Flavobacterium Heparinase I

Heparinase I

Heparinase I, also known as Heparin Lyase I, is endolytically and exolytically active on both heparin and highly sulfated domains of heparan sulfate. It cleaves selectively, via an elimination mechanism, highly sulfated polysaccharide chains containing 1-4 linkages between hexosamines and o-sulfated iduronic acid residues. The reaction yields oligosaccharide products (mainly disaccharides) containing unsaturated uronic acids which can be detected by UV spectroscopy at 232 nm. The enzyme also cleaves the antithrombin III binding pentasaccharide domain in the heparin molecule.

Recombinant enzyme homologous to its native counterpart and expressed in the natural host, Pedobacter heparinus (formerly, Flavobacterium heparinum)

Heparinase I is O-glycosylated at Ser-39.
EC Number: 4.2.2.7
Purity: ≥ 95 % by reversed phase HPLC analysis.
IBEX liquid heparinase I has optimal activity and stability for up to three years when stored frozen at -70°C.
IBEX lyophilized heparinase I has optimal activity and stability for up to two years when stored at 2-8^oC.
Calcium dependent enzyme and has optimal activity in the presence of 1.0-5.0 mM CaCl₂. Optimal concentration is 1 mM

HEPARINASE II

HEPARINASE III

HEPARINASE I for DIAGNOSTICS

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APPLICATIONS

REFERENCES

Data Sheet - LIQUID


	DATA SHEET	Revised FEB 2023, R. 05
Heparinase I	Research Grade	Part No.
Synonyms	Heparinase; heparin lyase; heparin eliminase	50-010 (0.5 IU/vial)
Source	Flavobacterium heparinum (recombinant)	50-010-001 (0.1 IU/vial)
EC Number	4.2.2.7	50-010-002 (Bulk)
CAS Number	9025-39-2
Purity	≥95 % by reversed phase HPLC analysis.
Product Format	Heparinase I is presented in a phosphate buffered saline pH 7.0 containing a disaccharide as cryoprotectant. Supplied as frozen solution. No bovine serum albumin (BSA), glycerol or preservatives added.

Storage and Shipping Information	Storage Temperature: -70 °C Transport Condition: Product shipped on dry ice
Catalyzed Reaction	The enzyme cleaves selectively (via an elimination mechanism) highly sulfated polysaccharide chains containing 1-4 linkages between hexosamines & O-sulfated iduronic acid residues. The reaction yields oligosaccharide products (mainly disaccharides) containing unsaturated uronic acids, which can be detected by UV spectroscopy at 232 nm. The enzyme also cleaves the antithrombin III binding pentasaccharide domain in the heparin molecule.
Substrate Specificity	Heparin; heparan sulfate (specific activity with heparin is approx. three times higher than with heparan sulfate).
Properties	• O-glycosylated at Ser-39 • Isoelectric point: 9.3 – 9.5 • Molecular weight: 42,508 Da • Calcium ion is a cofactor and an activator
Activity	• One International Unit (IU) is defined as the amount of enzyme that will liberate 1.0 µmole unsaturated oligosaccharides from porcine mucosal heparin per minute at 30°C & pH 7.0. (Activity depends on the assay temperature, the buffer, the source & the type of Heparin used). • One Unit (U) is also defined in other preparation as 1 U that liberates 0.1 µmol of unsaturated uronic acid per hour at 25°C and pH 7.5; 1 IU is equivalent to 600 U.

