CN101637492A - Preparation method of total flavonoids in persimmon leaves and application of total flavonoids in liver-protecting drugs and health foods - Google Patents
Preparation method of total flavonoids in persimmon leaves and application of total flavonoids in liver-protecting drugs and health foods Download PDFInfo
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Abstract
A preparation method of total flavonoids in persimmon leaves comprises the following steps: using 70% ethanol to be heated in water bath at a temperature of 75 DEG C to carry out reflux and extraction, using 70% ethanol to be heated by microwave radiation to carry out reflux and extraction or using 70% ethanol to be heated by ultrasonic waves to carry out reflux and extraction; combining extract,using 2% HCl aqueous solution to regulate the pH value to 5.5, enabling the extract to pass through LX-28 type large pore resin absorption columns or LX-38 type large pore resin absorption columns, washing until effluent is colorless, using 70% of ethanol solution for elution and collecting eluent; decompressing and concentrating the eluent at a temperature of 60 DEG C to obtain extract with relative density being 1.30-1.40g/cm<3>, placing the extract into a freeze-drying oven and drying the extract at a temperature of minus 50 DEG C to obtain the total flavonoids in persimmon leaves. The invention also provides application of the active ingredients- total flavonoids in liver-protecting drugs and health foods. Drug effect tests show that the total flavonoids can reduce the contents of ALTand AST in the serums and MDA in the livers, increase the activities of SOD and CAT in the livers, reduce the injury degrees of the liver cells and play an important role in preventing and treating liver diseases.
Description
Technical field
The invention belongs to the medicinal preparation technical field of the product that contains raw material or itself and not clear structure, be specifically related to derive from the material of plant.
Background technology
Folium Kaki is the fresh or dried leaves of Ebenaceae calamander Fructus Kaki (Diospyros kaki L.f), bitter in the mouth, cold in nature, and its beginning of being used as medicine sees bright " the southern regions of the Yunnan Province book on Chinese herbal medicine " record and " applies the swelling skin ulcer through red leaves ", and " book on Chinese herbal medicine is new again " record is used for " control cough and spit blood, slake thirst and help produce saliva ".Folium Kaki has been made into Chinese patent medicine, is used for the treatment of coronary heart disease, angina pectoris etc., also has multiple medical health care functions such as anti-inflammation, promoting the production of body fluid to quench thirst, heat-clearing and toxic substances removing, lung moistening heart tonifying, antitussive hemostasis, anti-cancer and cancer-preventing.China's Folium Kaki aboundresources, in recent years,, more and more deep to the research of calamander along with the develop rapidly of modern extraction and separation technology and modern pharmacology.
Main active in the Folium Kaki is a Flavonoid substances, the Folium Kaki total flavonoids substance of having found has isoquercitin, kaempferol, Quercetin, rutin, astragaloside, Rhizoma Kaempferiae phenolic glycoside, Quercetin glycosides, hyperin and astragalin etc., and the composition and the content that adopt diverse ways to extract the gained Flavonoid substances all exist than big-difference.
As everyone knows, liver has the function of biotransformation sugar, lipid, protein, vitamin, hormone and generation, secretion of bile and picked-up, conversion, the bilirubinic effect of drainage.Liver is intravital detoxifcation organ of animal and important metabolism place.Current, because the influence of each side such as environmental pollution and medicine, hepatic disease has become one of primary root of serious threat human health, and the hepatic injury due to the various harmful factors can show as hepatic necrosis, fatty liver, cholestasis, hepatic fibrosis, liver cirrhosis and hepatocarcinoma etc.The control of hepatic injury is still a global severe problem at present.Therefore, by setting up experimental liver damage animal model, the mechanism of research hepatopathy is screened hepatic, explores the hepatoprotective effect principle, is of great practical significance.
Summary of the invention
A technical problem to be solved by this invention is to provide a kind of extraction preparation method of Folium Kaki total flavones.
Another technical problem to be solved by this invention is to provide a kind of new purposes for the Folium Kaki total flavones.
Solving the problems of the technologies described above the technical scheme that is adopted is made up of following step:
1, take by weighing the Folium Kaki dried powder, the mass concentration that adds 20 times of amounts is 70% ethanol in 75 ℃ of heating in water bath reflux, extract, 1 hour, filters, and filtering residue is respectively 70% ethanol extraction twice again with the mass concentration of 10 times of amounts, each 1 hour.
