Determination method for contents of 17 chemical components in dry extract of Qingxiaoku drink

Abstract

The invention discloses a method for determining the contents of 17 chemical components in dry extract of Qingxiaoqiao drink, which adopts a UHPLC-MS / MS (Ultra High Performance Liquid Chromatography-Mass Spectrometry / Mass Spectrometry) method to simultaneously determine the contents of the 17 chemical components in the dry extract of Qingxiaoqiao drink. The method comprises the following steps: (1) establishing a standard curve of 17 chemical components; (2) acquiring selective ion detection chromatograms of 17 chemical components in the dry extract of the Qingxiaoku drink; and (3) determining the contents of the 17 chemical components in the dry extract of the Qingxiaodrink for children. The method disclosed by the invention is simple to operate, high in sensitivity, strong in specificity and easy to analyze and evaluate. According to the method, the contents of 17 chemical components in the dry extract of the Qingxiaoku drink are measured, the quality of the Qingxiaoku drink can be controlled, and the safety, effectiveness and stability of clinical medication of the Qingxiaoku drink are ensured.

Description

Technical Field

[0001] The invention relates to the field of quality analysis of traditional Chinese medicines, and in particular to a method for determining the contents of 17 chemical components in a Qinggan Tongyin dry extract. Background Art

[0002] Qinggan Tongyin is mainly used to prevent and treat respiratory infectious diseases, with significant clinical effects. The formula consists of ten medicinal materials, including burdock seed, coix seed, platycodon, red peony root, perilla leaf, forsythia, charred hawthorn, licorice, chrysanthemum, and mint. Burdock seed and forsythia are the main medicines. Burdock seed is pungent and cold in nature, and is good at dispersing wind-heat, promoting lung function and clearing rash, and detoxifying and relieving sore throat. Forsythia is bitter and slightly cold in nature, and is good at clearing heat and detoxifying, and dispersing wind-heat. It is combined with the pungent and warm perilla to strengthen the evacuation of wind and cold. Platycodon opens and promotes lung qi to help the evil to be resolved from the surface. Together, they play the role of promoting lung qi, moistening the lungs and promoting fluid production, dispersing wind and resisting anti-toxicity. Licorice and coix seed strengthen the spleen and eliminate dampness to help transportation and transformation. Chrysanthemum, mint, and red peony root clear away heat and detoxify. Summary of the invention

[0003] The purpose of the present invention is to provide a method for determining the contents of 17 chemical components in the Qinggan Tongyin dry extract, so as to achieve more reliable and comprehensive quality control of the Qinggan Tongyin extract. The specific technical scheme is as follows:

[0004] A method for determining the content of chemical components in a Qinggan Tongyin extract, using a UHPLC-MS / MS method, and simultaneously determining the content of chemical components in the Qinggan Tongyin extract, wherein the chemical components include arctigenin, forsythiaside A, arctiin, baicalin, hyperoside, forsythiaside, platycodonin D, rosmarinic acid, caffeic acid, luteolin, apigenin, quercetin, chlorogenic acid, gallic acid, paeoniflorin, catechins, and paeoniflorin. The method comprises:

[0005] (1) Establishment of standard curves for 17 chemical components in Qinggan Tongyin extract

[0006] Accurately weigh the reference substances arctigenin, forsythiaside A, arctiin, baicalin, hyperoside, forsythiaside, platycodonoside D, rosmarinic acid, caffeic acid, luteolin, apigenin, quercetin, chlorogenic acid, gallic acid, paeoniflorin, catechin, and paeoniflorin, and add methanol to make up to volume to prepare stock solutions of each reference substance with known concentrations, and store at 4°C for future use.

