Mineralization of Trichloroethylene by Heterotrophic Enrichment Cultures

Microbial consortia capable of aerobically degrading more than 99% of exogenous trichloroethylene (TCE) (50 mg/liter) were collected from TCE-contaminated subsurface sediments and grown in enrichment cultures. TCE at concentrations greater than 300 mg/liter was not degraded, nor was TCE used by the consortia as a sole energy source. Energy sources which permitted growth included tryptone-yeast extract, methanol, methane, and propane. The optimum temperature range for growth and subsequent TCE consumption was 22 to 37°C, and the pH optimum was 7.0 to 8.1. Utilization of TCE occurred only after apparent microbial growth had ceased. The major end products recovered were hydrochloric acid and carbon dioxide. Minor products included dichloroethylene, vinylidine chloride, and, possibly, chloroform.     

Continuous vapor-phase trichloroethylene biofiltration using hydrocarbon-enriched compost as filtration matrix

Two sources of finished compost material were examined for the capacity to support trichloroethylene(TCE)-degrading microbial populations in a gas-phase bioreactor. Gaseous hydrocarbon was passed through the bioreactor to stimulate cometabolic oxidation of TCE. Significant differences in TCE removal efficiencies were observed between the two compost types, and between hydrocarbon-stimulated and non-stimulated compost. At an average column retention time of 5.6 min, deciduous leaf debris compost removed more than 95% of a 5–50 ppm (by vol.) TCE gas stream, whereas less than 15% removal was observed under similar conditions with a woodchip and bark compost. Trichloroethylene removal efficiency varied with the hydrocarbon-stimulation regime employed, although propane and methane stimulated TCE degradation equally well. Amendment of compost with granular activated carbon substantially increased biological TCE removal. Differences in TCE removal efficiencies observed between the two compost types and between hydrocarbon-stimulated and non-stimulated composts were investigated in terms of changes in the overall heterotrophic microbial populations by using community-level physiological profile analysis. Received: 14 April 1997 / Received revision: 21 July 1997 / Accepted: 25 August 1997     

Aerobic Mineralization of Trichloroethylene, Vinyl Chloride, and Aromatic Compounds by Rhodococcus Species

Two Rhodococcus strains which were isolated from a trichloroethylene (TCE)-degrading bacterial mixture and Rhodococcus rhodochrous ATCC 21197 mineralized vinyl chloride (VC) and TCE. Greater than 99.9% of a 1-mg/liter concentration of VC was degraded by cell suspensions. [1,2-¹⁴C]VC was degraded by cell suspensions, with the production of greater than 66% ¹⁴CO₂ and 20% ¹⁴C-aqueous phase products and incorporation of 10% of the ¹⁴C into the biomass. Cultures that utilized propane as a substrate were able to mineralize greater than 28% of [1,2-¹⁴C]TCE to ¹⁴CO₂, with approximately 40% appearing in ¹⁴C-aqueous phase products and another 10% of ¹⁴C incorporated into the biomass. VC degradation was oxygen dependent and occurred at a pH range of 5 to 10 and temperatures of 4 to 35°C. Cell suspensions degraded up to 5 mg of TCE per liter and up to 40 mg of VC per liter. Propane competitively inhibited TCE degradation. Resting cell suspensions also degraded other chlorinated aliphatic hydrocarbons, such as chloroform, 1,1-dichloroethylene, and 1,1,1-trichloroethane. The isolates degraded a mixture of aromatic and chlorinated aliphatic solvents and utilized benzene, toluene, sodium benzoate, naphthalene, biphenyl, and n-alkanes ranging in size from propane to hexadecane as carbon and energy sources. The environmental isolates appeared more catabolically versatile than R. rhodochrous ATCC 21197. The data report that environmental isolates of Rhodococcus species and R. rhodochrous ATCC 21197 have the potential to degrade TCE and VC in addition to a variety of aromatic and chlorinated aliphatic compounds either individually or in mixtures.     

