A dynamic river model for biodegradability studies

The objective of this publication is to present a new dynamic aerobic biodegradation test method simulating a river. A laboratory cascade test system and standardized batch shake flask tests were used for biodegradation studies with the non-volatile and non-sorbing model compounds 2,4-dinitrophenol, naphthalene-1-sulphonic acid and sulphanilic acid. To be closer to the often very low concentrations of substances in the environment the concentrations of the compounds used were standard test concentrations and lower. ¹⁴C labelled compounds were measured at 50 g/l, capillary electrophoresis at 5000 g/l and the removal of dissolved organic carbon at 50000 g/l. The test results obtained confirmed the known ultimate biodegradability of the test compounds and showed that biodegradation degrees, rates and degradation durations depended on the test systems, the concentrations of test compounds and the inocula. The river model is a suitable simulation test for natural dynamic surface waters which can be used to perform biodegradability studies at low test concentrations if adequate analytical tools, preferably radioactive-labelled substances, are available.Abbreviations BOD biochemical oxygen demand - DAWT DOC-die-away test - DNP 2,4-dinitrophenol - DOC dissolved organic carbon - E effluent of laboratory wastewater treatment plants - MOST modified OECD screening test - NSA naphthalene-1-sulphonic acid - P pond water - SAA sulphanilic acid (=4-amino benzene sulphonic acid)     

Biodegradation of phthalic acid esters in river water and activated sludge.

The primary and ultimate biodegradability of phthalic acid, monobutyl phthalate, and five structurally diverse phthalic acid ester plasticizers in river water and activated sludge samples were determined via ultraviolet spectrophotometry, gas chromatography, and CO2 evolution. The compounds studied underwent rapid primary biodegradation in both unacclimated river water and acclimated activated sludge. When activated sludge acclimated to phthalic acid esters was used as the inoculum for the CO2 evolution procedure, greater than 85% of the total theoretical CO2 was evolved. These studies demonstrate that the phthalic acid ester plasticizers and intermediate degradation products readily undergo ultimate degradation in different mixed microbial systems at concentrations ranging from 1 to 83 mg/liter.     

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.     

Degradation of phenol and phenolic compounds by a defined denitrifying bacterial culture

Phenol, a major pollutant in several industrial waste waters is often used as a model compound for studies on biodegradation. This study investigated the anoxic degradation of phenol and other phenolic compounds by a defined mixed culture of Alcaligenes faecalis and Enterobacter species. The culture was capable of degrading high concentrations of phenol (up to 600 mg/l) under anoxic conditions in a simple minimal mineral medium at an initial cell mass of 8 mg/l. However, the lag phase in growth and phenol removal increased with increase in phenol concentration. Dissolved CO₂ was an absolute requirement for phenol degradation. In addition to nitrate, nitrite and oxygen could be used as electron acceptors. The kinetic constants, maximum specific growth rate _max; inhibition constant, K _i and saturation constant, K _s were determined to be 0.206 h^–1, 113 and 15 mg phenol/l respectively. p-Hydroxybenzoic acid was identified as an intermediate during phenol degradation. Apart from phenol, the culture utilized few other monocyclic aromatic compounds as growth substrates. The defined culture has remained stable with consistent phenol-degrading ability for more than 3 years and thus shows promise for its application in anoxic treatment of industrial waste waters containing phenolic compounds.     

Effect of Concentration of Organic Chemicals on Their Biodegradation by Natural Microbial Communities

The effect of concentration on the biodegradation of synthetic organic chemicals by natural microbial communities was investigated by adding individual ¹⁴C-labeled organic compounds to stream water at various initial concentrations and measuring the formation of ¹⁴CO₂. The rate of degradation of p-chlorobenzoate and chloroacetate at initial concentrations of 47 pg/ml to 47 μg/ml fell markedly with lower initial concentrations, although half or more of the compound was converted to CO₂ in 8 days or less. On the other hand, little mineralization of 2,4-dichlorophenoxyacetate and 1-naphthyl-N-methylcarbamate, or the naphthol formed from the latter, occurred when these compounds were present at initial concentrations of 2 to 3 ng/ml or less, although 60% or more of the chemical initially present at higher concentrations was converted to CO₂ in 6 days. It is concluded that laboratory tests of biodegradation involving chemical concentrations greater than those in nature may not correctly assess the rate of biodegradation in natural ecosystems and that low substrate concentration may be important in limiting biodegradation in natural waters.     

