CN106309523A - Total flavonoid extract of acanthopanax senticosus harms, as well as preparation method and application thereof - Google Patents
Total flavonoid extract of acanthopanax senticosus harms, as well as preparation method and application thereof Download PDFInfo
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Abstract
The invention relates to a total flavonoid extract of acanthopanax senticosus harms, as well as a preparation method and application thereof. The extract contains 55-100 percent of total flavonoids, including 10-99 percent of rutin, 0.05-10 percent of quercetin, 0.002-0.5 percent of kaempferol and 0.5-70 percent of hyperoside. The preparation method comprises the following steps: extracting acanthopanax senticosus harms with a solvent, extracting and removing impurities with liquid, concentrating, standing and collecting residue to obtain a crude extract of total flavonoids; dissolving a proper amount of solvent, and passing through a chromatographic column to obtain the total flavonoid extract of acanthopanax senticosus harms. The invention further discloses application of the acanthopanax senticosus harms extract in preparation of medicines for treating myocardial and cerebral ischemia, stopping bleeding and relieving cough and preparation of sunscreen and whitening cosmetics. The preparation method is reasonable in process, simple and feasible in operation, has low production cost, and small environmental pollution, the flavonoid compound has high yield and purity, and wide application, and the utilization value of acanthopanax senticosus harms can be greatly improved.
Description
Technical field
The present invention relates to a kind of Folium Acanthopanacis Radicis extractive of general flavone and its production and use.
Background technology
Cardio-cerebralvascular diseases is one of difficult diseases that harm human body is healthy at present, and its sickness rate worldwide and mortality rate are the most very
High.Although having developed a lot of cardio-cerebralvascular medicine at present, but owing to the toxic and side effects of medicine itself is big, cause a lot of medicine in clinic
It is restricted during use.Therefore the most important thing that the little cardiovascular and cerebrovascular diseases medicament of toxic and side effects is research at present is found.
In recent years, serious along with environmental pollution, the ozone layer in earth overhead is constantly destroyed, accelerated skin ageing and some skins thus caused
Property disease showed increased, such as solar dermatitis, farmersskin, polymorphous light eruption etc., the most sun-proof delaying aging that becomes, prevents and controls
Treat one of main measure of disease.Having a variety of sunscreen using in the market, wherein main Types is the oiliness sunscreen of cinnamate derivative,
And the skin care item of high sun protection factor add the sunscreen of higher concentration mostly, there is light or heavy side effect and the zest serious to skin.
Chinese medicine sunscreen is from the natural trend not only conforming with cosmetics " back to nature ", and because of the wide spectrum of its uniqueness, sun-proof result efficient, safe, is subject to
Consumers mostly.
Studies on plants of Acanthopanax Miq. in Araliaceae has a variety of, and wherein acanthopanax gracilistylus and Radix Et Caulis Acanthopanacis Senticosi are clinical conventional Chinese medicines, respectively with its root bark and rhizome be embodied in " in
State's pharmacopeia " in；Rough leaf Cortex Acanthopancis is not only used for medicinal purpose, and among the people is also often used as food materials and brew medicated wine, belongs to the plant of medicine-food two-purpose.But pass
The leaf of slender acanthopanax is generally removed and abandons by system processing, and this not only causes the great wasting of resources, also can increase production cost and operation, also may be used environment
Can pollute.Plant chemical ingredient research shows, the main chemical compositions of Folium Acanthopanacis Radicis is flavone compound, and flavone compound has regulation
Cardio-cerebrovascular, antioxidation, scavenging activated oxygen etc. acts on.Through retrieval, not yet find that Folium Acanthopanacis Radicis total flavones is used for cardiac-cerebral ischemia, stops blooding and stop
Cough medicine and prepare the report of sun-proof skin-lightening cosmetic.Therefore patent of the present invention by the leaf of slender acanthopanax through suitably processing, it is provided that one has treatment heart and brain
The preparation method of the Folium Acanthopanacis Radicis extractive of general flavone of ischemia, hemostasis and antitussive action, it is simple to operate, safely, be prone to industrialized production, fully profit
With bioactive substance and the pharmacologically active thereof of wherein flavonoid, not only reducing the wasting of resources, and protect environment, having market widely should
Use prospect.
Summary of the invention
One of the object of the invention is to provide the Folium Acanthopanacis Radicis extractive of general flavone of a kind of high flavones content.
The two of the object of the invention are to provide the new preparation method of above-mentioned Folium Acanthopanacis Radicis extractive of general flavone.
The three of the object of the invention are to provide the purposes of above-mentioned Folium Acanthopanacis Radicis extractive of general flavone.
