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Multifunctional endoglucanase, RfGH5_4 from Ruminococcus flavefaciens showed (β/α)

Ionic liquids (ILs) have been widely used in the pretreatment of biomass. However, the effects of residual ILs on the enzymolysis and fermentation of biomass are still unknown. Therefore, a large quantity of water-washing is usually followed after biomass pretreatment to eliminate the inhibition of residual ILs on subsequent hydrolysis and fermentation steps. In this work, the effect of choline glycine ([Ch][Gly]) concentration on the activity of cellulase and Bacillus sp. strain P38 was systematically investigated to explore the impacts of residual ILs on enzymolysis and fermentation. The results confirmed that the activities of them were almost not inhibited in low concentrations (less than 0.5 wt%) of [Ch][Gly]. Under optimal pretreatment conditions, the maximum cellulose digestibility was 99.23%. Enzymatic hydrolysate was suitable for l -lactic acid fermentation without appreciable inhibition, and the highest sugar-acid conversion rate of 96.33% was obtained by simplified detoxification. This work provides an economic route to produce fermentable sugar and l -lactic acid, which shows an industrial application prospect in lignocellulosic biorefinery.

Enzyme-mediated isolation of cellulose nanocrystals (CNCs) is a promising environment friendly method with expected lower capital and operating expenditures compared to traditional processes. However, it is still poorly understood. In this study, an endoxylanase was applied as accessory enzyme to assess its potential to increase the selectivity of an endoglucanase during cellulose hydrolysis to isolate CNCs with improved properties. Only combinations of the enzymes with xylanase activity equal to or higher than the endoglucanase activity resulted in CNCs with improved properties (i.e., crystallinity, thermostability, uniformity, suspension stability and aspect ratio). The beneficial effects of the accessory enzyme are related to its hydrolytic (xylan and cellulose hydrolysis) and non-hydrolytic action (swelling of cellulose fibers and fiber porosity) and on the ratio of the enzymes, which in turn allows to tailor the properties of the CNCs. In conclusion, compared to the traditional sulfuric acid hydrolysis method, accessory enzymes help to isolate cellulose nanomaterials with improved and customized (sizes, aspect ratio and morphology) properties that may allow for new applications.

Polar soils under different vegetation cover from Livingston Island (Antarctica) were studied analyzing indicators of carbohydrates decomposition in soils: organic carbon, biomass carbon, total nitrogen, C:N ratio, cellulase, amylase, and invertase activity. The highest values of microbial biomass in soils were indicated for sites with vegetation cover while the lowest values in soils without vegetation, which correlate with the content of total organic carbon and the C:N ratio. The highest percentage of biomass carbon compared to the total organic biomass carbon is obtained for two sites with mosses, and the lowest in sites without vegetation, followed by the sites with lichens. Cellulase activity is highest in polar soils with moss cover. Amylase activity depends more strongly on the type of vegetation. The highest amylase activity is detected in soils under algae cover and the lowest in soils without vegetation. Invertase activity is limited by the extreme soil and climatic conditions of Antarctica. A very strong, positive correlation is found between total carbon and total nitrogen. The relationship between total carbon and the C:N ratio, as well as between biomass carbon and amylase activity, is moderate, and positive. The higher dependence of amylase activity from organic carbon with microbial origin correlates with higher values of the enzyme amylase compared to the enzyme cellulase. There is a strong (cellulase) and very strong (amylase) positive relationship between the activity of enzymes and the combination of factors: total carbon, total nitrogen, C: N ratio and biomass carbon.

This study aimed to reveal the roles of the protein kinase A catalytic subunit 1 (pkac1) and carbon catabolite repressor cre1 genes in cellulase production by Trichoderma reesei wild-type strain QM6a. Our strategy might be useful to construct a high-yielding cellulase strain for its wide application.This paper describes cellulase activity, plate conidiation, and yellow pigment synthesis assays of QM6a with the disruption of pkac1 and cre1.Deletion of pkac1 (Δpkac1) had no effect on cellulase production or transcript levels of major cellulase genes in the presence of cellulose. Disruption of cre1 (Δcre1) resulted in a remarkable increase in cellulase production and expression of the four major cellulase genes. Double disruption of pkac1 and cre1 significantly improved enzyme activity and protein production. The double disruption also resulted in a significant reduction in yellow pigment production and abrogated conidial production.Double deletion of pkac1 and cre1 led to increased hydrolytic enzyme production in T. reesei using cellulose as a carbon source.

Human industrial activities have caused environmental uranium (U) pollution, resulting in uranium(VI) had radiotoxicity and chemical toxicity. Here, a cellulase-producing Penicillium fungus was screened and characterized by X-ray fluorescence (XRF), and Fourier transform infrared reflection (FT-IR), as well as by GC/MS metabolomics analysis, to study the response to uranium(VI) stress. The biomass of Penicillium decreased after exposure to 100 mg/L U. Uranium combined with carboxyl groups, amino groups, and phosphate groups to form uranium mineralized deposits on the surface of this fungal strain. The α-activity concentration of uranium in the strain was 2.57×10

One crucial step in processing the recalcitrant lignocellulosic biomass is the fast hydrolysis of natural cellulose to fermentable sugars that can be subsequently converted to biofuels and bio-based chemicals. Recent studies have shown that lytic polysaccharide monooxygenase (LPMOs) with auxiliary activity family 9 (AA9) are capable of efficiently depolymerizing the crystalline cellulose via regioselective oxidation reaction. Intriguingly, the catalysis by AA9 LPMOs requires reductant to provide electrons, and lignin and its phenolic derivatives can be oxidized, releasing reductant to activate the reaction. The activity of AA9 LPMOs can be enhanced by in-situ generation of H

This study presents a potential approach to enhance integrated sequential biohydrogen production from waste biomass using magnetite nanoparticle (Fe3O4 NPs) which is synthesized through waste seeds of Syzygium cumini. Consequences of 0.5% Fe3O4 NPs have been investigated on the thermal and pH stability of fungal crude cellulase. It is noticed that Fe3O4 NPs treated enzyme and control exhibits 100% activity in the temperature range of 45–60 °C and 45–55 °C, respectively. Moreover, Fe3O4 NPs treated enzyme showed extended thermal stability in the temperature range of 50–60 °C up to 12 h. Beside this, Fe3O4 NPs treated enzyme possesses 100% stability in the pH range of 5.0–7.0 whereas control exhibited only at pH 6.0. Enzymatic hydrolysis via Fe3O4 NPs treated enzyme has been employed which produces ∼68.0 g/L reducing sugars from sugarcane bagasse. Subsequently, sugar hydrolyzate has been utilized as substrate in the sequential integrated fermentation that produces ∼3427.0 mL/L cumulative hydrogen after 408 h. This approach may have potential for the pilot scale production of biohydrogen from waste biomass at low-cost in an eco-friendly manner.

