CN109280090A - One kind having the active aspongopus class Pentatomidae Insects extraction method of polysaccharides of warming kidney and enhancing body and application - Google Patents
One kind having the active aspongopus class Pentatomidae Insects extraction method of polysaccharides of warming kidney and enhancing body and application Download PDFInfo
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Abstract
There is the active aspongopus class Pentatomidae Insects extraction method of polysaccharides of warming kidney and enhancing body the invention discloses one kind and application, step are: A, drying aspongopus class insect body, crush, with petroleum ether refluxing extraction, filtering, solid-liquor separation；The grease dregs of a decoction must be removed；B, the dregs of a decoction are volatilized, with distilled water refluxing extraction, filtering, merging filtrate is concentrated, obtains water total extract；C, distilled water is added slightly to dilute water total extract, it is stirring while adding with alcoholic solution precipitate polysaccharides, it stands, filtering obtains Insect Polysaccharide alcohol hypostasis；D, polysaccharide polyol hypostasis is diluted with water, dissolved, isolated and purified with DEAE- sephadex cellulose chromatography, rinsed with distilled water, sodium chloride solution, obtain aspongopus class Insect Polysaccharide.Application of the polysaccharide in the preparation treatment or prevention deficiency of yang, impotence, erectile dysfunction drug is further related to, easy to implement the method, easy to operate, purity is up to 95% polysaccharide compound, has powerful warming kidney and enhancing body, antifatigue, recovery body immunity effect.
Description
Technical field
The invention belongs to Pentatomiddae aspongopus class Insect Polysaccharide extractive technique fields, are more particularly to a kind of with warming kidney and enhancing body
Active aspongopus class Pentatomidae Insects extraction method of polysaccharides further relates to a kind of with the active aspongopus class Pentatomiddae elder brother of warming kidney and enhancing body
The purposes of worm polysaccharide.
Background technique
Erectile dysfunction is one of most common sex dysfunction of male, current research suggests that the age is erection function
Strongest independent factor in obstacle Related Risk Factors.Male erectile is a complicated process, be related to brain, hormone, emotion,
The various problems such as nerve, muscle, blood vessel.
Aspongopus is the dry of Semiptera Pentatomiddae aspongopus subfamily insect aspongopus Aspongopus chinensis Dallas
Dry body, the black pocket worm of alias, the black stinkbug of melon, smelly wind worm, wind bar worm etc. are Chinese unique wheat." Chinese Pharmacopoeia " (version in 2015) is recorded
Aspongopus has regulating qi-flowing for relieving pain, and effect that middle benefit gas is supporing yang is used for stomach cold distending pain, stomachache due to emotional depression and the hyperactive liver-qi attacking the stomach, impotence due to deficiency of the kidney, soreness of waist and knee joint.Begin
It is loaded in Compendium of Material Medica: " salty temperature is nontoxic, is used for diaphragm gastral cavity stagnant gas, and spleen-kidney deficiency strengthens person's Yuanyang ".It is recorded in " book on Chinese herbal medicine is newly organized ": " worm
In to good person, also the object of emerging sun, weak most suitable to help ".Civil is even more to have: " rich man eats pilose antler, and people out of funds eats wind bar worm "
Say.Although establishing-Yang effect of aspongopus is on the books repeatly, the experimental study of its establishing-Yang effect is had not been reported.In conjunction with establishing-Yang row
To be closely related with the fatigue of body, the object of the invention provides a kind of Pentatomiddae aspongopus class Insect Polysaccharide extracting method and application,
And the Insect Polysaccharide toxicity is higher than commercially available Yang invigorating medicine drug effect, Small side effects.Western medicine silaenafil or phentolamine on the market
The curative effect phase is short, and taking for a long time has certain side effect, palliative, and generates the influence of patient's dependence.
Summary of the invention
There is warming kidney and enhancing body, antifatigue, recovery body's immunity activity the purpose of the invention is to provide a kind of
Aspongopus class Pentatomidae Insects polysaccharide extracting method, easy to implement the method, easy to operate, which can prepare
Prevent and treat the Insect Polysaccharide of the deficiency of yang, the aspongopus class Pentatomiddae of impotence.And have to the rat of physiology, pathology, mouse obvious
Yang invigorating functions, and drug effect has potential industry development value, to Yang invigorating medicine better than general medea drug effect in the market
Research and development are of great significance.
