CN103439425A - Screening method of active pharmaceutical ingredient - Google Patents
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Abstract
The invention discloses a screening method of active pharmaceutical ingredient. The screening method comprises the following steps: 1) primarily determining the active pharmaceutical ingredient: 1.1) obtaining the pathological pharmacodynamic indexes of a pathological body; 1.2) obtaining the maximal pharmacodynamic point moment and the minimal pharmacodynamic point moment of a medicament with active pharmaceutical ingredient to be determined; 1.3) obtaining the whole blood and/or body fluid of the pathological body; 1.4) creating full-fingerprints; 1.5) comparing the full-fingerprints, wherein the part having the maximal contrast in the full-fingerprints is the primarily determined active pharmaceutical ingredient, and the part having the maximal contrast is the part having the maximal chromatographic peak change in the full-fingerprint of the maximal pharmacodynamic point moment and the full-fingerprint of the minimal pharmacodynamic point moment; 2) obtaining the primarily determined active pharmaceutical ingredient; and 3) detecting, screening and determining the active pharmaceutical ingredient. The screening method of active pharmaceutical ingredient can be used for accurately and quickly determining the active ingredient in a traditional Chinese medicine.
Description
Technical field
The invention belongs to medical domain, relate to a kind of screening technique of active constituents of medicine, relate in particular to and a kind ofly take drug effect and show and take the active constituents of medicine screening technique that micro emulsion is separating medium as leading.
Background technology
Traditional Chinese medicine has a long history and significant curative effect, but the basic research of Chinese medicine is weaker, and the chemical composition of most Chinese medicines is not yet touched, and study of active components has no way of carrying out especially.Wherein restricting Chinese medicine is to lack effective active ingredient of Chinese herbs research method in the main cause of international market developing and competition.The research method of generally using now is extract the Chinese medicine single component or extract a certain cut, then carry out pharmacological evaluation and determine its active component, research aspect and the degree of depth of this method are often limited to according to researcher's cognition, also greatly reduce the probability of success simultaneously.
Most representative in the current research strategy to the Chinese medicine active ingredient screening and hypothesis is full composition opinion and active component opinion.Full composition opinion thinks that why traditional Chinese medicine can produce curative effect is that wherein all chemical compositions all work, if any composition is removed to the organic conception of all running counter to the traditional Chinese medical science.The understanding of this theory is confined to " organic conception ", active component developing thought in Chinese medicine is narrowed down, active component opinion has two kinds of viewpoints: 1. serum pharmacological: within 1988, Japanese scholars Tian Daizhen is once having proposed serum pharmacological, this theory is based on the human research, if and think that medicine plays therapeutic action and must can enter blood, so these compositions are likely just real active components, this theory has been dwindled the scope of finding active ingredient of Chinese herbs greatly, but lacks the method for determining active component from Contained Serum; 2. traditional Natural Medicine Chemistry method: according to clinical efficacy, in vitro Chinese medicine compound prescription being extracted, separate, identified of " specific aim " arranged, thereby obtain its active component, but due to its can not rediscover internal milieu, and be confined to people's cognitive range, so the active component obtained is difficult to realize whole effects of former Chinese medicine compound prescription, even occur that drug effect and attending scope disconnect, thereby cause the phenomenons such as active stronger composition is lost.Previously the bibliographical information active component all is transported to target organ by blood, if Chinese medicine compound prescription can produce clear and definite curative effect, may be its contained active component be transported to target organ and with receptors bind generation effect; If after an effective Chinese medicine compound prescription medication, drug effect fades away, may be that the active component that is transported to target organ disappears drug effect because metabolism causes concentration to descend.
Be not difficult to find, the particular problem of restriction active ingredient of Chinese herbs screening is as follows: after 1. medicine enters in body, collected body fluid example need pass through pre-treatment mostly, and very likely attend to one thing and lose sight of another in pre-treatment: such as hydrophilic compounds and lipophilic compound, can only take into account a kind ofly, acid compound, alkali compounds and neutral compound can not be present in body fluid example simultaneously.2. medicine can be transformed into material of different nature according to the difference of interior environment after entering in body, or acid or alkali or neutrality.Prior art causes the kind of separating substances parameter limited, is difficult to these materials in biological fluid samples are detected simultaneously and effectively separates, and becomes the bottleneck problem of active ingredient of Chinese herbs research.Only have application new method and new technology, strengthen active ingredient of Chinese herbs research, could accelerate modernization of Chinese medicine paces.
