CN105147804A - Total ziyuglycoside extract and preparation method and application thereof - Google Patents
Total ziyuglycoside extract and preparation method and application thereof Download PDFInfo
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Abstract
The invention provides total ziyuglycoside extract and a preparation method and application thereof. The total ziyuglycoside extract has total saponins content up to 98.50%. Compared with existing methods, the preparation method has the advantage that content of total ziyuglycoside in the total ziyuglycoside extract is significantly increased, with activity of rising white blood cells being significantly improved. The preparation method is simple to operate, high in efficiency and low in production cost, and industrial production of the total ziyuglycoside is benefited. The rising white blood cells in the total ziyuglycoside extract prepared by the invention are high in activity, and a new pharmaceutical for the clinical treatment of leukopenia is provided.
Description
Technical field
The present invention relates to a kind of radix sanguisorbae total saponin extract and its production and use, belong to drug world.
Background technology
Often there is myelosuppression in tumor Radiotherapy chemotherapy process, thus cause leukopenia.Leukocyte count reduces, and can weaken human body immunity, the infection that clinical normal appearance is serious and hemorrhage.Therefore, the leukopenia that treatment tumor Radiotherapy chemotherapy causes needs to set about from protection hemopoietic function of bone marrow.
Radix Sanguisorbae is used for as prescribed preparation the clinical history that myelosuppressive treatment has more than 20 year, there is the advantages such as taking dose is little, evident in efficacy, safety non-toxic, wide material sources, preparation technology are simple, cheap, be therefore expected to be developed to clinical prevention and the irreplaceable natural drug for the treatment of bone marrow depression.
Confirm at present, Radix Sanguisorbae urgees material base mainly saponin and the tannin two class material of hemopoietic.The method for separating and preparing of tannin and technology relative maturity, and a kind of method not having radix sanguisorbae total saponin of good separation and purification high level at present on producing.
Application number: 200910082470.6, denomination of invention: the preparation method of radix sanguisorbae total saponin and Ziyuglycoside I, the method needs to carry out separation and purification by macroporous resin, and cost is higher, and efficiency is low, is unfavorable for commercial production.Application number: 201210129134.4 denominations of invention: the isolation and purification method of radix sanguisorbae total saponin, the method needs Radix Sanguisorbae extracting solution is carried out to defat, extraction, precipitation, extracts, in preparation process, extracting solution is through repeatedly shifting, radix sanguisorbae total saponin loss is large, complex steps, is also unfavorable for commercial production.
Therefore, provide a kind of technological operation is easy, cost is low, efficiency is high, prepare radix sanguisorbae total saponin content high and the Application and Development of preparation method to Radix Sanguisorbae resource being suitable for industrialized great production has very important meaning.
Summary of the invention
Technical scheme of the present invention there is provided a kind of radix sanguisorbae total saponin extract and its production and use.
The invention provides a kind of radix sanguisorbae total saponin extract, in this extract, Ziyuglycoside I content is: 50%-97%, and Ziyuglycoside I I content is: 2%-10%.
Preferably, in this extract, Ziyuglycoside I content is: 92.34%, and Ziyuglycoside I I content is: 6.16%.
Wherein, in this extract, total saponins weight percentage is 60-99%.
Preferably, in this extract, total saponins weight percentage is 98.50%.
Wherein, described extract, is prepared as follows:
A. get Radix Sanguisorbae medical material, add organic solvent extraction, extracting solution is for subsequent use;
B. get the extracting solution that step a is obtained, adjust pH to 12 ~ 14, leave standstill, centrifugal, sucking filtration obtains supernatant, supernatant concentrating under reduced pressure, for subsequent use;
C. get the obtained supernatant of step b and adjust pH to 11 ~ 12, leave standstill, centrifugal, collecting precipitation; Supernatant continues to adjust pH to 12 ~ 13, leaves standstill, centrifugal, collecting precipitation; Merge twice precipitation;
The precipitation of d. getting step c obtained adds organic solvent extraction, filters, and collect filtrate, concentrating filter liquor, drying, obtain radix sanguisorbae total saponin.
The invention provides a kind of method preparing radix sanguisorbae total saponin extract, it comprises the steps:
A. get Radix Sanguisorbae medical material, add organic solvent extraction, extracting solution is for subsequent use;
B. get the extracting solution that step a is obtained, adjust pH to 12 ~ 14, leave standstill, centrifugal, sucking filtration obtains supernatant, supernatant concentrating under reduced pressure, for subsequent use;
C. get the obtained supernatant of step b and adjust pH to 11 ~ 12, leave standstill, centrifugal, collecting precipitation; Supernatant continues to adjust pH to 12 ~ 13, leaves standstill, centrifugal, collecting precipitation; Merge twice precipitation;
The precipitation of d. getting step c obtained adds organic solvent extraction, filters, and collect filtrate, concentrating filter liquor, drying, obtain radix sanguisorbae total saponin.
