CN115316319B - Breeding method for improving antithrombin activity of leeches - Google Patents
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Classifications
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K61/00—Culture of aquatic animals
- A01K61/40—Culture of aquatic animals of annelids, e.g. lugworms or Eunice
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
- A23K10/16—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/20—Animal feeding-stuffs from material of animal origin
- A23K10/24—Animal feeding-stuffs from material of animal origin from blood
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/30—Animal feeding-stuffs from material of plant origin, e.g. roots, seeds or hay; from material of fungal origin, e.g. mushrooms
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/142—Amino acids; Derivatives thereof
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/142—Amino acids; Derivatives thereof
- A23K20/147—Polymeric derivatives, e.g. peptides or proteins
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- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/174—Vitamins
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/80—Feeding-stuffs specially adapted for particular animals for aquatic animals, e.g. fish, crustaceans or molluscs
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management
- Y02A40/81—Aquaculture, e.g. of fish
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Abstract
In order to solve the problem that the antithrombin activity is seriously lower than that of wild leech in the existing artificial culture of leech, the invention provides a culture method for improving the antithrombin activity of leech, which comprises the following steps: s1, setting a net cage; s2, seedling throwing; s3, water quality management: the water for cultivation adopts well water, the water temperature is kept in the range of 22-28 ℃, the PH value is kept in the range of 6.8-7.2, and the water body in the cultivation box adopts micro-flowing water; s4, bait feeding: mixing the nutrient solution and fresh pig blood according to a proportion, then filling the mixture into a cleaned pig casing, and bundling two ends to ensure no leakage; putting the leeches into a leech culture box, sucking the leeches, and feeding the leeches again at intervals of 7 days; s5, controlling the illumination intensity: the ambient light intensity is controlled to be 20-50Lux at 8-20 points per day, and the ambient light intensity is controlled to be 0Lux at other times. The cultivation method greatly improves the antithrombin activity and growth rate of the leech.
Description
Technical Field
The invention belongs to the technical field of leech cultivation, and particularly relates to a cultivation method for improving antithrombin activity of leeches.
Background
Leech is used for treating diseases and has quite long history at home and abroad. Leech in China is used as a traditional Chinese medicine, and is originally carried in Shennong Ben Cao Jing, namely, the leech can "dispel blood stasis" and break "blood mass", and is the only leech medicinal material type recorded in Chinese pharmacopoeia at present and taking animal blood as food source. Along with the continuous and deep clinical research of leeches in recent years, the leeches also have the effects of resisting thrombus, inflammation, tumors, atherosclerosis and the like, and particularly show great superiority and broad prospect in the aspect of treating cardiovascular and cerebrovascular diseases, cancers and the like which seriously threaten human health, so that the leech cultivation has good market prospect and economic benefit.
Because a large amount of pesticides and disinfectants are used and manually and wantonly captured, the wild resources of the leech are seriously destroyed, and the increasingly-rising demands of the market cannot be met at all. Although the artificial culture of the leeches relieves the contradiction between supply and demand of the leeches to a certain extent, the traditional culture method and the culture mode generally have the problem of low survival rate, and meanwhile, under the artificial culture condition, the antithrombin activity of the leeches is lower than that of the wild leeches, so that the utilization value in the process of pharmacy is reduced and even the leeches cannot be used. The blood water produced by leeches during feeding can also cause certain pollution to the local environment. These problems seriously obstruct the large-scale cultivation of leeches, and can not fundamentally solve the contradiction between supply and demand of leeches in domestic and foreign markets.
Disclosure of Invention
In order to solve the problem that the antithrombin activity is seriously lower than that of wild leeches in the existing artificial culture of leeches, the invention provides a culture method for improving the antithrombin activity of leeches.
