A method for detecting the content of active ingredients in Feire Qingjie Guogan powder

Abstract

This invention discloses a method for detecting the content of active ingredients in a lung-heat clearing and detoxifying decoction, comprising the following steps: S1. Accurately weigh the reference standards of morin A, ferulic acid, p-coumaric acid, and rutin, dissolve and dilute them in methanol to prepare a mixed reference solution; S2. Accurately weigh the lung-heat clearing and detoxifying decoction sample, add solvent for extraction, filter or centrifuge the extract, and take the filtrate as the test solution; S3. Detect the sample using high-performance liquid chromatography (HPLC), with the following chromatographic conditions: a C18 column packed with octadecylsilane-bonded silica gel; a mobile phase consisting of methanol as phase A and 0.1% phosphoric acid aqueous solution as phase B, with gradient elution; a detection wavelength of 315 nm for 0-18 minutes and 257 nm for 18-40 minutes; a column temperature of 25-35℃; and a flow rate of 0.8-1.2 ppm. mL / min; S4. Accurately pipette the reference solution and the test solution separately and inject them into the high performance liquid chromatograph for determination. Qualitative identification is performed based on the retention time of the chromatographic peaks, and the content of each component is calculated using the external standard method based on the peak area.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical testing and analysis technology, specifically relating to a method for detecting the content of active ingredients in a lung heat clearing and detoxifying decoction. Background Technology

[0002] The Lung Heat Clearing Formula originates from a clinically proven prescription by veteran traditional Chinese medicine practitioners at Huangpu People's Hospital in Zhongshan City, Guangdong Province. It consists of ten Chinese herbs: mulberry bark, fritillaria bulb, reed rhizome, licorice root, earthworm, trichosanthes peel, winter melon seed, bamboo shavings, wild chrysanthemum, and plantain seed. It has the effects of clearing heat and ventilating the lungs, relieving cough and asthma, and is mainly used to treat cough due to lung heat. Clinically, it is primarily used for the treatment of acute bronchitis and lung infections. Decoction, as a special form of traditional decoction, involves pulverizing the herbs into coarse particles before decocting, which has advantages such as saving herbs, increasing the dissolution rate of active ingredients, and shortening decoction time. Developing the Lung Heat Clearing Formula into a decoction form has excellent clinical application prospects.

[0003] To effectively control the intrinsic quality of Lung-Clearing and Detoxifying Decoction and ensure the stability and consistency of its clinical efficacy, a reliable quality control method needs to be established. Traditional Chinese medicine fingerprinting technology can comprehensively reflect the overall chemical composition of the compound, while multi-index component content determination can accurately quantify key pharmacodynamic substances. In the Lung-Clearing and Detoxifying Decoction, mulberry bark is the principal ingredient, and its main active ingredient, morin A, has anti-inflammatory and anti-tumor effects; reed rhizome is the assistant ingredient, containing organic acids such as ferulic acid and p-coumaric acid, which have antibacterial, anti-inflammatory, and antioxidant activities; simultaneously, many herbs in the formula contain flavonoids such as rutin, which also have broad pharmacological activities. Currently, there are no reported methods for simultaneously determining the content of these four active ingredients in Lung-Clearing and Detoxifying Decoction. Therefore, developing a simple, accurate, reproducible, and specific detection method is essential for the quality control and widespread application of Lung-Clearing and Detoxifying Decoction. Summary of the Invention

[0004] The purpose of this invention is to address the shortcomings of existing technologies and provide a method for detecting the content of active ingredients in Lung Heat Clearing and Detoxifying Decoction. This method employs high-performance liquid chromatography (HPLC), which can simultaneously, rapidly, and accurately detect the content of four active ingredients in Lung Heat Clearing and Detoxifying Decoction: morin A, ferulic acid, p-coumaric acid, and rutin.

[0005] To achieve the above objectives, the present invention adopts the following technical solution:

[0006] A method for detecting the content of active ingredients in a lung-heat-clearing decoction includes the following steps:

[0007] S1. Preparation of reference solution

[0008] Accurately weigh the reference standards of morin A, ferulic acid, p-coumaric acid and rutin, dissolve and dilute them in methanol to prepare a mixed reference solution;

[0009] S2. Preparation of the test solution

[0010] Take the lung heat clearing and detoxifying powder as the test sample, accurately weigh it, add solvent to extract it, filter or centrifuge the resulting extract, and take the filtrate as the test sample solution.

