CN106511528A - Chaenomeles speciosa extract, chaenomeles speciosa extract composition and uses of chaenomeles speciosa extract and chaenomeles speciosa extract composition - Google Patents
Chaenomeles speciosa extract, chaenomeles speciosa extract composition and uses of chaenomeles speciosa extract and chaenomeles speciosa extract composition Download PDFInfo
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- CN106511528A CN106511528A CN201610899003.2A CN201610899003A CN106511528A CN 106511528 A CN106511528 A CN 106511528A CN 201610899003 A CN201610899003 A CN 201610899003A CN 106511528 A CN106511528 A CN 106511528A
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- wrinkled papaya
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- singer
- bull
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- A—HUMAN NECESSITIES
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- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/06—Fungi, e.g. yeasts
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- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/23—Apiaceae or Umbelliferae (Carrot family), e.g. dill, chervil, coriander or cumin
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- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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Abstract
The invention provides a chaenomeles speciosa extract and a composition thereof, wherein the real-time chaenomeles speciosa extract is the alcohol extract of chaenomeles speciosa, and the composition is the alcohol extract of chaenomeles speciosa and Laetiporus sulphureus, or the alcohol extract of chaenomeles speciosa, Laetiporus sulphureus and celery. According to the present invention, the Laetiporus sulphureus extract has a xanthine oxidase inhibition effect and can regulate uric acid metabolism so as to prevent and treat hyperuricemia and gout; and the combination of chaenomeles speciosa, Laetiporus sulphureus and celery can enhance the xanthine oxidase inhibition and regulate the uric acid metabolism.
Description
Technical field
The present invention provides a kind of wrinkled papaya extract and compositionss, and specially wrinkled papaya extract is wrinkled papaya alcohol
Extract, combinations thereof are wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract or wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. and Herba Apii graveolentis ethanol extract.Belong to food
Product, pharmaceutical field.
Background technology
Elephant skin wrinkled papaya is the dry of rosaceous plant chaenomeles lagenaria Chaenomeles speciosa (Sweet) Nakai
Dry mature fruit, main product, in Hubei Province Tujia Autonomous County of Changyang, are the famous product for food and medicine purposes of our province.With relaxing muscles and tendons and activating QI and blood in the collateral,
Effect of stomach dampness elimination, for treating the diseases such as pain in the low back and legs, rheumatic arthritis, muscular spasm of the limbs, big vomiting and diarrhoea, tinea pedis edema.
Also there are various effects such as antiinflammatory, antibacterial, lipid-loweringing, anti-.Wrinkled papaya not only has unique medical value, also with good
Nutrition, health-care effect.Its nutritive value is no less than Fructus actinidiae chinensis, among the people to have saying for " one benefit of Fructus Pruni, two benefit of pears, wrinkled papaya hundred being beneficial ",
With good Development volue.Wrinkled papaya is containing a large amount of organic acid, vitamin, multiple protein enzyme and a large amount of flavone, saponinss
Compound.Not only it is widely used in producing various Chinese patent medicines such as wrinkled papaya ball, wind network pain piece, wrinkled papaya wine, is additionally operable to production
The cosmetic bath articles such as wrinkled papaya skin care item, wrinkled papaya fancy soap, cleansing milk, bath gel, while and production wrinkled papaya
A series of raw material of health foods such as piece, beverage.
Existing reported in literature Fructus Chaenomeliss composition compound of Chinese medicine hyperuricemia and gout, its effect is mainly from anti-inflammatory mechanisms
Studied.We demonstrate that Fructus Chaenomeliss, Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer., by suppressing xanthine oxidase activity, reach and prevent and treat hyperuricemia and gout
Effect.Fructus Chaenomeliss functional food is provided from food angle for preventing and treating hyperuricemia and gout first.Define elephant skin wood
Melon prevents and treats the compositionss of hyperuricemia and gout with Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. or wrinkled papaya with Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. and Herba Apii graveolentis.It was found that sulfur mykol
Extract can be by suppressing xanthine oxidase activity, for preventing and treating hyperuricemia and gout.