The invention relates to a cultivation method for improving antithrombin activity of leeches, which comprises the following steps:
s1, setting a net cage;
s2, seedling throwing;
s3, water quality management: the water for cultivation adopts well water, the water temperature is kept in the range of 22-28 ℃, the PH value is kept in the range of 6.8-7.2, and the water body in the cultivation box adopts micro-flowing water;
s4, bait feeding: mixing the nutrient solution with fresh pig blood according to a ratio of 5-10:2-5, then filling into a cleaned pig casing, and bundling two ends to ensure no leakage; putting the leeches into a leech culture box, sucking the leeches, and feeding the leeches again at intervals of 7 days;
s5, controlling the illumination intensity: the ambient light intensity is controlled to be 20-50Lux at 8-20 points per day, and the ambient light intensity is controlled to be 0Lux at other times.
In the step S1, the length, the width and the height of the net cage are 120cm, 60cm and 45cm respectively, a water outlet of 25cm multiplied by 25cm is arranged in the middle of the bottom of the net cage, and a 20-mesh escape-preventing net is installed.
Further, in the step S2, the seedling throwing density is set according to 1500 g/box, namely, the specification is 0.3 g/leech 5000 pieces/box.
Further, in the step S4, the nutrient solution is an aqueous solution of nutrient substances consisting of protein, compound vitamin, compound amino acid, compound enzyme and preventive medicine, and the concentration of the nutrient solution is 20-30mg/ml.
Further, in the step S4, the nutrient substances consist of the following substances in mass ratio:
further, in the step S4, the nutrient substances consist of the following substances in mass ratio:
further, in the step S4, the protein is soluble fish paste protein; the compound vitamin is a compound of vitamins A, B1, B2 and C, D, E; the compound amino acid is a compound of lysine, cysteine and histidine; the complex enzyme is a complex of cellulase, amylase, pectase, xylanase, mannanase, neutral protease and a carrier.
In step S4, the preventive medicine is prepared from radix Isatidis, coptidis rhizoma, cortex Phellodendri, scutellariae radix, and fructus crataegi at a mass ratio of 3-5:0.5-1.5:0.5-1.5:0.5-1.5:2-5, adding water, decocting with strong fire, and making into medicinal liquid with concentration of 120-180 mg/ml.
In step S4, the preventive medicine is prepared from radix Isatidis, coptidis rhizoma, cortex Phellodendri, scutellariae radix, and fructus crataegi according to a mass ratio of 4:1:1:1:3 adding water and decocting with strong fire to obtain 150mg/ml medicinal liquid.
Further, in the step S4, the nutrient solution and fresh pig blood are mixed according to a ratio of 7:3, mixing evenly.
Further, in the step S5, the controlling the ambient light intensity is: 40Lux in the morning, 25Lux in the afternoon and 0Lux in the other times.
Compared with the prior art, the invention has the following advantages:
1. the illumination intensity of the living environment of the leech is accurately controlled, and the antithrombin activity and the growth speed of the leech are greatly improved.
2. The addition of the nutrient solution into the bait solves the problems of insufficient nutrition and unbalanced nutrition caused by a single blood bait source, solves the problems of dyspepsia caused by blood sucking of leeches or related diseases caused by bacterial infection, and improves the antithrombin activity and the growth speed of the leeches.
3. Fresh pig blood is used as a phagostimulant, and the optimal proportion is determined, so that the problem of poor ingestion of the artificial compound feed is solved.
4. Greatly relieves the problem of hindering the large-scale cultivation of the leeches caused by insufficient fresh blood in partial areas.
Detailed Description
The examples are presented for better illustration of the present invention, but are not intended to limit the scope of the present invention to the examples. Those skilled in the art will appreciate from the foregoing disclosure that various modifications and adaptations of the embodiments described herein are possible and can be made without departing from the scope of the invention.
Example 1
A cultivation method for improving antithrombin activity of leeches comprises the following steps:
s1, setting a net cage: setting the length, width and height of the net cage to be 120cm, 60cm and 45cm respectively, arranging a 25cm multiplied by 25cm water outlet in the middle of the bottom of the net cage, and installing a 20-mesh escape-preventing net;
s2, seedling throwing: the seedling throwing density is 1500 g/box, namely 0.3 g/leech 5000 pieces/box;
s3, water quality management: the water for cultivation adopts well water, the water temperature is kept in the range of 22-28 ℃, the PH value is kept in the range of 6.8-7.2, and the water body in the cultivation box adopts micro-flowing water;
s4, bait feeding: mixing the nutrient solution with fresh pig blood according to a proportion of 5:2, uniformly mixing the components in proportion, then filling the mixture into a cleaned pig casing, and bundling two ends to ensure no leakage; putting the leeches into a leech culture box, sucking the leeches, and feeding the leeches again at intervals of 7 days; the nutrient solution is an aqueous solution of nutrient substances consisting of protein, compound vitamin, compound amino acid, compound enzyme and preventive medicine, and the concentration of the nutrient solution is 20mg/ml.
