CN103585239B - A kind of preparation method of Boraginaceae extract and application thereof - Google Patents
A kind of preparation method of Boraginaceae extract and application thereof Download PDFInfo
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Abstract
The present invention relates to a kind of preparation method and application thereof of Boraginaceae extract, belong to field of medicaments.Its preparation method take Flos et Folium Eritrichii Rupestris as raw material, after extracting, obtained, comprise Boraginaceae extract A, Boraginaceae extract B, Boraginaceae extract C, Boraginaceae extract D by twice purification with macroreticular resin by different concentration ethanol.This Boraginaceae extract, can combine with pharmaceutically acceptable carrier or other proper excipient, conventionally make the dosage form of oral administration.Boraginaceae extract of the present invention can be used for preparation treatment AIDS-treating medicine.
Description
Technical field
The invention belongs to biomedicine field, relate to a kind of preparation method and application thereof of Chinese medicinal material extract, specifically relate to a kind of preparation method and application thereof of Boraginaceae extract.
Background technology
Acquired immune deficiency syndrome (AIDS) is the new virus sexually transmitted disease found in 1981, and the traditional Chinese medical science past is not on the books to it.The traditional Chinese medical science is dialectical to acquired immune deficiency syndrome (AIDS), thinks that acquired immune deficiency syndrome (AIDS) belongs to epidemic disease category.The cause of disease is the scorchingly hot heresy invasion and attack of epidemic disease, deficiency of vital QI and causing.Pathogenesis is mainly weakened defensive QI, and yang-energy damages, and internal organs are got involved, and causes that the deficiency of the lung is insufficiency of the spleen suffers from a deficiency of the kidney, secondary infection.Thermal burn venation, coagulation of QI-blood, Xing Cheng mass in the abdomen lump tumor.
Syndrome Differentiation of Chinese Medicine " card ": mainly distinguish ebb and flow between pathogen and vital QI and deficiency syndrome excess syndrome.HIV (human immunodeficiency virus) is evil poison, and the change of disease in evil poison enters, the dynamic change of just resisting with the heresy of body healthy energy is advancing of disease process.Infect early stage based on the domination of pathogen, mid-term " causes void because of real ", late period " excess caused by deficiency ".The acquired immune deficiency syndrome (AIDS) state of an illness copies changeable, and acquired immune deficiency syndrome (AIDS) and secondary infection thereof all can be formed " disease ", and has difference " type " and " phase ".The traditional Chinese medical science has dialectical differential diagnosis of diseases to acquired immune deficiency syndrome (AIDS), the typing different diagnosis and treatment project such as by stages.
The Chinese medicine of acquired immune deficiency syndrome (AIDS) still belongs to conceptual phase at present, at home and abroad there is no at present generally acknowledged curative effect certainly go through produce AIDS virus resisting Chinese medicine or Chinese medicine monomer compound.AIDS virus resisting plant amedica and Chinese medicine research mainly divide 3 kinds of approach, and the AIDS virus resisting Chinese medicinal plant medicine that smoking set obtains and composition thereof, some is in clinic trial.
Since HIV (human immunodeficiency virus) finds, use HIV (human immunodeficiency virus) cell culture and reverse transcriptase in the world, the ill vitro test method such as protease and intergrase, while extensive SCREENED COMPOUND, also plant and Chinese herbal medicine extract and composition thereof is screened in a large number, also studied some compound Chinese medicinal preparation, find the active drug of inhibiting HIV in vitro, the viremia of the suppressed simian immunodeficiency virus had (SIV) infected monkey.Some is in clinic trial, not yet clear and definite effect.
