ENGLISH ABSTRACT: The need to mitigate the twin crises of peak oil and climate change has driven a headlong
rush to biofuels. This study was aimed at the development of a process to efficiently
convert steam explosion pretreated (STEX) sugarcane bagasse into ethanol by using
combinations of commercial enzyme cocktails and recombinant Saccharomyces
cerevisiae strains. Though enzymatic saccharification is promising in obtaining sugars
from lignocellulosics, the low enzymatic accessibility of the cellulose and hemicellulose
is a key impediment thus necessitating development of an effective pretreatment scheme
and optimized enzyme mixtures with essential accessory activities. In this context, the
effect of uncatalysed and SO2 catalysed STEX pretreatment of sugarcane bagasse on the
composition of pretreated material, digestibility of the water insoluble solids (WIS)
fraction and overall sugar recovery was investigated. STEX pretreatment with water
impregnation was found to result in a higher glucose recovery (28.1 g/ 100 bagasse) and
produced WIS with a higher enzymatic digestibility, thus was used in the optimization of
saccharification and fermentation. Response surface methodology (RSM) based on the 33
factorial design was used to optimize the composition of the saccharolytic enzyme
mixture so as to maximize glucose and xylose production from steam exploded bagasse.
It was established that a combination of 20 FPU cellulase/ g WIS and 30 IU
-glucosidases/ g WIS produced the highest desirability for glucose yield. Subsequently
the optimal enzyme mixture was used to supplement enzyme activities of recombinant
yeast strains co-expressing several cellulases and xylanases in simultaneous
saccharification and fermentations SSFs. In the SSFs, ethanol yield was found to be
inversely proportional to substrate concentration with the lowest ethanol yield of 70%
being achieved in the SSF at a WIS concentration of 10% (w/v). The ultimate process
would however be a one-step “consolidated” bio-processing (CBP) of lignocellulose to
ethanol, where hydrolysis and fermentation of polysaccharides would be mediated by a
single microorganism or microbial consortium without added saccharolytic enzymes. The
cellulolytic yeast strains were able to autonomously multiply on sugarcane bagasse and
concomitantly produce ethanol, though at very low titres (0.4 g/L). This study therefore
confirms that saccharolytic enzymes exhibit synergism and that bagasse is a potential substrate for bioethanol production. Furthermore the concept of CBP was proven to be
feasible.
AFRIKAANSE OPSOMMING: Die behoefte om die twee krisisse van piek-olie en klimaatsverandering te versag, het
veroorsaak dat mense na biobrandstof as alternatiewe energiebron begin kyk het. Hierdie
studie is gemik op die ontwikkeling van 'n proses om stoomontplofde voorafbehandelde
(STEX) suikerriet bagasse doeltreffend te omskep in etanol deur die gebruik van
kombinasies van kommersiële ensiem mengsels en rekombinante Saccharomyces
cerevisiae stamme. Alhoewel ensiematiese versuikering belowend is vir die verkryging
van suikers vanaf lignosellulose, skep die lae ensiematiese toeganklikheid van die
sellulose en hemisellulose 'n hindernis en dus is die ontwikkeling van' n effektiewe
behandelingskema en optimiseerde ensiemmengsels met essensiële bykomstige
aktiwiteite noodsaaklik. In hierdie konteks, was die effek van ongekataliseerde en SO2
gekataliseerde stoomontploffing voorafbehandeling van suikerriet bagasse op die
samestelling van voorafbehandelde materiaal, die verteerbaarheid van die (WIS) breuk
van onoplosbare vastestowwe in water (WIS), en die algehele suikerherstel ondersoek.
Daar was bevind dat stoomontploffing behandeling (STEX) met water versadiging lei tot
'n hoër suikerherstel (21.8 g/ 100g bagasse) en dit het WIS met ‘n hoër ensimatiese
verteerbaarheid vervaardig en was dus gebruik in die optimalisering van versuikering en
fermentasie. Reaksie oppervlak metodologie (RSM), gebasseer op die 33 faktoriële
ontwerp, was gebruik om die samestelling van die ‘saccharolytic’ ensiemmengsel te
optimaliseer om sodoende die maksimering van glukose en ‘xylose’ produksie van
stoomontplofde bagasse te optimaliseer. Daar was bevestig dat ‘n kombinasie van 20
FPU sellulase/ g WIS en 30 IU ‘ -glucosidases/ g’ WIS die hoogste wenslikheid vir
glukose-opbrengs produseer het. Daarna was die optimale ensiemmengsel gebruik om
ensiemaktiwiteit van rekombinante gisstamme aan te vul, wat gelei het tot die medeuitdrukking
van verskillende ‘cellulases’ en ‘xylanases’ in gelyktydige versuikering en
fermentasie SSFs. In die SSFs was daar bevind dat die etanol-produksie omgekeerd
proporsioneel is tot substraat konsentrasie, met die laagste etanolopbrengs van 70% wat
bereik was in die SSF by ‘n WIS konsentrasie van 10% (w/v). Die uiteindelike proses sal
egter 'n eenmalige "gekonsolideerde" bioprosessering (CBP) van lignosellulose na etanol
behels, waar die hidrolise en fermentasie van polisakkariede deur' n enkele mikroorganisme
of mikrobiese konsortium sonder bygevoegde ‘saccharolytic’ ensieme bemiddel sal word. Die ‘cellulolytic’ gisstamme was in staat om vanself te vermeerder op
suikerriet bagasse en gelyktydig alkohol te produseer, al was dit by baie lae titres (0.4
g/L). Hierdie studie bevestig dus dat ‘saccharolytic’ ensieme sinergisme vertoon en dat
bagasse 'n potensiële substraat is vir bio-etanol produksie. Daar was ook onder meer
bewys dat die konsep van CBP uitvoerbaar is.