Another object of the present invention is to be the provision of one kind to have warming kidney and enhancing body, antifatigue, recovery immunity of organism function
It can active aspongopus class Pentatomidae Insects polysaccharide answering in the preparation treatment or prevention deficiency of yang, impotence, erectile dysfunction drug
With.Using the action character of Chinese medicine multiple target point, the mechanism of corresponding erection function complexity, simultaneous treatment of principal and subordinate symptoms, curative effect is superior, cure rate
Height, integration of drinking and medicinal herbs take the Chinese medical extract preparation method of the treatment erectile dysfunction without obvious toxic-side effects for a long time.
In order to achieve the above purpose, the present invention with following technical proposals by being realized:
A kind of extracting method with the active aspongopus class Pentatomidae Insects polysaccharide of warming kidney and enhancing body comprising the steps of:
A, it by 50 DEG C of -80 DEG C of drying 10h-14h of aspongopus class insect body, crushes, cross 40 meshes, with petroleum ether or ether
Powder 2h-4h is infiltrated, dry powder complete swelling is made, carries out refluxing extraction, degreasing 2-6 times, 0.5~2h, filtering, liquid divide admittedly every time
From；Obtain the dregs of a decoction of removal grease.
B, the obtained dregs of a decoction of step A are volatilized into unrecovered oil ether, with distilled water immersion 2-4h, is impregnated with the dregs of a decoction completely, returned
Stream extracts 2-6 times, and 0.5~2h, filtering, solid-liquor separation, merging filtrate fling to solvent every time, is concentrated, obtains water total extract；
C, add distilled water slightly to dilute the water total extract that step B is obtained, with alcoholic solution precipitate polysaccharides, it is stirring while adding extremely
Determining alcohol is protected from light to 50~80%, stands 12h-24h, filtering obtains Insect Polysaccharide alcohol hypostasis in solution.
D, water is added slightly to dilute, dissolve the polysaccharide polyol hypostasis that step C is obtained, with DEAE- sephadex cellulose column layer
Analysis isolates and purifies, and is rinsed with distilled water, sodium chloride (NaCl) solution, collects eluent, and filtering volatilizes solvent, obtains a kind of tool
There are warming kidney and enhancing body, antifatigue, the recovery active aspongopus class Pentatomidae Insects polysaccharide of body immunity.
Aspongopus class insect in the step A is aspongopus, black pocket worm, Cyclopelta parva Distana, any without piercement bug.
Alcoholic solution in the step C is any in methanol solution or ethanol solution；
The condition being heated to reflux in the step B is 65~85 DEG C of progress.
The concentration of the ethanol solution is that percent by volume is 50~80%；
The concentration of the methanol solution is that percent by volume is 50~80%；
The elution volume is 10-20 times of column volume；
Eluent sodium chloride (NaCl) the solution wash-out concentration is 0.1-0.9mol/L.
The invention provides a kind of method for extraction and purification of aspongopus class Chinese medical extract, fragrant by nine by the method for alcohol precipitation
Thick many candies in worm medicinal material extract, and wherein the ingredients such as fat oil, deposition activity position Insect Polysaccharide pass through DEAE- for removal
Sephadex cellulose elutes Thick many candies, obtains the aspongopus polysaccharide that purity reaches 95% or more, changes extracting method health ring
It protects pollution-free, is eliminated as much as unwanted contributions, effective component purity is high.The Chinese medical extract has warming kidney and enhancing body, antifatigue, extensive
Body immunity function of answering a pager's call is active, and aspongopus class Pentatomidae Insects polysaccharide treats or prevents the deficiency of yang, impotence, erectile dysfunction in preparation
Application in drug.The invention is directed to Western medicine silaenafil on the market or the phentolamine curative effect phase is short, and taking for a long time has one
Determine side effect, it is palliative, and the influence of patient's dependence is generated, the present invention utilizes the action character of Chinese medicine multiple target point, corresponding
The mechanism of erection function complexity, simultaneous treatment of principal and subordinate symptoms, curative effect is superior, and cure rate is high, integration of drinking and medicinal herbs, takes for a long time without obvious toxic-side effects
Treatment erectile dysfunction Chinese medical extract preparation method.
There is one kind warming kidney and enhancing body, antifatigue, the recovery active aspongopus class Pentatomidae Insects polysaccharide of body immunity to exist
Preparation treats or prevents the deficiency of yang, impotence, the application in erectile dysfunction drug, the steps include:
1, the influence of Serum lactic acid content；
2, the influence of blood urea nitrogen；
3, to the influence of swimming time；
4, the influence of normal mouse mating ability；
5, the influence of deficiency of yang mouse mating ability；
6, the influence of deficiency of yang rat mating ability.
By above-mentioned experiment, treatment group's significant effect of the present invention is all effective to model of yang asthenia and intact animal.With bright
Aobvious tonifying kidney and strengthening yang is antifatigue, improves immunity of organisms function.