Wherein, in step a, add the organic solvent reflux, extract, 2 times of Radix Sanguisorbae medical material 8 times amount, each 1.5 hours; Described organic solvent is ethanol or methanol, and concentration of alcohol is 30% ~ 95%v/v.
Preferably, described organic solvent is ethanol, and concentration of alcohol is 90%.
Wherein, in step b, the aqueous slkali that extracting solution adds 10% adjusts pH to 12 ~ 14, hold over night, and centrifugal, sucking filtration obtains supernatant, and supernatant is evaporated to proper volume, add water in right amount to concentration of alcohol be 20%v/v, for subsequent use; Described aqueous slkali is NaOH, KOH or Ca (OH)
2solution.
Preferably, described aqueous slkali is NaOH solution.
In step c, time of repose is 12h, collection be deposited in 70 DEG C of drying under reduced pressure.
In steps d, get the precipitation that step c is obtained, to reflux 45min with organic solvent, filter, collect filtrate, reclaim under reduced pressure organic solvent is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, obtains radix sanguisorbae total saponin; Wherein, described organic solvent is dehydrated alcohol or absolute methanol.
Preferably, described organic solvent is dehydrated alcohol.
The invention provides described radix sanguisorbae total saponin extract and prepare the purposes had in the medicine of function of increasing leukocyte.
Present invention also offers a kind of pharmaceutical composition of leukocyte increasing, it is active component by described radix sanguisorbae total saponin extract, adds pharmaceutically acceptable adjuvant or complementary composition is prepared into pharmaceutically conventional preparation.
Wherein, described preparation is oral formulations.
The present invention, by the improvement to radix sanguisorbae total saponin extract preparation method, substantially increases the content of radix sanguisorbae total saponin, makes its content reach 98.50%.With existing Measures compare, radix sanguisorbae total saponin significantly increases in Ziyuglycoside I and Ziyuglycoside I I content, and the activity of leukocyte increasing have also been obtained remarkable lifting.Present invention process is easy and simple to handle, and efficiency is high, and production cost is low, is conducive to the industrialized great production of radix sanguisorbae total saponin.Radix sanguisorbae total saponin prepared by the present invention is that the clinical treatment of leukopenia provides a kind of new medicine.
Below by way of detailed description of the invention, the present invention is described in further detail, but do not limit the present invention, the various change that those skilled in the art make according to the present invention and replacement, only otherwise depart from spirit of the present invention, all should belong to the scope of claims of the present invention.
Detailed description of the invention
The preparation method of embodiment 1 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, after suitably pulverizing, add 8 times amount 90% alcohol reflux 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, the NaOH solution adding 10% adjusts pH to 12 ~ 14, hold over night, centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Supernatant concentrating under reduced pressure becomes to proper volume, and add water appropriate to concentration of alcohol about 20%, the NaOH solution with 10% adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; The NaOH solution that supernatant continues with 10% adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with dehydrated alcohol, filter, collect filtrate, decompression recycling ethanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
The preparation method of embodiment 2 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, add 8 times amount 90% methanol eddies after suitably pulverizing and extract 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, the NaOH solution adding 10% adjusts pH to 12 ~ 14, hold over night, centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Supernatant concentrating under reduced pressure becomes to proper volume, and add water appropriate to methanol concentration about 20%, the NaOH solution with 10% adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; The NaOH solution that supernatant continues with 10% adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with absolute methanol, filter, collect filtrate, reclaim under reduced pressure methanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
The preparation method of embodiment 3 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, after suitably pulverizing, add 8 times amount 90% alcohol reflux 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, the KOH solution adding 10% adjusts pH to 12 ~ 14, hold over night, centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Supernatant concentrating under reduced pressure becomes to proper volume, and add water appropriate to concentration of alcohol about 20%, the KOH solution with 10% adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; The KOH solution that supernatant continues with 10% adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with dehydrated alcohol, filter, collect filtrate, decompression recycling ethanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
The preparation method of embodiment 4 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, after suitably pulverizing, add 8 times amount 90% alcohol reflux 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, add the Ca (OH) of 10%
2solution adjusts pH to 12 ~ 14, hold over night, and centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Supernatant concentrating under reduced pressure becomes to proper volume, adds water appropriate to concentration of alcohol about 20%, the Ca (OH) with 10%
2solution adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; Supernatant continues the Ca (OH) with 10%
2solution adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with dehydrated alcohol, filter, collect filtrate, decompression recycling ethanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
