Detection system for multiple medicinal components in medicine, pharmacokinetic analysis method and application of detection system in gynecological Qianjin prescription medicine

Abstract

The invention discloses a detection system for multiple symbolic medicinal components in a medicine, a pharmacokinetic analysis method and application of the detection system in a gynecological Qianjin prescription medicine. The detection system comprises a sample pretreatment module and a detection module, the detection module comprises ultra-high performance liquid chromatography-mass spectrometry equipment and is used for carrying out ultra-high performance liquid chromatography-mass spectrometry detection on the test sample solution; the pharmaceutical composition is characterized in that the pharmaceutical composition is prepared from a plurality of symbolic medicinal components, and the symbolic medicinal components comprise genistin, formononin, andrographolide, palmatine, magnoflorine, jateorhizine, berberine, dehydroandrographolide, genistein, formononetin, ligusticum chuanxiong hort lactone A and Z-ligustilide. According to the invention, the UPLC-MS / MS method is adopted to simultaneously determine the various symbolic medicinal components and the contents thereof in the traditional Chinese medicine, and the change rule of the various symbolic medicinal components in the body can be revealed, so that the metabolic difference and the absorption and transformation conditions of different patients after medication can be determined.

Description

Technical Field

[0001] The present invention belongs to the technical field of drug analysis, and more specifically, relates to a detection system for multiple active ingredients in drugs, a pharmacokinetic analysis method and its application in gynecological Qianjinfang drugs. Background Art

[0002] A large number of clinical studies have shown that the efficacy of drugs is not only closely related to the absorption of drugs in the body, but also to the distribution of drugs in tissues. The in vivo process is the basis for drugs to exert pharmacological effects and produce therapeutic effects. There are obvious advantages in studying the material basis and mechanism of the efficacy of traditional Chinese medicine from the in vivo process. The substance that exerts the efficacy of traditional Chinese medicine depends on whether it can eventually be absorbed into the systemic circulation and has suitable pharmacokinetic characteristics. There is a certain correlation between its in vivo pharmacokinetic process and efficacy. Since the content of traditional Chinese medicine ingredients is generally very low, the prototype drug ingredients and metabolites that enter the body are often only at the ng or pg level. Conventional analytical methods such as thin layer chromatography (TLC), high performance liquid chromatography (HPLC) and gas chromatography (GC) are difficult to detect, and the pretreatment operation is cumbersome, and the separation from the matrix is ​​also very difficult. There is a lack of effective means to solve the problems caused by the complex ingredients of traditional Chinese medicine, the relatively low content entering the body, and the large detection interference.

[0003] Genistin, formononetin, andrographolide, palmatine, magnolamine, jatrorrhizine, berberine, dehydroandrographolide, genistein, formononetin, tangelolide A and Z-ligustilide are important active ingredients in many Chinese herbal medicines and Chinese medicine preparations, many of which are standard ingredients in the quality control of Chinese medicine preparations or in the pharmacopoeia standards, and therefore can also be called iconic active ingredients. Currently, there is no system and method for simultaneous detection of the above multiple active ingredients that is fast, convenient, highly sensitive and highly specific.

[0004] The drugs of the Gynecological Qianjin Prescription include Gynecological Qianjin Tablets and Gynecological Qianjin Capsules, which are composed of Radix Achyranthis Bidentatae, Radix Chrysanthemi Indici, Andrographis paniculata, Chinese Mahoniae, Chinese Herba Atractylodes, Chinese Angelicae Sinensis, Millettia Spatholobi and Codonopsis Pilosulae. They are mainly used for chronic pelvic inflammatory disease, endometritis, chronic cervicitis, etc., with definite clinical efficacy, high market share and significant clinical and commercial value. Gynecological Qianjin Capsules are composed of Radix Achyranthis Bidentatae, Radix Achyranthis Bidentatae, Radix Achyranthis Bidentatae, Chinese Mahoniae Indici, Chinese Herba Achyranthis Bidentatae and Codonopsis Pilosulae. In this prescription, Radix Achyranthis Bidentatae and Chinese Mahoniae Indici clear away heat and detoxify, dry dampness and stop leukorrhea, and are the main drugs. Andrographis paniculata and Chinese Herba Atractylodes Pilosulae clear away heat and detoxify, cool blood and reduce swelling, dry dampness and stop leukorrhea, and are the minister drugs. Millettia Spatholobi and Chinese Angelicae Sinensis nourish blood and activate blood circulation; Codonopsis Pilosulae invigorates qi and strengthens spleen, promotes water and moisture transportation and stops leukorrhea; Radix Achyranthis Bidentatae consolidates essence and stops leukorrhea, and are the adjuvant drugs. All the drugs are combined to play the role of clearing heat and removing dampness, invigorating qi and removing blood stasis.

[0005] All drugs related to the Gynecological Qianjin Prescription involve the above-mentioned multiple active ingredients. Drugs related to the Gynecological Qianjin Prescription are used to treat gynecological diseases, which are common diseases among women. Gynecological inflammation is one of the gynecological diseases, including cervicitis, vaginitis, pelvic inflammatory disease, endometritis, etc., involving multiple tissues and organs.

[0006] At present, the research on the drugs related to Fuke Qianjinfang mainly focuses on the pharmacological effects, clinical applications and chemical components of single herbs, and the pharmacokinetic analysis method is relatively lacking. The quality detection of its active ingredients is of great significance for drug quality control.

[0007] Elucidating the absorption and transformation of Gynecological Qianjin Fang-related drugs in animals can provide a certain scientific basis for the exploration of the drugability of improved new drugs related to Gynecological Qianjin Fang and the development of new drugs, and can also provide guidance for the subsequent auxiliary treatment of Gynecological Qianjin Fang-related drugs. Therefore, how to simultaneously detect the characteristic drug components in Gynecological Qianjin Fang-related drugs and apply them to explore and clarify the metabolism and specific distribution of the characteristic pharmacological components in Gynecological Qianjin Fang-related drugs in the body (blood and various tissues) has become a technical problem that needs to be solved urgently, and will also provide valuable technical references and references for drug research methods in this field. Summary of the invention

[0008] In view of the above-mentioned existing technical problems, the primary purpose of the present invention is to provide a simultaneous detection system for multiple characteristic pharmacological ingredients in a drug. The detection system adopts the UPLC-MS / MS method to quickly, sensitively, accurately and simply determine multiple characteristic pharmacological ingredients and their contents in the drug at the same time. The detection system can also be used in the blood or tissue of animals that have taken the drug in vitro to reveal the changing patterns of multiple characteristic pharmacological ingredients in the body, so as to clarify the metabolic differences, absorption and conversion conditions of different patients after taking the drug.