Intended Use, Reference & Precaution	• These products are for in vitro R&D use only & not for therapeutic or other uses. • Refer to the respective lot-specific certificate of analysis for the actual activity and the shelf life. • Once thawed, aliquot as needed & freeze at -70°C to avoid multiple freeze-thaw cycles.
Applications	• In vitro neutralization of heparin in blood & plasma samples before analysis. • Preparation of disaccharides of heparin & the preparation of oligosaccharide libraries. • Measurement of heparin in blood & plasma using the in vitro thromboelastography (TEG) tests. • Coagulation & anticoagulation efficacy studies. • Production of low- & ultra-low molecular weight heparins (LMWH & ULMWH) from unfractionated heparin & immobilization of heparinase I for such use. • In-process, quality control, & compendial testing of heparins, heparan sulfate (HS), heparin- & HS-derived products. • Structural analysis, mass spectral analysis & characterization of heparin, heparan sulfate (HS), low molecular weight heparins, & synthetic heparin pentasaccharides & oligosaccharides. • Depolymerization of heparin, HS & chemically modified heparins, & molecular weight profiling of heparins. • Quantification of contaminants in heparin such as over-sulfated chondroitin sulfate & persulfonated heparin & quantification of process-related impurities. • Glycobiology & cancer biology research. • Identification of the biological properties of HS that depend on the integrity of the S-domains & determination of the spacing between S-domains. • In vitro host-pathogen interactions in viral infections, virusadhesion inhibition studies, virus-plaque inhibition assays, cell culture experiments, etc. • In vivo inhibition studies of neovascularization & proliferation of capillary endothelial cells. • Circumventing the inhibitory effects of heparin in PCR, RT-PCR, real-time RT-qPCR reaction& Western Blot. • In vitro histochemistry, immunohistochemistry, immunocytochemistry & flow cytometry, etc

Safety Data Sheet - LIQUID


	Safety Data Sheet - Heparinase I
SECTION 1 – IDENTIFICATION
Release Date	May 2019 R. 01
Product Identity	Heparinase I, recombinant in sucrose, Research Grade
Part Number	50-010
Manufacturer	IBEX Pharmaceuticals Inc. 5485 Rue Paré, Suite 100 Montréal, Québec, H4P 1P7 Canada Tel : (514) 344-4004 Fax : (514) 344-8827
Product use	Research

SECTION 2 – HAZARD IDENTIFICATION
Emergency overview	Solution in water. Will not support burning. Not explosive.
Potential Health Effects
Skin	May cause skin irritation
Eyes	May cause irritation or burning sensation
Inhalation	May cause irritation to the respiratory tract
Swallowed	Not known to cause toxic effects
Injection	Not known to cause toxicity with acute exposure, but may produce toxicity with repeated injections.
Carcinogenicity	Not determined
SECTION 3 – COMPOSITION/INFORMATION ON INGREDIENTS
CAS No.	9025-39-2
EC No.	4.2.2.7
Systematic name	Heparin lyase
Synonyms	Heparin eliminase Heparinase I
Origin	Recombinant, non-pathogenic microorganism
Ingredients	No components need to be disclosed according to the applicable regulations.
SECTION 4 – FIRST AID MEASURES
Skin contact	Flush with copious amounts of water.
Eye contact	Rinse opened eye for several minutes under running water. Then consult a doctor.
Inhalation	Supply the victim with fresh air; consult doctor in case of complaints.
Swallowing	Rinse mouth with plenty of water.
On all of the above	Consult a doctor if symptoms develop.
SECTION 5 – FIRE FIGHTING MEASURES
Suitable extinguishing agents	Noncombustible.
Protective equipment	No special measures required. Prevent contact with skin and eyes
SECTION 6 – ACCIDENTAL RELEASE MEASURES
Person-related safety precautions	Prevent contact with skin and eyes. Ventilate area.
Measures for environmental protection	No special measures required.
Measures for cleaning/collecting	Spilled material should be carefully wiped up and disposed as biomedical waste as per applicable waste disposal regulations. Wash spill site after material pickup is complete.
SECTION 7 – HANDLING AND STORAGE
Handling
Information for safe handling	Prevent contact with skin and eyes and prevent inhalation and swallowing. No other special precautions are necessary if used correctly.
Information about protection against explosions and fires	No special measures required.
Storage
Requirements to be met by storerooms and receptacles	No special requirements except that product should be stored frozen (preferably at -70 °C).
Information about storage in one common storage facilit	Not required.
Further information about storage conditions	None
Storage class	None assigned
Class according to regulation on flammable liquids	Void
SECTION 8 – EXPOSURE CONTROLS / PERSONAL PROTECTION
General protective and hygienic measures	The usual precautionary measures for handling chemicals and potentially biohazardous materials should be followed. Use clothing sufficient to avoid skin contact. Safety shower and eye wash should be available in proximity.
Protection of hands	Use synthetic water resistant gloves.
Eye protection	Use safety glasses.
SECTION 9 – PHYSICAL AND CHEMICAL PROPERTIES
Physical form	Frozen, colorless, odorless aqueous solution. (Heparinase I is a protein)
Melting point	0 °C (melting point of aqueous solution)
Boiling point	Not applicable
Flash point	Not applicable
Flammability	Product is not flammable
Danger of explosion	Not determined
Solubility	Heparinase I and ingredients are water soluble
Organic solvents	0 %
Solids content	0 %
SECTION 10 – STABILITY AND REACTIVITY
Thermal decomposition / conditions to be avoided	No decomposition if used according to specifications.
Dangerous reactions	No dangerous reactions known.
Dangerous products of decomposition	No dangerous decomposition products known.
SECTION 11 – TOXICOLOGICAL INFORMATION
Effects of Acute Exposure	Low acute toxicity expected following inhalation, ingestion, injection or skin exposure. The potential eye irritation effects of Heparinase I are unknown and it should be considered to be potentially irritating to the eyes.
Effects of Chronic Exposure	Not determined for oral ingestion or skin exposure
Development Toxicity	Not determined
Reproductive Toxicity	Not determined
Carcinogenicity	Not determined
SECTION 12 – ECOLOGICAL INFORMATION
Toxicity	No data available
Persistence and degradability	No data available
Bioaccumulative potential	No data available
Mobility in soil	No data available
PBT and vPvB assessment	No data available
Other adverse effects	No data available
SECTION 13 – DISPOSAL INFORMATION
Waste Disposal	Incinerate or bury as a solid in a licensed facility. Do not discharge into waterways or sewer systems without proper authority.
Container Disposal	Dispose of in a licensed facility. Recommend crushing or other means to prevent unauthorized reuse.
SECTION 14 – TRANSPORT INFORMATION
DOT (US)	Not dangerous goods
IMDG	Not dangerous goods
IATA	Not dangerous goods
SECTION 15 – REGULATORY INFORMATION
Not a dangerous substance or mixture according to the Globally Harmonised System (GHS).
SECTION 16 – OTHER INFORMATION
This information is given without any warranty or representation. It is believed to be correct but does not claim to be all inclusive and shall be used only as a guide. IBEX Pharmaceuticals Inc. shall not be held liable for any damage resulting from handling or contact with the above product. It is offered solely for your consideration, investigation and verification.

