Oxidation of catechol in plants : III. Purification and properties of the diphenylene dioxide 2,3-quinone-forming enzyme system from Tecoma leaves

In attempting to determine the nature of the enzyme system mediating the conversion of catechol to diphenylenedioxide 2,3-quinone, in Tecoma leaves, further purification of the enzyme was undertaken. The crude enzyme from Tecoma leaves was processed further by protamine sulfate precipitation, positive adsorption on tricalcium phosphate gel, and elution and chromatography on DEAE-Sephadex. This procedure yielded a 120-fold purified enzyme which stoichiometrically converted catechol to diphenylenedioxide 2,3-quinone. The purity of the enzyme system was assessed by polyacrylamide gel electrophoresis. The approximate molecular weight of the enzyme was assessed as 200,000 by gel filtration on Sephadex G-150. The enzyme functioned optimally at pH 7.1 and at 35 °C. The Km for catechol was determined as 4 × 10−4 Image . The enzyme did not oxidize o-dihydric phenols other than catechol and it did not exhibit any activity toward monohydric and trihydric phenols and flavonoids. Copper-chelating agents did not inhibit the enzyme activity. Copper could not be detected in the purified enzyme preparations. The purified enzyme was not affected by extensive dialysis against copper-complexing agents. It did not show any peroxidase activity and it was not inhibited by catalase. Hydrogen peroxide formation could not be detected during the catalytic reaction. The enzymatic conversion of catechol to diphenylenedioxide 2,3-quinone by the purified Tecoma leaf enzyme was suppressed by such reducing agents as GSH and cysteamine. The purified enzyme was not sensitive to carbon monoxide. It was not inhibited by thiol inhibitors. The Tecoma leaf was found to be localized in the soluble fraction of the cell. Treatment of the purified enzyme with acid, alkali, and urea led to the progressive denaturation of the enzyme.

The Influence of the Val158Met Catechol-O-Methyltransferase Polymorphism on the Personality Traits of Bipolar Patients

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Catechol 2,3-dioxygenase-assisted cleavage of aromatics by “anaerobic” termite gut spirochetes and genomic evidence of a complete meta-pathway

The termite hindgut microbial ecosystem functions like a miniature lignocellulose-metabolizing natural bioreactor, has significant implications to nutrient cycling in the terrestrial environment, and represents an array of microbial metabolic diversity. Deciphering the intricacies of this microbial community to obtain as complete a picture as possible of how it functions as a whole, requires a combination of various traditional and cutting-edge bioinformatic, molecular, physiological, and culturing approaches. Isolates from this ecosystem, including Treponema primitia str. ZAS-1 and ZAS-2 as well as T. azotonutricium str. ZAS-9, have been significant resources for better understanding the termite system. While not all functions predicted by the genomes of these three isolates are demonstrated in vitro, these isolates do have the capacity for several metabolisms unique to spirochetes and critical to the termite system’s reliance upon lignocellulose. In this thesis, work culturing, enriching for, and isolating diverse microorganisms from the termite hindgut is discussed. Additionally, strategies of members of the termite hindgut microbial community to defend against O₂-stress and to generate acetate, the “biofuel” of the termite system, are proposed. In particular, catechol 2,3-dioxygenase and other meta-cleavage catabolic pathway genes are described in the “anaerobic” termite hindgut spirochetes T. primitia str. ZAS-1 and ZAS-2, and the first evidence for aromatic ring cleavage in the phylum (division) Spirochetes is also presented. These results suggest that the potential for O₂-dependent, yet nonrespiratory, metabolisms of plant-derived aromatics should be re-evaluated in termite hindgut communities. Potential future work is also illustrated.

Electrocatalytic oxidation of catechol at multi-walled carbon nanotubes modified electrode

In 0.05 mol/L phosphate buffer solution (pH 7.0), carbon nanotubes modified electrode exhibits rapid response, strong catalytic activity with high stability toward the electrochemical oxidation of catechol. The electrochemical behavior of catechol on both the multi-walled and single-walled carbon nanotubes modified electrode was investigated. The experimental conditions, such as pH of the solution and scan rate were optimized. The currents (measured by constant potential amperometry) increase linearly with the concentrations of catechol in the range of 2.0 x 10(-5) - 1.2 x 10(-3) mol/L. Moreover, at the multi-walled carbon nanotubes modified electrode the electrochemical responses of catechol and ascorbic acid can be separated clearly.

DETERMINATION OF CATECHOL DERIVATIVES ON PRETREATMENT AND COPOLYMER COATED GLASSY-CARBON ELECTRODE

A glassy carbon electrode was pretreated electrochemically and was coated with a copolymer of maleic acid anhydride attached with Eastman-AQ55D (MA/AQ). The voltammetric behavior of a series of biologically important compounds, such as dopamine, L-DOPA, D

LIQUID-CHROMATOGRAPHY AMPEROMETRIC DETECTION OF CATECHOL, RESORCINOL, AND HYDROQUINONE WITH A COPPER-BASED CHEMICALLY MODIFIED ELECTRODE

A copper-based chemically modified electrode (CME) has been constructed and characterized for flow-through amperometric detection of catechol, resorcinol, and hydroquinone. Novel potential dependence of the detector response was first obtained for these analytes at the Cu CME, where negative peaks together with positive ones were observed in one definite chromatogram using amperometric detection. Its advantages in chromatographic applications were demonstrated. From these observations it is proposed that the detector response was governed by formation of copper complexes with the solutes. A dynamic linear range over two orders of magnitude was obtained, when operating the detector at +0.10 V vs. SCE, from which ng detection limits were achieved.

