Hydrolysis Of 4-nitrophenyl Phosphate Research Articles

Acid phosphomonoesterase activity in floodplain forest soils

The 4-nitrophenyl phosphate method for acid phosphomonoesterase (EC 3.1.3.2; acid orthophosphoric monoester phosphohydrolase) determination was slightly modified to increase the sensitivity and the stability of the 4-nitrophenol coloured complex in samples rich in humic substances and easily soluble organic compounds. Based on an approach used for mycorrhizal roots, the new analytical protocol was tested on samples taken from a single forest site with a large variation in soil types. Distinctive properties of forest soils and the accuracy/repeatability of the optimised technique were considered in the selection of the most appropriate analytical steps. Finally, the acid phosphomonoesterase activity was expressed in μg of 4-nitrophenol/g fresh soil recalculated dry matter/h released after hydrolysis of 4-nitrophenyl phosphate in sodium tetraborate/succinic acid buffer at pH 4.8. The modified method proposed was statistically compared with other procedures and the results obtained were found to give merit to these slight modifications. The rationale behind the slight modifications and their comparative significance was reported. Due to high soil complexity, it is suggested that a single laboratory measurement of chemical and biochemical properties is of limited application with forest soil and can be only one component of a broadly focused ecological analysis.

Synthesis, spectral, magnetic, electrochemical and kinetic studies of copper(II), nickel(II) and zinc(II) acetate complexes derived from phenol based ‘end-off’ ligands: Effect of p-substituents

A new class of symmetric, end-off, N-methyl piperazine armed binucleating ligands 2,6-bis(4-methyl piperazin-1-yl-methyl)-4-acetyl phenol (HL 1) and 2,6-bis[(4-methyl piperazin-1-yl-methyl)]-(4-methylcarboxy) phenol (HL 2) were synthesized by the Mannich reaction. Their mononuclear and binuclear Cu(II), Ni(II) and Zn(II) complexes have been synthesized. These complexes were characterized by elemental analysis, infra-red and electronic spectral analysis. In the electronic spectra, the lower electron withdrawing nature of the C(O)CH 3 p-substituent (HL 1) compared with the C(O)OCH 3 p-substituent (HL 2) of the phenolic ring causes a red shift in the LMCT-charge transfer band. The mononuclear Cu(II) complexes 1 and 7 have a magnetic moment value close to the spin only value with four hyperfine EPR signals. The binuclear Cu(II) complexes 4 and 10 illustrate an antiferromagnetic interaction ( μ eff 1.56 and 1.55 BM) at 298 K with a broad EPR signal. A variable temperature magnetic moment study of the binuclear copper(II) complexes shows that the extent of antiferromagnetic coupling increases in the order: CHO [K. Shanmuga Bharathi, A. Kalilur Rahiman, K. Rajesh, S. Sreedaran, P.G. Aravindan, D. Velmurugan, V. Narayanan, Polyhedron 25 (2006) 2859] < C(O)CH 3 < C(O)OCH 3 (−2 J values 134 [Shanmuga Bharathi et al., mentioned above], 149 and 158 cm −1, respectively). The mononuclear Ni(II) complexes 2 and 8 are square planar and diamagnetic. The six coordinated binuclear Ni(II) complexes 5 and 11 show a magnetic moment value of 2.96 and 2.95 BM, respectively. Electrochemical studies of the complexes reveal that all the mononuclear complexes show a single irreversible one-electron transfer reduction wave and the binuclear complexes show two irreversible one-electron transfer reduction waves in the cathodic region. There is an anodic shift in the reduction of the metal centres when the electron withdrawing nature of the p-substituent of the phenolic ring increases. The catecholase activity of the mono and binuclear copper(II) complexes, using pyrocatechol as a model substrate, and the hydrolysis of 4-nitrophenyl phosphate using the mono and binuclear copper(II), nickel(II) and zinc(II) complexes as catalysts showed that the binuclear complexes have higher rate constant values than those of the corresponding mononuclear complexes. A comparison of the spectral, electrochemical and magnetic behaviour of the complexes derived from the ligands is discussed on the basis of the substituent at the para position of the phenolic ring.

