Total Cellulase Activity Research Articles

Ganoderma lucidum driven fermentation of Rosa roxburghii pomace: Effects on noodle physicochemical properties, digestion, and gut microbiota

Rosa roxburghii pomace (RRP) is a high dietary fiber byproduct that is underutilized. This study investigated the effects of Ganoderma lucidum fermented Rosa roxburghii pomace (FRRP) on noodle cooking characteristics, texture, structure, in vitro digestion, and fermentation. The results showed that soluble dietary fiber, active components (polysaccharides, triterpenoids and dietary flavonoids), and the total cellulase (Filter paper enzymes, Carboxymethyl cellulase), β-glucosidase and laccase activity were significantly increased in FRRP (p < 0.05). FRRP improved the cooking and sensory properties of noodles and inhibited starch hydrolysis during in vitro digestion. The resistant starch content in noodles with 15 % FRRP increased by 12.63 %, and the predicted glycemic index decreased by 9.34 %. Moreover, the intestinal microbiota structure was significantly improved, promoting the growth of beneficial bacteria such as Bifidobacterium and Lactobacillus. This study contributed to the high-value and environmentally friendly utilization of RRP and provided new insights into the development of efficient noodles.

Production of Cellulolytic Enzymes by Paecilomyces TP08 Using Grape Stalk and Wheat Bran as Substrates in Solid State Culture

Objective: The objective of this study is to investigate the use of grape stems and wheat bran as substrates in solid state fermentation, for the production of cellulolytic enzymes using the fungus Paecilomyces TP08 sp. Theoretical Framework: In grape production, driven by increased consumption and exports, by-products represent 30% of the total volume of grapes processed for wine, requiring efficient management to ensure sustainability. Method: The methodology adopted for this research involves solid-state cultivation, with a substrate made up of 50% grape stems and 50% wheat bran, with samples taken every 24 hours. Each sample taken went through a filtration process, followed by enzyme activity analysis using the DNS method. Results and Discussion: The results obtained showed that the cultivation with the association of wheat bran and grape stalks presented good total cellulase activity during the process, value of 0.846 U/g in 144h, followed by the activity of 0.663U/g observed in 24h. Research Implications: The practical and theoretical implications of this research are discussed, providing insights into how the results can be applied or influence practices in the field of industrial biotechnology. These implications could cover the food, textile and pharmaceutical industries. Originality/Value: This study contributes to the literature by associating grape stalks with wheat bran for the production of fungal enzymes. The relevance and value of this research are evidenced by the use of an innovative substrate (grape stalks) associated with a conventional substrate (wheat bran) for induction of the microorganism little used in the literature for enzyme production.

Exploring the cellulolytic activity of environmental mycobacteria

Although studies on non-tuberculous mycobacteria have increased in recent years because they cause a considerable proportion of infections, their cellulolytic system is still poorly studied. This study presents a characterization of the cellulolytic activities of environmental mycobacterial isolates derived from soil and water samples from the central region of Argentina, aimed to evaluate the conservation of the mechanism for the degradation of cellulose in this group of bacteria. The molecular and genomic identification revealed identity with Mycolicibacterium septicum. The endoglucanase and total cellulase activities were assessed both qualitatively and quantitatively and the optimal enzymatic conditions were characterized. A specific protein of around 56 kDa with cellulolytic activity was detected in a zymogram. Protein sequences possibly arising from a cellulase were identified by mass spectrometry-based shotgun proteomics. Results showed that M. septicum encodes for cellulose- and hemicellulose-related degrading enzymes, including at least an active β-1,4 endoglucanase enzyme that could be useful to improve its survival in the environment. Given the important health issues related to mycobacteria, the results of the present study may contribute to the knowledge of their cellulolytic system, which could be important for their ability to survive in many different types of environments.

Lignocellulolytic Enzymes Production by Four Wild Filamentous Fungi for Olive Stones Valorization: Comparing Three Fermentation Regimens.

