Optimization of PHB production in Methylosinus trichosporium OB3b

• Author / Creator

  Zaldivar Carrillo, Jorge Adalberto

• The utilization of waste materials to generate value-added products is an appealing prospect that can generate economic and environmental benefits. When the end product of this process is itself environmentally friendly, the benefits escalate. Such is the case of the present work. M. trichosporium OB3b is a bacterium that can metabolize methane—a potent greenhouse gas and common industrial waste—and methanol—another common industrial waste—to produce polyhydroxybutyrate (PHB)—a naturally occurring polymer. PHB is the focus of intense research because it is completely biodegradable into harmless compounds and exhibits properties similar to those of other relevant polymers.
  The growth of Methylosinus trichosporium OB3b and its production of PHB was studied and characterized. Empirical second order response surface models were developed to predict three responses—cell dry weight, PHB concentration and PHB cell content—as a function of three independent variables—the carbon source, the nitrogen source and the nitrogen-to-carbon ratio. Methane, methanol and mixtures of them were investigated as carbon sources. Ammonium, nitrate and mixtures of them were investigated as nitrogen sources. Nitrogen-to-carbon ratios in the molar range from 0.005 to 1.0 were investigated. The effect of oxygen was also assessed. A maximum response for PHB concentration was predicted at a composition of 30%mol of methane and 70%mol of methanol as carbon source, 100% nitrate as nitrogen source, and a 0.017 nitrogen-to-carbon ratio, with a predicted concentration of 40 mg/L of PHB. The prediction was confirmed experimentally obtaining a PHB concentration of 48.7 ± 8.3 mg/L, with a PHB cell content of 52.5% ± 6.3%. The predictions of the models for cell dry weight and for PHB cell content failed experimental confirmation due to the low signal-to-noise ratio that resulted from the low cell densities worked with, but important behavioural trends were predicted and confirmed. Greater cell dry weights were predicted and observed with ammonium at high nitrogen-to-carbon ratios. The cell dry weight was predicted and observed to be independent of the carbon source used when the nitrogen source was nitrate. On the other hand, the preferred nitrogen source for PHB accumulation was found to vary depending on the carbon source used. For methane, ammonium was preferred; for methanol, nitrate was preferred. Nitrogen-to-carbon ratios of 0.017 were also found to promote PHB accumulation. A synergistic effect of the carbon source was found on PHB concentration, in that certain mixtures of methane and methanol resulted in greater accumulation of PHB than when either of the carbon sources was used separately. An important effect of oxygen was found, in that the availability of a great stoichiometric excess of it favored the accumulation of PHB. In the absence of this excess, accumulated PHB was metabolized and depleted to various levels.

• Subjects / Keywords

  • Methylotroph
  • PHB
  • Methylosinus trichosporium OB3b
  • Response surface methodology
  • Polyhydroxybutyrate

• Graduation date

  Fall 2015

• Type of Item

  Thesis

• Degree

  Master of Science

• DOI

  https://doi.org/10.7939/R3D795G74

• License

  This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.