Iron Promoted Decontamination Studies of Nitrophenylphosphate in Aqueous and Microemulsion Media: A Model for Phosphate Ester Decontamination in the Environment

The hydrolysis of 4-Nitrophenylphosphate (NPP) in the presence of Ferrous and Ferric ions in Oil in water microemulsion media was investigated. The reaction was monitored by measuring the absorbance of the nitrophenolate ion produced in the reaction aliquots with time. The order of effectiveness of the microemulsion system towards the hydrolysis of NPP was found to be cationic > anionic> aqueous in the presence of Ferrous, Ferric and Turnbull’s blue at neutral pH. The result of the present investigation enriches our understanding of the possible roles polymetallic ions play in hydrolysis reactions. The reactions mimic the roles of purple acid phosphatases in the hydrolysis of phosphate esters. The application of the above systems for environmental decontamination of organophosphates is also envisoned.

Structural and kinetic alterations of constitutive conidial alkaline phosphatase from the osmotically-sensitive mutant ofNeurospora crassa

The osmotically-sensitive os-1 mutant of Neurospora crassa overproduced conidial alkaline phosphatase. The enzyme was purified by Phenyl-Sepharose CL-4B chromatography and Sephadex G-200 gel filtration. PAGE analysis of the purified enzyme suggested the occurrence of aggregation and/or disaggregation phenomena. The enzyme is a glycoprotein containing 16% saccharide, with apparent molar mass of 137 kDa. Two protein bands (36 and 62 kDa) were observed in SDS-PAGE, suggesting that the native enzyme was a trimer. The pI was estimated to be 2.7, and optima of pH and temperature were 9.5 and 65 degrees C, respectively. The enzyme showed broad substrate specificity, hydrolyzing preferentially 4-nitrophenyl phosphate, O-phosphoamino-acids and 2-phosphoglycerate. The hydrolysis of 4-nitrophenyl phosphate was stimulated by Co(II) (26%), Ni(II) (23%) and Mg(II) ions (80%). The enzyme was stable for up to 6 months at 4 degrees C in 5 mmol/L Tris-HCl buffer and also upon storage at 25 degrees C for 10 d. The kinetic and structural properties of the conidial enzyme purified from the os-1 mutant were quite different from those of the wild type strain. The enzyme overproduction observed in the mutant may be related to cell wall alterations that affect the process of enzyme secretion.

Synthesis of new ‘end-off’ μ-phenoxo and bis-μ-acetato tri-bridged copper(II), nickel(II) and zinc(II) complexes: Spectral, magnetic, electrochemical and catalytic studies

A new end-off type acyclic ligand with two N-methyl piperazine arms, 2,6-bis[(4-methyl piperazin-1-yl)]-4-formyl phenol [L], has been synthesized by a simple Mannich reaction. The mono and binuclear complexes of Cu(II), Ni(II) and Zn(II) have been prepared. The complexes were characterized by elemental and spectral analysis. The EPR spectrum of the mononuclear copper complex shows four hyperfine splittings and the binuclear complex shows a broad signal due to an anti-ferromagnetic interaction. The room temperature magnetic moment of the mono and binuclear copper complexes are found to be 1.72 μ B ( μ eff) and 1.58 μ B ( μ eff), respectively. The mononuclear Ni(II) complex is square planar and diamagnetic, whereas the six-coordinated binuclear Ni(II) complex shows a magnetic moment value of 2.98 μ B ( μ eff). Cyclic voltammetric studies evidenced one irreversible reduction wave for all the mononuclear complexes and two irreversible one-electron reduction waves for the binuclear complexes are obtained in the cathodic region. In the anodic region, one irreversible oxidation wave for the mononuclear nickel(II) complex and two irreversible oxidation waves for the binuclear nickel(II) complexes are obtained. Kinetic studies on the oxidation of pyrocatechol to o-quinone using the mono and binuclear copper(II) complexes as catalysts and on the hydrolysis of 4-nitrophenylphosphate using the mono and binuclear copper(II), nickel(II) and zinc(II) complexes as catalysts were carried out. The binuclear complexes have higher rate constant values than those of the corresponding mononuclear complexes. The rate constant values for the complexes for the hydrolysis are in the order nickel(II) > copper(II) > zinc(II).