In order to realize foregoing invention purpose, the present invention adopts the following technical scheme that
A kind of Folium Acanthopanacis Radicis extractive of general flavone that the present invention provides, it has the feature that the amount containing total flavones is 55%～100%, and wherein rutin contains
Amount is 10%～99%, and quercetin content is 0.05%～10%, and the content of kaempferol is 0.002%～0.5%, and Determination of Hyperoside is 0.5%～70%.
The present invention provides the preparation method of a kind of above-mentioned Folium Acanthopanacis Radicis extractive of general flavone, it is characterised in that comprise the following steps:
(1) preparation of crude extract
Take Folium Acanthopanacis Radicis, with the solvent extraction 1 of 4～15 times of volumes～3 times, united extraction liquid, recycling design；Residue is fully dispersed with appropriate distilled water,
Petroleum ether extraction 3～8 times；Water layer is concentrated into appropriate volume, stand at low temperature 24h, takes residue, obtains total flavones crude extract.
(2) purification
Fully being dissolved by crude extract solvent, upper chromatographic column, methanol aqueous solution or ethanol water in varing proportions are eluting solvent, and eluting is appropriate
Volume, collects fraction, merges the fraction containing flavones ingredient, decompression and solvent recovery, is dried, obtains Folium Acanthopanacis Radicis extractive of general flavone.
Folium Acanthopanacis Radicis described in step (1) is rough leaf Folium Acanthopanacis Radicis, acanthopanax gracilistylus leaf, Fructus Evodiae Folium Acanthopanacis Radicis, acanthopanax leucorrhizus Harms leaf, Cortex Acanthopanacis Giraldii leaf, short
Stalk Folium Acanthopanacis Radicis, pubescence Folium Acanthopanacis Radicis, Folium Acanthopanacis Senticosi, Panax sessiliflorus leaf, white hair Folium Acanthopanacis Radicis, Tang's Folium Acanthopanacis Radicis, island Folium Acanthopanacis Radicis, different post Folium Acanthopanacis Radicis, home
One or more in the Folium Acanthopanacis Radicis of family name's Folium Acanthopanacis Radicis, intelligence different mountain.
Folium Acanthopanacis Radicis preferred rough leaf Folium Acanthopanacis Radicis described in step (1).
Extraction solvent described in step (1) is methanol, ethanol, water or its mixture.
Extracting method described in step (1) is one or more in heating and refluxing extraction, homogenate extraction, supersound extraction, merceration extraction.
In step (2), the adsorbent of institute's chromatographic column is polyamide, or D101, HPD-100, AB-8, X-5, NKA-9 type macroporous adsorbent resin.
Drying means described in step (2) is one or more in drying, vacuum drying, microwave drying, lyophilization.
The invention provides a kind of Folium Acanthopanacis Radicis extract and at preparation treatment cardiac-cerebral ischemia, hemostasis and cough suppressing medicine and prepare answering in sun-proof skin-lightening cosmetic
With.
The present invention has the following advantages and beneficial effect:
1. in the Folium Acanthopanacis Radicis extractive of general flavone that the present invention prepares, the amount containing total flavones is 55%～100%, and wherein rutin content is 10%～99%, Mongolian oak
Skin cellulose content is 0.05%～10%, and kaempferol content is 0.002%～0.5%, and Determination of Hyperoside is 0.5%～70%；Gained extractive of general flavone is used for
Prepare cardiac-cerebral ischemia, hemostasis and cough suppressing medicine it can also be used to prepare sun-proof skin-lightening cosmetic.
The preparation technology of Folium Acanthopanacis Radicis flavone extract the most of the present invention is reasonable, and operation is simple and feasible, and production cost is low, and environmental pollution is little,
The yield of flavone compound and purity are high, can be used for industrialized mass.
Accompanying drawing explanation
Fig. 1 is the HPLC collection of illustrative plates of hybrid standard product rutin, Quercetin, kaempferol, hyperin.
Fig. 2 is the HPLC collection of illustrative plates of Folium Acanthopanacis Radicis extractive of general flavone.
Fig. 3 is that Folium Acanthopanacis Radicis extractive of general flavone causes the impact of mice ECG ST section to Iso.
Detailed description of the invention
In Folium Acanthopanacis Radicis extractive of general flavone obtained by the following embodiment of the present invention, the assay method of total flavones is with reference to " Chinese Pharmacopoeia " version in 2010
Annex determined by ultraviolet spectrophotometry, with rutin as reference substance, measures trap at 500nm, and calculates the content of total flavones.