[0009] In order to achieve the above object, the present invention is implemented by the following technical solutions:

[0010] A system for simultaneously detecting multiple characteristic pharmacodynamic components in a drug, comprising: a sample pretreatment module and a detection module;

[0011] The detection module includes an ultra-high performance liquid chromatography-mass spectrometry device, and the detection module performs ultra-high performance liquid chromatography-mass spectrometry detection on the test solution to obtain the compound content of multiple characteristic medicinal ingredients in the test solution;

[0012] The detection conditions of the ultra-high performance liquid chromatography are as follows: chromatographic column: ACQUITY UPLC BEH C18, the specification of the chromatographic column is 2.1mm×100mm, 1.7μm; the flow rate is 0.3-0.4mL / min; the column temperature is 30-45°C; the injection volume is 4uL; the mobile phase is composed of phase A and phase B, phase A is acetonitrile, and phase B is 0.1% formic acid water;

[0013] At the beginning of gradient elution, 0-0.5 min, 95% phase B;

[0014] 0.5-10min, 95%-70% phase B;

[0015] 10-15 min, 70%-50% phase B;

[0016] 15-20 min, 50%-10% phase B;

[0017] 20-22 min, 10% phase B;

[0018] 22min~22.1min, 10%~95% phase B;

[0019] 22.1min~24min, 95% phase B;

[0020] The mass spectrometry detection conditions are as follows: acquisition in the positive ion mode using the multiple reaction monitoring mode, the capillary voltage is 1.5-2KV, the cone voltage is 20V, the ion source temperature is 150°C, the desolvation gas temperature is 400-500°C, the desolvation gas flow rate is 800-1000L / h, and the cone gas flow rate is 150L / h;

[0021] The drug contains a variety of characteristic pharmacological ingredients including genistein, formononetin, andrographolide, palmatine, magnolamine, jatrorrhizine, berberine, dehydroandrographolide, genistein, formononetin, ligustilide A and Z-ligustilide.

[0022] Because the content of Chinese medicine ingredients is generally low, the prototype drug ingredients and metabolites entering the body are often only at the ng or pg level. Conventional analytical methods such as thin layer chromatography (TLC) and gas chromatography (GC) are difficult to detect, and the pre-treatment operation is cumbersome, and the difficulty of separation from the matrix is ​​also very large. Based on this, the present invention adopts the detection technology of ultra-high performance liquid chromatography and mass spectrometry in series, combining the high separation ability of ultra-high performance liquid chromatography technology with the high sensitivity and high specificity of mass spectrometry technology, avoiding the above problems. Further, the inventor optimized the ultra-high performance liquid chromatography conditions and mass spectrometry conditions through research, and adopted the ultra-high pressure short chromatographic column of 1.7 μm bridged ethyl idene hybrid particles (Bridged ethyl idene hybridparticles, BEH) for ultra-high performance liquid chromatography analysis, and used 0.1% formic acid water and acetonitrile as mobile phases for gradient elution. Under this detection condition, the multiple characteristic active ingredients in the drug achieve good sensitivity and high separation, and the chromatogram has a good peak shape, which is easy to identify. In addition, the present invention also adopts the multi-reactive ion monitoring (MRM) mode instead of the single ion monitoring (SRM) mode for monitoring, which reduces the interference of other impurity components in the sample, and selects positive ions to synchronously monitor and determine all analytes and internal standards, thereby enhancing specificity and improving sensitivity.

[0023] The detection system of the present invention uses UPLC-MS / MS method to rapidly, sensitively, accurately and conveniently simultaneously determine each component and its content in the drug containing multiple characteristic pharmacological ingredients of the present invention, and can be applied to the blood and / or tissue of animals that have taken the drug in vitro, and obtain its pharmacokinetic parameters through analysis to reveal the changing rules of multiple characteristic pharmacological ingredients in vivo. The detection system can monitor patients after taking the drug to clarify the metabolic differences, absorption and conversion of different patients after taking the drug, and then provide guidance and direction for the subsequent medication plan of the drug and the combined treatment plan of the drug and other drugs.

[0024] Preferably, the sample of the test solution is selected from one or more of the drug, the blood of an animal in vitro that has taken the drug, or the tissue of an animal in vitro that has taken the drug; the detection module also includes ultra-high performance liquid chromatography-mass spectrometry detection of a reference solution, and the sample of the reference solution is selected from a standard solution, the blood or tissue of a blank control animal containing a standard solution, and the blank control animal has not taken the drug; the tissue includes one or more of heart tissue, liver tissue, spleen tissue, lung tissue, kidney tissue, brain tissue and uterine tissue.

[0025] Preferably, when the test solution or the reference solution is a drug, the pretreatment module includes an ultrasonic unit and a centrifugal unit; the ultrasonic unit includes an ultrasonic component, and the centrifugal unit includes a centrifugal component; the operation method of using the sample pretreatment module is: dissolving the drug in water, ultrasonicating with an ultrasonic component, and then centrifuging in a centrifugal component to obtain the test solution or the reference solution;

[0026] Or, when the sample of the test solution or the reference solution is blood, the sample pretreatment module includes an anticoagulation unit, a centrifugal unit, a protein precipitation unit, and a protein removal unit; the anticoagulation unit includes an anticoagulant, the centrifugal unit includes a centrifugal component, the protein precipitation unit includes a protein precipitant containing an internal standard solution, and the protein removal unit includes a vortex mixing component, a centrifugal component, an ultrasonic component, and a protein precipitant;

[0027] The operation method of using the sample pretreatment module is as follows: blood is mixed with an anticoagulant, and then input into a centrifugal component for centrifugation to obtain a plasma sample; the plasma sample is mixed with a protein precipitant containing an internal standard solution, and then input into a vortex mixing component and a centrifugal component in sequence for vortex mixing and centrifugation to obtain a residue; the residue is mixed with a protein precipitant, and then input into a vortex mixing component, an ultrasonic component, and a centrifugal component again for vortex mixing, ultrasonication, and centrifugation to obtain the test solution or the reference solution;

[0028] Or, when the sample of the test solution or the reference solution is tissue, the sample pretreatment module includes a homogenization unit, a protein precipitation unit, a centrifugal unit, a vortex mixing unit, and an ultrasonic unit; the homogenization unit includes a homogenization component, the protein precipitation unit includes a protein precipitant containing an internal standard solution, the centrifugal unit includes a centrifugal component, the vortex mixing unit includes a vortex mixing component, and the ultrasonic unit includes an ultrasonic component;

[0029] The operation method of using the sample pretreatment module is as follows: homogenizing the tissue in a homogenization component, then mixing it with a protein precipitant containing an internal standard solution, then homogenizing it using the homogenization component and centrifuging it using the centrifugation component, taking the supernatant after centrifugation and mixing it with the protein precipitant, then vortexing, sonicating, and centrifuging in a vortex mixing component, an ultrasonic component, and a centrifugation component in sequence to obtain the test solution or the reference solution.

[0030] Preferably, the detection system and method of the present invention are particularly suitable for drugs related to the Gynecological Qianjin Recipe, including but not limited to Gynecological Qianjin Tablets and Capsules.

[0031] More specifically, 10 pills of the drug were taken, dissolved in 10 mL of pure water, sonicated for 10 min using an ultrasonic component, and then centrifuged in a centrifugal component, and the supernatant was taken to obtain a test solution.

[0032] Preferably, the centrifugal speed is 2000-4000 rpm and the centrifugal time is 8-15 min.