Lignocellulolytic enzymes play a crucial role in efficiently converting lignocellulose into valuable platform molecules in various industries. However, they are limited by their production yields, costs, and stability. Consequently, their production by producers adapted to local environments and the choice of low-cost raw materials can address these limitations. Due to the large amounts of olive stones (OS) generated in Morocco which are still undervalued, Penicillium crustosum, Fusarium nygamai, Trichoderma capillare, and Aspergillus calidoustus, are cultivated under different fermentation techniques using this by-product as a local lignocellulosic substrate. Based on a multilevel factorial design, their potential to produce lignocellulolytic enzymes during 15 days of dark incubation was evaluated. The results revealed that P. crustosum expressed a maximum total cellulase activity of 10.9 IU/ml under sequential fermentation (SF) and 3.6 IU/ml of β-glucosidase activity under submerged fermentation (SmF). F. nygamai recorded the best laccase activity of 9 IU/ml under solid-state fermentation (SSF). Unlike T. capillare, SF was the inducive culture for the former activity with 7.6 IU/ml. A. calidoustus produced, respectively, 1,009 μg/ml of proteins and 11.5 IU/ml of endoglucanase activity as the best results achieved. Optimum cellulase production took place after the 5th day under SF, while ligninases occurred between the 9th and the 11th days under SSF. This study reports for the first time the lignocellulolytic activities of F. nygamai and A. calidoustus. Furthermore, it underlines the potential of the four fungi as biomass decomposers for environmentally-friendly applications, emphasizing the efficiency of OS as an inducing substrate for enzyme production.

Response of soil microbial glycoside hydrolase family 6 cellulolytic population to lignocellulosic biochar reveals biochar stability toward microbial degradation.

Biochar produced from lignocellulosic biomass offers an opportunity to recycle waste into a valuable soil amendment. The application of biochar has been proposed to mitigate climate change by sequestering carbon in the soil. However, the field impact of biochar treatment on the cellulolytic microbial populations involved in the earlier steps of cellulose degradation is poorly understood. A field trial spanning three consecutive crop cycles of Zea mays was conducted in a degraded tropical Ultisol of Peninsular Malaysia. The soil was amended with two contrasting biochar made from oil palm kernel shells (pyrolyzed at 400°C) and rice husks (gasified at 800°C) with or without fertilizer supplementation. Soil samples were taken at each harvesting stage and analyzed for total organic carbon, labile active organic carbon, total cellulase, and β-glucosidase. Microbial glycoside hydrolase family 6 (GH6) cellulase genes and transcripts, involved in the early steps of cellulose degradation, were quantified from the extracted soil deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), respectively. Total organic carbon, labile active organic carbon, and β-glucosidase activity were significantly increased, while no effect on total cellulase activity was found. Both biochars stimulated the total population (DNA-derived) abundance of soil microorganisms harboring the GH6 cellulase genes. The biochar amendment did not affect the active population (RNA-derived) of the GH6 cellulolytic community, showing no significant changes in transcript expression. This indirectly corroborates the role of biochar as a potential carbon sequester in the soil.

Different Putative Methyltransferases Have Different Effects on the Expression Patterns of Cellulolytic Genes.