[0007] Diluting the reference substance mixed stock solution with methanol having a volume fraction of 70-100% to form a series of reference substance mixed solutions having 17 chemical components with different known concentrations;

[0008] Under the same preset chromatographic conditions and mass spectrometry conditions, the mixed solutions of the reference substances of different concentrations were respectively injected into the ultra-high performance liquid chromatography-mass spectrometry instrument to obtain the multi-reaction ion chromatograms of each reference substance at different concentrations;

[0009] The standard curve of each chemical component was obtained by linear regression calculation, with the peak area y of the chromatographic peak of each reference substance as the ordinate and the concentration x of each reference substance as the abscissa;

[0010] (2) Obtaining the multiple reaction ion monitoring chromatograms of 17 chemical components in Qinggan Tongyin extract

[0011] Accurately weigh Qinggan Tongyin, add 70% methanol to the fixed volume, extract by ultrasonic, take out and cool to room temperature, make up for the weight loss. Take the supernatant to extract the drug solution, filter it with a 0.22μm filter membrane, dilute it with methanol, and store it at 4℃ for testing.

[0012] Under the same preset chromatographic conditions and mass spectrometry conditions as in step (1) above, a volume V1 of the test solution was injected into an ultra-high performance liquid chromatography-mass spectrometer to obtain a multiple reaction ion monitoring chromatogram of each chemical component in the Qinggan Tongyin extract;

[0013] (3) Determine the content of 17 chemical components in Qinggan Tongyin extract

[0014] The content of each chemical component in Qinggan Tongyin extract was determined based on the peak area of ​​the multiple reaction ion monitoring chromatogram of each chemical component in Qinggan Tongyin extract and the standard curve of each chemical component.

[0015] In a specific embodiment of the present invention, the determination of the content of each chemical component in the Qinggan Tongyin extract comprises:

[0016] The concentration C corresponding to the peak area of ​​each chemical component in Qinggan Tongyin extract was determined from the standard curve of each chemical component, and the contents of 17 chemical components in the test sample were calculated according to the following formulas;

[0017]

[0018] In a specific embodiment of the present invention, the standard curve of each chemical component is obtained by linear regression operation, specifically:

[0019] The standard curve is obtained by linear regression calculation using weighted least square method, and the weight coefficient is 1 / X. In a specific embodiment of the present invention, the preset chromatographic conditions include:

[0020] Chromatographic column: octadecylsilane bonded silica gel as the stationary phase; mobile phase: phase A: water, phase B: acetonitrile; gradient elution method; column temperature: 25-40°C; flow rate: 0.2-0.5mL / min; injection volume: 2-5μL.

[0021] In a specific embodiment of the present invention, the preset chromatographic conditions include:

[0022] Chromatographic column: Mobile phase: Phase A is water, phase B is acetonitrile; gradient elution method is adopted; column temperature: 40°C; flow rate: 0.2mL / min; injection volume: 3μL.

[0023] The elution program was: 0-2 min, 20% B; 2-3 min, 20-26% B; 3-5 min, 26-34% B; 5-7 min, 34-56% B; 7-10 min, 56-78% B; 10-15 min, 78-80% B; 15-15.10 min, 80-20% B; 15.10-18.10 min, 20% B.

[0024] In a specific embodiment of the present invention, the preset mass spectrometry conditions include:

[0025] Ionization mode: electrospray ion source (ESI); multiple reaction ion monitoring mode, positive ion scanning, negative ion scanning; drying gas temperature: 400-500°C; drying gas flow rate 7-15L / min; spray gas pressure 5-20psi; drying gas and nebulizing gas: nitrogen.

[0026] In a specific embodiment of the present invention, the preset mass spectrometry conditions include:

[0027] Ionization method: electrospray ion source (ESI); mass spectrometry multiple reaction monitoring (MRM) mode and positive and negative ion fast switching mode are used for monitoring; ion source temperature: 500°C; drying gas flow rate: 10L / min; spray gas pressure: 10psi; drying gas and nebulizing gas: nitrogen.

