CN85101294A - From the ripe fish of property, obtain the method for the property product that is suitable for being fertilized - Google Patents
From the ripe fish of property, obtain the method for the property product that is suitable for being fertilized Download PDFInfo
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- CN85101294A CN85101294A CN85101294.9A CN85101294A CN85101294A CN 85101294 A CN85101294 A CN 85101294A CN 85101294 A CN85101294 A CN 85101294A CN 85101294 A CN85101294 A CN 85101294A
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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
The present invention be described in natural spawning season of fish irrelevant during in, from the sexual maturity fish, obtain the method for the sexual product that is suitable for being fertilized.Adopt this method, the actual maturing stage is measured by the property surveyed product (ovum or sperm); The fish that ripe fish can not be ovulated with prematurity still separates; Inject compound with hormonal action to the fish body; After the ovulation, the property obtained product from this fish body or its environment.
According to method of the present invention, with nonapeptide-C
1-4Alkylamide or decapeptide amide compound and salt thereof or metal complex inject the fish body.
Description
The present invention be described in natural spawning season of fish irrelevant during in, from the sexual maturity fish, obtain the method for the property product that is suitable for being fertilized.
We know, the sexuality of raun and milter is regulated by the gonadotropin that the adenohypophysis of fish discharges.The form of this parahormone restriction sexual gland and the formation of function, the maturation of sexual gland, and the secretion of active (generation of ovum and sperm) (sexual gland) and gonadal hormone takes place in the gamete of sexual gland.
In the individuality of two kinds of sexes, adenohypophysis all discharges same two kinds of gonadotropin.One of them is called the FSH(follicle stimulating hormone), another is called the LH(luteinizing hormone), perhaps be called the ICSH(interstitial cell-stimulating hormone according to its effect in male body).
We also know, these two kinds of hormones do not show the specificity to sex, because same FSH is influential to the activity that produces gamete in male body and the female body, and same LH or ICSH will induce in the male body secretion of oestrogenic hormone in the androgenic secretion and female body or progesterone.
FSH and LH are glycoprotein, and they show very strong species specificity to fish.
(brain) hypophysis of fish makes the many telotismizations of endocrine system of fish, should also have some coordination function by (brain) hypophysis between endocrine system and nervous system, and this mainly rolls into a ball by autonomic nuclear in the hypothalamus zone of diencephalon and realizes.In addition, (brain) hypophysis is regulated the cyclostage under the assistance with species specificity trophic hormone.
Under normal operation, the ovulation of fish occurs among certain ichthyomorphic egg-laying environment.Ripe ovum is not ovulated automatically, and they still were in long or short latent period, forms until suitable condition.This reproduction adaptive character of fish is that hypophysis and the central nervous system by fish jointly controls.In latent period, the body of fish is in poised state, and the ability that discharges gonadotropin is very low, therefore, only has very a spot of gonadotropin at this moment to be released to blood.The lay eggs correct time of condition of the best of measuring fish is impossible because the condition of laying eggs be formed on during certain section in (as late spring or early summer) more or less have contingency.This makes its cyclostage can adapt to external environment with regard to the regulation mechanism that means fish reproduction.This adjusting in the fish body is by the hypothalamus one hypophysis system coordination of fish.
Research according to recent years it has been recognized that, neurohormone is regulated the release of some trophic hormone in the adenohypophysis.Neurohormone comes out from nerve ending, passes to some nuclear group place of hypothalamus, is absorbed by so-called portal vein capillary immediately.Thus, these neurohormones directly arrive adenohypophysis Dou Chu by blood.
Be oligopeptides or polypeptide with regard to the neurohormone so far.Induce the neurohormone of trophic hormone secretion to be called " releasing factor " (RF), and the neurohormone of inhibition trophic hormone secretion is called " inhibiting factor " (IF).According to nearest nomenclature,, use " releasing hormone " (RH) or " release inhibting hormone " (RIH) this class term for the neurohormone of known its 26S Proteasome Structure and Function.
All neurohormones all have such feature, i.e. their effects in removing the hypophysis animal body are similar to observed effect after adding trophic hormone.