[0011] S3. Detection was performed using high-performance liquid chromatography (HPLC). The chromatographic conditions included:

[0012] Chromatographic column: C18 column packed with octadecylsilane-bonded silica gel;

[0013] Mobile phase: Methanol as phase A and 0.1% phosphoric acid aqueous solution as phase B, gradient elution was performed;

[0014] Detection wavelength: The wavelength switching method is adopted. The detection wavelength is 315nm within 0-18 minutes and 257nm within 18-40 minutes.

[0015] Column temperature: 25-35℃;

[0016] Flow rate: 0.8-1.2 mL / min;

[0017] S4. Measurement and Calculation

[0018] Accurately pipette the reference solution and the test solution separately and inject them into the high-performance liquid chromatograph for determination. Qualitative identification is performed based on the retention time of the chromatographic peaks, and the content of each component is calculated using the external standard method based on the peak area.

[0019] Preferably, in the mixed reference solution described in step S1, the concentrations of morin A (0.0120-0.0840 μg / mL), ferulic acid (0.0072-0.0504 μg / mL), coumaric acid (0.0144-0.1008 μg / mL), and rutin (0.0112-0.0784 μg / mL) are as follows.

[0020] Preferably, the test solution in step S2 is prepared by: taking the extract of Lung Heat Clearing and Detoxifying Decoction, adding methanol, sonicating, centrifuging, and filtering the supernatant through a microporous membrane. More preferably, the volume ratio of the Lung Heat Clearing and Detoxifying Decoction extract to methanol is 1:1; the sonication power is 200W, the frequency is 50kHz, and the time is 1 min; the centrifugation speed is 10000 r / min, and the time is 15 min; the pore size of the microporous membrane is 0.22 μm or 0.45 μm.

[0021] Preferably, the gradient elution procedure in step S3 is as follows:

[0022] 0-10 min, mobile phase A is 15%, and mobile phase B is 85%;

[0023] Over 10-22 minutes, the mobile phase A decreased from 15% to 44%, and the mobile phase B decreased from 85% to 56%.

[0024] Between 22 and 26 minutes, the mobile phase A decreased from 44% to 44.5%, and the mobile phase B decreased from 56% to 55.5%.

[0025] Over 26-30 minutes, the mobile phase A decreased from 44.5% to 60%, and the mobile phase B decreased from 55.5% to 40%.

[0026] Over 30-40 minutes, the mobile phase A decreased from 60% to 15%, and the mobile phase B decreased from 40% to 85%.

[0027] Preferably, the chromatographic column in step S3 is a SHIMADZU Shim-pack GIS C18 column (4.6 mm × 250 mm, 5 μm) or a Hypersil ODS2 C18 column (250 mm × 4.6 mm, 5 μm); the column temperature is 30 °C; and the flow rate is 1.0 mL / min.

[0028] The main function of the Lung Heat Clearing and Detoxifying Decoction of this invention is to clear lung heat, supplemented by expectoration, cough relief, and asthma relief. Mulberry bark is the principal ingredient, and its main component, morin A, has anti-inflammatory and anti-lung cancer effects and is easily soluble during decoction. It also contains rutin, which has anti-inflammatory, antioxidant, anti-apoptotic, antibacterial, and antiviral pharmacological activities, and is therefore used as an indicator component. Reed rhizome, as an assistant ingredient, clears heat and purges fire, promotes fluid production and quenches thirst, and is mainly used to treat feverish thirst, lung heat cough, and lung abscess with purulent sputum. It mainly contains organic acids, such as ferulic acid and p-coumaric acid, which have various biological activities such as antibacterial, anti-inflammatory, and antioxidant effects. Related technologies indicate that the Lung Heat Clearing and Detoxifying Decoction has antitussive, expectorant, anti-inflammatory, asthma-relieving, antibacterial, antiviral, and antipyretic effects. Therefore, morin A, ferulic acid, p-coumaric acid, and rutin are selected as fingerprint spectroscopy markers to identify the main chemical components for controlling the quality of the Lung Heat Clearing and Detoxifying Decoction.

[0029] This invention also provides the application of the aforementioned detection method in the quality control of Lung Heat Clearing and Decoction.

[0030] Compared with the prior art, the present invention has the following beneficial effects:

[0031] 1. This invention establishes for the first time an HPLC method for simultaneously detecting the contents of four active ingredients—morrhagic acid A, ferulic acid, p-coumaric acid, and rutin—in Lung Heat Clearing and Detoxifying Decoction, filling a gap in the existing technology.