2, each time extracting solution is merged, with mass concentration is that 2% HCl aqueous solution is regulated pH value to 5.5, crosses LX-28 type macroporous adsorptive resins or LX-38 type macroporous adsorptive resins, washes with water to effluent colourless, with mass concentration is 70% alcoholic solution eluting, collects eluent.
3, eluent being evaporated to 60 ℃ of relative densities is 1.30～1.40g/cm
3, placing in the freeze drying box ,-50 ℃ of dryings obtain the Folium Kaki total flavones.
In above-mentioned steps 1, can be that 2000～3000MHz, power are to reflux 10 minutes under the carry out microwave radiation heating of 400～450W also, filter that filtering residue is respectively 70% ethanol microwave extraction twice again with the mass concentration of 20 times of amounts, each 10 minutes in frequency.
Can also be that 40～50KHz, power are to reflux 30 minutes under the ultrasound wave heating of 220W in frequency in above-mentioned steps 1, filtering residue be respectively 70% ethanol ultrasonic extraction twice again with the mass concentration of 20 times of amounts, each 30 minutes.
The outward appearance of above-mentioned extract Folium Kaki total flavones is the pale yellow powder shape, contains isoquercitin, kaempferol, Quercetin, rutin, astragaloside, Rhizoma Kaempferiae phenolic glycoside, Quercetin glycosides, hyperin, Herba Astragali Melilotoidis (Herba Astragali Sinici) glycosides etc.
The application of Folium Kaki total flavones in preparation hepatic and health food.
The medicine that the preparation of effective ingredient Folium Kaki total flavones protects the liver uses with the form of conventional pharmaceutical formulation.Described conventional pharmaceutical formulation contains the Folium Kaki total flavones as active component, and this active component mixes as solid or the liquid excipient that is suitable for the organic or inorganic of administration in the gastrointestinal with pharmaceutically acceptable carrier in preparation.This pharmaceutical formulation can be solid form such as tablet, granule, capsule; Also can be liquid form such as suspending agent, syrup, Emulsion etc.
Can contain auxiliary substance, stabilizing agent, wetting agent, solubilizing agent and other additive commonly used in the above-mentioned preparation, as lactose, Pulvis Talci, cellulose, polyvinylpyrrolidone, starch, pectin, tween 80, polyvinyl alcohol etc.
Above-mentioned preparation can be made according to the preparation technology of various preparation routines.
Containing Folium Kaki total flavones in the Folium Kaki total flavones hepatic, to contain mass ratio be 1%～50% active component, preferably contains mass ratio and be 5%～20% active component.
Active ingredient Folium Kaki total flavones also can add in the food such as milk product, beverage, Folium Camelliae sinensis, bread, makes dissolving liver-protecting healthy food.
Active component Folium Kaki total flavones of the present invention is made the oral drugs of various dosage forms, becomes human oral, 200mg of common dose, and 2 times on the 1st, one after each meal, children is taken the circumstances into consideration decrement.In health food every day consumption 400mg, but life-time service.
Indication of the present invention: treatment hepatopathy and protecting the liver.
The medicinal preparation for oral administration that said extracted thing Folium Kaki total flavones is made according to a conventional method is said tablet or granule or capsule or an oral liquid on the galenic pharmacy.
Medicine of the present invention shows through the test of pesticide effectiveness, and the Folium Kaki total flavones can suppress effectively that the activity of glutamic oxaloacetic transaminase, GOT (ALT) and glutamate pyruvate transaminase (AST) rises in the carbon tetrachloride induced mice serum, promptly reduces the level of Serum ALT and AST; Can obviously reduce CCl
4Cause the content of malonaldehyde in the mouse liver injury hepatic tissue (MDA), improve total superoxide dismutase (SOD) and catalase (CAT) activity.The electron microscopic observation result shows that the Folium Kaki total flavones can reduce hepatocellular damage.
Animal vivo test studies show that the Folium Kaki total flavones can be removed the free radical that is compromised cell, antagonism CCl effectively
4The hepatocyte free radical and the lipid peroxidation injury that bring out, have significant protection and the treatment hepatic injury effect, do not observe in test significant side effects.Can be used as hepatic and health food.Folium Kaki total flavones acute toxicity test in mice shows that the administration of Folium Kaki total flavones mice can not asked LD50.