[0028] In a specific embodiment of the present invention, the mass spectrometry parameters are as follows:

[0029]

[0030]

[0031] The embodiments of the present invention have the following advantages:

[0032] The present invention adopts UHPLC-MS / MS method to simultaneously determine the contents of 17 chemical components in Qinggan Tongyin extract, which has the characteristics of high sensitivity, strong reliability, high efficiency and rapidity. By determining the contents of 17 chemical components in Qinggan Tongyin extract, the quality of Qinggan Tongyin can be more effectively controlled, thereby ensuring the clinical efficacy of Qinggan Tongyin. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] In order to more clearly illustrate the implementation methods of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for the implementation methods or the description of the prior art. Obviously, the drawings in the following description are only exemplary, and for ordinary technicians in this field, other implementation drawings can be derived from the provided drawings without creative work.

[0034] The structures, proportions, sizes, etc. illustrated in this specification are only used to match the contents disclosed in the specification so as to facilitate understanding and reading by persons familiar with the technology. They are not used to limit the conditions under which the present invention can be implemented, and therefore have no substantial technical significance. Any structural modification, change in proportion or adjustment of size shall still fall within the scope of the technical contents disclosed in the present invention without affecting the effects and purposes that can be achieved by the present invention.

[0035] Figure 1 Figure A is the multiple reaction ion monitoring chromatogram of 9 chemical component reference substances at a certain concentration.

[0036] Figure 1 Figure B is the multi-reaction ion detection chromatogram of the chemical components in Qinggan Tongyin extract;

[0037] Figure 1 Figure C in the figure is the multi-reaction ion detection chromatogram of the blank solution;

[0038] Figure 1 In the figure, each number represents:

[0039] 1. Arctigenin; 2. Forsythiaside A; 3. Arctiin; 4. Baicalin; 5. Hyperoside; 6. Forsythiaside;

[0040] 7. Platycodon saponin D; 8. Rosmarinic acid; 9. Caffeic acid;

[0041] Figure 2 Figure A in the figure is the multiple reaction ion monitoring chromatogram of 8 chemical component reference substances at a certain concentration.

[0042] Figure 2 Figure B is the multi-reaction ion detection chromatogram of the chemical components in Qinggan Tongyin extract;

[0043] Figure 2 Figure C in the figure is the multi-reaction ion detection chromatogram of the blank solution;

[0044] Figure 2 In the figure, each number represents:

[0045] 10. Luteolin; 11. Apigenin; 12. Quercetin; 13. Chlorogenic acid; 14. Paeoniflorin; 15. Gallic acid; 16. Catechin; 17. Peony lactone glycosides. DETAILED DESCRIPTION

[0046] The present invention adopts UHPLC-MS / MS method (ultra-high performance liquid chromatography-tandem mass spectrometry) to simultaneously determine the contents of 17 chemical components in Qinggan Tongyin extract.

[0047] The technical solution of the present invention will be clearly and completely described below in conjunction with specific embodiments. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0048] First of all, it should be noted that the term "precision weighing" mentioned in the present invention means that the weighing weight should be accurate to one thousandth of the weight taken. "Precision suction" means that the accuracy of the measured volume should meet the accuracy requirements of the measuring instrument in the national standard.

[0049] 1. Instruments, reagents and materials

[0050] 1.1 Instrument

[0051] 4500 LC / MS system, including Shimadzu UHPLC system (Shimadzu Corporation, Kyoto, Japan), AB Sciex 4500 triple quadrupole mass spectrometer and application software Analyst software version 1.7 and OS version WIN 7 (AB Sciex). SORVALL LEGEND MICRO21R high-speed refrigerated centrifuge (Thermo Scientific); Avanti JE desktop centrifuge (Beckman); SVANTSPD2010 vacuum concentrator (Thermo Electron); BS210S electronic balance (Sartorius); Milli-Q automatic water purifier (Millipore); -20℃ refrigerator (Qingdao Haier Company); Forma-80℃ ultra-low temperature refrigerator (Thermo Electron); XW-80A vortex mixer (Shanghai Medical University Instrument Factory); MX-S mixer (Beijing Dalong Xingchuang Experimental Instrument Co., Ltd.).