Rutin in Folium Acanthopanacis Radicis extractive of general flavone, Quercetin, kaempferol, assay method reference " Chinese Pharmacopoeia " version one in 2010 of hyperin
Portion's annex high effective liquid chromatography for measuring.Chromatographic condition and system suitability: employing 18 alkyl silica gel is filler；With methanol-0.4% phosphorus
Acid solution (50:50) is flowing phase, and detection wavelength is 360nm；Column temperature is 30 DEG C；Flow velocity is 1.0mL/min；Theoretical cam curve is with rutin
Meter is not less than 2000；Hybrid standard product HPLC collection of illustrative plates and sample HPLC collection of illustrative plates such as Fig. 1 and Fig. 2.
By following example, the present invention is further elaborated.
Embodiment 1
The preparation method of Folium Acanthopanacis Radicis extractive of general flavone, step is as follows:
(1) preparation of crude extract
Take rough leaf Folium Acanthopanacis Radicis, with the 80% ethanol water heating and refluxing extraction 3 times of 10 times of volumes, united extraction liquid, recycling design；Residue is with suitable
Amount distilled water is fully dispersed, petroleum ether extraction 5 times；Water layer is concentrated into appropriate volume, stand at low temperature 24h, takes residue, obtains total flavones crude extract.
(2) purification
Fully being dissolved by crude extract aqueous solution, upper D101 macroporous resin chromatography, ethanol water in varing proportions is eluting solvent, eluting
Appropriate volume, collects fraction, merges the fraction containing flavones ingredient, decompression and solvent recovery, is dried, obtains rough leaf Folium Acanthopanacis Radicis extractive of general flavone
A。
In dried rough leaf Folium Acanthopanacis Radicis extractive of general flavone A, the amount of total flavones is 95%, and wherein rutin content is 80%, and quercetin content is 5.2%,
Kaempferol content is 0.25%, and Determination of Hyperoside is 9.55%.
Embodiment 2
The preparation method of Folium Acanthopanacis Radicis extractive of general flavone, step is as follows:
(1) preparation of crude extract
Take acanthopanax gracilistylus leaf, with 12 times of volume 50% methanol aqueous solution supersound extraction 3 times, united extraction liquid, recycling design；Residue is with appropriate distillation
Water is fully dispersed, petroleum ether extraction 8 times；Water layer is concentrated into appropriate volume, stand at low temperature 24h, takes residue, obtains total flavones crude extract.
(2) purification
It is eluting solvent by crude extract with water-soluble abundant dissolving, upper polyamide chromatographic column, methanol aqueous solution in varing proportions, eluting appropriate volume,
Collect fraction, merge the fraction containing flavones ingredient, decompression and solvent recovery, be dried, obtain acanthopanax gracilistylus leaf total flavone extract B.
In dried acanthopanax gracilistylus leaf total flavone extract B, the amount of total flavones is 92.6%, and wherein rutin content is 76.8%, and quercetin content is 8.2%,
Kaempferol content is 0.45%, and Determination of Hyperoside is 7.15%.
Embodiment 3
The preparation method of Folium Acanthopanacis Radicis extractive of general flavone, step is as follows:
(1) preparation of crude extract
Take Fructus Evodiae Folium Acanthopanacis Radicis, with the 95% ethanol water homogenate extraction 2 times of 15 times of volumes, united extraction liquid, recycling design；Residue is with in right amount
Distilled water is fully dispersed, petroleum ether extraction 6 times；Water layer is concentrated into appropriate volume, stand at low temperature 24h, takes residue, obtains total flavones crude extract.
(2) purification
Fully being dissolved by crude extract solvent, upper HPD-100 macroporous resin chromatography, ethanol water in varing proportions is eluting solvent, eluting
Appropriate volume, collects fraction, merges the fraction containing flavones ingredient, decompression and solvent recovery, is dried, and obtains Fructus Evodiae Folium Acanthopanacis Radicis total flavones and extracts
Thing C.
In dried Fructus Evodiae Folium Acanthopanacis Radicis extractive of general flavone C, the amount of total flavones is 81.9%, and wherein rutin content is 75%, and quercetin content is
2.1%, kaempferol content is 0.32%, and Determination of Hyperoside is 4.48%.
Embodiment 4
Folium Acanthopanacis Radicis extractive of general flavone is presented herein below and carries out animal experiment in order to activity and the application of extract of the present invention to be described.
One, the Folium Acanthopanacis Radicis extractive of general flavone impact on mouse cardiac muscle hypoxic-ischemic
1. laboratory animal Kunming mouse 110, male and female half and half, Hunan Si Laike Jing Da laboratory animal company limited provide.