Comparison of TCE Transformation Abilities of Methane- and Propane-Utilizing Microorganisms

Subsurface microorganisms from McClellan Air Force Base (AFB) were grown in batch aquifer microcosms on methane, propane, and butane to evaluate the potential for aerobic trichloroethylene (TCE) cometabolism. Microorganisms stimulated on all three substrates indicated the existence of a subsurface microbial community capable of utilizing alkanes as growth substrates. Initial growth substrate utilization lag periods of 2 weeks for methane and 3 weeks for propane and butane were observed. Methane- and propane-utilizers were active toward TCE cometabolism, whereas butane-utilizers showed no ability to transform TCE. Gradually increasing TCE concentrations were effectively transformed with uniform additions of methane and propane for up to 1 year. TCE was transformed most rapidly during active methane utilization, and continued at a slower rate for approximately 1 week after methane was consumed. Propane microcosms maintained first-order TCE transformation for up to 4 weeks after propane was consumed. The microbial communities remained active toward primary substrate utilization as the TCE concentration was gradually increased. Both methane- and propane-utilizers showed positive correlations between TCE transformation rates and primary substrate utilization rates. Observed maximum TCE transformation yields were 0.068 g TCE/g methane and 0.048 g TCE/g propane. The methane-utilizers also transformed chloroform (CF) but not 1,1,1-trichloroethane (1,1,1-TCA). Propane-utilizers transformed both CF and 1,1,1-TCA, indicating they were better suited for cometabolizing chlorinated aliphatic hydrocarbon mixtures in the McClellan AFB subsurface.     

Effects of hydrocarbon enrichment on trichloroethylene biodegradation and microbial populations in finished compost

This study focused on the capacity of finished compost, often used as packing material in biofiltration units, to support microbial biodegradation of trichloroethylene (TCE). Finished compost was enriched with methane or propane (10% head space) to stimulate cometabolic biodegradation of gaseous TCE. Successful hydrocarbon enrichment, as indicated by rapid depletion of hydrocarbon gas and measurable growth of hydrocarbon-utilizing micro-organisms, occurred within a week. Within batch reactor flasks, approximately 75% of head space TCE (1–40 ppmv) was rapidly sorbed onto compost material. Up to 99% of the remaining head space TCE was removed via biodegradation in compost enriched with either hydrocarbon. Hydrocarbon enrichment with methane or propane corresponded to 10-fold increases in methanotrophic or propanotrophic populations, respectively. Based on growth assessment under different nutritional regimes, there appeared to be complex metabolic interactions within the microbial community in enriched compost. Five separate bacterial cultures were derived from the hydrocarbon-enriched compost and assayed for the ability to degrade TCE.     

TCE degradation in a methanotrophic attached-film bioreactor

Trichloroethene was degraded in expanded-bed bioreactors operated with mixed-culture methanotrophic attached films. Biomass concentrations of 8 to 75 g volatile solids (VS) per liter static bed (L(sb)) were observed. Batch TCE degradation rates at 35 degrees C followed the Michaelis-Menten model, and a maximum TCE degradation rate (q(max)) of 10.6 mg TCE/gVS . day and a half velocity coefficient (K(S)) of 2.8 mg TCE/L were predicted. Continuous-flow kinetics also followed the Michaelis-Menten model, but other parameters may be limiting, such as dissolved copper and dissolved methane-q(max) and K(S) were 2.9 mg TCE/gVS . day and 1.5 mg TCE/L, respectively, at low copper concentrations (0.003 to 0.006 mg Cu/L). The maximum rates decreased substantially with small increases in dissolved copper. Methane consumption during continuous-flow operation varied from 23 to 1200 g CH(4)/g TCE degraded. Increasing the influent dissolved methane concentration from 0.01 mg/L to 5.4 mg/L reduced the TCE degradation rate by nearly an order of magnitude at 21 degrees C. Exposure of biofilms to 1.4 mg/L tetrachloroethene (PCE) at 35 degrees C resulted in the loss of methane utilization ability. Tests with methanotrophs grown on granular activated carbon indicated that lower effluent TCE concentrations could be obtained. The low efficiencies of TCE removal and low degradation rates obtained at 35 degrees C suggest that additional improvements will be necessary to make methanotrophic TCE treatment attractive. (c) 1993 John Wiley & Sons, Inc.     

Transformation capacities of chlorinated organics by mixed cultures enriched on methane, propane, toluene, or phenol