[0032] 2. This invention optimizes chromatographic conditions, particularly by using methanol-0.1% phosphoric acid aqueous solution as the mobile phase and performing gradient elution, combined with wavelength switching technology (315nm and 257nm), to achieve good separation of the four target components within 40 minutes, with symmetrical peaks, no interference, and strong specificity.

[0033] 3. Methodological verification showed that the method of the present invention exhibited good linearity within its respective linear range (r≥0.9999), with RSDs of less than 3% in precision, repeatability, and stability tests, and recovery rates between 98.99% and 100.04%, indicating that the method is accurate and reliable and suitable for quantitative analysis of lung heat clearing and decoction.

[0034] 4. This method provides a scientific basis for the overall quality evaluation of Lung Heat Clearing and Detoxifying Decoction, and can be used for quality control in its production process to ensure the consistency of quality between different batches of products, which is of great significance for ensuring the safety and effectiveness of clinical medication. Attached Figure Description

[0035] Figure 1 This is the HPLC chromatogram for the specificity test;

[0036] Figure 2 High-performance liquid chromatography (HPLC) chromatograms at 315 nm for the test solution and mixed reference solution of Lung Heat Clearing and Detoxifying Decoction;

[0037] Figure 3 High-performance liquid chromatography (HPLC) chromatograms of the test solution and mixed reference solution of Lung Heat Clearing and Detoxifying Powder at a wavelength of 257 nm;

[0038] In the figure: 7. Morcinol A; 12. Ferulic acid; 13. Coumaric acid; 16. Rutin. Detailed Implementation

[0039] The present invention will be further described below with reference to specific embodiments, but is not limited thereto.

[0040] Example 1: A method for detecting the content of active ingredients in a lung-heat clearing and detoxifying decoction.

[0041] 1. Instruments and reagents

[0042] Instruments: The main instruments are a high-performance liquid chromatograph (Uitimate 3000, Thermo Fisher Scientific (China) Co., Ltd.), an electronic balance of 0.0001 g (XS205DU, Shanghai Qiuhe Instrument Co., Ltd.), an ultrasonic cleaner (SB-800DTD, Ningbo Xinzhi Biotechnology Co., Ltd., power 200W, frequency 50kHz), a UV-Vis spectrophotometer (T3202S, Shanghai Youke Instrument Co., Ltd.), and an electronic balance of 0.1 g (JA500, Shanghai Youke Instrument Co., Ltd.).

[0043] Reagents and reagents: Morcinol A reference standard (batch number 112086-202402, purity ≥98%), ferulic acid reference standard (batch number 110773-202316, purity ≥98%), p-coumaric acid reference standard (batch number 112037-202102, purity ≥98%), and rutin reference standard (batch number 100080-202513, purity ≥98%) were all purchased from the China National Institutes for Food and Drug Control. Methanol (chromatographic grade, batch number: 1123QH086, Tianjin Biaoshiqi Technology Development Co., Ltd.), methanol (analytical grade, batch number: 2024060110, Guangzhou Chemical Reagent Factory), phosphoric acid (analytical grade, batch number: 2024050209, Guangzhou Chemical Reagent Factory), and ultrapure water were used. Ten batches of samples of Lung Heat Clearing and Detoxifying Decoction (numbered S1-S10) were prepared by Huangpu People's Hospital of Zhongshan City according to an optimized process (prescription dosage, particle size 3.0mm, material-to-liquid ratio 1:25, decoction twice, 50 min each time). The main ingredients were the following Chinese herbal medicines in parts by weight: 0.99 parts of reed rhizome, 0.90 parts of fritillaria bulb, 0.90 parts of winter melon seed, 0.90 parts of mulberry bark, 0.75 parts of wild chrysanthemum, 0.75 parts of bamboo shavings, 0.75 parts of trichosanthes peel, 0.60 parts of plantain seed, 0.60 parts of earthworm, and 0.345 parts of licorice.

[0044] 2. Solution preparation

[0045] (1) Preparation of reference solution:

[0046] Accurately weigh 4.10, 2.20, 3.60, and 4.40 mg of morin A, ferulic acid, p-coumaric acid, and rutin reference standards, respectively, and dissolve them in methanol to prepare single reference stock solutions with concentrations of 0.820, 0.440, 0.720, and 0.880 mg / mL. Accurately transfer 1.463, 1.636, 2.000, and 1.273 mL of the above stock solutions to 200 mL volumetric flasks, dilute to the mark with methanol, and shake well to obtain a mixed reference solution containing 6.000 μg / mL morin A, 3.600 μg / mL ferulic acid, 7.200 μg / mL p-coumaric acid, and 5.600 μg / mL rutin.