The electrochemical oxidation of catechol and dopamine on platinum in 1-Ethyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide ([C(2)mim][NTf2]) and 1-Butyl-3-methylimidazolium tetrafluoroborate ([C(4)mim][BF4]): Adsorption effects in ionic liquid voltammetry

The electrochemical oxidation of catechol and dopamine has been studied at a platinum micro-electrode (10 pm diameter) in two room temperature ionic liquids (RTILs): 1-Ethyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide ([C(2)mim][NTf2]) and 1-Butyl-3-methylimidazolium tetrafluoroborate ([C(4)mim][BE4]). For catechol in [C(2)mim][NTf2], an electrochemically quasi-reversible oxidation peak was observed at 1.1 V vs. Pt with a back peak at 0.4 V vs. Pt. This is assigned to the two-electron oxidation of catechol to doubly protonated o-benzoquinone. Double-step chronoamperometry gave a diffusion coefficient for the catechol and the oxidised species which is 3.8 x 10(-11) m(2) s(-1) for both. For catechol in [C(4)mim][BF4], a two-electron oxidation wave was observed at 1.0 V vs. Pt with no back peak. Another peak at less positive potential was also observed at 0.6 V vs. Pt in [C(4)mim][BF4] but not in [C(2)mim][NTf2] which is assigned to the adsorption of electrochemically formed neutral o-benzoquinone on the platinum electrode. The oxidised protonated o-benzoquinone is suggested to be deprotonated by the [BF4](-) anion, but not by the [NTf2](-) anion: hence adsorption of the neutral species at the platinum electrode, not the charged species. For dopamine in both RTILs, two chemically irreversible oxidation peaks were observed at 0.75 V and 1.1 V vs. Pt, and assigned to the oxidation of dopamine to the corresponding semi-quinone and the quinone. Potential-step chronoamperometry was carried out on the oxidation waves of dopamine in [C(2)mim][NTf2] and the diffusion coefficient of species in solution was calculated to be 6.85 x 10(-12) m(2) s(-1) and confirmed that the waves corresponded to one and two electron processes. A third wave was observed at 1.8 V vs. Pt which is attributed to the oxidation of the amine group to a radical cation with likely subsequent follow up chemistry. In [C(4)mim][BF4] a peak at less positive potential was observed for dopamine, similar to catechol which is assigned to the adsorption of the neutral quinone species on the platinum electrode formed by the reaction of the removal of protons from the oxidised dopamine with the [BF4](-) anion. (C) 2009 Elsevier B.V. All rights reserved.

Variants in the catechol-o-methyltransferase (COMT) gene are associated with schizophrenia in Irish high-density families

The enzyme catechol-o-methyltransferase (COMT) transfers a methyl group from adenosylmethionine to catecholamines including the neurotransmitters dopamine, epinephrine and norepinephrine. This methylation results in the degradation of catecholamines. The involvement of the COMT gene in the metabolic pathway of these neurotransmitters has made it an attractive candidate gene for many psychiatric disorders. In this article, we reported our study of association of COMT with schizophrenia in Irish families with a high density of schizophrenia. Three single nucleotide polymorphisms (SNPs) were genotyped for the 274 such families and within-family transmission disequilibrium tests were performed. SNP rs4680, which is the functional Val/Met polymorphism, showed modest association with the disease by the TRANSMIT, FBAT and PDT programs, while the other two SNPs were negative. These SNPs showed lower level of LDs with each other in the Irish subjects than in Ashkenazi Jews. Haplotype analysis indicated that a haplotype, haplotype A-G-A for SNPs rs737865-rs4680-rs165599, was preferentially transmitted to the affected subjects. This was different from the reported G-G-G haplotype found in Ashkenazi Jews, but both haplotypes shared the Val allele. We concluded that COMT gene is associated with schizophrenia and carries a small but significant risk to the susceptibility in the Irish subjects.

Catechol-O-methyltransferase and the clinical features of psychosis

A functional polymorphism (Val-158-Met) at the Catechol-O-methyltransferase (COMT) locus has been identified as a potential etiological factor in schizophrenia. Yet the association has not been convincingly replicated across independent samples. We hypothesized that phenotypic heterogeneity might be diluting the COMT effect. To clarify the putative association, we performed an exploratory analysis to test for association between COMT and five psychosis symptom scales. These were derived through factor analysis of the Operational Criteria Checklist for Psychiatric Illness. Our sample was the Irish Study of High Density Schizophrenia Families, a large collection consisting of 268 multiplex families. This sample has previously shown a small but significant effect of the COMT Val allele in conferring risk for schizophrenia. We tested for preferential transmission of COMT alleles from parent to affected offspring (n = 749) for each of the five factor-derived scales (negative symptoms, delusions, hallucinations, mania, and depression). Significant overtransmission of the Val allele was found for mania (P <0.05) and depression (P = 0.01) scales. Examination of odds ratios (ORs) revealed a heterogeneous effect of COMT, whereby it had no effect on Negative Symptoms, but largest impact on Depression (OR = 1.4). These results suggest a modest affective vulnerability conferred by this allele in psychosis, but will require replication.