China removes Africa treatment AIDS patients from 1987 Nian Pai combination of Chinese and Western medicine medical teams.Domestic before 2002, lack specially good effect Anti-AIDS Drugs, Chinese and western medicine, combines with dialectical to AIDS patients differential diagnosis of diseases according to theory of Chinese medical science clinical, with heat-clearing and toxic substances removing, strengthening the body resistance, or removing heat from blood the kidney invigorating, the rule for the treatment of such as blood circulation promoting and blood stasis dispelling, by the property of Chinese materia medica, select Chinese medicine, prescribed treatment.By clinical practice, think that some Medical Complex Prescription in Treating AIDS can improve symptom, mitigate the disease.In recent years to the AIDS patients that laboratory diagnosis is made a definite diagnosis, adopt treatment by Chinese herbs, if matched group, at treatments period, periodic detection blood viral charge (HIV-RNA) and cd4 cell level, evaluation effect.There is report to improve symptom, improve immunity, mostly still in clinical verification.Enter clinical AntiHIV1 RT activity single medicinal material or plant amedica and Chinese medicine compound as follows:
Glycyrrhizin: glycyrrhizin (glycyrrhizin, GL) be the glycyrrhizin that glycyrrhiza uralensis fisch (Glycyrrhiza glabra L) extracts, there is broad-spectrum disease resistance toxic action, vitro inhibition HIV1-RT, HIV cell fusion is suppressed in cell culture, also resisiting influenza virus and SARS virus, the dead and pneumonopathy change of the pneumonia that experiment in vivo proves to protect influenza infection mice to cause; The CCl of protection mice
4toxic liver injury; Induce gamma interferon; There is parahormone effect.Japan's its intravenous administrations chronic hepatitis B conventional, also once tried out treatment AIDS patients, can suppress serum HIV-1-P24 antigen, raised cd4 cell.Lv Weibai professor 1988-1992 can treat AIDS patients African Tanzania application glycyrrhizin oral formulations gram Chinese mugwort can improve symptom, and cd4 cell slightly rises.Its effect is not yet verified.
Trichosanthin: trichosanthin (Trichosanthin, TCS, GLQ223, Q material) belong to Cucurbitaceae (Cucurbitaceae) for Chinese medicine Fructus Trichosanthis (Trichosanthes kirilowii), the main Chinese medicine ingredients of China's Chinese medicine compound for miscarrying among the people is Fructus Trichosanthis, the effective ingredient of China scientist isolation identification device antiearly pregnancy and induced labor is trichosanthin, molecular weight is 27kD, one, secondary structure and three dimensional structure are understood fully completely, for the toxalbumin of Profilin synthesis, ribosome inactivating protein (the Ribosome Inactivating Protein of strand, RIP).The U.S. is when screening anti-HIV-1 medicines, find that it suppresses HIV-1 cell fusion in lymphocyte is cultivated, thereafter find P24 antigen and the RNA of HIV-1 in mononuclear phagocyte is cultivated, blood plasma HIV-1 can be suppressed in the intravenous injection of 3 simian acquired immunodeficiency syndrome virus (SIV) infected monkeys.The U.S. uses intravenous administrations AIDS patients in forest farm, and HIV-1-P24 antigen in the T lymphocyte that peripheral blood HIV-1 can be suppressed to infect and mononuclear phagocyte, makes CD4/ CD8 ratio slightly raise.Due to anaphylactic reaction can be caused dead, neurovirulent Mental Subnormality and nephrotoxicity reaction, clinical experiment stops.From Radix Trichosanthis, isolate other albumen TAP29 in recent years, molecular weight, at 29 kD, also have the effect suppressing HIV-1, but toxicity is less, not in clinic trial.In addition the bitter melon protein of Fructus Momordicae charantiae (MOmordica charantia) and the Radix Phytolaccae albumen (Pokeweed protein) etc. of Radix Phytolaccae (Phytolacca americana) have the effect suppressing HIV.
Hypericin: plant Herba Hyperici Monogyni (St. John Wort, Hypericum perforatum) its extract for treating depression of folk tradition, the active component proposed from flower is hypericin (hypericin), it is the multi-ring dimerization anthraquinone analog compound of aromatic series, for anti-depression drug, complete synthesis.Hypericin has multiple antiviral activity, in cell culture, suppress HIV, and its mechanism of action is undressing of blocking virus, and assembling and release, the effect of killing the virus, by blood brain barrier.Murine leukemia virus can be suppressed in cell culture to breed.The splenomegaly that murine leukemia virus infecting mouse causes can be suppressed in vivo, duck hepatitis B virus infection Sanguis Anas domestica also can be suppressed to ask viral DNA, and have broad spectrum antibiotic active.Once in treatment on probation of clinical I/II phase HIV sufferers, test had been stopped because there being light sensitization.