2. prepared by the present embodiment 1-3 respectively for reagent thing Folium Acanthopanacis Radicis total flavones A, B, C；Breviscapine, by YUNNAN BAIYAO, Group Co., Ltd gives birth to
Produce, isoproterenol, urethane.
3. experimental technique
(1) mouse cardiac muscle ischemia model sets up Kunming mouse 60, is randomly divided into 6 groups, respectively normal group, model group, positive control drug group
(breviscapine EB, 10mg/kg), Folium Acanthopanacis Radicis flavone extract A, B, C group, often group 10.Mouse peritoneal injection urethane (112mg/kg)
Anesthesia, traces standard limb II with BL-420E+ biological functional system and leads normal ECG, and then tail vein injection (iv) is administered, wherein,
Normal group and the normal saline containing DMSO of model group mice iv equivalent, mice iv is accordingly by reagent for other groups.Iv is administered 1h, subcutaneous
Injection (sc) isoproterenol (Iso) 20mg/kg, record injection Iso after 1,2,5,15,30, the electrocardiogram of each time point of 60min
(ECG) change of ST section, is observed.Test after terminating, the blood remained in taking out rapidly heart normal saline flushing extrusion heart, then with filtering
Paper blots after making homogenate, is placed in-20 DEG C of Refrigerator stores.Strict test kit of pressing operates LDH, MDA, SOD and GSH-PX etc. in detection cardiac muscular tissue
Biochemical indicator.
(2) mice trachea folder closed model Kunming mouse 50, male and female half and half, be randomly divided into 5 groups, respectively blank group, positive controls
(breviscapine EB, 10mg/kg) and Folium Acanthopanacis Radicis flavone extract A, B, C group, often group 10.Each group mouse peritoneal injection urethane (112g/kg)
Anesthesia, fixing, trace standard limb II lead electrocardiogram with BL-420E+ biological functional system, then iv is administered, separating mouse after 1h
Trachea folder close, and record folder is held one's breath the time that Guan Houzhi electrocardiogram is wholly absent.
(3) statistical procedures experimental data uses SPSS 13.0 statistical software to carry out one factor analysis of variance.The neat person of variance uses LSD inspection to carry out group
Between compare, heterogeneity of variance person use Tamhanest2 inspection organize between compare.
4. experimental result
(1) Iso is caused impact that myocardial ischemia mice ECG ST section changes from figure 3, it can be seen that model group mice 60min upon administration
In each observing time put its ST section and all have raising in various degree.And Folium Acanthopanacis Radicis extractive of general flavone can cause suppression Iso in various degree
Raising of ST section, notable (P ＜ 0.01) with model group comparing difference.Positive control drug can also suppress raising of the ST section that Iso causes.
(2) Iso is caused the impact of LDH activity in myocardial ischemia mouse cardiac muscle ischemic myocardial tissue and the results are shown in Table 1.In model group murine myocardium
The activity of LDH significantly reduces, and has significance with normal group comparing difference.Identical with 10mg/kg EB, Folium Acanthopanacis Radicis extractive of general flavone A, B, C
Group has significant inhibitory action (P ＜ 0.01) to the reduction of LDH vigor in cardiac muscular tissue.
(3) Iso is caused the impact of MDA content and SOD and GSH-Px activity in myocardial ischemia mouse cardiac muscle ischemic myocardial tissue and the results are shown in Table 3.Mould
In type group murine myocardium, MDA content dramatically increases, and the activity of SOD and GSH-Px significantly reduces, and all has significantly with normal group comparing difference
Property.Identical with 10mg/kg EB, Folium Acanthopanacis Radicis extractive of general flavone can significantly reduce MDA content in murine myocardium, raises the work of GSH-PX
Property；Notable increased SOD activity.
Table 1 Folium Acanthopanacis Radicis extractive of general flavone is on LDH, SOD, GSH-Px activity and the impact of MDA content in Iso Myocardial Ischemia mouse cardiac muscle
5. experiment conclusion
By this experimentation, show that Folium Acanthopanacis Radicis extractive of general flavone has certain protective role to mouse cardiac muscle hypoxic-ischemic.
Two, the Folium Acanthopanacis Radicis extractive of general flavone impact on mice anastalsis
1. laboratory animal health Kunming mouse 80, body weight 19～21g, male and female half and half, Hunan Si Laike Jing Da laboratory animal company limited carry
Supply.
2. prepared by the present embodiment 1-3 respectively for reagent thing Folium Acanthopanacis Radicis extractive of general flavone A, B, C；Prednisolone acetate, Shanghai Xinyi Pharmaceutical Factory has
Limit company.