Summary of the invention
In order to solve the above-mentioned technical matters existed in background technology, the invention provides a kind of active constituents of medicine screening technique of can be accurately and determining rapidly active component in Chinese medicine.
Technical solution of the present invention is: the invention provides a kind of active constituents of medicine screening technique, its special character is: said method comprising the steps of:
1) tentatively determine active constituents of medicine:
1.1) obtain the pathology pharmacodynamics index of pathology body;
1.2) according to step 1.1) the pathology pharmacodynamics index of resulting pathology body obtains moment of maximum drug effect point of medicine of active constituents of medicine to be determined and the moment of minimum drug effect point;
1.3) obtain respectively whole blood and/or the body fluid of pathology body in the moment of moment of the maximum drug effect point of the medicine of active constituents of medicine to be determined and minimum drug effect point;
1.4) according to the Microemulsion liquid chromatography method to step 1.3) whole blood and/or the body fluid of resulting pathology body sets up respectively full finger-print;
1.5) according to drug effect differential serum chromatography by step 1.4) the full finger-print obtained of setting up contrasted, described full finger-print medium contrast maximum be preliminary definite active constituents of medicine; Described contrast maximum is that chromatographic peak changes the best part in the full finger-print of the full finger-print of maximum EDD point and minimum EDD point;
2) obtain preliminary definite active constituents of medicine;
3) detection, screening definite active constituents of medicine.
Above-mentioned steps 1.1) specific implementation is:
1.1.1) set up the pathological model of pathology body;
1.1.2) the pathology body is injected to the medicine of active constituents of medicine to be determined;
1.1.3) detect every pathology pharmacodynamics index of the pathology body of the medicine injected active constituents of medicine to be determined according to the conventional sense means.
Above-mentioned steps 2) specific implementation is:
Determining step 1) preliminary definite active constituents of medicine whether be standard items; If standard items, directly adopt standard items; If not, adopt routine techniques means preparation process 1) preliminary definite active constituents of medicine.
Above-mentioned steps 3) specific implementation is:
Determining step 2) obtaining preliminary definite active constituents of medicine and whether adopt standard items, if directly confirm step 2) institute's accepted standard product are exactly the active constituents of medicine of medicine to be determined; If not, according to the routine techniques means, detect, screen and determine the active constituents of medicine of medicine to be measured.
Said method also comprises after step 3):
4) the determined active constituents of medicine of step 3) is verified.
Above-mentioned steps 4) specific implementation is: step 3) is determined to the pharmacodynamic index of the active constituents of medicine obtained and the pharmacodynamic index of the medicine that the pathology body is injected are contrasted, if step 3) is determined the pharmacodynamic index of the active constituents of medicine obtained and is not less than 1/2 of the pharmacodynamic index of pathology body institute injectable drug, confirms that the resulting active constituents of medicine of step 3) is the active constituents of medicine to pathology body institute injectable drug; If step 3) determines that the pharmacodynamic index of the active constituents of medicine obtained is lower than 1/2 of the pharmacodynamic index to pathology body institute injectable drug, repeating step 1)-step 3), until the pharmacodynamic index of the active constituents of medicine finally obtained is not less than 1/2 of the pharmacodynamic index of pathology body institute injectable drug.
