Structural changes of lignocelluloses by physico-chemical treatment and improving saccharification using cellulose microfibril swelling (Cms) enzyme
Structural changes Physico-chemical treatment cellulose microfibril swellase enzyme wood cellulose;
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In this study, reed straw structural changes during mild alkali pretreatment were evaluated. Therefore, various concentrations of NaOH (1% to 2.5%) were used for pretreatment of reed straw at 105°C for 10 min. Scanning electron microscopy, atomic force microscopy, and Fourier transform infrared spectroscopy studies showed that 2% and 2.5% NaOH pretreated sample exposed more cellulose fibers compared to other treatments. The cellulose crystalline index was increased by the 1% to 2% NaOH-treatments and slightly lowered by the 2.5% NaOH-treatment due to destructing cellusose fibres. 2% NaOH pretreatment caused 69.9% lignin removal while 2.5% NaOH pretreatment removed 72.4% lignin. Besides, reed straw, when pretreated at 2% and 2.5% NaOH, resulted 56.4% and 60.5% hemicellulose removal, respectively. However, the difference in removal of lignin and hemicellulose between 2% and 2.5% NaOH treated reed straw was very marginal. In addition, very negligible increase of cellulose level was estimated, amounting 78.8% and 76.6% in 2.5% and 2% NaOH treated sample, respectively. Moreover, after 72 h, reducing sugar yield was 81.2% and 83.3% using enzyme loading of 15 FPU (g dry biomass)- 1 and 30 IU (g dry biomass)- 1 and xylanase 4 FXU (g dry biomass)- 1 from 2% and 2.5% NaOH pretreated reed straw, respectively. Reducing sugar yield was increased very marginally when NaOH concentration increased from 2% to 2.5% for reed straw pretreatment. Therefore, 2% NaOH is supposed to be effective for reed straw pretreatment at this mentioned condition. Secondly, woody biomass attracts the attention as wood is the bulk source of lignocellulosic materials. In near future it might be the staple substrate in the biofuel field. Therefore, we pretreated pine wood using NaOH and used Congo red (CR) for chosing the pretreatment condition. Actually this is the new approach may be helpful in bioethanol research. The basis of this study was that cellulose free -OH groups can bind with CR while in contact with CR solution. Therefore, structural change of pine wood caused by different NaOH (0 to 20%) treatment at 130°C for 30 min was evaluated in terms of Congo red adsorption. It was observed that 10% NaOH treated sample adsorb the highest CR. This might be due to the more cellulose exposure compared to other treatments. Scanning electron microscopy, and Fourier transform infrared spectroscopy studies showed that 10% NaOH pretreated sample exposed more cellulose fibers compared to other treatments leading to more CR binding. The cellulose crystalline index was increased with increasing NaOH-treatments and lowered by the 20% NaOH-treatment due to degradation of cellusose fibres. Moreover, after 72 h, reducing sugar yield was 76.5% and 70.7% using enzyme loading of of 15 FPU (g dry biomass)- 1 and 30 IU (g dry biomass)- 1 from 10% and 20% NaOH pretreated pine wood samples, respectively. Reducing sugar yield was decreased from samples while treating more than 10% NaOH solution. Therefore, 10% NaOH treatment can be considered as an effective concentration for pine wood pretreatment at this mentioned condition, evaluated by CR adsorption approach. Finally, in this study, hydrothermally pretreated pine wood saccharification was enhanced using cellulose microfibril swelling (Cms) enzyme from Bacillus sp. AY8. For doing so, pine wood were pretreated hydrothermally at 150°C, 170°C and 190°C for 20, 40 and 60 min. Among the samples, the 170°C and 40 min pretreated samples showed suitable amount of cellulose and adsorb substantial amount of Congo red. Therefore, this sample was selected for subsequent Cms enzyme treatment. Bacillus sp. AY8 crude enzyme [100 µg (0.01g sample)-1] treatment causes disruption of hydrogen bond in pine wood cellulose leading to more space for cellulase enzyme binding. The application of Bacillus sp. AY8 crude enzyme increased the reducing sugar yield from 380 mg g-1 of cellulose to 590 mg g-1 of wood cellulose, yield increased by almost 1.5 fold. This might be due to the Cms enzyme which reduces the crystallinity of cellulose materials by disrupting hydrogen bonds. The disruption of hydrogen bond was evidenced by FT-IR analysis. Therefore, Cms enzyme from Bacillus sp. AY8 is helpful for boosting up enzymatic saccharification and producing bioethanol precursor.