Gerald E. Speitel

Kinetics of Aerobic Cometabolism of Chlorinated Solvents

The objectives of this paper are to review the wide range of kinetic models that have been introduced to describe the cometabolic oxidation of chlorinated solvents, to compare modeling approaches and associated experimental data, and to discuss knowledge gaps in the general topic of cometabolism kinetics. To begin, a brief description of the mechanism of oxygenase enzyme metabolism and its qualitative effects on cometabolic degradation kinetics is given. Next, a variety of kinetic expressions that have been used to describe cometabolism, ranging from adaptations of simple metabolic relationships to the development of complex equations that account for intracellular concentrations of key reaction species, are presented. A large number of kinetic coefficients published for a variety of oxygenase populations degrading a broad range of chlorinated solvents are categorized and compared. The discussion section of the paper contains an exploration of knowledge gaps that exist in our understanding of the kinetics of aerobic chlorinated solvent cometabolism. Specific topics covered include: • the use of half saturation constants (Ksc and Ksg) as estimates for inhibition constants (Kisc and Kisg) in saturation modeling expressions, • the specific nature of chlorinated solvent induced product toxicity and the capability for cells to recover from toxic effects, and • methods for incorporating reducing energy limitations into cometabolism models. Finally, the applicability of the broad range of kinetic modeling approaches to scale-up and field applications for in situ bioremediation of chlorinated solvents is discussed.

Kinetics of Methyl t-Butyl Ether Cometabolism at Low Concentrations by Pure Cultures of Butane-Degrading Bacteria

ABSTRACT Butane-oxidizing Arthrobacter (ATCC 27778) bacteria were shown to degrade low concentrations of methylt-butyl ether (MTBE; range, 100 to 800 μg/liter) with an apparent half-saturation concentration (Ks) of 2.14 mg/liter and a maximum substrate utilization rate (kc) of 0.43 mg/mg of total suspended solids per day. Arthrobacter bacteria demonstrated MTBE degradation activity when grown on butane but not when grown on glucose, butanol, or tryptose phosphate broth. The presence of butane, tert-butyl alcohol, or acetylene had a negative impact on the MTBE degradation rate. NeitherMethylosinus trichosporium OB3b nor Streptomyces griseus was able to cometabolize MTBE.

Cometabolism of chlorinated solvents and binary chlorinated solvent mixtures using M. trichosporium OB3b PP358.

The mutant methanotroph, Methylosinus trichosporium OB3b PP358, which constitutively expresses soluble methane monooxygenase (sMMO), was used to study the degradation kinetics of individual chlorinated solvents and binary solvent mixtures. Although sMMOs broad specificity permits a wide range of chlorinated solvents to be degraded, it creates the potential for competitive inhibition of degradation rates in mixtures because multiple chemicals are simultaneously available to the enzyme. To effectively design both ex-situ and in-situ groundwater bioremediation systems using strain PP358, kinetic parameters for chlorinated solvent degradation and accurate kinetic expressions to account for inhibition in mixtures are required. Toward this end, the degradation parameters for six prevalent chlorinated solvents and the verification of enzyme competition model for binary mixtures were the focus of this investigation. M. trichosporium OB3b PP358 degraded trichloroethylene (TCE), chloroform, cis-1,2-dichloroethylene (c-DCE), trans-1,2-dichloroethylene (t-DCE), and 1, 1-dichloroethylene (1,1-DCE) rapidly, with maximum substrate transformation rates of >20.8, 3.1, 9.5 24.8, and >7.5 mg/mg-day, respectively. 1,1,1-trichloroethane (TCA) was not significantly degraded. Half-saturation coefficients ranged from 1 to greater than 10 mg/L. Competition experiments were carried out to observe the effect of a second solvent on degradation rates and to verify the applicability of the Monod model adjusted for competitive inhibition. Binary mixtures of 0.3->0.5 mg/L TCE with up to 5 mg/L c-DCE and up to 7 mg/L 1,1,1-TCA were studied with 20 mM of formate and no growth substrate. No competition was observed at any of these concentrations. Additional competition experiments, using binary mixtures of t-DCE with TCE and t-DCE with c-DCE, were conducted at higher concentrations (i.e., 7-18 mg/L) and enzyme competition was observed. Predictions from a competitive inhibition model compared well with experimental data for these mixtures.

