Enzymatic hydrolysis of oil palm biomass for fermentable sugar using polyethylene glycol immobilized cellulase

In this work, enzymatic hydrolysis using cellulase both in solution and immobilized form was studied to convert lignocellulosic biomass from empty fruit bunch into fermentable sugars. The cellulase was covalently immobilized with activated and functionalized polyethylene glycol via glutaraldehyde coupling. To determine sample enzyme activity, the equivalent reducing sugars released during hydrolysis reaction with free cellulase and immobilized cellulase respectively, were quantified using 3,5- dinitrosalicylic acid (DNS) method. As a whole, the immobilized cellulase displayed 50% higher efficiency over free cellulase, in reducing sugar recovery during hydrolysis reactions. From the kinetic study, it showed that Michaelis constant (Km) and limiting velocity(Vm«) of immobilized cellulase were 179.2 mg/ml and 33.5mg/ml.min respectively, whereas that of free cellulase were 171.8mg/ml and 34.5mg/ml.min respectively. The higher Km value of immobilized cellulase could be attributed to the polyethylene glycol interference with the binding of cellulase to expose substrate, and enables free interaction of cellulase to hydrolyse cellulose maximally

A kinetic study of enzymatic hydrolysis of oil palm biomass for fermentable sugar using polyethylene glycol (PEG) immobilized cellulase

In this work, enzymatic hydrolysis by cellulase in a soluble and an immobilized form was studied to convert lignocellulosic oil palm empty fruit bunch (EFB) biomass into fermentable sugars as a feedstock for bioethanol production. The cellulase was covalently immobilized with activated and functionalized polyethylene glycol (PEG) via glutaraldehyde coupling method. As a whole, the immobilized cellulase displayed 50 higher efficiency over free cellulase, in reducing sugar recovery during hydrolysis reactions at pH of 4.8 and temperature of 50°C. From the kinetic study, it showed that Michaelis constant (Km) and limiting velocity (Vmax) of immobilized cellulase were 179.2 mg/ml and 33.5 mg/ml.min respectively, comparable with the value for free cellulose, 171.8 mg/ml and 34.5 mg/ml.min respectively. This result could be attributed to the effect of PEG on the binding cellulase to substrate desorb substrates, and enables free interaction of cellulase to hydrolyse cellulose maximally

Effect of immobilized cellulase enzyme treatment on properties of ramie fabric

In this study, Eudragit S-100 has been covalently bound to the cellulase enzyme to form immobilized cellulase enzyme and then the effect of the treatment on ramie fabric properties is studied. The ramie fabrics treated with immobilized cellulase enzyme show lower quantities of reducing sugar, weight loss, and higher tensile strength than native cellulase enzyme-treated fabrics. Scanning electron microscopic analysis shows that the surface of ramie fabrics treated with cellulase enzyme is smoother than that of the untreated sample. Furthermore, treatment by the immobilized cellulase enzyme is less damaging to the fibres. X-ray diffraction studies show that there is hardly any loss in the crystallinity of ramie fabrics. Low-stress mechanical properties evaluated by the Kawabata Evaluation System for Fabric indicate that immobilized cellulase enzyme treatment improves the softness, flexibility, and elastic recovery of the ramie fabrics

Immobilization of cellulase and yeast for the hydrolysis and fermentation of pre-treated bagasse for ethanol production

