Catechol Oxidase Enzyme Research Articles

Structural characterization of benzoate anion-supported supramolecular assemblages of Co(III) and Ni(II) cyclam compounds with catecholase-like and antibacterial activities

Two mononuclear compounds, cis-[Co(cyclam)(Bz)2](Bz)·H2O (1), and trans-[Ni(cyclam)(H2O)2](MOBA)2·4H2O (2) (where Bz = benzoate and MOBA = o-methoxy benzoate) have been synthesized and characterized by elemental analyses, spectroscopy, X-ray diffraction, and thermogravimetric techniques. Compounds 1 and 2 crystallize in a monoclinic crystal system with P21/c and P21/n space groups, respectively. The carboxylate ligand in compound 1 enables the cyclam ligand to adopt a cis-V conformation, while in compound 2, cyclam adopts a stable trans-III conformation. Hirshfeld surface analyses were carried out to elucidate the intricate intermolecular forces shaping the crystal structure. The ability of compounds 1 and 2 to imitate the activities of the catechol oxidase enzyme was evaluated using 3,5-di-tert-butyl catechol (3,5-DTBC) as the model substrate. The antibacterial activity was investigated using the Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli bacterial strains. Interestingly, compound 1 exhibited higher catecholase-like activity than compound 2 and enhanced antibacterial activity.

Effect of structural variation on enzymatic activity in tetranuclear (Cu4) clusters with defective cubane core

The stimulus to the modeling of enzyme functioning sites comes from their potential to give insight into the natural enzyme’s mechanistic pathways, ascertain the role of that different metal ion in the active site and construct better catalysts motivated by nature. The presence of metal ion leads to the activation of molecular oxygen in the metalloenzymes. The metalloenzymes such as the catechol oxidase (CO) enzyme that oxidizes the catechol to corresponding quinones which eventually protect damage tissues from plant and pathogen. Thus, the design and characterization of catalysts used as selectively and efficiently oxidation reactions have grown to be unique challenges for modern inorganic chemists. In this work, two novel tetranuclear complexes (1 and 2) have been synthesized in excellent yield. The complexes were characterized using various spectroscopic techniques such as FTIR, UV–Visible and PXRD pattern. The structure of 1 and 2 was elucidated by SC-XRD (single crystal X-ray diffraction) analysis. The magnetic study reveals the presence of the antiferromagnetic nature of 1 and 2. Both 1 and 2 shows a very good catecholase-like activity by oxidizing the catechol to analogous quinone in methanolic solution. Thus, a structure-activity relationship can further help us design other substituted tetranuclear complexes with enhanced catecholase like activity. Communicated by Ramaswamy H. Sarma

Structure, DFT studies and evaluation of catechol oxidase (CO) mimic activity of mononuclear Co(II) complexes derived from aminoalcohols: an experimental and theoretical approach

The impetus to modeling of enzyme mimics comes from their potential to provide insight to the alternate mechanistic pathways of the native enzymes. The present study demonstrates the syntheses and characterization of two different cobalt(II) complexes, [Co(pdm)(Phen)Cl]Cl·H2O (1) and [Co(pmmH)2(SCN)2] (2) with the aminoalcohol ligands such as pyridine-2,6-dimethanol (pdmH2) or 2-pyridinemonomethanol (pmmH) and their assessment as catechol oxidase (CO) enzyme mimic. Single Crystal X-ray diffraction and powder X-ray diffraction data suggest the octahedral environment around the Co(II) ion and the complexes form extensive 1D or 2D propagating network as a result of non-covalent interactions (O···H and C-H···π). TD-DFT calculations were used to explain the spectral bands obtained during the UV-Vis absorption studies and it is ascertained that the transitions were mainly of the intra-ligand charge transfer (ILCT) type. The catecholase biomimetic catalytic activity of the synthesized complexes has been investigated in detail and the kinetics is also performed. The results obtained show that both the complexes catalyze the aerobic oxidation of catechol to the corresponding o-quinone. The K cat value for 1 is 106.99 h‒1 and for 2 is 90.32 h‒1 in methanol. It may be mentioned here that 1 and 2 are effective catalysts, with the order of activity being 1 > 2. The order of enzymatic activity is well justified by CV and DFT studies. Communicated by Ramaswamy H. Sarma

Design and catalytic studies of structural and functional models of the catechol oxidase enzyme.

