Abstract
Plastic is a ubiquitous material abundant in everyday human life due to its high convenience and low cost. However, over-production and the incorrect disposal of plastic has resulted in the once-revolutionary material becoming a destructive environmental pollutant. Consequently, there has been an increased demand for readily biodegradable and environmentally friendly alternatives. Lactic acid (LA) and polyhydroxyalkanoates (PHA) are two microbially produced molecules that have been identified to have the potential of forming plastics, termed bioplastics, with faster degradation and lower environmental damage. Polylactic acid (PLA) is manufactured from LA monomers, which can be derived from bacterial metabolism. PHA is synthesised in PHA-accumulating bacteria as an energy source in stressful conditions, which can be extracted for bioplastic manufacturing. The aim of this study was to identify bacteria with high LA yield, particularly in the D- enantiomer, that can be utilised for commercial-scale production of LA, and ultimately bioplastics. This study focused on five Lactobacillus species: L. plantarum R0202, L. delbrueckii NCDO1489, L. delbrueckii R1087, L. acidophilus 1899, and L. reuteri DSM20016. Here the LA production and cell viability of these five isolates in various environmental conditions and different substrate compositions is reported. It was observed that all five strains grew equally well under anaerobic and aerobic conditions (with 5% CO2). Growth of the selected strains was substrate limited up to 20 g/L glucose, and was then further limited by acidic pH, due to the high LA concentrations produced by these isolates. In an attempt to overcome this limitation, adaptive evolution was used to produce low pH- adapted mutants (AK4.5 and AK3.7) of L. plantarum R0202. Whole genome sequencing was used to identify the mutation points in AK4.5 and AK3.7, relative to the wild-type. In the second part of the thesis, the growth kinetics and PHA production of Cupriavidus necator in strains DSM531 and DSM545 in nutrient broth and fructose-nitrogen minimal media was investigated. Nile red staining and transmission electron microscopy was used to visualise and quantify PHA produced in strains DSM531 and DSM545. Overall, the study presents good candidates for LA and PHA production, forming the basis for further studies to optimise their use for bioplastic production.