Hydrolysis Of 4-nitrophenyl Acetate Research Articles

Engineering the Hydrophobic Microenvironment in Polystyrene-Supported Artificial Catalytic Triad Nanocatalysts: An Effective Strategy for Improving Catalytic Performance

Hydrophobic environments have been identified as one of the main parameters affecting the catalytic performance of artificial catalytic triads but are often ignored as an approach to engineering these catalysts. Here, we have developed a simple yet powerful strategy to engineer the hydrophobic environment in polystyrene-supported artificial catalytic triad (PSACT) nanocatalysts. Hydrophobic copolymers containing either oligo(ethylene glycol) side chains or hydrocarbon side chains were synthesized and used for the preparation of nanocatalysts through nanoprecipitation in aqueous media. By using the hydrolysis of 4-nitrophenyl acetate (4NA) as a model reaction, we studied the influence of chemical structures and effective constituent ratios of hydrophobic copolymers on the catalytic performance of PSACT nanocatalysts. Additionally, PSACT nanocatalysts could catalyze the hydrolysis of a few carboxylic esters, even polymers, and be reused for five consecutive runs without significant loss of catalytic activity. This strategy may open an avenue for engineering other artificial enzymes, and these PSACT nanocatalysts have potential applications for the hydrolysis of carboxylic esters.

A photo-modulated organometallic nanozyme based on β-cyclodextrin-capped gold nanoparticles for the detection of Zn(II) and cascade catalysis

Organometallic nanozymes, organometallics of which mimic the cofactor of natural enzymes, have the advantages of tunable catalytic activity, high stability, and material variety. By assembling organometallics (catalytic site) on inorganic nanoparticles (support), organometallic nanozymes have been rationally designed, but modulating their assembly and activity remains a tough challenge. Here, we construct a photo-modulated organometallic nanozyme by anchoring azobenzene-modified triethylenetetramine (Azo-N4) to the surface of β-cyclodextrin-capped gold nanoparticles (AuNP@CDs) through host–guest interaction, followed by Zn(II) coordination to form organometallics. The catalytic site derives from the outer organometallic layer. The nanozyme catalyzes the hydrolysis of 4-nitrophenyl acetate to 4-nitrophenol and the catalytic signal is used to detect Zn(II). Due to the photoisomerization of azobenzene, the supramolecular assembly of host–guest interaction is modulated by light. The disassembly leads to cascade catalysis of AuNP@CDs-Azo-N4-Zn and the recovery of AuNP@CDs. Our study provides a new idea and reference for the rational design and activity modulation of organometallic nanozymes.

Automated monitoring the kinetics of homogeneous and heterogeneous chemical processes using a smartphone

Heterogeneous chemical processes occupy a pivotal position in many fields of applied chemistry. Monitoring reaction kinetics in such heterogeneous systems together with challenges associated with ex-situ analytical methodologies can lead to inaccurate information about the nature of the catalyst surfaces as well as information about the steps involved. The present work explores the possibility of kinetic measurements of chemical reactions and adsorption processes of homogeneous and heterogeneous systems through the variation of RGB intensities of digital images using a smartphone combined with a program written in Python to accelerate and facilitate data acquisition. In order to validate the method proposed, the base promoted hydrolysis of 4-nitrophenyl acetate was initially investigated. The rate constants obtained through RGB analysis (0.01854 min−1) is almost identical to that using traditional UV–Vis spectroscopy (0.01848 min−1). The proposed method was then applied to monitor the kinetics of three heterogeneous processes: (1) reduction of 4-nitrophenolate in the presence of dispersed Pd/C; (2) decomposition of methyl orange with TiO2; and (3) adsorption of rhodamine on montmorillonite. In general, the method via digital images showed high reproducibility and analytical frequency, allowing the execution of simultaneous analyses, with an accuracy comparable to UV–Vis spectrophotometry. The method developed herein is a practical and valuable alternative for obtaining kinetic data of heterogeneous reactions and processes where a color change is involved, bypassing sampling collection and processing which decreases analytical frequency and may lead to data errors.

