Cellulase production by thermophilic Bacillus licheniformis 2D55 and its application for recovery of fermentable sugar from rice husk
Fermentable sugar production from various agro-waste biomass via enzymatic saccharification using mesophilic fungi cellulase has been the issue of interest for many researchers. However, research carried out using thermophilic bacterial cellulase for sugar production are limited. Cellulases, chem...
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| Format: | Thesis |
| Language: | English |
| Published: |
2018
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| Online Access: | http://psasir.upm.edu.my/id/eprint/68510/1/FBSB%202018%204%20IR.pdf http://psasir.upm.edu.my/id/eprint/68510/ |
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| Summary: | Fermentable sugar production from various agro-waste biomass via enzymatic
saccharification using mesophilic fungi cellulase has been the issue of interest for
many researchers. However, research carried out using thermophilic bacterial
cellulase for sugar production are limited. Cellulases, chemical pretreatment and
enzymatic saccharification required for sugar production are very costly and often
result in low sugar yield, thereby affecting the overall economics of the bioconversion
process. Hence, the main aim of this study was to isolate, characterize and enhance
the production of cellulolytic enzyme complex (CMCase, FPase and β-glucosidase)
and Xylanase from agro-waste mixture (a mixture of untreated sugarcane bagasse and
pretreated rice husk) by a locally isolated thermophilic Bacillus licheniformis 22D55
and utilize the crude thermostable cellulase enzyme for sugar production through
enzymatic saccharification of rice husk subjected to high pressure steam pretreatment.
Cellulose degrading bacterium isolated from oil palm empty fruit bunch-chicken
manure compost was identified by morphological, biochemical and 16S rRNA test
and known as B. licheniformis 2D55. In basal medium with microcrystalline cellulose
as carbon source, the bacterium produced cellulase at 50C within 18-24 h. Among
the various untreated and sodium hydroxide (NaOH) pretreated agro-waste biomass
used, agro-waste mixture comprising of untreated sugarcane bagasse and pretreated
rice husk results in overall improvement in the cellulolytic enzyme complex with
CMCase at 0.37 U/mL, FPase at 0.29 U/ml, β-glucosidase at 0.006 U/mL and xylanase
at 0.98 U/mL. The SEM image reveal deformity in bacteria cell grown on NaOH
pretreated sugarcane bagasse only, which resulted in the low performance of the
bacteria for cellulase production on the substrate.
Effect of nutritional and physicochemical factors were investigated for enhancing
cellulase production. The CMCase, FPase, and β-glucosidase activities increased by
77.4-folds, 44.3-folds, and 10-folds, respectively. The crude enzyme was highly active
and stable over broad temperature (50 to 80°C) and pH (3.5 to 10.0) ranges with
optimum temperature at 65°C and 80ºC for CMCase and FPase, respectively. While
the optimum pH for CMCase and FPase was 7.5 and 6.0, respectively.
An operational condition was developed for high pressure steam pretreatment (HPSP)
of rice husk for fermentable sugar recovery through enzymatic saccharification. The
pretreatment at 200°C/1.85 MPa for 7 min was found to effectively enhance cellulose
content of the rice husk. The scanning electron micrograph (SEM), fourier transform
infrared (FTIR) and x-ray diffraction (XRD) analysis expressed effectiveness of the
pretreatment. A two-step saccharification of pretreated rice husk at 60°C yielded
reducing sugar at 0.581 g/g substrate that was equivalent to 73.5% saccharification.
Therefore, the strain B. licheniformis 2D55 has the potential of utilizing agro-waste
mixture for higher cellulolytic enzyme production. The thermostable nature of the
cellulase contributed to the efficient release of fermentable sugars at higher
temperature. The application of high pressure steam pretreatment and two-step
enzymatic saccharification provides greener technology and improve the fermentable
sugar production. The results of this study could contribute to future research in
thermophilic bioprocessing rice husk with the application of thermostable cellulase
producing bacterium for improving biomass-sugar conversion. |
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