Methanotrophic biodegradation of trichloroethylene in a hollow fiber membrane bioreactor.

Biodegradation of trichloroethlyene (TCE) in a hollow fiber membrane bioreactor was investigated using a mutant of the methanotrophic bacteria, Methylosinus trichosporium OB3b. Contaminated water flowed through the lumen (i.e., fiber interior), and the bacteria circulated through the shell side of the membrane module and an external growth reactor. In mass transfer studies with a radial cross-flow membrane module, 78.3-99.9% of the TCE was removed from the lumen at hydraulic residence times of 3-15 min in the lumen and the shell. In biodegradation experiments, 80-95% of the TCE was removed from the lumen at hydraulic residence times of 5-9 min in the lumen. The TCE transferred to the shell was rapidly biodegraded, with rate constants ranging from 0.16 to 0.9 L (mg of TSS) -1 day -1 . Radiochemical data showed that over 75% of the transferred TCE was biodegraded in the shell, with the byproducts being approximately equally divided between carbon dioxide and nonvolatiles. This study shows that a hollow fiber membrane bioreactor system coupled with the mutant strain PP358 of M. trichosporium OB3b is a very promising technology for chlorinated solvent biodegradation.

Fate and Transport of High Explosives in a Sandy Soil: Adsorption and Desorption

Several areas of the Massachusetts Military Reservation (MMR) have soils with significant levels of high explosives (HE) contamination because of a long history of training and range activities (such as open burning, open detonation, disposal, and artillery and mortar firing). Site-specific transport and attenuation mechanisms were assessed in sandy soils for three contaminants of concern: the nitramine hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), and the nitroaromatics 2,4-dinitrotolune (2,4-DNT) and 2,4,6-trinitrotoluene (TNT). For all three contaminants, linear distribution coefficients (Kd) were dependent on the fraction of organic carbon in soil. The nitroaromatics sorbed much more strongly than RDX in both soils. Over 120 hours, the desorption rate of RDX from field contaminated surface soil was much slower than its sorption rate, with the desorption Kd (1.5 L/kg) much higher than Kd for sorption (0.37 L/kg). Desorption of 2,4-DNT was negligible over 120 hours. Thus, applying sorption-derived Kd values for transport modeling may significantly overestimate the flux of explosives from MMR soils. Based on multiple component column transport tests, RDX will be the most mobile of these contaminants in MMR soils. In saturated columns packed with uncontaminated soil, RDX broke through rapidly, whereas the nitroaromatics were significantly attenuated by irreversible sorption or abiotic transformations.

Biodegradation kinetics of Methylosinus trichosporium OB3b at low concentrations of chloroform in the presence and absence of enzyme competition by methane

Abstract The kinetics of chloroform and trichloroethylene (TCE) degradation by the methanotroph, Methylosinus trichosporium OB3b, were measured in batch kinetic experiments. The experiments focused on initial concentrations of around 100 μg/l for chloroform and around 1 mg/l for TCE. The pseudo first-order rate constants ranged from 0.2 to 0.41/mg TSS-day for chloroform and from 0.5 to 3.31/mg TSS-day for TCE in the absence of methane. Comparison of the TCE rate constants to other work indicates that the organisms were producing the soluble form of their methane monooxygenase. The presence of methane caused significant enzyme competition at methane concentrations as low as 0.35 mg/l, resulting in smaller chloroform rate constants. The decreases in the rate constant were consistent with the predictions of an enzyme competitive inhibition model, which indicated a half saturation coefficient for methane in the order of 0.3 mg/l. The predominant degradation products from chloroform was carbon dioxide.

Cometabolism of trihalomethanes by Nitrosomonas europaea.

ABSTRACT The ammonia-oxidizing bacterium Nitrosomonas europaea (ATCC 19718) was shown to degrade low concentrations (50 to 800 μg/liter) of the four trihalomethanes (trichloromethane [TCM], or chloroform; bromodichloromethane [BDCM]; dibromochloromethane [DBCM]; and tribromomethane [TBM], or bromoform) commonly found in treated drinking water. Individual trihalomethane (THM) rate constants (\batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(k_{1_{THM}}\) \end{document}) increased with increasing THM bromine substitution, with TBM > DBCM > BDCM > TCM (0.23, 0.20, 0.15, and 0.10 liters/mg/day, respectively). Degradation kinetics were best described by a reductant model that accounted for two limiting reactants, THMs and ammonia-nitrogen (NH3-N). A decrease in the temperature resulted in a decrease in both ammonia and THM degradation rates with ammonia rates affected to a greater extent than THM degradation rates. Similarly to the THM degradation rates, product toxicity, measured by transformation capacity (Tc), increased with increasing THM bromine substitution. Because both the rate constants and product toxicities increase with increasing THM bromine substitution, a waters THM speciation will be an important consideration for process implementation during drinking water treatment. Even though a given water sample may be kinetically favored based on THM speciation, the resulting THM product toxicity may not allow stable treatment process performance.