3. experimental technique
(1) animal selects and 80 mices are randomly divided into 8 groups by packet, often group 10, and respectively blank group, Folium Acanthopanacis Radicis total flavones extracts
The high dose group of thing A, B, C group and low dose group, positive controls.Testing each group to fill respectively and raise relative medicine, blank group gives equivalent and steams
Distilled water, continuous 7 days.
(2) after cutting tail method mensuration bleeding time last administration 1h, being fixed by mice, dew rat-tail is in outward, with knife blade at Mouse Tail-tip 1/2
Cross-section (strictly controlling depth of cut), treats that blood overflows beginning timing voluntarily and sucks drop of blood once every 30s filter paper, until blood flow stops naturally
Till (depletion of blood when filter paper is inhaled), it is the bleeding time.
After slide method measures clotting time of mice last administration 1h, using internal diameter 1mm, the glass capillary of long 10cm is from mice endocanthion ball rear vein beard
Take blood, when blood is full of glass tubing, start timing, then fracture two ends glass capillary (about 0.5cm) every 30s, and slowly draw to the left and right
Opening, observe the place of fractureing and occur with or without solidifying silk, the time occurring to blood clotting silk is then clotting time.
(3), after total number of blood platelet measures last administration 1h, after inserting each Mus endocanthion ball respectively with the capillary glass tube of a diameter of 1mm, arteries and veins clump takes blood,
Detection mouse platelets sum.
(4), after abdominal cavity capillary permeability measures last administration 1h, mouse peritoneal liquid optical density (OD value) is measured with 721 type optical density instrument.
(5) data process and use SPSS17.0 statistical software to carry out statistical analysis, and experimental data represents with mean ± standard deviation, carries out variance analysis.
4. experimental result
The impact on mice bleeding and cotting time, platelet count and capillary permeability of Folium Acanthopanacis Radicis flavone extract A, B, C group, is shown in Table 2.Wherein
Prednisolone acetate, Folium Acanthopanacis Radicis flavone extract A, B, C group high dose group and low dose group all can obviously reduce mice bleeding time and clotting time
(PHigh＜ 0.01, PLow＜ 0.05)；Have a significant effect (P to total number of blood plateletHigh＜ 0.01, PLow＜ 0.05)；All can significantly reduce mouse peritoneal capillary
Vascular permeability, difference all has significance (PHigh＜ 0.01, PLow＜ 0.05).
Table 2 Folium Acanthopanacis Radicis extractive of general flavone is hemorrhage on mice, clotting time, platelet and the impact of capillary permeability
Note: compare with matched group, * P ＜ 0.05, * * P ＜ 0.01
5. experiment conclusion
By this research experiment, show that Folium Acanthopanacis Radicis extractive of general flavone has obvious anastalsis.
Three, mice antitussive action is studied by Folium Acanthopanacis Radicis extractive of general flavone
1. laboratory animal Male Kunming strain mice 80, are provided by Hunan Si Laike Jing Da laboratory animal company limited.
2. prepared by the present embodiment 1-3 respectively for reagent thing Folium Acanthopanacis Radicis extractive of general flavone A, B, C；Normal saline.
3. experimental technique
(1) take 80 male mice in kunming, be randomly divided into 8 groups, respectively matched group, positive controls, Folium Acanthopanacis Radicis extractive of general flavone A, B,
C respectively organizes high dose group and low dose group, often group 10.The corresponding solution of ig respectively, matched group ig equal-volume normal saline, every day 1 time, even
Continuous 6 days.Ammonia is at the uniform velocity sprayed into case with soniclizer with constant pressure by after only putting into bell jar after being administered 1h by mice by last, spraying
Time is 70s, observe cough number of times in its cough latent period (i.e. from the time starting to be sprayed to cause the 1st cough of mice) and 3min (with
Mice contraction of abdominal muscle, magnifies mouth and is as the criterion with cough simultaneously).And this experiment is carried out twice checking.
(2) data process: application SPSS 17.0 software carries out statistical analysis, and result, to represent, compares employing one factor analysis of variance between group.
4. experimental result
The results are shown in Table 3.Test result indicate that for twice, Folium Acanthopanacis Radicis extractive of general flavone high dose group and low dose group all can significantly extend ammonia and cause mice to cough
Cough incubation period.All can substantially reduce cough number of times in mice 3min.
Table 3 Folium Acanthopanacis Radicis extractive of general flavone causes the impact (n=10) of mouse cough to ammonia
Note: compare with matched group, * P ＜ 0.05, * * P ＜ 0.01
5. experiment conclusion
This experimental studies results shows, Folium Acanthopanacis Radicis extractive of general flavone has good antitussive action.
