CN104926958B - Method for extracting herba lycopi polysaccharide from herba lycopi and application of herba lycopi polysaccharide - Google Patents
Method for extracting herba lycopi polysaccharide from herba lycopi and application of herba lycopi polysaccharide Download PDFInfo
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Abstract
The invention relates to a method for extracting herba lycopi polysaccharide from herba lycopi and the application of the herba lycopi polysaccharide. The extraction and separation aims of herba lycopi polysaccharide components can be effectively achieved. According to the technical scheme, herba lycopi medicine decoction pieces are taken, water is added, extraction is carried out one to three times at the temperature of 70 DEG C to 90 DEG C, extraction is carried out for 1-3 h every time, filtering is carried out, filter liquor is condensed, ethyl alcohol of more than 95 percent is added into concentrated liquor till the alcohol content can be 75 percent to 85 percent, the concentrated liquor stands still overnight, filtering is carried out to obtain crude polysaccharide precipitate, and then HPD-100 macroreticular resin is used for purification. The herba lycopi polysaccharide can be used for development of drugs and healthcare products with antioxidant functions, the new application prospect of herba lycopi medicinal materials is developed, and the innovation on extraction and separation of the herba lycopi polysaccharide components is achieved.
Description
Technical field
The present invention relates to field of medicaments, particularly a kind of method that Herba Lycopi's polysaccharide is extracted from Herba Lycopi and its application.
Background technology
Glucide is one of three major nutrient, and developing rapidly with analytical technology and experimental technique, it is existing big
Quantity research shows that the polysaccharide separated from natural product has antitumor, antioxidation, blood fat reducing, blood sugar lowering, raising immunity
The pharmacologically actives such as power, the medicine of various polysaccharides is clinically applied.
Herba Lycopi is labiate hair leaf lycopus lucidus Lycopus lucidus Turcz var.hirtus Regel's
Dry aerial parts.Beginning is loaded in《Sheng Nong's herbal classic》, history tree is on the books.Herba Lycopi's bitter in the mouth, pungent, tepor, Return liver, spleen channel.
Work(with blood circulation promoting and blood stasis dispelling, line water detumescence.For the diseases such as menoxenia, amenorrhea, dysmenorrhea, postnatal blood stasis stomachache, edema, Ze Lanzhu
Will be containing triterpene acids, phenolic acids, flavonoid, volatile oil, saccharide etc., and the extraction separation to Herba Lycopi's polysaccharide component and pharmacology are lived
The research of property has no report.
The content of the invention
It is the defect for solving prior art for above-mentioned situation, the purpose of the present invention is just to provide one kind and carries from Herba Lycopi
Take method and its application of Herba Lycopi's polysaccharide, can effectively solving Herba Lycopi's polysaccharide component extraction separation problem.
The technical scheme that the present invention is solved is to comprise the following steps：1) Herba Lycopi's pharmaceutical decocting piece, plus Herba Lycopi's pharmaceutical decocting piece weight are taken
The water of 20～30 times of amount, 70～90 DEG C are extracted 1～3 time, and every time 1～3h, filters, and relative density is when filtrate is concentrated into 60 DEG C
1.10～1.25 polysaccharide concentrated solution, the ethanol for adding volumetric concentration to be more than 95%, makes alcohol content reach volumetric concentration 75%
～85%, stand overnight, filter, precipitate is obtained, precipitate absolute ethanol washing is colourless to cleaning mixture, is dried, and obtains Herba Lycopi thick
Polysaccharide；
2) take Herba Lycopi 0.3～0.5g of crude polysaccharides to be dissolved in 10ml water, be splined on HPD-100 macroporous adsorptive resins, use 200
～400mL water elutions, flow velocity is 0.5～1.5ml/min, merges effluent, be further concentrated to when 40 DEG C relative density be 1.02～
1.06 extractum, the ethanol for adding volumetric concentration to be more than 95%, to alcohol content volumetric concentration 80% is reached, and is stood overnight, mistake
Filter, obtains precipitate, and precipitate is respectively washed 1～2 time successively with dehydrated alcohol, acetone, the absolute ether of 50 times of weight of precipitate, is done
It is dry, obtain refined polysaccharide.
Herba Lycopi's polysaccharide of the present invention can be used for the medicine of anti-oxidation function and the exploitation of health product, open up Herba Lycopi's medical material it is new should
It is the innovation in the extraction separation of Herba Lycopi's polysaccharide component with prospect.
Specific embodiment
The specific embodiment of the present invention is described in further detail with reference to embodiments.