The content of the invention
Elephant skin wrinkled papaya (Yichang City, Hubei Province Tujia Autonomous County of Changyang), Herba Apii graveolentis (commercially available), Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. source is (for examination
Strain Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. is separated by natural product key lab of Hubei Province of SanXia University and preservation).
Wrinkled papaya used is rosaceous plant chaenomeles lagenaria (Chaenomeles speciosa (Sweet) Nakai)
Dry mature fruit；Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. is On Polyporaceae Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. (Laetiporus sulphureus (Fr.) Murrill)
Mycelium or sporophore；Stem and leaf of the Herba Apii graveolentis for category samphire Herba Apii graveolentis (Apium graveolens L.).
The present invention provides a kind of wrinkled papaya extract and compositionss, and wrinkled papaya extract is wrinkled papaya ethanol extract,
Combinations thereof are wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract or wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. and Herba Apii graveolentis ethanol extract.
Described wrinkled papaya extract is to crush after wrinkled papaya is dried at 60-80 DEG C and cross 20-100 mesh, 50%-
95% alcohol reflux, concentrating under reduced pressure obtain final product wrinkled papaya ethanol extract after 30-60 DEG C of vacuum drying.
Described wrinkled papaya is wrinkled papaya and the drying at 60-80 DEG C respectively of sulfur mycelium with Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract
After crush and cross 20-100 mesh, 50%-95% alcohol reflux three times every time plus the 6-15 times of ethanol for measuring (w/w), merges filter
Liquid, concentrating under reduced pressure obtain final product wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract after 30-60 DEG C of vacuum drying；Wherein, wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer.
The mass ratio of filament is 1：0.1-0.5, more preferably wrinkled papaya are 1 with the mycelial mass ratio of sulfur：0.2.
Described wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. and Herba Apii graveolentis ethanol extract are wrinkled papaya, sulfur mycelium, Herba Apii graveolentis respectively in 60-
20-100 mesh is crushed and is crossed after drying at 80 DEG C, and 50%-95% alcohol reflux three times adds 6-15 times every time and measures (w/w)
Ethanol, merging filtrate, concentrating under reduced pressure obtain final product wrinkled papaya with Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. and Herba Apii graveolentis ethanol extract after 30-60 DEG C of vacuum drying；Its
In, wrinkled papaya, sulfur mycelium, the mass ratio of Herba Apii graveolentis are 1：0.1-0.3：0.4-0.8.More preferably wrinkled papaya,
Sulfur mycelium, the mass ratio of Herba Apii graveolentis are 1：0.2：0.5.
The mycelial preparation method of described sulfur is as follows：
The preparation of fluid medium：By 15：15：2：100 ratio weighs glucose, analysis for soybean powder, KH respectively2PO4, elephant skin
Wrinkled papaya proper amount of boiling water is first boiled 30min, filtered through gauze by Fructus Chaenomeliss, and wrinkled papaya residue adds water filtration once, takes filtrate
Plus glucose, analysis for soybean powder, KH2PO4And the constant volume that adds water is to 1000mL, after fully dissolving, it is sub-packed in 500mL triangular flasks, loading amount is
Take out after sterilizing 20 minutes under 121 DEG C of steams of 200mL, 1.15Mpa, store after cooling standby；
Thalline is activated：Sulfur bacteria strain standby on -4 DEG C of test tube slants will be stored in and accessed by sterile working and prepared in advance
In good PDA plate, cultivate 3-7 days in 28 DEG C of thermostatic chambers；
Amplification culture：Cultured sulfur bacteria strain is accessed in ready-made fluid medium by sterile working, connect
300 bottles are planted, quiescent culture 60 days under the conditions of 25 DEG C, culture are filtered with double gauze after terminating, and obtain Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. mycelium.
Described wrinkled papaya extract and compositionss are being prepared suppression in vitro to xanthine oxidase activity by the present invention
Medicine on application.
The present invention answering on the medicine for preparing treatment hyperuricemia by described wrinkled papaya extract and compositionss
With.
The present invention is by described wrinkled papaya extract and compositionss on the medicine for preparing the acute gouty arthritis for the treatment of
Application.
