scholarly journals Immobilized Horseradish Peroxidase on Discs of Polyvinyl Alcohol-Glutaraldehyde Coated with Polyaniline

2012 ◽  
Vol 2012 ◽  
pp. 1-8 ◽  
Author(s):  
Samantha Salomão Caramori ◽  
Kátia Flávia Fernandes ◽  
Luiz Bezerra de Carvalho Junior
Keyword(s):  
Horseradish Peroxidase ◽  
Polyvinyl Alcohol ◽  
Soluble Enzyme ◽  
Initial Activity

Discs of network polyvinyl alcohol-glutaraldehyde (PVAG) were synthesized and coated with polyaniline (PANI) using glutaraldehyde as a chemical arm (PVAG-PANIG-HRP disc). The best conditions for the immobilization were established as about 1.0 mg mL−1of protein, for 60 min and pH 5.5. The soluble enzyme lost all of its activity after incubation at 70°C for 15 min, whereas the PVAG-PANIG-HRP disc retained about half of the initial activity for pyrogallol. The same PVAG-PANIG-HRP disc was used consecutively three times without any activity lossbut presented 25% of the initial activity after the 7th use. PVAG-PANIG-HRP disc retained approximately 80% and 60% of its initial activity after 60 and 80 days of storage, respectively. Resorcinol, m-cresol, catechol, pyrogallol,α-naphthol,βnaphthol, and 4, 4′-diaminodiphenyl benzidine were efficiently oxidized by the PVAG-PANIG-HRP disc (from about 70% to 90%), and it was less efficient towards aniline, phenol, and 2-nitrosonaphthol.

Competitive enzyme-liked immunoassay with use of soluble enzyme/antibody immune complexes for labeling. I. Measurement of human choriogonadotropin.

1976 ◽  
Vol 22 (8) ◽  
pp. 1372-1377 ◽  
Author(s):  
D E Yorde ◽  
E A Sasse ◽  
T Y Wang ◽  
R O Hussa ◽  
J C Garancis
Keyword(s):  
Horseradish Peroxidase ◽  
Peroxidase Activity ◽  
Quantitative Measurement ◽  
Original Sample ◽  
Soluble Enzyme ◽  
Soluble Complex ◽  
Human Choriogonadotropin ◽  
Preliminary Study ◽  
Free Antigen ◽  
Serum And Urine

Abstract We described the principle of a new enzyme-immunoassay, competitive enzyme-liked immunoassay (CELIA), for quantitative measurement of soluble antigens and haptens. In the assay, binding of antibody to antigen-immunosorbent is competitively inhibited by the free antigen to be measured. The amount of first antibody bound to the immunosorbent is measured by an enzymatic technique in which a heterologous bridging antibody and a soluble antibody/enzyme immune complex are applied in sequence. The soluble complex we used was rabbit antiperoxidase/horseradish peroxidase. Peroxidase activity is inversely proportional to the concentration in the original sample of the substance to be assayed. The enzyme-linked reagents are potentially widely applicable to any substance to be measured. To demonstrate the feasibility of CELIA, we report a preliminary study of its application to the measurement of human chloriogonadotropin in serum and urine. The assay described for this hormone has a working range of 1 to 50 int. units per milliliter of sample. The technique obviates the disadvantages associated with measurement and handling of radioisotopes in radioimmunoassays and the only major instrumentation required is a centrifuge and a conventional spectrophotometer.

scholarly journals Preparation of Stable Cross-Linked Enzyme Aggregates (CLEAs) of a Ureibacillus thermosphaericus Esterase for Application in Malathion Removal from Wastewater

Catalysts ◽  
2018 ◽  
Vol 8 (4) ◽  
pp. 154 ◽  
Author(s):  
Yuliya Samoylova ◽  
Ksenia Sorokina ◽  
Alexander Piligaev ◽  
Valentin Parmon
Keyword(s):  
Response Surface Methodology ◽  
Bovine Serum Albumin ◽  
High Stability ◽  
Municipal Wastewater ◽  
Maximal Activity ◽  
Cross Linking ◽  
Soluble Enzyme ◽  
Initial Activity ◽  
E Coli ◽  
Ureibacillus Thermosphaericus

