Thesis (PhD)--Stellenbosch University, 2008. / ENGLISH ABSTRACT: This study strives to improve two current industrial processes by making them more cost effective
through the use of hydrolytic enzymes or microbial systems. The first process targeted is the
industrial conversion of starch to ethanol. In the second process, hydrolytic enzymes are applied to
the manufacturing of instant coffee.
The engineering of microbial systems to convert starch to bio-ethanol in a one-step process may
result in large cost reductions in current industrial processes. These reductions will be due to
decreased heating energy requirements, as well as a decrease in money spent on the purchase of
commercial enzymes for liquefaction and saccharification. In this study, a recombinant
Saccharomyces cerevisiae strain was engineered to express the wild-type Aspergillus awamori
glucoamylase (GA I) and α-amylase (AMYL III) as well as the Aspergillus oryzae glucoamylase
(GLAA) as separately secreted polypeptides. The recombinant strain that secreted functional GA I
and AMYL III was able to utilise raw corn starch as carbon source, and converted raw corn starch
into bio-ethanol at a specific production rate of 0.037 grams per gram dry weight cells per hour. The
ethanol yield of 0.40 gram ethanol per gram available sugar from starch translated to 71% of the
theoretical maximum from starch as substrate. A promising raw starch converter was therefore
generated.
In the second part of this study, soluble solid yields were increased by hydrolysing spent coffee
ground, which is the waste generated by the existing coffee process, with hydrolytic enzymes.
Recombinant enzymes secreted from engineered Aspergillus strains (β-mannanase, β-endoglucanase
1, β-endo-glucanase 2, and β-xylanase 2), enzymes secreted from wild-type organisms
(β-mannanases) and commercial enzyme cocktails displaying the necessary activities
(β-mannanase, cellulase, and pectinase) were applied to coffee spent ground to hydrolyse the
residual 42% mannan and 51% cellulose in the substrate. Hydrolysis experiments indicated that an
enzyme cocktail containing mainly β-mannanase increased soluble solids extracted substantially,
and a soluble solid yield of 23% was determined using the optimised enzyme extraction process.
Soluble solid yield increases during the manufacturing of instant coffee will result in; (i) an increase
in overall yield of instant coffee product, (ii) a decrease in amount of coffee beans important for the
production of the product, and (iii) a reduction in the amount of waste product generated by the
process. / AFRIKAANSE OPSOMMING: Hierdie studie poog om twee huidige industriële prosesse te verbeter deur die prosesse meer kosteeffektief
met behulp van hidroltiese ensieme en mikrobiese sisteme te maak. Die eerste industrie
wat geteiken word, is die omskakeling van rou stysel na etanol, en die tweede om hidrolities
ensieme in die vervaardiging van kitskoffie te gebruik.
Die skep van mikrobiese sisteme om rou-stysel in ’n ’een-stap’ proses om te skakel na bio-etanol
sal groot koste besparing tot gevolg hê. Hierdie besparings sal te wyte wees aan die afname in
verhittingsenergie wat tydens die omskakelingsproses benodig word, asook ’n afname in die koste
verbonde aan die aankoop van duur kommersiële ensieme om die stysel na fermenteerbare suikers
af te breek. In hierdie studie is ’n rekombinante Saccharomyces cerevisiae-gis gegenereer wat die
glukoamilase (GA I) and α-amilase (AMYL III) van Aspergillus awamori, asook die glukoamilase
van Aspergillus oryzae (GLAA) as aparte polipeptide uit te druk. Die rekombinante gis wat die
funksionele GA I en AMYL III uitgeskei het, was in staat om op die rou-stysel as koolstofbron te
groei, en het roustysel na bio-etanol teen ’n spesifieke tempo van 0.037 gram per gram droë gewig
biomassa per uur omgeskakel. Die etanolopbrengs van 0.40 gram per gram beskikbare suiker vanaf
stysel was gelykstaande aan 71% van die teoretiese maksimum vanaf stysel as substraat. ’n
Belowende gis wat roustysel kan omskakel na bio-etnaol was dus geskep.
In die tweede deel van hierdie studie is die opbrengs in oplosbare vastestowwe vermeerder deur die
koffie-afval wat tydens die huidige industrieële proses genereer word, met hidrolitiese ensieme te
behandel. Rekombinante ensieme afkomstig vanaf Aspergillus-rasse (β-mannanase,
β-endoglukanase 1, β-endo-glukanase 2 en β-xilanase 2), ensieme deur wilde-tipe organismes
uitgeskei (β-mannanase), asook kommersiële ensiempreparate wat die nodige ensiemaktiwiteite
getoon het (β-mannanase, sellulase en pektinase) is gebruik om die oorblywende 42% mannaan en
51% sellulose in koffie-afval te hidroliseer. Hidrolise eksperimente het getoon dat ’n
ensiempreparaat wat hoofsaaklik mannanase bevat, die oplosbare vastestofopbrengs grootliks kan
verbeter, met ’n verhoogde opbrengs van 23% tydens geöptimiseerde ensiembehandelings. ’n
Verhoogde opbrengs in oplosbare vastestowwe tydens die vervaardiging van kitskoffie sal die
volgende tot gevolg hê: (i) ’n toename in totale opbrengs van kitskoffie produk, (ii) ’n afname in die
hoeveelheid koffiebone wat vir die produksie ingevoer moet word, en (iii) ’n afname in die
hoeveelheid afval wat tydens die vervaardigingsproses produseer word.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/21457 |
| Date | 03 1900 |
| Creators | De Villiers, Tania |
| Contributors | Van Zyl, W. H., Gorgens, J. F., Stellenbosch University. Faculty of Science. Dept. of Microbiology. |
| Publisher | Stellenbosch : Stellenbosch University |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | English |
| Type | Thesis |
| Format | xvii, 208 leaves : ill. |
| Rights | Stellenbosch University |
Page generated in 0.002 seconds