Native Lipoprotein Lipase from Microorganism

Products

Native Lipoprotein Lipase from Microorganism

Product Specifications

Cat#	NATE-1005
Descriptions	Lipoprotein Lipase effectively breaks down triglycerides bound to serum lipoproteins into glycerol and fatty acids, the foundational catalytic step for full triglyceride quantitative diagnosis. It tolerates high-lipid turbid serum samples without activity inhibition, cooperating with glycerol kinase and glycerophosphate oxidase to form a complete triglyceride detection system for metabolic disorder and hyperlipidemia screening.
Applications	This product can be combined with corresponding supporting enzymes to perform enzymatic determination of triglycerides in serum for clinical diagnosis.
Synonyms	Triacylglycerol acylhydrolase
EC Number	EC 3.1.1.3
Catalysis	Triglyceride + 3H₂O → Glycerol + 3RCOOH
Activity	≥ 1,000 U/g (Lipoprotein lipase activity)
Appearance	White powder, lyophilized
Additive	Glycine
Molecular Weight	33,000 (Gel filtration)
Isoelectric Point	4.3
Km	3.95 × 10⁻³M
Optimum pH	7.0
pH Stability	5.0-9.0 (30°C, 24 h)
Optimum Temperature	50°C
Thermal Stability	up to 40°C (pH 7.0, 30 min)
Inhibitors	Sodium Cholate
Stabilizers	Ca²⁺
Unit Definition	1 unit is defined as the enzyme quantity which liberates 1 μ mole of fatty acid per minute under the conditions described below.
Storage	24 months from the date of analysis when stored at -20°C or below in a dry place under sealed conditions.

General Properties

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Assay Principle

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Activity Assay

Unit Definition

1 unit is defined as the enzyme quantity which liberates 1 μ mole of fatty acid per minute under the conditions described below.

Reagents

A. 2/15 M Phosphate buffer (KH2PO4-Na2HPO4, pH 7.0)
B. 1/15 M Phosphate buffer (KH2PO4-Na2HPO4, pH 7.0)
C. Substrate solution: Weigh 1.60 g of bovine serum albumin and dissolve in 16 mL of 2/15 M Phosphate buffer (A). Add 24 mL of 10% Intralipos into the solution and stir well. (Can be used only on the day of preparation if refrigerated)
D. Heptane -2-propanol-sulfuric acid solution: Mix 75 mL of heptane, 300 mL of 2-propanol, and 7.5 mL of 1 mol/L sulfuric acid successively. (Can be used for 3 months at room temperature)
E. Cresol red indicator: Weigh 50 mg of cresol red and dissolve in 50 mL of ethanol. Filtrate with filter paper. (Can be used for 1 month if stored in a brown bottle at room temperature or in a refrigerator)
F. Triton X-100 stock solution (5g/100 mL deionized water)
G. Diluent: Mix approx. 900 mL of 1/15 M Phosphate buffer (B) and 2 mL of Triton X-100 stock solution (F), then fill up to 1000 mL with 1/15 M Phosphate buffer (B).
H. Enzyme solution: Weigh out Lipoprotein Lipase and gently dissolve in chilled Diluent (G). Enzyme solution should be prepared so that the value of (T20-T0) becomes in the range of 0.25-0.50 mL. Do not shake the Enzyme solution vigorously, or the enzyme becomes inactive.
I. 0.01N KOH-ethanol solution Pipette 5 mL of 0.1 mol/L potassium hydroxide ethanolic solution (Commercial product, for volumetric analysis) into a volumetric flask (50 mL) and fill up to 50 mL with ethanol.

Procedure

Pipette 1.0 mL of 2/15 M Phosphate buffer (A) and 2.0 mL of Substrate solution (C) respectively into a test tube (Φ25 x 200 mm) with a ground stopper and mix well. Stand the solution in a water bath shaker at 37 ± 0.5℃ for more than 10 minutes, then add 1.0 mL of Enzyme solution (H) and incubate with shaking. Exactly 20 minutes after addition of Enzyme solution (H), add 10.0 mL of Heptane-2-propanpol-sulfuric acid solution (D), then shake and mix well. Add 6.0 mL of heptane and 4.0 mL of deionized water, then cap the stopper and shake using shaker for 5 minutes. After shaking, stand the reaction mixture in tap water for more than 20 minutes to make heptane layer separate from water layer. Pipette 6.0 mL of upper layer (heptane layer) into a test tube (Φ15 x 105 mm) and add 2 drops of Cresol red indicator (E). Then, titrate the solution with 0.01N KOH-ethanol solution (I) in nitrogen currents while stirring (T20 mL). It is the end point that the color of the solution turns violet. As a blank, pipette 1.0 mL of 2/15 M Phosphate buffer (A) and 2.0 mL of Substrate solution (C) into another test tube (Φ25 x 200 mm) with ground stopper, then add 10.0 mL of Heptane-2-propanol-sulfic acid solution (D) and mix well. Add 1 mL of Enzyme solution (H) and take the same procedure described above (T0 mL)

Calculation

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Safe Handling Avoid inhalation; in the event of direct skin or eye contact, immediately flush the affected area with abundant water, and consult the SDS for further information.