Microbial conversion of glycerol to value-added products: 1,3-propanediol and polyhydroxyalkanoates
Date of Issue2016-07-08
School of Civil and Environmental Engineering
Residues and Resource Reclamation Centre
Growing biodiesel market resulted in oversupplied glycerol because crude glycerol was produced as waste stream at the ratio of glycerol-to-biodiesel 1:10 (w/w). To achieve monetary and environmental benefits, conversion of glycerol to value-added products is a promising approach. Therefore, this dissertation focused on the microbial conversion of low value crude glycerol to two commercially viable products: 1,3-propanediol (1,3-PDO) and polyhydroxyalkanoates (PHAs). 1,3-PDO is selected as the main target product, because it is a valuable chemical intermediate widely used in polymer materials and cosmetic, etc. Besides, 1,3-PDO yield in microbial process is high. This provides an attractive solution to oversupplied glycerol. However, almost all isolated 1,3-PDO microbial producers are obligate anaerobes, requiring sterilization and strict anaerobic condition. To simplify the procedure, this study enriched 1,3-PDO producing mixed culture from activated sludge, freshwater sediment, rainforest soil, and mangrove sediment. All tested inoculants produced 1,3-PDO with yields around 0.50 mol/mol, and dominant Gammaproteobacteria and Firmicutes were responsible for 1,3-PDO production. Except 1,3-PDO, carboxylic acids such as acetate, propionate, butyrate, and lactate were produced concurrently. In regard to microbial community, products spectrum, and potential salt tolerance, mangrove sediment was perceived as the best tested inoculant. After anaerobic fermentation, 1,3-PDO is co-produced with carboxylic acids to achieve redox balance and effective approach to separate 1,3-PDO from this mixture is necessary. As the first attempt, the microbial approach to remove carboxylic acids from 1,3-PDO in glycerol anaerobic digestion effluent (ADE) by PHA-producing consortium was investigated. The consortium of B. megaterium and C. hydrocarbooxydans achieved higher cell density on glycerol ADE than single strain. Kinetic study showed the consortium depleted carboxylic acids and converted them into PHAs comprising unit of 3-hydroxybutyrate (3-HB) while preserved over 80% of fed 1,3-PDO. As PHAs are commercially valuable biocompatible and biodegradable polymers with numerous potential industrial applications, this provided a win-win solution to remove carboxylic acids from 1,3-PDO and convert them into PHAs as a secondary value-added product.As an inexpensive and highly reduced carbon source, crude glycerol might fit the niches of high PHAs production cost associated with substrate price and low yield. When glycerol was fed as sole substrate, B. megaterium had the highest cell density among tested strains including P. putida, C. hydrocarbooxydans, B. megaterium, C. glutamicum, C. necator, N. lucida, and B. thuringiensis. PHAs synthesis in B. megaterium occurred in the condition of abundant nitrogen source. Kinetic study showed that for consumed glycerol, 20% was present as unidentified components in liquid while merely 10% was channeled in PHAs. Compared to 1,3-PDO production, aerobic PHAs production from glycerol had a lower target product yield, a slower substrate utilization rate, and a higher biomass yield. Hence, in the scope of this dissertation, anaerobic 1,3-PDO production with subsequent aerobic carboxylic acids removal for PHAs production was deemed as a preferred bioprocess. The dissertation also dedicated to the application of selected bioprocess with pretreated crude glycerol. After pretreatment with acidification to pH 3.0 plus filtration, pretreated crude glycerol had almost identical performance to that of pure glycerol at 20 g/L in serum bottle test. Increasing fed concentration of pretreated crude glycerol from 20 g/L to 100 g/L shifted carboxylic acids spectrum and also changed the relative abundances of Enterobacteria and Clostridia in microbial community. Severe substrate inhibition was observed when fed glycerol concentration was 140 g/L. Coupled with anaerobic process, 20 g/L and 100 g/L glycerol ADE with distinct strengths and constituents were fed to the consortium of B. megaterium and C. hydrocarbooxydans. After depletion of carboxylic acids, carbon contribution from 1,3-PDO over total organic carbon accounted for more than 70% in 20 g/L glycerol ADE and approached to almost 100% in 100 g/L glycerol ADE, respectively. The results showed the pretreated crude glycerol could be applied to the combined anaerobic and aerobic bioprocesses to harvest 1,3-PDO as the main target product and PHAs as the secondary target product.