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P8250 Sigma

Peroxidase from horseradish

Type II, essentially salt-free, lyophilized powder, 150-250 units/mg solid (using pyrogallol)

Synonym: Donor:hydrogen-peroxide oxidoreductase, Horseradish peroxidase

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Description

Analysis Note

The RZ (Reinheitszahl) is the absorbance ratio A403/A275 determined at 0.5-1.0 mg/ml in deionized water. It is a measure of hemin content, not enzymatic activity. Even preparations with high RZ may have low enzymatic activity.

Other Notes

View more information on peroxidase at www.sigma-aldrich.com/enzymeexplorer.

Packaging

5000, 25000, 50000, 100000, 200000 units in glass bottle

Quality

Preliminary work shows it to contain at least five isoenzymes.

Unit Definition

One pyrogallol unit will form 1.0 mg purpurogallin from pyrogallol in 20 sec at pH 6.0 at 20 °C.

Application

Horseradish peroxidase (HRP) is isolated from horseradish roots (Amoracia rusticana). It is used in biochemistry applications such as western blots, ELISA and Immunohistochemistry. Horseradish peroxidase is used to amplify a weak signal and increase detectability of a target molecule, such as a protein. Horseradish peroxidase, product P8250, has been used to study nonoral antigens in inflamed gingiva and Ebola virus glycoprotein toxicity.

The enzyme has been used as a comparison during the peroxidase assay of extract from mature tall fescue leaf blades. It has also been used to measure H2O2 production.

Biochem/physiol Actions

HRP readily combines with hydrogen peroxide (H2O2) and the resultant [HRP-H2O2] complex can oxidize a wide variety of hydrogen donors. The optimal pH is 6.0-6.5 and the enzyme is most stable in the pH range of 5.0-9.0. HRP can be conjugated to antibodies by several different methods including glutaraldehyde, periodate oxidation, through disulfide bonds, and also via amino and thiol directed cross-linkers. It is smaller and more stable than the enzyme labels β-galactosidase and alkaline phosphatase. Hence, it is the most desired label. Also, its glycosylation leads to lower non-specific binding. It is also used for the determination of glucose and peroxides in solution. Sodium azide, cyanide, L-cystine, dichromate, ethylenethiourea, hydroxylamine, sulfide, vanadate, p-aminobenzoic acid, and Cd2+, Co2+, Cu2+, Fe3+, Mn2+, Ni2+, Pb2+ ions are found to inhibit the enzyme activity.

When incubated with a substrate, horseradish peroxidase produces a coloured, fluorimetric, or luminescent derivative of the labeled molecule, allowing quantification. Horseradish peroxidase has been shown to slightly reduce the level of inhibition in a cydAB mutant.

General description

Horseradish peroxidase is isolated from horseradish roots (Amoracia rusticana) and belongs to the ferroprotoporphyrin group of peroxidases. HRP is a single chain polypeptide containing four disulfide bridges. It is a glycoprotein containing 18% carbohydrate. The carbohydrate composition consists of galactose, arabinose, xylose, fucose, mannose, mannosamine, and galactosamine depending upon the specific isozyme. Its molecular weight (~44 kDa) includes the polypeptide chain (33,890 Daltons), hemin plus Ca2+ (~700 Daltons), and carbohydrate (~9,400 Daltons). At least seven isozymes of HRP exist. The isoelectric point for horseradish peroxidase isozymes ranges from 3.0 - 9.0.

Price and Availability

Suggested Laboratory Gloves


Laboratory GlovesThis substance has been tested against several types of hand protection for CE compliance. Click below to find the recommended gloves for handling this product.




All labs need water
Safety & Documentation

Safety Information

RIDADR 
NONH for all modes of transport
WGK Germany 
3

Documents

Certificate of Analysis

Certificate of Origin

Protocols & Articles

Protocols

Enzymatic Assay of Alcohol Oxidase (EC 1.1.3.13)

The continuous spectrophotometric rate determination (A405, Light path = 1 cm) is based on the following reactions:
Keywords: Extinction coefficient

Enzymatic Assay of Cholesterol Oxidase

To standardize a procedure for the determination of the enzymatic assay of Cholesterol Oxidase.
Keywords: Enzyme activity, Extinction coefficient, Isomerizations

Enzymatic Assay of Diamine Oxidase (E.C. No. 1.4.3.6)

Replacing OP SPPUTR01. New document number assigned to conform to current document numbering system.
Keywords: Carcinogens, Extinction coefficient

Enzymatic Assay of Lipase Type XIII from Pseudomonas Species and Pseudomonas Cepacia using a coupled enzyme system of Glycerol Kinase and Glycerophosphate Oxidase (EC 3.1.1.3)

To standardize a procedure for the enzymatic assay of Lipase Type XIII from Pseudomonas Species, Sigma Product Number L9518 , and Pseudomonas Cepacia, Sigma Product Number L9156, using a coupled enzy...
Keywords: Centrifugation, Extinction coefficient, High performance liquid chromatography

Enzymatic Assay of Peroxidase (EC 1.11.1.7)

This procedure is for the determination of Peroxidase enzymatic activity using Pyrogallol as the substrate. Products using this method include, but are not limited to, P1709, P2088, P6140, P6782, P81...
Keywords: Enzyme activity, Extinction coefficient

Spectrophotometric Determination of Reinheitszahl (RZ) for Peroxidase (EC 1.11.1.7)

Reinheitszahl (RZ) is the ratio of absorbance due to hemin (A403, Soret region) to absorbance due to protein (A275). This ratio (A403/A275) is not a measure of enzyme activity. Products using this me...
Keywords: Enzyme activity

Related Content

Enzymes & Proteins

Application Index | Enzyme Index | Substrate Index | Inhibitor Index | Cofactor Index | Lectin Index
Keywords: Cell signaling, Diagnostic, Drug discovery, Molecular biology

Peer-Reviewed Papers
15

References

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