CN104458964B - The tracking detection method of Bulbus Fritillariae Pallidiflorae drug activity composition and drug activity composition thereof - Google Patents
The tracking detection method of Bulbus Fritillariae Pallidiflorae drug activity composition and drug activity composition thereof Download PDFInfo
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Abstract
The invention discloses tracking detection method and the drug activity composition thereof of a kind of Bulbus Fritillariae Pallidiflorae drug activity composition, belong to active constituents of medicine triage techniques field.The method following steps: 1) extraction of composition: extract the various compositions in Bulbus Fritillariae Pallidiflorae；2) multicomponent coordinates: is combined by various compositions obtained above, obtains at least one compositions；3) mensuration of Partition coefficients: measure the Partition coefficients of component in above-mentioned various composition and compositions respectively；4) tracking of active component: by Partition coefficients obtained above as activity rating index, carry out comparison and detection with standard oil water partition coefficients.The method both considered active component absorption in vivo, distribution situation, it is also contemplated that the situation of the interphase interaction of active constituents of medicine, was the tracking detection method of a kind of good drug activity composition.
Description
Technical field
The method that the present invention relates to the screening of a kind of active constituents of medicine, particularly relates to a kind of Bulbus Fritillariae Pallidiflorae drug effect and lives
The tracking detection method of property composition and drug activity composition thereof.
Background technology
Chinese medicine is the Chinese medicine product through clinical practice in thousand of years, and its curative effect is unquestionable, but owing to Chinese medicine becomes
The multiformity divided and the complexity of mechanism of action, bioactive components research always researcher grinds
The focus studied carefully and difficult point.
Up to now, bioactive components research mainly uses phytochemistry progressively to separate, pharmacology is lived
The pattern of property spike.This research mode achieves certain achievement, but, in research process it has also been found that
Many Chinese medicines or plant amedica are often that crude extract is effective, but along with the most isolated and purified, the activity of effective site
More and more weak, the most even can not get activated monomer component.This gives the research of bioactive components
Cause puzzlement greatly.The material base of this phenomenon explanation Chinese medicine onset is not that single component is to single
The effect of target spot, and jointly act on the result of organism often after multicomponent and interphase interaction thereof.Cause
This, multicomponent and interphase interaction thereof have important impact for the performance of herbal medicine efficacy, and by tradition
Chemical composition of Chinese materia medica is carried out being separately separated one by one, then the method carrying out drug effect tracking has certain limitation
Property.
Bulbus Fritillariae Pallidiflorae (Bulbus fritillariae Pallidiflorae) is liliaceous plant Fritillaria walujewii Regel or Fritillaria pallidiflora Schrenk
Dry bulb, be distributed mainly on Xinjiang Yili of China river valley.Owing to Fritillaria pallidiflora Schrenk yield is high, containing of alkaloid
Measuring of a relatively high in similar Bulbus Fritillariae Uninbracteatae, premunition is strong and price of medicinal material is cheap, and particularly it eliminates the phlegm and antitussive etc.
Physiologically active relatively Bulbus Fritillariae Cirrhosae is slightly strong, the most extremely concern of people.
And the present inventor is just found that analogue during the study of active components of Bulbus Fritillariae Pallidiflorae, her shellfish of Chinese medicine
Mother is demonstrated by the strongest cough-relieving curative effect, but by the principal monomer material imperialine of plant isolated but than her
The activity of Bulbus Fritillariae Uninbracteatae total extract is weak a lot, illustrates traditional chemical composition of Chinese materia medica to carry out being separately separated one by one,
The method carrying out drug effect tracking active component again is not appropriate for Bulbus Fritillariae Pallidiflorae.
Summary of the invention
Based on this, it is an object of the invention to overcome the defect of prior art, it is provided that a kind of Bulbus Fritillariae Pallidiflorae drug effect is lived
The tracking detection method of property composition, this drug effect tracking detection method can be good at finding in Bulbus Fritillariae Pallidiflorae and has relatively
The active component of strong cough suppressing effect.
