Bis P-nitrophenyl Phosphate Research Articles

Presence, enzymatic activity, and subcellular localization of paraoxonases 1, 2, and 3 in human lung tissues

AimsThe human paraoxonases family (PONs) includes three calcium-dependent esterases: PON1, PON2, and PON3. The presence of PONs mRNA in human lungs is known, however, their enzymatic activity and subcellular localization have not been sufficiently explored. Main methodsIn this work, the presence of PONs in human lung tissues, at both mRNA and protein levels, was confirmed by Real-Time RT-PCR and Western blot analysis. Moreover, the activities of PONs were determined in cytosol and microsomes of 30 subjects and in mitochondria of 8 representative lung tissues using selective and non-selective substrates. Besides, to exclude the possible contribution of other esterases on PON1 organophosphate activity, the effect of bis-p-nitrophenyl phosphate (BNPP) and phenylmethylsulfonyl fluoride (PMSF), esterase inhibitors, and ethylenediaminetetraacetic acid (EDTA), a general paraoxonase inhibitor, was tested. Finally, the presence and activities of PONs in the A549 pulmonary cell line were also evaluated in order to be used as a model for studies on paraoxonases' metabolism. Key findingsOur results demonstrated high interindividual variability in both PONs mRNA/protein levels and enzymatic activities and pointed out the presence of all PONs in human lungs and their subcellular distribution in the cytosol, microsomes, and mitochondria. SignificanceThese findings add further information to our knowledge of pulmonary metabolism and, given that PON1 can metabolize some drugs used for respiratory diseases, the presence of PON1 activity in the lung tissue should no longer be ignored in the development of treatment plans and the design of new drugs.

Tetranuclear iron(III) complexes with a carboxylate-rich ligand as synthetic mimics of phosphoesterases in aqueous media

The characterization and catalytic activities of two tetranuclear Fe(III) complexes of a carboxylate-rich ligand, N,N’-Bis[2-carboxybenzomethyl]-N,N’ -Bis[carboxymethyl]-1,3-diaminopropan-2-ol (H5ccdp) ligand towards the hydrolysis of p-nitrophenyl phosphate (PNPP) and bis(p-nitrophenyl) phosphate (BNPP) substrates in aqueous systems are described. Kinetic investigations were carried out using UV–vis spectrophotometric techniques at 25 °C and 37 °C and varied pH conditions (7 – 9.5). The kinetic studies showed that the turnover rate (kcat) values for the hydrolysis of PNPP by [Fe4(ccdp)2(μ-O)(μ-OH)(d-GluA)2]3- were marginally higher than [Fe4(ccdp)2(OH)2(o-phth)2]4- at 9.24 × 10-6 s−1 (pH 8 and 37 °C) and 4.27 × 10-7 s−1 (pH 8 and 25 °C), respectively. However, similar comparisons among the BNPP hydrolysis showed the turnover rate values for [Fe4(ccdp)2(OH)2(o-phth)2]4- was marginally better than [Fe4(ccdp)2(μ -O)(μ –OH)(d-GluA)2]3- with values of 1.85 × 10-6 s−1 (pH 8 and 37 °C) and 6.23 × 10-7 (pH 8 and 37 °C). ESI-MS techniques detected catalyst-substrate adduct species for both metal complexes with PNPP. Additionally, both complexes also showed evidence for complex-substrate-nucleophile adduct formation with both BNPP and PNPP substrates via ESI-MS. Based on the kinetic activity and structural information obtained in this study, a reaction mechanism for the hydrolysis of phosphorester has been proposed.

Structural and functional identification of the uncharacterized metallo-β-lactamase superfamily protein TW9814 as a phosphodiesterase with unique metal coordination.

