Production of a cellulolytic enzyme system in mixed-culture solid-state fermentation of soybean hulls supplemented with wheat bran (original) (raw)
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Journal of the Taiwan Institute of Chemical Engineers, 2017
In-house production of saccharifying enzymes using lignocellulosic biomass as inducer of enzyme production can yield (hemi)cellulolytic enzymes with more specificity and efficiency for hydrolyzing the same lignocellulosic substrates. In the present study, production of (hemi)cellulolytic enzymes was carried out using microwave assisted alkali (MAA) treated wheat straw and the crude enzyme was evaluated for hydrolysis of the same substrate. Co-production of cellulolytic and hemicellulolytic enzymes by Aspergillus niger ADH-11 was optimized using MAA treated wheat straw as a substrate and corn steep liquor (CSL) as moistening medium under solid state fermentation employing response surface methodology. Under optimized conditions viz. inoculum 30% (v/v of moistening agent), CSL 7.1% and incubation time of 4.99 days, 2.34 U/g of FP activity, 308.16 U/g endo-glucanase, 96.61 U/g of β-glucosidase, 3815.96 U/g of xylanase and 174.42 U/g of β-xylosidase activity were produced. By statistical optimization, FP activity and xylanase yield were enhanced by 2.0 and 14.22 fold respectively and time for production was reduced significantly. It was found that supplementation of in-house produced enzyme to commercial cellulase can improve the levels of xylanase, β-glucosidase and β-xylosidase significantly. Enzyme cocktail containing 5 FPU/g of SIGMA cellulase and 5 FPU/g in-house produced enzyme yielded 610.35 mg/g of reducing sugars in 72 h with 68.41% saccharification and released more glucose as FP activity: β-glucosidase ratio was enhanced. The cocktail was also assessed for its efficacy at high substrate loading and lower temperature for its use in simultaneous saccharification and fermentation (SSF) process for bioethanol production.
2016
Production of cellulolytic enzymes like CMCase (endoglucanase), FPase, and xylanase by Aspergillus fumigatus SKH2 under solid state fermentation was carried out employing wheat bran as low cost substrate. Fermentation time, medium pH and incubation temperature were optimized at 48 h, pH 5.0 and 35 °C, respectively. At optimized state, CMCase (endoglucanase), FPase and xylanase of 826, 102 and 1130 U/gds yield was noticed, respectively. Crude enzyme cocktail was assayed at varied pH and temperature, and pH 5.0 and 35 °C were proved to be optimal for the studied enzyme activities. Fourier transform infrared spectroscopic FTIR analysis attested that NaOH was a good delignifying agent for sugarcane bagasse and grass Aristida sp., which enhanced subsequent saccharification efficiency of cellulolytic enzyme cocktail. By correlating FTIR analysis with saccharification profile it was found that highest saccharification was achieved after 16 h and 48 h after treating with 1M and 3M NaOH for ...
Enzyme Research, 2011
We investigated the effect of pretreatment on the physicochemical characteristics—crystallinity, bed porosity, and volumetric specific surface of soybean hulls and production of cellulolytic enzymes in solid-state fermentation ofTrichoderma reeseiandAspergillus oryzaecultures. Mild acid and alkali and steam pretreatments significantly increased crystallinity and bed porosity without significant change inholocellulosic composition of substrate. Crystalline and porous steam-pretreated soybean hulls inoculated withT. reeseiculture had 4 filter paper units (FPU)/g-ds, 0.6 IU/g-ds β-glucosidase, and 45 IU/g-ds endocellulase, whereas untreated hulls had 0.75 FPU/g-ds, 0.06 IU/g-ds β-glucosidase, and 7.29 IU/g-ds endocellulase enzyme activities. InA. oryzaesteam-pretreated soybean hulls had 47.10 IU/g-ds endocellulase compared to 30.82 IU/g-ds in untreated soybean hulls. Generalized linear statistical model fitted to enzyme activity data showed that effects of physicochemical characteristi...