Synthesis and Antioxidant Activity of New Norcantharidin Analogs

New norcantharidin analogs were designed and obtained as compounds with biological activity. As a starting material, exo‐7‐oxabicyclo[2.2.1]heptane‐2,3‐dicarboxylic acid anhydride was used. Three groups of compounds: dicarboximides, triazoles and thiazolidines were obtained in multistep reactions. The ¹H‐ and ¹³C‐NMR spectra were used to confirm the structures of all obtained products and they were in agreement with the proposed structure of substances. All derivatives were screened for their antioxidant activity. The most promising group was dicarboximides ( 1 – 4 , 6 ). Derivatives 2–4 displayed antioxidant activity with EC₅₀=7.75–10.89 μg/ml, which may be comparable to strong antioxidant Trolox (EC₅₀=6.13 μg/ml). Excellent activity with EC₅₀=10.75 μg/ml also presented norcantharidin analog with 1,2,4‐triazole system ( 12 ).     

High-performance liquid chromatographic method for guanylhydrazone compounds

A high-performance liquid chromatographic method has been developed for a series of aromatic guanylhydrazones that have demonstrated therapeutic potential as anti-inflammatory agents. The compounds were separated using octadecyl or diisopropyloctyl reversed-phase columns, with an acetonitrile gradient in water containing heptane sulfonate, tetramethylammonium chloride, and phosphoric acid. The method was used to reliably quantify levels of analyte as low as 785 ng/ml, and the detector response was linear to at least 50 μg/ml, using a 100 μl injection volume. The assay system was used to determine the basic pharmacokinetics of a lead compound, CNI-1493, from serum concentrations following a single intravenous injection in rats.     

Biodegradation kinetics of LAS in river water

This article reports the biodegradation kinetics of linear alkylbenzene sulfonates (LAS) in river water. The authors used the ‘river die-away' test method and high performance liquid chromatography to monitor LAS concentrations as functions of time in a series of tests systems. Controlled variables included initial LAS concentrations and incubation temperature. The kinetic parameters computed from the experimental data demonstrated strong correlations (r²>0.99) with theoretical values computed from the kinetic model presented in this paper. The proposed model accurately predicts concentrations of non-biodegradable substrate and the maximum specific microbial growth rate.     

Biodegradation process and the nature of metabolism of metalaxyl in soil

The enhanced biodegradation of metalaxyl was studied in tobacco, citrus, avocado and corn soils. The most rapid degradation of metalaxyl occurred in a tobacco soil in which the half-life (50% degradation) of metalaxyl was 6 days. The main breakdown product of metalaxyl in all soils was the acid metabolite. Ring labelled [¹⁴C]metalaxyl incubated for 4 wk in 6 soils demonstrated a low rate of ¹⁴CO₂ evolution ranging from 2.1% to 11.3% which was unrelated to the biodegradation properties of the soil. A relationship between the concentration of metalaxyl and the subsequent rate of biodegradation was found in the tobacco soils. Higher concentrations of metalaxyl resulted in faster biodegradation rates. A single exposure of tobacco and corn soils to metalaxyl (100 μg/ml or 200 μg/g dry weight of soil) significantly increased their subsequent capacity to degrade the fungicide. Addition of the fungicide thiram or the antibiotics streptomycin and chloramphenicol to an avocado soil resulted in 75% and 51% inhibition of metalaxyl degradation, respectively. A combination of the fungicide and antibiotics resulted in 89% inhibition. The results indicate that enhanced microbial degradation of metalaxyl can occur in a wide range of soils. Under experimental conditions using soil solutions or soil systems, a single application of the fungicide may trigger this event. A wide range of fungi and bacteria appear to take part in degrading metalaxyl.     

