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Method for preparing ACE inhibitory peptide by enzymolysis of squid viscera with magnetic nanoparticle immobilized cells

A technology of magnetic nanoparticles and immobilized cells, applied in the biological field, can solve the problems of poor operation stability of tissue enzyme autolysis method, high cost of exogenous enzyme enzymolysis, and high process complexity, so as to improve applicability and flexibility. The effect of improving the enzymatic hydrolysis efficiency and reducing the cost

Active Publication Date: 2021-05-14
HANSHAN NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to solve the existing technology of preparing squid visceral waste ACE inhibitory peptides by enzymatic hydrolysis. The disadvantages of the method are long cycle time, high process complexity, and poor operational stability of the tissue enzyme autolysis method

Method used

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  • Method for preparing ACE inhibitory peptide by enzymolysis of squid viscera with magnetic nanoparticle immobilized cells
  • Method for preparing ACE inhibitory peptide by enzymolysis of squid viscera with magnetic nanoparticle immobilized cells
  • Method for preparing ACE inhibitory peptide by enzymolysis of squid viscera with magnetic nanoparticle immobilized cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] This implementation illustrates the method and beneficial effects of preparing squid visceral active peptides by using nano-magnetic microspheres to immobilize cells compared with free cells.

[0043] The specific implementation steps are as follows:

[0044] Step 1: Mix 500g of squid viscera with 500mL of deionized water, heat at 100°C for 30 minutes, remove the massive fish bones, grind the heated slurry in a homogenizer, and then place it in a shallow pan at 80°C Drying for 18 hours, collecting the dried bulk powder, grinding into a fine powder in a pulverizer, and passing through a 60-mesh sieve to obtain a fine powder of the squid viscera extract.

[0045] The second step: the preparation of magnetic ferric iron tetroxide (Fe 3 o 4 ) Nanoparticles: Dissolve 21.6g of ferric chloride hexahydrate and 8g of ferrous chloride tetrahydrate in 100mL of deionized water, gradually raise the temperature to 60°C under the protection of nitrogen, and stir at constant temperat...

Embodiment 2

[0069] The specific implementation steps are as follows:

[0070] Step 1: Mix 500g of squid viscera with 1000mL of deionized water, heat at 90°C for 20 minutes, remove the massive fish bones, grind the heated slurry in a homogenizer, and then place it in a shallow pan at 90°C Dry it under the hood for 20 hours, collect the dried bulk powder, grind it into a fine powder in a pulverizer, and pass through an 80-mesh sieve to obtain a fine powder of the squid viscera extract.

[0071] The second step: the preparation of magnetic ferric iron tetroxide (Fe 3 o 4 ) Nanoparticles: Dissolve 25g of ferric chloride hexahydrate and 10g of ferrous chloride tetrahydrate in 120mL of deionized water, gradually raise the temperature to 60°C under the protection of nitrogen, and stir at constant temperature for 3 hours. Then add 80 mL of ammonia water with a concentration of 25%, raise the temperature to 80° C. and react for 2 hours, stop stirring and cool to room temperature, and let stand u...

Embodiment 3

[0080] In this example, magnetic microsphere immobilization and sodium alginate, polyethylene glycol, chitosan and other common media immobilization methods were used to study the effects on proteolysis rate and ACE inhibitory activity.

[0081] The magnetic microspheres were immobilized as described in Example 1, using common media such as sodium alginate, polyethylene glycol, and chitosan to immobilize references (International Journal of Peptide Research and Therapeutics, 25 (2), 681-689. ) method was implemented, the immobilized cells carried out enzymolysis for 8 hours according to the same operation as the enzymolysis process described in Implementation 1, and adopted the same detection method as in Example 1 to measure proteolysis rate and ACE inhibitory activity. The results are as follows figure 2 shown.

[0082] From figure 2It can be seen that the degree of proteolysis after immobilization with magnetic microspheres is significantly higher than that of immobiliza...

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Abstract

The invention relates to the technical field of biology, in particular to a method for preparing ACE inhibitory peptide by adopting polyethyleneimine (PEI) modified silicon dioxide coated ferroferric oxide (Fe3O4) magnetic nano-microspheres and utilizing nano-microsphere immobilized bacillus subtilis cells to carry out enzymolysis on squid viscera leftovers. The method comprises the following steps: preparing squid viscera extract dry powder, preparing the magnetic nanoparticles, activating and culturing bacterial strains, immobilizing cells, performing enzymolysis and separation, performing ultrafiltration and drying. The method disclosed by the invention is high in enzymolysis efficiency, and an enzymolysis product is easy to separate. The immobilized cells can still achieve good protein hydrolysis degree and ACE inhibitory activity after being repeatedly used for multiple times, and the cost of the method is reduced. The prepared polypeptide product has good ACE inhibitory activity, and the IC50 value reaches 0.109 mg / mL.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for preparing ACE inhibitory peptides by enzymolyzing squid viscera with immobilized cells using magnetic nanoparticles. Background technique [0002] Squid viscera is the main by-product of squid processing, accounting for more than 20% of the fresh weight of squid. Squid offal has high moisture content and active tissue enzymes, making it difficult to store. After processing, most of them are discarded directly, or simply processed into feed for breeding, which not only pollutes the environment, but also causes a waste of resources. Squid viscera is a kind of protein-rich biological resource, which contains rich protein and other nutrients, and most of these proteins are water-soluble proteins that are easy to degrade. Therefore, squid viscera is a good raw material for preparing bioactive peptides. At present, there are three main methods for preparing bioactive peptides...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N11/14C12P21/06C07K1/34C12R1/125
CPCC12N11/14C12P21/06C07K1/34
Inventor 李云周飞朱慧
Owner HANSHAN NORMAL UNIV
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