Adsorption of cellulose by Trichoderma viride

Adsorption of cellulase by Trichoderma viride QM 9414 has been studied with resting and growing cells and equations have been derived to describe the process quantitatively. It has been observed that the adsorption is a purely physical process being dependent only on cell and cellulose concentrations. It has also been demonstrated that adsorption isrequired for the induction of cellulases; some discussions are devoted to this point.     

Adsorption of cellulase fromTrichoderma viride on microcrystalline cellulose

Summary The adsorption behaviour of cellulase fromTrichoderma viride on microcrystalline celluloses with different specific surface areas was studied. The adsorption was found to fit a Langmuir isotherm. There was an increase in the maximum adsorption amount (A_max) as the specific surface area of microcrystalline cellulose increased. The values of A_max and adsorption equilibrium constant (K) decreased with increasing temperature. Thermodynamic parameters in adsorption were calculated from K. It was found from the enthalpy of adsorption, that van der Waals-Type interaction was responsible for adsorption of cellulase on microcrystalline cellulose. The adsorption process was exothermic and an adsorption enthalpy-controlled reaction.     

Conversion of cellulose to glucose with cellulase of Trichoderma viride ITCC-1433

Trichoderma viride ITCC-1433 secretes a cellulase complex that is rich in β-glucosidase and therefore well suited for the saccharification of cellulosic materials. The cellulase was investigated with respect to optimum conditions of reaction and enzyme stability. Avicelase, CMCase, and β-glucosidase differed considerably in their physicochemical properties. At temperatures above 50°C, β-glucosidase is not very stable. Therefore, as a compromise the conditions of hydrolysis were chosen to be 50°C and pH 4.5. With the crude culture filtrate of T. viride ITCC-1433 a nearly pure glucose solution of 4% is reached from a 10% cellulose suspension. Wood pulp and newsprint are hydrolyzed to a much smaller extent. With an enzyme concentrate up to 8% glucose accumulated in the reaction fluid within 48 hr. At this time the glucose-cellobiose ratio was 75:1. Glucose was demonstrated to be the most potent inhibitor of total hydrolysis. The addition of glucose to the enzyme-substrate solution at zero time completely stopped its own formation and cellobiose and reducing groups (oligosaccharides) accumulated. By removing glucose through an ultrafilter device about 90% saccharification of cellulose to glucose was achieved in 48 hr without any accumulation of cellobiose.     

Biodegradation of wastepaper by cellulase from Trichoderma viride

Environmental issues such as the depletion of non-renewable energy resources and pollution are topical. The extent of solid waste production is of global concern and development of its bioenergy potential can combine issues such as pollution control and bioproduct development, simultaneously. Various wastepaper materials, a major component of solid waste, were treated with the cellulase enzyme from Trichoderma viride, thus bioconverting their cellulose component into fermentable sugars. All wastepaper materials exhibited different susceptibilities towards the cellulase as well as the production of non-similar sugar releasing patterns when increasing amounts of paper were treated with a fixed enzyme concentration. The hydrolysis of wastepaper with changing enzyme concentrations and incubation periods also resulted in dissimilar sugar-producing tendencies. A general decline in hydrolytic efficiency was observed when increasing sugar concentrations were produced during biodegradation of all wastepaper materials.     

Kinetic study on enzymatic hydrolysis of cellulose by cellulose from Trichoderma viride

Pure cellulose (Avicel) was hydrolyzed batchwise at 50 degrees C and pH 4.8 by cellulase from Trichoderma viride (Meicelase CEP). Then the effects of the crystallinity of cellulose as well as the thermal deactivation and product (cellubiose and glucose) inhibition to cellulose on the hydrolysis rate were quantitatively investigated. While these factor had evidently retarded the enzymatic hydrolysis of cellulose to a significant extent, the hydrolysis rates observed could not be explained. For practical purposes, an empirical, simple rate expression was developed which included only one parameter: a overall rate retardation constant. This empirical rate expression held for the hydrolysis of at least two kind of cellulosic materials: Avicel and tissue paper.     

