The role of microorganisms in the degradation of xenobiotics in the environment is well established. Bacteria from the genus Pseudomonas are particularly well adapted to the degradation of hydrocarbons, aromatics, and numerous other natural and introduced substrates. We have isolated a strain of Pseudomonas putida, designated PC2P15, that uses toluene, phenol, benzene, and a number of other substrates as its sole sources of carbon and energy.

Thesis (M.S.)--University of North Carolina at Wilmington, 2003. / Includes bibliographical references (leaves : [90]-91).

Thesis (Ph.D.)--Ohio University, November, 2007. / Title from PDF t.p. Includes bibliographical references.

Thesis (Ph. D.)--University of Canterbury, 2008. / Typescript (photocopy). Includes bibliographical references. Also available via the World Wide Web.

Thesis (Ph. D.)--Johns Hopkins University, 1923. / Biography.

Electro-organic reactions are often complicated by the need to add supporting electrolytes and co-solvents. In many cases these additives take part in side reactions causing low yields and hinder the purification stages. The solid polymer electrolyte (SPE) reactor uses an ion exchange membrane to transfer charged species between the electrodes and so eliminates the need for any additives. In this way improvements in electrochemical processing can be achieved. The SPE reactor has only been studied for model organic and aqueous based electrochemical reactions. The aims of this project were to develop the reactor for use as a suitable means of synthesising alcohols and acids based on substituted toluenes. This involved selection of suitable electrode material, polymer electrolyte pre-treatment and reactor modelling. According to published reports the direct electro-oxidation of toluene takes place with maximum yields of 19% with an acetic acid co-solvent and a nitric acid supporting electrolyte. Higher yields are possible with inorganic mediators such as Mn³⁺ and Cr⁶⁺. 30% yields of methoxylated products are possible from electrolysis in methanol although many non volatile by-products are formed. Initial research was spent investigating the oxidation of toluene in sulphuric acid at a lead dioxide rotating disk electrode. It was found that the reaction is mass transfer limited in the potential region below gas evolution. The order of reaction with respect to toluene was 0.5. Electrolysis of toluene on platinum mesh in nitric acid with and without acetic acid was found to produce benzyl alcohol and benzaldehyde with low current efficiencies. Without co-solvent the maximum current efficiency was 10% at 2S0Alm². An SPE reactor fabricated from glass with an active electrode area of Scm2 was used to perform electrode tests. Highest yields of benzaldehyde were obtained using nickel foam, graphite felt and palladium coated mesh electrodes. The current efficiencies were 52.4%, 20.3% and 10.7% respectively. This work highlighted the need for a good membrane-electrode contact. The oxidation of benzyl alcohol in the same reactor using nickel foam Abstract was accomplished with a current efficiency of 85.4% showing that the difficult step in the oxidation of toluene was the first one to benzyl alcohol. Pre-treatment of the membrane by swelling in solvents was considered to be an important factor in the performance of the SPE reactor. Several ion exchange membranes were pre-treated in a variety of aqueous and organic solvents including methanol, toluene, DMF, water and sulphuric acid. Nafion® 117 was found to increase in size more than the other tested membranes in all solvents except water and sulphuric acid. Many of the pre-treated membranes were tested in an SPE reactor made from steel with an active electrode area of 2lcm2 for the oxidation of toluene in methanol. The anode-membrane potential was measured as a function of time and current density with Nafion® 117 having the lowest values of potential. Selection of the pre-treatment method for future use was determined by assessing the performance in the reactor, contamination of products and chemical hazards. Swelling in aqueous solvents was the chosen procedure. The steel SPE reactor was operated in continuous mode with recycle for the oxidation of toluene in methanol. Galvanostatic electrolysis took place at several current densities, temperatures and feed concentrations. Two products were identified as ⍺-methoxytoluene and ⍺,⍺-dimethoxytoluene and these were formed at low current efficiencies between 1.4% and 9%. The main product was thought to be an oligomer of toluene. The gas generated was found to be mainly hydrogen with a small amount of oxygen thought to come from residual water in the pre-treated membrane. A computer simulation of the SPE reactor for toluene oxidation in methanol was based on two series and one parallel reaction. These were first order in reactant species and followed Tafel type kinetics. Mass transfer of dilute reactants was based on Fickian diffusion. Parameters not available in the literature such as membrane potential and electro-osmotic flow were correlated to applied variables using experimental data and multiple linear regression. The importance of electro-osmotic flow in the SPE reactor was demonstrated by considering its effect on product distribution. The model showed that the oligomerisation of toluene was the dominant reaction making the SPE reactor unsuitable for the oxidation of toluene.

