Preparation method of magnetic immobilized lipase capable of being used in organic media

A technology for immobilizing lipase and lipase, applied in the directions of biochemical equipment and methods, immobilization on or in inorganic carriers, enzymes, etc.

Active Publication Date: 2016-11-23
ANHUI UNIVERSITY OF TECHNOLOGY AND SCIENCE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although it has been reported that the magnetic Fe 3 o 4 , Fe 3 o 4 @SiO 2 , Fe 3 o 4 @C nanocarriers can improve the stability of lipase (Chem.Asian J.,2013,8,1447-1454; J.Mater.Chem.,2012,22,8385-8393; J.Mater.Chem.A,2014 ,2,18339-18344), bu

Method used

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  • Preparation method of magnetic immobilized lipase capable of being used in organic media
  • Preparation method of magnetic immobilized lipase capable of being used in organic media
  • Preparation method of magnetic immobilized lipase capable of being used in organic media

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] In the preparation method of magnetically immobilized lipase provided by the invention, the magnetic Fe 3 o 4 / P(St-AA) nanospheres were synthesized by the following method:

[0051] Step 1. Weigh 1059.5 mg of iron acetylacetonate and add it to 20 mL of diphenyl ether, then add 2 mL of oleylamine, 1 mL of ethylene glycol, and 2 mL of oleic acid, stir evenly, and react for 0.5 h at 200 ° C in a nitrogen atmosphere , and then reflux at 250° C. for 2 hours, stop heating, cool to room temperature, magnetically separate the black product, and wash with ethanol three times to obtain ferric oxide nanoparticles coated with oleic acid-oleylamine;

[0052] Step 2. Add 36mL sodium lauryl sulfate solution (mass concentration, 0.1mol / L) in the iron ferric oxide nano-particle of oleic acid-oleylamine wrapping obtained in step 1, after stirring and mixing, then Disperse under ultrasonic conditions for 15 minutes to prepare a magnetic ferric oxide emulsion;

[0053] Step 3. Add 90 m...

Embodiment 2

[0067] Magnetic Fe 3 o 4 The preparation of / P(St-AA) nanospheres is the same as in Example 1.

[0068] Magnetic Fe 3 o 4 The operation process of / P(St-AA) nanosphere immobilized lipase is as follows:

[0069] Step 1. The freeze-dried lipase powder derived from porcine pancreas is prepared into a 1 mg / mL lipase solution with pH=7.4 phosphate buffer, and stored in a refrigerator at 4°C for later use;

[0070] Step 2. take by weighing the magnetic Fe that 100mg above-mentioned method makes 3 o 4 / P(St-AA) nanospheres, sequentially added 191.7mg 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride, 57.6mg N-hydroxysuccinimide and 20mL morpholine Ethansulfonic acid buffer solution (0.1mol / L, pH=6.0), ultrasonically dispersed for 2min, and reacted at 25°C for 3h, the obtained product was magnetically separated and washed to obtain activated magnetic polystyrene nanospheres.

[0071] Step 3. Add 40mL of the above-mentioned lipase solution to the above-mentioned activat...

Embodiment 3

[0075] In the preparation method of magnetically immobilized lipase provided by the invention, the magnetic Fe 3 o 4 / PS-CH 2 Cl nanospheres were synthesized by the following method:

[0076] According to step 1 and step 2 of Example 1, a magnetic iron ferric oxide emulsion was prepared; 72 mg of sodium lauryl sulfate and 5 mL of p-chloromethyl styrene were added successively to 24 mL of water, and after stirring and mixing evenly, the Disperse for 20 minutes to prepare a p-chloromethylstyrene monomer emulsion; mix the obtained magnetic iron ferric oxide emulsion and p-chloromethylstyrene monomer emulsion evenly, and add 30mg Potassium persulfate initiator, after reacting at 80°C for 2 hours, then add 30mg of potassium persulfate initiator, and react at 80°C for another 20h, after the reaction is completed, naturally cool to room temperature, and the obtained precipitate is subjected to magnetic separation, Magnetic Fe after washing and drying 3 o 4 / PS-CH 2 Cl nanospher...

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Abstract

The invention discloses a preparation method of magnetic immobilized lipase capable of being used in organic media. A magnetic Fe3O4/P(St-AA) nanosphere or a magnetic Fe3O4/ PS-CH3Cl nanosphere is used as a carrier, and the lipase is fixed on the surface of the magnetic Fe3O4/P(St-AA) nanosphere or the magnetic Fe3O4/ PS-CH3Cl nanosphere through covalent binding. The method is simple and moderate, the carrier content of the lipase is high, and the magnetic immobilized lipase has the advantages of high recovery rate, good stability and wide application prospect and can be widely used in the organic media.

Description

technical field [0001] The invention relates to a preparation method of a magnetically immobilized lipase which can be used in an organic medium. Background technique [0002] Lipase is an important ester bond hydrolase, which can hydrolyze the ester bonds formed by glycerol and fatty acids, so it is also called triacylglycerol hydrolase. Studies have found that in addition to catalyzing the hydrolysis and synthesis of glycerol lipids, lipase can also catalyze esterification, transesterification, alcoholysis, acidolysis and ammonialysis in the organic phase, and has a high degree of regional and substrate and stereoselectivity. However, lipase is a water-soluble protease, which is almost insoluble in organic media and cannot fully contact with substrates such as esters, acids, alcohols, anhydrides and amides, which restricts the application of lipase in the field of organic catalysis. In addition, lipase is unstable to heat, strong acid, strong alkali, organic solvent, etc...

Claims

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Application Information

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IPC IPC(8): C12N11/14
CPCC12N9/20C12N11/14C12Y301/01003
Inventor 陈志明刘磊磊
Owner ANHUI UNIVERSITY OF TECHNOLOGY AND SCIENCE
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