The Folium Kaki total flavones can reduce the effect of Serum ALT and AST significantly, and can obviously reduce the content of liver MDA, improves liver SOD and CAT activity, significantly reduces hepatocellular degree of injury, and prevention and treatment hepatic disease are had important function.Medicine with the preparation of Folium Kaki total flavones can enter clinical trial.
Description of drawings
Fig. 1 is the high-efficient liquid phase chromatogram of the Folium Kaki total flavones of embodiment 1 extraction.
Fig. 2 is CCl
4The painted electron microscopic observation result of hepatic injury mouse liver cell HE.
The specific embodiment
The present invention is described in more detail below in conjunction with embodiment, but the invention is not restricted to these embodiment.
Embodiment 1
With dry Folium Kaki 10g is that example used other raw materials and extracting method thereof are as follows:
1, take by weighing Folium Kaki dried powder 10g, the mass concentration that adds 20 times of amounts is 70% ethanol in 75 ℃ of heating in water bath reflux, extract, 1 hour, filters, and filtering residue is respectively 70% ethanol extraction twice again with the mass concentration of 10 times of amounts, each 1 hour.
2, each time extracting solution being merged, is 2% HCl aqueous solution adjust pH to 5.5 with mass concentration, crosses semipolar LX-28 type macroporous adsorptive resins, washes with water to effluent colourlessly, is 70% alcoholic solution eluting with mass concentration, the collection eluent.
3, eluent being evaporated to relative density is 1.30～1.40g/cm
3(60 ℃) place in the freeze drying box, and-50 ℃ of dryings obtain Folium Kaki total flavones 391mg, and general flavone content in the extract (in rutin) is 72.1%.
The Folium Kaki total flavones that adopts the inventive method to extract is analyzed with high performance liquid chromatography, and chromatographic column is Phenonenex Luna C18 (250 * 4.6nm, 5 μ m); Mobile phase is methanol (A), 0.5% phosphoric acid (B); Gradient elution is 0-5 minute 13%A, 5-8 minute 13-18%A, 8-25 minute 18-19%A, 25-40 minute 19-20%A, 40-50 minute 20-60%A, 50-60 minute 60-13%; Flow velocity is 1.0ml/min; The detection wavelength is 360nm.High-efficient liquid phase chromatogram is seen Fig. 1.As seen from Figure 1, the Folium Kaki total flavones contains the composition more than 10 kinds.
Embodiment 2
With dry Folium Kaki 10g is that example used other raw materials and extracting method thereof are as follows:
1, take by weighing Folium Kaki dried powder 10g, the mass concentration that adds 20 times of amounts is 70% ethanol in being that 2450MHz, power are to reflux 10 minutes under the carry out microwave radiation heating of 400-450W in frequency, filters; The mass concentration of 20 times of amounts of filtering residue reuse be 70% ethanol microwave extraction 10 minutes once.
2, extracted twice liquid being merged, is 2% HCl aqueous solution adjust pH to 5.5 with mass concentration, crosses semipolar LX-28 type macroporous adsorptive resins, washes with water to effluent colourlessly, is 70% alcoholic solution eluting with mass concentration, the collection eluent.
3, with eluent decompression, be concentrated into to relative density be 1.30～1.40g/cm
3The extractum of (60 ℃) places in the freeze drying box, and-50 ℃ of dryings obtain Folium Kaki total flavones 586mg, and general flavone content in the extract (in rutin) is 72.5%.
Embodiment 3
With dry Folium Kaki 10g is that example used other raw materials and extracting method thereof are as follows:
1, takes by weighing Folium Kaki dried powder 10g, the mass concentration that adds 20 times of amounts is 70% ethanol in frequency is that 40～50KHz, power are to reflux 30 minutes under the ultrasound wave heating of 220W, filtering residue more respectively with the mass concentration of 20 times of amounts be 70% ethanol ultrasonic extraction 30 minutes once.
2, extracted twice liquid being merged, is 2% HCl aqueous solution adjust pH to 5.5 with mass concentration, crosses the LX-38 type macroporous adsorptive resins of low pole, washes with water to effluent colourlessly, is 70% alcoholic solution eluting with mass concentration, the collection eluent.
3, eluent being evaporated to relative density is 1.30～1.40g/cm
3The extractum of (60 ℃) places in the freeze drying box, and-50 ℃ of dryings get Folium Kaki total flavones 567mg, and general flavone content in the extract (in rutin) is 71.5%.