Embodiment 1
1) Herba Lycopi's pharmaceutical decocting piece, plus the water of 30 times of Herba Lycopi's pharmaceutical decocting piece weight are taken, 70 DEG C are extracted 2 times, each 2h is filtered,
Relative density is 1.12 polysaccharide concentrated solution when filtrate is concentrated into 60 DEG C, the ethanol for adding volumetric concentration to be 95%, is contained to solution
Alcohol amount is volumetric concentration 75%, is stood overnight, and is filtered, and obtains precipitate, and precipitate absolute ethanol washing is colourless to cleaning mixture, is done
It is dry, obtain Herba Lycopi's crude polysaccharides；
2) take Herba Lycopi crude polysaccharides 0.3g to be dissolved in 10ml water, be splined on HPD-100 macroporous adsorptive resins, use 300mL water
Eluting, flow velocity is 1.5ml/min, merges effluent, is further concentrated to the extractum of relative density 1.03 when 40 DEG C, adds volumetric concentration
For 95% ethanol, volumetric concentration 80% is reached to alcohol content, stood overnight, filtered, obtain precipitate, precipitate is successively with precipitation
Dehydrated alcohol, acetone, the absolute ether that 50 times of thing weight is respectively washed 1 time, is dried, and obtains refined polysaccharide.
Embodiment 2
1) Herba Lycopi's pharmaceutical decocting piece, plus the water of 25 times of Herba Lycopi's pharmaceutical decocting piece weight are taken, 80 DEG C are extracted 3 times, each 1h is filtered,
Relative density is 1.20 polysaccharide concentrated solution when filtrate is concentrated into 60 DEG C, the ethanol for adding volumetric concentration to be 97%, makes alcohol content
Volumetric concentration 80% is reached, is stood overnight, filtered, obtain precipitate, precipitate absolute ethanol washing is colourless to cleaning mixture, done
It is dry, obtain Herba Lycopi's crude polysaccharides；
2) take Herba Lycopi crude polysaccharides 0.4g to be dissolved in 10ml water, be splined on HPD-100 macroporous adsorptive resins, use 250mL water
Eluting, flow velocity is 1.0ml/min, merges effluent, and relative density is 1.04 extractum when being further concentrated to 40 DEG C, adds volume dense
The ethanol for 97% is spent, to alcohol content volumetric concentration 80% is reached, stood overnight, filtered, obtain precipitate, it is heavy that precipitate is used successively
Dehydrated alcohol, acetone, the absolute ether that 50 times of starch weight is respectively washed 2 times, is dried, and obtains refined polysaccharide.
Embodiment 3
1) Herba Lycopi's pharmaceutical decocting piece, plus the water of 20 times of Herba Lycopi's pharmaceutical decocting piece weight are taken, 90 DEG C are extracted 3 times, each 3h is filtered,
Relative density is 1.25 polysaccharide concentrated solution when filtrate is concentrated into 60 DEG C, the ethanol for adding volumetric concentration to be 100%, makes alcohol content
Volumetric concentration 85% is reached, is stood overnight, filtered, obtain precipitate, precipitate absolute ethanol washing is colourless to cleaning mixture, done
It is dry, obtain Herba Lycopi's crude polysaccharides；
2) take Herba Lycopi crude polysaccharides 0.5g to be dissolved in 10ml water, be splined on HPD-100 macroporous adsorptive resins, use 200mL water
Eluting, flow velocity is 0.5ml/min, merges effluent, and relative density is 1.05 extractum when being further concentrated to 40 DEG C, adds volume dense
The ethanol for 95% is spent, to alcohol content volumetric concentration 80% is reached, stood overnight, filtered, obtain precipitate, it is heavy that precipitate is used successively
Dehydrated alcohol, acetone, the absolute ether that 50 times of starch weight is respectively washed 2 times, is dried, and obtains refined polysaccharide.
Described Herba Lycopi's decoction pieces are the stems and leaf of Herba Lycopi's 1～2cm length.
Described water is one or more of tap water, pure water, distilled water or deionized water.
There is Herba Lycopi's polysaccharide that the inventive method is extracted antioxidative to act on, and animal experiment is fully proved, concrete examination
Test situation as follows：
1. test material
1.1 experimental animal：Kunming kind no-special pathogen level (SPF) healthy mice, male and female half and half, 4～6 week old, body weight
18～22g, purchased from Henan Province's animal experimental center.