Advantage of the present invention is:
The invention provides and take drug effect and show and take the active constituents of medicine screening technique that micro emulsion is separating medium as leading, for catching the active component of medicine, catch and be accompanied by that drug effect disappears and the active component that disappears by the difference of the mass spectral:mass spectrographic maximum drug effect of serum thing and minimum drug effect, simultaneously farthest separation, purifying, to identify these active components be that contain in former prescription or metabolic product body.Then adopt the verification modelling verification, the continuous feedback of drug effect result is adjusted to the differential chromatographic condition, until confirm active component.After active component is determined, dynamically carry out the variation of qualitative, quantitative Chinese medicine activity in vivo composition with mass spectrum and nuclear magnetic resonance spectrum, and the corresponding pharmacodynamics index of analysis-by-synthesis, finally confirm the active component of Chinese medicine.Wherein, drug effect differential serum chromatography of the present invention can be followed the disease combination, side demonstrate,proves the holistic approach thinking associated, that medicine is corresponding with card, principle-method-recipe-medicines is unified, especially challenge can be oversimplified, utilize the multiple technologies means such as modern pharmacology, Pharmaceutical Analysis, contrast the difference in change of corresponding body serum chemistry number of components and determine active component, the method can compare science, screen the active component in Chinese medicine exactly.The advantage of Microemulsion liquid chromatography method of the present invention exactly can make up existing bottleneck problem in the active ingredient of Chinese herbs screening, its concrete advantage is as follows: 1.. micro emulsion solubilized albumen, so can realize the direct injection analysis body fluid example while with micro emulsion, doing mobile phase.Biological fluid samples not only true reappearance internal milieu, and comprised all active components.So the mobile phase that the micro emulsion of take is separating medium is just one of active ingredient of Chinese herbs screening three large difficult points: realize that the biological fluid samples direct injected provides theoretical support.2.. from the micromechanism of micro emulsion, the micro emulsion drop has the biomembranous orderly hydrophilic area of class and hydrophobic region structure, so to water wettability and lipophilic compound are all had to stronger dissolving power.This just makes the Microemulsion liquid chromatography method measure water wettability simultaneously and lipophilic compound may become.3.. in the micro emulsion mobile phase, solute and micro emulsion drop and fixedly between the phase adsorbed layer, exist the multi-acting forces such as hydrophobic, hydrogen bond, static, so it is more to regulate and control parameter, be applicable to the separation of complex mixture in Chinese medicine.This is just one of active ingredient of Chinese herbs screening three large difficult points: active constituents of medicine can reach effective separation simultaneously provides theoretical support.The Microemulsion liquid chromatography method is promoted the use of in various fields.Such as: declaring study on the stability in new drug is an important content, institute's method for building up must be can will synthesize or impurity degraded from the major component peak with the auxiliary material peak separately and separates well.And the Microemulsion liquid chromatography method with unique separation selectivity and widely applicability in study on the stability, be used widely, represented the high potentiality of Microemulsion liquid chromatography method in stability study.
For feasibility and the science of verifying method provided by the present invention, adopt early stage drug effect differential serum chromatography successfully to determine in the representative single medicinal materials promoting blood circulation and removing blood stasis such as safflower, the red sage root and improve hemorheological active component.Any theory that is applied to that the present invention is simple by active ingredient screening all is difficult to the breakthrough of realization to existing research, and Pharmaceutical Analysis and pharmacodynamic study " integration " should be got up.By Pharmaceutical Analysis and pharmacodynamics organic combination, initiated and take pharmacodynamics index as leading the drug effect differential serum chromatography of showing to determine active ingredient of Chinese herbs.The present invention can realize the body fluid example Direct Analysis, and has accomplished the existing interior environment of reducing activity composition as far as possible.
Embodiment
The invention provides a kind of active constituents of medicine screening technique, the method comprises the following steps:
1) tentatively determine active constituents of medicine:
1.1) obtain the pathology pharmacodynamics index of pathology body:
1.1.1) set up the pathological model of pathology body;
1.1.2) the pathology body is injected to the medicine of active constituents of medicine to be determined;
1.1.3) detect every pathology pharmacodynamics index of the pathology body of the medicine injected active constituents of medicine to be determined according to the conventional sense means.
1.2) according to step 1.1) the pathology pharmacodynamics index of resulting pathology body obtains moment of maximum drug effect point of medicine of active constituents of medicine to be determined and the moment of minimum drug effect point;
1.3) obtain respectively whole blood and/or the body fluid of pathology body in the moment of moment of the maximum drug effect point of the medicine of active constituents of medicine to be determined and minimum drug effect point;
1.4) according to the Microemulsion liquid chromatography method to step 1.3) whole blood and/or the body fluid of resulting pathology body sets up respectively full finger-print;
1.5) according to drug effect differential serum chromatography by step 1.4) the full finger-print obtained of setting up contrasted, full finger-print medium contrast maximum be preliminary definite active constituents of medicine; The contrast maximum is that chromatographic peak changes the best part in the full finger-print of the full finger-print of maximum EDD point and minimum EDD point.