Embodiment 4
To produce Folium Kaki total flavones hepatic granule product 1000 grams is that the used Folium Kaki total flavones of example and adjuvant and proportioning thereof are:
Folium Kaki total flavones 20g
Sucrose 400g
Dextrin adds to 1000g.
The Folium Kaki total flavones is added ethanol make and contain the alcohol amount and reach 60%, left standstill 24 hours, get supernatant, decompression recycling ethanol is not to there being the alcohol flavor, and being concentrated into relative density is 1.30～1.40g/cm
3The clear paste of (60 ℃) adds dextrin, sucrose, and mixing is made granule, drying, pack, every bag heavy 10g, every gram contains Folium Kaki total flavones 20mg.
To produce 1000 of Folium Kaki total flavones hepatic tablet products is that the used Folium Kaki total flavones of example and adjuvant and proportioning thereof are:
Folium Kaki total flavones 100g
Starch adds to 400g.
Its preparation method is undertaken by the preparation method of galenic pharmacy tablet.Every heavy 0.4g, every contains Folium Kaki total flavones 100mg.
To produce 1000 of Folium Kaki total flavones hepatic capsule products is that the used Folium Kaki total flavones of example and adjuvant and proportioning thereof are:
Folium Kaki total flavones 100g
The 3-cyclodextrin adds to 300g.
Its preparation method is undertaken by the preparation method of galenic pharmacy capsule.Every heavy 0.3g, every contains Folium Kaki total flavones 100mg.
To produce Folium Kaki total flavones hepatic oral liquid 1000mL is that the used Folium Kaki total flavones of example and adjuvant and proportioning thereof are:
Folium Kaki total flavones 20g
Sucrose 400g
Distilled water adds to 1000mL.
Its preparation method is undertaken by the preparation method of galenic pharmacy oral liquid.Every bottle of 10mL, every milliliter contains Folium Kaki total flavones 20mg.
Embodiment 5
Get Folium Kaki total flavones 20g, with the dissolve with ethanol of 10 times of amounts, evenly be sprayed onto on the 980g Folium Camelliae sinensis, make that the Folium Kaki content of total flavone is 20mg in every gram Folium Camelliae sinensis, oven dry is distributed into the teabag of every bag of 10g, drinks 2 bags every day, but long-term drink.
Embodiment 6
Get Folium Kaki total flavones 1.6g, suspend with sodium carboxymethyl cellulose, add in the 970mL milk and make milk product with hepatoprotective effect with 0.3% food of 30mL, every 100mL contains Folium Kaki total flavones 160mg, sterilization back packing, every bag of 250mL, drink for each person every day and be no more than 250mL, but long-term drink.
Embodiment 7
Get Folium Kaki total flavones 4g, vegetable oil 50g, the 20g active yeast, standard flour adds to 1000g, carries out according to conventional breadmaking technology.Edible for each person every day 100g contains Folium Kaki total flavones 400mg.The edible for each person every day 100g that is no more than.
In other food, in food such as cookies, cake, also can add the Folium Kaki total flavones, be advisable by Folium Kaki total flavones amount 400mg for each person every day.
In order to verify the Folium Kaki total flavones in the effect aspect prevention, the treatment hepatic injury, the applicant adopts the Folium Kaki total flavones of the embodiment of the invention 1 preparation to carry out pharmacodynamics test, and various test situation are as follows:
Be subjected to the reagent thing: the Folium Kaki total flavones, self-control is mixed with mass concentration and is 3% Folium Kaki total flavones aqueous solution according to a conventional method during test.
Animal subject: Kunming mouse, body weight 25～30g, the male and female dual-purpose is provided by Xi'an Communications University's medical experiment animal center.The animal quality certification is SCXK (Shan) 2007-001.
1, Folium Kaki total flavones acute toxicity test in mice
Get 20 of mices, male and female half and half, mouse stomach (300mg/kg) 3 times are given in fasting 12 hours in one day, each 4 hours at interval, make its day cumulative amount reach 900mg/kg, administration relief mice freely ingests and drinks water, and observes continuously 7 days, the record animal appearance, appetite, behavioral activity, toxicity situations such as death.After 7 days mice is put to death, dissect perusal main organs lesion tissue situation.