Description of the drawings
Fig. 1 is eburicoic acid high-efficient liquid phase chromatogram.
Specific embodiment
Animal and cell strain Kunming male SPF level mices, weight 20g-25g are carried by SanXia University's Experimental Animal Center
For.
Animal productiong licensing number is：SYXK (Hubei Province) 2011-0012.In constant temperature (24 ± 2 DEG C), periodicity of illumination 12h:12h
Test after raising in environment 1 week, Laboratory Animal Facility uses credit number：SYXK (Hubei Province) 2011-0061, animal productiong licensing
Number it is：SCXK (Hubei Province) 2011-0012.
Sprague-Dawley male SPF level rats, weight 200g ± 20g are carried by SanXia University's Experimental Animal Center
For.Animal productiong licensing number is：SYXK (Hubei Province) 2011-0012.In constant temperature (24 ± 2 DEG C), periodicity of illumination 12h:In 12h environment
Test after raising 1 week, Laboratory Animal Facility uses credit number：SYXK (Hubei Province) 2011-0061.
Human umbilical vein endothelial cells (HUVEC) are purchased from China typical culture collection center cell bank, are passed by this laboratory
For preservation.
Reagent
Xanthine (Sigma), Oteracil Potassium (Aladdin), yeast extract (Angel Yeast), allopurinol (The Place) are yellow
Purine oxidase, uric acid (UA) test kit, xanthine oxidase (XOD) test kit (Nanjing is built up), other reagents are pure for analysis,
Uric acid (Shanghai source leaf), indomethacin (Sigma), (Gibco departments of U.S. product, using another plus 1.8g bicarbonates in liquid for DMEM powder
Sodium), 3- (4,5- dimethylthiazole -2) -2,5- diphenyltetrazolium bromide bromides (MTT), dimethyl sulfoxide (DMSO) are (Sigma), blue or green
Mycin, streptomycin (Huabei Pharmaceutic Co., Ltd), (Hangzhou Ilex purpurea Hassk.[I.chinensis Sims biological engineering material is limited for the new fetal calf serum of import
Company), rat interleukin-11 (IL-1 β) and tumor necrosis factor-alpha (TNF-α) ELISA detection kit (are biotechnology up to section
Company limited), other are pure for analysis.
Instrument
Superclean bench (Suzhou Decontamination Equipment Plant), vertical pressure steam sterilization pan (Japanese HIRAYAMA), JA2003 types
Electronic analytical balance (Shanghai balance equipment factory), 200 microplate reader of infinite (Tecan companies of Switzerland), 5415D mini desktops
High speed centrifuge (German Eppendof companies), pH meter (Shanghai instrument electricity science), ultra-pure water instrument (Ai Kepu companies), -80 DEG C surpass
Cryogenic refrigerator (Haier Group), vacuum freeze drier (LABCONCO companies of the U.S.), YLS-7B foot pawl toes pressure capacity testers
(the glad Soft Inform ation Science and Technology Ltd. in Shanghai), 96 orifice plates (Corning companies of the U.S.), KX41 type inverted microscopes (Olympus),
3111CO2Incubator (U.S. Thermo).
It is prepared by medicine process and crystallite Monosodium urate (MSU)
Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer.：The preparation of fluid medium presses 15:15:2:100 ratio weigh respectively glucose, analysis for soybean powder,
KH2PO4, wrinkled papaya, wrinkled papaya proper amount of boiling water is boiled into 30min first, filtered through gauze, wrinkled papaya residue add water filtration
Once, take filtrate and add glucose, analysis for soybean powder, KH2PO4And the constant volume that adds water is to 1000mL.After fully dissolving, 500mL triangles are sub-packed in
In bottle, loading amount is 200mL, (121 DEG C) the sterilizings rear taking-up in 20 minutes or so of 1.15Mpa steams, is stored standby after cooling；Thalline is lived
Change will be stored in sulfur bacteria strain standby on -4 DEG C of test tube slants and access in preprepared PDA plate by sterile working,
(28 DEG C) are cultivated 3 days in thermostatic chamber；Cultured sulfur bacteria strain is accessed ready-made by amplification culture by sterile working
In fluid medium, 300 bottles are inoculated with.Quiescent culture 60 days under the conditions of 25 DEG C, culture are filtered with double gauze after terminating, and obtain
Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. mycelium.