CHLOROTETRACYCLINE-ASSOCIATED FLUORESCENCE CHANGES DURING CALCIUM UPTAKE AND RELEASE BY RAT BRAIN SYNAPTOSOMES1

Abstract– The fluorescent divalent metal chelate-probe, chlorotetracycline (CTC), was used as a dynamic monitor of calcium association with rat brain snynaptosomes. The determined fluorescence excitation and emission maxima, 412 nm and 522 nm respectively, were used to monitor membrane-calcium interactions as a function of various parameters. Positive correlations were observed between increased or decreased fluorescence quantum yield and the uptake of both CTC and ⁴⁵Ca by synpatosomes. The divalent metal ionophore A23187 enhanced fluorescence as well as probe and ⁴⁵Ca uptake. Whereas, the polar chelator, EGTA, markedly reduced fluorescence, and the synaptosomal bound CTC and ⁴⁵Ca. The CTC fluorescence changes also demonstrated the saturable manner in which ⁴⁵Ca bound synaptosomes. At concentrations greater than 100μg/ml, CTC bound to the synaptosomes in a manner which quenched fluorescence at 522 nm. Also, CTC, at concentrations above 15 μg/ml, enhanced the uptake of ⁴⁵Ca. At CTC concentrations between 10 and 15 μg/ml the quenching and iono-phoretic properties of the probe were minimized without affecting the capability of using the probe to visualize calcium interactions with synaptosomal membranes. Also, at a low CTC concentration (12.5 μg/ml) the inhibition of calcium uptake by increasing monovalent ion concentrations was clearly demonstrated.     

Quantitative determination of naproxen in plasma by a simple high-performance liquid chromatographic method

A high-performance liquid chromatographic method for the determination of naproxen in plasma is described. The technique is based on the single extraction of the drug from acidified plasma with chloroform using 2-naphthalene acetic acid as internal standard. The chromatographic system consisted of a column packed with Spherisorb ODS (5 μm); the mobile phase was acetonitrile—phosphoric acid (pH 3) (45:55, v/v).The method can accurately measure plasma naproxen concentrations down to 1 μg/ml using 100 μl of sample, with no interference from endogenous compounds. The coefficients of variation of the method at 120 μg/ml and 1 μg/ml are 2.8 and 21.6%, respectively, and the calibration curve is linear. The method described is very suitable for routine clinical and pharmacokinetic studies.     