4-Nitrophenyl phosphoric acid and its four different potassium salts: a solid state structure and kinetic study

The structures of 4-nitrophenyl phosphoric acid, H2NPP (1), and its four different potassium salts, K(H2NPP)(HNPP) (2), K(HNPP)·CH3OH (3), K(HNPP)·2H2O (4), and K2(NPP)·4H2O (5), were determined by X-ray diffraction methods. These investigations, together with the study of the role of potassium ions in the hydrolysis of 4-nitrophenyl phosphate, provided information about interactions between the acid, monoanionic and dianionic forms of 4-nitrophenyl phosphate and the potassium cations. In the crystalline state, the H2NPP molecules form one-dimensional chains of bifurcated O–H⋯O hydrogen bonds between phosphate groups. A different network, derived from ladder-like chains of the O–H⋯O hydrogen bonds linking the phosphate group of the H2NPP molecules and (HNPP)− monoanions, is formed in 2. By contrast, in 4, the phosphate groups of the (HNPP)− monoanions are donors and acceptors of the O–H⋯O hydrogen bonds for the water molecules only. The 4-nitrophenyl phosphate ligands differ in their coordination modes. The potassium cations are chelated by the nitro group in 2 and 4, whereas in 3 and 5 the potassium cations are chelated by the phosphate group, with the ester oxygen atom engaging in the chelating mode.

Coordination properties of a dicompartmental ligand with tetra- and hexadentate coordination sites towards copper (II) and nickel (II) ions

A new dicompartmental ligand, N, N′-bis[2-iminomethyl-4-methyl-6-(4-methylpiperazin-1-ylmethyl)1-hydroxybenzene]1,8,naphthalene (H 2L) possessing contiguous tetra- and hexadentate coordination sites has been synthesized in simple route by the Schiff’s base condensation of the precursor compound (PC) 2-formyl-4-methyl-6[4-methyl-6-(4-methylpiperazin-1-yl)methyl]phenol with 1,8-diaminonaphthalene.The mononuclear and binuclear Cu(II) and Ni(II) complexes of this ligand have been prepared and characterized by elemental analysis, IR, NMR and UV–Vis spectra.The electronic spectra of the mono- and binuclear Cu(II) complexes and mononuclear Ni(II) complex revealed one d–d transition in the region 570–600 nm, whereas the binuclear Ni(II) complex showed three d–d transitions at 800 nm ( ε ∼ 95 M −1 cm −1), 620 nm ( ε ∼ 140 M −1 cm −1) and 560 nm ( ε ∼ 250 M −1 cm −1), which were assigned to 3A 2g → 3T 2g, 3A 2g → 3T 1g, 3A 2g → 3T 1g(P), and these spin allowed electronic transitions are characteristic of an octahedral Ni 2+ centre.The electrochemical studies evidenced for the mononuclear Cu(II) complex shows a single quasireversible one electron wave at −0.83 V ( E pc).The binuclear Cu(II) complex shows two quasireversible one electron reduction waves at - 0.68 V ( E pc 1 ) and - 1.35 V ( E pc 2 ) in the cathodic region.The mononuclear Ni(II) complex shows one quasireversible reduction potential at −0.80 V ( E pc) and the binuclear Ni(II) complex shows two irreversible reduction potentials at - 1.10 V ( E pc 1 ) and - 1.40 V ( E pc 2 ) in the cathodic region.The room-temperature magnetic moment value for the mononuclear Cu(II) complex is 1.72 μ B( μ eff) and for the binuclear Cu(II), 2.05 μ B( μ eff). The mononuclear Ni(II) complex is square planar and diamagnetic whereas the binuclear Ni(II) complex has a magnetic moment value of 3.05 μ B ( μ eff). Kinetic studies on the oxidation of pyrocatechol to o-quinone using the copper (II) complexes and hydrolysis of 4-nitrophenyl phosphate using the nickel (II) complexes as catalysts showed that the binuclear complexes have higher rate constant values than that of the corresponding mononuclear complexes.

New Tetraaza Macrobicyclic Ditopic Receptors for Metal Ions: Synthesis, Redox Response and Kinetics of Phosphate Hydrolysis of Unsymmetrical Macrobicyclic Mono- and Binuclear Nickel(II) Complexes

A new series of macrobicyclic ditopic receptors is derived from the precursor compound 3,4:10,11-dibenzo-1,13[N,N′-bis{(3-formyl-2-hydroxy-5-methyl)benzyl}di-aza]-5,9-dioxocyclohexadecane. Using this precursor, mono- and binuclear nickel(II) complexes of type [NiL](ClO4) and [Ni2L](ClO4)2 have been synthesized to undertake electrochemical and catalytic studies on the basis of macrocyclic ring size. The receptor is a tricompartmental macrocycle consisting of ether oxygen, tertiary nitrogen and imine nitrogen atoms. The redox studies of these systems show that the nickel(II) complexes undergo quasi-reversible one-electron reduction and oxidation. All the nickel(II) complexes have square planar geometry and are EPR silent. Examination of the kinetics of the hydrolysis of 4-nitrophenyl phosphate shows that the catalytic activities of the complexes increase with the macrocyclic ring size of the complexes. As the macrocyclic ring size of the complexes increases, the spectral, electrochemical and catalytic studies of the complexes show considerable variation due to distortion in the geometry around the nickel(II) centre.

