-
Heliyon Jan 2021Kinetic and physicochemical properties of peroxidase purified using a novel and cost efficient protocol was investigated with a view to providing information on its...
Kinetic and physicochemical properties of peroxidase purified using a novel and cost efficient protocol was investigated with a view to providing information on its possible biotechnological potentials. peroxidase was purified to homogeneity in two steps, involving ATPS and size exclusion chromatography on Sephadex G-100 with a yield of 84.12 %. In-gel activity staining revealed the presence of one isoform of peroxidase. The purified peroxidase is monomeric with native and subunits molecular weight of 38.9 and 43.5 kDa respectively. Kinetic parameters - , , of the purified enzyme were 2.5 units/mg protein, 0.020 ± 0.04 mM and 1.37 ± 0.18 mM respectively. Its optimum pH and temperature were 5 and 30 °C respectively. The purified enzyme cross-linked BSA into an insoluble matrix with the aid of caffeic acid. The study concluded that the purification scheme adopted is rapid and efficient, the purified enzyme exhibited some physiochemical properties that make it suitable for various biotechnological applications.
PubMed: 33521366
DOI: 10.1016/j.heliyon.2021.e06032 -
Physiology and Molecular Biology of... Apr 2011The Santalum peroxidase was extracted from the leaves and precipitated with double volume of chilled acetone. The optimum percent relative activity for the Santalum...
The Santalum peroxidase was extracted from the leaves and precipitated with double volume of chilled acetone. The optimum percent relative activity for the Santalum peroxidase was observed at pH 5.0 and 50 °C temperature. The Santalum peroxidase per cent relative activity was stimulated in the presence of phenolic compounds like ferrulic acid and caffeic acids; however, indole-3-acetic acid (IAA) and protocatechuic acid act as inhibitors. All divalent cations Fe(2+), Mn(2+), Mg(2+), Cu(2+) and Zn(2+) stimulate the relative activity of the Santalum peroxidase at concentration of 2.0 μM. Amino acids like L-alanine and L-valine activate the per cent relative activity, while L-proline and DL-methionine showed moderate inhibition for the Santalum peroxidase. However, a very low a concentration of cysteine acts as a strong inhibitor of Santalum peroxidase at the concentration of 0.4 mM. Native polyacrylamide gel electrophoresis (Native-PAGE) was performed for isoenzyme determination and two bands were observed. Km and Vmax values were calculated from Lineweaver-Burk graph. The apparent Vmax/Km value for O-dianisidine and H2O2 were 400 and 5.0 × 105 Units/min/mL respectively.
PubMed: 23573005
DOI: 10.1007/s12298-011-0054-x -
Acta Crystallographica. Section E,... Mar 2018The title compound, (systematic name: ,'-dibenzyl-3,3'-dimeth-oxy-1,1'-biphenyl-4,4'-di-amine), CHNO, was synthesized by the reduction of a Schiff base prepared a...
The title compound, (systematic name: ,'-dibenzyl-3,3'-dimeth-oxy-1,1'-biphenyl-4,4'-di-amine), CHNO, was synthesized by the reduction of a Schiff base prepared a condensation reaction between -dianisidine and benzaldehyde under acidic conditions. The mol-ecule lies on a crystallographic inversion centre so that the asymmetric unit contains one half-mol-ecule. The biphenyl moiety compound is essentially planar. Two intra-molecular N-H⋯O hydrogen bonds occur. The dihedral angle between the terminal phenyl and phenyl-ene rings of a benzidine unit is 48.68 (6)°. The methyl-ene C atom of the benzyl group is disordered over two sets of sites, with occupancy ratio 0.779 (18):0.221 (18). In the crystal, mol-ecules are connected by hydrogen bonding between -dianisidine O atoms and H atoms of the terminal benzyl groups, forming a one-dimensional ladder-like structure. In the data from DFT calculations, the central biphenyl showed a twisted conformation.
PubMed: 29765704
DOI: 10.1107/S2056989018001688 -
Access Microbiology 2019Honey is a natural product with many beneficial properties including antimicrobial action. Production of hydrogen peroxide (HO) in diluted honey is central to this...
Honey is a natural product with many beneficial properties including antimicrobial action. Production of hydrogen peroxide (HO) in diluted honey is central to this action. Here, we describe an optimized method for measuring levels of HO in honey. This method is based on established methods, with the level of dilution, the time between dilution and reading the assay, and aeration of the samples during the assay identified as critical points for ensuring reliability and reproducibility. The method is cost-effective and easy to perform using common laboratory equipment. Using this method, we quantified the hydrogen peroxide content of five different, unprocessed polyfloral honeys collected in NC, USA. Our results show that HO production by these honeys varies greatly, with some samples producing negligible levels of HO. We assessed the effect of colour on the assay by measuring the recovery of spiked HO from light and dark honey and from serially diluted dark corn syrup, and found the amount of HO that could be detected was lower in dark corn syrup and darker honey samples.