The degradation of trichloroethylene (TCE), chloroform (CF), and 1,2-dichloroethane (1,2-DCA) by four aerobic mixed cultures (methane, propane, toluene, and phenol oxidizers) grown under similar chemostat conditions was measured. Methane and propane oxidizers were capable of degrading both saturated and unsaturated chlorinated organics (TCE, CF, and 1,2-DCA). Toluene and phenol oxidizers degraded TCE but were not able to degrade CF, 1,2-DCA, or other saturated organics. None of the cultures tested were able to degrade perchloroethylene (PCE) or carbon tetrachloride (CC(4)). For the four cultures tested, degradation of each of the chlorinated organics resulted in cell inactivation due to product toxicity. In all cases, the toxic products were rapidly depleted, leaving no toxic residues in solution. Among the four tested cultures, the resting cells of methane oxidizers exhibited the highest transformation capacities (T(c)) for TCE, CF, and 1,2-DCA. The T(c) for each chlorinated organic was observed to be inversely proportional to the chlorine carbon ratio (Cl/C). The addition of low concentrations of growth substrate or some catabolic intermediates enhanced TCE transformation capacities and degradation rates, presumably due to the regeneration of reducing energy (NADH); however, addition of higher concentrations of most amendments reduced TCE transformation capacities and degradation rates. Reducing energy limitations and amendment toxicity may significantly affect T(c) measurements, causing a masking of the toxicity associated with chlorinated organic degradation. (c) 1995 John Wiley & Sons, Inc.     

Propane-Induced biodegradation of vapor phase trichoroethylene

Microbial degradation of trichloroethylene (TCE) has been demonstrated under aerobic conditions with propane. The primary objective of this research was to evaluate the feasibility of introducing a vapor phase form of TCE in the presence of propane to batch bioreactors containing a liquid phase suspension of Mycobacterium vaccae JOB5 to accomplish degradation. The reactor system consisted of three phases: a vapor phase introducing air, propane, and TCE; a liquid phase of the microbial suspension; and a solid phase in the form of the microorganisms. Long-term and initial rate experiments were conducted on three culture sets to evaluate microbial response. In two long-term test fed propane and approximately 0.1 mg/L and 1 mg/L of TCE, respectively, propane utilization was more efficient at the high TCE concentration (600 mmol propane/mmol TCE versus 11,900 mmol propane/mmol TCE), because the propane degradation rate was approximately the same for both tests (6.73 mg/L . h and 7.85 mg/L . h for the high and low tests). In addition, TCE utilization decreased after complete propane consumption. Initial rate tests on culture sets fed propane only revealed that cells with a history of exposure to a high concentration of TCE had the highest specific growth rate, but the lowest half-saturation constant (7.60e(-3) h(-1) and 0.10 mg/L, respectively). Tests fed variable TCE concentrations (0.031 to 5.378 mg/L in the liquid phase) with no propane showed TCE depletion but no biomass growth. The tests revealed that the TCE removal increased as the TCE concentration increased, indicating a greater removal efficiency at the higher concentrations. Tests with a constant initial propane concentration and variable liquid phase TCE concentration revealed that specific propane utilization was essentially the same. (c) 1995 John Wiley & Sons, Inc.     

Survey of microbial oxygenases: trichloroethylene degradation by propane-oxidizing bacteria

Microorganisms that biosynthesize broad-specificity oxygenases to initiate metabolism of linear and branched-chain alkanes, nitroalkanes, cyclic ketones, alkenoic acids, and chromenes were surveyed for the ability to biodegrade trichloroethylene (TCE). The results indicated that TCE oxidation is not a common property of broad-specificity microbial oxygenases. Bacteria that contained nitropropane dioxygenase, cyclohexanone monooxygenase, cytochrome P-450 monooxygenases, 4-methoxybenzoate monooxygenase, and hexane monooxygenase did not degrade TCE. However, one new unique class of microorganisms removed TCE from incubation mixtures. Five Mycobacterium strains that were grown on propane as the sole source of carbon and energy degraded TCE. Mycobacterium vaccae JOB5 degraded TCE more rapidly and to a greater extent than the four other propane-oxidizing bacteria. At a starting concentration of 20 microM, it removed up to 99% of the TCE in 24 h. M. vaccae JOB5 also biodegraded 1,1-dichloroethylene, trans-1,2-dichloroethylene, cis-1,2-dichloroethylene, and vinyl chloride.     

Treatment of trichloroethylene (TCE) in a membrane biofilter

This article reports on the biodegradation of trichloroethylene (TCE) in a hollow-fiber membrane biofilter. Air contaminated with TCE was passed through microporous hollow fibers while an oxygen-free nutrient solution was recirculated through the shell side of the membrane module. The biomass was attached to the outside surface of the microporous hollow fibers by initially supplying toluene in the gas phase that flows through the fibers. While studies on TCE biodegradation were conducted, there was no toluene present in the gas phase. At 20-ppmv inlet concentration of TCE and 36-s gas-phase residence time, based on total internal volume of the hollow fibers, 30% removal efficiency of TCE was attained. At higher air flow rates or lower gas-phase residence times, lower removal efficiencies were observed. During TCE degradation, the pH of the liquid phase on the shell side of the membrane module decreased due to release of chloride ions. A mathematical model was developed to describe the synchronous aerobic/anaerobic biodegradation of TCE. (c) 1996 John Wiley & Sons, Inc.     