The catechol oxidase activity of three copper/bicompartmental salen derivatives has been studied. One mononuclear, [CuL] (1), one homometallic, [Cu2L(NO3)2] (2), and one heterometallic, [CuMnL(NO3)2] (3) complexes were obtained using the ligand H2L = N,N'-bis(3-methoxysalicylidene)-1,3-propanediamine through different synthetic methods (electrochemical, chemical and solid state reaction). The structural data indicate that the metal ion disposition models the active site of type-3 copper enzymes, such as catechol oxidase. In this way, their ability to act as functional models of the enzyme has been spectrophotometrically determined by monitorization of the oxidation of 3,5-di-tert-butylcatechol (3,5-DTBC) to 3,5-di-tert-butyl-o-benzoquinone (3,5-DTBQ). All the complexes show significant catalytic activity with ratio constants (kobs) lying in the range (223-294) × 10-4min-1. A thorough kinetic study was carried out for complexes 2 and 3, since they show structural similarities with the catechol oxidase enzyme. The greatest catalytic activity was found for the homonuclear dicopper compound (2) with a turnover value (kcat) of (3.89 ± 0.05) × 106h-1, which it is the higher reported to date, comparable to the enzyme itself (8.25 × 106h-1).

A novel Cu(II) distorted cubane complex containing Cu4O4 core as the first tetranuclear catalyst for temperature dependent oxidation of 3,5-di-tert-butyl catechol and in interaction with DNA & protein (BSA).

The tri-dentate Schiff base ligand 3-(2-hydroxyethylimino)-1-phenylbut-1-en-1-ol (L) produced the tetra-nuclear Cu(II) distorted cubane complex which contain Cu4O4 core, upon reaction with Cu(II)acetate.H2O. The complex was structurally characterized by X-ray crystallography and found that, in this tetrameric and tetra-nuclear distorted cubane structure, each two-fold deprotonated Schiff base ligand coordinated to a Cu(II) center with their alcoholic oxygens and imine nitrogens and formed six and five-membered chelate rings. At the same time, each ligand bridged to a neighboring Cu(II) atom by its alcoholic oxygen, thus the metal centers became penta-coordinated. The copper(II) complex with μ-ɳ2-hydroxo bridges and Cu….Cu distance about 3 Å was structurally similar to the active site of natural catechol oxidase enzyme and exhibited excellent catecholase activity in aerobic oxidation of 3,5-di-tert-butyl catechol to its o-quinone. The kinetics and mechanism of the oxidation of 3, 5-DTBCH2 catalyzed by [CuL]4 complex, were studied at four different temperatures from 283 to 313K by UV-Vis spectroscopy. Interaction of [CuL]4 complex with FS-DNA was investigated by UV-Vis and fluorescence spectroscopy, viscosity measurements, cyclic voltammetry (CV), circular dichroism (CD) and agarose gel electrophoresis. The main mode of binding of the complexes with DNA was intercalation. The interaction between [CuL]4 complex and bovine serum albumin (BSA) was studied by UV-Vis, fluorescence and synchronous fluorescence spectroscopic techniques. The results indicated a high binding affinity of the complex to BSA. Invitro anticancer activity of the complex was evaluated against A549, Jurkat and Ragi cell lines by MTT assay. The complex was remarkably active against the cell lines and can be a good candidate for an anticancer drug. Theoretical docking studies were performed to further investigate the DNA and BSA binding interactions.

Inhibition of flash rusting of HY80 by a mussel adhesive protein: Characterizing the interaction of MeFP-5 with a high strength low alloy steel

