CN101411743A - Use of Chinese medicine jade screen powder as medicament for preventing and treating liver damage - Google Patents
Use of Chinese medicine jade screen powder as medicament for preventing and treating liver damage Download PDFInfo
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- CN101411743A CN101411743A CN 200810182203 CN200810182203A CN101411743A CN 101411743 A CN101411743 A CN 101411743A CN 200810182203 CN200810182203 CN 200810182203 CN 200810182203 A CN200810182203 A CN 200810182203A CN 101411743 A CN101411743 A CN 101411743A
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Abstract
The invention discloses application of a traditional Chinese medicine, namely Jade Screen Powder as a medicine for preventing and treating hepatic injury, which is characterized in that the Jade Screen Powder which is prepared from 1 to 4 portions of radix astragali, 1 to 4 portions of Atractylodes macrocephala and 1 to 3 portions of radix sileris in portion by weight is soaked in water at a temperature of between 50 and 100 DEG C; extract is concentrated; lower layer of extract is extracted after removal of lipid impurities, mixed with chloroform and n-butyl alcohol according to the volume ratio of 5 to 1, and subjected to oscillation, standing and delamination for removing an intermediate layer; upper layer of extract is mixed with lower layer of solvent, and subjected to oscillation, standing and delamination for removing an intermediate layer; the upper layer of the extract obtained after repeated operation is prepared into having ethanol concentration of 30 percent, and stands for removing deposits; supernatant obtained is prepared into having ethanol concentration of 50 percent; and deposits subjected to standing and separation are polysaccharide effective ingredients of the Jade Screen Powder which can be used as the medicine for preventing and treating hepatic injury, wherein the clinic dosage is 0.9 to 1.8 grams per day as expressed by the amount of drymeal, and the treatment period is at least one week. The application lays a foundation for further research of Jade Screen Powder and the polysaccharide effective ingredients of the Jade Screen Powder.
Description
Technical field
The invention belongs to the purposes technical field of Chinese medicine YUPINGFENG SAN, the polysaccharide active ingredient that particularly extracts from YUPINGFENG SAN is as the application process of control liver injury medicament.
Background technology
According to " pharmacology of Chinese medical formulae " (People's Health Publisher, version in 2002,817 pages) middle record, YUPINGFENG SAN comes from " studying carefully former side ", and record is the classics recipe of treatment exterior deficiency spontaneous perspiration from " hospital birdss of the same feather flock together " volume 150, the function of tool QI invigorating, consolidating superficial resistance, hidroschesis, the sweet temperature of the Radix Astragali in the side, the interior gas of spleen reinforcing lung greatly, but outer strengthening superficial resistance to stop perspiration; But Rhizoma Atractylodis Macrocephalae invigorating the spleen and benefiting QI helps the Radix Astragali to strengthen the power of benefiting QI for strengthening the superficies, is ministerial drug; Assistant is walked table and the imperial heresy of the wind that looses with Radix Saposhnikoviae.The Radix Astragali gets Radix Saposhnikoviae, consolidating superficial resistance and do not stay heresy then, and Radix Saposhnikoviae gets the Radix Astragali, then dispels the wind and does not just hinder.The record of YUPINGFENG SAN is also arranged in " danxi's experiential therapy ", the Golden Mirror of Medicine in addition.Report this side in the works such as " Standards of Diagnosis and Treatment ", " Chinese medicine voluminous dictionary " and come from Effective Formulas Tested by Physicians for Generations, but there is no this side in the Effective Formulas Tested by Physicians for Generations handed down from ancient times.The Pharmacopoeia of the People's Republic of China has all been included this ancient prescription prescribed preparation---Yupingfeng Koufuye in each version in the period of nineteen ninety to 2005, writes out a prescription by Radix Astragali 600g, and Radix Saposhnikoviae 200g, Rhizoma Atractylodis Macrocephalae (parched) 200g forms.
The YUPINGFENG SAN of being put down in writing in the document is all formed with the Radix Astragali, the Rhizoma Atractylodis Macrocephalae, Radix Saposhnikoviae three flavors at all times, but ratio is different.According to " YUPINGFENG SAN is formed examination and association " (the 4th phase of " Chinese patent medicine " nineteen ninety-five), surplus the Radix Astragali on the books, the Rhizoma Atractylodis Macrocephalae, Radix Saposhnikoviae mass fraction proportioning mode reach ten more than the kind, the mode of seeing at most is 1:1:1,1:2:1,3:1:1 and 2:2:1, the ratio difference, and the effect emphasis is also had any different.
According to " the YUPINGFENG SAN side of closing utilization simple analysis " (" time precious traditional Chinese medical science traditional Chinese medicines " 2006 the 17th volume the 5th phase 839-840 page or leaf), YUPINGFENG SAN now is usually used in the multiple treatment of diseases relevant with immune dysfunction clinically, as treat spontaneous perspiration, flu, cough with asthma, show effect repeatedly upper respiratory tract infection, chronic bronchitis, allergic rhinitis, glomerulonephritis of children's is brought out diseases such as sending out author, chronic urticaria and recurrent aphtha repeatedly because of diseases caused by exogenous pathogenic factor, and effect is remarkable.Summarized the effect of YUPINGFENG SAN in the control SARS in " YUPINGFENG SAN control ' SARS ' mechanism is inquired into " (" TCM Document magazine " 2003 the 3rd phase 35-37 pages or leaves) literary composition.Do not see at present the report of relevant YUPINGFENG SAN purposes aspect the control hepatic injury.
According to " the pharmaceutical research progress of YUPINGFENG SAN " (" Anhui medicine " 2003 the 7th volumes the 4th phase 241-242 page or leaf), YUPINGFENG SAN pharmacology aspect mainly contains immune dual regulation; Comprehensive function to anaphylactogen; Alleviate experimental nephritis pathological changes, promote the nephritis pathological repair; Inhibitory action to influenza virus; Resisting fatigue, low temperature resistant, anoxia enduring etc.Have not yet to see the report that relevant YUPINGFENG SAN has the active pharmacological research of control hepatic injury aspect.