1.2 experiment reagents：Herba Lycopi's polysaccharide of the present invention；Vc effervescent tablet (federal company limited of pharmaceutical factory)；Total protein is quantitatively tried
Agent box, total number born (T-SOD) test kit, catalase (CAT) test kit, Glutathione peroxidase
(GSH-PX) test kit, malonaldehyde (MDA) test kit are purchased from Nanjing and build up Bioengineering Research Institute.
2. test method
After animal feeding room is adapted to one week, 72 mices are divided into 6 groups to mice, every group of male and female half and half.First group, gavage life
Reason saline 0.1mL/ (10gd), is blank group.Second group, lumbar injection D- galactose 150mg/ (kgd), gavage physiology salt
Water 0.1mL/ (10gd), is model group.3rd group, lumbar injection D- galactose 150mg/ (kgd), and gavage vitamin C
150mg/ (kgd), is Vc group.4th group, lumbar injection D- galactose 150mg/ (kgd), and gavage Herba Lycopi's polysaccharide
100mg/ (kgd), is low dose group.5th group, lumbar injection D- galactose 150mg/ (kgd), and gavage Herba Lycopi's polysaccharide
200mg/ (kgd), is middle dose group.6th group, lumbar injection D- galactose 150mg/ (kgd), gavage Herba Lycopi's polysaccharide
300mg/ (kgd), is high dose group.Vc and Herba Lycopi's polysaccharide are aqueous solution, and D- galactose physiological saline solutions, mice fills
Body of stomach accumulated amount is 0.1mL/ (10gd), and after three weeks, fasting 12 hours, the de- neck of mice is put to death, and quickly removes liver, brain, accurately
Claim its quality, add normal saline glass homogenizer to be homogenized manually in frozen water with the ﹕ 9 (g ﹕ mL) of volume ratio 1 by quality, uniformly
10% tissue homogenate is afterwards, 3000 revs/min, after centrifugation 10min, supernatant is taken, and is freezed standby.Saying by test kit
Bright book operation, uses ultraviolet-uisible spectrophotometer mensuration absorbance, is calculated each desired value.
3. result of the test
The impact result table of the Herba Lycopi's Polysaccharides on Mice brain and liver tissues T-SOD vigor of table 1
Note：Compare * * with model group for p<0.01, * is p<0.05
As can be seen from Table 1, the T-SOD vigor of model group Mice brain tissues and hepatic tissue is substantially lower than blank group, table
Bright lumbar injection D- galactose 150mg/ (kgd) make the low model success of total number born vigor.
It can also be seen that, Herba Lycopi's polysaccharide is in positive dose relationship with the T-SOD vigor of brain and hepatic tissue.Herba Lycopi's polysaccharide is middle and high
Dosage group and Vc group can pole be obviously improved liver, cerebral tissue T-SOD vigor (P<0.01).
The impact result table of the Herba Lycopi's Polysaccharides on Mice brain and liver tissues GSH-PX vigor of table 2
Note：Compare * * with model group for p<0.01, * is p<0.05
As can be seen from Table 2, the GSH-PX vigor of model group Mice brain tissues and hepatic tissue is substantially lower than blank group, table
Bright lumbar injection D- galactose 150mg/ (kgd) make the low model success of Glutathione peroxidase (GSH-PX) vigor.
It can also be seen that, Herba Lycopi's polysaccharide is in positive agent with Glutathione peroxidase (GSH-PX) vigor of brain and hepatic tissue
Magnitude relation.The basic, normal, high dosage group of Herba Lycopi's polysaccharide and Vc group can pole be obviously improved murine liver tissue glutathion peroxidating
Hydrogen enzyme activity (P<0.01), Herba Lycopi's polysaccharide high dose group and Vc group can pole be obviously improved Mice brain tissues glutathion peroxide
Change hydrogen enzyme activity (P<0.01), and Herba Lycopi's polysaccharide middle dose group can be obviously improved Mice brain tissues Glutathione peroxidase work
Power (P<0.05), low dose group is compared with model group, without remarkable effect.
The impact result table of the Herba Lycopi's Polysaccharides on Mice brain and liver tissues CAT vigor of table 3
Note：Compare * * with model group for p<0.01, * is p<0.05
As can be seen from Table 3, the CAT vigor of model group Mice brain tissues and hepatic tissue is substantially lower than blank group, shows
Lumbar injection D- galactose 150mg/ (kgd) makes the low model success of catalase (CAT) vigor.