No description available.

The patterns, stoichiometry, and kinetics of microbial benzene, toluene, p-xylene, m-xylene, and o-xylene degradation by a denitrifying activated sludge consortium was investigated in a sequencing batch reactor (SBR) receiving a mixed waste. After six months of acclimation, toluene and m-xylene were routinely degraded to below detection. Both toluene and m-xylene could serve as sole carbon and energy sources. The removal of o-xylene was also possible; however, its transformation was dependent upon gratuitous metabolism during toluene degradation. Benzene and p-xylene were recalcitrant throughout the study. The first order decay coefficient (b) of the denitrifying biomass was determined to be 0.016 ± 0.006 h⁻¹ on a theoretical oxygen demand (thOD) basis. The true growth yields (Y) for the biogenic and toluene/m-xylene components of the mixed waste were determined to be 0.41 ± 0.02 and 0.35 ± 0.04 mg thOD biomass per mg thOD substrate, respectively. The Monod parameters, qmax and KS, for toluene ranged from 0.059 to 0.14 mg toluene/mg protein/h and 0.84 to 6.9 mg/L, respectively. For m-xylene, the qmax and KS parameters ranged from 0.034 to 0.041 mg m-xylene/mg protein/h and 0.28 to 3.7 mg/L, respectively. Some of the variation observed between kinetic experiments was attributed to the different accumulation levels of the denitrification intermediate nitrite (NO⁻) and the inhibitory effects of its conjugate acid, nitrous acid (HNO₂). Other evidence suggested that part of the variation was also due to a continuous acclimation and refinement towards higher affinity toluene- and m-xylene-degrading enzyme systems within the biomass. / Master of Science

