Degradation of 6-aminonaphthalene-2-sulphonic acid by mixed cultures: kinetic analysis

Summary The degradation of 6-aminonaphthalene-2-sulphonic acid (6A2NS) by mixed cultures via an interspecies transfer of 5-aminosalicylic acid (5AS) was investigated using a continuous chemostat culture. Two different bacterial communities were employed. Steady-state data were obtained from a multi-species culture only, but not from a defined two-species culture. Experimental data showed the conversion of 6A2NS into 5AS to be rate-determining for degradation. Both the Monod equation, and an extended model regarding the interspecies transfer of 5AS, were found to be suitable to describe the relationship between biomass and substrate concentration, depending on the flow rate of continuous culture. Substrate consumed for endogenous metabolism was considred according to Pirt (1975).     

Enantioselective degradation of the herbicide mecoprop [2-(2-methyl-4-chlorophenoxy) propionic acid] by mixed and pure bacterial cultures

Abstract A consortium of three bacteria was isolated from top soil through their capacity to utilise the chlorinated, aromatic herbicide mecoprop as a single growth substrate. The consortium constituted a tight association of Alcaligenes denitrificans, Pseudomonas glycinea and Pseudomonas marginalis . The culture exclusively degraded the ( R )-(+)-isomer of the herbicide while the ( S )-(−)-enantiomer remained unaffected. The mecoprop-degrading community could also degrade 2,4-dichlorophenoxyacetic acid, 2-methyl-4-chlorophenoxyacetic acid and racemic 2-phenoxypropionic acid. Initially, no single member of the consortium was able to degrade mecoprop as a pure culture but after prolonged incubation, A. denitrificans was able to grow on the herbicide as the sole source of carbon and energy.     

Mineralization of 2-aminobenzenesulfonate by a bacterial consortium

A variety of environmental inocula were tested for the development of 2-aminobenzenesulfonate (2-ABS, Orthanilic acid) degrading bacterial enrichment. A bacterial consortium (BC), which could utilize 2-ABS as the sole carbon and energy source, could only be developed from the sludge derived from a wastewater treatment unit of a large chemical industry manufacturing nitro and aminoaromatics. BC consisted of two bacterial strains. Based on 16S rDNA sequence analysis, these strains were identified to be belonging to the genus, Acinetobacter and Flavobacterium. The consortium could degrade 1,000 mg l⁻¹ 2-ABS within 40 h. Evidence for the extensive mineralization of 2-ABS, during the growth of BC, was derived from U.V-spectral and total organic carbon analysis. BC was highly specific for 2-ABS, as other benzene sulfonates tested in this study, including other ABS isomers, were not utilized as growth substrates. 2-ABS removal pattern in the presence of glucose was significantly influenced by acclimation characteristics of the culture. Consortium adapted to 2-ABS/glucose demonstrated the concomitant removal of both substrates, whereas glucose exerted catabolic repression on 2-ABS removal with glucose adapted culture. Presence of chloramphenicol inhibited 2-ABS degradation by cells, pregrown on succinate, indicating that the 2-ABS degrading enzymes are inducible in nature. Thus the presence of 2-ABS is essential for maintaining the high degradation potential. This enrichment culture can find an application in the treatment of 2-ABS containing wastewaters.     

Preliminary studies on the toxicity and mutagenicity of 1-amino-2-naphthol-4-sulphonic acid inDrosophila melanogaster

The toxicity and mutagenicity of 1-amino-2-naphtho-4-sulphonic acid were analysed inDrosphila melanogaster. Rate of development and viability were the two parameters employed to study the toxicity. The frequency of dominant lethals was scored to evaluate the mutagenic effect of the chemical on male and female germ cells. Concentrations of 250 mg and above/100 ml wheat cream agar medium were found to be significantly toxic. Significant number of dominant lethals was induced even by a concentration as low as 50 mg/100 ml medium. Male germ cells were more sensitive than female germ cells.     