Compared with prior art, the present invention having the following advantages that and effect:
(1) Pentatomidae Insects are many kinds of, have a vast market foreground.
(2) it being compared in conventional extraction petroleum ether-ethyl acetate-n-butanol method, the present invention mentioned by petroleum ether-water-
The polyoses extract that the method for alcohol precipitation obtains, preparation method is simple, obtained extract removes other unwanted contributions, extraction process
Environmental protection, cost is relatively low.
(3) the Insect Polysaccharide extract removes most of other compositions and obtains purity by isolating and purifying operation and be up to
95% polysaccharide compound, polysaccharide are mainly made of monosaccharide such as mannose, glucose, rhamnose, galactolipins.
(4) the alcohol hypostasis Insect Polysaccharide that the present invention obtains has powerful warming kidney and enhancing body, antifatigue, recovery immunity of organism energy
Power effect.
(5) the Insect Polysaccharide extract drug effect is high, and toxicity is low, Small side effects, can be used as warming kidney and enhancing body, antifatigue, recovery
Body immunity drug candidate.
(6) the Insect Polysaccharide extract takes out insect oils, retains warming kidney and enhancing body, antifatigue, recovery immunity of organism activity
Higher Insect Polysaccharide extract avoids fat equal other side effects.
Specific embodiment
Embodiment 1:
Experimental material
Hydrocortisone, Chengdu end Kodak chemical reagent Co., Ltd (purity 98%)；Rhodiola kirilowii Regel capsule, Jiang Sukang
Edge medicine company limited liability company；Chinkuei shin chewan pills, Beijing Tongrentang Technology Development Co.ltd. Pharmaceutical Factory；Progynova (penta
Sour estradiol), Bayer HealthCare Co；Physiological saline, Binghu Shuanghe Pharmaceutical Co., Ltd., Wuhan；Lactic acid test
Box, urea nitrogen kit, Bioengineering Research Institute is built up in Nanjing.Female, male kunming mice, weight 18-22g are purchased from Hubei Province's reality
Test Animal Research Center；Female, male SD rat, weight 180-220g are purchased from Hubei Province Animal Experimental Study center；Ultrapure water:
The dedicated ultrapure water machine in the laboratory Water purifier.Electronic analytical balance: Precisa, model: XB220A；Refrigerated centrifuge:
German Eppendorf, model: 5804 R of Centrifuge；Microplate reader: U.S. Bio-Tek, model: Synergy 2.
It is a kind of with warming kidney and enhancing body, it is antifatigue, restore body immunity active aspongopus class Pentatomidae Insects polysaccharide
Extracting method the steps include:
A, aspongopus class insect dry, pulverize, cross 40 meshes, carried out with the petroleum ether of liquid-solid ratio 6:1 volume or ether
Refluxing extraction degreasing refluxing extraction 2 or 4 or 6 times, every time 0.5 or 1 or 1.5 or 2h, filtering；Solid-liquor separation obtains removal grease
The dregs of a decoction.
B, the obtained dregs of a decoction of step B are volatilized into unrecovered oil ether, with the distilled water refluxing extraction 2 or 4 of liquid-solid ratio 8:1 volume
Or 6 times, 0.5 or 1 or 1.5 or 2h, filtering, solid-liquor separation obtain water total extract every time；
C, the water extract for obtaining step C alcoholic solution precipitate polysaccharides, it is stirring while adding to determining alcohol in solution to 50
Or 60 or 70 or 80%, it is protected from light, stands 12 or 18 or 22 or for 24 hours, filtering obtains alcohol hypostasis aspongopus class Pentatomidae Insects polysaccharide.
D, water is added slightly to dilute, dissolve the polysaccharide polyol hypostasis that step C is obtained, with DEAE- sephadex cellulose column layer
Analysis isolates and purifies, and is rinsed with distilled water, sodium chloride (NaCl) solution, collects eluent, and filtering volatilizes solvent, obtains a kind of tool
There are warming kidney and enhancing body, antifatigue, the recovery active aspongopus class Pentatomidae Insects polysaccharide of body immunity.
The condition being heated to reflux in the step A is 65 or 70 or 75 or 80 or 85 DEG C of progress.
Aspongopus class insect in the step B be aspongopus, black pocket worm, Cyclopelta parva Distana, without piercement bug any one.
Alcoholic solution in the step C is any in methanol solution or ethanol solution；
It is 50 or 60 or 70 or 80% that the concentration of the step C ethanol solution, which is percent by volume,；
It is 50 or 60 or 70 or 80% that the concentration of the step C methanol solution, which is percent by volume,.