Isolation and some physiological properties of natural plant growth inhibitors

Using paper chromatography and conventional methods of isolation, natural growth inhibitors were isolated from green leaves of different plants (Brassica oleracea, Zea mays, Pisum sativum andSalix rubra). All isolated inhibitors were found to be phenolic compounds and the chemical structure of most of them was determined; only the final structure of theBrassica inhibitor has not yet been ascertained. 500 mg of natural inhibitor ofPisum sativum was isolated from 1500 g of leaves and was identified as quercetin-glucosil-p-coumarate (QGC), described earlier byFuruya, Galston andStowe (1961). The structure of the natural inhibitor ofZea mays (4 mg from 100 g of leaves) was identical with p-coumaric acid and the chemical nature of the plant growth inhibitor fromSalix rubra (700 mg from 1,5 kg of leaves and young bark) was that of 2-chalconaringenin-glucoside or isosalipurposide, described earlier byCharaux andRabaté (1931) andHarborne (1966). All isolated substances had inhibiting properties in the straight growth test of wheat coleoptile sections and decreased the growth of isolated stem sections prepared from plants—donors of inhibitors. Thus, maximum growth inhibition (LG max) was attained, if wheat coleoptile sections were incubated with:Brassica inhibitor in the concentration of 0·5 mg/ml, withPisum inhibitor (QGC) in the concentration of 16 mg/ml, withZea inhibitor (p-coumaric acid)—0·35 mg/ml and with Salix inhibitor (isosalipurposide) in the concentration of 0·5 mg/ml. In small concentrations no mentioned substances were able to enhance the growth as actively as indolic auxins (on 250–300%); only slight growth activation in biotests was sometimes observed for low concentrations. Inhibition in p-coumaric acid was much more active in a free form than in the bound form as an acyl-rest of QGC. As a rule, the wheat coleoptile test was much more sensitive (3–5 times) to the plant growth inhibitors, than tests prepared from tissue and organs of plants—donors. The retardation activity of plant growth inhibitors is not correlated with their molecular weight. Dormin (or±abscissin II) was also tested on wheat coleoptile sections. In neither of the applied concentrations (10-0·05 μg/l range) was dormin able to depress straight growth of wheat coleoptile sections, but even in a 1·7 μg/l concentration it inhibited the IAA-activated growth of sections. However, additional experiments showed that dormin in higher concentrations (40 μg/l and more was able even to depress endogenous straight growth of wheat coleoptise sections. The differences between the properties of natural phenolic growth inhibitors and dormin were discussed.     

Degradation of polycyclic aromatic hydrocarbons and long chain alkanes at 6070 °C by Thermus and Bacillus spp

Although polycyclic aromatic hydrocarbons (PAH) and alkanesare biodegradable at ambient temperature, in some cases low bioavailabilities are thereason for slow biodegradation. Considerably higher mass transfer rates and PAH solubilities and hence bioavailabilities can be obtained at higher temperatures. Mixed and pure cultures of aerobic, extreme thermophilic microorganisms (Bacillus spp., Thermus sp.) were used to degrade PAH compounds and PAH/alkane mixtures at 65 °C. The microorganismsused grew on hydrocarbons as sole carbon and energy source. Optimal growthtemperatures were in the range of 60–70 °C at pH values of 6–7. The conversion of PAH with 3–5 rings (acenaphthene, fluoranthene, pyrene, benzo[e]pyrene) was demonstrated. Efficient PAH biodegradation required a second, degradable liquid phase. Thermus brockii Hamburg metabolized up to 40 mg (l h)^-1 pyrene and 1000 mg(1 h)^-1 hexadecane at 70 °C. Specific growth rates of 0.43 h^-1 were measured for this strain with hexadecane/pyrene mixtures as the sole carbon and energy source in a 2-liter stirred bioreactor. About 0.7 g cell dry weight were formed from 1 g hydrocarbon. The experiments demonstrate the feasibility and efficiency of extreme thermophilic PAH and alkane biodegradation.     

Removal of BTEX compounds by industrial sludge microbes in batch systems: statistical analysis of main and interaction effects

Biodegradation is an effective technique to remediate polluted soil and groundwater. In the present experimental study, a mixed microbial culture obtained from the wastewater treatment sludge of a chemical industry was used to degrade liquid phase benzene, toluene, ethyl benzene, and xylene (BTEX), at individual initial concentrations varying between 15 and 75 mg/l. Experiments were conducted according to 2 ^k−1 fractional factorial design at the low (15 mg/l) and high (75 mg/l) levels of BTEX concentrations, to identify the main and interaction effects of parameters and their influence on biodegradation of individual BTEX compounds in mixtures. The individual removals varied between 16% and 75% when the concentrations of B, T, E, and X were sufficiently low in the mixture. However, both synergistic (removal of ethyl benzene) and antagonistic (removal of benzene) behavior were noticed when the concentrations of toluene and xylene was increased to higher levels. The individual removals were greater than 67% at their center point levels. The total BTEX removal values were later statistically analyzed and based on the Fischer’s variance ratio (F) and Probability values (P) it was observed that the main effects for total BTEX removal were significant than the squared and interaction effects.     