2) obtain preliminary definite active constituents of medicine:
Determining step 1) preliminary definite active constituents of medicine whether be standard items; If standard items, directly adopt standard items; If not, adopt routine techniques means preparation process 1) preliminary definite active constituents of medicine.
3) detection, screening definite active constituents of medicine:
Determining step 2) obtaining preliminary definite active constituents of medicine and whether adopt standard items, if directly confirm step 2) institute's accepted standard product are exactly the active constituents of medicine of medicine to be determined; If not, according to the routine techniques means, detect, screen and determine the active constituents of medicine of medicine to be measured.
Method provided by the present invention also comprises after step 3):
4) the determined active constituents of medicine of step 3) is verified:
The pharmacodynamic index of step 3) being determined to the active constituents of medicine obtained is contrasted with the pharmacodynamic index of the medicine that the pathology body is injected, if step 3) is determined the pharmacodynamic index of the active constituents of medicine obtained and is not less than 1/2 of the pharmacodynamic index of pathology body institute injectable drug, confirms that the resulting active constituents of medicine of step 3) is the active constituents of medicine to pathology body institute injectable drug; If step 3) determines that the pharmacodynamic index of the active constituents of medicine obtained is lower than 1/2 of the pharmacodynamic index to pathology body institute injectable drug, repeating step 1)-step 3), until the pharmacodynamic index of the active constituents of medicine finally obtained is not less than 1/2 of the pharmacodynamic index of pathology body institute injectable drug.
Theoretical foundation of the present invention is: the key point of the modernization of Chinese medicine is to be difficult to clear and definite active component wherein, thereby can not further inquire into mechanism of action and the metabolic pathway of Chinese medicine.Illustrate the effective substance of Chinese medicine, the screening active component, what lack is effective investigative technique.At present, the difficult point of setting up active ingredient of Chinese herbs screening gordian technique is following 3 points: 1. realize the biological fluid samples direct injected, to guarantee to contain whole active constituents of medicine; 2. active constituents of medicine can reach effective separation simultaneously; 3. determine the active component in medicine.
For above difficult point, the present invention proposes a kind of brand-new research method first---drug effect differential serum chromatography.When Chinese medicine produces the pharmacodynamics variation in the verification body, in body serum, in the corresponding difference in change of chemical group umber, minimum zone is farthest being comprised to real active component.Based on this research method, successfully determined safflower, the red sage root, etc. improve hemorheological active component in representative single medicinal material promoting blood circulation and removing blood stasis.Result of study shows: drug effect differential serum chromatography can solve one of active ingredient of Chinese herbs screening three large difficult points: determine the active component in medicine.
The present invention is described in detail as example to take safflower and the red sage root: the main active in safflower is hydroxyl radical carthamin yellow carthamus A, and in the red sage root, main active is tanshin polyphenolic acid B.When being studied, DANHONG ZHUSHEYE finds: during according to DANHONG ZHUSHEYE prescription proportioning combination hydroxyl radical carthamin yellow carthamus A and tanshin polyphenolic acid B, and the curative effect of chemical drug combination group too late DANHONG ZHUSHEYE group (in Table 1).Also have other active components in this prompting DANHONG ZHUSHEYE, these active components may be to contain in the red sage root and safflower, or the red sage root and safflower are combined the novel substance that produces when chemical reaction occurs or even the metabolic product of body.The reason that these active components fail to be detected may have following 2 points: 1. in the pre-treatment step of sample, active component is lost.Although existing method is simplified pre-treatment as much as possible, due to the target that lacks gordian technique and still be difficult to realize the direct injection analysis body fluid example, active component may be lost in pre-treatment.2. existing triage techniques still can not be realized simultaneously the effective separation to complex mixture.Complicated component in Chinese medicine, water wettability, lipophilic compound exist simultaneously, and existing triage techniques does not possess the not only dissolving power of hydrophilic but also lipophilic; Have the compound of acid, alkalescence and the multiple character such as neutral in Chinese medicine, existing triage techniques can not be realized effective separation simultaneously.