Putative methyltranferase LaeA and LaeA-like proteins, conserved in many filamentous fungi, regulate fungal growth, development, virulence, the biosynthesis of secondary metabolites, and the production of cellulolytic enzymes. Penicillium oxaliucm is a typical fungus that produces cellulolytic enzymes. In this study, we reported the biological function of eight putative methyltransferases (PoMtr23C/D/E/F/G/H and PoMtr25A/B) containing a methyltransf_23 or methyltransf_25 domain, with a focus on their roles in the production of cellulolytic enzymes. In P. oxalicum, various methyltransferase genes displayed different transcriptional levels. The genes Pomtr23C and Pomtr25A exhibited high transcriptional levels, while Pomtr23D/E/F/G/H and Pomtr25B were transcribed constantly at low levels. The gene deletion mutants (Δmtr23C/D/E/F/G/H and Δmtr25A/B) were constructed. Various mutants have different patterns in cellulolytic enzyme production. Compared to the WT, the largest increase in filter paper activity (FPA, indicating total cellulase activity) was observed in the Δmtr23G mutant, the only mutant with a cellulolytic halo surrounding the colony. Three mutants (Δmtr23C/D and Δmtr25A) also showed increased cellulolytic enzyme production. The Δmtr23E and Δmtr25B mutants displayed decreased FPA activity, while the Δmtr23F and Δmtr23H mutants displayed similar patterns of cellulolytic enzyme production compared with the WT. The assay of transcriptional levels of cellobiohydrolase gene Pocbh1 and β-1,4-endoglucanase Poeg1 supported that higher cellulolytic gene transcription resulted in higher production of cellulolytic enzymes, and vice versa. The transcriptional levels of two transcription factors, activator XlnR and repressor CreA, were measured. The high transcription level of the PoxlnR gene in the Δmtr23D mutant should be one reason for the increased transcription of its cellulolytic enzyme gene. Both XlnR and CreA transcriptional levels increased in the Δmtr23G mutant, but the former showed a more significant increase than the latter, indicating that the activation effect predominated. The PoMtr25A is localized in the nucleus. The catalytic subunit SNF2 of the SWI/SNF chromatin-remodeling complex was found as one of the interacting proteins of PoMtr25A via tandem affinity purification coupled with mass spectrometry. PoMtr25A may affect not only the transcription of repressor CreA but also by recruiting SWI/SNF complexes that affect chromatin structure, thereby regulating the transcription of target genes.

Gut microbial communities and their potential roles in cellulose digestion and thermal adaptation of earthworms

Adaptations to temperature and food resources, which can be affected by gut microbiota, are two main adaptive strategies allowing soil fauna to survive in their habitats, especially for cold-blooded animals. Earthworms are often referred to as ecosystem engineers because they make up the biggest component of the animal biomass found in the soil. They are considered as an important indicator in the triangle of soil quality, health and functions. However, the roles of gut microbiota in the environmental adaptation of earthworms at a large scale remain obscure. We explored the gut bacterial communities and their functions in the environmental adaptation of two widespread earthworm species (Eisenia nordenskioldi Eisen and Drawida ghilarovi Gates) in Northeast China (1661 km). Based on our findings, the alpha diversity of gut bacterial communities decreased with the increase of latitude, and the gut bacterial community composition was shaped by both mean annual temperature (MAT) and cellulose. Actinobacteria, Proteobacteria, Firmicutes, and Planctomycetes, recognized as the predominant cellulose degraders, were keystone taxa driving gut bacterial interactions. Actinobacteria, Firmicutes, and Planctomycetes were influenced by MAT and cellulose, and had higher contributions to gut total cellulase activity. The optimal temperature for total cellulase in the gut of E. nordenskioldi (25-30 °C) was lower than that of D ghilarovi (40 °C). The gut microbiota-deleted earthworms had the lowest cellulose degradation rate (1.07 %). The cellulose was degraded faster by gut bacteria from the host they were derived, indicating the presence of home field advantage of cellulose decomposition. This study provides a foundation for understanding the biotic strategies adopted by earthworms when they enter a new habitat, with gut microbiota being central to food digestion and environmental adaptability.