[0052] 1.2 Reagents

[0053] Reference substances arctigenin, forsythiaside A, arctiin, baicalin, hyperoside, forsythiaside, platycodonin D, rosmarinic acid, luteolin, apigenin, quercetin and catechins were purchased from Chengdu Glip Biotechnology Co., Ltd. Chlorogenic acid, caffeic acid, gallic acid, paeoniflorin and paeoniflorin were purchased from Chengdu Pufede Biotechnology Co., Ltd. Methanol (chromatographic grade) and acetonitrile (chromatographic grade) were purchased from Merck, Germany.

[0054] 1.3 Materials

[0055] Qinggan Tongyin dry extract was provided by Baokang Hospital of Tianjin University of Traditional Chinese Medicine, batch number: 20072101.

[0056] 2. Chromatographic conditions and mass spectrometry conditions

[0057] 2.1 Chromatographic conditions Chromatographic column: Ultimate XB-C18 (2.1×100mm, 3μm); mobile phase: water (A): acetonitrile (B), gradient elution. 0-2min, 20% B; 2-3min, 20-26% B; 3-5min, 26-34% B; 5-7min, 34-56% B; 7-10min, 56-78% B; 10-15min, 78-80% B; 15-15.10min, 80-20% B; 15.10-18.10min, 20% B; flow rate: 0.2mL / min, column temperature: 40℃; injection volume: 3μL.

[0058] 2.2 Mass spectrometry conditions Ionization mode: Ionization mode: electrospray ionization source (ESI); mass spectrometry multiple reaction monitoring (MRM) mode and positive and negative ion rapid switching mode were used for monitoring; ion source temperature: 500°C; drying gas flow rate: 10L / min; spray gas pressure: 10psi; drying gas and nebulizing gas: nitrogen.

[0059] Table 1 Mass spectrometry parameters of 17 chemical components in Qinggan Tongyin dry extract

[0060]

[0061]

[0062] Solution preparation

[0063] 2.3.1 Preparation of reference solution

[0064] Accurately weigh 1 mg of each of the reference substances arctigenin, forsythiaside A, arctiin, baicalin, hyperoside, forsythiaside, platycodonoside D, rosmarinic acid, caffeic acid, luteolin, apigenin, quercetin, chlorogenic acid, gallic acid, paeoniflorin, catechin, and paeoniflorin, add methanol to make the volume to 1 ml, and prepare a 1 mg / mL reference substance stock solution, which are all stored in a 4°C refrigerator for future use.

[0065] 2.3.2 Preparation of test solution

[0066] Weigh 0.1g Qinggan Tongyin (5 batches, S1-S5) accurately, add 70% methanol to make up the volume, extract by ultrasonic, take out and cool to room temperature, make up the weight loss. Take the supernatant to extract the drug solution, filter it with a 0.22μm filter membrane, dilute it with methanol, and store it at 4℃ for testing.

[0067] 2.4 Methodological Review

[0068] 2.4.1 Standard curve, detection limit, and quantification limit

[0069] Accurately measure 50 μL of the series of working solutions of each component under "2.3.1" and mix them evenly to obtain a series of mixed working solutions including all 17 test components, namely the mixed standard solution. Accurately pipette 100 μL of each concentration of mixed standard solution, process according to the steps of "2.3.2", and then perform sample injection and detection according to the chromatographic conditions described in "2.1" and the mass spectrometry conditions under "2.2". The concentration gradients of arctigenin, hyperoside, and caffeic acid were 2, 5, 10, 20, 50, 100, 200, and 500 ng / mL; the concentration gradient of forsythiaside A was 2, 10, 50, 100, 500, 1000, 4000, and 8000 ng / mL; the concentration gradients of arctiin, chlorogenic acid, and paeoniflorin were 5, 10, 50, 100, 500, 1000, 2000, and 5000 ng / mL; The concentration gradient of baicalin, forsythin, rosmarinic acid, and paeoniflorin was 5, 10, 50, 100, 200, 500, 1000, 2000 ng / mL; the concentration gradient of platycodon saponin D and quercetin was 1, 2, 5, 10, 20, 50, 100, 200 ng / mL; the concentration gradient of luteolin, apigenin, gallic acid, and catechin was 2, 5, 10, 20, 50, 100, 200, 500 ng / mL. The chromatographic peak area was taken as the ordinate (y), the reference concentration was taken as the abscissa (x), and the detection limit (LOD, S / N=3) and the quantification limit (LOQ, S / N=10) were calculated by the ratio of each component to the surrounding baseline noise (S / N). All components had good linearity within the corresponding linear range, and r was greater than 0.995. The results are shown in Table 2.