In this study, the active and stable cross-linked enzyme aggregates (CLEAs) of the thermostable esterase estUT1 of the bacterium Ureibacillus thermosphaericus were prepared for application in malathion removal from municipal wastewater. Co-expression of esterase with an E. coli chaperone team (KJE, ClpB, and ELS) increased the activity of the soluble enzyme fraction up to 200.7 ± 15.5 U mg−1. Response surface methodology (RSM) was used to optimize the preparation of the CLEA-estUT1 biocatalyst to maximize its activity and minimize enzyme loss. CLEA-estUT1 with the highest activity of 29.4 ± 0.5 U mg−1 (90.6 ± 2.7% of the recovered activity) was prepared with 65.1% (w/v) ammonium sulfate, 120.6 mM glutaraldehyde, and 0.2 mM bovine serum albumin at 5.1 h of cross-linking. The biocatalyst has maximal activity at 80 °С and pH 8.0. Analysis of the properties of CLEA-estUT1 and free enzyme at 50–80 °C and pH 5.0–10.0 showed higher stability of the biocatalyst. CLEA-estUT1 showed marked tolerance against a number of chemicals and high operational stability and activity in the reaction of malathion hydrolysis in wastewater (up to 99.5 ± 1.4%). After 25 cycles of malathion hydrolysis at 37 °С, it retained 55.2 ± 1.1% of the initial activity. The high stability and reusability of CLEA-estUT1 make it applicable for the degradation of insecticides.

scholarly journals Expression and Characterization of a Dye-Decolorizing Peroxidase from Pseudomonas Fluorescens Pf0-1

Catalysts ◽  
2019 ◽  
Vol 9 (5) ◽  
pp. 463 ◽  
Author(s):  
Nikola Lončar ◽  
Natalija Drašković ◽  
Nataša Božić ◽  
Elvira Romero ◽  
Stefan Simić ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Pseudomonas Fluorescens ◽  
Alginate Beads ◽  
Sequence Information ◽  
Soluble Enzyme ◽  
Initial Activity ◽  
Alternative Substrate ◽  
65 H ◽  
Genome Sequence Information

The consumption of dyes is increasing worldwide in line with the increase of population and demand for clothes and other colored products. However, the efficiency of dyeing processes is still poor and results in large amounts of colored effluents. It is desired to develop a portfolio of enzymes which can be used for the treatment of colored wastewaters. Herein, we used genome sequence information to discover a dye-decolorizing peroxidase (DyP) from Pseudomonas fluorescens Pf-01. Two genes putatively encoding for DyPs were identified in the respective genome and cloned for expression in Escherichia coli, of which one (PfDyP B2) could be overexpressed as a soluble protein. PfDyP B2 shows some typical features known for DyPs which includes the ability to convert dyes at the expense of hydrogen peroxide. Interestingly, t-butyl hydroperoxide could be used as an alternative substrate to hydrogen peroxide. Immobilization of PfDyP B2 in calcium-alginate beads resulted in a significant increase in stability: PfDyP B2 retains 80% of its initial activity after 2 h incubation at 50 °C, while the soluble enzyme is inactivated within minutes. PfDyP B2 was also tested with aniline and ethyl diazoacetate as substrates. Based on GC-MS analyses, 30% conversion of the starting material was achieved after 65 h at 30 °C. Importantly, this is the first report of a DyP-catalyzed insertion of a carbene into an N-H bond.