一株金黄杆菌对2-吡啶甲酸的好氧生物降解

【背景】2-吡啶甲酸具有高毒性、致癌性,能长期稳定存在于水体中,从而对环境造成危害。【目的】开发一种能够高效经济处理含2-吡啶甲酸废水的技术。【方法】筛选一株在好氧条件下以2-吡啶甲酸为唯一碳、氮、能源的菌株,考察该菌株的降解性能,建立降解动力学模型。【结果】经过16S r RNA基因序列分析,该菌株被鉴定为金黄杆菌(Chryseobacterium sp.),命名为ZD2。当2-吡啶甲酸初始浓度为100、200、400、600和800 mg/L时,ZD2完全降解2-吡啶甲酸的时间分别为10、18、22、78和114 h。零级动力学模型较好地描述了2-吡啶甲酸的降解行为,当初始浓度为100-400 mg/L时,降解速率常数随着浓度的增加而增加,并于400 mg/L时达到最大;600-800 mg/L时,降解速率常数开始下降,呈现抑制作用。【结论】菌株ZD2对2-吡啶甲酸的降解效果较好,能够为处理实际的2-吡啶甲酸工业废水提供理论依据。     

Extractionless determination of 6-methoxy-2-naphthylacetic acid, a major metabolite of nabumetone, in human plasma by high-performance liquid chromatography

Following oral administration of the prodrug nabumetone, the major metabolite 6-methoxy-2-naphthylacetic acid (6-MNA) was determined in human plasma. Minimal sample preparation was followed by reversed-phase liquid chromatography and UV detection, affording high sample throughput. The lower limit of quantification (LLOQ) was 70 ng/ml, at a signal-to-noise ratio of 8:1. The assay method displayed good correlation (r=0.997), and can be readily employed in pharmacokinetic and bioequivalence studies.     

Syntrophic interactions during degradation of 4-aminobenzenesulfonic acid by a two species bacterial culture

During synthrophic growth of Hydrogenophaga palleronii (strain S1) and Agrobacterium radiobacter (strain S2) with 4-aminobenzene sulfonate (4ABS) only strain S1 desaminates 4ABS by regioselective 3,4-dioxygenation. The major part of the metabolite catechol-4-sulfonate (4CS) is excreted and further metabolized by strain S2. Although both organisms harbour activities of protocatechuate pathways assimilation of the structural analog 4CS requires first of all enzyme activities with broader substrate specificity: protocatechuate 3,4-dioxygenase and carboxymuconate cycloisomerase activities were identified which in addition to the natural substrates also convert 4CS requires first of all enzyme activities with Carboxymethyl-4-sulfobut-2-en-4-olide (4SL) was identifed as a metabolite. Its further metabolism requires a desulfonating enzyme which eliminates sulfite from (4SL) and generates maleylacetate. Convergence with the 3-oxoadipate pathway is catalyzed by a maleyl acetate reductase, which was identified in cell-free extracts of both organisms S1 and S2. Characteristically, only strain S1 can oxidize sulfite and thus contributes to the interdependence of the two bacteria during growth with 4ABS.     

Studies on the anaerobic degradation of benzoic acid and 2-aminobenzoic acid by a denitrifying Pseudomonas strain

The growth of a denitrifying Pseudomonas strain on benzoic acid and 2-aminobenzoic acid (anthranilic acid) has been studied. The organism grew aerobically on benzoate, 2-aminobenzoate, and gentisate, but not on catechol or protocatechuic acid. These and other findings suggest that aerobic degradation of benzoic acid was via gentisic acid. Under completely anaerobic conditions in the presence of nitrate, benzoate and 2-aminobenzoate (5 mM each) were oxidized to CO₂ with the concurrent reduction of NO ₃ ^- to NO ₂ ^- . Only after complete NO ₃ ^- consumption was NO ₂ ^- reduced to N₂. Cells contained a NADP-specific 2-oxoglutaate dehydrogenase, in contrast to a NAD-specific pyruvate dehydrogenase. During anaerobic metabolism of [carboxyl-¹⁴C]benzoic acid, 16% of the label of metabolized benzoic acid was incorporated into cell material; this excludes intermediary decarboxylation during anaerobic metabolism. Extracts catalysed the activation of benzoic acid and a variety of its derivatives to the respective aryl-coenzyme A thioesters, ATP being cleaved to AMP and PP_i; two synthetase activites were present. Extracts from 2-aminobenzoate-grown cells catalyzed a NADH-dependent reduction of 2-aminobenzoyl-CoA (100 nmol·min^-1·mg^-1 cell protein) to an unidentified CoA thioester, with a stoichiometric release of NH₃ and a stoichiometry of 3 mol NADH oxidized per mol 2-aminobenzyol-CoA reduced when tested under aerobic conditions. The 2-aminobenzoyl-CoA reductase activity was lacking in anaerobic benzoate-grown cells and in aerobic cells. This is taken as evidence that 2-aminobenzoyl-CoA reductase is a key enzyme in a novel reductive pathway of anaerobic 2-aminobenzoic acid metabolism.Dedicated to Prof. Charles W. Evans     