The preparation method of embodiment 5 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, after suitably pulverizing, add 8 times amount 30% alcohol reflux 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, the NaOH solution adding 10% adjusts pH to 12 ~ 14, hold over night, centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Add water appropriate to concentration of alcohol about 20%, the NaOH solution with 10% adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; The NaOH solution that supernatant continues with 10% adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with dehydrated alcohol, filter, collect filtrate, decompression recycling ethanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
The preparation method of embodiment 6 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, after suitably pulverizing, add 8 times amount 40% alcohol reflux 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, the NaOH solution adding 10% adjusts pH to 12 ~ 14, hold over night, centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Add water appropriate to concentration of alcohol about 20%, the NaOH solution with 10% adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; The NaOH solution that supernatant continues with 10% adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with dehydrated alcohol, filter, collect filtrate, decompression recycling ethanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
The preparation method of embodiment 7 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, after suitably pulverizing, add 8 times amount 50% alcohol reflux 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, the NaOH solution adding 10% adjusts pH to 12 ~ 14, hold over night, centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Supernatant concentrating under reduced pressure becomes to proper volume, and add water appropriate to concentration of alcohol about 20%, the NaOH solution with 10% adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; The NaOH solution that supernatant continues with 10% adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with dehydrated alcohol, filter, collect filtrate, decompression recycling ethanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
The preparation method of embodiment 8 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, after suitably pulverizing, add 8 times amount 60% alcohol reflux 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, the NaOH solution adding 10% adjusts pH to 12 ~ 14, hold over night, centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Supernatant concentrating under reduced pressure becomes to proper volume, and add water appropriate to concentration of alcohol about 20%, the NaOH solution with 10% adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; The NaOH solution that supernatant continues with 10% adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with dehydrated alcohol, filter, collect filtrate, decompression recycling ethanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
The preparation method of embodiment 9 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, after suitably pulverizing, add 8 times amount 70% alcohol reflux 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, the NaOH solution adding 10% adjusts pH to 12 ~ 14, hold over night, centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Supernatant concentrating under reduced pressure becomes to proper volume, and add water appropriate to concentration of alcohol about 20%, the NaOH solution with 10% adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; The NaOH solution that supernatant continues with 10% adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with dehydrated alcohol, filter, collect filtrate, decompression recycling ethanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
The preparation method of embodiment 10 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, after suitably pulverizing, add 8 times amount 80% alcohol reflux 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, the NaOH solution adding 10% adjusts pH to 12 ~ 14, hold over night, centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Supernatant concentrating under reduced pressure becomes to proper volume, and add water appropriate to concentration of alcohol about 20%, the NaOH solution with 10% adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; The NaOH solution that supernatant continues with 10% adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with dehydrated alcohol, filter, collect filtrate, decompression recycling ethanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
The preparation method of embodiment 11 radix sanguisorbae total saponin extract
Get Radix Sanguisorbae decoction pieces 1kg, after suitably pulverizing, add 8 times amount 95% alcohol reflux 2 times, each 1.5 hours, filter, merge extracted twice liquid, leave standstill after letting cool, the NaOH solution adding 10% adjusts pH to 12 ~ 14, hold over night, centrifugal going is precipitated, and filtrate sucking filtration obtains supernatant.Supernatant concentrating under reduced pressure becomes to proper volume, and add water appropriate to concentration of alcohol about 20%, the NaOH solution with 10% adjusts pH to 11 ~ 12, leaves standstill 12h, centrifugal collecting precipitation; The NaOH solution that supernatant continues with 10% adjusts pH to 12 ~ 13, leaves standstill 12h, centrifugal collecting precipitation; Merge both sides precipitation, be decompressed to dry in 70 DEG C, to reflux 45min with dehydrated alcohol, filter, collect filtrate, decompression recycling ethanol is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, to obtain final product.
Beneficial effect of the present invention is proved below by way of specific experiment.
The preparation method of experimental example 1 radix sanguisorbae total saponin extract of the present invention and the comparative experiments of patented method
1. experimental technique:
The preparation of 1.1 laboratory samples
Sample 1 is prepared according to embodiment 1.