PubMed: 32974499
DOI: 10.1099/acmi.0.000065 -
Indoor Air Mar 2021With an increasing use of indoor disinfectants such as chlorine (Cl ) and hypochlorous acid, a convenient sampler for estimating exposure to oxidants, such as effective...
With an increasing use of indoor disinfectants such as chlorine (Cl ) and hypochlorous acid, a convenient sampler for estimating exposure to oxidants, such as effective chlorine, is necessary. Here, we developed a personal passive air sampler (PPAS) composed of a redox dye, o-dianisidine, in a polydimethylsiloxane (PDMS) sheet. o-Dianisidine readily reacts with gaseous oxidants generated by bleach usage, and its color changes as the reaction progresses; hence, personal exposure to effective chlorine could be easily detected by the naked eye, while cumulative exposure could be determined by measuring concentrations of o-dianisidine reacting with it. The PPAS was calibrated, and a sampling rate of 0.00253 m /h was obtained using a small test chamber. The PPAS was tested with the help of ten volunteers whose personal exposure to Cl -equivalent gas was estimated after bathrooms were cleaned using spray and liquid-type household disinfection products, and the accumulated exposure-gas concentrations ranged from 69 to 408 ppbv and 148 to 435 ppbv, respectively. These PPAS-derived exposure concentrations were approximately two orders lower than those estimated using ConsExpo, suggesting a significant overestimation by prevailing screening models, possibly due to the ignorance of transformation reactions.
Topics: Air Pollutants; Air Pollution, Indoor; Chlorine; Dimethylpolysiloxanes; Disinfectants; Disinfection; Environmental Monitoring; Humans; Hypochlorous Acid; Inhalation Exposure
PubMed: 32978992
DOI: 10.1111/ina.12747 -
International Journal of Analytical... 2013One titrimetric and two spectrophotometric methods are described for the determination of ketotifen fumarate (KTF) in bulk drug and in tablets using cerium(IV) as the...
One titrimetric and two spectrophotometric methods are described for the determination of ketotifen fumarate (KTF) in bulk drug and in tablets using cerium(IV) as the oxidimetric agent. In titrimetry (method A), the drug was treated with a measured excess of cerium(IV) in H2SO4 medium and after a standing time of 10 min, the surplus oxidant was determined by back titration with iron(II). The spectrophotometric procedures involve addition of a known excess of cerium(IV) to KTF in acid medium followed by the determination of unreacted oxidant by reacting with either p-dimethyl amino benzaldehyde and measuring the resulting colour at 460 nm (method B) or o-dianisidine and subsequent measurement of the absorbance of coloured product at 470 nm (method C). Titrimetric assay is based on a 1 : 2 reaction stoichiometry between KTF and cerium(IV) and the method is applicable over 2-18 mg range. In spectrophotometry, regression analysis of Beer's law plots showed a good correlation in 0.4-8.0 and 0.4-10.0 g mL(-1) KTF ranges for method B and method C, respectively, and the corresponding molar absorptivity coefficients are calculated to be 4.0 × 10(4) and 3.7 × 10(4) L mol(-1) cm(-1).
PubMed: 24324496
DOI: 10.1155/2013/697651 -
The Journal of Biological Chemistry Dec 2023Prolyl hydroxylase domain (PHD)-containing enzyme 3 (PHD3) belongs to the Caenorhabditis elegans gene egl-9 family of prolyl hydroxylases. PHD3 catalyzes proline...
Prolyl hydroxylase domain (PHD)-containing enzyme 3 (PHD3) belongs to the Caenorhabditis elegans gene egl-9 family of prolyl hydroxylases. PHD3 catalyzes proline hydroxylation of hypoxia-inducible factor α (HIF-α) and promotes HIF-α proteasomal degradation through coordination with the pVHL complex under normoxic conditions. However, the relationship between PHD3 and the hypoxic response is not well understood. In this study, we used quantitative real-time PCR assay and O-dianisidine staining to characterize the hypoxic response in zebrafish deficient in phd3. We found that the hypoxia-responsive genes are upregulated and the number of erythrocytes was increased in phd3-null zebrafish compared with their wild-type siblings. On the other hand, we show overexpression of phd3 suppresses HIF-transcriptional activation. In addition, we demonstrate phd3 promotes polyubiquitination of zebrafish hif-1/2α proteins, leading to their proteasomal degradation. Finally, we found that compared with wild-type zebrafish, phd3-null zebrafish are more resistant to hypoxia treatment. Therefore, we conclude phd3 has a role in hypoxia tolerance. These results highlight the importance of modulation of the hypoxia signaling pathway by phd3 in hypoxia adaptation.
Topics: Animals; Hypoxia-Inducible Factor 1, alpha Subunit; Hypoxia-Inducible Factor-Proline Dioxygenases; Procollagen-Proline Dioxygenase; Proline; Zebrafish; Zebrafish Proteins; Gene Deletion; Oxygen
PubMed: 37923141
DOI: 10.1016/j.jbc.2023.105420 -
Veterinary Sciences Oct 2023Ceruloplasmin (Cp) assessment in biological samples exploits the oxidase activity of this enzyme against several substrates, such as -phenylenediamine (-P), -dianisidine...