At present few at the research of YUPINGFENG SAN active component, successively reported " YUPINGFENG SAN polysaccharide composition is to Immune Effects " (2006 the 22nd volume the 1st phase 2-4 pages or leaves) on " Pharmacology and Clinics of Chinese Materia Medica " magazine, " YUPINGFENG SAN total polysaccharides different modes of administration with time influence mutually the comparative study of immune function of mice " (2006 the 22nd volume the 2nd phase 5-7 pages or leaves), " the YUPINGFENG SAN total polysaccharides influences intestinal mucosa damage mice intestinal-respiratory tract IgA and secretes research " (2007 the 23rd volumes the 5th phase 43-45 page or leaf), above-mentioned studies show that, YUPINGFENG SAN total extract and polysaccharide composition thereof have immunological enhancement, and the YUPINGFENG SAN total polysaccharides may be by activating the effect of intestinal mucosa immunity performance local immunity." extraction and the pharmacological evaluation of jade screen wind crude polysaccharides " literary composition in " Strait Pharmaceutical Journal " (2004 the 16th volume the 2nd phase 38-40 page or leaf) has been introduced and has been adopted conventional method to extract crude polysaccharides, observes jade screen wind crude polysaccharides tumor growth is had certain inhibitory action.What more than report was used all is the YUPINGFENG SAN total polysaccharides, not further separation and purification; " YUPINGFENG SAN part polysaccharide is to the research of caused by cyclophosphamide immunocompromised mouse immune regulating action " (" Immunomodulating Effects of FractionedPolysaccharides Isolated from Yu-Ping-Feng-Powder in Cyclophosphamide-TreatedMice ") literary composition among " America Chinese medicine magazine " (" The American Journal of ChineseMedicine " 2006 the 34th volume the 4th phase 631641 pages) and Medical University Of Anhui 2006 doctorate paper " research of herbal mixture jade screen wind immuno-modulating effect of polysaccharides and mechanism " have reported the YUPINGFENG SAN polysaccharide position of the being extracted nonspecific immunity to immunosuppressed mice, the influence of specific immune function and mechanism, and to the influence and the mechanism of macrophage coherent signal transduction path in the rat adjuvant type arthritis model, but its research is influence and the mechanism of medicine to immunosuppressed mice immunologic function and rat adjuvant type arthritis model, does not see that it carries out the research for the hepatic injury preventive and therapeutic effect of YUPINGFENG SAN and active ingredient thereof.
Hepatic injury be all multifactor due to the pathological changes result of various hepatic disease; modern medicine has more in depth been inquired into its mechanism from many aspects; and hepatics such as research and development membrane protective agent, anti peroxidation of lipid agent, anti-immune response agent; but the still difficult people's will to the greatest extent of curative effect, and limited its clinical practice because of toxic and side effects.Chinese medicine has been gone through practice and development in several thousand, dialectical prescription under instruction of Chinese Medicine theory, strict processing is concocted, can bring into play the comprehensive function of many target spots, multipath, too many levels, so Chinese medicine is a kind of both effective and practicable treatment thinking aspect the treatment hepatic injury unique advantage being arranged.
Comprehensively " liver damage animal model and treatment by Chinese herbs research overview " (" contemporary Chinese Chinese medicine " the 8th volume o. 11th 25-28 page or leaf), " research of Chinese herbal medicine anti-hepatic fibrosis " (" Chinese crude drug " calendar year 2001 the 24th volume the 3rd phase 212-214 page or leaf), introduction in " Japanese Modern Chinese prescription medicine is to the prophylactic treatment progress of hepatopathy " documents such as (" journal of shanghai Chinese medicine " 2004 the 38th volumes the 1st phase 58-60 pages or leaves), the at present ongoing Chinese herbal medicine that is used to prevent and treat hepatic injury research mainly contains: (1) single medicinal material: Radix Angelicae Sinensis, Radix Salviae Miltiorrhizae, Rhizoma Curcumae, Semen Trigonellae, Radix Sophorae Flavescentis, Radix Gentianae, Herba Silybi mariani, Stigma Croci, Fructus Lycii, Rhizoma Picrorhizae, Folium Ginkgo extract, tetrandrine, Radix Glycyrrhizae, Rhizoma Curcumae Longae, Semen Persicae, Radix Notoginseng, the Radix Astragali, Herba Lycopi, Cordyceps etc.(2) herbal mixture: Herba Sidae Rhombifoliae soup, Sini San, bupleurum powder for relieving liver-qi, compound recipe 861, Han Danle, soft liver sheet, liver heart health, anti-fine compound recipe, hepatitis are flat, liver-strengthening capsule, Herba Sidae Rhombifoliae soup, BUZHONG YIQI TANG, RENSHEN YANGRONG TANG, decoction of ten powerful tonics, Chai Lingtang, CHAIHU GUIZHI TANG, GUIPI TANG etc.
So far do not see relevant YUPINGFENG SAN and active ingredient thereof report as control liver injury medicament and application process aspect thereof.
Summary of the invention
The object of the present invention is to provide the new purposes of a kind of Chinese medicine YUPINGFENG SAN as the control liver injury medicament.
Chinese medicine YUPINGFENG SAN of the present invention is as the using method of control liver injury medicament, it is characterized in that: press mass fraction with Radix Astragali 1-4: Rhizoma Atractylodis Macrocephalae 1-4: the YUPINGFENG SAN that Radix Saposhnikoviae 1-3 is formulated, water by this YUPINGFENG SAN of every gram and 10-30 milliliter is mixed, soak 0.5-10h, under this temperature, be incubated 1-6h after being heated to 50-100 ℃, carry out solid-liquid separation, isolated extracting solution is concentrated into the 1/5-1/10 of original volume; To hang down the extracting solution after the polar organic solvent extraction concentrates, keep lower floor's extract; 5:1 is mixed with mixed organic solvents with chloroform and n-butyl alcohol by volume, with this mixed organic solvents and this lower floor's extract by volume 1:2-2:1 mix, leave standstill after the vibration to layering, remove the intermediate layer; Upper layer of extraction liquid is mixed with the lower floor organic solvent, leave standstill to layering after the vibration again, remove the intermediate layer; Press the said process repeatable operation 3-15 time, keep the upper layer of extraction liquid that obtains at last; With this upper layer of extraction liquid with ethanol allocate to concentration of alcohol be 30%, discard precipitation after leaving standstill; With the gained supernatant again with ethanol allocate to concentration of alcohol be 50%, isolate precipitate after leaving standstill, will be crushed into powder after this drying precipitate, be YUPINGFENG SAN polysaccharide active ingredient, note is made YPF-P; As the control liver injury medicament, clinical dosage is shown 0.9-1.8g/ day with the dry powder scale, the course of treatment at least one week with this YPF-P dry powder.
" by mass fraction with Radix Astragali 1-4: Rhizoma Atractylodis Macrocephalae 1-4: the YUPINGFENG SAN that Radix Saposhnikoviae 1-3 is formulated " described in the present invention comprises traditional proportioning mode of 1:1:1,1:2:1,1:1:3,3:1:1,2:2:1,1:3:1,1.5:2:1,2:1.5:1,2:1:1 or 4:4:1.
Take among the present invention extracting solution " is incubated 1-6h after being heated to 50-100 ℃ " under this temperature, described " heating " method can comprise the mode of heating of extraction pot steam heated or ultrasound wave heating; If it is too short that the extracting solution heating-up temperature is lower than 50 ℃ or heat time heating time, then extraction efficiency is too low causes waste; And oversize if heating-up temperature is higher than 100 ℃ or heat time heating time, then can pharmaceutically active be caused than macrolesion, so the present invention takes extracting solution in 50-100 ℃ of heating 1-6h.
The residue that obtains after described " solid-liquid separation " also can carry out multiple extraction, merges the extracting solution that each time obtains, to make full use of the effective ingredient in the YUPINGFENG SAN medical material.
Described " with low polar organic solvent extraction extracting solution " can take repeatedly to extract the back combining extraction liquid, to improve the purity of effective ingredient; Described " low polar organic solvent " can select petroleum ether, normal hexane or ethyl acetate for use, is used for extraction and removes extracting solution lipid impurity.