Production of the Cellulase Enzyme System by Locally Isolated Trichoderma and Aspergillus Species Cultivated on Banana Pseudostem during Solid-State Fermentation

The production cost of cellulases is regarded as a limiting factor in the cellulosic ethanol production chain. Trichoderma and Aspergillus species were used to produce cellulases through solid-state fermentation (SSF) utilizing banana pseudostem (BPS) as a carbon source. The production of cellulases was investigated at various substrate moisture contents (65–80%), incubation temperatures (30–40 °C), substrate pre-treatment methods (3% w/v NaOH, 5% v/v H2SO4, and water), and with different co-culturing of microorganisms. Trichoderma longibrachiatum LMLSAUL 14-1 produced the maximum total cellulase (75 FPU/g d.s), endoglucanase (11.35 U/g d.s), and β-glucosidase (235.83 U/g d.s) activities at a 75% moisture content of the untreated BPS at 30 °C in static culture. Pre-treatment of BPS improved the production of specific enzymes. Aspergillus fumigatus LMLPS 13-4 produced more β-glucosidase (259.8 U/g d.s) when grown on acid-pre-treated BPS, whereas T. harzianum LMLBP07 13-5 produced the highest β-glucosidase activity (319.5 U/g d.s) on alkali-pre-treated BPS. Co-culturing of T. harzianum LMLBP07 13-5 and A. fumigatus LMLPS 13-4 improved the production of endoglucanase. These results suggest that banana pseudostem, a waste product of the banana industry, could be a potentially cheaper and abundant substrate for the production of the cellulase enzymes.

A study on critical associations of media components on enhanced cellulase production from wild Trichoderma viride and cellulase immobilization on iron-oxide magnetic nanoparticles

Aim: The current study is a preliminary step towards enhancing the cellulase productivity in wild Trichoderma viride which will enable robust valorization of non-edible lignocellulosic biomass through co-generative enzymatic saccharification, specifically concentrating on influence of individual media components on biomass growth and cellulase productivity. Further, cellulase immobilization on iron-oxide magnetic nanoparticles was also achieved that can increase the shelf life of the enzyme. Methodology: The cellulase production in the wild Trichoderma viride was enhanced using media design and formulation. EDC {1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide} functionalized iron-oxide nanoparticles were chosen to act as carriers for cellulase immobilization. The binding efficiency and relative activity were measured in addition to optimal pH and temperature for cellulase bound iron-oxide nanoparticles. Further, the hydrolysis efficiency of immobilized cellulases was also measured after which it was subjected to consecutive hydrolytic cycles to calculate the recycle rate. Results: A maximum growth rate of 60 PCV (Packed cell volume) and total cellulase activity of 7.4 U ml-1 was obtained on media design and formulation. 82.5% binding efficiency was achieved on EDC functionalized iron-oxide magnetic nanoparticles which showed good stability at 5pH and 500C. There was 44.4% activity loss after 5 consecutive hydrolytic cycles which showed steady decline with increased cycle number and finally at the end of the 10th hydrolytic cycle, 22.2% of total relative activity was retained. Interpretation: Unprecedented total cellulase activity from a wild strain was obtained through media design. The stability of cellulases was further enhanced using iron-oxide magnetic nanoparticle immobilization. Key words: Cellulases, Immobilization, Iron-oxide magnetic nanoparticles, Submerged fermentation, Trichoderma viride

Identification of a Bidirectional Promoter from Trichoderma reesei and Its Application in Dual Gene Expression.

The cellulolytic filamentous fungus Trichoderma reesei has a strong capability in protein synthesis and secretion and is increasingly used as a fungal chassis for the production of heterologous proteins or secondary metabolites. However, bidirectional promoters that would significantly facilitate multiple genes’ expression have not been characterized in T. reesei. Herein, we show that a 767-bp intergenic region between two polyketide synthase encoding genes that were involved in the biosynthesis of the typical yellow pigment served as a bidirectional promoter in T. reesei. This region was shown to be able to drive the simultaneous expression of two fluorescence reporter genes when fused to each end. Quantitative RT-PCR analysis demonstrated that the driving strength of this bidirectional promoter from each direction reached about half of that of the commonly used promoter PgpdA. Moreover, the co-expression of two cellulase genes driven by this bidirectional promoter enabled T. reesei to produce cellulases on glucose and improved the total cellulase activities with cellulose Avicel as the carbon source. Our work identified the first bidirectional promoter in T. reesei, which would facilitate gene co-expression and find applications in synthetic biology using fungal systems.

