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A convenient and rapid separation and extraction process of cobotide

An extraction process, cobotide technology, applied in the field of biomedical extraction, can solve problems such as difficult quality control, cumbersome operation, and long production cycle, and achieve the effects of convenient operation, simple extraction process, and increased production capacity

Active Publication Date: 2018-12-07
云南南诏药业有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The technical problem to be solved by the present invention is to overcome the long production cycle existing in the extraction method of the existing cobra snake venom neurotoxin, the operation is comparatively loaded down with trivial details, the shortcomings such as quality is difficult to control, Provide a more convenient, fast and safe separation and extraction process of cobotide

Method used

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  • A convenient and rapid separation and extraction process of cobotide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0105] Embodiment 1: the preparation of cobra neurotoxin:

[0106] (1) Sephadex G-50 is swelled with water for injection, and equilibrated with 0.01M phosphate buffer at pH 8.0 after packing.

[0107] (2) Swell CM-32 with water for injection, react with 0.5M sodium hydroxide for 30 minutes, and rinse until neutral; then react with 0.5M hydrochloric acid for 30 minutes, rinse until neutral, pack the column, and use pH8.0 Equilibrate with 0.01M phosphate buffer.

[0108] (3) Take 10g of crude cobra venom and add 4 times the amount of 0.01M phosphate buffer with pH 8.0 to dissolve, centrifuge at 3000rpm for 10min, and set aside.

[0109] (4) Put the centrifuged snake venom solution in step (3) on the equilibrated Sephadex G-50 column, elute with 0.01M phosphate buffer at pH 8.0 at a flow rate of 40ml / h, and collect the filtrate with an automatic partial collector.

[0110] (5) After 24 hours, detect and collect with an ultraviolet spectrophotometer, and collect peak E.

[0111...

Embodiment 2

[0112] Embodiment 2: the preparation of cobra venom neurotoxin:

[0113] (1) Sephadex G-50 is swelled with water for injection, and equilibrated with 0.01M phosphate buffer with pH 7.9 after packing.

[0114] (2) Swell CM-32 with water for injection, react with 0.5M sodium hydroxide for 30 minutes, and rinse until neutral; then react with 0.5M hydrochloric acid for 30 minutes, rinse until neutral, pack the column, and use pH7.9 Equilibrate with 0.01M phosphate buffer.

[0115] (3) Take 10g of crude cobra venom and add 4 times the amount of 0.01M phosphate buffer with pH7.9 to dissolve, centrifuge at 3000rpm for 10min, and set aside.

[0116] (4) The centrifuged snake venom solution in step (3) is put on the equilibrated Sephadex G-50 column, eluted with 0.01M phosphate buffer solution with pH 7.9, the flow rate is 30ml / h, and the filtrate is collected by an automatic partial collector.

[0117] (5) After 24 hours, detect and collect with an ultraviolet spectrophotometer, and...

Embodiment 3

[0119] Embodiment 3: the preparation of cobra venom neurotoxin:

[0120] (1) Sephadex G-50 is swollen with water for injection, and equilibrated with 0.01M phosphate buffer at pH 8.1 after packing.

[0121] (2) CM-32 was swelled with water for injection, reacted with 0.5M sodium hydroxide for 30 minutes, and washed to neutrality; then reacted with 0.5M hydrochloric acid for 30 minutes, washed to neutrality, packed into a column, and washed with pH8.1 Equilibrate with 0.01M phosphate buffer.

[0122] (3) Take 10g of crude cobra venom and add 4 times the amount of 0.01M phosphate buffer with pH 8.1 to dissolve, centrifuge at 3000rpm for 10min, and set aside.

[0123] (4) The centrifuged snake venom solution in step (3) is put on the equilibrated Sephadex G-50 column, eluted with 0.01M phosphate buffer solution with pH 8.1, the flow rate is 50ml / h, and the filtrate is collected by an automatic partial collector.

[0124] (5) After 24 hours, detect and collect with an ultraviole...

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Abstract

The invention belongs to the technical field of biological medicine extraction and relates to a convenient and fast cobratide separation extraction technology. The technology comprises separating treated snake venom stock solutions according to different relative mass-to-charge ratio molecule sizes from large to small through a SephadexG-50 column to obtain a desired E peak, carrying out desired E peak separation purification through a cation CM-32 exchange chromatography column and carrying out freeze drying to obtain cobratide. The technology has simple processes, can be operated easily, realizes good quality control, greatly shortens a production period, reduces a production cost, produces a finished product with purity greater than 97% and is suitable for industrial large-scale production.

Description

Technical field: [0001] The invention belongs to the technical field of biomedicine extraction, and in particular relates to a convenient and rapid kobotide separation and extraction process. Background technique: [0002] Snake venom is a liquid secreted by poisonous snakes from the venom glands. The main component is toxic protein, which accounts for about 90% to 95% of the dry weight. There are more than 20 kinds of enzymes and toxins. In addition, it also contains some small molecular peptides, amino acids, carbohydrates, lipids, nucleosides, biogenic amines and metal ions. The composition of snake venom is very complex, and the toxicity, pharmacology and toxicological effects of different snake venoms have their own characteristics. As far as cobra snake venom is concerned, it contains cytotoxin (CTX), neurotoxin (neurotoxin, NT), nerve growth factor (Nerve growth factor, NGF) and various enzymes. The content of neurotoxin (neurotoxin, NT) in snake venom is relativel...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/46C07K1/16A61K38/17
Inventor 窦啟玲苏凯
Owner 云南南诏药业有限公司