For achieving the above object, the present invention takes techniques below scheme:
The tracking detection method of a kind of Bulbus Fritillariae Pallidiflorae drug activity composition, comprises the following steps:
1) extraction of composition: extract the various compositions in Bulbus Fritillariae Pallidiflorae, described composition be one matter monomer and
/ or the mixture of at least two material composition；
2) multicomponent coordinates: is combined by various compositions obtained above, obtains at least one compositions；
3) mensuration of Partition coefficients: measure the profit of component in above-mentioned various composition and compositions respectively and divide
Cloth coefficient；
4) tracking of active component: by Partition coefficients obtained above as activity rating index, with mark
Quasi-Partition coefficients carries out comparison and detection.The composition or the compositions that meet predetermined oil water partition coefficients standard are
For Bulbus Fritillariae Pallidiflorae drug activity composition.
The tracking detection method of the Bulbus Fritillariae Pallidiflorae drug activity composition of the present invention, with Partition coefficients as activity
Evaluation index, had both considered active component absorption in vivo, distribution situation, again can be by single component or many
Planting composition totally as investigating object, the combination that can also be between heterogeneity is investigated, it is contemplated that
The situation of the interphase interaction of active constituents of medicine, is the trace detection side of a kind of good drug activity composition
Method.
Wherein in an embodiment, the assay method of described Partition coefficients is: by various compositions or group
Compound adds in the mixed solution of oil phase and aqueous phase so that it is dissolve and after between oil phase and aqueous phase, distribution is stable,
Measure component concentration in oil phase and the concentration in aqueous phase in this composition or compositions, calculate according to the following formula
Its Partition coefficients:
Partition coefficients=lg (CO/CW)
Wherein: COFor the concentration in oil phase, CWFor the concentration in aqueous phase.
Wherein in an embodiment, described oil phase is water saturated n-octyl alcohol, and described aqueous phase is that n-octyl alcohol is satisfied
The water of sum.With water saturated n-octyl alcohol as oil phase, due to the solubility parameter that n-butyl alcohol is overall with biomembrane
Close, therefore, it is possible to preferably simulate situation in organism, it is thus achieved that tracking activity effect more accurately.
Wherein in an embodiment, described standard oil water partition coefficients is-1 to 2.When Partition coefficients exists
In the range of this, this composition or compositions have preferable activity.
Wherein in an embodiment, described various compositions are: imperialine, Bulbus Fritillariae Pallidiflorae total flavones.
Wherein in an embodiment, the detection method of described imperialine concentration is: draw oil phase or aqueous phase is molten
Liquid, with the imperialine content in this solution of high effective liquid chromatography for measuring, the survey of described high performance liquid chromatography
Fixed condition is as follows:
Fixing phase: C18 reverse phase silica gel post；
Flowing phase: acetonitrile: the volume ratio of 0.005-0.02% triethylamine aqueous solution is 55-65: 35-45, flow velocity
For 0.8-1.2ml/min；
ELSD detector: drift tube temperature is 65-75 DEG C, nitrogen flow rate is 1.3-1.7L min-1。
The invention also discloses a kind of tracking detection method using above-mentioned Bulbus Fritillariae Pallidiflorae drug activity composition to obtain
Bulbus Fritillariae Pallidiflorae drug activity composition, combine the compositions obtained for imperialine and Bulbus Fritillariae Pallidiflorae total flavones.
The Bulbus Fritillariae Pallidiflorae drug activity composition of the present invention, is to use above-mentioned referring to Partition coefficients as evaluation
The active component that mark obtains, the two cooperation, its cough suppressing effect is than alone imperialine and alone Bulbus Fritillariae Pallidiflorae total flavones
The direct superposition of effect also to be got well, and has collaborative effect.
Wherein in an embodiment, described Bulbus Fritillariae Pallidiflorae total flavones is prepared by the following method and obtains: by her shellfish
Female pulverizing, adding percent by volume is the ethanol of 80-99%, and reflux, extract, obtains filtrate after filtration, reclaim
Filtrate obtains thick extractum, adds the distilled water thick extractum of suspension, then adjust pH value extremely with HCl in this thick extractum
2-4, stands 8-12 hour, is extracted with ethyl acetate, and i.e. obtains Bulbus Fritillariae Pallidiflorae total flavones after extract recycling design.
Wherein in an embodiment, the weight part ratio of described imperialine and Bulbus Fritillariae Pallidiflorae total flavones is 1:1-7.Will
Imperialine matches with this weight part ratio with Bulbus Fritillariae Pallidiflorae total flavones, has preferable cough suppressing effect.