[0047] (2) Preparation of the test solution:

[0048] Weigh out the following powdered herbs (particle size 2.5 mm): 0.990 g of reed rhizome, 0.900 g of fritillaria bulb, 0.900 g of winter melon seed, 0.900 g of mulberry bark, 0.750 g of wild chrysanthemum, 0.750 g of bamboo shavings, 0.750 g of trichosanthes peel, 0.600 g of plantain seed, 0.600 g of earthworm, and 0.345 g of licorice root. Place them in a 500 mL round-bottom flask. Add water at a ratio of 1:25 and decoct twice, 50 min each time. Filter with gauze, shake well, and obtain 0.069 g / mL of crude herb. -1 For the decoction extract of traditional Chinese medicine, take 5 mL of the extract, add methanol to a 10 mL volumetric flask and dilute to volume. Shake well and sonicate for 1 min (power 200 W, frequency 50 kHz). Remove, cool, and centrifuge at 10000 r / min for 15 min. Take the supernatant and filter it through a 0.22 μm microporous membrane. Take the filtrate as the test solution.

[0049] 3. Chromatographic conditions

[0050] Chromatographic column: SHIMADZU Shim-pack GIS C18 column (4.6mm×250mm, 5μm);

[0051] Mobile phase: methanol (A) - 0.1% phosphoric acid aqueous solution (B), gradient elution was performed under the following conditions, as shown in Table 1.

[0052] Table 1. Gradient elution conditions of the mobile phase

[0053]

[0054] Flow rate: 1.0 mL / min;

[0055] Column temperature: 30℃;

[0056] Detection wavelengths: 315 nm (0–18 min, morin A, ferulic acid, p-coumaric acid), 257 nm (18–40 min, rutin);

[0057] Injection volume: 5 μL.

[0058] 4. Determination method

[0059] Accurately pipette 5 μL each of the mixed reference solution and the test solution into the liquid chromatograph, and determine the chromatograms under the chromatographic conditions described above. The chromatogram of the test solution should show peaks with retention times (tR) consistent with those of morin A, ferulic acid, p-coumaric acid, and rutin in the reference solution chromatogram. Calculate the content of each component in the test solution based on peak area using the external standard method, and then convert the result to the content per gram of crude drug (mg / g).

[0060] Example 2: Methodological Investigation

[0061] 1) System suitability test

[0062] Under the above chromatographic conditions, the theoretical plate number calculated based on each component peak is not less than 5000, the resolution is greater than 1.5, and the tailing factor is between 0.95 and 1.05, indicating that the system has good suitability.

[0063] 2) Examination of linear relationships

[0064] Accurately weigh 2, 4, 6, 8, 10, 12, and 14 μL of the mixed reference solution from Example 1 and inject them according to the chromatographic conditions described in Example 1. Measure and record the peak areas. Plot the injection volume (μg) on ​​the x-axis (X) and the peak area on the y-axis (Y) to calculate the regression equation and correlation coefficient. The results show that the correlation coefficient r ≥ 0.999, indicating that morin A, ferulic acid, p-coumaric acid, and rutin have good linear relationships within their respective concentration ranges (see Table 2 below).

[0065] Table 2 Regression Equations and Linear Range

[0066]

[0067] 3) Precision test

[0068] Take 5 μL of the mixed control solution and inject it 6 times consecutively under the chromatographic conditions of Example 1. Record the peak area. Using peak 1 (morulin A) as a reference, calculate the relative standard deviation (RSD) of retention time for each common peak. The RSD of peak area is ≤ 0.11% (n=6) and ≤ 0.83% (n=6). The results show that the RSDs of peak area for morulin A, ferulic acid, p-coumaric acid, and rutin are 0.78%, 0.06%, 0.67%, and 0.80% (n=6), respectively, indicating that the instrument has good precision.

[0069] 4) Repeatability test

[0070] Six samples (S5) of the same batch of Lung Heat Clearing and Detoxifying Decoction were prepared in parallel according to the test solution preparation method in Example 1. Five μL of each sample was taken and injected for analysis according to the chromatographic conditions in Example 1. Using peak 1 (morrhuside A) as a reference, the retention time RSD of each common peak was calculated to be ≤ 0.28% (n=6); the peak area RSD was ≤ 2.33% (n=6). The RSDs of the peak areas of morrhuside A, ferulic acid, p-coumaric acid, and rutin were 1.27%, 1.23%, 1.43%, and 1.65% (n=6), respectively, indicating good reproducibility of the method.