Result of the test: do not occur dead behind the mouse stomach.After irritating stomach for the first time, for the second time, activity shows no obvious abnormalities; After irritating stomach for the third time, the part mice autonomic activities occurs and reduces, closes symptoms such as order reposes, bradykinesia, but does not have abnormal conditions such as perpendicular hair lazyness is moving, tic, overturning, and movable as usual after 1-3 hour, stool dark brown matter is soft, rare sample do not occur just.Observed 7 days all survivals, all no abnormal variations such as diet, activity, the mental status, the fur colour of skin, defecation continuously.After 7 days mice is put to death, dissect, main organs such as the perusal heart, liver, spleen, lung, kidney there is no morphological abnormalities and change.
Conclusion (of pressure testing): the administration of Folium Kaki total flavones mice, can not ask LD50, maximum dosage-feeding is 800mg/kg.Press 60kg adult consumption 400mg and calculate, be equivalent to 72 times of people's consumption per day (800mg/kg * 60%/400mg * 60kg).The toxicity of Folium Kaki total flavones is less.
2, Folium Kaki total flavones pharmacodynamics test
Reagent: glutamic oxaloacetic transaminase, GOT (ALT), glutamate pyruvate transaminase (AST), malonaldehyde (MDA), total superoxide dismutase (SOD) and peroxidating hydrogenation enzyme (CAT) test kit and protein reagent box build up bio-engineering research by Nanjing to be provided; Bifendate drop pill is produced by Zhejiang Prov WanBang Pharmaceutical Co., Ltd, and lot number is 081010, and specification is the 1.5mg/ ball; Carbon tetrachloride (AR) is produced by Tianjin Chemical Reagents Factory No.1, is made into 0.2% solution during experiment with vegetable oil; Other reagent is homemade analytical pure.
Preparation bifendate solution: being mixed with mass concentration according to a conventional method is 1% bifendate aqueous solution.
Test method: get 48 of Kunming mouses, male and female half and half are divided into 6 groups every group 8, normal control group (NCG), CCl at random
4Model group (MCG), positive controls (LCG), treat basic, normal, high group (DKLF1-3,3 groups) and place under the ventilation condition, all animals can freely drink water and diet.Normal control group (NCG) and CCl
4Model group (MCG) is irritated stomach distilled water once (10mL/kg) every day, positive controls (LCG) is irritated stomach bifendate (100mg/kg) once every day, treat basic, normal, high group of (DKLF1-3, dosage is respectively 50mg/kg, 100mg/kg, 300mg/kg Folium Kaki total flavones) every day gastric infusion once, handled continuously 21 days.Tested the 22nd day, except that normal control group (NCG) waits the dosage vegetable oil, all the other each groups give 0.2% carbon tetrachloride vegetable oil solution (in advance through the ultrasound wave mixing), prepare acute liver tissue injury model, irritate the stomach amount all by body weight 10ml/kg, fasting was subsequently got blood from eye socket after 24 hours, and dislocation claims gross weight.
(1) the Folium Kaki total flavones is to CCl
4Lure the exponential influence of acute liver damage Mouse Liver into
Mice administration last day claims gross weight, and dislocation causes death, and dissects and takes out liver, rinse with ice-cold normal saline and wash bloodstain, and dip in filter paper dried, to the weighing of liver difference.Calculate the liver index by following formula:
L (liver index)=m
1(weight of liver)/m
2(mice body weight) * 100%
Result of the test sees Table 1.
Table 1 Folium Kaki total flavones is to CCl
4The influence of hepatic injury Mouse Liver index, Serum ALT and the AST that lures into
| Group | Liver index (%) | Glutamic oxaloacetic transaminase, GOT (ALT, U/L) | Glutamate pyruvate transaminase (AST, U/L) |
| ??NCG | ??4.22 ▲▲ | ??28.31 ▲▲ | ??49.56 ▲▲ |
| ??MCG | ??6.66 **■■ | ??69.8 **■■ | ??81.92 **■■ |
| ??LCG | ??4.72 ▲▲ | ??42.98 *▲▲ | ??56.18 *▲▲ |
| ??DKLF1 | ??6.25 **■ | ??65.86 **■■ | ??76.52 **■ |
| ??DKLF2 | ??5.02 *▲ | ??43.61 *▲ | ??62.29 *▲ |
| ??DKLF3 | ??4.29 ▲▲ | ??29.57 ▲▲ | ??50.6 ▲▲ |
Annotate: DKLF1 is a Folium Kaki total flavones low dose group in the table 1, and DKLF2 is a dosage group in the Folium Kaki total flavones, and DKLF3 is a Folium Kaki total flavones high dose group.