Compared with prior art, the method have the advantages that
The tracking detection method of the Bulbus Fritillariae Pallidiflorae drug activity composition of the present invention, measure in Bulbus Fritillariae Pallidiflorae respectively is various
Composition and the Partition coefficients of compositions, with its Partition coefficients as activity rating index, to her shellfish
Active component in mother is tracked.Both considered active component absorption in vivo, distribution situation, examined again
Consider the situation of the interphase interaction having arrived active constituents of medicine, be the tracking of a kind of good drug activity composition
Detection method.Particularly, the method can will extract the Multiple components obtained, and multicomponent in Bulbus Fritillariae Pallidiflorae
Between combination detect as an entirety, contingent interaction between ingredient,
More accurately completely drug activity composition can be tracked detection.The method is as a kind of brand-new
Research Thinking, it is possible to play an important role in the research of herbal medicine efficacy.
The Bulbus Fritillariae Pallidiflorae drug activity composition of the present invention, is to use above-mentioned referring to Partition coefficients as evaluation
The active component that mark obtains, the two cooperation, its cough suppressing effect is than alone imperialine and alone Bulbus Fritillariae Pallidiflorae total flavones
The superposition of effect also to be got well, and has synergy.
Accompanying drawing explanation
Fig. 1 be before and after imperialine mixes with Bulbus Fritillariae Pallidiflorae total flavones in Caco-2 cell monolayer model AP to BL
The absorption curve in direction.
Detailed description of the invention
Describe the present invention in detail below in conjunction with the drawings and specific embodiments, but the present invention is not caused any
Limit.
Instrument used in following example and reagent are as follows:
Agilent1200 high performance liquid chromatograph, containing ELSD detector, Agilent company of the U.S..
YLS-8A lures to cough to draw and breathes heavily instrument, medical science equipment station, Shandong Province product.
Reference substance imperialine by this laboratory separate obtain (used UV, MS,1H and13The light such as C NMR
Spectral method determines its structure), its purity >=99%.
Bulbus Fritillariae Pallidiflorae total flavones (wherein the mass fraction of flavone is 85%) is made by oneself by this laboratory.
N-octyl alcohol, triethylamine and strong aqua ammonia (25%) are analytical pure, purchased from Guangzhou chemical reagent work.
Kunming mouse, male, weight 18～22g, medical faunae center, Guangdong Province provide, qualified
Card SCXK (Guangdong) 2008-00002.
Embodiment
The tracking detection method of a kind of Bulbus Fritillariae Pallidiflorae drug activity composition, comprises the following steps:
1) extraction of composition: extract the various compositions in Bulbus Fritillariae Pallidiflorae, obtain imperialine and Bulbus Fritillariae Pallidiflorae total flavones.
1, the preparation method of imperialine: pulverized by Bulbus Fritillariae Pallidiflorae, adding percent by volume is the ethanol of 95%, returns
Stream extracts 3 hours, extracts 3 times, sucking filtration, merging filtrate, decompression recycling ethanol, obtains thick extractum, past
Adding the distilled water thick extractum of suspension in this thick extractum, then adjust pH value to 10 with 1mol/L NaOH, alkali liquor is used
Chloroform extracts 3 times, and combining extraction liquid is vacuum dried and i.e. obtains Bulbus Fritillariae Pallidiflorae total alkaloids, through silica gel column chromatography essence
Prepare imperialine (use UV, MS,1H and13This product is characterized by the spectrographic techniques such as C NMR,
Determine that it is imperialine).
2, the preparation method of Bulbus Fritillariae Pallidiflorae total flavones: pulverized by Bulbus Fritillariae Pallidiflorae, adding percent by volume is the second of 95%
Alcohol, reflux, extract, 3 hours, extract 3 times, sucking filtration, merging filtrate, decompression recycling ethanol, obtain thick leaching
Cream, adds the distilled water thick extractum of suspension in this thick extractum, then with 2mol/L HCl tune pH value to 3, quiet
Putting 8-12 hour (overnight), repeatedly extract by ethyl acetate, extract merges final vacuum and concentrates, i.e. get Yi Bei
Female total flavones.
2) multicomponent coordinates: imperialine obtained above and Bulbus Fritillariae Pallidiflorae total flavones is combined, obtains compositions.
3) mensuration of Partition coefficients: measure the imperialine of above-mentioned individualism and the most yellow with Bulbus Fritillariae Pallidiflorae respectively
The Partition coefficients of imperialine after ketone mixing.
1, the preparation of reagent.