1992 year report the native compound that from Malaysian rainforest plant Calophyllum lanigerum be separated dextrorotation lane element A: dextrorotation lane element A(+Calanolide A), Mus coumarin kind compound, suppress HIV1-RT, but invalid to HIV-2.Belong to non-nucleoside HIV1-RT inhibitor, in cell culture, multi-to-multi lives in the responsive and drug resistance HIV-1 of the AZT of clinical separation effectively, and and AZT, NVP have synergism.In HIV hollow fiber pipe Mice Body, prove there is AIDS virus resisting effect in experimental model (HIV hollow fiber mouse model), can drug effect be strengthened with AZT use in conjunction.Complete synthesis, having carried out its clinical trial of I-II proves there is the therapeutic effect suppressing HIV-1 to infect, and toxicity is little, and side effect is light, has headache and inappetence etc., effective with AZT drug combination.Be the single plant component uniquely entering II phase clinical experiment at present, observing in checking curative effect, not yet ratify to produce.
Herba Sidae Rhombifoliae soup: Herba Sidae Rhombifoliae soup (Radix Bupleuri, Radix Ginseng, the Rhizoma Pinelliae, Radix Scutellariae, Radix Glycyrrhizae, Rhizoma Zingiberis Recens, Fructus Jujubae)
It is Zhang Zhongjing Treatise on Febrile Diseases side.Hepatitis B is treated with it by Japan, finds that it suppresses HIV in cell culture.The U.S. is used for the treatment of numerical example AIDS patients clinical, can reduce blood-serum P 24 antigen.We prove that it has the effect suppressing HIV1-RT and suppress HIV-1 in cell culture.Study the effect of its 7 kinds of composition Chinese medicines respectively, find that only Radix Scutellariae water extract and composition baicalin (baicalin) thereof have and suppress HIV-1 effect, HIV1-RT is suppressed in vitro in cell free system, HIV-1 cytopathy is suppressed in human T-cell and peripheral blood lymphocytes cultivate, FLuorescent antigen and P24 antigen, in body, oral splenomegaly and the blood leukocytes of suppressing of murine leukemia virus infecting mouse raises.Japan Ono reports that noroxylin (baicalien) suppresses HIV1-RT.
In grind No. I: in grind No. I commodity such as (principal agent has Herba Violae, Radix Trichosanthis composition) and be called: Chinese mugwort
Gram electuary take heat-clearing and toxic substances removing as the rule for the treatment of, by Chinese medicine research institute of China prescription 1992-1993 in African Tanzania to the clinical practice of early metaphase acquired immune deficiency syndrome (AIDS), think to have and necessarily improve symptom, improve the effect of immunity.Laboratory proves that this medicine vitro inhibition hiv reverse transcriptase is active, simian acquired immunodeficiency syndrome poison infected monkey experimental therapy result is shown in body, plasma viral titre can be reduced, and cd4 cell number and CD4/CD8 ratio and T can be improved, bone-marrow-derived lymphocyte numerical value, inducement interferon and generation immunoloregulation function.Once in Africa and China's clinic trial, not yet clinical verification was ratified.
Zhongyan No. 2: Zhongyan No. 2 (Radix Astragali, Radix Ginseng, Radix Angelicae Sinensis, Fructus Lycii and Radix Glycyrrhizae etc.) is China's traditional Chinese medical science
Academy medical team prescription.1992 at African Tanzania treatment AIDS patients.1998-1999 is at African special outpatient clinic to 29 routine AIDS patients treatments, and 2/16 routine HIV virus load declines, and the routine CD4 of CD4 8/29 raises.2001-2002 helps the routine AIDS patients of peace hospitalize 15 in Beijing, 3/15 routine virus load declines, and the routine CD4 of CD4 1/15 raises.Be improved the effect of symptom.Experiment proves that this medicine shows simian acquired immunodeficiency syndrome poison infected monkey experimental therapy in human body, and medication 4-12 week can reduce plasma viral titre, P27 antigen, and can improve cd4 cell number, has activation to infected monkey lymphocyte.