Electrospun polyvinyl alcohol/bovine serum albumin biocomposite membranes for horseradish peroxidase immobilization

2016 ◽  
Vol 93-94 ◽  
pp. 1-10 ◽  
Author(s):  
Ramin Fazel ◽  
Seyed-Fakhreddin Torabi ◽  
Pooya Naseri-Nosar ◽  
Salehe Ghasempur ◽  
Seyed-Omid Ranaei-Siadat ◽  
...  
Keyword(s):  
Bovine Serum Albumin ◽  
Horseradish Peroxidase ◽  
Polyvinyl Alcohol ◽  
Serum Albumin ◽  
Bovine Serum

scholarly journals Immobilization of Peroxidase onto Magnetite Modified Polyaniline

2012 ◽  
Vol 2012 ◽  
pp. 1-5 ◽  
Author(s):  
Eduardo Fernandes Barbosa ◽  
Fernando Javier Molina ◽  
Flavio Marques Lopes ◽  
Pedro Antonio García-Ruíz ◽  
Samantha Salomão Caramori ◽  
...  
Keyword(s):  
Horseradish Peroxidase ◽  
Initial Activity ◽  
Active Protein ◽  
Optimum Ph ◽  
Repeated Use

The present study describes the immobilization of horseradish peroxidase (HRP) on magnetite-modified polyaniline (PANImG) activated with glutaraldehyde. After the optimization of the methodology, the immobilization of HRP on PANImG produced the same yield (25%) obtained for PANIG with an efficiency of 100% (active protein). The optimum pH for immobilization was displaced by the effect of the partition of protons produced in the microenvironment by the magnetite. The tests of repeated use have shown that PANImG-HRP can be used for 13 cycles with maintenance of 50% of the initial activity.

scholarly journals Removal of Persistent Sulfamethoxazole and Carbamazepine from Water by Horseradish Peroxidase Encapsulated into Poly(vinyl chloride) Electrospun Fibers

2021 ◽  
Vol 23 (1) ◽  
pp. 272
Author(s):  
Jakub Zdarta ◽  
Oliwia Degórska ◽  
Katarzyna Jankowska ◽  
Agnieszka Rybarczyk ◽  
Adam Piasecki ◽  
...  
Keyword(s):  
Horseradish Peroxidase ◽  
Vinyl Chloride ◽  
Immobilized Enzymes ◽  
Stability Study ◽  
Process Conditions ◽  
Electrospun Fibers ◽  
Substrate Affinity ◽  
Initial Activity ◽  
Poly Vinyl Chloride ◽  
Negative Effect

Enzymatic conversion of pharmaceutically active ingredients (API), using immobilized enzymes should be considered as a promising industrial tool due to improved reusability and stability of the biocatalysts at harsh process conditions. Therefore, in this study horseradish peroxidase was immobilized into sodium alginate capsules and then trapped into poly(vinyl chloride) electrospun fibers to provide additional enzyme stabilization and protection against the negative effect of harsh process conditions. Due to encapsulation immobilization, 100% of immobilization yield was achieved leading to loading of 25 μg of enzyme in 1 mg of the support. Immobilized in such a way, enzyme showed over 80% activity retention. Further, only slight changes in kinetic parameters of free (Km = 1.54 mM) and immobilized horseradish peroxidase (Km = 1.83 mM) were noticed, indicating retention of high catalytic properties and high substrate affinity by encapsulated biocatalyst. Encapsulated horseradish peroxidase was tested in biodegradation of two frequently occurring in wastewater API, sulfamethoxazole (antibiotic) and carbamazepine (anticonvulsant). Over 80% of both pharmaceutics was removed by immobilized enzyme after 24 h of the process from the solution at a concentration of 1 mg/L, under optimal conditions, which were found to be pH 7, temperature 25 °C and 2 mM of H2O2. However, even from 10 mg/L solutions, it was possible to remove over 40% of both pharmaceuticals. Finally, the reusability and storage stability study of immobilized horseradish peroxidase showed retention of over 60% of initial activity after 20 days of storage at 4 °C and after 10 repeated catalytic cycles, indicating great practical application potential. By contrast, the free enzyme showed less than 20% of its initial activity after 20 days of storage and exhibited no recycling potential.