Described is in order to remove protide impurity with mixed organic solvents and " repeatable operation 3-15 time " of carrying out after extract mixes, be lower than 3 times then in the extracting solution protein retain morely, most protein is removed in the time of 15 times, increases having little significance of number of operations.
Described " drying " can select the drying mode that comprises drying under reduced pressure, drying with water bath, lyophilization or spray drying for use.
Described this YPF-P dry powder is meant the human dosage of YPF-P dry powder as the clinical dosage " 0.9-1.8g/ day " of control liver injury medicament, does not comprise the adjuvant weight of adding when making various dosage form.
Face with preceding the YPF-P dry powder of 0.9-1.8g/ daily dose can be able to be divided after with the appropriate solvent dissolving 2 times oral; Described " appropriate solvent " can be normal saline, glucose saline, deionized water or distilled water; Described " divide 2 times oral " is to formulate according to the dynamic metabolism characteristics of medicine, and this scheme is compared with 1 time the scheme of taking medicine every day, and the fluctuation of medicine bulk concentration is littler, and curative effect is better; Also YPF-P can be added adjuvant on the basis of effective dose 0.9-1.8g/ day and make various peroral dosage forms, comprise oral liquid, granule, tablet or capsule; Described " adjuvant " comprises the additive of wetting agent, lubricant, diluent, binding agent and/or coloring agent.
Beneficial effect of the present invention is:
In the purposes of existing YUPINGFENG SAN, do not have the purposes of official seal treating the liver damage aspect as yet; During the few research of active ingredient is reported at present at YUPINGFENG SAN, just only see and from YUPINGFENG SAN, extract total polysaccharides or total glycosides, do not see further separation and purification again; And the present invention carries out pharmacological research with the specific polysaccharide position that the YUPINGFENG SAN total polysaccharides carries out obtaining after the fractionated, and this way of discarding the dross and selecting the essential makes that research before goes a step further again to the drug activity material base in the YUPINGFENG SAN; Adopted the YUPINGFENG SAN polysaccharide position similar in " YUPINGFENG SAN part polysaccharide is to the research of caused by cyclophosphamide immunocompromised mouse immune regulating action " and the academic dissertation " research of herbal mixture jade screen wind immuno-modulating effect of polysaccharides and mechanism " with the present invention, but its research contents is influence and the mechanism of medicine to immunosuppressed mice immunologic function and rat adjuvant type arthritis model, does not carry out the research of YUPINGFENG SAN polysaccharide active ingredient for the hepatic injury preventive and therapeutic effect.The present invention has opened up another new field for the drug efficacy study of YUPINGFENG SAN polysaccharide active ingredient, has filled up this blank.
The present invention has obtained a kind of polysaccharide active ingredient in the YUPINGFENG SAN by extraction separation, and confirmed that by experiment this composition has significant curative effect aspect the control hepatic injury, tentatively inquired into the mechanism of this ingredients for preventing and treating hepatic injury effect, the present invention helps this ancient prescription is carried out further further investigation, for the clinical application range of expanding YUPINGFENG SAN, promote the development and use of YUPINGFENG SAN to lay the foundation, and in the mankind seek the road of control hepatic injury good medicine, found the another material that is rich in prospect.
Description of drawings
Fig. 1 is CCl
4Induce acute chemical hepatic injury experiment normal group mouse liver histopathology figure;
Fig. 2 is CCl
4Induce acute chemical hepatic injury model group mouse liver histopathology figure;
Fig. 3 is YPF-P50mgkg
-1The dosage group is to CCl
4Induce the figure that influences of acute chemical hepatic injury mouse liver histopathology;
Fig. 4 is YPF-P100mgkg
-1The dosage group is to CCl
4Induce the figure that influences of acute chemical hepatic injury mouse liver histopathology;
Fig. 5 is YPF-P200mgkg
-1The dosage group is to CCl
4Induce the figure that influences of acute chemical hepatic injury mouse liver histopathology;
Fig. 6 is that the bifendate matched group is to CCl
4Induce the figure that influences of acute chemical hepatic injury mouse liver histopathology.
Fig. 7 induces acute chemical hepatic injury experiment normal group mouse liver histopathology figure for D-Ga1;
Fig. 8 induces acute chemical hepatic injury model group mouse liver histopathology figure for D-Ga1;
Fig. 9 is YPF-P50mgkg
-1The dosage group is induced the figure that influences of acute chemical hepatic injury mouse liver histopathology to D-Ga1;
Figure 10 is YPF-P100mgkg
-1The dosage group is induced the figure that influences of acute chemical hepatic injury mouse liver histopathology to D-Ga1;
Figure 11 is YPF-P200mgkg
-1The dosage group is induced the figure that influences of acute chemical hepatic injury mouse liver histopathology to D-Ga1;
Figure 12 induces the figure that influences of acute chemical hepatic injury mouse liver histopathology for the bifendate matched group to D-Ga1.
Figure 13 unites lipopolysaccharide-induced acute immunologic liver injury experiment normal group mouse liver histopathology figure for bacillus calmette-guerin vaccine;
Figure 14 unites lipopolysaccharide-induced acute immunologic liver injury model group mouse liver histopathology figure for bacillus calmette-guerin vaccine;
Figure 15 is YPF-P50mgkg
-1The dosage group is to the figure that influences of acute immunologic liver injury mouse liver histopathology variation;
Figure 16 is YPF-P100mgkg
-1The dosage group is to the figure that influences of acute immunologic liver injury mouse liver histopathology variation;
Figure 17 is YPF-P200mgkg
-1The dosage group is to the figure that influences of acute immunologic liver injury mouse liver histopathology variation;
Figure 18 is the influence figure of bifendate matched group to acute immunologic liver injury mouse liver histopathology variation.
Figure 19 unites lipopolysaccharide-induced acute immunologic liver injury experiment normal group murine liver tissue TNF-alpha expression figure for bacillus calmette-guerin vaccine;
Figure 20 unites lipopolysaccharide-induced acute immunologic liver injury model group murine liver tissue TNF-alpha expression figure for bacillus calmette-guerin vaccine;
Figure 21 is YPF-P50mgkg
-1The dosage group is to the figure that influences of acute immunologic liver injury murine liver tissue TNF-alpha expression;
Figure 22 is YPF-P100mgkg
-1The dosage group is to the figure that influences of acute immunologic liver injury murine liver tissue TNF-alpha expression;
Figure 23 is YPF-P200mgkg
-1The dosage group is to the figure that influences of acute immunologic liver injury murine liver tissue TNF-alpha expression;
Figure 24 is the influence figure of bifendate matched group to acute immunologic liver injury murine liver tissue TNF-alpha expression.
Figure 25 unites lipopolysaccharide-induced acute immunologic liver injury experiment normal group murine liver tissue iNOS expression figure for bacillus calmette-guerin vaccine;
Figure 26 unites lipopolysaccharide-induced acute immunologic liver injury model group murine liver tissue iNOS expression figure for bacillus calmette-guerin vaccine;
Figure 27 is YPF-P50mgkg
-1The dosage group is to the figure that influences of acute immunologic liver injury murine liver tissue iNOS expression;
Figure 28 is YPF-P100mgkg
-1The dosage group is to the figure that influences of acute immunologic liver injury murine liver tissue iNOS expression;
Figure 29 is YPF-P200mgkg
-1The dosage group is to the figure that influences of acute immunologic liver injury murine liver tissue iNOS expression;
Figure 30 is the influence figure of bifendate matched group to acute immunologic liver injury murine liver tissue iNOS expression.
