CN114957506A - Extraction method of active polysaccharide of Vespa mandarinia - Google Patents
Extraction method of active polysaccharide of Vespa mandarinia Download PDFInfo
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- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/0003—General processes for their isolation or fractionation, e.g. purification or extraction from biomass
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Abstract
The invention discloses an extraction method of active polysaccharide of hornet wasps, belonging to the technical field of insect source polysaccharide extraction. The extraction method comprises the working procedures of insect pretreatment, degreasing, extraction and purification, and comprises the steps of freezing and killing the wasps aurantiaca, cleaning, drying at 55-65 ℃, crushing and sieving. And refluxing the insect powder in a degreasing agent at 75-85 ℃ for 3.5-4.5 h, degreasing and then drying. Degreasing insect powder, water and 0.01-0.04 mol/L alkali liquor according to the weight ratio of 1g:20mL of: mixing 180-220 mL of materials and liquid, and performing reflux extraction at 70-80 ℃ for 1-2.5 h. Adjusting the extraction stock solution to be neutral, adding a precipitator, standing in the dark, and filtering after polysaccharide is precipitated to obtain the target active polysaccharide of the hornet wasps. The extraction method provided by the invention makes up the technical blank of the extraction of the active polysaccharide of the wasp horneri, has high extraction rate, and well retains the natural conformation and biological activity of the wasp horneri polysaccharide. The process is simple and easy to implement, the general interchangeability of the equipment is good, the economic benefit is outstanding, and the popularization and application value is high.
Description
Technical Field
The invention belongs to the technical field of insect source polysaccharide extraction, and particularly relates to an extraction method of active polysaccharide of hornet wasps.
Background
The insect resources in Yunnan province are abundant and diverse, and the medicinal insects include 87 kinds of 34 families of 10 meshes, Wasp et al (Wasp.) kuwakameVespa mandarinia) The whole insect can be used as a medicine, belongs to hymenoptera and wasp families, can be used for treating rheumatic pain, arthritis and the like, and has a very long application history in China. The Vespa mandarinia polysaccharide is characterized in that a sugar chain is used as a basic skeleton, and is often combined with protein, lipid and the like to form a polysaccharide complex, and the special combination mechanism endows the Vespa mandarinia polysaccharide with very different macromolecular three-dimensional conformations with other plant or animal derived polysaccharides, so that the defects of low extraction rate, poor method stability and reproducibility and the like of the existing Vespa mandarinia polysaccharide are overcome. In recent years, with the maturity of the artificial breeding technology of the hornet and the steady increase of the yield, the research on the extraction process of the active polysaccharide in the hornet can provide corresponding technical basis for the comprehensive utilization and further deep processing of the hornet.
Disclosure of Invention
The invention aims to provide an extraction method of active polysaccharide of wasp horneri.
The purpose of the invention is realized by the following steps: the extraction method of the active polysaccharide of the hornet comprises the working procedures of pretreatment, degreasing, extraction and purification of the worm body, and specifically comprises the following steps:
A. pretreatment of polypide: freezing and killing the wasps aurantiaca, cleaning, drying at 55-65 ℃, crushing and sieving to obtain insect powder for later use.
B. Degreasing: and refluxing the insect powder in a degreasing agent at 75-85 ℃ for 3.5-4.5 h, degreasing, and drying to obtain the degreased insect powder.
C. Extraction: degreasing insect powder, water and 0.01-0.04 mol/L alkali liquor according to the weight ratio of 1g:20mL of: mixing 180-220 mL of materials and liquid, and performing reflux extraction at 70-80 ℃ for 1-2.5 hours to obtain an extraction stock solution.
D. And (3) purification: adjusting the extraction stock solution to be neutral, adding a precipitator, standing in a dark place, and filtering after polysaccharide is precipitated to obtain the target hornet active polysaccharide.
In the pretreatment procedure of the insect body, the freezing temperature is less than or equal to-20 ℃, and the drying temperature is preferably 60 ℃. The polypide powder needs to be sieved by a 60-mesh sieve.
In the degreasing step, the degreasing temperature is preferably 80 ℃ and the degreasing time is preferably 4 hours. The degreasing agent is any one of petroleum ether, diethyl ether or isopropyl ether.
In the extraction step, the alkali liquor is any one of sodium hydroxide and potassium hydroxide, and the concentration of the alkali liquor is preferably 0.02 mol/L. The feed-liquid ratio of the degreased insect powder, the water and the alkali liquor is preferably 1g to 20mL to 200 mL. The extraction temperature is preferably 72-75 ℃, and the extraction time is preferably 2 h.
