We test the solubility of this product at 10 mg/mL in a 0.1 M phosphate buffer (pH 6.0). In regards to solution stability of this product, the following information has been listed under Preparation Note on the product's webpage: Solutions of this product at 1 mg/mL in 0.1 M phosphate buffer (pH 6.0) remain active for at least two weeks at room temperature. The solution retains activity after 5 freeze-thaw cycles.
P6782
Peroxidase from horseradish
Type VI-A, essentially salt-free, lyophilized powder, 950-2000 units/mg solid (using ABTS), ≥250 units/mg solid (using pyrogallol)
Synonym(s):
Donor:hydrogen-peroxide oxidoreductase, Horseradish peroxidase
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About This Item
CAS Number:
eCl@ss:
32160410
UNSPSC Code:
12352204
NACRES:
NA.54
EC Number:
232-668-6
MDL number:
Specific activity:
≥250 units/mg solid (using pyrogallol), 950-2000 units/mg solid (using ABTS)
Pricing and availability is not currently available.
type
Type VI-A
Quality Segment
form
essentially salt-free, lyophilized powder
specific activity
≥250 units/mg solid (using pyrogallol), 950-2000 units/mg solid (using ABTS)
mol wt
~44 kDa
solubility
0.1 M phosphate buffer: soluble 10 mg/mL, clear, orange to red (pH 6.0), H2O: soluble
application(s)
diagnostic assay manufacturing
storage temp.
2-8°C
SMILES string
[O+H2]O[O-]
InChI
1S/H2O3/c1-3-2/h1-2H
InChI key
JSPLKZUTYZBBKA-UHFFFAOYSA-N
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.
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. Product P6782 is type VI-A and it is essentially a salt-free, lyophilized powder. It is used to quantify cholesterol and cholesterol esters[1] and to study in vitro lipid deposition on hydrogel and silicone hydrogel contact lenses[2].
The enzyme from Sigma has been used in the development of rapid and sensitive galactose oxidase-peroxidase biosensor for galactose detection with prolonged stability. It has been used as a component of a media (MRS agar) plate containing 3,3′,5,5′-tetramethylbenzidine (TMB) for the growth of Lactobacillus sp. TMB acts as a chromogenic substrate of peroxidase. Peroxidase generates O2 from H2O2 produced by the lactobacilli, and the TMB stains the colonies blue in the presence of O2. Thus, after 48 hours of incubation under 5% CO2 in air, the colonies that produce H2O2 on MRS agar appear dark blue. H2O2 nonproducers are colorless.[3] It has been used to study the stabilization effect of polyvinyl alcohol on horseradish peroxidase. It is also used for the determination of glucose and peroxides in solution.
Biochem/physiol Actions
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.
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 that include the use of glutaraldehyde, periodate oxidation, 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. Sodium azide, cyanide, L-cystine, dichromate, ethylenethiourea, hydroxylamine, sulfide, vanadate, p-aminobenzoic acid, as well as Cd2+, Co2+, Cu2+, Fe3+, Mn2+, Ni2+, and Pb2+ ions are found to inhibit the enzyme activity.
Packaging
Packaged in mg solid
Preparation Note
Solutions of this product at 1 mg/mL in 0.1 M phosphate buffer (pH 6.0) remain active for at least two weeks at room temperature. The solution retains activity after 5 freeze-thaw cycles.
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.
This product is assayed using ABTS for easy comparison to other suppliers′ unit: approx. 1,000 units per mg solid
Other Notes
One ABTS unit will oxidize 1 μmole of ABTS per minute at 25 °C at pH 5.0
One pyrogallol unit will form 1.0 mg purpurogallin from pyrogallol in 20 sec at pH 6.0 at 20 °C.
Similar to P8375
View more information on peroxidase at www.sigma-aldrich.com/enzymeexplorer.
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This Item | |||
|---|---|---|---|
| specific activity ≥250 units/mg solid (using pyrogallol), 950-2000 units/mg solid (using ABTS) | specific activity ≥250 units/mg solid (using pyrogallol) | specific activity 150-250 units/mg solid (using pyrogallol) | specific activity ≥250 units/mg solid (using pyrogallol) |
| form essentially salt-free, lyophilized powder | form essentially salt-free, lyophilized powder | form essentially salt-free, lyophilized powder | form essentially salt-free, lyophilized powder |
| application(s) diagnostic assay manufacturing | application(s) diagnostic assay manufacturing | application(s) diagnostic assay manufacturing | application(s) - |
| mol wt ~44 kDa | mol wt ~44 kDa | mol wt ~44 kDa | mol wt ~44 kDa |
| storage temp. 2-8°C | storage temp. 2-8°C | storage temp. 2-8°C | storage temp. 2-8°C |
| solubility 0.1 M phosphate buffer: soluble 10 mg/mL, clear, orange to red (pH 6.0), H2O: soluble | solubility 0.1 M phosphate buffer: soluble 10 mg/mL, clear, orange to red (pH 6.0) | solubility 0.1 M phosphate buffer: soluble (pH 6.0), H2O: soluble | solubility 0.1 M phosphate buffer: soluble (pH 6.0), H2O: soluble |
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signalword
Danger
hcodes
pcodes
Hazard Classifications
Resp. Sens. 1
Storage Class
11 - Combustible Solids
ppe
Eyeshields, Gloves, type N95 (US)
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Can I dissolve 10mg HRP powder by splitting into certain concentrations like 1mg/ml,4mg/ml and 5mg/ml by weighing 1mg,4mg and 5mg amount and dissolving it separately instead of made it to 10mg/ml in phosphate buffer as mention catalog. Will it work?
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