Treatment of high-concentration gaseous benzene streams using a novel bioreactor system

A continuous treatment system combining a packed-bed column and a two-phase partitioning bioreactor has been designed to treat high-concentration benzene-containing gas streams. 1-Octadecene was used in a closed loop as an absorbant to scrub benzene in the counter-current column, after which it was transferred to the two-phase partitioning bioreactor to partition benzene into the 1 l aqueous phase for degradation by Klebsiella sp. The solvent was then recirculated back to the absorber. A gas stream containing 20 mg l^–1 benzene at a flow rate of 60 l h^–1 was introduced to the system, and the benzene was degraded at a biological removal efficiency of 87% at steady state.     

Biodegradation of trichloroethylene and toluene by indigenous microbial populations in soil.

The biodegradation of trichloroethylene (TCE) and toluene, incubated separately and in combination, by indigenous microbial populations was measured in three unsaturated soils incubated under aerobic conditions. Sorption and desorption of TCE (0.1 to 10 micrograms ml-1) and toluene (1.0 to 20 micrograms ml-1) were measured in two soils and followed a reversible linear isotherm. At a concentration of 1 micrograms ml-1, TCE was not degraded in the absence of toluene in any of the soils. In combination, both 1 microgram of TCE ml-1 and 20 micrograms of toluene ml-1 were degraded simultaneously after a lag period of approximately 60 to 80 h, and the period of degradation lasted from 70 to 90 h. Usually 60 to 75% of the initial 1 microgram of TCE ml-1 was degraded, whereas 100% of the toluene disappeared. A second addition of 20 micrograms of toluene ml-1 to a flask with residual TCE resulted in another 10 to 20% removal of the chemical. Initial rates of degradation of toluene and TCE were similar at 32, 25, and 18 degrees C; however, the lag period increased with decreasing temperature. There was little difference in degradation of toluene and TCE at soil moisture contents of 16, 25, and 30%, whereas there was no detectable degradation at 5 and 2.5% moisture. The addition of phenol, but not benzoate, stimulated the degradation of TCE in Rindge and Yolo silt loam soils, methanol and ethylene slightly stimulated TCE degradation in Rindge soil, glucose had no effect in either soil, and dissolved organic carbon extracted from soil strongly sorbed TCE but did not affect its rate of biodegradation.     

Selective desorption of carbon dioxide from sewage sludge for in situ methane enrichment--part I: pilot-plant experiments

In situ methane enrichment in anaerobic digestion of sewage sludge has been investigated by experiments and by modeling. In this first part, the experimental work on the desorption of carbon dioxide and methane from sewage sludge is reported. The bubble column, had a diameter of 0.3 m and a variable height up to 1.8 m. At operation the dispersion height in the column was between 1 and 1.3 m. Outdoor air was used. The column was placed close to a full-scale sewage sludge digester, at a municipal wastewater treatment plant. The digester was operated at mesophilic conditions with a hydraulic retention time of about 20 days. The bubble column was operated to steady-state, at which carbon dioxide concentration and alkalinity were determined on the liquid side, and the concentration of carbon dioxide and methane on the gas side. Thirty-eight experiments were performed at various liquid and gas flow rates. The experimental results show that the desorption rates achieved for carbon dioxide ranges from 0.07 to 0.25 m(3) CO(2)/m(3) sludge per day, which is comparable to the rate of generation by the anaerobic digestion. With increasing liquid flow rate and decreasing gas flow rate the amount of methane desorbed per amount of carbon dioxide desorbed increases. The lowest methane loss achieved is approximately 2% of the estimated methane production in the digestion process.     

Kinetics of chlorinated hydrocarbon degradation by Methylosinus trichosporium OB3b and toxicity of trichloroethylene.