[0033] Preferably, after the plasma sample is mixed with the protein precipitant or homogenized using a homogenization component, it is then centrifuged at a speed of 12000-13000 rpm; the centrifugation temperature is 2-6° C.; and the centrifugation time is 8-15 min.

[0034] Preferably, the frequency of the homogenization is 50 to 80 Hz, and the time of the homogenization is 8 to 12 minutes.

[0035] Further preferably, the anticoagulant may be a blood anticoagulant conventionally used in the art for ex vivo blood or various tissues; more specifically, the anticoagulant includes but is not limited to sodium heparin, lithium heparin, and the like.

[0036] Further preferably, the centrifugal component may be a device or apparatus conventionally used in the art for sample centrifugation; more specifically, the centrifugal component includes but is not limited to a centrifuge, a refrigerated centrifuge, etc. The vortex mixing component may be a device or apparatus conventionally used in the art for sample vortex mixing; more specifically, the vortex mixing component includes but is not limited to a vortex mixer, etc. The ultrasonic component may be a device or apparatus conventionally used in the art for sample ultrasonication; more specifically, the ultrasonic component includes but is not limited to an ultrasonic cleaning machine, etc. The homogenization component may be a device or apparatus conventionally used in the art for sample homogenization; more specifically, the homogenization component includes but is not limited to a homogenizer, a handheld high-speed homogenizer, etc.

[0037] Further preferably, the ultra-high performance liquid chromatography-mass spectrometry device can be a device or apparatus conventionally used for sample analysis in the art. More specifically, the ultra-high performance liquid chromatography-mass spectrometry device includes but is not limited to an ultra-high performance liquid chromatography-tandem mass spectrometer, an ultra-high performance liquid chromatography-mass spectrometer, an ultra-high performance liquid chromatography-triple quadrupole mass spectrometer, and the like.

[0038] Preferably, the standard solution is a methanol solution of the characteristic medicinal ingredient.

[0039] Preferably, the protein precipitant is selected from one or more of acetonitrile, methanol or ethanol.

[0040] Preferably, when the sample is blood, the volume ratio of plasma to protein precipitant is 1:(2-6); when the sample is tissue, the mass volume ratio of tissue to protein precipitant is 1:(2-6) mg / mL.

[0041] Further preferably, when the sample is blood, the volume ratio of plasma to protein precipitant is 1:4; when the sample is tissue, the mass volume ratio of tissue to protein precipitant is 1:4 mg / mL. At this preferred ratio, the protein precipitant has better separation and extraction effects, more stable repetition rate and recovery rate for multiple characteristic active ingredients in plasma or tissue. Further preferably, when the sample is blood, the protein precipitant is selected from acetonitrile; when the sample is tissue, the protein precipitant is selected from methanol.

[0042] Preferably, the internal standard solution is a mixed solution of psoralen and methanol.

[0043] The present invention also claims a method for analyzing the pharmacokinetic properties of multiple characteristic pharmacodynamic components in a drug, wherein the drug contains multiple characteristic pharmacodynamic components, including: genistein, formononetin, andrographolide, palmatine, magnolamine, jatrorrhizine, berberine, dehydroandrographolide, genistein, formononetin, tangelolide A and Z-ligustilide; the method for analyzing the pharmacokinetic properties includes a sample pretreatment step and a detection step;

[0044] S1. Sample pretreatment: taking samples for pretreatment to obtain test sample solutions; the samples are selected from animal samples collected at different time points, isolated from the body, and taken with the drug, the animal samples include animal blood and animal tissues; the animal tissues include one or more of heart tissue, liver tissue, spleen tissue, lung tissue, kidney tissue, brain tissue and uterine tissue;

[0045] S2. Detection: The test solution is tested using ultra-high performance liquid chromatography-mass spectrometry to obtain the compound content of multiple characteristic active ingredients in the test solution;

[0046] The detection conditions of the ultra-high performance liquid chromatography are as follows: chromatographic column: ACQUITY UPLC BEH C18, the specification of the chromatographic column is 2.1mm×100mm, 1.7μm; the flow rate is 0.3-0.4mL / min; the column temperature is 30-45°C; the injection volume is 4uL; the mobile phase is composed of phase A and phase B, phase A is acetonitrile, and phase B is 0.1% formic acid water;

[0047] At the beginning of gradient elution, 0-0.5 min, 95% phase B;

[0048] 0.5-10min, 95%-70% phase B;

[0049] 10-15 min, 70%-50% phase B;

[0050] 15-20 min, 50%-10% phase B;

[0051] 20-22 min, 10% phase B;

[0052] 22min~22.1min, 10%~95% phase B;

[0053] 22.1min~24min, 95% phase B;

[0054] The mass spectrometry detection conditions are as follows: acquisition in the positive ion mode using the multiple reaction monitoring mode, the capillary voltage is 1.5-2KV, the cone voltage is 20V, the ion source temperature is 150°C, the desolvation gas temperature is 400-500°C, the desolvation gas flow rate is 800-1000L / h, and the cone gas flow rate is 150L / h;

[0055] Preferably, when the sample is the blood of an animal that has taken the drug in vitro, the pretreatment is: mixing the blood with an anticoagulant, then inputting the blood into a centrifugal component for centrifugation to obtain a plasma sample; mixing the plasma sample with a protein precipitant containing an internal standard solution, sequentially inputting the blood into a vortex mixing component and a centrifugal component for vortex mixing and centrifugation to obtain a residue; mixing the residue with a protein precipitant, and again inputting the blood into a vortex mixing component, an ultrasonic component, and a centrifugal component for vortex mixing, ultrasonication, and centrifugation to obtain the test solution.

[0056] Preferably, when the sample is an animal tissue isolated from a subject that has taken the drug, the pretreatment is as follows: the tissue is homogenized in a homogenization component, and then mixed with a protein precipitant containing an internal standard solution, and then homogenized by a homogenization component and centrifuged by a centrifugation component, and after centrifugation, the supernatant is taken and mixed with a protein precipitant, and then vortexed, sonicated, and centrifuged in a vortex mixing component, an ultrasonic component, and a centrifugation component in sequence to obtain the test solution.

[0057] When the sample is animal blood, the different time points are: 0.083h, 0.167h, 0.33h, 0.5h, 0.75h, 1h, 1.5h, 2h, 4h, 6h, 8h, 12h, and 24h after the animal is administered the drug.

[0058] When the sample is animal tissue, the different time points are: 0.5h, 1h, 4h, 8h, and 16h.

[0059] In the above scheme, the drug can be a drug formed by mixing extracts of any one or more crude drugs of Herba Lycopodii, Radix Chrysanthemi Indici, Herba Andrographis, Herba Mahoniae, Herba Lycopodii, Radix Angelicae Sinensis, Herba Millettiae, or Herba Codonopsis Pilosulae. Further preferably, the drug can be Gynecological Qianjin Capsules or Gynecological Qianjin Tablets.

[0060] Furthermore, the present invention also requests protection for the application of the detection system or method in the detection and quality control of the drug ingredients related to the Gynecological Qianjin Prescription, or in the screening and determination of medication regimens and dosages, or in drug development.