<p>The mass transfer of ethanol and toluene from air stream to liquid phase, and bioremediation of contaminated air streams containing either ethanol or toluene have been investigated using a stirred tank bioreactor. This investigation was conducted in six phases: </p> 1) mass transfer experiments involving the transport of toluene and ethanol from contaminated air streams into the liquid phase,</p> 2) study of air stripping effects of ethanol and toluene out of the liquid phase,</p> 3) batch growth experiments to determine growth kinetic models and model parameters,</p> 4) bioremediation of ethanol or toluene as the sole substrate to determine the capacity of Pseudomonas putida (P. putida) (ATCC 23973) growth on these substrates,</p> 5) toluene removal from contaminated air streams using ethanol and benzyl alcohol as co-substrates, and</p> 6) modelling the above studies using metabolic pathways to better understand the bioremediation process.</p> <p>Preliminary oxygen mass transfer studies showed that the presence of ethanol in the liquid phase enhances the overall oxygen mass transfer coefficients. Increasing the ethanol concentration from 0 to 8 g/L caused the oxygen mass transfer coefficients to increase from 0.015 to 0.049 s-1, and from 0.017 to 0.076 s-1, for impeller speeds of 450 and 600 rpm, respectively. Mass transfer studies using ethanol vapor in the air stream demonstrated complete absorption into the aqueous phase of the bioreactor at all operating conditions investigated (air flowrates up to 2.0 L/min and inlet concentrations up to 95.0 mg/L) and therefore mass transfer coefficients for ethanol absorption could not be determined. On the other hand, toluene mass transfer coefficients could be measured and were found to be 8.3x10-4, 8.8x10-4 and 1.0x10-3 s-1 at agitation speeds of 300, 450 and 600 rpm, respectively. The ethanol air stripping parameters (b values) were determined (at initial ethanol liquid concentration of 8.6 g/L) to be 0.002 and 0.007 h-1 for air flow rates of 0.4 L/min (0.3 vvm) and 1.4 L/min (1 vvm), respectively. The toluene air stripping rates, at initial liquid toluene concentration of 440 mg/L, were found to be 1.9, 5.3, 10.4, and 12.6 h-1 for air flow rates of 0.4, 0.9, 1.4, 2.1 L/min, respectively, which is much higher than those of ethanol at the same air flow rates and stirring speed of 450 rpm. It was also observed that benzyl alcohol was not stripped to any detectable level at any of the operating conditions used in this study.</p> <p>The growth of <i>P. putida</i> using toluene as sole substrate was carried out at several operating conditions by varying the dilution rates (D) from 0.01 to 0.1 h-1, the toluene air inlet concentration from 4.5 to 23.0 mg/L and air flow rates of 0.25 to 0.37 L/min (resulting in inlet toluene loadings from 70 to 386 mg/L-h). Steady state operation could not be achieved with toluene as the sole substrate. Ethanol and benzyl alcohol were therefore used as co-substrates for the toluene removal process. In order to understand the kinetics of P. putida growing on ethanol or benzyl alcohol, batch growth experiments were carried out at different initial substrate concentrations. The specific growth rates determined from the batch runs showed that ethanol had no inhibition effect on the growth of P. putida. The growth on ethanol followed the Monod equation with the maximum growth rate of 0.56 h-1 and yield of 0.59. The results from the batch growth experiments on benzyl alcohol showed that benzyl alcohol inhibits the growth of P. putida when the initial concentration of benzyl alcohol in the growth media is increased. The maximum growth rate was 0.42 h-1 in the inhibition model and the yield value was 0.45. </p><p>By operating the bioreactor in continuous mode using a pure strain of <i>P. putida</i>, it was possible to continuously convert ethanol into biomass without any losses to the gas phase or accumulation in the bioreactor at inlet ethanol concentrations of 15.9 and 19.5 mg/L. With ethanol as a co-substrate, toluene was efficiently captured in the bioreactor and readily degraded by the same strain of P. putida. A toluene removal efficiency of 89% was achieved with an ethanol inlet concentration of 15.9 mg/L and a toluene inlet concentration of 4.5 mg/L. With the introduction of benzyl alcohol as co-substrate at a feed rate of 0.12 g/h, the toluene removal efficiency reached 97% at toluene inlet concentrations up to 5.7 mg/L. All the experimental results at steady state were obtained when the bioreactor operated in a continuous mode at a dilution rate of 0.1 h-1, an air flowrate of 0.4 L/min, an agitation speed of 450 rpm and a reactor temperature of 25.0oC. The results of this study indicate that the well-mixed bioreactor is a suitable technology for the removal of VOCs with both high and low water solubility from polluted air streams. The results were achieved at higher inlet pollutant concentrations compared to existing biofilter treatments.</p><p>A metabolic model has been developed to simulate the bioremediation of ethanol, benzyl alcohol and toluene. For continuous steady state operations, ethanol as a sole substrate required less maintenance for biomass growth (0.010 C-mol/C-mol-h) than bioremediations in the presence of toluene, as seen with the ethanol/toluene mixture (0.027 C-mol/C-mol-h), and the benzyl alcohol/toluene mixture (0.069 C-mol/C-mol-h).</p>