Flos et Folium Eritrichii Rupestris is raw Flos et Folium Eritrichii Rupestris (Eritrichium rupestre(Pall.) Bunge of comfrey stone) dry aerial parts.Gather when summer, the flowers are in blossom, removing root and impurity, dry in the shade.Record in 1998 " Drug Standard of Ministry of Public Health of the Peoples Republic of China mongolian medicine fascicle ", it is cool in nature, bitter in the mouth, sweet.Have and kill " gluing ", heat clearing away, removing toxic substances.For warm influenza, " association's day " human connection disease, " association's day " heat.The common drug of the minority areas such as illiteracy is hidden for China.But about the basic research of Flos et Folium Eritrichii Rupestris is still very limited, the follow-up promotion and application of this medical material are restricted.Modern study rarely has report about the chemical composition of Flos et Folium Eritrichii Rupestris and pharmacological action.
Domestic patent search result, has no Flos et Folium Eritrichii Rupestris Patents.
Above-mentioned document and patent etc., there is not yet the relevant report of Boraginaceae extract preparation method and application thereof.
Summary of the invention
The object of the present invention is to provide a kind of preparation method and application thereof of Boraginaceae extract.
The present invention is achieved through the following technical solutions:
The raw materials used Flos et Folium Eritrichii Rupestris of the present invention is raw Flos et Folium Eritrichii Rupestris (Eritrichium rupestre(Pall.) Bunge of comfrey stone) dry aerial parts.
The preparation method of a kind of Boraginaceae extract of the present invention and application thereof; The preparation method of described Boraginaceae extract is:
(1) Flos et Folium Eritrichii Rupestris, soaks 4-12 hour, adopts extraction by steam distillation volatile oil, extraction time 1-4 hour, extraction time 1-3 time, and amount of water is 8-15 times of Flos et Folium Eritrichii Rupestris weight, collects volatile oil, obtains Boraginaceae extract A; Flos et Folium Eritrichii Rupestris aqueous extract is filtered, obtains medicinal liquid A and medicinal residues A respectively;
(2) step (1) being obtained medicinal residues A concentration 50%-95% ethanol is solvent, heating and refluxing extraction, extraction time is 1-3 time, each extraction time is 1-3 hour, and each solvent load is 8-15 times of Flos et Folium Eritrichii Rupestris weight, filters, extracting solution reclaims ethanol, concentrated, dry, obtain Boraginaceae extract B;
(3) by the medicinal liquid A that step (1) obtains, be concentrated into relative density d=1.10-1.20, filter, medicinal liquid, by nonpolar macroporous adsorption resin, first washes with water, and water elution liquid is directly by polar macroporous resin; Use the alcoholic solution eluting nonpolar macroporous adsorption resin of concentration 30%-70% again, collect different concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract C; Use the alcoholic solution eluting polar macroporous adsorption resin of concentration 50%-95% again, collect different concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract D;
(4) above-mentioned Boraginaceae extract A, Boraginaceae extract B, Boraginaceae extract C, Boraginaceae extract D, wherein one or both or three kinds or four kinds mix by a certain percentage, obtain Boraginaceae extract of the present invention.
A kind of preparation method of Boraginaceae extract is:
(1) Flos et Folium Eritrichii Rupestris, soaks 4-12 hour, adopts extraction by steam distillation volatile oil, extraction time 1-4 hour, extraction time 1-3 time, and amount of water is 8-15 times of Flos et Folium Eritrichii Rupestris weight, collects volatile oil, obtains Boraginaceae extract A; Flos et Folium Eritrichii Rupestris aqueous extract is filtered, obtains medicinal liquid A and medicinal residues A respectively;
(2) step (1) being obtained medicinal residues A concentration 50%-95% ethanol is solvent, heating and refluxing extraction, extraction time is 1-3 time, each extraction time is 1-3 hour, and each solvent load is 8-15 times of Flos et Folium Eritrichii Rupestris weight, filters, extracting solution reclaims ethanol, concentrated, dry, obtain Boraginaceae extract B;
(3) by the medicinal liquid A that step (1) obtains, be concentrated into relative density d=1.10-1.20, filter, medicinal liquid, by nonpolar macroporous adsorption resin, first washes with water, and water elution liquid is directly by polar macroporous resin; Use the alcoholic solution eluting nonpolar macroporous adsorption resin of concentration 30%-70% again, collect different concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract C; Use the alcoholic solution eluting polar macroporous adsorption resin of concentration 50%-95% again, collect different concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract D;
(4) by above-mentioned Boraginaceae extract A, Boraginaceae extract B, Boraginaceae extract C, Boraginaceae extract D mixing, Boraginaceae extract of the present invention is obtained.