Assessment of biostimulation and bioaugmentation for removing chlorinated volatile organic compounds from groundwater at a former manufacture plant

Site in a former chemical manufacture plant in China was found contaminated with high level of chlorinated volatile organic compounds (CVOCs). The major contaminants chloroform (CF), 1,2-dichloroethane (1,2-DCA) and vinyl chloride (VC) in groundwater were up to 4.49 × 10⁴, 2.76 × 10⁶ and 4.35 × 10⁴ μg/L, respectively. Ethene and methane were at concentrations up to 2219.80 and 165.85 μg/L, respectively. To test the hypothesis that the CVOCs in groundwater at this site could be removed via biodegradation, biomarker analyses and microcosm studies were conducted. Dehalococcoides 16S rRNA gene and VC-reductase gene vcrA at densities up to 1.5 × 10⁴ and 3.2 × 10⁴ copies/L were detected in some of the groundwater samples, providing strong evidence that dechlorinating bacteria were present in the aquifer. Results from the microcosm studies showed that at moderate concentrations (CF about 4000 μg/L and 1,2-DCA about 100 μg/L), CF was recalcitrant under natural condition but was degraded under biostimulation and bioaugmentation, while 1,2-DCA was degraded under all the three conditions. At high concentration (CF about 1,000,000 μg/L and 1,2-DCA about 20,000 μg/L), CF was recalcitrant under all the three treatments and 1,2-DCA was only degraded under bioaugmentation, indicating that high concentrations of contaminants were inhibitory to the bacteria. Electron donors had influence on the degradation of contaminants. Of the four fatty acids (pyruvate, acetate, propionate and lactate) examined, all could stimulate the degradation of 1,2-DCA at both moderate and high concentrations, whereas only pyruvate and acetate could stimulate the degradation of CF at moderate concentration. In the microcosms, the observed first-order degradation rates of CF and 1,2-DCA were up to 0.12 and 0.11/day, respectively. Results from the present study provided scientific basis for remediating CVOCs contaminated groundwater at the site.     

Stimulative effect of prostaglandins on production of hexosamine-containing substances by cultured fibroblasts (2) early effect of various prostaglandins at various doses

Early effects of various prostaglandins on the production of hexosamine-containing substances by cultured fibroblasts, which were derived from a rat carrageenin granuloma, were studied. At the stationary phase, the cells were exposed for 6 h to one of the prostaglandin A₁ (PGA₁), A₂, B₁, B₂, D₂, F_1α, F_2α, E₁, E₂ or arachidonic acid in various concentrations ranging from 0.01 to 10 μg/ml for all the stimuli and from 10 pg to 10 μg/ml for PGF_2α. The activity of the cells in incorporating ³H-glucosamine into hexosamine-containing substances (acidic) glycosaminoglycans and glycoproteins) during this period was compared with that of control cells. All the stimuli tested showed more or less stimulative effect on the synthesis of hexosamine-containing substances at their specific concentrations. PGF_2α was found to be the most potent stimulant and its stimulative effect was found significant even at the low concentration of 100 pg/ml. PGD₂, F_1α and E₂ were the next potent stimuli. Their optimum dose were around 1 μg/ml but they still had significant stimulation at the concentration of 0.01 μg/ml. Effect of PGE₂ was rather mild. Stimulation by PGA₁, A₂, B₁ and B₂ or arachidonic acid was seen at high dose, and its seemed to be non-specific. The results suggested that these prostaglandins such as PGF_2α, D₂, F_1α and E₂ play some important role on regulating the production of intercellular ground substances.     