Four, the Folium Acanthopanacis Radicis extractive of general flavone anti-sunlight function to sunlight direct irradiation shaving mice
1. laboratory animal regular grade female mice 90, weight 17～20g, Hunan Si Laike Jing Da laboratory animal company limited provide.
2. prepared by the present embodiment 1-3 respectively for reagent thing Folium Acanthopanacis Radicis extractive of general flavone A, B, C；Positive control: SPF20 (PA++) fourth man is the most beautiful
White sunscreen cream.
3. experimental technique
(1) animal packet and model are set up and 90 mices are randomly divided into 9 groups, often group 10, and blank group, model group, Folium Acanthopanacis Radicis are the most yellow
Ketone extract A group, B group, the high dose group of C group and low dose group, positive controls.The coarse wool of Mus back is cut off with shears, then with disposable
Remaining fine hair is shaved except clean by Medical shaving knife, makes skin of back fully expose, shaving area about 2cm*1cm.Depilation 1 time weekly, after depilation
Clean with temperature clear water.
(2) it is administered and ultraviolet irradiates and normally feeds every day, give moisture and the food of abundance.Folium Acanthopanacis Radicis extractive of general flavone A group, B group, C group
High dose group and low dose group 30min before solarization press 0.4g/cm respectively in shaving mouse back2And 0.1g/cm2Smear relative medicine, comparison
Group 30min before solarization presses 0.5g/cm in shaving mouse back2Smearing the suitable Whitening sunscreen cream of SPF20 (PA++) fourth man, model group is not coated with any sun-proof
Frost, above 6 groups (segmentation completes to expose to mice face every time and perspires about 30min, will 6～11 o'clock July to 3 o'clock sun exposure 1h
Mice is moved from indoor rest 30min, then irradiates 30min again by upper method, and accumulative irradiation dose reaches 1h) Continuous irradiation 4 weeks.Blank group is not
Give sun exposure, be not the most coated with any sunscreen cream.Testing for the 4th weekend, mouse back loses hair or feathers again, and area is 2cm*1cm.Disconnected neck is put to death all
After mice, take off full thickness skin at the depilation of back immediately.
(3) each mice skin tissue specimen of preparing of skin histology homogenate takes 0.1～0.2g, rinses in ice normal saline, removes blood, filter paper
Wipe dry, shred piece of tissue with ophthalmology is little, pour in glass homogenate tube；Measure 9 times of cold salines with graduated cylinder, pour in homogenizer and be homogenized；
Centrifugation 10min with 3000r/min；Take supernatant and carry out index determining in strict accordance with SOD and MDA test kit requirement.
(4) data process and use SPSS17.0 statistical software to carry out statistical analysis, and experimental data represents with mean ± standard deviation, carries out variance analysis.
4. during experimental result respectively organizes laboratory animal skin, SOD activity and MDA changes of contents are shown in Table 4.Model group compares with blank group, little Corium Mus
SOD activity decrease in skin tissue, MDA content raises, and both differences the most statistically significant (P ＜ 0.01).Comparing with model group, total flavones carries
Take thing high dose group and low dose group is respectively organized SOD activity in mice skin tissue and all risen, and (P the most statistically significant with model group differenceHigh
＜ 0.01, PLow＜ 0.05), and respectively organize MDA content in mice skin tissue and decline, and (P the most statistically significant with model group differenceHigh＜ 0.01,
PLow＜ 0.05).Comparing with matched group, extractive of general flavone high dose group and low dose group respectively organize SOD activity difference in mice skin tissue statistics
Meaning (PHigh＜ 0.01, PLow＜ 0.05)；And respectively organize the statistically significant (P of MDA content difference in mice skin tissueHigh＜ 0.01, PLow＜ 0.05).
Table 4 is respectively organized SOD activity and MDA changes of contents in laboratory animal skin and is compared
Note: compare with model group,#P ＜ 0.05,##P ＜ 0.01；Compare with matched group,*P ＜ 0.05,**P ＜ 0.01.
5. experiment conclusion
This experimentation proves that Folium Acanthopanacis Radicis extractive of general flavone has anti-sunlight function to daylight induced mice shaving skin of back, and the sun-proof result of high dose group is slightly
Higher than the suitable Whitening sunscreen cream of SPF20 (PA++) fourth man.
Five, the experimentation that Folium Acanthopanacis Radicis extractive of general flavone suppression melanin is formed
1. Study on Its Antioxidant Activity in Vitro
(1) the preparation precision of DPPH solution weighs DPPH sample 0.1g in 50mL measuring bottle, in light protected environment, adds anhydrous alcohol solution also
It is settled to scale, obtains the DPPH storing solution of concentration 2mg/mL.From DPPH storing solution, pipette 5mL, be settled to 100mL with dehydrated alcohol,
To the DPPH solution that concentration is 0.1mg/mL.