Compare with normal group (NCG), * * P＜0.01, compare with model group (MCG) * P＜0.05,
▲ ▲P＜0.01,
▲Compare with bifendate matched group (LCG) P＜0.05,
■ ■P＜0.01,
■P＜0.05.
By table 1 as seen, the liver index of model group (MCG) mice all has significant increase than normal group (NCG) and matched group (LCG), and has significant difference (P＜0.01), shows that the inflammation of model group mouse liver cell is serious, CCl
4Induced the damage of mouse liver cell; The liver index of matched group (LCG) mice is more not remarkable than normal group difference, Folium Kaki total flavones low dose group (DKLF1,50mg/kg) the liver index of mice is apparently higher than matched group, middle dosage group and its difference are little, when administration concentration reaches 300mg/kg, the liver index of hepatocyte injury mice returns to normal level basically, with normal group and matched group there are no significant difference.
Get leftlobe of liver 0.5 * 0.5 * 0.5m
3Fritter is put mass concentration and is in 4% the neutral buffered formalin, conventional dehydration, and paraffin embedding, HE dyeing is done in section, examines under a microscope hepatic tissue pathology and changes.Histopathologic examination the results are shown in Figure 2.(A) normal group (NCG), (B) model group (MCG), (C) bifendate matched group (LCG), (D) Folium Kaki total flavones low dose group (DKLE1), (E) dosage group (DKLE2) in the Folium Kaki total flavones, (F) Folium Kaki total flavones high dose group (DKLE3).
Fig. 2 shows that Folium Kaki flavone in treating group and model group compare, and low dose group does not have significant difference, middle dosage group, high dose group have obviously alleviated hepatocyte piecemeal necrosis, spotty necrosis and hepatocyte cloudy swelling hydropic degeneration, and the normal liver cell number increases, and is suitable with matched group, no significant difference.The result shows that the Folium Kaki total flavones is to CCl
4Lure that acute liver damage has tangible drug effect into.
(2) the Folium Kaki total flavones is to CCl
4Hepatic injury mice serum ALT that lures into and the influence of AST
CCl
4Fasting is 24 hours after the modeling, wins eyeball and gets blood, and separation of serum according to the test kit requirement, is surveyed Serum ALT and AST index.Result of the test sees Table 1.As shown in Table 1, CCl
4The model group serum alt is compared with matched group with normal group with AST after the modeling all significant difference (P＜0.01).Compare with model group, and Folium Kaki total flavones low dose group (DKLF1,50mg/kg) level of mice serum ALT and AST all has reduction, but difference is not remarkable; In dosage group (100mg/kg) mice serum ALT and AST obviously descend (P＜0.05); Its ALT of high dose group (300mg/kg) and AST level and model group significant difference (P＜0.01).Middle and high dosage group all with matched group (LCG) there was no significant difference.It is normal that the ALT of high dose group and AST level have reached substantially.Experimental result shows that the Folium Kaki total flavones can obviously reduce hepatic injury mice serum ALT and AST level, has the effect avoiding damaging of protection cell, the effect that has the identical acute carbon tetrachloride hepatic injury of protection with bifendate.
(3) the Folium Kaki total flavones is to CCl
4The influence of the hepatic injury mice MDA content of luring into
After animal dislocation is put to death, dissect and take out liver,, remove blood, wipe away driedly with filter paper, prepare liver homogenate with rinsing in the ice-cold normal saline.Survey the content of lipid peroxide (MDA) in the mouse liver according to the requirement of test kit.Result of the test sees Table 2.
Table 2 Folium Kaki total flavones is to CCl
4The influence of SOD, MDA and CAT in the hepatic injury Mouse Liver of luring into
| Group | Lipid peroxide (MDA, nmol/mg prot) | Total superoxide dismutase (SOD, U/mgprot) | Catalase (CAT, U/mgprot) |
| ??NCG | ??7.18 ▲▲ | ??182.99 ▲▲ | ??35.13 ▲▲ |
| ??MCG | ??14.08** ■■ | ??135.25** ■■ | ??15.15** ■■ |
| ??LCG | ??8.6 ▲▲ | ??176.11 ▲▲ | ??30.72 ▲▲ |
| ??DKLF1 | ??13.26** ■ | ??140.34** ■■ | ??16.73** ■■ |
| ??DKLF2 | ??9.86* ▲ | ??167.81* ▲ | ??26.16* ▲■ |
| ??DKLF3 | ??7.42 ▲▲ | ??184.84 ▲▲ | ??34.06 ▲▲■ |
Annotate: DKLF1 is a Folium Kaki total flavones low dose group in the table 2, and DKLF2 is a dosage group in the Folium Kaki total flavones, and DKLF3 is a Folium Kaki total flavones high dose group.