Endophytic fungi in plant tissues produce a wide range of secondary metabolites and enzymes, which exhibit a variety of biological activities. In the present study, litter endophytic fungi were isolated from a fire-prone forest and screened for thermostable cellulases. Among nine endophytic fungi tested, two isolates, Bartalinia pondoensis and Phoma sp., showed the maximum cellulase activity. Bartalinia pondoensis was further selected for its cellulase production and characterization. Among the carbon and nitrogen sources tested, maximum cellulase production was observed with maltose and yeast extract, and the eucalyptus leaves and rice bran served as the best natural substrates. The cellulase activity increased with increasing temperature, with maximum activity recorded at 100 °C. The maximum CMCase activity was observed between pH 6.0 and 7.0 and retained 80% of its activity in the pH range of 8-10. Partially purified cellulase of B. pondoensis retained 50% of its activity after 2 h of incubation at 60 °C, 80 °C and 100 °C. These results suggest that litter endophytic fungus B. pondoensis is a potential source for the production of thermostable and alkali-tolerant cellulase.

In this study, indigo-dyed denim fabric was decolorized by washing and printing with separate and simultaneous applications of laccase enzyme, sodium hydrosulfite, and cellulase enzyme. In this regard, the surface reflectance and color coordinates of the discolored fabrics were analyzed, and SEM photographs of the treated fabrics were prepared to analyze their surfaces. Finally, the effects of the discoloration process and materials on various parameters of the treated samples were investigated, including moisture content, creaserecovery angle, air permeability, and abrasion resistance. The color experiments showed that the discoloration mechanism with the combined use of laccase enzyme, sodium hydrosulfite, and cellulose enzyme had a significant effect on the improvement of the lightness (L*) of the samples, as the lightness of the treated samples was improved by 101.18 percent and 55.79 percent in both printing and washing, respectively. Furthermore, examination of specimen color coordinates revealed that the hue of the treated samples was changed to blue and green, and the purity of color (C*) was improved. The increased moisture content and air permeability of the treated specimens suggested that the comfort of the jeans clothing provided by these treatment methods had improved. As a result, it should be noted that the mediating action of sodium hydrosulfite was significantly influential for discoloration of denim with the laccase enzyme.

Black rice contains a variety of bioactive substances that contribute to the high nutritional value of black rice wine (BRW). However, the dense bran layer of black rice retards the fermentation rate and reduces the dissolution of active components. Hence, this study aims to investigate the effects of pre-gelatinization (PG) before cooking and cellulase (CE) addition during fermentation on the fermentation performance of BRW and its antioxidant activity.PG combined with CE treatments (PGCE) increases the alcohol content, free amino acid content, volatile flavor content and total antioxidant activity of BRW by 90.81%, 15.36%, 38.05% and 19.56%, respectively, compared with the control group. Scanning electron microscopy, low-field nuclear magnetic resonance and texture properties analysis indicate that PG treatment increases gelatinization degree of starch during cooking, decreases bound water content in cooked black rice and promotes unbound water release. CE destroys the aleurone layer structure, facilitates the release of unbound water and the exposure of rice starch, thus increasing the reaction area and extravasation content significantly, which is beneficial to microbial growth and fermentation. Incomplete aleurone layer also promotes the dissolution of anthocyanins, phenols and other active substances, increasing the antioxidant activities of BRW.PG and CE treatments reduce the fermentation time and improve the quality of BRW by destroying the black rice structure. © 2022 Society of Chemical Industry.

The genus Aloe (family: Xanthorrhoeaceae) encompasses 490 shrubby-succulent-perennial species which are native to Africa. Among these, Aloe vera has grabbed the limelight in the arena of herbal medicine due to the presence of unique phytochemicals such as Aloin A and B, Homonataloin, Aloe emodin, acemannan etc. in its latex. Moreover, its leaf extract contains 99% water, and 75 active compounds including vitamins (A, B1, B2, B3, B6, B9, B12, C, and E), minerals (Zn, Se, Na, Mn, Mg, K, Cu, Cr, Ca), amino acids, and enzymes (peroxidase, lipase, cellulase, catalase, carboxypeptidase, bradykinase, amylase, alkaline phosphatase). Thus, Aloe vera is revered as a ‘wonder plant’ as it possesses multiple pharmaceutical properties such as anti-oxidant, anti-inflammatory, anti-tumor, anti-fungal, anti-diabetic, anti-ulcer, anti-hypertensive and its juice helps to cure gastrointestinal disorders, and dermal diseases, it also strengthens the immune system. Being a xerophyte, Aloe vera is commercially cultivated in many countries which include South Africa, America, Australia, India etc. The Aloe gel, is meticulously extracted, processed and stabilized before using it in preparation of various health care products. This review focuses on the explicit information on origin, botanical description, phytochemistry, nutritional and medicinal benefits, with special emphasis on value-added products of Aloe vera. The review revealed 46 value-added Aloe-products that are being manufactured by different companies worldwide. Thus, sustainable Aloe-cultivation must be ensured to meet the rising global demands for Aloe vera gel and allied products.

The characterization of bacteria with hydrolytic potential significantly contributes to the industries. Six cellulose-degrading bacteria were isolated from mixture soil samples collected at Kingfisher Lake and the University of Manitoba campus by Congo red method using carboxymethyl cellulose agar medium and identified as Paenarthrobacter sp. MKAL1, Hymenobacter sp. MKAL2, Mycobacterium sp. MKAL3, Stenotrophomonas sp. MKAL4, Chryseobacterium sp. MKAL5, and Bacillus sp. MKAL6. Their cellulase production was optimized by controlling different environmental and nutritional factors such as pH, temperature, incubation period, substrate concentration, nitrogen, and carbon sources using the dinitrosalicylic acid and response surface methods. Except for Paenarthrobacter sp. MKAL1, all strains are motile. Only Bacillus sp. MKAL6 was non-salt-tolerant and showed gelatinase activity. Sucrose enhanced higher cellulase activity of 78.87 ± 4.71 to 190.30 ± 6.42 U/mL in these strains at their optimum pH (5-6) and temperature (35-40 °C). The molecular weights of these cellulases were about 25 kDa. These bacterial strains could be promising biocatalysts for converting cellulose into glucose for industrial purposes.

The genomes of two Penicillium strains were sequenced and studied in this study: strain 2HH was isolated from the digestive tract of Anobium punctatum beetle larva in 1979 and the cellulase hypersecretory strain S1M29, derived from strain 2HH by a long-term mutagenesis process. With these data, the strains were reclassified and insight is obtained on molecular features related to cellulase hyperproduction and the albino phenotype of the mutant. Both strains were previously identified as Penicillium echinulatum and this investigation indicated that these should be reclassified. Phylogenetic and phenotype data showed that these strains represent a new Penicillium species in series Oxalica, for which the name Penicillium ucsense is proposed here. Six additional strains (SFC101850, SFCP10873, SFCP10886, SFCP10931, SFCP10932 and SFCP10933) collected from the marine environment in the Republic of Korea were also classified as this species, indicating a worldwide distribution of this new taxon. Compared to the closely related strain Penicillium oxalicum 114-2, the composition of cell wall-associated proteins of P. ucsense 2HH shows five fewer chitinases, considerable differences in the number of proteins related to β-D-glucan metabolism. The genomic comparison of 2HH and S1M29 highlighted single amino-acid substitutions in two major proteins (BGL2 and FlbA) that can be associated with the hyperproduction of cellulases. The study of melanin pathways shows that the S1M29 albino phenotype resulted from a single amino-acid substitution in the enzyme ALB1, a precursor of the 1,8-dihydroxynaphthalene (DHN)-melanin biosynthesis. Our study provides important knowledge towards understanding species distribution, molecular mechanisms, melanin production and cell wall biosynthesis of this new Penicillium species.

