Rebecca Smith

Document Type

Thesis - University Access Only

Award Date

2006

Degree Name

Master of Science (MS)

Department / School

Biology and Microbiology

First Advisor

William Gibbons

Abstract

Polyhydroxyalkanoate (PHA) is a naturally-occurring, biodegradable polymer with plasticlike characteristics. PHA has potential applications in the manufacturing of biodegradable plastics and providing an alternative to petroleum-based plastic products. Rhodospirillum rubrum produces PHA as a means of storing carbon and energy when excess carbon is present and another nutrient (N, S, phosphate, Fe, Mg, Kor oxygen) is limiting. R rubrum is typically cultured on an expensive medium (SMN), and our objective was to develop a less expensive alternative using condensed corn solubles (CCS), a byproduct of ethanol fermentation. In serum bottle trials a basal medium, composed of 240 g/L CCS (wet basis), performed as well as SMN and tryptic soy broth, achieving a maximum cell density of 1.6E9 CFU / ml and growth rate of 0. 9 5 h-1 • Illumination had no effect on growth, and supplementation of CCS with nickel or biotin also had no stimulatory effect. CCS concentrations of 80-320 g/L also supported similar growth. The presence of a small amount of oxygen in the headspace resulted in significantly higher (P<0.05) maximum cell densities and growth rates, compared to strictly anaerobic cultures. In bioreactor trials growth was significantly better in 240 g/L CCS (2.3E9 CFU / ml) than in the defined mSMN medium (1.9E9 CFU/ml). Active aeration of the CCS medium greatly increased maximum cell density to 1.3E 10 CFU / ml. Nitrogen supplementation of 240 g/L CCS increased the C:N ratio from 6.7:1 to 5.7:1 which dramatically increased maximum cell density (9.6E10 CFU / ml) and carbon utilization when the pH of the media was controlled. Decreasing the concentration of the CCS to 80 g/L supplemented with nitrogen allowed good cell growth with complete utilization of carbon and nitrogen in the growth phase, not P(HB-HV) production phase. This carbon and nitrogen utilization would ensure that P(HB-HV) would be produced under nitrogen-limiting conditions with an additional source of carbon at that time. Syngas (produced from the gasification of dry distillers grain) was tested as a potential source of carbon for autotrophic P(HB-HV) synthesis after initial growth in CCS. Optimal timing of syngas addition was found to be between 6 h and 17 h when most of the nutrients are used up in the CCS and the culture has reached a high density so it starts using the syngas carbon for P(HB-HV) synthesis. Centrifuging and switching to autotrophic growth conditions with a basal medium was not a sufficient trigger to induce R rubrum to accumulate a significant amount of P(HB-HV). Utilization of CCS and syngas for P(HBHV) production would provide the ethanol industry with an additional high value product.

Library of Congress Subject Headings

Poly-beta-hydroxyalkanoates Poly-beta-hydroxybutyrate Rhodospirillum rubrum Synthesis gas Corn products industry -- By-products Polymers Biodegradable plastics

Publisher

South Dakota State University

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In Copyright