Figure 31 gel electrophoresis figure that acute immunologic liver injury murine liver tissue TNF-α mRNA expresses for YPF-P influences;
Figure 32 gel electrophoresis sxemiquantitative cartogram that acute immunologic liver injury murine liver tissue TNF-α mRNA expresses for YPF-P influences.
The specific embodiment
Below in conjunction with embodiment, the invention will be further described.
Embodiment 1:
The Radix Astragali, Rhizoma Atractylodis Macrocephalae (parched), Radix Saposhnikoviae are pressed the formulated YUPINGFENG SAN of mass fraction 3:1:1, crushed after being dried in 50 ℃ of baking ovens, cross 20 mesh sieves, this YUPINGFENG SAN of 5kg is placed extraction pot, pour 50L distilled water immersion 30min into after, under this temperature, be incubated 2h after being heated to 95 ℃, carry out solid-liquid separation, add the 50L distilled water in the residue that after solid-liquid separation, obtains again, under this temperature, be incubated 2h after being heated to 95 ℃, merge extracted twice liquid after the solid-liquid separation and be concentrated into 20L; With 1L petroleum ether ultrasonic extraction extracting solution 2 times, each 30min keeps lower floor's extract; 5:1 is mixed with mixed organic solvents with chloroform and n-butyl alcohol by volume, with this mixed organic solvents and this lower floor's extract by volume 1:1 in separatory funnel, mix, vibrate and leave standstill to layering after 20 seconds, remove the intermediate layer; Upper layer of extraction liquid is mixed with the lower floor mixed organic solvents, left standstill to layering after vibrating again 20 seconds, remove the intermediate layer; Press the said process repeatable operation 10 times, keep the upper layer of extraction liquid that obtains at last; With this upper layer of extraction liquid under alcohol meter monitoring with ethanol allocate to concentration of alcohol be 30%, discard precipitation after leaving standstill 12h; With the gained supernatant again under alcohol meter monitoring with ethanol allocate to concentration of alcohol be 50%, isolate precipitate after leaving standstill 12h, this precipitate is crushed into powder behind 50 ℃ of drying under reduced pressure through Rotary Evaporators, is YUPINGFENG SAN polysaccharide active ingredient, note is made YPF-P; The polyoses content that adopts the phenol sulfuric acid process to record among this YPF-P is 88%.
Pharmacological testing and result with YPF-P illustrates its new purposes in pharmaceutical field below.
YPF-P is as follows to the preventive and therapeutic effect test of hepatic injury:
(1) material
Animal: Kunming mouse, male, body weight 20 ± 2g provides the (quality certification number: real moving accurate No. 02 of Anhui doctor) by Medical University Of Anhui's Experimental Animal Center.
Main medicine and reagent: crude drug Inner Mongolia Astragalis; the Jilin Radix Saposhnikoviae; the Anhui Rhizoma Atractylodis Macrocephalae (parched) is all available from Hefei and justice hall prepared slices of Chinese crude drugs Co., Ltd; YPF-P (self-control; polyoses content is 88%); bifendate drop pill (Beijing XieHe medicine Factory's product; lot number: 06040202); carbon tetrachloride (Shantou Xilong Chemical Factory; lot number: 060707); D-Gal; inducible nitric oxide synthase test kit (Sigma company product; lot number: 085k1321; v-4651); bacillus calmette-guerin vaccine (Shanghai Vaccine and Serum Institute's product; lot number: 04049701); lipopolysaccharide test kit (Sigma company product; lot number: L2880); AST; ALT measures test kit (Shanghai Rongsheng Bioisystech Co., Ltd's product; lot number: 20070304; 20070401); MDA; NO; SOD; GSH measures test kit, and (bio-engineering research institute is built up in Nanjing; lot number: A003-2; A012; A001-1; A006); formaldehyde; dehydrated alcohol (Shanghai chemical reagent company limited; lot number: 200602152; 200610206); glacial acetic acid (Chemical Reagent Co., Ltd., Sinopharm Group; lot number: 20050928); sheep anti mouse TNF-alpha monoclonal antibodies; anti-sheep of the rabbit of horseradish peroxidase-labeled or rabbit goat-anti rabbit two anti-(Santa Cruz Biotechnology products; lot number: sc-3092; sc-2445); Trizol reagent (Invitrogen company product; lot number: 1121068); (PiJi Biology Engineering Co., Ltd., Shenzhen City produces the PCR test kit; lot number: 20050901); agarose (Spain's import packing; lot number: BI00821); 2000bp DNA marker (TaKaRa company product; lot number: B4601A); reverse transcription Superscript II RT (Promega company product; lot number: 242806); ethidium bromide (Sigma company product; lot number: E8751); glycerol (Bangbu chemical reagent factory product; lot number: 031026); chloroform; hydrochloric acid (analytical pure) (build letter chemical industry company limited chemical reagent work and produce by Shanghai; lot number: 041205; 041221); isopropyl alcohol (analytical pure) (Huainan City, Anhui chemical reagent factory product, lot number: 041101).
Key instrument: Ts14-2 type optical readings balance (Hunan instrument balance equipment factory product); BP211D electronic balance (German Saftarius produces); ultramicroscope XDS-1B (Chongqing Photoelectric Instruments Corp.'s product); the medical numerical control supersonic washer of KQ-500DE type (Kunshan Ultrasonic Instruments Co., Ltd.); DL-5M low speed refrigerated centrifuge (Hunan, Changsha instrument centrifuge instrument company limited product); TGL-16H high speed centrifuge (Heima Medical Instrument Co., Ltd., Zhuhai City's product); 722S spectrophotometer (Shanghai exact science instrument company product); GSY-8 electric-heated thermostatic water bath (Beijing Medical Equipment Plant anticipates into company's product); MDF-382E Sanyo cryogenic refrigerator (SANYO GS group product); XW-80A vortex agitator (going up Industrial Co., Ltd. of Nereid section produces); the automatic dual pure water distillator of SZ-2 (Shanghai Hu Xi analytical tool Co., Ltd., Factory); gene-amplificative instrament (Hema8000 Heima Medical Instrument Co., Ltd., Zhuhai City); ultraviolet transilluminator (Heima Medical Instrument Co., Ltd., Zhuhai City), ultraviolet gel imaging system BioSens SC 720 types (Shanghai Bioisystech Co., Ltd).
