635-78-9

Basic information

• Product Name: RESORUFIN
• Synonyms: RESORUFIN;7-HYDROXY-3H-PHENOXAZIN-3-ONE;7-HYDROXY-3-ISO-PHENOXAZIN-3-ONE;LABOTEST-BB LT00847597;7-hydroxy-3h-phenoxazin-3-on;resorufine;RESORUFIN (DYE CONTENT 95%);Resorufin, pract.
• CAS NO: 635-78-9
• Molecular Formula: C₁₂H₇NO₃
• Molecular Weight: 213.19
• EINECS: 211-241-8
• Product Categories: Isolabel;Fluorescent;Heterocyclic;Oxazone
• Mol File: 635-78-9.mol
• Article Data: 97

Chemical Properties

• Melting Point: >300 °C(lit.)
• Boiling Point: 412.967 °C at 760 mmHg
• Flash Point: 203.555 °C
• Appearance: Purple/powder
• Density: 1.46 g/cm³
• Storage Temp.: ?20°C
• Solubility: Solubility Sparingly soluble in water; soluble in ethanol, N,N-d
• PKA: 6.6(at 25℃)
• BRN: 174850
• CAS DataBase Reference: RESORUFIN(CAS DataBase Reference)
• NIST Chemistry Reference: RESORUFIN(635-78-9)
• EPA Substance Registry System: RESORUFIN(635-78-9)

Safety Data

• Hazard Codes: Xi
• Statements: 36/37/38
• Safety Statements: 26-37
• WGK Germany: 3
• RTECS: SP7698000
• Hazardous Substances Data: 635-78-9(Hazardous Substances Data)

Usage

Uses

Resorufin is a pink fluorescent dye which has been used as a marker for microfilaments in the leading lamella of moving cells.

Purification Methods

Wash it with water and recrystallise it several times from EtOH. [Beilstein 27 II 108, 27 III/IV 2263.]

InChI:InChI=1/C12H7NO3/c14-7-1-3-9-11(5-7)16-12-6-8(15)2-4-10(12)13-9/h1-6,14H

Upstream product

• 74683-21-9 3-hydroxy-2'-nitrodiphenyl ether 
• 34994-50-8 7-hydroxy-3H-phenoxazin-3-one sodium salt 
• 921-01-7 rac-cysteine 
• 70-18-8 glutathion 
• 62758-13-8 resazurin sodium salt 
• 109055-80-3 lacmoid, reduced product 
• 13066-95-0 4-aminoresorcinol 
• 108-95-2 phenol 
• 698-31-7 4-nitroso-1,3-benzenediol 
• 123-30-8 4-amino-phenol 
• 108-46-3 recorcinol 

Downstream Products

• 5725-89-3 7-methoxyresorufin 
• 136565-96-3 resorufinyl β-D-galactopyranoside 

Relevant articles and documents

Building three-dimensional nanostructures with active enzymes by surface templated layer-by-layer assembly

We show that well-defined three-dimensional nanostructures of functional enzymes can be controllably fabricated by layer-by-layer assembly of avidin and biotinylated horseradish peroxidase on micro-contact printing patterned surface templates. The Royal S

Increased potassium conductance of brain mitochondria induces resistance to permeability transition by enhancing matrix volume