The preparation method of preferred a kind of Boraginaceae extract is:
(1) Flos et Folium Eritrichii Rupestris, soaks 6 hours, and adopt extraction by steam distillation volatile oil, 2 hours extraction times, extraction time 2 times, amount of water is 12 times of Flos et Folium Eritrichii Rupestris weight, collects volatile oil, obtains Boraginaceae extract A; Flos et Folium Eritrichii Rupestris aqueous extract is filtered, obtains medicinal liquid A and medicinal residues A respectively;
(2) step (1) being obtained medicinal residues A concentration 60% ethanol is solvent, heating and refluxing extraction, and extraction time is 2 times, and each extraction time is 2 hours, each solvent load is 12 times of Flos et Folium Eritrichii Rupestris weight, filters, and extracting solution reclaims ethanol, concentrated, dry, obtain Boraginaceae extract B;
(3) by the medicinal liquid A that step (1) obtains, be concentrated into relative density d=1.12, filter, medicinal liquid, by D101 nonpolar macroporous adsorption resin, first washes with water, the polar macroporous resin of the DM130 that water elution liquid directly passes through; Use the alcoholic solution eluting D101 nonpolar macroporous adsorption resin of concentration 60% again, collect 60% concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract C; Use the alcoholic solution eluting DM130 polar macroporous adsorption resin of concentration 75% again, collect 75% concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract D;
(4) by above-mentioned Boraginaceae extract A, Boraginaceae extract B, Boraginaceae extract C, Boraginaceae extract D mixing, Boraginaceae extract of the present invention is obtained.
The preparation method of a kind of Boraginaceae extract of the present invention, is characterized in that: the nonpolar macroporous adsorption resin adopted is D101 macroporous adsorbent resin, AB-8 macroporous adsorbent resin; The polar macroporous adsorption resin adopted is DM130 macroporous adsorbent resin, ADS-17 macroporous adsorbent resin.
The preparation method of a kind of Boraginaceae extract of the present invention and application, the described application be applied as in preparation treatment AIDS-treating medicine.
The treatment AIDS-treating medicine that Boraginaceae extract of the present invention and chemical drugs or Chinese medicine or natural drug form.
Boraginaceae extract A of the present invention, Boraginaceae extract B, Boraginaceae extract C, the Boraginaceae extract D application in preparation treatment AIDS-treating medicine.
The treatment AIDS-treating medicine that Boraginaceae extract A of the present invention, Boraginaceae extract B, Boraginaceae extract C, Boraginaceae extract D and chemical drugs or Chinese medicine or natural drug form.
Boraginaceae extract of the present invention, by adding the various adjuvants that pharmaceutics allows, makes the peroral dosage forms such as the tablet on pharmaceutics, granule, capsule.
The present invention's exploratory development is first with raw Flos et Folium Eritrichii Rupestris (Eritrichium rupestre(Pall.) Bunge of comfrey stone) dry aerial parts be that raw material extracts preparation treatment acquired immune deficiency syndrome (AIDS) extract.This experimentation shows, Boraginaceae extract of the present invention can be used for preparation treatment AIDS-treating medicine, the toxicity of Boraginaceae extract to C8166 cell is little, induces C8166 cell to form syncytium have significant inhibit activities to HIV-1, can apply in preparation treatment AIDS-treating medicine.Clinical treatment HIV sufferers is evident in efficacy.
Detailed description of the invention
Extremely apply below by specific experiment example and the preparation method of embodiment to a kind of Boraginaceae extract of the present invention and be described further, but be not limited to the present invention.
Be described in further detail below by specific embodiment.