<p>The mass transfer of ethanol and toluene from air stream to liquid phase, and bioremediation of contaminated air streams containing either ethanol or toluene have been investigated using a stirred tank bioreactor. This investigation was conducted in six phases: </p> 1) mass transfer experiments involving the transport of toluene and ethanol from contaminated air streams into the liquid phase,</p> 2) study of air stripping effects of ethanol and toluene out of the liquid phase,</p> 3) batch growth experiments to determine growth kinetic models and model parameters,</p> 4) bioremediation of ethanol or toluene as the sole substrate to determine the capacity of Pseudomonas putida (P. putida) (ATCC 23973) growth on these substrates,</p> 5) toluene removal from contaminated air streams using ethanol and benzyl alcohol as co-substrates, and</p> 6) modelling the above studies using metabolic pathways to better understand the bioremediation process.</p> <p>Preliminary oxygen mass transfer studies showed that the presence of ethanol in the liquid phase enhances the overall oxygen mass transfer coefficients. Increasing the ethanol concentration from 0 to 8 g/L caused the oxygen mass transfer coefficients to increase from 0.015 to 0.049 s-1, and from 0.017 to 0.076 s-1, for impeller speeds of 450 and 600 rpm, respectively. Mass transfer studies using ethanol vapor in the air stream demonstrated complete absorption into the aqueous phase of the bioreactor at all operating conditions investigated (air flowrates up to 2.0 L/min and inlet concentrations up to 95.0 mg/L) and therefore mass transfer coefficients for ethanol absorption could not be determined. On the other hand, toluene mass transfer coefficients could be measured and were found to be 8.3x10-4, 8.8x10-4 and 1.0x10-3 s-1 at agitation speeds of 300, 450 and 600 rpm, respectively. The ethanol air stripping parameters (b values) were determined (at initial ethanol liquid concentration of 8.6 g/L) to be 0.002 and 0.007 h-1 for air flow rates of 0.4 L/min (0.3 vvm) and 1.4 L/min (1 vvm), respectively. The toluene air stripping rates, at initial liquid toluene concentration of 440 mg/L, were found to be 1.9, 5.3, 10.4, and 12.6 h-1 for air flow rates of 0.4, 0.9, 1.4, 2.1 L/min, respectively, which is much higher than those of ethanol at the same air flow rates and stirring speed of 450 rpm. It was also observed that benzyl alcohol was not stripped to any detectable level at any of the operating conditions used in this study.</p> <p>The growth of <i>P. putida</i> using toluene as sole substrate was carried out at several operating conditions by varying the dilution rates (D) from 0.01 to 0.1 h-1, the toluene air inlet concentration from 4.5 to 23.0 mg/L and air flow rates of 0.25 to 0.37 L/min (resulting in inlet toluene loadings from 70 to 386 mg/L-h). Steady state operation could not be achieved with toluene as the sole substrate. Ethanol and benzyl alcohol were therefore used as co-substrates for the toluene removal process. In order to understand the kinetics of P. putida growing on ethanol or benzyl alcohol, batch growth experiments were carried out at different initial substrate concentrations. The specific growth rates determined from the batch runs showed that ethanol had no inhibition effect on the growth of P. putida. The growth on ethanol followed the Monod equation with the maximum growth rate of 0.56 h-1 and yield of 0.59. The results from the batch growth experiments on benzyl alcohol showed that benzyl alcohol inhibits the growth of P. putida when the initial concentration of benzyl alcohol in the growth media is increased. The maximum growth rate was 0.42 h-1 in the inhibition model and the yield value was 0.45. </p><p>By operating the bioreactor in continuous mode using a pure strain of <i>P. putida</i>, it was possible to continuously convert ethanol into biomass without any losses to the gas phase or accumulation in the bioreactor at inlet ethanol concentrations of 15.9 and 19.5 mg/L. With ethanol as a co-substrate, toluene was efficiently captured in the bioreactor and readily degraded by the same strain of P. putida. A toluene removal efficiency of 89% was achieved with an ethanol inlet concentration of 15.9 mg/L and a toluene inlet concentration of 4.5 mg/L. With the introduction of benzyl alcohol as co-substrate at a feed rate of 0.12 g/h, the toluene removal efficiency reached 97% at toluene inlet concentrations up to 5.7 mg/L. All the experimental results at steady state were obtained when the bioreactor operated in a continuous mode at a dilution rate of 0.1 h-1, an air flowrate of 0.4 L/min, an agitation speed of 450 rpm and a reactor temperature of 25.0oC. The results of this study indicate that the well-mixed bioreactor is a suitable technology for the removal of VOCs with both high and low water solubility from polluted air streams. The results were achieved at higher inlet pollutant concentrations compared to existing biofilter treatments.</p><p>A metabolic model has been developed to simulate the bioremediation of ethanol, benzyl alcohol and toluene. For continuous steady state operations, ethanol as a sole substrate required less maintenance for biomass growth (0.010 C-mol/C-mol-h) than bioremediations in the presence of toluene, as seen with the ethanol/toluene mixture (0.027 C-mol/C-mol-h), and the benzyl alcohol/toluene mixture (0.069 C-mol/C-mol-h).</p>