Table 1 active component group is on the hemorheological impact of rat's blood stasis model
With normal group, compare,
Δp ﹤ 0.01; With the DANHONG ZHUSHEYE group, compare,
*p ﹤ 0.05.
How to solve other difficult points in the active ingredient of Chinese herbs screening? the separation selectivity that micro emulsion is unique with it and broad applicability can solve this two difficult points.
Micro emulsion is to be scattered in formed dispersion system of colloid in another kind of not miscible liquid (external phase) by a kind of liquid with the drop (disperse phase) of nanosized.Generally water, oil phase, surfactant, consisting of, is a kind of optical clear, thermodynamically stable system.Because micro emulsion has very high surface area, can be used as separating medium and be applied in high performance liquid chromatography, be i.e. Microemulsion liquid chromatography.First micro emulsion is introduced to high efficiency liquid phase as far back as nineteen ninety Berthod etc., along with updating, the separation selectivity that Microemulsion liquid chromatography is unique with it and widely applicability cause extensive attention.The chromatographic column that the method adopts is identical with conventional liquid phase chromatography with detecting device, and the reservation of solute is determined in the fixing partition equilibrium between phase, micro emulsion drop and water by it.Micro emulsion is a kind of stable dispersion system of colloid, has the following advantages: 1.. micro emulsion solubilized albumen, so can realize the target of direct injection analysis body fluid example while with micro emulsion, doing mobile phase.2.. the micro emulsion drop has the biomembranous orderly hydrophilic area of class and hydrophobic region, so water wettability and lipophilic compound are all had to stronger dissolving power, and solute and micro emulsion drop and fixedly between the phase adsorbed layer, exist the multi-acting forces such as hydrophobic, hydrogen bond, static, parameter can be regulated and controled more, so can be used for the separation of compound mixture.Existing drug effect serum differential chromatography can solve the problem of determining active component in medicine, and the advantage of micro emulsion can realize biological fluid samples direct injected and the purpose of separate complex potpourri simultaneously effectively, if the two can be organically combined and can set up a kind of gordian technique for the active constituents of medicine screening.
The present invention intends in earlier stage pioneering drug effect differential serum chromatography basis, the single medicinal material active component proportioning combination of finding when DANHONG ZHUSHEYE is developed can not replace the particular problems such as DANHONG ZHUSHEYE, set up a kind of novel gordian technique that is applicable to active ingredient of Chinese herbs screening, and inquire into active component in DANHONG ZHUSHEYE and the drug action of each composition with this.The present invention not only provides the foundation for the research and development of DANHONG ZHUSHEYE, also provides new thinking for the screening of active component in Chinese medicine simultaneously.
Result of study shows, drug effect differential serum chromatography can be followed the disease combination, side demonstrate,proves the holistic approach thinking associated, that medicine is corresponding with card, principle-method-recipe-medicines is unified, especially challenge can be oversimplified, utilize the multiple technologies means such as modern pharmacology, Pharmaceutical Analysis, contrast the difference in change of corresponding body serum chemistry number of components and determine active component, the method can compare science, screen the active component in Chinese medicine exactly.Early-stage Study has confirmed science and the feasibility of the method in determining active ingredient of Chinese herbs, and this is with regard to one of three large problems of having solved the active ingredient of Chinese herbs screening: determine the active component in Chinese medicine.This just lays a good foundation for the active ingredient of Chinese herbs screening gordian technique of setting up that the present invention intends studying.