Cellulase and Xylanase production by Ganoderma lucidum in solid-state fermentation using amazonian lignocellulosic wastes

Ganoderma lucidum is a basidiomycete whose production is of great interest due to its medicinal properties, and analyzing the potential of Amazonian lignocellulosic residues in the cultivation of G. lucidum is a way to enable the use of this material in mushroom cultivation and the production of biomolecules, including enzymes of commercial interest. The objective of this study was to evaluate the activity of cellulases and xylanases produced by G. lucidum when cultivated in açaí seed (Euterpe sp.) residues and marupá (Simarouba amara) sawdust under supplemented and non-supplemented conditions. Solid-state cultivation was carried out in 250 mL flasks containing 50 g of lignocellulosic residues, plus 2% of CaCO3, under supplemented (18% of rice, wheat and corn bran) and non-supplemented conditions. The flasks were incubated in a BOD incubator at 25 ºC. Enzyme extraction was performed from the fungus growth substrate over 30 days of cultivation, with collections every two days. The enzymatic extracts had their endoglucanase (CMCase), total cellulase (FPase) and xylanase activities determined. The highest enzymatic activity of CMCase and xylanase were 5.97 U/g and 1.90 U/g, respectively, on the 26th day of cultivation in the supplemented marupá sawdust substrate. While the maximum FPase content was 0.24 U/g, which was observed in the 8th day for the supplemented açaí substrate. Thus, the tested residues proved to be promising for the mycelial development of G. lucidum, with the supplementation with corn, rice and wheat bran being favorable to the production of enzymes with biotechnological interest.

Production and properties of Irpex lacteus cellulase and xylanase

This study aimed to characterize the cellulolytic system of Irpex lacteus BCC 104 isolated from the forest of Georgia. The results obtained showed that the expression of endoglucanase and xylanase synthesis in this fungus is inducible in the presence of cellulose-containing growth substrates. Among them, crystalline cellulose appeared to be the best carbon source providing the highest endoglucanase (48.0 U/mL), total cellulase (7.6 U/mL), and xylanase (68.4 U/mL) activities. Supplementation of the Avicel-induced culture with 0.5 or 1% glycerol caused catabolite repression of the cellulase and xylanase formation by I.lacteus BCC 104. The enzyme synthesis resumed only after the depletion of easily metabolizable carbon source from the medium. The crude enzyme obtained after the cultivation of I. lacteus in the fermenter and used for saccharification of pretreated wheat straw (40 mg/mL, FPA load 20 U/g substrate) yielded 10.7 mg reducing sugars/mL (32.2% from the theoretically possible). CMCase and xylanase were purified 31- and 34-fold and characterized. The optimum pH for both enzyme activities was found to be 5.0; at pH 5-6, they retained 60% of their initial activity after incubation at 30°C for 48 h. CMCase expressed maximum activity at 50°C while xylanase – at 55°C. Half-inactivation periods of endoglucanase and xylanase during incubation at 60°C were 60 min and 150 min, respectively.

Regulation of Schizophyllum commune BCC632 cellulase and xylanase activities

The polysaccharide-hydrolyzing enzymes of Schizophyllum commune BCC 632 have been analyzed in submerged fermentation conditions to elucidate the effect of chemically and structurally different carbon sources on the expression of cellulases and xylanase. Production of cellulase and xylanase by S. commune BCC 632 requires the presence of inducing substrate in the culture medium. Crystalline cellulose appeared to be the best carbon source providing the highest endoglucanase (53.5 U/mL) and total cellulase (9.2 U/mL) activities. In particular, this fungus appeared to be a super-producer of xylanase (636.1 U/mL). Among complex lignocellulosic substrates, mandarin pomace provided high enzyme activity whereas wheat straw appeared to be a poor substrate for both cellulases and xylanase secretion. However, the supplementation of wheat straw-based medium with mandarin pomace or glycerol resulted in a significant secretion of cellulase and xylanase by the fungus. Nevertheless, the addition of 0.5% glucose to the Avicel-containing medium caused short-term catabolite repression of the synthesis of both cellulase and xylanase while the addition of α-deoxy-D-glucose completely prevented enzyme secretion. Extracellular protein electrophoresis and inhibitory assay using cycloheximide and actinomycin D has shown that both cellulase and xylanase are inducible enzymes and subject to catabolite repression by readily metabolizable sugars.

