Production of Bioethanol from Brewery Spent Grain Using Cellulase Enzyme Hydrolysis
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Date
2018-07
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Addis Ababa University
Abstract
Bioethanol is a clear, colorless and mobile flammable liquid chemical compound with acceptable
odour, and produced from biomass feed stocks through fermentation. The objective of this study
was production of bioethanol from Barley spent grain by using cellulase enzyme hydrolysis.
Decreasing of world petroleum reserves, fluctuation of petroleum price and environmental
concerns, low yield cellulose acid hydrolysis and formation of inhibitory were the problem
statement. Enzymatic hydrolysis method was used for cellulose hydrolysis, producing monomeric
C6 sugars (glucose); enzymatic hydrolysis is more specific and high conversion rates can be
achieved in long reaction times obtaining very high yields. Raw material characterization of BSG
were, hemicelluloses content 28.4 %, cellulose content 27 %, and dry matter content 98.2 %. The
aim of pretreatment is to disrupt recalcitrant structures of cellulosic biomass to make cellulose
more accessible to the enzymes that convert carbohydrate polymers into fermentable sugars. The
pretreated feedstock was mixed with 1.25 % (v/v) sulfuric acid solutions with a liquid-to-solid ratio
of 20 %( w/w) and it was occurred on the temperature 121 ºC and time of 17 min, the severity
factor coefficient was 0.198. Cellulase enzyme was produced from BSG using bacillus subtilus,
under liquid state fermentation for 5 days at temperature of 37 ºC
with pH of 5.0, in controlled
incubator. For the enzyme hydrolysis, 26 full factorial central composite design (CCD) was
applied to investigate the effect of temperature (40-50 ºC), pH (4.0-5.0), reaction time (24-72 hr.)
and enzyme loading (1-2 %) using Design expert® 6 software. RSM was applied to investigate the
interaction effect of hydrolysis process variables on the yield of ethanol from BSG. After hydrolysis
process sugar content of the hydrolysate was quantified using spectrophotometer measuring its
absorbance. Fermentation of the hydrolysate were performed 2.5 % Saccharomyces cerevisiae
and 2.5 % fusarium oxysporum at 30 ºC temperature, pH 5.0 and 72 hr. fermentation time for all
samples. Significance of the process variables were analyzed using analysis of variance. As the
result of RSM optimization, the best yield of TRS was found at 40 ºC hydrolysis temperature, 4.5
PH, 48 hr. reaction time and 1.5 % v/v enzyme loading. Under these condition 96.55 %w/w and
53.68 % per 6.58 g dry barley spent grain of TRS and bio-ethanol respectively obtained.
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Keywords
Bioethanol, Barley spent grains, bacillus subtilus, cellulase, enzyme hydrolysis, fermentation, fusarium oxysporum, Saccharomyces cerevisiae