Kinetics of inhibition in the biodegradation of monoaromatic hydrocarbons in presence of heavy metals

The toxicity and inhibitory effects of heavy metals such as cadmium, nickel and zinc on alkylbenzene removal were evaluated with a Bacillus strain. The kinetics of alkylbenzene biodegradation with the different heavy metals at various concentrations were modeled using the Andrews equation which yielded a good fit between model and experimental data. Additional experiments undertaken with a Pseudomonas sp. in presence of nickel confirmed a good fit between experimental data and the Andrews model for this strain as well. The heavy metals inhibition constants (Ki) were calculated for different combinations of volatile organic compounds (VOC) and heavy metals. The present approach provides a method for evaluating and quantifying the inhibition effect of heavy metals on the biodegradtion of pollutants by specific microbial strains.     

Kinetics of biodegradation of diethylketone by Arthrobacter viscosus

The performance of an Arthrobacter viscosus culture to remove diethylketone from aqueous solutions was evaluated. The effect of initial concentration of diethylketone on the growth of the bacteria was evaluated for the range of concentration between 0 and 4.8 g/l, aiming to evaluate a possible toxicological effect. The maximum specific growth rate achieved is 0.221 h⁻¹ at 1.6 g/l of initial diethylketone concentration, suggesting that for higher concentrations an inhibitory effect on the growth occurs. The removal percentages obtained were approximately 88%, for all the initial concentrations tested. The kinetic parameters were estimated using four growth kinetic models for biodegradation of organic compounds available in the literature. The experimental data found is well fitted by the Haldane model (R ² = 1) as compared to Monod model (R ² = 0.99), Powell (R ² = 0.82) and Loung model (R ² = 0.95). The biodegradation of diethylketone using concentrated biomass was studied for an initial diethylketone concentration ranging from 0.8–3.9 g/l in a batch with recirculation mode of operation. The biodegradation rate found followed the pseudo-second order kinetics and the resulting kinetic parameters are reported. The removal percentages obtained were approximately 100%, for all the initial concentrations tested, suggesting that the increment on the biomass concentration allows better results in terms of removal of diethylketone. This study showed that these bacteria are very effective for the removal of diethylketone from aqueous solutions.     

Kinetics of biodegradation of p-nitrophenol by different bacteria

Three bacterial species, i.e., Ralstonia sp. SJ98, Arthrobacter protophormiae RKJ100, and Burkholderia cepacia RKJ200, have been examined for their efficiency and kinetics behavior toward PNP degradation. All the three bacteria utilized PNP as the sole source of carbon, nitrogen, and energy. The rates of radiolabeled [U-(14)C]PNP degradation by all the bacteria were higher in the nitrogen-free medium compared to the medium with nitrogen. The apparent K(m) values of PNP degradation by SJ98, RKJ100, and RKJ200 were 0.32, 0.28, and 0.23 mM, respectively, as determined from the Michaelis-Menten curves. The maximum rates of PNP degradation (V(max)) according to Lineweaver-Burk's plots were 11.76, 7.81, and 3.84 micromol PNP degraded/min/mg dry biomass, respectively. The interpretation drawn from the Lineweaver-Burk's plots showed that the PNP degradation by SJ98 was stimulated by 4-nitrocatechol and 1, 2,4-benzenetriol. Benzoquinone and hydroquinone inhibited PNP degradation by RKJ100 noncompetitively and competitively, respectively, whereas in the case of RKJ200, benzoquinone and hydroquinone inhibited PNP degradation in an uncompetitive manner. beta-Ketoadipate did not affect the rate of PNP degradation in any case.     

Kinetics of competitive inhibition and cometabolism in the biodegradation of benzene, toluene, and p-xylene by two Pseudomonas isolates

Two Pseudomonas species (designated strains B1 and X1) were isolated from an aerobic pilot-scale fluidized bed reactor treating groundwater containing benzene, toluene, and p-xylene (BTX). Strain B1 grew with benzene and toluene as the sole sources of carbon and energy, and it cometabolized p-xylene in the presence of toluene. Strain X1 grew on toluene and p-xylene, but not benzene. In single substrate experiments, the appearance of biomass lagged the consumption of growth substrates, suggesting that substrate uptake may not be growth-rate limiting for these substrates. Batch tests using paired substrates (BT, TX, or BX) revealed competitive inhibition and cometabolic degradation patterns. Competitive inhibition was modeled by adding a competitive inhibition term to the Monod expression. Cometabolic transformation of nongrowth substrate (p-xylene) by strain B1 was quantified by coupling xylene transformation to consumption of growth substrate (toluene) during growth and to loss of biomass during the decay phase. Coupling was achieved by defining two transformation capacity terms for the cometabolizing culture: one that relates consumption of growth substrate to the consumption of nongrowth substrate, and second that relates consumption of biomass to the consumption of nongrowth substrate. Cometabolism increased decay rates, and the observed yield for strain B1 decreased in the presence of p-xylene. (c) 1993 Wiley & Sons, Inc.     