The basophilic cell of fish hypophysis is seasonal variety, and its quantity reduced between the egg-laying period.In the procedure of adaptation to environment, produced releasing hormone (RH), these hormones flow into hypophysis by blood, and regulating wherein, trophic hormone is discharged into blood.Thus, fish obtains existing the suitable external world information of condition of laying eggs.The result, luteinizing hormone releasing hormone (LH-RH) (luteinizing hormone adjusting hormone) activates hypophysis, and make it in blood, discharge gonadotropin, thereby begin the process of laying eggs (Bretton B and Wei Er cl, Paris, and academy of sciences's newspaper (C.R.Acad.Sc., Paris), 1973 277 phases, 2061～2064 pages).
Be delivered to endocrine system by this way by the sensory nerve impulsion that produces in the place of laying eggs, this mechanism has activated to be in always forces preclinical genital system.According to the viewpoint of filial generation existence, can detect best external environment (the lattice Bilsky is with above-mentioned) this moment.
Neurohormone has highly characteristic between kind.The order of luteinizing hormone and releasing hormone is different from mammal in the fish body.Therefore, when two kinds of hormones exchange, there is not usefulness.
Owing to found the hypophysis technology, and this technology worldwide enlarges use, and artificial induction's breeding of fish has become possible (the lattice Bilsky is with above-mentioned).
The hypophysis of collecting from the common bream of ripe or approaching maturation in order to lay eggs with hormone induction through after the suitable preservation, is commonly used.Other natural or synthetic hormone of being tried out does not so far then still have consistent experimental result.
The common breed of this common bream, reserves from all parts of the world are very big.Therefore can collect the hypophysis of q.s easily, to satisfy the production purpose.The hypophysis of common bream also can be successfully used to breed the fish of other several breed.The fingerling of edge far away (as sturgeon) belongs to exception in the classification.This class fish is then used the hormone suspension for preparing by its hypophysis of the same race.
Collection pituitary and preservation all use the same method, and be irrelevant with its donor kind.Hypophysis should be collected from ripe (being sexually matured as far as possible) fish, but this possibility is very little.Hypophysis is littler in the small fish body because when using by weight, so when small fish is collected hypophysis on one's body, need a large amount of donor fishes.
According to known method, the hypophysis acetone drying.Soak 3 times in acetone, after each 8 to 12 hours, hypophysis is sloughed water and fat.After the sclerosis, hypophysis at room temperature dry 24 hours, acetone is evaporated out simultaneously.Acetone is without detriment to the short sexual gland active component of hypophysis.Gan Zao hypophysis can be stored the long time and not loss in the above described manner.
In order to activate gonadotropin, body of gland is ground in porcelain mortar, hormone is dissolved in the normal saline solution of fish (containing 0.65%NaCl).
Hormone solution will inject the muscle of fish, comes out or any other loss to avoid refluence.If small fish or fish of flaccid muscles, parenteral solution will inject the body cavity of fish.The pituitrin of injection is with identical in the hormone role of finding the release of egg-laying environment macrura reevesii itself, and they all cause the process of laying eggs.
Although the hypophysis technology is extensively adopted in the breed of fish, be not all can prove effective in all cases.Its main cause is as follows:
A) owing to not knowing the maturing stage of donor fish (collecting hypophysis on one's body), so do not know the gonadotropin content of hypophysis from these fishes yet.Even if in addition various restrictions when therefore calculating usage amount, its effect is still uncertain.
B) hormone of common bream hypophysis is only effective to the fingerling that non-edge far away is gone up in general cyprinid fish and classification.Therefore, see very important several fishes from economic point of view, its reproduction rate is very low or irreproducible at all.
C) use the hypophysis of common bream (or any other kind), under big working condition, can only obtain the effect of luteinizing hormone.By this method, can not produce the effect of follicle stimulating hormone, this may be because a) in the section due to the reason mentioned.
D) owing to there is not the effect of follicle stimulating hormone, the hypophysis technology can only be used to induce the steck ovum of the maturation of ovum.
E) adopt the hypophysis technology, artificial induction's breeding can only be carried out at spawning season, and the possibility that this means breeding only limits in 1 year within these few days.Therefore the use of production equipment also is restricted.
F) can only from jejune fish, collect hypophysis in a large number.The feature of this hypophysis is that short sexual gland activity is low.Only kill valuable donor fish, just may from ripe fish, collect hypophysis.
The present invention seeks to describe in detail a method, this method can obtain to be used for the property product of natural insemination or artificial insemination from any ripe fingerling.
The present invention is based on following understanding, if promptly be applied to the organism of ripe fish with the derivative of new gonadotropin, then above-mentioned purpose can reach fully.We have found that, can successfully replace with above-claimed cpd from oozy follicle stimulating hormone/luteinizing hormone one releasing hormone of the hypothalamus of fish.In other words, inject generation and the ovulation that these compounds can successfully be induced sperm and ovum.