Enhanced Cellulase Production by Talaromyces amestolkiae CMIAT055 Using Banana Pseudostem

Cellulases are a complex of enzymes necessary for the complete solubilization of cellulose in sugars, thus playing a key role in the natural carbon cycle through the hydrolysis of lignocellulosic structures. The aim of this study was to evaluate the increase in the capacity of Talaromyces amestolkiae CMIAT 055 to produce cellulases by optimizing the components of the culture medium containing banana pseudostem as an inducer, as well as in different agitation configurations in a bioreactor. Optimization was performed through statistical experimental design (Plackett–Burman and DCCR), a study of pH control in bioreactors, and a study of the agitation system by comparing impellers with different flow profiles in the liquid medium. For this purpose, a wild strain of Talaromyces amestolkiae CMIAT 055 was used. In the Plackett–Burman and DCCR statistical design, four components of the culture medium were significant and optimized for greater synthesis of FPase: banana pseudostem, CaCl2, KH2PO4, and urea. In bioreactors tests, these parameters were beneficial for greater enzyme activities: maintenance of pH at 5.0, use of Pitched blade impeller, and rotation speed at 300 rpm. Comparing the first test using banana pseudostem in an Erlenmeyer flask to the last fermentation process in bioreactors, it was observed that the total cellulase activity increased from 424.7 to 2172.8 FPU/L. This fact showed that the strategies adopted in this study are a pertinent way to reduce the cost of enzyme production through the use of lignocellulosic materials.Graphical Abstract

Development of a powerful synthetic hybrid promoter to improve the cellulase system of Trichoderma reesei for efficient saccharification of corncob residues

BackgroundThe filamentous fungus Trichoderma reesei is a widely used workhorse for cellulase production in industry due to its prominent secretion capacity of extracellular cellulolytic enzymes. However, some key components are not always sufficient in this cellulase cocktail, making the conversion of cellulose-based biomass costly on the industrial scale. Development of strong and efficient promoters would enable cellulase cocktail to be optimized for bioconversion of biomass.ResultsIn this study, a synthetic hybrid promoter was constructed and applied to optimize the cellulolytic system of T. reesei for efficient saccharification towards corncob residues. Firstly, a series of 5’ truncated promoters in different lengths were established based on the strong constitutive promoter Pcdna1. The strongest promoter amongst them was Pcdna1-3 (− 640 to − 1 bp upstream of the translation initiation codon ATG), exhibiting a 1.4-fold higher activity than that of the native cdna1 promoter. Meanwhile, the activation region (− 821 to − 622 bp upstream of the translation initiation codon ATG and devoid of the Cre1-binding sites) of the strong inducible promoter Pcbh1 was cloned and identified to be an amplifier in initiating gene expression. Finally, this activation region was fused to the strongest promoter Pcdna1-3, generating the novel synthetic hybrid promoter Pcc. This engineered promoter Pcc drove strong gene expression by displaying 1.6- and 1.8-fold stronger fluorescence intensity than Pcbh1 and Pcdna1 under the inducible condition using egfp as the reporter gene, respectively. Furthermore, Pcc was applied to overexpress the Aspergillus niger β-glucosidase BGLA coding gene bglA and the native endoglucanase EG2 coding gene eg2, achieving 43.5-fold BGL activity and 1.2-fold EG activity increase, respectively. Ultimately, to overcome the defects of the native cellulase system in T. reesei, the bglA and eg2 were co-overexpressed under the control of Pcc promoter. The bglA-eg2 double expression strain QPEB70 exhibited a 178% increase in total cellulase activity, whose cellulase system displayed 2.3- and 2.4-fold higher saccharification efficiency towards acid-pretreated and delignified corncob residues than the parental strain, respectively.ConclusionsThe synthetic hybrid promoter Pcc was generated and employed to improve the cellulase system of T. reesei by expressing specific components. Therefore, construction of synthetic hybrid promoters would allow particular cellulase genes to be expressed at desired levels, which is a viable strategy to optimize the cellulolytic enzyme system for efficient biomass bioconversion.

