Bioprocessing of corn stover for bioethanol production
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Date
2022
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Punjab Agricultural University, Ludhiana
Abstract
The present study was conducted with the objective of standardization and evaluation of an
efficient and ecofriendly, method for the pretreatment of corn stover and its subsequent conversion to
bioethanol. The proximate analysis of corn stover (500 µ) revealed a composition of cellulose (57.4%),
hemicellulose (17.5%), lignin (14.4) and ash (2.2%) in the raw corn stover. Among the different chemical
pretreatment methods, acid (1.0% H2SO4) –autoclave (15 psi for 90 min) followed by alkali (2.0% NaOH)
pretreatment resulted in 86.8 % and 79.4% decrease in lignin and hemicellulose, respectively and 66.0%
increase in relative proportion of cellulose. Among the green solvent methods of pretreatment, organosolv
pretreatment methods, acetic acid (40:60, 40%) resulted in 84.6 % and 53.6% decrease in lignin and
hemicellulose, respectively and 53.7% increase in relative proportion of cellulose whereas deep eutectic
solvent pretreatment, choline chloride-lactic acid (1:8 molar, ratio) resulted in 82.5 % and 33.0% decrease in
lignin and hemicellulose, respectively and 69.3% increase in relative proportion of cellulose. For biological
pretreatment of corn stover, a two fungal consortium of strains viz. Pleurotus ostreatus PAU03 and
Phanerochaete chrysosporium MTCC787 was screened for ligninolytic enzyme production by plate assay on
lignin modifying enzyme (LME)- basal medium (LBM), supplemented with 0.01 % (w/v) Azure B and 0.02
% (w/v) Remazole brilliant blue dye and inoculated with agar discs ( 10 mm) of active mycelia.
Decolourization of the respective dyes was observed with ligninolytic index of 1.5 and 1.22 for Azure B and
Remazole brilliant blue dyes, respectively. The consortium culture of two fungal strains viz. P. ostreatus and
P. chrysosporium was used for ligninolytic enzyme production, by using corn stover as substrate. Maximum
enzyme activity (U/ml) was recorded on 10th day as 48.33, 61.85 and 40.8 for LiP, MnP, laccase,
respectively. The enzyme production was scaled upto 4000ml and the crude extract was concentrated (6.7
times) using acetone and 600 ml concentrated enzyme was produced having enzyme activities (U/ml) 68.89,
41.13 and 110.08 for Lacc, LiP and MnP enzymes, respectively. The latter was partially purified by Fast
Performing Liquid Chromatograpgy (FPLC) technique. The enzyme activity (U/ml) of 131.96, 130.38 and
94.21 was recorded for Lacc, LiP and MnP enzymes, respectively in the partially purified enzyme, which
was further concentrated (1.5 times) and the enzyme activities (U/ml) of 140.9, 168.53 and 98.05 for Lacc,
LiP and MnP enzymes, respectively. This partially purified ligninozyme was used for nanoligninozyme
synthesis. In the nanologninozyme (enzyme: sodium:silicate nanohydrate, 1:1) showed the enzyme activities
(U/ml) of 128.93, 187.09 and 116.94 for for Lacc, LiP and MnP enzymes respectively. The pretreatment of
corn stover with nanoligninolzyme under shake flask conditions (50 ml reaction volume) using optimized
physico-chemical parameters viz. corn stover concentration, 2.5 g; enzyme volume, 8.0 ml; Mn2+ ions (0.5
mM) and incubation temperature, 45°C in 72 h of enzymatic action resulted in 87.2% and 67.4% decrease in
lignin and hemicellulose, respectively and 80.3% increase in relative proportion of cellulose. The
saccharification of pretreated corn stover at different concentrations of corn stover (1.0-10mg) with
Arrowzyme (commercial cellulase) at enzyme loading of 30 FPU, resulted in maximum release of reducing sugars (0.396 g/gds) at 2.5g concentration of corn stover. Under these optimized conditions, saccharification
of organosolv (acetic acid, 40:60) and biological (nanoligninozyme) pretreated corn stover resulted in release
of 0.395 and 0.439 g/g, reducing sugars, respectively. The fermentation of organosolv as well as biological
pretreated, and Arrowzyme saccharified corn stover hydrolysate resulted in 0.112 and 0.132 g/gds ethanol,
respectively. The fermentation efficiency of 66.54 % [33.9 % yield (yps)] and 70.91 % [36.1 % yield (yps)]
was recorded for organosolv and biological pretreatment, respectively.
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Citation
Mandeep Kaur (2022). Bioprocessing of corn stover for bioethanol production (Unpublished Ph.D. Dissertation). Punjab Agricultural University, Ludhiana, Punjab, India.