Modulation of K+ conductance of the inner mitochondrial membrane has been proposed to mediate preconditioning in ischemia-reperfusion injury. The mechanism is not entirely understood, but it has been linked to a decreased activation of mitochondrial permeability transition (mPT). In the present study K+ channel activity was mimicked by picomolar concentrations of valinomycin. Isolated brain mitochondria were exposed to continuous infusions of calcium. Monitoring of extramitochondrial Ca2+ and mitochondrial respiration provided a quantitative assay for mPT sensitivity by determining calcium retention capacity (CRC). Valinomycin and cyclophilin D inhibition separately and additively increased CRC. Comparable degrees of respiratory uncoupling induced by increased K+ or H+ conductance had opposite effects on mPT sensitivity. Protonophores dose-dependently decreased CRC, demonstrating that so-called mild uncoupling was not beneficial per se. The putative mitoKATP channel opener diazoxide did not mimic the effect of valinomycin. An alkaline matrix pH was required for mitochondria to retain calcium, but increased K+ conductance did not result in augmented ΔpH. The beneficial effect of valinomycin on CRC was not mediated by H2O2-induced protein kinase C∈ activation. Rather, increased K+ conductance reduced H2O2 generation during calcium infusion. Lowering the osmolarity of the buffer induced an increase in mitochondrial volume and improved CRC similar to valinomycin without inducing uncoupling or otherwise affecting respiration. We propose that increased potassium conductance in brain mitochondria may cause a direct physiological effect on matrix volume inducing resistance to pathological calcium challenges.

Cu2+-Modified Metal–Organic Framework Nanoparticles: A Peroxidase-Mimicking Nanoenzyme

The synthesis and characterization of UiO-type metal–organic framework nanoparticles (NMOFs) composed of Zr4+ ions bridged by 2,2′-bipyridine-5,5′-dicarboxylic acid ligands and the postmodification of the NMOFs with Cu2+ ions are described. The resulting Cu2+-modified NMOFs, Cu2+-NMOFs, exhibit peroxidase-like catalytic activities reflected by the catalyzed oxidation of Amplex-Red to the fluorescent Resorufin by H2O2, the catalyzed oxidation of dopamine to aminochrome by H2O2, and the catalyzed generation of chemiluminescence in the presence of luminol/H2O2. Also, the Cu2+-NMOFs mimic NADH peroxidase functions and catalyze the oxidation of dihydronicotinamide adenine dinucleotide, NADH, to nicotinamide adenine dinucleotide, NAD+, in the presence of H2O2. The Cu2+-NMOFs-catalyzed generation of chemiluminescence in the presence of luminol/H2O2 is used to develop a glucose sensor by monitoring the H2O2 formed by the aerobic oxidation of glucose to gluconic acid in the presence of glucose oxidase. Furthermore, loading the Cu2+-NMOFs with fluorescein and activating the catalyzed generation of chemiluminescence in the presence of luminol/H2O2 yield an efficient chemiluminescence resonance energy transfer (CRET) process to the fluorescein reflected by the activation of the fluorescence of the dye (λ = 520 nm, CRET efficiency 35%).

A 3,7-Dihydroxyphenoxazine-based Fluorescent Probe for Selective Detection of Intracellular Hydrogen Peroxide

A novel N-borylbenzyloxycarbonyl-3,7-dihydroxyphenoxazine fluorescent probe (NBCD) for detecting H2O2 in living cells is described. The probe could achieve high selectivity for detecting H2O2 over other biological reactive oxygen species (ROS). In addition, upon addition of H2O2, NBCD exhibited color change from colorless to pink, which makes it a "naked-eye" probe for H2O2 detection. NBCD could not only be used to detect enzymatically generated H2O2 but also to detect H2O2 in living systems by using fluorescence spectroscopy, with a detection limit of 2 μm. Importantly, NBCD enabled the visualization of epidermal growth factor (EGF)-stimulated H2O2 generation inside the cells.

Development of an improved Amplex Red peroxidation activity assay for screening cytochrome P450 variants and identification of a novel mutant of the thermophilic CYP119