Lignocellulose ethanol promises to be the cheapest form of fuel, however, the drawback in the production is in the pretreatment process to remove lignin and the efficient hydrolysis of free cellulose. This research work is designed to delignify sugarcane bagasse, hydrolyze and ferment it with immobilized cellulase from the snail gut isolates and yeast respectively. The biomass were pretreated with Ca(OH)2 and then placed in the water-bath with temperature of 200C, 400C, 600C, 800C,1000C and 1200C. The pretreated biomass was hydrolysed with free and immobilized cellulase at 500C for 5-48hrs. The activity, optimum pH, optimum temperature, substrate concentration profile and kinetic parameters, Vmax and Km of cellulase were also determined. The optimum pH for free and immobilized cellulase ranged from 4.0-5.5 and optimum temperature was recorded at 450C and 550Cfor free and immobilized cellulase respectively. The effect of temperature on both free and immobilized cellulases showed that immobilized cellulase has higher resistance to temperature than the free cellulase. Also the yield of glucose (40mg/ml) was higher with immobilized enzyme after 24hrs. The results obtained has also shown that immobilized cellulase has a higher Km when compared with free cellulase The maximum reaction rate (Vmax) obtained from Michaelis Menten plots was lower for immobilized cellulase than for the free enzyme. Higher value of Vmax for free enzyme indicated that the enzyme converted more substrate to product per unit time upon saturation with substrate. The biomass was fermented for 48hrs with immobilized Saccharomyces cerevisiae and the results showed the ethanol yield of 31.75% at 24hrs and 70.84% at 48hrs. The initial glucose concentration was 40mg/ml and this significantly reduced to 6.21mg/ml after 24hrs and 1.25mg/ml after 48hrs of the fermentation process. These results showed a proportional increase in ethanol yield against a depleting concentration of glucose which is being used up in the fermentation reaction revealing the maximum efficiency of the immobilized yeast cells. In this study, it has shown that the entrapped cellulase cells produced high levels of reducing sugars in hydrolysis compared with their native counterparts and immobilized yeast cells also gave a high yield of ethanol. The immobilization process therefore obtained more thermostable biocatalysts with increased productivity which is more economical for biofuel production.Keywords: Immobilization, Cellulase, Saccharomyces cerevisiae, biocatalysts, delignificatio

Cellulase immobilization on superparamagnetic nanoparticles for reuse in cellulosic biomass conversion

Current cellulosic biomass hydrolysis is based on the one-time use of cellulases. Cellulases immobilized on magnetic nanocarriers offer the advantages of magnetic separation and repeated use for continuous hydrolysis. Most immobilization methods focus on only one type of cellulase. Here, we report co-immobilization of two types of cellulases, β-glucosidase A (BglA) and cellobiohydrolase D (CelD), on sub-20 nm superparamagnetic nanoparticles. The nanoparticles demonstrated 100% immobilization efficiency for both BglA and CelD. The total enzyme activities of immobilized BglA and CelD were up to 67.1% and 41.5% of that of the free cellulases, respectively. The immobilized BglA and CelD each retained about 85% and 43% of the initial immobilized enzyme activities after being recycled 3 and 10 times, respectively. The effects of pH and temperature on the immobilized cellulases were also investigated. Co-immobilization of BglA and CelD on MNPs is a promising strategy to promote synergistic action of cellulases while lowering enzyme consumption

Immobilization of cellulase for large scale reactors to reduce cellulosic ethanol cost

Cellulosic ethanol is an alternative renewable energy source. Cellulase used in the production of cellulosic ethanol is very expensive. The difficulty in separating cellulase from the cellulose solution after the hydrolysis process limits the reusability of the cellulase, which highly precludes the scales of this application because of the high cost of the enzyme. Immobilization of cellulase provides a promising approach to allow the enzyme to be recycled, thus reducing the production cost. This research focused on immobilizing cellulase for reuse to reduce the cellulosic ethanol cost. Four immobilization techniques were explored for the immobilization of cellulase on four different immobilization carriers. The immobilized cellulases by Layer-by-Layer Nano-Assembly (LbL) and Ca2+-Al(OH)X modification methods had high initial activities but low reusabilities. Enzyme desorption was observed during the hydrolysis of cellulose solutions by the immobilized cellulases for both LbL and Ca2+-Al(OH)x modification methods. Efforts were focused on improving the reusability of the immobilized cellulase, yet rarely worked. For the immobilized cellulase by the combination of Ca2+-Al(OH)x modification and LbL, the reusability of the immobilized cellulase was improved with the number of enzyme layers. Unfortunately, for the present moment, the initial activity decreased with the number of enzyme layers. Immobilization of cellulase on silica gel by 3-APTES and glutaraldehyde modification showed the highest reusability. No enzyme desorption was observed during the hydrolysis of cellulose solution. It indicated that the cellulase molecules firmly covalent bound to the silica gel. The immobilized cellulase on silica gel by 3-APTES and glutaraldehyde modification had the highest activity per unit mass of immobilization carriers because of the porous structure of the silica gel

Magnetic nickel nanostructure as cellulase immobilization surface for the hydrolysis of lignocellulosic biomass

In this research, a magnetic reusable nickel nanoparticle (NiNPs) supporting materials were prepared for cellulase enzyme immobilization. The immobilized cellulase showed high activity recovery, large & fast immobilization capacity and improved pH & temperature 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

Cellulase Production by Wild-type Aspergillus niger, Penicillium chrysogenum and Trichoderma harzianum Using Waste Cellulosic Materials