scholarly journals Horseradish Peroxidase-Carrying Electrospun Nonwoven Fabrics for the Treatment of o-Methoxyphenol

2015 ◽  
Vol 2015 ◽  
pp. 1-6
Author(s):  
Chao Pan ◽  
Ran Ding ◽  
Li Dong ◽  
Jing Wang ◽  
Yucai Hu
Keyword(s):  
Horseradish Peroxidase ◽  
Storage Stability ◽  
Storage Period ◽  
Industrial Applications ◽  
Initial Activity ◽  
Nonwoven Fabrics ◽  
Base Stability ◽  
Phenolic Wastewater ◽  
20 Nm ◽  
And Storage

The carboxyl-functionalized polystyrene (poly(styrene-co-methacrylic acid), PSMAA) nanofibers with average diameters of 250 ± 20 nm was prepared by electrospinning. PSMAA nanofibrous membrane were employed for immobilization of horseradish peroxidase (HRP) enzyme on the fibrous surface by a chemical method. The parameters about immobilizing HRP on the PSMAA nanofibers were studied and the influence on the activity of the HRP is discussed. This study showed that soap-free emulsion method is an ideal technology to modify the polystyrene surface and ultimately achieve enzyme immobilization on electrospun PSMAA nanofibers surfaces. Compared with free HRP, the acid-base stability, thermal stability, and storage stability of HRP were increased after the immobilization. The immobilized HRP maintained about 60% of its initial activity during a 20-day storage period. However, the free HRP maintained only 40% of its initial activity. The removal percentages of o-methoxyphenol (OMP) reached 80.2% after 120 min for immobilized HRP. These results suggest that the proposed scheme for immobilization of HRP has potential in industrial applications for the treatment of phenolic wastewater.

scholarly journals Stabilization of b-Glucuronidase by Immobilization in Magnetic-Silica Hybrid Supports

Catalysts ◽  
2020 ◽  
Vol 10 (6) ◽  
pp. 669
Author(s):  
Sonali Correa ◽  
Magdalena Ripoll ◽  
Erienne Jackson ◽  
Valeria Grazú ◽  
Lorena Betancor
Keyword(s):  
Residual Activity ◽  
Covalent Attachment ◽  
Hydrodynamic Diameter ◽  
Individual Effect ◽  
Soluble Enzyme ◽  
Initial Activity ◽  
Magnetic Silica ◽  
Low Probability ◽  
The Individual ◽  
Silica Hybrid

β-Glucuronidases are a class of enzymes that catalyze the breakdown of complex carbohydrates. They have well documented biocatalytic applications in synthesis, therapeutics, and analytics that could benefit from enzyme immobilization and stabilization. In this work, we have explored a number of immobilization strategies for Patella vulgata β-Glucuronidase that comprised a tailored combination of biomimetic silica (Si) and magnetic nanoparticles (MNPs). The individual effect of each material on the enzyme upon immobilization was first tested. Three different immobilization strategies for covalent attachment on MNPs and different three catalysts for the deposition of Si particles were tested. We produced nine different immobilized preparations and only two of them presented negligible activity. All the preparations were in the micro-sized range (from 1299 ± 52 nm to 2101 ± 67 nm of hydrodynamic diameter). Their values for polydispersity index varied around 0.3, indicating homogeneous populations of particles with low probability of agglomeration. Storage, thermal, and operational stability were superior for the enzyme immobilized in the composite material. At 80 °C different preparations with Si and MNPs retained 40% of their initial activity after 6 h of incubation whereas the soluble enzyme lost 90% of its initial activity within 11 min. Integration of MNPs provided the advantage of reusing the biocatalyst via magnetic separation up to six times with residual activity. The hybrid material produced herein demonstrated its versatility and robustness as a support for β-Glucuronidases immobilization.

Sign in / Sign up

Export Citation Format

Share Document