The kinetics of the degradation of trichloroethylene (TCE) and seven other chlorinated aliphatic hydrocarbons by Methylosinus trichosporium OB3b were studied. All experiments were performed with cells grown under copper stress and thus expressing soluble methane monooxygenase. Compounds that were readily degraded included chloroform, trans-1,2-dichloroethylene, and TCE, with Vmax values of 550, 330, and 290 nmol min-1 mg of cells-1, respectively. 1,1-Dichloroethylene was a very poor substrate. TCE was found to be toxic for the cells, and this phenomenon was studied in detail. Addition of activated carbon decreased the acute toxicity of high levels of TCE by adsorption, and slow desorption enabled the cells to partially degrade TCE. TCE was also toxic by inactivating the cells during its conversion. The degree of inactivation was proportional to the amount of TCE degraded; maximum degradation occurred at a concentration of 2 mumol of TCE mg of cells-1. During conversion of [14C]TCE, various proteins became radiolabeled, including the alpha-subunit of the hydroxylase component of soluble methane monooxygenase. This indicated that TCE-mediated inactivation of cells was caused by nonspecific covalent binding of degradation products to cellular proteins.     

Kinetics of chlorinated hydrocarbon degradation by Methylosinus trichosporium OB3b and toxicity of trichloroethylene

The kinetics of the degradation of trichloroethylene (TCE) and seven other chlorinated aliphatic hydrocarbons by Methylosinus trichosporium OB3b were studied. All experiments were performed with cells grown under copper stress and thus expressing soluble methane monooxygenase. Compounds that were readily degraded included chloroform, trans-1,2-dichloroethylene, and TCE, with Vmax values of 550, 330, and 290 nmol min-1 mg of cells-1, respectively. 1,1-Dichloroethylene was a very poor substrate. TCE was found to be toxic for the cells, and this phenomenon was studied in detail. Addition of activated carbon decreased the acute toxicity of high levels of TCE by adsorption, and slow desorption enabled the cells to partially degrade TCE. TCE was also toxic by inactivating the cells during its conversion. The degree of inactivation was proportional to the amount of TCE degraded; maximum degradation occurred at a concentration of 2 mumol of TCE mg of cells-1. During conversion of [14C]TCE, various proteins became radiolabeled, including the alpha-subunit of the hydroxylase component of soluble methane monooxygenase. This indicated that TCE-mediated inactivation of cells was caused by nonspecific covalent binding of degradation products to cellular proteins.     

Evaluation of Toxic Effects of Aeration and Trichloroethylene Oxidation on Methanotrophic Bacteria Grown with Different Nitrogen Sources

In this study we evaluated specific and nonspecific toxic effects of aeration and trichloroethylene (TCE) oxidation on methanotrophic bacteria grown with different nitrogen sources (nitrate, ammonia, and molecular nitrogen). The specific toxic effects, exerted directly on soluble methane monooxygenase (sMMO), were evaluated by comparing changes in methane uptake rates and naphthalene oxidation rates following aeration and/or TCE oxidation. Nonspecific toxic effects, defined as general cellular damage, were examined by using a combination of epifluorescent cellular stains to measure viable cell numbers based on respiratory activity and measuring formate oxidation activities following aeration and TCE transformation. Our results suggest that aeration damages predominantly sMMO rather than other general cellular components, whereas TCE oxidation exerts a broad range of toxic effects that damage both specific and nonspecific cellular functions. TCE oxidation caused sMMO-catalyzed activity and respiratory activity to decrease linearly with the amount of substrate degraded. Severe TCE oxidation toxicity resulted in total cessation of the methane, naphthalene, and formate oxidation activities and a 95% decrease in the respiratory activity of methanotrophs. The failure of cells to recover even after 7 days of incubation with methane suggests that cellular recovery following severe TCE product toxicity is not always possible. Our evidence suggests that generation of greater amounts of sMMO per cell due to nitrogen fixation may be responsible for enhanced TCE oxidation activities of nitrogen-fixing methanotrophs rather than enzymatic protection mechanisms associated with the nitrogenase enzymes.     

Effect of Chemical Oxidation on Subsurface Microbiology and Trichloroethene (TCE) Biodegradation

Research was conducted to determine the effect of chemical oxidation on subsurface microbiology and cometabolic biodegradation capacity in a trichloroethene (TCE)/perchloroethene (PCE)-contaminated aquifer previously treated with Fenton's reagent. Groundwater pH declined from 5 to 2.4 immediately after the treatment, and subsequently rose to a range of 3.4 to 4.0 after 17 months. Limited microbial growth and TCE degradation were detected in the treated zone (pH 3.37 and TCE 5 to 21 mg/L) with carbon addition (i.e., methane and phenol). Methane addition resulted in the enrichment of yeast and fungi in microcosms at low pH. In contrast, methane addition to groundwater from the control well (pH 4.9 and TCE ca. 0.7 mg/L) stimulated methanotrophic growth, indicated by methane consumption, fluorescent antibody analysis, phospholipid-based markers, and rDNA probes. TCE degradation was measured in the control microcosms, but only when phenol was added. Although higher TCE concentrations in the treated zone might have inhibited TCE cometabolism, these results also indicate that low groundwater pH resulting from the chemical oxidation process (pH 3.37 versus 4.9) inhibited TCE degradation. Methanotrophic growth and TCE biodegradation may be possible as pH increases both in the treated zone and at the leading edge of plume, as long as the local soil is able to buffer the groundwater pH. Moreover, the Fenton's reagent process could be designed to operate at a higher pH (e.g., ≥ 4.5) and/or lower hydrogen peroxide concentration to minimize detrimental effects, providing an optimal environment to couple advanced oxidation processes with bioremediation technologies.     