[0070] Table 2 Regression equations, quantification limits and detection limits of standard curves of 17 chemical components

[0071]

[0072]

[0073] 2.4.2 Precision test

[0074] Take 100 μL of the test solution (S1) prepared under "2.3.2", inject it 5 times continuously according to the chromatographic conditions under "2.1" and the mass spectrometry conditions under "2.2", record the chromatogram, and conduct precision investigation. The RSD of the peak area of ​​each peak is not higher than 2.87%. The experimental results show that the precision of this method is good, see Table 3 for details.

[0075] 2.4.3 Repeatability test

[0076] Take 100 μL of the same test solution (S1), process it according to the steps of "2.3.2", prepare 5 copies in parallel, record the peak area of ​​each component according to the chromatographic conditions under "2.1" and the mass spectrometry conditions under "2.2", and calculate the RSD value. The RSD value of the peak area of ​​each peak is not higher than 2.85%. The results show that the method has good repeatability, see Table 3 for details.

[0077] 2.4.4 Stability test

[0078] Take 100 μL of the same sample solution (S1), process it according to the steps of "2.3.2", prepare 5 copies in parallel, and analyze the samples at 0, 6, and 24 hours according to the chromatographic conditions under "2.1" and the mass spectrometry conditions under "2.2", and record the chromatogram. The RSD value of the peak area of ​​each peak is not higher than 2.87%. The results show that the sample has good stability within 24 hours, see Table 3 for details.

[0079] Sample recovery test

[0080] Accurately measure the known content of Qinggan Tongyin test solution (S1), add equal amounts of each component and the sample concentration of the mixed reference solution, mix well and accurately pipette 100 μL, prepare 5 copies in parallel according to "2.3.2", inject and detect according to the chromatographic conditions under "2.1" and the mass spectrometry conditions under "2.2", record the peak area of ​​each component, and calculate the RSD value. The results show that the sample recovery rate of 17 components is 97.09% to 102.96%, see Table 3 for details.

[0081] Table 3 Methodological investigation results of 17 components in Qinggan Tongyin dry extract

[0082]

[0083]

[0084] 2.5 Sample determination

[0085] The Qinggan Tongyin dry extract was used to prepare a backup test solution in parallel according to the method under "2.3.2", and the test was carried out according to the chromatographic conditions under "2.1" and the mass spectrometry conditions under "2.2". The concentration C corresponding to the peak area of ​​each chemical component in the Qinggan Tongyin dry extract was determined from the standard curve of each chemical component in Table 2, and the contents of 17 chemical components in the test sample were calculated according to the following formulas. The results are shown in Table 4.

[0086] The content of 5 batches of Qinggan Tongyin was measured, among which the ingredients with higher content were forsythiaside A, arctiin, chlorogenic acid, baicalin, gallic acid, rosmarinic acid, paeoniflorin, and forsythiaside.

[0087] Table 4 Contents of 17 chemical components in Qinggan Tongyin dry extract (μg / g)

[0088]

[0089]

[0090] The present invention adopts UHPLC-MS / MS method to simultaneously determine the contents of 17 chemical components in Qinggan Tongyin, which has the characteristics of high sensitivity, strong reliability, high efficiency and rapidity. By determining the contents of 17 chemical components in Qinggan Tongyin, the quality of Qinggan Tongyin can be more effectively controlled, and the contents of the main components in Qinggan Tongyin can be explored, thereby ensuring the clinical efficacy of Qinggan Tongyin.