Isolation and Identification of Cellulase Producing and Sugar Fermenting Bacteria for Second-Generation Bioethanol Production

Over the last decades, the negative impacts of fossil fuel on the environment and increasing demand for energy due to the unavoidable depletion of fossil fuels, has transformed the world’s interests towards alternative fuels. In particular, bioethanol production from cellulosic biomass for the transportation sector has been incrementing since the last decade. The bacterial pathway for bioethanol production is a relatively novel concept and the present study focused on the isolation of potential “cellulase-producing” bacteria from cow dung, compost soil, and termite gut and isolating sugar fermenting bacteria from palm wine. To select potential candidates for cellulase enzyme production, primary and secondary assays were conducted using the Gram’s iodine stain in Carboxy Methyl Cellulose (CMC) medium and the Dinitrosalicylic acid (DNS) assays, respectively. Durham tube assay and Solid-Phase Micro-Extraction (SPME) coupled with Gas Chromatography-Mass Spectrometry (GC-MS) was used to evaluate the sugar fermenting efficiency of the isolated bacteria. Out of 48 bacterial isolates, 27 showed cellulase activity where Nocardiopsis sp. (S-6) demonstrated the highest extracellular crude enzyme activity of endoglucanase (1.56±0.021 U) and total cellulase activity (0.93±0.012 U). The second-highest extracellular crude enzyme activity of endoglucanase (0.21±0.021 U) and total cellulase activity (0.35±0.021 U) was recorded by Bacillus sp. (T-4). Out of a total of 8 bacterial isolates, Achromobacter sp. (PW-7) was positive for sugar fermentation resulting in 3.07% of ethanol in broth medium at 48 h incubation. The results of the study revealed that Nocardiopsis sp. (S-6) had the highest cellulase enzyme activity. However, the highest ethanol percentage was achieved with by having both Bacillus sp. (T-4) and Achromobacter sp. (PW-7) for the simultaneous saccharification and fermentation (SSF) method, as compared to separate hydrolysis and fermentation (SHF) methodologies.

The Carbon Source Controls the Secretion and Yield of Polysaccharide-Hydrolyzing Enzymes of Basidiomycetes.

In the present study, the polysaccharide-hydrolyzing secretomes of Irpex lacteus (Fr.) Fr. (1828) BCC104, Pycnoporus coccineus (Fr.) Bondartsev and Singer (1941) BCC310, and Schizophyllum commune Fr. (1815) BCC632 were analyzed in submerged fermentation conditions to elucidate the effect of chemically and structurally different carbon sources on the expression of cellulases and xylanase. Among polymeric substrates, crystalline cellulose appeared to be the best carbon source providing the highest endoglucanase, total cellulase, and xylanase activities. Mandarin pomace as a growth substrate for S. commune allowed to achieve comparatively high volumetric activities of all target enzymes while wheat straw induced a significant secretion of cellulase and xylanase activities of I. lacteus and P. coccineus. An additive effect on the secretion of cellulases and xylanases by the tested fungi was observed when crystalline cellulose was combined with mandarin pomace. In I. lacteus the cellulase and xylanase production is inducible in the presence of cellulose-rich substrates but is suppressed in the presence of an excess of easily metabolizable carbon source. These enzymes are expressed in a coordinated manner under all conditions studied. It was shown that the substitution of glucose in the inoculum medium with Avicel provides accelerated enzyme production by I. lacteus and higher cellulase and xylanase activities of the fungus. These results add new knowledge to the physiology of basidiomycetes to improve cellulase production.