Abstract: Biocatalysts are increasingly utilized in the synthesis of drugs and agrochemicals as an alternative to chemical catalysis. They are preferred in the synthesis of enantiopure products due to their high regioselectivity and enantioselectivity. Cytochrome P450 (P450) oxygenases are valuable biocatalysts, since they catalyze the oxidation of carbon–hydrogen bonds with high efficiency and selectivity. However, practical use of P450s is limited due to their need for expensive cofactors and electron transport partners. P450s can employ hydrogen peroxide (H2O2) as an oxygen and electron donor, but the reaction with H2O2 is inefficient. The development of P450s that can use H2O2 will expand their applications. Here, an assay that utilizes Amplex Red peroxidation, to rapidly screen H2O2-dependent activity of P450 mutants in cell lysate was developed. This assay was employed to identify mutants of CYP119, a thermophilic P450 from Sulfolobus acidocaldarius, with increased peroxidation activity. A mutant library of CYP119 containing substitutions in the heme active site was constructed via combinatorial active-site saturation test and screened for improved activity. Screening of 158 colonies led to five mutants with higher activity. Among improved variants, T213R/T214I was characterized. T213R/T214I exhibited fivefold higher kcat for Amplex Red peroxidation and twofold higher kcat for styrene epoxidation. T213R/T214I showed higher stability towards heme degradation by H2O2. While the Km for H2O2 and styrene were not altered by the mutation, a fourfold decrease in the affinity for another substrate, lauric acid, was observed. In conclusion, Amplex Red peroxidation screening of CYP119 mutants yielded enzymes with increased peroxide-dependent activity. Graphic abstract: [Figure not available: see fulltext.].

Covalent hemin-DNA adducts for generating a novel class of artificial heme enzymes

(Chemical Equation Presented) Well-defined conjugates between DNA and heme enzymes are accessible by the reconstitution of apo-enzymes with covalent hemin-DNA adducts (see picture). This concept allows access to novel redox catalysts with programmable binding properties that may be applied as biocatalysts, sensors, and biomaterials.

Heterolytic reduction of fatty acid hydroperoxides by cytochrome c/cardiolipin complexes: Antioxidant function in mitochondria

(Chemical Equation Presented) Cytochrome c (cyt c), a mitochondrial intermembrane electron shuttle between complexes III and IV, can, upon binding with an anionic phospholipid, cardiolipin (CL), act as a peroxidase that catalyzes cardiolipin oxidation. H

Monitoring of Heparin Activity in Live Rats Using Metal-Organic Framework Nanosheets as Peroxidase Mimics

Metal-organic framework (MOF) nanosheets are a class of two-dimensional (2D) porous and crystalline materials that hold promise for catalysis and biodetection. Although 2D MOF nanosheets have been utilized for in vitro assays, ways of engineering them into diagnostic tools for live animals are much less explored. In this work, a series of MOF nanosheets are successfully engineered into a highly sensitive and selective diagnostic platform for in vivo monitoring of heparin (Hep) activity. The iron-porphyrin derivative is selected as a ligand to synthesize a series of archetypical MOF nanosheets with intrinsic heme-like catalytic sites, mimicking peroxidase. Hep-specific AG73 peptides as recognition motifs are physically adsorbed onto MOF nanosheets, blocking active sites from nonspecific substrate-catalyst interaction. Because of the highly specific interaction between Hep and AG73, the activity of AG73-MOF nanosheets is restored upon the binding of Hep, but not Hep analogues and other endogenous biomolecules. Furthermore, by taking advantages of biocompatibility and diagnostic property enabled by AG73-MOF nanosheets, the elimination process of Hep in live rats is quantitatively monitored by coupling with microdialysis technology. This work expands the biomedical applications of 2D MOF nanomaterials and provides access to a promising in vivo diagnostic platform.

Single Molecule Investigation of Nanoconfinement Hydrophobicity in Heterogeneous Catalysis

Nanoconfinement imposes physical constraints and chemical effects on reactivity in nanoporous catalyst systems. In the present study, we lay the groundwork for quantitative single-molecule measurements of the effects of chemical environment on heterogeneous catalysis in nanoconfinement. Choosing hydrophobicity as an exemplary chemical environmental factor, we compared a range of essential parameters for an oxidation reaction on platinum nanoparticles (NPs) confined in hydrophilic and hydrophobic nanopores. Single-molecule experimental measurements at the single particle level showed higher catalytic activity, stronger adsorption strength, and higher activation energy in hydrophobic nanopores than those in hydrophilic nanopores. Interestingly, different dissociation kinetic behaviors of the product molecules in the two types of nanopores were deduced from the single-molecule imaging data.