According to following patent, (application number: 200910082470.6) method a, prepares sample 2.
According to following patent, (application number: 201210129134.4) method b, prepares sample 3.
Patented method a: get Radix Sanguisorbae decoction pieces 1kg, add 8 times amount 30% alcohol heating reflux and extract 1h, filtering residue extracts 1h with 6 times amount 30% alcohol heating reflux again, merging filtrate, add 20 times of water gaging dilutions, sodium hydroxide solution (3.5kg sodium hydroxide is dissolved in 24L water) is added under stirring, adjust pH is 14.2, leave standstill, centrifugal filtrate, by D101 type macroporous adsorbent resin, flow velocity is 1.5 column volumes/h, 4 column volume 30% ethanol elutions are used again after having gone up medicinal liquid, and then wash with water to effluent colourless, finally use 70% ethanol elution of 3 times of bed volumes, collect 70% ethanol stream fluid, vacuum concentration to have measured quantity of material separate out after, stop concentrated, leave standstill more than 2 hours, decompress filter, use 40% washing with alcohol, drain, take out filter cake, vacuum drying, pulverize, obtain.
Patented method b: get Radix Sanguisorbae decoction pieces 1kg, with 8 times amount 70% alcohol reflux 3 times, each 1.5h, 70 DEG C of concentrating under reduced pressure medicinal liquids are to appropriate (without ethanol taste), be transferred to separatory funnel, first with water saturated ether defatting 2 times, the ether that mother solution 70 DEG C of reclaim under reduced pressure are mixed into, then add water and be dissolved to, again with water saturated n-butanol extraction 2 times, extract 80 DEG C of concentrating under reduced pressure, add certain density ethanol solid content is dissolved, be transferred in beaker, add water appropriate, adjust pH to 10-14, place, centrifugal, take out precipitation, 70 DEG C of vacuum drying 2h, be transferred in tool plug triangular flask, add dehydrated alcohol supersound extraction 30 minutes, filter, filtrate 70 DEG C of decompression recycling ethanols are to just having solid content to separate out, be transferred in evaporating dish, 80 DEG C of water-baths volatilize ethanol, 70 DEG C of vacuum drying 12h, take out, grinding, obtain radix sanguisorbae total saponin.
The mensuration of 1.2 sanguisorbin I content:
Adopt the method for high performance liquid chromatography, its condition is SapphireC
18post (5 μm, 150 × 4.6mm) chromatographic column is separated, and is filler with octadecylsilane chemically bonded silica; With methanol-water (64:36) for mobile phase; Flow velocity 1.0mL/min, ELSD (evaporative light scattering detector) detect, drift tube temperature 85 DEG C, gas flow rate 2.8L/min.Theoretical cam curve is pressed sanguisorbin I peak and is calculated, and should be not less than 4000.
The preparation of reference substance solution in experiment: it is appropriate that precision takes sanguisorbin I reference substance, dissolves respectively with 50% ethanol and makes the solution of every 1mL containing 0.0565mg and 0.226mg, for subsequent use.
The preparation of need testing solution: get extractum (doing) 10g or 30, preparation, accurately weighed weight, porphyrize, get appropriate (being about equivalent to the weight of 10) totally 2 parts, accurately weighed, put in 150mL tool plug conical flask, precision adds 50% ethanol 20mL, shakes up, weighs, ultrasonic 10 minutes, put to room temperature, supply the weight of less loss with 50% ethanol, shake up, centrifugal, get supernatant, 0.22 μm of filter membrane filters, and gets subsequent filtrate as need testing solution.Then get reference substance solution and each 20 μ l of need testing solution respectively, measure by above-mentioned chromatographic condition, and press sanguisorbin I content in external standard two-point method calculation sample.
The mensuration of 1.3 sanguisorbin II content:
Adopt the method for high performance liquid chromatography, its condition is Yi Lite HypersilC
18chromatographic column (4.6mm × 200mm, 5 μm); Volume flow 1.2mL/min; Mobile phase A is methanol-acetonitrile (1: 1), and Mobile phase B is 0.5% phosphoric acid solution, gradient elution (0 ~ 10min, 45.0%A; 10 ~ 27min, 45.0%-60.0%A; 27 ~ 40min, 60.0%A; 40 ~ 45min, 60.0%-45.0%A); Determined wavelength (0 ~ 27min, λ
1=203nm; 27 ~ 35min, λ
2=289nm; 35 ~ 45min, λ
3=306nm); Sample size 10 μ L.Theoretical cam curve must not lower than 3000, and separating degree is all greater than 2.