Molecular cloning of genetic determinants for inhibition of fungal growth by a fluorescent pseudomonad

Pseudomonas fluorescens HV37a inhibits growth of the fungus Pythium ultimum in vitro. Optimal inhibition is observed on potato dextrose agar, a rich medium. Mutations eliminating fungal inhibition were obtained after mutagenesis with N-methyl-N'-nitro-N-nitrosoguanidine. Mutants were classified by cosynthesis and three groups were distinguished, indicating that a minimum of three genes are required for fungal inhibition. Cosmids that contain wild-type alleles of the genes were identified in an HV37a genomic library by complementation of the respective mutants. This analysis indicated that three distinct genomic regions were required for fungal inhibition. The cosmids containing these loci were mapped by transposon insertion mutagenesis. Two of the cosmids were found to contain at least two genes each. Therefore, at least five genes in HV37a function as determinants of fungal inhibition.     

Kinetics of anaerobic biodegradation of glycerol by sulfate-reducing bacteria

A kinetic model has been developed and kinetic parameters of anaerobic degradation of glycerol, an abundant by-product of biofuel manufacturing, by a consortium of sulfate reducing bacteria (SRB) in a closed system have been determined. The following main species of SRB has been identified in the consortium: Desulfovibrio baarsii, Desulfomicrobium sp., and Desufatomaculum sp. The proposed model included processes of glycerol degradation, sulfate reduction, and inhibition by metabolic products, as well as effects of pH and temperature. The suggested equation for the anaerobic glycerol degradation was based on Edward and Andrew’s equation. The following kinetic parameters of the anaerobic glycerol degradation were obtained for the initial glycerol concentration from 0.15 to 4 ml/l and sulfate concentration of 2760 mg/l at 22°C: maximum specific growth rate of SRB μ_max = 0.56 day⁻¹, economic coefficient of ashless biomass from glycerol of 0.08 mol SRB/mol COC, and yield of ashless biomass from sulfate of 0.020 mol SRB/mol SO₄. It was shown that the optimum molar ratio of $ {{C_{Gl} } \mathord{\left/ {\vphantom {{C_{Gl} } {C_{SO_4 } }}} \right. \kern-\nulldelimiterspace} {C_{SO_4 } }} $ {{C_{Gl} } \mathord{\left/ {\vphantom {{C_{Gl} } {C_{SO_4 } }}} \right. \kern-\nulldelimiterspace} {C_{SO_4 } }} for SRB growth was 0.8. Initial boundary concentration of inhibition by undissociated hydrogen sulfide was 70 mg/l. Dependence of the specific growth rate of bacteria on the temperature was approximated by the Arrhenius equation in the temperature range of 20–30°C with the goodness of fit R² = 0.99.     

Metabolism of beta-methylaspartate by a pseudomonad

A bacterium was isolated from soil which utilizes threo-beta-methyl-l-aspartate, certain other amino acids, and a variety of organic substances as single energy sources. It is, or closely resembles, Pseudomonas putida biotype B. The ability of this organism to rapidly decompose such amino acids is dependent on inducible enzyme systems. Dialyzed cell-free extracts of this bacterium metabolize beta-methylaspartate only when catalytic amounts of alpha-ketoglutarate, or pyruvate, and pyridoxal phosphate are also present. The main products formed from beta-methylaspartate under these conditions are alpha-aminobutyrate, carbon dioxide, and alpha-ketobutyrate. When l-aspartate is substituted for beta-methylaspartate in this system, it is converted mainly to alanine and carbon dioxide. beta-Methyloxalacetate is decarboxylated, and the resulting alpha-ketobutyrate is converted enzymatically in the presence of glutamate to alpha-aminobutyrate which accumulates. The added keto acids are converted, in part, to the corresponding amino acids probably by transamination. The data indicate that beta-methylaspartate is converted to alpha-aminobutyrate, and aspartate to alanine, by a circuitous transamination-beta-decarboxylation-transamination sequence rather than by a direct beta-decarboxylation.     

Production of the siderophore aerobactin by a halophilic pseudomonad.