[0061] Compared with the prior art, the present invention has the following beneficial effects:

[0062] The present invention provides a detection system and a pharmacokinetic analysis method for multiple characteristic pharmacodynamic components in a drug. The system and method use UPLC-MS / MS method to rapidly, sensitively, accurately and conveniently simultaneously determine multiple components and their contents in a drug, and obtain their pharmacokinetic parameters through analysis to reveal the changing patterns of multiple characteristic pharmacodynamic components in the body. The detection system can monitor patients after taking the drug to clarify the metabolic differences, absorption and conversion conditions of different patients after taking the drug, and then provide guidance and direction for the subsequent auxiliary treatment plan of the drug and the combined treatment plan of the drug and other drugs. In addition, the detection system can also be used to identify and evaluate the specific content of each characteristic pharmacodynamic component in the drug, and can be used for the stability evaluation of the drug. BRIEF DESCRIPTION OF THE DRAWINGS

[0063] Figure 1 Chromatograms of the pharmacodynamic components of each marker after adding 4 times the volume of acetonitrile solution to the plasma sample.

[0064] Figure 2 Chromatograms of the pharmacological components of each marker after adding 4 times the volume of methanol solution to the plasma sample.

[0065] Figure 3 Chromatograms of the pharmacodynamic components of each marker after adding 2 times the volume of acetonitrile solution to the plasma sample.

[0066] Figure 4 Chromatograms of the pharmacodynamic components of each marker after adding 6 times the volume of acetonitrile solution to the plasma sample.

[0067] Figure 5 This is the chromatogram of heart tissue 0.5 hours after oral administration of Fuke Qianjin Capsule.

[0068] Figure 6 This is the chromatogram of liver tissue 0.5 hours after oral administration of Fuke Qianjin Capsule.

[0069] Figure 7 This is the chromatogram of spleen tissue 0.5 hours after oral administration of Fuke Qianjin Capsule.

[0070] Figure 8 This is the chromatogram of lung tissue 0.5 hours after oral administration of Fuke Qianjin Capsule.

[0071] Fig. 9 This is the chromatogram of kidney tissue 0.5 hours after oral administration of Fuke Qianjin Capsule.

[0072] Fig.10 This is the chromatogram of brain tissue 0.5 hours after oral administration of Fuke Qianjin Capsule.

[0073] Fig.11This is the chromatogram of uterine tissue 0.5 hours after oral administration of Fuke Qianjin Capsule.

[0074] Fig.12 The chromatogram is of blank plasma containing internal standard solution.

[0075] Fig.13 The chromatograms are of blank plasma spiked with various marker active ingredients and internal standard solutions.

[0076] Fig.14 This is the chromatogram of plasma 0.5 hours after oral administration of Fuke Qianjin Capsule.

[0077] Fig.15 It is the linear range, standard curve, correlation coefficient, precision, accuracy, recovery rate, matrix effect and stability diagram of multiple marker active components in plasma.

[0078] Fig.16 These are the linear range, standard curve, correlation coefficient, precision, accuracy, recovery rate, matrix effect and stability diagram of multiple marker active components in cardiac tissue.

[0079] Fig.17 These are the linear range, standard curve, correlation coefficient, precision, accuracy, recovery rate, matrix effect and stability diagram of various marker active components in liver tissue.

[0080] Fig.18 These are the linear range, standard curve, correlation coefficient, precision, accuracy, recovery rate, matrix effect and stability diagram of various marker active components in spleen tissue.

[0081] Fig.19 These are the linear range, standard curve, correlation coefficient, precision, accuracy, recovery rate, matrix effect and stability diagram of multiple marker active components in lung tissue.

[0082] Fig. 20 These are the linear range, standard curve, correlation coefficient, precision, accuracy, recovery rate, matrix effect and stability diagram of various marker active components in renal tissue.

[0083] Fig.21 These are the linear range, standard curve, correlation coefficient, precision, accuracy, recovery rate, matrix effect and stability diagram of various marker active components in brain tissue.

[0084] Fig. 22 These are the linear range, standard curve, correlation coefficient, precision, accuracy, recovery rate, matrix effect and stability diagram of various marker active components in uterine tissue.

[0085] Fig.23This is a curve chart of the concentration of marker active ingredients in the blood versus time after taking Fuke Qianjin Capsule.

[0086] Fig.24 These are the pharmacokinetic parameters of multiple marker active ingredients after oral administration of 4g / kg dose of Fuke Qianjin Capsule.

[0087] Fig.25 This is a distribution rectangle diagram of the same marker active ingredient in different organs at different time points after Fuke Qianjin Capsule was gavage-administered to rats. DETAILED DESCRIPTION

[0088] The present invention is further described below in conjunction with the accompanying drawings and specific examples of the specification, but the examples do not limit the present invention in any form. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the field of the present technology. For the convenience of explanation, the following examples are described with Gynecological Qianjin Capsules as medicines, but obviously, the design concept of the present invention is applicable to medicines containing genistein, formononetin, andrographolide, palmatine, magnolamine, jatrorrhizine, berberine, dehydroandrographolide, genistein, formononetin, yangchuanqiong lactone A and Z-ligustilide, among which multiple active ingredients are not limited to Gynecological Qianjin Capsules.

[0089] The instruments and equipment used in the embodiments of the present invention are as follows.

[0090] Handheld high-speed homogenizer (model: HR-6; source: Shanghai Huxi); vortex mixer (model: VORTEX 2S025; source: IKA); ultrasonic cleaner (model: SB-800DTD; source: Ningbo Xinzhi); nitrogen blower (model: MD 200-2; source: IKA); electronic balance (model: XSE105DU; source: Mettler Toledo); ultra-high performance liquid chromatography-triple quadrupole mass spectrometer (model: Waters XEVO TQ-XS; source: Waters); refrigerated centrifuge (model: 5430R; source: Eppendorf Shanghai Huxi).

[0091] Example 1 A detection system for multiple characteristic active ingredients in a drug

[0092] A detection system for multiple characteristic active ingredients in a drug comprises: a sample pretreatment module and a detection module, wherein the detection module comprises an ultra-high performance liquid chromatography-triple quadrupole mass spectrometer, and the detection module performs ultra-high performance liquid chromatography-mass spectrometry detection on a test solution and a reference solution to obtain the compound contents of multiple active ingredients in the test solution.

[0093] In this embodiment, samples of the test solution are selected from the blood and tissues of animals that have taken Fuke Qianjin Capsules in vitro; samples of the reference solution are selected from the blood and tissues of blank control animals containing standard solutions, and the blank control animals have not taken Fuke Qianjin Capsules; the tissues include heart tissue, liver tissue, spleen tissue, lung tissue, kidney tissue, brain tissue and uterine tissue.

[0094] When the test solution or the reference solution is the Gynecological Qianjin Capsule, the pretreatment module includes an ultrasonic unit and a centrifugal unit; the ultrasonic unit includes an ultrasonic component, and the centrifugal unit includes a centrifugal component; the operation method of using the sample pretreatment module is: dissolve the Gynecological Qianjin Capsule in water, use the ultrasonic component to sonicate, and then centrifuge in the centrifugal component to obtain the test solution or the reference solution.