Low and high acetate amendments are equally as effective at promoting complete dechlorination of trichloroethylene (TCE)

Experiments with trichloroethylene-contaminated aquifer material demonstrated that TCE, cis-DCE, and VC were completely degraded with concurrent Fe(III) or Fe(III) and sulfate reduction when acetate was amended at stoichiometric concentration; competing TEAPs did not inhibit ethene production. Adding 10× more acetate did not increase the rate or extent of TCE reduction, but only increased methane production. Enrichment cultures demonstrated that ~90 μM TCE or ~22 μM VC was degraded primarily to ethene within 20 days with concurrent Fe(III) or Fe(III) + sulfate reduction. The dechlorination rates were comparable between the low and high acetate concentrations (0.36 vs 0.34 day^?1, respectively), with a slightly slower rate in the 10× acetate amended incubations. Methane accumulated to 13.5 (±0.5) μmol/tube in the TCE-degrading incubations with 10× acetate, and only 1.4 (±0.1) μmol/tube with low acetate concentration. Methane accumulated to 16 (±1.5) μmol/tube in VC-degrading enrichment with 10× acetate and 2 (±0.1) μmol/tube with stoichiometric acetate. The estimated fraction of electrons distributed to methanogenesis increased substantially when excessive acetate was added. Quantitative PCR analysis indicated that 10× acetate did not enhance Dehalococcoides biomass but rather increased the methanogen abundance by nearly one order of magnitude compared to that with stoichiometric acetate. The data suggest that adding low levels of substrate may be equally if not more effective as high concentrations, without producing excessive methane. This has implications for field remediation efforts, in that adding excess electron donor may not benefit the reactions of interest, which in turn will increase treatment costs without direct benefit to the stakeholders.     

Numerical modeling and uncertainties in rate coefficients for methane utilization and TCE cometabolism by a methane-oxidizing mixed culture

The rates of methane utilization and trichloroethylene (TCE) cometabolism by a methanotrophic mixed culture were characterized in batch and pseudo-steady-state studies. Procedures for determination of the rate coefficients and their uncertainties by fitting a numerical model to experimental data are described. The model consisted of a system of differential equations for the rates of Monod kinetics, cell growth on methane and inactivation due to TCE transformation product toxicity, gas/liquid mass transfer of methane and TCE, and the rate of passive losses of TCE. The maximum specific rate of methane utilization (k(CH(4) )) was determined by fitting the numerical model to batch experimental data, with the initial concentration of active methane-oxidizing cells (X(0) (a)) also used as a model fitting parameter. The best estimate of k(CH(4) ) was 2.2 g CH(4)/g cells-d with excess copper available, with a single-parameter 95% confidence interval of 2.0-2.4 mg/mg-d. The joint 95% confidence region for k(CH(4) ) and X(0) (a) is presented graphically. The half-velocity coefficient (K(S,CH(4) )) was 0.07 mg CH(4)/L with excess copper available and 0.47 mg CH(4)/L under copper limitation, with 95% confidence intervals of 0.02-0.11 and 0.35-0.59 mg/L, respectively. Unique values of the TCE rate coefficients k(TCE) and K(S,TCE) could not be determined because they were found to be highly correlated in the model fitting analysis. However, the ratio k(TCE)/K(S,TCE) and the TCE transformation capacity (T(C)) were well defined, with values of 0.35 L/mg-day and 0.21 g TCE/g active cells, respectively, for cells transforming TCE in the absence of methane or supplemental formate. The single-parameter 95% confidence intervals for k(TCE)/K(S,TCE) and T(C) were 0.27-0.43 L/mg-d and 0.18-0.24 g TCE/g active cells, respectively. The joint 95% confidence regions for k(TCE)/K(S,TCE) and T(C) are presented graphically. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 53: 320-331, 1997.