Evaluation of the Impact of Esterases and Lipases from the Circulatory System against Substrates of Different Lipophilicity.

Esterases and lipases can process amphiphilic esters used as drugs and prodrugs and impact their pharmacokinetics and biodistribution. These hydrolases can also process ester components of drug delivery systems (DDSs), thus triggering DDSs destabilization with premature cargo release. In this study we tested and optimized assays that allowed us to quantify and compare individual esterase contributions to the degradation of substrates of increased lipophilicity and to establish limitations in terms of substrates that can be processed by a specific esterase/lipase. We have studied the impact of carbonic anhydrase; phospholipases A1, A2, C and D; lipoprotein lipase; and standard lipase on the hydrolysis of 4-nitrophenyl acetate, 4-nitrophenyl palmitate, DGGR and POPC liposomes, drawing structure–property relationships. We found that the enzymatic activity of these proteins was highly dependent on the lipophilicity of the substrate used to assess them, as expected. The activity observed for classical esterases was diminished when lipophilicity of the substrate increased, while activity observed for lipases generally increased, following the interfacial activation model, and was highly dependent on the type of lipase and its structure. The assays developed allowed us to determine the most sensitive methods for quantifying enzymatic activity against substrates of particular types and lipophilicity.

Inside or outside the box? Effect of substrate location on coordination-cage based catalysis.

In this work we compare and contrast the hydrolysis of two different aromatic esters using an octanuclear cubic Co8 coordination cage host as the catalyst. Diacetyl fluorescein (DAF) is too large to bind inside the cage cavity, but in aqueous solution it interacts with the exterior surface of the cage via a hydrophobic interaction with K = 1.5(2) × 104 M−1. This is sufficient to bring it into close proximity to the layer of hydroxide ions which also surrounds the 16+ cage surface even at modest pH values, accelerating the hydrolysis of DAF to fluorescein with kcat/kuncat (the rate acceleration for that fraction of DAF in contact with the cage surface in the equilibrium) ≈50. This is far smaller than many known examples of catalysis inside a cage cavity, but at the exterior surface it is potentially general with no cavity-imposed size/shape limitations for guest binding. In contrast 4-nitrophenyl acetate (4NPA) binds inside the cage cavity with K = 3.5(3) × 103 M−1 and as such is surrounded in solution by the hydroxide ions which accumulate around the cage surface. However its hydrolysis is actually inhibited: either because of a geometrically unfavourable geometry of the bound substrate which makes it inaccessible to surface-bound hydroxide, or because the necessary volume expansion/geometry change associated with formation of a tetrahedral intermediate cannot be accommodated inside the cavity. Any 4NPA that is free in solution as part of the equilibrium undergoes catalysed hydrolysis at the cage exterior surface in the same way as DAF, but the effect is limited by the low affinity of 4NPA for the exterior surface. We conclude that exterior-surface catalysis can be effective and potentially general; and that cavity-binding of guests can result in negative, rather than positive, catalysis.

Peptide-anchored gold nanoparticles with bicatalytic sites for photo-switchable cascade catalysis

Due to the difference in multiple catalytic sites of cascade catalysts, it is still a challenge to accurately adjust different catalytic sites to adapt to each step of the catalytic process in the cascade reaction. Here, we report a peptide = anchored gold nanoparticle cascade catalyst self-assembled by host–guest (cyclodextrin–azobenzene) interaction. The catalyst can complete photo-switchable cascade catalysis in one pot. The self-assembled peptides can be used as an esterase mimic to catalyze the hydrolysis of 4-nitrophenyl acetate to 4-nitrophenol. Under UV irradiation, the self-assembled peptides disassemble and gold nanoparticles then catalyze the reduction of 4-nitrophenol to 4-aminophenol. This study provides a new way to design catalytically active peptide–gold nanoparticle conjugates as cascade catalysts.