Eburicoic acid：95% alcohol reflux of Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. mycelium, concentrating under reduced pressure remove solvent, as crude extract, Jing second
Alcohol-water recrystallization and normal-phase silica gel column chromatography petroleum ether：Ethyl acetate (3:1) isolate and purify, obtain eburicoic acid, Jing HPLC analysis teeth
Hole acid purity >=95%, such as Fig. 1.
Described wrinkled papaya extract is to crush and cross 20-100 mesh after wrinkled papaya is dried at 60-80 DEG C, 95%
Alcohol reflux, concentrating under reduced pressure obtain final product wrinkled papaya ethanol extract after 30-60 DEG C of vacuum drying.
Described wrinkled papaya is wrinkled papaya and the drying at 60-80 DEG C respectively of sulfur mycelium with Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract
After crush and cross 20-100 mesh, 95% alcohol reflux three times, every time plus the 6-15 times of ethanol for measuring (w/w), merging filtrate subtracts
Pressure concentration, obtains final product wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract after 30-60 DEG C of vacuum drying；Wherein, wrinkled papaya and sulfur mycelium
Mass ratio be 1：0.2.
Described wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. and Herba Apii graveolentis ethanol extract are wrinkled papaya, sulfur mycelium, Herba Apii graveolentis respectively in 60-
20-100 mesh is crushed and is crossed after drying at 80 DEG C, and 95% alcohol reflux three times adds the 6-15 times of ethanol for measuring (w/w) every time,
Merging filtrate, concentrating under reduced pressure obtain final product wrinkled papaya with Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. and Herba Apii graveolentis ethanol extract after 30-60 DEG C of vacuum drying；Wherein, elephant skin
Fructus Chaenomeliss, sulfur mycelium, the mass ratio of Herba Apii graveolentis are 1：0.2：0.5.
It is prepared by crystallite Monosodium urate (MSU)：Under aseptic condition, 1g uric acid is dissolved in the distilled water that 97mL boils, and adds 3mL
1M NaOH hydrotropies, it is 8.9,4 DEG C of Refrigerator store 24h to adjust pH value in 60 DEG C, is filtered, distilled water wash, drying, obtains crystallite urine
Sour sodium.
Vitro Drug is to xanthine oxidase activity inhibitory action
The preparation buffer of solvent:KH is weighed accurately2PO4、K2HPO4·3H2O, EDTA-2Na, ultrapure water dissolution are prepared
The phosphate buffer of 0.2mol/L (pH=7.50).Substrate solution is prepared:Xanthine is weighed accurately, ultrapure water dissolution is obtained
1.5mmol/L storing solutions, are diluted to the xanthine solution of variable concentrations before use.Enzyme liquid is prepared:Buffer is into 0.16U's
XOD storing solutions, take the work enzyme liquid that appropriate enzyme storing solution buffer is diluted to variable concentrations, before use by work in experimentation
Ice-water bath preservation is placed in as enzyme liquid.Prepare for reagent thing:Experiment in vitro, precision weigh appropriate amount of sample, dimethyl sulfoxide：Without water beetle
Alcohol (1:9) variable concentrations are made.
Impact xanthine oxidase of the medicine to xanthine oxidase vigor can be catalyzed xanthine generation under optimum conditions
Uric acid, uric acid have characteristic absorption peak at 295nm.Enzyme reaction initial velocity is certain, records the increase of absorbance at 295nm per minute
Xanthine oxidase enzyme activity (dA/min) can be calculated.By 100 μ L of sample and blank solution (phosphate buffer), concentration it is
50 μ l of 0.04U/mL xanthine oxidases liquid, sequentially add 96 orifice plates, and 37 DEG C of incubation 3min add concentration to be that 0.48mmol/L is yellow
Adenine solution starts reaction, records absorbance A, altogether 5min under 295nm wavelength every 15s readings once.Calculate and suppress
Rate：Suppression ratio (%)=[(dA/dt) blank-(dA/dt) sample]/(dA/dt) blank × 100.Wherein (dA/dt) is blank for sky
The reaction rate of white group, (dA/dt) sample is sample sets reaction rate.This test is prepared using allopurinol as positive control
The sample of variable concentrations is tested, and calculates suppression ratio of the medicine to xanthine oxidase activity.