S5, controlling the illumination intensity: the ambient light intensity is controlled to be 20Lux at 8-20 points per day, and the ambient light intensity is controlled to be 0Lux at other times.
In the step S4, the nutrient substances consist of the following substances in mass ratio:
wherein the protein is soluble fish paste protein; the compound vitamin is a compound of vitamins A, B1, B2 and C, D, E; the compound amino acid is a compound of lysine, cysteine and histidine; the complex enzyme is a complex of cellulase, amylase, pectase, xylanase, mannanase, neutral protease and a carrier.
Wherein the preventive medicine is prepared from radix isatidis, rhizoma coptidis, phellodendron, radix scutellariae and hawthorn according to the mass ratio of 3:1.5:0.5:1.5:2 adding water and decocting with strong fire to obtain medicinal liquid with concentration of 120 mg/ml.
Example 2
A cultivation method for improving antithrombin activity of leeches comprises the following steps:
s1, setting a net cage: setting the length, width and height of the net cage to be 120cm, 60cm and 45cm respectively, arranging a 25cm multiplied by 25cm water outlet in the middle of the bottom of the net cage, and installing a 20-mesh escape-preventing net;
s2, seedling throwing: the seedling throwing density is 1500 g/box, namely 0.3 g/leech 5000 pieces/box;
s3, water quality management: the water for cultivation adopts well water, the water temperature is kept in the range of 22-28 ℃, the PH value is kept in the range of 6.8-7.2, and the water body in the cultivation box adopts micro-flowing water;
s4, bait feeding: mixing the nutrient solution with fresh pig blood according to a ratio of 7:3, uniformly mixing the components in proportion, then filling the mixture into a cleaned pig casing, and bundling two ends to ensure no leakage; putting the leeches into a leech culture box, sucking the leeches, and feeding the leeches again at intervals of 7 days; the nutrient solution is an aqueous solution of nutrient substances consisting of protein, compound vitamin, compound amino acid, compound enzyme and preventive medicine, and the concentration of the nutrient solution is 25mg/ml.
S5, controlling the illumination intensity: the illumination intensity of the environment is controlled as follows: 40Lux in the morning, 25Lux in the afternoon and 0Lux in the other times.
In the step S4, the nutrient substances consist of the following substances in mass ratio:
wherein the protein is soluble fish paste protein; the compound vitamin is a compound of vitamins A, B1, B2 and C, D, E; the compound amino acid is a compound of lysine, cysteine and histidine; the complex enzyme is a complex of cellulase, amylase, pectase, xylanase, mannanase, neutral protease and a carrier.
Wherein the preventive medicine is prepared from radix isatidis, rhizoma coptidis, phellodendron, radix scutellariae and hawthorn according to the mass ratio of 4:1:1:1:3 adding water and decocting with strong fire to obtain 150mg/ml medicinal liquid.
Example 3
A cultivation method for improving antithrombin activity of leeches comprises the following steps:
s1, setting a net cage: setting the length, width and height of the net cage to be 120cm, 60cm and 45cm respectively, arranging a 25cm multiplied by 25cm water outlet in the middle of the bottom of the net cage, and installing a 20-mesh escape-preventing net;
s2, seedling throwing: the seedling throwing density is 1500 g/box, namely 0.3 g/leech 5000 pieces/box;
s3, water quality management: the water for cultivation adopts well water, the water temperature is kept in the range of 22-28 ℃, the PH value is kept in the range of 6.8-7.2, and the water body in the cultivation box adopts micro-flowing water;
s4, bait feeding: mixing the nutrient solution with fresh pig blood according to a ratio of 10:5, uniformly mixing the components in proportion, then filling the mixture into a cleaned pig casing, and bundling two ends to ensure no leakage; putting the leeches into a leech culture box, sucking the leeches, and feeding the leeches again at intervals of 7 days; the nutrient solution is an aqueous solution of nutrient substances consisting of protein, compound vitamin, compound amino acid, compound enzyme and preventive medicine, and the concentration of the nutrient solution is 30mg/ml.