Embodiment 1: the preparation of Boraginaceae extract
(1) Flos et Folium Eritrichii Rupestris 19kg, soaks 6 hours, and adopt extraction by steam distillation volatile oil, 2 hours extraction times, extraction time 2 times, amount of water is 12 times of Flos et Folium Eritrichii Rupestris weight, collects volatile oil, obtains Boraginaceae extract A; Flos et Folium Eritrichii Rupestris aqueous extract is filtered, obtains medicinal liquid A and medicinal residues A respectively;
(2) step (1) being obtained medicinal residues A concentration 60% ethanol is solvent, heating and refluxing extraction, and extraction time is 2 times, and each extraction time is 2 hours, each solvent load is 12 times of Flos et Folium Eritrichii Rupestris weight, filters, and extracting solution reclaims ethanol, concentrated, dry, obtain Boraginaceae extract B;
(3) by the medicinal liquid A that step (1) obtains, be concentrated into relative density d=1.12, filter, medicinal liquid, by D101 nonpolar macroporous adsorption resin, first washes with water, and water elution liquid is directly by the polar macroporous resin of DM130; Use the alcoholic solution eluting D101 nonpolar macroporous adsorption resin of concentration 60% again, collect 60% concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract C; Use the alcoholic solution eluting DM130 polar macroporous adsorption resin of concentration 75% again, collect 75% concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract D;
(4) by above-mentioned Boraginaceae extract A, Boraginaceae extract B, Boraginaceae extract C, Boraginaceae extract D mixing, Boraginaceae extract of the present invention is obtained.
Embodiment 2: the preparation of Boraginaceae extract
(1) Flos et Folium Eritrichii Rupestris 21kg, soaks 4 hours, and adopt extraction by steam distillation volatile oil, 1 hour extraction time, extraction time 3 times, amount of water is 8 times of Flos et Folium Eritrichii Rupestris weight, collects volatile oil, obtains Boraginaceae extract A; Flos et Folium Eritrichii Rupestris aqueous extract is filtered, obtains medicinal liquid A and medicinal residues A respectively;
(3) step (1) being obtained medicinal residues A concentration 50% ethanol is solvent, heating and refluxing extraction, and extraction time is 1 time, and each extraction time is 3 hours, each solvent load is 8 times of Flos et Folium Eritrichii Rupestris weight, filters, and extracting solution reclaims ethanol, concentrated, dry, obtain Boraginaceae extract B;
(2) by the medicinal liquid A that step (1) obtains, be concentrated into relative density d=1.20, filter, medicinal liquid, by AB-8 nonpolar macroporous adsorption resin, first washes with water, and water elution liquid is directly by the polar macroporous resin of ADS-17; Use the alcoholic solution eluting AB-8 nonpolar macroporous adsorption resin of concentration 30% again, collect 30% concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract C; Use the alcoholic solution eluting ADS-17 polar macroporous adsorption resin of concentration 50% again, collect 50% concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract D;
(4) by above-mentioned Boraginaceae extract A, Boraginaceae extract B, Boraginaceae extract C, Boraginaceae extract D mixing, Boraginaceae extract of the present invention is obtained.
Embodiment 3: the preparation of Boraginaceae extract
(1) Flos et Folium Eritrichii Rupestris 25kg, soaks 12 hours, and adopt extraction by steam distillation volatile oil, 4 hours extraction times, extraction time 1 time, amount of water is 15 times of Flos et Folium Eritrichii Rupestris weight, collects volatile oil, obtains Boraginaceae extract A; Flos et Folium Eritrichii Rupestris aqueous extract is filtered, obtains medicinal liquid A and medicinal residues A respectively;
(3) step (1) being obtained medicinal residues A concentration 95% ethanol is solvent, heating and refluxing extraction, and extraction time is 3 times, and each extraction time is 1 hour, each solvent load is 15 times of Flos et Folium Eritrichii Rupestris weight, filters, and extracting solution reclaims ethanol, concentrated, dry, obtain Boraginaceae extract B;
(2) by the medicinal liquid A that step (1) obtains, be concentrated into relative density d=1.10, filter, medicinal liquid, by D101 nonpolar macroporous adsorption resin, first washes with water, and water elution liquid is directly by the polar macroporous resin of DM130; Use the alcoholic solution eluting D101 nonpolar macroporous adsorption resin of concentration 70% again, collect 70% concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract C; Use the alcoholic solution eluting DM130 polar macroporous adsorption resin of concentration 95% again, collect 95% concentration ethanol eluent, concentrate drying, obtains Boraginaceae extract D;
(4) by above-mentioned Boraginaceae extract A, Boraginaceae extract B, Boraginaceae extract C, Boraginaceae extract D mixing, Boraginaceae extract of the present invention is obtained.