A proteinaceous biopolymer, mussel adhesive protein #5 (MeFP-5), isolated from the common blue mussel (Mytilus edulis L) has been investigated as a model for designing an aqueously soluble candidate corrosion inhibitor system that is non-toxic and environmentally friendly and is capable of inhibiting the flash rusting of exposed high strength low alloy (HSLA) steel surfaces during the paint removal process. In a previous study it was found that a significant amount of corrosion inhibition is possible (nearly 100% inhibition after 7 days) on HY80 steel in a 100% relative humidity environment at 40 °C. In an attempt to determine the possible mechanisms of corrosion inhibition, surface characterization of HY80 steel treated with various solutions containing the MeFP-5 biopolymer were performed. Fourier transform infrared (FT-IR) spectroscopy using attenuated total reflectance (ATR) of the protein adsorbed onto glass and metal surfaces indicated that complexation of the iron atoms on the oxyhydroxide covered metal surface involved the amino acid l-dopa and the primary amine of lysine. The involvement of these amino acids in the adsorption of the protein onto glass were much less evident. When the adsorbed protein was treated with a catechol oxidase enzyme, vibration bands consistent with metal complex formation involving l-dopa were not observed, indicating that the treatment with enzyme resulted in mess metal chelation at the surface oxide-protein film interface. Energy dispersive x-ray spectroscopy (EDS) findings indicate that iron content is highest where the MeFP-5 biopolymer is adsorbed onto the steel substrate at pH 5.5, 50% higher than the content of the iron on the steel surface alone. When the biopolymer is enzymatically cross-linked on the steel surface at pH 5.5, the iron content is decreased by 15% of that of the adsorbed biopolymer, suggesting that the iron at the steel surface is undergoing complexation and possible metal-mediated cross-linking, whereas the enzyme cross-linked protein on the steel surface complexes less of the iron. Three-dimensional modeling efforts of MeFP-5 suggest that specific domains of MeFP-5 where two amino acids, l-Dopa and lysine, are concentrated may be involved in adsorption to a metal oxyhydroxide film. These results indicate that it is possible to utilize the biochemistry of a naturally occurring biopolymer isolated from the marine mussel, Mytilus edulis (L), to develop a non-toxic and environmentally friendly corrosion inhibitor.

Catechol oxidase mimetic activity of cadmium sulfide nanoparticles for the oxidation of L- 3,4 -dihydroxyphenylalanine

In the present work we have investigated the oxidation of L-3,4 -dihydroxyphenylalanine (L-Dopa) using colloidal cadmium sulfide nanoparticles (CdS NPs) as a photocatalyst. The CdS NPs were synthesized and characterized using UV-visible absorption spectroscopy, transmission electron microscopy (TEM) and powder x-ray diffraction (XRD). It was observed that the CdS NPs oxidized L-Dopa within 30 min of irradiation using a 200 W Hg(Xe) arc lamp. The oxidation product was identified using high performance liquid chromatography (HPLC). The HPLC analyses were carried out on a reverse-phase C18 column under isocratic conditions using methanol/water (10/90) as the mobile phase. The retention time of the product matched with that of dopachrome. The mechanistic studies indicated the participation of hydroxyl radicals (·OH) in the photocatalytic oxidation of L-Dopa. The kinetic analysis was carried out using the Michaelis-Menten equation. The results demonstrated that the as-prepared CdS NPs mimicked the activity of the catechol oxidase enzyme. To the best of our knowledge, this is the first report on the oxidation of L-Dopa using semiconductor NPs.

Square planar versus square pyramidal copper(II) complexes containing N3O moiety: Synthesis, structural characterization, kinetic and catalytic mimicking activity

In quest of copper complexes having [CuN3O] cores, [Cu(phen)(l–Val)(H2O)]NO3 (1) and [Cu(bpy)(l–Val)]ClO4 (2) complexes have been synthesized and structurally characterized (phen = 1,10–phenanthroline; bpy = 2,2′–bipyridine). Complex 1 possesses a distorted square-pyramidal, whereas 2 has a distorted square-planar coordination geometry. Structures of 1 and 2 have supramolecular networks formed via inter- and intra-molecular hydrogen bonding interactions. The kinetics and mechanism of ligand substitution of 1 and 2 by thiourea (TU) were studied in detail and showed a biphasic process in which an initial fast reaction is followed by a slower one. The activation parameters for the fast reaction: ΔH# = 68 ± 4 and 73 ± 5 kJ mol−1, ΔS# = 43 ± 10 and 54 ± 9 J K−1 mol−1 for 1 and 2, respectively, supports a dissociative substitution mechanism. Whereas for the slow reaction: ΔH# = 33 ± 6 and 43 ± 3 kJ mol−1, ΔS# = −77 ± 10 and −56 ± 9 J K−1 mol−1 for 1 and 2, respectively, support an associative substitution mechanism. It is concluded from the activation parameters that the difference in structure does not affect the mechanism. Complexes 1 and 2 have also been evaluated as functional models for the catechol oxidase enzyme and phenoxazinone synthase. The model complexes 1 and 2 show catecholase activity of Kcat = 10.9 × 103 and 11.4 × 103 h−1 and phenoxazinone synthase activity of Kcat = 2.1 × 103 and 4.3 × 103 h−1, respectively. Compared to the enzyme itself (Kcat = 8.3 h−1), the model complexes 1 and 2 are promising candidates as functional mimics for catechol oxidase and phenoxazinone synthase.