Metallo-β-lactamase (MBL) superfamily proteins have a common αβ/βα sandwich fold and perform a variety of functions through metal-mediated catalysis. However, because of the enormous scale of this superfamily, only a small percentage of the proteins belonging to the superfamily have been annotated structurally or functionally to date. Therefore, much remains unknown about the MBL superfamily proteins. Here, TW9814, a hypothetical MBL superfamily protein, was structurally and functionally investigated. Guided by the crystal structure of dimeric TW9814, it was demonstrated that TW9814 functions as a phosphodiesterase (PDE) in the presence of divalent metal ions such as manganese(II) or nickel(II). A docking model between TW9814 and the substrate bis(p-nitrophenyl)phosphate (bpNPP) showed the importance of the dimerization of TW9814 for its bpNPP-hydrolyzing activity and for the interaction between the enzyme and the substrate. TW9814 showed outstanding catalytic efficiency (kcat/Km) under alkaline conditions compared with other PDEs. The activity of TW9814 appears to be regulated through a disulfide bond, which is a feature that is not present in other MBL superfamily members. This study provides a platform for the functional characterization of other hypothetical proteins of the MBL or other superfamilies.

Homodinuclear copper(II) and zinc(II) complexes of a carboxylate-rich ligand as synthetic mimics of phosphoester hydrolase in aqueous solutions

The synthesis, characterization and catalytic activities of two homodinuclear Cu(II) and Zn(II) complexes of a carboxylate-rich ligand, N,N′-Bis[2-carboxybenzomethyl]-N,N′ -Bis[carboxymethyl]-1,3-diaminopropan-2-ol (H5ccdp) ligand towards the hydrolysis of (p-nitrophenyl phosphate) (PNPP) and bis(p-nitrophenyl) phosphate (BNPP) substrates in aqueous systems are described. Kinetic investigations were carried out using UV–Vis spectrophotometric techniques at 25 °C and 37 °C and different pH (7–10) conditions. The kinetic studies revealed that the turnover rate (kcat) values among the PNPP hydrolysis systems, the highest and the lowest kcat values were displayed by [Cu2(ccdp)(μ-OAc)]2− at 2.34 × 10−6 s−1 (pH 8 and 37 °C) and 2.13 × 10−8 s−1 (pH 8 and 25 °C), respectively. However, similar comparisons among the BNPP hydrolysis revealed that highest and the lowest kcat values were displayed by [Zn2(ccdp)(μ-OAc)]2− at 4.64 × 10−8 s−1 (pH 9 and 37 °C) and 2.38 × 10−9 (pH 9 and 25 °C). Significantly enough, the catalyst−substrate adduct species containing a metal bound PNPP and BNPP have been detected by ESI-MS techniques. Additionally, a PNPP-bound copper complex has been isolated and crystalized using single crystal X-ray diffraction technique. Based on the structural and activity information obtained in this study, reaction mechanisms for the hydrolysis of PNPP have been proposed.

2-Aminoterephthalic acid co-coordinated Co-MOF fluorescent probe for highly selective detection of the organophosphorus pesticides with p-nitrophenyl group in water systems

It has drawn extensive research interest to integrate luminescent ligands into metal-organic frameworks as luminescent materials due to striking luminescent properties. In this work, a Co-MOF with weak fluorescence emission, [Co(OBA)(2,2′-BPY)], was synthesized by a mixed solvothermal route with Co(NO3)2 as the Co2+ ion source, 4,4′-Oxybis(benzoic acid) (OBA) and 2,2′-Bipyridyl (2,2′-BPY) as the ligands. Its fluorescence was obviously enhanced through integrating a small amount of luminescent ligands: 2-aminoterephthalic acid. Interestingly, the above enhanced fluorescence could be selectively quenched by the organophosphorus pesticides with p-nitrophenyl group such as p-nitrophenyl phosphate (PNPP) and bis(p-nitrophenyl) phosphate (BNPP) in water systems. As a case, the detection limit towards PNPP was up to 352 nM (0.07 mg kg−1), which is far lower than the standards for similar pesticide residues in fruits in China (0.1–0.6 mg kg−1) [1]. At the same time, the above phenomenon of the fluorescence quenching was not interfered from some other pesticides, metallic ions, amino acids and aromatic nitrocompounds, implying the present fluorescent material has potential application as a highly selective probe for detecting PNPP pesticide residues in food and domestic water. Furthermore, a possible quenching mechanism was discussed.