Major involvement of rabbit liver cytochrome P4501A in thiabendazole 5-hydroxylation

1. Thiabendazole is a widely used food preservative and anthelmintic drug for breeding animal species. In order to characterize precisely the cytochrome P450 isozyme(s) involved in its major route of metabolism, a rapid and sensitive spectrofluorimetric method was developed for the simultaneous determination of thiabendazole and its main hepatic metabolite 5-hydroxythiabendazole. 2. The kinetics of thiabendazole 5-hydroxylation were determined in microsomal preparations from control rabbits or animals previously treated with either β-naphthoflavone, isosafrole, phenobarbital, rifampicin or clofibrate. These treatments led to specific induction of CYP1A1, 1A2, 2B4, 3A6 and 4A1 respectively. 3. By considering this panel of characterised microsomal preparations, only those obtained from BNF-treated rabbits exhibited an increase in thiabendazole 5-hydroxylase activity. Ethoxyresorufin O-deethylation in these microsomes was solely inhibited by thiabendazole. These argue for a specific involvement of the CYP1A, subfamily. 4. In the CYP1A subfamily, CYP1A2 appears to be responsible for basal 5-hydroxylation and further unidentified metabolism of thiabendazole in control livers. However, the major involvement of CYP1A1 is supported by the following characteristics of 5-hydroxylation of thiabendazole: (1) the correlation with CYP1A1 expression and (2) the inhibition by ellipticine and not by furafylline, inhibitors of CYP1A1 and CYP1A2 respectively. 5. All these data demonstrated that the rabbit cytochrome P4501A is predominantly involved in thiabendazole 5-hydroxylation which has been suspected to be critical in terms of safety of the parent drug.

Antioxidant capacity of two novel bioactive Fe(III)-cyclophane complexes

The cyclophanes 2,9,25,32-tetraoxo-4,7,27,30-tetrakis(carboxymethyl)-1,4,7, 10, 24,27,30,33-octaaza-17,40-dioxa[10.1.10.1]paracyclophane (PO) and 2,9,25,32-tetraoxo- 4,7,27,30-tetrakis(carboxymethyl)-1,4,7,10,24,27,30,33- octaaza[10.1.10.1]paracyclophane

Measurement of enzyme kinetics using a continuous-flow microfluidic system

This paper describes a microanalytical method for determining enzyme kinetics using a continuous-flow microfluidic system. The analysis is carried out by immobilizing the enzyme on microbeads, packing the microbeads into a chip-based microreactor (volume ～1.0 nL), and flowing the substrate over the packed bed. Data were analyzed using the Lilly-Hornby equation and compared to values obtained from conventional measurements based on the Michaelis-Menten equation. The two different enzyme-catalyzed reactions studied were chosen so that the substrate would be nonfluorescent and the product fluorescent. The first reaction involved the horseradish peroxidase-catalyzed reaction between hydrogen peroxide and N-acetyl-3,7-dihydroxyphenoxazine (amplex red) to yield fluorescent resorufin, and the second the β-galactosidase-catalyzed reaction of nonfluorescent resorufin-β-D-galactopyranoside to yield D-galactose and fluorescent resorufin. In both cases. the microfluidics-based method yielded the same result obtained from the standard Michaelis-Menten treatment. The continuous-flow method required ～10μL of substrate solution and 109 enzyme molecules. This approach provides a new means for rapid determination of enzyme kinetics in microfluidic systems, which may be useful for clinical diagnostics, and drug discovery and screening.