In this research, a magnetic reusable nickel nanoparticle (NiNPs) supporting materials were prepared for cellulase enzyme immobilization. The immobilized cellulase showed high activity recovery, largefast immobilization capacity and improved pHtemperature tolerance. The excellent stability and reusability enabled the immobilized cellulase to retain 84% of its initial activity after ten cycles. At 2 mg/mL enzyme concentration, highest 93% immobilization efficiency was achieved within two hours of immobilization. When the treatment temperature reached 40 °C and pH 5, the immobilized cellulase exhibited highest residual activity. The immobilized cellulase could be separated from the solution by a magnetic force. This study introduced a novel supporting material for cellulase immobilization, and the immobilized cellulase poses a great potential in the hydrolysis of lignocellulosic biomass which can used as an easily applicable and sustainable pre-treatment step for advanced biofuel production.

Bacterial cellulase enzymes are potent candidates for the efficient production of bioethanol, a promising alternative to fossil fuels, from cellulosic biomass. These enzymes catalyze the breakdown of cellulose in plant biomass into simple sugars and then to bioethanol. In the absence of the enzyme, the cellulosic biomass is recalcitrant to decomposition due to fermentation-resistant lignin and pectin coatings on the cellulose surface, which make them inaccessible for hydrolysis. Cellobiohydrolase CelS is a microbial enzyme that binds to cellulose fiber and efficiently cleaves it into a simple sugar (cellobiose) by a repeated processive chopping mechanism. The two contributing factors to the catalytic reaction rate and the yield of cellobiose are the efficient product expulsion from the product binding site of CelS and the movement of the substrate or cellulose chain into the active site. Despite progress in understanding product expulsion in other cellulases, much remains to be understood about the molecular mechanism of processive action of these enzymes. Here, nonequilibrium molecular dynamics simulations using suitable reaction coordinates are carried out to investigate the energetics and mechanism of the substrate dynamics and product expulsion in CelS. The calculated free energy barrier for the product expulsion is three times lower than that for the processive action indicating that product removal is relatively easier and faster than the sliding of the substrate to the catalytic active site. The water traffic near the active site in response to the product expulsion and the processive action is also explored.

Background The filamentous fungus Trichoderma reesei has been used as a host organism for the production of lignocellulosic biomass-degrading enzymes. Although this microorganism has high potential for protein production, it has not yet been widely used for heterologous recombinant protein production. Transcriptional induction of the cellulase genes is essential for high-level protein production in T. reesei; however, glucose represses this transcriptional induction. Therefore, cellulose is commonly used as a carbon source for providing its degraded sugars such as cellobiose, which act as inducers to activate the strong promoters of the major cellulase (cellobiohydrolase 1 and 2 (cbh1 and cbh2) genes. However, replacement of cbh1 and/or cbh2 with a gene encoding the protein of interest (POI) for high productivity and occupancy of recombinant proteins remarkably impairs the ability to release soluble inducers from cellulose, consequently reducing the production of POI. To overcome this challenge, we first used an inducer-free biomass-degrading enzyme expression system, previously developed to produce cellulases and hemicellulases using glucose as the sole carbon source, for recombinant protein production using T. reesei. Results We chose endogenous secretory enzymes and heterologous camelid small antibodies (nanobody) as model proteins. By using the inducer-free strain as a parent, replacement of cbh1 with genes encoding two intrinsic enzymes (aspartic protease and glucoamylase) and three different nanobodies (1ZVH, caplacizumab, and ozoralizumab) resulted in their high secretory productions using glucose medium without inducers such as cellulose. Based on signal sequences (carrier polypeptides) and protease inhibitors, additional replacement of cbh2 with the nanobody gene increased the percentage of POI to about 20% of total secreted proteins in T. reesei. This allowed the production of caplacizumab, a bivalent nanobody, to be increased to 9.49-fold (508 mg/L) compared to the initial inducer-free strain. Conclusions In general, whereas the replacement of major cellulase genes leads to extreme decrease in the degradation capacity of cellulose, our inducer-free system enabled it and achieved high secretory production of POI with increased occupancy in glucose medium. This system would be a novel platform for heterologous recombinant protein production in T. reesei.

Protein adsorption to solid carbohydrate interfaces is critical to many biological processes, particularly in biomass deconstruction. To engineer more-efficient enzymes for biomass deconstruction into sugars, it is necessary to characterize the complex protein–carbohydrate interfacial interactions. A carbohydrate-binding module (CBM) is often associated with microbial surface-tethered cellulosomes or secreted cellulase enzymes to enhance substrate accessibility. However, it is not well known how CBMs recognize, bind, and dissociate from polysaccharides to facilitate efficient cellulolytic activity, due to the lack of mechanistic understanding and a suitable toolkit to study CBM–substrate interactions. Our work outlines a general approach to study the unbinding behavior of CBMs from polysaccharide surfaces using a highly multiplexed single-molecule force spectroscopy assay. Here, we apply acoustic force spectroscopy (AFS) to probe a Clostridium thermocellum cellulosomal scaffoldin protein (CBM3a) and measure its dissociation from nanocellulose surfaces at physiologically relevant, low force loading rates. An automated microfluidic setup and method for uniform deposition of insoluble polysaccharides on the AFS chip surfaces are demonstrated. The rupture forces of wild-type CBM3a, and its Y67A mutant, unbinding from nanocellulose surfaces suggests distinct multimodal CBM binding conformations, with structural mechanisms further explored using molecular dynamics simulations. Applying classical dynamic force spectroscopy theory, the single-molecule unbinding rate at zero force is extrapolated and found to agree with bulk equilibrium unbinding rates estimated independently using quartz crystal microbalance with dissipation monitoring. However, our results also highlight critical limitations of applying classical theory to explain the highly multivalent binding interactions for cellulose–CBM bond rupture forces exceeding 15 pN.

Anthracnose disease caused by Colletotrichum lindemuthianum (Sacc. and Magnus) Lams- -Scrib is one of the most devastating seed-borne diseases of common bean (Phaseolus vulgaris L.). In the present study, we evaluated the antifungal activity of Bunium persicum essential oil (EO) and its main constituents on mycelial growth, sporulation and spore germination inhibition of C. lindemuthianum. The main objective of this study was to investigate the effect of EO and its main constituents on decreasing the activity of cell wall degrading enzymes (CWDEs) produced by C. lindemuthianum, which are associated with disease progress. Also, the effects of seed treatment and foliar application of EO and its main constituent, cuminaldehyde, on anthracnose disease severity was investigated. The essential oil of B. persicum, was obtained by using a clevenger apparatus and its major constituents were identified by gas chromatography-mass spectrometry (GC-MS). The EO was characterized by the presence of major compounds such as cuminaldehyde (37.7%), γ-terpinene (17.1%) and β-pinene (15.4%), which indicated antifungal effects against C. lindemuthianum. This pathogen did not grow in the presence of EO, cuminaldehyde and γ-terpinene, β-pinene at 1,500; 1,010 and 1,835 ppm concentrations, respectively. Also, sporulation and spore germination of C. lindemuthianum was completely inhibited by EO and cuminaldehyde. Synergistic effects of the main constituents showed that combing γ-terpinene with cuminaldehyde induced a synergistic activity against C. lindemuthianum and in combination with β-pinene caused an additive effect. Activities of pectinase, cellulase and xylanase, as main CWDEs, were decreased by EO and its main constituents at low concentration without affecting mycelial growth. Seed treatment and foliar application of peppermint EO and/or cuminaldehyde significantly reduced the development of bean anthracnose. We introduced B. persicum EO and constituents, cuminaldehyde and γ-terpinene, as possible control agents for bean anthracnose.