Bifunctional TiO2/CeO2 reactive adsorbent/photocatalyst for degradation of bis-p-nitrophenyl phosphate and CWAs

TiO2/CeO2 nanocomposites were prepared by simple refluxing of peroxo titanium and cerium complexes in water at ambient air and they were used for degradation of several organophosphorus compounds (OPC). The combination of high surface reactivity of ceria with photoactivity of titania led to i) rapid reactive adsorption of bis-p-nitrophenyl phosphate (BNPP) forming p-nitrophenyl phosphate (NP) and ii) subsequent photo-induced degradation of both BNPP and NP. The composites were also three and four times more efficient for rapid degradation of chemical warfare agents (CWAs) soman and VX in nonane compared to bare ceria and titania, respectively. The strong interaction of Ti with Ce within the composites led to the formation of Ce3+ and Ti<4+ states, reduction of titania crystallite size, change of acid-base and surface properties and synergetic effects that are all responsible for highly improved degradation efficiency of these bifunctional nanocomposites with considerable potential of application for abatement of various toxic compounds.

Phosphatase-like Activity of Tetranuclear Iron(III) and Zinc(II) Complexes.

Three new tetranuclear iron(III) and zinc(II) complexes, [Fe4(cpdp)2(phth)2(OH)2]·8H2O (1), [Fe4(cpdp)2(terephth)2(OH)2] (2), and [Zn4(Hcpdp)2(suc)]Br2·12H2O (3), have been synthesized as models for the active site of phosphoester hydrolases by utilizing a polydentate ligand, N, N'-bis[2-carboxybenzomethyl]- N, N'-bis[2-pyridylmethyl]-1,3-diaminopropan-2-ol (H3cpdp) in combination with exogeneous phthalate (phth), terephthalate (terephth), and succinate (suc). Single crystal X-ray analyses reveal that the metallic core of complex 1 consists of four distorted octahedral iron(III) ions with average intraligand Fe---Fe separation of 3.656(2) Å, while the structure 3 represents a tetranuclear metallic core containing four distorted trigonal bipyramidal zinc(II) ions with average intraligand Zn---Zn separation of 3.472(2) Å. The molecular structure of complex 2 has been optimized by the DFT method which shows that its core arrangement is similar to that of 1. Complex 1 has a very interesting centrosymmetric structure that includes two crystallographically equivalent [Fe2(cpdp)]3+ dinuclear units, connected together by a pair of syn-syn bridging phthalates and a pair of bridging hydroxides to generate a "dimer of dimers" structural motif. In complex 3, a succinate group connects two crystallographically equivalent [Zn2(Hcpdp)]2+ dinuclear units in a syn-syn bidentate manner forming a "dimer of dimers" structural design. All three complexes show phosphatase-like activity that has been examined in methanol-water (1:1; v/v) using bis( p-nitrophenyl) phosphate (BNPP) as model substrate by applying the UV-vis spectrophotometric technique. In each case, the kinetic data have been analyzed by the Michaelis-Menten approach. The order of catalytic efficiency for the conversion of substrate to product follows the trend 1 > 2 > 3 with turnover rates ( kcat) of (2.73 ± 0.13) × 10-5 for 1, (1.06 ± 0.07) × 10-5 for 2, and (2.33 ± 0.18) × 10-6 s-1 for 3. These kcat values are comparable to, albeit slightly lower than, the values reported for similar iron(III)- and zinc(II)-based model complexes in the literature. DFT calculations have been carried out to support the proposed mechanism for phosphatase-like activity.