(2) Folium Acanthopanacis Radicis extractive of general flavone A, B, C solution and the vitamin c solution of sample determination compound concentration 2mg/mL respectively, respectively takes 2mL
Adding 2mL DPPH solution, after mix homogeneously, at room temperature dark, lucifuge stands 30min, with the sample under 2mL dehydrated alcohol and this concentration of 2mL
Product solution is blank zeroing, measures absorbance At at 517nm., return to zero for blank with 4mL dehydrated alcohol meanwhile, measure 2mL anhydrous
Ethanol and 2mL DPPH solution lucifuge at room temperature dark stands the absorbance A 0 after 30min, the DPPH free radical scavenging activity computing formula of sample
For:
Clearance rate=(1-At/A0) * 100%.Wherein At is the absorbance of sample, and A0 is 2mL dehydrated alcohol and the absorbance of 2mL DPPH solution.
(3) experimental result the results are shown in Table 5.
Table 5 DPPH free radical scavenging activity results contrast
2. suppress the mensuration of tyrosinase activity
(1) preparation of solution
The preparation precision of TYR solution weighs TYR 0.0212g, and phosphate buffer is pre-dispersed, and ultrasonic 30min uses phosphate-buffered
Liquid is dissolved in 100mL volumetric flask surely.
The preparation of tyrosinase solution is that 200U/mL weighs a certain amount of tryrosinase according to enzyme activity unit, with phosphate buffer, constant volume in
In volume required volumetric flask, it is stored in refrigerator standby,.
The preparation of phosphate buffered saline(PBS) weighs Na2HPO4·12H2O7.5g, NaH2PO4·2H2O3.2g, fully dissolves constant volume in 250 with deionized water
In mL volumetric flask, it is configured to the phosphate buffer of the 0.2mol/L that pH value is 6.8.
(2) assay method
By " table 6 reactant liquor composition " with micropipettor pipette TYR, PBS and Folium Acanthopanacis Radicis extractive of general flavone A, B, the reactant liquor of C sample divide
It is not placed in 8 PE pipes, mixing, 37 DEG C of constant temperature 10min, is then respectively adding the tryrosinase of 1mL, react 10min, quickly use
Ultraviolet spectrophotometer measures its absorbance at 475nm and is labeled as A1, A2, Aa1、Aa2、Ab1、Ab2、Ac1、Ac2.Calculate sample as follows
The activity of product suppression tryrosinase:
Inhibitory activity against tyrosinase=[1-(ASample 2-ASample 1)/(A2-A1)] × 100% wherein, A1: do not add sample and do not add the reactant liquor of tryrosinase at 475nm
The absorbance that place records；A2: do not add sample and add the absorbance that the reactant liquor of tryrosinase records at 475nm；ASample 1: add sample and with cheese ammonia
The absorbance recorded at the reactant liquor 475nm of acid enzyme；ASample 2: add sample and add the absorbance that the reactant liquor of tryrosinase records at 475nm
Table 6 reactant liquor forms
(3) experimental result
3. experiment conclusion
In conjunction with removing the experiment of DPPH free radical and tyrosinase inhibition test, can come by the way of scavenging activated oxygen, suppression tryrosinase respectively
Suppress melanic generation, result display Folium Acanthopanacis Radicis extractive of general flavone that oxygen-derived free radicals and tryrosinase are respectively provided with stronger clearance rate and suppression ratio,
So it can be used for the preparation of skin-lightening cosmetic.
Above example has been shown and described the ultimate principle of the present invention and principal character and advantages of the present invention.The technical staff of the industry should
Solving, the present invention is not restricted to the described embodiments, and described in above-described embodiment and description is the principle of the present invention to be described rather than with any
Mode limits the scope of the present invention, and without departing from the scope of the invention, the present invention also has various changes and modifications, these changes and improvements
Both fall within claimed scope.Claimed scope is defined by appending claims and equivalent thereof.
Claims (10)
1. a Folium Acanthopanacis Radicis extractive of general flavone, it is characterised in that in described extract, the amount containing total flavones is 55%～100%, and wherein rutin content is
10%～99%, quercetin content is 0.05%～10%, and kaempferol content is 0.002%～0.5%, and Determination of Hyperoside is 0.5%～70%.