A bacterial strain, isolated from a cyanobacterial culture, was identified as Pseudomonas sp. strain X40. Under iron-limiting conditions, the Pseudomonas sp. produced aerobactin, a dihydroxamate siderophore previously found only in the family Enterobacteriaceae. Aerobactin was identified by electrophoretic mobility, spectrophotometric titration, proton nuclear magnetic resonance spectroscopy, mass spectrometry, acid hydrolysis, and biological activity. Aerobactin was used as a siderophore in the Pseudomonas sp. and Escherichia coli. Two iron-repressed outer membrane proteins were observed in the Pseudomonas sp., neither of which had electrophoretic mobility identical to that of the aerobactin outer membrane receptor protein from E. coli. DNA hybridization assays showed no hybridization to the aerobactin genes from the E. coli plasmid pColV, indicating that the genetic determinants for aerobactin production by Pseudomonas strain X40 differ substantially from those found in the archetypic enteric plasmid pColV-K30.     

Production of the siderophore aerobactin by a halophilic pseudomonad.

A bacterial strain, isolated from a cyanobacterial culture, was identified as Pseudomonas sp. strain X40. Under iron-limiting conditions, the Pseudomonas sp. produced aerobactin, a dihydroxamate siderophore previously found only in the family Enterobacteriaceae. Aerobactin was identified by electrophoretic mobility, spectrophotometric titration, proton nuclear magnetic resonance spectroscopy, mass spectrometry, acid hydrolysis, and biological activity. Aerobactin was used as a siderophore in the Pseudomonas sp. and Escherichia coli. Two iron-repressed outer membrane proteins were observed in the Pseudomonas sp., neither of which had electrophoretic mobility identical to that of the aerobactin outer membrane receptor protein from E. coli. DNA hybridization assays showed no hybridization to the aerobactin genes from the E. coli plasmid pColV, indicating that the genetic determinants for aerobactin production by Pseudomonas strain X40 differ substantially from those found in the archetypic enteric plasmid pColV-K30.     

Kinetics study of pyridine biodegradation by a novel bacterial strain,Rhizobium sp. NJUST18

Biodegradation of pyridine by a novel bacterial strain, Rhizobium sp. NJUST18, was studied in batch experiments over a wide concentration range (from 100 to 1,000 mg l^?1). Pyridine inhibited both growth of Rhizobium sp. NJUST18 and biodegradation of pyridine. The Haldane model could be fitted to the growth kinetics data well with the kinetic constants μ* = 0.1473 h^?1, K _s = 793.97 mg l^?1, K _i = 268.60 mg l^?1 and S _m = 461.80 mg l^?1. The true μ _max, calculated from μ*, was found to be 0.0332 h^?1. Yield coefficient Y _X/S depended on S _i and reached a maximum of 0.51 g g^?1 at S _i of 600 mg l^?1. V _max was calculated by fitting the pyridine consumption data with the Gompertz model. V _max increased with initial pyridine concentration up to 14.809 mg l^?1 h^?1. The q _S values, calculated from $V_{ \hbox{max} }$ , were fitted with the Haldane equation, yielding q _Smax = 0.1212 g g^?1 h^?1 and q* = 0.3874 g g^?1 h^?1 at S _m′ = 507.83 mg l^?1, K _s′ = 558.03 mg l^?1, and K _i′ = 462.15 mg l^?1. Inhibition constants for growth and degradation rate value were in the same range. Compared with other pyridine degraders, μ _max and S _m obtained for Rhizobium sp. NJUST18 were relatively high. High K _i and K _i′ values and extremely high K _s and K _s′ values indicated that NJUST18 was able to grow on pyridine within a wide concentration range, especially at relatively high concentrations.     

Kinetics and modeling of autotrophic thiocyanate biodegradation

The biokinetic parameters for autotrophic systems are difficult to obtain and are often mistakenly determined because the size of the autotrophic population in mixed (i.e., heterotrophic and autotrophic) cultures cannot be accurately estimated. This article presents a systematic approach, combining bioenergetic calculations and experimental data, to obtain values of the biokinetic parameters pertinent to the aerobic, autotrophic biodegradation of thiocyanate. Nonlinear regression techniques were employed using both initial thiocyanate utilization rate data and single thiocyanate depletion curves. Both types of data were necessary to overcome the problems arising from the linear nature of the substrate depletion curves and the high correlation of the biokinetic model parameters inherent in nonlinear regression analysis. The aerobic biodegradation of thiocyanate followed a substrate inhibition pattern that was successfully described by the Haldane-Andrews model. Although regression analysis did not yield unique biokinetic parameter estimates, the following parameter value ranges were obtained: maximum specific substrate utilization rate (k), 0.26 to 0.44 mg SCN-/mg biomass h; half-saturation coefficient (Ks), 2.3 to 7.1 mg SCN-/L; and inhibition coefficient (Ki), 28 to 109 mg SCN-/L. Based on the estimated biokinetic parameter values, a design and operation diagram was constructed that depicts the steady-state thiocyanate concentration as a function of solids retention time for a completely mixed, continuous-flow reactor.     