[0095] When the sample of the test solution or the reference solution is blood, the sample pretreatment module includes an anticoagulation unit, a centrifugation unit, a protein precipitation unit, and a protein removal unit; the anticoagulation unit includes an anticoagulant, the centrifugation unit includes a centrifugal component, the protein precipitation unit includes a protein precipitant containing an internal standard solution, and the protein removal unit includes a vortex mixing component, a centrifugal component, an ultrasonic component, and a protein precipitant.

[0096] When the sample of the test solution or the reference solution is tissue, the sample pretreatment module includes a homogenization unit, a protein precipitation unit, a centrifugal unit, a vortex mixing unit, and an ultrasonic unit; the homogenization unit includes a homogenization component, the protein precipitation unit includes a protein precipitant containing an internal standard solution, the centrifugal unit includes a centrifugal component, the vortex mixing unit includes a vortex mixing component, and the ultrasonic unit includes an ultrasonic component.

[0097] In this embodiment, the centrifugal component is a refrigerated centrifuge, the vortex mixing component is a vortex mixer, the ultrasonic component is an ultrasonic cleaning machine, and the homogenization component is a handheld high-speed homogenizer.

[0098] In this embodiment, the standard solution is a methanol solution of multiple active ingredients, the protein precipitant is acetonitrile or methanol, and the internal standard solution is a mixed solution of psoralen and methanol.

[0099] In this embodiment, the detection conditions of ultra-high performance liquid chromatography are as follows: chromatographic column: ACQUITY UPLC BEH C18, the specifications of the chromatographic column are 2.1 mm×100 mm, 1.7 μm; the flow rate is 0.4 mL / min; the column temperature is 40° C.; the injection volume is 4 uL; the mobile phase is composed of phase A and phase B, phase A is acetonitrile, and phase B is 0.1% formic acid water; the program elution conditions are shown in Table 1 below:

[0100] Table 1

[0101] Time (min) A％ B％ 0.0 5.0 95 0.5 5.0 95 10.0 30 70 15.0 50 50 20.0 90 10 22.0 90 10 22.1 5.0 95 24.0 5.0 95

[0102] The detection conditions of mass spectrometry are as follows: acquisition in multiple reaction monitoring mode in positive ion mode, capillary voltage of 2KV, cone voltage of 20V, ion source temperature of 150°C, desolvation gas temperature of 500°C, desolvation gas flow rate of 800L / h, and cone gas flow rate of 150L / h.

[0103] In this embodiment, the multiple characteristic medicinal ingredients include: genistein, formononetin, andrographolide, palmatine, magnolamine, jatrorrhizine, berberine, dehydroandrographolide, genistein, formononetin, ligustilide A and Z-ligustilide.

[0104] In this embodiment, the use and operation method of the detection system for simultaneously detecting multiple medicinal ingredients in Fuke Qianjin Capsule includes the following steps:

[0105] (1) Sample acquisition

[0106] Blood sample acquisition: After three days of adaptive feeding, 8 healthy female SD rats were fasted for 12 hours, with free access to water during the period. Gynecological Qianjin Capsule (source: Zhuzhou Qianjin Pharmaceutical Co., Ltd.) was administered at a dose of 4g / kg, and blood samples of about 400μL were collected from the eye sockets of the 8 rats before and 0.5h after administration (the blood samples obtained before administration were blank blood samples).

[0107] Tissue sample acquisition: After three days of adaptive feeding, 48 healthy female SD rats were randomly divided into 6 groups, with 8 rats in each group. The 48 rats were fasted for 12 hours and had free access to water during the period. Except for the blank group, the other groups were given 4g / kg of Fuke Qianjin Capsule, and then each group was anesthetized with 10% chloral hydrate 0.5h after administration, and blood was taken into EP tubes with anticoagulant added, placed in a 4℃ refrigerator, centrifuged at 3000rpm for 10min, and the upper plasma was taken into centrifuge tubes, and then heart tissue, liver tissue, spleen tissue, lung tissue, kidney tissue, brain tissue and uterine tissue were respectively removed, and the residual blood on the surface was washed with 0.9% saline, the filter paper was blotted and weighed, and the liquid nitrogen was quickly frozen and stored at -80℃ for use. The blank group was anesthetized with 10% chloral hydrate, blood was collected and heart tissue, liver tissue, spleen tissue, lung tissue, kidney tissue, brain tissue and uterine tissue were removed, and the residual blood on the surface was washed with 0.9% saline, dried with filter paper, weighed, quick-frozen in liquid nitrogen, and stored at -80°C for use, which was the blank tissue sample.

[0108] (2) Sample pretreatment

[0109] Among them, the sample pretreatment steps of the test solution are as follows:

[0110] The blood sample of the animal that has taken Fuke Qianjin Capsule obtained in step (1) is placed in a dried heparin tube (the tube contains anticoagulant heparin lithium or heparin sodium) or a heparin anticoagulant tube with a small amount of liquid, so that the blood sample and the anticoagulant are fully mixed. After centrifugation at 3000 rpm for 10 minutes, the supernatant obtained is the plasma sample.

[0111] 4 volumes of acetonitrile solution and internal standard solution (the internal standard solution is a methanol solution of psoralen with a psoralen content of 10 ng / mL) were added to the plasma sample, vortexed for 5 minutes, and centrifuged at 12700 rpm and 4°C for 10 minutes. 400 μL of the supernatant was taken and dried at room temperature to obtain the residue. The residue was re-dissolved in 150 μL of acetonitrile solution and vortexed. After ultrasonication for 5 minutes, the supernatant was centrifuged for 10 minutes and injected to obtain the drug-containing plasma analysis sample (test solution).

[0112] After SD rats were gavaged with Fuke Qianjin Capsule, the heart tissue, liver tissue, spleen tissue, lung tissue, kidney tissue, brain tissue and uterine tissue collected were added with magnetic beads and homogenized at 65 Hz for 10 minutes. 300 mg of each tissue was weighed, and 4 volumes (1200 mL) of methanol solution and internal standard solution (the internal standard solution was a methanol solution of psoralen; the psoralen content was 10 ng / mL) were added and homogenized at 65 Hz for 10 minutes. Subsequently, the mixture was centrifuged at 12700 rpm and 4°C for 10 minutes, and 800 μL of supernatant was taken and dried at room temperature; the mixture was reconstituted with 200 μL of pure methanol and vortexed, and the mixture was ultrasonicated for 5 minutes and centrifuged for 10 minutes, and the supernatant was taken for sampling to obtain drug-containing tissue analysis samples (test solution).