Impact of the medicine to hyperuricemia
Hyperuricemia Animal Model
66 male mouse of kunming of packet are randomly divided into 11 groups：Blank group, hyperuricemia model group, allopurinol group
(5mg/kg), eburicoic acid low dose group (80mg/kg), eburicoic acid high dose group (160mg/kg), wrinkled papaya ethanol extract are low
(80mg/kg), wrinkled papaya ethanol extract height (160mg/kg), wrinkled papaya ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract low (80mg/kg), elephant skin wood
Melon ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract height (160mg/kg), wrinkled papaya ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ＆ Herba Apii graveolentis ethanol extract low (80mg/kg), wrinkled papaya ＆ sulfur
Sulphur bacterium ＆ Herba Apii graveolentis ethanol extract height (160mg/kg), wherein both wrinkled papaya ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract groups ethanol extract are 1:0.2, elephant skin
Fructus Chaenomeliss ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ＆ Herba Apii graveolentis ethanol extract group three's ethanol extract ratio is 1:0.2:0.5, precision weighs appropriate amount of sample, 0.5% carboxylic
Methylcellulose sodium solution (CMC-Na) is configured to the suspension of desired concn, gastric infusion.
Modeling and administering mode gastric infusion, administered volume is：10mL/kg, 6 per group.Once a day, it is continuous 7 days, empty
White group and model group rats give the 0.5%CMC-Na of respective volume, and the 8th day starts modeling, and modeling after being administered 1 hour is removed
Blank group is given outside normal saline, and the equal gavage of other each groups gives 21g/kg yeast, 250mg/kg Oteracil Potassium modelings, and continuous 5
My god.Experiment mice is put to death front fasting in 12 hours and can't help water, and after modeling in the 12nd day 1 hour, eyeball takes blood, and it is little that blood is stored at room temperature 1
When, 3000r/min is centrifuged 15 minutes, separates serum.Take each group mouse liver tissue, -80 DEG C of Refrigerator stores.
Liver, renal index are calculated, and after eyeball takes blood, solution takes liver, kidney and claims
Weight, gauge index：Liver/renal index (%)=(viscera weights/body weight) × 100%
Impact of the medicine to hyperuricemia animal model blood uric acid (UA), blood XOD, liver XOD builds up life according to Nanjing
The kit specification operation of thing Graduate School of Engineering, detection each group uric acid in serum, xanthine oxidase activity.During experiment, respectively
Group mice takes 50mg liver glass homogenizers by tissue：Normal saline=1:9 ratio, grinds in ice bath, obtains final product liver
Tissue homogenate, 20000r/m centrifugation 15min, takes supernatant, determines liver xanthine oxidase activity.
Impact of the medicine to acute gout cell model
Cell culture common agents and medicine prepare DMEM culture fluid：Take one bag of DMEM powder culture mediums, plus 950mL tri-
Water is steamed, the penicillin and streptomycin of 5mL 100U/mL is added, is mixed, constant volume is to 1000mL, 0.22 μm of filter membrane vacuum of Jing after dissolving
Filtration sterilization, subpackage, 4 DEG C of preservations, used time add 10% import hyclone.PBS buffer solution：Weigh Sodium Chloride 8g, phosphoric acid
Potassium dihydrogen 0.2g, disodium hydrogen phosphate 1.42g and potassium chloride 0.2g, add 900mL tri-distilled waters, and after fully dissolving, constant volume is arrived
1000mL, 121 DEG C of sterilizing 30min, 4 DEG C of preservations.MTT solution：It is accurate to claim MTT powder 0.25g, it is in adding 50mLPBS, fully molten
Solution, 0.22 μm of filter membrane vacuum filter of Jing are degerming, subpackage, and 4 DEG C keep in dark place.Eburicoic acid and indomethacin storing solution：Accurately weigh
Eburicoic acid 10.0mg, adds 40 μ LDMSO, adds 40 μ L dehydrated alcohol and 20 μ L ethyl acetate, mixes concussion dissolving；Accurately
Claim indomethacin 10.0mg, add 40 μ LDMSO, add 60 μ L sterilized water, mix concussion dissolving.In superclean bench, use
Storing solution filtration sterilization, room temperature preservation are diluted to aimed concn with the culture medium containing 10% serum during use by 0.22 μm of filter membrane.