N-octyl alcohol that aqueous solution is saturated and the saturated aqueous solution of n-octyl alcohol: after n-octyl alcohol and water are mixed, shaking
Swing vibration 24h on device so that it is the most saturated, after stratification, two-phase laminated flow, save backup respectively.
Partition coefficients laboratory sample is prepared: to weigh imperialine, imperialine the most yellow with Bulbus Fritillariae Pallidiflorae for precision respectively
The mixture of ketone as in beaker, dissolves with n-octyl alcohol saturation water respectively, obtain certain density imperialine,
Imperialine and the aqueous phase solution of Bulbus Fritillariae Pallidiflorae total flavones mixture, precision pipettes above-mentioned 2 groups of aqueous phase solution 3mL respectively
It is placed in 10mL centrifuge tube, the accurate water saturated n-octyl alcohol of 3mL that adds, mixing, seals pipe with sealed membrane
Mouthful, centrifuge tube is placed in water bath with thermostatic control agitator vibration, temperature is 37 DEG C, and frequency of oscillation is 100r/min,
Operation repetitive 3 parts.Take out centrifuge tube, 4500r/min, centrifugal 10min after 24h, make two phase stratification, point
Not taking oil phase (0) and aqueous phase (W), with methanol dilution to debita spissitudo, 0.45 μm microporous filter membrane is standby after filtering.
2, the mensuration of Partition coefficients.
(1) chromatographic condition.
Chromatographic column: Phenomenex Luna C18 (250mm × 4.6mm, 5 μm).
Flowing phase: acetonitrile-0.01% triethylamine aqueous solution (60: 40), flow velocity: 1.0mL min-1。
Detector parameters: drift tube temperature: 70 DEG C, nitrogen flow rate: 1.5L min-1。
(2) standard curve: accurate absorption imperialine reference substance storing solution 0.1,0.2,0.4,0.8 and 1.0mL,
With methanol dilution and be settled to 5mL, prepared concentration is respectively 20,40,80,160 and 200 μ g mL-1
Reference substance solution, respectively sample introduction 20 μ L (n=3), record peak area, with imperialine peak area to imperialine concentration
Returning, obtain both standard curves: regression equation is Y=8.26 × 132X-30.56, r=0.9996, wherein X is west
The concentration of Bei Su, Y is the peak area of imperialine, and the range of linearity is 20-200 μ g mL-1。
(3) measure: measure above-mentioned prepared Partition coefficients laboratory sample with above-mentioned chromatographic condition, and press
Its Partition coefficients (lgPow) is calculated according to following formula:
Partition coefficients=lg (CO/CW)
Wherein: COFor the concentration in oil phase, CWFor the concentration in aqueous phase.
3, result.
Measurement result is as shown in the table.
Table 1 Partition coefficients measurement result
| Group | Imperialine (individually) | Imperialine (after mixing with Bulbus Fritillariae Pallidiflorae total flavones) |
| Pow | 110.6 | 58.2 |
| lgPow | 2.04 | 1.74 |
Above-mentioned Pow=CO/CW。
Pass through the above results, it can be seen that after being mixed with Bulbus Fritillariae Pallidiflorae total flavones by imperialine, its oil and water zonation
Coefficient lgPow there occurs change, is reduced to 1.74 by 2.04.
4) tracking of active component: by Partition coefficients obtained above as evaluation index, with standard oil
Water partition coefficients (-1 to 2) contrasts, and the imperialine of conformance with standard Partition coefficients is the most yellow with Bulbus Fritillariae Pallidiflorae
The compositions of ketone is Bulbus Fritillariae Pallidiflorae drug activity composition.
In order to the above results is verified, also carry out the following test of pesticide effectiveness.
One, imperialine is tested at Caco-2 single cell model permeability
1, cell is cultivated
Caco-2 cell is incubated at 5%CO2, in 37 DEG C of cell culture incubators of relative humidity 95%.Caco-2
Support in the DMEM culture medium containing 10%FBS, 1%NEAA, 100U/mL Pen .-Strep,
Within every 2 days, change liquid once, cultivate 15-20 days, detect the cross-film resistance (TEER) of cell monolayer with resistance instrument,
Resistance value >=400 Ω cm2, meet requirement of experiment.
