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Method for entrapping and cross-linking phosphatidase A1 aggregates

A technology of cross-linking phospholipids and enzyme aggregates, applied in the direction of immobilization on/in the organic carrier, which can solve the problem of low activity of phospholipase A

Inactive Publication Date: 2012-02-01
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The present invention is aimed at immobilized phospholipase A 1 low activity, and the proposed use of entrapped-cross-linked enzyme aggregates for phospholipase A 1 immobilization method

Method used

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Experimental program
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Effect test

specific Embodiment approach 1

[0007] Embodiment 1: Immobilization of Phospholipase A by Embedding-Cross-Linking Enzyme Aggregates 1 The method is realized by the following steps: 1. Phospholipase A 1 Immobilization: 1. Phospholipase A 1 Preparation of aggregates: 1 mL of enzyme was added to a certain amount of sodium dihydrogen phosphate and citric acid buffer with a pH value of 5.0, and absolute ethanol was slowly added at 0°C with continuous stirring to ensure uniform particles of enzyme aggregates. The reaction was carried out in an ice bath for a period of time, and then centrifuged at 10,000 rpm to collect the pellet. 2. Cross-linked phospholipase A 1 Preparation and embedding process of aggregates: add a certain concentration of glutaraldehyde to the aggregate suspension, stir for a period of time, remove the supernatant by centrifugation, wash the cross-linking enzyme aggregates with buffer several times until the washing solution No viability was detected, and the pellet was collected and suspen...

specific Embodiment approach 2

[0008] Embodiment 2: The difference between this embodiment and Embodiment 1 is that in step 3, phospholipase A is carried out under the condition that the concentration of the enzyme solution is 0.03-0.07 g / mL. 1 fixed. Other steps are the same as in the first embodiment.

specific Embodiment approach 3

[0009] Embodiment 3: The difference between this embodiment and Embodiment 1 is that in step 3, phospholipase A is carried out under the condition of 70-85% saturation of the precipitant. 1 fixed. Other steps are the same as in the first embodiment.

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Abstract

The invention provides a method for immobilizing phosphatidase A1. The method is improved on the technical basis of cross-linked enzyme aggregates and comprises the steps of: preparing free phosphatidase A1 into cross-linked phosphatidase aggregates; and then entrapping the obtained cross-linked phosphatidase aggregates to obtain immobilized phosphatidase A1. The influences of concentration of a phosphatidase liquid, saturation degree of a precipitating agent, precipitation Ph value, precipitation time, concentration of a cross-linking agent, cross-linking time, concentration of sodium alginate, concentration of calcium ions and other different immobilizing conditions on the immobilizing efficiency and the catalytic effect of the phosphatidase are researched, and the enzymology nature of an immobilized phosphatidase membrane is discussed. The advantage of immobilizing the phosphatidase by adopting the method for entrapping and cross-linking aggregates is further explained through an SEM (Scanning Electron Microscope) image of the immobilized phosphatidase membrane. The phosphatidase is immobilized by utilizing the excellent chemical stability, mechanical property and other properties of the entrapped and cross-linked phosphatidase aggregates, the activity and the stability of the phosphatidase in a reaction system are improved, the activity and the selectivity of the phosphatidase are adjusted and controlled, so as to be favorable for the recovery of the phosphatidase and the maintenance of higher activity recovery rate of the phosphatidase; and the activity recovery rate of the finally obtained immobilized phosphatidase is 80.2 percent, and the relative activity of the phosphatidase after being repeatedly used for seven times is kept at 65 percent above.

Description

Technical field: [0001] The present invention relates to a method for immobilizing phospholipase. Background technique: [0002] Phospholipase A 1 It is widely used in phospholipid modification, oil refining, etc., but free phospholipase A 1 It is expensive, difficult to recycle, and the conditions for catalyzing hydrolysis are harsh; compared with free enzymes, immobilized enzymes maintain their high efficiency, specificity and mild enzyme catalytic reaction characteristics, while also showing high storage stability, easy separation and recovery, and high reliability. It has the advantages of repeated use, continuous and controllable operation, and simple process. [0003] Cross-linked enzyme aggregates (CLEAs) are a new type of immobilized enzyme technology. They have the advantages of simple preparation, high recovery rate of enzyme activity, and strong operational and storage stability. They are widely used in the preparation of immobilized enzymes. In recent years, a...

Claims

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Application Information

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IPC IPC(8): C12N11/10C12N11/04
Inventor 于殿宇胡立志王立琦杜晶王妍李越张佳宁时敏陈晓慧李志平王雪宋云花周旋奚会松
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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