[0113] (3) Ultra-high performance liquid chromatography-mass spectrometry detection

[0114] The drug-containing plasma analysis samples and drug-containing tissue analysis samples in step (2) were subjected to ultra-high performance liquid chromatography-mass spectrometry detection according to the above-mentioned chromatography and mass spectrometry detection conditions. Among them, the MRM information of the 12 active ingredients and psoralen is shown in Table 2 below:

[0115] Table 2

[0116]

[0117] The chromatograms of each active ingredient in the drug-containing plasma analysis sample are as follows: Figure 1 The chromatograms of the active ingredients of the drug-containing tissue analysis samples are shown in Figures 5 to 11 As shown. Figure 1 and Figures 5 to 11 As shown, the detection system of the present invention can rapidly, sensitively, accurately and conveniently determine the 12 medicinal ingredients and their contents in Fuke Qianjin Capsule, and can clearly determine the distribution of the 12 medicinal ingredients in the blood and / or tissues of animals that have taken Fuke Qianjin Capsule.

[0118] Example 2 Investigation of different protein precipitants and their ratios

[0119] (1) Investigation of plasma samples with different types and volume ratios of protein precipitants

[0120] The "plasma sample is added with 4 times the volume of acetonitrile solution" in step (2) of Example 1 was replaced with "plasma sample is added with 4 times the volume of methanol solution", "plasma sample is added with 2 times the volume of acetonitrile solution", and "plasma sample is added with 6 times the volume of acetonitrile solution", respectively. The remaining steps were the same and the investigation was carried out. Different drug-containing plasma analysis samples were prepared and tested using the ultra-high performance liquid chromatography-mass spectrometry detection conditions in step (3) of Example 1. The results are shown in FIG. Figures 1 to 4 shown.

[0121] Depend on Figures 1 to 4 It can be seen that the peak shape of the target compound detected by adding 4 times the volume of acetonitrile solution to the plasma sample is better, the signal-to-noise ratio and peak height are higher, and the extraction effect of the target compound in the sample is better than the extraction effect using 2 times the volume of acetonitrile solution, better than the extraction effect using 6 times the volume of acetonitrile solution, and better than the extraction effect using 4 times the volume of methanol solution.

[0122] (2) Investigation of tissue samples (liver tissue) and protein precipitants of different types and volume ratios

[0123] In step (2) of Example 1, "adding 4 times the volume of methanol solution to the tissue sample" was replaced by "adding 2 times the volume of methanol solution to the tissue sample", "adding 6 times the volume of methanol solution to the tissue sample", "adding 2 times the volume of acetonitrile solution to the tissue sample", "adding 4 times the volume of acetonitrile solution to the tissue sample", and "adding 6 times the volume of acetonitrile solution to the tissue sample", and the remaining steps were the same for investigation. Different drug-containing tissue analysis samples were prepared and tested using the ultra-high performance liquid chromatography-mass spectrometry detection conditions in step (3) of Example 1.

[0124] Table 3 shows the peak areas of drug components in drug-containing tissue analysis samples (liver tissue) after administration after being subjected to protein precipitants of different types and volume ratios.

[0125] Table 3

[0126]

[0127]

[0128] It can be seen from Table 3 that when the mass volume ratio of the tissue sample to the protein precipitant (methanol) is 1:4, the detection system generally has a larger peak area for a variety of active ingredients, that is, when the mass volume ratio of the tissue sample to the protein precipitant (methanol) is 1:4, the tissue sample has a better extraction effect, and the detection system can achieve a stable repetition rate and an optimal recovery rate.

[0129] Example 3 Methodological investigation of ultra-high performance liquid chromatography-mass spectrometry detection, pharmacokinetic and tissue distribution experimental results

[0130] The compounds of 12 kinds of active ingredients in Fuke Qianjin Capsule (genistein, formononetin, andrographolide, palmatine, magnolamine, jatrorrhizine, berberine, dehydroandrographolide, genistein, formononetin, yangchuanqiolide A and ligustilide) were accurately weighed and dissolved in methanol to obtain standard solutions with concentrations of 1.0020 mg / mL, 1.0070 mg / mL, 1.0020 mg / mL, 1.0010 mg / mL, 1.0000 mg / mL, 0.99000 mg / mL, 1.0060 mg / mL, 1.0000 mg / mL, 1.0050 mg / mL, 0.9800 mg / mL, 1.0000 mg / mL, and 1.0000 mg / mL, respectively. The standard solutions of the 12 kinds of marker active ingredients were accurately measured respectively, and gradiently diluted with methanol to the required concentration to obtain the reference substance stock solution.

[0131] Take the blank plasma sample and each blank tissue sample in Example 1, and add the reference substance stock solutions of 12 kinds of active ingredients and the internal standard solution (the internal standard solution is a methanol solution of psoralen, and the psoralen content is 10 ng / mL) to the blank plasma sample and each blank tissue sample, respectively. The remaining steps are the same to prepare the low, medium and high concentration quality control samples shown in Table 4 below for subsequent analysis.

[0132] Table 4

[0133]

[0134]

[0135] (1) Specificity inspection

[0136] Take a blank plasma sample, and except for not adding the internal standard solution, the rest of the operation is carried out according to the "sample pretreatment step of the test solution" in step (2) of Example 1 to obtain a blank sample chromatogram; a certain concentration of mixed standard solution (genistein, formononetin, andrographolide, palmatine, magnolamine, jatrorrhizine, berberine, dehydroandrographolide, genistein, formononetin, tangelolide A and ligustilide mixed series reference solution concentrations are: 0.50 ng / mL, 0.05 ng / mL, 0.38 ng / mL, 0.38 ng / mL, 0.38 ng / mL, 0.10 ng / mL, 0.75 ng / mL, 37.50 ng / mL, 2.00 ng / mL, 0.08 ng / mL, 0.20 ng / mL, 12.50 ng / mL) and internal standard solution (10 ng / mL) were added to the blank plasma sample, and the corresponding chromatograms were obtained according to the above operation method; the plasma sample in step (2) of Example 1 was taken and the corresponding chromatogram was obtained according to the above method.

[0137] (2) Investigation of standard curve, linear range and limit of quantitation

[0138] Six blank matrices (blank plasma samples and blank tissue samples) were taken, and the internal standard solution (10 ng / mL) and a certain concentration of mixed standard solution (the concentrations of the mixed series of reference solutions of genistein, formononetin, andrographolide, palmatine, magnolamine, jatrorrhizine, berberine, dehydroandrographolide, genistein, formononetin, ligustilide A and ligustilide were 0.50 ng / mL, 0.05 ng / mL, 0.38 ng / mL, 0.38 ng / mL, 0.10 ng / mL, respectively) were added to the blank matrices. g / mL, 0.75ng / mL, 37.50ng / mL, 2.00ng / mL, 0.08ng / mL, 0.20ng / mL, 12.50ng / mL) were added to the blank matrix respectively, and the rest was operated according to the "sample pretreatment step of the test solution" in Example 1. The internal standard method was used for quantification. The ratio of the peak area of ​​the analyte to the peak area of ​​the internal standard (A / Ai) was used as the ordinate Y, and the concentration of each substance (C) was used as the abscissa X for linear regression. The weight coefficient was 1 / X2, and the linear equation was obtained, which was the standard curve. The lowest point concentration of the calibration curve was used as the limit of quantification (LOQ), and the LOQ met the relative error (RE≤20%) and relative standard deviation (RSD≤20%), as well as the signal-to-noise ratio ≥10).