Cofactor-free organic nanozyme with assembly-induced catalysis and light-regulated activity

Organic nanozymes that mimic cofactor-free enzymes avoid the use of metal ions and other cofactors, and show great advantages in the field of biomimetic catalysis. The assembly of organics such as peptides is usually necessary for their catalysis and is also a major challenge for organic nanozymes. Here we constructed cofactor-free organic nanozyme by self-assembly of cyclodextrins modified gold nanoparticle and azobenzene modified peptide chain through host–guest interaction. At room temperature, free peptide chain has negligible catalytic activity. Organic nanozyme reduces the activation energy and increases the reaction rate of hydrolysis of 4-nitrophenyl acetate. The catalysis of organic nanozyme derives from the ordered assembly of peptide chains on the surface of support and the deprotonation of the imidazole groups of neighboring histidines that facilitates the splitting of a proton from water, and thus promotes the nucleophilic attack on the incoming substrate. Photoisomerization of azobenzene group leads to the assembly and disassembly of nanozyme, causing the change of catalytic activity. These findings may contribute to the design of advanced biomimetic catalysts and provide a model for cofactor-free nanozymes.

Heterochiral tetrapeptide self-assembly into hydrogel biomaterials for hydrolase mimicry.

Self-assembling short peptides have attracted great interest as enzyme mimics, especially if the catalytic activity resides solely in the supramolecular structure so that it can be switched on/off as needed by controlling assembly/disassembly. Among the various enzyme classes, hydrolases find wide application in biomaterials, and their mimetics often contain His residues, in addition to either divalent cations or other amino acids to mimic the catalytic site. This work reports two self-assembling tetrapeptides based on the Ser-His motif for catalysis and the Phe-Phe motif to drive amyloid structure formation. Both peptides form thermoreversible hydrogels in phosphate buffer at neutral pH that display a mild esterase-like activity, as demonstrated on the hydrolysis of 4-nitrophenyl acetate as a model substrate, although presence of Ser did not enhance catalytic activity. The systems are characterised by circular dichroism, transmission electron microscopy, oscillatory rheology and Thioflavin T fluorescence as an amyloid stain, to provide further insights that may assist the future design of improved supramolecular catalysts.

Peptide–Gold Nanoparticle Conjugates as Artificial Carbonic Anhydrase Mimics

We herein describe the design and synthesis of a catalytically active peptide–gold nanoparticle conjugate (Pep-Au-NP) that binds Zn(II) within its peptide monolayer and develops carbonic anhydrase activity. Specifically, a modified variant of the β-sheet forming IHIHIQI-peptide (IHQ), which forms an interstrand 3-His Zn(II)-binding site, was used as a ligand for spherical gold nanoparticles (Au-NPs). The resulting immobilized peptide maintains its ability to form β-sheets, as determined by circular dichroism (CD)-spectroscopy and, thus, maintains its ability to form Zn(II)-binding sites. The addition of Zn(II)-ions to the peptide–gold nanoparticle conjugates (Au@IHQ-NP) resulted in significant improvements in rates of ester hydrolysis of 4-nitrophenyl acetate (4-NPA) and the hydration of CO2 compared to the unconjugated peptide variants. Recycling of the catalyst revealed that Au@IHQ-NP remains intact with at least 94% of its initial activity after five rounds of CO2 hydration. The herein reported results reveal that Pep-Au-NPs are able to perform reactions catalyzed by natural metalloenzymes and open up new possibilities for the implementation of these conjugates.

Metalloenzyme Mimicry at the Nodes of Metal-Organic Frameworks

In this issue of Chem, Dincă and co-workers show that the node of the metal-organic framework MFU-4l can be modified by anion exchange to serve as a biomimetic model of carbonic anhydrase given the similarity in their ligand fields. The resulting material, which has reactivity characteristics of the enzyme, gives great promise for CO2-related applications.