Data Sheet - LYOPHILIZED

DATA SHEET

Revised October, 2022, R. 02

Heparinase I Lyophilized

Research Grade

PN 60-010 (0.5 IU/vial)
PN 60-012 (2 IU/vial)
PN 60-014 (10 IU/vial)

Storage Temperature

2 – 8°C

Synonyms

Heparinase; heparin lyase; heparin eliminase

Source

Flavobacterium heparinum (recombinant)

EC Number

4.2.2.7

CAS Number

9025-39-2

Catalyzed Reaction

The enzyme cleaves selectively (via an elimination mechanism) highly sulfated polysaccharide chains containing 1-4 linkages between hexosamines & O-sulfated iduronic acid residues. The reaction yields oligosaccharide products (mainly disaccharides) containing unsaturated uronic acids, which can be detected by UV spectroscopy at 232 nm. The enzyme also cleaves the antithrombin III binding pentasaccharide domain in the heparin molecule

Substrate Specificity

Heparin; heparan sulfate (specific activity with heparin is approx. three (3) times higher than with heparan sulfate).

Properties

• O-glycosylated at Ser-39
• Molecular weight: 42,508 Da
• Isoelectric point: 9.3 – 9.5

Parameters (enzymatic activity assay)

	Range	Optimum
pH	4.0 - 9.0	7.0
Temperature	20 - 37°C	30°C
Calcium Concentration	1.0 - 5.0 mM	2.5mM

Product Format

Heparinase I is formulated in 0.33 mg Sodium Phosphate & 0.29 mg Sodium Chloride & 12.5 mg Sucrose (pH 7.0) as lyophilisate in a vacuum-sealed vial. No BSA or preservative

Shipping

Shipped at ambient temperature.