EG5C-1, processive endoglucanase from Bacillus subtilis, is a typical bifunctional cellulase with endoglucanase and exoglucanase activities. The engineering of processive endoglucanase focuses on the catalytic pocket or carbohydrate-binding module tailoring based on sequence/structure information. Herein, a computational strategy was applied to identify the desired mutants in the enzyme molecule by evolutionary-coupling analysis; subsequently, four residue pairs were selected as evolutionary mutational hotspots. Based on iterative-saturation mutagenesis and subsequent enzymatic activity analysis, a superior mutant K51T/L93T has been identified away from the active center. This variant had increased specific activity from 4170 U/µmol of wild-type (WT) to 5678 U/µmol towards carboxymethyl cellulose-Na and an increase towards the substrate Avicel from 320 U/µmol in WT to 521 U/µmol. In addition, kinetic measurements suggested that superior mutant K51T/L93T had a high substrate affinity (K

Lignocellulosic materials are composed of three main structural polymers: hemicellulose, cellulose, and lignin. Cellulose is a long chain molecule of glucose requiring a small number of enzymes for degradation due to its simple structure while lignin is a complex polymer of phenylpropane making its biochemical decomposition difficult. Under anaerobic conditions, lignocellulose breakdown is much easier and more rapid than aerobic conditions. Various studies have been carried out to estimate the rate of degradation of lignocellulosic materials. Microorganisms play a key role in the degradation of lignocellulosic materials because they produce a variety of hydrolytic enzymes including cellulase, proteases, xylanases, lipases, laccase, and phosphatases during the degradation of lignocellulosic materials. Based on the body of literature, microorganismal activity can provide useful information about the process of organic matter decomposition.

Aims Optimize the production of Aspergillus brasiliensis endoglucanase in a solid-phase bioprocess using cupuaçu shell as substrate. Methods and Results The shells were supplemented with nitrogen and phosphorous and used as a substrate. The centesimal and inorganic composition of the residue was determined, and found to be rich in fibres, and possessed essential elements for fungal growth. In the initial cultivation of A. brasiliensis, endoglucanase activity of 7.35 U g−1 was obtained. A factorial experimental design was used to determine the most significant variables for the bioprocess. The interactions between moisture, temperature and nitrogen source were noteworthy (p < 0.05). From the rotational central composite design, the optimization of temperature and nitrogen supplementation was obtained, and this reached 40.50 U g−1, which is an increase of more than five times the value obtained initially. The enzymatic extract was applied as the biocatalyst in the hydrolysis of cupuaçu shells and, after 48 h, it was possible to observe the production of reducing sugars. Conclusions Cupuaçu shell can be used as a substrate for endoglucanase production by A. brasiliensis. The process was optimized for the cultivation temperature and the nitrogen source. The enzymatic extract can be applied in the hydrolysis of lignocellulosic biomass. Significance and Impact of the Study Cupuaçu shells can be used to produce cellulases, a product of high added value that can generate economic and environmental benefits for communities and companies producing derivatives of the cupuaçu fruit.

Resistance against antibiotics mainly due to their misuse and overuse is an emerging health issue, worldwide. Antibiotics release active antibiotic residues in the environment during their production. Bacteria encounter these active antibiotic residues and the genes present in them; resultantly, they acquire resistance against antibiotics. The current study was conducted to determine the prevalence of multidrug-resistant (MDR) bacterial strains, isolated from hospital wastewater. Using standard procedures, bacterial resistance patterns against different classes of antibiotics were analysed and their species level identification was made. The disc diffusion method was used to determine the bacterial activity against antimicrobial agents. Clear zones were measured separately in millimeters around each disc. Five wastewater samples were collected from different drainage regions of hospitals situated in Multan. A total of 45 bacterial strains were isolated. Out of these 45 bacterial strains, 13 (29%) were found resistant against two or more than two classes of antibiotics. All the bacterial strains (100%) isolated from samples 2 and 3 were MDR. Twenty-five bacterial strains (55.5%) belonged to the Bacillus species and others belonged to Enterococcus species, Micrococcus species, Staphylococcus species, and Streptococcus species, respectively. The presence of resistant bacterial strains in hospital waste demands the availability of effectual treatment plants to treat the waste before it is disposed of into hospital waste lines.

In this study, cross-linked cellulase aggregates (C-CLEAs) were synthesized by precipitation of cellulase with ammonium sulfate and then cross-linking with glutaraldehyde. The results revealed that the optimal pH of C-CLEAs shifted toward a more acidic environment by 2.0 pH units, and the optimal temperature shifted toward higher temperature by 20 °C after immobilization. The half-life (t

Transglycosylating glycoside hydrolases (GHs) offer great potential for the enzymatic synthesis of oligosaccharides. Although knowledge is progressing, there is no unique strategy to improve the transglycosylation yield. Obtaining efficient enzymatic tools for glycan synthesis with GHs remains dependent on an improved understanding of the molecular factors governing the balance between hydrolysis and transglycosylation. This enzymatic and structural study of RBcel1, a transglycosylase from the GH5_5 subfamily isolated from an uncultured bacterium, aims to unravel such factors. The size of the acceptor and donor sugars was found to be critical since transglycosylation is efficient with oligosaccharides at least the size of cellotetraose as the donor and cellotriose as the acceptor. The reaction pH is important in driving the balance between hydrolysis and transglycosylation: hydrolysis is favored at pH values below 8, while transglycosylation becomes the major reaction at basic pH. Solving the structures of two RBcel1 variants, RBcel1_E135Q and RBcel1_Y201F, in complex with ligands has brought to light some of the molecular factors behind transglycosylation. The structure of RBcel1_E135Q in complex with cellotriose allowed a +3 subsite to be defined, in accordance with the requirement for cellotriose as a transglycosylation acceptor. The structure of RBcel1_Y201F has been obtained with several transglycosylation intermediates, providing crystallographic evidence of transglycosylation. The catalytic cleft is filled with (i) donors ranging from cellotriose to cellohexaose in the negative subsites and (ii) cellobiose and cellotriose in the positive subsites. Such a structure is particularly relevant since it is the first structure of a GH5 enzyme in complex with transglycosylation products that has been obtained with neither of the catalytic glutamate residues modified.