(2) method
1. carbon tetrachloride (CCl
4) preparation of inducing mouse acute chemical hepatic injury model, grouping and specimen take
Kunming mouse, male, body weight 20 ± 2g is divided into normal group at random, model group, YPF-P50mgkg
-1Dosage group, 100mgkg
-1Dosage group, 200mgkg
-1(Bifendate pill, BDP) matched group, BDP dosage are 200mgkg for dosage group and bifendate
-1, 10 every group.Before the administration YPF-P, BDP are pressed corresponding dosage with the normal saline wiring solution-forming, gastric infusion, normal group and model group give normal saline, every day 1 time, continuous 7 days.Except that normal group mouse peritoneal injection respective amount normal saline, all the other respectively organize mice disposable celiac injection 0.1%CCl after the last administration
4Peanut oil solution 10mlkg
-1After fasting be can't help drinking 16h, extract eyeball of mouse and collect blood, centrifugal 10min isolates serum under rotating speed 3000rpm, detects serum alanine aminotransferase (ALT), aspartate amino transferase (AST), nitric oxide (NO) content; The taking-up liver is weighed; Get the hepatic tissue of the about 1cm of leftlobe of liver * 1cm size, place 10% formalin, wait until histopathologic examination, and get 0.5g in the liver same area and organize, be used to measure superoxide dismutase (SOD), malonaldehyde (MDA), glutathion (GSH) content with the conventional preparation of cold saline 10% liver homogenate.
2.D-the preparation of aminogalactose (D-Ga1) inducing mouse acute chemical hepatic injury model, grouping and specimen are taked
Laboratory animal condition and divide into groups the same.Medication is the same, continuous 7 days.After the last administration, except that normal group mouse peritoneal injection respective amount normal saline, all the other respectively organize mice disposable celiac injection D-Ga1800mgkg
-1After fasting be can't help drinking 16h, specimen taked method the same.
3. the preparation of bacillus calmette-guerin vaccine (BCG) associating lipopolysaccharide (LPS) inducing mouse immunologic liver injury model, grouping and specimen are taked
Laboratory animal condition and divide into groups the same.Administration group normal saline wiring solution-forming, by the body weight gastric infusion, model group and normal group give the normal saline of respective volume, gastric infusion, every day 1 time, continuous 10 days.Administration first day, except that normal group, all the other only respectively organize tail vein injection BCG2.5mg/0.2mg/, behind the 10th day gastric infusion, tail vein injection LPS7.5 μ g/0.2ml/ only, fasting afterwards can't help drinking 16h, extract eyeball of mouse and collect blood, centrifugal 10min isolates serum under rotating speed 3000rpm, detects serum alt, AST content; Taking-up liver, spleen are weighed respectively; Get the about 1cm of leftlobe of liver * 1cm hepatic tissue, place 10% formalin, wait until histopathologic examination; In addition take by weighing the 0.5g hepatic tissue, in ice bath, add the 4.5ml normal saline and be prepared into 10% liver homogenate, under the rotating speed 2500rpm behind the centrifugal 10min, get supernatant and put-75 ℃ of refrigerators and preserve to be checked with refiner in the liver same area.
4. assay method
The liver spleen index: put to death after mice is weighed, take out the liver spleen, blot surface moisture and bloodstain with filter paper, weigh, computing formula is as follows: the weight/body weight of organ index=internal organs * 100%
Serum biochemistry index determining: press the described method of test kit and measure serum alanine aminotransferase (ALT), aspartate amino transferase (AST).
The mensuration of liver biochemical indexes and cytokine: press the described method of test kit and measure superoxide dismutase (SOD), malonaldehyde (MDA), glutathion (GSH), nitric oxide (NO) content, the SABC method is measured tumor necrosis factor-alpha (TNF-α) and inducible nitric oxide synthase (iNOS) content in the homogenate, and the RT-PCR method is measured TNF-α mRNA.
The liver organization pathological examination: liver organization is fixed with 10% formalin, and specimens paraffin embedding slices is carried out conventional H E dyeing, observes and respectively organizes the murine liver tissue pathological change.
(3) result
1.YPF-P to CCl
4Induce the influence of acute chemical hepatic injury mice
1.1 YPF-P is to CCl
4Induce the influence of acute chemical hepatic injury Mouse Liver index, ALT, AST
Table 1 has provided YPF-P to CCl
4Induce the exponential influence of acute chemical hepatic injury mouse liver.As shown in table 1, compare YPF-P200mgkg with model group
-1Dosage group and BDP group can significantly reduce the mouse liver index.
Table 1.YPF-P is to CCl
4Induce the exponential influence of acute chemical hepatic injury mouse liver (x ± s, n=10)
#Compare with normal group P＜0.01;
*Compare with model group P＜0.05
Table 2 has provided YPF-P to CCl
4Induce acute chemical hepatic injury mice serum ALT, AST influence.As shown in table 2, compare with model group, each dosage group of YPF-P all can reduce acute chemical hepatic injury mice serum ALT, AST level, 200mgkg
-1Dosage group difference has significance, shows that it has the effect of stable liver plasma membrane and mitochondrial membrane.
Table 2.YPF-P is to CCl
4Induce acute chemical hepatic injury mice serum ALT, AST the influence (x ± s, n=10)
#Compare with normal group P＜0.01;
*P＜0.05,
*Compare with model group P＜0.01
1.2 YPF-P is to CCl
4Induce the influence of acute chemical hepatic injury mice MDA, SOD, GSH
Table 3 has provided YPF-P to CCl
4Induce acute chemical hepatic injury mouse liver even slurry GSH, SOD, MDA influence.As shown in table 3, compare with model group, YPF-P can significantly reduce the MDA concentration that increases in each treatment group, the activity of SOD and GSH in the raising liver homogenate, wherein, YPF-P100,200mgkg
-1The dosage group has significance.Show that YPF-P may have certain inhibition peroxidating material and form and promote the effect that antioxidant generates, and can be by improving hepatic tissue SOD, GSH vigor, the development of checking hepatic injury with lapse to.
Table 3.YPF-P is to CCl
4Induce acute chemical hepatic injury mouse liver even slurry GSH, SOD, MDA influence (x ± s, n=10)
#Compare with normal group P＜0.01;
*P＜0.05,
*Compare with model group P＜0.01
1.3YPF-P to CCl
4The influence (HE dyeing, * 200) of inducing acute chemical hepatic injury mouse liver histopathology to change
Fig. 1 is CCl
4Induce acute chemical hepatic injury experiment normal group mouse liver histopathology figure, visible lobules of liver clear in structure among the figure, hepatic cords, hepatic sinusoid are that the center is radial arrangement with the central vein; Fig. 2 is CCl
4Induce acute chemical hepatic injury model group mouse liver histopathology figure, visible lobules of liver normal configuration disappears among the figure, and hepatocyte is big lamellar necrosis around the portal area, companion's massive inflammatory cells infiltrated; Fig. 3, Fig. 4, Fig. 5 are respectively YPF-P50mgkg
-1, 100mgkg
-1, 200mgkg
-1The dosage group is to CCl
4Induce the figure that influences of acute chemical hepatic injury mouse liver histopathology variation, show as among the figure with the increase of YPF-P dosage, hepatocellular degeneration, degree of necrosis alleviate gradually, and inflammatory cell infiltration reduces; Fig. 6 is that the BDP matched group is to CCl
4That induces that acute chemical hepatic injury mouse liver histopathology changes influences figure, pathological manifestations and YPF-P200mgkg among the figure
-1The dosage group is similar.