[0071] 5) Stability test

[0072] The same test solution was taken and incubated at room temperature for 0, 4, 8, 12, 16, and 24 hours for analysis. Using peak 1 (morulin A) as a reference, the retention time RSD of each common peak was calculated to be ≤ 0.08% (n=6); the peak area RSD was ≤ 2.77% (n=6). The results showed that the RSDs of the peak areas for morulin A, ferulic acid, p-coumaric acid, and rutin were 2.18%, 2.17%, 2.45%, and 2.13% (n=6), respectively, indicating that the test solution was stable within 24 hours.

[0073] 6) Recovery test

[0074] Six test solutions were prepared using the same batch of Lung Heat Clearing and Detoxifying Decoction samples with known content. Appropriate amounts of morin A, ferulic acid, p-coumaric acid, and rutin were added respectively, following the method in Example 1. The solutions were injected and analyzed, and the peak areas of each component were recorded. The recovery rates were calculated. The average recoveries of morin A, ferulic acid, p-coumaric acid, and rutin were 99.73%, 99.98%, 100.04%, and 98.99%, respectively, with RSDs of 0.76%, 1.54%, 1.34%, and 1.01% (n=6), indicating good method accuracy. (See Table 3.)

[0075] Table 3 Results of the recovery test of four components in Lung Heat Clearing Oral Liquid (n=6)

[0076]

[0077] 7) Specificity test

[0078] Take 5 μL each of the blank solvent (methanol), the test solution of Lung Heat Clearing and Detoxifying Powder, the mixed reference solution, and the negative control solution of Lung Heat Clearing and Detoxifying Powder (lacking mulberry bark, prepared in Example 1), and inject them according to the chromatographic conditions in Example 1. Record the chromatogram. Figure 1 As shown, the results indicate that the test solution has corresponding chromatographic peaks at the same retention time as the mixed reference solution, the negative test sample has no interference, and the resolution of each component peak in the chromatogram of the test solution is >1.5.

[0079] Example 3: Establishment of fingerprint spectrum and identification of common peaks for Lung Heat Clearing and Detoxifying Decoction

[0080] Take 10 batches of lung heat-clearing herbs.

[0081] Lung-heat clearing and detoxifying decoction (S1-S10) was prepared using the same procedure as in Example 1. The samples were injected under the chromatographic conditions of Example 1, and the HPLC chromatograms were recorded and imported into the "Traditional Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2012 Version)" software. Using the chromatogram of sample S1 as a reference chromatogram, the time window width was set to 0.1 min. Multiple points were calibrated using the median method to generate a control fingerprint chromatogram R and an HPLC fingerprint chromatogram superimposed from 10 batches of test solutions (e.g., ...). Figure 1-2 A total of 17 common peaks were identified. The similarity of 10 batches of Lung Heat Clearing Decoction was between 0.990 and 0.999, as shown in Table 4. The chemical composition of each batch of samples was highly consistent, the overall quality difference was not significant, and the similarity was good.

[0082] Table 4. Similarity of fingerprint patterns of 10 batches of Lung Heat Clearing and Detoxifying Decoction

[0083]

[0084] Example 4: Sample Content Determination

[0085] Ten batches of Lung Heat Clearing and Detoxifying Decoction samples (S1-S10) were taken, and test solutions were prepared according to the method in Example 1. Then, 5 μL of the solution was injected under the chromatographic conditions in Example 1 to determine four index components. The peak areas were recorded, and the contents of the four components (calculated as mg / g crude drug) were calculated. The contents of these four index components in different batches of Lung Heat Clearing and Detoxifying Decoction are shown in Table 5 below. The contents of the four components in the 10 batches of samples showed certain differences, but the overall content was stable. This method can effectively perform quantitative analysis on them.

[0086] Table 5. Results of mass concentration determination of four components in 10 batches of Lung Heat Clearing and Detoxifying Decoction

[0087]

[0088] In summary, the method for detecting the content of morin A, ferulic acid, p-coumaric acid and rutin in the Lung Heat Clearing and Detoxifying Decoction established in this invention is simple to operate, accurate and reliable, highly specific and reproducible, and can be used for the quality control of Lung Heat Clearing and Detoxifying Decoction, providing technical support for ensuring the stability of its clinical efficacy.