Production of cellulase in solid-state fermentation withTrichoderma reesei MCG 80 on wheat straw
Applied Biochemistry and Biotechnology, 1996
It is an accepted fact that ethanol production from lignocellulosic materials is not economical as yet because of the high cost of cellulase production. To reduce the cost of cellulase production, lignocellulosic material (wheat straw [WS]), a comparatively much cheaper substrate, was used instead of costly substrates (pure cellulose or lactose). A pan bioreactor was developed for solid-state fermentation (SSF) that required a small capital investment. High yields of complete cellulase system were obtained compared to that in the liquid-state fermentation (LSF) from WS, when treated with 4.25% NaOH at 121~ for 1 h and mixed with Mandels' medium. A complete cellulase system is defined as one in which the ratio of Bglucosidase activity to filter paper activity in the enzyme solution is close to 1.0. The cellulase system derived from SSF using the pan bioreactor gave more than 85% hydrolysis of delignified WS. The prototype pan bioreactor requires further improvements so that optimum quantity of substrate can be fermented to obtain high yields of complete cellulase system per unit space. The SSF process provides a means for the production of complete cellulase system for the economical bioconversion of renewable biomass into ethanol.
Applied Biochemistry and Biotechnology
We evaluated various agricultural lignocellulosic biomass and variety of fungi to produce cellulolytic enzymes cocktail to yield high amount of reducing sugars. Solid-state fermentation was performed using water hyacinth, paddy straw, corn straw, soybean husk/tops, wheat straw, and sugarcane bagasse using fungi like Nocardiopsis sp. KNU, Trichoderma reesei, Trichoderma viride, Aspergillus flavus, and Phanerochaete chrysosporium alone and in combination to produce cellulolytic enzymes. Water hyacinth produced (U ml −1) endoglucanase (51.13) and filter paperase (0.55), and corn straw produced (U ml −1) β-glucosidase (4.65), xylanase (113.32), and glucoamylase (41.27) after 7-day incubation using Nocardiopsis sp. KNU. Production of cellulolytic enzymes was altered due to addition of various nitrogen sources, metal ions, vitamins, and amino acids. The maximum cellulolytic enzymes were produced by P. chrysosporium (endoglucanase; 166.32 U ml −1 and exoglucanase; 12.20 U ml −1), and by T. viride (filter paperase; 1.57 U ml −1). Among all, co-culture of T. reesei, T. viride, A. flavus, and P. chrysosporium showed highest β-glucosidase (17.05 U ml −1). The highest xylanase (1129 U ml −1) was observed in T. viride + P. chrysosporium co-culture. This study revealed the dependency on substrate and microorganism to produce good quality enzyme cocktail to obtain maximum reducing sugars.
Cellulases can be used for biofuel production to decrease the fuel crises in the world. Microorganisms cultured on lignocellulosic wastes can be used for the production of cellulolytic enzymes at large scale. In the current study, cellulolytic enzyme production potential of Aspergillus fumigatus was explored and optimized by employing various cultural and nutritional parameters. Maximum endoglucanase production was observed after 72 h at 55 °C, pH 5.5, and 70 % moisture level. Addition of 0.3 % of fructose, peptone, and Tween-80 further enhanced the production of endoglucanase. Maximum purification was achieved with 40 % ammonium sulfate, and it was purified 2.63-fold by gel filtration chromatography. Endoglucanase has 55 °C optimum temperature, 4.8 optimum pH, 3.97 mM K m , and 8.53 μM/mL/min V max. Maximum exoglucanase production was observed at 55 °C after 72 h, at pH 5.5, and 70 % moisture level. Further addition of 0.3 % of each of fructose, peptone, and Tween-80 enhances the secretion of endoglucanase. It was purified 3.30-fold in the presence of 40 % ammonium sulfate followed by gel filtration chromatography. Its optimum temperature was 55 °C, optimum pH was 4.8, 4.34 mM K m , and 7.29 μM/mL/min V max. In the case of β-glucosidase, maximum activity was observed after 72 h at 55 °C, pH 5.5, and 70 % moisture level. The presence of 0.3 % of fructose, peptone, and Tween-80 in media has beneficial impact on β-glucosidase production. A 4.36-fold purification was achieved by 40 % ammonium sulfate precipitation and gel filtration chromatography. Optimum temperature of β-glucosidase was 55 °C, optimum pH was 4.8, K m was 4.92 mM, and V max 6.75 μM/mL/min. It was also observed that fructose is better than glucose, and peptone is better than urea for the growth of A. fumigatus. The K m and V max values indicated that endoglucanase, exoglucanase, and β-glucosidase have good affinity for their substrates.