[0091] Although the present invention has been described in detail above by general description and specific embodiments, it is obvious to those skilled in the art that some modifications or improvements can be made to the present invention. Therefore, these modifications or improvements made without departing from the spirit of the present invention all belong to the scope of protection claimed by the present invention.

[0092] The terms such as "upper", "lower", "left", "right", "middle", etc. cited in this specification are only for the convenience of description and are not intended to limit the scope of implementation of the present invention. Changes or adjustments to their relative relationships should be regarded as within the scope of implementation of the present invention without substantially changing the technical content.

Claims

1. A method for determining the contents of 17 chemical components in the dry extract of Qinggan Tongyin, characterized in that: The contents of 17 chemical components in the dry extract of Qinggan Tongyin were simultaneously determined by UHPLC-MS / MS method.

2. The method for determining the contents of 17 chemical components in the Qinggan Tongyin dry extract according to claim 1, characterized in that: The 17 chemical components include: 1 arctigenin, 2 forsythiaside A, 3 arctiin, 4 baicalin, 5 hyperoside, 6 forsythiaside, 7 platycodonosposide D, 8 rosmarinic acid, 9 caffeic acid, 10 luteolin, 11 apigenin, 12 quercetin, 13 chlorogenic acid, 14 gallic acid, 15 paeoniflorin, 16 catechins, and 17 paeoniflorin.

3. The method for determining the contents of 17 chemical components in the Qinggan Tongyin dry extract according to claim 1, characterized in that: The following steps are involved: Step 1: Establishing the standard curves of 17 chemical components, the method is as follows: accurately weighing 17 chemical component reference substances respectively and dissolving them in methanol to prepare reference substance stock solutions with known concentrations, mixing, diluting with methanol to prepare reference substance mixed stock solutions with known concentrations of 17 chemical components, then diluting the reference substance mixed stock solutions with a volume fraction of 70%-100% methanol into a series of reference substance mixed solutions with different known concentrations of 17 chemical components, respectively injecting V1 volumes of reference substance mixed solutions of various concentrations into an ultra-high performance liquid chromatography-mass spectrometry instrument, obtaining multiple reaction ion monitoring chromatograms of the reference substances at different concentrations, then using the peak area of ​​the chromatogram of each reference substance as the coordinate axis (y) and the concentration of each reference substance as the horizontal axis (x), obtaining the standard curve of each chemical component by linear regression operation; Step 2: obtaining the multi-reaction ion monitoring chromatograms of 17 chemical components in the dry extract of Qinggan Tongyin, the method is: dissolving the dry extract of Qinggan Tongyin with a mass M into a volume of V2 with 70% by volume of methanol, ultrasonically extracting for 30 minutes, filtering the extract through a membrane as a test solution, and then injecting a volume of V1 of the test solution into an ultra-high performance liquid chromatography-mass spectrometer to obtain the multi-reaction ion monitoring chromatograms of the chemical components in the dry extract of Qinggan Tongyin; Step three: Determine the content of 17 chemical components in the dry extract of Qinggan Tongyin. The method is: determine the content of each chemical component in the extract of the dry extract of Qinggan Tongyin based on the peak area of ​​the multi-reaction ion chromatogram of each chemical component in the dry extract of Qinggan Tongyin and the annotation curve of each chemical component.

4. The method for determining the contents of 17 chemical components in the Qinggan Tongyin dry extract according to claim 1, characterized in that: The method of determining the content of each chemical component in the Qinggan Tongyin dry extract comprises: determining the concentration C corresponding to the peak area of ​​each chemical component in the Qinggan Tongyin dry extract from the standard curve of each chemical component, and calculating the content of 17 chemical components in the test sample according to the following formulas; 5. The method for determining the contents of 17 chemical components in the Qinggan Tongyin dry extract according to claim 1, characterized in that: The standard curve of each chemical component is obtained by linear regression operation. Specifically, the standard curve is obtained by linear regression operation using weighted least square method, and the weight coefficient is 1 / X.