The final concentration of DMSO and dehydrated alcohol in cell culture fluid is made to remain less than 0.1% and 1% respectively.
Acute gout cell model sets up Human umbilical vein endothelial cells (HUVEC), and adherent growth is based on DMEM cultures
37 DEG C, 5%CO2, aseptic culture in cell culture incubator under the conditions of saturated humidity.Culture fluid is blue or green containing 10%NBS, 100U/mL
The complete DMEM cell culture mediums of mycin and 100U/mL streptomycins, cell monolayer are passed on when growing to culture bottle 80%.Acute gout
Cell model sets up cultured cells by 1 × 105The density of individual/mL is planted in 96 well culture plates, adds final concentration of 100 μ g/
ML MSU, 37 DEG C, 5%CO224h is cultivated in incubator, sets up acute gout cell model.
Medicine affects tetramethyl azo azoles salt micro enzyme reaction colorimetry (MTT) acute gout cell model：Set up anxious
Property gout cell model, different acute drug samples is added in 96 well culture plates, and each Concentraton gradient sets 6 multiple holes,
And blank well (celliferous culture fluid+MTT+DMSO), high dose medicament hole (+100 μ g/mL medicines of celliferous culture fluid are set
Thing+MTT+DMSO), middle dosage medicine hole (+50 μ g/mL medicine+MTT+DMSO of celliferous culture fluid), low-dose drugs hole
(+25 μ g/mL medicine+MTT+DMSO of celliferous culture fluid) and zeroing hole (culture fluid+MTT+DMSO).After continuing culture 48h,
MTT reagents (5mg/mL) develop the color, and survey its OD value under microplate reader 490nm wavelength.
Impact of the medicine to acute gouty arthritises rat model
The structure and administering mode of acute gouty arthritises model
66 male mouse of kunming are randomly divided into 11 groups：Blank group, hyperuricemia model group, indomethacin group (3mg/
Kg), eburicoic acid low dose group (40mg/kg), eburicoic acid high dose group (80mg/kg), the low (40mg/ of wrinkled papaya ethanol extract
Kg), wrinkled papaya ethanol extract height (80mg/kg), wrinkled papaya ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract low (40mg/kg), wrinkled papaya ＆ sulfur
Mykol extract height (80mg/kg), wrinkled papaya ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ＆ Herba Apii graveolentis ethanol extract low (40mg/kg), wrinkled papaya ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ＆ celerys
Dish ethanol extract height (80mg/kg), wherein both wrinkled papaya ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract groups ethanol extract are 1:0.2, wrinkled papaya ＆ sulfur
Sulphur bacterium ＆ Herba Apii graveolentis ethanol extract group three's ethanol extract ratio is 1:0.2:0.5, precision weighs appropriate amount of sample, 0.5% carboxymethyl cellulose
Plain sodium solution (CMC-Na) is configured to the suspension of desired concn, gastric infusion.Successive administration 7 days, after administration in the 5th day, 2 is little
When, MSU (normal saline suspension) 10mg/ is only injected into rat right hind leg articular cavity, and only, entry point is right hind to 0.05mL/
Sufficient internal malleolus trailing edge, pin are right hind Ankle lateral trailing edge to articulatio tibiotarsalis chamber, entry point, and pin mouth inclined-plane is towards front upper place and tibia
Angle at 45 ° is worn to ankle joint chamber, is heaved as injection standard with joint capsule offside, while normal group gives the physiology of equal volume
Saline.