Degradation of triphenyltin by a fluorescent pseudomonad

Triphenyltin (TPT)-degrading bacteria were screened by a simple technique using a post-column high-performance liquid chromatography using 3,3',4',7-tetrahydroxyflavone as a post-column reagent for determination of TPT and its metabolite, diphenyltin (DPT). An isolated strain, strain CNR15, was identified as Pseudomonas chlororaphis on the basis of its morphological and biochemical features. The incubation of strain CNR15 in a medium containing glycerol, succinate, and 130 microM TPT resulted in the rapid degradation of TPT and the accumulation of approximately 40 microM DPT as the only metabolite after 48 h. The culture supernatants of strain CNR15, grown with or without TPT, exhibited a TPT degradation activity, whereas the resting cells were not capable of degrading TPT. TPT was stoichiometrically degraded to DPT by the solid-phase extract of the culture supernatant, and benzene was detected as another degradation product. We found that the TPT degradation was catalyzed by low-molecular-mass substances (approximately 1,000 Da) in the extract, termed the TPT-degrading factor. The other fluorescent pseudomonads, P. chlororaphis ATCC 9446, Pseudomonas fluorescens ATCC 13525, and Pseudomonas aeruginosa ATCC 15692, also showed TPT degradation activity similar to strain CNR15 in the solid-phase extracts of their culture supernatants. These results suggest that the extracellular low-molecular-mass substance that is universally produced by the fluorescent pseudomonad could function as a potent catalyst to cometabolite TPT in the environment.     

Degradation of 2-fluorobenzoate by a pseudomonad

Pseudomonas (spp), isolated from a complex petrochemical sludge, was able to utilize 2-fluorobenzoate as its sole source of carbon and energy. At the end of the growth phase, about 42% of the organically bound fluoride was released. Catechol, 3-fluorocatechol, and 6-fluorodihydrodihydroxybenzoate were confirmed as intermediates by chromatographic and spectral analyses. During 2-fluorobenzoate metabolism, fluoride is eliminated before the aromaticity of the ring is lost. Twofold higher levels of catechol 1,2-oxygenase were detected in 2-fluorobenzoate-grown cells compared with cells grown on benzoate. When used as assay substrates, 3-chlorocatechol showed less catechol 1,2-oxygenase activity than catechol or 4-chlorocatechol. The ability to degrade 4-fluorobenzoate could be transferred toPseudomonas (spp) by the conjugal transfer of plasmid pWR1 fromPseudomonas sp. B13.     

N-Alkane oxidation enzymes of a pseudomonad.

A nicotinamide adenine dinucleotide (NAD)-dependent n-alkane dehydrogenase and an NAD phosphate (reduced form)-dependent alkane hydroxylase have been purified from cell-free extracts of Pseudomonas sp. strain 196Aa grown anaerobically on n-alkane. The n-alkane dehydrogenase (fraction R-3), obtained as a single peak from Bio-Gel P-60, showed an overall 135-fold purification and was demonstrated by infrared spectroscopy and gas chromatography to convert n-decane to 1-decene. The alkene hydroxylase activity in the S-3 fraction, purified 167 times from diethylaminoethyl-cellulose, was shown by the same methodology to convert decene to decanol. Commercial ferredoxin has been shown to increase the alkane dehydrogenase activity. An NAD-, flavine adenine dinucleotide-, and iron-dependent alcohol dehydrogenase was demonstrated in the R-3 fraction. A mechanism for the anaerobic conversion of n-alkane to fatty acid has been proposed.     

Kinetics of biodegradation of free gossypol by Candida tropicalis in solid-state fermentation

In the present work experiments were carried out to study the effect of free gossypol on the growth of Candida tropicalis ZAU-1, evaluate its ability in biodegrading free gossypol, analyze the time course of solid-state fermentation, and model the microbial growth by determining the kinetics of dry matter weight loss, total carbohydrate concentration and the free gossypol content during solid-state fermentation. Results showed that the biomass in inorganic salts glucose medium were unaffected by free gossypol at 500 and 1000 mg/l levels, compared with the control group (p > 0.05); degradation of free gossypol reached 95.12% and 94.12%, respectively. A logistic equation (R² = 0.9922), describing the growth model of C. tropicalis ZAU-1 was obtained, with the maximum values of u_m and X_m at 0.0970 h⁻¹ and 21.8631% of dry matter weight loss, respectively. A good-fit curvilinear regression model was achieved to describe the change pattern of total carbohydrate concentration (R² = 0.9910), and the biodegradation pattern of free gossypol (R² = 0.9825). These models could be used to predict the fermentation course by C. tropicalis ZAU-1 under solid-state fermentation.