As mentioned above, the present invention be disclosed in natural spawning season of fish irrelevant during in, from the sexual maturity fish, obtain the method for the property product that is suitable for being fertilized.Adopt this method, the actual maturing stage of the property product (sperm and ovum) of sexual maturity fish is measured by detection method; Still prematurity and the fish that can not ovulate branch away from ripe fish; The compound administration that plays hormonal action is in the fish body; After ovulation, acquired product from fish body or its environment.
According to the present invention, will have the nonapeptide-C of general molecular formula (I)
1-
4Alkyl-acid amides and decapeptide acid amides and salt thereof and metal complex (as the compound that plays hormonal action) are introduced in the fish body, and dosage is 0.1 microgram～5 milligram, and 2～500 micrograms are relatively good, preferably 5～100 micrograms.Still can not ovulate if fish is immature, above-mentioned dosage divides secondary to inject at least, is no more than ten secondaries at most.Wherein last dosage is identical or preferably once big 1.5 times than preceding at least with last time at least.
General formula (I)
Pyroglutamyl-histidyl--tryptophanyl-seryl-tyrosyl-
X
1-X
2-X
3-prolyl-X
4
Wherein, X
1Be the D-isomer of glycyl or any natural or synthesizing amino acid residue,
X
2Be illustrated in the L-amino acid residue, L-phenylalanyl or the L-tryptophanyl that contain 1～4 carbon atom on the side chain,
X
3The expression side chain is to contain 1～4 carbon atom alkyl or side chain is the L-amino acid residue that contains 2～4 carbon atom alkoxy-acid amides,
X
4Be glycine-acid amides or 1～4 carbon atom alkyl-amide groups.
The abbreviation of above-mentioned molecular formula meets approval of chemistry of peptides circle and published nomenclature, and for example " journal of biological chemistry " (volume was 527 pages in 1966 the 241st, and volume was 977 pages in 1972 the 247th) were once reported this nomenclature.
This compound is nontoxic, even if excessive use does not also have the effect of paying.
The fish that can not ovulate if handle immature, frequency injection depends on the time length of this fish apart from the mature ovulation phase.Shorter during this period of time, frequency injection is fewer.
The fish that can ovulate for maturation if genus is male, can move when then its sperm contacts with water; If genus is female, then its cell nucleus is attached to the edge of egg cell.
According to method of the present invention, among the compound of general formula (I), use decapeptide compound that special benefits is arranged with formula II and formula III:
Pyroglutamyl-histidyl--tryptophanyl-seryl-tyrosyl-D-phenylalanyl-leucyl-glutamine-prolyl-glycine-NH
2
（Ⅱ）
Pyroglutamyl-histidyl--tryptophanyl-seryl-tyrosyl-D-phenylalanyl-tryptophanyl-leucyl-prolyl-glycine-NH
2
（Ⅲ）
The employed compound with general formula (I) of preparation the inventive method can adopt technology for solid phase synthesis of peptide (the meeting Chi of Merifield R.B. american chemical man association, volume was 2149～2151 pages in 1963 85).According to the structure of required compound, if the preparation alkyl peptide amide then preferentially uses benzhydrylamine resin.Adopt dicyclohexyl carbonizer's imines (DCC), N, N '-diisopropyl carbodiimides (DIC) or Acibenzolar method can be attached to amino acid on the above-mentioned resin, as their N-α-uncle-Ding oxygen carbonyl acyl derivative.Final products usable acid or alkaline lysis technology are separated from solid support.
According to the chemical property of various aminoacid ingredients, the synthetic step by step and fragment condensation by appropriate combination, available suitable protection amino acid prepares needed final products with general formula (I).
The major advantage of the inventive method is as follows:
A) this method makes the breeding with common bream fingerling of edge far away in classification become possibility.Owing to lack species specific hypophysis, this class fish did not breed in the past.
B) this method can be used in very wide ecological limit because general formula (I) though compound under diverse biotic factor, also can bring into play physiological action.
C) this method can be bred fingerling in season at non-natural spawning, can make full use of production equipment simultaneously.
D) this method can be eliminated because of hypophysis consumption inaccuracy and cause anovular situation.
E) active ingredient of the inventive method use can synthesize in the laboratory, therefore can overcome the unsettled shortcoming of the peculiar effect of known hypophysis technology.