Development of simultaneous quantification method of loteprednol etabonate (LE) and its acidic metabolites, and analysis of LE metabolism in rat

Loteprednol etabonate (LE) is a soft corticosteroid with two labile ester bonds at 17α- and 17β-positions. Its corticosteroidal activity disappears upon hydrolysis of either ester bond. Hydrolysis of both ester bonds produces the inactive metabolite, Δ1-cortienic acid (Δ1-CA).The simple high-performance liquid chromatography method using acetic acid gradient was developed for the simultaneous determination of LE and its acidic metabolites.LE was hydrolyzed in rat plasma with a half-life of 9 min. However, LE hydrolysis was undetectable in rat liver and intestine. LE hydrolysis in rat plasma was completely inhibited by paraoxon and bis(p-nitrophenyl) phosphate, thus identifying carboxylesterase as the LE hydrolase. Rat plasma carboxylesterase had a Km of 6.7 μM for LE.In contrast to the disappearance rate of LE in rat plasma, the formation rate of 17α-monoester and Δ1-CA was markedly low, and a main hydrolysate of LE was not detected in rat plasma.The metabolism of LE proceeded via different pathways in human and rat plasma. LE was slowly hydrolyzed by paraoxonase in human plasma to 17α-monoester with a half-life of 12 h, but by carboxylesterase in rat plasma to yield undetectable products, presumed to include the unstable 17β-monoester.

Synthesis, characterization and biological properties of mixed ligand complexes of cobalt(II/III) valproate with 2,9‐dimethyl‐1,10‐phenanthroline and 1,10‐phenanthroline

The synthesis of mononuclear cobalt(II/III) complexes with two different ligands (complex 2: [Co(valp)2(2,9‐dmp)] and complex 3: [Co(valp)2(H2O)(1,10‐phen)]) was investigated and the characterization of both complexes was achieved using IR, UV–Vis, and single crystal X‐ray diffraction. Using single crystal X‐ray diffraction, the crystal structure of each of the complexes was determined. Additionally, the biological activity of these complexes was studied in five gram‐positive and four gram‐negative bacterial strains. Whereas in all gram‐negative bacteria tested, cobalt valproate complexes did not show any anti‐bacterial activity, both complexes had effects on gram positive bacteria. Complex 2 demonstrated good anti‐bacterial activity against all gram‐positive bacteria with inhibition zone diameter (IZD) ranging between 15–28 mm. Complex 3 exhibited low inhibition activity against all gram‐positive bacteria except E. faecalis with IZD ranging between 11.3–13.7 mm. Moreover, as an indication of its uses as industrial catalyst, the rate of bis(p‐nitrophenyl) phosphate (BNPP) hydrolysis when catalyzed by these complexes was measured at different temperatures, concentrations and pH. Complex 2 proved to be a better catalyst to induce the hydrolysis of BNPP.

An atypical phosphodiesterase capable of degrading haloalkyl phosphate diesters from Sphingobium sp. strain TCM1

Sphingobium sp. strain TCM1 can degrade tris(2-chloroethyl) phosphate (TCEP) to inorganic phosphate and 2-chloroethanol. A phosphotriesterase (PTE), phosphodiesterase (PDE) and phosphomonoesterase (PME) are believed to be involved in the degradation of TCEP. The PTE and PME that respectively catalyze the first and third steps of TCEP degradation in TCM1 have been identified. However, no information has been reported on a PDE catalyzing the second step. In this study, we identified, purified, and characterized a PDE capable of hydrolyzing haloalkyl phosphate diesters. The final preparation of the enzyme had a specific activity of 29 µmol min−1 mg−1 with bis(p-nitrophenyl) phosphate (BpNPP) as the substrate. It also possessed low PME activity with p-nitrophenyl phosphate (pNPP) as substrate. The catalytic efficiency (kcat/Km) with BpNPP was significantly higher than that with pNPP, indicating that the enzyme prefers the organophosphorus diester to the monoester. The enzyme degraded bis(2,3-dibromopropyl) phosphate, bis(1,3-dichloro-2-propyl) phosphate and bis(2-chloroethyl) phosphate, suggesting that it is involved in the metabolism of haloalkyl organophosphorus triesters. The primary structure of the PDE from TCM1 is distinct from those of typical PDE family members and the enzyme belongs to the polymerase and histidinol phosphatase superfamily.

Phosphoryl Transfer Processes Promoted by a Trifunctional Calix[4]arene Inspired by DNA Topoisomerase I.