Embodiment 4: the preparation of Boraginaceae extract tablet
Example 1 Boraginaceae extract 365g, adds starch 110g, mixing, granulates, dry, sieves, adds microcrystalline Cellulose 15g, magnesium stearate 5g, and mixing, is pressed into 1000, obtains Boraginaceae extract tablet.
Embodiment 5: the preparation of Boraginaceae extract granule
Example 2 Boraginaceae extract 365g, adds dextrin 160g, mixing, granulates, and dry, granulate, obtains Boraginaceae extract granule.
Embodiment 6: the preparation of Boraginaceae extract capsule
Example 3 Boraginaceae extract 345g, adds starch 85g, mixing, and granulate, granulate, obtains Boraginaceae extract capsule by encapsulated 1000.
Experimental example 1: the clinical observation of Boraginaceae extract capsule for treating acquired immune deficiency syndrome (AIDS) is tested.
1, money so-and-so, man, 43 years old, Changchun, Jilin, low grade fever January, with pharyngalgia, went out acquired immune deficiency syndrome (AIDS) to examination in hospital, treats three months by Western medicine system, do not take a turn for the better, and occurred that tumor is itched dermatitis, and band blood streak diarrhoea appears in the later stage.On June 10th, 2010 starts to take Boraginaceae extract capsule of the present invention, each serving with 5, takes 3 every day, stopping of suffering from diarrhoea after three days, and take dermatitis after three months and disappear, can also go to work in the fields so far, health is fine.
2, grandson so-and-so, female, 32 years old, people from Liaoning, Shenyang, whole body tumor was itched, and skin ulcer face appears in perineal position, is acquired immune deficiency syndrome (AIDS) to examination in hospital.Fail to take effect January through western medicine, there is dysentery of having blood in stool on a small quantity on the contrary, take Boraginaceae extract capsule of the present invention, each serving with 5, take 3 every day, within 4 days, dysentery stops, and fully recover in skin ulcer face, 4 months posterior perineum portions, also simultaneously by Boraginaceae extract washout of the present invention, within 3 days, wash once, wash nine times altogether, healthy so far, normal work can be participated in
Experimental example 2 Boraginaceae extract In Vitro Anti HIV-1 activity test
Testing sample: medicine: Boraginaceae extract, Boraginaceae extract A, Boraginaceae extract B, Boraginaceae extract C, Boraginaceae extract D, is and prepares by the embodiment of the present invention 1 method.Positive reference compound nitrine breast former times (AZT, 3'-Azido-3'-deoxythymidine) available from Sigma.Testing sample is dissolved in dimethyl sulfoxide (DMSO), 4 DEG C of preservations after subpackage; AZT is dissolved in RPMI-1640 complete medium, and 0.22um membrane filtration is degerming ,-20 DEG C of preservations after subpackage.
Reagent: HEPES (4-light ethyl croak prolixity ethyl sulfonic acid), MTT (3-(4; 5 one dimethyl expose to the sun miles-2)-2; 5 one diphenyl four nitrogen mile smelling salts), DMF (N, N mono-dimethyl methyl phthalein amine), penicillin (Penicillin), streptomycin sulfate (Streptomycin sulfate), paddy ammonia phthalein amine (Glutamine) all available from Sigma; 2-coloured glaze base ethanol (2-ME, 2-Mercaptoethanol) is Bio-Rad Products.RPMI-1640 and newborn calf serum are Gibco Products.
Culture medium: RPMI-1640 complete medium, containing 10% newborn calf serum, 2mM L-paddy ammonia phthalein amine, 10mM HEPES, 50uM 2-coloured glaze base ethanol, 100000IU penicillin, 100 μ g/ml streptomycins.
Cell and virus: human T lymphocyte system C8166, HIV-1 test strain HIV-1
b.Be so kind as to give by Britain MedicalResearch Council, A work DS Reagent Project.All cells and virus are all cultivated containing RPM work-1640 complete medium of 10% calf serum.Prepare HIV-1 according to a conventional method
b, titration also calculates viral TCID
50.After virus stock solution subpackage, put-70 DEG C of preservations.Cell and virus freezing and thawing according to a conventional method.