Polymetallic complexes in microemulsions for the hydrolysis of 4-nitrophenyl phosphate: a bio-mimetic model for decontamination of organophosphates in the environment

Polymetallic complexes in microemulsions for the hydrolysis of 4-nitrophenyl phosphate: a bio-mimetic model for decontamination of organophosphates in the environment

Synthesis, characterization and electrochemical behavior of mononuclear, binuclear nickel(II) and heterobinuclear zinc(II)nickel(II) complexes derived from novel macrobicyclic tricompartmental ligands: kinetics of the catalytic hydrolysis of 4-nitrophenyl phosphate by the complexes

Mononuclear, binuclear NiII and heterobinuclear ZnIINiII complexes have been derived from lateral macrobicyclic tricompartmental ligands embracing three different donor sets: (i) O2N2-donor set, derived from ether oxygens and tertiary amine nitrogens; (ii) N2O2-donor set, derived from tertiary amine nitrogens and phenolic oxygens; (iii) O2N2-donor set, derived from phenolic oxygens and azomethine nitrogens. Cyclic voltammograms of the mononuclear NiII complexes showed irreversible one-electron reduction processes in the −1.2 to −1.3 V region and an irreversible oxidation process in the range +0.8 V potential region. The binuclear complexes showed quasireversible two-step single electron reduction processes around the −1.3 and −1.7 V potential regions. The anodic potential region showed an irreversible oxidation process at +1.0 V. The heterobinuclear ZnIINiII complex showed an irreversible reduction of the NiII species at −1.55 V. The catalytic hydrolysis towards 4-nitrophenyl phosphate by the mononuclear, binuclear NiII complexes and the heterobinuclear complex were found to be appreciable. The pseudo-first order rate constant for the catalytic hydrolysis catalyzed by the binuclear and heterobinuclear complexes were found to be higher (9.8 × 10−4 s−1) than that of the corresponding mononuclear complexes (1.3 × 10−5 s−1), which ascertain the requirement of two metal ions in close proximity for the binding of the nucleophilic OH and the phosphate group.

Hydrolysis of 4-nitrophenyl phosphate by magnetite or fly ash: A model for the fate

Reaction of fly ash or magnetite towards hydrolysis of 4-nitrophenyl phosphate (NPP) was investigated in both the solid and solution phases at 10-3 M concentration in oil-in-water microemulsions and aqueous media. The reactions were monitored by measuring the absorbance of the nitrophenolate ion produced in the reaction aliquots with time. The possible application of the above systems for environmental decontamination of organophosphates is discussed. The study provides the basis for increasing our knowledge of the fate of persistent organophosphates in the soil and provides a possible application to environmental clean-up problems. (Bulletin of The Chemical Society of Ethiopia: 2002 16 (2): 133-140)

Selective Hydrolysis of 4-Nitrophenyl Phosphate by a Dinuclear Cu(II) Complex

Abstract A flexible dinuclear Cu(II) complex with 1,3-bis(1,4,7-triaza-1-cyclononyl)propane, hydrolyzes 4-nitrophenyl phosphate under mild conditions, with 300 times rate enhancement over the corresponding mononuclear Cu(II) complex. In contrast, there is no difference in the reactivity in hydrolyzing bis(4-nitrophenyl)phosphate.

Catalytic hydrolysis of phosphate monoesters by lanthanide(III) cryptate (2.2.1) complexes

The hydrolysis of 4-nitrophenyl phosphate is catalysed by lanthanide(III)(2.2.1) complexes, and kinetic studies reveal that the europium(III)(2.2.1) chloride (c= 4.0 mmol dm–3) hydrolyses the phosphate monoester at an enhanced rate (kp1= 1.7 × 10–4 s–1 at pH 7.5 and 50 °C), with a turnover number of 4; the pH dependence of the rate constants is well correlated with the concentration of the lanthanide (2.2.1) hydroxides.