The cone-calix[4]arene derivative (1H3)2+, decorated at the upper rim with two guanidinium units and a phenolic hydroxyl in an ABAH functionalization pattern, effectively promotes the cleavage of the DNA model compound bis(p-nitrophenyl) phosphate (BNPP) in 80% DMSO solution at pH values in the range 8.5-12.0. The pH dependence of the kinetics was found to be fully consistent with the results of the potentiometric titration of the triprotic acid (1H3)2+. At pH 9.5, the rate enhancement of p-nitrophenol liberation from BNPP relative to background hydrolysis is 6.5 × 104-fold at 1 mM concentration of the calix[4]arene derivative. Experimental data clearly point to the effective cooperation of the three active units and to the involvement of the phenolate moiety as a nucleophile in the phosphoryl transfer step. Subsequent liberation of a second equivalent of p-nitrophenol from the phosphorylated calixarene intermediate is conceivably promoted by the "built-in" guanidine/guanidinium catalytic dyad.

Synthesis and phosphate ester cleavage properties of copper(II) complexes of guanidinium-bridged bis(1,4,7-triazacyclononane) ligands

The synthesis and characterization of three new bis(1,4,7-triazacyclononane) ligands, L1, L2 and L3, featuring a bridging guanidinium group between two macrocyclic units, is reported. The corresponding binuclear copper(II) complexes have been studied as agents to accelerate the cleavage of P–O bonds within two model phosphodiesters, namely bis(p-nitrophenyl)phosphate (BNPP) and 2-hydroxypropyl-p-nitrophenylphosphate (HPNPP). The results of a comparative study of cleavage rates, using the mononuclear copper(II)–tacn complexes bearing single alkylguanidinium groups as a reference, revealed that the binuclear copper(II) complexes are generally less effective cleavage agents, which may be related to a tendency to form hydrolytically inactive hydroxo-bridged species at near-neutral pH and above. However, at pH 7, these complexes produced 4–18-fold increases in the rate of BNPP hydrolysis compared to the parent complex, [Cu(tacn)(OH2)2]2+. Likewise, at pH 6, the complexes cleaved HPNP 5–130-fold faster than [Cu(tacn)(OH2)2]2+, suggesting some degree of cooperative interplay between the two proximal copper(II) centers and the protonated guanidinium bridging group in promoting phosphodiester hydrolysis under these conditions.

Applications of Coordination Chemistry in Biological Systems

A novel 2,2′:6′,2″-terpyridine–picolylamine-based bridging ligand has been synthesized and fully characterized using a variety of analysis techniques including single crystal X-ray diffraction. As shown in figure (a), the ligand has both tridentate and bidentate metal binding sites available to coordinate with various metal ions. By varying the size of anions both dinuclear complexes and supramolecular assemblies have been produced. Addition of metal salts containing small anions like halides result in formation of Cu2L and Zn2L dinuclear complexes, figure (b), where one metal ion binds at each of the binding sites of the ligand. The metal ions in these complexes mimic active site of the hydrolytic enzymes and promote phosphatediester hydrolysis of model DNA/RNA compounds. Nearly ten times increase in the rate of hydrolysis of bis(p-nitrophenyl)phosphate (BNPP) is observed in comparison to the parent terpyridine and picolylamine complexes under physiological conditions. Larger anions like PF6-, ClO4-, SO42- , NO3- result in formation of Zn4L4 type squares via. head-to-head and tail-to-tail, HH-TT, (H=tridentate site, T=bidentate site) coordination of the ligand. The octahedrally bound Zn(II) ion between two tridentate sites can be replaced with Fe(II) to prepare Fe2Zn2L4 squares. A flat molecule of terephthalic acid was also deliberately encapsulated in the middle of the Fe2Zn2L4 square as shown in figure (c). The head-to-tail, HT, coordination of the ligand in case of Ni(II) results in formation of decanickel wheels, like [Ni10L10Cl4(H2O)6](Cl)15Br·~140H2O shown in figure (d). Due to the large structure of the molecule X-ray crystallographic studies rather have been quite challenging.

Tripodal molecules for the promotion of phosphoester hydrolysis.