Construction of self-assembled vesicle nanoenzyme using cucurbit[8]uril-based supra-amphiphiles

Self-assembled vesicles are constructed based on supra-amphiphiles formed by cucurbit[8]uril and its two guest molecules. By modifying one of the guest molecules, an artificial nanohydrolase is constructed. An enhancement of catalytic activity for hydrolysis of 4-nitrophenyl acetate is observed, which is attributed to the enrichment effect of substrate around catalytic sites through the molecular recognition.

Micro-reactor mixing unit interspacing for fast liquid-liquid reactions leading to a generalized scale-up methodology

The effect of geometrical arrangements of a residence time channel (RTC) positioned in between LL-Rhombus static mixing elements on interphase mass transfer rates is investigated. Four variations of a micro-reactor plate are tested with four immiscible aqueous-organic systems: water coupled with n-butanol, n-hexanol, methyl tert-butyl ether and toluene. The alkaline hydrolysis of 4-nitrophenyl acetate is used to determine the overall volumetric mass transfer coefficient (Kda). For all solvent pairs, the micro-reactor without a RTC has the greatest Kda at a given flow rate due to a greater energy dissipation rate (ɛ) and the fact that drops can re-coalesce leading to reduced advective transfer in the RTC pending the flow regime. An increase in flow rate results in a rise in conversion up to a maximum, after which viscous dissipation impedes further turbulent breakdown of the droplets as they near the Kolmogorov length scale. When the resulting flow is finely dispersed, interphase mass transfer rates no longer depend on reactor geometry but was correlated to the solvent system and rate of energy dissipated. The negative impact of operating at greater ɛ values than at maximum conversion is shown via the required pressure loss to achieve a certain conversion or residence time. Lastly, relevant to production rates in the fine chemical and pharmaceutical industry, an algorithm for scaling a generic mass transfer-limited liquid-liquid reaction at identified optimal operating conditions is presented.

Microreactor Mixing-Unit Design for Fast Liquid-Liquid Reactions

Abstract Based on previous work studying complex microreactors, it was desired to further improve the mixing efficiency by varying the mixing unit design for fast liquid-liquid reactions. Different flow regimes were studied, including slug flow, parallel flow, and drop flow. The two-phase hydrolysis of 4-nitrophenyl acetate in sodium hydroxide solution was used to evaluate the overall volumetric mass transfer coefficients (K org a) as a function of the average rate of energy dissipation (ε) for each microreactor design and all flow regimes. The liquid-liquid systems investigated used n-butanol or toluene as the organic phase solvent and a 0.5-M NaOH aqueous solution. The use of surfactant was also investigated with the toluene- water system. All microreactor geometry designs were based on contraction–expansion repeating units with asymmetric obstacles to aid the breakup of slugs and desynchronize the recombination of split streams. The investigated designs were chosen to avoid the formation of the parallel flow regime, contrary to curvature-based mixing-unit designs. The microreactor design can then be optimized to reduce the ε required to reach drop flow, since K org a has been found to be constant at equal ε for a given solvent system in this flow regime, regardless of the reactor selection. Additionally, the “3/7th” scaleup rule was applied and confirmed with the LL-Triangle mixer. It was found that, for low interfacial-tension systems (i.e., n-butanol-water), the onset of drop flow occurred at a lower ε for the LL-Triangle mixer when compared with the Sickle or LL-Rhombus mixers.