6. The method for determining the contents of 17 chemical components in the Qinggan Tongyin dry extract according to claim 1, characterized in that: The preset chromatographic conditions include: the chromatographic column is an octadecylsilane bonded silica gel stationary phase; the mobile phase: phase A is water, phase B is acetonitrile; the column temperature: 40°C; the flow rate: 0.2mL / min; the injection volume is 3μL; the gradient elution method is adopted, and the elution program is: 0-2min, 20% B; 2-3min, 20-26% B; 3-5min, 26-34% B; 5-7min, 34-56% B; 7-10min, 56-78% B; 10-15min, 78-80% B; 15-15.10min, 80-20% B; 15.10-18.10min, 20% B.

7. The method for determining the contents of 17 chemical components in the Qinggan Tongyin dry extract according to claim 1, characterized in that the preset mass spectrometry conditions include: Ionization mode: electrospray ionization; multiple reaction ion monitoring mode: positive ion scanning and negative ion scanning; drying gas temperature: 500°C; drying gas flow rate: 10L / min; spray gas pressure: 10psi; drying gas and nebulizing gas: nitrogen; ionization mode: electrospray ionization.

8. The method for determining the contents of 17 chemical components in the Qinggan Tongyin dry extract according to claim 1, characterized in that: The mass spectrometry parameters are as follows: The compounds in negative ion mode are:

1. Arctigenin, parent ion is 371.2m / z, daughter ion is 136.0m / z, collision voltage is -122.75V, collision energy is -38.58V; 2. Arctiin, parent ion is 579.1m / z, daughter ion is 371m / z, collision voltage is -34.86V, collision energy is -22.98V; 3. Scutellarin, parent ion is 463.2m / z, daughter ion is 285.1m / z, collision voltage is -115.79V, collision energy is -29.42V; 4. Hyperoside, parent ion is 463.2m / z, daughter ion is 300.0m / z, collision voltage is -153.68V, collision energy is -41.25V; 5. Platycodon saponin D, parent ion is 1223.6m / z, daughter ion is 469.1m / z, collision voltage is -200.31V, collision energy is -75.85V; 6. Rosmarinic acid, parent ion is 359.1 m / z, daughter ion is 161.1 m / z, collision voltage is -89.21 V, collision energy is -21.00 V; 7. Caffeic acid, parent ion is 179.0m / z, daughter ion is 135.0m / z, collision voltage is -60.01V, collision energy is -26.79V; 8. Luteolin, parent ion is 463.2m / z, daughter ion is 300.0m / z, collision voltage is -153.68V, collision energy is -41.25V; 9. Apigenin, parent ion is 269.0m / z, daughter ion is 117.2m / z, collision voltage is -122.67V, collision energy is -48.90V; 10. Quercetin, parent ion is 301.2m / z, daughter ion is 151.2m / z, collision voltage is -113.18V, collision energy is -29.86V; 11. Chlorogenic acid, parent ion is 191.0 m / z, daughter ion is 127.0 m / z, collision voltage is -126.20 V, collision energy is -25.10 V; 12. Paeoniflorin, parent ion is -525.4m / z, daughter ion is 449.5m / z, collision voltage is -45.63V, collision energy is -20.31V; 13. Gallic acid, parent ion is 169.0m / z, daughter ion is 125.0m / z, collision voltage is -71.40V, collision energy is -22.60V; 14. Catechin, parent ion is -289.2m / z, daughter ion is 245.2m / z, collision voltage is -107.91V, collision energy is -19.07V; 15. Paeoniflorin, parent ion is -479.2m / z, daughter ion is 121.1m / z, collision voltage is -125.80V, collision energy is -23.97V. The compounds in positive ion mode are:

1. Forsythiaside A, parent ion is 642.2m / z, daughter ion is 325.1m / z, collision voltage is 75.01V, collision energy is 23.89V; 2. Forsythiaside, parent ion is 552.3m / z, daughter ion is 249.2m / z, collision voltage is 69.79V, and collision energy is 20.50V.

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