A series of low molecular weight tripodal amide/histidine-containing compounds (1-2) have been synthesised and shown to increase the rate of bis-(p-nitrophenyl) phosphate (BNPP) and soman (GD) breakdown in buffered aqueous solution.

Hydrolysis of BNPP Catalyzed by Metallomicelles Made of Pr(III) Complexes

A macrocyclic ligand was synthesized and characterized. The kinetics of hydrolysis of bis(p-nitrophenyl)phosphate (BNPP) in the catalytic system containing macrocyclic ligand and praseodymium(III) was investigated. The analysis of specific absorption spectrums of the hydrolytic reaction systems indicated that key intermediates made up of BNPP and praseodymium(III) complexes are formed in the reaction process of BNPP catalytic hydrolysis. In this, the mechanism of BNPP catalytic hydrolysis proposed is based on the analytic result of specific absorption spectrum, and the corresponding kinetic constants are calculated. The results showed that the praseodymium(III) complexes as hydrolase mimics exhibit good catalytic activity and similar catalytic character to natural enzyme.

Impact of Endogenous Esterase Activity on In Vitro P-Glycoprotein Profiling of Dabigatran Etexilate in Caco-2 Monolayers

Dabigatran etexilate, a double prodrug of dabigatran, is a reversible, competitive, direct thrombin inhibitor that has been approved for use in many countries. A recent guideline from the European Medicines Agency on drug-drug interactions proposed dabigatran etexilate as a sensitive in vivo and in vitro probe substrate for intestinal P-glycoprotein (P-gp) inhibition. We therefore performed a series of in vitro studies to determine the best experimental conditions for evaluation of P-gp involvement on the transport process of dabigatran etexilate across colorectal adenocarcinoma Caco-2 cell monolayers. Experiments using expressed carboxylesterase 1 (CES1) and CES2 bactosomes revealed that dabigatran etexilate was hydrolyzed into BIBR 1087 by CES1 expressed in our Caco-2 cells. The impact of CES1-mediated BIBR 1087 formation during transcellular transport experiments was assessed by comparing several combinations of three experimental approaches: radioactivity detection using [(14)C]dabigatran etexilate as substrate, liquid chromatography-tandem mass spectrometry (LC-MS/MS) quantification of dabigatran etexilate, and in the presence and absence of a CES inhibitor bis(p-nitrophenyl) phosphate (BNPP). The experimental approach that was based on the use of nonlabeled dabigatran etexilate together with LC-MS/MS quantification and the addition of BNPP was selected as the most favorable condition in which to correctly evaluate the permeability coefficient (Papp) of dabigatran etexilate and its transcellular transport by P-gp. The in vitro Caco-2 study at the selected condition revealed that dabigatran etexilate is a P-gp substrate with an efflux ratio of 13.8 and an intrinsic Papp, which is the Papp under the condition of complete blockage of P-gp by P-gp inhibitor, of 29 × 10(-6) cm/s.

Multiple Cytochrome P450 Isoforms Are Involved in the Generation of a Pharmacologically Active Thiol Metabolite, whereas Paraoxonase 1 and Carboxylesterase 1 Catalyze the Formation of a Thiol Metabolite Isomer from Ticlopidine