2, dosing experiment
Successful for cultivation Caco-2 cell monolayer is placed in Transwell culture plate, with 37 DEG C of preheatings
PBS solution cleans cell monolayer film 2 times, puts and hatches 30min in incubator.Enteric cavity side is separately added into PBS
The hybrid medicine solution of imperialine, imperialine and the Bulbus Fritillariae Pallidiflorae total flavones of solution preparation, substrate side adds PBS
Solution, parallel 3 parts of every kind of drug solution.Hatch in being placed in 37 DEG C of cell culture incubators, respectively at 30,60,
90, the 120 and 150min solution 100 μ L taking substrate side, uses above-mentioned liquid phase by each group sample collected
Chromatography determination.
3, data process
The diluting effect produced for fluid infusion after eliminating every sub-sampling, uses formula 1 penetrating to the accumulation of medicine
Amount is corrected.
Wherein An is the measured value of the n-th penetrating amount of sample；Vsn is the sampling volume of the n-th sample；VR
Volume for acceptance pool.
Utilize the penetrating amount of accumulation after correcting, calculate apparent permeability coefficients P according to the following formulaapp。
In formula: dQ/dt: infiltration rate (μ g/s), the dose i.e. passed through within the dt time period；A: cell
Monolayer surface amasss (cm2), i.e. in membrane area, this experiment, A is 1.12cm2；C0: it is administered side (enteric cavity side)
The initial concentration (μ g/ml) of medicine.
Imperialine mix with Bulbus Fritillariae Pallidiflorae total flavones before and after total absorptivity and apparent permeability coefficients see table and Fig. 1.
Absorbance before and after the mixing of table 2 imperialine and apparent permeability coefficients (n=3)
**Represent through statistical test, p < 0.01
From in the above results, after mixing with Bulbus Fritillariae Pallidiflorae total flavones, imperialine absorbance and apparent
Infiltration coefficient compares with time independent, all dramatically increases (p < 0.01).
Fig. 1 be before and after imperialine mixes with Bulbus Fritillariae Pallidiflorae total flavones in Caco-2 cell monolayer model AP (intestinal
Side, chamber) to the absorption curve in BL (substrate side) direction, understand from this curve, mix with Bulbus Fritillariae Pallidiflorae total flavones
After conjunction, imperialine starts just to show higher absorbance from experiment.These results suggest that total with Bulbus Fritillariae Pallidiflorae
After flavone mixing, imperialine absorption in intestinal dramatically increases, and the two has collaborative effect.
Two, ammonia draws and coughs experiment
1, animal is administered and draws and coughs
Take Kunming mouse 40, be randomly divided into blank group, imperialine group, Bulbus Fritillariae Pallidiflorae total flavones group and western shellfish
Element adds Bulbus Fritillariae Pallidiflorae flavonid composition group, often group 10, labelling.Before experiment, fasting 8 hours, presses respectively
Table 3 dosage gastric infusion, every day 1 time, for three days on end, is administered in last and within latter 1 hour, starts experiment, will
Mice is placed in draw and coughs in instrument, adds 25% strong aqua ammonia 1ml in atomizing cup, is atomized 15s (atomization rates
0.15ml/min), the cough number in counting 3min.
2, result
Cough number of times and the antitussive rate of each group are shown in Table 3, carry out statistical analysis with SPSS 12.0 software, test number
X ± s represents according to this, compares and check with t between group.P ＜ 0.05 is that difference is statistically significant.
Table 3 ammonia draws coughs test dose and result
* compares p < 0.01 with blank group；* p < 0.05 is compared with blank group；# compares p < 0.05 with imperialine group
It can be seen that individually dosed group of imperialine in from the above, cough number of times is variant relative to blank group,
Illustrate that imperialine has certain antitussive effect, but its antitussive rate is only 25.8%；And Bulbus Fritillariae Pallidiflorae total flavones does not show
Antitussive effect is shown.After imperialine mixes with Bulbus Fritillariae Pallidiflorae total flavones, the cough number of times of compositions group substantially reduces,
Antitussive rate significantly improves, and compares with blank group, and there were significant differences, and compositions group compares with imperialine group, also
There is statistical difference.
Above-mentioned experiment demonstrates when imperialine and Bulbus Fritillariae Pallidiflorae total flavones being share, and said composition shows higher
Drug effect, demonstrates the effectiveness of the tracking detection method of the Bulbus Fritillariae Pallidiflorae drug activity composition of the present invention, also confirms that
The Research Thinking affecting Chinese medicine entirety drug effect based on multicomponent Partition coefficients change that the present invention proposes
Feasibility.This is a brand-new Research Thinking, it is possible to between compositions various in medicine and various composition
Combination after cooperating is studied, and the method is used for the research of herbal medicine efficacy, necessarily can explain more
Share rear drug effect or phenomenon that toxicity changes, this Research Thinking can be in the research of herbal medicine efficacy more
Play an important role.