F) effect of the synthetic hormone of the present invention's use is identical with the effect of the gonadotropin of fish release itself.
G) therefore specificity between the synthetic hormone used of this method does not have kind goes up the big fingerling of difference to classifying, and this hormone is also effective.
Following example present invention will be further described method, but do not limit protection domain.
Example 1
Artificial induction's elementary corpuscle sturgeon (Acipenser ruthenus L.)
From the population in a certain place of production, select ripe fish, then it is transported to breeding place.Milter and raun are separated, measure the maturing stage of female fish-egg.If cell nucleus attached to the edge of egg cell, then think fish maturation can ovulate.The sperm of milter needn't check, because in fact the property product of male body and female body is in the same maturing stage in the population of a certain place of production.If fish maturation can ovulate, every fish (comprising milter) is once injected the decapeptide of 70 micromolar formulas (II).
Still can not ovulate if fish is immature, then this fish should be kept under the temperature of natural spawning, and should inject above-claimed cpd, each consumption 10 micrograms, secondary at least, maximum ten secondaries move on to the edge with cell nucleus from cell centre and are as the criterion.Number of processes depended on " ovulation the is ripe " phase compares the actual period of property product maturation.This maturation continues 100 day degrees at least, is no more than 630 day degrees at most.Temperature range is 12～18 ℃.(day degree equals the mean temperature that fate is multiplied by ripening period institute water).
Inject last dosage after 24～32 hours, ovulation begins to carry out.Afterwards can be in 40～60 minutes obtaining property product.
Example 2
The artificial induction breeds common carp (Cyprinus Carpio L.)
With the breeding fish of maturation in 1 year any season and arbitrary temp under from its colony, choose, be transported to the breeding place.The fish that separates two kinds of sexes.Then according to maturing stage of example 1 property measured product.If the ovulation maturation of fish is then once injected the decapeptide compound of 70 microgram formulas (III), comprise milter.
Still immature and the fish that can not ovulate should remain under the temperature of natural spawning, should inject above-claimed cpd simultaneously, each dosage 5 micrograms at least 2 times, maximum 15 times, move on to edge with cell nucleus from the egg cell center and are as the criterion.The number of times that injects processing depended on " ovulation the is ripe " phase compares the actual maturing stage of property product.This maturation continues 100 day degrees at least, 750 day degrees at the most, and temperature range is 20～40 ℃.
Last inject 240～260 o'clock degree after, ovulation process takes place.Fissility product in 40～60 minutes then.
Example 3
The artificial induction breeds perch (Perca fluviatilis L.)
From a certain population, select ripe fish, be transported to breeding place.According to the maturity of example 1 property measured product, but the fish of not separated two kinds of sexes.To the fish (comprising milter) that maturation can be ovulated, be the decapeptide compound treatment of 20 microgram formula II with dose.
Still immature and fish that can not ovulate should remain under the natural spawning temperature, the dosage with 2 micrograms injects above-claimed cpd at every turn, at least 2 times, maximum 10 times, moves on to the edge with the cell nucleus of egg cell from cell centre and is as the criterion.Number of processes depended on " ovulation the is ripe " phase compares the actual maturing stage of property product.Maturation continues 100 day degrees at least, to few situation, continues nearly 400 day degrees, and water temperature range is 12～16 ℃.
After injecting, last dosage promptly laid eggs in 24 to 36 hours.Perch more easily carries out artificial spawning in the pond.In this case, there is no need the fissility product.
Example 4
The artificial induction breeds jack mackerel (Trachurus trachurus L.)
From the November the jack mackerel alives that catch in the Adriatic, selecting the fish height is 240 millimeters sample, it is transported to breed the place.(fish of this specification, sexual maturity).Formula (II) compound of 10 micrograms dose is once injected every fish.Ovulate after 24 hours, water temperature is 18～24 ℃.I.e. obtaining property product after two hours.
For jack mackerel, there is no need to measure maturing stage and sex, because Cheng Shu fish non-quantitative in fact.
Example 5
The artificial induction breeds pike (Esox lucius L.)
Among a certain population, select the sexual maturity fish, transport to the breeding spot then.The fish that separates two kinds of sexes is according to the maturing stage of example 1 property measured product.The ripe fish (comprising milter) that can ovulate carries out single treatment with the decapeptide compound of the formula (II) of 100 micrograms dose.