Receipt

Store refrigerated upon receipt.

Purity

≥95 % by reversed phase HPLC analysis, made from Heparinase I (PN 50-009).

Activity

• One international unit (IU) is defined as the amount of enzyme that will liberate 1.0 µmole unsaturated oligosaccharides from porcine mucosal heparin per minute at 30 ± 0.5°C & pH 7.0 ± 0.1. (Activity depends on the assay temperature, the buffer source & the type of Heparin used)
• One Unit (U) is also defined in other preparation as 1 U that liberates 0.1 µmol of unsaturated uronic acid per hour; 1 IU is equivalent to 600 U

Stability & Activity

Refer to the lot-specific Certificate of Analysis (CoA) for the shelf life when the products are stored as lyophilized vials (without reconstitution) at 2 – 8o C and the actual activity post-reconstitution.

Reconstitution & Concentration Post-reconstitution

Part No	Purified Water	Concentration	Activity
60-010	250 µL	≥ 2 IU/mL	≥ 0.5 IU/vial
60-012	250 µL	≥ 8 IU/mL	≥ 2 IU/vial
60-014	250 µL	≥ 40 IU/mL	≥ 10 IU/vial

Precautions & Disclaimer

• These products are for in vitro R&D use only & not for therapeutic or other uses.
• Reconstitute just before use
• DO NOT freeze the reconstituted enzyme

Applications

• In vitro neutralization of heparin in blood & plasma samples before analysis.
• Preparation of disaccharides of heparin & the preparation of oligosaccharide libraries.
• Measurement of heparin in blood & plasma using the in vitro thromboelastography (TEG) tests.
• Coagulation & anticoagulation efficacy studies.
• Production of low- & ultra-low molecular weight heparins from unfractionated heparin & immobilization of heparinase I for such use.
• Structural analysis, mass spectral analysis & characterization of heparin, heparan sulfate (HS), low molecular weight heparins, & synthetic heparin pentasaccharides & oligosaccharides.
• Depolymerization of heparin, HS & chemically modified heparins, & molecular weight profiling of heparins.
• Quantification of contaminants in heparin such as oversulfated chondroitin sulfate, persulfonated heparin & process-related impurities.
• Glycobiology & cancer biology research.
• Identification of the biological properties of HS that depend on the integrity of the S-domains & determination of the spacing between S-domains.
• In vitro host-pathogen interactions in viral infections, virusadhesion inhibition studies, virus-plaque inhibition assays, cell culture experiments, etc.
• In vivo inhibition studies of neovascularization & proliferation of capillary endothelial cells.
• Circumventing the inhibitory effects of heparin in PCR, RTPCR, real-time RT-qPCR reaction& Western Blot.
• In vitro histochemistry, immunohistochemistry, immunocytochemistry & flow cytometry, etc