2-amino-4-acetylaminobutyric acid, 2,4-diaminobutyric acid and 2-amino-6N-oxalylureidopropionic acid (oxalylalbizziine) in seeds of acacia angustissima

A new amino acid previously detected in 17 species of Acacia has been isolated from seeds of Acacia angustissima and identified as oxalylalbizziine. These seeds also contain more than 6% dry weight of 2-amino-4-acetylaminobutyric acid, which has not been reported previously in a legume, and lower concentrations of 2,4-diaminobutyric acid.     

Rapid degradation of ferulic acid via 4-vinylguaiacol and vanillin by a newly isolated strain of bacillus coagulans

A new strain Bacillus coagulans BK07 was isolated from decomposed wood-bark, based on its ability to grow on ferulic acid as a sole carbon source. This strain rapidly decarboxylated ferulic acid to 4-vinylguaiacol, which was immediately converted to vanillin and then oxidized to vanillic acid. Vanillic acid was further demethylated to protocatechuic acid. Above 95% substrate degradation was obtained within 7 h of growth on ferulic acid medium, which is the shortest period of time reported to date. The major degradation products, was isolated and identified by thin-layer chromatography, high performance liquid chromatography and ¹H-nuclear magnetic resonance spectroscopy were 4-vinylguaiacol, vanillin, vanillic acid and protocatechuic acid.     

Rat brain arachidonic acid metabolism is increased by a 6-day intracerebral ventricular infusion of bacterial lipopolysaccharide

In a rat model of acute neuroinflammation, produced by a 6-day intracerebral ventricular infusion of bacterial lipopolysaccharide (LPS), we measured brain activities and protein levels of three phospholipases A2 (PLA2) and of cyclo-oxygenase-1 and -2, and quantified other aspects of brain phospholipid and fatty acid metabolism. The 6-day intracerebral ventricular infusion increased lectin-reactive microglia in the cerebral ventricles, pia mater, and the glial membrane of the cortex and resulted in morphological changes of glial fibrillary acidic protein (GFAP)-positive astrocytes in the cortical mantel and areas surrounding the cerebral ventricles. LPS infusion increased brain cytosolic and secretory PLA2 activities by 71% and 47%, respectively, as well as the brain concentrations of non-esterified linoleic and arachidonic acids, and of prostaglandins E2 and D2. LPS infusion also increased rates of incorporation and turnover of arachidonic acid in phosphatidylethanolamine, plasmenylethanolamine, phosphatidylcholine, and plasmenylcholine by 1.5- to 2.8-fold, without changing these rates in phosphatidylserine or phosphatidylinositol. These observations suggest that selective alterations in brain arachidonic acid metabolism involving cytosolic and secretory PLA2 contribute to early pathology in neuroinflammation.     

Isolation of new 6-methylnicotinic-acid-degrading bacteria, one of which catalyses the regioselective hydroxylation of nicotinic acid at position C2

2-Hydroxynicotinic acid is an important building block for herbicides and pharmaceuticals. Enrichment strategies to increase the chances of finding microorganisms capable of hydroxylating at the C2 position and to avoid the degradation of nicotinic acid via the usual intermediate, 6-hydroxynicotinic acid, were used. Three bacterial strains (Mena 23/3–3c, Mena 25/4–1, and Mena 25/ 4–3) were isolated from enrichment cultures with 6-methylnicotinic acid as the sole source of carbon and energy. Partial characterization of these strains indicated that they represent new bacterial species. All three strains completely degraded 6-methylnicotinic acid, and evidence is presented that the first step in the degradation pathway of strain Mena 23/3–3c is hydroxylation at the C2 position. Resting cells of this strain grown on 6-methylnicotinic acid also hydroxylated nicotinic acid at the C2 position, but did not further degrade the product. Strain Mena 23/ 3–3c showed the highest degree of 16S rRNA sequence similarity to members of the genera Ralstonia and Burkholderia. Received: 4 April 1997 / Accepted: 10 June 1997     

Isolation and characterization of a meta-cleavage product in the degradation of quinaldic acid by Azotobacter sp.