Copper(II) and Sodium(I) Complexes based on 3,7-Diacetyl-1,3,7-triaza-5-phosphabicyclo[3.3.1]nonane-5-oxide: Synthesis, Characterization, and Catalytic Activity.

The reaction of 3,7-diacetyl-1,3,7-triaza-5-phosphabicyclo[3.3.1]nonane (DAPTA) with metal salts of CuII or NaI /NiII under mild conditions led to the oxidized phosphane derivative 3,7-diacetyl-1,3,7-triaza-5-phosphabicyclo[3.3.1]nonane-5-oxide (DAPTA=O) and to the first examples of metal complexes based on the DAPTA=O ligand, that is, [CuII (μ-CH3 COO)2 (κO-DAPTA=O)]2 (1) and [Na(1κOO';2κO-DAPTA=O)(MeOH)]2 (BPh4 )2 (2). The catalytic activity of 1 was tested in the Henry reaction and for the aerobic 2,2,6,6-tetramethylpiperidin-1-oxyl (TEMPO)-mediated oxidation of benzyl alcohol. Compound 1 was also evaluated as a model system for the catechol oxidase enzyme by using 3,5-di-tert-butylcatechol as the substrate. The kinetic data fitted the Michaelis-Menten equation and enabled the obtainment of a rate constant for the catalytic reaction; this rate constant is among the highest obtained for this substrate with the use of dinuclear CuII complexes. DFT calculations discarded a bridging mode binding type of the substrate and suggested a mixed-valence CuII /CuI complex intermediate, in which the spin electron density is mostly concentrated at one of the Cu atoms and at the organic ligand.

Unprecedented hetero-geometric discrete copper(II) complexes: Crystal structure and bio-mimicking of Catecholase activity

An unprecedented solid of coper(II) complexes, [Cu(dpa)2NCS]2[Cu(dpa)2(NCS)2](ClO4)2 (1) [dpa = 2,2′-dipyridylamine; SCN = thiocyanate], has been synthesized and crystallographically characterized with the aim to study the catecholase activity. The Cu(II) complex mimics the full catalytic cycle of the active site of catechol oxidase enzyme in acetonitrile medium with a turnover number of 4.788×103 h −1 along with the production of semiquinone radical and hydrogen peroxide. In situ generation of Cu(I) species in the catalytic pathway of catechol oxidation was established by electrochemical study and further confirmed by electron paramagnetic resonance (EPR) spectroscopy. This paper reports an X-ray crystallographically characterized Cu(II) complexes which mimics the full catalytic cycle of functional model of catechol oxidase enzyme with significant turnover number (4.788×103 h-1) along with the production of semiquinone radical and hydrogen peroxide.

Design of a mononuclear copper(II)-phenanthroline complex: Catechol oxidation, DNA cleavage and antitumor properties

A mononuclear copper(II) complex [Cu(phen)(OH2)2(NO3)](NO3) (1) [phen=1,10-phenanthroline] has been synthesized and structurally characterized by different spectroscopic characterization methods including single crystal X-ray diffraction study. X-ray crystal structure analysis shows that 1 adopts square pyramidal geometry with CuN2O3 chromophore and the molecule crystallises in P21/n space group. 1 has been evaluated as model system for the catechol oxidase enzyme by using 3,5-di-tert-butylcatechol (DTBC) as the substrate in methanol medium, revealing that 1 exhibits greater catalytic activity with Kcat value 3.91×103h−1 compared to a few copper(II) complexes of the same class. Electrochemical analysis suggests that the mononuclar Cu(II) complex mimics the catechol oxidase enzyme in methanolic medium through radical pathway. The complex cleaves the double strand of pBR 322 plasmid DNA in a concentration-dependent manner so as for the DNA to change from super coiled form to both nicked circular and linear forms. The complex is cytotoxic to the human hepatocarcinoma cell HepG2, as revealed in MTT assay. AO/EB and Hoechst 33528 fluorescent staining techniques were used to find the mode of cell death. Interestingly, AO/EB and Hoechst stained cells observed at IC50 concentration shows that 1 brings about conformational change on DNA to induce apoptosis which would be the basis underlying its cytotoxic property.