Gas-phase formaldehyde adsorption isotherm studies on activated carbon: Correlations of adsorption capacity to surface functional group density

Formaldehyde (HCHO) adsorption isotherms were developed for the first time on three activated carbons representing one activated carbon fiber (ACF) cloth, one all-purpose granular activated carbon (GAC), and one GAC commercially promoted for gas-phase HCHO removal. The three activated carbons were evaluated for HCHO removal in the low-ppm(v) range and for water vapor adsorption from relative pressures of 0.1-0.9 at 26 °C where, according to the IUPAC isotherm classification system, the adsorption isotherms observed exhibited Type V behavior. A Type V adsorption isotherm model recently proposed by Qi and LeVan (Q-L) was selected to model the observed adsorption behavior because it reduces to a finite, nonzero limit at low partial pressures and it describes the entire range of adsorption considered in this study. The Q-L model was applied to a polar organic adsorbate to fit HCHO adsorption isotherms for the three activated carbons. The physical and chemical characteristics of the activated carbon surfaces were characterized using nitrogen adsorption isotherms, X-ray photoelectron spectroscopy (XPS), and Boehm titrations. At low concentrations, HCHO adsorption capacity was most strongly related to the density of basic surface functional groups (SFGs), while water vapor adsorption was most strongly influenced by the density of acidic SFGs.

Sustained trichloroethylene cometabolism by phenol-degrading bacteria in sequencing biofilm reactors

Bench-scale studies of a sequencing packed-bed bioreactor were conducted for the treatment of waters contaminated with trichloroethylene (TCE). The reactor contained a mixed-culture biofilm grown aerobically on phenol; the biofilm cometabolized TCE in the absence of phenol. These studies focused on the effect of phenol-feeding strategies in relation to sustaining high levels of TCE removal efficiency. The reactor was cycled between a rejuvenation stage, during which phenol at 5, 25, or 100 mg/L was supplied in a nutrient water, and a degradation stage, during which TCE was supplied as a contaminant at 100 μg/L in the absence of phenol. Two to three hours of rejuvenation per 24 hours of TCE degradation were required for stable operation. Pseudo-first-order rate constants for TCE degradation between 50 and 100 L/g volatile suspended solids.d were sustained for up to 29 days. The successful feeding strategies yielded average TCE removals between 70 and 90% at a packed-bed hydraulic residence time of 14 minutes. Conceivably, a full-scale sequencing reactor could operate indefinitely with the proper feeding strategy.

Trichloroethylene degradation by Methylosinus trichosporium OB3b mutants in a sequencing biofilm reactor

Abstract Methylosinus trichosporium OB3b was used in a sequencing biofilm reactor to degrade trichloroethylene (TCE) in water at approximately 100 μg/l influent concentration. The reactor consisted of biofilms grown on diatomaceous earth pellets or glass beads, and was sequenced between growth cycle, in which methane and air were fed in a gas phase, and degradation cycle, in which the column was completely liquid-filled. Biomass loading on the support media was only 10 mg cell dry weight/g support media, giving a calculated biofilm thickness of 0.04 cm. Apparent psuedo-first order degradation rate constants in the system were improved by more than an order of magnitude, from 0.008 l/mg d to 0.1 l/mg d, through the use of antibiotic-resistant strains of copper-resistant organisms that express only the soluble methane monooxygenase, the enzyme responsible for trichloroethylene degradation. Also, biomass accumulation was improved through the use of a cationic coagulating polymer. However, TCE degradation was sustained only for short times, and cycling back to growth mode did not completely regenerate TCE degradation ability. Thus, biofilms of Methylosinus trichosporium OB3b did not give rise to sustained high degradation rates.