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Method for extracting castor crude toxalbumin

An extraction method and a technology for toxic proteins, which are applied in the preparation methods of peptides, chemical instruments and methods, plant peptides, etc., can solve the problems of long time required, high preparation cost, low extraction efficiency, etc., and achieve the maintenance of biological activity and operation. Simple, non-toxic effect of reagents

Inactive Publication Date: 2012-06-06
ZHONGKAI UNIV OF AGRI & ENG +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the steps of the above-mentioned methods are relatively cumbersome, the required time is long, the extraction efficiency is low, the preparation cost is high, and it is difficult to industrialize

Method used

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  • Method for extracting castor crude toxalbumin
  • Method for extracting castor crude toxalbumin
  • Method for extracting castor crude toxalbumin

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Select fresh castor beans, put the castor bean kernels obtained after shelling or the castor bean cakes obtained after oil extraction into a mortar or colloid mill or masher; according to the castor bean or castor bean cakes and solvent The mass ratio is 1:3, and 20% by mass NaCl aqueous solution is added as a solvent for grinding; then, centrifugation (20000 rpm, 4 min) at -20°C to remove the grease layer and impurity protein precipitation, and collect the clear liquid to obtain Castor crude toxic protein solution. Using isoelectric point precipitation method, the pH value of the crude ricin solution is adjusted to the isoelectric point of ricin (i.e. PI6.7 and PI6.1) through the acetic acid-sodium acetate buffer with pH value of 4.6. Then centrifuge (20000rpm, 4min) at -20°C to precipitate the ricin, discard the supernatant, and take the resulting precipitate as crude ricin.

Embodiment 2

[0021] Select fresh castor beans, put the castor bean kernels obtained after shelling or the castor bean cakes obtained after oil extraction into a mortar or colloid mill or masher; according to the castor bean or castor bean cakes and solvent The mass ratio is 1:30, and 15% by mass NaCl aqueous solution is added as a solvent for grinding; subsequently, centrifugation (20000rpm, 4min) is performed at 0°C to remove the grease layer and impurity protein precipitation, and the clear liquid is collected to obtain castor Hemp protein solution. Using the isoelectric point precipitation method, the pH value of the crude ricin solution was adjusted to the isoelectric point of ricin (i.e. PI6.7 and PI6.1) through an acetic acid-sodium acetate buffer with a pH of 6.0. Then centrifugation (20000 rpm, 4 min) was performed at 0°C to precipitate the ricin, the supernatant was discarded, and the resulting precipitate was the crude ricin.

Embodiment 3

[0023] Select fresh castor beans, put the castor bean kernels obtained after shelling or the castor bean cakes obtained after oil extraction into a mortar or colloid mill or masher; according to the castor bean or castor bean cakes and solvent The mass ratio is 1:3, and 20% by mass NaCl aqueous solution is added as a solvent for grinding; subsequently, centrifugation (20000rpm, 4min) is performed at 20°C to remove the grease layer and impurity protein precipitation, and the clear liquid is collected to obtain castor Hemp protein solution. Using isoelectric point precipitation method, the pH value of the crude ricin solution is adjusted to the isoelectric point of ricin (i.e. PI6.7 and PI6.1) through the sodium dihydrogen phosphate buffer with pH value of 5.0. Then centrifugation (20000rpm, 4min) was performed at 20°C to precipitate the ricin, the supernatant was discarded, and the resulting precipitate was the crude ricin.

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Abstract

The invention discloses a method for simply and rapidly extracting castor crude toxalbumin. The method comprises the following steps of: 1) shelling castor seeds or expelling oil, and backing up the castor kernel after shelling or the castor cake after expelling oil, 2) taking the castor kernel or the castor cake and adding solvent so as to grind, 3) centrifuging, removing the oil layer and the foreign protein sediment, and collecting the clear liquid so as to acquire castor crude toxalbumin solution, and 4) separating and purifying the acquired castor crude toxalbumin solution by using the isoelectric point sedimentation method so as to acquire the castor crude toxalbumin. The extracting method has simple operation, the required agent is non-toxic, environment-friendly and low-cost, and the method is fit for commercial process. The inorganic salt solution such as NaCl water solution is used as solvent to extract the castor crude toxalbumin so the bioactivity of the castor crude toxalbumin is maintained. Besides, the extracted castor crude toxalbumin is dissolved in the NaCl water solution in order to store and apply the castor crude toxalbumin.

Description

Technical field [0001] The present invention relates to a method for extracting protein, more specifically, the present invention relates to a method for extracting crude ricin. Background technique [0002] Ricin is a ribosome-inactivating protein. It was first isolated from castor bean by Dixson in 1887. Its molecular weight is about 64,000 to 80,000. The crystalline forms that have been discovered are BI type and TL. Type, type CI and type D, of which type D is the most toxic. Ricin with a molecular weight of 64,000 is covalently connected by two peptide chains A and B by disulfide bonds. With the assistance of chain B (RTB), Ricin easily penetrates the cell membrane and destroys the 60S sub-nucleoprotein body. The unit inhibits protein synthesis and causes cell death. The primary structure of Ricin shows that the A chain is composed of 263 amino acid residues and has a molecular weight of 32,000. It is an active chain. The 10th residue, Asparagine (Asn), has been glycosylat...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K1/30C07K1/14C07K14/415
Inventor 宋光泉胡良成孙媚华陈迁梁关生张越华
Owner ZHONGKAI UNIV OF AGRI & ENG