Liquid–liquid flow regimes and mass transfer in various micro-reactors

The flow regimes and mass transfer rates in five complex micro-reactors with different mixing mechanisms were investigated using the two-phase alkaline hydrolysis of 4-nitrophenyl acetate. n-Butanol and toluene were used as organic solvents. Using n-butanol in curvature-based micro-mixers, the flow regime evolved from slug to parallel to drop/dispersed flow with increasing flow rates. In obstacle-based micro-mixers, no parallel flow was observed. Using toluene, no parallel flow was observed for all reactors. The conversion of 4-nitrophenyl acetate was found to be strongly dependent on the flow regime. In slug and parallel flow, the conversion generally decreased with an increase in flow rate whereas it typically increased in drop flow and was constant or slightly decreased in dispersed flow. The different micro-mixers were compared using the overall volumetric mass transfer coefficient, Korga, which was primarily a function of the rate of energy dissipation within the dispersed flow regime. The geometry itself impacts the resulting flow regime and rate of energy dissipation at a given flow rate. The micro-reactors were then compared using modified Damköhler’s numbers. Curvature-based reactors were found to be inadequate for liquid–liquid reactions under the studied conditions, as they favor parallel flow patterns and yield relatively low interphase mass transfer rates.

Liquid–Liquid Mass Transfer in an Oscillatory-Flow Mesoscale Coil Reactor without Baffles

Interphase mass transfer rates of an immiscible liquid–liquid system are investigated in an oscillatory-flow coil reactor without baffles. A baffle-less system is chosen since much research has been completed in baffled tubes with little performance comparison to baffle-less tubes, which offer lower operating and capital costs. In our experiments, interphase mass transfer rates are evaluated via the two-phase alkaline hydrolysis of 4-nitrophenyl acetate (4-NPA) in a toluene–water biphasic system. The mass transfer rate increased 7-fold at the maximum tested oscillation of 5000 oscillatory-flow Reynolds number. The best application for the oscillatory-flow coil reactor is determined using a comparison to other liquid–liquid flow platforms in the presented toolbox approach. The oscillatory-flow coil reactor becomes a clear complement to a plate microreactor for gaining volume.

Liquid–liquid mass transfer in a serpentine micro-reactor using various solvents

The flow regimes are identified and mass transfer rates measured in a serpentine micro-reactor with a hydraulic diameter of 0.7mm using the two-phase alkaline hydrolysis of 4-nitrophenyl acetate between flow rates of 0.8 and 32mL/min. Four different organic solvents (n-butanol, n-hexanol, MTBE and toluene) were used in order to investigate the effect of phase physical properties. The continuous phase overall volumetric mass transfer coefficient, Kca, was greatest in the drop flow regime and for organic solvents with lower interfacial tension resulting in larger interfacial area, a. In the parallel flow regime, Kca values increased linearly with the continuous phase capillary number. In the drop flow regime, two correlations for the prediction of the drop size were developed with fitted parameters falling within the expected values for turbulent break-up of drops in conventional static mixers.

Preparation, crystal structures and properties of zinc complexes containing Bis(oxazoline) ligand

Three zinc complexes ([(DMOX)ZnX2], X = Cl (1), Br (2) or I (3)), where DMOX is 4,5-dihydro-2-(4,5-dihydro-4,4-dimethyloxazol-2-yl)-4,4-dimethyloxazole), have been fully characterized by single crystal X-ray diffraction, UV–vis, NMR and IR spectroscopy. Moreover, all zinc complexes showed activities in catalyzing hydrolysis of 4-nitrophenyl acetate (4-NA). And active species have been determined by pH titrations of [(DMOX)ZnI2] in catalyzing hydrolysis of 4-NA.

Purification and Characterization of Carbonic Anhydrase from Ağrı Balık Lake Trout Gill (Salmo trutta labrax) and Effects of Sulfonamides on Enzyme Activity

Carbonic anhydrase (CA) was purified from Ağrı Balık Lake trout gill (fCA) by affinity chromatography on a sepharose 4B-tyrosine-sulfanilamide column. The fCA enzyme was purified with about a 303.9 purification factor, a specific activity 4130.4 EU (mg-protein)(-1), and a yield of 79.3 by using sepharose-4B-L tyrosine-sulfanilamide affinity gel chromatography. The molecular weight determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was found to be about 29.9 kDa. The kinetic parameters, K(M) and V(max) were determined for the 4-nitrophenyl acetate hydrolysis reaction. Some sulfonamides were tested as inhibitors against the purified CA enzymes. The Ki constants for mafenide (1), p-toluenesulfonamide (2), 2-bromo-benzene sulfonamide (3), 4-chlorobenzene sulfonamide (4), 4-amino-6-chloro-1-3 benzenedisulfonamide (5), sulfamethazine (6), sulfaguanidine (7), sulfadiazine (8), and acetozazolamide (9) were in the range of 7.5-108.75 μM.