Ceruloplasmin (Cp) assessment in biological samples exploits the oxidase activity of this enzyme against several substrates, such as -phenylenediamine (-P), -dianisidine (-D) and, most recently, ammonium iron(II) sulfate (AIS). Once developed in humans, these assays are often used in veterinary medicine without appropriately optimizing in the animal species of interest. In this study, two assays using AIS and -D as substrates have been compared and validated for Cp oxidase activity assessment in horse's plasma. The optimization of the assays was performed mainly by varying the buffer pH as well as the buffer and the substrate molar concentration. Under the best analytical conditions obtained, the horse blood serum samples were treated with sodium azide, a potent Cp inhibitor. In the -D assay, 500 µM sodium azide treatment completely inhibits the enzymatic activity of Cp, whereas, using the AIS assay, a residual analytical signal was still present even at the highest (2000 µM) sodium azide concentration. Even though the analytical values obtained from these methods are well correlated, the enzymatic activity values significantly differ when expressed in Units L. A disagreement between these assays has also been detected with the Bland-Altman plot, showing a progressive discrepancy between methods with increasing analytical values.
PubMed: 37888575
DOI: 10.3390/vetsci10100623 -
Frontiers in Cell and Developmental... 2022Tissue factor (TF) is crucial for embryogenesis, as mice lacking TF are embryonically lethal (E10.5). This lethality may be attributed to defects in vascular development...
Tissue factor (TF) is crucial for embryogenesis, as mice lacking TF are embryonically lethal (E10.5). This lethality may be attributed to defects in vascular development and circulatory failure, suggesting additional roles for TF in embryonic development beyond coagulation. In this study, we characterized the role of one of the TF paralogs () using a zebrafish model. The expression of during embryonic developmental stages was determined by RT-PCR. Spatiotemporal expression pattern of revealed (high expression from 28 to 36 hpf) the role of in the development of the yolk sac, circulation, and fins. Morpholinos (MO), an antisense-based oligonucleotide strategy, was used to knockdown and examined for defects in morphological appearance, bleeding, and vascular patterning. MO-injected embryos showed morphological abnormalities, including shorter body lengths and crooked tails. O-dianisidine staining showed MO-injected embryos exhibited bleeding in the trunk (5.44%) and head (9.52%) regions. Imaging of endothelial-specific transgenic lines () showed a 3-fold decreased caudal vein plexus (CVP) in morphants versus controls at 48 hpf, suggesting a potential role for in angiogenesis. These findings confirm that is essential for angiogenesis, in addition to its involvement in hemostasis.
PubMed: 35386206
DOI: 10.3389/fcell.2022.852989 -
The Science of the Total Environment Nov 2020Surfactants are widely used in the industry of detergents, household products, and cosmetics. SAPDMA is a cationic surfactant that is used mostly in cosmetics,...
Surfactants are widely used in the industry of detergents, household products, and cosmetics. SAPDMA is a cationic surfactant that is used mostly in cosmetics, conditioning agents and has recently gained attention as a corrosion inhibitor in the sea pipelines industry. In this regard, literature concerning the ecotoxicological classification of SAPDMA on aquatic animals is lacking. This study aims to evaluate the potential ecotoxicity of SAPDMA using the aquatic zebrafish embryo model. The potential toxic effects of SAPDMA were assessed by different assays. This includes (i) mortality/survival assay to assess the median lethal concentration (LC); (ii) teratogenicity assay to assess the no observed effect concentration (NOEC); (iii) organ-specific toxicity assays including cardiotoxicity, neurotoxicity (using locomotion assay), hematopoietic toxicity (hemoglobin synthesis using o-dianisidine staining), hepatotoxicity (liver steatosis and yolk retention using Oil Red O (ORO) stain); (iv) cellular cytotoxicity (mitochondrial membrane potential) by measuring the accumulation of JC-1 dye into mitochondria. Exposure of embryos to SAPDMA caused mortality in a dose-dependent manner with a calculated LC of 2.3 mg/L. Thus, based on the LC value and according to the Fish and Wildlife Service (FWS) Acute Toxicity Rating Scale, SAPDMA is classified as "moderately toxic". The No Observed Effect Concentration (NOEC) concerning a set of parameters including scoliosis, changes in body length, yolk, and eye sizes was 0.1 mg/L. At the same NOEC concentration (0.1 mg/L), no organ-specific toxicity was detected in fish treated with SAPDMA, except hepatomegaly with no associated liver dysfunctions. However, higher SAPDMA concentrations (0.8 mg/L) have dramatic effects on zebrafish organ development (eye, heart, and liver development). Our data recommend a re-evaluation of the SAPDMA employment in the industry setting and its strictly monitoring by environmental and public health agencies.
Topics: Animals; Dimethylamines; Embryo, Nonmammalian; Lethal Dose 50; Surface-Active Agents; Water Pollutants, Chemical; Zebrafish
PubMed: 32886985
DOI: 10.1016/j.scitotenv.2020.140450