Still immature and fish that can not ovulate should remain under the natural spawning temperature, should inject the compound of the above-mentioned appointment of 5 micrograms at every turn, inject at least 2 times, be no more than at most 10 times, move on to edge with the cell nucleus of egg cell from cell centre and be as the criterion.Number of processes depended on " ovulation the is ripe " phase compares the actual maturing stage of property product.Maturation continues 70 day degrees at least, reaches 350 day degrees under few situation, and water temperature is 8～14 ℃.
Example 6
The artificial induction breeds white chain (Hypophthalmichthys molitrix Val.)
From the colony of this fish, select the sexual maturity fish, it is transported to the breeding place.Separate milter and raun.According to the maturing stage of example 1 property measured product.The ripe fish (comprising male fish) that can ovulate carries out single treatment with the decapeptide compound of the formula III of 100 micrograms dose.
Still immature and fish that can not ovulate should remain under the temperature of natural spawning, and above-claimed cpd should be injected the fish body, each dosage is 10 micrograms, injects secondary at least, maximum ten times, moves on to edge with cell nucleus in the egg cell from cell centre and is as the criterion.Number of processes depended on " ovulation the is ripe " phase compares the actual maturing stage of property product.This maturation continues 150 day degrees at least, assigns 850 day degrees in few situation, and water temperature range is 20～26 ℃.
Last potion is ovulated, then the fissility product after injecting 240～260 o'clock degree.
In the literary composition,
The formula I:
Glp-His-Trp-Ser-Tyr-X
1-X
2-X
3-Pro-X
4
The formula II:
Glp-His-Trp-Ser-Tyr-D-Phe-Leu-Gln-Pro-Gly-NH
2
The formula III:
Glp-His-Trp-Ser-Tyr-D-Phe-Trp-Leu-Pro-Gly-NH
2
First page the 7th section referring to " present situation that the neurohormone in fish sexual cycles is regulated and biotechnology are bred the present situation of hormonal effects in the fish the artificial induction ", the 8th page, outstanding Bilsky original work,, Leningrad, publisher: Gos in 1966.University.
Claims (1)
- A method, with natural spawning season of fish irrelevant during in, from the sexual maturity fish, obtain the method for the property product that is suitable for being fertilized.Adopt this method, the actual maturing stage is measured by the property surveyed product (ovum or sperm); The fish that ripe fish can not be ovulated with prematurity still separates; Inject compound with hormonal action to the fish body; After the ovulation, the property obtained product from this fish body or its environment; The feature of this method is the nonapeptide-C of general formula (I) 1-4Alkylamide and decapeptide acid amides and salt thereof or its metal complex are the compound with hormonal action,Pyroglutamyl-histidyl--tryptophanyl-seryl-cruel aminoacyl-X 1-X 2-X 3-prolyl-X 4(I)Wherein, X 1Be glycyl, or the D-isomer of any natural or synthesizing amino acid residue,X 2Be illustrated in the L-amino acid residue that contains 1～4 carbon atom on the side chain, or L-phenyl alanyl, or L-look acyl residue,X 3The expression side chain is to contain 1～4 carbon atom or side chain is the L-amino acid residue that contains 2～4 carbon atom alkoxy-acid amides,X 4Be glycine-acid amides or 1～4 carbon atom alkyl-amide groups,These compounds are injected in fish bodies, and dosage is 0.1 microgram to 5 milligram, and 2 to 500 micrograms are better, are preferably 5 to 100 micrograms; For the immature and fish that can not ovulate still, above-mentioned dosage is divided into two parts at least, be divided into 12 parts at the most to inject the fish bodies, wherein last doses at least with its before a identical or preferably big 1.5 times than preceding portion at least.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN85101294A CN1014672B (en) | 1985-04-01 | 1985-04-01 | Method for obtaining sexual product suitable for fertilization from sexual mature fish |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN85101294A CN1014672B (en) | 1985-04-01 | 1985-04-01 | Method for obtaining sexual product suitable for fertilization from sexual mature fish |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN85101294A true CN85101294A (en) | 1987-01-24 |
| CN1014672B CN1014672B (en) | 1991-11-13 |
Family
ID=4791752
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN85101294A Expired CN1014672B (en) | 1985-04-01 | 1985-04-01 | Method for obtaining sexual product suitable for fertilization from sexual mature fish |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1014672B (en) |
-
1985
- 1985-04-01 CN CN85101294A patent/CN1014672B/en not_active Expired
Also Published As
| Publication number | Publication date |
|---|---|
| CN1014672B (en) | 1991-11-13 |
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