Immobilization of lipase from Burkholderia cepacia into calcium carbonate microcapsule and its use for enzymatic reactions in organic and aqueous media

We disclosed a useful procedure for the immobilization of a lipase from Burkholderia cepacia based on its encapsulation into vaterite calcium carbonate microcapsule. The immobilized enzyme had an adequate activity for the hydrolysis of 4-nitrophenyl acetate, which was preserved over 2 years in storage at 4°C. The enantioselective esterification of (±)-1-phenylethanol with vinyl acetate using the immobilized lipase smoothly progressed to afford the corresponding optically active compounds, where the enzyme was easily recovered for reuse. Although the hydrolysis of (±)-2-acetoxyhexyl tosylate in aqueous solution was catalyzed by the enzyme, the phase transition of calcium carbonate from vaterite to poorly porous calcite during the reaction prevented the access of the reactant to the active site of the lipase, resulting in the significant decrease of the activity of the recovered enzyme. However, the treatment of the immobilized enzyme with zinc chloride solution inhibited the phase transition under the reaction conditions. The enzyme after the zinc treatment was recyclable for the hydrolysis of the substrate without significant decrease of the reactivity even after the uses in aqueous media.

Reversible Inhibition of Human Carboxylesterases by Acyl Glucuronides

Carboxylesterases hydrolyze esters, amides, and thioesters to produce carboxylic acids and resulting alcohols, amines, and thiols, respectively. Uridine 5'-diphosphate- glucuronosyltransferases are colocalized with carboxylesterases and have the potential to further metabolize carboxylic acids to acyl glucuronides, but it is currently unknown if acyl glucuronides, being esters, also interact with carboxylesterases. This study explores the ability of acyl glucuronides to act as substrates or inhibitors of human carboxylesterases 1 (hCES1) and 2 (hCES2). The stability of six acyl glucuronides in the presence of hCES1, hCES2, and buffer alone (100 mM potassium phosphate, pH 7.4, 37°C) were investigated. Reversible inhibition of 4-nitrophenyl acetate hydrolysis by the acyl glucuronides was also studied. Diclofenac-β-d-glucuronide was used to explore potential time-dependent inactivation. The chemical stability half-life values for CGP 47292-β-d-glucuronide, diclofenac-β-d-glucuronide, (R)-naproxen-β-d-glucuronide, (S)-naproxen-β-d-glucuronide, ibuprofen-β-d-glucuronide (racemic), clopidogrel-β-d-glucuronide, and valproate-β-d-glucuronide were found to be 0.252, 0.537, 0.996, 1.77, 3.67, 5.02, and 15.2 hours, respectively. Diclofenac-β-d-glucuronide, clopidogrel-β-d-glucuronide, ibuprofen-β-d-glucuronide, (R)-naproxen-β-d-glucuronide, and (S)-naproxen-β-d-glucuronide selectively inhibited hCES1, with Ki values of 4.32 ± 0.47, 24.8 ± 4.2, 355 ± 38, 468 ± 21, 707 ± 64 µM, respectively, but did not significantly inhibit hCES2. Valproate-β-d-glucuronide and CGP 47292-β-d-glucuronide did not inhibit either hCES. Time-dependent inactivation of hCES1 by diclofenac-β-d-glucuronide was not observed. Lastly, both hCES1 and hCES2 were shown not to catalyze the hydrolysis of the acyl glucuronides studied. Drug-drug interaction studies may be warranted for drugs that metabolize to acyl glucuronides due to the potential inhibition of hCESs.