S5, controlling the illumination intensity: the ambient light intensity was controlled at 50Lux at 8-20 points per day, and at 0Lux at other times.
In the step S4, the nutrient substances consist of the following substances in mass ratio:
wherein the protein is soluble fish paste protein; the compound vitamin is a compound of vitamins A, B1, B2 and C, D, E; the compound amino acid is a compound of lysine, cysteine and histidine; the complex enzyme is a complex of cellulase, amylase, pectase, xylanase, mannanase, neutral protease and a carrier.
Wherein the preventive medicine is prepared from radix Isatidis, coptidis rhizoma, cortex Phellodendri, scutellariae radix and fructus crataegi according to a mass ratio of 5:0.5:1.5:0.5:5 adding water and decocting with strong fire to obtain the medicinal liquid with concentration of 180 mg/ml.
Example 4
A cultivation method for improving antithrombin activity of leeches comprises the following steps:
s1, setting a net cage: setting the length, width and height of the net cage to be 120cm, 60cm and 45cm respectively, arranging a 25cm multiplied by 25cm water outlet in the middle of the bottom of the net cage, and installing a 20-mesh escape-preventing net;
s2, seedling throwing: the seedling throwing density is 1500 g/box, namely 0.3 g/leech 5000 pieces/box;
s3, water quality management: the water for cultivation adopts well water, the water temperature is kept in the range of 22-28 ℃, the PH value is kept in the range of 6.8-7.2, and the water body in the cultivation box adopts micro-flowing water;
s4, bait feeding: mixing the nutrient solution with fresh pig blood according to a ratio of 10:2, uniformly mixing the components in proportion, then filling the mixture into a cleaned pig casing, and bundling two ends to ensure no leakage; putting the leeches into a leech culture box, sucking the leeches, and feeding the leeches again at intervals of 7 days; the nutrient solution is an aqueous solution of nutrient substances consisting of protein, compound vitamin, compound amino acid, compound enzyme and preventive medicine, and the concentration of the nutrient solution is 28mg/ml.
S5, controlling the illumination intensity: the ambient light intensity is controlled to be 30Lux at 8-20 points per day, and the ambient light intensity is controlled to be 0Lux at other times.
In the step S4, the nutrient substances consist of the following substances in mass ratio:
wherein the protein is soluble fish paste protein; the compound vitamin is a compound of vitamins A, B1, B2 and C, D, E; the compound amino acid is a compound of lysine, cysteine and histidine; the complex enzyme is a complex of cellulase, amylase, pectase, xylanase, mannanase, neutral protease and a carrier.
Wherein the preventive medicine is prepared from radix isatidis, rhizoma coptidis, phellodendron, radix scutellariae and hawthorn according to the mass ratio of 4:1:1:1:2.5 adding water and decocting with strong fire to obtain 160mg/ml medicinal liquid.
Comparative example 1 (example 2 without nutrient solution)
A cultivation method for improving antithrombin activity of leeches comprises the following steps:
s1, setting a net cage: setting the length, width and height of the net cage to be 120cm, 60cm and 45cm respectively, arranging a 25cm multiplied by 25cm water outlet in the middle of the bottom of the net cage, and installing a 20-mesh escape-preventing net;
s2, seedling throwing: the seedling throwing density is 1500 g/box, namely 0.3 g/leech 5000 pieces/box;
s3, water quality management: the water for cultivation adopts well water, the water temperature is kept in the range of 22-28 ℃, the PH value is kept in the range of 6.8-7.2, and the water body in the cultivation box adopts micro-flowing water;
s4, bait feeding: filling fresh pig blood into the cleaned pig casing, and bundling the two ends to ensure no leakage; putting the leeches into a leech culture box, sucking the leeches, and feeding the leeches again at intervals of 7 days;
s5, controlling the illumination intensity: the illumination intensity of the environment is controlled as follows: 40Lux in the morning, 25Lux in the afternoon and 0Lux in the other times.