2.YPF-P D-Ga1 is induced the influence of acute chemical hepatic injury mice
2.1 YPF-P induces the influence of acute chemical hepatic injury Mouse Liver index, ALT, AST to D-Ga1
Table 4 has provided YPF-P D-Ga1 has been induced the exponential influence of acute chemical hepatic injury Mouse Liver.As shown in table 4, to compare with model group, each dosage group of YPF-P increases with dosage, and the Mouse Liver index reduces gradually, YPF-P200mgkg
-1Dosage group and BDP matched group tool significant difference.
The table 4.YPF-P to D-Ga1 induce the exponential influence of acute chemical hepatic injury Mouse Liver (x ± s, n=10)
#Compare with normal group P＜0.01;
*Compare with model group P＜0.05
Table 5 has provided YPF-P D-Ga1 has been induced acute chemical hepatic injury mice serum ALT, AST influence.As shown in table 5, compare YPF-P200mgkg with model group
-1Dosage group and BDP control group mice Serum ALT, AST level significantly reduce.
The table 5.YPF-P to D-Ga1 induce acute chemical hepatic injury mice serum ALT, AST the influence (x ± s, n=10)
#Compare with normal group P＜0.01;
*P＜0.05,
*Compare with model group P＜0.01
2.2 YPF-P induces the influence of acute chemical hepatic injury mice MDA, SOD, GSH to D-Ga1
Table 6 has provided YPF-P D-Ga1 has been induced acute chemical hepatic injury mouse liver even slurry MDA, SOD, GSH influence.As shown in table 6, compare with model group, each treatment group of YPF-P is with the increase of dosage, and MDA concentration reduces gradually in the liver homogenate, and the activity of SOD and GSH increases gradually.Wherein, YPF-P100,200mgkg
-1Dosage group and BDP matched group difference have significance.
Table 6.YPF-P to D-Ga1 induce acute chemical hepatic injury mouse liver even slurry MDA, SOD, GSH influence (x ± s, n=10)
#Compare with normal group P＜0.01;
*P＜0.05,
*Compare with model group P＜0.01
2.3 the influence (HE dyeing, * 200) that YPF-P induces acute chemical hepatic injury murine liver tissue histopathology to change to D-Ga1
Fig. 7 induces acute chemical hepatic injury experiment normal group mouse liver histopathology figure for D-Ga1, showing as hepatocyte among the figure is that the center is radial arrangement with the central vein, hepatic cords, hepatic sinusoid queueing discipline, the lobules of liver structural integrity is not seen pathological changes such as hepatocellular degeneration, necrosis; Fig. 8 is a model group, shows as the hepatocyte cloudy swelling, and Cytoplasm is loose, and the kiernan's space boundary is unclear, and hepatocyte has spotty necrosis and focal necrosis, and the portal area inflammatory cell infiltration is obvious; Fig. 9 is YPF-P50mgkg
-1The dosage group, it is remarkable to show as hepatocellular degeneration, necrosis and inflammatory cell infiltration; Figure 10, Figure 11, Figure 12 are respectively YPF-P100mgkg
-1, 200mgkg
-1Dosage group and BDP matched group are induced the figure that influences of acute chemical hepatic injury murine liver tissue histopathology variation to D-Ga1, show as hepatocellular degeneration among the figure, degree of necrosis significantly alleviates, inflammatory cell infiltration reduces, and the liver tissue injury pathological change obviously alleviates.
Table 7.YPF-P induces acute chemical hepatic injury murine liver tissue pathological grading (n=5) to D-Ga1
#Compare with normal group P＜0.01;
*Compare with model group P＜0.05
3.YPF-P BLG is united the influence that LPS induces acute immunologic liver injury mice
3.1 the influence that YPF-P induces acute immunologic liver injury Mouse Liver spleen index, ALT, AST to BLG associating LPS
Table 8 has provided the influence of YPF-P to BLG associating LPS induction of immunity liver damage Mouse Liver spleen index.As shown in table 8, each treatment group of YPF-P all can reduce liver, the spleen index of immunologic liver injury mice, compares with model group, and middle and high dosage group difference has significance.
Table 8 YPF-P to the influence of BLG associating LPS induction of immunity liver damage Mouse Liver spleen index (x ± s, n=10)
#Compare with normal group P＜0.01;
*P＜0.05,
*Compare with model group P＜0.01
Table 9 has provided the influence of YPF-P to BLG associating LPS induction of immunity liver damage mice serum ALT and AST.As shown in table 9, in the damage of BCG+LPS inducing mouse generation immunity, the model group blood plasma level obviously raises; Each treatment group is prevented administration during modeling, YPF-P50,100,200mgkg
-The dosage group
1All can reduce ALT, the AST level that raises in the blood plasma with the BDP matched group, with YPF-P100,200mgkg
-1Dosage group and the effect of BDP group are the most remarkable.
Table 9 YPF-P to the influence of BLG associating LPS induction of immunity liver damage mice serum ALT and AST (x ± s, n=10)
#Compare with normal group P＜0.01;
*P＜0.05,
*Compare with model group P＜0.01
3.2 YPF-P is to the influence of acute immunologic liver injury mouse liver even slurry SOD, MDA, NO level
Table 10 has provided the influence of YPF-P to immunologic liver injury mouse liver even slurry SOD, MDA, NO level.As shown in table 10, model group liver homogenate SOD level reduces, and MDA, NO level raise simultaneously, the SOD level that each dosage group of YPF-P and BDP matched group all can raise and reduce, and reduce MDA, the NO level that raises, with YPF-P100,200mgkg
-1Dosage group effect is obvious.
Table 10 YPF-P to the influence of immunologic liver injury mouse liver even slurry SOD, MDA, NO level (x ± s, n=10)
#Compare with normal group P＜0.01;
*P＜0.05,
*Compare with model group P＜0.01
3.3 the influence that YPF-P changes acute immunologic liver injury mouse liver histopathology (HE dyeing, * 200)
Figure 13 unites lipopolysaccharide-induced acute immunologic liver injury experiment normal group mouse liver histopathology figure for bacillus calmette-guerin vaccine, shows as the hepatic tissue clear in structure among the figure, and hepatic cords is radial proper alignment around central vein, and the portal area does not have cell infiltration; Figure 14 unites lipopolysaccharide-induced acute immunologic liver injury model group mouse liver histopathology figure for bacillus calmette-guerin vaccine, and it is obvious to show as hepatic tissue cell swelling among the figure, and sees a large amount of spotty necrosis kitchen ranges, and massive inflammatory cells infiltrated is arranged around liver parenchyma and the portal area; Figure 15, Figure 16, Figure 17 are respectively YPF-P50mgkg
-1, 100mgkg
-1, 200mgkg
-1The dosage group is induced the figure that influences that acute chemical hepatic injury murine liver tissue histopathology changes to D-Ga1, shows as swelling of liver cell among the figure and degree of necrosis is light than model group, and with the increase of dosage, tissue injury's recovery extent more obviously; Figure 18 induces the figure that influences of acute chemical hepatic injury murine liver tissue histopathology variation, demonstration pathological change and degree and YPF-P200mgkg among the figure for the BDP matched group to D-Ga1
-1The dosage group is similar.