Ticlopidine is a first-generation thienopyridine antiplatelet drug that prevents adenosine 5'-diphosphate (ADP)-induced platelet aggregation. We identified the enzymes responsible for the two-step metabolic bioactivation of ticlopidine in human liver microsomes and plasma. Formation of 2-oxo-ticlopidine, an intermediate metabolite, was NADPH dependent and cytochrome P450 (CYP) 1A2, 2B6, 2C19, and 2D6 were involved in this reaction. Conversion of 2-oxo-ticlopidine to thiol metabolites was observed in both microsomes (M1 and M2) and plasma (M1). These two metabolites were considered as isomers, and mass spectral analysis suggested that M2 was a thiol metabolite bearing an exocyclic double bond, whereas M1 was an isomer in which the double bond was migrated to an endocyclic position in the piperidine ring. The conversion of 2-oxo-ticlopidine to M1 in plasma was significantly increased by the addition of 1 mM CaCl2. In contrast, the activity in microsomes was not changed in the presence of CaCl2. M1 formation in plasma was inhibited by EDTA but not by other esterase inhibitors, whereas this activity in microsomes was substantially inhibited by carboxylesterase (CES) inhibitors such as bis-(p-nitrophenyl)phosphate (BNPP), diisopropylphosphorofluoride (DFP), and clopidogrel. The conversion of 2-oxo-ticlopidine to M1 was further confirmed with recombinant paraoxonase 1 (PON1) and CES1. However, M2 was detected only in NADPH-dependent microsomal incubation, and multiple CYP isoforms were involved in M2 formation with highest contribution of CYP2B6. In vitro platelet aggregation assay demonstrated that M2 was pharmacologically active. These results collectively indicated that the formation of M2 was mediated by CYP isoforms whereas M1, an isomer of M2, was generated either by human PON1 in plasma or by CES1 in the human liver.

Kinetic and Mechanistic Investigations of the Catalysed Hydrolysis of Bis(p-Nitrophenyl) Phosphate by a Macrocyclic Neodymium(III) Complex

The macrocyclic ligand 5,5,7,12,12,14-hexamethyl-1,4,8,11-tetraazacyclo-tetradecane (L) was synthesised and characterised. The kinetics of hydrolysis of bis( p-nitrophenyl) phosphate (BNPP) in a catalytic system comprising the macrocyclic ligand and neodymium(III) was investigated. The analysis of specific absorption spectra of the hydrolytic reaction system indicated that key intermediates containing BNPP and the Nd(III) complex are formed in the reaction. The mechanism proposed for the catalytic hydrolysis proposed is based on the analytical results of the specific absorption spectrum, and the corresponding kinetic constants are calculated. The results showed that the Nd(III) complex as a mimic hydrolase exhibits good catalytic activity, i.e. similar to that of the natural enzyme.

Hydrolysis of BNPP Catalyzed by the Dodecyliminodiacetate Nickel (II) Complexes

One novel dodecyliminodiacetate nickel (II) complex were synthesized and characterized, and these complexes were used as mimic hydrolytic in catalytic hydrolysis of bis (p-nitrophenyl) phosphate (BNPP). The analysis of specific absorption spectra of the hydrolytic reaction systems indicated that the catalytic hydrolysis involved the key intermediates formed by BNPP with nickel (II) complexes. The kinetic parameter of BNPP catalytic hydrolysis has been calculated and the activation energy for the catalytic hydrolysis is 95.26kJ·mol-1.

Hydrolysis of phosphodiester catalyzed by metallomicelles with histidine residue: Kinetics and mechanism

A new 2-amino-N-dodecyl-3-(1H-imidazole-4-yl)propanamide (L2) surfactant was synthesized and its metallomicelles made of La(III), Cu(II), Co(II) and Zn(II) complexes were used as hydrolytic metalloenzyme mimics to catalyze the hydrolysis of phosphodiester bis(p-nitrophenyl) phosphate (BNPP) in buffer solution at 35°C. The effects of long alkyl ligand, metal ions, temperature and pH on the catalytic hydrolysis of phosphodiester were investigated. The ternary complex kinetic model for metallomicellar catalysis was suggested to analyze the experimental data. The kinetic and thermodynamic parameters k′N, KM, KT, Km and pKa were obtained. The effect of pH was especially analyzed. The pH-independent rate constant of the reaction catalyzed by metallomicelles LaL2, CuL2, CoL2 and ZnL2 was evaluated as 7.03×10−4, 6.51×10−5, 4.28×10−5 and 1.78×10−5s−1, respectively. The different association manners between metallomicelles and substrate were proposed. A hydroxide ion OH− associated metallomicelle, which was detected by ESI-MS spectral analysis, was suggested as the real active species to catalyze the hydrolysis of phosphodiester. Imidazole group might accelerate the hydrolysis by activating H2O associated the metal into metal-OH−. Effect of microenvironment of micelle on the catalysis hydrolysis of BNPP was also discussed.