Synthesis and bio-catalytic activity of isostructural cobalt(III)-phenanthroline complexes

We have synthesized two isostructural mononuclear cobalt(III) complexes [1]NO3⋅3H2O and [1]NO3⋅CH3CO2H⋅H2O {[1] + = [Co(1,10-phenanthroline)2Cl2]+} and characterized by single crystal X-ray structural analyses. Mass spectral studies of the complexes indicate both the compounds to produce identical cationic species viz., [Co(phen)2Cl2]+ in methanol solution. [1]+ has been evaluated as model system for the catechol oxidase enzyme by using 3,5-di-tert-butylcatechol (3,5-DTBC) as the substrate in methanol medium, which revealed that the cationic complex efficiently inhibits catalytic activity with kcat value 9.65 ×102 h−1. [1]+ cleaved pBR 322 DNA without addition of an activating agent. Further, the anti-cancer activity of [1]+ on human hepatocarcinoma cell line (HepG2) has been examined. The induction of apoptosis induced in the cell line was assessed base on the changes in cell morphology, which showed the efficacy of [1] + to induce apoptosis in 53% of cells during 24 h treatment. Interestingly, the observed IC50 values reveal that [1]+ brings about conformational change on DNA strongly and exhibits remarkable cytotoxicity. The cationic cobalt(III) complex exhibits good catalytic activity in saturated oxygen environment towards 3,5-ditertbutylcatechol with kcat value 9.65×102 h-1. It also exhibits important cytotoxicity against human hepatocarcinoma cell line (HepG2).

A mononuclear cobalt(iii) complex and its catecholase activity

A mononuclear Co(iii) complex mimics the catechol oxidase enzyme with appreciably high turnover numbers in different solvents.

Zero‐Order Catechol Oxidase Activity by a Mononuclear Manganese(III) Complex Showing High Turnover Comparable to Catechol Oxidase Enzyme

Enzymatic mimics are designed by using small molecules to impart insight into the mechanistic pathways of the enzymes and to generate catalysts that could be as efficient as the enzyme itself. Among the copper-based polyphenol oxidase enzymes, the copper(II)-containing enzyme catechol oxidase catalyses exclusively the oxidation of ortho-diphenols to orthodiquinones (Scheme 1) while reducing oxygen to water. Such

New mononuclear copper(I) and copper(II) complexes containing N4 donors; crystal structure and catechol oxidase biomimetic catalytic activity

The synthesis of new tetradentate Schiff-base containing N4 donors is described along with the preparation of a series of copper complexes derived from this ligand. These complexes have the form [CuLX]X′, X = OH−, X′ = (1) or X = X′ = Cl− (2); and [CuL]H2O(ClO4)2 (3), in addition to a copper(I) complex [Cu2L2](ClO4)2 (4). The synthesized compounds were spectroscopically characterized, showing the N4 donor ligand. The single-crystal X-ray structural analysis of 4 demonstrated the dimeric structure and univalent copper. This dimer consists of , two uncoordinated , and one acetonitrile. Each ligand is tetradentate via pyridyl and azomethine nitrogen atoms, providing a strongly distorted tetrahedron around copper(I) despite the pseudo-macrocyclic skeleton of the ligand. These complexes have been evaluated as functional model systems for catechol oxidase enzyme using 3,5-di-tert-butylcatechol (3,5-DTBC) as the test substrate. The catalytic performance of the air oxidation of 3,5-DTBC to the corresponding light absorbing 3,5-di-tert-butylquinone (3,5-DTBQ) at ambient conditions was studied using UV-Vis absorption spectra. Complex 4 exhibits the highest catalytic activity with turnover number of 33 h−1. A kinetic treatment on the basis of the Michaelis–Menten model was applied. Correlation among reactivity, binding constants, electrochemical properties, and the geometry was determined. These correlations showed that the rate of oxidation is linearly correlated with the binding constants for the five coordinate 1 and 2. The catalytic investigations demonstrate that geometrical effects are only one facet of the activity. The probable mechanistic implications of the catalyzed oxidation reactions are discussed.