[0139] (3) Accuracy and precision

[0140] Intra-day and inter-day accuracy and precision were evaluated by analyzing quality control samples at low, medium, and high concentrations on one and three consecutive days.

[0141] (4) Extraction recovery and matrix effect

[0142] The extraction recoveries were evaluated using quality control samples at low, medium, and high concentrations and calculated by comparing the IS-normalized peak areas of blank samples spiked with the 12 active ingredients before and after extraction. Matrix effects were evaluated by comparing the IS-normalized peak areas of the active ingredients in the blank biological matrix to the IS-normalized peak areas of the active ingredients dissolved in methanol but not in the biological matrix.

[0143] (5) Stability

[0144] Short-term stability was evaluated by analyzing quality control samples stored at room temperature (25°C) for 8 hours. Long-term stability was evaluated by analyzing samples after 30 days of storage at -80°C. Freeze-thaw stability was evaluated by analyzing quality control samples after three freeze-thaw cycles (-80°C).

[0145] Under the conditions of ultra-high performance liquid chromatography-mass spectrometry, the results of the specificity investigation are as follows Figures 12-14 As shown, Figures 12-14 They are the chromatograms of blank plasma containing internal standard solution, blank plasma with 12 active ingredients and internal standard solution added, and plasma 0.5 hours after intragastric administration of Fuke Qianjin Capsule.

[0146] The linear range, standard curve, correlation coefficient, precision, accuracy, recovery rate, matrix effect and stability of 12 active ingredients in plasma and different tissues (heart tissue, liver tissue, spleen tissue, lung tissue, kidney tissue, brain tissue, uterine tissue) are shown in the figure below. Figures 15 to 22 shown.

[0147] Figures 12-14 and Figures 15 to 22 This indicates that the 12 active ingredients are not significantly interfered by endogenous substances. The 12 active ingredients in blank plasma and blank tissue have a good linear relationship within the linear range, and the correlation coefficient (R 2 )>0.99. The RSD% of accuracy, precision, matrix effect and extraction recovery was <20%. Meanwhile, the intra-day accuracy was between 84.35% and 118.92%, and the inter-day accuracy was between 82.82% and 118.82%. The extraction recovery and matrix effect of the analytes at low, medium and high QC concentrations were 83.5%-117.37% and 83.83%-119.25%, respectively, and the RSD% of room temperature, freeze-thaw and long-term stability was <20%. The above results show that the method has good applicability.

[0148] (6) Pharmacokinetic data processing

[0149] Referring to the sample acquisition method (1) in Example 1, about 400 μL of blood samples were collected from the eye sockets of rats at various time points (0.083 h, 0.16 h, 0.33 h, 0.5 h, 0.75 h, 1.0 h, 1.5 h, 2 h, 3 h, 4 h, 6 h, 8 h, 12 h, 24 h) after administration of the drug (Gynecological Qianjin Capsule). Referring to the sample acquisition method (1) in Example 1, various tissues were collected at various time points (0.5 h, 1 h, 4 h, 8 h and 16 h) after administration of the drug (Gynecological Qianjin Capsule). The ultra-high performance liquid chromatography-mass spectrometry detection and analysis method established in Example 1 was used to determine the blood drug concentrations of the 12 active ingredients at various time points. All raw data were processed on the MassLynx v4.1 workstation. The non-compartmental model analysis method in the DAS2.0 data processing software was used to fit the pharmacokinetic parameters.

[0150] Fig.23 This is a curve diagram of the concentration of the active ingredients in the blood and time after taking Fuke Qianjin Capsule. After processing by DAS2.0 software, the relevant pharmacokinetic parameters are shown in Fig.24 After the Fuke Qianjin Capsule was administered to rats by gavage, the distribution of the same active ingredient in different organs at different time points is shown in the rectangular diagram. Fig.25 .

[0151] Depend on Figures 23 to 25 It can be seen that the t of 12 active ingredients in plasma 1 / 2 0.54-13.22h, T max 0.08-7.72h, C max 0.60-668.79 ng / mL, AUC 0-t The results showed that all 12 active ingredients were rapidly absorbed and their plasma concentrations could be measured after 5 minutes. The drug concentrations of most compounds except for formononetin in plasma reached T within 1 hour. max , which means that these ingredients are quickly absorbed in the body, stay in the body for a short time, and are quickly eliminated. max The gastrointestinal absorption rate of formononetin was lower than that of other drugs after 7.72±4.34h. 1 / 2 The values ​​were 10.87±9.48h, 11.48±7.14h, 13.22±8.89h and 13.21±13.9h respectively. The results showed that these compounds were eliminated slowly in plasma and could still be detected until 10 hours after administration. In addition, some compounds (genistein, jatrorrhizine, berberine, genistein and Z-ligustrin lactone) showed a double peak phenomenon, which may be due to the pH conversion of enterohepatic circulation or different absorption areas of the intestine. This phenomenon is also conducive to prolonging the retention time of the drug in the body and maintaining the efficacy.

[0152] According to the experimental results of pharmacokinetic, 0.5h, 1h, 4h, 8h and 16h were selected for tissue distribution study, including distribution phase, equilibrium phase and elimination phase. After 0.5h and 1h of administration, the intake of drug components in organs with rich blood flow (including liver, kidney and lung) was high. Among them, phthalide compounds (Z-ligustilide and ligustilide A) and flavonoids (formononetin and formononetin) were rapidly distributed and highly enriched in the liver after 0.5h and 1h of Fuke Qianjin Capsule administration, followed by kidney, lung, spleen and uterus. Alkaloid compounds (jatrorrhizine, berberine and bamipine) and andrographolide were highly present in the lung, followed by liver and kidney, indicating that lung, liver and kidney may be the target organs of these alkaloids. In addition, genistein and dehydroandrographolide were mainly distributed in the kidney. The distribution of these components in rats reflects the characteristics of multi-component, multi-target and multi-action mechanism of traditional Chinese medicine to a certain extent.

[0153] The foregoing examples are merely illustrative and are used to explain some features of the method of the present invention. The appended claims are intended to require the widest possible range that can be imagined, and the embodiments presented herein are demonstrated by the applicant's actual test results. Therefore, the applicant's intention is that the appended claims are not limited by the selection of examples that illustrate the features of the present invention. Some numerical ranges used in the claims also include sub-ranges therein, and changes in these ranges should also be interpreted as being covered by the appended claims where possible.