By examples detailed above, a kind of more pure Insect Polysaccharide is obtained, removes the impurity such as grease in insect bodies, this is mentioned
Take method easy to implement the method, easy to operate, environment protection health.Aspongopus has preferable warming kidney and enhancing body, antifatigue, recovery immunity of organism
The effect of function, but its grease contained will lead to the side effects such as obesity, and this method removes in aspongopus by simple method
Grease obtains the higher Insect Polysaccharide of pharmacological activity.
Embodiment 2:
There is one kind warming kidney and enhancing body, antifatigue, the recovery active aspongopus class Pentatomidae Insects polysaccharide of body immunity to exist
Preparation treats or prevents the deficiency of yang, impotence, the application in erectile dysfunction drug, the steps include:
1, the influence of Serum lactic acid content
Male mouse of kunming is randomly divided into 4 groups, every group animal 10, respectively blank control group (physiological saline), the positive
Control group (rhodiola kirilowii Regel capsule, 590mg/kg), each extract low dose group (1.4g/kg, based on crude drug), each extract
Middle dosage (7g/kg, based on crude drug) each extract high dose (14g/kg, based on crude drug) is administered 15 days.Last dose
30min posterior orbit endocanthion 20 μ L of blood sampling, do not bear a heavy burden after 30 ± 1 DEG C of water tank went swimming 10min, eye socket endocanthion blood sampling immediately 20
After μ L, rest 10min, 20 μ L of eye socket endocanthion blood sampling.EDTA is anticoagulant, and 2500rpm/min centrifugation takes supernatant, tests according to lactic acid
Kit tests lactic acid content.
Table 1: influence (n=10) of the polyoses extract to mouse swimming Plasma Before And After lactic acid content
2, the influence of blood urea nitrogen
Dosage, administration and the same 2.2.1 of grouping.After last dose 30min, do not bear a heavy burden in 30 ± 1 DEG C of water tank went swimming
90min, the blood sampling of rest 60min posterior orbit endocanthion, about 0.5ml.4 DEG C of refrigerators prevent 3h, and 2500rpm/min centrifugation takes supernatant, presses
Urea detection kit detects urea nitrogen content.
Table 2: influence (n=10) of the polyoses extract to mice plasma urea nitrogen content
3, to the influence of swimming time
Dosage, administration and the same 2.2.1 of grouping.After last dose 30min, by the weight bearing iron wire of mouse weight 5%, 30 ± 1
DEG C water tank went swimming, start timing, until the mouse 8s that sinks under water is not floating, record swimming time.
Table 3: influence (n=10) of the polyoses extract to mouse swimming time
4, the influence of normal mouse mating ability
Male mouse of kunming is randomly divided into 5 groups, every group animal 10, respectively blank control group (physiological saline), the positive
Control group (levodopa, 150mg/kg), each extract low dose group (1.4g/kg, based on crude drug), each extract middle dosage
(7g/kg, based on crude drug), each extract high dose (14g/kg, based on crude drug) are administered 30 days.Female mice with mate before
48h、
It is synchronous to be allowed to estrus time for 4h stomach-filling Estradiol Valerate (0.4mg/kg).Mouse in 20min is observed after stomach-filling 1h to hand over
It (captures incubation period with ability, capture number, mating incubation period, number of copulations).
Table 4: influence (n=10) of the polyoses extract to normal mouse mating ability
5, the influence of deficiency of yang mouse mating ability
Male mouse of kunming is randomly divided into 6 groups, every group animal 10, respectively blank control group (physiological saline), model
Group (physiological saline) positive controls (chinkuei shin chewan pills, 7g/kg), each extract low dose group (1.4g/kg, by crude drug
Meter), each extract middle dosage (7g/kg, based on crude drug), each extract high dose (14g/kg, based on crude drug) administration 30
It.Since administration the 4th day, except for the normal group, remaining group subcutaneous injection hydrocortisone 25mg/kg, one time a day, continuous 7
It, modeling is administered simultaneously.Female mice with mate before 48h, 4h stomach-filling Estradiol Valerate (0.4mg/kg), it is synchronous to be allowed to estrus time.
Mouse mating ability in 30min (capture incubation period, capture number, mating incubation period, number of copulations) is observed after stomach-filling 1h.