Claims

1. A method for detecting the content of active ingredients in a lung-heat-clearing and detoxifying decoction, characterized in that, The active ingredients are morin A, ferulic acid, p-coumaric acid, and rutin, and the method includes the following steps: S1. Preparation of reference solution Accurately weigh the reference standards of morin A, ferulic acid, p-coumaric acid and rutin, dissolve and dilute them in methanol to prepare a mixed reference solution; S2. Preparation of the test solution Take the lung heat clearing and detoxifying powder as the test sample, accurately weigh it, add solvent to extract it, filter or centrifuge the resulting extract, and take the filtrate as the test sample solution. S3. Detection was performed using high-performance liquid chromatography (HPLC). The chromatographic conditions included: Chromatographic column: C18 column packed with octadecylsilane-bonded silica gel; Mobile phase: Methanol as phase A and 0.1% phosphoric acid aqueous solution as phase B, gradient elution was performed; Detection wavelength: The wavelength switching method is adopted. The detection wavelength is 315nm within 0-18 minutes and 257nm within 18-40 minutes. Column temperature: 25-35 ℃; Flow rate: 0.8-1.2 mL / min; S4. Measurement and Calculation Accurately pipette the reference solution and the test solution separately and inject them into the high-performance liquid chromatograph for determination. Qualitative identification is performed based on the retention time of the chromatographic peaks, and the content of each component is calculated using the external standard method based on the peak area.

2. The method for detecting the content of active ingredients in the lung-heat clearing and detoxifying decoction according to claim 1, characterized in that, The conditions for gradient elution in step S3 are as follows: 0-10 min, mobile phase A is 15%, and mobile phase B is 85%; Over 10-22 minutes, the mobile phase A decreased from 15% to 44%, and the mobile phase B decreased from 85% to 56%. Between 22 and 26 minutes, the mobile phase A decreased from 44% to 44.5%, and the mobile phase B decreased from 56% to 55.5%. Over 26-30 minutes, the mobile phase A decreased from 44.5% to 60%, and the mobile phase B decreased from 55.5% to 40%. Over 30-40 minutes, the mobile phase A decreased from 60% to 15%, and the mobile phase B decreased from 40% to 85%.

3. The method for detecting the content of active ingredients in the lung-heat clearing and detoxifying decoction according to claim 1, characterized in that, The chromatographic column in step S3 has the following specifications: 4.6 mm × 250 mm, 5 μm; column temperature: 30 °C; flow rate: 1.0 mL / min; injection volume: 5 μL.

4. The method for detecting the content of active ingredients in the lung-heat clearing and detoxifying decoction according to claim 1, characterized in that, The preparation method of the test solution in step S2 is as follows: take the extract of lung heat clearing and decoction, add methanol, sonicate, centrifuge, take the supernatant and filter it with a microporous membrane to obtain the solution.

5. The method for detecting the content of active ingredients in the lung-heat clearing and detoxifying decoction according to claim 4, characterized in that, In step S1, the volume ratio of the lung heat clearing and decoction extract to methanol is 1:1; the ultrasonic treatment power is 200W, the frequency is 50kHz, and the time is 1min; the centrifugation speed is 10000 r / min, and the time is 15min; the microporous filter membrane has a pore size of 0.22μm or 0.45μm.

6. The method for detecting the content of active ingredients in the lung-heat clearing and detoxifying decoction according to claim 1, characterized in that, In the mixed reference solution described in step S1, the concentrations of morin A, ferulic acid, coumaric acid, and rutin are 0.0120-0.0840 μg / mL, 0.0072-0.0504 μg / mL, 0.0144-0.1008 μg / mL, and 0.0112-0.0784 μg / mL.

7. The method for detecting the content of active ingredients in the lung-heat clearing and detoxifying decoction according to claim 6, characterized in that... The Lung Heat Clearing and Detoxifying Decoction is made from the following proportions of Chinese medicinal herbs by weight: Reed rhizome 0.5–1.5 parts, Fritillaria cirrhosa 0.5–1.2 parts, winter melon seed 0.5–1.2 parts, mulberry bark 0.5–1.2 parts, wild chrysanthemum 0.4–1.0 parts, bamboo shavings 0.4–1.0 parts, trichosanthes peel 0.4–1.0 parts, plantain seed 0.3–0.9 parts, earthworm 0.3–0.9 parts, licorice root 0.2–0.6 parts.

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