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Method for extracting high-purity metallothionein (MT) from oratosquilla oratoria

A technology for metallothionein and mantis shrimp, which is applied in the field of extracting high-purity metallothionein from mantis shrimp, can solve the problems of harsh extraction objects, short extraction period, long extraction period and the like, and reduces the complexity of the preparation process. , increased preparation, mild composition effect

Active Publication Date: 2019-08-27
MARINE FISHERIES RES INST OF ZHEJIANG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The present invention provides a method for extracting high-purity metallothionein from Mantis Shrimp in order to overcome the problems of demanding extraction objects, long extraction period, high cost and low p

Method used

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  • Method for extracting high-purity metallothionein (MT) from oratosquilla oratoria
  • Method for extracting high-purity metallothionein (MT) from oratosquilla oratoria
  • Method for extracting high-purity metallothionein (MT) from oratosquilla oratoria

Examples

Experimental program
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Effect test

Embodiment 1

[0041] (1) Shrimp shells are removed, the edible part is taken, and Tris-HCl buffer solution with a pH of 7.8 and a concentration of 0.02mol / L is added at a liquid-to-material ratio of 4:1, stirred and extracted in a water bath at 51°C for 1 hour, and the extract Refrigerate and centrifuge at 4°C and 13000r / min for 25min at high speed, discard the precipitate; collect the supernatant a and heat it in a water bath at 80°C for 5min to remove thermally unstable macromolecular proteins. Centrifuge at 1 / min for 25 min; after collecting the supernatant b, add 3 times the volume of pre-cooled (temperature 4°C) absolute ethanol, and precipitate overnight at -20°C. After centrifugation at 4°C and 13000r / min for 25min, take the precipitate and add 2-3mL ultrapure water to fully dissolve it, then centrifuge at 4°C, collect the supernatant c and store it at -18°C to obtain Mantis shrimp metallothionein crude extract, referred to as crude extract MT freeze-dried powder;

[0042] (2) Sepha...

Embodiment 2

[0053](1) Shrimp shells are removed, the edible part is taken, and Tris-HCl buffer solution with a pH of 7.5 and a concentration of 0.05mol / L is added according to the liquid-to-material ratio of 3:1, stirred and extracted in a water bath at 45°C for 2 hours, and the extract Refrigerate and centrifuge at 10°C and 10000r / min for 20min at high speed, discard the precipitate; collect the supernatant a and heat it in a water bath at 70°C for 7min to remove thermally unstable macromolecular proteins. Centrifuge at 20 min / min for 20 min; after collecting the supernatant b, add 3 times the volume of pre-cooled (temperature 4°C) absolute ethanol, and precipitate overnight at -20°C. After centrifugation at 10°C and 10000r / min for 20min, take the precipitate and add 2-3mL ultrapure water to fully dissolve it, then centrifuge at 4°C, collect the supernatant c and store it at -18°C to obtain Mantis shrimp metallothionein crude extract, referred to as crude extract MT freeze-dried powder; ...

Embodiment 3

[0056] (1) Remove the shells of the shrimps, take the edible part, add the Tris-HCl buffer solution with a pH of 8.0 and a concentration of 0.04mol / L according to the ratio of liquid to material 6:1, stir and extract in a water bath at 55°C for 1.5h, and extract The solution was refrigerated and centrifuged at 6°C and 12000r / min for 30 minutes at high speed, and the precipitate was discarded; the collected supernatant a was heated to 75°C in a water bath for 3 minutes to remove thermally unstable macromolecular proteins, and then rapidly cooled to room temperature and then placed at 6°C, Centrifuge at 12000r / min for 25min; after collecting the supernatant b, add 3 times the volume of pre-cooled (temperature 4°C) absolute ethanol, and precipitate overnight at -20°C. After centrifugation at 6°C and 12000r / min for 25min, take the precipitate and add 2-3mL ultrapure water to fully dissolve it, then centrifuge at 4°C, collect the supernatant c and store it at -18°C to obtain Mantis...

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Abstract

The invention relates to the technical field of extraction of metallothionein (MT), aims to solve the problems that an extraction target is demanding, an extraction cycle is long, the cost is high, and the purity is low in an existing MT extraction process, and provides a method for extracting high-purity MT from oratosquilla oratoria. The method comprises the following steps: (1) removing the shell of the oratosquilla oratoria, taking an edible part, performing homogenate, and performing centrifugal extraction and freeze drying; (2) performing gel filtration chromatography; (3) performing anion exchange chromatography; (4) performing desalination treatment to obtain a first subtype MT and a second subtype MT. According to the method provided by the invention, the edible part of the oratosquilla oratoria is taken as a raw material; an extraction process of the high-purity MT is designed specific to the self specific matrix characteristics of the oratosquilla oratoria; the extraction cycle of the process is short; the extraction amount is large; the cost is low; the purity is high; the process is suitable for industrialized and large-scale production.

Description

technical field [0001] The invention relates to the technical field of metallothionein extraction, in particular to a method for extracting high-purity metallothionein from mantis shrimp. Background technique [0002] Metallothionein (MT) is a low-molecular-weight protein (6000–7000 Da) rich in cysteine ​​(Cys), which has the ability to bind heavy metals through its cysteine ​​thiol group, and can bind up to 20 Monovalent or 7 divalent heavy metal ions. MT plays an important physiological role in the living body, mainly for the storage, transportation and metabolism of trace metal elements in the living body and the chelation and detoxification of non-essential heavy metal elements. Tissue oxidative damage, antagonism of physiological and biochemical reactions are also of great significance, and provide active centers for other metalloproteins and metalloenzymes, so MT has a very wide range of applications in the fields of environmental monitoring, health food and medicine....

Claims

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Application Information

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IPC IPC(8): C07K14/825C07K1/36C07K1/34C07K1/16C07K1/18
CPCC07K14/825
Inventor 梅光明严国张小军陈思孟春英
Owner MARINE FISHERIES RES INST OF ZHEJIANG
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