HIV-1 infectious titration: HIV-1
bcarry out titration by Johnson & Byington (1990) described method improvement, be summarized as follows: HIV-1 stock solution is done 4 times of dilutions on 96 orifice plates, 10 gradients, every gradient 6 repeating holes, arrange control wells 6 hole simultaneously.Every hole adds C8166 cell 50u1 (4 × 10
5/ ml), every hole final volume is 200u1.37 DEG C, CO
2cultivate.Within 3rd day, add fresh RPMI-1640 complete medium 100u1, under inverted microscope, within 7th day, observe the cytopathic effect (CytopathicEffect of HIV-1 induction in every hole, CPE), the formation of syncytium (Syncytium) whether is had to determine with every hole.The TCID of virus is calculated by Reed & Muench method
50(50% Tissue Culture Infection Dose, tissue culture infective dose).
Toxicity test to C8166 cell: by 4 × 10
5/ ml C8166 cell suspension 100u1 mixes from different drug solutions to be measured, if three repeating holes.The control wells of not drug containing is set simultaneously.37 DEG C, CO
2cultivate 3 days, adopt MTT colorimetric determination cytotoxicity.ELx800 microplate reader measures OD value, and mensuration wavelength is 595nm, and reference wavelength is 630nm.Calculate CC
50value (50%Cytotoxic Concentration), namely to 50% normal T-lymphocytes system C8166 produce toxicity time drug level.Experimental result is as table 1.
Table 1 Boraginaceae extract is to the experiment number of the toxic action of C8166 cell.
Test example 3 Boraginaceae extract is to HIV-1
bthe inhibition test of cytopathogenic effect (CPE)
By 8 × 10
5/ ml C8166 cell 50u1/ hole is inoculated on the 96 porocyte culture plates containing 100u1/ gradient pores doubling dilution medicine, then adds the HIV-1 of 50u1
bdilution supernatant, 1300TCID
50/ hole.If 3 repeating holes.The normal cell controls hole of not drug containing is set simultaneously.AZT is positive drug control.37 DEG C, 5% CO
2cultivate 3 days, under inverted microscope, (100 ×) count plasmodial formation.EC
50(50% Effective Concentration) is for suppressing drug level during Syncytium formation 50%.Experimental result is as shown in table 2.
Table 2 Boraginaceae extract is to HIV-1
bthe inhibitory action of induction C8166 cytopathy (CPE)
Result shows, Boraginaceae extract is to HIV-1
binduction C8166 cytopathy has good inhibitory action.
Positive drug AZT(azidothymidine AZT) be the inverase used at first clinically for 1987, single or bigeminy treats HIV, because namely this treatment occurred obvious drug resistance usually in 6 months, and it is not good to the immunologic reconstitution of body, and also have the toxicities such as bone marrow depression, leukopenia, total lymphocyte number minimizing and megaloblastic anemia, therefore comprehensive therapeutic effect is unsatisfactory clinically.The toxicity of Boraginaceae extract to C8166 cell is little, induces C8166 cell to form syncytium have significant inhibit activities to HIV-1, can apply in preparation treatment AIDS-treating medicine.
Claims (3)
1. the application of Boraginaceae extract in preparation treatment AIDS-treating medicine, the preparation method of described Boraginaceae extract is: Flos et Folium Eritrichii Rupestris, soak 4-12 hour, adopt extraction by steam distillation volatile oil, extraction time 1-4 hour, extraction time 1-3 time, amount of water is 8-15 times of Flos et Folium Eritrichii Rupestris weight, collect volatile oil, obtain Boraginaceae extract A, Boraginaceae extract A is Boraginaceae extract.
2. application according to claim 1, it is characterized in that the preparation method of described Boraginaceae extract is: Flos et Folium Eritrichii Rupestris, soak 6 hours, adopt extraction by steam distillation volatile oil, 2 hours extraction times, extraction time 2 times, amount of water is 14 times of Flos et Folium Eritrichii Rupestris weight, collect volatile oil, obtain Boraginaceae extract A, Boraginaceae extract A is Boraginaceae extract.
3. application according to claim 1, is characterized in that the treatment AIDS-treating medicine that described Boraginaceae extract and chemical drugs or Chinese medicine or natural drug form.
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| 紫草药理作用及有效成分提取研究进展;韦新成等;《包头医学院学报》;20111231;第27卷(第3期);125-126 * |
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