Comparative example 2 (light conditions different from those of example 2, light of low intensity was maintained)
A cultivation method for improving antithrombin activity of leeches comprises the following steps:
s1, setting a net cage: setting the length, width and height of the net cage to be 120cm, 60cm and 45cm respectively, arranging a 25cm multiplied by 25cm water outlet in the middle of the bottom of the net cage, and installing a 20-mesh escape-preventing net;
s2, seedling throwing: the seedling throwing density is 1500 g/box, namely 0.3 g/leech 5000 pieces/box;
s3, water quality management: the water for cultivation adopts well water, the water temperature is kept in the range of 22-28 ℃, the PH value is kept in the range of 6.8-7.2, and the water body in the cultivation box adopts micro-flowing water;
s4, bait feeding: mixing the nutrient solution with fresh pig blood according to a ratio of 7:3, uniformly mixing the components in proportion, then filling the mixture into a cleaned pig casing, and bundling two ends to ensure no leakage; putting the leeches into a leech culture box, sucking the leeches, and feeding the leeches again at intervals of 7 days; the nutrient solution is an aqueous solution of nutrient substances consisting of protein, compound vitamin, compound amino acid, compound enzyme and preventive medicine, and the concentration of the nutrient solution is 25mg/ml.
S5, controlling the illumination intensity: the day time at 8-20 points is kept at 0-10Lux, and the other time is 0Lux.
In the step S4, the nutrient substances consist of the following substances in mass ratio:
wherein the protein is soluble fish paste protein; the compound vitamin is a compound of vitamins A, B1, B2 and C, D, E; the compound amino acid is a compound of lysine, cysteine and histidine; the complex enzyme is a complex of cellulase, amylase, pectase, xylanase, mannanase, neutral protease and a carrier.
Wherein the preventive medicine is prepared from radix isatidis, rhizoma coptidis, phellodendron, radix scutellariae and hawthorn according to the mass ratio of 4:1:1:1:3 adding water and decocting with strong fire to obtain 150mg/ml medicinal liquid.
Comparative example 3 (higher intensity illumination was maintained, unlike example 2 illumination conditions)
A cultivation method for improving antithrombin activity of leeches comprises the following steps:
s1, setting a net cage: setting the length, width and height of the net cage to be 120cm, 60cm and 45cm respectively, arranging a 25cm multiplied by 25cm water outlet in the middle of the bottom of the net cage, and installing a 20-mesh escape-preventing net;
s2, seedling throwing: the seedling throwing density is 1500 g/box, namely 0.3 g/leech 5000 pieces/box;
s3, water quality management: the water for cultivation adopts well water, the water temperature is kept in the range of 22-28 ℃, the PH value is kept in the range of 6.8-7.2, and the water body in the cultivation box adopts micro-flowing water;
s4, bait feeding: mixing the nutrient solution with fresh pig blood according to a ratio of 7:3, uniformly mixing the components in proportion, then filling the mixture into a cleaned pig casing, and bundling two ends to ensure no leakage; putting the leeches into a leech culture box, sucking the leeches, and feeding the leeches again at intervals of 7 days; the nutrient solution is an aqueous solution of nutrient substances consisting of protein, compound vitamin, compound amino acid, compound enzyme and preventive medicine, and the concentration of the nutrient solution is 25mg/ml.
S5, controlling the illumination intensity: the illumination intensity of the environment is controlled as follows: the day time is 8-20 points and kept at 150-200Lux, and the other time is 0Lux.
In the step S4, the nutrient substances consist of the following substances in mass ratio:
wherein the protein is soluble fish paste protein; the compound vitamin is a compound of vitamins A, B1, B2 and C, D, E; the compound amino acid is a compound of lysine, cysteine and histidine; the complex enzyme is a complex of cellulase, amylase, pectase, xylanase, mannanase, neutral protease and a carrier.