Use of green light to improve the production of lignocellulose-decay enzymes by Pleurotus spp. in liquid cultivation

The influence of green light on mycelium biomass growth and extracellular enzyme activities of edible mushrooms from the Pleurotus genus, which is popularly cultivated all over the world, were investigated. The mycelium of seven strains of five species of Pleurotus (P. citrinopileatus, P. djamor, P. eryngii, P. ostreatus, and P. pulmonarius) was grown in liquid medium at 28 °C in the dark or under green light (515−530 nm). The light source was light-emitting diodes (LED) with photon flux density adjusted to 20 μmol m−2 s-1 that was kept on throughout the cultivation period. After 12 days of growth, the mycelium was recovered and used for biomass determination and the cultivation medium was used to total cellulase, endoglucanase, xylanase, and laccase activities determination. Green light reduced the mycelial biomass growth of Pleurotus spp. but increased the cellulolytic and xylanolytic activities. The cellulolytic activity of most strains increased in the presence of green light with increases ranging from 1.5 times (P. ostreatus endoglucanase) to 8 times (P. citrinopileatus total cellulase and endoglucanase). Green light reduced laccase activity for most strains with the greatest reduction for P. eryngii (2.2 times lower). The specific enzymatic activity of cellulase and endoglucanase from P. citrinopileatus, increased by 31 times and 30 times, respectively, compared to the dark. Also, the specific laccase and xylanase activities of P. pulmonarius increased 4.4 times and 6.8 times, respectively, under green light. The use of light at particular wavelengths can be a viable strategy to increase the production of enzymes for different biotechnological applications and species of Pleurotus are particularly interesting for this purpose.

Effects of the Transcription Factor Ace2 from Trichoderma reesei on Cellulase and Hemicellulase Expression in Trichoderma orientalis EU7-22.

Trichoderma orientalis (T. orientalis) EU7-22 has a complete cellulase system and shows a remarkable enzyme activity with high potential in the industry. Ace2 is an important transcriptional factor for cellulase and hemicellulase expression in Trichoderma reesei (T. reesei). However, the ace2 gene cannot be found in the genome of T. orientalis. Researches show that the mechanism of cellulase transcriptional regulation in T. orientalis keeps high similarity with T. reesei up till now. So, in this study, the ace2 of Trichoderma reesei QM9414 was heterologous expressed in T. orientalis EU7-22. As a result, xylanase activity and β-glucosidase activity of ace2 heterogeneous expression strains are improved and total cellulase activity is decreased. The result of qPCR is in accordance with enzyme activities. This study provides a reference for an in-depth study on transcriptional regulation mechanisms of T. orientalis.

Evaluation of the production of cellulases by Penicillium sp. FSDE15 using corncob and wheat bran as substrates

This study aimed to evaluate the production of cellulases by Penicillium sp. FSDE15, from agroindustrial waste. These enzymes are able to convert part of the lignocellulosic biomass into fermentable sugars. Through fermentative assays, it was possible to study the enzymatic kinetics over 240 h. In the tests, moisture content remained between 50 and 70%, while pH varied widely, from 3 to 8. In addition, maximum CMCase (endoglucanase) and FPase (total cellulase) activities of 21.11 + 0.74 U/g and 1.29 + 0.27 U/g, respectively, were achieved in 216 h of fermentation, for cultivation carried out with wheat bran and corncob in the proportion of 50% as substrates. The highest β-glucosidase activity was 8.72 + 0.42 U/g in 216 h for cultivation using wheat bran only. These results demonstrate that Penicillium sp. FSDE15 is a good cellulase producer and also that corncob can be used effectively as a substrate for the production of enzymatic cocktails.