The medicinal plant Juniperus oxycedrus is less recognized for the diversity of its fungal endophytes and their potential to produce extracellular enzymes. The present study is the first report on the isolation and identification of a mesophilic endophytic strain JO-A, Preussia africana, from fresh stems of the J. oxycedrus endemic tree in the Ifrane region-Morocco, and the evaluation of its ability to produce cellulases. A one-time multi-parameter one-factor screening was optimized to select factors that enhance cellulase production in P. africana. The maximum production of both CMCase and FPase activities were 1.913 IU.mL

Significant anaerobic fermentation occurs in silage through the action of anaerobic bacteria. The objective of this study was to evaluate the effects of cellulase and Lactobacillus plantarum on the fermentation quality and bacterial community of whole-plant corn and peanut vine mixed silage. Mixed silage was tested with no addition (CK), addition of Lactobacillus plantarum (LP), addition of cellulase (CE), and the simultaneous addition of Lactobacillus plantarum and cellulase (LPCE). LPCE samples exhibited decreased pH; decreased content of acetic acid, propionic acid, and butyric acid; and increased content of lactic acid. LP and LPCE had better effects on chemical composition than CK and CE, especially in decreasing acid detergent fiber and neutral detergent fiber content. High-throughput sequencing identified Lactobacillus, Klebsiella, Serratia, and Weissella as the main microorganisms. LP and CE increased the abundance of Acetobacter, and LPCE decreased the abundance of Acetobacter. All additives decreased the abundance of Weissella, Leuconostoc, and Lactococcus, and increased the abundance of Pantoea. Overall, simultaneous addition of cellulase and Lactobacillus plantarum helped to improve the quality of mixed silage of whole-plant corn and peanut vines.

The quest for novel broad spectrum bioactive compounds is needed continuously because of the rapid advent of pathogenic multi drug resistant organisms. Actinomycetes, isolated from unexplored habitats can be a solution of this problem. The motive of this research work was isolation of actinomycetes having potential antimicrobial activities from unexplored regions of Devbag and Tilmati beach. The isolated actinomycetes were screened against pathogenic microbes for antimicrobial activities through cross streak method. Enzyme production activity was checked for these actinomycetes for amylase, protease, cellulase and lipase enzymes. Further antimicrobial activity of ethyl acetate extract of the potent strain KS46 was performed. The strain KS46 was identified with 16S rRNA gene sequencing and secondary structure was analysed. Gas chromatography-Mass spectrometry (GC-MS) profiling was conducted to ascertain the presence of bioactive metabolites in the ethyl acetate extract. The collected samples were pre-treated and 70 actinomycetes were isolated. The Streptomyces sp. strain KS46 showed the best antimicrobial activity in primary screening. Ethyl acetate extract of the strain KS46 revealed antimicrobial activity against S. aureus, B. subtilis, B. cereus, E. faecalis, K. pneumoniae, E. coli, S. flexneri, C. albicans and C. glabrata. The 16S rRNA gene sequencing identified the strain KS46 as Streptomyces levis strain KS46. The GC-MS metabolite profiling of the ethyl acetate extract revealed the availability of 42 compounds including fatty acid esters, fatty acid anhydrides, alkanes, steroids, esters, alcohols, carboxylic ester, etc. having antibacterial, antifungal, antiproliferative, antioxidant activities. This study indicated that Devbag and Tilmati beaches being untapped habitats have enormous diversity of promising antimicrobial metabolite producing actinomycetes. Therefore, further exploration should be carried out to characterize the potential actinomycetes, which can be optimistic candidates for generation of novel antimicrobial drugs.

Distillers' dried grains with solubles (DDGS) is a by-product of dry-mill corn ethanol production comprising a high nutritional value due to residual fiber, protein, and lipid contents. The fiber content of DDGS is high enough to be considered a valuable source for the production of hydrolytic enzymes, such as cellulase and xylanases, which can be used for hydrolysis of lignocellulosic feedstock during ethanol production. The DDGS-based medium prepared after acid hydrolysis provides adequate sugars for enzyme production, while additional macronutrients, such as salts and nitrogen sources, can enhance the enzyme production. Therefore, this study was undertaken to evaluate the effect of salts (KH2PO4, CaCl2 center dot 2H(2)O, MgSO4 center dot 7H(2)O, FeSO4 center dot 7H(2)O, CoCl2 center dot 6H(2)O, and MnSO4 center dot H2O), peptone, and yeast extract on enzyme secretion by four different Aspergillus niger strains and to optimize the nitrogen source for maximum enzyme production. Yeast extract improved the cellulase production (0.38 IU/ml) for A. niger (NRRL 1956) as compared to peptone (0.29 IU/ml). However, maximum cellulase productions of 0.42 IU/ml and 0.45 IU/ml were obtained by A. niger (NRRL 330) and A. niger (NRRL 567), respectively, in presence of ammonium sulfate. The optimized nitrogen amounts resulted in a significant increase in the cellulase production from 0.174 to 0.63 IU/ml on day 9 of the fermentation with A. niger (NRRL 330). The composite model improved both cellulase and xylanase production. In conclusion, the optimization of all three nitrogen sources improved both cellulase and xylanase production in the DDGS-based media.

To achieve an economically viable substitutive strategy for lignocellulosic residues use, cellulose degradation by efficient Trichoderma strains become a key alternative. This study aimed to explore the effects of various cultural conditions on the degradation of different lignocellulosic substrates (corn straw and licorice residues) by Trichoderma longibrachiatum and Trichoderma afroharzianum in solid state fermentation. The strains were isolated from the rhizospheric soils of medicinal plants. In addition, the best fermentation combination was selected to evaluate the degradation results. This study confirmed that fermentation conditions and different components of corn straw and licorice residues were both crucial factors affecting the cellulase activity and spore production of two Trichoderma strains. A combination of T. afroharzianum degrading corn straw was selected for Response Surface Methodology optimization. The best fermentation conditions were fermentation time of 5.72 days, initial material liquid ratio of 1:4.58, initial pH of 6.32 and fermentation temperature of 28.8 ℃, with a filter paper cellulase activity of 0.91 U/mL. After continuous fermentation, the spore production reached 6.18 × 109 spores/g, the crude fiber degradation rate reached 23.5% with the reducing sugar content maintained at 11 mg/g. These results highlight the great potential of T. afroharzianum degrading corn straw for organic amendment.

Polyphenols display potential functional activities especially for rosmarinic acid (RosA) and caffeic acid (CafA). In this study, two different methods including ultrasonic-assisted ethanol extraction (60%) and ultrasound-assisted cellulase (≥ 15000 U/g, 2%) hydrolysis were applied for the extraction of total phenolics from 44 species of Perilla frutescens. Folin-Ciocalteu method detection showed that the content of total phenolics extracted by cellulase hydrolysis was the highest and attained to 28.00 mgGAE/gextracts for ZB1. Continuously, the extracts were purified by XDA-8 macroporous resin and medium-pressure liquid chromatography (MPLC), and the content of total phenolics improved to 66.62 mgGAE/gextract. High-performance liquid chromatography (HPLC) assay showed that the total polyphenols were mainly composed of gallic acid, caffeic acid, rosmarinic acid, luteolin and apigenin. Besides, a sequential XDA-8 macroporous resin combined with high-speed counter-current chromatography (HSCCC)/MPLC system was established for simultaneously isolation and preparation of RosA (purity 98.29%) and CafA (purity 97.01%) from the extracts. Further, antibacterial activities of the total polyphenols were evaluated by disc diffusion method and scanning electron microscope (SEM) observation. The results verified that the total polyphenols had effective antibacterial capacity on three kinds of bacteria including E. coli, S. aureus, and B. subtilis in a concentration-dependent manner. All these results demonstrated that the ultrasound-assisted cellulase hydrolysis extraction of total polyphenols and proposed three-step separation of RosA and CafA had a high yield and good purity, and they exhibited an effective antibacterial ability.