The most according to claim 1, the preparation method of Folium Acanthopanacis Radicis extractive of general flavone, it is characterised in that comprise the following steps:
(1) preparation of crude extract
Take Folium Acanthopanacis Radicis, with the solvent extraction 1 of 4～15 times of volumes～3 times, united extraction liquid, recycling design；Residue is fully dispersed with appropriate distilled water, stone
Oil ether extracts 3～8 times；Water layer is concentrated into appropriate volume, stand at low temperature 24h, takes residue, obtains total flavones crude extract.
(2) purification
Fully being dissolved by crude extract solvent, upper chromatographic column, methanol aqueous solution or ethanol water in varing proportions are eluting solvent, eluting appropriate volume,
Collect fraction, merge the fraction containing flavones ingredient, decompression and solvent recovery, be dried, obtain Folium Acanthopanacis Radicis extractive of general flavone.
The preparation method of Folium Acanthopanacis Radicis extractive of general flavone the most according to claim 2, it is characterised in that described Folium Acanthopanacis Radicis is rough leaf Folium Acanthopanacis Radicis, thin
Post Folium Acanthopanacis Radicis, Fructus Evodiae Folium Acanthopanacis Radicis, acanthopanax leucorrhizus Harms leaf, Cortex Acanthopanacis Giraldii leaf, Cortex Acanthopancis leaf, pubescence Folium Acanthopanacis Radicis, Folium Acanthopanacis Senticosi, Panax sessiliflorus leaf,
One or more in white hair Folium Acanthopanacis Radicis, Tang's Folium Acanthopanacis Radicis, island Folium Acanthopanacis Radicis, different post Folium Acanthopanacis Radicis, Min Jiashi Folium Acanthopanacis Radicis, intelligence different mountain Folium Acanthopanacis Radicis.
4. according to the preparation method of the Folium Acanthopanacis Radicis extractive of general flavone described in Claims 2 or 3, it is characterised in that described Folium Acanthopanacis Radicis preferred rough leaf slender acanthopanax
Leaf.
The preparation method of Folium Acanthopanacis Radicis extractive of general flavone the most according to claim 2, it is characterised in that: the Extraction solvent described in step (1) is
Methanol, ethanol, water or its mixture.
The preparation method of Folium Acanthopanacis Radicis extractive of general flavone the most according to claim 2, it is characterised in that: the extracting method described in step (1) is
One or more in the extraction of heating and refluxing extraction, homogenate extraction, supersound extraction, merceration.
The preparation method of Folium Acanthopanacis Radicis extractive of general flavone the most according to claim 2, it is characterised in that: the absorption of chromatographic column described in step (2)
Agent is polyamide, or D101, HPD-100, AB-8, X-5, NKA-9 type macroporous adsorbent resin.
The preparation method of Folium Acanthopanacis Radicis extractive of general flavone the most according to claim 2, it is characterised in that: the drying means described in step (2) is
One or more in drying, vacuum drying, microwave drying, lyophilization.
Folium Acanthopanacis Radicis extractive of general flavone the most according to claim 1, for preparing treatment cardiac-cerebral ischemia, hemostasis and the medicine of cough-relieving.
Folium Acanthopanacis Radicis extractive of general flavone the most according to claim 1, is used for preparing sun-proof skin-lightening cosmetic.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108276375A (en) * | 2018-02-02 | 2018-07-13 | 陇东学院 | A method of isolating Stilbene effective constituents from acanthopanax leucorrhizus Harms stem skin |
| CN109730947A (en) * | 2018-11-04 | 2019-05-10 | 长沙博海生物科技有限公司 | With the active rough leaf slender acanthopanax method for preparing extractive of whitening anti-wrinkling and the application in cosmetics |
| CN116965337A (en) * | 2023-09-22 | 2023-10-31 | 东北林业大学 | Method for synchronously promoting growth and flavone accumulation of acanthopanax embryogenic callus |
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| CN1813834A (en) * | 2006-03-03 | 2006-08-09 | 中国科学院长春应用化学研究所 | Method for preparing total flavone extract of many prickle acanthopanax |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108276375A (en) * | 2018-02-02 | 2018-07-13 | 陇东学院 | A method of isolating Stilbene effective constituents from acanthopanax leucorrhizus Harms stem skin |
| CN109730947A (en) * | 2018-11-04 | 2019-05-10 | 长沙博海生物科技有限公司 | With the active rough leaf slender acanthopanax method for preparing extractive of whitening anti-wrinkling and the application in cosmetics |
| CN116965337A (en) * | 2023-09-22 | 2023-10-31 | 东北林业大学 | Method for synchronously promoting growth and flavone accumulation of acanthopanax embryogenic callus |
| CN116965337B (en) * | 2023-09-22 | 2023-12-01 | 东北林业大学 | Method for synchronously promoting growth and flavone accumulation of acanthopanax embryogenic callus |
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