Claims

1. A system for simultaneously detecting multiple characteristic active ingredients in a drug, characterized in that: include: Sample pre-treatment module and detection module; The detection module includes an ultra-high performance liquid chromatography-mass spectrometry device, and the detection module performs ultra-high performance liquid chromatography-mass spectrometry detection on the test solution to obtain the compound content of multiple characteristic medicinal ingredients in the test solution; The detection conditions of the ultra-high performance liquid chromatography are as follows: chromatographic column: ACQUITY UPLC BEHC18, the specifications of the chromatographic column are 2.1 mm×100 mm, 1.7 μm; the flow rate is 0.3-0.4 mL / min; the column temperature is 30-45° C.; the injection volume is 4 uL; the mobile phase is composed of phase A and phase B, phase A is acetonitrile, and phase B is 0.1% formic acid water; At the beginning of gradient elution, 0-0.5 min, 95% phase B; 0.5-10min, 95%-70% phase B; 10-15 min, 70%-50% phase B; 15-20 min, 50%-10% phase B; 20-22 min, 10% phase B; 22min~22.1min, 10%~95% phase B; 22.1min~24min, 95% phase B; The mass spectrometry detection conditions are as follows: acquisition in the positive ion mode using the multiple reaction monitoring mode, the capillary voltage is 1.5-2KV, the cone voltage is 20V, the ion source temperature is 150°C, the desolvation gas temperature is 400-500°C, the desolvation gas flow rate is 800-1000L / h, and the cone gas flow rate is 150L / h; The drug contains a variety of characteristic pharmacological ingredients including genistein, formononetin, andrographolide, palmatine, magnolamine, jatrorrhizine, berberine, dehydroandrographolide, genistein, formononetin, ligustilide A and Z-ligustilide.

2. The detection system according to claim 1, characterized in that: The sample of the test solution is selected from one or more of the drug, isolated blood of an animal that has taken the drug, or isolated tissue of an animal that has taken the drug; The detection module further comprises ultra-high performance liquid chromatography-mass spectrometry detection of a reference solution, wherein the sample of the reference solution is selected from a standard solution, blood or tissue of a blank control animal containing a standard solution, and the blank control animal does not take the drug; The tissue comprises one or more of heart tissue, liver tissue, spleen tissue, lung tissue, kidney tissue, brain tissue and uterine tissue.

3. The detection system according to claim 2, characterized in that: When the test solution or the reference solution is the drug, the pretreatment module includes an ultrasonic unit and a centrifugal unit; the ultrasonic unit includes an ultrasonic component, and the centrifugal unit includes a centrifugal component; the operation method of using the sample pretreatment module is: dissolving the drug in water, ultrasonicating with an ultrasonic component, and then centrifuging in a centrifugal component to obtain the test solution or the reference solution; Or, when the sample of the test solution or the reference solution is blood, the sample pretreatment module includes an anticoagulation unit, a centrifugal unit, a protein precipitation unit, and a protein removal unit; the anticoagulation unit includes an anticoagulant, the centrifugal unit includes a centrifugal component, the protein precipitation unit includes a protein precipitant containing an internal standard solution, and the protein removal unit includes a vortex mixing component, a centrifugal component, an ultrasonic component, and a protein precipitant; The operation method of using the sample pretreatment module is as follows: blood is mixed with an anticoagulant, and then input into a centrifugal component for centrifugation to obtain a plasma sample; the plasma sample is mixed with a protein precipitant containing an internal standard solution, and then input into a vortex mixing component and a centrifugal component in sequence for vortex mixing and centrifugation to obtain a residue; the residue is mixed with a protein precipitant, and then input into a vortex mixing component, an ultrasonic component, and a centrifugal component again for vortex mixing, ultrasonication, and centrifugation to obtain the test solution or the reference solution; Or, when the sample of the test solution or the reference solution is tissue, the pre-treatment module includes a homogenization unit, a protein precipitation unit, a centrifugal unit, a vortex mixing unit, and an ultrasonic unit; the homogenization unit includes a homogenization component, the protein precipitation unit includes a protein precipitant containing an internal standard solution, the centrifugal unit includes a centrifugal component, the vortex mixing unit includes a vortex mixing component, and the ultrasonic unit includes an ultrasonic component; The operation method of using the sample pretreatment module is as follows: homogenizing the tissue in a homogenization component, then mixing it with a protein precipitant containing an internal standard solution, then homogenizing it using the homogenization component and centrifuging it using the centrifugation component, taking the supernatant after centrifugation and mixing it with the protein precipitant, then vortexing, sonicating, and centrifuging in a vortex mixing component, an ultrasonic component, and a centrifugation component in sequence to obtain the test solution or the reference solution.

4. The detection system according to claim 2 or 3, characterized in that: The standard solution is a methanol solution of the characteristic medicinal ingredient.

5. The detection system according to claim 3, characterized in that: The protein precipitant is selected from one or more of acetonitrile, methanol or ethanol.

6. The detection system according to claim 3, characterized in that: When the sample is blood, the volume ratio of plasma to protein precipitant is 1:(2-6); when the sample is tissue, the mass volume ratio of tissue to protein precipitant is 1:(2-6) mg / mL.

7. The detection system according to claim 6, characterized in that: When the sample is blood, the protein precipitant is selected from acetonitrile; when the sample is tissue, the protein precipitant is selected from methanol.

8. The detection system according to claim 3, characterized in that: The internal standard solution is a mixed solution of psoralen and methanol.

9. A method for analyzing the pharmacokinetic properties of multiple characteristic pharmacodynamic components in a drug, wherein the drug contains multiple characteristic pharmacodynamic components, including: Genistin, formononetin, andrographolide, palmatine, magnolamine, jatrorrhizine, berberine, dehydroandrographolide, genistein, formononetin, ligustilide A and Z-ligustilide; characterized in that the pharmacokinetic analysis method includes a sample pretreatment step and a detection step; S1. Sample pretreatment: taking samples for pretreatment to obtain test sample solutions; the samples are selected from animal samples collected at different time points, isolated from the body, and taken with the drug, the animal samples include animal blood and animal tissues; the animal tissues include one or more of heart tissue, liver tissue, spleen tissue, lung tissue, kidney tissue, brain tissue and uterine tissue; S2. Detection: The test solution is tested using ultra-high performance liquid chromatography-mass spectrometry to obtain the compound content of multiple characteristic active ingredients in the test solution; The detection conditions of the ultra-high performance liquid chromatography are as follows: chromatographic column: ACQUITY UPLC BEHC18, the specifications of the chromatographic column are 2.1 mm×100 mm, 1.7 μm; the flow rate is 0.3-0.4 mL / min; the column temperature is 30-45° C.; the injection volume is 4 uL; the mobile phase is composed of phase A and phase B, phase A is acetonitrile, and phase B is 0.1% formic acid water; At the beginning of gradient elution, 0-0.5 min, 95% phase B; 0.5-10min, 95%-70% phase B; 10-15 min, 70%-50% phase B; 15-20 min, 50%-10% phase B; 20-22 min, 10% phase B; 22min~22.1min, 10%~95% phase B; 22.1min~24min, 95% phase B; The detection conditions of the mass spectrometer are: acquisition in multiple reaction monitoring mode in positive ion mode, capillary voltage of 1.5-2KV, cone voltage of 20V, ion source temperature of 150°C, desolvation gas temperature of 400-500°C, desolvation gas flow rate of 800-1000L / h, and cone gas flow rate of 150L / h.

10. Application of the detection system or method according to any one of claims 1 to 9 in the detection and quality control of drug ingredients related to Fu Ke Qian Jin Fang, or in the screening and determination of medication regimens and dosages, or in drug development.

Images