Medicine affects 4h after modeling to the foot swelling of acute gouty arthritises model, measures right hind with sufficient volumetric method little
The same site volume of lower limb ankle joint (taking average), calculates swelling rate.
Swelling rate=(foot swelling before foot swelling-cause is scorching after cause inflammation)/cause foot swelling × 100% before inflammation
Medicine to acute gouty arthritises rat model blood uric acid (UA), blood XOD, liver XOD impact according to Nanjing
Build up the kit specification operation of Bioengineering Research Institute, detection each group uric acid in serum, xanthine oxidase activity.Experiment
When, each group mice takes 50mg liver glass homogenizers by tissue：Normal saline=1:9 ratio, grinds in ice bath, obtains final product
Hepatic homogenate, 20000r/m centrifugation 15min, takes supernatant, determines liver xanthine oxidase activity.
Vitro Drug is to xanthine oxidase inhibitory action
Xanthine Oxidase by Allopurinol has inhibitory action, its IC50For 12.44 μ g/mL, tooth close with described in document
Hole acid, Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract, wrinkled papaya ethanol extract, wrinkled papaya ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract, wrinkled papaya ＆ Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ＆ Herba Apii graveolentis alcohol
Extract has inhibitory action to xanthine oxidase activity, its IC50Respectively 201.31 μ g/mL, 319.45 μ g/mL, 181.29 μ
G/mL, 82.59 μ g/mL, is shown in Table 1.
The impact of 1 medicine xanthine oxidase activity of table
Impact of the medicine to hyperuricemia animal model
Medicine is right to hyperuricemia animal model liver, the impact hyperuricemia model group of renal index and blank
According to a group ratio, liver index, renal index are raised, and have significant difference (P<0.01), allopurinol and each sample group drop
Low hyperuricemia animal model liver index, compared with model group, difference has significance (P<0.01 or P<0.05) 2, are shown in Table.
2 medicine of table is to hyperuricemia animal model liver, the impact of renal index
Note：* the P compared with model group<0.05；* P compared with model group<0.01
Medicine to hyperuricemia animal model blood uric acid (UA), blood XOD, liver XOD impact hyperuricemia model
Group and blank control group ratio, blood uric acid, blood XOD, liver XOD are raised, and have significant difference (P<0.01), allopurinol and various kinds
Product group reduces blood uric acid, blood XOD, liver XOD levels, and compared with model group, difference has significance (P<0.01 or P<0.05),
It is shown in Table 3.
3 medicine of table to hyperuricemia animal model serum UA, serum XOD, liver XOD impact
Note：* the P compared with model group<0.05；* P compared with model group<0.01
Impact of the medicine to acute gout cell model
With residue HUVEC cell viabilities after thiazole blue laws (MTT) determination experiment, as shown in the figure.MTT detection MSU stimulate 48h
Each group cell OD values, calculate cell viability afterwards：Cell viability=each group cell OD values/blank group × 100%.
4 medicine of table is to the impact to acute gout cell model
Impact of the medicine to acute gouty arthritises animal model
4h after impact MSU modelings of the medicine to acute gouty arthritises animal model foot swelling, foot swelling reach highest
Peak, detects foot swelling behind the now each group animal right side, calculates paw swelling.
5 medicine of table is affected on the foot swelling of acute gouty arthritises animal model
Note：* the P compared with model group<0.05；* P compared with model group<0.01
Medicine to acute gouty arthritises animal model blood uric acid (UA), blood XOD, liver XOD impact acute gout
Property arthritis model group and blank control group ratio, blood uric acid, blood XOD, liver XOD raise, and has significant difference (P<0.01), not
Purine alcohol and each sample group reduce blood uric acid, blood XOD, liver XOD levels, and compared with model group, difference has significance (P<
0.01 or P<0.05) 6, are shown in Table.
6 medicine of table to hyperuricemia animal model serum UA, serum XOD, liver XOD impact
Note：* the P compared with model group<0.05；* P compared with model group<0.01.