The intake of moderate oils and fats is necessary to maintain the body's energy balance, and the fatty acid composition of different oils and fats varies in their nutrition and function. The study aimed to investigate the effects of lard and vegetable blend oil on gut microbiota, intestinal enzyme activities, and blood routine. Kunming mice were assigned to the three groups: (1) Control group (CK) was gavage administration with distilled water, (2) Plant oil group (ZWY) was gavage administration with edible vegetable blend oil, (3) Lard group (DWY) was gavage administration with lard. After 42 days, microbiological, digestive enzymes, and blood routine were performed. Compared with the CK group, Escherichia coli, Lactobacilli, and Bifidobacteria were significantly decreased (p0.05), the activities of protease, cellulase, amylase, and xylanase were markedly reduced (p0.05), the hemoglobin was significantly increased (p0.05) in the ZWY group and DWY groups, and the hematocrit was increased in the ZWY group (p0.05), while other routine blood indices were increased (p0.05). Compared to the ZWY group, the activity of cellulase and amylase were significantly increased (p0.05), the intestinal microorganism and the routine blood indexes had no significant difference in the DWY group. Lard and vegetable blend oil diet affected the composition of the intestinal microorganisms, and the functions of digestive enzymes. Meanwhile, the levels of digestive enzymes may be correlated with the intestinal microbiota.

Enhanced hydrogen evolution was pursued in this work. Rhodobacter sp. (Rb) and Rhodopseudomonas palustris (Rp), single or mixed were used to extract hydrogen molecules from Chlorella fusca biomass. To elevate their fermentable contents, Chlorella was grown at nitrogen and/or phosphorus deprivation. Besides, cellulase and/or macerozyme, Triton X100 or sonicated yeast were applied for further biohydrogen fermentation. Utilizing hydrolysates of mineral deprived Chlorella cultures, Rb exhibited relatively higher cumulative hydrogen (4200 ml L−1) than Rp (2500 ml L−1) while mixed cultures attained significantly higher levels (4700 ml L−1). Triton or enzymes significantly enhanced hydrogen evolution, with more effectiveness of macerozyme than cellulase. A novel use of sonicated yeast, as enzymes pool, induced the highest significant collective H2 (up to 47 times that of microalgal supernatant). Sonicated yeast induced a remarkable hydrolysis of algae, as inferred from increased reducing sugars. However, hydrogen evolution efficiency exhibited poor proportionality with reducing sugars, indicating fermentation of other metabolites.

This study provides a detailed analysis of the lignocellulolytic activity of a new isolate Stereum gausapatum F28, a Serbian autochthonous fungi, on beechwood sawdust supplemented with cheap waste, sugar beet molasses still age. Advanced multiple response optimization techniques were applied to improve ligninolytic and reduce hydrolytic activity as a requirement for potential biorefinery use. The applied techniques were supposed to select cultivation conditions that would give manganese peroxidase and laccase activities above 0.84 and 0.12 U g(-1) substrate, respectively, and cellulase and xylanase activities below 1.12 and 1.4 U g(-1) substrate. The optimal cultivation conditions that met the set requirements included molasses stillage concentration of 10 %, substrate moisture content of 53 %, incubation temperature of 23.5 degrees C, and pH 5.2. The research showed that the addition of molasses stillage had a positive effect on enzyme production and that the optimal stillage concentration differed depending on the enzyme type (for laccase it was lt 5 %, manganese peroxidase approximate to 12 %, cellulase approximate to 21 % and xylanase approximate to 16 %), which should be taken into consideration when optimizing the desired process..

Labyrinthulomycetes are a group of ubiquitous and diverse unicellular Stramenopiles and have long been known for their vital role in ocean carbon cycling. However, their ecological function from the perspective of organic matter degradation remains poorly understood. This study reports high-quality genomes of two newly isolated Labyrinthulomycetes strains, namely, Botryochytrium sp. strain S-28 and Oblongichytrium sp. strain S-429, and provides molecular analysis of their ecological functions using comparative genomics and a biochemical assay. Our results suggest that Labyrinthulomycetes may occupy multiple ecological niches in marine ecosystems because of the significant differences in gene function among different genera. Certain strains could degrade wheat bran independently by secreting cellulase. The key glycoside hydrolase families (GH1, GH5, and GH9) related to cellulase and the functional domains of carbohydrate-active enzymes (CAZymes) were more enriched in their genomes. This group can actively participate in marine biochemical cycles as decomposers. In contrast, other strains that could not produce cellulase may thrive as "leftover scavengers" and act as a source of nutrients to the higher-trophic-level plankton. In addition, our findings emphasize the dual roles of endoglucanase, acting as both exo- and endoglucanases, in the process of cellulose degradation. Using genomic, biochemical, and phylogenetic analyses, our study provides a broader insight into the nutritional patterns and ecological functions of Labyrinthulomycetes. IMPORTANCE Unicellular heterotrophic eukaryotes are an important component of marine ecosystems. However, their ecological functions and modes of nutrition remain largely unknown. Our current understanding of marine microbial ecology is incomplete without integrating these heterotrophic microeukaryotes into the food web models. This study focuses on the unicellular fungus-like protists Labyrinthulomycetes and provides two high-quality genomes of cellulase-producing Labyrinthulomycetes. Our study uncovers the basis of their cellulase production by deciphering the results of genomic, biochemical, and phylogenetic analyses. This study instigates a further investigation of the molecular mechanism of organic matter utilization by Labyrinthulomycetes in the world's oceans.

The research was conducted to isolate soil fungi and screen them for cellulase production using the zone of hydrolysis technique. Several fungi were isolated and characterised from soil environments of different locations using conventional microbiological methods. A total of six isolates were confirmed to be Penicillium chrysogenum, Emericella rogulosus, Aspergillus terreus, Aspergillus flavus, Aspergillus niger, Aspergillus fumigatus, all coded as BG1, BG2, BG3, BG4, BG5 and BG6, respectively. Fungal isolate BG5 has the highest percentage of occurrence (34.30 %), followed by SBG3 (22.86 %). The isolates were screened for cellulase production using the carboxymethyl cellulose (CMC) agar plate method. All the fungal isolates demonstrated cellulase production ability, with fungal isolates BG5 (18 mm) and BG3 (15 mm) having the highest diameter of zone of cellulose hydrolysis. The research reveals the potentiality of using locally isolated soil fungi for cellulase production.