It can also be seen that, Herba Lycopi's polysaccharide is in positive dose relationship with catalase (CAT) vigor of brain and hepatic tissue.Herba Lycopi
The basic, normal, high dosage group of polysaccharide and Vc group can pole be obviously improved Mice brain tissues catalase (CAT) vigor (P<
0.01), the middle and high dosage group of Herba Lycopi's polysaccharide and Vc group can pole be obviously improved murine liver tissue catalase (CAT) vigor (P
<0.01), and Herba Lycopi's polysaccharide low dose group can be obviously improved murine liver tissue catalase (CAT) vigor (P<0.05)
The impact result table of the Herba Lycopi's Polysaccharides on Mice brain and liver tissues MDA contents of table 4
Note：Compare * * with model group for p<0.01, * is p<0.05
As shown in Table 4, model group Mouse Liver, the MDA contents of cerebral tissue are all significantly higher than blank group, show lumbar injection D-
Galactose 150mg/ (kgd) modeling success.
Compare with model group, the middle and high dosage group of Herba Lycopi's polysaccharide and Vc group can pole significantly reduce Mice brain tissues MDA contents
(P<0.01), Herba Lycopi's polysaccharide low dose group can significantly reduce Mice brain tissues MDA content (P<0.05), Herba Lycopi's polysaccharide is low, high agent
Amount group can significantly reduce murine liver tissue MDA content (P<0.05), but middle dose group and Vc group effect it is unobvious.
Four forms can draw more than, and the Testing index of model group and blank group has pole significant difference (p<
0.01), i.e., lumbar injection D- galactose successfully causes aging model；Vc group compares with model group can be obtained, and Vc is to hepatic tissue MDA
Content do not improve significantly outer, to liver cerebral tissue, other indexs have extremely significant improvement result (p<0.01)；And
Herba Lycopi's polysaccharide high dose group belongs to significant difference (p compared with model group except the content reduction to hepatic tissue MDA is acted on<
0.05) outward, to the improvement result of other indices, there is pole significant difference (p<0.01).
The aging of human body, mainly due to the oxidation institute caused by free radical extremely, the strong oxidizing property of free radical can damage machine
The tissue and cell of body, and then cause aging effect and various chronic diseases, such as alzheimer disease, cardiovascular and cerebrovascular disease, cataract,
Parkinson disease, diabetes, tumor etc..Therefore, the oxidation resistance for wanting good health and a long life enhancing body is the important ring of defying age
Section.
Results of animal shows that the Herba Lycopi's polysaccharide extracted in the present invention can improve body SOD, GSH-Px, CAT etc. and resist
Oxidasic vigor, so as to promote the elimination of machine interior free yl, mitigates lipid peroxidation and occurs, and protects biomembrane, thus has
Antioxidation and defying age class health product or medicine are developed in prestige, open up the new application prospect of Herba Lycopi's medical material, there is actual promotion price
Value.
Claims (7)
1. it is a kind of from Herba Lycopi extract Herba Lycopi's polysaccharide method, it is characterised in that comprise the following steps：
1）Take Herba Lycopi's pharmaceutical decocting piece, plus the water of 20～30 times of Herba Lycopi's pharmaceutical decocting piece weight, 70～90 DEG C are extracted 1～3 time, every time 1
～3h, filters, and relative density is 1.10～1.25 polysaccharide concentrated solution when filtrate is concentrated into 60 DEG C, adds volumetric concentration to be 95%
Ethanol above, makes alcohol content reach volumetric concentration 75%～85%, stands overnight, and filters, and obtains precipitate, the anhydrous second of precipitate
Alcohol washs colourless to cleaning mixture, is dried, and obtains Herba Lycopi's crude polysaccharides；
2）Take Herba Lycopi 0.3～0.5g of crude polysaccharides to be dissolved in 10ml water, be splined on HPD-100 macroporous adsorptive resins, with 200～
400mL water elutions, flow velocity is 0.5～1.5ml/min, merges effluent, be further concentrated to when 40 DEG C relative density be 1.02～
1.06 extractum, the ethanol for adding volumetric concentration to be more than 95%, to alcohol content volumetric concentration 80% is reached, and is stood overnight, and is filtered,
Precipitate, precipitate respectively washs 1～2 time successively with dehydrated alcohol, acetone, the absolute ether of 50 times of weight of precipitate, is dried,
Obtain refined polysaccharide.