Inhibition of human cytochrome P450 enzymes by 1,2-dithiole-3-thione, oltipraz and its derivatives, and sulforaphane

Recent studies have demonstrated that two chemoprotective agents, oltipraz (OPZ), a synthetic derivative of the natural compound 1,2-dithiole- 3-thione (D3T), and sulforaphane (SF), an isothiocyanate, are not only inducers of glutathione S-transferases but also inhibitors of some major cytochrome P450 enzymes (P450s) involved in xenobiotic metabolism. We examined P450 inhibition by the two compounds and compared two OPZ metabolites (OPZ M3 and M8) and D3T using human P450s expressed in Escherichia coli membranes. OPZ was a more potent inhibitor than D3T or SF, in the following order of inhibition: P450 1A2 > 3A4 > 1A1 ~ 1B1 > 2E1. OPZ M3 also inhibited P450s 1A2, 1A1, 1B1, and 3A4 but not more effectively than OPZ. OPZ Ms was not inhibitory. OPZ behaved as a competitive inhibitor of P450 1A2, with a K(i) of 1.5 μM. Incubation of P450 1A2 with OPZ and NADPH led to a first-order loss of the P450 spectrum, and the loss was not blocked by glutathione. No such time-dependent loss of P450 was seen with P450 1A2 and D3T, P450 1A2 and OPZ M3, P450 1A2 and SF, P450 3A4 and OPZ, P450 3A4 and D3T, P450 2E1 and OPZ, or P450 2E1 and D3T. The time- and concentration- dependent loss of P450 1A2 activity in the presence of OPZ was characterized with a K(i) of 9 μM and a k(inactivation) of 0.19 min-1. The activation of 2-amino-3,5- dimethylimidazo[4,5-f]quinoline (MeIQ) in an E. coli lac-based mutagenicity tester system containing functional human P450 1A2 was inhibited by OPZ (IC50 1 μM) but not appreciably by 40 μM D3T. Our results indicate that OPZ is a competitive and mechanism-based inhibitor of P450 1A2, and the extent of this inhibition is significantly greater than that of other chemoprotective chemicals with P450 1A2 or other human P450s.

Facet Effects of Ag3PO4 on Charge-Carrier Dynamics: Trade-Off Between Photocatalytic Activity and Charge-Carrier Lifetime

Silver phosphate (Ag3PO4) is a promising visible-light-driven photocatalyst with a strong oxidation power and exceptionally high apparent quantum yield of O2 evolution. Although engineering Ag3PO4 facets is widely known to enhance its photocatalytic activity, most studies have explained its facet effect by calculating surface energies. Herein, the charge carrier dynamics in three kinds of Ag3PO4 crystals (mixed facets, cubic, and tetrahedral structures) were first investigated using single-particle photoluminescence microscopy and femtosecond time-resolved diffuse reflectance spectroscopy. As a result, we clarified that the disagreement between the photocatalytic activities (dye degradation and O2 evolution) of different Ag3PO4 facets are the consequence of trade-off between catalytic activity and lifetime of photogenerated charge carriers in addition to surface energy.

Hydrogel-Immobilized Coacervate Droplets as Modular Microreactor Assemblies

Immobilization of compartmentalized microscale objects in 3D hydrogels provides a step towards the modular assembly of soft functional materials with tunable architectures and distributed functionalities. Herein, we report the use of a combination of micro-compartmentalization, immobilization, and modularization to fabricate and assemble hydrogel-based microreactor assemblies comprising millions of functionalized polysaccharide–polynucleotide coacervate droplets. The heterogeneous hydrogels can be structurally fused by interfacial crosslinking and coupled as input and output modules to implement a UV-induced photocatalytic/peroxidation nanoparticle/DNAzyme reaction cascade that generates a spatiotemporal fluorescence read-out depending on the droplet number density, intensity of photoenergization, and chemical flux. Our approach offers a route to heterogeneous hydrogels with endogenous reactivity and reconfigurable architecture, and provides a step towards the development of soft modular materials with programmable functionality.