3.4 the influence that YPF-P expresses acute immunologic liver injury murine liver tissue TNF-α, iNOS (SP dyeing * 200)
Figure 19-Figure 24, Figure 25-Figure 30 are respectively immunohistochemical staining that TNF-α, iNOS express figure as a result.Figure 19, Figure 25 are respectively bacillus calmette-guerin vaccine and unite lipopolysaccharide-induced acute immunologic liver injury experiment normal group murine liver tissue TNF-α, iNOS expression figure, show as murine liver tissue TNF-α, iNOS among the figure and all are slight expression; Figure 20, Figure 26 are respectively bacillus calmette-guerin vaccine and unite lipopolysaccharide-induced acute immunologic liver injury model group murine liver tissue TNF-α, iNOS expression figure, show as murine liver tissue TNF-α, iNOS among the figure and express significantly enhancing, show to occur tangible yellow or brown yellow granule in cell cytoplasm, the nucleus; Figure 21,22,23 and Figure 27,28,29 be followed successively by YPF-P50mgkg respectively
-1, 100mgkg
-1, 200mgkg
-1The dosage group is to the figure that influences of acute immunologic liver injury murine liver tissue TNF-α, iNOS expression, and showing as among the figure with dosage increases, and TNF-α, iNOS express and weaken gradually; Figure 24, Figure 30 are for being the influence figure of bifendate matched group to acute immunologic liver injury murine liver tissue TNF-α, iNOS expression, performance and YPF-P200mgkg among the figure
-1The dosage group is similar.What table 11 was YPF-P to mouse immune liver damage model iNOS, TNF-alpha expression influences SABC graphical analysis result.
Table 11 number YPF-P (YPF-P) to mouse immune liver damage model iNOS, TNF-alpha expression influence SABC graphical analysis result (x ± s, n=6)
#Compare with normal group P＜0.01;
*P＜0.05,
*Compare with model group P＜0.01
3.5 the influence that YPF-P expresses acute immunologic liver injury murine liver tissue TNF-α mRNA
RT-PCR detects TNF-α mRNA changes of expression level in the hepatic tissue, the results are shown in Figure 31 and Figure 32.Figure 31 gel electrophoresis figure that acute immunologic liver injury murine liver tissue TNF-α mRNA expresses for YPF-P influences is compared with normal group, and model group is expressed obviously and strengthened, and each treatment group of YPF-P is compared with model group to express reducing tendency, and is dose dependent.The BDP matched group is expressed and YPF-P200mgkg
-1The dosage group is similar.Figure 32 gel electrophoresis sxemiquantitative cartogram that acute immunologic liver injury murine liver tissue TNF-α mRNA expresses for YPF-P influences.
4. result
Above-mentioned experimental result is arranged as seen, YPF-P200mgkg
-1The dosage group can significantly reduce the liver index of acute chemical hepatic injury mice, reduces third ALT, the AST level that raise in the serum; YPF-P100,200mgkg
-1The dosage group can significantly reduce the MDA content that raises in the mouse liver even slurry, improves the activity of SOD, GSH in the liver homogenate, proves that thus YPF-P has preventive and therapeutic effect to chemical liver injury.YPF-P100,200mgkg
-1The dosage group can significantly reduce liver, the index and spleen index of acute immunologic liver injury mice, reduce MDA, NO content in Serum ALT, AST level and the liver homogenate that raises, improve the activity of SOD in the liver homogenate, significantly reduce iNOS, TNF-α and the expression of mRNA in hepatic tissue thereof, prove that thus YPF-P has remarkable preventive and therapeutic effect to immunologic liver injury.
Below under the constant situation of other conditions, the mass fraction ratio of the Radix Astragali, Rhizoma Atractylodis Macrocephalae (parched) and Radix Saposhnikoviae is adjusted into 1:1:1,1:2:1,1:1:3,2:2:1,1:3:1,1.5:2:1,2:1.5:1,2:1:1 and 4; 4:1 produces respectively and obtains YPF-P.
Adopt as above-mentioned same pharmacological experimental method then and carry out the test of mouse liver injury preventive and therapeutic effect to producing each YPF-P that obtains respectively, the result who obtains proves: above-mentioned all have similar preventive and therapeutic effect with producing each YPF-P that obtains respectively after the Radix Astragali, Rhizoma Atractylodis Macrocephalae (parched) and the adjustment of Radix Saposhnikoviae mass fraction ratio to mouse liver injury.Proof thus, by mass fraction with Radix Astragali 1-4: Rhizoma Atractylodis Macrocephalae 1-4: the formulated YUPINGFENG SAN of Radix Saposhnikoviae 1-3 all can adopt above-mentioned test method to extract YUPINGFENG SAN polysaccharide active ingredient YPF-P dry powder as the medicine of preventing and treating hepatic injury.
In an embodiment under the constant situation of other conditions, every gram YUPINGFENG SAN is heated in 15 milliliters, 30 ml waters, the YPF-P that makes is roughly the same with the YPF-P that operates gained by embodiment aspect polysaccharide yield and the content; But prove that through control mouse liver injury pharmacological evaluation this YPF-P has similar control hepatic injury effect equally.
Under the constant situation of other conditions, when soak time was 0.5h, 10h, the YPF-P that makes was roughly the same with the YPF-P that operates gained by embodiment in an embodiment; Prove that through above-mentioned same pharmacological evaluation this YPF-P has similar control hepatic injury effect equally.
Under the constant situation of other conditions, when mode of heating was the ultrasound wave heating, the YPF-P that makes was lower than the YPF-P yield of operating gained by embodiment in an embodiment; Prove that through above-mentioned same pharmacological evaluation this YPF-P has similar control hepatic injury effect equally.
Under the constant situation of other conditions, when heating-up temperature was 50 ℃, the YPF-P yield that makes was lower in an embodiment, and is roughly the same when yield and 95 ℃ in the time of 100 ℃; Prove that through above-mentioned same pharmacological evaluation this YPF-P has similar control hepatic injury effect equally.
Under the constant situation of other conditions, when be 1h heat time heating time, the YPF-P yield that makes was lower in an embodiment, and yield is higher during 6h; Prove that through above-mentioned same pharmacological evaluation be that the YPF-P that 1h makes has similar control hepatic injury effect heat time heating time; Be that the YPF-P that 6h makes also has control hepatic injury effect heat time heating time, but curative effect is slightly poorer.
Under the constant situation of other conditions, when extraction time was 1 time, 3 times, the YPF-P that makes was roughly the same with the YPF-P that operates gained by embodiment aspect polysaccharide yield and the content in an embodiment; Prove to have similar control hepatic injury effect through above-mentioned same pharmacological evaluation.
In an embodiment under the constant situation of other conditions, when extracting solution be concentrated into original volume 1/7,1/10 the time, the YPF-P that makes is roughly the same with the YPF-P that operates gained by embodiment aspect polysaccharide yield and the content; Prove to have similar control hepatic injury effect through above-mentioned same pharmacological evaluation.
In an embodiment under the constant situation of other conditions, when with normal hexane, when ethyl acetate is made extractant, the YPF-P that makes is roughly the same with the YPF-P that operates gained by embodiment aspect polysaccharide yield and the content; Prove to have similar control hepatic injury effect through above-mentioned same pharmacological evaluation.