A sustainable and integrated biorefinery strategy was achieved by a facile biphasic system, ferric trichloride (FeCl3) solution assisted by methyl isobutyl ketone (MIBK), yielding multiple products: furfural, sugars, and lignin nano-particles simultaneously. For the highest yield of furfural, the conditions were optimized resulting in 75.55% and over 69.38% of the yields from xylose and some typical biomasses respectively. Both the catalyst and MIBK can be recycled. Eucalyptus was lucubrated on the valorizations of three major-components with 75.18% of furfural yield, 97.54% of cellulase digestibility from the residue and lignin nano-particles. Meanwhile, the regularities of biomass degradation under this biphasic system were summarized as well as high digestibility which was resulted by hemicellulose and lignin removal in cooperation with the amount of the cellulase binding sites. Based on the proposed biorefinery strategy, 65.82% of the initial mass can be valorized into high-value end-products.

Saccharification of lignocellulose is prerequisite for the biorefinery. Clostridium thermocellum is a promising candidate for the hydrolysis of lignocellulose. A study on the saccharification by Clostridium thermocellum combined with surfactant and cellulase addition was investigated. The reducing sugars content increased with increasing substrate loading in the fermentation of C. thermocllum with isooctyl alcohol polyoxyethylene ether (IAPE) addition. Based on IAPE addition, the glucose content of two-step saccharification was higher than that of the batch saccharification. Besides, the addition of IAPE and Triton X-100 had the same effect on saccharification. The saccharification performance was related to the substrate, especially the lignin content. For the fed-batch hydrolysis of 20 % (w/v) rice straw pretreated by hydrogen peroxide-acetic acid (HPAC), 66.9 g/L glucose and 19.5 g/L xylose were obtained by using C. thermocellum DSM 1313 and 1 FPU/g substrate cellulase with 0.1 % (w/v) IAPE addition, and the glucose and xylose yield reached 63.2 % and 37.4 % respectively.

Endoglucanases are carbohydrate-degrading enzymes widely used for bioethanol production as part of the enzymatic cocktail. However, family 5 subfamily 5 (GH5_5) endoglucanases are still poorly explored in depth. The Trichoderma reesei representative is the most studied enzyme, presenting catalytic activity in acidic media and mild temperature conditions. Though biochemically similar, its modular structure and synergy with other components vary greatly compared to other GH5_5 members and there is still a lack of specific studies regarding their interaction with other cellulases and application on novel and better mixtures. In this regard, the threedimensional structure elucidation is a highly valuable tool to both uncover basic catalytic mechanisms and implement engineering techniques, proved by the high success rate GH5_5 endoglucanases show. GH5_5 enzymes must be carefully evaluated to fully uncover their potential in biomass-degrading cocktails: the optimal industrial conditions, synergy with other cellulases, structural studies, and enzyme engineering approaches. We aimed to provide the current understanding of these main topics, collecting all available information about characterized GH5_5 endoglucanases function, structure, and bench experiments, in order to suggest future directions to a better application of these enzymes in the industry.

Low nutritional value and digestive efficiency restrict the feeding application of brewer's spent grain (BSG). Bacillus velezensis has been increasingly considered a promising fermenting agent in feed production. This study aimed to investigate the fermentative characteristics of Bacillus velezensis K8 on degrading lignocellulose and increasing soluble sugar and protein in BSG. The lignocellulose contents (including cellulose, hemicellulose and lignin) decreased by 36.85 % and 27.68 % in BSG and ultrasonic-pretreated BSG (UBSG), respectively. The reducing sugar contents increased by 226.8 % and 198.1 %, and the soluble protein contents increased by 260.7 % and 258.3 % in BSG and UBSG, respectively. The arabinose, cellobiose, xylose, glucose, fructose, and xylooligosaccharides contents were significantly increased, indicating effective conversion of cellulose and hemicellulose into soluble sugars. High cellulase activity was obtained in BSG fermentation, especially CMCase and filter paper activities. Protease secretion increased but cellulase decreased, and the activities of xylanase and fructofuranosidase remained stable in UBSG fermentation. Genome analysis revealed that B. velezensis K8 possessed more amino acid-metabolism genes than carbohydrate-metabolism genes, illustrating that B. velezensis K8 preferentially utilized protein nutrients but left more sugars. These results provide a promising strategy for improving the value of grain residue in food and feed industrial applications.

PCWDEs are considered important components of the virulence arsenal of necrotrophic plant pathogens. However, despite intensive research, the role of PCWDEs in the pathogenicity of necrotrophic phytopathogenic fungi remains ambiguous.

Numerous transcription factors (TFs) in ascomycete fungi play crucial roles in cellular processes; however, how most of them function is poorly understood. Here, we identified and characterized a novel TF, CxrC (POX01387), acting downstream of the key TF CxrA, which is essential for plant-biomass-degrading-enzyme (PBDE) production in Penicillium oxalicum. Deletion of cxrC in P. oxalicum significantly affected the production of PBDEs, as well as mycelial growth and conidiospore production. CxrA directly repressed the expression of cxrC after about 12 hr following switch to Avicel culture. CxrC bound the promoters of major PBDE genes and genes involved in conidiospore development. CxrC was found to bind the TSSGTYR core sequence (S: C and G; Y: T and C; R: G and A) of the important cellulase genes cbh1 and eg1. Both N- and C-terminal peptides of CxrC and the CxrC phosphorylation were found to mediate its homodimerization. The conserved motif LPSVRSLLTP (65-74) in CxrC was found to be required for regulating cellulase production. This study reveals novel mechanisms of TF-mediated regulation of the expression of PBDE genes and genes involved in cellular processes in an ascomycete fungus.

Invasive plants offer an interesting and unconventional source of protein and the considerable investment made towards their eradication can potentially be salvaged through their revalorisation. To identify viable sources, effective and high-throughput screening methods are required, as well as efficient procedures to isolate these components. Rigorous assessment of low-cost, high-throughput screening assays for total sugar, phenolics and protein was performed, and ninhydrin, Lever and Fast Blue assays were found to be most suitable owing to high reliability scores and false positive errors less than 1%. These assays were used to characterise invasive Scottish plants such as Gorse (Ulex europeans), Broom (Cystisus scoparius) and Fireweed (Chamaenerion angustifolium). Protein extraction (alkali-, heat- and enzyme assisted) were tested on these plants, and further purification (acid and ethanol precipitation, as well as ultrafiltration) procedures were tested on Gorse, based on protein recovery values. Cellulase treatment and ethanol precipitation gave the highest protein recovery (64.0 ± 0.5%) and purity (96.8 ± 0.1%) with Gorse. The amino acid profile of the purified protein revealed high levels of essential amino acids (34.8 ± 0.0%). Comparison of results with preceding literature revealed a strong association between amino acid profiles and overall protein recovery with the extraction method employed. The final purity of the protein concentrates was closely associated to the protein content of the initial plant mass. Leaf protein extraction technology can effectively raise crop harvest indices, revalorise underutilised plants and waste streams.