Biologically relevant chemical properties of peroxymonophosphate ({double bond, long}O3POOH)

It has been suggested that peroxymonophosphate could serve as an endogenous hydrogen peroxide-derived regulator of cellular protein tyrosine phosphatase activity under physiological or pathophysiological conditions. To facilitate further consideration of

The excited-state interaction of resazurin and resorufin with amines in aqueous solutions. Photophysics and photochemical reactions.

The photophysics and photochemical behavior of the phenoxazin-3-one dyes, resazurin and resorufin, have been studied in aqueous solutions. The irradiation of resazurin in the presence of amines leads to deoxygenation of the N-oxide group, giving resorufin. This photoreaction is highly dependent on the amine structure and is efficient only in the presence of tertiary aliphatic amines. The absorption and fluorescence properties of these dyes are dependent on pH. At pH above 7.5 both dyes are in their anionic form. For resorufin this form is highly fluorescent (phiF = 0.75). At lower pH the fluorescence is strongly reduced. The N-oxide dye presents a very weak fluorescence quantum yield (0.11), which also is reduced at low pH. Flash photolysis experiments allowed characterization of the triplet state and the transients formed after irradiation of these dyes in the absence and presence of amines. The triplet quantum yields are 0.08 for resazurin and 0.04 for resorufin. The photodeoxygenation of N-oxide in the presence of amines occurs from the triplet state.

A GGCT fluorogenic probe: Design, synthesis and application to cancer-related cells

Cancer-related γ-glutamyl cyclotransferase (GGCT) specifically converts γ-glutamyl amino acids (γ-Glu-Xaa) into pyroglutamate and the corresponding amino acids (Xaa). Here we report a novel GGCT fluorogenic probe LISA-101 containing a masked O-acylated fluorophore resorufin on the side chain of the P′1 amino acid (Xaa). Upon GGCT treatment, the P′1 amino acid was liberated and spontaneously released the intact fluorophore. Thus, the fluorescence was regained. LISA-101 will expand the strategies for cancer studies.

Multiplex analysis of enzyme kinetics and inhibition by droplet microfluidics using picoinjectors

Enzyme kinetics and inhibition is important for a wide range of disciplines including pharmacology, medicine and industrial bioprocess technology. We present a novel microdroplet-based device for extensive characterization of the reaction kinetics of enzyme substrate inhibitor systems in a single experiment utilizing an integrated droplet picoinjector for bioanalysis. This device enables the scanning of multiple fluorescently-barcoded inhibitor concentrations and substrate conditions in a single, highly time-resolved experiment yielding the Michaelis constant (Km), the turnover number (kcat) and the enzyme inhibitor dissociation constants (ki, k i′). Using this device we determine Km and k cat for β-galactosidase and the fluorogenic substrate Resorufin β-d-galactopyranoside (RBG) to be 442 μM and 1070 s-1, respectively. Furthermore, we examine the inhibitory effects of isopropyl-β-d-thiogalactopyranoside (IPTG) on β-galactosidase. This system has a number of potential applications, for example it could be used to screen inhibitors to pharmaceutically relevant enzymes and to characterize engineered enzyme variants for biofuels production, in both cases acquiring detailed information about the enzyme catalysis and enzyme inhibitor interaction at high throughput and low cost.

PVP stabilized Pt nano particles catalyzed de-oxygenation of phenoxazine group by hydrazine in physiological buffer media: Surfactant competes with reactants for the same surface sites