Wherein the preventive medicine is prepared from radix isatidis, rhizoma coptidis, phellodendron, radix scutellariae and hawthorn according to the mass ratio of 4:1:1:1:3 adding water and decocting with strong fire to obtain 150mg/ml medicinal liquid.
Examples 1-4 and comparative examples 1, 2 were tested for antithrombin activity, survival rate, weight gain rate:
antithrombin activity assay method:
directly beating Hirudo after 60 days of cultivation into Hirudo pulp, accurately weighing Hirudo pulp about 1g, accurately adding 5ml of 0.9% sodium chloride solution with a 5ml large-belly pipette, fully stirring and mixing, covering a plug, leaching for 30 minutes, shaking from time to time, placing into a centrifuge, and centrifuging for 5 minutes at a rotation speed of 2000 r/min. The solution was poured into another 10ml centrifuge tube and placed in a centrifuge and centrifuged at 2000r/min for 5 minutes. 100. Mu.l of the supernatant was precisely measured with a 100. Mu.l syringe, placed in a small test tube of 9X 110mm, 200. Mu.l of a tris (hydroxymethyl) aminomethane hydrochloride buffer (prepared as a coagulate) containing 0.5% (bovine) fibrinogen (calculated as a coagulate) was added with a 500. Mu.l syringe, shaken well, and placed in a water bath (37.+ -. 0.5 ℃ C.) of a disintegrator for 5 minutes. A100. Mu.l sample injector was used to drop 56 units or 43 units of thrombin solution (drop every 1 minute, 5. Mu.l each time) until clotting, and the volume of thrombin solution consumed was recorded as follows:
wherein: u-containing antithrombin activity units per 1g, U/g;
c1-concentration of thrombin solution, U/ml;
c2-concentration of sample solution, g/ml;
v1-volume of thrombin solution consumed, μl;
v2-amount of sample solution added, μl.
The antithrombin activity, survival rate, and weight gain rate test results for examples 1-4 and comparative examples 1, 2 were as follows:
| antithrombin Activity (u/g) | Survival rate (%) | Weight gain Rate (%) | |
| Example 1 | 129 | 95 | 433 |
| Example 2 | 152 | 98 | 526 |
| Example 3 | 133 | 91 | 439 |
| Example 4 | 145 | 96 | 502 |
| Comparative example 1 | 90 | 65 | 133 |
| Comparative example 2 | 80 | 78 | 166 |
| Comparative example 3 | 92 | 80 | 172 |
By comparison of experimental data, the culture period of the invention is 60 days, the culture leech antithrombin reaches 129-152u/g, the survival rate reaches 91-98%, the weight gain rate reaches 433-526%, and all the data are obviously superior to the traditional culture method.
The above description is only of the preferred embodiments of the present invention and is not intended to limit the present invention, but it should be understood that any modifications, equivalents, improvements, etc. falling within the spirit and principles of the present invention will fall within the scope of the present invention.
Claims (7)
1. A cultivation method for improving antithrombin activity of leeches comprises the following steps:
s1, setting a net cage;
s2, seedling throwing;
s3, water quality management: the water for cultivation adopts well water, the water temperature is kept in the range of 22-28 ℃, the pH value is kept in the range of 6.8-7.2, and the water body in the cultivation box adopts micro-flowing water;
s4, bait feeding: mixing the nutrient solution with fresh pig blood according to a ratio of 5-10:2-5, then filling into a cleaned pig casing, and bundling two ends to ensure no leakage; putting the leeches into a leech culture box, sucking the leeches, and feeding the leeches again at intervals of 7 days;
s5, controlling the illumination intensity: the illumination intensity of the environment is controlled to be 20-50Lux at 8-20 points per day, and the illumination intensity of the environment is controlled to be 0Lux at other times;
in the step S4, the nutrient solution is an aqueous solution of nutrient substances consisting of protein, compound vitamin, compound amino acid, compound enzyme and preventive medicine, and the concentration of the nutrient solution is 20-30mg/ml;
the nutrient substance consists of the following substances in percentage by mass:
proteins 85-95
Vitamin 2-5 complex
Complex amino acids 1-5
Complex enzyme 1-5
1-4 parts of a preventive medicine;
wherein the protein is soluble fish paste protein; the compound vitamin is a compound of vitamins A, B1, B2 and C, D, E; the compound amino acid is a compound of lysine, cysteine and histidine; the complex enzyme is a complex of cellulase, amylase, pectase, xylanase, mannanase, neutral protease and a carrier;
wherein the preventive medicine is prepared from radix Isatidis, coptidis rhizoma, cortex Phellodendri, scutellariae radix, and fructus crataegi at mass ratio of 3-5:0.5-1.5:0.5-1.5:0.5-1.5:2-5, adding water, decocting with strong fire, and making into medicinal liquid with concentration of 120-180 mg/ml.