Compare * * P＜0.01, * P＜0.05 with normal group (NCG); Compare with model group (MCG),
▲ ▲P＜0.01,
▲P＜0.05; Compare with bifendate matched group (LCG),
■ ■P＜0.01,
■P＜0.05.
As can be seen from Table 2, model group (MCG) hepatic tissue MDA content is compared with normal group (NCG), significant rising (P＜0.01) is arranged, promptly at CCl
4Inducing under, caused the lipid peroxidation injury of mouse liver, caused the generation of MDA.Matched group (LCG) is higher slightly than normal group (NCG) MDA content, but there was no significant difference.(MCG) compares with model group, Folium Kaki total flavones (DKLF) is along with its hepatic tissue of increase MDA content of drug dose also significantly reduces, its MDA content of high dose group (300mg/kg) has dropped to normal level, and it is also not remarkable that high dose and middle dosage group and matched group (LCG) are compared difference.The result shows that the Folium Kaki total flavones can reduce CCl
4Induce the content of lipid peroxide MDA in the murine liver tissue of initiation, the Folium Kaki total flavones has been brought into play effect in the mice body, can alleviate the degree of hepatocyte damaged.
(4) the Folium Kaki total flavones is to CCl
4The active influence of SOD in the hepatic injury Mouse Liver of luring into
After animal dislocation is put to death, dissect and take out liver,, remove blood, wipe away driedly with filter paper, prepare liver homogenate with rinsing in the ice-cold normal saline.SOD vigor in the liver is surveyed in requirement according to test kit.Result of the test sees Table 2.Total superoxide dismutase (SOD) vigor is starkly lower than normal group (NCG) and matched group (LCG) in model group (MCG) hepatic tissue as can be seen from Table 2, and significant difference (P＜0.01) is all arranged, and illustrates that hepatocyte injury is serious.(MCG) compares with model group, three dosage groups of Folium Kaki total flavones (DKLF) SOD activities of liver also increases significantly, learn Treatment Analysis by statistics, except that low dose group (50mg/kg), other two groups (100,300mg/kg) all have significant difference (P＜0.05 or P＜0.01), and have tangible dose-effect relationship.Middle dosage group (DKLF2) is compared with matched group (LCG) with high dose group (DKLF3) SOD vigor, and difference is not remarkable.The result shows that the Folium Kaki total flavones can suppress CCl
4Due to hepatic SOD descend; be that the Folium Kaki total flavones can be by a series of indirect Mechanism of Physiological and Biochemical; promote the biosynthesis or the activation of antioxidases such as body SOD; improve the oxidation resistance of body; remove superoxide radical protection cell and avoid damage; alleviate the damage of coercing of cell, play the effect of protection cell.
(5) the Folium Kaki total flavones is to CCl
4The influence of the hepatic injury mice CAT vigor of luring into
Test CCl
4Induce the variation that causes damage Mouse Liver CAT, the results are shown in Table 2, compare, model group (MCG) CAT significantly descend (P＜0.01) with normal group (NCG) hepatic tissue catalase (CAT) vigor; (MCG) compares with model group, the treatment group except that the low dosage (50mg/kg), the CAT vigor of all the other two groups of mouse livers all has increase in various degree, and there is tangible dose-effect relationship, when administration concentration reaches 300mg/kg, Mouse Liver CAT vigor returns to normal level substantially, and not remarkable with matched group difference.The result shows that the Folium Kaki total flavones can improve ring CCl
4Induced mice hepatic injury effect, the influence of removing free radical, the vigor of CAT in the raising mice body.
(6) conclusion (of pressure testing)
The activity that the Folium Kaki total flavones can suppress glutamic oxaloacetic transaminase, GOT (ALT) and glutamate pyruvate transaminase (AST) in the carbon tetrachloride induced mice serum effectively rises, and promptly reduces the level of Serum ALT and AST; Can obviously reduce CCl
4Cause the content of malonaldehyde in the mouse liver injury hepatic tissue (MDA), improve total superoxide dismutase (SOD) and catalase (CAT) activity.The electron microscopic observation result shows that also the Folium Kaki total flavones can reduce hepatocellular damage.