The preparation of reference substance solution in experiment: it is appropriate that precision takes sanguisorbin-II reference substance, be placed in 5 20mL measuring bottles respectively, add 60% dissolve with methanol and be diluted to scale, again respectively accurate draw above-mentioned solution 10,5,2,10,5mL is placed in 50mL measuring bottle, add 60% methanol dilution to scale, shake up, obtain sanguisorbin-II reference substance solution of 0.0374mg/mL.
The preparation of need testing solution: get extractum (doing) appropriate, porphyrize, mixing, take about 1.0g, accurately weighed, precision adds 60% methanol 50mL, close plug, weighed quality, ultrasound wave (power 160W, frequency 50kHz) supersound process 30min, let cool, more weighed quality, the quality of less loss is supplied with 60% methanol, shake up, filter, get subsequent filtrate as need testing solution.Then accurate absorption reference substance solution and need testing solution, measure by above-mentioned chromatographic condition, and press sanguisorbin-II content in external standard two-point method calculation sample.
2. experimental result:
The comparision contents of table 1 radix sanguisorbae total saponin
3. experiment conclusion:
Experimental data shows, compared with two kinds of patented methods, the Ziyuglycoside I that the present invention prepares, Ziyuglycoside I I and radix sanguisorbae total saponin and yield are all higher, sufficient proof the inventive method substantially increases the content of radix sanguisorbae total saponin, Ziyuglycoside I and Ziyuglycoside I I, is obviously better than method disclosed in patent.
The pharmacological evaluation of experimental example 2 radix sanguisorbae total saponin extract of the present invention compares
1. experiment material, instrument
1.1 test medicine: sample 1, sample 2, the sample 3 of preparation in experimental example 1.
1.2 tool drugs: cyclophosphamide.
1.3 laboratory animals: KM mice: body weight 18.5 ~ 22.5g.
1.4 experimental apparatus: full-automatic blood cell analysis machine; BS-600L electronic balance: specification: 600g/0.1g, Shanghai Yousheng Balance Co., Ltd..
1.5 statistical method: carry out statistical analysis with SPSS17.0 software.Data are with mean ± standard deviation
represent, adopt one factor analysis of variance between group, carry out LSD inspection between the neat person's group of variance, heterogeneity of variance person carries out Tamhane ' sT2 and checks.
2. experimental technique
2.1 laboratory animal groupings and model preparation:
All Animal adaptabilities are divided at random by body weight after feeding 1 week: normal group, model group, sample 1 group: 15mgkg-1 dosage gavage sample 1 aqueous solution; Sample 2 groups: 15mgkg-1 gavage sample 2 aqueous solution; Sample 3 groups: 15mgkg-1 dosage gavage sample 3 aqueous solution; DIYU SHENGBAI PIAN group: 200mgkg-1 dosage gavage DIYU SHENGBAI PIAN aqueous solution, often organizes 10.Test the 1st day, except blank group, all the other respectively organize mice by 50mgkg-1 dosage intraperitoneal injection of cyclophosphamide normal saline solution, for three days on end, and naive mice lumbar injection equal-volume normal saline.
2.2 experiment grouping and administrations:
Each experimental group starts according to dosage from experiment on the 1st day, administering mode gives relative medicine, normal group and model group mouse stomach equal-volume pure water, continuous 7 days.
2.3 collections of specimens:
Test the 8th day, each experimental mice eye socket gets blood, collects to be measured with the 0.5mlEP pipe that EDTA anticoagulant is housed.
2.4 Testing index and method:
Peripheral hemogram detects: adopt full-automatic blood counting instrument to count each experimental mice peripheral blood leucocyte (WBC).
3. experimental result
Table 2 each experimental mice peripheral white blood cell amount
Note: compare with model group, * P<0.05, * * P<0.01; Compare with DIYU SHENGBAI PIAN group,
▲p<0.05,
▲ ▲p<0.01.
4. experiment conclusion
Table 2 shows, and sample 1 group of mouse peripheral blood leukocyte has remarkable rising (P<0.05) than model group; Even also high than positive group DIYU SHENGBAI PIAN group, show that the activity of radix sanguisorbae total saponin leukocyte increasing prepared by the present invention is best.
To sum up, compared with existing patented method, preparation method of the present invention significantly improves the content of radix sanguisorbae total saponin, Ziyuglycoside I and Ziyuglycoside I I, and drug effect is also significantly better than total saponins prepared by existing method.Preparation method technological operation of the present invention is easy, cost is low, efficiency is high, is suitable for industrialized great production.The activity of radix sanguisorbae total saponin extract leukocyte increasing prepared by the present invention is strong, and the clinical treatment for leukopenia provides a kind of new medicine.