PVP capped platinum nano particles (PNP) of 5 nm diameter were prepared and characterized as homogeneous and of spherical nature. At physiological pH range (6.0-8.0), these PNP catalyze the deoxygenation of phenoxazine group containing resazurin (1) by hydrazine. The observed rate constants (ko), increase linearly with [PNP] at constant [1] and [Hydrazine]; but first increase and then after reaching a maximum it decrease with increase in [1] as well as in [Hydrazine]. The ko values increase linearly with 1/[H +] indicating N2H4 as the reducing species that generates from the PNP assisted deprotonation of N2H 5+. The kinetic observations suggest Langmuir-Hinshelwood type surface reaction mechanism where both 1 and hydrazine are adsorbed on nano particles surface and compete for the same sites. Interestingly, the surfactant molecules, polyvinylpyrrolidone (PVP), though do not take part into reduction reaction but having same type of functional groups as reactants, competes with them for the same surface sites. Adsorption on PNP with same type of functional group is further supported by the FTIR spectra of Pt-PVP and Pt-1. Thus on increasing [PVP], ko decreases linearly and only when [PVP] is held constant, the plot of kovs. [PNP] passes through the origin indicating the insignificance of uncatalyzed reaction. The plot of ln k ovs. [1] or [Hydrazine] shows two different linear zones with different exponent values with respect to [1] and [Hydrazine]. This indicates that along with the complex heterogeneous surface adsorption processes, the mutual interactions between the reactants are also changing with the relative concentrations of reactants or, in general, with the molar ratio ([Hydrazine]/[1]).

Modification of the heme active site to increase the peroxidase activity of thermophilic cytochrome P450: A rational approach

The site specific mutants of the thermophilic P450 (P450 175A1 or CYP175A1) were designed to introduce residues that could act as acid-base catalysts near the active site to enhance the peroxidases activity. The Leu80 in the distal heme pocket of CYP175A1 was located at a position almost equivalent to the Glu183 that is involved in stabilization of the ferryl heme intermediate in chloroperoxidase (CPO). The Leu80 residue of CYP175A1 was mutated with histidine (L80H) and glutamine (L80Q) that could potentially form hydrogen bond with hydrogen peroxide and facilitate formation and stabilization of the putative redox intermediate of the peroxidase cycle. The mutants L80H and L80Q of CYP175A1 showed higher peroxidase activity compared to that of the wild type (WT) CYP175A1 enzyme at 25°C. The activity constants (kcat) for the L80H and L80Q mutants of CYP175A1 were higher than those of myoglobin and wild type cytochrome b562 at 25°C. The optimum temperature for the peroxidase activity of the WT and mutants of CYP175A1 was ~70°C. The rate of catalysis at temperatures above ~70°C was higher for L80Q mutant of CYP175A1 compared to that of the well known natural peroxidase, horseradish peroxidase (HRP) that denatures at such high temperature. The peroxidase activities of the mutants of CYP175A1 were maximum at pH 9, unlike that of HRP which is at pH ~5. The results have been discussed in the light of understanding the structure-function relationship of the peroxidase properties of these thermostable heme proteins.

Multifunctional linker for orthogonal decoration of gold nanoparticles with DNA and protein

A novel trifunctional linker was prepared to allow multiple orthogonal functionalization of gold nanoparticles (AuNPs) with biomolecules. Thiooctic groups contained within the linker act as AuNP surface anchors, while two additional orthogonal functional groups are used for attachment of heme and DNA using amide and copper catalyzed Huisgen cycloaddition, respectively. We demonstrate that heme can act as a platform for reconstitution of a functional heme protein such as myoglobin. Using multifunctional linker, fully functional Au-DNA-Mb hybrids were obtained and such multifunctional constructs expand the synthetic toolbox for nanomaterial tailoring and design of biosensors and novel catalytic materials.

Selective fluorometric detection of aromatic thiols by a chemosensor containing two electrophilic sites with different local softness

A resorufin-dinitrophenyl ether conjugate (1) shows emission enhancement for aromatic thiols in aqueous media with a neutral-basic pH, while being nonemissive for aliphatic thiols. This is achieved by two electrophilic sites with different local softness on compound 1; the respective sites selectively react with aromatic or aliphatic thiolate anions.