Waste cellulosic materials (corncob, sawdust and sugarcane pulp) and crystalline cellulose induced cellulase production in wild strains of Aspergillus niger, Penicillium chrysogenum and Trichoderma harzianum isolated from a wood-waste dump in Lagos, Nigeria. Cellulose-supplemented media gave the maximum cellulase activity of 0.54, 0.67 and 0.39 units mg Protein-1 for A. niger, P. chrysogenum and T. harzianum respectively. The maximum enzyme activity for A. niger was obtained at 36 h of cultivation, while P. chrysogenum and T. harzianum gave their maximum enzyme activities at 12 and 60 h respectively. For the cellulosic wastes, highest enzyme activity was obtained with sawdust where A. niger, P. chrysogenum and T. harzianum gave the maximum enzyme activity of 0.30, 0.24 and 0.20 units mg Protein-1 respectively after 144 h of cultivation. A. niger recorded the highest enzyme activity with any of the three cellulosic materials followed by P. chrysogenum. It thus appears that the use of sawdust presents the best option for low-cost commercial production of cellulase using A. niger and P. chrysogenum as discussed herewith

Immobilization of Cellulase on Zinc Oxide Deposited on Zeolite Pellets for Enzymatic Saccharification of Cellulose

The consumption of fossil fuels to fulfill the global energy demand can cause global warming issues. Renewable energy, i.e., bioethanol, from lignocellulosic biomass, is a promising source of alternative energy to fossil fuels. The conversion of lignocellulosic biomass into bioethanol requires the release of fermentable sugars during the saccharification process using cellulase. However, the utilization of this enzyme on an industrial scale is not feasible due to its difficult separation, instability, and high cost. Here, we present a method for cellulase immobilization on functionalized zinc oxide prepared from either zinc nitrate hexahydrate (ZnO(I)) or zinc acetate dihydrate (ZnO(II)) solutions on zeolite (ZEO) pellets. The immobilized cellulase on ZnO-ZEO structures was characterized by scanning electron microscopy, Xray diffraction spectroscopy, and Fourier transform infrared spectroscopy. The immobilization efficiencies of immobilized cellulase either on ZnO(I)-ZEO or ZnO(II)-ZEO were determined as 58.17 ± 0.75% and 55.51 ± 0.81%, respectively. The immobilized cellulase on ZnO-ZEO was capable of catalyzing microcrystalline cellulose breakdown, releasing reducing sugars. The immobilized cellulase on these structures could be recycled up to four repetitive runs. Based on kinetic data, both the Michaelis constants (Km) and maximum reaction velocity (Vmax) of the immobilized cellulase on the ZnO-ZEO structures were lower than those of free cellulase. This suggests that immobilized cellulase has a higher affinity toward the substrate, but a lower reaction rate than the free enzyme

Influence of compost and digestates on plant growth and health: potentials and limits

Composts can influence soil fertility and plant health. These influences can be positive or negative, depending of the quality of the composts. In order to estimate the potential of Swiss composts to influence soil fertility and plant health, one hundred composts representative of the different composting systems and qualities available on the market were analyzed. The organic substance and the nutrient content of the composts varied greatly between the composts; the materials of origin were the major factor influencing these values. The respiration rate and enzyme activities also varied greatly, particularly in the youngest composts. These differences decreased when the composts become more mature. Maturity, the degradation stage of the organic matter, depended not only on the age of the compost, but also on the management of the process. The Nmineralization potential of compost added to soil showed that a high proportion of young composts immobilized the nitrogen in the soil. Two compost parameters allow to predict the risk of nitrogen immobilization in soil: the NO3- and the humic acids contents. The phytotoxicity of the composts varied very much even in mature composts, showing that the storage of the compost plays a decisive role. While the majority of composts protected cucumber plants against Pythium ultimum, only a few composts suppressed Rhizoctonia solani in basil. With respect to disease suppression, the management of the maturation process seems to play a major role. In field experiments, some biologically immature composts immobilized nitrogen in soil and reduced growth of maize. With additional fertilization, however, it was possible to compensate this effect. Digestates and composts increased the pH-value and the biological activity of soil. These effects were observable also one maize season after compost application. In conclusion, big differences were observed in the quality of composts and digestates, and in their impact on soil fertility and plant health. The management of the composting process seems to influence the quality of the composts to a higher extent than the materials of origin or the composting system. More attention should be paid to biological quality of composts, in order to produce composts with more beneficial effects on crops