Interest in the aliphatic carbon compound 1,3-propanediol (1,3-PDO) has risen over the past 15 years. In part, this interest is due to the ability of 1,3-PDO to generate a variety of industrially relevant products such as the biodegradable polymer, polytrimethylene terephtalate. Our research group previously reported the identification of a novel Lactobacillus panis PM1 isolate capable of converting glycerol to 1,3-PDO. In this body of work, the effects of various process parameters and the ability of the novel L. panis isolate to produce 1,3-PDO in static and fed-batch cultures were examined. Data collected indicated that the concentrations of glycerol, and glucose, and pH, play a vital role in the optimized production of 1,3-PDO. Optimal conditions for the production of 1,3-PDO were determined to include: i) carbon-limited culture, defined as below 50 mM glucose and ii) growth at 37°C without agitation in the presence of glycerol (150 – 250 mM) at an elevated pH of 9 – 10. Factors such as inoculum size and temperature (OD600 in the range of 0.5 – 2 and a temperature range from 15° - 37°C) in a two-step fermentation showed insignificant variance in the production of 1,3-PDO. Initial fed-batch trials reflected the importance of pH on culture viability. A pH of 8 was determined to be necessary within culture parameters for the fed-batch production of 1,3-PDO. Further, the molar concentrations of 1,3-PDO produced were found to vary only slightly between fed batch culture and a static culture. The variance of 1,3-PDO production between the static and fed-batch trials was found to be 9.1 ± 4.9 mM for an average culture producing 85.3 ± 12.0 mM of 1,3-PDO. However, the mol concentrations of 1,3-PDO produced were found to be significantly higher with 22.3 ± 1.6 versus 5.3 ± 0.7 mmol 1,3-PDO produced for the fed batch versus the static cultures, respectively. The duration of 1,3-PDO production was found to be extended in the fed-batch model of production with increased levels of 1,3-PDO being produced over 120 hours. The cloning and characterization of the recombinant 1,3-PDO NAD+-dependent oxidoreductase also were explored to gain further insight into the native production of 1,3-PDO. Initial kinetic studies determined a Km value of 1.28 ± 0.57 mM for NAD+ versus 23.8 ± 1.1 mM for 1,3-PDO. The Km values demonstrated that the availability of NAD+/NADH may be a determining factor in 1,3-PDO concentration. These findings support the literature and the conclusion that the bottleneck in 1,3-PDO production lies in maintaining an available pool of NAD+/NADH while mitigating negative effects associated with the accumulation of toxic byproducts.
| Identifer | oai:union.ndltd.org:USASK/oai:ecommons.usask.ca:10388/ETD-2012-04-399 |
| Date | 2012 April 1900 |
| Contributors | Tanaka, Takuji |
| Source Sets | University of Saskatchewan Library |
| Language | English |
| Detected Language | English |
| Type | text, thesis |
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