Catechol Oxidase Activity of a Series of New Dinuclear Copper(II) Complexes with 3,5-DTBC and TCC as Substrates: Syntheses, X-ray Crystal Structures, Spectroscopic Characterization of the Adducts and Kinetic Studies

A series of dinuclear copper(II) complexes has been synthesized with the aim to investigate their applicability as potential structure and function models for the active site of catechol oxidase enzyme. They have been characterized by routine physicochemical techniques as well as by X-ray single-crystal structure analysis: [Cu 2(H 2L2 (2))(OH)(H 2O)(NO 3)](NO 3) 3.2H 2O ( 1), [Cu(HL1 (4))(H 2O)(NO 3)] 2(NO 3) 2.2H 2O ( 2), [Cu(L1 (1))(H 2O)(NO 3)] 2 ( 3), [Cu 2(L2 (3))(OH)(H 2O) 2](NO 3) 2, ( 4) and [Cu 2(L2 (1))(N 3) 3] ( 5) [L1 = 2-formyl-4-methyl-6R-iminomethyl-phenolato and L2 = 2,6-bis(R-iminomethyl)-4-methyl-phenolato; for L1 (1) and L2 (1), R = N-propylmorpholine; for L2 (2), R = N-ethylpiperazine; for L2 (3), R = N-ethylpyrrolidine, and for L1 (4), R = N-ethylmorpholine]. Dinuclear 1 and 4 possess two "end-off" compartmental ligands with exogenous mu-hydroxido and endogenous mu-phenoxido groups leading to intermetallic distances of 2.9794(15) and 2.9435(9) A, respectively; 2 and 3 are formed by two tridentate compartmental ligands where the copper centers are connected by endogenous phenoxido bridges with Cu-Cu separations of 3.0213(13) and 3.0152(15) A, respectively; 5 is built by an end-off compartmental ligand having exogenous mu-azido and endogenous mu-phenoxido groups with a Cu-Cu distance of 3.133(2) A (mean of two independent molecules). The catecholase activity of all of the complexes has been investigated in acetonitrile and methanol medium by UV-vis spectrophotometric study using 3,5-di- tert-butylcatechol (3,5-DTBC) and tetrachlorocatechol (TCC) as substrates. In acetonitrile medium, the conversion of 3,5-DTBC to 3,5-di- tert-butylbenzoquinone (3,5-DTBQ) catalyzed by 1- 5 is observed to proceed via the formation of two enzyme-substrate adducts, ES1 and ES2, detected spectroscopically for the first time. In methanol medium no such enzyme-substrate adduct has been detected, and the 3,5-DTBC to 3,5-DTBQ conversion is observed to be catalyzed by 1- 5 very efficiently. The substrate TCC forms an adduct with 2- 5 without performing further oxidation to TCQ due to the high reduction potential of TCC (in comparison with 3,5-DTBC). But most interestingly, 1 is observed to be effective even in TCC oxidation, a process never reported earlier. Kinetic experiments have been performed to determine initial rate of reactions (3,5-DTBC as substrate, in methanol medium) and the activity sequence is 1 > 5 > 2 = 4 > 3. A treatment on the basis of Michaelis-Menten model has been applied for kinetic study, suggesting that all five complexes exhibit very high turnover number, especially 1, which exhibits turnover number or K cat of 3.24 x 10 (4) (h (-1)), which is approximately 3.5 times higher than the most efficient catalyst reported to date for catecholase activity in methanol medium.

Catalytic oxidation of 3,5-di- tert-butylcatechol by a manganese(III) 18-azametallacrown-6 compound: Synthesis, crystal structure, fluorescence, magnetic and kinetic investigation

A new macrocyclic hexanuclear manganese(III) 18-azametallacrown-6 compound, [Mn 6(ashz) 6(CH 3OH) 3(H 2O) 3] · 3H 2O · 3DMF ( 1), has been prepared using a trianionic pentadentate ligand N-acetylsalicylhydrazide (ashz 3−) and characterised by various techniques such as elemental analysis, IR, UV-vis and fluorescence spectroscopy, cyclic voltammetry and X-ray diffraction. Six ashz 3− ligands connect six metal ions to form the cyclic skeleton based on the M–N–N–M linkage. Due to the meridional coordination of the ligand to the Mn 3+ ion, the ligand enforces the stereochemistry of the Mn 3+ ions as a propeller configuration with alternating Δ/Λ forms. The kinetic studies on catecholase activity of 1 for the oxidation of 3,5-di- tert-butylcatechol (3,5-DTBC) by O 2 were done using UV–Vis absorption spectra method. Compound 1 has been evaluated as a model system for the catechol oxidase enzyme and it is found that the compound shows high catecholase activity. It exhibits the activity with a turnover number of 270 h −1. A kinetic treatment on the basis of the Michaelis–Menten model has been applied. The magnetic susceptibility (300–5 K) study indicates antiferromagnetic exchange interactions with J = −2.6 cm −1 between the adjacent Mn 3+ ions.