2. The method for raising antithrombin activity of leech as claimed in claim 1, wherein in said step S1, the length, width and height of the net cage are 120cm, 60cm and 45cm respectively, and a water outlet of 25cm x 25cm is provided at the middle position of the bottom of the net cage, and a 20 mesh escape preventing net is installed.
3. The method according to claim 1, wherein in the step S2, the seedlings are placed at a density of 1500 g/box, i.e., 0.3 g/strip of leech 5000 pieces/box.
4. The method according to claim 1, wherein in the step S4, the nutrient substances comprise the following substances in mass ratio:
protein 90
Vitamin complex 3.5
Complex amino acid 2.5
Complex enzyme 2.5
Preventive medicine 1.5.
5. The method for improving antithrombin activity of leech as claimed in claim 1, wherein in the step S4, the preventive medicine is radix Isatidis, rhizoma Coptidis, cortex Phellodendri, radix Scutellariae, fructus crataegi, in a mass ratio of 4:1:1:1:3 adding water and decocting with strong fire to obtain 150mg/ml medicinal liquid.
6. The method according to any one of claims 1 to 5, wherein in step S4, the nutrient solution and fresh pig blood are mixed according to a ratio of 7:3, mixing evenly.
7. The method according to any one of claims 1 to 5, wherein in the step S5, the environmental light intensity is controlled as follows: 40Lux in the morning, 25Lux in the afternoon and 0Lux in the other times.
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| FR2834292A1 (en) * | 2002-01-03 | 2003-07-04 | Ricarimpex | Extracts from bacterial symbionts of leeches, useful e.g. as anticoagulant, antiinflammatory or thrombolytic agents, prepared from cultured digestive tract bacteria |
| CN105594666A (en) * | 2015-12-25 | 2016-05-25 | 贵州信邦制药股份有限公司 | Breeding method for fast increasing activity of antithrombase of leeches |
| CN105707015A (en) * | 2016-03-03 | 2016-06-29 | 王文渊 | Method for feeding bait to medicinal blood-sucking leeches |
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| CN105558313A (en) * | 2015-12-25 | 2016-05-11 | 贵州信邦制药股份有限公司 | Raising bait for sucking-blood leeches as well as preparation method and application of raising bait |
| CN106577372B (en) * | 2016-11-04 | 2019-04-16 | 宁波大学 | A kind of overwintering and artificial fecundation method of sea otter |
| CN106804491A (en) * | 2016-12-14 | 2017-06-09 | 五河县茂源水蛭生态养殖专业合作社 | A kind of cultural method of leech |
| CN114557298A (en) * | 2022-02-28 | 2022-05-31 | 重庆多普泰制药股份有限公司 | Opening method for artificially breeding young leeches of Japanese medicina |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| FR2834292A1 (en) * | 2002-01-03 | 2003-07-04 | Ricarimpex | Extracts from bacterial symbionts of leeches, useful e.g. as anticoagulant, antiinflammatory or thrombolytic agents, prepared from cultured digestive tract bacteria |
| CN105594666A (en) * | 2015-12-25 | 2016-05-25 | 贵州信邦制药股份有限公司 | Breeding method for fast increasing activity of antithrombase of leeches |
| CN105707015A (en) * | 2016-03-03 | 2016-06-29 | 王文渊 | Method for feeding bait to medicinal blood-sucking leeches |
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