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Leech-specific nutritional induction solution, and method for extracting natural hirudin

An extraction method and specific technology are applied in the field of extraction of leech-specific nutrient-inducing liquid and natural hirudin, which can solve the problems of unfavorable separation and purification, activity loss, leech damage, etc. Loss of activity, effect of protecting activity

Pending Publication Date: 2019-03-29
SUZHOU UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method also has the damage of extruding physical stimulation to the leech, and in addition, the leech has too many other impurities (such as interstitial fluid, animal blood, etc.) in the whole tissue, which finally leads to a very low purity and recovery rate of the extracted hirudin.
Although "a large-scale production method of natural hirudin" (CN104151425A) records the use of acetic acid, citric acid, sodium chloride and carbonate solution as inducing substances, while using zinc sulfate or a mixture of zinc sulfate and ethanol as an emetic , but the amount of food absorbed by leeches and the emetic effect are not good, especially for the crude product liquid (primary extract) spit out by leeches, only two simple processes of filtering and drying are used to extract hirudin. It is obviously difficult to obtain high activity and high Pure Natural Hirudin
[0009] 4. At present, there is still a considerable degree of activity loss in the use of freeze-drying technology for hirudin extracts, and a large amount of impurities such as host animal blood remaining in the overall homogenate of leeches is not conducive to further separation and purification

Method used

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  • Leech-specific nutritional induction solution, and method for extracting natural hirudin
  • Leech-specific nutritional induction solution, and method for extracting natural hirudin
  • Leech-specific nutritional induction solution, and method for extracting natural hirudin

Examples

Experimental program
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Effect test

Embodiment 1

[0078] Take 30g of adult Philippine bull leeches fasted and hungry for about a month, and rinse with sterile water. Filling with casing, preheated to 37℃, containing 150mmol / L NaCl, 10mmol / L compound amino acid, 1mmol / L glucose and 0.03% trans-4,5-epoxy-2-decenal specific nutrition induction 300mL of liquid, feeding for 2h. Soak the fed philippines leeches in 8% ethanol for 15 minutes to spit out the body fluid; remove the leeches and move them to sterile water to soak and clean them repeatedly; put the leeches in mineral water to continue feeding for the next time It is used to extract natural hirudin; meanwhile, the exhaled liquid in the collection container is the primary extract (contains specific nutrient-inducing fluid, salivary gland secretions, emetics and other mucus mixtures). Use 30%-80% saturated ammonium sulfate to fractionate and remove impurities. Centrifuge at 8000 rpm for 20 minutes, dialyze to remove salt, concentrate, collect the precipitate, and dissolve th...

Embodiment 2

[0080] Take 30g of adult Philippine bull leeches fasted and hungry for about a month, and rinse with sterile water. Filling with casing, preheated to 37℃, containing 150mmol / L NaCl, 10mmol / L compound amino acid, 1mmol / L glucose and 0.03% trans-4,5-epoxy-2-decenal specific nutrition induction 300mL of liquid, feeding for 2h. Soak the fed philippines leeches in 8% ethanol for 20 minutes to spit out the body fluid; remove the leeches and transfer them to sterile water to soak and clean them repeatedly; put the leeches in mineral water to continue feeding for the next time It is used to extract natural hirudin; at the same time, the liquid in the container is collected as the primary extract (containing specific nutrient-inducing fluid, salivary gland secretions, emetics and other mucus mixtures). Use 30%-80% saturated ammonium sulfate to fractionate and remove impurities. Centrifuge at 8000 rpm for 20 minutes, dialyze to remove salt, concentrate, collect the precipitate, and diss...

Embodiment 3

[0082] Take 30g of adult Philippine bull leeches fasted and hungry for about a month, and rinse with sterile water. Filling with casing, preheated to 37℃, containing 100mmol / L NaCl, 15mmol / L compound amino acid, 0.5mmol / L glucose and 0.04% trans-4,5-epoxy-2-decenal specific nutrition 300mL of induction solution, fed for 2h. Soak the fed philippines leeches in 8% ethanol for 20 minutes to spit out the body fluid; remove the leeches and transfer them to sterile water to soak and clean them repeatedly; put the leeches in mineral water to continue feeding for the next time It is used to extract natural hirudin; at the same time, the liquid in the collection container is the primary extract (containing specific nutrient-inducing fluid, salivary gland secretions, emetics and other mucus mixtures). Use 30%-80% saturated ammonium sulfate to fractionate and remove impurities. Centrifuge at 8000 rpm for 20 minutes, dialyze to remove salt, concentrate, collect the precipitate, and dissol...

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Abstract

The invention relates to the field of biotechnologies, and especially relates to a leech-specific nutritional induction solution, and a method for extracting natural hirudin. The leech-specific nutritional induction solution includes sodium chloride, compound amino acids, glucose, trans-4,5-epoxy-(E)-2-decenal and sterile water. A technology for satiating Poecilobdella manillensis Lesson by usingliving body induction is adopted to avoid large-scale killing of leeches and make the leeches reused. Feeding of the specific nutritional induction solution instead of whole bovine blood and other induction substances with poor palatability can induce the full intake of leeches and reduce the mixing of a large amount of blood and other impurities, so the subsequent separation and purification of natural hirudin are benefited.

Description

Technical field [0001] The present invention relates to the field of biotechnology, in particular to a leech specific nutrient induction liquid and a method for extracting natural hirudin. Background technique [0002] Leeches are commonly known as leeches, horse turtles, etc. They belong to the phylum Annelida and the class of Hirudinidae. It is estimated that there are about 500 species in the world, and there are about 100 species in my country, and 89 species have been discovered. Leeches can be divided into two categories according to their feeding habits: sucking blood and not sucking blood. At present, blood-sucking leeches that suck the blood of mammals or other vertebrates are collectively referred to as medical leeches in the world. [0003] The Philippine cow leech (Poecilobdella manillensis Lesson) is commonly known as the Phnom Penh leech, and belongs to the phylum Annelida, Hirudinidae, Hirudeidae, Medical Hiruidae, and Bovine Hiru. Among the blood-sucking leeches,...

Claims

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Application Information

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IPC IPC(8): A23K50/80A23K20/22A23K20/142A23K20/163A23K20/121A01K61/40C07K14/815C07K1/30C07K1/14
CPCA23K20/121A23K20/142A23K20/163A23K20/22A23K50/80A01K61/40C07K14/815
Inventor 许维岸卢天才韩宏岩郭桃桃李阳
Owner SUZHOU UNIV
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