Structural and functional models for the dinuclear copper active site in catechol oxidases: Synthesis, X-ray crystal structures, magnetic and spectroscopic properties of μ-methoxo-bridged dinuclear copper(II) complexes with N-substituted sulfonamide ligands

Two new μ-methoxo-bridged dinuclear copper(II) complexes with a N-substituted sulfonamide, [Cu(μ-OMe)(L)(NH 3)] 2 ( 1) and [Cu(μ-OMe)(L)(DMSO)] 2 ( 2) [HL, N-2-(4-methylbenzothiazole)benzenesulfonamide], have been prepared and characterized by single-crystal X-ray difraction analyses. Compound 1 crystallizes in the monoclinic space group C 2/ c with a=22.0678(18), b=7.9134(7), c=21.1186(18)Å, β=113.788(4)° and Z=8. Compound 2 crystallizes in the monoclinic space group C 2/ c with a=18.0900(10), b=9.5720(10), c=24.2620(10) Å, β=98.7120(10)° and Z=8. In both complexes the copper atoms have square–planar environments bridged by two oxygen atoms from methoxide groups. Magnetic susceptibility measurements indicate a very strong antiferromagnetic coupling between the copper(II) ions in both complexes (2 J<−1000 cm −1). Electronic Paramagnetic Resonance (EPR) spectra of the two complexes both in solid and in solution are silent. 13C NMR spectra of the complexes in solid state have been studied. The complexes have been evaluated as model systems for the catechol oxidase enzyme using 3,5-di- tert-butylcatechol as the test substrate. Complex 2 is slightly more active than complex 1.

Tuning the activity of catechol oxidase model complexes by geometric changes of the dicopper core.

Dicopper(II) complexes of a series of different pyrazolate-based dinucleating ligands [L1](-)-[L4](-) have been synthesized and characterized structurally and spectroscopically. A major difference between the four complexes is the individual metal-metal separation that is enforced by the chelating side arms of the pyrazolate ligand scaffold: it varies from 3.45 A in 2 x (BF4)4 to 4.53 A in 4 x (ClO4)2. All complexes have been evaluated as model systems for the catechol oxidase enzyme by using 3,5-di-tert-butylcatechole (DTBC) as the test substrate. They were shown to exhibit very different catecholase activities ranging from very efficient to poor catalysts (k(obs) between 2430+/-202 and 22.8+/-1.2 h(-1)), with an order of decreasing activity 2 x (ClO4)4 > 1 x (ClO4)2 > 3 x (ClO4)2 >> 4 x (ClO4)2. A correlation of the catecholase activities with the variation in Cu...Cu distances, as well as other effects resulting from the distinct redox potentials, neighboring groups, and the individual coordination spheres are discussed. Saturation behavior for the rate dependence on substrate concentration was observed in only two cases, that is, for the most active 2 x (ClO4)4 and for the least active 4 x (ClO4)2, whereas a catalytic rate that is almost independent of substrate concentration (within the range studied) was observed for 1 x (ClO4)2 and 3 x (ClO4)2. H2O2 was detected as the product of O2 reduction in the catecholase reaction of the three most active systems. The structures of the adducts of "L3Cu2" and "L4Cu2" with a substrate analogue (tetrachlorocatecholate, TCC) suggest a bidentate substrate coordination to only one of the copper ions for those catalysts that feature short ligand side arms and correspondingly exhibit larger metal-metal separations; this possibly contributes to the lower activity of these systems. TCC binding is supported by several H-bonding interactions to water molecules at the adjacent copper or to ligand-side-arm N-donors; this emphasizes the importance of functional groups in proximity to the bimetallic active site.

A Novel Catechol Oxidase Enzyme Electrode for the Specific Determination of Catechol.

An enzyme electrode for the specific determination of catechol was developed by using catechol oxidase (EC 1.10.3.1) from eggplant (Solanum melangena L.) in combination with a dissolved oxygen probe. Optimization studies of the prepared catechol oxidase enzyme electrode established a phosphate buffer 50 mM at pH 7.0 and 35°C to provide the optimum conditions for affirmative electrode response. The enzyme electrode response depended linearly on a catechol concentration range of 5•10(-7)-30•10(-5) M with a response time of 25 sec and substrate specificity of the catechol oxidase electrode of 100%. The biosensor retained its enzyme activity for at least 70 days.