Embodiment described above only have expressed the several embodiments of the present invention, and it describes more concrete and detailed,
But therefore can not be interpreted as the restriction to the scope of the claims of the present invention.It should be pointed out that, for this area
Those of ordinary skill for, without departing from the inventive concept of the premise, it is also possible to make some deformation and
Improving, these broadly fall into protection scope of the present invention.Therefore, the protection domain of patent of the present invention should be with appended
Claim is as the criterion.
Claims (4)
1. the tracking detection method of a Bulbus Fritillariae Pallidiflorae drug activity composition, it is characterised in that comprise the following steps:
1) extraction of composition: extracting the various compositions in Bulbus Fritillariae Pallidiflorae, described composition is imperialine and/or Bulbus Fritillariae Pallidiflorae total flavones；
2) multicomponent coordinates: is combined by various compositions obtained above, obtains at least one compositions；
3) mensuration of Partition coefficients: measure the Partition coefficients of component in above-mentioned various composition and compositions respectively；Specific as follows:
(1) chromatographic condition；
Chromatographic column: Phenomenex Luna C18, its specification is 250mm × 4.6mm, 5 μm；
Flowing phase: acetonitrile-0.01% triethylamine aqueous solution, its ratio is 60: 40, flow velocity: 1.0mL min-1；
Elsd detector detector, parameter is: drift tube temperature: 70 DEG C, nitrogen flow rate: 1.5L min-1；
(2) standard curve: accurate draw imperialine reference substance storing solution 0.1,0.2,0.4,0.8 and 1.0mL, with methanol dilution and be settled to 5mL, prepared concentration is respectively 20,40,80,160 and 200 μ g mL-1Reference substance solution, sample introduction 20 μ L respectively, repeat sample introduction 3 times, record peak area, with imperialine peak area to imperialine concentration returns, obtains both standard curves: regression equation is Y=8.26 × 132X-30.56, r=0.9996, wherein X is the concentration of imperialine, and Y is the peak area of imperialine, and the range of linearity is 20-200 μ g mL-1；
(3) measure: measure above-mentioned prepared Partition coefficients laboratory sample with above-mentioned chromatographic condition, and calculate its Partition coefficients, i.e. lgPow according to the following formula:
Partition coefficients=lg (CO/CW)
Wherein: COFor the concentration in oil phase, CWFor the concentration in aqueous phase；
4) tracking of active component: by Partition coefficients obtained above as evaluation index, carrying out comparison and detection with standard oil water partition coefficients, the imperialine of conformance with standard Partition coefficients is Bulbus Fritillariae Pallidiflorae drug activity composition with the compositions of Bulbus Fritillariae Pallidiflorae total flavones.
The tracking detection method of Bulbus Fritillariae Pallidiflorae drug activity composition the most according to claim 1, it is characterised in that described oil phase is water saturated n-octyl alcohol, described aqueous phase is the water that n-octyl alcohol is saturated.
3. use the Bulbus Fritillariae Pallidiflorae drug activity composition that the tracking detection method of the Bulbus Fritillariae Pallidiflorae drug activity composition described in any one of claim 1-2 obtains, it is characterized in that, this Bulbus Fritillariae Pallidiflorae drug activity composition is that imperialine and Bulbus Fritillariae Pallidiflorae total flavones combine the compositions obtained, and described imperialine is 1:1-7 with the weight part ratio of Bulbus Fritillariae Pallidiflorae total flavones.
Bulbus Fritillariae Pallidiflorae drug activity composition the most according to claim 3, it is characterized in that, described Bulbus Fritillariae Pallidiflorae total flavones is prepared by the following method and obtains: pulverized by Bulbus Fritillariae Pallidiflorae, and adding percent by volume is the ethanol of 80-99%, reflux, extract, obtain filtrate after filtration, reclaim filtrate and obtain thick extractum, in this thick extractum, add the distilled water thick extractum of suspension, again with HCl tune pH value to 2-4, stand 8-12 hour, be extracted with ethyl acetate, after extract recycling design, i.e. obtain Bulbus Fritillariae Pallidiflorae total flavones.
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