In the purification step, the precipitant is any one of methanol, ethanol, and acetone. And the dark standing time is 10-12 h.
Compared with the substantive characteristics and the outstanding progress of the prior art, the extraction method of the active polysaccharide of the wasp horneri is shown as follows:
the Vespa mandarinia polysaccharide is combined with protein through covalent bond rather than physical or ionic bond, and the dilute alkali extractant is favorable to breaking the bond between polysaccharide and protein and raising the polysaccharide separating rate. Secondly, the ingredients of the polysaccharide of the hornet are complex, and the selection of an extracting agent and the optimization of an extracting process have decisive influence on the extraction rate, the properties, the components and the biological activity of the polysaccharide. The extraction process selected by the invention can improve the extraction rate of the hornet polysaccharide to more than 15 percent, which is more than twice of the highest extraction rate of the same sample when hot water extraction and protease extraction are adopted, and the natural conformation and the biological activity of the hornet polysaccharide are well maintained. In conclusion, the extraction method disclosed by the invention fills up the technical blank of the active polysaccharide extraction of the wasp horneri, is simple and feasible in process, good in general interchangeability of equipment, outstanding in economic benefit and extremely high in popularization and application value.
Detailed Description
The present invention is further illustrated but not limited in any way by the following description, and any alterations or substitutions based on the teachings of the present invention are intended to fall within the scope of the present invention.
Example 1
A. Pretreatment of worm bodies: freezing and killing Vespa mandarinia at-20 deg.C, cleaning, drying at 60 deg.C, pulverizing, and sieving with 60 mesh sieve to obtain insect powder.
B. Degreasing: refluxing the insect powder in petroleum ether at 80 deg.C for 4h, defatting, and air drying to obtain defatted insect powder.
C. Extraction: degreasing insect powder, water and 0.02mol/L sodium hydroxide are mixed according to the weight ratio of 1g:20mL of: mixing 200mL of materials and liquid, and extracting under reflux at 72.5 ℃ for 2h to obtain an extraction stock solution.
D. And (3) purification: adjusting the extraction stock solution to be neutral, adding ethanol, standing for 11h in a dark place, and filtering after polysaccharide is precipitated to obtain the target hornet active polysaccharide.
Example 2
A. Pretreatment of worm bodies: freezing and killing Vespa mandarinia at-25 deg.C, cleaning, drying at 55 deg.C, pulverizing, and sieving with 60 mesh sieve to obtain insect powder.
B. Degreasing: refluxing the insect powder in ethyl ether at 85 deg.C for 3.5h, defatting, and air drying to obtain defatted insect powder.
C. Extraction: degreasing insect powder, water and 0.03mol/L sodium hydroxide are mixed according to the weight ratio of 1g:20mL of: mixing 220mL of materials and liquid, and extracting under reflux at 70 ℃ for 2.5h to obtain an extraction stock solution.
D. And (3) purification: adjusting the extraction stock solution to be neutral, adding methanol, standing for 12h in a dark place, and filtering after polysaccharide is precipitated to obtain the target hornet active polysaccharide.
Example 3
A. Pretreatment of polypide: freezing and killing Vespa mandarinia at-30 deg.C, cleaning, drying at 65 deg.C, pulverizing, and sieving with 60 mesh sieve to obtain insect powder.
B. Degreasing: refluxing the insect powder in isopropyl ether at 75 deg.C for 4.5h, defatting, and air drying to obtain defatted insect powder.
C. Extraction: degreasing insect powder, water and 0.04mol/L potassium hydroxide are mixed according to the weight ratio of 1g:20mL of: mixing 180mL of materials and liquid, and extracting under reflux at 80 ℃ for 1h to obtain an extraction stock solution.
D. And (3) purification: adjusting the extraction stock solution to be neutral, adding acetone, standing for 10h in a dark place, and filtering after polysaccharide is precipitated to obtain the target hornet active polysaccharide.
Example 4
-extracting polysaccharide from Wasp Sinapis cerana of 14 different regions and comparing the extracted polysaccharide with the content
Wasp's larva (A. Ex Fr.) with golden ringVespa Mandarinia) The samples were from 14 different regions of Yunnan province, and the specific information of the samples is shown in Table 1.
The extraction process was the same as in example 1.
TABLE 1 Vespa mandarinia sample information
The content of the polysaccharide in the Vespa mandarinia is determined by adopting a spectrophotometric method, and the content of the polysaccharide is calculated according to a content calculation formula in DB 22/T1685 + 2012 Jilin province local standard ginseng polysaccharide determination spectrophotometric method.