Hydrogen peroxide supports human and rat cytochrome P450 1A2-catalyzed 2-amino-3-methylimidazo[4,5-f]quinoline bioactivation to mutagenic metabolites: Significance of cytochrome P450 peroxygenase

We show that the naturally occurring hydroperoxide hydrogen peroxide is highly effective in supporting the cytochrome P450 1A2 peroxygenase-catalyzed metabolic activation of the heterocyclic aromatic amine 2-amino-3- methylimidazo[4,5-f]quinoline (IQ) to genetoxic metabolites. Mutagenicity was assessed by the Ames assay with Salmonella typhimurium strain YG1012 and an activation system consisting of hydroperoxides plus either 3- methylcholanthrene-induced rat liver microsomes (rP4501A) or human P450 1A2- containing microsomes (hP4501A2). The mutagenic response was dependent on the concentration of microsomal protein, IQ, and hydroperoxides. The addition of hydrogen peroxide or tert-butyl hydroperoxide to rP4501A greatly enhanced the yield of histidine prototrophic (His+) revertants. This increase was inhibited, in a concentration-dependent manner, by α-naphthoflavone, a P450 1A inhibitor. Hydrogen peroxide was the most effective peroxygenase cofactor, particularly with hP4501A2 (K(m) =0.1 μM). The hydroperoxide-supported activation of IQ produced reactive intermediates which bound to 2'- deoxyguanosine; LC/MS analysis of the adducts revealed the same major (protonated) adduct at m/z = 464.4 as previously reported for the DNA adduct formed (in vivo or in vitro) by the mixed function-catalyzed bioactivation system. None of the peroxidase-catalyzed IQ metabolites (nitro-, azo-, or azoxy-IQ) were detected. In conclusion, hydrogen peroxide in the physiological/pathological concentration range may be able to support the metabolic activation of arylamines to genotoxic products through the cytochrome P450 peroxygenase pathway.

Silicalite-1/glass fibre substrates for enhancing the photocatalytic activity of TiO2

Silicalite-1 (S1) coatings were prepared on silica wool substrates by hydrothermal synthesis and subsequently immersed into a Ti-containing sol at a steady rate of 30 cm min-1. The material was annealed in a furnace at 90 ?°C for 2 h and 550 ?°C for 2 h to create a silica fibre core surrounded by concentric layers of silicalite-1 and TiO2. The resulting samples were characterised by X-ray diffraction (XRD), Raman spectroscopy, scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS) and Brunauer-Emmett-Teller (BET) surface area measurement. The photocatalytic activity of the samples was evaluated using the intelligent ink test and during degradation of stearic acid under UVA light (?? = 365 nm). The new coated-fibres were shown to be substantially better photocatalysts than comparable TiO2 coatings on plain glass fibres. The TiO2/S1/glass fibres have potential use in air/water cleaning applications.

Synthetic Silica Nano-Organelles for Regulation of Cascade Reactions in Multi-Compartmentalized Systems

In eukaryotic cells, enzymes are compartmentalized into specific organelles so that different reactions and processes can be performed efficiently and with a high degree of control. In this work, we show that these features can be artificially emulated in

A specifically triggered turn-on fluorescent probe platform and its visual imaging of HClO in cells, arthritis and tumors

Understanding disease-related processes at the molecular level is of great importance for the prevention and treatment of diseases. However, due to the lack of effective analytical tools, it is challenging to gain insight into the relationships between a specific bioactive molecule and the associated disease. Herein, a rapid turn-on resorufin-based fluorescent probe platform utilizing the HClO-specific oxidative cleavage of the amide was constructed, allowing the visualization of HClO in vitro and in vivo. These probes could quickly respond to HClO ( 50 s) with high selectivity and sensitivity (12–153 nM). The probe REClO-6 had the fastest response (30 s) and the highest sensitivity (12 nM), and was successfully used for the imaging of endogenous and exogenous HClO in cells and zebrafish. Notably, it was also successfully applied to the imaging of HClO in mouse arthritis and solid tumors. This study provided a rapid imaging analysis tool, which would be used to investigate the relationship between HClO and the disease-related physiological processes.