Isolement,action biologique et evolution des substances paragoniales contenues dans le spermatophore d'Acanthoscelides obtectus (Coleoptère)

In Acanthoscelides obtectus, some male secretions deposited in the spermatophore during mating reach the blood of the females and stimulate oögenesis. Water extracts from spermatophores injected into a female abdomen stimulate oögenesis but do not influence egg-laying or sexual receptivity. After column chromatograph of spermatophores, aqueous extracts on Sephadex G 25 Coarse, G 25 Superfine, and G 15, an active fraction has been isolated. This injected into the abdomen of virgin females stimulates oögenesis at low concentrations, but it is toxic at higher concentrations. This fraction was examined by paper electrophoresis at low voltage and then chromatographed on G 10 Sephadex. Two peaks were obtained: the first corresponds to the paragonial substance A which stimulates oögenesis at 0,2 10^?3 μg/μl concentration. The second contains the paragonial substance B. At a 0,3 10^?3 ug/μl concentration this substance is toxic. First this toxicity inhibits oögenesis and then causes the death of most females at higher concentrations. The toxic effect appears 2 or 3 days after injection. These two substances are purified on paper chromatography and the biological activities are contained in a zone of R_f 0.25 to 0.45 (paragonial substance A) and in a zone of 0.16–0.30 R_f (paragonial substance B).The paragonial substances disappear from the spermatophore after mating. Aqueous extracts of spermatophores obtained 6 hr after mating do not stimulate oögenesis and do not have any toxic effect. The chemical nature of these both fractions is not yet determined because the quantity of extracts obtained at the end of the purification is very low.The action of both paragonial substances is similar to the action of hormones. The paragonial substances influence unknown receptors at low concentration after a latent period. The origin of the paragonial B substance was not determined, but this substance which inhibits oögenesis at low concentrations could be an antagonist of paragonial A substance.     

Effects of microcosm preparation on rates of toluene biodegradation under denitrifying conditions

Microcosms were prepared with subsurface material from two aquifers to examine the effects of preparation methods on rates of toluene biodegradation under denitrifying conditions. In both cases, the data fit a zero-order kinetics plot. However, rates of removal were generally proportional to initial toluene concentrations, resulting in similar half-lives. Increasing the solid/liquid mass ratio resulted in decreased lag times in one aquifer material, although in both cases the specific toluene mass removal rate (g toluene g sediment ⁻¹ day⁻¹) also decreased. Varying either the initial toluene concentrations or the solid/liquid ratios by two to three orders of magnitude resulted in a half-life variation of only a factor of two, with most of the differences occurring at the extreme ranges of the test variables. These data indicate that similar biodegradation rates might be expected from microcosms prepared with different contaminant concentrations and solid/liquid ratios, which is useful for design of microcosm studies to evaluate biodegradation at field sites. Received 14 March 1996/ Accepted in revised form 24 July 1996     

Effects of Adaptation on Biodegradation Rates in Sediment/Water Cores from Estuarine and Freshwater Environments

Experiments were devised to determine whether exposure to xenobiotics would cause microbial populations to degrade the compounds more rapidly during subsequent exposures. Studies were done with water/sediment systems (ecocores) taken from a salt marsh and a river. Systems were tested for adaptation to the model compounds methyl parathion and p-nitrophenol. ¹⁴CO₂ released from radioactive parent compounds was used as a measure of mineralization. River populations preexposed to p-nitrophenol at concentrations as low as 60 μg/liter degraded the nitrophenol much faster than did control populations. River populations preexposed to methyl parathion also adapted to degrade the pesticide more rapidly, but higher concentrations were required. Salt marsh populations did not adapt to degrade methyl parathion. p-Nitrophenol-degrading bacteria were isolated from river samples but not from salt marsh samples. Numbers of nitrophenol-degrading bacteria increased 4 to 5 orders of magnitude during adaptation. Results indicate that the ability of populations to adapt depends on the presence of specific microorganisms. Biodegradation rates in laboratory systems can be affected by concentration and prior exposure; therefore, adaptation must be considered when such systems are used to predict the fate of xenobiotics in the environment.