Abstract From different samples of soil seventeen strains were isolated which grew aerobically in mineral salts medium with quinaldic acid as sole carbon source. Mutants were induced with N -methyl- N '-nitro- N -nitrosoguanidine. One mutant could be isolated which accumulated a yellow compound. The properties of this purified compound were those expected for 2-oxo-3-(4'-hydroxy-2'-oxo-3',4'-en-butyrate)-pyridine-6-carboxylic acid.     

Cloning and characterization of a bacterial iterative type I polyketide synthase gene encoding the 6-methylsalicyclic acid synthase

Unusual polyketide synthases (PKSs), that are structurally type I but act in an iterative manner for aromatic polyketide biosynthesis, are a new family found in bacteria. Here we report the cloning of the iterative type I PKS gene chlB1 from the chlorothricin (CHL) producer Streptomyces antibioticus DSM 40725 by a rapid PCR approach, and characterization of the function of the gene product as a 6-methylsalicyclic acid synthase (6-MSAS). Sequence analysis of various iterative type I PKSs suggests that the resulting aromatic or aliphatic structure of the products might be intrinsically determined by a catalytic feature of the paired KR-DH domains in the control of the double bond geometry. The finding of ChlB1 as a 6-MSAS not only enriches the current knowledge of aromatic polyketide biosynthesis in bacteria, but will also contribute to the generation of novel polyketide analogs via combinatorial biosynthesis with engineered PKSs.     

Interactions studies of CYP2D6 with quercetin and hyperoside by spectral analysis and molecular dynamics simulations

Some ingredients from herbal medicine can significantly affect the activity of CYP2D6, thus leading to serious interactions between herbs and drugs. Quercetin and hyperoside are active ingredients widely found in vegetables, fruits, and herbal medicines. Quercetin and hyperoside have many biological activities. In this work, the characteristic bindings of CYP2D6 with quercetin/hyperoside are revealed by multi-spectroscopy analysis, molecular docking, and molecular dynamics simulations. The fluorescence of CYP2D6 is statically quenched by quercetin and hyperoside. The binding constant (K_a) values of CYP2D6–quercetin/hyperoside range from 10⁴ L mol⁻¹, which indicates that these two flavonoids bind moderately to CYP2D6. Meanwhile, quercetin has a stronger quenching ability to CYP2D6 than that of hyperoside. The secondary structure of CYP2D6 is obviously changed by binding with quercetin/hyperoside. The docking results reveal that the quercetin/hyperoside enters the active site of CYP2D6 near heme and binds to CYP2D6 by hydrogen bonds and van der Waals forces. The molecular dynamics simulation results indicate that the binding of quercetin/hyperoside can stabilize the two complexes, enhance the flexibility of CYP2D6 backbone atoms, and make a more unfolded and looser structure of CYP2D6.     

Effects of sulfate on lactate and C2-, C3- volatile fatty acid anaerobic degradation by a mixed microbial culture

The effects of sulfate on the anaerobic degradation of lactate, propionate, and acetate by a mixed bacterial culture from an anaerobic fermenter fed with wine distillery waste water were investigated. Without sulfate and with both sulfate and molybdate, lactate was rapidly consumed, and propionate and acetate were produced; whereas with sulfate alone, only acetate accumulated. Propionate oxidation was strongly accelerated by the presence of sulfate, but sulfate had no effect on acetate consumption even when methanogenesis was inhibited by chloroform. The methane production was not affected by the presence of sulfate. Counts of lactate- and propionate-oxidizing sulfate-reducing bacteria in the mixed culture gave 4.5×10⁸ and 1.5×10⁶ viable cells per ml, respectively. The number of lactate-oxidizing fermentative bacteria was 2.2×10⁷ viable cells per ml, showing that sulfate-reducing bacteria outcompete fermentative bacteria for lactate in the ecosystem studied. The number of acetoclastic methanogens was 3.5×10⁸ viable cells per ml, but only 2.5×10⁴ sulfate reducers were counted on acetate, showing that acetotrophic methanogens completely predominated over acetate-oxidizing sulfate-reducing bacteria. The contribution of acetate as electron donor for sulfate reduction in the ecosystem studied was found to be minor.     

Stereoselective aliphatic hydroxylation of 6-n-propylchromone-2-carboxylic acid by female Dutch rabbits.