Hydrolysis of 4-Nitrophenyl Phosphate by (Amino Acidato)zinc Complexes

Simple zinc complexes of glycine, histidine, and cysteine are proposed as evolutionary models for hydrolytic enzymes. Catalytic activity of (amino acidato)zinc complexes has been studied towards hydrolysis of 4-nitrophenyl phosphate at 40 °C over the pH range 5.0—10.0 by monitoring release of p-nitrophenol at 405 nm. Detailed kinetic investigations on hydrolysis of ester catalyzed by zinc complexes are discussed through formation of (amino acidato)zinc-substrate complex. Mechanistic and evolutionary aspect of this study are discussed in relation to the esterase property of the zinc metalloenzymes.

Human placental alkaline phosphatase. An improved purification procedure and kinetic studies.

An improved method for the purification of human placental alkaline phosphatase is described. The partially purified enzyme from Sigma was further purified by successive Concanavalin A-Sepharose and Q-Sepharose chromatography. The whole procedure may be completed in one working day. Highly purified enzyme was obtained with a 39% yield. The intrinsic fluorescence of the enzyme decreased at elevated temperature. The conformation of the enzyme molecule was studied by the fluorescence quenching technique. Upward Stern-Volmer plots were obtained for the quenching data which suggested that, in addition to collisional quenching, static quenching was involved in the quenching mechanism. The dynamic and static quenching constants were found to be 0.7 +/- 0.16 M-1 and 0.44 +/- 0.1 M-1, respectively, using acrylamide as the quenching agent. The corresponding values were 0.43 +/- 0.23 M-1 and 0.84 +/- 0.18 M-1, respectively, with KI as the quenching agent. Mg2+ and PO4(3-) induced protein conformational changes which altered both the dynamic and static quenching constants. Mg2+ was found to be a non-essential activator for the placental alkaline-phosphatase-catalyzed hydrolysis of 4-nitrophenyl phosphate. At pH 9.8, Mg2+ increased Vmax by 1.2-fold without affecting the Kd of the substrate. The tetranitromethane-modified enzyme showed slower migration toward the anode on electrophoresis and increased Kd for Mg2+.

Separation and properties of the haemolysins and extracellular enzymes of Listeria monocytogenes and L. ivanovii

Desalted ammonium-sulphate (0-65%) precipitates from the cell-free supernates of 16-24-h cultures of Listeria monocytogenes Boldy and L. ivanovii (previously L. monocytogenes) Type 5 were eluted through Sephadex G-200. The enzyme activities gave rise to two main peaks. The first peak (approximate mol. wt of protein 150,000) contained only phosphatase activity (assayed by hydrolysis of 4-nitrophenylphosphate at pH 5.0 and 7.0). The second peak (approximate mol. wts of proteins 40,000-60,000) contained the haemolysin activity and the following hydrolytic activities (assay substrates are given in parentheses): phospholipase C (phosphatidyl choline and 4-nitrophenyl-phosphoryl-choline); phosphodiesterase (bis-4-nitrophenyl-phosphate); acid phosphatase (4-nitrophenylphosphatase); and esterases and lipases (4-nitrophenyl acetate, naphthyl-acetate and -oleate, triacetin and triolein). DEAE-Sephadex chromatography of appropriate fractions from the Sephadex G-200 purification step separated the first peak into two phosphatases and resolved the second peak into its constituent activities. Polyacrylamide gel electrophoresis showed that the individual fractions from the DEAE-Sephadex step consisted of mixtures of protein. The effects of pH and potential activators and inhibitors on the active proteins purified by DEAE-Sephadex chromatography were examined.

Zirconium-hydroxide-mediated alkaline hydrolysis of phosphate esters

Under mildly alkaline conditions (pH 7–9) the hydrolysis of phosphate monoesters such as 4-nitrophenyl phosphate, glucose 1-phosphate, and glycerol 1-phosphate proceeds slowly, if at all. However, in the presence of gelatinous zirconium hydroxide the rate of hydrolysis of 4-nitrophenyl phosphate is increased by a factor of ca. 30, although the rates of hydrolysis of a range of phosphate esters and related compounds such as adenosine mono- and tri-phosphates, nitrophenyl glycosides, and nitrophenyl sulphates and acetates are not altered. The effects of pH, zirconium hydroxide concentration, phosphate ion, and zirconium phosphate upon this catalysis are described. Titanium(IV) hydroxide exhibited only a slight catalytic effect. A mechanism for the selective catalytic reaction of the zirconium hydroxide is suggested.