Claims (11)
1. a radix sanguisorbae total saponin extract, is characterized in that: in this extract, the weight percentage of Ziyuglycoside I is: the weight percentage of 50%-97%, Ziyuglycoside I I is: 2%-10%.
2. extract according to claim 1, is characterized in that: in this extract, the weight percentage of Ziyuglycoside I is: the weight percentage of 92.34%, Ziyuglycoside I I is: 6.16%.
3. extract according to claim 1 and 2, is characterized in that: in this extract, total saponins weight percentage is 60-99%.
4. extract according to claim 3, is characterized in that: in this extract, total saponins weight percentage is 98.50%.
5. the extract according to claim 1-4 any one, is characterized in that: it is prepared by the following method:
A. get Radix Sanguisorbae medical material, add organic solvent extraction, extracting solution is for subsequent use;
B. get the extracting solution that step a is obtained, adjust pH to 12 ~ 14, leave standstill, centrifugal, sucking filtration obtains supernatant, supernatant concentrating under reduced pressure, for subsequent use;
C. get the obtained supernatant of step b and adjust pH to 11 ~ 12, leave standstill, centrifugal, collecting precipitation; Supernatant continues to adjust pH to 12 ~ 13, leaves standstill, centrifugal, collecting precipitation; Merge twice precipitation;
The precipitation of d. getting step c obtained adds organic solvent extraction, filters, and collect filtrate, concentrating filter liquor, drying, obtain radix sanguisorbae total saponin.
6. prepare the method for the extract described in claim 1-5 any one, it comprises the steps:
A. get Radix Sanguisorbae medical material, add organic solvent extraction, extracting solution is for subsequent use;
B. get the extracting solution that step a is obtained, the aqueous slkali adding 10% adjusts pH to 12 ~ 14, and leave standstill, centrifugal, sucking filtration obtains supernatant, supernatant concentrating under reduced pressure, for subsequent use;
C. get the aqueous slkali that supernatant that step b obtains adds 10% and adjust pH to 11 ~ 12, leave standstill, centrifugal, collecting precipitation; Supernatant continues aqueous slkali tune pH to 12 ~ 13 adding 10%, leaves standstill, centrifugal, collecting precipitation; Merge twice precipitation;
The precipitation of d. getting step c obtained adds organic solvent extraction, filters, and collect filtrate, concentrating filter liquor, drying, obtain radix sanguisorbae total saponin.
7. method according to claim 6, is characterized in that:
In step a, add the organic solvent reflux, extract, 2 times of Radix Sanguisorbae medical material 8 times amount, each 1.5 hours; Wherein, organic solvent is ethanol or methanol, and concentration of alcohol is 30% ~ 95%v/v;
In step b, the aqueous slkali that extracting solution adds 10% adjusts pH to 12 ~ 14, hold over night, and centrifugal, sucking filtration obtains supernatant, and supernatant is evaporated to proper volume, add water in right amount to concentration of alcohol be 20%v/v, for subsequent use; Wherein, described aqueous slkali is NaOH, KOH or Ca (OH)
2solution;
In step c, time of repose is 12h, collection be deposited in 70 DEG C of drying under reduced pressure;
In steps d, get the precipitation that step c is obtained, to reflux 45min with organic solvent, filter, collect filtrate, reclaim under reduced pressure organic solvent is separated out to there being solid content, volatilizes, and gained solid content drying under reduced pressure 12 hours, obtains radix sanguisorbae total saponin; Wherein, described organic solvent is dehydrated alcohol or absolute methanol.
8. method according to claim 7, is characterized in that:
In step a, described organic solvent is ethanol, and concentration of alcohol is 90%v/v;
In step b and step c, described aqueous slkali is NaOH solution;
In steps d, described organic solvent is dehydrated alcohol.
9. the radix sanguisorbae total saponin extract described in claim 1-5 any one is preparing the purposes had in the medicine of function of increasing leukocyte.
10. a pharmaceutical composition for leukocyte increasing, is characterized in that: it is active component by the radix sanguisorbae total saponin extract described in claim 1-5 any one, adds pharmaceutically acceptable adjuvant or complementary composition is prepared into pharmaceutically conventional preparation.
11. pharmaceutical compositions according to claim 10, is characterized in that: described preparation is oral formulations.
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