses on the isolation of novel bacteria and their enzymes with lignolytic potential and their application in bioconversion of lignocellulosic biomass into ethanol.

A Reusable Biomimetic Magnetic Nanoenzyme for Cellulosic Biomass Degradation

Synthesis of reusable biomimetic enzymes was generated with the goal of cost-effective production of the enzyme for the application in bioconversion of lignocellulosic biomass. The current report describes the preparation and characterization of a biomimetic magnetic nanoenzyme (BMNE), functionalized with a carboxylic acid group that hydrolyzes β-1,4-glycosidic bonds with high catalytic efficiency. The nanometer scale of a BMNE allows it to structurally mimic the active sites of biomass-degrading enzymes. The BMNE functioned optimally at pH 3. The combination of BMNE with natural enzymes had a synergistic action on rice straw, increasing sugar release by 29.7 % and reducing hydrolysis time relative to the natural enzyme alone. The BMNE retained significant activity after five consecutive reaction cycles.

Hydration treatments increase the biodegradability of native wheat straw for hydrogen production by a microbial consortium

Enhancing substrate accessibility is one of the prerequisites for efficient pretreatment and bioconversion of lignocellulosic biomass, for which reducing particle size and increasing porosity have been implemented. Biomass porosity, characterized as the water retention capacity (WRC), increases significantly with the use of a culture filtrate of an H2-producing microbial consortium, causing native lignocellulosic fibers to hydrate and swell extensively. This study describes the effects of hydration treatment and particle size on the biodegradability of native wheat straw fibers with regard to direct H2 production by a microbial consortium. A culture filtrate and tap water were used as hydration media, and particle sizes of 0.212 and 3.35 mm were tested. As a result, biodegradability, measured as H2 production, doubled when the lignocellulosic substrate was hydrated with the culture filtrate. Also, hydration had a significant impact on H2 yield, increasing it from 6.8 to 86.5 mL H2/g total consumed sugars when the 0.212 mm fibers were first hydrated. H2 production was higher with coarser particles (3.35 > 0.212 mm), but hydrogen yield improved with finer particles (0.212 > 3.35 mm). Based on our results, particle size and hydration improve the biodegradability of native wheat straw for H2 production, although hydration with a culture filtrate increased the biomass porosity (measured as WRC), which influenced the biodegradability more than a reduction in particle size.

Production and characterization of cellulolytic enzymes by isolated<br>Klebsiella sp. PRW-1 using agricultural waste biomass

The efficient cellulolytic microorganism was isolated from soil samples collected from Shivaji University campus. Biochemical test and phylogenetic analysis of isolated culture identified as Klebsiella sp. PRW-1. The isolated culture could utilize pure cellulosic substrates (carboxymethylcellulose (CMC) and avicel) and different agricultural wastes like sugarcane bagasse, sugarcane barbojo, sorghum husks, grass powder, corn straw and paddy straw by producing a large amount of endoglucanase, exoglucanase, β-glucosidase, filter paperase (FPU), xylanase and glucoamylase. The reducing sugar production was found higher in the presence of grass powder and sugarcane barbojo. Effects of different physic-chemical parameters to achieve maximum cellulolytic enzymes production were systematically investigated. The effects of supplementation of different metals additives as well as the stability in the presence of higher temperature, pH and commercial detergents, on cellulolytic enzymes were also studied. The foregoing result increases the applicability of the strain for the utilization and bioconversion of lignocellulosic biomass that could be used for bioenergy production.

EUCALYPTUS- A LIGNOCELLULOSIC BIOMASS WASTE AS A MAJOR SOURCE IN BIOETHANOL PRODUCTION

Bioconversion of lignocellulosic biomass like eucalyptus leaves to ethanol production depends on the chemical complexity of biomass. Thus, such biomasses play a challenge to be used as feed stocks for cellulosic ethanol production. The complexity of the biomass can be simplified by pretreatment which may remove or modify the surrounding matrix of lignin and hemicellulose prior to the enzymatic hydrolysis of the polysaccharides in the biomass. Pretreatment is a process that converts lignocellulosic biomass from its native form to a form were cellulose hydrolysis is much more effective. In general, pretreatment methods can be classified into three categories, including physical, chemical, and biological pretreatment. The present investigation deals with the isolation of commercially important bacteria from marine sources which have the potential to breakdown lignocellulosic biomasses. The isolates were screened and the organism exhibiting maximum biodegradation capacity against the substrate was identified as Bacillus pumilus. Different agrowaste were degraded using the isolate and eucalyptus was found to be one of the substrates having the potential to yield high amount of simple sugars which could be used for the production of ethanol by S. cerevisiae. This ability of degradation was enhanced by the pretreatment of Eucalyptus leaves in preparation for enzymatic hydrolysis and microbial fermentation for cellulosic ethanol production.