Claims (10)
1. a kind of wrinkled papaya extract and compositionss, it is characterised in that wrinkled papaya extract is wrinkled papaya ethanol extract, its
Compositionss are wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract or wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. and Herba Apii graveolentis ethanol extract.
2. the wrinkled papaya extract described in claim 1 and compositionss, it is characterised in that described wrinkled papaya extract is
Wrinkled papaya is crushed after drying at 60-80 DEG C and mistake 20-100 mesh, 50-95% alcohol reflux, concentrating under reduced pressure, 30-60 DEG C
Wrinkled papaya ethanol extract is obtained final product after vacuum drying.
3. the wrinkled papaya extract described in claim 1 and compositionss, it is characterised in that described wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer.
Ethanol extract is to crush after wrinkled papaya is dried at 60-80 DEG C respectively with sulfur mycelium and cross 20-100 mesh, 50-95% ethanol
Reflux, extract, three times, adds 6-15 times every time and measures（w/w）Ethanol, merging filtrate, concentrating under reduced pressure, after 30-60 DEG C of vacuum drying i.e.
Obtain wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. ethanol extract；Wherein, wrinkled papaya is 1 with the mycelial mass ratio of sulfur：0.1-0.5.
4. the wrinkled papaya extract described in claim 3 and compositionss, it is characterised in that wrinkled papaya is mycelial with sulfur
Mass ratio is 1：0.2.
5. the wrinkled papaya extract described in claim 1 and compositionss, it is characterised in that described wrinkled papaya and Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer.
It is to crush and cross 20-100 mesh after wrinkled papaya, sulfur mycelium, Herba Apii graveolentis are dried at 60-80 DEG C respectively with Herba Apii graveolentis ethanol extract,
50-95% alcohol reflux three times, adds 6-15 times every time and measures（w/w）Ethanol, merging filtrate, concentrating under reduced pressure, 30-60 DEG C is true
Sky obtains final product wrinkled papaya with Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. and Herba Apii graveolentis ethanol extract after being dried；Wherein, wrinkled papaya, sulfur mycelium, the quality of Herba Apii graveolentis
Than for 1：0.1-0.3：0.4-0.8.
6. the wrinkled papaya extract described in claim 5 and compositionss, it is characterised in that wrinkled papaya, sulfur mycelium, celery
The mass ratio of dish is 1：0.2：0.5.
7. the wrinkled papaya extract described in claim 3-6 any one and compositionss, it is characterised in that described Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer.
The preparation method of filament is as follows：
The preparation of fluid medium：By 15：15：2：100 ratio weighs glucose, analysis for soybean powder, KH respectively2PO4, wrinkled papaya,
Wrinkled papaya proper amount of boiling water is boiled into 30 min, filtered through gauze first, wrinkled papaya residue adds water filtration once, takes filtrate and add Portugal
Grape sugar, analysis for soybean powder, KH2PO4And the constant volume that adds water is to 1000 mL, after fully dissolving, it is sub-packed in 500 mL triangular flasks, loading amount is
200 mL, take out after sterilizing 20 minutes, store standby after cooling under 1.15 121 DEG C of Mpa steams；
Thalline is activated：Sulfur bacteria strain standby on -4 DEG C of test tube slants will be stored in preprepared is accessed by sterile working
In PDA plate, cultivate 3-7 days in 28 DEG C of thermostatic chambers；
Amplification culture：Cultured sulfur bacteria strain is accessed in ready-made fluid medium by sterile working, inoculation 300
Bottle, quiescent culture 60 days under the conditions of 25 DEG C, culture are filtered with double gauze after terminating, and obtain Laetiporus sulphureus (Bull. Ex Fr.) Bond. Et Singer. mycelium.
8. wrinkled papaya extract and compositionss described in claim 1-7 any one is preparing suppression in vitro to xanthine oxidase
Change the application on the medicine of enzymatic activity.
9. wrinkled papaya extract and compositionss described in claim 1-7 any one treats the medicine of hyperuricemia in preparation
Application on thing.
10. wrinkled papaya extract and compositionss described in claim 1-7 any one is preparing the acute ventilation joint for the treatment of
Application on scorching medicine.
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