Continuous cultivation of Trichoderma viride on cellulose

Using ball milled cellulose as the only carbon source Trichoderma viride was grown in a continuous flow culture at pH = 5.0 and T = 30°C. Steady-state values for cell protein, cellulose, and cellulase for different substrate concentrations (4–11 g/liter) and dilution rates (0.033–0.080 hr^?1) were obtained. Under steady-state conditions, 50–75% of the cellulose was consumed indicating a critical dilution rate on 0.17 hr^?1. Cellulase activity (U/ml) in the fermentation broth increased slightly with increasing substrate concentration and decreased with increasing dilution rate, while the specific cellulase productivity (U/mg cell protein·hr) was fairly independent of the dilution rate, with a maximum around D = 0.05 hr^?1. Following step changes in substrate concentration and dilution rate, new steady-state values were reached after three to five residence times (cell protein and cellulose) and four to six residence times (celullase activity).     

Production of cellulase and protein from barley straw by Trichoderma viride

The cellulase production by two strains of the cellulolytic fungus Trichoderma viride was examined. The fungi were grown on different preparations of barley straw pretreated with NaOH under high pressure. The production of cellulases and microbial protein by the better strain (QM 9123) was investigated in an aerated 5-liter fermenter under varying stirring rates (200-350 rpm) and straw concentrations (1–2%). The pH was kept between 3.5 and 4.5. The growth of the fungus was followed by measuring the quantity of CO₂ produced and the cell protein. After 2–6 days growth ceased, the lag phases lasting 0–2 days, increasing with increasing straw concentrations. The maximum enzyme yields were reached after 4–10 days. The protein content of the product was 21–26% and up to 70% of the straw was utilized. The yield constants were calculated to be 0.40–0.56; of the same order as those which can be obtained by growing the fungus on glucose.     

Kinetics of cellulase production intrichoderma viride on microcrystalline cellulose

During the cultivation of a wild strain ofT.viride on microcrystalline cellulose the synthesis of cell-bound FP cellulases precedes cell growth. During the growth they are released into the medium as extracellular enzymes. The rate of synthesis of extracellular FP cellulases increases during cell growth, reaching a maximum at the beginning of transition to the stationary phase when the cell growth rate decreases. In contrast to extracellular enzymes, the rate of synthesis of bound cellulases during active growth is almost constant. In the stationary phase the rate of synthesis of both FP cellulases drops sharply, ceasing well before cell lysis sets in and before the maximum level of extracellular cellulases is attained.     

A method for increasing cellulase production by Trichoderma viride

A doubling of cellulase production by Trichoderma viride (QM9414) is possible by increasing the cellulose concentration in the medium from 0.75 to 2%, increasing the nitrogen concentration, and controlling pH during growth. A four-to fivefold increase in β -glucosidase is found with the higher cellulose concentration. Culture filtrates from 2% cellulose cultures can reduce the hydrolysis time in a practical saccharification to one-half that required by culture filtrates from 0.75% cellulose cultures.     

Transglycosylation of cellobiose by partially purified Trichoderma viride cellulase.

A commercial cellulase from Trichoderma viride was fractionated into three fractions, F1, F2, and F3, in order to investigate transglycosylation activities. Among these fractions, F3, which demonstrated highly hydrolytic activity toward p-nitrophenyl beta-D-glucopyranoside and Avicel, most effectively catalyzed the transglycosylation of cellobiose and converted cellobiose into beta-Glc-(1-->6)-beta-glc-(1-->4)-Glc and beta-Glc-(1-->6)-beta-Glc-(1-->6)-beta-Glc(1-->4)-Glc. The F3 fraction contained the enzyme to catalyze beta-glucosyl transfer toward only the C-6 position of the sugar acceptor, and thus it is expected to be of use for syntheses of functional oligosaccharides.     

Strain improvement for enhanced production of cellulase in Trichoderma viride

The filamentous fungi Trichoderma species produce extracellular cellulase. The current study was carried out to obtain an industrial strain with hyperproduction of cellulase. The wild-type strain, Trichoderma viride TL-124, was subjected to successive mutagenic treatments with UV irradiation, low-energy ion beam implantation, atmospheric pressure non-equilibrium discharge plasma (APNEDP), and N-methyl-N'-nitro-N-nitrosoguanidine to generate about 3000 mutants. Among these mutants, T. viride N879 strain exhibited the greatest relevant activity: 2.38-fold filter paper activity and 2.61-fold carboxymethyl cellulase, 2.18-fold beta-glucosidase, and 2.27-fold cellobiohydrolase activities, compared with the respective wild-type activities, under solid-state fermentation using the inexpensive raw material wheat straw as a substrate. This work represents the first application of APNEDP in eukaryotic microorganisms.     