2. it is according to claim 1 from Herba Lycopi extract Herba Lycopi's polysaccharide method, it is characterised in that comprise the following steps：
1）Herba Lycopi's pharmaceutical decocting piece, plus the water of 30 times of Herba Lycopi's pharmaceutical decocting piece weight are taken, 70 DEG C are extracted 2 times, each 2h is filtered, filtrate
Relative density is 1.12 polysaccharide concentrated solution when being concentrated into 60 DEG C, the ethanol for adding volumetric concentration to be 95%, is to solution alcohol content
Volumetric concentration 75%, stands overnight, and filters, and obtains precipitate, and precipitate absolute ethanol washing is colourless to cleaning mixture, is dried, and obtains damp
Blue crude polysaccharides；
2）Take Herba Lycopi crude polysaccharides 0.3g to be dissolved in 10ml water, be splined on HPD-100 macroporous adsorptive resins, use 300mL water elutions,
Flow velocity is 1.5ml/min, merges effluent, is further concentrated to the extractum of relative density 1.03 when 40 DEG C, adds volumetric concentration to be 95%
Ethanol, reach volumetric concentration 80% to alcohol content, stand overnight, filter, obtain precipitate, precipitate uses successively weight of precipitate
50 times of dehydrated alcohol, acetone, absolute ether is respectively washed 1 time, is dried, and obtains refined polysaccharide.
3. it is according to claim 1 from Herba Lycopi extract Herba Lycopi's polysaccharide method, it is characterised in that comprise the following steps：
1）Herba Lycopi's pharmaceutical decocting piece, plus the water of 25 times of Herba Lycopi's pharmaceutical decocting piece weight are taken, 80 DEG C are extracted 3 times, each 1h is filtered, filtrate
Relative density is 1.20 polysaccharide concentrated solution when being concentrated into 60 DEG C, the ethanol for adding volumetric concentration to be 97%, makes alcohol content reach body
Product concentration 80%, stands overnight, and filters, and obtains precipitate, and precipitate absolute ethanol washing is colourless to cleaning mixture, is dried, and obtains Herba Lycopi
Crude polysaccharides；
2）Take Herba Lycopi crude polysaccharides 0.4g to be dissolved in 10ml water, be splined on HPD-100 macroporous adsorptive resins, use 250mL water elutions,
Flow velocity is 1.0ml/min, merges effluent, and relative density is 1.04 extractum when being further concentrated to 40 DEG C, and addition volumetric concentration is
97% ethanol, to alcohol content volumetric concentration 80% is reached, and is stood overnight, and is filtered, and obtains precipitate, and precipitate uses successively precipitate weight
Dehydrated alcohol, acetone, the absolute ether of 50 times of amount is respectively washed 2 times, is dried, and obtains refined polysaccharide.
4. it is according to claim 1 from Herba Lycopi extract Herba Lycopi's polysaccharide method, it is characterised in that comprise the following steps：
1）Herba Lycopi's pharmaceutical decocting piece, plus the water of 20 times of Herba Lycopi's pharmaceutical decocting piece weight are taken, 90 DEG C are extracted 3 times, each 3h is filtered, filtrate
Relative density is 1.25 polysaccharide concentrated solution when being concentrated into 60 DEG C, the ethanol for adding volumetric concentration to be 100%, reaches alcohol content
Volumetric concentration 85%, stands overnight, and filters, and obtains precipitate, and precipitate absolute ethanol washing is colourless to cleaning mixture, is dried, and obtains damp
Blue crude polysaccharides；
2）Take Herba Lycopi crude polysaccharides 0.5g to be dissolved in 10ml water, be splined on HPD-100 macroporous adsorptive resins, use 200mL water elutions,
Flow velocity is 0.5ml/min, merges effluent, and relative density is 1.05 extractum when being further concentrated to 40 DEG C, and addition volumetric concentration is
95% ethanol, to alcohol content volumetric concentration 80% is reached, and is stood overnight, and is filtered, and obtains precipitate, and precipitate uses successively precipitate weight
Dehydrated alcohol, acetone, the absolute ether of 50 times of amount is respectively washed 2 times, is dried, and obtains refined polysaccharide.
5. according to claim 1 or any one of 2-4 from Herba Lycopi extract Herba Lycopi's polysaccharide method, it is characterised in that institute
The Herba Lycopi's decoction pieces stated are the stems and leaf of Herba Lycopi's 1～2cm length；Described water is tap water, pure water, distilled water or deionized water
One or more.
6. Herba Lycopi's polysaccharide that prepared by claim 1 or the method described in any one of 2-4 is preparing anti-oxidation medicine or antioxidation is protected
Application in strong product.
7. Herba Lycopi's polysaccharide that prepared by the method described in claim 5 answering in anti-oxidation medicine or antioxidant health-care product is prepared
With.
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