Under the constant situation of other conditions, when extraction times was 1 time, 3 times, the YPF-P that makes was roughly the same with the YPF-P that operates gained by embodiment aspect polysaccharide yield and the content in an embodiment; Prove to have similar control hepatic injury effect through above-mentioned same pharmacological evaluation.
Under the constant situation of other conditions, when the volume ratio of mixed organic solvents and extract was 1:2,2:1, the YPF-P that makes was roughly the same with the YPF-P that operates gained by embodiment aspect polysaccharide yield and the content in an embodiment; Prove to have similar control hepatic injury effect through above-mentioned same pharmacological evaluation.
In an embodiment under the constant situation of other conditions, when duration of oscillation was 10 seconds, the time of leaving standstill will lack, and when duration of oscillation is 30 seconds, the time of leaving standstill will be grown, and the YPF-P that makes is roughly the same with the YPF-P that operates gained by embodiment aspect polysaccharide yield and the content; Prove to have similar control hepatic injury effect through above-mentioned same pharmacological evaluation.
Under the constant situation of other conditions, when the repeatable operation number of times was 3 times, the YPF-P content that makes was lower in an embodiment, and the YPF-P content that makes in the time of 15 times is higher; Prove to have similar control hepatic injury effect through above-mentioned same pharmacological evaluation.
In an embodiment under the constant situation of other conditions, when drying mode is drying with water bath, the more difficult dissolving of the YPF-P that makes, the very easy dissolving again of the YPF-P that lyophilization and spray drying make, and outward appearance and character are better; Prove to have similar control hepatic injury effect through above-mentioned same pharmacological evaluation.
When YPF-P dry powder is used as control hepatic injury medicine clinically, face with preceding that YPF-P dry powder is oral after with normal saline, glucose saline, deionized water or dissolved in distilled water, clinical dosage is shown 0.9-1.8g/ day with the dry powder scale, divide 2 times oral, in the course of treatment at least one week, have and prevent and treat the hepatic injury effect significantly.
YPF-P dry powder is gone into various adjuvants such as wetting agent, lubricant, diluent, binding agent and/or coloring agent and make oral liquid, granule, tablet, capsule, dosage is shown 0.9-1.8g/ day with YPF-P dry powder scale, divide 2 times oral, in the course of treatment at least one week, have control hepatic injury effect equally.
Claims (7)
1, a kind of Chinese medicine YUPINGFENG SAN is as the using method of control liver injury medicament, it is characterized in that: press mass fraction with Radix Astragali 1-4: Rhizoma Atractylodis Macrocephalae 1-4: the YUPINGFENG SAN that Radix Saposhnikoviae 1-3 is formulated, water by this YUPINGFENG SAN of every gram and 10-30 milliliter is mixed, soak 0.5-10h, under this temperature, be incubated 1-6h after being heated to 50-100 ℃, carry out solid-liquid separation, isolated extracting solution is concentrated into the 1/5-1/10 of original volume; To hang down the extracting solution after the polar organic solvent extraction concentrates, keep lower floor's extract; 5:1 is mixed with mixed organic solvents with chloroform and n-butyl alcohol by volume, with this mixed organic solvents and this lower floor's extract by volume 1:2-2:1 mix, leave standstill after the vibration to layering, remove the intermediate layer; Upper layer of extraction liquid is mixed with the lower floor organic solvent, leave standstill to layering after the vibration again, remove the intermediate layer; Press the said process repeatable operation 3-15 time, keep the upper layer of extraction liquid that obtains at last; With this upper layer of extraction liquid with ethanol allocate to concentration of alcohol be 30%, discard precipitation after leaving standstill; With the gained supernatant again with ethanol allocate to concentration of alcohol be 50%, isolate precipitate after leaving standstill, be crushed into powder after the drying, be YUPINGFENG SAN polysaccharide active ingredient; Used as the control liver injury medicament, clinical dosage is shown 0.9-1.8g/ day with the dry powder scale, the course of treatment at least one week.
2, according to claim 1 the Chinese medicine YUPINGFENG SAN as the control liver injury medicament using method, be characterised in that the residue that will obtain after the described solid-liquid separation carries out multiple extraction, merge the extracting solution that each time obtains, the extracting solution after will merging again is concentrated into the 1/5-1/10 of original volume.
3, according to claim 1 the Chinese medicine YUPINGFENG SAN as the control liver injury medicament using method, be characterised in that described to hang down the extracting solution after the polar organic solvent extraction concentrates, after repeatedly extracting with low polar organic solvent again, keep the lower floor's extract that obtains at last.
4, the Chinese medicine YUPINGFENG SAN is characterised in that as the using method of control liver injury medicament described low polar organic solvent selects petroleum ether, normal hexane or ethyl acetate for use according to claim 1.
5, the Chinese medicine YUPINGFENG SAN is characterised in that as the using method of control liver injury medicament described drying selects drying under reduced pressure, drying with water bath, lyophilization or spray drying for use according to claim 1.
6, according to claim 1 the Chinese medicine YUPINGFENG SAN as the using method of control liver injury medicament, be characterised in that face with preceding YUPINGFENG SAN polysaccharide active ingredient dry powder with the 0.9-1.8g/ daily dose oral with appropriate solvent dissolving back branch 2 times; Described appropriate solvent is normal saline, glucose saline, deionized water or distilled water.
7, according to claim 1 the Chinese medicine YUPINGFENG SAN as the control liver injury medicament using method, be characterised in that to face YUPINGFENG SAN polysaccharide active ingredient is added the peroral dosage form that adjuvant is made on the basis of effective dose 0.9-1.8g/ day, comprise oral liquid, granule, tablet or capsule with preceding; Described adjuvant comprises wetting agent, lubricant, diluent, binding agent and/or coloring agent.
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| CN102885879A (en) * | 2012-11-02 | 2013-01-23 | 江苏吉贝尔药业有限公司 | Method for producing yupingfeng lyophilized powder capsule |
| CN105254777A (en) * | 2015-12-04 | 2016-01-20 | 佛山科学技术学院 | Extraction method of Yupingfeng polysaccharide |
| CN105920083A (en) * | 2016-04-27 | 2016-09-07 | 新乡医学院 | Application of Yupingfengsan to immunity improvement |
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| CN1836686A (en) * | 2005-03-23 | 2006-09-27 | 刘凯 | Chinese traditional effective part compound preparation for promoting organic body immune function, and its preparation method |
| CN1985875B (en) * | 2006-12-14 | 2011-07-20 | 天津科技大学 | Preparing process of Jade-screen total polyose |
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| CN102885879A (en) * | 2012-11-02 | 2013-01-23 | 江苏吉贝尔药业有限公司 | Method for producing yupingfeng lyophilized powder capsule |
| CN105254777A (en) * | 2015-12-04 | 2016-01-20 | 佛山科学技术学院 | Extraction method of Yupingfeng polysaccharide |
| CN105920083A (en) * | 2016-04-27 | 2016-09-07 | 新乡医学院 | Application of Yupingfengsan to immunity improvement |
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