Cell growth and cellulase production inTrichoderma viride on microcrystalline cellulose

The production of CM and FP cellulases was studied during the growth of a wild strain ofTrichoderma viride on microcrystalline cellulose. Part of the enzymes was found to be released into the medium while another part remained bound to the cell. Bound cellulases are released into the medium at the stage of cell lysis which takes place in the post-stationary phase. In this period extracellular CM and FP cellulases attain maximum activities. When the hyphae are subjected to a cold shock, maximum cellulase activity is detected already at the beginning of the stationary phase. An indirect method of dry cell mass determination showed that during exponential growth of cells on microcrystalline cellulose the μmax was 0.23 and the yield coefficient was 41 %.     

Enzymatic treatment of primary municipal sludge with Trichoderma viride cellulase

Enzymatic hydrolysis of the cellulose in raw primary settled municipal sludge by Trichoderma viride cellulase achieved conversions of up to 75% of the cellulose, primarily to cellobiose, in 24 hr. Simultaneously the gel-like characteristic of raw primary sludge was changed to that of a slurry of fine particles in less than 2 hr, causing a radical change in the ability to ultrafilter the sludge. The use of raw primary sludge as a growth medium for T. viride cellulase production was also investigated. It was possible, with nitrogen supplements, to obtain an enzyme with a filter paper activity (FPA) of two compared to over four which is attainable on defined medium of similar strength. The potential use of cellulase treatment as a pretreatment or integral part of waste treatment processes is discussed and the alternatives evaluated.     

Sugar production from agricultural woody wastes by saccharification with Trichoderma viride cellulase.

The saccharification of agricultural woody wastes was studied using a commercial enzyme preparation, Cellulase onozuka, derived from Trichoderma viride or the solid culture extracts of the fungus. With the intention of producing sugar at low cost, a simple procedure of enzymatic saccharification of rice straw, bagasse, and sawdust was studied. Delignifying methods of these wastes were investigated using dilute sodium hydroxide solution and dilute peracetic acid. Rice straw and bagasse were effectively delignified by boiling in a 1% sodium hydroxide solution for 3 hr or by autoclaving at 120 degrees C in a 1% sodium hydroxide solution for 1 hr. The sawdust from a broad leaved tree (Machilus thunbergii) was delignified by autoclaving at 120 degrees C in a 1% sodium hydroxide solution for 1 hr and by subsequent boiling in diluted 1/5 peracetic acid for 1 hr. This type of sawdust was also delignified by boiling in 1/5 peracetic acid for 1 hr and by subsequent autoclaving at 120 degrees C in a 1% sodium hydroxide solution for 1 hr. The sawdust from a coniferous tree (Cryptomeria japonica) was delignified by boiling in 1/5 peracetic acid for 1 hr and by subsequent autoclaving at 120 degrees C in a 1% sodium hydroxide solution for 1 hr; however, the successive treatment by autoclaving with alkali solution and subsequent boiling with diluted peracetic acid failed to bring about the desired effect. The saccharification of delignified rice straw, bagasse, and sawdust was examined using Cellulase onozuka, wheat bran or rice straw solid culture at various substrate concentrations, resulting in the formation of 5 to 10% sugar solutions after incubation at pH 5.0, 45 degrees C for 48 hr. The optimum substrate concentration existed at around 10%. Reuse of cellulase solution and resaccharification of residual sawdust were considered to be inadequate.     

Adsorption of Trichoderma reesei cellulase on cellulose: experimental data and their analysis by different equations

The adsorption of cellulase from Trichoderma reesei MCG 77 on Avicel was measured at varying cellulase (2-8 g/L) and Avicel (10-200 g/L) concentrations at pH 4.8 and 50 degrees C. Different mathematical equations were derived for the evaluation of the experimental data. The fraction of cellulase protein that can maximally be adsorbed is 0.96, and 1 g Avicel can bind maximally 0.092 g cellulase protein. The Michaelis constant for the adsorption equilibrium [cellulase] + [Avicel] right harpoon over left harpoon [cellulase Avicel] complex is between 2.0 and 2.3 . 10(-5) mol/L. This value is based on the assumption that cellulase has an average molecular weight of 48.000. The apparent molecular weight of Avicel, i.e., that amount in grams that can bind 1 mol cellulase, is 520,000. Under maximum binding the enzyme covers on Avicel a surface of 42 m(2)/g, and the occupied volume is 0.186 cm(3)/g Avicel.