Result of the test show the Folium Kaki total flavones can remove effectively some the infringement cells free radical, antagonism CCl
4The hepatocyte free radical and the lipid peroxidation injury that bring out have significant protection and treatment hepatic injury effect, can be used as treatment hepatopathy and hepatic and health food.
The function of Folium Kaki total flavones medicine with cure mainly: treatment hepatopathy or protect the liver.
The specification of Folium Kaki total flavones medicine: every bag heavy 10g of Folium Kaki total flavones medicinal granule, every gram contains Folium Kaki total flavones 20mg; Every heavy 0.4g of Folium Kaki total flavones medicinal tablet, every contains Folium Kaki total flavones 100mg; Every heavy 0.3g of Folium Kaki total flavones medicine capsule, every contains Folium Kaki total flavones 100mg; Every bottle of 10mL of Folium Kaki total flavones medicine oral liquid, every milliliter contains Folium Kaki total flavones 20mg.
The usage and the consumption of Folium Kaki total flavones medicine: the oral drugs granule, boiled water is taken after mixing it with water, one time 1 bag, each 1 time sooner or later; The oral drugs tablet, one time 2,2 times on the one; The oral drug capsule agent, one time 2,2 times on the one; The oral drugs oral liquid, one time 1 bottle, 2 times on the one.
Claims (2)
1, a kind of preparation method of Folium Kaki total flavones is characterized in that being made up of following step:
(1) take by weighing the Folium Kaki dried powder, the mass concentration that adds 20 times of amounts is 70% ethanol in 75 ℃ of heating in water bath reflux, extract, 1 hour, filters, and filtering residue is respectively 70% ethanol extraction twice again with the mass concentration of 10 times of amounts, each 1 hour; Or take by weighing the Folium Kaki dried powder, the mass concentration that adds 20 times of amounts is 70% ethanol in being that 2000～3000MHz, power are to reflux 10 minutes under the carry out microwave radiation heating of 400～450W with frequency, filter, filtering residue is respectively 70% ethanol microwave extraction twice again with the mass concentration of 20 times of amounts, each 10 minutes; Or take by weighing the Folium Kaki dried powder, the mass concentration that adds 20 times of amounts is 70% ethanol in frequency is that 40～50KHz, power are to reflux 30 minutes under the ultrasound wave heating of 220W, filtering residue is respectively 70% ethanol ultrasonic extraction twice again with the mass concentration of 20 times of amounts, each 30 minutes;
(2) each time extracting solution is merged, with mass concentration is 2% HCl aqueous solution adjust pH to 5.5, crosses LX-28 type macroporous adsorptive resins or LX-38 type macroporous adsorptive resins, washes with water to effluent colourless, with mass concentration is 70% alcoholic solution eluting, collects eluent;
(3) 60 ℃ of eluents being evaporated to relative density is 1.30～1.40g/cm
3, placing in the freeze drying box ,-50 ℃ of dryings obtain the Folium Kaki total flavones.
2, the application of effective ingredient Folium Kaki total flavones in preparation hepatic and health food.
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| CN102068483A (en) * | 2011-01-07 | 2011-05-25 | 河南中医学院 | Persimmon leaf extract for improving brain ischemic tolerance |
| CN102178722A (en) * | 2011-05-06 | 2011-09-14 | 陕西科技大学 | Persimmon leaf general flavone sustained-release micropills and preparation method thereof |
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| CN104689600A (en) * | 2015-03-10 | 2015-06-10 | 沈阳东新药业有限公司 | Extraction and purification method for persimmon leaf total flavonoids |
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| CN103284148B (en) * | 2013-05-20 | 2015-04-15 | 国家林业局泡桐研究开发中心 | Processing method of persimmon leaf concentrated powder rich in natural vitamin C |
| CN104689600A (en) * | 2015-03-10 | 2015-06-10 | 沈阳东新药业有限公司 | Extraction and purification method for persimmon leaf total flavonoids |
| CN104689600B (en) * | 2015-03-10 | 2016-07-06 | 沈阳东新药业有限公司 | The extraction of Folium Kaki total flavones and purification process |
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| CN108279278B (en) * | 2017-12-29 | 2020-09-04 | 广州白云山和记黄埔中药有限公司 | Method for separating flavonoid components and application thereof |
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