In the treatment of traditional Chinese medicine to the cardiovascular and cerebrovascular stasis syndrome, play an important role, wherein definite with the result for the treatment of of DANHONG ZHUSHEYE again.DANHONG ZHUSHEYE is prepared from through strict aqueous extraction-alcohol precipitation technology by the red sage root and safflower two flavor medicinal materials, and wherein red sage root master cold in nature is heavy, safflower main liter warm in nature, and one falls one liter, and the effect of playing altogether removing blood stasis for promoting tissue regeneration is eliminating evil and just do not hinder.The DANHONG ZHUSHEYE annual sales amount has reached 1,700,000,000 yuan at present, and the accumulative total number of users reaches more than 5,000 ten thousand.But DANHONG ZHUSHEYE is Chinese medicine preparation, also may there be other materials that can cause the body spinoff in the ingredient complexity except the active component that really plays in vivo therapeutic action, and this is also that DANHONG ZHUSHEYE is at the clinical reason place that has spinoff.If the maximum retention activity composition of energy is removed the composition that causes spinoff, can make the curative effect of DANHONG ZHUSHEYE more definite, greatly reduce clinical side effects.In conjunction with DANHONG ZHUSHEYE, be that water extract-alcohol precipitation is prepared from, active component with drug effect serum differential chromatography to the single medicinal material material is studied, and proves that the active component that plays function promoting blood circulation and removing blood stasis in the red sage root, safflower is respectively tanshin polyphenolic acid B and hydroxyl radical carthamin yellow carthamus A.But by after tanshin polyphenolic acid B and hydroxyl radical carthamin yellow carthamus A mixing, its drug effect is but not as good as DANHONG ZHUSHEYE (in Table 1) according to DANHONG ZHUSHEYE prescription proportioning.This prompting: also have other active components in DANHONG ZHUSHEYE, these active components may be to contain in the crude drug red sage root and safflower, also or the red sage root and safflower combine the novel substance that produces when chemical reaction occurs or even the metabolic product of body.Realize maximum retention activity composition if want and remove other compositions that cause spinoff, at first will determine the active component in DANHONG ZHUSHEYE comprehensively.
The Microemulsion liquid chromatography method has been carried out to feasibility study, it is investigated index and has comprised the indexs such as post pressure, peak shape, degree of separation and appearance time, experimental result shows: common stratographic analysis post be take micro emulsion during as mobile phase, and that its post is pressed is stable, peak shape is good, degree of separation is good, appearance time is desirable.This prompting, the Microemulsion liquid chromatography method is not only theoretical feasible, and practical operation practical (in Table 2).
Two kinds of micro emulsion mobile phase high performance liquid chromatography parameters of table 2 are investigated result
The advantage of Microemulsion liquid chromatography method exactly can make up existing bottleneck problem in the active ingredient of Chinese herbs screening, and its concrete advantage is as follows: 1.. micro emulsion solubilized albumen, so can realize the direct injection analysis body fluid example while with micro emulsion, doing mobile phase.Biological fluid samples not only true reappearance internal milieu, and comprised all active components.So the mobile phase that the micro emulsion of take is separating medium is just one of active ingredient of Chinese herbs screening three large difficult points: realize that the biological fluid samples direct injected provides theoretical support.2.. from the micromechanism of micro emulsion, the micro emulsion drop has the biomembranous orderly hydrophilic area of class and hydrophobic region structure, so to water wettability and lipophilic compound are all had to stronger dissolving power.This just makes the Microemulsion liquid chromatography method measure water wettability simultaneously and lipophilic compound may become.3.. in the micro emulsion mobile phase, solute and micro emulsion drop and fixedly between the phase adsorbed layer, exist the multi-acting forces such as hydrophobic, hydrogen bond, static, so it is more to regulate and control parameter, be applicable to the separation of complex mixture in Chinese medicine.This is just one of active ingredient of Chinese herbs screening three large difficult points: active constituents of medicine can reach effective separation simultaneously provides theoretical support.The Microemulsion liquid chromatography method is promoted the use of in various fields.Such as: declaring study on the stability in new drug is an important content, institute's method for building up must be can will synthesize or impurity degraded from the major component peak with the auxiliary material peak separately and separates well.And the Microemulsion liquid chromatography method with unique separation selectivity and widely applicability in study on the stability, be used widely, represented the high potentiality of Microemulsion liquid chromatography method in stability study.
The stability of micro emulsion is the difficult point of its development of restriction always, but the prior art theory can solve the stability of micro emulsion.Why unstable micro emulsion is is because micro emulsion is comprised of oil phase, water, thus water-oil interface can exist than high tension, this be also surfactant be why form micro emulsion the reason of essential composition.As a rule, independent surfactant molecule can not make oil water interfacial tension be reduced to can form stable micro emulsion, if be applied in the high pressure liquid phase, tension force must produce because of the existence of high pressure demulsifying phenomenon.But current research shows, the alcohol of little molecular weight can be inserted in the middle of the surfactant of drop as cosurfactant, reduce the repulsive interaction between surfactant molecule, thereby can further reduce surface tension to the state that approaches " zero ", make micro emulsion keep stable.