6-n-Alkylchromone-2-carboxylic acids are metabolized solely by aliphatic oxidation. In the rabbit, the 6-n-propyl congener (PCCA) undergoes omega-1 hydroxylation exclusively. Following administration of PCCA to female Dutch rabbits (500 mumol/kg), some 77% of the dose was excreted in the urine, 41% as PCCA and 36% as 6-(2'-hydroxy-n-propyl)chromone-2-carboxylic acid. Since this metabolite is chiral, we have examined the stereochemistry of the excreted material. Diastereoisomeric (as camphanate and alpha-methoxy-alpha-(trifluoro-methyl)phenylacetate esters) and direct chiral HPLC and chiral lanthanide shift NMR have each shown the S:R ratio of the excreted metabolite to be 76:24. When rabbits were dosed with the racemic metabolite, excretion of the compound was not stereoselective. The regio- and stereo-selectivity of the aliphatic hydroxylation of PCCA are thus reflections of the selectivities of the enzyme systems responsible for its formation and suggest PCCA to be an appropriate probe compound for the study of prochiral-chiral hydroxylations.     

Rapid degradation of ferulic acid via 4-vinylguaiacol and vanillin by a newly isolated strain of Bacillus coagulans

A new strain Bacillus coagulans BK07 was isolated from decomposed wood-bark, based on its ability to grow on ferulic acid as a sole carbon source. This strain rapidly decarboxylated ferulic acid to 4-vinylguaiacol, which was immediately converted to vanillin and then oxidized to vanillic acid. Vanillic acid was further demethylated to protocatechuic acid. Above 95% substrate degradation was obtained within 7 h of growth on ferulic acid medium, which is the shortest period of time reported to date. The major degradation products, was isolated and identified by thin-layer chromatography, high performance liquid chromatography and ¹H-nuclear magnetic resonance spectroscopy were 4-vinylguaiacol, vanillin, vanillic acid and protocatechuic acid.     

双歧杆菌脂磷壁酸对结肠癌细胞CD44v6和MMP-2表达的影响

目的研究双歧杆菌脂磷壁酸(LTA)对结肠癌细胞中CD44v6与基质金属蛋白酶2(MMP-2)表达的影响,探讨其在抑制结肠癌转移中的作用。方法结肠癌LoVo细胞及HT-29细胞用含50 mg/L双歧杆菌LTA的培养液培养24 h后,RT-PCR和免疫细胞化学染色检测CD44v6和MMP-2在结肠癌细胞中的表达变化。结果结肠癌LoVo细胞及HT-29细胞中CD44v6和MMP-2的mRNA和蛋白质均呈高表达,经双歧杆菌LTA处理后,其表达均明显下降,与对照组比较,差异有非常显著性(P0.01)。结论双歧杆菌LTA可能通过下调CD44v6和MMP-2的表达来抑制结肠癌的转移。     

Anaerobic metabolism of 2-hydroxybenzoic acid (salicylic acid) by a denitrifying bacterium

The anaerobic metabolism of 2-hydroxybenzoic acid (salicylic acid) was studied in a denitrifying bacterium. Cells grown with 2-hydroxybenzoate were simultaneously adapted to degrade benzoate. Extract of these cells formed benzoate or benzoyl-CoA when incubated under reducing conditions with salicylate, MgATP, and coenzyme A, suggesting a degradation of 2-hydroxybenzoate via benzoate or benzoyl-CoA. This suggestion was supported by enzyme activity measurements. In extracts of 2-hydroxybenzoate-grown cells, the following enzyme activities were detected: two CoA ligases, one specific for 2-hydroxybenzoate, the other for benzoate, and two different enzyme activities catalyzing the reductive transformation of 2-hydroxybenzoyl-CoA. These findings suggest a degradation of salicylic acid by two new enzymes, 2-hydroxybenzoate-CoA ligase (AMP-forming) and 2-hydroxybenzoyl-CoA reductase (dehydroxylating), catalyzing (1) 2-hydroxybenzoate + MgATP + CoASH → 2-hydroxybenzoyl-CoA + MgAMP + PP_i (2) 2-hydroxybenzoyl-CoA + 2[H] → benzoyl-CoA + H₂O Benzoyl-CoA was dearomatized by reduction of the ring. This represents another case in which benzoyl-CoA is a central intermediate in anaerobic aromatic metabolism. Received: 1 February 1996 / Accepted: 24 February 1996