Safety Data Sheet - LYOPHILIZED


	Safety Data Sheet - Heparinase I - Lyophilized
SECTION 1 – IDENTIFICATION
Release Date	February 28, 2018 R. 01
Product Identity	Heparinase I, Lyophilized
Part Number	60-010, 60-012, 60-014, 60-017
Manufacturer	IBEX Pharmaceuticals Inc. 5485 Rue Paré, Suite 100 Montréal, Québec, H4P 1P7 Canada Tel : (514) 344-4004 Fax : (514) 344-8827
Product use	Research
SECTION 2 – HAZARD IDENTIFICATION
Emergency overview	Will not support burning. Not explosive.
Potential Health Effects
Skin	May cause skin irritation
Eyes	May cause irritation or burning sensation
Inhalation	May cause irritation to the respiratory tract
Swallowed	Not known to cause toxic effects
Injection	Not known to cause toxicity with acute exposure, but may produce toxicity with repeated injections.
Carcinogenicity	Not determined
SECTION 3 – COMPOSITION/INFORMATION ON INGREDIENTS
CAS No.	9025-39-2
EC No.	4.2.2.7
Systematic name	Heparin lyase
Synonyms	Heparin eliminase Heparinase I
Origin	Recombinant, non-pathogenic microorganism
Ingredients	No components need to be disclosed according to the applicable regulations.
SECTION 4 – FIRST AID MEASURES
Skin contact	Flush with copious amounts of water.
Eye contact	Rinse opened eye for several minutes under running water. Then consult a doctor.
Inhalation	Supply the victim with fresh air; consult doctor in case of complaints.
Swallowing	Rinse mouth with plenty of water.
On all of the above	Consult a doctor if symptoms develop.
SECTION 5 – FIRE FIGHTING MEASURES
Suitable extinguishing agents	Noncombustible.
Protective equipment	No special measures required. Prevent contact with skin and eyes
SECTION 6 – ACCIDENTAL RELEASE MEASURES
Person-related safety precautions	Prevent contact with skin and eyes. Ventilate area.
Measures for environmental protection	No special measures required.
Measures for cleaning/collecting	Spilled material should be carefully wiped up and disposed as biomedical waste as per applicable waste disposal regulations. Wash spill site after material pickup is complete.
SECTION 7 – HANDLING AND STORAGE
Handling
Information for safe handling	Prevent contact with skin and eyes and prevent inhalation and swallowing. No other special precautions are necessary if used correctly.
Information about protection against explosions and fires	No special measures required.
Storage
Requirements to be met by storerooms and receptacles	No special requirements except that product should be stored stored refrigerated 2 - 8 °C
Information about storage in one common storage facilit	Not required.
Further information about storage conditions	None
Storage class	None assigned
Class according to regulation on flammable liquids	Void
SECTION 8 – EXPOSURE CONTROLS / PERSONAL PROTECTION
General protective and hygienic measures	The usual precautionary measures for handling chemicals and potentially biohazardous materials should be followed. Use clothing sufficient to avoid skin contact. Safety shower and eye wash should be available in proximity.
Protection of hands	Use synthetic water resistant gloves.
Eye protection	Use safety glasses.
SECTION 9 – PHYSICAL AND CHEMICAL PROPERTIES
Physical form	Lyophilized white cake in amber glass vial closed with cap covered with aluminum seal.
Melting point	Not applicable
Boiling point	Not applicable
Flash point	Not applicable
Flammability	Product is not flammable
Danger of explosion	Not determined
Solubility	Heparinase I and ingredients are water soluble
Organic solvents	0 %
Solids content	100 %
SECTION 10 – STABILITY AND REACTIVITY
Thermal decomposition / conditions to be avoided	No decomposition if used according to specifications.
Dangerous reactions	No dangerous reactions known.
Dangerous products of decomposition	No dangerous decomposition products known.
SECTION 11 – TOXICOLOGICAL INFORMATION
Effects of Acute Exposure	Low acute toxicity expected following inhalation, ingestion, injection or skin exposure. The potential eye irritation effects of Heparinase I are unknown and it should be considered to be potentially irritating to the eyes.
Effects of Chronic Exposure	Not determined for oral ingestion or skin exposure
Development Toxicity	Not determined
Reproductive Toxicity	Not determined
Carcinogenicity	Not determined
SECTION 12 – ECOLOGICAL INFORMATION
Toxicity	No data available
Persistence and degradability	No data available
Bioaccumulative potential	No data available
Mobility in soil	No data available
PBT and vPvB assessment	No data available
Other adverse effects	No data available
SECTION 13 – DISPOSAL INFORMATION
Waste Disposal	Incinerate or bury as a solid in a licensed facility. Do not discharge into waterways or sewer systems without proper authority.
Container Disposal	Dispose of in a licensed facility.
SECTION 14 – TRANSPORT INFORMATION
DOT (US)	Not dangerous goods
IMDG	Not dangerous goods
IATA	Not dangerous goods
SECTION 15 – REGULATORY INFORMATION
Not a dangerous substance or mixture according to the Globally Harmonised System (GHS).
SECTION 16 – OTHER INFORMATION
This information is given without any warranty or representation. It is believed to be correct but does not claim to be all inclusive and shall be used only as a guide. IBEX Pharmaceuticals Inc. shall not be held liable for any damage resulting from handling or contact with the above product. It is offered solely for your consideration, investigation and verification.
Date of revision	February 28, 2018

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