Micro emulsion is that a kind of surface tension can be reduced to the stabilising system of " zero ", can realize the biological fluid samples direct injected as separating medium, and the separation selectivity that it is unique and broad applicability and active ingredient of Chinese herbs screening gordian technique requires to match, this makes the Microemulsion liquid chromatography method can be applicable to the active ingredient of Chinese herbs screening to become and more may.
Claims (6)
1. an active constituents of medicine screening technique is characterized in that: said method comprising the steps of:
1) tentatively determine active constituents of medicine:
1.1) obtain the pathology pharmacodynamics index of pathology body;
1.2) according to step 1.1) the pathology pharmacodynamics index of resulting pathology body obtains moment of maximum drug effect point of medicine of active constituents of medicine to be determined and the moment of minimum drug effect point;
1.3) obtain respectively whole blood and/or the body fluid of pathology body in the moment of moment of the maximum drug effect point of the medicine of active constituents of medicine to be determined and minimum drug effect point;
1.4) according to the Microemulsion liquid chromatography method to step 1.3) whole blood and/or the body fluid of resulting pathology body sets up respectively full finger-print;
1.5) according to drug effect differential serum chromatography by step 1.4) the full finger-print obtained of setting up contrasted, described full finger-print medium contrast maximum be preliminary definite active constituents of medicine; Described contrast maximum is that chromatographic peak changes the best part in the full finger-print of the full finger-print of maximum EDD point and minimum EDD point;
2) obtain preliminary definite active constituents of medicine;
3) detection, screening definite active constituents of medicine.
2. active constituents of medicine screening technique according to claim 1, it is characterized in that: specific implementation described step 1.1) is:
1.1.1) set up the pathological model of pathology body;
1.1.2) the pathology body is injected to the medicine of active constituents of medicine to be determined;
1.1.3) detect every pathology pharmacodynamics index of the pathology body of the medicine injected active constituents of medicine to be determined according to the conventional sense means.
3. active constituents of medicine screening technique according to claim 1 and 2, it is characterized in that: specific implementation described step 2) is:
Determining step 1) preliminary definite active constituents of medicine whether be standard items; If standard items, directly adopt standard items; If not, adopt routine techniques means preparation process 1) preliminary definite active constituents of medicine.
4. active constituents of medicine screening technique according to claim 3, it is characterized in that: the specific implementation of described step 3) is:
Determining step 2) obtaining preliminary definite active constituents of medicine and whether adopt standard items, if directly confirm step 2) institute's accepted standard product are exactly the active constituents of medicine of medicine to be determined; If not, according to the routine techniques means, detect, screen and determine the active constituents of medicine of medicine to be measured.
5. active constituents of medicine screening technique according to claim 4, it is characterized in that: described method also comprises after step 3):
4) the determined active constituents of medicine of step 3) is verified.
6. active constituents of medicine screening technique according to claim 5, it is characterized in that: the specific implementation of described step 4) is: step 3) is determined to the pharmacodynamic index of the active constituents of medicine obtained and the pharmacodynamic index of the medicine that the pathology body is injected are contrasted, if step 3) is determined the pharmacodynamic index of the active constituents of medicine obtained and is not less than 1/2 of the pharmacodynamic index of pathology body institute injectable drug, confirms that the resulting active constituents of medicine of step 3) is the active constituents of medicine to pathology body institute injectable drug; If step 3) determines that the pharmacodynamic index of the active constituents of medicine obtained is lower than 1/2 of the pharmacodynamic index to pathology body institute injectable drug, repeating step 1)-step 3), until the pharmacodynamic index of the active constituents of medicine finally obtained is not less than 1/2 of the pharmacodynamic index of pathology body institute injectable drug.
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| 杨静等: "红花的药效差示血清色谱法研究", 《第四军医大学学报》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108680654A (en) * | 2018-02-08 | 2018-10-19 | 杭州师范大学 | A kind of novel separation method of Chinese medicine |
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