In the formula:
x-content of polysaccharide of wasp, unit: microgram per gram (mu g/g)
C 1 -obtaining the content of hornet's polysaccharide in units according to a standard regression equation: microgram per milliliter (mug/mL)
f-dilution factor of the test solution
V is the constant volume of the liquid to be measured, unit: milliliter (mL)
M-sample mass to be measured, unit: gram (g)
The results of extraction and content calculation are shown in table 2.
TABLE 214 active polysaccharide content of Vespa mandarinia in different regions
The results show that the content of active polysaccharides of the hornet wasps in 14 different areas is arranged from high to low as follows: 7H >14H >3H >6H >9H >5H >8H >4H >10H >12H >2H >13H >1H >11H, the highest is No. 7 sample, the content of active polysaccharide reaches 1535.0 mu g/g, and the extraction rate reaches 15.35%.
Claims (9)
1. The extraction method of the active polysaccharide of the hornet is characterized by comprising the working procedures of pretreatment, degreasing, extraction and purification of the polypide, and specifically comprises the following steps:
A. pretreatment of worm bodies: freezing and killing the wasps aurantiaca, cleaning, drying at 55-65 ℃, crushing and sieving to obtain insect powder for later use;
B. degreasing: refluxing the insect powder in a degreasing agent at 75-85 ℃ for 3.5-4.5 h, degreasing, and drying to obtain degreased insect powder;
C. extraction: degreasing insect powder, water and 0.01-0.04 mol/L alkali liquor according to the weight ratio of 1g:20mL of: mixing 180-220 mL of materials and liquid, and performing reflux extraction at 70-80 ℃ for 1-2.5 hours to obtain an extraction stock solution;
D. and (3) purification: adjusting the extraction stock solution to be neutral, adding a precipitator, standing in a dark place, and filtering after polysaccharide is precipitated to obtain the target hornet active polysaccharide.
2. The extraction method according to claim 1, wherein the freezing temperature is less than or equal to-20 ℃ and the drying temperature is 60 ℃ in the insect pretreatment process.
3. The extraction method according to claim 1, wherein in the degreasing step, the degreasing temperature is 80 ℃ and the degreasing time is 4 hours.
4. The extraction method according to claim 1, wherein in the degreasing step, the degreasing agent is any one of petroleum ether, diethyl ether and isopropyl ether.
5. The extraction method according to claim 1, wherein in the extraction step, the alkali solution is one of sodium hydroxide and potassium hydroxide, and the concentration of the alkali solution is 0.02 mol/L.
6. The extraction method according to claim 1, wherein in the extraction step, the feed-liquid ratio of the defatted insect powder, the water and the alkali liquor is 1g:20mL:200 mL.
7. The extraction method according to claim 1, wherein the extraction temperature is 72 to 75 ℃ and the extraction time is 2 hours.
8. The extraction method according to claim 1, wherein in the purification step, the precipitant is any one of methanol, ethanol, and acetone.
9. The extraction method according to claim 1, wherein the light-shielding standing time in the purification step is 10 to 12 hours.
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Citations (3)
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| CN105646728A (en) * | 2016-03-11 | 2016-06-08 | 江苏农牧科技职业学院 | Honeycomb spleen polysaccharide extract and preparation method thereof |
| CN109280090A (en) * | 2018-11-22 | 2019-01-29 | 湖北中医药大学 | One kind having the active aspongopus class Pentatomidae Insects extraction method of polysaccharides of warming kidney and enhancing body and application |
| CN111285938A (en) * | 2020-03-02 | 2020-06-16 | 贵州大学 | Extraction method and application of aspongopus crude polysaccharide |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105646728A (en) * | 2016-03-11 | 2016-06-08 | 江苏农牧科技职业学院 | Honeycomb spleen polysaccharide extract and preparation method thereof |
| CN109280090A (en) * | 2018-11-22 | 2019-01-29 | 湖北中医药大学 | One kind having the active aspongopus class Pentatomidae Insects extraction method of polysaccharides of warming kidney and enhancing body and application |
| CN111285938A (en) * | 2020-03-02 | 2020-06-16 | 贵州大学 | Extraction method and application of aspongopus crude polysaccharide |
Non-Patent Citations (2)
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| OLUREMI M. DAUDU等: "Bee Pollen Extracts as Potential Antioxidants and Inhibitors of α-Amylase and α-Glucosidase Enzymes In Vitro Assessment" * |
| 王丽华等: "蜜蜂壳聚糖提取与抑菌研究", 《中国蜂业》 * |
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