International Journal of Environment, Agriculture and Biotechnology, 2021
The use of the I-Optimal mixture design technique of agro-industrial residues in cultivation submerged at 28°C for 15 days with Trichoderma reesei QM 9414, complemented with nutrients, was used to optimize the mixture for the production of fibrolytic enzymes. The results demonstrated that the use of 100% (m/v) of brewer's spent grain was promising for the production of total cellulases (0.42 FPU/mL) and xylanase (39.60 U/mL), as well as the use of 33.3% citrus pulp and 66.7% brewer's spent grain for the production of xylanase (40.2 U/mL). The combination of 16.67% wheat bran, 16.67% citrus pulp, and 66.7% brewer's spent grain was the most promising for the production of endoglucanase (2.03 U/mL), exoglucanase (3.20 U/mL), and β-glycosidase (0.12 U/mL). The study on the demand for minerals, sucrose, and yeast extract (as a vitamin and amino acid source) revealed that 0.1% yeast extract, 0.11% dibasic potassium phosphate, 0.0028% zinc, and 1% of sucrose in 12 days of culture were sufficient to maximize the production of cellulases, increasing by 2.38 times (1.0 FPU/mL) compared to the initial culture (0.42 FPU/mL). Cellulolytic production remained the same with the use of 0.01% tween 80 in citrus pulp (0.40 FPU/mL) compared to that obtained in the design with a brewer's spent grain without tween 80, however it reduced substantially (from 15 to 9 days) the cultivation time. On the other hand, the use of tween 80 dramatically inhibited the fungal production of xylanases (2.96 U/mL). The best combination of salts was combined with tween 80 to obtain 1.12 FPU / mL in 9 days of fermentation. An enzymatic hydrolysis of cassava bagasse was carried out by combining cellulases and amylases, reaching 48 g / L of reducing sugar. Thus, this work shows that by studying the influence of residues, kind of salts and concentration of tween 80, a more efficient and economical bioprocess was possible to obtain, as well as the association between fibrolytic enzymes.
Comparative study of saccharification of biomass by various cellulolytic enzymes
In this paper a comparative study of saccharification of cellulose-rich switchgrass biomass by various enzyme preparations (Accelerase 1500, GC 220, NZ50013 and CTec 2) has been carried out. Furthermore, an effect of supplementary β-glucosidase on activity of enzymes and yield of sugars has been studied. It was found that adding of β-glucosidase to enzyme preparations improves cellulolytic activity and increases the yield of glucose after enzymatic hydrolysis of the cellulosic biomass. In order to evaluate the yield of glucose (Y) from cellulosic substrate after its cleavage by different enzymatic preparations, a combined parameter of cellulolytic activity, P=L2/D, has been proposed; where L is used level of cellulolytic activity expressed in FPU per 1g substrate; and D is used doze of the enzyme expressed in mg proteins per 1g substrate. There is a directly proportional dependence between yield of glucose and combined parameter of cellulolytic activity. Due to high squared correlation coefficient (R2 = 0.934), the regression equation Y = k P can be used to predict the yield of glucose from the biomass hydrolyzed by different enzyme preparations.