The present study reports the characterization of an extracellular polymer produced by a strain of Lactobacillus helveticus in potato waste medium and the subsequent application of the biopolymer for protecting farm-bruised pomegranates against fungal infection following harvest. Chemical, thermogravimetric analysis, FTIR and scanning electron micrographs of the purified biopolymer revealed it to be a polysaccharide with good thermal stability and a compact structure. Purified biopolymer exhibited strong antifungal properties against the fungal pathogen Penicillium implicatum. In tissues of pomegranates, subjected to fungal challenge, pectinase, cellulase and xylanase activity indicative of infectivity was not observed upon storage for 14 days at 28 °C. Besides visual discoloration of pomegranates, characteristic of soft rot and fungal growth of tissue extracts could not be detected on PDA plates. Postharvest parameters, physiological loss of weight, TA, TSS, TSS/TA ratio, ascorbic acids, total sugars and sensory attributes in the biopolymer dipped bruised pomegranates challenged with P. implicatum, showed no significant (p > 0.05) change following storage. P. implicatum challenged bruised fruits which lacked dipping treatment was spoiled by 48 h. Farm-bruised pomegranates dipped with biopolymer offered complete protection of the bruised pomegranates upon storage for 14 days at ambient temperature (28 °C). Overall, the results of this study suggest a sustainable use of potato processing wastes for the reutilization of 5–16% of damaged pomegranates, economically beneficial to farmers. Besides, the process will have strong importance in reducing and recycling potato industry wastes for innovative postharvest applications for other horticultural produces.

PurposeThe purpose of this research is to produce “cempedak” juice using enzyme aided-liquefaction by examining the effects of enzyme types (Pectinex® Ultra SP-L, Celluclast® 1.5 L and Fungamyl® 800 L), enzyme concentrations (0–1.5% v/w), incubation time (0–2.5 h) and incubation temperature (35–60 °C) on juice yield and viscosity, total soluble solids and color of fruit puree.Design/methodology/approachRipe “cempedak” pulp from CH28 fruit was first pureed in a blender and then homogenized with water at 1:2 ratio. The diluted puree was then liquefied with the enzymes separately to reduce its viscosity. Analyses such as juice yield, viscosity, total soluble solids and color of the liquefied “cempedak” puree were then carried out.FindingsResults indicated that the optimized use of 1.2% (v/w) Celluclast® 1.5 L (Novozymes, Denmark), a cellulase preparation, at 45 °C and 1 h produced juice with the lowest viscosity (349.4 cP) and the highest juice yield (82.3% v/w). Liquefied “cempedak” juice was darker (with L* value of 51.17) and more yellowish (b* value of 38.88) compared to “cempedak” juice without liquefaction (control). When compared to untreated “cempedak” juice, the droplet size of “cempedak” juice obtained after liquefaction under optimized conditions was found to be lower, regardless of whether the juice was filtered (with a total reduction of 23% of droplet size) or not filtered (with a total reduction of 16% of droplet size). The results indicate the possibility of employing Celluclast® 1.5 L to produce “cempedak” juice that can be further processed such as for the production of “cempedak” fruit powder.Originality/valueThis paper provides information on the enzyme concentration, incubation time and temperature for liquefying “cempedak” pulp such that the liquefied material produced can be used as a base feed for spray-drying to produce “cempedak” fruit powder.

Effective pretreatment is crucial for cellulase production from sugarcane bagasse. Pretreatment with tamarind extract could reduce the hazardous effect associated with chemical pretreatment. The present work investigated tamarind (Tamarindus indica) extract in combination with H2SO4 and thermal pretreatment of sugarcane bagasse for cellulase production by Aspergillus flavus. The sugarcane bagasse was pretreated with tamarind extract pH 2 and pH 4, followed by 1% H2SO4 and thermal treatment at 121°C for 15 min. The pretreatment slurry was analysed for reducing sugar while solid bagasse was analysed for weight loss. Aspergillus flavus grew on sugarcane bagasse under solid state fermentation and the Carboxy Methyl Cellulase (CMCase) and Filter Paper Assay (FPA) activities were compared on the various pretreatments. The pretreatments changed the visible morphology of the sugarcane bagasse observed by the swelling, fibrous appearance and colour change. Pretreatment slurry yielded highest soluble reducing sugar at 60.01 mg/ml in tamarind extract (pH 4/1% H2SO4 ) and highest weight loss of solids at 73.70% in tamarind extract (pH 2/1% H2SO4 /thermal 121°C). Aspergillus flavus performed better on tamarind extract (pH 2/1% H2SO4 ) by producing optimal CMCase and FPA activities at 0.100 U/ml and 0.409 U/ml respectively after 3 days of fermentation. Cellulase was maximally active at temperature of 50 °C. The tamarind extract pretreatment successfully proved to be an alternative organo-chemical pretreatment of sugarcane bagasse as evidenced by the physical properties, soluble reducing sugars and cellulase activities. Keywords: Aspergillus flavus, Cellulase, Pretreatment, Sugarcane bagasse, Tamarind extract

La cinética microbiana y enzimática son factores importantes durante la degradación aerobia de la fracción orgánica de los residuos sólidos urbanos, estas dependen principalmente de la temperatura de incubación y las tasas de aireación. El objetivo de esta investigación fue evaluar el proceso de degradación aerobia, por múltiples variables y su combinación para comprender las interacciones entre las tasas de aireación en la degradación aerobia y sus respuestas. Las tasas de aireación se fijaron en 0.032, 0.064, 0.125, 0.251 y 0.392 L de aire húmedo kg-1 min-1 a 35 °C con inóculo. La actividad microbiana se evaluó de forma indirecta por medio de la respirometria; es decir, la generación de CO2 y el consumo de O2. Las actividades enzimáticas extracelulares (es decir, pectinasas, celulasas, xilanasas y proteasas) se cuantificaron mediante la liberación de los azúcares reductores. Los diferentes ensayos se realizaron en la Universidad Autónoma Metropolitana Unidad Iztapalapa en septiembre de 2019. Encontrando una fuerte relación positiva entre la actividad enzimática xilanasa y pectinasa con la pérdida de peso en seco, junto con el aumento de las actividades celulasas y xilanasas a mayores tasas de aireación.

Management of lignocellulosic wastes in and around the municipality area requires special consideration. Continuous deposition of these wastes to the nearby areas led to gradual deterioration of the environment. The objective of this study was to produce cellulase from the bacteria isolated from the unexplored rainforest of NE-India for lignocellulosic waste hydrolysis. Based on carboxymethyl cellulose utilization and the congo red test, Novosphingobium sp. Cm1 was found to be the most promising strain out of 114 bacterial isolates and the strain was selected for further study. The optimization of the fermentative conditions for maximum enzyme activity was carried out using one factor-at-a-time strategy and the optimum pH, temperature and incubation time was recorded as pH 5, 37 °C and 96 h respectively. The maximum β-1,4-endoglucanase activity was observed with 1.5% CMC (5.1 ± 0.05 U/mL) and 0.25% yeast extract (7.6 ± 0.72 U/mL). The bacterial waste hydrolysis ability was investigated using three wastes where vegetable waste showed maximum activity of 3.4 ± 0.48 U/mL. Bacterial interaction and waste utilization were verified using Scanning Electron Microscope and Fourier-Transform infrared spectroscopy analysis. The present study confirmed the promising ability of Novosphingobium sp. to waste hydrolysis. Further investigations may lead to new possibilities for low-cost enzyme production that will help to meet the rising cellulase demand.