Electrochemical sensor for catechol and dopamine based on a catalytic molecularly imprinted polymer-conducting polymer hybrid recognition element

One of the difficulties with using molecularly imprinted polymers (MIPs) and other electrically insulating materials as the recognition element in electrochemical sensors is the lack of a direct path for the conduction of electrons from the active sites to the electrode. We have sought to address this problem through the preparation and characterization of novel hybrid materials combining a catalytic MIP, capable of oxidizing the template, catechol, with an electrically conducting polymer. In this way a network of "molecular wires" assists in the conduction of electrons from the active sites within the MIP to the electrode surface. This was made possible by the design of a new monomer that combines orthogonal polymerizable functionality; comprising an aniline group and a methacrylamide. Conducting films were prepared on the surface of electrodes (Au on glass) by electropolymerization of the aniline moiety. A layer of MIP was photochemically grafted over the polyaniline, via N,N'-diethyldithiocarbamic acid benzyl ester (iniferter) activation of the methacrylamide groups. Detection of catechol by the hybrid-MIP sensor was found to be specific, and catechol oxidation was detected by cyclic voltammetry at the optimized operating conditions: potential range -0.6 V to +0.8 V (vs Ag/AgCl), scan rate 50 mV/s, PBS pH 7.4. The calibration curve for catechol was found to be linear to 144 µM, with a limit of detection of 228 nM. Catechol and dopamine were detected by the sensor, whereas analogues and potentially interfering compounds, including phenol, resorcinol, hydroquinone, serotonin, and ascorbic acid, had minimal effect (=3%) on the detection of either analyte. Nonimprinted hybrid electrodes and bare gold electrodes failed to give any response to catechol at concentrations below 0.5 mM. Finally, the catalytic properties of the sensor were characterized by chronoamperometry and were found to be consistent with Michaelis-Menten kinetics. © 2009 American Chemical Society.

Assessment of toxicological interactions of benzene and its primary degradation products (catechol and phenol) using a lux-modified bacterial bioassay

A bacterial bioassay has been developed to assess the relative toxicities of xenobiotics commonly found in contaminated soils, rivers, waters, and ground waters. The assay utilized decline in luminescence of lux- marked Pseudomonas fluorescens on exposure to xenobiotics. Pseudomonas fluorescens is a common bacterium in the terrestrial environment, providing environmental relevance to soil, river, and ground water systems. Three principal environmental contaminants associated with benzene degradation were exposed to the luminescence-marked bacterial biosensor to assess their toxicity individually and in combination. Median effective concentration (EC50) values for decline in luminescence were determined for benzene, catechol, and phenol and were found to be 39.9, 0.77, and 458.6 mg/L, respectively. Catechol, a fungal and bacterial metabolite of benzene, was found to be significantly more toxic to the biosensor than was the parent compound benzene, showing that products of xenobiotic biodegradation may be more toxic than the parent compounds. Combinations of parent compounds and metabolites were found to be significantly more toxic to the bioassay than were the individual compounds themselves. Development of this bioassay has provided a rapid screening system suitable for assessing the toxicity of xenobiotics commonly found in contaminated soil, river, and ground-water environments. The assay can be utilized over a wide pH range and is therefore more applicable to such environmental systems than bioluminescence-based bioassays that utilize marine organisms and can only be applied over a limited pH and salinity range.

Characterization and Catalytic Activity of Polymer Supported Ruthenium Schiff Base Complexes Towards Catechol Oxidation

Ruthenium(III) complexes of the Schiff bases formed by the condensation of polymer bound aldehyde and the amines, such as 1,2-phenylenediamine (PS-opd), 2-aminophenol (PS-ap), and 2-aminobenzimidazole (PS-ab) have been prepared. The magnetic moment, EPR and electronic spectra suggest an octahedral structure for the complexes. The complexes of PS-opd, PS-ap, and PS-ab have been assigned the formula [PS-opdRuCl3(H2O)], [PS-apRuCl2(H2O)2], [PS-ab- RuCl3(H2O)2], respectively. These complexes catalyze oxidation of catechol using H2O2 selectively to o-benzoquinone. The catalytic activity of the complexes is in the order [PS-ab- RuCl3(H2O)2] . [PS-opdRuCl3(H2O)] [PS-apRuCl2(H2O)2]. Mechanism of the catalytic oxidation of catechol by ruthenium( III) complex is suggested to take place through the formation of a ruthenium(II) complex and its subsequent oxidation by H2O2 to the ruthenium(III) complex.