Table 5: influence (n=10) of the polyoses extract to deficiency of yang mouse mating ability
6, the influence of deficiency of yang rat mating ability
SD male rat is randomly divided into 6 groups, every group animal 10.Respectively blank control group (physiological saline), model group
(physiological saline) positive controls (chinkuei shin chewan pills, 7g/kg), each extract low dose group (1.4g/kg, based on crude drug),
Each extract low dose group (7g/kg, based on crude drug), each extract high dose (14g/kg, based on crude drug) administration 30
It.Since administration the 4th day, except for the normal group, remaining group subcutaneous injection hydrocortisone 25mg/kg, one time a day, continuous 7
It, modeling is administered simultaneously.Female mice cuts off ovary in the operation in first 14 days that mates, and Estradiol Valerate (20 μ are subcutaneously injected in 48h before mating
G/ is only) and 4h subcutaneous injection progesterone (500 μ g/ are only), it is synchronous to be allowed to estrus time.Rat in 30min is observed after stomach-filling 1h to hand over
It (captures incubation period with ability, capture number, mating incubation period, number of copulations).Rat anesthesia, abdominal aortic blood, room temperature is certainly
Right blood coagulation, 2500rpm/min centrifugation, detects IL-2, IL-6, TNF-α content in serum.
Table 6: influence (n=10) of the polyoses extract to deficiency of yang immune functions of rats
Table 7: influence (n=10) of the polyoses extract to deficiency of yang rat mating ability
2.4 polysaccharide and monosaccharide detection:
By phend-sulphuric acid detection polyoses content after DEAE- cellulose column chromatography, detection purity of polysaccharide is
95.6%；Polysaccharide acidification hydrolization is analyzed through HPLC after derivatization, mainly single by mannose, glucose sugar, rhamnose, galactolipin etc.
Sugar composition.
Claims (4)
1. a kind of extracting method with the active aspongopus class Pentatomidae Insects polysaccharide of warming kidney and enhancing body, which is characterized in that its step
It is:
A, it by 50-80 DEG C of drying 10h-14h of aspongopus class insect body, crushes, cross 40 meshes, infiltrate powder with petroleum ether or ether
Last 2h-4h, is swollen dry powder, carries out refluxing extraction, degreasing 2-6 times, 0.5~2h, filtering, solid-liquor separation are removed every time
The dregs of a decoction of grease；
B, the dregs of a decoction of step (A) are volatilized into unrecovered oil ether, with distilled water immersion 2-4h, is impregnated with the dregs of a decoction, refluxing extraction 2-6
Secondary, 0.5~2h, filtering, solid-liquor separation, merging filtrate fling to solvent every time, are concentrated, obtain water total extract；
C, distilled water is added to dilute the water total extract of step (B), it is stirring while adding to alcohol in solution with alcoholic solution precipitate polysaccharides
Concentration is protected from light to 50~80%, stands 12h-24h, filtering obtains Insect Polysaccharide alcohol hypostasis；
D, the polysaccharide polyol hypostasis of step (C) is diluted with water, dissolved, separated with DEAE- sephadex cellulose chromatography pure
Change, rinsed with distilled water, sodium chloride solution, collect eluent, filtering volatilizes solvent, obtain it is a kind of have warming kidney and enhancing body, resist it is tired
Labor restores the active aspongopus class Pentatomidae Insects polysaccharide of body immunity；
Aspongopus class insect in the step (A) is aspongopus, black pocket worm, Cyclopelta parva Distana, any without piercement bug；
The condition being heated to reflux in the step (B) is 65~85 DEG C；
Alcoholic solution in the step (C) is any in methanol solution or ethanol solution；
It is 50~80% that the concentration of the ethyl alcohol or methanol solution, which is percent by volume,；
The elution volume is 10-20 times of column volume；
The eluent sodium chloride solution wash-out concentration is 0.1-0.9mol/L.
A kind of have the active aspongopus class Pentatomidae Insects polysaccharide of warming kidney and enhancing body in preparation treatment or pre- 2. described in claim 1
Application in anti-deficiency of yang drug.
A kind of have the active aspongopus class Pentatomidae Insects polysaccharide of warming kidney and enhancing body in preparation treatment or pre- 3. described in claim 1
Application in anti-impotence drug.
A kind of have the active aspongopus class Pentatomidae Insects polysaccharide of warming kidney and enhancing body in preparation treatment or pre- 4. described in claim 1
Application in anti-erectile dysfunction drug.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111285938A (en) * | 2020-03-02 | 2020-06-16 | 贵州大学 | Extraction method and application of aspongopus crude polysaccharide |
| CN111285938B (en) * | 2020-03-02 | 2021-07-06 | 贵州大学 | Extraction method and application of aspongopus crude polysaccharide |
| CN114957506A (en) * | 2022-07-11 | 2022-08-30 | 德宏师范高等专科学校 | Extraction method of active polysaccharide of Vespa mandarinia |
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